Skip to main content

Advertisement

Log in

Altered FoxO3 expression and apoptosis in granulosa cells of women with polycystic ovary syndrome

  • Gynecologic Endocrinology and Reproductive Medicine
  • Published:
Archives of Gynecology and Obstetrics Aims and scope Submit manuscript

Abstract

Purpose

To determine the level of apoptosis, and alteration of FoxO3 (forkhead box O3 transcription factor) expression and phosphorylation in human granulosa cells amongst polycystic ovary syndrome (PCOS) patients and control group.

Methods

We recruited infertile women with PCOS (n = 14) and compared them with infertile women due to tubal blockage or male factor infertility (n = 14, controls). GnRH agonist and gonadotropins were used for ovarian stimulation. Follicular fluids from large follicles (>16 mm) were pooled and granulosa cells (GCs) were isolated using cell strainer methodology. Apoptosis of purified GCs was measured by flow cytometry using Annexin V and propidium iodide. Quantitative real-time PCR and western blotting were performed to assess alteration of FoxO3 expression and phosphorylation in GCs.

Results

There were higher percentages of early and late apoptosis in GCs of PCOS patients than in the control group. FoxO3 mRNA level and total FoxO3 protein were significantly higher in PCOS group than in the control group. The ratio of p-FoxO3/total FoxO3 decreased significantly in PCOS than in the control group. It was inferred that unphosphorylated (active form) FoxO3 was higher in GCs of PCOS patients. Apoptosis was significantly and positively correlated with the total FoxO3 and negatively correlated with the p-FoxO3 protein levels in PCOS patients.

Conclusions

Activation and overexpression of FoxO3 in granulosa cells of PCOS women correlated with higher apoptosis levels in these cells suggesting that FoxO3 may be a candidate for the higher apoptosis in granulosa cells from women with PCOS.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Ehrmann DA (2005) Polycystic ovary syndrome. New Engl J Med 352:1223–1236

    Article  CAS  PubMed  Google Scholar 

  2. Moran L, Teede H (2009) Metabolic features of the reproductive phenotypes of polycystic ovary syndrome. Hum Reprod Update 15:477–488

    Article  CAS  PubMed  Google Scholar 

  3. Köninger A, Koch L, Edimiris P, Enekwe A, Nagarajah J, Kasimir-Bauer S, Kimmig R, Strowitzki T, Schmidt B (2014) Anti-Mullerian Hormone: an indicator for the severity of polycystic ovarian syndrome. Arch Gynecol Obstet 290:1023–1030

    Article  PubMed  Google Scholar 

  4. Honnma H, Endo T, Henmi H, Nagasawa K, Baba T, Yamazaki K, Kitajima Y, Hayashi T, Manase K, Saito T (2006) Altered expression of Fas/Fas ligand/caspase 8 and membrane type 1-matrix metalloproteinase in atretic follicles within dehydroepiandrosterone-induced polycystic ovaries in rats. Apoptosis 11:1525–1533

    Article  CAS  PubMed  Google Scholar 

  5. Yu YS, Sui HS, Han ZB, Wei L, Luo MJ, Tan JH (2004) Apoptosis in granulosa cells during follicular atresia: relationship with steroids and insulin-like growth factors. Cell Res 14:341–346

  6. Maeda A, Inoue N, Matsuda-Minehata F, Goto Y, Cheng Y, Manabe N (2007) The role of interleukin-6 in the regulation of granulosa cell apoptosis during follicular atresia in pig ovaries. J Reprod Dev 53:481–490

    Article  CAS  PubMed  Google Scholar 

  7. Dijkers PF, Birkenkamp KU, Lam EW-F, Thomas NSB, Lammers J-WJ, Koenderman L, Coffer PJ (2002) FKHR-L1 can act as a critical effector of cell death induced by cytokine withdrawal protein kinase B—enhanced cell survival through maintenance of mitochondrial integrity. J Cell Biol 156:531–542

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Stahl M, Dijkers PF, Kops GJ, Lens SM, Coffer PJ, Burgering BM, Medema RH (2002) The forkhead transcription factor FoxO regulates transcription of p27Kip1 and Bim in response to IL-2. J Immunol 168:5024–5031

    Article  CAS  PubMed  Google Scholar 

  9. Accili D, Arden KC (2004) FoxOs at the crossroads of cellular metabolism, differentiation, and transformation. Cell 117:421–426

    Article  CAS  PubMed  Google Scholar 

  10. Obexer P, Geiger K, Ambros P, Meister B, Ausserlechner M (2007) FKHRL1-mediated expression of Noxa and Bim induces apoptosis via the mitochondria in neuroblastoma cells. Cell Death Differ 14:534–547

    Article  CAS  PubMed  Google Scholar 

  11. Suh CS, Jee BC, Choi YM, Kim JG, Lee JY, Moon SY, Kim SH (2002) Prognostic implication of apoptosis in human luteinized granulosa cells during IVF–ET. J Assist Reprod Gen 19:209–214

    Article  Google Scholar 

  12. Urman B, Tiras B, Yakin K (2004) Assisted reproduction in the treatment of polycystic ovarian syndrome. Reprod Biomed Online 8:419–430

    Article  PubMed  Google Scholar 

  13. Iavazzo C, Vitoratos N (2010) Polycystic ovarian syndrome and pregnancy outcome. Arch Gynecol Obstet 282:235–239

    Article  CAS  PubMed  Google Scholar 

  14. Castrillon DH, Miao L, Kollipara R, Horner JW, DePinho RA (2003) Suppression of ovarian follicle activation in mice by the transcription factor Foxo3a. Science 301:215–218

    Article  CAS  PubMed  Google Scholar 

  15. Rotterdam ESHRE/ASRM sponsored consensus workshop group (2004) Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome (PCOS). Hum Reprod 19:41–47

    Article  Google Scholar 

  16. Nikolettos N, Kupker W, Al-Hasani S, Demirel L, Schöpper B, Sturm R, Diedrich K (2000) ICSI outcome in patients of 40 years age and over: a retrospective analysis. Eur J Obstet Gyn R B 91:177–182

    Article  CAS  Google Scholar 

  17. Catteau-Jonard S, Jamin SP, Leclerc A, Gonzalès J, Dewailly D, di Clemente N (2008) Anti-Mullerian hormone, its receptor, FSH receptor, and androgen receptor genes are overexpressed by granulosa cells from stimulated follicles in women with polycystic ovary syndrome. J Clin Endocrinol Metab 93:4456–4461

    Article  CAS  PubMed  Google Scholar 

  18. Smith LP, Nierstenhoefer M, Yoo SW, Penzias AS, Tobiasch E, Usheva A (2009) The bile acid synthesis pathway is present and functional in the human ovary. PLoS One 4:e7333

    Article  PubMed  PubMed Central  Google Scholar 

  19. Wolffe AP, Tata JR (1984) Primary culture, cellular stress and differentiated function. Febs L 176:8–15

    Article  CAS  Google Scholar 

  20. Ferrero H, Delgado-Rosas F, Garcia-Pascual CM, Monterde M, Zimmermann RC, Simón C, Pellicer A, Gómez R (2012) Efficiency and purity provided by the existing methods for the isolation of luteinized granulosa cells: a comparative study. Hum Reprod 27:1781–1789

    Article  CAS  PubMed  Google Scholar 

  21. Fedorcsák P, Ráki M, Storeng R (2007) Characterization and depletion of leukocytes from cells isolated from the pre-ovulatory ovarian follicle. Hum Reprod 22:989–994

    Article  PubMed  Google Scholar 

  22. Figenschau Y, Sundsfjord J, Yousef M, Fuskevåg O, Sveinbjörnsson B, Bertheussen K (1997) A simplified serum-free method for preparation and cultivation of human granulosa-luteal cells. Hum Reprod 12:523–531

    Article  CAS  PubMed  Google Scholar 

  23. Al-Gubory KH, Fowler PA, Garrel C (2010) The roles of cellular reactive oxygen species, oxidative stress and antioxidants in pregnancy outcomes. Int J Biochem Cell Biol 42:1634–1650

    Article  CAS  PubMed  Google Scholar 

  24. Billig H, Furuta I, Hsueh A (1993) Estrogens inhibit and androgens enhance ovarian granulosa cell apoptosis. Endocrinol 133:2204–2212

    CAS  Google Scholar 

  25. Kaneko T, Saito H, Takahashi T, Ohta N, Saito T, Hiroi M (2000) Effects of controlled ovarian hyperstimulation on oocyte quality in terms of the incidence of apoptotic granulosa cells. J Assist Reprod Gen 17:580–585

    Article  CAS  Google Scholar 

  26. Karuputhula NB, Chattopadhyay R, Chakravarty B, Chaudhury K (2013) Oxidative status in granulosa cells of infertile women undergoing IVF. Syst Biol Reprod Med 59:91–98

    Article  CAS  PubMed  Google Scholar 

  27. Nisenblat V, Norman RJ (2009) Androgens and polycystic ovary syndrome. Curr Opin Endocrinol Diabetes Obes 16:224–231

    Article  CAS  PubMed  Google Scholar 

  28. Balen AH, Tan SL, Jacobs HS (1993) Hypersecretion of luteinising hormone: a significant cause of infertility and miscarriage. BJOG 100:1082–1089

    Article  CAS  Google Scholar 

  29. Nakahara K, Saito H, Saito T, Ito M, Ohta N, Takahashi T, Hiroi M (1997) The incidence of apoptotic bodies in membrana granulosa can predict prognosis of ova from patients participating in in vitro fertilization programs. Fertil Steril 68:312–317

    Article  CAS  PubMed  Google Scholar 

  30. Zhang J, Zhu G, Wang X, Xu B, Hu L (2007) Apoptosis and expression of protein TRAIL in granulosa cells of rats with polycystic ovarian syndrome. J Huazhong Univ Sci Technol 27:311–314

    Article  CAS  Google Scholar 

  31. Onalan G, Selam B, Baran Y, Cincik M, Onalan R, Gündüz U, Ural AU, Pabuccu R (2005) Serum and follicular fluid levels of soluble Fas, soluble Fas ligand and apoptosis of luteinized granulosa cells in PCOS patients undergoing IVF. Hum Reprod 20:2391–2395

    Article  CAS  PubMed  Google Scholar 

  32. Das M, Djahanbakhch O, Hacihanefioglu B, Saridogan E, Ikram M, Ghali L, Raveendran M, Storey A (2008) Granulosa cell survival and proliferation are altered in polycystic ovary syndrome. J Clin Endocrinol Metab 93:881–887

    Article  CAS  PubMed  Google Scholar 

  33. Almahbobi G, Anderiesz C, Hutchinson P, McFarlane JR, Wood C, Trounson AO (1996) Functional integrity of granulosa cells from polycystic ovaries. Clin Endocrinol 44:571–580

    Article  CAS  Google Scholar 

  34. Wang X-L, Wu Y, Tan L-B, Tian Z, Liu J-H, Zhu D-S, Zeng S-M (2012) Follicle-stimulating hormone regulates pro-apoptotic protein Bcl-2-interacting mediator of cell death-extra long (BimEL)-induced porcine granulosa cell apoptosis. J Biol Chem 287:10166–10177

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Matsuda F, Inoue N, Maeda A, Cheng Y, Sai T, Gonda H, Goto Y, Sakamaki K, Manabe N (2011) Expression and function of apoptosis initiator FOXO3 in granulosa cells during follicular atresia in pig ovaries. J Reprod Develop 57:151–158

    Article  CAS  Google Scholar 

  36. Inoue N, Maeda A, Matsuda-Minehata F, Fukuta K, Manabe N (2006) Expression and localization of Fas ligand and Fas during atresia in porcine ovarian follicles. J Reprod Develop 52:723–730

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The present study was funded by Deputy Ministry for Research, Tehran University of Medical Sciences; Grant No. 91-03-30-19007. We would like to thank Dr. Ensieh Tehraninejad (Vali-e-Asr Reproductive Health Research Centre, Tehran University of Medical Sciences, Tehran, Iran) and Parisa Hayat (Cellular and Molecular Research Center, Iran University of Medical Sciences, Tehran, Iran) for their kind help with the present research.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Mehdi Abbasi.

Ethics declarations

Conflict of interest

The authors declare that they have no competing interests associated with this publication.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Mikaeili, S., Rashidi, B.H., Safa, M. et al. Altered FoxO3 expression and apoptosis in granulosa cells of women with polycystic ovary syndrome. Arch Gynecol Obstet 294, 185–192 (2016). https://doi.org/10.1007/s00404-016-4068-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00404-016-4068-z

Keywords

Navigation