Mouse model for endometriosis is characterized by proliferation and inflammation but not epithelial-to-mesenchymal transition and fibrosis

Endometriosis is a common disorder of unknown etiology, and non-surgical therapies are still a challenge. To understand the pathogenesis and preclinical testing of drugs for endometriosis, animal models are highly desirous. Herein, we carried out longitudinal characterization of a mouse model for endometriosis where uterine tissue was transplanted onto the intestinal mesentery. During the course of lesion development from day 15 to 60 post-induction, the ectopic endometrium became pale, fluid-filled and the animals developed peritoneal adhesions. Most lesions resembled a well-differentiated type of endometriosis and ~ 13% of animals had mixed type of lesions. There was extensive stromal compaction in the ectopic tissue. During the progression of endometriosis, there was increased proliferation of epithelial and stromal cells as evident by PCNA staining. Cyp19a1 (aromatase) mRNA was detected in the ectopic lesions on day 15 and 30 post-induction of endometriosis, by day 60 the expression was reduced. As compared to the control endometrium, the mRNA levels of Esr1 progressively reduced while the levels of inflammation associated genes (Esr2, Ifng, Tnf and Il1b) increased in the ectopic lesions. Infiltration of macrophages and polymorphonuclear leucocytes was also observed in the ectopic lesions indicative of inflammation. As compared to control, there was no change in levels of Cytokeratin and E-cadherin in the epithelial cells of ectopic endometrium. We did not observe excessive collagen deposition or α-SMA positive myofibroblasts in the stroma of the ectopic endometrium. Thus, epithelial-to-mesenchymal transition and fibrosis are not detected in the mouse model of endometriosis. Our results show that the mouse model of endometriosis mimics some but not all the features of human endometriosis. Similar content being viewed by others

Abrao MS, Neme RM, Carvalho FM, Aldrighi JM and Pinotti JA 2003 Histological classification of endometriosis as a predictor of response to treatment. Int. J. Gynaecol. Obstet. 82 31–40 Barcena de Arellano ML, Gericke J, Reichelt U, Okuducu AF, Ebert AD, et al. 2011 Immunohistochemical characterization of endometriosis-associated smooth muscle cells in human peritoneal endometriotic lesions. Hum. Reprod. 26 2721–2730 Bernuit D, Ebert AD, Halis G, Strothmann A, Gerlinger C, et al. 2011 Female perspectives on endometriosis: Findings from the Uterine Bleeding and Pain Women’s Research Study. J. Endometriosis 3 73–85 Bhutda S, Surve MV, Anil A, Kamath KG, Singh N et al. 2017 Histochemical staining of collagen and identification of its subtypes by picrosirius red dye in mouse reproductive tissues. Bio-protocol 7 e2592 Bilotas M, Meresman G, Stella I, Sueldo C and Baranao RI 2010 Effect of aromatase inhibitors on ectopic endometrial growth and peritoneal environment in a mouse model of endometriosis. Fertil. Steril. 93 2513–2518 Bozdag G 2015 Recurrence of endometriosis: risk factors, mechanisms and biomarkers. Womens Health 11 693–699 Braundmeier AG and Fazleabas AT 2009 The non-human primate model of endometriosis: research and implications for fecundity. Mol. Hum. Reprod. 15 577–586 Bruner-Tran KL, Mokshagundam S, Herington JL, Ding T and Osteen KG 2018 Rodent models of experimental endometriosis: identifying mechanisms of disease and therapeutic targets. Curr Womens Health Rev. 14 173–188 Bullon P and Navarro JM 2017 Inflammasome as a key pathogenic mechanism in endometriosis. Curr. Drug Targets 18 997–1002 Chehna-Patel N, Sachdeva G, Gajbhiye R, Warty N and Khole V 2010 “Spot”-ting differences between the ectopic and eutopic endometrium of endometriosis patients. Fertil. Steril. 94 1964–1971, 1971 e1961 Cummings AM and Metcalf JL 1995 Induction of endometriosis in mice: a new model sensitive to estrogen. Reprod. Toxicol. 9 233–238 Debrock S, Vander Perre S, Meuleman C, Moerman P, Hill JA, et al. 2002 In-vitro adhesion of endometrium to autologous peritoneal membranes: effect of the cycle phase and the stage of endometriosis. Hum. Reprod. 17 2523–2528 Dutta M, Anitha M, Smith PB, Chiaro CR, Maan M, et al. 2016 Metabolomics reveals altered lipid metabolism in a mouse model of endometriosis. J. Proteome Re.s 15 2626–2633 Fazleabas AT, Brudney A, Chai D, Langoi D and Bulun SE 2003 Steroid receptor and aromatase expression in baboon endometriotic lesions. Fertil. Steril. 80 820–827 Fazleabas AT, Brudney A, Gurates B, Chai D and Bulun S 2002 A modified baboon model for endometriosis. Ann. NY Acad. Sci. 955 308–317; discussion 340–302, 396–406 Gajbhiye R, McKinnon B, Mortlock S, Mueller M and Montgomery G 2018 Genetic variation at chromosome 2q13 and its potential influence on endometriosis susceptibility through effects on the IL-1 family. Reprod. Sci. 25 1307–1317 Galvankar M, Singh N and Modi D 2017 Estrogen is essential but not sufficient to induce endometriosis. J. Biosci. 42 251–263 Godbole GB, Modi DN and Puri CP 2007 Regulation of homeobox A10 expression in the primate endometrium by progesterone and embryonic stimuli. Reproduction 134 513–523 Gordts S, Koninckx P and Brosens I 2017 Pathogenesis of deep endometriosis. Fertil. Steril. 108 872–885 e871 Greaves E, Cousins FL, Murray A, Esnal-Zufiaurre A, Fassbender A et al. 2014 A novel mouse model of endometriosis mimics human phenotype and reveals insights into the inflammatory contribution of shed endometrium. Am. J. Pathol. 184 1930–1939 Greaves E, Critchley HOD, Horne AW and Saunders PTK 2017 Relevant human tissue resources and laboratory models for use in endometriosis research. Acta Obstet. Gynecol. Scand. 96 644–658 Han SJ, Jung SY, Wu SP, Hawkins SM, Park MJ, et al. 2015 Estrogen receptor beta modulates apoptosis complexes and the inflammasome to drive the pathogenesis of endometriosis. Cell 163 960–974 Jha U, Asad M, Asdaq SM, Das AK and Prasad VS 2010 Fertility inducing effect of aerial parts of Coccinia cordifolia L. in female rats. J. Ethnopharmacol. 127 561–564 Jones RL, Lang SA, Kendziorski JA, Greene AD and Burns KA 2018 Use of a mouse model of experimentally induced endometriosis to evaluate and compare the effects of bisphenol A and bisphenol AF exposure. Environ. Health Perspect. 126 127004 Laheri S, Modi D and Bhatt P 2017 Extra-oviductal expression of oviductal glycoprotein 1 in mouse: Detection in testis, epididymis and ovary. J. Biosci. 42 69–80 Langoi D, Pavone ME, Gurates B, Chai D, Fazleabas A, et al. 2013 Aromatase inhibitor treatment limits progression of peritoneal endometriosis in baboons. Fertil. Steril. 99 656–662 e653 Lin YH, Chen YH, Chang HY, Au HK, Tzeng CR, et al. 2018 Chronic niche inflammation in endometriosis-associated infertility: current understanding and future therapeutic strategies. Int. J. Mol. Sci. 19 2385 Lin YJ, Lai MD, Lei HY and Wing LY 2006 Neutrophils and macrophages promote angiogenesis in the early stage of endometriosis in a mouse model. Endocrinology 147 1278–1286 Maia H, Jr., Haddad C and Casoy J 2012 Correlation between aromatase expression in the eutopic endometrium of symptomatic patients and the presence of endometriosis. Int. J. Womens Health 4 61–65 Matsuzaki S and Darcha C 2012 Epithelial to mesenchymal transition-like and mesenchymal to epithelial transition-like processes might be involved in the pathogenesis of pelvic endometriosis. Hum. Reprod. 27 712–721 McKinnon B, Mueller M and Montgomery G 2018 Progesterone resistance in endometriosis: an acquired property? Trends Endocrinol. Metab. 29 535–548 Mehedintu C, Plotogea MN, Ionescu S and Antonovici M 2014 Endometriosis still a challenge. J. Med. Life 7 349–357 Mishra A, Galvankar M, Singh N and Modi D 2020 Spatial and temporal changes in the expression of steroid hormone receptors in mouse model of endometriosis. J. Assist. Reprod. Genet. 37 1069–1081 Mishra VV, Gaddagi RA, Aggarwal R, Choudhary S, Sharma U, et al. 2015 Prevalence; characteristics and management of endometriosis amongst infertile women: a one year retrospective study. J. Clin. Diagn. Res. 9 QC01–03 Munoz-Hernando L, Munoz-Gonzalez JL, Marqueta-Marques L, Alvarez-Conejo C, Tejerizo-Garcia A, et al. 2015 Endometriosis: alternative methods of medical treatment. Int. J. Womens Health 7 595–603 Nair HB, Baker R, Owston MA, Escalona R, Dick EJ, et al. 2016 An efficient model of human endometriosis by induced unopposed estrogenicity in baboons. Oncotarget 7 10857–10869 Nishimoto-Kakiuchi A, Netsu S, Okabayashi S, Taniguchi K, Tanimura H, et al. 2018 Spontaneous endometriosis in cynomolgus monkeys as a clinically relevant experimental model. Hum. Reprod. 33 1228–1236 Park JS, Lee JH, Kim M, Chang HJ, Hwang KJ, et al. 2009 Endometrium from women with endometriosis shows increased proliferation activity. Fertil. Steril. 92 1246–1249 Pelch KE, Sharpe-Timms KL and Nagel SC 2012 Mouse model of surgically-induced endometriosis by auto-transplantation of uterine tissue. J. Vis. Exp. https://doi.org/10.3791/3396.e3396 Pellegrini C, Gori I, Achtari C, Hornung D, Chardonnens E, et al. 2012 The expression of estrogen receptors as well as GREB1, c-MYC, and cyclin D1, estrogen-regulated genes implicated in proliferation, is increased in peritoneal endometriosis. Fertil. Steril. 98 1200–1208 Pereira FE, Almeida PR, Dias BH, Vasconcelos PR, Guimaraes SB, et al. 2015 Development of a subcutaneous endometriosis rat model. Acta Cir. Bras. 30 6–12 Qi Q-M, Guo S-W and Liu X-S 2017 Estrogen biosynthesis and its regulation in endometriosis. Reprod. Dev. Med. 1 55–61 Rahimi S, Akaev I, Marani C, Chopra M and Yeoh CC 2019 Immunohistochemical expression of different subtypes of cytokeratins by endometrial stromal sarcoma. Appl. Immunohistochem. Mol. Morphol. 27 466–470 Rogers PA, D’Hooghe TM, Fazleabas A, Gargett CE, Giudice LC, et al. 2009 Priorities for endometriosis research: recommendations from an international consensus workshop. Reprod. Sci. 16 335–346 Schindelin J, Arganda-Carreras I, Frise E, Kaynig V, Longair M,et al. 2012 Fiji: an open-source platform for biological-image analysis. Nat. Methods 9 676–682 Somigliana E, Vigano P, Benaglia L, Busnelli A, Vercellini P, et al. 2012 Adhesion prevention in endometriosis: a neglected critical challenge. J. Minim. Invasive Gynecol. 19 415–421 Soni UK, Chadchan SB, Kumar V, Ubba V, Khan MTA, et al. 2019 A high level of TGF-B1 promotes endometriosis development via cell migration, adhesiveness, colonization, and invasivenessdagger. Biol. Reprod. 100 917–938 Symons LK, Miller JE, Kay VR, Marks RM, Liblik K, et al. 2018 The immunopathophysiology of endometriosis. Trends Mol. Med. 24 748–762 Umezawa M, Tanaka N, Tainaka H, Takeda K, Ihara T, et al. 2009 Microarray analysis provides insight into the early steps of pathophysiology of mouse endometriosis model induced by autotransplantation of endometrium. Life Sci. 84 832–837 Varghese F, Bukhari AB, Malhotra R and De A 2014 IHC Profiler: an open source plugin for the quantitative evaluation and automated scoring of immunohistochemistry images of human tissue samples. PLoS One 9 e96801 Vercellini P, Somigliana E, Vigano P, De Matteis S, Barbara G, et al. 2009 The effect of second-line surgery on reproductive performance of women with recurrent endometriosis: a systematic review. Acta Obstet. Gynecol. Scand. 88 1074–1082 Vigano P, Candiani M, Monno A, Giacomini E, Vercellini P, et al. 2018 Time to redefine endometriosis including its pro-fibrotic nature. Hum. Reprod. 33 347–352 Wingfield M, Macpherson A, Healy DL and Rogers PA 1995 Cell proliferation is increased in the endometrium of women with endometriosis. Fertil. Steril. 64 340–346 Xiong W, Zhang L, Liu H, Li N, Du Y, et al. 2019 E2 -mediated EMT by activation of beta-catenin/Snail signalling during the development of ovarian endometriosis. J. Cell Mol. Med. 23 8035–8045 Yang YM and Yang WX 2017 Epithelial-to-mesenchymal transition in the development of endometriosis. Oncotarget 8 41679–41689 Yilmaz BD and Bulun SE 2019 Endometriosis and nuclear receptors. Hum. Reprod. Update 25 473–485 Zhang Q, Dong P, Liu X, Sakuragi N and Guo SW 2017 Enhancer of Zeste homolog 2 (EZH2) induces epithelial-mesenchymal transition in endometriosis. Sci. Rep. 7 6804 Zhang Q, Duan J, Liu X and Guo SW 2016 Platelets drive smooth muscle metaplasia and fibrogenesis in endometriosis through epithelial-mesenchymal transition and fibroblast-to-myofibroblast transdifferentiation. Mol. Cell Endocrinol. 428 1–16 Zhao Y, Gong P, Chen Y, Nwachukwu JC, Srinivasan S, et al. 2015 Dual suppression of estrogenic and inflammatory activities for targeting of endometriosis. Sci. Transl. Med. 7 271ra279

We express our gratitude to Dr Lalita Shankar Savardekar for sparing the Artisan™ Masson’s Trichrome Stain Kit. We thank the staff of the Animal House (NIRRH) for their help during surgeries and animal maintenance. The help of Dr Atahar Husein in editing the manuscript is gratefully acknowledged. The manuscript bears the NIRRH ID: RA/791/08–2019. DM’s lab is funded by grants from ICMR, India. The study was funded by grants from the Department of Biotechnology (DBT), India (BT/OR6587/MED/30/886/2012) to DM. AM is the recipient of the Junior and Senior Research fellowship from the University Grants Commission (UGC), India. MG was the recipient of the ICMR postdoctoral fellowship (sixth batch). Author information Authors and Affiliations Corresponding author Additional information Communicated by Ullas Kolthur-Seetharam. Corresponding editor: Ullas Kolthur-Seetharam Electronic supplementary material Below is the link to the electronic supplementary material. Rights and permissions About this article Cite this article Mishra, A., Galvankar, M., Vaidya, S. et al. Mouse model for endometriosis is characterized by proliferation and inflammation but not epithelial-to-mesenchymal transition and fibrosis. J Biosci 45, 105 (2020). https://doi.org/10.1007/s12038-020-00073-y Received: Accepted: Published: Version of record: DOI: https://doi.org/10.1007/s12038-020-00073-y