Gonadotropin-Induced Molecular Alterations in Experimental Endometriosis: Downregulation of Tumor Suppressor Genes and Inflammatory Activation

Background Endometriosis is a hormonally responsive inflammatory disease with a recognized association with certain ovarian cancer subtypes. Gonadotropins are widely used in assisted reproductive technologies; however, their direct molecular effects on endometriotic tissue remain insufficiently characterized.

This study aimed to investigate the effects of gonadotropin treatment on molecular markers potentially associated with early carcinogenesis-related processes in endometriotic lesions, using a surgically induced rat model.

Twenty-two female Wistar Albino rats underwent surgical induction of endometriosis via autologous peritoneal implantation. The animals were randomly assigned to two groups: a control group receiving saline and a treatment group receiving gonadotropin (2 IU/kg/day) for 14 days. Following treatment, endometriotic lesions were excised and analyzed histologically and immunohistochemically for phosphatase and tensin homolog (PTEN), tumor protein 53 (TP53), and tumor necrosis factor-alpha (TNF-α) expression. Statistical analyses were performed using the Mann–Whitney U test based on non-parametric data distribution.

Histopathological evaluation revealed no significant morphological differences between the groups. However, quan- titative immunohistochemical analysis demonstrated that gonadotropin-treated rats exhibited markedly decreased PTEN (p < 0.001) and TP53 (p = 0.001) expression, alongside a significant increase in TNF-α expression (p = 0.035) compared with controls (Table 1, Figs.  5, 6). These alterations reflect early molecular changes involving tumor suppressor gene downregula- tion and pro-inflammatory cytokine upregulation.

These findings suggest that gonadotropin exposure may induce early molecular alterations in endometriotic tis- sue. Further translational and clinical studies are warranted to validate these findings in human populations.

Endometriosis · Gonadotropins · PTEN · TP53 · TNF-α 1 Introduction Endometriosis is a chronic, estrogen-dependent inflamma- tory disorder characterized by the presence of endometrial- like tissue outside the uterine cavity [1 –3]. Affecting up to 15% of women of reproductive age, it represents a major cause of chronic pelvic pain, dysmenorrhea, dyspareu - nia, and infertility [4 , 5]. Although histologically benign, endometriosis exhibits several malignant-like characteris- tics—including local invasion, hormone responsiveness, and recurrence—and is associated with an increased risk of certain ovarian cancers, particularly the endometrioid and clear cell subtypes [6, 7]. * Erol Karakaş [email protected] 1 Department of Obstetrics and Gynecology, Op. Dr. Erol Karakaş Gynecology And Obstetrics Clinic, Alpaslan Mah. Aşık Veysel Boulevard Road Number: 21/A Melikgazi, 38140 Kayseri, Turkey 2 Department of Obstetrics and Gynecology, Faculty of Medicine, Erciyes, University, Kayseri, Turkey 3 Department of Biochemistry, Faculty of Veterinary Medicine, Erciyes University, Kayseri, Turkey 4 Department of Pathology, Faculty of Medicine, Erciyes, University, Kayseri, Turkey 5 Department of Histology and Embryology, Faculty of Medicine, Erciyes, University, Kayseri, Turkey 1133Bratislava Medical Journal (2026) 127:1132–1141 Approximately one-third of women with endometriosis experience infertility [8 ]. Assisted reproductive technolo- gies (ART), such as ovulation induction with gonadotropins, are commonly employed to address endometriosis-related subfertility [9 ]. However, growing concern has emerged regarding the potential tumorigenic effects of gonadotro- pins on hormonally sensitive tissues [10–13]. Gonadotropins stimulate ovarian follicular growth and elevate circulating estrogen levels, potentially exacerbating proliferative and inflammatory activity within ectopic endometrial implants [14–16]. While epidemiological findings remain inconclu- sive, mechanistic evidence elucidating the direct molecular effects of gonadotropin exposure on endometriotic tissues is notably scarce. This study was designed to determine whether gonado- tropin administration induces molecular alterations relevant to pathways implicated in malignant transformation in sur - gically induced endometriotic lesions in rats. Specifically, we examined the expression patterns of the tumor suppres- sor genes phosphatase and tensin homolog (PTEN) and tumor protein 53 (TP53), along with the pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-α), which are key regulators implicated in tumorigenesis [17– 19]. The findings of this study may provide mechanistic insights into the molecular effects of gonadotropin exposure in endome- triotic tissue and contribute to ongoing discussions regard- ing treatment safety [20– 22]. These markers were selected due to their established involvement in endometriosis-asso- ciated carcinogenesis and inflammation-driven malignant transformation. 2 Materıals and Methods 2.1 Study Design and Animals This experimental study was conducted at the Experimental Research Center of Erciyes University following approval by the Institutional Animal Ethics Committee (Protocol No: 17/085). Twenty-two female Wistar Albino rats (8 weeks old, 180–260 g) were used. The animals were housed indi- vidually under standard laboratory conditions (12-h light/ dark cycle, 22 ± 2 °C, and 55–60% humidity) with ad libitum access to standard chow and water. 2.2 Induction of Endometriosis A surgical endometriosis model was established according to the method described by Vernon and Wilson [ 1] and is illustrated in Fig. 1. Under general anesthesia, 1 cm segment of the left uterine horn was excised, opened longitudinally, and sutured onto the peritoneal surface with the endome- trial side facing inward [1 ]. Cefazolin sodium (50 mg/kg) was administered preoperatively and for three consecutive Fig. 1 Rat Endometriosis Model Setup. A schematic representation of the surgically induced endometriosis model in Wistar-Albino rats. Endo- metrial tissue was excised from the uterine horn and sutured onto the peritoneal surface under anesthesia. The model successfully simulates ectopic endometrial implantation sites after four weeks of recovery 1134 Bratislava Medical Journal (2026) 127:1132–1141 days postoperatively to prevent infection. After four weeks, endometriotic implants were confirmed in all animals. The peritoneal autologous transplantation model was selected because it allows controlled evaluation of early molecular changes in ectopic endometrial tissue without confound - ing cystic degeneration, making it suitable for mechanistic assessment. 2.3 Experimental Groups Rats were randomly allocated into two groups (n = 11 per group): • Control group Received subcutaneous physiological saline at an injection volume equivalent to that of the treatment group for 14 days. • Gonadotropin group Received subcutaneous recombinant follicle-stimulating hormone (rFSH; Gonal-F®, Merck Serono) at a dose of 2 IU/kg/day for 14 days. This for - mulation provides FSH activity only. [2]. The gonadotropin dose of 2 IU/kg/day was derived from body surface area (BSA) conversion of standard human clinical doses (150–225 IU/day) used for controlled ovarian stimulation. Based on the Food and Drug Administration (FDA)-recommended formula and the method of Reagan- Shaw et al., the human equivalent dose (HED) was translated to the rat model using Km factors (human: 37; rat: 6). This conversion yielded an approximate dose of 2 IU/kg/day in rats, representing a moderate ovarian stimulation regimen comparable to clinical practice [23]. Characteristic endometriotic lesions observed after four weeks are shown in Fig.  2. All 22 rats completed the study protocol without mortality or exclusion during the modeling or treatment phases. All animals were included in the final histological and immunohistochemical analyses. 2.4 Tissue Collection and Histopathology At the end of the treatment period, rats were euthanized, and endometriotic implants were excised. Tissues were fixed in 10% neutral buffered formalin, embedded in paraffin, and sectioned for hematoxylin and eosin (H&E) and Masson’s trichrome staining. The epithelial integrity of endometrial lesions was assessed and scored using standard histopatho- logical criteria. 2.5 Immunohistochemistry Immunohistochemical staining for PTEN, TP53, and TNF-α was performed using the standard avidin–biotin–peroxidase complex method. The primary antibodies were as follows: anti-PTEN (rabbit polyclonal, Abcam, ab32199, 1:200), anti-TP53 (mouse monoclonal, Santa Cruz Biotechnol- ogy, sc-47698, 1:100), and anti-TNF-α (rabbit polyclonal, Abcam, ab6671, 1:100). Rat liver served as the positive con- trol for PTEN and TP53, and rat spleen for TNF-α. Negative controls were prepared by omitting the primary antibody. Antigen retrieval, blocking, and antibody incubation steps were followed by diaminobenzidine (DAB) chromogen visu- alization and hematoxylin counterstaining. Staining inten- sity was quantified using ImageJ software in ten randomly selected high-power fields (HPFs) per sample. Thresholds Fig. 2 Gross view of induced endometriotic lesions. Rep- resentative gross images of ectopic endometriotic lesions observed on the peritoneal surface four weeks after surgi- cal induction. Images were proportionally cropped to main- tain consistent magnification across panels. Endometriotic lesions appear as well-defined, vascularized nodules and are indicated by arrows and circles. A scale bar is included to dem- onstrate lesion size. Comparable gross lesion morphology is observed between the control and gonadotropin-treated groups 1135Bratislava Medical Journal (2026) 127:1132–1141 for positive staining were standardized across all images. All scorers were blinded to treatment allocation. Although inter-observer variability was not statistically analyzed, all slides were independently evaluated by two histopathologists and finalized by consensus. 2.6 Statistical Analysis All statistical analyses were performed using SPSS ver - sion 22. Data normality was assessed using the Shap- iro–Wilk test. Group comparisons were performed using the Mann–Whitney U test based on non-parametric data distribution. A p -value < 0.05 was considered statistically significant (Figs.  3, 4, 5, 6, 7, 8, 9). 3 Results 3.1 Histological Evaluation Histological scoring based on epithelial integrity showed no statistically significant difference between the control and gonadotropin groups (p = 0.127), indicating comparable lesion formation across both groups [24]. The distribution of histological epithelial integrity scores is shown in Fig.  8. Although no statistically significant difference was observed, graphical presentation allows clearer visualization of score variability between groups. H&E staining demonstrated inflammatory infiltration, edema, and capillary dilation in both groups (Fig.  3), while Masson’s trichrome staining revealed collagen deposition in both the control and gonadotropin-treated groups. To objectively evaluate fibrosis, we quantified the percentage of collagen-positive area using *ImageJ* software across 10 randomly selected high-power fields per lesion, for all animals (n = 11 per group). The median fibrosis area was **21.3% (IQR: 17.0–25.7%)** in the control group and **18.6% (IQR: 14.2–22.4%)** in the gonadotropin-treated group, with **no statistically significant difference** (p = 0.43, Mann–Whitney U test). These results support the initial histological impression that gonadotropin treatment did not significantly affect fibrotic remodeling of ectopic lesions (Fig.  4). Approximate gross lesion size comparison is presented in Fig. 9. Lesion dimensions were obtained dur- ing macroscopic excision and are provided as supportive, semi-quantitative data. Lesion volume was not quantitatively measured; however, gross inspection did not reveal apparent differences between groups. 3.2 Immunohistochemical Findings Immunohistochemical analysis revealed significant differ - ences in the expression levels of PTEN, TP53, and TNF-α between the control and gonadotropin-treated groups (Table  1). Representative immunohistochemical staining Control H&E * * x20 x40 Gonadotropin x20 x40 ** * * * Epithelium * Mononuclear cell infiltration S S S Stroma Fig. 3 Histological Features of Ectopic Lesions (H&E Staining). Representative hematoxylin–eosin (H&E) stained sections of ectopic endometrial lesions from the control and gonadotropin-treated groups. Both groups show glandular structures surrounded by stroma, with visible inflammation, stromal edema, and increased capillary density. Magnification: × 400. Scale bars (100  μm and 200  μm) are shown in the images 1136 Bratislava Medical Journal (2026) 127:1132–1141 patterns of PTEN and TP53 in both groups are shown in Fig.  6. Quantitative ImageJ-based analysis demonstrated that PTEN and TP53 expression was significantly reduced in the gonadotropin-treated group, whereas TNF-α expres- sion was significantly increased compared with controls. These findings indicate that gonadotropin administration is associated with downregulation of tumor suppressor proteins (PTEN and TP53) and upregulation of the pro-inflammatory cytokine TNF-α, suggesting enhanced inflammatory and proliferative activity within ectopic endometriotic tissue. All 22 rats completed the experimental protocol without mortality or exclusion, and molecular analyses included all samples (n = 11 per group). 4 Discussion Infertility affects approximately one in seven couples world- wide, and the increasing reliance on assisted reproductive technologies (ART) has raised concerns regarding their long-term safety [25, 26]. Among these interventions, gon- adotropins are widely used for ovulation induction [27]. However, accumulating evidence suggests a potential onco- genic risk, particularly in hormonally responsive conditions such as endometriosis [28–30]. In this study, gonadotropin administration in a rat model of surgically induced endometriosis resulted in significant downregulation of the tumor suppressor proteins PTEN and Fig. 4 Quantitative assessment of fibrosis in ectopic endome- triotic lesions. Representative Masson’s trichrome–stained sections of ectopic endometri- otic lesions from the control and gonadotropin-treated groups. Collagen fibers are stained blue, while cytoplasm is stained red. Images are shown at × 400 mag- nification; scale bar = 100 μm. Quantitative analysis of fibrosis expressed as collagen-positive area percentage, measured using ImageJ software. Fibrosis was analyzed in 10 randomly selected high-power fields per animal (n = 11 per group). Data are presented as box- and-whisker plots showing median and interquartile range. No statistically significant difference in fibrosis area was observed between the control and gonadotropin-treated groups (Mann–Whitney U test, p = 0.43) 1137Bratislava Medical Journal (2026) 127:1132–1141 TP53, accompanied by increased expression of the pro- inflammatory cytokine TNF-α [18– 20]. These molecular alterations are consistent with pathways implicated in early neoplastic transformation, which have been described in association with endometriosis-related malignancies. PTEN and TP53 play pivotal roles in maintaining genomic stability and regulating apoptosis. Their suppres- sion may facilitate uncontrolled cellular proliferation and impair DNA repair, both hallmarks of carcinogenesis. Con- currently, elevated TNF-α levels indicate an inflammatory microenvironment that may favor tumor initiation and pro- gression. TNF-α is known to promote angiogenesis, immune evasion, and epithelial-to-mesenchymal transition (EMT), all of which are central to malignant transformation [21–24, 31–34]. These results align with previous reports describing similar molecular alterations in atypical endometriosis and endometriosis-associated ovarian cancer. The present study provides experimental evidence suggesting that exogenous hormonal stimulation via gonadotropins may influence molecular pathways that have been implicated in endome- triosis-related carcinogenesis [14, 35, 36]. A methodological consideration is the use of saline as a control, while the gonadotropin formulation may contain stabilizing excipients. Future experiments should include a vehicle control prepared with the identical solvent composi- tion to exclude potential non-specific effects. Furthermore, the present study focused exclusively on TNF-α as a representative inflammatory cytokine. Expand- ing future analyses to include IL-6, IL-8, and IL-1β would provide a more comprehensive understanding of the cytokine TNF-α ControlGonadotropin Fig. 5 TNF-α Immunohistochemical Expression. Immunohistochemi- cal staining for TNF-α in ectopic lesions. Gonadotropin-treated rats exhibit more intense cytoplasmic TNF-α expression in both epithelial and stromal compartments compared to controls. Increased staining correlates with enhanced inflammatory activity. Magnification: × 400. Scale bars (100 μm) are shown in the images Fig. 6 Expression of PTEN and TP53 Tumor Suppressor Proteins. Immunohistochemi- cal staining of PTEN and TP53 in endometriotic lesions. Both markers show reduced nuclear expression in the gonadotro- pin-treated group relative to controls, indicating suppression of tumor suppressor pathways. Magnification: × 400. Scale bars (100 μm) are shown in the images P53 ControlGonadotropin PTEN 1138 Bratislava Medical Journal (2026) 127:1132–1141 network implicated in endometriosis-associated inflamma- tion and malignancy. As illustrated schematically in Fig.  7, gonadotropin stimulation may act through receptor-mediated pathways, potentially involving activation of the PI3K/AKT and NF-κB signaling cascades, which may contribute to downregulation of PTEN and TP53 and upregulation of TNF-α in endometri- otic tissue. Although the underlying signaling mechanisms were not directly examined, it is biologically plausible that gonadotropin stimulation acts through receptor-mediated pathways. Follicle-stimulating hormone (FSH) and luteiniz- ing hormone (LH) bind to their respective G protein–coupled receptors (FSHR and LHR), activating downstream cascades such as the cyclic AMP/protein kinase A (cAMP–PKA) and phosphoinositide 3-kinase/protein kinase B (PI3K–AKT) pathways [37–40]. Activation of PI3K–AKT signaling has been shown to suppress PTEN activity, which may explain the reduced PTEN expression observed in the present study [41]. Additionally, TP53 function may be indirectly modu- lated through these pathways, thereby contributing to a pro- proliferative cellular milieu [42]. Gonadotropin-induced inflammation may also involve the nuclear factor kappa B (NF-κB) pathway, a key regu- lator of inflammatory gene expression including TNF-α [43]. NF-κB activation in response to hormonal stimula- tion has been demonstrated in endometrial and ovarian tissues [44], with downstream effects on cytokine secre- tion, angiogenesis, immune modulation, and EMT—all hallmarks of early oncogenesis [45, 46]. These mecha- nistic hypotheses are consistent with the molecular pro- file observed in our gonadotropin-treated group—spe- cifically, concurrent suppression of PTEN and TP53 and Fig. 7 Hypothetical schematic representation of potential molecular pathways affected by gonadotropin treatment in endometriotic tissue. This schematic illustrates a hypothetical, literature-based framework summarizing potential signaling pathways that may be involved in gonadotropin-associated molecular alterations in endometriotic tis- sue. Gonadotropin stimulation may activate the follicle-stimulating hormone receptor (FSHR), potentially leading to activation of the PI3K/AKT signaling pathway and reduced expression of the tumor suppressor proteins PTEN and TP53. In parallel, gonadotropin signal- ing may contribute to activation of the NF-κB pathway, resulting in increased expression of the pro-inflammatory cytokine TNF-α. This diagram is intended for conceptual purposes only and does not repre- sent a mechanistic pathway directly demonstrated by the experimental data presented in this study Fig. 8 Distribution of histological epithelial integrity scores in con- trol and gonadotropin-treated groups presented as box-and-whisker plots (median and interquartile range) Fig. 9 Approximate gross lesion size comparison between groups based on macroscopic measurements obtained during tissue excision. Measurements are presented as semi-quantitative values due to tech- nical limitations 1139Bratislava Medical Journal (2026) 127:1132–1141 upregulation of TNF-α. Future studies incorporating gene expression profiling, pathway-specific inhibitors, and longer observation periods are warranted to further elucidate these signaling relationships. From a translational perspective, these findings under - score the need for further investigation into the molecu- lar effects of gonadotropin exposure before any defini- tive clinical conclusions can be drawn. Considering the established association between PTEN and TP53 altera- tions and malignant transformation, the potential clinical implications of these molecular findings warrant further investigation in well-designed human studies before any preventive strategies can be recommended [36, 47]. Addi- tional clinical studies are required to determine whether these molecular alterations translate into increased cancer incidence among patients undergoing ovulation induction [48–51]. Until such data become available, individualized treatment planning and careful risk–benefit assessment are essential, particularly for women with severe, recur - rent, or long-standing endometriosis [12, 18]. Although fibrosis was initially evaluated qualitatively, subsequent quantitative analysis using *ImageJ* con- firmed that gonadotropin exposure did not significantly alter collagen deposition in ectopic endometriotic lesions. This suggests that the molecular alterations induced by gonadotropin treatment (e.g., TNF-α upregulation, PTEN and TP53 downregulation) occur independently of changes in fibrotic tissue remodeling within the short experimental window. The absence of significant histo- pathological changes despite marked molecular altera- tions may be attributable to the relatively short exposure period. Alternatively, gonadotropin-induced stress signal - ing could provoke transient inflammatory or pro-neoplas- tic molecular events preceding detectable structural alter - ations. These possibilities warrant further investigation. 4.1 Limitations This study has several limitations. The relatively short treatment and observation period (14 days) may explain the absence of overt morphological changes despite significant molecular alterations. The sample size was relatively small, which may limit the statistical power and generalizability of the findings. Functional assays such as Ki-67 or Caspase-3 immunostaining were not performed, and molecular analy - sis was limited to three markers: PTEN, TP53, and TNF-α. Broader cytokine panels (e.g., IL-6, IL-8, IL-1β), transcrip- tomic analyses, and extended follow-up durations should be considered in future studies. Additionally, the eutopic endo- metrium was not evaluated; thus, systemic effects of gon- adotropins cannot be excluded. The findings are restricted to ectopic lesions. The absence of a positive control group and a vehicle control group using the same excipients as the gonadotropin formulation may also limit the ability to distinguish specific hormonal effects from potential solvent- related influences. 5 Conclusion This study provides experimental evidence that gonadotro- pin administration is associated with early molecular alter - ations in surgically induced endometriotic tissue. Specifi- cally, decreased expression of PTEN and TP53, along with increased expression of the pro-inflammatory cytokine TNF- α, was observed in ectopic lesions in rats. These molecular changes may reflect a shift toward a pro-inflammatory and pro-proliferative microenvironment in hormonally respon- sive tissues. While these findings offer mechanistic insights into the potential effects of gonadotropins on endometriotic Table 1 Semi-quantitative histological scores and ImageJ-based quantitative immunohistochemical measurements of endometriotic tissues Data are presented as median (interquartile range, IQR). Histological epithelial integrity was evaluated using a semi-quantitative scoring system on a 0–3 scale. PTEN, TP53, and TNF-α values represent ImageJ- derived quantitative measurements of staining intensity and/or positive staining area obtained from digital image analysis. For each marker, multiple microscopic high-power fields were analyzed per lesion. n indi- cates the total number of microscopic fields evaluated. Statistical comparisons were performed using the Mann–Whitney U test. p < 0.05 was considered statistically significant. Superscript letters (a, b) indicate statistically significant differences between groups Control Gonadotropin p SCORE (Histological epithelial integrity score) (n = 30) 1.00 (0.00–2.00)a 2.00 (1.00–2.25)a 0.127 PTEN (n = 60) 77.69 (70.30–88.38)a 64.89 (60.60–73.16)b 0.001* TP53 (n = 60) 63.20 (59.01–69.80)a 57.13 (55.48–60.58)b 0.001* TNF-α (n = 60) 87.79 (76.67–103.54)a 95.13 (85.79–101.31)b 0.035* 1140 Bratislava Medical Journal (2026) 127:1132–1141 lesions, they are based on a short-term experimental model and should be interpreted with caution. No morphological evidence of malignant transformation was observed, and the current results do not establish a causal link to cancer development. Therefore, the results of this study should be considered preliminary and hypothesis-generating. Further experimen- tal studies with longer observation periods, broader molecu- lar profiling, and eventual validation in human models are warranted to better understand the long-term implications of gonadotropin exposure in the context of endometriosis. 5.1 Translational Perspective This experimental study suggests that gonadotropin expo- sure may be associated with early molecular alterations in ectopic endometriotic tissue, including reduced PTEN and TP53 expression and increased TNF-α. These findings offer mechanistic insights into potential pathways relevant to endometriosis-associated carcinogenesis. However, as they are derived from a short-term animal model, they do not provide direct evidence of cancer risk in humans. Further research is needed to explore the molecular effects of gonadotropins in endometriosis using long-term experimental designs and human studies. Until such data become available, these results should be interpreted as preliminary and hypothesis-generating.

This study was supported by the Scientific Research Projects Coordination Unit of Erciyes University (Project No: TTU-2021-11050). The funding period has concluded, and no active financial support is currently available. Authors Contribution E.K. (Erol Karakaş): Supervision, Methodology, Data Curation, Writing—Original Draft, Writing—Review & Editing. M.D.: Data Curation, Funding Acquisition, Formal Analysis, Writ- ing—Original Draft Preparation. M.E.: Investigation, Data Curation. H.A.: Writing—Original Draft Preparation, Formal Analysis. E.K. (Enes Karaman): Investigation, Data Curation, Writing. A.H.Y. and E.K. (Eda Köseoğlu): Funding Acquisition, Formal Analysis, Supervi- sion, Validation. Funding Open access funding provided by the Scientific and Techno- logical Research Council of Türkiye (TÜBİTAK). This research was supported by the Scientific Research Projects Coordination Unit of Erciyes University (Project No: TTU-2021-11050). The funding period has concluded, and no active financial support is currently available. Data Availability The datasets generated and/or analyzed during the current study are available from the corresponding author upon rea- sonable request. Declarations Ethics Approval and Consent to Participate All experimental proce- dures were approved by the Ethics Committee of Erciyes University (Approval No: 17/085). All methods were conducted in accordance with relevant institutional and international guidelines and regulations. Patient Consent for Publication Not applicable. This study did not involve human participants. Competing interests The authors declare that they have no competing interests. Open Access This article is licensed under a Creative Commons Attri- bution 4.0 International License, which permits use, sharing, adapta- tion, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

1. Vernon MW, Wilson EA. Studies on the surgical induction of endometriosis in the rat. Fertil Steril. 1985;44(5):684–94. 2. Ohba T, Ujioka T, Ishikawa K, et al. Ovarian hyperstimulation syndrome-model rats; the manifestation and clinical implication. Mol Cell Endocrinol. 2003;202(1–2):47–52. 3. Homburg R, Insler V. Ovulation induction in perspective. Hum Reprod Update. 2002;8(5):449–62. 4. Sato N, Tsunoda H, Nishida M, et  al. Loss of heterozygo- sity on 10q23.3 and mutation of the tumor suppressor gene PTEN in benign endometrial cyst of the ovary. Cancer Res. 2000;60(24):7052–6. 5. Martini M, Cenci T, D’ Amico F, et al. Possible involvement of hMLH1, p16INK4a and PTEN in the malignant transformation of endometriosis. Int J Cancer. 2002;102(4):398–406. 6. Schuijer M, Berns EMJJ. TP53 and ovarian cancer. Hum Mutat. 2003;21(3):285–91. 7. Nezhat F, Datta MS, Hanson V, et al. Comparative immunohisto- chemical studies of bcl-2 and p53 in benign and malignant ovarian endometriotic cysts. Cancer. 2002;94(11):2935–40. 8. Wang XM, Ma ZY, Song N. Inflammatory cytokines IL-6, IL-10, IL-13, TNF-α and peritoneal fluid flora were associated with infertility in patients with endometriosis. Eur Rev Med Pharmacol Sci. 2018;22(9):2513–8. 9. Riccio LdaGC, Santulli P, Marcellin L, et al. Immunology of endo- metriosis. Best Pract Res Clin Obstet Gynaecol. 2018;50:39–49. 10. Kvaskoff M, Mu F, Terry KL, et al. Endometriosis: a high-risk population for major chronic diseases? Hum Reprod Update. 2015;21(4):500–16. 11. Brinton LA, Trabert B, Anderson GL, et  al. Relationship of benign gynecologic diseases to subsequent risk of ovar - ian and uterine tumors. Cancer Epidemiol Biomarkers Prev. 2005;14(12):2929–35. 12. Anglesio MS, Bashashati A, Wang YK, et al. Multifocal endome- triotic lesions associated with cancer are clonal and carry a high mutation burden. J Pathol. 2015;236(2):201–9. 13. Suginami H. A reappraisal of the coelomic metaplasia theory by reviewing endometriosis occurring in unusual sites and instances. Am J Obstet Gynecol. 1991;165(1):214–8. 14. Burney RO, Giudice LC. Pathogenesis and pathophysiology of endometriosis. Fertil Steril. 2012;98(3):511–9. 1141Bratislava Medical Journal (2026) 127:1132–1141 15. Mantovani A, Allavena P, Sica A, Balkwill F. Cancer-related inflammation. Nature. 2008;454(7203):436–44. 16. Jiang X, Hitchcock A, Bryan EJ, et al. Microsatellite analysis of endometriosis reveals loss of heterozygosity at candidate ovarian tumor suppressor gene loci. Cancer Res. 1996;56(15):3534–9. 17. Cramer DW, Welch WR. Determinants of ovarian cancer risk II. Inferences regarding pathogenesis. J Natl Cancer Inst. 1983;71(4):717–21. 18. Brosens I, Benagiano G. Is neonatal uterine bleeding involved in the pathogenesis of endometriosis as a source of stem cells? Fertil Steril. 2013;100(3):622–3. 19. Canis M, Bourdel N, Houlle C, et al. Endometriosis may not be a chronic disease: an alternative theory offering more optimistic prospects. Fertil Steril. 2016;105(1):32–4. 20. Chapron C, Marcellin L, Borghese B, et al. Rethinking mecha- nisms, diagnosis and management of endometriosis. Nat Rev Endocrinol. 2019;15(11):666–82. 21. Arici A. Local cytokines in endometrial tissue: the role of inter - leukins and growth factors in the pathogenesis of endometriosis. Ann N Y Acad Sci. 1997;828:79–89. 22. Gazvani R, Templeton A. Peritoneal environment, cytokines and angiogenesis in the pathophysiology of endometriosis. Reproduc- tion. 2002;123(2):217–26. 23. Reagan-Shaw S, Nihal M, Ahmad N. Dose translation from animal to human studies revisited. FASEB J. 2008;22(3):659–61. https:// doi. org/ 10. 1096/ fj. 07- 9574L SF. 24. Eskenazi B, Warner ML. Epidemiology of endometriosis. Obstet Gynecol Clin North Am. 1997;24(2):235–58. 25. Inhorn MC, Patrizio P. Infertility around the globe: new thinking on gender, reproductive technologies and global movements in the 21st century. Hum Reprod Update. 2015;21(4):411–26. 26. Mascarenhas MN, et al. National, regional, and global trends in infertility prevalence. PLoS Med. 2012;9(12):e1001356. 27. Fauser BCJM, et al. Assisted reproductive technology: trends and safety. Hum Reprod Update. 2011;17(3):334–53. 28. Brinton LA, et al. Ovulation induction and cancer risk. Fertil Steril. 2005;83(2):261–74. 29. Vlahos NF, et al. Endometriosis and ovarian cancer risk: a review. Eur J Clin Invest. 2010;40(6):576–81. 30. Siristatidis CS, et al. Controlled ovarian hyperstimulation and risk of ovarian, endometrial and breast cancer. Hum Reprod Update. 2013;19(2):105–23. 31. Halme J, Hammond MG, Hulka JF, et al. Retrograde menstrua- tion in healthy women and in patients with endometriosis. Obstet Gynecol. 1984;64(2):151–4. 32. Adamson GD, Pasta DJ. Endometriosis fertility index: the new, validated endometriosis staging system. Fertil Steril. 2010;94(5):1609–15. 33. Vercellini P, Viganò P, Somigliana E, Fedele L. Endome- triosis: pathogenesis and treatment. Nat Rev Endocrinol. 2014;10(5):261–75. 34. Lebovic DI, Mueller MD, Taylor RN. Immunobiology of endo- metriosis. Fertil Steril. 2001;75(1):1–10. 35. Stewart LM, Holman CDJ, Hart R, et  al. In  vitro fertiliza- tion and breast cancer: is there cause for concern? Fertil Steril. 2012;98(2):334–40. 36. Vlahos NF, Kalampokas T, Lavranos G. Histological and molecu- lar aspects of endometrial carcinogenesis. Ann N Y Acad Sci. 2010;1205:38–44. 37. Dinulescu DM, Ince TA, Quade BJ, et al. Role of mutant PTEN and K-ras in the development of endometriosis-associated ovarian carcinoma. Proc Natl Acad Sci U S A. 2005;102(49):18644–9. 38. Simoni M, Gromoll J, Nieschlag E. The follicle-stimulating hor - mone receptor: biochemistry, molecular biology, physiology, and pathophysiology. Endocr Rev. 1997;18(6):739–73. 39. Casarini L, Simoni M. Mechanism of action of FSH. Front Endo- crinol (Lausanne). 2014;5:134. 40. Ma X, Dong Y, Matzuk MM, Kumar TR. Targeted disruption of luteinizing hormone beta-subunit leads to hypogonadism, defects in gonadal steroidogenesis, and infertility. Proc Natl Acad Sci U S A. 2004;101(49):17294–9. 41. Vasudevan KM, Garraway LA. AKT signaling in physiology and disease. Curr Top Microbiol Immunol. 2010;347:105–33. 42. Mayo LD, Donner DB. A phosphatidylinositol 3-kinase/Akt path- way promotes translocation of Mdm2 from the cytoplasm to the nucleus. Proc Natl Acad Sci U S A. 2001;98(20):11598–603. 43. Karin M, Greten FR. NF-κB: linking inflammation and immu- nity to cancer development and progression. Nat Rev Immunol. 2005;5(10):749–59. 44. Khazaei M, Montaseri A, Khazaei MR. Role of NF-κB in endo- metriosis pathogenesis. Iran Biomed J. 2009;13(4):183–90. 45. Balkwill F, Mantovani A. Inflammation and cancer: back to Vir- chow? Lancet. 2001;357(9255):539–45. 46. Aggarwal BB. Nuclear factor-κB: the enemy within. Cancer Cell. 2004;6(3):203–8. 47. Wiegand KC, Shah SP, Al-Agha OM, et al. ARID1A mutations in endometriosis-associated ovarian carcinomas. N Engl J Med. 2010;363(16):1532–43. 48. Koshiyama M, Matsumura N, Konishi I. Subtypes of ovar - ian cancer and ovarian cancer screening. Diagnostics (Basel). 2017;7(1):12. 49. Ness RB. Endometriosis and ovarian cancer: thoughts on shared pathophysiology. Am J Obstet Gynecol. 2003;189(1):280–94. 50. Anglesio MS, Bashashati A, Wang YK, et al. Cancer-associ- ated mutations in endometriosis without cancer. N Engl J Med. 2017;376(19):1835–48. 51. Kvaskoff M, Mu F, Terry KL, Harris HR, Poole EM, Farland L, et al. Endometriosis: a high-risk population for major chronic diseases? Hum Reprod Update. 2015;21(4):500–16. Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.