Repurposing ramipril to mitigate EMT-like transition in endometriosis by PI3K/AKT/S6K1 signalling pathway: a study in endometriosis induced rats

Endometriosis, an atypical benign disorder, may disrupt epithelial-mesenchymal transition (EMT) due to a dysregulated balance between matrix metalloproteinases (MMPs) and their inhibitors. Ramipril, an angiotensin converting enzyme (ACE) inhibitor, is crucial in mediating angiogenesis, inflammation, oxidative stress, and apoptosis. In the current work, we investigated how ramipril modulates EMT in endometriosis rats. Five adult virgin female Wistar rats were donor rats, and thirty rats were randomly divided into three groups following peritoneal uterine tissue transplantation (group 2 and group 3). The sham control group was group 1. Morphological alterations were predominantly assessed through hematoxylin–eosin (H-E) staining, succeeded by the immunoreactivity analysis of MMP9, tissue inhibitor of metalloproteinases 1 (TIMP1), reversion-inducing cysteine-rich protein with Kazal motifs (RECK), epithelial cadherin (E-cadherin), neural cadherin (N-cadherin), and vimentin in the uteri and ectopic lesions of the specified groups. Zymography was conducted to assess the activities of MMP9 and MMP2. Immunoblotting was subsequently conducted for MMP9, TIMP1, snail, E-cadherin, N-cadherin, and vimentin in both uteri and ectopic lesions. Immunoblotting was conducted for phosphoinositide 3-kinase (PI3K), protein kinase B (Akt), mechanistic target of rapamycin (mTOR), p70 ribosomal S6 kinase 1 (S6K1), and hypoxia-inducible factor 1-alpha (HIF-1ɑ) in the ectopic lesions. A significant reduction in the average quantity of ectopic endometrial glands was recorded in group 3. A significant decrease in the expressions of MMP9:TIMP1 (RECK) ratios (p = 0.00012; p = 0.001), snail, p-PI3K, p-AKT, p-mTOR, p-S6K1, and HIF-1ɑ (p < 0.05) proteins was observed in the ectopic lesions of group 3. A significant increase in the E-cadherin to N-cadherin (vimentin) ratios (p = 0.001) was observed in group 3 ectopic lesions. In conclusion, the administration of ramipril led to the reversal of EMT-like process by disrupting the MMP9/PI3K/AKT/S6K1 pathway in rats induced with endometriosis. Similar content being viewed by others Data availability The corresponding author can provide the datasets used in this study upon reasonable request. Abbreviations - ACE : - Angiotensin converting enzyme - AP : - Alkaline phosphatase - BCA : - Bicinchoninic acid method - BCIP : - 5-Bromo-4-chloro-3-indoyl-phosphate - BSA : - Bovine serum albumin - EMT : - Epithelial-mesenchymal transition (EMT) - ECM : - Extracellular matrix - H-E : - Hematoxylin-eosin - HRP : - Horseradish peroxidase - HIF-1ɑ : - Hypoxia-inducible factor 1-alpha - MMPs : - Matrix metalloproteinases - mTOR : - Mechanistic target of rapamycin - NBT : - Nitro blue tetrazolium - OVX : - Ovariectomy - PBS : - Phosphate buffered saline - PI3K : - Phosphoinositide 3-kinase - RECK : - Reversion-inducing cysteine-rich protein with Kazal motifs - RIPA : - Radioimmunoprecipitation assay buffer - TIMP1 : - Tissue inhibitor of metalloproteinases 1 - TFs : - Transcription factors - S6K1 : - P70 ribosomal S6 kinase 1

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The first author, Piyali Mazumdar, is thankful to the University Grants Commission (UGC), New Delhi, India, for providing the research fellowship, UGC-Ref. No.: UGCES-22-GE-WES-F-SJSGC-4185. Funding This work was supported by a grant from the scientific project {1198(Sanc.)/STBT-11012(15)}, funded by the Department of Science and Technology and Biotechnology, West Bengal, India. We are thankful to DBT-BUILDER (BT/INF/22/SP45088/2022) for instrumental and infrastructural facilities and DST-FIST, Govt. of India for other infrastructural facilities to the Life Sciences department (SR/FST/LSI-560/2013). Author information Authors and Affiliations Contributions PM has conceptualised, researched, designed, executed the tests, analysed the results, and prepared the original draft. SSB has conceptualised, overseen, secured funding and laboratory resources, and revised the initial draft. The authors declare that all data were generated in-house and that no paper mill was used. Corresponding author Ethics declarations Ethics approval and consent to participate All animal procedures followed the Committee for Control and Supervision of Experiments on Animals (CCSEA) guidelines for the Humane Care of Laboratory Animals. Presidency University, Kolkata, India Institutional Animals Care and Use Committee has authorized all the procedures. Consent for publication Not applicable. Competing interests The authors declare that they have no competing interests. Additional information Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Supplementary Information Below is the link to the electronic supplementary material. ESM 1 (download PDF ) (PDF 134 KB) Rights and permissions Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law. About this article Cite this article Mazumdar, P., Biswas, S.S. Repurposing ramipril to mitigate EMT-like transition in endometriosis by PI3K/AKT/S6K1 signalling pathway: a study in endometriosis induced rats. Naunyn-Schmiedeberg's Arch Pharmacol 399, 9923–9936 (2026). https://doi.org/10.1007/s00210-026-05030-4 Received: Accepted: Published: Version of record: Issue date: DOI: https://doi.org/10.1007/s00210-026-05030-4