Journal of Reproduction & Infertility

Journal of Reproduction & Infertility

Detailed Investigation of Downstream TLR Signaling in the Follicular Cells of Women with Endometriosis

Authors
Reza Jafari 1
Seyed Abdolvahab Taghavi 2
Elham Amirchaghmaghi 3
Reza Salman Yazdi 4
Leili Karimian 5
Mahnaz Ashrafi 6
Reza Aflatoonian 3
1 School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran
2 Department of Gynecology and Obstetrics, School of Medicine, Yasuj University of Medical Sciences, Yasuj, Iran
3 Department of Endocrinology and Female Infertility, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran
4 Department of Andrology, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran
5 Department of Embryology, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran
6 Department of Endocrinology and Female Infertility, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran; Department Gynecology and Obstetrics, School of Medicine, Iran Unive
10.18502/jri.v21i4.4325
Abstract
Background: Inflammatory responses within the peritoneal cavity may result in endometrial dysfunction in women with endometriosis. The true causes of this disease remain poorly understood. It is hypothesized that downstream toll-like receptors (TLRs) inflammatory cytokines in response to pathogens may be associated with endometriosis. So, this study was aimed at evaluating the expression of TLRs signaling and endometriosis-associated inflammatory responses. Methods: Totally, 20 infertile endometriosis patients and 20 normal women undergoing controlled ovarian stimulation were enrolled. The cellular pellet and supernatant were obtained by centrifugation of follicular fluid (FF). Evaluation of TLRs and their signaling pathway gene expression was performed on cellular pellets using quantitative-PCR. The supernatant was used for determination of cytokine protein expression by ELISA. The results are expressed as mean±SEM and a p<0.05 was considered statistically significant. Results: Quantitative-PCR analysis suggested that TLR1, 5, 6, 7, 8, 10, MYD88, NF-ĸB, IL-10 and TGF-β genes expression significantly increased in patients compared to the control group (p<0.05). TLR3, 9, INF-β genes expression was significantly lower in endometriosis than control group (p<0.05). There was no significant difference in the expression of TLR2, TLR4, TIRAP, TRIF, TRAM, and IRF3 between two groups. Also, significant increase in the levels of IL-6, IL-8 and MIF protein in FF of endometriosis group was detected in comparison with normal women (p<0.05). Conclusion: The expression of TLR downstream signaling in the follicular cells can initiate inflammatory responses and changes in the FF cytokine profile which in turn may induce endometriosis and infertility disorder.
Keywords
Endometriosis
Follicular cells
Infertility
Inflammation
TLR
  1. Jones R. Bisphenol A and Bisphenol AF potentiate endometriosis differently based on hormonal status in female mice and disrupt normal ovarian function [dissertation]: [USA]: University of Cincinnati; 2018. 54 p.
  2. Stilley JAW, Birt JA, Sharpe-Timms KL. Cellular and molecular basis for endometriosis-associated infertility. Cell Tissue Res. 2012;349(3):849-62.
  3. Kajihara H, Yamada Y, Kanayama S, Furukawa N, Noguchi T, Haruta S, et al. New insights into the pathophysiology of endometriosis: from chronic inflammation to danger signal. Gynecol Endocrinol. 2011;27(2):73-9.
  4. Burney RO, Giudice LC. Pathogenesis and pathophysiology of endometriosis. Fertil Steril. 2012;98(3):511-9.
  5. Sidell N, Han SW, Parthasarathy S. Regulation and modulation of abnormal immune responses in endometriosis. Ann N Y Acad Sci. 2002;955:159-73.
  6. Berbic M, Fraser I. Regulatory T cells and other leukocytes in the pathogenesis of endometriosis. J Reprod Immunol. 2011;88(2):149-55.
  7. Ahn SH, Monsanto SP, Miller C, Singh SS, Thomas R, Tayade C. Pathophysiology and immune dysfunction in endometriosis. Biomed Res Int. 2015;2015:795976.
  8. Latha M, Vaidya S, Movva S, Chava S, Govindan S, Govatati S, et al. Molecular pathogenesis of endometriosis; Toll-like receptor-4 A896G (D299G) polymorphism: a novel explanation. Genet Test Mol Biomarkers. 2011;15(3):181-4.
  9. Khan KN, Kitajima M, Fujishita A, Nakashima M, Masuzaki H. Toll‐like receptor system and endometriosis. J Obstet Gynaecol Res. 2013;39(8):1281-92.
  10. Redegeld FA, Yu Y, Kumari S, Charles N, Blank U. Non‐IgE mediated mast cell activation. Immunol Rev. 2018;282(1):87-113.
  11. Amirchaghmaghi E, Taghavi SA, Shapouri F, Saeidi S, Rezaei A, Aflatoonian R. The role of toll like receptors in Pregnancy. Int J Fertil Steril. 2013;7(3):147-54.
  12. McKernan DP. Toll-like receptors and immune cell crosstalk in the intestinal epithelium. AIMS Allergy Immunol. 2019;3(1):13.
  13. Takeda K, Kaisho T, Akira S. Toll-like receptors. Annu Rev Immunol. 2003;21:335-76.
  14. Jafari R, Aflatoonian R, Falak R, Pourmand G, Dehghani S, Mortazavi M, et al. Down-regulation of inflammatory signaling pathways despite up-regulation of Toll-like receptors; the effects of corticosteroid therapy in brain-dead kidney donors, a double-blind, randomized, controlled trial. Mol Immunol. 2018;94:36-44.
  15. Kawai T, Akira S. TLR signaling. Semin Immunol. 2007;19(1):24-32.
  16. Fazeli A, Bruce C, Anumba DO. Characterization of Toll-like receptors in the female reproductive tract in humans. Hum Reprod. 2005;20(5):1372-8.
  17. Aflatoonian R, Tuckerman E, Elliott SL, Bruce C, Aflatoonian A, Li TC, et al. Menstrual cycle-dependent changes of Toll-like receptors in endometrium. Hum Reprod. 2007;22(2):586-93.
  18. Aboussahoud W, Aflatoonian R, Bruce C, Elliott S, Ward J, Newton S, et al. Expression and function of Toll-like receptors in human endometrial epithelial cell lines. J Reprod Immunol. 2010;84(1):41-51.
  19. Taghavi SA, Ashrafi M, Mehdizadeh M, Karimian L, Joghataie MT, Aflatoonian R. Toll-like receptors expression in follicular cells of patients with poor ovarian response. Int J Fertil Steril. 2013;8(2):183-92.
  20. Gholamnezhadjafari R, Tajik N, Falak R, Aflatoonian R, Dehghan S, Rezaei A. Innate inflammatory gene expression profiling in potential brain-dead donors: detailed investigation of the effect of common corticosteroid therapy. Innate Immun. 2017;23(5):440-8.
  21. Agic A, Xu H, Finas D, Banz C, Diedrich K, Hornung D. Is endometriosis associated with systemic subclinical inflammation? Gynecol Obstet Invest. 2006;62(3):139-47.
  22. Zarmakoupis PN, Rier SE, Maroulis GB, Becker JL. Uterus and endometrium: inhibition of human endometrial stromal cell proliferation by interleukin 6. Hum Reprod. 1995;10(9):2395-9.
  23. Rier SE, Zarmakoupis PN, Hu X, Becker JL. Dysregulation of interleukin-6 responses in ectopic endometrial stromal cells: correlation with decreased soluble receptor levels in peritoneal fluid of women with endometriosis. J Clin Endocrinol Metabol. 1995;80(4):1431-7.
  24. [No authors listed]. Revised American society for reproductive medicine classification of endometriosis: 1996. Fertil Steril. 1997;67(5):817-21.
  25. Rezazadeh Valojerdi M, Eftekhari-Yazd P, Karimian L, Hassani F, Movaghar B. Vitrification versus slow freezing gives excellent survival, post warming embryo morphology and pregnancy outcomes for human cleaved embryos. J Assist Reprod Genet. 2009;26(6):347-54.
  26. Mönkkönen KS, Aflatoonian R, Lee KF, Yeung WS, Tsao SW, Laitinen JT, et al. Localization and variable expression of G alpha(i2) in human endometrium and fallopian tubes. Hum Reprod. 2007;22(5):1224-30.
  27. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25(4):402-8.
  28. Herreros-Villanueva M, Chen CC, Tsai EM, Er TK. Endometriosis-associated ovarian cancer: What have we learned so far? Clin Chim Acta. 2019;493:63-72.
  29. Allhorn S, Böing C, Koch AA, Kimmig R, Gashaw I. TLR3 and TLR4 expression in healthy and diseased human endometrium. Reprod Biol Endocrinol. 2008;6:40.
  30. Kyama CM, Overbergh L, Mihalyi A, Meuleman C, Mwenda JM, Mathieu C, et al. Endometrial and peritoneal expression of aromatase, cytokines, and adhesion factors in women with endometriosis. Fertil Steril. 2008;89(2):301-10.
  31. Xu H, Schultze-Mosgau A, Agic A, Diedrich K, Taylor RN, Hornung D. Regulated upon activation, normal T cell expressed and secreted (RANTES) and monocyte chemotactic protein 1 in follicular fluid accumulate differentially in patients with and without endometriosis undergoing in vitro fertilization. Fertil Steril. 2006;86(6):1616-20.
  32. Bedaiwy M, Shahin AY, AbulHassan AM, Goldberg JM, Sharma RK, Agarwal A, et al. Differential expression of follicular fluid cytokines: relationship to subsequent pregnancy in IVF cycles. Reprod Biomed Online. 2007;15(3):321-5.
  33. Kats R, Collette T, Metz CN, Akoum A. Marked elevation of macrophage migration inhibitory factor in the peritoneal fluid of women with endometriosis. Fertil Steril. 2002;78(1):69-76.
  34. Morin M, Bellehumeur C, Therriault MJ, Metz C, Maheux R, Akoum A. Elevated levels of macrophage migration inhibitory factor in the peripheral blood of women with endometriosis. Fertil Steril. 2005;83(4):865-72.
  35. Carli C, Leclerc P, Metz CN, Akoum A. Direct effect of macrophage migration inhibitory factor on sperm function: possible involvement in endometriosis-associated infertility. Fertil Steril. 2007;88(4 Suppl):1240-7.
  36. Xu X, Wang B, Ye C, Yao C, Lin Y, Huang X, et al. Overexpression of macrophage migration inhibitory factor induces angiogenesis in human breast cancer. Cancer Lett. 2008;261(2):147-57.
  37. Beswick EJ, Reyes VE. Macrophage migration inhibitory factor and interleukin-8 produced by gastric epithelial cells during Helicobacter pylori exposure induce expression and activation of the epidermal growth factor receptor. Infect Immun. 2008;76(7):3233-40.
  38. Soni UK, Chadchan SB, Kumar V, Ubba V, Khan MTA, Vinod BSV, et al. A high level of TGF-B1 promotes endometriosis development via cell migration, adhesiveness, colonization, and invasiveness. Biol Reprod. 2018;100(4):917-38.
  39. Yang HL, Zhou WJ, Chang KK, Mei J, Huang LQ, Wang MY, et al. The crosstalk between endometrial stromal cells and macrophages impairs cytotoxicity of NK cells in endometriosis by secreting IL-10 and TGF-β. Reproduction. 2017;154(6):815-25.
  40. Young VJ, Brown JK, Saunders PT, Duncan WC, Horne AW. The peritoneum is both a source and target of TGF-β in women with endometriosis. PloS one. 2014;9(9):e106773.
  41. Young VJ, Ahmad S, Duncan WC, Horne AW. The role of TGF-β in the pathophysiology of peritoneal endometriosis. Hum Reprod Update. 2017;23(5):548-59.
  42. da Silveira JC, Winger QA, Bouma GJ, Carnevale EM. Effects of age on follicular fluid exosomal microRNAs and granulosa cell transforming growth factor-β signalling during follicle development in the mare. Reprod Fertil Dev. 2015;27(6):897-905.
Journal of Reproduction & Infertility
Volume 21, Issue 4
Autumn 2020
Pages 231-239