Investigation of NLRP3, GSDMD, IL-1β and IL-18 expression in pediatric Crohn's disease with perianal lesions

Investigation of NLRP3, GSDMD, IL-1β and IL-18 expression in pediatric Crohn's disease with perianal lesions | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Investigation of NLRP3, GSDMD, IL-1β and IL-18 expression in pediatric Crohn's disease with perianal lesions Hongding Wang, Zhongqin Jin This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7536822/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 4 You are reading this latest preprint version Abstract Introduction : Crohn's disease(CD) is a complex inflammatory disease with perianal lesions as one of its serious complications. Pyroptosis is an inflammatory form of programmed cell death, and the role of pyroptosis in CD intestinal epithelial cells is unknown. Methods : Between September 2022 and August 2024, we selected 60 children with a primary diagnosis of Crohn's disease from the Department of Gastroenterology at the Children's Hospital Affiliated to Soochow University as the case group, and divided them into a group with perianal lesions (PCD group) and a group without perianal lesions (Non-PCD group). Additionally, 40 children diagnosed with simple juvenile intestinal polyps during the same period were selected as the control group. All subjects underwent immunohistochemistry (IHC) to quantitatively and locally detect the expression of NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue. Result :Clinical samples confirmed that the staining scores for NLRP3, GSDMD, IL-1β, and IL-18 were significantly higher in the PCD group than in the Non-PCD group and the control group. Immunohistochemical localization revealed that NLRP3, GSDMD, IL-1β, and IL-18 were primarily expressed in epithelial cells and immune cells of the intestinal mucosal tissue. Compared with the control group and the non-PCD group, the staining extent and intensity of these markers were significantly increased in the PCD group. Conclusion : Our findings identify NLRP3, GSDMD, IL-1β, and IL-18 as key pyroptosis-associated biomarkers in PCD, contributing to the understanding of pyroptosis in the pathogenesis of PCD. Crohn’s disease Perianal lesions Pyroptosis Immunohistochemistry Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 1 Introduction Crohn's disease (CD) is an immune-mediated, chronic relapsing and remitting disease characterized by inflammation of the gastrointestinal tract, with a segmental distribution of lesions that can involve the entire GI tract from the mouth to the anus.(Elmasry and Ha)The epidemiological characteristics of CD have significant geographical differences. In China, although its incidence and prevalence are still lower than those in developed countries in Europe and the United States and in other parts of Asia, with the rapid development of the social economy and the improvement of the quality of life and the changes in dietary structure, coupled with the rapid increase in mental stress caused by the acceleration of the pace of modern life, these factors have jointly contributed to the rapid increase in the incidence of the disease. Compared with adults, CD in children is characterized by more severe clinical manifestations, more extensive lesions, longer disease duration, and a large genetic role, with as many as half of the children requiring surgery within 10 years of diagnosis(Kerur et al.; Mohan, Deswal and Bhardwaj). Perianal disease (PD) is an aggressive and disabling phenotype of Crohn's disease, and the combination of Crohn's disease with perianal lesions is collectively referred to as Perianal Crohn's disease (PCD)(Tarrant et al.). Children with PCD are often associated with a greater disease burden, lower quality of life, and more expensive medical care(Johnston, Hwang and Mattei). Perianal lesions may be the first symptom of Crohn's disease, whereas PCD often goes undetected due to its insidious onset, and it is difficult to distinguish PCD from common perianal disorders, which can easily lead to misdiagnosis. The cumulative incidence of perianal lesions in Crohn's disease increases with disease duration; in a large multicenter prospective database cohort of 6,600 pediatric Crohn's disease patients, perianal disease was reported in 21% of cases, and the estimated probability of perianal disease occurring over time at 2 months, 1 year, 3 years, and 5 years after diagnosis was 4%, 9%, 17%, and 26%(Yamamoto et al.), respectively.Recent studies have shown that the pathogenesis of PCD not only involves a sustained inflammatory response, but may also be closely related to genetic polymorphisms, imbalanced immune response, and epithelial-mesenchymal transition, among other complex factors(Donnelly et al.). Pyroptosis, a form of programmed cell death carried out by the gasdermin (GSDM) family of proteins, is central to the innate immune response and is characterized by activation of the NOD-like receptor pyrin structural domain 3 (NLRP3), cleavage of gasdermin D (GSDMD) proteins, and the formation of cell membrane pores(Zheng and Li). However, excessive pyroptosis leads to the release of large amounts of inflammatory mediators, which ultimately leads to cell death, tissue damage, and possibly autoimmune inflammation or septic shock(Mulvihill et al.). There is growing evidence that excessive focal death is involved in the pathogenesis of CD. Studies have revealed that increased levels of intrinsic cytokines are seen in both UC and CD patients, involving factors such as Interleukin-8 (IL-8), Interleukin-6 (IL-6), IL-1β and IL-18. During the active phase of IBD, the production of IL-1β and IL-18 in the intestinal mucosa is significantly increased, and caspase-1 activity in intestinal tissues and macrophages is also enhanced(Thinwa et al.).PCD shares many inflammatory and cytokine-mediated pathologic mechanisms with CD, however, the exact mechanisms are not fully understood. One study found that in the rectal mucosa, the expression of IL-1β and IL-6 was higher in PCD patients than in CD patients or healthy controls, while at the same time, in the perianal fistula tissues of PCD patients, the expression of interleukin-12p40 (IL-12p40), tumor necrosis factor-α (Tumor necrosis factor-α (TNF-α), IL-1β and IL-8 were up-regulated(Meima-van Praag, van Rijn, et al.).The structure of IL-18 is closest to that of IL-1β, and the current study showed that both molecules are involved in inflammatory bowel diseases.(Borthwick) Pyroptosis, as a specific form of necrosis, is closely related to the pathogenesis of inflammatory bowel disease. Existing studies have mainly focused on the construction of animal models of colitis and have led to inconsistent conclusions due to differences in experimental design and conditions.Therefore, the aim of the present study was to investigate whether the expression levels of NLRP3, GSDMD, IL-1β, and IL-18 in the intestinal mucosal tissues of perianal children with Crohn's disease The aim of this study is to investigate whether the expression levels of NLRP3, GSDMD, IL-1β and IL-18 in the intestinal mucosa of children with perianal Crohn's disease are different from those in the Crohn's disease group and the control group, and to explore the role of the NLRP3/GSDMD pathway in the pathogenesis of perianal Crohn's disease, so as to search for the potential therapeutic target of perianal Crohn's disease, and to provide theoretical evidence for the clinical treatment. 2 Materials and methods 2.1 Subjects of study Sixty children with Crohn's disease who were first diagnosed in the Department of Gastroenterology of Children's Hospital of Soochow University between September 2022 and August 2024 and met the following inclusion and exclusion criteria were selected as a case group.The children with Crohn's disease strictly conformed to the “Expert Consensus on the Diagnosis and Treatment of Inflammatory Bowel Disease in Childhood (2019 edition)”, “European Association of Pediatric Gastroenterology, Hepatology, and Nutrition Inflammatory Bowel Disease in Children and Adolescents Diagnostic Criteria of the Modified Porto Criteria for Diagnosis (2014 Edition)” diagnostic criteria. Eligible children were categorized into a group with perianal Crohn's disease (Perianal Crohn's disease, PCD group) and a group without perianal Crohn's disease (Non-perianal Crohn's disease, Non-PCD group) according to the 2003 American Gastroenterological Association (AGA) criteria(Sandborn et al.). Meanwhile, 40 children with simple juvenile intestinal polyps who were hospitalized during the same period were used as a control group, and there was no statistically significant difference in terms of gender and age compared to the case group (P > 0.05). All children included in the study and their families gave informed consent, and the study strictly adhered to the ethical guidelines of the Ethics Committee of the Affiliated Children's Hospital of Soochow University, and formal ethical approval was obtained from this committee (2023CS292). 2.1.1 Conditions for inclusion (1) CD group: children and adolescents aged 18 years and below; children with a first diagnosis of Crohn's disease who meet the above diagnostic criteria and are diagnosed and followed up in our hospital;children who have not been treated before colonoscopy, such as anti-inflammatory drugs, glucocorticoids, immunosuppressants, biologics, or enteral nutrition; and children who have complete data on all cases. (2) Control group: Given that simple juvenile intestinal polyps are a common condition for colonoscopy in children, and there is usually no sign of inflammation in the normal mucosal area, we selected 40 cases of normal intestinal polyps. Therefore, we selected 40 normal intestinal mucosal tissue samples from children with simple juvenile intestinal polyps who underwent endoscopic resection from their unaffected intestinal mucosal tissue areas as a control group. 2.1.2 Exclusion criteria (1) CD group: various kinds of enteritis of unknown diagnosis; combined with Clostridium difficile, intestinal tuberculosis and other bacterial infections;the presence of autoimmune disease children; suffering from serious heart, liver, kidney, lung and other diseases, unable to tolerate enteroscopy; the presence of serious complications, such as intestinal obstruction, perforation, toxic giant colon, etc.; not yet completed the proctoscopic diagnosis or enteroscopy (colonoscopy, small colonoscopy), ultrasound, perianal magnetic resonance and other examinations to exclude perianal lesions, Patients who have not completed rectal diagnosis or enteroscopy (colonoscopy, small enteroscopy), ultrasound, perianal nuclear magnetic examination to exclude perianal lesions. (2) Control group: children with blood in stool due to other gastrointestinal diseases; children with recent signs of respiratory, gastrointestinal, or other systemic infections; children with multiple polyps detected by colonoscopy, or children diagnosed with Peutz-Jeghers syndrome (PJS) and familial adenomatous polyposis (FAP); children with multiple polyps detected by colonoscopy, or children diagnosed with PJS and FAP. Children with specific polyp syndromes such as Peutz-Jeghers syndrome (PJS) and Familial adenomatous polyposis (FAP). 2.1.3 Grouping (1) Sixty children with Crohn's disease who were first admitted to the Department of Gastroenterology of Affiliated Children's Hospital of Soochow University between September 2022 and August 2024 were selected as the case group. Children with simple juvenile intestinal polyps admitted to the hospital during the same period were taken as the control group. (2) According to the criteria of the American Gastroenterological Association (AGA) in 2003, the children in the case group who met the criteria were categorized into the perianal lesion group or the no perianal lesion group. 2.2 Research methodology Intestinal mucosal specimen collection: All children and their families signed an informed consent for the procedure prior to gastroenteroscopy, and all underwent routine bowel preparation after admission to the hospital, followed by colonoscopy two to three days later. During the examination, two samples were collected from the intestinal mucosa with lesions in children with Crohn's disease and two samples were collected from the normal intestinal mucosa in children with juvenile intestinal polyps alone. The samples were treated in formalin solution and paraffin sections were made for subsequent immunohistochemical staining analysis. 2.3Immunohistochemistry (1) Paraffin sections: Pathology biopsies of children with CD and juvenile intestinal polyps were borrowed from the Department of Pathology of the Children's Hospital of Soochow University between September 2022 and August 2024, and then placed on ice, fixed one by one on a paraffin slicer, and then sliced continuously with a thickness of about 4 um. The CD group, the group without perianal lesions, and the control group were labeled. CD group, CD group without perianal lesions and control group. (2) In the first step, the sections were sequentially immersed in eco-friendly dewaxing solvents I, II, and III, with each eco-friendly dewaxing solvent being immersed for 10 minutes; in the second step, the sections were then sequentially transferred to anhydrous ethanol solutions I, II, and III, with each anhydrous ethanol solution being immersed for 5 minutes; and in the third step, the sections were thoroughly washed and treated with distilled water. (3) Antigen repair: pre-prepared citric acid antigen repair buffer (pH = 6.0), completely submerge the tissue sections in this buffer, and use the microwave oven to carry out antigen repair, the repair program is set as follows: heating for 10 minutes on medium heat, pause for 5 minutes; then continue to heat for 5 minutes on medium-low heat, pause again for 2 minutes; and finally heat for 5 minutes on medium-low heat, the whole process should be careful to prevent the sections from The whole process should be careful to prevent the slices from drying out. After the slices were cooled to room temperature, they were placed in a container containing phosphate buffer (pH = 7.4) and washed using a shaker for a total of 3 times, each time for 5 minutes. 4) Blocking endogenous peroxidase: The samples were placed in 3% hydrogen peroxide solution and incubated in a shaded environment for 25 minutes at room temperature. Afterwards, the slides were immersed in phosphate (PBS) buffer (pH 7.4) and washed by gentle shaking on a decolorizing shaker, repeating this step 3 times for 5 minutes each time. (5) Serum confinement: Add 3% bovine serum albumin (BSA) to the histochemical reaction system, ensuring that it evenly covers the tissue surface. Next, place the system at room temperature and allow the sealing action to continue for 30 minutes. (During this process, rabbit serum should be used for primary antibodies from goats, and BSA should be used for primary antibodies from other sources.) (6) Addition of primary antibody: pre-prepare antibodies (NLRP3, GSDMD, IL-1β, IL-18) diluted in the ratio of 1:200, gently shake the sealing solution, and carefully add the diluted primary antibody solution dropwise onto the tissue area of each slide, and in order to ensure that the antibody binds to the tissue adequately, place the slides in a wet box at 4°C for overnight incubation. (7) Addition of secondary antibody: Immerse the slides in PBS buffer at pH 7.4 and rinse with shaking for 5 minutes. After shaking the slide dry, HRP-labeled secondary antibody corresponding to the primary antibody was added dropwise, ensuring that the tissue was completely covered, and incubated for 50 minutes at room temperature. (8) DAB color development: Place the slide in PBS buffer at pH 7.4 and wash three times by shaking the bed, each time for 5 minutes. After washing, the excess liquid was shaken off and freshly prepared DAB reagent was added dropwise. The color development reaction was closely monitored under a microscope until the tissue sections showed brownish yellow color and then the slides were immediately rinsed with tap water to terminate the color development process. (9) Re-staining: In the first step, the slides were re-stained in hematoxylin staining solution for 3 minutes and then washed; in the second step, the slides were treated with differentiation solution for a few seconds to remove the excess staining and then washed again; and in the third step, the slides were thoroughly rinsed with water after being treated with re-bluing using re-bluing solution. (10) Dehydration and sealing: In the first step, the sections were sequentially immersed in 75% and 85% ethanol, and the duration of immersion in each type of ethanol was 5 minutes; in the second step, the sections were transferred to anhydrous ethanol solution I, anhydrous ethanol solution II, n-butanol, and xylene I, and the sections were immersed in each type of solution for 5 minutes in order to realize complete dehydration and transparency. After the slices were dried, they were sealed using sealing adhesive. Microscopic examination: Interpret the results under a white light microscope. Determination of results: Quantitative evaluation of tissue sections was performed using an intelligent image processing system, and the stained areas and their degree of staining were transformed into quantifiable data indices by using H-score (Histochemistry Score) to perform semi-quantitative analysis of tissue staining [31, 32]. h-score=∑(pi×i)=(weakly positive areas) Percentage × 1) + (Percentage of medium positive areas × 2) + (Percentage of strong positive areas × 3) (where pi denotes the proportion of positive signals to the total number of pixels or cells, and i denotes the intensity of different degrees of staining, which accurately reflects the staining characteristics of the sample). 2.3Statistical analysis SPSS 27.0 statistical software was applied for data processing, while GraphPadPrism9 and Adobe Illustrator 2024 software were utilized for graphing. Count data were expressed as number of cases and percentage [n(%)], and comparisons between two groups were made by chi-square test. Independent measurement data were first tested for normality, and data that conformed to normal distribution were described by mean ± standard deviation (± S), and comparisons between two groups were made by t-test, and comparisons between multiple groups that conformed to chi-square were analyzed by analysis of variance (ANOVA), while those that did not conform to chi-square were analyzed by Welch's ANOVA; those that did not conform to normal distribution were described by median (25th percentile, 75th percentile) [M(P25, P75 )] described by the Mann-Whitney rank-sum test for comparisons between two groups, the Kruskal-Wallis rank-sum test for comparisons between multiple groups, and the Bonferroni correction for all two-by-two comparisons after tests between multiple groups. For non-independent measures, the normality test for two-group differences was performed first, and the paired t-test was used for comparisons between two groups whose differences conformed to normal distribution; the Wilcoxon signed rank test was used for differences that did not conform to normal distribution. For the correlation test, normal distribution and linear trend analysis were performed first, and Pearson correlation analysis was used if both were consistent with each other, while Spearman correlation analysis was used if they were not consistent with each other to explore the correlation between the indicators. All results were considered statistically different at P < 0.05. Receiver Operating Characteristic (ROC) analysis was performed, and the area under curve (AUC) was calculated, the optimal critical value was selected based on the Jordon index, and the sensitivity and specificity were calculated. 3 Results 3.1General information on the subjects of the study The results of clinical data analysis showed that there were 60 cases in the case group, of which 36 cases (60.00%) were male and 24 cases (40.00%) were female, with an average age of (12.34 ± 2.14) years old; there were 40 cases in the control group, of which 22 cases (55.00%) were male and 18 cases (45.00%) were female, with an average age of (11.58 ± 1.77) years old; no statistically significant difference was found between the case and control groups in terms of gender and age ( P all > 0.05).There were 30 cases in the PCD group, of which 24 (80.00%) were male and 6 (20.00%) were female; there were 30 cases in the Non-PCD group, of which 12 (40.00%) were male and 18 (60.00%) were female cases; There was no statistically significant difference between the PCD group and the Non-PCD group in terms of PCDAI score, number of days of first hospitalization, and age ( P all > 0.05), and the difference between the PCD group and the Non-PCD group in terms of gender was statistically significant (P < 0.05). (For details, see Tables 1 and 2). Table 3 − 1 General information between the two groups, control and case group General information Control group (n = 40) Case group (n = 60) t/χ 2 P Sex[n(%)] male 22(55.00%) 36(60.00%) 0.246 0.620 female 18(45.00%) 24(40.00%) Age(year, \(\:\stackrel{-}{\varvec{X}}\) ±S) 11.58 ± 1.77 12.34 ± 2.14 1.848 0.068 Table 3 − 2 General information between the two groups of perianal lesions group and no perianal lesions group Group Sex [n(%)] Age (year, \(\:\stackrel{-}{\varvec{X}}\) ±S) First hospitalization days PCDAI PCD(n = 30) male 24(80.00%) 11.97 ± 1.90 13.26 ± 2.30 24.28 ± 8.01 female 6 (20.00%) Group Sex [n(%)] Age (year , \(\:\stackrel{-}{\varvec{X}}\) ±S) First hospitalization days PCDAI Non-PCD(n = 30) male 12(40.00%) 12.71 ± 2.34 13.40 ± 2.47 22.41 ± 6.43 female 18(60.00%) χ 2 /t 10.000 -1.338 -0.216 0.995 P 0.002 ** 0.186 0.830 0.324 3.2Types of perianal lesions The two most common types of perianal lesions in the PCD group were perianal abscess and anal fistula in 14 (46.67%) and 8 (26.67%) cases, respectively, and the remaining in order of prevalence were dermatomal and rectovaginal fistulae. All the children with perianal involvement were treated with simple medication in 22 cases and incision and drainage in 8 cases as shown in Fig. 1. 3.3Clinical typing In terms of lesion distribution: The PCD group and the Non-PCD group had 1 case (3.33%) and 6 cases (20.00%) of L1 type (ileal type), respectively, 2 cases (6.67%) and 2 cases (6.67%) of L2 type (colonic type), respectively, L3 type (ileocolonic type) were 11 cases (36.67%) and 8 cases (26.66%), respectively, and L4 type (upper gastrointestinal type) were 16 cases (53.33%) and 14 cases (46.67%), respectively. In terms of disease behavior: The PCD group and the Non-PCD group had 10 cases (33.33%) and 22 cases (73.33%) of Type B1 (non-stenotic/non-penetrating), respectively; 8 cases (26.67%) and 6 cases (20.00%) of Type B2 (stenotic), respectively; and 12 cases (40.00%) and 2 cases (6.67%) for type B3 (perforating), respectively. There was no statistically significant difference in lesion distribution between the two groups ( P > 0.05), but there was a statistically significant difference in lesion behavior ( P < 0.05), as shown in Table 3 – 3 . Table 3 3 Comparison of Montreal classification between the anal region lesion group and the non-anal region lesion group [n (%)] Group Location of the lesion Lesion behavior L1 L2 L3 L4 B1 B2 B3 PCD(n = 30) 1 (3.33%) 2 (6.67%) 11 (36.67%) 16 (53.33%) 10 (33.33%) 8 (26.67%) 12 (40.00%) Non-PCD (n = 30) 6 (20.00%) 2 (6.67%) 8 (26.66%) 14 (46.67%) 22 (73.33%) 6 (20.00%) 2 (6.67%) χ 2 4.154 ▼ 11.929 ▲ P 0.276 0.002 ** 3.4Clinical manifestation Comparative analysis of the initial clinical manifestations of each group: in the PCD group and Non-PCD group, in terms of digestive symptoms, abdominal pain was found in 10 cases (33.33%) and 15 cases (50.00%), diarrhea was found in 16 cases (53.33%) and 12 cases (40.00%), and blood in the stools was found in 3 cases (10.00%) and 6 cases (20.00%), respectively; and in terms of systemic symptoms, fever were 3 (10.00%) and 4 (13.33%), weight loss was 8 (26.67%) and 5 (16.67%), and anemia was 2 (6.67%) and 4 (13.33%), respectively; for extra-intestinal manifestations, oral ulcers were 10 (33.33%) and 7 (23.33%), respectively. Arthralgia was found in only 1 case (3.33%) in the PCD group. Initial symptoms were not statistically different between the PCD and Non-PCD groups (both P > 0.05), see Table 3 –4. Table 3 4 Comparison of initial symptoms between the two groups [n (%)] Group[n(%)] PCD(n = 30) Non-PCD(n = 30) χ 2 P digestive symptoms abdominal pain 10(33.33%) 15(50.00%) 1.714 ▲ 0.190 diarrhea 16(53.33%) 12(40.00%) 1.071 ▲ 0.301 blood in stool 3(10.00%) 6(20.00%) —— ▼ 0.472 systemic symptoms fever 3(10.00%) 4(13.33%) —— ▼ 1.000 weight loss 8(26.67%) 5(16.67%) 0.884 ▲ 0.347 anemia 2(6.67%) 4(13.33%) —— ▼ 0.671 extra-intestinal manifestations oral ulcers 10(33.33%) 7(23.33%) 0.739 ▲ 0.390 joint pain 1(3.33%) 0(0) —— ▼ 1.000 3.5Imaging Examination A total of 47 cases underwent small bowel CT angiography, with 25 cases in the PCD group and 22 cases in the Non-PCD group (83.33% vs. 73.33%). In the PCD group and Non-PCD group, the most common intestinal wall thickening was observed in 21 cases and 17 cases, respectively (84.00% vs. 77.27%). followed by mesenteric lymph node enlargement in 10 and 13 cases (40.00% vs. 59.09%), intestinal lumen narrowing in 9 and 4 cases (36.00% vs. 18.18%), and abdominal fluid accumulation in 5 and 3 cases (20.00% vs. 13.63%). intestinal wall enhancement in 7 and 6 cases (28.00% vs. 27.27%), and comb sign in 4 and 5 cases (16.00% vs. 22.72%) as shown in Table 3 –5. Table 3 5 Imaging comparison between the two groups [n (%)] Group PCD Non-PCD thickening of the intestinal wall 21(84.00%) 17(77.27%) multiple mesenteric lymph node enlargement 10(40.00%) 13(59.09%) intestinal stenosis 9(36.00%) 4(18.18%) abdominal fluid 5(20.00%) 3(13.63%) intestinal wall strengthening 7(28.00%) 6(27.27%) comb 4(16.00%) 5(22.72%) 3.6Comparison of immunohistochemical staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissues of the three groups of pediatric patients NLRP3 staining scores were 53.16 (49.60, 70.69) in the PCD group (Group A), 32.57 (28.30, 41.28) in the non-PCD group (Group B), and 16.34 (11.33, 19.15) in the control group (Group C); GSDMD staining scores were 43.94 (39.49, 51.69) in the PCD group, 35.76 (25.46, 42.07) in the Non-PCD group, and 15.67 (11.07, 21.04) in the control group ; IL-1β staining scores were 45.50 (39.47, 51.50) in the PCD group, 28.60 (21.29, 40.90) in the Non-PCD group, and 16.72 (9.29, 21.83) in the control group; IL-18 staining scores were 62.20 (50.06, 71.78) in the PCD group, 52.05 (37.47, 63.43) in the Non-PCD group, and 17.27 (9.04, 22.63) in the control group. There were statistically significant differences in the staining scores of NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue among the PCD group, Non-PCD group, and control group ( P < 0.01 for all). Further pairwise comparisons between groups revealed that in the PCD group, compared with both the Non-PCD group and the control group, the staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue were significantly elevated (P < 0.05 for all comparisons); in the Non-PCD group, compared with the control group, the staining scores for all four factors were significantly higher than those in the control group (P < 0.05). as shown in Table 3 –6. Table 3 6 Expression analysis of NLRP3, GSDMD, IL-1β, and IL-18 staining scores in three groups of pediatric patients ([M(P25,P75)]) Group A(n = 30) Group B(n = 30) Group C(n = 40) H P NLRP3 53.16(49.60,70.69) 32.57(28.30,41.28) 16.34(11.33,19.15) 75.549 ♦ <0.001 ** 0.004 ab** <0.001 ac** <0.001 bc** GSDMD 43.94(39.49,51.69) 35.76(25.46,42.07) 15.67(11.07,21.04) 68.300 ♦ <0.001 ** 0.006 ab** <0.001 ac** <0.001 bc** IL-1β 45.50(39.47,51.50) 28.60(21.29,40.90) 16.72(9.29,21.83) 63.776 ♦ <0.001 ** <0.001 ab** <0.001 ac** <0.001 bc** IL-18 62.20(50.06,71.78) 52.05(37.47,63.43) 17.27(9.04,22.63) 66.638 ♦ <0.001 ** 0.048 ab* <0.001 ac** <0.001 bc** 3.7Immunohistochemical staining localization of intestinal mucosal tissue specimens NLRP3 was expressed in intestinal epithelial cells, macrophages, and lymphocytes in both the case group and control group children, but the staining intensity was lower in the control group; GSDMD was expressed in intestinal epithelial cells and immune cells (such as macrophages, neutrophils, monocytes, T cells, and B cells) in both the case group and control group children, but GSDMD in the control group was mainly expressed as scattered cytoplasmic expression, specifically appearing as sparse yellow or brown granules scattered throughout the cytoplasm. In the case group, both the staining extent and intensity were increased compared to the control group; IL-1β and IL-18 were primarily expressed in intestinal epithelial cells, monocytes, macrophages, and lymphocytes of the lamina propria in both the case group and control group children. In the case group, strong staining of IL-1β and IL-18 was observed in intestinal mucosal specimens, with lymphocytes, macrophages, and intestinal epithelial cells showing moderate to strong positive expression, monocytes showed weak positive expression, while the control group exhibited overall lower staining intensity, with weak positive expression. Further comparison revealed that NLRP3, GSDMD, IL-1β, and IL-18 were enhanced in the CD group with perianal lesions compared to the CD group without perianal lesions, as shown in Figs. 3 − 2, 3–3, 3–4 and 3–5 (left-side magnification: X200; right-side magnification: X1000). Positive pyroptotic cells (arrows) 3.7 Correlation analysis of immunohistochemical staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in children with PCD A correlation analysis of immunohistochemical scores in the PCD group revealed that the immunohistochemical staining scores for GSDMD, IL-1β, and IL-18 were positively correlated with the NLRP3 immunohistochemical staining score (P < 0.01), with Spearman correlation coefficients of 0.798, 0.695, and 0.847, respectively. The immunohistochemical staining scores for IL-1β and IL-18 were positively correlated with the immunohistochemical staining score for GSDMD (P < 0.01), with Spearman correlation coefficients of 0.709 and 0.801, respectively; The immunohistochemical staining scores for IL-1β and IL-18 were positively correlated ( P < 0.01), with a correlation coefficient of 0.654, as shown in Table 3 –7. Table 3 7 Correlation analysis of immunohistochemical staining scores Speraman r P GSDMD and NLRP3 0.798 <0.001 ** IL-1β and NLRP3 0.695 <0.001 ** IL-18 and NLRP3 0.847 <0.001 ** IL-1β and GSDMD 0.709 <0.001 ** IL-18 and GSDMD 0.801 <0.001 ** IL-1β and IL-18 0.654 <0.001 ** 3.8 The independent predictive efficacy of immunohistochemical staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue of the PCD group on the occurrence of PCD Using children in the Non-PCD group as controls, we established ROC curves for NLRP3, GSDMD, IL-1β, and IL-18 immunohistochemical staining scores for the diagnosis of PCD. The AUC for NLRP3 staining scores was 0.878 (95% CI: 0.779–0.997), with an optimal cutoff value of 49.33, sensitivity of 83.30%, and specificity of 79.30%; The AUC for GSDMD staining scores was 0.770 (95% CI: 0.649–0.891), with an optimal cutoff value of 39.42, sensitivity of 66.70%, and specificity of 75.90%; The AUC of IL-1β staining scores was 0.870 (95% CI: 0.781–0.960), with an optimal cutoff value of 38.13, sensitivity of 73.30%, and specificity of 82.80%; The AUC for IL-18 staining scores was 0.680 (95% CI: 0.543–0.818), with an optimal cutoff value of 57.63, sensitivity of 65.50%, and specificity of 70.00%, as shown in Table 3 –8. Table 3 8 The independent predictive efficacy of NLRP3, GSDMD, IL-1β, and IL-18 immunohistochemical scores in intestinal mucosal tissue for the occurrence of PCD AUC 95% CI cutoff value sensitivity (%) specificity (%) P NLRP3 0.878 0.779–0.997 49.33 83.30 79.30 <0.001 * GSDMD 0.770 0.649–0.891 39.42 66.70 75.90 <0.001 ** IL-1β 0.870 0.781–0.960 38.13 73.30 82.80 <0.001 ** IL-18 0.680 0.543–0.818 57.63 65.50 70.00 0.017 * 3.9 The independent predictive efficacy of immunohistochemical staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue of the PCD group on the occurrence of PCD Based on a single indicator, ROC curves were constructed using multiple indicators to predict PCD. The results showed that: Area under the curve (AUC), sensitivity, and specificity were as follows: NLRP3 staining score + GSDMD staining score: 0.880 (95% CI: 0.780–0.981), 83.30%, 79.30%; NLRP3 staining score + IL-1β staining score: 0.957 (95% CI: 0.915–1.000), 90.00%, 86.20%; NLRP3 staining score + IL-18 staining score: 0.905 (95% CI: 0.821–0.988), 93.10%, 83.30%; NLRP3 + GSDMD + IL-1β + IL-18 staining score: 0.966 (95% CI: 0.924–1.000), 96.60%, 86.70%, see Table 3 –9. Table 3 9 Combined predictive efficacy of NLRP3, GSDMD, IL-1β, and IL-18 immunohistochemical scores in intestinal mucosal tissue for the occurrence of PCD joint inspection AUC 95%CI sensitivity (%) specificity (%) P NLRP3 + GSDMD 0.880 0.780–0.981 83.30 79.30 <0.001 ** NLRP + IL-1β 0.957 0.915-1.000 90.00 86.20 0.022 * NLRP3 + IL-18 0.905 0.821–0.988 93.10 83.30 <0.001 ** NLRP3 + GSDMD + IL-1β + IL-18 0.966 0.924-1.000 96.60 86.70 <0.001 ** 3.10 Treatment 3.10.1 Analysis of clinical remission rates in groups with and without perianal lesions This study evaluated the clinical treatment efficacy of pediatric Crohn's disease using the Pediatric Crohn's Disease Activity Index (PCDAI) score (Hyams et al.). During the disease remission phase, the PCDAI score typically ranges from 1 to 10 points. We scored the disease activity index of enrolled patients before and after treatment. The results showed that the PCDAI scores of patients with perianal lesions and those without perianal lesions before treatment were (24.28 ± 8.01) points and (22.41 ± 6.43) points, respectively, with no statistically significant difference ( P > 0.05). All patients were followed up for 3–6 months. Both groups showed a decreasing trend in PCDAI scores compared to pre-treatment levels, but there was no statistically significant difference in PCDAI scores between the two groups post-treatment (P > 0.05). There was a significant difference in clinical remission rates between the CD group with perianal lesions and the CD group without perianal lesions (73.33% vs. 96.67%, P < 0.05), see Table 3 –10 and Table 3 –11. Table 3 10 Comparison of PCDAI scores before and after treatment in the PCD group and Non-PCD group Group before treatment Post-treatment t P PCD(n = 30) 24.28 ± 8.01 8.58 ± 4.89 9.162 <0.001 ** Non-PCD(n = 30) 22.41 ± 6.43 6.16 ± 4.81 11.073 <0.001 ** t 0.995 1.927 P 0.324 0.059 Table 3 11 Comparison of clinical remission rates between PCD group and Non-PCD group Group relieve[n(%)] unrelieved[n(%)] PCD(n = 30) 22(73.33%) 8(26.67%) Non-PCD(n = 30) 29(96.67%) 1(3.33%) χ 2 ____ ▼ P 0.026 * 3.10.2Efficacy evaluation of IFX at week 14 in patients with or without perianal lesions In the group with perianal lesions, a total of 23 patients received IFX treatment, while in the group without perianal lesions, 18 patients received IFX treatment. Prior to IFX treatment, the PCDAI scores for the groups with and without perianal lesions were (22.04 ± 6.66) points and (21.22 ± 6.16) points, respectively. There was no statistically significant difference between the groups (P > 0.05). After 14 weeks of IFX treatment, the PCDAI scores in both groups decreased compared to pre-treatment levels. There was no statistically significant difference in PCDAI scores between the two groups post-treatment (P > 0.05). Among patients treated with IFX, there was no difference in clinical remission rates between the CD group with perianal lesions and the CD group without perianal lesions (82.60% vs 94.44%, P = 0.363). see Table 3 –12 and Table 3 –13. Table 3 12 Comparison of PCDAI scores before and after IFX treatment PCD group and Non-PCD group Group before treatment Post treatment t P PCD(n = 30) 22.04 ± 6.66 6.19 ± 5.10 9.057 <0.001 ** Non-PCD(n = 30) 21.22 ± 6.16 5.13 ± 3.97 9.308 <0.001 ** t 0.405 0.723 P 0.688 0.474 Table 3 13 Comparison of clinical remission rates between groups with and without perianal lesions treated with IFX Group relieve[n(%)] unrelieved[n(%)] PCD(n = 23) 19(82.60%) 4(17.40%) Non-PCD (n = 18) 17(94.44%) 1(5.56%) χ 2 ____ ▼ P 0.363 4. DISCUSSION Crohn's disease is a chronic transmural inflammatory bowel disease(Feroz et al.; Wasmann et al.; de Groof et al.). The incidence of Crohn's disease in children has risen rapidly over the past few decades. Compared to adults, the clinical course of Crohn's disease in childhood is particularly aggressive, with approximately 20–25% of patients developing the disease during childhood or adolescence (Wewer et al.)Perianal lesions are common in Crohn's disease. According to reports, the incidence of perianal lesions in Crohn's disease patients ranges from 20–40% globally (Tsai et al.; Williams and Shaffer), with higher rates in Asian countries, ranging from 30.3–58.8% (Agrawal and Jess). Anal lesions may manifest before or concurrently with intestinal symptoms. Some patients may not exhibit symptoms of anal lesions until several years after being diagnosed with Crohn's disease. A study found that approximately 26% of CD patients developed anal fistulas within 20 years of diagnosis(Tsai et al.), which is associated with higher rates of hospitalization, surgical intervention, immunosuppressive therapy, and reduced quality of life(Adegbola et al.). The presence of perianal lesions, particularly fistulas, in CD patients is a hallmark of the chronic and invasive nature of the disease (Lukin). The etiology of perianal lesions in Crohn's disease is diverse and complex, with its pathogenesis involving genetic polymorphisms, dysregulated immune responses, and epithelial-mesenchymal transition. 4.1 Clinical characteristics and influencing factors of PCD Regarding risk factors associated with PCD, current research findings have not yet reached a consensus. Previous literature has suggested that factors such as gender, age, disease duration, initial location of lesions, lesion location, and smoking history may be associated with the occurrence and development of PCD(Lukin). In our study, the PCD group had a higher proportion of males, with a male-to-female ratio of approximately 4:1. This ratio differed significantly from that of the Crohn's disease group without perianal lesions (P < 0.05). Our findings align with those of Mutanen A (Mutanen and Pakarinen), who also noted that gender is a risk factor for PCD, with male patients being more prone to the condition. This may be related to racial influences on the phenotype of inflammatory bowel disease. Western reports indicate a higher proportion of female PCD patients compared to males, while Asian countries show a higher prevalence of male PCD patients (Meima-van Praag, Buskens, et al.). In terms of the distribution characteristics of intestinal lesions, adult Crohn's disease patients in Western countries exhibit a relatively balanced trend in the proportions of L1, L2, and L3 lesions. In contrast, Chinese CD patients more commonly present with L3 lesions, while L4 lesions are relatively rare (Zeng et al.). Additionally, pediatric Crohn's disease differs significantly from adult cases in terms of disease phenotype and behavior (de Bie et al.). Currently, the Montreal classification system is widely used for categorizing anal lesions in pediatric Crohn's disease. In pediatric patients, the disease progression is characterized by greater invasiveness and diffuseness, resulting in a higher probability of lesions in the colon (L2) and upper gastrointestinal tract (L4) compared to adults. Statistically, approximately half of children and adolescents with Crohn's disease have upper gastrointestinal tract damage at the time of diagnosis(Assa, Rinawi and Shamir), and the data from this study is consistent with this. The proportion of L4 lesions was higher in both the perianal lesion group (53.33%) and the non-perianal lesion group (46.67%). SB Ingle's study noted that colonic involvement is an important predictive factor for Crohn's disease with perianal lesions, with a significantly increased risk of perianal lesions in patients with colonic Crohn's disease(Ingle and Loftus). The data from this study aligns with this finding, with a higher proportion of L3 cases in the perianal lesion group (36.67%) compared to the non-perianal lesion group (26.66%). It is generally believed that the B1 phenotype (non-penetrating/non-stricturing) at diagnosis is the most common presentation of Crohn's disease in both Eastern and Western patients(Dolinger, Torres and Vermeire). However, in our study, only 33.33% of children in the perianal lesion group had the B1 phenotype, while the B1 phenotype was predominant in the group without perianal lesions, accounting for 73.33%. A study by Braithwaite GC(Braithwaite et al.)found that the primary characteristic of the B3 type (penetrating) is a high incidence of fistulas. Compared with other types, patients with the B3 type have a significantly higher incidence of anal fistulas, and the incidence of anal fistula lesions also increases accordingly. In this study, Type B3 was more common in the anal perianal lesion group than in the group without anal perianal lesions, with proportions of 40.00% and 6.67%, respectively. The difference between the two groups was statistically significant (P < 0.05). This may be related to the classification of perianal fistulas as B3 type in this study. The specific impact of disease behavior on perianal lesions in CD requires further validation through large-scale, ongoing, and extensive research using a broader sample size. The typical initial presentation of Crohn's disease is often nonspecific, with common clinical manifestations including chronic diarrhea, abdominal pain, and weight loss. In children and adolescents, abdominal pain is typically the primary complaint (Moon). In this study, the Crohn's disease group without perianal lesions primarily presented with abdominal pain, while the PCD group exhibited a significant increase in diarrhea frequency, which may be related to the frequent stimulation and pressure on the perianal area caused by diarrhea(Bolshinsky and Church). Children in the PCD group exhibited relatively more extraintestinal manifestations, a finding consistent with previous relevant studies(Seyfried and Herold). However, there was no statistically significant difference in the initial clinical manifestations between the two groups (P > 0.05), which may be attributed to the diverse and nonspecific nature of Crohn's disease symptoms. 4.2Pyroptosis-related protein NLRP3 and its association with PCD Mutations in the NLRP3 gene are considered an important risk factor for the onset of IBD. Abnormally activated NLRP3 inflammasomes play a key role in the pathogenesis of UC (Klughammer et al.) and are also involved in the onset of CD (Sun et al.). In recent years, an increasing number of studies have suggested that uncontrolled activation of the NLRP3 inflammasome plays a key role in chronic intestinal inflammation, and the pathogenesis of colitis is associated with elevated levels of IL-1β and IL-18 (Chen et al.). Reduced NLRP3 production is associated with a lower risk of colitis [63, 64]. In animal studies, mice with elevated NLRP3 levels were more susceptible to dextran sulfate sodium salt (DSS) and 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis, but other experiments found that NLRP3-deficient mice exhibited more severe colitis after oral administration of DSS (Bauer et al.). In our study, to investigate the expression levels of NLRP3 and its mRNA in the intestinal mucosa of CD patients, we used a combined analysis of immunohistochemistry (IHC) and qRT-PCR. The results showed that the IHC staining scores and mRNA expression levels in the case group were higher than those in the control group (P < 0.05); Further intergroup analysis revealed that IHC staining scores and mRNA expression levels were higher in the CD group without perianal lesions and the PCD group compared to the control group, and also higher in the PCD group compared to the CD group without perianal lesions (P < 0.05 for all). Previous studies have shown that NLRP3 mRNA and protein expression is elevated in the intestinal mucosa of adult CD patients and positively correlated with disease severity(Ranson et al.) In rectal mucosa, IL-1β and IL-6 expression was higher in Crohn's disease patients with anal fistulas than in patients with luminal CD or healthy controls (Ruffolo et al.). Meanwhile, in perianal fistula tissue from Crohn's disease patients with anal fistulas, IL-12p40, TNF-α, IL-1β, and IL-8 expression was up-regulated (van Onkelen et al.). The expression of NLRP3 in the intestinal mucosa of the PCD group was higher than that in the CD group without perianal lesions, which may be related to the fact that NLRP3 can indirectly regulate the secretion of IL-1β and IL-18, promoting their synthesis and release (Fu and Wu). The results of this experiment are consistent with those of basic animal experiments and adult studies. Additionally, immunohistochemical staining of intestinal mucosa in this study showed that NLRP3 was sporadically expressed in the cytoplasm of intestinal epithelial cells, macrophages, and lymphocytes in the control group, with yellow or brown granules also visible in the cytoplasm. Compared with the control group, the staining range and intensity were increased in the case group, suggesting elevated NLRP3 levels in intestinal tissues of patients in the case group and indicating that NLRP3 is expressed in various cells of the immune system. Further comparison showed that NLRP3 staining range and intensity were enhanced in the CD group with perianal lesions compared with the CD group without perianal lesions (P > 0.05). 4.2Pyroptosis-related protein GSDMD and its association with PCD GSDMD is a key executor of pyroptosis and plays a crucial regulatory role in host immune defense, inflammatory responses, autoimmune diseases, and various other systemic disorders (Li et al.). The role of GSDMD in colitis remains controversial. On one hand, elevated GSDMD levels in mucosal biopsies from IBD patients suggest a pathogenic role for this protein in the disease (Bulek et al.). In TNBS-induced mouse colitis, high expression of GSDMD and GSDME in colonic epithelial cells supports this observation(Tan et al.). However, this is inconsistent with the severe colitis phenotype observed in another mouse strain lacking GSDMD (Ma et al.). This study aims to investigate the expression levels of GSDMD in intestinal mucosa using immunohistochemistry to address these knowledge gaps. Our findings revealed statistically significant differences in GSDMD intestinal mucosa IHC staining scores among the PCD group, Non-PCD group, and control group (P < 0.05 for all); Further intergroup analysis revealed that the GSDMD intestinal mucosa IHC staining scores in the Non-PCD and PCD groups were higher than those in the control group (P < 0.05), and the GSDMD intestinal mucosa IHC staining scores in the PCD group were higher than those in the Non-PCD group (P < 0.05). The expression of GSDMD in the intestinal mucosa of the PCD group was higher than that in the Non-PCD group. Considering that perianal lesions are predictive factors for the progression of Crohn's disease, this suggests a possible rapid progression from inflammation to stenosis or perforation phenotypes(Tarrant et al.). Additionally, immunohistochemical localization experiments showed that GSDMD was expressed in intestinal epithelial cells and immune cells (such as macrophages, neutrophils, monocytes, T cells, and B cells) in the intestinal mucosa of patients in the case group, while the control group showed minimal expression in intestinal epithelial cells and immune cells under the microscope. Further comparison revealed that GSDMD staining intensity and extent were enhanced in the CD group with perianal lesions compared to the CD group without perianal lesions, consistent with previous studies reporting GSDMD expression in immune cells and intestinal epithelial cells (Wang, Ding and Shao). 4.3Pyroptosis-related protein IL-1β and IL-18 and its association with PCD Interleukin-1β (IL-1β) is a key mediator of the inflammatory response, playing a role in both innate and adaptive immunity. It is crucial for the host's response to and resistance against pathogens and may exacerbate tissue damage in acute and chronic diseases(Cao et al.). IL-18 is a ubiquitous pro-inflammatory cytokine within the IL-1 family, structurally similar to the inflammatory factor IL-1β. IL-18 activates T cells, enhances immune responses, and plays a significant role as an inflammatory marker in inflammatory diseases (Detry et al.). Studies have shown that both UC and CD exhibit elevated levels of innate cytokines, including IL-8, IL-6, IL-1β, and IL-18. The production of IL-1β and IL-18 in the intestinal mucosa of patients with active IBD is increased (Thinwa et al.). The results of this study showed that the serum concentrations of IL-1β and IL-18, intestinal mucosa IHC staining scores, and mRNA expression levels were higher in the case group than in the control group (P < 0.05 for all), Further intergroup analysis showed that the IHC staining scores for IL-1β and IL-18 were higher in both the Non-PCD group and the PCD group compared to the control group, and also higher in the PCD group compared to the Non-PCD group (P < 0.05). This is consistent with clinical study results, which indicate that IL-1 secretion is increased in the colon tissue and macrophages of IBD patients and is positively correlated with disease severity (Perera et al.). Regarding the higher expression of IL-1β in the intestinal mucosa of the PCD group compared to the CD group without perianal lesions, we analyzed that this may be related to the following aspects: (1) Studies have shown that IL-1β and IL-6 expression levels in rectal mucosal tissue of Crohn's disease fistula patients are significantly higher than in patients with luminal-type Crohn's disease and healthy individuals (Ruffolo et al.). This study also found the same results, with IL-1β intestinal mucosa IHC staining scores in the PCD group being higher than in the Non-PCD group (P < 0.05). (2) Through an in-depth analysis of perianal fistula tissue in Crohn's disease patients with anal fistulas, researchers observed that inflammatory factors such as IL-12p40, TNF-α, IL-1β, and IL-8 showed a significant upward trend (van Onkelen et al.). IL-18 expression was higher in the PCD group than in the Non-PCD group, which may be associated with perianal lesions being an important predictive factor for disease progression in Crohn's disease, suggesting a rapid progression from inflammation to stricture or perforation phenotypes. IL-18 was highly expressed in intestinal epithelial cells of Crohn's disease patients in remission, indicating its primary protective function in the early stages of colitis. The absence of IL-18 may increase host susceptibility to chemically induced colitis. As disease severity increases, the number of IL-18-associated macrophages in active CD patients increases, and IL-18 expression in intestinal epithelial cells correspondingly rises (Lei-Leston, Murphy and Maloy). Immunohistochemical staining localization showed that the staining range and intensity of IL-1β and IL-18 were enhanced in the PCD group compared to the Non-PCD group. Previous studies have suggested that IL-1β is primarily produced by activated mononuclear phagocytes and can also be produced by macrophages, endothelial cells, and vascular smooth muscle cells(Wang et al.). The precursor of IL-18 is primarily present in the cytoplasm of monocytes, macrophages, and dendritic cells, as well as in endothelial cells, keratinocytes, and intestinal epithelial cells of the gastrointestinal tract (Ihim et al.). Our findings are consistent with previous results. 4.4 NLRP3, GSDMD, IL-1β, and IL-18 individually and in combination predict PCD efficacy In terms of intestinal mucosal immunohistochemical staining scores, NLRP3 had the largest area under the curve (AUC) of 0.878. When the NLRP3 staining score was 49.33, the diagnostic sensitivity was 83.30% and the specificity was 79.30%. The diagnostic performance of NLRP3 was superior to that of GSDMD, IL-1β, and IL-18. Mona(Watany, Elmazny and Nasif) et al. found in their study on the diagnosis of septic shock using NLRP3, IL-1β, and IL-18 that IL-1β had higher diagnostic efficacy than NLRP3; this aspect has not been explored in the present study. We conducted further combined diagnostic analysis of these indicators. The results showed that the four factors in the immunohistochemical staining score demonstrated higher efficacy in predicting the occurrence of PCD, with an AUC value of 0.966, a sensitivity of 96.60%, and a specificity of 86.70%. This suggests that the above indicators have a certain predictive efficacy for the occurrence of PCD and may be used as screening indicators for reference. 4.5NLRP3, GSDMD, IL-1β, IL-18 correlation NLRP3, GSDMD, IL-1β, and IL-18 are four factors located on the NLRP3/GSDMD pathway. A study found (Garnacho-Montero et al.)that the expression of NLRP3 on this pathway is positively correlated with IL-1β levels. Therefore, this study analyzed their correlation. The results showed that in our experiment, the key proteins NLRP3, GSDMD, IL-1β, and IL-18, the key proteins in the pyroptosis pathway, are pairwise correlated. In terms of immunohistochemical staining scores: the immunohistochemical staining scores of GSDMD, IL-1β, and IL-18 were positively correlated with the NLRP3 immunohistochemical staining score (P < 0.01), with Spearman correlation coefficients of 0.798, 0.695, and 0.847, respectively; The immunohistochemical staining scores for IL-1β and IL-18 were positively correlated with the immunohistochemical staining score for GSDMD (P < 0.01), with Spearman's correlation coefficients of 0.709 and 0.801, respectively; The immunohistochemical staining scores for IL-1β and IL-18 were positively correlated (P < 0.01), with a correlation coefficient of 0.654. By analyzing the associations among the four factors, this study not only validated that the pathogenesis of PCD indeed involves complex cascading signal transduction processes but also proposed new directions for exploration: the specific mechanisms underlying PCD have not yet been fully elucidated. Whether compensatory regulatory mechanisms exist between upstream and downstream factors within the same signal transduction pathway, and how these mechanisms regulate inflammatory responses, remain unclear and warrant further investigation in the future. 4.6 Efficacy evaluation The treatment goal for CD patients is to achieve and maintain disease remission. Short-term goals include alleviating clinical symptoms and restoring serum and fecal inflammatory markers to normal levels; long-term goals aim for complete resolution of clinical symptoms, sustained normalization of inflammatory markers, and endoscopic mucosal healing(Turner et al.). During the induction phase of remission, a combination of treatment modalities is typically employed, with corticosteroids, biologics, and enteral nutrition being the most commonly used approaches (Ong et al.). For children with fistulizing lesions, considering that fistula formation may exacerbate the condition and impair the child's quality of life, clinicians often prefer to initiate IFX therapy early. In contrast, the use of corticosteroids is more common in children without fistulizing lesions (Gecse et al.). In this study, 23 cases (76.67%) in the PCD group received IFX, which had a higher biologic agent usage rate compared to the 18 cases (60.00%) in the Non-PCD group, consistent with previous studies. This study used a standardized PCDAI scoring method before and after treatment to comprehensively assess the severity of anal lesions and the efficacy of drug therapy in pediatric patients. There was no statistically significant difference in PCDAI scores between the PCD group and the non-PCD group before treatment. After 3–6 months of treatment, PCDAI scores decreased significantly in both the PCD and non-PCD groups, but there was no statistically significant difference in PCDAI scores between the PCD and non-PCD groups after treatment (P > 0.05). Meanwhile, in this study, after 3–6 months of treatment, there was a difference in clinical remission rates between the PCD group and the Non-PCD group (73.33% vs. 96.67%, P = 0.026), which may be related to the more aggressive nature of PCD. We further analyzed that in children who achieved remission with IFX induction, the clinical remission rate was higher in the PCD group, and there was no significant difference in clinical remission rates between the two groups with or without perianal lesions (82.60% vs. 94.44%, P = 0.363). This is consistent with previous studies—a meta-analysis investigated the efficacy differences between the “early step-down” treatment strategy, which involves initiating biologic agents within two years of Crohn's disease diagnosis, and the traditional “step-up” therapy (i.e., considering biologic agent use more than two years after diagnosis). The results showed that compared with the traditional “step-up” therapy, the early introduction of biologics significantly improved patient outcomes, specifically manifested as higher clinical remission rates, significantly reduced disease recurrence rates, and a substantial increase in mucosal healing rates (Wasmann et al.). Additional data support that early initiation of IFX therapy not only leads to more patients achieving endoscopic disease remission but also effectively reduces the incidence of stricturing lesions(Schnitzler et al.). Therefore, for Crohn's disease patients, regardless of the presence of perianal lesions, infliximab therapy is recommended at the time of diagnosis. Conclusions In the PCD group, the expression levels of NLRP3, GSDMD, IL-1β, and IL-18 were all increased in intestinal mucosal tissue, and there was a positive correlation among the four factors, suggesting that the NLRP3/GSDMD signaling pathway may be involved in the pathogenesis of Crohn's disease with perianal lesions. The expression levels of NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue have certain predictive value for the diagnosis of Crohn's disease with perianal lesions in children, and the combination of these four factors can improve the predictive value of diagnosis. Declarations Competing interests The authors declare no competing interests. Department of Gastroenterology at the Children's Hospital Affiliated to Soochow University, Suzhou, China Funding sources he Jiangsu Provincial Key Medical Discipline (No. ZDXKA2016013) Author Contribution Wang Hongding writes the revised manuscript for Jin Zhongqin Data Availability Data used to support findings reported herein are available from the corresponding author upon request. References Adegbola SO et al (2020) Burden of Disease and Adaptation to Life in Patients with Crohn's Perianal Fistula: A Qualitative Exploration. Health Qual Life Outcomes 18(1):370 Print Agrawal M, Jess T (2022) Implications of the Changing Epidemiology of Inflammatory Bowel Disease in a Changing World. United Eur Gastroenterol J 10:1113–1120 Print Assa A, Rinawi F, Shamir R (2018) The Long-Term Predictive Properties of the Paris Classification in Paediatric Inflammatory Bowel Disease Patients. 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Am J Gastroenterol 103(12):3082–3093 Print Thinwa J et al Integrin-Mediated First Signal for Inflammasome Activation in Intestinal Epithelial Cells. J Immunol 193.3 (2014): 1373–1382. Print. Tsai L et al (2022) Epidemiology and Natural History of Perianal Crohn's Disease: A Systematic Review and Meta-Analysis of Population-Based Cohorts. Inflamm Bowel Dis 28:1477–1484 Print Turner D et al (2021) Stride-Ii: An Update on the Selecting Therapeutic Targets in Inflammatory Bowel Disease (Stride) Initiative of the International Organization for the Study of Ibd (Ioibd): Determining Therapeutic Goals for Treat-to-Target Strategies in Ibd. Gastroenterology 160(5):1570–1583 Print van Onkelen RS et al (2016) Pro-Inflammatory Cytokines in Cryptoglandular Anal Fistulas. Tech Coloproctol 20(9):619–625 Print Wang K, Ding J, Shao F (2023) Determination of Gasdermin Pores. Methods Mol Biol 2696:149–167 Print Wang P et al (2023) Role of Signal Transduction Pathways in Il-1β-Induced Apoptosis: Pathological and Therapeutic Aspects. Immun Inflamm Dis 11(1):e762 Print Wasmann KA et al (2020) Treatment of Perianal Fistulas in Crohn's Disease, Seton Versus Anti-Tnf Versus Surgical Closure Following Anti-Tnf [Pisa]: A Randomised Controlled Trial. J Crohns Colitis 14:1049–1056 Print Watany MM, Elmazny MI, Nasif EM (2020) Interleukin-31 Interaction with Inflammasome: A Promising Diagnostic and Prognostic Panel for Early Sepsis Identification in Critically Ill Patients. Cytokine 131:155102 Print Wewer MD et al (2021) The Incidence and Disease Course of Perianal Crohn's Disease: A Danish Nationwide Cohort Study, 1997–2015. J Crohns Colitis 15(1):5–13 Print Williams JL, Shaffer VO (2021) Modern Management of Perianal Crohn's Disease: A Review. Am Surg 87:1361–1367 Print Yamamoto T et al (2023) Diagnosis and Clinical Features of Perianal Lesions in Newly Diagnosed Crohn's Disease: Subgroup Analysis from Inception Cohort Registry Study of Patients with Crohn's Disease (Icrest-Cd). J Crohns Colitis 17:1193–1206 Print Zeng Z et al (2013) Incidence and Clinical Characteristics of Inflammatory Bowel Disease in a Developed Region of Guangdong Province, China: A Prospective Population-Based Study. J Gastroenterol Hepatol 28:1148–1153 Print Zheng Z, Li G (2020) Mechanisms and Therapeutic Regulation of Pyroptosis in Inflammatory Diseases and Cancer. Int J Mol Sci 21.4 Print. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Reviewers invited by journal 02 Jan, 2026 Editor assigned by journal 10 Sep, 2025 Submission checks completed at journal 09 Sep, 2025 First submitted to journal 04 Sep, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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10:52:34","extension":"html","order_by":14,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":191747,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-7536822/v1/75c442ae1a67287251645864.html"},{"id":99602151,"identity":"b99bb89d-a088-4d22-9047-7e15d210b632","added_by":"auto","created_at":"2026-01-06 10:52:34","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":110740,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eFigure 3-1 Distribution of perianal lesion types in the PCD group\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Picture1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7536822/v1/c3c61755e1527c74cee9d4f5.jpg"},{"id":99602159,"identity":"6856c6ae-49ab-44b9-b498-ba1a915727e6","added_by":"auto","created_at":"2026-01-06 10:52:34","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":2313341,"visible":true,"origin":"","legend":"\u003cp\u003eFigure 3-2 NLRP3 expression results under the microscope in three groups (a) perianal lesion group, (b) no perianal lesion group, (c) control group.\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7536822/v1/3d89397bb398fea342a45344.jpeg"},{"id":99602153,"identity":"d88d1f11-cf7a-4bad-bb1e-36a508ccdc32","added_by":"auto","created_at":"2026-01-06 10:52:34","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":2096714,"visible":true,"origin":"","legend":"\u003cp\u003eFigure 3-3 GSDMD expression results under three groups of microscopes (a) perianal lesion group, (b) no perianal lesion group, (c) control group\u003c/p\u003e","description":"","filename":"floatimage3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7536822/v1/c52741048aa300e29752c1e4.jpeg"},{"id":99602160,"identity":"a2469f51-c1a9-4b43-a451-4a2520203687","added_by":"auto","created_at":"2026-01-06 10:52:34","extension":"jpeg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":2383339,"visible":true,"origin":"","legend":"\u003cp\u003eFigure 3-4 IL-1β expression results under the microscope in three groups (a) perianal lesion group, (b) no perianal lesion group, and (c) control group.\u003c/p\u003e","description":"","filename":"floatimage4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7536822/v1/96038e72809ceff6731f870a.jpeg"},{"id":99602157,"identity":"f283f92a-19d9-4aee-8d00-1c2947d57ccc","added_by":"auto","created_at":"2026-01-06 10:52:34","extension":"jpeg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":2309740,"visible":true,"origin":"","legend":"\u003cp\u003eFigure 3-5 IL-18 expression results under the microscope in three groups (a) perianal lesion group, (b) no perianal lesion group, (c) control group\u003c/p\u003e","description":"","filename":"floatimage5.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7536822/v1/a1a30cfe25075be124edc4ea.jpeg"},{"id":99804097,"identity":"404ae511-7e33-4a02-9b10-522d7dd9cb23","added_by":"auto","created_at":"2026-01-08 14:11:45","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":10917826,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7536822/v1/5a39071a-0c5e-471a-b041-b96d798bbf84.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Investigation of NLRP3, GSDMD, IL-1β and IL-18 expression in pediatric Crohn's disease with perianal lesions","fulltext":[{"header":"1 Introduction","content":"\u003cp\u003eCrohn's disease (CD) is an immune-mediated, chronic relapsing and remitting disease characterized by inflammation of the gastrointestinal tract, with a segmental distribution of lesions that can involve the entire GI tract from the mouth to the anus.(Elmasry and Ha)The epidemiological characteristics of CD have significant geographical differences. In China, although its incidence and prevalence are still lower than those in developed countries in Europe and the United States and in other parts of Asia, with the rapid development of the social economy and the improvement of the quality of life and the changes in dietary structure, coupled with the rapid increase in mental stress caused by the acceleration of the pace of modern life, these factors have jointly contributed to the rapid increase in the incidence of the disease. Compared with adults, CD in children is characterized by more severe clinical manifestations, more extensive lesions, longer disease duration, and a large genetic role, with as many as half of the children requiring surgery within 10 years of diagnosis(Kerur et al.; Mohan, Deswal and Bhardwaj).\u003c/p\u003e \u003cp\u003ePerianal disease (PD) is an aggressive and disabling phenotype of Crohn's disease, and the combination of Crohn's disease with perianal lesions is collectively referred to as Perianal Crohn's disease (PCD)(Tarrant et al.). Children with PCD are often associated with a greater disease burden, lower quality of life, and more expensive medical care(Johnston, Hwang and Mattei). Perianal lesions may be the first symptom of Crohn's disease, whereas PCD often goes undetected due to its insidious onset, and it is difficult to distinguish PCD from common perianal disorders, which can easily lead to misdiagnosis. The cumulative incidence of perianal lesions in Crohn's disease increases with disease duration; in a large multicenter prospective database cohort of 6,600 pediatric Crohn's disease patients, perianal disease was reported in 21% of cases, and the estimated probability of perianal disease occurring over time at 2 months, 1 year, 3 years, and 5 years after diagnosis was 4%, 9%, 17%, and 26%(Yamamoto et al.), respectively.Recent studies have shown that the pathogenesis of PCD not only involves a sustained inflammatory response, but may also be closely related to genetic polymorphisms, imbalanced immune response, and epithelial-mesenchymal transition, among other complex factors(Donnelly et al.).\u003c/p\u003e \u003cp\u003ePyroptosis, a form of programmed cell death carried out by the gasdermin (GSDM) family of proteins, is central to the innate immune response and is characterized by activation of the NOD-like receptor pyrin structural domain 3 (NLRP3), cleavage of gasdermin D (GSDMD) proteins, and the formation of cell membrane pores(Zheng and Li). However, excessive pyroptosis leads to the release of large amounts of inflammatory mediators, which ultimately leads to cell death, tissue damage, and possibly autoimmune inflammation or septic shock(Mulvihill et al.). There is growing evidence that excessive focal death is involved in the pathogenesis of CD. Studies have revealed that increased levels of intrinsic cytokines are seen in both UC and CD patients, involving factors such as Interleukin-8 (IL-8), Interleukin-6 (IL-6), IL-1β and IL-18. During the active phase of IBD, the production of IL-1β and IL-18 in the intestinal mucosa is significantly increased, and caspase-1 activity in intestinal tissues and macrophages is also enhanced(Thinwa et al.).PCD shares many inflammatory and cytokine-mediated pathologic mechanisms with CD, however, the exact mechanisms are not fully understood. One study found that in the rectal mucosa, the expression of IL-1β and IL-6 was higher in PCD patients than in CD patients or healthy controls, while at the same time, in the perianal fistula tissues of PCD patients, the expression of interleukin-12p40 (IL-12p40), tumor necrosis factor-α (Tumor necrosis factor-α (TNF-α), IL-1β and IL-8 were up-regulated(Meima-van Praag, van Rijn, et al.).The structure of IL-18 is closest to that of IL-1β, and the current study showed that both molecules are involved in inflammatory bowel diseases.(Borthwick)\u003c/p\u003e \u003cp\u003ePyroptosis, as a specific form of necrosis, is closely related to the pathogenesis of inflammatory bowel disease. Existing studies have mainly focused on the construction of animal models of colitis and have led to inconsistent conclusions due to differences in experimental design and conditions.Therefore, the aim of the present study was to investigate whether the expression levels of NLRP3, GSDMD, IL-1β, and IL-18 in the intestinal mucosal tissues of perianal children with Crohn's disease The aim of this study is to investigate whether the expression levels of NLRP3, GSDMD, IL-1β and IL-18 in the intestinal mucosa of children with perianal Crohn's disease are different from those in the Crohn's disease group and the control group, and to explore the role of the NLRP3/GSDMD pathway in the pathogenesis of perianal Crohn's disease, so as to search for the potential therapeutic target of perianal Crohn's disease, and to provide theoretical evidence for the clinical treatment.\u003c/p\u003e"},{"header":"2 Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Subjects of study\u003c/h2\u003e \u003cp\u003e Sixty children with Crohn's disease who were first diagnosed in the Department of Gastroenterology of Children's Hospital of Soochow University between September 2022 and August 2024 and met the following inclusion and exclusion criteria were selected as a case group.The children with Crohn's disease strictly conformed to the \u0026ldquo;Expert Consensus on the Diagnosis and Treatment of Inflammatory Bowel Disease in Childhood (2019 edition)\u0026rdquo;, \u0026ldquo;European Association of Pediatric Gastroenterology, Hepatology, and Nutrition Inflammatory Bowel Disease in Children and Adolescents Diagnostic Criteria of the Modified Porto Criteria for Diagnosis (2014 Edition)\u0026rdquo; diagnostic criteria. Eligible children were categorized into a group with perianal Crohn's disease (Perianal Crohn's disease, PCD group) and a group without perianal Crohn's disease (Non-perianal Crohn's disease, Non-PCD group) according to the 2003 American Gastroenterological Association (AGA) criteria(Sandborn et al.). Meanwhile, 40 children with simple juvenile intestinal polyps who were hospitalized during the same period were used as a control group, and there was no statistically significant difference in terms of gender and age compared to the case group (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05). All children included in the study and their families gave informed consent, and the study strictly adhered to the ethical guidelines of the Ethics Committee of the Affiliated Children's Hospital of Soochow University, and formal ethical approval was obtained from this committee (2023CS292).\u003c/p\u003e \u003cdiv id=\"Sec4\" class=\"Section3\"\u003e \u003ch2\u003e2.1.1 Conditions for inclusion\u003c/h2\u003e \u003cp\u003e \u003cb\u003e(1)\u003c/b\u003e CD group: children and adolescents aged 18 years and below; children with a first diagnosis of Crohn's disease who meet the above diagnostic criteria and are diagnosed and followed up in our hospital;children who have not been treated before colonoscopy, such as anti-inflammatory drugs, glucocorticoids, immunosuppressants, biologics, or enteral nutrition; and children who have complete data on all cases.\u003c/p\u003e \u003cp\u003e \u003cb\u003e(2)\u003c/b\u003e Control group: Given that simple juvenile intestinal polyps are a common condition for colonoscopy in children, and there is usually no sign of inflammation in the normal mucosal area, we selected 40 cases of normal intestinal polyps. Therefore, we selected 40 normal intestinal mucosal tissue samples from children with simple juvenile intestinal polyps who underwent endoscopic resection from their unaffected intestinal mucosal tissue areas as a control group.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section3\"\u003e \u003ch2\u003e2.1.2 Exclusion criteria\u003c/h2\u003e \u003cp\u003e(1) CD group: various kinds of enteritis of unknown diagnosis; combined with Clostridium difficile, intestinal tuberculosis and other bacterial infections;the presence of autoimmune disease children; suffering from serious heart, liver, kidney, lung and other diseases, unable to tolerate enteroscopy; the presence of serious complications, such as intestinal obstruction, perforation, toxic giant colon, etc.; not yet completed the proctoscopic diagnosis or enteroscopy (colonoscopy, small colonoscopy), ultrasound, perianal magnetic resonance and other examinations to exclude perianal lesions, Patients who have not completed rectal diagnosis or enteroscopy (colonoscopy, small enteroscopy), ultrasound, perianal nuclear magnetic examination to exclude perianal lesions.\u003c/p\u003e \u003cp\u003e(2) Control group: children with blood in stool due to other gastrointestinal diseases; children with recent signs of respiratory, gastrointestinal, or other systemic infections; children with multiple polyps detected by colonoscopy, or children diagnosed with Peutz-Jeghers syndrome (PJS) and familial adenomatous polyposis (FAP); children with multiple polyps detected by colonoscopy, or children diagnosed with PJS and FAP. Children with specific polyp syndromes such as Peutz-Jeghers syndrome (PJS) and Familial adenomatous polyposis (FAP).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section3\"\u003e \u003ch2\u003e2.1.3 Grouping\u003c/h2\u003e \u003cp\u003e(1) Sixty children with Crohn's disease who were first admitted to the Department of Gastroenterology of Affiliated Children's Hospital of Soochow University between September 2022 and August 2024 were selected as the case group. Children with simple juvenile intestinal polyps admitted to the hospital during the same period were taken as the control group.\u003c/p\u003e \u003cp\u003e(2) According to the criteria of the American Gastroenterological Association (AGA) in 2003, the children in the case group who met the criteria were categorized into the perianal lesion group or the no perianal lesion group.\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.2 Research methodology\u003c/h2\u003e \u003cp\u003eIntestinal mucosal specimen collection: All children and their families signed an informed consent for the procedure prior to gastroenteroscopy, and all underwent routine bowel preparation after admission to the hospital, followed by colonoscopy two to three days later. During the examination, two samples were collected from the intestinal mucosa with lesions in children with Crohn's disease and two samples were collected from the normal intestinal mucosa in children with juvenile intestinal polyps alone. The samples were treated in formalin solution and paraffin sections were made for subsequent immunohistochemical staining analysis.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e2.3Immunohistochemistry\u003c/h2\u003e \u003cp\u003e(1) Paraffin sections: Pathology biopsies of children with CD and juvenile intestinal polyps were borrowed from the Department of Pathology of the Children's Hospital of Soochow University between September 2022 and August 2024, and then placed on ice, fixed one by one on a paraffin slicer, and then sliced continuously with a thickness of about 4 um. The CD group, the group without perianal lesions, and the control group were labeled. CD group, CD group without perianal lesions and control group.\u003c/p\u003e \u003cp\u003e(2) In the first step, the sections were sequentially immersed in eco-friendly dewaxing solvents I, II, and III, with each eco-friendly dewaxing solvent being immersed for 10 minutes; in the second step, the sections were then sequentially transferred to anhydrous ethanol solutions I, II, and III, with each anhydrous ethanol solution being immersed for 5 minutes; and in the third step, the sections were thoroughly washed and treated with distilled water.\u003c/p\u003e \u003cp\u003e(3) Antigen repair: pre-prepared citric acid antigen repair buffer (pH\u0026thinsp;=\u0026thinsp;6.0), completely submerge the tissue sections in this buffer, and use the microwave oven to carry out antigen repair, the repair program is set as follows: heating for 10 minutes on medium heat, pause for 5 minutes; then continue to heat for 5 minutes on medium-low heat, pause again for 2 minutes; and finally heat for 5 minutes on medium-low heat, the whole process should be careful to prevent the sections from The whole process should be careful to prevent the slices from drying out. After the slices were cooled to room temperature, they were placed in a container containing phosphate buffer (pH\u0026thinsp;=\u0026thinsp;7.4) and washed using a shaker for a total of 3 times, each time for 5 minutes.\u003c/p\u003e \u003cp\u003e4) Blocking endogenous peroxidase: The samples were placed in 3% hydrogen peroxide solution and incubated in a shaded environment for 25 minutes at room\u003c/p\u003e \u003cp\u003etemperature. Afterwards, the slides were immersed in phosphate (PBS) buffer (pH 7.4) and washed by gentle shaking on a decolorizing shaker, repeating this step 3 times for 5 minutes each time.\u003c/p\u003e \u003cp\u003e(5) Serum confinement: Add 3% bovine serum albumin (BSA) to the histochemical reaction system, ensuring that it\u003c/p\u003e \u003cp\u003eevenly covers the tissue surface. Next, place the system at room temperature and allow the sealing action to continue for 30 minutes. (During this process, rabbit serum should be used for primary antibodies from goats, and BSA should be used for primary antibodies from other sources.)\u003c/p\u003e \u003cp\u003e(6) Addition of primary antibody: pre-prepare antibodies (NLRP3, GSDMD, IL-1β, IL-18) diluted in the ratio of 1:200, gently shake the sealing solution, and carefully add the diluted primary antibody solution dropwise onto the tissue area of each slide, and in order to ensure that the antibody binds to the tissue adequately, place the slides in a wet box at 4\u0026deg;C for overnight incubation.\u003c/p\u003e \u003cp\u003e(7) Addition of secondary antibody: Immerse the slides in PBS buffer at pH 7.4 and rinse with shaking for 5 minutes. After shaking the slide dry, HRP-labeled secondary antibody corresponding to the primary antibody was added dropwise, ensuring that the tissue was completely covered, and incubated for 50 minutes at room temperature.\u003c/p\u003e \u003cp\u003e(8) DAB color development: Place the slide in PBS buffer at pH 7.4 and wash three times by shaking the bed, each time\u003c/p\u003e \u003cp\u003efor 5 minutes. After washing, the excess liquid was shaken off and freshly prepared DAB reagent was added dropwise. The color development reaction was closely monitored under a microscope until the tissue sections showed brownish yellow color and then the slides were immediately rinsed with tap water to terminate the color development process.\u003c/p\u003e \u003cp\u003e(9) Re-staining: In the first step, the slides were re-stained in hematoxylin staining solution for 3 minutes and then washed; in the second step,\u003c/p\u003e \u003cp\u003ethe slides were treated with differentiation solution for a few seconds to remove the excess staining and then washed again; and in the third step, the slides were thoroughly rinsed with water after being treated with re-bluing using re-bluing solution.\u003c/p\u003e \u003cp\u003e(10) Dehydration and sealing: In the first step, the sections were sequentially immersed in 75% and 85% ethanol, and the duration of immersion in each type of ethanol\u003c/p\u003e \u003cp\u003ewas 5 minutes; in the second step, the sections were transferred to anhydrous ethanol solution I, anhydrous ethanol solution II, n-butanol, and xylene I, and the sections were immersed in each type of solution for 5 minutes in order to realize complete dehydration and transparency. After the slices were dried, they were sealed using sealing adhesive.\u003c/p\u003e \u003cp\u003e \u003col\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eMicroscopic examination: Interpret the results under a white light microscope.\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eDetermination of results: Quantitative evaluation of tissue sections was performed using an intelligent image processing system, and the stained areas and their degree of staining were transformed into quantifiable data indices by using H-score (Histochemistry Score) to perform semi-quantitative analysis of tissue staining [31, 32]. h-score=\u0026sum;(pi\u0026times;i)=(weakly positive areas) Percentage \u0026times; 1) + (Percentage of medium positive areas \u0026times; 2) + (Percentage of strong positive areas \u0026times; 3) (where pi denotes the proportion of positive signals to the total number of pixels or cells, and i denotes the intensity of different degrees of staining, which accurately reflects the staining characteristics of the sample).\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003c/ol\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e2.3Statistical analysis\u003c/h2\u003e \u003cp\u003eSPSS 27.0 statistical software was applied for data processing, while GraphPadPrism9 and Adobe Illustrator 2024 software were utilized for graphing. Count data were expressed as number of cases and percentage [n(%)], and comparisons between two groups were made by chi-square test. Independent measurement data were first tested for normality, and data that conformed to normal distribution were described by mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation (\u0026plusmn;\u0026thinsp;S), and comparisons between two groups were made by t-test, and comparisons between multiple groups that conformed to chi-square were analyzed by analysis of variance (ANOVA), while those that did not conform to chi-square were analyzed by Welch's ANOVA; those that did not conform to normal distribution were described by median (25th percentile, 75th percentile) [M(P25, P75 )] described by the Mann-Whitney rank-sum test for comparisons between two groups, the Kruskal-Wallis rank-sum test for comparisons between multiple groups, and the Bonferroni correction for all two-by-two comparisons after tests between multiple groups. For non-independent measures, the normality test for two-group differences was performed first, and the paired t-test was used for comparisons between two groups whose differences conformed to normal distribution; the Wilcoxon signed rank test was used for differences that did not conform to normal distribution. For the correlation test, normal distribution and linear trend analysis were performed first, and Pearson correlation analysis was used if both were consistent with each other, while Spearman correlation analysis was used if they were not consistent with each other to explore the correlation between the indicators. All results were considered statistically different at P\u0026thinsp;\u0026lt;\u0026thinsp;0.05.\u003c/p\u003e \u003cp\u003eReceiver Operating Characteristic (ROC) analysis was performed, and the area under curve (AUC) was calculated, the optimal critical value was selected based on the Jordon index, and the sensitivity and specificity were calculated.\u003c/p\u003e \u003c/div\u003e"},{"header":"3 Results","content":"\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003e3.1General information on the subjects of the study\u003c/h2\u003e \u003cp\u003eThe results of clinical data analysis showed that there were 60 cases in the case group, of which 36 cases (60.00%) were male and 24 cases (40.00%) were female, with an average age of (12.34\u0026thinsp;\u0026plusmn;\u0026thinsp;2.14) years old; there were 40 cases in the control group, of which 22 cases (55.00%) were male and 18 cases (45.00%) were female, with an average age of (11.58\u0026thinsp;\u0026plusmn;\u0026thinsp;1.77) years old; no statistically significant difference was found between the case and control groups in terms of gender and age (\u003cem\u003eP\u003c/em\u003e all \u0026gt;\u0026thinsp;0.05).There were 30 cases in the PCD group, of which 24 (80.00%) were male and 6 (20.00%) were female; there were 30 cases in the Non-PCD group, of which 12 (40.00%) were male and 18 (60.00%) were female cases; There was no statistically significant difference between the PCD group and the Non-PCD group in terms of PCDAI score, number of days of first hospitalization, and age (\u003cem\u003eP\u003c/em\u003e all \u0026gt;\u0026thinsp;0.05), and the difference between the PCD group and the Non-PCD group in terms of gender was statistically significant (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05). (For details, see Tables\u0026nbsp;1 and 2).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e\u0026thinsp;\u0026minus;\u0026thinsp;1 General information between the two groups, control and case group\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGeneral information\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eControl group\u003c/p\u003e \u003cp\u003e(n\u0026thinsp;=\u0026thinsp;40)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eCase group\u003c/p\u003e \u003cp\u003e(n\u0026thinsp;=\u0026thinsp;60)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003et/χ\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSex[n(%)]\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e22(55.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e36(60.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e0.246\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e0.620\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003efemale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e18(45.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e24(40.00%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAge(year, \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\stackrel{-}{\\varvec{X}}\\)\u003c/span\u003e\u003c/span\u003e\u0026plusmn;S)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e11.58\u0026thinsp;\u0026plusmn;\u0026thinsp;1.77\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e12.34\u0026thinsp;\u0026plusmn;\u0026thinsp;2.14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.848\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.068\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e\u0026thinsp;\u0026minus;\u0026thinsp;2 General information between the two groups of perianal lesions group and no perianal lesions group\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGroup\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSex\u003c/p\u003e \u003cp\u003e[n(%)]\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAge\u003c/p\u003e \u003cp\u003e(year, \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\stackrel{-}{\\varvec{X}}\\)\u003c/span\u003e\u003c/span\u003e\u0026plusmn;S)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eFirst hospitalization days\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003ePCDAI\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003ePCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emale 24(80.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e11.97\u0026thinsp;\u0026plusmn;\u0026thinsp;1.90\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e13.26\u0026thinsp;\u0026plusmn;\u0026thinsp;2.30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e24.28\u0026thinsp;\u0026plusmn;\u0026thinsp;8.01\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003efemale 6 (20.00%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eGroup\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003eSex\u003c/b\u003e\u003c/p\u003e \u003cp\u003e\u003cb\u003e[n(%)]\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003eAge\u003c/b\u003e\u003c/p\u003e \u003cp\u003e\u003cb\u003e(year\u003c/b\u003e, \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\stackrel{-}{\\varvec{X}}\\)\u003c/span\u003e\u003c/span\u003e\u003cb\u003e\u0026plusmn;S)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003eFirst hospitalization days\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003ePCDAI\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eNon-PCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emale 12(40.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e12.71\u0026thinsp;\u0026plusmn;\u0026thinsp;2.34\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e13.40\u0026thinsp;\u0026plusmn;\u0026thinsp;2.47\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e22.41\u0026thinsp;\u0026plusmn;\u0026thinsp;6.43\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003efemale 18(60.00%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eχ\u003csup\u003e2\u003c/sup\u003e/t\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e-1.338\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-0.216\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.995\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.002\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.186\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.830\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.324\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003e3.2Types of perianal lesions\u003c/h2\u003e \u003cp\u003eThe two most common types of perianal lesions in the PCD group were perianal abscess and anal fistula in 14 (46.67%) and 8 (26.67%) cases, respectively, and the remaining in order of prevalence were dermatomal and rectovaginal fistulae. All the children with perianal involvement were treated with simple medication in 22 cases and incision and drainage in 8 cases as shown in Fig.\u0026nbsp;1.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e3.3Clinical typing\u003c/h2\u003e \u003cp\u003eIn terms of lesion distribution: The PCD group and the Non-PCD group had 1 case (3.33%) and 6 cases (20.00%) of L1 type (ileal type), respectively, 2 cases (6.67%) and 2 cases (6.67%) of L2 type (colonic type), respectively, L3 type (ileocolonic type) were 11 cases (36.67%) and 8 cases (26.66%), respectively, and L4 type (upper gastrointestinal type) were 16 cases (53.33%) and 14 cases (46.67%), respectively. In terms of disease behavior: The PCD group and the Non-PCD group had 10 cases (33.33%) and 22 cases (73.33%) of Type B1 (non-stenotic/non-penetrating), respectively; 8 cases (26.67%) and 6 cases (20.00%) of Type B2 (stenotic), respectively; and 12 cases (40.00%) and 2 cases (6.67%) for type B3 (perforating), respectively. There was no statistically significant difference in lesion distribution between the two groups (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;0.05), but there was a statistically significant difference in lesion behavior (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05), as shown in Table\u0026nbsp;\u0026lt;link rid=\"tb13\"\u0026gt;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u0026lt;/link\u0026gt;\u003c/span\u003e\u0026ndash;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e3 Comparison of Montreal classification between the anal region lesion group and the non-anal region lesion group [n (%)]\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"9\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colspan=\"2\" morerows=\"1\" nameend=\"c2\" namest=\"c1\" rowspan=\"2\"\u003e \u003cp\u003eGroup\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c6\" namest=\"c3\"\u003e \u003cp\u003eLocation of the lesion\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"3\" nameend=\"c9\" namest=\"c7\"\u003e \u003cp\u003eLesion behavior\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eL1\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eL2\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eL3\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eL4\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eB1\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eB2\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eB3\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003ePCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1\u003c/p\u003e \u003cp\u003e(3.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2\u003c/p\u003e \u003cp\u003e(6.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e11\u003c/p\u003e \u003cp\u003e(36.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e16\u003c/p\u003e \u003cp\u003e(53.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e10\u003c/p\u003e \u003cp\u003e(33.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e8\u003c/p\u003e \u003cp\u003e(26.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e12\u003c/p\u003e \u003cp\u003e(40.00%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eNon-PCD (n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6\u003c/p\u003e \u003cp\u003e(20.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2\u003c/p\u003e \u003cp\u003e(6.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e8\u003c/p\u003e \u003cp\u003e(26.66%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e14\u003c/p\u003e \u003cp\u003e(46.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e22\u003c/p\u003e \u003cp\u003e(73.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e6\u003c/p\u003e \u003cp\u003e(20.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e2\u003c/p\u003e \u003cp\u003e(6.67%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eχ\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"4\" nameend=\"c6\" namest=\"c3\"\u003e \u003cp\u003e4.154\u003csup\u003e▼\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"3\" nameend=\"c9\" namest=\"c7\"\u003e \u003cp\u003e11.929\u003csup\u003e▲\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"4\" nameend=\"c5\" namest=\"c2\"\u003e \u003cp\u003e0.276\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"3\" nameend=\"c8\" namest=\"c6\"\u003e \u003cp\u003e0.002\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c9\" namest=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e3.4Clinical manifestation\u003c/h2\u003e \u003cp\u003eComparative analysis of the initial clinical manifestations of each group: in the PCD group and Non-PCD group, in terms of digestive symptoms, abdominal pain was found in 10 cases (33.33%) and 15 cases (50.00%), diarrhea was found in 16 cases (53.33%) and 12 cases (40.00%), and blood in the stools was found in 3 cases (10.00%) and 6 cases (20.00%), respectively; and in terms of systemic symptoms, fever were 3 (10.00%) and 4 (13.33%), weight loss was 8 (26.67%) and 5 (16.67%), and anemia was 2 (6.67%) and 4 (13.33%), respectively; for extra-intestinal manifestations, oral ulcers were 10 (33.33%) and 7 (23.33%), respectively. Arthralgia was found in only 1 case (3.33%) in the PCD group. Initial symptoms were not statistically different between the PCD and Non-PCD groups (both \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;0.05), see Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;4.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e4 Comparison of initial symptoms between the two groups [n (%)]\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGroup[n(%)]\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eNon-PCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eχ\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003edigestive symptoms\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eabdominal pain\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10(33.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e15(50.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1.714\u003csup\u003e▲\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.190\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ediarrhea\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e16(53.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e12(40.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1.071\u003csup\u003e▲\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.301\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eblood in stool\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3(10.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6(20.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026mdash;\u0026mdash;\u003csup\u003e▼\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.472\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003esystemic symptoms\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003efever\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3(10.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4(13.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026mdash;\u0026mdash;\u003csup\u003e▼\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eweight loss\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8(26.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5(16.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.884\u003csup\u003e▲\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.347\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eanemia\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2(6.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4(13.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026mdash;\u0026mdash;\u003csup\u003e▼\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.671\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003eextra-intestinal manifestations\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eoral ulcers\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10(33.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e7(23.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.739\u003csup\u003e▲\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.390\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ejoint pain\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1(3.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0(0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026mdash;\u0026mdash;\u003csup\u003e▼\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003e3.5Imaging Examination\u003c/h2\u003e \u003cp\u003eA total of 47 cases underwent small bowel CT angiography, with 25 cases in the PCD group and 22 cases in the Non-PCD group (83.33% vs. 73.33%). In the PCD group and Non-PCD group, the most common intestinal wall thickening was observed in 21 cases and 17 cases, respectively (84.00% vs. 77.27%). followed by mesenteric lymph node enlargement in 10 and 13 cases (40.00% vs. 59.09%), intestinal lumen narrowing in 9 and 4 cases (36.00% vs. 18.18%), and abdominal fluid accumulation in 5 and 3 cases (20.00% vs. 13.63%). intestinal wall enhancement in 7 and 6 cases (28.00% vs. 27.27%), and comb sign in 4 and 5 cases (16.00% vs. 22.72%) as shown in Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;5.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e5 Imaging comparison between the two groups [n (%)]\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGroup\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePCD\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eNon-PCD\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ethickening of the intestinal wall\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e21(84.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e17(77.27%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003emultiple mesenteric lymph node enlargement\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10(40.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e13(59.09%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eintestinal stenosis\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e9(36.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4(18.18%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eabdominal fluid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5(20.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3(13.63%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eintestinal wall strengthening\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7(28.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6(27.27%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ecomb\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4(16.00%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5(22.72%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003e3.6Comparison of immunohistochemical staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissues of the three groups of pediatric patients\u003c/b\u003e \u003c/p\u003e \u003cp\u003eNLRP3 staining scores were 53.16 (49.60, 70.69) in the PCD group (Group A), 32.57 (28.30, 41.28) in the non-PCD group (Group B), and 16.34 (11.33, 19.15) in the control group (Group C); GSDMD staining scores were 43.94 (39.49, 51.69) in the PCD group, 35.76 (25.46, 42.07) in the Non-PCD group, and 15.67 (11.07, 21.04) in the control group ; IL-1β staining scores were 45.50 (39.47, 51.50) in the PCD group, 28.60 (21.29, 40.90) in the Non-PCD group, and 16.72 (9.29, 21.83) in the control group; IL-18 staining scores were 62.20 (50.06, 71.78) in the PCD group, 52.05 (37.47, 63.43) in the Non-PCD group, and 17.27 (9.04, 22.63) in the control group. There were statistically significant differences in the staining scores of NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue among the PCD group, Non-PCD group, and control group (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.01 for all).\u003c/p\u003e \u003cp\u003eFurther pairwise comparisons between groups revealed that in the PCD group, compared with both the Non-PCD group and the control group, the staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue were significantly elevated (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 for all comparisons); in the Non-PCD group, compared with the control group, the staining scores for all four factors were significantly higher than those in the control group \u003cem\u003e(P\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). as shown in Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;6.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab6\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e6 Expression analysis of NLRP3, GSDMD, IL-1β, and IL-18 staining scores in three groups of pediatric patients ([M(P25,P75)])\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGroup A(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eGroup B(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eGroup C(n\u0026thinsp;=\u0026thinsp;40)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eH\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNLRP3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e53.16(49.60,70.69)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e32.57(28.30,41.28)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e16.34(11.33,19.15)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e75.549\u003csup\u003e\u0026diams;\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.004\u003csup\u003eab**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003eac**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003ebc**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGSDMD\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e43.94(39.49,51.69)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e35.76(25.46,42.07)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e15.67(11.07,21.04)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e68.300\u003csup\u003e\u0026diams;\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.006\u003csup\u003eab**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003eac**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003ebc**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-1β\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e45.50(39.47,51.50)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e28.60(21.29,40.90)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e16.72(9.29,21.83)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e63.776\u003csup\u003e\u0026diams;\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003eab**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003eac**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003ebc**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e62.20(50.06,71.78)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e52.05(37.47,63.43)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e17.27(9.04,22.63)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e66.638\u003csup\u003e\u0026diams;\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.048\u003csup\u003eab*\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003eac**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003ebc**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e3.7Immunohistochemical staining localization of intestinal mucosal tissue specimens\u003c/h2\u003e \u003cp\u003eNLRP3 was expressed in intestinal epithelial cells, macrophages, and lymphocytes in both the case group and control group children, but the staining intensity was lower in the control group; GSDMD was expressed in intestinal epithelial cells and immune cells (such as macrophages, neutrophils, monocytes, T cells, and B cells) in both the case group and control group children, but GSDMD in the control group was mainly expressed as scattered cytoplasmic expression, specifically appearing as sparse yellow or brown granules scattered throughout the cytoplasm. In the case group, both the staining extent and intensity were increased compared to the control group; IL-1β and IL-18 were primarily expressed in intestinal epithelial cells, monocytes, macrophages, and lymphocytes of the lamina propria in both the case group and control group children. In the case group, strong staining of IL-1β and IL-18 was observed in intestinal mucosal specimens, with lymphocytes, macrophages, and intestinal epithelial cells showing moderate to strong positive expression, monocytes showed weak positive expression, while the control group exhibited overall lower staining intensity, with weak positive expression. Further comparison revealed that NLRP3, GSDMD, IL-1β, and IL-18 were enhanced in the CD group with perianal lesions compared to the CD group without perianal lesions, as shown in Figs.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026thinsp;\u0026minus;\u0026thinsp;2, 3\u0026ndash;3, 3\u0026ndash;4 and 3\u0026ndash;5 (left-side magnification: X200; right-side magnification: X1000). Positive pyroptotic cells (arrows)\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003e3.7 Correlation analysis of immunohistochemical staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in children with PCD\u003c/b\u003e \u003c/p\u003e \u003cp\u003eA correlation analysis of immunohistochemical scores in the PCD group revealed that the immunohistochemical staining scores for GSDMD, IL-1β, and IL-18 were positively correlated with the NLRP3 immunohistochemical staining score (P\u0026thinsp;\u0026lt;\u0026thinsp;0.01), with Spearman correlation coefficients of 0.798, 0.695, and 0.847, respectively. The immunohistochemical staining scores for IL-1β and IL-18 were positively correlated with the immunohistochemical staining score for GSDMD (P\u0026thinsp;\u0026lt;\u0026thinsp;0.01), with Spearman correlation coefficients of 0.709 and 0.801, respectively; The immunohistochemical staining scores for IL-1β and IL-18 were positively correlated (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.01), with a correlation coefficient of 0.654, as shown in Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;7.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab7\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e7 Correlation analysis of immunohistochemical staining scores\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c3\" namest=\"c2\"\u003e \u003cp\u003eSperaman\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003er\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGSDMD and NLRP3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.798\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-1β and NLRP3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.695\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-18 and NLRP3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.847\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-1β and GSDMD\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.709\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-18 and GSDMD\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.801\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-1β and IL-18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.654\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003e3.8 The independent predictive efficacy of immunohistochemical staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue of the PCD group on the occurrence of PCD\u003c/b\u003e \u003c/p\u003e \u003cp\u003eUsing children in the Non-PCD group as controls, we established ROC curves for NLRP3, GSDMD, IL-1β, and IL-18 immunohistochemical staining scores for the diagnosis of PCD. The AUC for NLRP3 staining scores was 0.878 (95% CI: 0.779\u0026ndash;0.997), with an optimal cutoff value of 49.33, sensitivity of 83.30%, and specificity of 79.30%; The AUC for GSDMD staining scores was 0.770 (95% CI: 0.649\u0026ndash;0.891), with an optimal cutoff value of 39.42, sensitivity of 66.70%, and specificity of 75.90%; The AUC of IL-1β staining scores was 0.870 (95% CI: 0.781\u0026ndash;0.960), with an optimal cutoff value of 38.13, sensitivity of 73.30%, and specificity of 82.80%; The AUC for IL-18 staining scores was 0.680 (95% CI: 0.543\u0026ndash;0.818), with an optimal cutoff value of 57.63, sensitivity of 65.50%, and specificity of 70.00%, as shown in Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;8.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab8\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e8 The independent predictive efficacy of NLRP3, GSDMD, IL-1β, and IL-18 immunohistochemical scores in intestinal mucosal tissue for the occurrence of PCD\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"7\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAUC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e95% CI\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ecutoff value\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003esensitivity\u003c/p\u003e \u003cp\u003e(%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003especificity\u003c/p\u003e \u003cp\u003e(%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNLRP3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.878\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.779\u0026ndash;0.997\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e49.33\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e83.30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e79.30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGSDMD\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.770\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.649\u0026ndash;0.891\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e39.42\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e66.70\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e75.90\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-1β\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.870\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.781\u0026ndash;0.960\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e38.13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e73.30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e82.80\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIL-18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.680\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.543\u0026ndash;0.818\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e57.63\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e65.50\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e70.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.017\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003e3.9 The independent predictive efficacy of immunohistochemical staining scores for NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue of the PCD group on the occurrence of PCD\u003c/b\u003e \u003c/p\u003e \u003cp\u003eBased on a single indicator, ROC curves were constructed using multiple indicators to predict PCD. The results showed that: Area under the curve (AUC), sensitivity, and specificity were as follows: NLRP3 staining score\u0026thinsp;+\u0026thinsp;GSDMD staining score: 0.880 (95% CI: 0.780\u0026ndash;0.981), 83.30%, 79.30%; NLRP3 staining score\u0026thinsp;+\u0026thinsp;IL-1β staining score: 0.957 (95% CI: 0.915\u0026ndash;1.000), 90.00%, 86.20%; NLRP3 staining score\u0026thinsp;+\u0026thinsp;IL-18 staining score: 0.905 (95% CI: 0.821\u0026ndash;0.988), 93.10%, 83.30%; NLRP3\u0026thinsp;+\u0026thinsp;GSDMD\u0026thinsp;+\u0026thinsp;IL-1β\u0026thinsp;+\u0026thinsp;IL-18 staining score: 0.966 (95% CI: 0.924\u0026ndash;1.000), 96.60%, 86.70%, see Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;9.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab9\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e9 Combined predictive efficacy of NLRP3, GSDMD, IL-1β, and IL-18 immunohistochemical scores in intestinal mucosal tissue for the occurrence of PCD\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003ejoint inspection\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAUC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e95%CI\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003esensitivity\u003c/p\u003e \u003cp\u003e(%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003especificity\u003c/p\u003e \u003cp\u003e(%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNLRP3\u0026thinsp;+\u0026thinsp;GSDMD\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.880\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.780\u0026ndash;0.981\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e83.30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e79.30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNLRP\u0026thinsp;+\u0026thinsp;IL-1β\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.957\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.915-1.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e90.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e86.20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.022\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNLRP3\u0026thinsp;+\u0026thinsp;IL-18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.905\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.821\u0026ndash;0.988\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e93.10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e83.30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNLRP3\u0026thinsp;+\u0026thinsp;GSDMD\u0026thinsp;+\u0026thinsp;IL-1β\u0026thinsp;+\u0026thinsp;IL-18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.966\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.924-1.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e96.60\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e86.70\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003e3.10 Treatment\u003c/h2\u003e \u003cdiv id=\"Sec18\" class=\"Section3\"\u003e \u003ch2\u003e3.10.1 Analysis of clinical remission rates in groups with and without perianal lesions\u003c/h2\u003e \u003cp\u003eThis study evaluated the clinical treatment efficacy of pediatric Crohn's disease using the Pediatric Crohn's Disease Activity Index (PCDAI) score (Hyams et al.). During the disease remission phase, the PCDAI score typically ranges from 1 to 10 points. We scored the disease activity index of enrolled patients before and after treatment. The results showed that the PCDAI scores of patients with perianal lesions and those without perianal lesions before treatment were (24.28\u0026thinsp;\u0026plusmn;\u0026thinsp;8.01) points and (22.41\u0026thinsp;\u0026plusmn;\u0026thinsp;6.43) points, respectively, with no statistically significant difference (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;0.05). All patients were followed up for 3\u0026ndash;6 months. Both groups showed a decreasing trend in PCDAI scores compared to pre-treatment levels, but there was no statistically significant difference in PCDAI scores between the two groups post-treatment (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05). There was a significant difference in clinical remission rates between the CD group with perianal lesions and the CD group without perianal lesions (73.33% vs. 96.67%, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05), see Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;10 and Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;11.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab10\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e10 Comparison of PCDAI scores before and after treatment in the PCD group and Non-PCD group\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGroup\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ebefore treatment\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePost-treatment\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003et\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e24.28\u0026thinsp;\u0026plusmn;\u0026thinsp;8.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e8.58\u0026thinsp;\u0026plusmn;\u0026thinsp;4.89\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e9.162\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNon-PCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e22.41\u0026thinsp;\u0026plusmn;\u0026thinsp;6.43\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6.16\u0026thinsp;\u0026plusmn;\u0026thinsp;4.81\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e11.073\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003et\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.995\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.927\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.324\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.059\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab11\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e11 Comparison of clinical remission rates between PCD group and Non-PCD group\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGroup\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003erelieve[n(%)]\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eunrelieved[n(%)]\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e22(73.33%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e8(26.67%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNon-PCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e29(96.67%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1(3.33%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eχ\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c3\" namest=\"c2\"\u003e \u003cp\u003e____\u003csup\u003e▼\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c3\" namest=\"c2\"\u003e \u003cp\u003e0.026\u003csup\u003e*\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section3\"\u003e \u003ch2\u003e3.10.2Efficacy evaluation of IFX at week 14 in patients with or without perianal lesions\u003c/h2\u003e \u003cp\u003eIn the group with perianal lesions, a total of 23 patients received IFX treatment, while in the group without perianal lesions, 18 patients received IFX treatment. Prior to IFX treatment, the PCDAI scores for the groups with and without perianal lesions were (22.04\u0026thinsp;\u0026plusmn;\u0026thinsp;6.66) points and (21.22\u0026thinsp;\u0026plusmn;\u0026thinsp;6.16) points, respectively. There was no statistically significant difference between the groups (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05). After 14 weeks of IFX treatment, the PCDAI scores in both groups decreased compared to pre-treatment levels. There was no statistically significant difference in PCDAI scores between the two groups post-treatment (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05). Among patients treated with IFX, there was no difference in clinical remission rates between the CD group with perianal lesions and the CD group without perianal lesions (82.60% vs 94.44%, P\u0026thinsp;=\u0026thinsp;0.363). see Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;12 and Table\u0026nbsp;\u003cspan refid=\"Tab13\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u0026ndash;13.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab12\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e12 Comparison of PCDAI scores before and after IFX treatment PCD group and Non-PCD group\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGroup\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ebefore treatment\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePost treatment\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003et\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e22.04\u0026thinsp;\u0026plusmn;\u0026thinsp;6.66\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6.19\u0026thinsp;\u0026plusmn;\u0026thinsp;5.10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e9.057\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNon-PCD(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e21.22\u0026thinsp;\u0026plusmn;\u0026thinsp;6.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5.13\u0026thinsp;\u0026plusmn;\u0026thinsp;3.97\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e9.308\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;0.001\u003csup\u003e**\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003et\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.405\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.723\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.688\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.474\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab13\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e13 Comparison of clinical remission rates between groups with and without perianal lesions treated with IFX\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGroup\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003erelieve[n(%)]\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eunrelieved[n(%)]\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePCD(n\u0026thinsp;=\u0026thinsp;23)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e19(82.60%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4(17.40%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNon-PCD (n\u0026thinsp;=\u0026thinsp;18)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e17(94.44%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1(5.56%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eχ\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c3\" namest=\"c2\"\u003e \u003cp\u003e____\u003csup\u003e▼\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c3\" namest=\"c2\"\u003e \u003cp\u003e0.363\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003c/div\u003e"},{"header":"4. DISCUSSION","content":"\u003cp\u003eCrohn's disease is a chronic transmural inflammatory bowel disease(Feroz et al.; Wasmann et al.; de Groof et al.). The incidence of Crohn's disease in children has risen rapidly over the past few decades. Compared to adults, the clinical course of Crohn's disease in childhood is particularly aggressive, with approximately 20\u0026ndash;25% of patients developing the disease during childhood or adolescence (Wewer et al.)Perianal lesions are common in Crohn's disease. According to reports, the incidence of perianal lesions in Crohn's disease patients ranges from 20\u0026ndash;40% globally (Tsai et al.; Williams and Shaffer), with higher rates in Asian countries, ranging from 30.3\u0026ndash;58.8% (Agrawal and Jess). Anal lesions may manifest before or concurrently with intestinal symptoms. Some patients may not exhibit symptoms of anal lesions until several years after being diagnosed with Crohn's disease. A study found that approximately 26% of CD patients developed anal fistulas within 20 years of diagnosis(Tsai et al.), which is associated with higher rates of hospitalization, surgical intervention, immunosuppressive therapy, and reduced quality of life(Adegbola et al.). The presence of perianal lesions, particularly fistulas, in CD patients is a hallmark of the chronic and invasive nature of the disease (Lukin). The etiology of perianal lesions in Crohn's disease is diverse and complex, with its pathogenesis involving genetic polymorphisms, dysregulated immune responses, and epithelial-mesenchymal transition.\u003c/p\u003e \u003cdiv id=\"Sec21\" class=\"Section2\"\u003e \u003ch2\u003e4.1 Clinical characteristics and influencing factors of PCD\u003c/h2\u003e \u003cp\u003eRegarding risk factors associated with PCD, current research findings have not yet reached a consensus. Previous literature has suggested that factors such as gender, age, disease duration, initial location of lesions, lesion location, and smoking history may be associated with the occurrence and development of PCD(Lukin). In our study, the PCD group had a higher proportion of males, with a male-to-female ratio of approximately 4:1. This ratio differed significantly from that of the Crohn's disease group without perianal lesions (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05). Our findings align with those of Mutanen A (Mutanen and Pakarinen), who also noted that gender is a risk factor for PCD, with male patients being more prone to the condition. This may be related to racial influences on the phenotype of inflammatory bowel disease. Western reports indicate a higher proportion of female PCD patients compared to males, while Asian countries show a higher prevalence of male PCD patients (Meima-van Praag, Buskens, et al.).\u003c/p\u003e \u003cp\u003eIn terms of the distribution characteristics of intestinal lesions, adult Crohn's disease patients in Western countries exhibit a relatively balanced trend in the proportions of L1, L2, and L3 lesions. In contrast, Chinese CD patients more commonly present with L3 lesions, while L4 lesions are relatively rare (Zeng et al.). Additionally, pediatric Crohn's disease differs significantly from adult cases in terms of disease phenotype and behavior (de Bie et al.). Currently, the Montreal classification system is widely used for categorizing anal lesions in pediatric Crohn's disease. In pediatric patients, the disease progression is characterized by greater invasiveness and diffuseness, resulting in a higher probability of lesions in the colon (L2) and upper gastrointestinal tract (L4) compared to adults. Statistically, approximately half of children and adolescents with Crohn's disease have upper gastrointestinal tract damage at the time of diagnosis(Assa, Rinawi and Shamir), and the data from this study is consistent with this. The proportion of L4 lesions was higher in both the perianal lesion group (53.33%) and the non-perianal lesion group (46.67%). SB Ingle's study noted that colonic involvement is an important predictive factor for Crohn's disease with perianal lesions, with a significantly increased risk of perianal lesions in patients with colonic Crohn's disease(Ingle and Loftus). The data from this study aligns with this finding, with a higher proportion of L3 cases in the perianal lesion group (36.67%) compared to the non-perianal lesion group (26.66%). It is generally believed that the B1 phenotype (non-penetrating/non-stricturing) at diagnosis is the most common presentation of Crohn's disease in both Eastern and Western patients(Dolinger, Torres and Vermeire). However, in our study, only 33.33% of children in the perianal lesion group had the B1 phenotype, while the B1 phenotype was predominant in the group without perianal lesions, accounting for 73.33%. A study by Braithwaite GC(Braithwaite et al.)found that the primary characteristic of the B3 type (penetrating) is a high incidence of fistulas. Compared with other types, patients with the B3 type have a significantly higher incidence of anal fistulas, and the incidence of anal fistula lesions also increases accordingly. In this study, Type B3 was more common in the anal perianal lesion group than in the group without anal perianal lesions, with proportions of 40.00% and 6.67%, respectively. The difference between the two groups was statistically significant (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05). This may be related to the classification of perianal fistulas as B3 type in this study. The specific impact of disease behavior on perianal lesions in CD requires further validation through large-scale, ongoing, and extensive research using a broader sample size.\u003c/p\u003e \u003cp\u003eThe typical initial presentation of Crohn's disease is often nonspecific, with common clinical manifestations including chronic diarrhea, abdominal pain, and weight loss. In children and adolescents, abdominal pain is typically the primary complaint (Moon). In this study, the Crohn's disease group without perianal lesions primarily presented with abdominal pain, while the PCD group exhibited a significant increase in diarrhea frequency, which may be related to the frequent stimulation and pressure on the perianal area caused by diarrhea(Bolshinsky and Church). Children in the PCD group exhibited relatively more extraintestinal manifestations, a finding consistent with previous relevant studies(Seyfried and Herold). However, there was no statistically significant difference in the initial clinical manifestations between the two groups (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05), which may be attributed to the diverse and nonspecific nature of Crohn's disease symptoms.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec22\" class=\"Section2\"\u003e \u003ch2\u003e4.2Pyroptosis-related protein NLRP3 and its association with PCD\u003c/h2\u003e \u003cp\u003eMutations in the NLRP3 gene are considered an important risk factor for the onset of IBD. Abnormally activated NLRP3 inflammasomes play a key role in the pathogenesis of UC (Klughammer et al.) and are also involved in the onset of CD (Sun et al.). In recent years, an increasing number of studies have suggested that uncontrolled activation of the NLRP3 inflammasome plays a key role in chronic intestinal inflammation, and the pathogenesis of colitis is associated with elevated levels of IL-1β and IL-18 (Chen et al.). Reduced NLRP3 production is associated with a lower risk of colitis [63, 64]. In animal studies, mice with elevated NLRP3 levels were more susceptible to dextran sulfate sodium salt (DSS) and 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis, but other experiments found that NLRP3-deficient mice exhibited more severe colitis after oral administration of DSS (Bauer et al.). In our study, to investigate the expression levels of NLRP3 and its mRNA in the intestinal mucosa of CD patients, we used a combined analysis of immunohistochemistry (IHC) and qRT-PCR. The results showed that the IHC staining scores and mRNA expression levels in the case group were higher than those in the control group (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05); Further intergroup analysis revealed that IHC staining scores and mRNA expression levels were higher in the CD group without perianal lesions and the PCD group compared to the control group, and also higher in the PCD group compared to the CD group without perianal lesions (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 for all). Previous studies have shown that NLRP3 mRNA and protein expression is elevated in the intestinal mucosa of adult CD patients and positively correlated with disease severity(Ranson et al.) In rectal mucosa, IL-1β and IL-6 expression was higher in Crohn's disease patients with anal fistulas than in patients with luminal CD or healthy controls (Ruffolo et al.). Meanwhile, in perianal fistula tissue from Crohn's disease patients with anal fistulas, IL-12p40, TNF-α, IL-1β, and IL-8 expression was up-regulated (van Onkelen et al.). The expression of NLRP3 in the intestinal mucosa of the PCD group was higher than that in the CD group without perianal lesions, which may be related to the fact that NLRP3 can indirectly regulate the secretion of IL-1β and IL-18, promoting their synthesis and release (Fu and Wu). The results of this experiment are consistent with those of basic animal experiments and adult studies. Additionally, immunohistochemical staining of intestinal mucosa in this study showed that NLRP3 was sporadically expressed in the cytoplasm of intestinal epithelial cells, macrophages, and lymphocytes in the control group, with yellow or brown granules also visible in the cytoplasm. Compared with the control group, the staining range and intensity were increased in the case group, suggesting elevated NLRP3 levels in intestinal tissues of patients in the case group and indicating that NLRP3 is expressed in various cells of the immune system. Further comparison showed that NLRP3 staining range and intensity were enhanced in the CD group with perianal lesions compared with the CD group without perianal lesions (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec23\" class=\"Section2\"\u003e \u003ch2\u003e4.2Pyroptosis-related protein GSDMD and its association with PCD\u003c/h2\u003e \u003cp\u003eGSDMD is a key executor of pyroptosis and plays a crucial regulatory role in host immune defense, inflammatory responses, autoimmune diseases, and various other systemic disorders (Li et al.). The role of GSDMD in colitis remains controversial. On one hand, elevated GSDMD levels in mucosal biopsies from IBD patients suggest a pathogenic role for this protein in the disease (Bulek et al.). In TNBS-induced mouse colitis, high expression of GSDMD and GSDME in colonic epithelial cells supports this observation(Tan et al.). However, this is inconsistent with the severe colitis phenotype observed in another mouse strain lacking GSDMD (Ma et al.). This study aims to investigate the expression levels of GSDMD in intestinal mucosa using immunohistochemistry to address these knowledge gaps. Our findings revealed statistically significant differences in GSDMD intestinal mucosa IHC staining scores among the PCD group, Non-PCD group, and control group (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 for all); Further intergroup analysis revealed that the GSDMD intestinal mucosa IHC staining scores in the Non-PCD and PCD groups were higher than those in the control group (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05), and the GSDMD intestinal mucosa IHC staining scores in the PCD group were higher than those in the Non-PCD group (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05). The expression of GSDMD in the intestinal mucosa of the PCD group was higher than that in the Non-PCD group. Considering that perianal lesions are predictive factors for the progression of Crohn's disease, this suggests a possible rapid progression from inflammation to stenosis or perforation phenotypes(Tarrant et al.). Additionally, immunohistochemical localization experiments showed that GSDMD was expressed in intestinal epithelial cells and immune cells (such as macrophages, neutrophils, monocytes, T cells, and B cells) in the intestinal mucosa of patients in the case group, while the control group showed minimal expression in intestinal epithelial cells and immune cells under the microscope. Further comparison revealed that GSDMD staining intensity and extent were enhanced in the CD group with perianal lesions compared to the CD group without perianal lesions, consistent with previous studies reporting GSDMD expression in immune cells and intestinal epithelial cells (Wang, Ding and Shao).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec24\" class=\"Section2\"\u003e \u003ch2\u003e4.3Pyroptosis-related protein IL-1β and IL-18 and its association with PCD\u003c/h2\u003e \u003cp\u003eInterleukin-1β (IL-1β) is a key mediator of the inflammatory response, playing a role in both innate and adaptive immunity. It is crucial for the host's response to and resistance against pathogens and may exacerbate tissue damage in acute and chronic diseases(Cao et al.). IL-18 is a ubiquitous pro-inflammatory cytokine within the IL-1 family, structurally similar to the inflammatory factor IL-1β. IL-18 activates T cells, enhances immune responses, and plays a significant role as an inflammatory marker in inflammatory diseases (Detry et al.). Studies have shown that both UC and CD exhibit elevated levels of innate cytokines, including IL-8, IL-6, IL-1β, and IL-18. The production of IL-1β and IL-18 in the intestinal mucosa of patients with active IBD is increased (Thinwa et al.). The results of this study showed that the serum concentrations of IL-1β and IL-18, intestinal mucosa IHC staining scores, and mRNA expression levels were higher in the case group than in the control group (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 for all), Further intergroup analysis showed that the IHC staining scores for IL-1β and IL-18 were higher in both the Non-PCD group and the PCD group compared to the control group, and also higher in the PCD group compared to the Non-PCD group (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05). This is consistent with clinical study results, which indicate that IL-1 secretion is increased in the colon tissue and macrophages of IBD patients and is positively correlated with disease severity (Perera et al.). Regarding the higher expression of IL-1β in the intestinal mucosa of the PCD group compared to the CD group without perianal lesions, we analyzed that this may be related to the following aspects: (1) Studies have shown that IL-1β and IL-6 expression levels in rectal mucosal tissue of Crohn's disease fistula patients are significantly higher than in patients with luminal-type Crohn's disease and healthy individuals (Ruffolo et al.). This study also found the same results, with IL-1β intestinal mucosa IHC staining scores in the PCD group being higher than in the Non-PCD group (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05). (2) Through an in-depth analysis of perianal fistula tissue in Crohn's disease patients with anal fistulas, researchers observed that inflammatory factors such as IL-12p40, TNF-α, IL-1β, and IL-8 showed a significant upward trend (van Onkelen et al.). IL-18 expression was higher in the PCD group than in the Non-PCD group, which may be associated with perianal lesions being an important predictive factor for disease progression in Crohn's disease, suggesting a rapid progression from inflammation to stricture or perforation phenotypes. IL-18 was highly expressed in intestinal epithelial cells of Crohn's disease patients in remission, indicating its primary protective function in the early stages of colitis. The absence of IL-18 may increase host susceptibility to chemically induced colitis. As disease severity increases, the number of IL-18-associated macrophages in active CD patients increases, and IL-18 expression in intestinal epithelial cells correspondingly rises (Lei-Leston, Murphy and Maloy).\u003c/p\u003e \u003cp\u003eImmunohistochemical staining localization showed that the staining range and intensity of IL-1β and IL-18 were enhanced in the PCD group compared to the Non-PCD group. Previous studies have suggested that IL-1β is primarily produced by activated mononuclear phagocytes and can also be produced by macrophages, endothelial cells, and vascular smooth muscle cells(Wang et al.). The precursor of IL-18 is primarily present in the cytoplasm of monocytes, macrophages, and dendritic cells, as well as in endothelial cells, keratinocytes, and intestinal epithelial cells of the gastrointestinal tract (Ihim et al.). Our findings are consistent with previous results.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec25\" class=\"Section2\"\u003e \u003ch2\u003e4.4 NLRP3, GSDMD, IL-1β, and IL-18 individually and in combination predict PCD efficacy\u003c/h2\u003e \u003cp\u003eIn terms of intestinal mucosal immunohistochemical staining scores, NLRP3 had the largest area under the curve (AUC) of 0.878. When the NLRP3 staining score was 49.33, the diagnostic sensitivity was 83.30% and the specificity was 79.30%. The diagnostic performance of NLRP3 was superior to that of GSDMD, IL-1β, and IL-18. Mona(Watany, Elmazny and Nasif) et al. found in their study on the diagnosis of septic shock using NLRP3, IL-1β, and IL-18 that IL-1β had higher diagnostic efficacy than NLRP3; this aspect has not been explored in the present study. We conducted further combined diagnostic analysis of these indicators. The results showed that the four factors in the immunohistochemical staining score demonstrated higher efficacy in predicting the occurrence of PCD, with an AUC value of 0.966, a sensitivity of 96.60%, and a specificity of 86.70%. This suggests that the above indicators have a certain predictive efficacy for the occurrence of PCD and may be used as screening indicators for reference.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec26\" class=\"Section2\"\u003e \u003ch2\u003e4.5NLRP3, GSDMD, IL-1β, IL-18 correlation\u003c/h2\u003e \u003cp\u003eNLRP3, GSDMD, IL-1β, and IL-18 are four factors located on the NLRP3/GSDMD pathway. A study found (Garnacho-Montero et al.)that the expression of NLRP3 on this pathway is positively correlated with IL-1β levels. Therefore, this study analyzed their correlation. The results showed that in our experiment, the key proteins NLRP3, GSDMD, IL-1β, and IL-18, the key proteins in the pyroptosis pathway, are pairwise correlated. In terms of immunohistochemical staining scores: the immunohistochemical staining scores of GSDMD, IL-1β, and IL-18 were positively correlated with the NLRP3 immunohistochemical staining score (P\u0026thinsp;\u0026lt;\u0026thinsp;0.01), with Spearman correlation coefficients of 0.798, 0.695, and 0.847, respectively; The immunohistochemical staining scores for IL-1β and IL-18 were positively correlated with the immunohistochemical staining score for GSDMD (P\u0026thinsp;\u0026lt;\u0026thinsp;0.01), with Spearman's correlation coefficients of 0.709 and 0.801, respectively; The immunohistochemical staining scores for IL-1β and IL-18 were positively correlated (P\u0026thinsp;\u0026lt;\u0026thinsp;0.01), with a correlation coefficient of 0.654. By analyzing the associations among the four factors, this study not only validated that the pathogenesis of PCD indeed involves complex cascading signal transduction processes but also proposed new directions for exploration: the specific mechanisms underlying PCD have not yet been fully elucidated. Whether compensatory regulatory mechanisms exist between upstream and downstream factors within the same signal transduction pathway, and how these mechanisms regulate inflammatory responses, remain unclear and warrant further investigation in the future.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec27\" class=\"Section2\"\u003e \u003ch2\u003e4.6 Efficacy evaluation\u003c/h2\u003e \u003cp\u003eThe treatment goal for CD patients is to achieve and maintain disease remission. Short-term goals include alleviating clinical symptoms and restoring serum and fecal inflammatory markers to normal levels; long-term goals aim for complete resolution of clinical symptoms, sustained normalization of inflammatory markers, and endoscopic mucosal healing(Turner et al.). During the induction phase of remission, a combination of treatment modalities is typically employed, with corticosteroids, biologics, and enteral nutrition being the most commonly used approaches (Ong et al.). For children with fistulizing lesions, considering that fistula formation may exacerbate the condition and impair the child's quality of life, clinicians often prefer to initiate IFX therapy early. In contrast, the use of corticosteroids is more common in children without fistulizing lesions (Gecse et al.). In this study, 23 cases (76.67%) in the PCD group received IFX, which had a higher biologic agent usage rate compared to the 18 cases (60.00%) in the Non-PCD group, consistent with previous studies. This study used a standardized PCDAI scoring method before and after treatment to comprehensively assess the severity of anal lesions and the efficacy of drug therapy in pediatric patients. There was no statistically significant difference in PCDAI scores between the PCD group and the non-PCD group before treatment. After 3\u0026ndash;6 months of treatment, PCDAI scores decreased significantly in both the PCD and non-PCD groups, but there was no statistically significant difference in PCDAI scores between the PCD and non-PCD groups after treatment (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05). Meanwhile, in this study, after 3\u0026ndash;6 months of treatment, there was a difference in clinical remission rates between the PCD group and the Non-PCD group (73.33% vs. 96.67%, P\u0026thinsp;=\u0026thinsp;0.026), which may be related to the more aggressive nature of PCD. We further analyzed that in children who achieved remission with IFX induction, the clinical remission rate was higher in the PCD group, and there was no significant difference in clinical remission rates between the two groups with or without perianal lesions (82.60% vs. 94.44%, P\u0026thinsp;=\u0026thinsp;0.363). This is consistent with previous studies\u0026mdash;a meta-analysis investigated the efficacy differences between the \u0026ldquo;early step-down\u0026rdquo; treatment strategy, which involves initiating biologic agents within two years of Crohn's disease diagnosis, and the traditional \u0026ldquo;step-up\u0026rdquo; therapy (i.e., considering biologic agent use more than two years after diagnosis). The results showed that compared with the traditional \u0026ldquo;step-up\u0026rdquo; therapy, the early introduction of biologics significantly improved patient outcomes, specifically manifested as higher clinical remission rates, significantly reduced disease recurrence rates, and a substantial increase in mucosal healing rates (Wasmann et al.). Additional data support that early initiation of IFX therapy not only leads to more patients achieving endoscopic disease remission but also effectively reduces the incidence of stricturing lesions(Schnitzler et al.). Therefore, for Crohn's disease patients, regardless of the presence of perianal lesions, infliximab therapy is recommended at the time of diagnosis.\u003c/p\u003e \u003c/div\u003e"},{"header":"Conclusions","content":"\u003cp\u003e \u003col\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eIn the PCD group, the expression levels of NLRP3, GSDMD, IL-1β, and IL-18 were all increased in intestinal mucosal tissue, and there was a positive correlation among the four factors, suggesting that the NLRP3/GSDMD signaling pathway may be involved in the pathogenesis of Crohn's disease with perianal lesions.\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eThe expression levels of NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue have certain predictive value for the diagnosis of Crohn's disease with perianal lesions in children, and the combination of these four factors can improve the predictive value of diagnosis.\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003c/ol\u003e \u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e \u003ch2\u003eCompeting interests\u003c/h2\u003e \u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e \u003c/p\u003e\u003cp\u003eDepartment of Gastroenterology at the Children\u0026apos;s Hospital Affiliated to Soochow University, Suzhou, China\u003c/p\u003e\u003ch2\u003eFunding sources\u003c/h2\u003e \u003cp\u003ehe Jiangsu Provincial Key Medical Discipline (No. ZDXKA2016013)\u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eWang Hongding writes the revised manuscript for Jin Zhongqin\u003c/p\u003e\u003ch2\u003eData Availability\u003c/h2\u003e\u003cp\u003eData used to support findings reported herein are available from the corresponding author upon request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAdegbola SO et al (2020) Burden of Disease and Adaptation to Life in Patients with Crohn's Perianal Fistula: A Qualitative Exploration. 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J Gastrointest Surg 11(1):16\u0026ndash;21 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSandborn WJ et al (2003) Aga Technical Review on Perianal Crohn's Disease. Gastroenterology 125(5):1508\u0026ndash;1530 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSchnitzler F et al (2021) Early Start of Infliximab in Crohn's Disease Increases Rates of Endoscopic Remission and Decreases Stenosis Formation: Experiences from a Single Center Cohort. Crohns Colitis 360 3(3):otab060 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSeyfried S, Herold A (2019) Management of Perianal Fistulas in Crohn's Disease. Visc Med 35(6):338\u0026ndash;343 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSun Y et al (2015) Wogonoside Protects against Dextran Sulfate Sodium-Induced Experimental Colitis in Mice by Inhibiting Nf-Κb and Nlrp3 Inflammasome Activation. 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Inflamm Bowel Dis 28:1477\u0026ndash;1484 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTurner D et al (2021) Stride-Ii: An Update on the Selecting Therapeutic Targets in Inflammatory Bowel Disease (Stride) Initiative of the International Organization for the Study of Ibd (Ioibd): Determining Therapeutic Goals for Treat-to-Target Strategies in Ibd. Gastroenterology 160(5):1570\u0026ndash;1583 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003evan Onkelen RS et al (2016) Pro-Inflammatory Cytokines in Cryptoglandular Anal Fistulas. Tech Coloproctol 20(9):619\u0026ndash;625 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang K, Ding J, Shao F (2023) Determination of Gasdermin Pores. Methods Mol Biol 2696:149\u0026ndash;167 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang P et al (2023) Role of Signal Transduction Pathways in Il-1β-Induced Apoptosis: Pathological and Therapeutic Aspects. 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Am Surg 87:1361\u0026ndash;1367 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYamamoto T et al (2023) Diagnosis and Clinical Features of Perianal Lesions in Newly Diagnosed Crohn's Disease: Subgroup Analysis from Inception Cohort Registry Study of Patients with Crohn's Disease (Icrest-Cd). J Crohns Colitis 17:1193\u0026ndash;1206 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZeng Z et al (2013) Incidence and Clinical Characteristics of Inflammatory Bowel Disease in a Developed Region of Guangdong Province, China: A Prospective Population-Based Study. J Gastroenterol Hepatol 28:1148\u0026ndash;1153 Print\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZheng Z, Li G (2020) Mechanisms and Therapeutic Regulation of Pyroptosis in Inflammatory Diseases and Cancer. Int J Mol Sci 21.4 Print.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"international-journal-of-colorectal-disease","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"ijcd","sideBox":"Learn more about [International Journal of Colorectal Disease](http://link.springer.com/journal/384)","snPcode":"384","submissionUrl":"https://submission.nature.com/new-submission/384/3","title":"International Journal of Colorectal Disease","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Crohn’s disease, Perianal lesions, Pyroptosis, Immunohistochemistry","lastPublishedDoi":"10.21203/rs.3.rs-7536822/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7536822/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e \u003cb\u003eIntroduction\u003c/b\u003e: Crohn's disease(CD) is a complex inflammatory disease with perianal lesions as one of its serious complications. Pyroptosis is an inflammatory form of programmed cell death, and the role of pyroptosis in CD intestinal epithelial cells is unknown.\u003c/p\u003e \u003cp\u003e \u003cb\u003eMethods\u003c/b\u003e: Between September 2022 and August 2024, we selected 60 children with a primary diagnosis of Crohn's disease from the Department of Gastroenterology at the Children's Hospital Affiliated to Soochow University as the case group, and divided them into a group with perianal lesions (PCD group) and a group without perianal lesions (Non-PCD group). Additionally, 40 children diagnosed with simple juvenile intestinal polyps during the same period were selected as the control group. All subjects underwent immunohistochemistry (IHC) to quantitatively and locally detect the expression of NLRP3, GSDMD, IL-1β, and IL-18 in intestinal mucosal tissue.\u003c/p\u003e \u003cp\u003e \u003cb\u003eResult\u003c/b\u003e:Clinical samples confirmed that the staining scores for NLRP3, GSDMD, IL-1β, and IL-18 were significantly higher in the PCD group than in the Non-PCD group and the control group. Immunohistochemical localization revealed that NLRP3, GSDMD, IL-1β, and IL-18 were primarily expressed in epithelial cells and immune cells of the intestinal mucosal tissue. Compared with the control group and the non-PCD group, the staining extent and intensity of these markers were significantly increased in the PCD group.\u003c/p\u003e \u003cp\u003e \u003cb\u003eConclusion\u003c/b\u003e: Our findings identify NLRP3, GSDMD, IL-1β, and IL-18 as key pyroptosis-associated biomarkers in PCD, contributing to the understanding of pyroptosis in the pathogenesis of PCD.\u003c/p\u003e","manuscriptTitle":"Investigation of NLRP3, GSDMD, IL-1β and IL-18 expression in pediatric Crohn's disease with perianal lesions","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-01-06 10:52:28","doi":"10.21203/rs.3.rs-7536822/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewersInvited","content":"","date":"2026-01-02T09:14:13+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-09-10T09:23:43+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-09-10T00:14:17+00:00","index":"","fulltext":""},{"type":"submitted","content":"International Journal of Colorectal Disease","date":"2025-09-04T13:48:11+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"international-journal-of-colorectal-disease","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"ijcd","sideBox":"Learn more about [International Journal of Colorectal Disease](http://link.springer.com/journal/384)","snPcode":"384","submissionUrl":"https://submission.nature.com/new-submission/384/3","title":"International Journal of Colorectal Disease","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"1d66fcaa-74f0-46bc-be27-f120f4297caf","owner":[],"postedDate":"January 6th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-01-06T10:52:29+00:00","versionOfRecord":[],"versionCreatedAt":"2026-01-06 10:52:28","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7536822","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7536822","identity":"rs-7536822","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}