Comparıson of cumulus cells and follıcular fluıd obtaıned from ınfertile ındıvıduals dıagnosed wıth polycystıc ovary syndrome (PCOS) and endomethrıosıs wıth samples obtaıned from healthy ındıvıduals

Year 2023, , 576 - 586, 30.09.2023

Duygu Dayanır , Halil Ruso , Ziya Kalem , Rabia Tural , Gülistan Sanem Sarıbaş , Aylin Sepici Dinçel , Timur Gürgan , Candan Özoğul

https://doi.org/10.18663/tjcl.1339043

Abstract

Aim: Investigating the relationship between Growth differentiation factor-9 (GDF-9), Bone morphogenetic protein-15 (BMP-15) markers, apoptosis levels in cumulus cells and total oxidant (TOS)/ anti-oxidant (TAS) stress levels, inflammation parameters (interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-alpha)) in follicular fluid belonging to patients with polycystic ovary syndrome (PCOS), endometriosis (END) and male factor (MF) (control) groups.
Material and Methods: GDF-9 and BMP-15 markers are determined by immunohistochemical methods, apoptosis levels are studied with TUNEL. TOS and TAS statuses are investigated with spectrophotometry, IL-6 and TNF – alpha levels are examined by Enzyme-Linked Immuno Sorbent Assay (ELISA).
Results: According to the data obtained in the study; GDF-9 and BMP-15 levels are found to be lower in PCOS and END groups and apoptosis levels of cumulus cells were significantly higher at these groups . TOS levels were significantly higher in PCOS and END groups whereas follicular fluid TAS levels were not statistically significant for these groups. IL-6 and TNF – alpha levels of follicular fluid was significantly higher in PCOS. These parameters were also higher for END group, however the difference was not found to be significant.
Conclusion: Our results imply that correlation between GDF-9, BMP-15 markers, apotosis levels, oxidative status, inflammation levels may be interpreted with improper environment for oocyte maturation for patients diagnosed with PCOS or END.
The need for further studies on subject proceeds. However, if similar datas are obtained in further studies, it is thought that evaluation of cumulus cell properties together with especially follicular fluid oxidative stress levels will contribute to the selection of the best oocyte.

Keywords

cumulus cell, follicular fluid, polycystic ovary syndrome, endometriosis, oxidative stress

Polikistik over sendromu (PCOS) ve endometriozis tanili infertil bireylerden elde edilen kumulus hücreleri ve foliküler sivinin sağlikli bireylerden elde edilen örneklerle karşilaştirilmasi

Abstract

Amaç: Polikistik over sendromu (PKOS), endometriozis (END) ve erkek faktör (MF) (kontrol) gruplarında bulunan hastalara ait kumulus hücrelerinde Büyüme farklılaşma faktörü-9 (GDF-9), Kemik morfogenetik protein-15 (BMP-15) belirteçleri, apoptoz seviyeleri ile foliküler sıvı inflamasyon parametreleri (interlökin-6 (IL-6), tümör nekroz faktör alfa (TNF-alfa), total oksidan (TOS)/anti-oksidan (TAS) stres seviyeleri arasındaki ilişkinin araştırılması.
Gereç ve Yöntemler: Kumulus hücrelerinde büyüme farklılaşma faktörü-9 (GDF-9) ve kemik morfogenetik protein-15 (BMP-15) belirteçleri immünohistokimyasal yolla değerlendirilmiş olup; hücre ölümü TUNEL yöntemi kullanılarak araştırılmıştır. Folikül sıvısı örneklerinde toplam oksidatif stres (TOS) ve toplam anti-oksidan düzey (TAS) spektrofotometrik olarak araştırılmış, interlökin-6 (IL-6) ve tümör nekrozis faktör alfa (TNF-alfa) düzeyleri ELISA (Enzyme-Linked ImmunoSorbent Assay) yöntemi ile incelenmiştir
Bulgular: GDF-9 ve BMP-15 düzeyleri sağlıklı gruba kıyasla PCOS ve END gruplarında düşük seviyede saptanırken, hücre ölümüne ilişkin veriler bu gruplarda daha yüksek gözlenmiştir. Endometriozis grubunda GDF-9, BMP-15 değerleri en düşük, hücre ölümü düzeyleri ise en yüksek olarak bulunmuştur. Sağlıklı gruba kıyasla PCOS ve endometriozis gruplarında folikül sıvısı TOS düzeyleri istatistiksel olarak anlamlı yüksek bulunmuştur. Folikül sıvısı TAS düzeyleri ise sağlıklı gruba kıyasla PCOS ve endometriozis gruplarında daha yüksek bulunmuş ancak gruplar arasındaki fark istatistiksel olarak anlamlı bulunmamıştır.
Sonuç: Sonuçlarımız, GDF-9, BMP-15 belirteçleri, apotoz seviyeleri, oksidatif durum, inflamasyon seviyeleri arasındaki korelasyonun PCOS veya END tanılı hastalarda oosit olgunlaşması için uygun olmayan mikroçevre ile yorumlanabileceğini düşündürmektedir.
Konu ile ilgili ileri çalışmalara ihtiyaç devam etmektedir. İleri çalışmalarda benzer verilerin elde edilmesi halinde kumulus hücre özelliklerinin, özellikle foliküler sıvı oksidatif stres düzeyleri ile birlikte değerlendirilmesinin, oosit seçimine katkı sağlayacağı düşünülmektedir.

Keywords

kumulus hücresi, folikül sıvısı, polikistik over sendromu, endometriozis, oksidatif stres

References

• Gupta S, Ghulmiyyah J, Sharma R, Halabi J, Agarwal A. Power of proteomics in linking oxidative stress and female infertility. BioMed research international. 2014;2014.
• Chachamovich JR, Chachamovich E, Ezer H, Fleck MP, Knauth D, Passos EP. Investigating quality of life and health-related quality of life in infertility: a systematic review. Journal of Psychosomatic Obstetrics & Gynecology. 2010;31(2):101-10.
• Lin D, Ran J, Zhu S, Quan S, Ye B, Yu A, et al. Effect of GOLPH3 on cumulus granulosa cell apoptosis and ICSI pregnancy outcomes. Scientific Reports. 2017;7(1):7863.
• Raman RS, Chan PJ, Corselli JU, Patton WC, Jacobson JD, Chan SR, et al. Comet assay of cumulus cell DNA status and the relationship to oocyte fertilization via intracytoplasmic sperm injection. Human Reproduction. 2001;16(5):831-5.
• Zhang Y, Liu L, Yin T-L, Yang J, Xiong C-L. Follicular metabolic changes and effects on oocyte quality in polycystic ovary syndrome patients. Oncotarget. 2017;8(46):80472.
• Gleicher N, Kushnir VA, Darmon SK, Wang Q, Zhang L, Albertini DF, et al. Suspected ontogeny of a recently described hypo-androgenic PCOS-like phenotype with advancing age. Endocrine. 2018;59(3):661-76.
• Zagrodzki P, Krzyczkowska-Sendrakowska M, Nicol F, Wietecha-Posłuszny R, Milewicz T, Kryczyk-Kozioł J, et al. Selenium status parameters in patients with polycystic ovary syndrome. Journal of Trace Elements in Medicine and Biology. 2017;44:241-6.
• Artimani T, Karimi J, Mehdizadeh M, Yavangi M, Khanlarzadeh E, Ghorbani M, et al. Evaluation of pro-oxidant-antioxidant balance (PAB) and its association with inflammatory cytokines in polycystic ovary syndrome (PCOS). Gynecological endocrinology. 2018;34(2):148-52.
• Kolesnikova L, Kolesnikov S, Darenskaya M, Grebenkina L, Nikitina O, Lazareva L, et al. Activity of LPO processes in women with polycystic ovarian syndrome and infertility. Bulletin of experimental biology and medicine. 2017;162(3):320-2.
• Da Broi M, Giorgi V, Wang F, Keefe D, Albertini D, Navarro P. Influence of follicular fluid and cumulus cells on oocyte quality: clinical implications. Journal of assisted reproduction and genetics. 2018;35(5):735-51.
• Scutiero G, Iannone P, Bernardi G, Bonaccorsi G, Spadaro S, Volta CA, et al. Oxidative stress and endometriosis: a systematic review of the literature. Oxidative medicine and cellular longevity. 2017;2017.
• Park S, Lim W, Bazer FW, Song G. Apigenin induces ROS‐dependent apoptosis and ER stress in human endometriosis cells. Journal of cellular physiology. 2018;233(4):3055-65.
• Donabela FC, Meola J, Padovan CC, de Paz CCP, Navarro PA. Higher SOD1 gene expression in cumulus cells from infertile women with moderate and severe endometriosis. Reproductive Sciences. 2015;22(11):1452-60.
• Shaeib F, Khan SN, Ali I, Thakur M, Saed MG, Dai J, et al. The defensive role of cumulus cells against reactive oxygen species insult in metaphase II mouse oocytes. Reproductive Sciences. 2016;23(4):498-507.
• Liu S, Jiang L, Zhong T, Kong S, Zheng R, Kong F, et al. Effect of acrylamide on oocyte nuclear maturation and cumulus cells apoptosis in mouse in vitro. PloS one. 2015;10(8):e0135818.
• Sanfins A, Rodrigues P, Albertini DF. GDF-9 and BMP-15 direct the follicle symphony. Journal of assisted reproduction and genetics. 2018:1-10.
• Kawabe S, Yamashita Y, Saito N, Kokunai K, Hayashi A, Hayashi M, et al. The effect of moderate to severe endometriosis on expression of growth differentiation factor-9 mRNA in human granulosa cells under controlled ovarian hyperstimulation. Reproductive medicine and biology. 2015;14(4):179-84.
• ZHANG Y-m, LIN X-n, ZHOU F, WEI M-l, ZHANG S-y. Influence of Bone Morphogenetic Protein (BMP)-15 on the IVF Outcomes of Endometriosis——A Clinical Trail and Experimental Research. Reproduction & Contraception. 2012(11):5.
• McNatty KP, Juengel JL, Reader KL, Lun S, Myllymaa S, Lawrence SB, et al. Bone morphogenetic protein 15 and growth differentiation factor 9 co-operate to regulate granulosa cell function in ruminants. Reproduction. 2005;129(4):481-7.
• Peng J, Li Q, Wigglesworth K, Rangarajan A, Kattamuri C, Peterson RT, et al. Growth differentiation factor 9: bone morphogenetic protein 15 heterodimers are potent regulators of ovarian functions. Proceedings of the National Academy of Sciences. 2013;110(8):E776-E85.
• Juengel J, McNatty K. The role of proteins of the transforming growth factor-β superfamily in the intraovarian regulation of follicular development. Human reproduction update. 2005;11(2):144-61.
• Yoshino O, McMahon HE, Sharma S, Shimasaki S. A unique preovulatory expression pattern plays a key role in the physiological functions of BMP-15 in the mouse. Proceedings of the National Academy of Sciences. 2006;103(28):10678-83.
• Su Y-Q, Sugiura K, Wigglesworth K, O'Brien MJ, Affourtit JP, Pangas SA, et al. Oocyte regulation of metabolic cooperativity between mouse cumulus cells and oocytes: BMP15 and GDF9 control cholesterol biosynthesis in cumulus cells. Development. 2008;135(1):111-21.
• Pocar P, Nestler D, Risch M, Fischer B. Apoptosis in bovine cumulus–oocyte complexes after exposure to polychlorinated biphenyl mixtures during in vitro maturation. Reproduction. 2005;130(6):857-68.
• De La Fuente R, Eppig JJ. Transcriptional activity of the mouse oocyte genome: companion granulosa cells modulate transcription and chromatin remodeling. Developmental biology. 2001;229(1):224-36.
• Salehi E, Aflatoonian R, Moeini A, Yamini N, Asadi E, Khosravizadeh Z, et al. Apoptotic biomarkers in cumulus cells in relation to embryo quality in polycystic ovary syndrome. Archives of gynecology and obstetrics. 2017;296(6):1219-27.
• Agarwal A, Aponte-Mellado A, Premkumar BJ, Shaman A, Gupta S. The effects of oxidative stress on female reproduction: a review. Reproductive biology and endocrinology. 2012;10(1):49.
• Nakagawa K, Hisano M, Sugiyama R, Yamaguchi K. Measurement of oxidative stress in the follicular fluid of infertility patients with an endometrioma. Archives of gynecology and obstetrics. 2016;293(1):197-202.
• Luddi A, Capaldo A, Focarelli R, Gori M, Morgante G, Piomboni P, et al. Antioxidants reduce oxidative stress in follicular fluid of aged women undergoing IVF. Reproductive Biology and Endocrinology. 2016;14(1):57.
• Agarwal A, Gupta S, Sharma RK. Role of oxidative stress in female reproduction. Reproductive biology and endocrinology. 2005;3(1):28.
• Yan C, Wang P, DeMayo J, DeMayo FJ, Elvin JA, Carino C, et al. Synergistic roles of bone morphogenetic protein 15 and growth differentiation factor 9 in ovarian function. Molecular endocrinology. 2001;15(6):854-66.
• Karagül Mİ, Aktaş S, YILMAZ BC, Yilmaz M, Temel GÖ. GDF9 and BMP15 expressions and fine structure changes during folliculogenesis in polycystic ovary syndrome. Balkan Medical Journal. 2018;35(1):43-54.
• Zhao S-Y, Qiao J, Chen Y-J, Liu P, Li J, Yan J. Expression of growth differentiation factor-9 and bone morphogenetic protein-15 in oocytes and cumulus granulosa cells of patients with polycystic ovary syndrome. Fertility and sterility. 2010;94(1):261-7.
• Hreinsson JG, Scott JE, Rasmussen C, Swahn ML, Hsueh AJ, Hovatta O. Growth differentiation factor-9 promotes the growth, development, and survival of human ovarian follicles in organ culture. The Journal of Clinical Endocrinology & Metabolism. 2002;87(1):316-21.
• Vireque A, Santana L, Moreno D, Ferriani R, Scrideli C, Reis R. Single-cell expression analysis of BMP15 and GDF9 in mature oocytes and BMPR2 in cumulus cells of women with polycystic ovary syndrome undergoing controlled ovarian hyperstimulation. Journal of assisted reproduction and genetics. 2012;29(10):1057-65.
• Juengel JL, Hudson NL, Heath DA, Smith P, Reader KL, Lawrence SB, et al. Growth differentiation factor 9 and bone morphogenetic protein 15 are essential for ovarian follicular development in sheep. Biology of reproduction. 2002;67(6):1777-89.
• Galloway SM, McNatty KP, Cambridge LM, Laitinen MP, Juengel JL, Jokiranta TS, et al. Mutations in an oocyte-derived growth factor gene (BMP15) cause increased ovulation rate and infertility in a dosage-sensitive manner. Nature genetics. 2000;25(3):279.
• Su Y-Q, Wu X, O'Brien MJ, Pendola FL, Denegre JN, Matzuk MM, et al. Synergistic roles of BMP15 and GDF9 in the development and function of the oocyte–cumulus cell complex in mice: genetic evidence for an oocyte–granulosa cell regulatory loop. Developmental biology. 2004;276(1):64-73.
• de Resende LOT, Vireque AA, Santana LF, Moreno DA, de Sá Rosa e Silva ACJ, Ferriani RA, et al. Single-cell expression analysis of BMP15 and GDF9 in mature oocytes and BMPR2 in cumulus cells of women with polycystic ovary syndrome undergoing controlled ovarian hyperstimulation. Journal of assisted reproduction and genetics. 2012;29:1057-65.
• Cui X, Jing X, Wu X, Bi X, Liu J, Long Z, et al. Abnormal expression levels of BMP15/Smad1 are associated with granulosa cell apoptosis in patients with polycystic ovary syndrome. Molecular medicine reports. 2017;16(6):8231-6.
• Song W-J, Shi X, Zhang J, Chen L, Fu S-X, Ding Y-L. Akt-mTOR signaling mediates abnormalities in the proliferation and apoptosis of ovarian granulosa cells in patients with polycystic ovary syndrome. Gynecologic and obstetric investigation. 2018;83(2):124-32.
• Ding L, Gao F, Zhang M, Yan W, Tang R, Zhang C, et al. Higher PDCD4 expression is associated with obesity, insulin resistance, lipid metabolism disorders, and granulosa cell apoptosis in polycystic ovary syndrome. Fertility and sterility. 2016;105(5):1330-7. e3.
• Nasiri N, Moini A, Eftekhari-Yazdi P, Karimian L, Salman-Yazdi R, Arabipoor A. Oxidative stress statues in serum and follicular fluid of women with endometriosis. Cell Journal (Yakhteh). 2017;18(4):582.
• Choi YS, Cho S, Seo SK, Park JH, Kim SH, Lee BS. Alteration in the intrafollicular thiol-redox system in infertile women with endometriosis. Reproduction. 2015;149(2):155-62.
• Rudnicka E, Duszewska AM, Kucharski M, Tyczyński P, Smolarczyk R. Oxidative stress and reproductive function: Oxidative stress in polycystic ovary syndrome. Reproduction. 2022; 164(6):145-154
• Zaha I, Muresan M, Tulcan C, Huniadi A, Naghi P, Sandor M, Tripon R, Gaspar C, Klaudia-Melinda M, Sachelarie L, Stefan L. The Role of Oxidative Stress in Infertility. J Pers Med. 2023;13(8):1264..
• Goud PT, Goud AP, Joshi N, Puscheck E, Diamond MP, Abu-Soud HM. Dynamics of nitric oxide, altered follicular microenvironment, and oocyte quality in women with endometriosis. Fertility and sterility. 2014;102(1):151-9. e5.
• Hussein TS, Froiland DA, Amato F, Thompson JG, Gilchrist RB. Oocytes prevent cumulus cell apoptosis by maintaining a morphogenic paracrine gradient of bone morphogenetic proteins. Journal of cell science. 2005;118(22):5257-68.