Association of Podoplanin Expression with Histological Grade in Oral Squamous Cell Carcinoma in a Tertiary Care Hospital in Bangladesh

Association of Podoplanin Expression with Histological Grade in Oral Squamous Cell Carcinoma in a Tertiary Care Hospital in Bangladesh | 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 Association of Podoplanin Expression with Histological Grade in Oral Squamous Cell Carcinoma in a Tertiary Care Hospital in Bangladesh Mst.Rubeyatul Jannat, Nafisa Abedin, Sayeed Kishwara kashfi, Sadia Shirin, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5815305/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background Oral carcinogenesis is a complex process driven by genetic alterations affecting proto-oncogenes and tumor suppressor genes, with key players including P16, P53, H-ras, cyclinD1, and EGFR. Podoplanin, a transmembrane glycoprotein, has emerged as a biomarker implicated in tumor progression and prognosis in oral squamous cell carcinoma (OSCC). This study aimed to evaluate the immunohistochemical expression of podoplanin across different histological grades of OSCC and analyze its association with clinicopathological parameters such as age, gender, anatomical site, personal habits, lymphovascular and perineural invasion, and tumor-infiltrating lymphocytes. Methods A cross-sectional observational study was conducted at the Department of Pathology, Bangladesh Institute of Research and Rehabilitation in Diabetes, Endocrine and Metabolic Disorders (BIRDEM) Hospital, Dhaka, Bangladesh, over two years. Fifty-six cases of OSCC were analyzed using hematoxylin and eosin-stained slides and paraffin-embedded tissue blocks. Podoplanin expression was assessed immunohistochemically, and staining intensity and extent were categorized as high or low. Results The results revealed high podoplanin expression in 33.90% of cases and low expression in 66.10%. High podoplanin expression was more frequently observed in moderately and poorly differentiated OSCC compared to well-differentiated tumors, with a statistically significant correlation between podoplanin expression and histological grade. However, no significant associations were identified between podoplanin expression and other factors, including age, gender, personal habits, anatomical site, lymphovascular invasion, perineural invasion, or tumor-infiltrating lymphocytes. Conclusion Podoplanin expression was significantly associated with the histological grade of OSCC, suggesting its potential role as a prognostic marker. High podoplanin expression may indicate aggressive tumor behavior and poor prognosis, underscoring its relevance in OSCC evaluation and management. Podoplanin Oral Squamous Cell Carcinoma Immunohistochemistry Histological Grade Prognosis Biomarker Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Introduction Oral cancer is the sixth most common malignancy worldwide and is one of the major public health problems¹. Oral squamous cell carcinoma (OSCC) is the most common type of malignancy, representing up to 90 percent of all oral neoplasms². Oral cancer ranks 3rd, 2nd, and 3rd in five-year prevalence, incidence, and mortality rate respectively in Bangladesh³. Common risk factors for developing OSCC are tobacco smoking, betel nut chewing, and alcohol consumption. Other predisposing factors are viral infection, chronic irritation, oral candidiasis, poor oral hygiene, ionizing radiation, etc⁴. Podoplanin (PDPN) is known to be expressed in numerous human tumors, including squamous cell carcinoma of the oral cavity, mesothelioma, testicular seminoma, soft tissue tumors, thymoma, and brain tumors⁵. PDPN seems to be expressed in aggressive tumors, with higher invasive and metastatic potential⁶. PDPN consists of an extracellular or transmembrane domain and a cytoplasmic domain. The extracellular domain is longer and carries four platelet aggregation-stimulating (PLAG) domains with plenty of potential O-glycosylation sites, crucial for interaction with platelets. The interaction between PDPN and c-type lectin (CLEC2) may regulate tumor invasion and metastasis⁷. The shorter cytoplasmic domain is associated with ezrin/radixin/moesin (ERM) protein that bridges plasma membrane proteins and the actin cytoskeleton⁸. PDPN overexpression causes marked phosphorylation of ERM proteins. It causes adhesion and cancer cell migration through modulating the actin cytoskeleton, Rho A, and epithelial-mesenchymal transition. PDPN causes down-regulation of E-cadherin and overexpression of matrix metalloproteinase⁶. It is upregulated in the invasive front of OSCC. It causes upregulation of TGF-β1 secretion and increases Epidermal Growth Factor Receptor phosphorylation as well as downstream effectors of AKT and ERK. PDPN also causes OSCC progression by interacting with matrix metalloproteinase and induces cytoskeletal remodelling, extracellular matrix degradation, and invasion⁹. PDPN in oral SCC causes tumor cell motility and metastasis by activation of endogenous lectin that binds to extracellular carbohydrate moieties. These cancer cells are remarkably resistant to currently available chemotherapy. Thus, PDPN becomes a chemotherapeutic target for oral SCC¹⁰. Anti-human Podoplanin antibody (NZ-1) would be an important target for the prevention of metastasis in the future for OSCC⁶. Materials and Methods 1. Study Design and Subjects This cross-sectional observational study was conducted at the Department of Pathology, Bangladesh Institute of Research and Rehabilitation in Diabetes, Endocrine and Metabolic Disorder (BIRDEM), during the period of March 2022 to February 2024. A total of 56 histologically diagnosed cases of oral squamous cell carcinoma (OSCC) were included in this study, meeting the following inclusion criteria: small and resected biopsy cases of OSCC. Exclusion criteria included carcinoma in situ, dysplastic lesions of the oral cavity, and patients who received neoadjuvant therapy or radiotherapy before surgery. Demographic and clinical information, such as age, sex, clinical presentation, and histopathological diagnosis (including grade), were collected from the departmental records of the pathology department. 2. Sample Collection The 56 cases of oral SCC were retrieved from the Department of Pathology, BIRDEM, including H&E stained slides and paraffin-embedded blocks. All slides were reviewed to assess tumor grade and other prognostic factors, including tumor-infiltrating lymphocytes (TILs), and the presence of lymphovascular and perineural invasion. 3. Immunohistochemical Staining for Podoplanin After confirming the diagnosis of OSCC, immunohistochemical (IHC) staining for podoplanin was performed at Bangabandhu Sheikh Mujib Medical University (BSMMU), using an appropriate positive control. Paraffin-embedded tissue blocks were sectioned at 3 µm thickness. The slides were gently lowered onto a water bath set to 45°C, spread without wrinkles onto slides coated with 0.1% poly-L-lysine, and air-dried. Slides were then baked at 60°C for 30 minutes on a hot plate. Dewaxing was performed by treating the slides in xylene, followed by rehydration through a graded series of alcohols. For antigen retrieval, the slides were immersed in preheated citrate buffer in a pressure cooker, boiled, and allowed to cool naturally. To block endogenous enzyme activity, hydrogen peroxide was added in a moist chamber at room temperature. 4. Primary and Secondary Antibodies Primary antibody : Monoclonal Rabbit d2-40 (pre-diluted, ready-to-use). Secondary antibody : DAKO REAL™ Envision™ (HRP Rabbit/Mouse) (ENV). Positive control : Appedicular tissue was used as a positive control. Chromogen : 3,3’-diaminobenzidine (DAB) was used for visualizing the antigen-antibody complex. Finally, slides were counterstained with hematoxylin. 5. Scoring of Podoplanin Expression Podoplanin expression was assessed by reviewing 10 representative areas from each slide. For each area, 100 tumor cells were counted, and both the percentage of positive cells and the intensity of podoplanin expression were evaluated. The following scoring system, as introduced by Yuan et al. (2006), was applied: Quantitative Score: 0 : 0% positive cells 1 : 1–10% positive cells 2 : 11–30% positive cells 3 : 31–50% positive cells 4 : 51–80% positive cells 5 : 81–100% positive cells Intensity Score: 0 : Negative (no staining) 1 : Weak (faint staining) 2 : Moderate (staining between weak and dark brown) 3 : Strong (dark brown staining) The immunoreactive score (IRS) for podoplanin expression was calculated by multiplying the quantitative score by the intensity score, yielding a final score ranging from 0 to 15 . According to this score, the expression was classified as follows: 0–7 : Low expression ≥ 8 : High expression 6. Statistical Analysis All collected data were compiled and analyzed using SPSS (Statistical Package for Social Sciences), version 25 . The Chi-square test was used to assess statistical significance between different clinicopathologic variables. A p-value of < 0.05 was considered statistically significant. Results A total of 56 oral squamous cell carcinoma (OSCC) patients were included in the study, and their clinicopathologic characteristics along with Podoplanin expression were analyzed. The association between Podoplanin expression and various clinicopathologic parameters, as well as histological grading, was evaluated (Table 1 ). Podoplanin Expression and Clinicopathologic Parameters Podoplanin expression was classified as high or low based on immunohistochemical staining. High Podoplanin expression was observed in 19 cases (33.90%) of OSCC. The distribution of high and low Podoplanin expression did not show any statistically significant association with age, gender, clinical presentation, or personal habits (tobacco use, betel quid use). Specifically, no significant differences in Podoplanin expression were noted across different age groups (P = 0.58), between genders (P = 0.58), or based on the clinical presentation (P = 0.58). Furthermore, Podoplanin expression did not correlate with tobacco use (P = 0.57), betel quid use (P = 1.0), or both tobacco and betel quid use (P = 1.0). Similarly, no significant association was found between Podoplanin expression and the anatomic location of the tumors (P = 0.24) (Table 1 ). Podoplanin Expression and Histological Grading The association between Podoplanin expression and the histological grading of OSCC was found to be statistically significant (P = 0.00) (Table 2 ). Podoplanin expression varied considerably across different grades of differentiation. In well-differentiated OSCC (n = 25), no cases exhibited high Podoplanin expression, with all tumors showing low expression. In moderately differentiated OSCC (n = 27), 17 cases (63.0%) exhibited high Podoplanin expression, while 10 cases (37.0%) showed low expression. In poorly differentiated OSCC (n = 4), high Podoplanin expression was observed in 2 cases (50.0%) and low expression in the remaining 2 cases (50.0%). This significant correlation between Podoplanin expression and histological grading suggests that Podoplanin expression is higher in less differentiated OSCC, with well-differentiated tumors showing predominantly low expression. The findings suggest that Podoplanin may serve as a useful marker of tumor differentiation, with higher expression being indicative of greater aggressiveness and poorer differentiation. Table 1 Clinicopathologic parameters of OSCC patients and their association with Podoplanin expression (n = 56) Clinicopathologic Parameter Podoplanin Expression P value High Low Age distribution Up to 40 years 2(33.3) 4(66.7) 0.58 41–50 years 3 (42.9) 4(57.1) 51–60 years 4(25.0) 12(75.0) 61–70 years 7(50.0) 7(50.0) Above 70 years 3(23.1) 10(76.9) Gender Female 8(29.6) 19(70.4) 0.58 Male 11(37.9) 18(62.1) Clinical Presentation Ulcer 15(35.7) 27(64.3) 0.58 Red patches 2(33.3) 4(66.7) White patches 1 (33.3) 2(66.7) Exophytic growth 1 (20.0) 4(66.1) Characteristics Tobacco Yes 9(39.1) 14(60.9) 0.57 No 6(46.2) 7(53.8) Betel quid Yes 12 (34.3) 23(65.7) 1 No 7(33.3) 14(63.8) Tobacco & betel quid both Yes 3 (30.0) 7(70.0) 1 No 16(34.8) 30(66.1) Anatomic Location Tongue 7(46.7) 8(53.3) 0.24 Buccal mucosa 11(34.4) 21(65.6) Others 1 (11.1) 8(88.9) Table 1 Clinicopathologic Parameters of OSCC Patients and Their Association with Podoplanin Expression (n = 56) . The table summarizes the distribution of Podoplanin expression (high vs. low) based on various clinicopathologic parameters, including age, gender, clinical presentation, tobacco use, betel quid use, and anatomic location of the tumor. The P values indicate the statistical significance of the associations between Podoplanin expression and each parameter. No significant associations were observed between Podoplanin expression and age, gender, clinical presentation, or personal habits. The only significant association found was between Podoplanin expression and histological grading (not shown in this table). Table 2 Histological grading of OSCC and immune-histochemical expression of podoplanin (n = 56) Histological grading Expression of Podoplanin P Value* High Low Well differentiated 0(0.0) 25(100) 0.00 Moderately differentiated 17(63.0) 10(37) Poorly differentiated 2(50.0) 2(50.0) Table 2 Histological Grading of OSCC and Immunohistochemical Expression of Podoplanin (n = 56) . This table shows the distribution of high and low Podoplanin expression across different histological grades of OSCC, including well-differentiated, moderately differentiated, and poorly differentiated tumors. A statistically significant association was observed between histological grading and Podoplanin expression (P = 0.00), with high expression predominantly found in moderately and poorly differentiated OSCC, while well-differentiated tumors exhibited predominantly low expression Figure:1 Gender distribution of the patients with Oral SCC (n =56) The pie chart illustrates the gender distribution among the study participants. The results indicate a higher prevalence of oral SCC in males, with a male-to-female ratio of 1.07:1. Figure: 2 Locations of SCC in the oral cavity (n =56) The figure shows the distribution of oral squamous cell carcinoma (SCC) cases based on their anatomical location within the oral cavity. Among the 56 patients, the majority of biopsies (57.1%) were taken from the buccal mucosa, followed by the tongue. The remaining cases were found in the retromolar trigone (16.1%), alveolus (10.7%), and gingiva (3.6%). A small percentage of cases (1.8%) were located in other areas of the oral cavity. Figure 3: Histological grade of oral SCC (n=56) The histological grade of oral SCC was assessed as per criteria set by WHO 2017. Among 56 patients, 48.2% were moderately differentiated (grade 2) followed by 44.6% of well differentiated (grade 1). Only 7.1% showed poorly differentiated (grade 3). Discussion The present study demonstrates that Podoplanin expression is present in nearly all cases of oral squamous cell carcinoma (OSCC), with a statistically significant association observed with histological grading. Specifically, the expression of Podoplanin was found to be higher in moderately and poorly differentiated OSCC cases, while all well-differentiated tumors exhibited low expression. This finding supports the hypothesis that Podoplanin could serve as an important biomarker for the aggressive nature of OSCC, with its expression correlating with poorer histological differentiation. In this study, 33.9% of the cases showed high Podoplanin expression, while the remaining 66.1% exhibited low expression. These findings are consistent with those of other studies, although variability in the levels of Podoplanin expression has been reported in different populations globally. Previous studies have documented a range of Podoplanin expression, with high expression varying from 20–60% and low expression ranging from 21.6–80%. For instance, Yuan et al. (2006) reported that 60% of oral SCC cases exhibited high Podoplanin expression, while other investigators, such as Parhar et al. (2015), Patil et al. (2015), Kim et al. (2015), and Sgaramella et al. (2016), reported high expression rates of 56.6%, 20%, 51.3%, and 51%, respectively [ 12 , 14 – 17 ]. The considerable variability in these findings can be attributed to several factors, including genetic and ethnic diversity, personal habits such as tobacco and betel quid use, and environmental influences that contribute to the development of oral cancer [ 1 , 4 ]. It is important to note that while the current study found no significant association between Podoplanin expression and factors such as age, gender, anatomic site, clinical presentation, or the use of tobacco and betel quid, other studies have suggested that Podoplanin expression correlates with tumor size, nodal involvement, advanced stage, poor treatment response, and reduced survival rates [ 5 , 7 , 19 ]. These associations indicate the potential of Podoplanin as a prognostic marker in OSCC, which could aid in predicting clinical outcomes and guiding therapeutic decisions. Furthermore, the observed relationship between Podoplanin expression and histological grading is in line with studies by Kim et al. (2015), Pradhan et al. (2019), and Patil et al. (2015), who also reported significant associations between high Podoplanin expression and poorly differentiated or moderately differentiated tumors [ 16 , 18 , 19 ]. However, our results differ from studies by Yuan et al. (2006), Aiswariya et al. (2019), and Prasad et al. (2015), who did not observe the same level of correlation between Podoplanin expression and histological grade [ 12 , 13 , 19 ]. The reasons for these discrepancies can likely be attributed to differences in study design, sample sizes, and the specific characteristics of the populations studied. For instance, genetic and environmental factors, as well as variations in diagnostic and histopathological assessment methods, may contribute to differing findings. Therefore, while the relationship between Podoplanin expression and OSCC is becoming increasingly recognized, further research involving larger and more diverse cohorts is necessary to better understand the complex interplay of factors that influence its expression and its potential as a prognostic biomarker. In conclusion, the current study reinforces the notion that Podoplanin expression is closely linked to the histological grading of OSCC, particularly with higher expression observed in more aggressive, poorly differentiated tumors. Despite some inconsistencies in the literature, the potential of Podoplanin as a prognostic marker for OSCC, especially in predicting tumor progression and patient outcomes, remains promising and warrants further exploration. Conclusion In recent years, numerous studies have sought to elucidate the role of Podoplanin in oral squamous cell carcinoma (OSCC), yet significant controversies remain in the scientific literature. In this study, we observed that Podoplanin expression was present in nearly all OSCC cases in Bangladesh, with high expression predominantly associated with higher-grade tumors. Furthermore, tumors originating from the tongue and those exhibiting lymphovascular and perineural invasion showed elevated levels of Podoplanin expression. While these findings are promising, the study's interpretation is limited by the small sample size, reliance on small biopsy specimens, and the absence of follow-up data. To solidify these results, further research with a larger cohort, utilizing resected specimens, is essential. Additionally, future studies should explore the role of Podoplanin in tumor staging, nodal metastasis, overall survival, and its potential as a target for novel therapies. By advancing our understanding of Podoplanin’s prognostic value, we can pave the way for more precise diagnostic and therapeutic strategies in OSCC management. Declarations Data Availability Statement: The data supporting the findings of this study are available from the corresponding author upon reasonable request. Funding Statement: No specific funding was received for this study. Conflict of Interest Disclosure: The authors declare that there are no financial, personal, or professional conflicts of interest that could have influenced the work presented in this manuscript. All authors have disclosed any potential conflicts, and no competing interests exist. Ethics Approval Statement: This study was approved by the institutional ethics committee of Bangladesh Institute of Research and Rehabilitation in Diabetes, Endocrine and Metabolic Disorders (BIRDEM) General Hospital, Dhaka, Bangladesh Patient Consent Statement: Informed consent was obtained from all patients or their legal guardians prior to participation in the study. Contribution Statement The project was designed by the Principal Investigator, Mst. Rubeyatul Jannat. The data collection and article writing were primarily conducted by Mst. Rubeyatul Jannat, Nafisa Abedin, Sayed Kishwara Kashfi, Shamim Ahamed, and Shahana Sultana. Sayed Kishwara Kashfi also contributed to the literature review. Sadia Shirin provided essential photographic documentation for the study. Shamim Ahamed played a significant role in preparing the manuscript for publication. Shahana Sultana contributed extensively to the discussion section. Prof. Mousumi Ahmed provided crucial intellectual input, offered critical feedback on the manuscript drafts, and guided the team through key methodological and analytical aspects of the study. Prof. Rita Rani Barua offered critical advice regarding the publication process. Nafisa Abedin performed the final proofreading and served as the corresponding author. Acknowledgement: The study was guided and supported by Dr. Nazma Afroze, Professor of Pathology, whose expert advice and encouragement were invaluable throughout the research. Gratitude is extended to the medical technologists and staff at BIRDEM General Hospital, Dhaka, and Bangabandhu Sheikh Mujib Medical University (BSMMU) for their diligent assistance. Special recognition is given to Md. Joynal Abedin, Md. Fazle Elahi, Mrs. Shimu Akhter, Mr. Sojib Sorder, Md. Aorango Jeb Sheikh, and Mr. Ruhul Amin Bhuiyan for their contributions. Appreciation is also expressed to all the patients and their attendants who participated in this study, and to the colleagues and contemporary MD student Dr. Bilkis for their cooperation and support. Finally, sincere thanks are given to the families, including first author’s parents, in-laws, husband Md. A. Rahim, and daughter Manha, for their unwavering support and encouragement. References Sah R, Akhter M (2020) Oral Cancer Senario in Multiple Centers of Dhaka, Bangladesh. Biomedical J Sci Tech Res 32(2) Hossain MA, Ahmed MM, Rahman AFMS, Haider MN, Alam AK (2015) M. S. Epidemiological study of oral squamous cell carcinoma: A hospital based study in Dhaka city . The Journal of Teachers Association RMC (Vol. 28) Ferlay J, Ervik M, Lam F, Laversanne M, Colombet M, Mery L, Piñeros M, Znaor A, Soerjomataram I, Bray F (2024) Global Cancer Observatory: Cancer Today. Lyon, France: International Agency for Research on Cancer. 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Oncol Rep 34(2):833–842 Sgaramella N, Jonsson EL, Boldrup L, Califano L, Coates PJ, Tartaro G, Muzio L, Lo, Fåhraeus R, Colella G, Dell’ G, Orabona A, Loljung L, Santagata M, Rossiello R, Wilms T, Danielsson K, Laurell O, G., and, Nylander K (2016) High expression of podoplanin in squamous cell carcinoma of the tongue occurs predominantly in patients £ 40 years but does not correlate with tumour spread. 2:3–8 Pradhan S, Guddattu V, Solomon MC (2019) Association of the co-expression of SOX2 and podoplanin in the progression of oral squamous cell carcinomas - An immunohistochemical study. J Appl Oral Sci 27 Prasad B, Kashyap B, Babu G, Kumar G, Manyam R (2015) Expression of podoplanin in different grades of oral squamous cell carcinoma. Annals Med Health Sci Res 5(4):299 Additional Declarations The authors declare no competing interests. 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Bangladesh.","correspondingAuthor":false,"prefix":"","firstName":"Shahana","middleName":"","lastName":"Sultana","suffix":""},{"id":401191976,"identity":"11c07b3e-ff0b-4825-a014-6fdcea5ce210","order_by":6,"name":"Mousumi Ahmed","email":"","orcid":"","institution":"Bangladesh Institute of Research and Rehabilitation in Diabetes, Endocrine and Metabolic Disorders (BIRDEM) Hospital, Dhaka, Bangladesh","correspondingAuthor":false,"prefix":"","firstName":"Mousumi","middleName":"","lastName":"Ahmed","suffix":""},{"id":401191977,"identity":"dcf38428-1e7a-4cf7-b830-0b6c6f59561f","order_by":7,"name":"Rita Rani Barua","email":"","orcid":"","institution":"Dr. Sirajul islam medical college, Dhaka, Bangladesh.","correspondingAuthor":false,"prefix":"","firstName":"Rita","middleName":"Rani","lastName":"Barua","suffix":""}],"badges":[],"createdAt":"2025-01-12 20:08:44","currentVersionCode":1,"declarations":{"humanSubjects":true,"vertebrateSubjects":false,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":true,"humanSubjectConsent":true,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":false},"doi":"10.21203/rs.3.rs-5815305/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5815305/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":73760822,"identity":"597cab35-6002-4a53-ad9e-38d0ded6c870","added_by":"auto","created_at":"2025-01-14 11:25:55","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":43248,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGender distribution of the patients with Oral SCC (n =56)\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/e0c884dfc41884a1f45e8c14.png"},{"id":73760821,"identity":"d4bb47d8-9386-4149-af01-5c3c76e41a5a","added_by":"auto","created_at":"2025-01-14 11:25:55","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":11213,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eLocations of SCC in the oral cavity (n =56)\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/9c964e6b64399c1760d00fe7.png"},{"id":73760823,"identity":"40632ac6-973b-4d03-bb54-18c7c7611e61","added_by":"auto","created_at":"2025-01-14 11:25:56","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":63236,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ePie diagram showing Histological Grade of OSCC\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/c79d3f3b7bf390e764989bca.png"},{"id":73759696,"identity":"1c311335-5d43-423e-923b-7ba8bab3b630","added_by":"auto","created_at":"2025-01-14 11:17:56","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":645223,"visible":true,"origin":"","legend":"\u003cp\u003ePhotomicrograph showing Podoplanin high expression, IRS 12(40X)\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/ec7a0284262c87b5c214e57b.png"},{"id":73759694,"identity":"84564fc7-b6bc-454f-aaf0-354e9f81054d","added_by":"auto","created_at":"2025-01-14 11:17:56","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":497551,"visible":true,"origin":"","legend":"\u003cp\u003ePhotomicrograph showing \u0026nbsp;showing Podoplanin high expression, IRS 12(40 X) Podoplanin high expression, IRS 15 (40X)\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/fb4269909128966fee90db57.png"},{"id":73759709,"identity":"8c6ce6d1-cdbf-459d-a64b-5980e30db520","added_by":"auto","created_at":"2025-01-14 11:17:56","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":376432,"visible":true,"origin":"","legend":"\u003cp\u003ePhotomicrograph showing low expression of Podoplanin, IRS- 4 (40X)\u003c/p\u003e","description":"","filename":"6.png","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/373df143e4ca36bb567ee438.png"},{"id":73759697,"identity":"505b0127-9d12-4e63-9b5e-1f47e830b649","added_by":"auto","created_at":"2025-01-14 11:17:56","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":434718,"visible":true,"origin":"","legend":"\u003cp\u003ePhotomicrograph showing low expression of Podoplanin, IRS- 0( 40 X)\u003c/p\u003e","description":"","filename":"7.png","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/085d701685410a733b7dd821.png"},{"id":73761139,"identity":"2a73861a-75ad-4d04-8927-460e8bd48579","added_by":"auto","created_at":"2025-01-14 11:33:57","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3371601,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/52c4532e-1ba2-444a-aa11-1b0908decdde.pdf"},{"id":73759686,"identity":"f679554d-7e6d-40d3-9919-bc7534c4f3b0","added_by":"auto","created_at":"2025-01-14 11:17:55","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":36649,"visible":true,"origin":"","legend":"","description":"","filename":"mastersheetRub.docx","url":"https://assets-eu.researchsquare.com/files/rs-5815305/v1/d48f7afe938fc00bd265125e.docx"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003eAssociation of Podoplanin Expression with Histological Grade in Oral Squamous Cell Carcinoma in a Tertiary Care Hospital in Bangladesh\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eOral cancer is the sixth most common malignancy worldwide and is one of the major public health problems\u0026sup1;. Oral squamous cell carcinoma (OSCC) is the most common type of malignancy, representing up to 90 percent of all oral neoplasms\u0026sup2;. Oral cancer ranks 3rd, 2nd, and 3rd in five-year prevalence, incidence, and mortality rate respectively in Bangladesh\u0026sup3;. Common risk factors for developing OSCC are tobacco smoking, betel nut chewing, and alcohol consumption. Other predisposing factors are viral infection, chronic irritation, oral candidiasis, poor oral hygiene, ionizing radiation, etc⁴.\u003c/p\u003e \u003cp\u003ePodoplanin (PDPN) is known to be expressed in numerous human tumors, including squamous cell carcinoma of the oral cavity, mesothelioma, testicular seminoma, soft tissue tumors, thymoma, and brain tumors⁵. PDPN seems to be expressed in aggressive tumors, with higher invasive and metastatic potential⁶.\u003c/p\u003e \u003cp\u003ePDPN consists of an extracellular or transmembrane domain and a cytoplasmic domain. The extracellular domain is longer and carries four platelet aggregation-stimulating (PLAG) domains with plenty of potential O-glycosylation sites, crucial for interaction with platelets. The interaction between PDPN and c-type lectin (CLEC2) may regulate tumor invasion and metastasis⁷. The shorter cytoplasmic domain is associated with ezrin/radixin/moesin (ERM) protein that bridges plasma membrane proteins and the actin cytoskeleton⁸. PDPN overexpression causes marked phosphorylation of ERM proteins. It causes adhesion and cancer cell migration through modulating the actin cytoskeleton, Rho A, and epithelial-mesenchymal transition. PDPN causes down-regulation of E-cadherin and overexpression of matrix metalloproteinase⁶.\u003c/p\u003e \u003cp\u003eIt is upregulated in the invasive front of OSCC. It causes upregulation of TGF-β1 secretion and increases Epidermal Growth Factor Receptor phosphorylation as well as downstream effectors of AKT and ERK. PDPN also causes OSCC progression by interacting with matrix metalloproteinase and induces cytoskeletal remodelling, extracellular matrix degradation, and invasion⁹.\u003c/p\u003e \u003cp\u003ePDPN in oral SCC causes tumor cell motility and metastasis by activation of endogenous lectin that binds to extracellular carbohydrate moieties. These cancer cells are remarkably resistant to currently available chemotherapy. Thus, PDPN becomes a chemotherapeutic target for oral SCC\u0026sup1;⁰. Anti-human Podoplanin antibody (NZ-1) would be an important target for the prevention of metastasis in the future for OSCC⁶.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e1. Study Design and Subjects\u003c/h2\u003e \u003cp\u003eThis cross-sectional observational study was conducted at the Department of Pathology, Bangladesh Institute of Research and Rehabilitation in Diabetes, Endocrine and Metabolic Disorder (BIRDEM), during the period of March 2022 to February 2024. A total of 56 histologically diagnosed cases of oral squamous cell carcinoma (OSCC) were included in this study, meeting the following inclusion criteria: small and resected biopsy cases of OSCC. Exclusion criteria included carcinoma in situ, dysplastic lesions of the oral cavity, and patients who received neoadjuvant therapy or radiotherapy before surgery.\u003c/p\u003e \u003cp\u003eDemographic and clinical information, such as age, sex, clinical presentation, and histopathological diagnosis (including grade), were collected from the departmental records of the pathology department.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003e2. Sample Collection\u003c/h3\u003e\n\u003cp\u003eThe 56 cases of oral SCC were retrieved from the Department of Pathology, BIRDEM, including H\u0026amp;E stained slides and paraffin-embedded blocks. All slides were reviewed to assess tumor grade and other prognostic factors, including tumor-infiltrating lymphocytes (TILs), and the presence of lymphovascular and perineural invasion.\u003c/p\u003e\n\u003ch3\u003e3. Immunohistochemical Staining for Podoplanin\u003c/h3\u003e\n\u003cp\u003eAfter confirming the diagnosis of OSCC, immunohistochemical (IHC) staining for podoplanin was performed at Bangabandhu Sheikh Mujib Medical University (BSMMU), using an appropriate positive control. Paraffin-embedded tissue blocks were sectioned at 3 \u0026micro;m thickness. The slides were gently lowered onto a water bath set to 45\u0026deg;C, spread without wrinkles onto slides coated with 0.1% poly-L-lysine, and air-dried. Slides were then baked at 60\u0026deg;C for 30 minutes on a hot plate. Dewaxing was performed by treating the slides in xylene, followed by rehydration through a graded series of alcohols.\u003c/p\u003e \u003cp\u003eFor antigen retrieval, the slides were immersed in preheated citrate buffer in a pressure cooker, boiled, and allowed to cool naturally. To block endogenous enzyme activity, hydrogen peroxide was added in a moist chamber at room temperature.\u003c/p\u003e\n\u003ch3\u003e4. Primary and Secondary Antibodies\u003c/h3\u003e\n\u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003ePrimary antibody\u003c/b\u003e: Monoclonal Rabbit d2-40 (pre-diluted, ready-to-use).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eSecondary antibody\u003c/b\u003e: DAKO REAL\u0026trade; Envision\u0026trade; (HRP Rabbit/Mouse) (ENV).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003ePositive control\u003c/b\u003e: Appedicular tissue was used as a positive control.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eChromogen\u003c/b\u003e: 3,3\u0026rsquo;-diaminobenzidine (DAB) was used for visualizing the antigen-antibody complex. Finally, slides were counterstained with hematoxylin.\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e\n\u003ch3\u003e5. Scoring of Podoplanin Expression\u003c/h3\u003e\n\u003cp\u003ePodoplanin expression was assessed by reviewing 10 representative areas from each slide. For each area, 100 tumor cells were counted, and both the percentage of positive cells and the intensity of podoplanin expression were evaluated. The following scoring system, as introduced by Yuan et al. (2006), was applied:\u003c/p\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eQuantitative Score:\u003c/h2\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e0\u003c/b\u003e: 0% positive cells\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e1\u003c/b\u003e: 1\u0026ndash;10% positive cells\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e2\u003c/b\u003e: 11\u0026ndash;30% positive cells\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e3\u003c/b\u003e: 31\u0026ndash;50% positive cells\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e4\u003c/b\u003e: 51\u0026ndash;80% positive cells\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e5\u003c/b\u003e: 81\u0026ndash;100% positive cells\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eIntensity Score:\u003c/h3\u003e\n\u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e0\u003c/b\u003e: Negative (no staining)\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e1\u003c/b\u003e: Weak (faint staining)\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e2\u003c/b\u003e: Moderate (staining between weak and dark brown)\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e3\u003c/b\u003e: Strong (dark brown staining)\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eThe \u003cb\u003eimmunoreactive score (IRS)\u003c/b\u003e for podoplanin expression was calculated by multiplying the quantitative score by the intensity score, yielding a final score ranging from \u003cb\u003e0 to 15\u003c/b\u003e. According to this score, the expression was classified as follows:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e0\u0026ndash;7\u003c/b\u003e: Low expression\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003e\u0026ge;\u0026thinsp;8\u003c/b\u003e: High expression\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e\n\u003ch3\u003e6. Statistical Analysis\u003c/h3\u003e\n\u003cp\u003eAll collected data were compiled and analyzed using \u003cb\u003eSPSS (Statistical Package for Social Sciences), version 25\u003c/b\u003e. The Chi-square test was used to assess statistical significance between different clinicopathologic variables. A \u003cb\u003ep-value of \u0026lt;\u0026thinsp;0.05\u003c/b\u003e was considered statistically significant.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eA total of 56 oral squamous cell carcinoma (OSCC) patients were included in the study, and their clinicopathologic characteristics along with Podoplanin expression were analyzed. The association between Podoplanin expression and various clinicopathologic parameters, as well as histological grading, was evaluated (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003ePodoplanin Expression and Clinicopathologic Parameters\u003c/h2\u003e \u003cp\u003ePodoplanin expression was classified as high or low based on immunohistochemical staining. High Podoplanin expression was observed in 19 cases (33.90%) of OSCC. The distribution of high and low Podoplanin expression did not show any statistically significant association with age, gender, clinical presentation, or personal habits (tobacco use, betel quid use). Specifically, no significant differences in Podoplanin expression were noted across different age groups (P\u0026thinsp;=\u0026thinsp;0.58), between genders (P\u0026thinsp;=\u0026thinsp;0.58), or based on the clinical presentation (P\u0026thinsp;=\u0026thinsp;0.58). Furthermore, Podoplanin expression did not correlate with tobacco use (P\u0026thinsp;=\u0026thinsp;0.57), betel quid use (P\u0026thinsp;=\u0026thinsp;1.0), or both tobacco and betel quid use (P\u0026thinsp;=\u0026thinsp;1.0). Similarly, no significant association was found between Podoplanin expression and the anatomic location of the tumors (P\u0026thinsp;=\u0026thinsp;0.24) (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003ePodoplanin Expression and Histological Grading\u003c/h2\u003e \u003cp\u003eThe association between Podoplanin expression and the histological grading of OSCC was found to be statistically significant (P\u0026thinsp;=\u0026thinsp;0.00) (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Podoplanin expression varied considerably across different grades of differentiation. In well-differentiated OSCC (n\u0026thinsp;=\u0026thinsp;25), no cases exhibited high Podoplanin expression, with all tumors showing low expression. In moderately differentiated OSCC (n\u0026thinsp;=\u0026thinsp;27), 17 cases (63.0%) exhibited high Podoplanin expression, while 10 cases (37.0%) showed low expression. In poorly differentiated OSCC (n\u0026thinsp;=\u0026thinsp;4), high Podoplanin expression was observed in 2 cases (50.0%) and low expression in the remaining 2 cases (50.0%).\u003c/p\u003e \u003cp\u003eThis significant correlation between Podoplanin expression and histological grading suggests that Podoplanin expression is higher in less differentiated OSCC, with well-differentiated tumors showing predominantly low expression. The findings suggest that Podoplanin may serve as a useful marker of tumor differentiation, with higher expression being indicative of greater aggressiveness and poorer differentiation.\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 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e\u003cb\u003eClinicopathologic parameters of OSCC patients and their association with Podoplanin\u003c/b\u003e expression (n\u0026thinsp;=\u0026thinsp;56)\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\" colspan=\"2\" morerows=\"1\" nameend=\"c2\" namest=\"c1\" rowspan=\"2\"\u003e \u003cp\u003eClinicopathologic Parameter\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c4\" namest=\"c3\"\u003e \u003cp\u003ePodoplanin Expression\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eP value\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eHigh\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eLow\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003eAge distribution\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\u003eUp to 40 years\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2(33.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4(66.7)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"4\" rowspan=\"5\"\u003e \u003cp\u003e0.58\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003e41\u0026ndash;50 years\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3 (42.9)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4(57.1)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003e51\u0026ndash;60 years\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4(25.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e12(75.0)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003e61\u0026ndash;70 years\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e7(50.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e7(50.0)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eAbove 70 years\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3(23.1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e10(76.9)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eGender\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eFemale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e8(29.6)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e19(70.4)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e0.58\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eMale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11(37.9)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e18(62.1)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eClinical Presentation\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eUlcer\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e15(35.7)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e27(64.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003e0.58\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eRed patches\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2(33.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4(66.7)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eWhite patches\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1 (33.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2(66.7)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eExophytic growth\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1 (20.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4(66.1)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eCharacteristics\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\u003eTobacco\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eYes\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9(39.1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e14(60.9)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e0.57\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6(46.2)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e7(53.8)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eBetel quid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eYes\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e12 (34.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e23(65.7)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e7(33.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e14(63.8)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eTobacco \u0026amp; betel quid both\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eYes\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3 (30.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e7(70.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e16(34.8)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e30(66.1)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eAnatomic Location\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eTongue\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e7(46.7)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8(53.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003e0.24\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eBuccal mucosa\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11(34.4)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e21(65.6)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eOthers\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1 (11.1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8(88.9)\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 \u003cstrong\u003eTable\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e\u003c/strong\u003e \u003cp\u003e \u003cem\u003eClinicopathologic Parameters of OSCC Patients and Their Association with Podoplanin Expression (n\u0026thinsp;=\u0026thinsp;56)\u003c/em\u003e. The table summarizes the distribution of Podoplanin expression (high vs. low) based on various clinicopathologic parameters, including age, gender, clinical presentation, tobacco use, betel quid use, and anatomic location of the tumor. The P values indicate the statistical significance of the associations between Podoplanin expression and each parameter. No significant associations were observed between Podoplanin expression and age, gender, clinical presentation, or personal habits. The only significant association found was between Podoplanin expression and histological grading (not shown in this table).\u003c/p\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 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eHistological grading of OSCC and immune-histochemical expression of podoplanin (n\u0026thinsp;=\u0026thinsp;56)\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\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 \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eHistological grading\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c3\" namest=\"c2\"\u003e \u003cp\u003eExpression of Podoplanin\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eP Value*\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eHigh\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eLow\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eWell differentiated\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0(0.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e25(100)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003e0.00\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eModerately differentiated\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e17(63.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10(37)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePoorly differentiated\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2(50.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2(50.0)\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 \u003cstrong\u003eTable\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e\u003c/strong\u003e \u003cp\u003e \u003cem\u003eHistological Grading of OSCC and Immunohistochemical Expression of Podoplanin (n\u0026thinsp;=\u0026thinsp;56)\u003c/em\u003e. This table shows the distribution of high and low Podoplanin expression across different histological grades of OSCC, including well-differentiated, moderately differentiated, and poorly differentiated tumors. A statistically significant association was observed between histological grading and Podoplanin expression (P\u0026thinsp;=\u0026thinsp;0.00), with high expression predominantly found in moderately and poorly differentiated OSCC, while well-differentiated tumors exhibited predominantly low expression\u003c/p\u003e \u003c/p\u003e \n\u003cp\u003e\u003cstrong\u003eFigure:1 Gender\u0026nbsp;distribution\u0026nbsp;of the\u0026nbsp;patients with\u0026nbsp;Oral\u0026nbsp;SCC\u0026nbsp;(n =56)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe pie chart illustrates the gender distribution among the study participants. The results indicate a higher prevalence of oral SCC in males, with a male-to-female ratio of 1.07:1.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFigure: 2 Locations\u0026nbsp;of\u0026nbsp;SCC\u0026nbsp;in the\u0026nbsp;oral\u0026nbsp;cavity (n =56)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe figure shows the distribution of oral squamous cell carcinoma (SCC) cases based on their anatomical location within the oral cavity. Among the 56 patients, the majority of biopsies (57.1%) were taken from the buccal mucosa, followed by the tongue. The remaining cases were found in the retromolar trigone (16.1%), alveolus (10.7%), and gingiva (3.6%). A small percentage of cases (1.8%) were located in other areas of the oral cavity.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFigure 3: Histological\u0026nbsp;grade\u0026nbsp;of\u0026nbsp;oral SCC (n=56)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe histological grade of oral SCC was assessed as per criteria set by WHO 2017. Among 56 patients, 48.2% were moderately differentiated (grade 2) followed by 44.6% of well differentiated (grade 1). Only 7.1% showed poorly differentiated (grade 3).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe present study demonstrates that Podoplanin expression is present in nearly all cases of oral squamous cell carcinoma (OSCC), with a statistically significant association observed with histological grading. Specifically, the expression of Podoplanin was found to be higher in moderately and poorly differentiated OSCC cases, while all well-differentiated tumors exhibited low expression. This finding supports the hypothesis that Podoplanin could serve as an important biomarker for the aggressive nature of OSCC, with its expression correlating with poorer histological differentiation.\u003c/p\u003e \u003cp\u003eIn this study, 33.9% of the cases showed high Podoplanin expression, while the remaining 66.1% exhibited low expression. These findings are consistent with those of other studies, although variability in the levels of Podoplanin expression has been reported in different populations globally. Previous studies have documented a range of Podoplanin expression, with high expression varying from 20\u0026ndash;60% and low expression ranging from 21.6\u0026ndash;80%. For instance, Yuan et al. (2006) reported that 60% of oral SCC cases exhibited high Podoplanin expression, while other investigators, such as Parhar et al. (2015), Patil et al. (2015), Kim et al. (2015), and Sgaramella et al. (2016), reported high expression rates of 56.6%, 20%, 51.3%, and 51%, respectively [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan additionalcitationids=\"CR15 CR16\" citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. The considerable variability in these findings can be attributed to several factors, including genetic and ethnic diversity, personal habits such as tobacco and betel quid use, and environmental influences that contribute to the development of oral cancer [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIt is important to note that while the current study found no significant association between Podoplanin expression and factors such as age, gender, anatomic site, clinical presentation, or the use of tobacco and betel quid, other studies have suggested that Podoplanin expression correlates with tumor size, nodal involvement, advanced stage, poor treatment response, and reduced survival rates [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. These associations indicate the potential of Podoplanin as a prognostic marker in OSCC, which could aid in predicting clinical outcomes and guiding therapeutic decisions.\u003c/p\u003e \u003cp\u003eFurthermore, the observed relationship between Podoplanin expression and histological grading is in line with studies by Kim et al. (2015), Pradhan et al. (2019), and Patil et al. (2015), who also reported significant associations between high Podoplanin expression and poorly differentiated or moderately differentiated tumors [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. However, our results differ from studies by Yuan et al. (2006), Aiswariya et al. (2019), and Prasad et al. (2015), who did not observe the same level of correlation between Podoplanin expression and histological grade [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe reasons for these discrepancies can likely be attributed to differences in study design, sample sizes, and the specific characteristics of the populations studied. For instance, genetic and environmental factors, as well as variations in diagnostic and histopathological assessment methods, may contribute to differing findings. Therefore, while the relationship between Podoplanin expression and OSCC is becoming increasingly recognized, further research involving larger and more diverse cohorts is necessary to better understand the complex interplay of factors that influence its expression and its potential as a prognostic biomarker.\u003c/p\u003e \u003cp\u003eIn conclusion, the current study reinforces the notion that Podoplanin expression is closely linked to the histological grading of OSCC, particularly with higher expression observed in more aggressive, poorly differentiated tumors. Despite some inconsistencies in the literature, the potential of Podoplanin as a prognostic marker for OSCC, especially in predicting tumor progression and patient outcomes, remains promising and warrants further exploration.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eIn recent years, numerous studies have sought to elucidate the role of Podoplanin in oral squamous cell carcinoma (OSCC), yet significant controversies remain in the scientific literature. In this study, we observed that Podoplanin expression was present in nearly all OSCC cases in Bangladesh, with high expression predominantly associated with higher-grade tumors. Furthermore, tumors originating from the tongue and those exhibiting lymphovascular and perineural invasion showed elevated levels of Podoplanin expression. While these findings are promising, the study's interpretation is limited by the small sample size, reliance on small biopsy specimens, and the absence of follow-up data. To solidify these results, further research with a larger cohort, utilizing resected specimens, is essential. Additionally, future studies should explore the role of Podoplanin in tumor staging, nodal metastasis, overall survival, and its potential as a target for novel therapies. By advancing our understanding of Podoplanin\u0026rsquo;s prognostic value, we can pave the way for more precise diagnostic and therapeutic strategies in OSCC management.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eData Availability Statement:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data supporting the findings of this study are available from the corresponding author upon reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding Statement:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNo specific funding was received for this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of Interest Disclosure:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that there are no financial, personal, or professional conflicts of interest that could have influenced the work presented in this manuscript.\u003c/p\u003e\n\u003cp\u003eAll authors have disclosed any potential conflicts, and no competing interests exist.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics Approval Statement:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was approved by the institutional ethics committee of Bangladesh Institute of Research and Rehabilitation in Diabetes, Endocrine and Metabolic Disorders (BIRDEM) General Hospital, Dhaka, Bangladesh\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePatient Consent Statement:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eInformed consent was obtained from all patients or their legal guardians prior to participation in the study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eContribution Statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe project was designed by the Principal Investigator, Mst. Rubeyatul Jannat. The data collection and article writing were primarily conducted by Mst. Rubeyatul Jannat, Nafisa Abedin, Sayed Kishwara Kashfi, Shamim Ahamed, and Shahana Sultana. Sayed Kishwara Kashfi also contributed to the literature review. Sadia Shirin provided essential photographic documentation for the study. Shamim Ahamed played a significant role in preparing the manuscript for publication. Shahana Sultana contributed extensively to the discussion section. Prof. Mousumi Ahmed provided crucial intellectual input, offered critical feedback on the manuscript drafts, and guided the team through key methodological and analytical aspects of the study. Prof. Rita Rani Barua offered critical advice regarding the publication process. Nafisa Abedin performed the final proofreading and served as the corresponding author.\u003c/p\u003e\n\u003cp\u003e\u003cb\u003eAcknowledgement:\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe study was guided and supported by Dr. Nazma Afroze, Professor of Pathology, whose expert advice and encouragement were invaluable throughout the research.\u003c/p\u003e \u003cp\u003eGratitude is extended to the medical technologists and staff at BIRDEM General Hospital, Dhaka, and Bangabandhu Sheikh Mujib Medical University (BSMMU) for their diligent assistance. Special recognition is given to Md. Joynal Abedin, Md. Fazle Elahi, Mrs. Shimu Akhter, Mr. Sojib Sorder, Md. Aorango Jeb Sheikh, and Mr. Ruhul Amin Bhuiyan for their contributions.\u003c/p\u003e \u003cp\u003eAppreciation is also expressed to all the patients and their attendants who participated in this study, and to the colleagues and contemporary MD student Dr. Bilkis for their cooperation and support.\u003c/p\u003e \u003cp\u003eFinally, sincere thanks are given to the families, including first author\u0026rsquo;s parents, in-laws, husband Md. A. Rahim, and daughter Manha, for their unwavering support and encouragement.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eSah R, Akhter M (2020) Oral Cancer Senario in Multiple Centers of Dhaka, Bangladesh. Biomedical J Sci Tech Res 32(2)\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHossain MA, Ahmed MM, Rahman AFMS, Haider MN, Alam AK (2015) M. S. \u003cem\u003eEpidemiological study of oral squamous cell carcinoma: A hospital based study in Dhaka city\u003c/em\u003e. \u003cem\u003eThe Journal of Teachers Association RMC\u003c/em\u003e (Vol. 28)\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFerlay J, Ervik M, Lam F, Laversanne M, Colombet M, Mery L, Pi\u0026ntilde;eros M, Znaor A, Soerjomataram I, Bray F (2024) Global Cancer Observatory: Cancer Today. Lyon, France: International Agency for Research on Cancer. Availablefrom: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://gco.iarc.who.int/today\u003c/span\u003e\u003cspan address=\"https://gco.iarc.who.int/today\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHannan MA, Rahman MA, Hossain S, Rahman QB (2018) Role Of Habitual Risk Factors On Oral Squamous Cell Carcinoma. Update Dent Coll J (8)\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLee HY, Yu NY, Lee SH, Tsai HJ, Wu CC, Cheng JC, Chen DP, Wang YR, Tseng CP (2020) Podoplanin promotes cancer-associated thrombosis and contributes to the unfavorable overall survival in an ectopic xenograft mouse model of oral cancer. Biomedical J 43(2):146\u0026ndash;162\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMARIUS R, CIMPEAN A M (2008) and DOMENICO R The Role of Podoplanin in Tumor Progression and Metastasis. \u003cem\u003eANTICANCER RESEARCH 28: 2997\u0026ndash;3006 (2008)\u003c/em\u003e 28: 2997\u0026ndash;3006\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHwang BO, Park SY, Cho ES, Zhang X, Lee SK, Ahn HJ, Chun KS, Chung WY, Song NY (2021/15/December) Platelet CLEC2-Podoplanin Axis as a Promising Target for Oral Cancer Treatment. \u003cem\u003eFrontiers in Immunology\u003c/em\u003e. Frontiers Media S.A\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAstarita JL, Acton SE, Turley SJ (2012) Podoplanin: Emerging functions in development, the immune system, and cancer. Frontiers in Immunology\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLi Y, yin, Zhou CX, Gao Y (2018) Interaction between oral squamous cell carcinoma cells and fibroblasts through TGF-β1 mediated by podoplanin. Exp Cell Res 369(1):43\u0026ndash;53\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOchoa-Alvarez JA, Krishnan H, Pastorino JG, Nevel E, Kephart D, Lee JJ et al (2015) Antibody and lectin target podoplanin to inhibit oral squamous carcinoma cell migration and viability by distinct mechanisms\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOliveira LR, Ribeiro-Silva A (2011) /March) Prognostic significance of immunohistochemical biomarkers in oral squamous cell carcinoma. Int J Oral Maxillofac Surg\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYuan P, Temam S, El-Naggar A, Zhou X, Liu DD, Lee JJ, Mao L (2006) Overexpression of podoplanin in oral cancer and its association with poor clinical outcome. Cancer 107(3):563\u0026ndash;569\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAiswarya A, Suresh R, Janardhanan M, Savithri V, Aravind T, Mathew L (2019) An immunohistochemical evaluation of podoplanin expression in oral leukoplakia and oral squamous cell carcinoma to explore its potential to be used as a predictor for malignant transformation. J Oral Maxillofacial Pathol 23(1):159\u0026ndash;160\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eParhar S, Kaur H, Vashist A, Verma S (2015) Role of podoplanin in potentially malignant disorders and oral squamous cell carcinoma and its correlation with lymphangiogenesis. In \u003cem\u003eIndian Journal of Cancer\u003c/em\u003e (Vol. 52). Medknow Publications\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePatil A, Patil K, Tupsakhare S, Gabhane M, Sonune S, Kandalgaonkar S (2015) Evaluation of Podoplanin in Oral Leukoplakia and Oral Squamous Cell Carcinoma. \u003cem\u003eScientifica\u003c/em\u003e 2015: 1\u0026ndash;6\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKim HY, Rha KS, Shim GA, Kim JH, Kim JM, Huang SM, Koo BS (2015) Podoplanin is involved in the prognosis of head and neck squamous cell carcinoma through interaction with VEGF-C. Oncol Rep 34(2):833\u0026ndash;842\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSgaramella N, Jonsson EL, Boldrup L, Califano L, Coates PJ, Tartaro G, Muzio L, Lo, F\u0026aring;hraeus R, Colella G, Dell\u0026rsquo; G, Orabona A, Loljung L, Santagata M, Rossiello R, Wilms T, Danielsson K, Laurell O, G., and, Nylander K (2016) High expression of podoplanin in squamous cell carcinoma of the tongue occurs predominantly in patients \u0026pound; 40 years but does not correlate with tumour spread. 2:3\u0026ndash;8\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePradhan S, Guddattu V, Solomon MC (2019) Association of the co-expression of SOX2 and podoplanin in the progression of oral squamous cell carcinomas - An immunohistochemical study. J Appl Oral Sci 27\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePrasad B, Kashyap B, Babu G, Kumar G, Manyam R (2015) Expression of podoplanin in different grades of oral squamous cell carcinoma. Annals Med Health Sci Res 5(4):299\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"BIRDEM","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Podoplanin, Oral Squamous Cell Carcinoma, Immunohistochemistry, Histological Grade, Prognosis, Biomarker","lastPublishedDoi":"10.21203/rs.3.rs-5815305/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5815305/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003eOral carcinogenesis is a complex process driven by genetic alterations affecting proto-oncogenes and tumor suppressor genes, with key players including P16, P53, H-ras, cyclinD1, and EGFR. Podoplanin, a transmembrane glycoprotein, has emerged as a biomarker implicated in tumor progression and prognosis in oral squamous cell carcinoma (OSCC). This study aimed to evaluate the immunohistochemical expression of podoplanin across different histological grades of OSCC and analyze its association with clinicopathological parameters such as age, gender, anatomical site, personal habits, lymphovascular and perineural invasion, and tumor-infiltrating lymphocytes.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eA cross-sectional observational study was conducted at the Department of Pathology, Bangladesh Institute of Research and Rehabilitation in Diabetes, Endocrine and Metabolic Disorders (BIRDEM) Hospital, Dhaka, Bangladesh, over two years. Fifty-six cases of OSCC were analyzed using hematoxylin and eosin-stained slides and paraffin-embedded tissue blocks. Podoplanin expression was assessed immunohistochemically, and staining intensity and extent were categorized as high or low.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eThe results revealed high podoplanin expression in 33.90% of cases and low expression in 66.10%. High podoplanin expression was more frequently observed in moderately and poorly differentiated OSCC compared to well-differentiated tumors, with a statistically significant correlation between podoplanin expression and histological grade. However, no significant associations were identified between podoplanin expression and other factors, including age, gender, personal habits, anatomical site, lymphovascular invasion, perineural invasion, or tumor-infiltrating lymphocytes.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003ePodoplanin expression was significantly associated with the histological grade of OSCC, suggesting its potential role as a prognostic marker. High podoplanin expression may indicate aggressive tumor behavior and poor prognosis, underscoring its relevance in OSCC evaluation and management.\u003c/p\u003e","manuscriptTitle":"Association of Podoplanin Expression with Histological Grade in Oral Squamous Cell Carcinoma in a Tertiary Care Hospital in Bangladesh","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-01-14 11:17:51","doi":"10.21203/rs.3.rs-5815305/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"06ea24cd-86c5-4361-b3da-b4191183f6b2","owner":[],"postedDate":"January 14th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-01-14T11:17:51+00:00","versionOfRecord":[],"versionCreatedAt":"2025-01-14 11:17:51","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-5815305","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5815305","identity":"rs-5815305","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}