Serum Trefoil Factor 3 Levels in Women with Polycystic Ovary Syndrome: A Cross- Sectional Study

Serum Trefoil Factor 3 Levels in Women with Polycystic Ovary Syndrome: A Cross- Sectional Study | 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 Serum Trefoil Factor 3 Levels in Women with Polycystic Ovary Syndrome: A Cross- Sectional Study Fulya Cagli, Merve Ozel Yetkin, Varol Gulseren, Onur Balkaya, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9258003/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 5 You are reading this latest preprint version Abstract Background Polycystic ovary syndrome (PCOS) is associated with chronic anovulation and prolonged unopposed estrogen exposure, which may contribute to endometrial dysfunction. Trefoil factor 3 (TFF3) is an estrogen-responsive peptide involved in epithelial repair and mucosal integrity. However, data on circulating TFF3 levels in women with PCOS are limited. Methods This exploratory cross-sectional study included 41 women with PCOS and 34 age-matched healthy controls. Serum TFF3 levels were measured and compared between groups. The association between TFF3 levels and menstrual cycle patterns was also evaluated. Receiver operating characteristic (ROC) analysis was performed to assess the discriminatory performance of TFF3 for PCOS. Results Serum TFF3 levels were significantly higher in women with PCOS compared with controls (1742 ± 582 vs. 1185 ± 704, p < 0.001). TFF3 levels were not associated with menstrual cycle pattern or oligomenorrhea. ROC analysis showed that TFF3 had a moderate ability to discriminate PCOS (AUC: 0.728), with a sensitivity of 80.5% and a specificity of 61.8% at the optimal cut-off value. No independent associations were identified in multivariate regression analysis. Conclusion Serum TFF3 levels are elevated in women with PCOS, suggesting a possible link between TFF3 and PCOS-related hormonal and endometrial changes. However, the lack of association with menstrual patterns and the moderate diagnostic performance indicate that TFF3 alone is unlikely to serve as a reliable biomarker. Further studies are needed to clarify its biological and clinical significance. Polycystic ovary syndrome Trefoil factor 3 Endometrium Estrogen AMH Figures Figure 1 Figure 2 Introduction Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders affecting women of reproductive age, with a reported prevalence of approximately 8–13%, and represents a major public health concern due to its reproductive, metabolic, and psychological consequences. The syndrome is characterized by hyperandrogenism, polycystic ovarian morphology, chronic anovulation, and metabolic disturbances [ 1 ]. Menstrual dysfunction in women with PCOS commonly presents as irregular, infrequent, or absent menstrual bleeding and is primarily attributable to chronic anovulation. One of the most important long-term consequences of chronic anovulation is prolonged exposure of the endometrium to unopposed estrogen, leading to sustained mitogenic stimulation. This persistent estrogen exposure contributes not only to abnormal endometrial proliferation but also to alterations in endometrial molecular signaling pathways. If left untreated, this hormonal imbalance may result in endometrial hyperplasia and progression to atypical hyperplasia or endometrial adenocarcinoma. Approximately 85% of endometrial cancers are estrogen-dependent, well-differentiated lesions [ 2 ], and women with PCOS have been reported to have an approximately threefold increased risk of developing endometrial cancer compared with the general population. Chronic anovulation in PCOS results in sustained estrogen exposure without cyclical progesterone opposition, which may contribute to altered endometrial molecular signaling. In addition, anti-Müllerian hormone (AMH), which is elevated in PCOS, has also been detected in endometrial tissue, suggesting a potential role in endometrial function [ 2 ]. Despite these findings, the molecular mechanisms underlying endometrial alterations in PCOS remain incompletely understood. Trefoil factor peptides (TFFs) are low-molecular-weight peptides involved in maintaining mucosal integrity and facilitating epithelial repair. Among them, trefoil factor 3 (TFF3) has been implicated in epithelial regeneration and has been shown to be overexpressed in several hormone-related and epithelial malignancies, including breast, prostate, and colorectal cancers [ 3 – 5 ]. TFF3 expression has also been identified in the female reproductive tract, including endocervical glandular epithelium, and has been associated with endometrial cancer [ 6 ]. Experimental studies have demonstrated a close interaction between estrogen signaling and TFF3 expression. Estrogen receptor activation has been shown to upregulate TFF3 expression, while TFF3 may enhance estrogen receptor–mediated transcriptional activity, suggesting that TFF3 is an estrogen-responsive peptide [ 7 ]. Given the chronic unopposed estrogen exposure in PCOS, TFF3 may represent a biologically plausible mediator linking hormonal imbalance to endometrial remodeling. In this context, circulating TFF3 levels may reflect estrogen-responsive pathways and epithelial repair processes associated with endometrial changes in PCOS. To date, no study has specifically evaluated circulating TFF3 levels in women with PCOS or their relationship with menstrual dysfunction. We hypothesized that serum trefoil factor 3 (TFF3) levels would be altered in women with PCOS compared with healthy controls and would be associated with menstrual irregularity. Therefore, this study aimed to evaluate serum TFF3 levels in women with PCOS and to investigate whether TFF3 levels differ according to menstrual patterns, particularly in patients with oligomenorrhea. Materials and Methods Study Design This study was designed as an exploratory cross-sectional study to investigate circulating trefoil factor 3 (TFF3) levels in women with polycystic ovary syndrome (PCOS) and to compare them with those of healthy controls. The primary outcome was the difference in serum TFF3 levels between women with PCOS and healthy controls. Study Population This study included 41 women aged 20–30 years diagnosed with PCOS and 34 age-matched healthy controls who presented to the Department of Obstetrics and Gynecology at Erciyes University. Participants in the PCOS group were recruited from women presenting to the outpatient clinic with complaints of menstrual irregularity and/or hirsutism, who were subsequently diagnosed with PCOS according to the Rotterdam diagnostic criteria [8]. Ovulatory dysfunction was defined and classified in accordance with the FIGO ovulatory disorders classification system [9], and clinical hyperandrogenism was assessed using the modified Ferriman–Gallwey (mFG) score. Polycystic ovarian morphology (PCOM) was defined as the presence of ≥20 follicles measuring 2–9 mm in diameter on transvaginal ultrasonography, in accordance with current guideline recommendations [10]. Women with diabetes mellitus, autoimmune diseases, inflammatory or rheumatologic disorders, known malignancy, or current medication use that could affect hormonal parameters were excluded from the study. The control group consisted of women attending the same outpatient clinic for routine gynecological evaluation, who had regular menstrual cycles, no clinical or biochemical features of PCOS, no chronic medical conditions, and were not using any medications. Ethical Approval The study protocol was approved by the Erciyes University Clinical Research Ethics Committee (approval date: December 7, 2022; approval number: 2022/788). The study was conducted in accordance with the principles of the Declaration of Helsinki. Written informed consent was obtained from all participants prior to enrollment. Biochemical Analysis Venous blood samples (8 mL) were collected from both PCOS patients and control subjects during the early follicular phase (days 2–5) of the menstrual cycle. In patients with irregular cycles, blood sampling was performed following a spontaneous or progesterone-induced withdrawal bleed.Blood samples were collected into serum separator tubes (8 mL, Vacuette) without anticoagulant and centrifuged at 2000 × g for 10 minutes. The separated serum was aliquoted and stored at −80°C until further analysis. Serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol were measured using an electrochemiluminescence immunoassay (ECLIA) on the Cobas e801 analyzer (Roche Diagnostics, Mannheim, Germany). Anti-Müllerian hormone (AMH) concentrations were measured using the same platform. Serum concentrations of trefoil factor 3 (TFF3) were measured using commercially available enzyme-linked immunosorbent assay (ELISA) kits according to the manufacturer’s instructions (Catalog No: RE1622H). All samples were analyzed in duplicate to minimize analytical variability. Statistical Analysis Statistical analyses were performed using IBM SPSS Statistics for Windows, Version 17.0 (IBM Corp., Armonk, NY, USA). Normality of distribution was assessed using the Shapiro–Wilk test. Continuous variables with normal distribution were expressed as mean ± standard deviation (SD), whereas non-normally distributed variables were presented as median (minimum–maximum). Comparisons between groups were performed using the independent samples t-test or the Mann–Whitney U test, as appropriate. Categorical variables were compared using the chi-square test. Correlation analyses were performed using Pearson or Spearman correlation coefficients depending on data distribution and were considered exploratory due to multiple comparisons. Univariate and multivariate logistic regression analyses were performed to identify factors associated with PCOS. Receiver operating characteristic (ROC) curve analysis was used to evaluate the discriminatory performance of serum TFF3 levels, and the area under the curve (AUC) was calculated. The optimal cut-off value was determined using the Youden index. A p-value of <0.05 was considered statistically significant. Results A total of 41 women with PCOS and 34 healthy controls were included in the study. The median age was similar between the groups [23 (18–34) vs. 23 (20–30), p = 0.780]. Body mass index was significantly higher in the PCOS group compared to controls (26.1 ± 6.0 vs. 20.5 ± 3.5, p < 0.001). Serum TFF3 levels were significantly higher in women with PCOS than in controls (1742 ± 582 vs. 1185 ± 704, p < 0.001). Similarly, AMH levels were higher in the PCOS group (5.7 ± 2.5 vs. 3.7 ± 2.0, p = 0.001). Detailed demographic and clinical characteristics are presented in Table 1. Correlation analysis showed no significant relationship between TFF3 and AMH levels (r = −0.046, p = 0.729) (Table 2). Receiver operating characteristic (ROC) analysis demonstrated that the optimal cut-off value of TFF3 for predicting PCOS was 1234, with a sensitivity of 80.5% and a specificity of 61.8% (AUC: 0.728, p < 0.001) (Figure 1). In the analysis of menstrual patterns, the same cut-off value yielded a sensitivity of 86.7% and a specificity of 55.6% (AUC: 0.671, p = 0.012) (Figure 2). Comparison of PCOS patients according to menstrual patterns is shown in Table 3. TFF3 levels did not differ between patients with oligomenorrhea and those with normal menstrual cycles (p = 0.620). AMH levels were slightly higher in patients with oligomenorrhea (p = 0.049). In regression analysis, although several variables were significant in univariate analysis, none remained independently associated with PCOS in the multivariate model (Table 4). Table 1 . Demographic and clinical characteristics of the study population Parameter PCOS (n = 41) Control (n = 34) p-value Age* 23 (18–34) 23 (20–30) 0.780 Menstrual pattern <0.001 – Regular 11 (26.8%) 34 (100%) – Oligomenorrhea 30 (73.2%) – Height† 161.0 ± 5.3 163.4 ± 5.6 0.062 Weight† 67.7 ± 15.1 54.7 ± 9.1 <0.001 Body mass index† 26.1 ± 6.0 20.5 ± 3.5 <0.001 Gravidity* 0 (0–1) 0 (0–1) 0.672 Parity* 0 (0–1) 0 (0–1) 0.272 TFF3† 1742 ± 582 1185 ± 704 <0.001 AMH† 5.7 ± 2.5 3.7 ± 2.0 0.001 Follicle-stimulating hormone† 6.3 ± 2.8 3.9 ± 1.9 <0.001 Luteinizing hormone† 9.7 ± 5.3 8.4 ± 6.2 0.386 Estradiol† 66.3 ± 36.7 48.7 ± 15.3 0.030 Values are presented as mean ± standard deviation or median (minimum–maximum), as appropriate. † indicates normally distributed variables, and * indicates non-normally distributed variables. Table 2. Correlation between TFF3 levels and clinical and laboratory parameters Parameter Correlation coefficient (r) p-value Age* -0.061 0.604 Body mass index† 0.091 0.437 AMH† -0.046 0.729 Ferriman–Gallwey score† -0.179 0.283 Antral follicle count† -0.295 0.120 Correlation coefficients were calculated using Pearson or Spearman tests according to data distribution. † Pearson correlation; * Spearman correlation Table 3. Comparison of clinical and laboratory parameters according to menstrual patterns in women with PCOS Parameter Oligomenorrhea (n = 30) Normal cycle (n = 11) p-value Age* 23 (18–29) 23 (18–34) 0.689 Height† 161.1 ± 5.5 160.8 ± 5.1 0.884 Weight† 65.7 ± 13.0 73.2 ± 19.4 0.159 Body mass index† 25.2 ± 4.5 28.6 ± 8.7 0.117 Gravidity* 0 (0–1) 0 (0–0) 0.752 Parity* 0 (0–1) 0 (0–0) 0.367 TFF3† 1714 ± 492 1818 ± 802 0.620 AMH† 6.4 ± 2.5 4.5 ± 2.1 0.049 Ferriman–Gallwey score† 19.8 ± 6.1 21.7 ± 4.5 0.392 Antral follicle count† 39.0 ± 7.5 36.5 ± 11.1 0.509 Follicle-stimulating hormone† 6.4 ± 3.1 6.3 ± 1.6 0.941 Luteinizing hormone† 9.7 ± 5.5 9.6 ± 4.9 0.956 Estradiol† 75.0 ± 39.3 45.0 ± 16.3 0.020 Values are presented as mean ± standard deviation or median (minimum–maximum), as appropriate. † indicates normally distributed variables, and * indicates non-normally distributed variables. Table 4. Univariate and multivariate logistic regression analyses for factors associated with PCOS Variable OR 95% CI p-value OR 95% CI p-value Univariate Multivariate Age 1.0 0.9–1.2 0.591 2.8 0.1–29.4 0.345 BMI 1.3 1.1–1.5 <0.001 13.2 0.3–54.1 0.172 TFF3 1.0 1.0–1.1 0.001 0.9 0.9–1.1 0.934 AMH 1.5 1.1–1.9 0.004 9.6 0.1–10.6 0.345 FSH 2.0 1.3–2.9 <0.001 6.5 0.1–54.9 0.207 LH 1.0 0.9–1.1 0.380 0.6 0.1–1.9 0.408 Estradiol 1.0 1.0–1.1 0.046 1.3 0.8–2.2 0.234 BMI: body mass index; FSH: follicle-stimulating hormone; LH: luteinizing hormone. Discussion Polycystic ovary syndrome (PCOS) is a complex endocrine and metabolic disorder characterized by chronic anovulation, hyperandrogenism, and polycystic ovarian morphology. Although endometrial dysfunction is recognized as an important long-term consequence of PCOS, the molecular mechanisms underlying these alterations remain incompletely understood. In the present study, serum trefoil factor 3 (TFF3) levels were found to be significantly higher in women with PCOS compared with healthy controls. In contrast, circulating TFF3 concentrations were not associated with menstrual cycle pattern or oligomenorrhea. Trefoil factor proteins are known to play a role in maintaining mucosal integrity and supporting epithelial repair [11]. Previous studies have shown that TFF3 expression varies throughout the menstrual cycle, with significantly higher levels during the proliferative phase compared with the secretory phase, suggesting a role in postmenstrual endometrial regeneration and epithelial proliferation [12]. These findings indicate that TFF3 is closely associated with estrogen-dominant endometrial states and may contribute to endometrial remodeling processes. The interaction between estrogen signaling and TFF3 expression has been demonstrated in both experimental and clinical studies. Estrogen receptor activation has been shown to increase TFF3 expression, while TFF3 may enhance estrogen receptor–mediated transcriptional activity [7,13,14]. Given that chronic anovulation in PCOS results in prolonged exposure of the endometrium to unopposed estrogen, the elevated TFF3 levels observed in this study may reflect increased estrogen-responsive epithelial activity or ongoing endometrial repair. This interpretation is consistent with the known effects of sustained estrogen stimulation on endometrial tissue. The endometrium in women with PCOS is frequently exposed to prolonged unopposed estrogen, which is a recognized risk factor for endometrial hyperplasia and carcinoma. Previous studies have shown increased expression of TFF3 in endometrial carcinoma, particularly in endometrioid subtypes [6,15]. Although the participants in the present study were young and had no clinical evidence of endometrial pathology, the higher serum TFF3 levels may represent an early molecular response to chronic estrogen exposure. However, this possibility remains speculative and would require confirmation in studies including endometrial tissue evaluation. Receiver operating characteristic analysis in the present study showed that TFF3 had a moderate ability to discriminate between women with PCOS and healthy controls (AUC: 0.728). While sensitivity was relatively high, specificity was limited, suggesting that TFF3 alone is unlikely to serve as a reliable diagnostic marker. Instead, it may have a potential role as a supportive or adjunct parameter when interpreted alongside established clinical and biochemical findings. In addition to reproductive features, PCOS is commonly associated with metabolic abnormalities such as increased body mass index, insulin resistance, and hyperinsulinemia [16–19]. In the present study, body mass index was significantly higher in the PCOS group. Previous reports have suggested that circulating TFF3 levels may also be influenced by metabolic factors, including obesity and glucose metabolism [20]. Therefore, the elevated TFF3 levels observed in this study may partly reflect metabolic differences between groups rather than PCOS-related mechanisms alone. This is further supported by the regression analysis, in which none of the variables, including TFF3, remained independently associated with PCOS after adjustment. No significant association was observed between TFF3 levels and menstrual cycle pattern. This finding suggests that circulating TFF3 may not directly reflect cycle-dependent menstrual dysfunction. Instead, it may be more closely related to underlying hormonal or metabolic factors. It is also possible that the relatively small number of patients in the subgroup analysis limited the ability to detect subtle differences. Interestingly, no correlation was found between TFF3 and AMH levels. Given that AMH reflects ovarian follicular reserve, this lack of association suggests that TFF3 may not be directly related to ovarian function. Rather, it may be more closely linked to endometrial or epithelial processes, which is consistent with its known biological role. Limitations This study has several limitations that should be considered. First, the relatively small sample size may have limited statistical power, particularly in subgroup analyses comparing women with oligomenorrhea to those with regular menstrual cycles. Second, the cross-sectional design does not allow for causal inferences regarding the relationship between serum TFF3 levels and hormonal or menstrual abnormalities associated with PCOS. Third, the difference in body mass index between groups may have influenced the observed TFF3 levels. In addition, TFF3 was measured only in serum, and endometrial tissue expression was not assessed; therefore, the relationship between circulating TFF3 levels and local endometrial function remains unclear. Metabolic parameters such as insulin resistance indices were also not directly evaluated, which may have limited the ability to assess the contribution of metabolic factors to TFF3 levels. Furthermore, correlation analyses should be interpreted with caution, as multiple comparisons increase the risk of type I error. Finally, the study population consisted of relatively young women, which may limit the generalizability of the findings to older populations or to patients with established endometrial pathology. Conclusion This study demonstrated that serum trefoil factor 3 (TFF3) levels are significantly elevated in women with polycystic ovary syndrome compared with healthy controls. To our knowledge, this is one of the first studies to investigate circulating TFF3 levels in women with PCOS. The observed increase in TFF3 levels may reflect underlying hormonal and metabolic alterations associated with chronic anovulation and prolonged estrogen exposure. Given the involvement of TFF3 in estrogen-responsive tissues and epithelial regeneration, these findings suggest a possible link between PCOS-related endocrine disturbances and altered TFF3 expression. However, no association was observed between TFF3 levels and menstrual cycle pattern. Further prospective studies with larger sample sizes, including endometrial tissue evaluation and comprehensive metabolic assessment, are needed to clarify the biological role of TFF3 in PCOS and its potential implications for long-term endometrial outcomes. Declarations Statements and Declarations Ethics approval and consent to participate This study was approved by the Erciyes University Clinical Research Ethics Committee (approval date: December 7, 2022; approval number: 2022/788). Written informed consent was obtained from all participants prior to enrollment. Consent for publication Not applicable. Availability of data and materials The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request. Competing interests The authors declare that they have no competing interests. Funding This study was supported by the Erciyes University Scientific Research Projects Coordination Unit (Project No: THD-2023-12537). Authors’ contributions F.C. contributed to the study design and manuscript preparation. E.M.A. and G.B. contributed to the study design. G.Bu. and O.B. were responsible for data collection. M.O.Y. and V.G. performed the data analysis. All authors read and approved the finalmanuscript. Acknowledgements Not applicable. References Joshi A. PCOS stratification for precision diagnostics and treatment. Front Cell Dev Biol. 2024;12:1358755. 10.3389/fcell.2024.1358755 . PMID: 38389707; PMCID: PMC10881805. Pandita P, Wang X, Jones DE, Collins K, Hawkins SM. Unique molecular features in high-risk histology endometrial cancers. Cancers (Basel). 2019;11(11):1665. 10.3390/cancers11111665 . Yamachika T, Werther JL, Bodian C, Babyatsky M, Tatematsu M, Yamamura Y, et al. Intestinal trefoil factor: a marker of poor prognosis in gastric carcinoma. Clin Cancer Res. 2002;8:1092–9. Faith DA, Isaacs WB, Morgan JD, Fedor HL, Hicks JL, Mangold LA, et al. Trefoil factor 3 overexpression in prostatic carcinoma: prognostic importance using tissue microarrays. Prostate. 2004;61:1–13. Babyatsky M, Lin J, Yio X, Chen A, Zhang JY, Zheng Y, et al. Trefoil factor-3 expression in human colon cancer liver metastasis. 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Association of serum trefoil factor 3 and leptin levels with obesity: a case-control study. Cytokine. 2024;181:156690. 10.1016/j.cyto.2024.156690 . Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Reviewers invited by journal 07 Apr, 2026 Editor invited by journal 01 Apr, 2026 Editor assigned by journal 31 Mar, 2026 Submission checks completed at journal 31 Mar, 2026 First submitted to journal 29 Mar, 2026 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. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-9258003","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":619664949,"identity":"682683de-3b8d-4328-aafd-5b2926605a19","order_by":0,"name":"Fulya 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University","correspondingAuthor":false,"prefix":"","firstName":"Ercan","middleName":"Mustafa","lastName":"Aygen","suffix":""},{"id":619664955,"identity":"257e1aea-e761-4467-81bc-59026a8335f2","order_by":6,"name":"Gülden Baskol","email":"","orcid":"","institution":"Erciyes University","correspondingAuthor":false,"prefix":"","firstName":"Gülden","middleName":"","lastName":"Baskol","suffix":""}],"badges":[],"createdAt":"2026-03-29 10:23:17","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-9258003/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-9258003/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":107019460,"identity":"3b3c47f7-e1ef-4dce-bfc6-88902fecc7ef","added_by":"auto","created_at":"2026-04-15 20:59:26","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":45274,"visible":true,"origin":"","legend":"\u003cp\u003eROC curve analysis of TFF3 for discriminating women with PCOS from controls.\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9258003/v1/11643f8a421f805914e5615e.jpeg"},{"id":108005841,"identity":"4ed80208-7ad0-4e51-ba1b-8a1edc9d38f5","added_by":"auto","created_at":"2026-04-28 12:49:14","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":39437,"visible":true,"origin":"","legend":"\u003cp\u003eROC curve analysis of TFF3 according to menstrual pattern in women with PCOS.\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9258003/v1/efa52dfd267c082630ee9b4c.jpeg"},{"id":108008449,"identity":"a437cbb5-9bfb-467c-bdc7-97adb5b10cc5","added_by":"auto","created_at":"2026-04-28 13:06:45","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":364802,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-9258003/v1/d8272055-0678-43e8-a3d0-b0c5fdf5a9ce.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Serum Trefoil Factor 3 Levels in Women with Polycystic Ovary Syndrome: A Cross- Sectional Study","fulltext":[{"header":"Introduction","content":"\u003cp\u003ePolycystic ovary syndrome (PCOS) is one of the most common endocrine disorders affecting women of reproductive age, with a reported prevalence of approximately 8\u0026ndash;13%, and represents a major public health concern due to its reproductive, metabolic, and psychological consequences. The syndrome is characterized by hyperandrogenism, polycystic ovarian morphology, chronic anovulation, and metabolic disturbances [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Menstrual dysfunction in women with PCOS commonly presents as irregular, infrequent, or absent menstrual bleeding and is primarily attributable to chronic anovulation.\u003c/p\u003e \u003cp\u003eOne of the most important long-term consequences of chronic anovulation is prolonged exposure of the endometrium to unopposed estrogen, leading to sustained mitogenic stimulation. This persistent estrogen exposure contributes not only to abnormal endometrial proliferation but also to alterations in endometrial molecular signaling pathways. If left untreated, this hormonal imbalance may result in endometrial hyperplasia and progression to atypical hyperplasia or endometrial adenocarcinoma. Approximately 85% of endometrial cancers are estrogen-dependent, well-differentiated lesions [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e], and women with PCOS have been reported to have an approximately threefold increased risk of developing endometrial cancer compared with the general population.\u003c/p\u003e \u003cp\u003eChronic anovulation in PCOS results in sustained estrogen exposure without cyclical progesterone opposition, which may contribute to altered endometrial molecular signaling. In addition, anti-M\u0026uuml;llerian hormone (AMH), which is elevated in PCOS, has also been detected in endometrial tissue, suggesting a potential role in endometrial function [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Despite these findings, the molecular mechanisms underlying endometrial alterations in PCOS remain incompletely understood.\u003c/p\u003e \u003cp\u003eTrefoil factor peptides (TFFs) are low-molecular-weight peptides involved in maintaining mucosal integrity and facilitating epithelial repair. Among them, trefoil factor 3 (TFF3) has been implicated in epithelial regeneration and has been shown to be overexpressed in several hormone-related and epithelial malignancies, including breast, prostate, and colorectal cancers [\u003cspan additionalcitationids=\"CR4\" citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. TFF3 expression has also been identified in the female reproductive tract, including endocervical glandular epithelium, and has been associated with endometrial cancer [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eExperimental studies have demonstrated a close interaction between estrogen signaling and TFF3 expression. Estrogen receptor activation has been shown to upregulate TFF3 expression, while TFF3 may enhance estrogen receptor\u0026ndash;mediated transcriptional activity, suggesting that TFF3 is an estrogen-responsive peptide [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Given the chronic unopposed estrogen exposure in PCOS, TFF3 may represent a biologically plausible mediator linking hormonal imbalance to endometrial remodeling. In this context, circulating TFF3 levels may reflect estrogen-responsive pathways and epithelial repair processes associated with endometrial changes in PCOS.\u003c/p\u003e \u003cp\u003eTo date, no study has specifically evaluated circulating TFF3 levels in women with PCOS or their relationship with menstrual dysfunction. We hypothesized that serum trefoil factor 3 (TFF3) levels would be altered in women with PCOS compared with healthy controls and would be associated with menstrual irregularity. Therefore, this study aimed to evaluate serum TFF3 levels in women with PCOS and to investigate whether TFF3 levels differ according to menstrual patterns, particularly in patients with oligomenorrhea.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003e\u003cstrong\u003eStudy Design\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was designed as an exploratory cross-sectional study to investigate circulating trefoil factor 3 (TFF3) levels in women with polycystic ovary syndrome (PCOS) and to compare them with those of healthy controls. The primary outcome was the difference in serum TFF3 levels between women with PCOS and healthy controls.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eStudy Population\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study included 41 women aged 20–30 years diagnosed with PCOS and 34 age-matched healthy controls who presented to the Department of Obstetrics and Gynecology at Erciyes University. Participants in the PCOS group were recruited from women presenting to the outpatient clinic with complaints of menstrual irregularity and/or hirsutism, who were subsequently diagnosed with PCOS according to the Rotterdam diagnostic criteria [8]. Ovulatory dysfunction was defined and classified in accordance with the FIGO ovulatory disorders classification system [9], and clinical hyperandrogenism was assessed using the modified Ferriman–Gallwey (mFG) score.\u003c/p\u003e\n\u003cp\u003ePolycystic ovarian morphology (PCOM) was defined as the presence of ≥20 follicles measuring 2–9 mm in diameter on transvaginal ultrasonography, in accordance with current guideline recommendations [10]. Women with diabetes mellitus, autoimmune diseases, inflammatory or rheumatologic disorders, known malignancy, or current medication use that could affect hormonal parameters were excluded from the study.\u003c/p\u003e\n\u003cp\u003eThe control group consisted of women attending the same outpatient clinic for routine gynecological evaluation, who had regular menstrual cycles, no clinical or biochemical features of PCOS, no chronic medical conditions, and were not using any medications.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthical Approval\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe study protocol was approved by the Erciyes University Clinical Research Ethics Committee (approval date: December 7, 2022; approval number: 2022/788). The study was conducted in accordance with the principles of the Declaration of Helsinki. Written informed consent was obtained from all participants prior to enrollment.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eBiochemical Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eVenous blood samples (8 mL) were collected from both PCOS patients and control subjects during the early follicular phase (days 2–5) of the menstrual cycle. In patients with irregular cycles, blood sampling was performed following a spontaneous or progesterone-induced withdrawal bleed.Blood samples were collected into serum separator tubes (8 mL, Vacuette) without anticoagulant and centrifuged at 2000 × g for 10 minutes. The separated serum was aliquoted and stored at −80°C until further analysis.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eSerum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol were measured using an electrochemiluminescence immunoassay (ECLIA) on the Cobas e801 analyzer (Roche Diagnostics, Mannheim, Germany). Anti-Müllerian hormone (AMH) concentrations were measured using the same platform.\u003c/p\u003e\n\u003cp\u003eSerum concentrations of trefoil factor 3 (TFF3) were measured using commercially available enzyme-linked immunosorbent assay (ELISA) kits according to the manufacturer’s instructions (Catalog No: RE1622H).\u003c/p\u003e\n\u003cp\u003eAll samples were analyzed in duplicate to minimize analytical variability.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eStatistical Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eStatistical analyses were performed using IBM SPSS Statistics for Windows, Version 17.0 (IBM Corp., Armonk, NY, USA). Normality of distribution was assessed using the Shapiro–Wilk test. Continuous variables with normal distribution were expressed as mean ± standard deviation (SD), whereas non-normally distributed variables were presented as median (minimum–maximum).\u003c/p\u003e\n\u003cp\u003eComparisons between groups were performed using the independent samples t-test or the Mann–Whitney U test, as appropriate. Categorical variables were compared using the chi-square test. Correlation analyses were performed using Pearson or Spearman correlation coefficients depending on data distribution and were considered exploratory due to multiple comparisons.\u003c/p\u003e\n\u003cp\u003eUnivariate and multivariate logistic regression analyses were performed to identify factors associated with PCOS.\u003c/p\u003e\n\u003cp\u003eReceiver operating characteristic (ROC) curve analysis was used to evaluate the discriminatory performance of serum TFF3 levels, and the area under the curve (AUC) was calculated. The optimal cut-off value was determined using the Youden index. A p-value of \u0026lt;0.05 was considered statistically significant.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eA total of 41 women with PCOS and 34 healthy controls were included in the study. The median age was similar between the groups [23 (18–34) vs. 23 (20–30), p = 0.780]. Body mass index was significantly higher in the PCOS group compared to controls (26.1 ± 6.0 vs. 20.5 ± 3.5, p \u0026lt; 0.001).\u003c/p\u003e\n\u003cp\u003eSerum TFF3 levels were significantly higher in women with PCOS than in controls (1742 ± 582 vs. 1185 ± 704, p \u0026lt; 0.001). Similarly, AMH levels were higher in the PCOS group (5.7 ± 2.5 vs. 3.7 ± 2.0, p = 0.001). Detailed demographic and clinical characteristics are presented in Table 1.\u003c/p\u003e\n\u003cp\u003eCorrelation analysis showed no significant relationship between TFF3 and AMH levels (r = −0.046, p = 0.729) (Table 2).\u003c/p\u003e\n\u003cp\u003eReceiver operating characteristic (ROC) analysis demonstrated that the optimal cut-off value of TFF3 for predicting PCOS was 1234, with a sensitivity of 80.5% and a specificity of 61.8% (AUC: 0.728, p \u0026lt; 0.001) (Figure 1). In the analysis of menstrual patterns, the same cut-off value yielded a sensitivity of 86.7% and a specificity of 55.6% (AUC: 0.671, p = 0.012) (Figure 2).\u003c/p\u003e\n\u003cp\u003eComparison of PCOS patients according to menstrual patterns is shown in Table 3. TFF3 levels did not differ between patients with oligomenorrhea and those with normal menstrual cycles (p = 0.620). AMH levels were slightly higher in patients with oligomenorrhea (p = 0.049).\u003c/p\u003e\n\u003cp\u003eIn regression analysis, although several variables were significant in univariate analysis, none remained independently associated with PCOS in the multivariate model (Table 4).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1\u003c/strong\u003e. Demographic and clinical characteristics of the study population\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"3\" cellpadding=\"0\" width=\"627\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eParameter\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003ePCOS (n = 41)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eControl (n = 34)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003ep-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAge*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e23 (18–34)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e23 (20–30)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.780\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eMenstrual pattern\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e– Regular\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e11 (26.8%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e34 (100%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e– Oligomenorrhea\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e30 (73.2%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e–\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eHeight†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e161.0 ± 5.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e163.4 ± 5.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.062\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eWeight†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e67.7 ± 15.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e54.7 ± 9.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eBody mass index†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e26.1 ± 6.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e20.5 ± 3.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eGravidity*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0 (0–1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0 (0–1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.672\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eParity*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0 (0–1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0 (0–1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.272\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eTFF3†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1742 ± 582\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1185 ± 704\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAMH†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e5.7 ± 2.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.7 ± 2.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eFollicle-stimulating hormone†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e6.3 ± 2.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e3.9 ± 1.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eLuteinizing hormone†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e9.7 ± 5.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e8.4 ± 6.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.386\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eEstradiol†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e66.3 ± 36.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e48.7 ± 15.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.030\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eValues are presented as mean ± standard deviation or median (minimum–maximum), as appropriate. † indicates normally distributed variables, and * indicates non-normally distributed variables.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2.\u0026nbsp;\u003c/strong\u003eCorrelation between TFF3 levels and clinical and laboratory parameters\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"3\" cellpadding=\"0\" width=\"609\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eParameter\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eCorrelation coefficient (r)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003ep-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAge*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.061\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.604\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eBody mass index†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.091\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.437\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAMH†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.046\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.729\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eFerriman–Gallwey score†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.179\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.283\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAntral follicle count†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.295\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.120\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eCorrelation coefficients were calculated using Pearson or Spearman tests according to data distribution. † Pearson correlation; * Spearman correlation\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 3.\u003c/strong\u003e Comparison of clinical and laboratory parameters according to menstrual patterns in women with PCOS\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"3\" cellpadding=\"0\" width=\"611\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eParameter\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eOligomenorrhea (n = 30)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eNormal cycle (n = 11)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003ep-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAge*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e23 (18–29)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e23 (18–34)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.689\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eHeight†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e161.1 ± 5.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e160.8 ± 5.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.884\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eWeight†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e65.7 ± 13.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e73.2 ± 19.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.159\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eBody mass index†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e25.2 ± 4.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e28.6 ± 8.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.117\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eGravidity*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0 (0–1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0 (0–0)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.752\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eParity*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0 (0–1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0 (0–0)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.367\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eTFF3†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1714 ± 492\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1818 ± 802\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.620\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAMH†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e6.4 ± 2.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e4.5 ± 2.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.049\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eFerriman–Gallwey score†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e19.8 ± 6.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e21.7 ± 4.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.392\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAntral follicle count†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e39.0 ± 7.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e36.5 ± 11.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.509\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eFollicle-stimulating hormone†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e6.4 ± 3.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e6.3 ± 1.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.941\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eLuteinizing hormone†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e9.7 ± 5.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e9.6 ± 4.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.956\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eEstradiol†\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e75.0 ± 39.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e45.0 ± 16.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.020\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eValues are presented as mean ± standard deviation or median (minimum–maximum), as appropriate. † indicates normally distributed variables, and * indicates non-normally distributed variables.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 4.\u003c/strong\u003e Univariate and multivariate logistic regression analyses for factors associated with PCOS\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"3\" cellpadding=\"0\" width=\"617\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eVariable\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eOR\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003e95% CI\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003ep-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eOR\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003e95% CI\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003ep-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eUnivariate\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eMultivariate\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAge\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.9–1.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.591\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.1–29.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.345\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eBMI\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.1–1.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e13.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.3–54.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.172\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eTFF3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.0–1.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.9–1.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.934\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAMH\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.1–1.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.004\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e9.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.1–10.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.345\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eFSH\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e2.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.3–2.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e6.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.1–54.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.207\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eLH\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.9–1.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.380\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.1–1.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.408\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eEstradiol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.0–1.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.046\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e1.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.8–2.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.234\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eBMI: body mass index; FSH: follicle-stimulating hormone; LH: luteinizing hormone.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003ePolycystic ovary syndrome (PCOS) is a complex endocrine and metabolic disorder characterized by chronic anovulation, hyperandrogenism, and polycystic ovarian morphology. Although endometrial dysfunction is recognized as an important long-term consequence of PCOS, the molecular mechanisms underlying these alterations remain incompletely understood. In the present study, serum trefoil factor 3 (TFF3) levels were found to be significantly higher in women with PCOS compared with healthy controls. In contrast, circulating TFF3 concentrations were not associated with menstrual cycle pattern or oligomenorrhea.\u003c/p\u003e\n\u003cp\u003eTrefoil factor proteins are known to play a role in maintaining mucosal integrity and supporting epithelial repair [11]. Previous studies have shown that TFF3 expression varies throughout the menstrual cycle, with significantly higher levels during the proliferative phase compared with the secretory phase, suggesting a role in postmenstrual endometrial regeneration and epithelial proliferation [12]. These findings indicate that TFF3 is closely associated with estrogen-dominant endometrial states and may contribute to endometrial remodeling processes.\u003c/p\u003e\n\u003cp\u003eThe interaction between estrogen signaling and TFF3 expression has been demonstrated in both experimental and clinical studies. Estrogen receptor activation has been shown to increase TFF3 expression, while TFF3 may enhance estrogen receptor–mediated transcriptional activity [7,13,14]. Given that chronic anovulation in PCOS results in prolonged exposure of the endometrium to unopposed estrogen, the elevated TFF3 levels observed in this study may reflect increased estrogen-responsive epithelial activity or ongoing endometrial repair. This interpretation is consistent with the known effects of sustained estrogen stimulation on endometrial tissue.\u003c/p\u003e\n\u003cp\u003eThe endometrium in women with PCOS is frequently exposed to prolonged unopposed estrogen, which is a recognized risk factor for endometrial hyperplasia and carcinoma. Previous studies have shown increased expression of TFF3 in endometrial carcinoma, particularly in endometrioid subtypes [6,15]. Although the participants in the present study were young and had no clinical evidence of endometrial pathology, the higher serum TFF3 levels may represent an early molecular response to chronic estrogen exposure. However, this possibility remains speculative and would require confirmation in studies including endometrial tissue evaluation.\u003c/p\u003e\n\u003cp\u003eReceiver operating characteristic analysis in the present study showed that TFF3 had a moderate ability to discriminate between women with PCOS and healthy controls (AUC: 0.728). While sensitivity was relatively high, specificity was limited, suggesting that TFF3 alone is unlikely to serve as a reliable diagnostic marker. Instead, it may have a potential role as a supportive or adjunct parameter when interpreted alongside established clinical and biochemical findings.\u003c/p\u003e\n\u003cp\u003eIn addition to reproductive features, PCOS is commonly associated with metabolic abnormalities such as increased body mass index, insulin resistance, and hyperinsulinemia [16–19]. In the present study, body mass index was significantly higher in the PCOS group. Previous reports have suggested that circulating TFF3 levels may also be influenced by metabolic factors, including obesity and glucose metabolism [20]. Therefore, the elevated TFF3 levels observed in this study may partly reflect metabolic differences between groups rather than PCOS-related mechanisms alone. This is further supported by the regression analysis, in which none of the variables, including TFF3, remained independently associated with PCOS after adjustment.\u003c/p\u003e\n\u003cp\u003eNo significant association was observed between TFF3 levels and menstrual cycle pattern. This finding suggests that circulating TFF3 may not directly reflect cycle-dependent menstrual dysfunction. Instead, it may be more closely related to underlying hormonal or metabolic factors. It is also possible that the relatively small number of patients in the subgroup analysis limited the ability to detect subtle differences.\u003c/p\u003e\n\u003cp\u003eInterestingly, no correlation was found between TFF3 and AMH levels. Given that AMH reflects ovarian follicular reserve, this lack of association suggests that TFF3 may not be directly related to ovarian function. Rather, it may be more closely linked to endometrial or epithelial processes, which is consistent with its known biological role.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eLimitations\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study has several limitations that should be considered. First, the relatively small sample size may have limited statistical power, particularly in subgroup analyses comparing women with oligomenorrhea to those with regular menstrual cycles. Second, the cross-sectional design does not allow for causal inferences regarding the relationship between serum TFF3 levels and hormonal or menstrual abnormalities associated with PCOS. Third, the difference in body mass index between groups may have influenced the observed TFF3 levels.\u003c/p\u003e\n\u003cp\u003eIn addition, TFF3 was measured only in serum, and endometrial tissue expression was not assessed; therefore, the relationship between circulating TFF3 levels and local endometrial function remains unclear. Metabolic parameters such as insulin resistance indices were also not directly evaluated, which may have limited the ability to assess the contribution of metabolic factors to TFF3 levels. Furthermore, correlation analyses should be interpreted with caution, as multiple comparisons increase the risk of type I error.\u003c/p\u003e\n\u003cp\u003eFinally, the study population consisted of relatively young women, which may limit the generalizability of the findings to older populations or to patients with established endometrial pathology.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis study demonstrated that serum trefoil factor 3 (TFF3) levels are significantly elevated in women with polycystic ovary syndrome compared with healthy controls. To our knowledge, this is one of the first studies to investigate circulating TFF3 levels in women with PCOS.\u003c/p\u003e\n\u003cp\u003eThe observed increase in TFF3 levels may reflect underlying hormonal and metabolic alterations associated with chronic anovulation and prolonged estrogen exposure. Given the involvement of TFF3 in estrogen-responsive tissues and epithelial regeneration, these findings suggest a possible link between PCOS-related endocrine disturbances and altered TFF3 expression. However, no association was observed between TFF3 levels and menstrual cycle pattern.\u003c/p\u003e\n\u003cp\u003eFurther prospective studies with larger sample sizes, including endometrial tissue evaluation and comprehensive metabolic assessment, are needed to clarify the biological role of TFF3 in PCOS and its potential implications for long-term endometrial outcomes.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eStatements and Declarations\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was approved by the Erciyes University Clinical Research Ethics Committee (approval date: December 7, 2022; approval number: 2022/788). Written informed consent was obtained from all participants prior to enrollment.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was supported by the Erciyes University Scientific Research Projects Coordination Unit (Project No: THD-2023-12537).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors’ contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eF.C. contributed to the study design and manuscript preparation. E.M.A. and G.B. contributed to the study design. G.Bu. and O.B. were responsible for data collection. M.O.Y. and V.G. performed the data analysis. All authors read and approved the finalmanuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eJoshi A. PCOS stratification for precision diagnostics and treatment. Front Cell Dev Biol. 2024;12:1358755. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.3389/fcell.2024.1358755\u003c/span\u003e\u003cspan address=\"10.3389/fcell.2024.1358755\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e. PMID: 38389707; PMCID: PMC10881805.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePandita P, Wang X, Jones DE, Collins K, Hawkins SM. Unique molecular features in high-risk histology endometrial cancers. 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Endometrial abnormality in women with polycystic ovary syndrome. Reprod Sci. 2012;19(7):674\u0026ndash;83. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1177/1933719111430993\u003c/span\u003e\u003cspan address=\"10.1177/1933719111430993\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePathare ADS, Hinduja I, Mahadik RC. Basic aspects of endometrial receptivity in PCOS patients. Mol Biol Rep. 2022;49(2):1519\u0026ndash;28. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1007/s11033-021-06976-9\u003c/span\u003e\u003cspan address=\"10.1007/s11033-021-06976-9\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePalomba S, Piltonen TT, Giudice LC. Endometrial function in women with polycystic ovary syndrome: a comprehensive review. 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Cytokine. 2024;181:156690. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1016/j.cyto.2024.156690\u003c/span\u003e\u003cspan address=\"10.1016/j.cyto.2024.156690\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\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":"bmc-womens-health","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"bmwh","sideBox":"Learn more about [BMC Women's Health](http://bmcwomenshealth.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/bmwh/default.aspx","title":"BMC Women's Health","twitterHandle":"","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Polycystic ovary syndrome, Trefoil factor 3, Endometrium, Estrogen, AMH","lastPublishedDoi":"10.21203/rs.3.rs-9258003/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-9258003/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e \u003cb\u003eBackground\u003c/b\u003e \u003c/p\u003e \u003cp\u003ePolycystic ovary syndrome (PCOS) is associated with chronic anovulation and prolonged unopposed estrogen exposure, which may contribute to endometrial dysfunction. Trefoil factor 3 (TFF3) is an estrogen-responsive peptide involved in epithelial repair and mucosal integrity. However, data on circulating TFF3 levels in women with PCOS are limited.\u003c/p\u003e \u003cp\u003e \u003cb\u003eMethods\u003c/b\u003e \u003c/p\u003e \u003cp\u003eThis exploratory cross-sectional study included 41 women with PCOS and 34 age-matched healthy controls. Serum TFF3 levels were measured and compared between groups. The association between TFF3 levels and menstrual cycle patterns was also evaluated. Receiver operating characteristic (ROC) analysis was performed to assess the discriminatory performance of TFF3 for PCOS.\u003c/p\u003e \u003cp\u003e \u003cb\u003eResults\u003c/b\u003e \u003c/p\u003e \u003cp\u003eSerum TFF3 levels were significantly higher in women with PCOS compared with controls (1742\u0026thinsp;\u0026plusmn;\u0026thinsp;582 vs. 1185\u0026thinsp;\u0026plusmn;\u0026thinsp;704, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001). TFF3 levels were not associated with menstrual cycle pattern or oligomenorrhea. ROC analysis showed that TFF3 had a moderate ability to discriminate PCOS (AUC: 0.728), with a sensitivity of 80.5% and a specificity of 61.8% at the optimal cut-off value. No independent associations were identified in multivariate regression analysis.\u003c/p\u003e \u003cp\u003e \u003cb\u003eConclusion\u003c/b\u003e \u003c/p\u003e \u003cp\u003eSerum TFF3 levels are elevated in women with PCOS, suggesting a possible link between TFF3 and PCOS-related hormonal and endometrial changes. However, the lack of association with menstrual patterns and the moderate diagnostic performance indicate that TFF3 alone is unlikely to serve as a reliable biomarker. Further studies are needed to clarify its biological and clinical significance.\u003c/p\u003e","manuscriptTitle":"Serum Trefoil Factor 3 Levels in Women with Polycystic Ovary Syndrome: A Cross- Sectional Study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-04-15 20:59:22","doi":"10.21203/rs.3.rs-9258003/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewersInvited","content":"","date":"2026-04-08T01:49:12+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2026-04-01T06:11:22+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-03-31T06:27:16+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-03-31T06:26:24+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Women's Health","date":"2026-03-29T10:18:24+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-womens-health","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"bmwh","sideBox":"Learn more about [BMC Women's Health](http://bmcwomenshealth.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/bmwh/default.aspx","title":"BMC Women's Health","twitterHandle":"","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"6b41b5fd-033f-4b98-b3a4-6cb1798b492d","owner":[],"postedDate":"April 15th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-04-15T20:59:22+00:00","versionOfRecord":[],"versionCreatedAt":"2026-04-15 20:59:22","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-9258003","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-9258003","identity":"rs-9258003","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}