Effect of a dienogest for an experimental three-dimensional endometrial culture model for endometriosis

The pathogenesis of endometriosis remains poorly understood at least in part because early stages of the disease process are difficult to investigate. Previous studies have proposed a three-dimensional fibrin matrix culture model to study human endometriosis. We examined the ultrastructural features of the endometriosis in this model and assessed the effect of a progestin on endometrial outgrowth and apoptosis in this culture system. Endometrial explants were placed in three-dimensional fibrin matrix culture and treated with and without various concentrations of the progestin dienogest. By the second week, endometrial gland-like formation was established in outgrowths both attached to and at a distance from the explants. These cells formed a combination of clumps and tubular monolayers surrounding a central cavity. Electron microscopy demonstrated that these cells are polarized with microvilli on the apical surface, desmosome-like structures, and basement membrane; features consistent with glandular epithelial cells. Outgrowth of endometrial stromal cells and glandular formation was impaired in response to dienogest in a dose-dependent manner. Our study shows that the human endometrial explants cultured in three-dimensional fibrin matrix establish outgrowths that ultrastructurally resemble ectopic endometrial implants. This model may provide insight into the cellular processes leading to endometriosis formation and enables screening of therapeutic compounds. Similar content being viewed by others

Holoch KJ, Lessey BA (2010) Endometriosis and infertility. Clin Obstet Gynecol 53:429–438 Coccia ME, Rizzello F, Palagiano A, Scarselli G (2011) Long-term follow-up after laparoscopic treatment for endometriosis: multivariate analysis of predictive factors for recurrence of endometriotic lesions and pain. Eur J Obstet Gynecol Reprod Biol 157:78–83 Porpora MG, Pallante D, Ferro A, Crisafi B, Bellati F, Benedetti Panici P (2010) Pain and ovarian endometrioma recurrence after laparoscopic treatment of endometriosis: a long-term prospective study. Fertil Steril 93:716–721 Lee DY, Bae DS, Yoon BK, Choi D (2010) Post-operative cyclic oral contraceptive use after gonadotrophin-releasing hormone agonist treatment effectively prevents endometrioma recurrence. Hum Reprod 25:3050–3054 Fourquet J, Baez L, Figueroa M, Iriarte RI, Flores I (2011) Quantification of the impact of endometriosis symptoms on health-related quality of life and work productivity. Fertil Steril 96:107–112 Braundmeier AG, Fazleabas AT (2009) The non-human primate model of endometriosis: research and implications for fecundity. Mol Hum Reprod 15:577–586 Fasciani A, Bocci G, Xu J, Bielecki R, Greenblatt E, Leyland N, Casper RF (2003) Three-dimensional in vitro culture of endometrial explants mimics the early stages of endometriosis. Fertil Steril 80:1137–1143 Sulaiman H, Dawson L, Laurent GJ, Bellingan GJ, Herrick SE (2002) Role of plasminogen activators in peritoneal adhesion formation. Biochem Soc Trans 30:126–131 Esfandiari N, Ai J, Nazemian Z, Javed MH, Gotlieb L, Casper RF (2007) Expression of glycodelin and cyclooxygenase-2 in human endometrial tissue following three-dimensional culture. Am J Reprod Immunol 7:49–54 Esfandiari N, Khazaei M, Ai J, Bielecki R, Gotlieb L, Ryan E, Casper RF (2007) Effect of a statin on an in vitro model of endometriosis. Fertil Steril 87:257–262 Kuhl H (1996) Comparative pharmacology of newer progestogens. Drugs 51:188–215 Foster RH, Wilde MI (1998) Dienogest. Drugs 56:825–833 Bunge RP, Wood P (1973) Studies on the transplantation of spinal cord tissue in the rat. I. The development of a culture system for hemisections of embryonic spinal cord. Brain Res 57:261–276 Witz CA, Monotoya-Rodriguez IA, Schenken RS (1999) Whole explants of peritoneum and endometrium: a novel model of the early endometriosis lesion. Fertil Steril 71:56–60 Witz CA, Dechaud H, Montoya-Rodriguez IA, Thomas MR, Nair AS, Centonze VE, Schenken RS (2002) An in vitro model to study the pathogenesis of the early endometriosis lesion. Ann N Y Acad Sci 955:296–307 Ohama Y, Harada T, Iwabe T, Taniguchi F, Takenaka Y, Terakawa N (2008) Peroxisome proliferator-activated receptor-gamma ligand reduced tumor necrosis factor-alpha-induced interleukin-8 production and growth in endometriotic stromal cells. Fertil Steril 89:311–317 Schweppe KW, Wynn RM (1981) Ultrastructural changes in endometriotic implants during the menstrual cycle. Obstet Gynecol 58:465–473 Schweppe KW, Wynn RM, Beller FK (1984) Ultrastructural comparison of endometriotic implants and eutopic endometrium. Am J Obstet Gynecol 148:1024–1039 Gordon M (1975) Cyclic changes in the fine structure of the epithelial cells of human endometrium. Int Rev Cytol 42:127–172 Boxberger HJ, Meyer TF, Grausam MC, Reich K, Becker HD, Sessler MJ (1997) Isolating and maintaining highly polarized primary epithelial cells from normal human duodenum for growth as spheroid-like vesicles. In Vitro Cell Dev Biol Anim 33:536–545 Jacquot J, Spilmont C, Burlet H, Fuchey C, Buisson AC, Tournier JM, Gaillard D, Puchelle E (1994) Glandular-like morphogenesis and secretory activity of human tracheal gland cells in a three-dimensional collagen gel matrix. J Cell Physiol 161:407–418 Strowitzki T, Marr J, Gerlinger C, Faustmann T, Seitz C (2010) Dienogest is as effective as leuprolide acetate in treating the painful symptoms of endometriosis: a 24-week, randomized, multicentre, open-label trial. Hum Reprod 25:633–641 Harada T, Momoeda M, Taketani Y, Aso T, Fukunaga M, Hagino H, Terakawa N (2009) Dienogest is as effective as intranasal buserelin acetate for the relief of pain symptoms associated with endometriosis—a randomized, double-blind, multicenter, controlled trial. Fertil Steril 91:675–681 Okada H, Nakajima T, Yoshimura T, Yasuda K, Kanzaki H (2001) The inhibitory effect of dienogest, a synthetic steroid, on the growth of human endometrial stromal cells in vitro. Mol Hum Reprod 7:341–347 Fu L, Osuga Y, Morimoto C, Hirata T, Hirota Y, Yano T, Taketani Y (2008) Dienogest inhibits BrdU uptake with G0/G1 arrest in cultured endometriotic stromal cells. Fertil Steril 89:1344–1347 Horie S, Harada T, Mitsunari M, Taniguchi F, Iwabe T, Terakawa N (2005) Progesterone and progestational compounds attenuate tumor necrosis factor alpha-induced interleukin-8 production via nuclear factor kappa B inactivation in endometriotic stromal cells. Fertil Steril 83:1530–1535 Okada H, Okamoto R, Tsuzuki T, Tsuji S, Yasuda K, Kanzaki H (2011) Progestins inhibit estradiol-induced vascular endothelial growth factor and stromal cell-derived factor 1 in human endometrial stromal cells. Fertil Steril 96:786–791 Sasagawa S, Shimizu Y, Nagaoka T, Tokado H, Imada K, Mizuguchi K (2008) Dienogest, a selective progestin, reduces plasma estradiol level through induction of apoptosis of granulosa cells in the ovarian dominant follicle without follicle-stimulating hormone suppression in monkeys. J Endocrinol Invest 31:636–641 Mattioli F, Garbero C, Gosmar M, Manfredi V, Carrozzino R, Martelli A, Brambilla G (2004) DNA fragmentation, DNA repair and apoptosis induced in primary rat hepatocytes by dienogest, dydrogesterone and 1,4,6-androstatriene-17beta-ol-3-one acetate. Mutat Res 564:21–29 Banu SK, Lee J, Speights VO Jr, Starzinski-Powitz A, Arosh JA (2008) Cyclooxygenase-2 regulates survival, migration, and invasion of human endometriotic cells through multiple mechanisms. Endocrinology 149:1180–1189 Yamanaka K, Xu B, Suganuma I, Kusuki I, Mita S, Shimizu Y, Mizuguchi K, Kitawaki J (2012) Dienogest inhibits aromatase and cyclooxygenase-2 expression and prostaglandin E2 production in human endometriotic stromal cells in spheroid culture. Fertil Steril 97:477–482 Hever A, Roth RB, Hevezi PA, Lee J, Willhite D, White EC, Marin EM, Herrera R, Acosta HM, Acosta AJ, Zlotnik A (2006) Molecular characterization of human adenomyosis. Mol Hum Reprod 12:737–748 Acknowledgments T.K. received grant-in-aid from Scientific Research from the Ministry of Education, Science, and Culture, Japan (no. 23592412). J.J.B. was supported by the Biomedical Research Unit in Reproductive Health, a joint initiative between the University Hospitals Coventry and Warwickshire National Health Service Trust and Warwick Medical School. Author information Authors and Affiliations Corresponding author Rights and permissions About this article Cite this article Prechapanich, J., Kajihara, T., Fujita, K. et al. Effect of a dienogest for an experimental three-dimensional endometrial culture model for endometriosis. Med Mol Morphol 47, 189–195 (2014). https://doi.org/10.1007/s00795-013-0059-3 Received: Accepted: Published: Issue date: DOI: https://doi.org/10.1007/s00795-013-0059-3