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Evaluation of Z-VAD-FMK as an anti-apoptotic drug to prevent granulosa cell apoptosis and follicular death after human ovarian tissue transplantation

  • Fertility Preservation
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Journal of Assisted Reproduction and Genetics Aims and scope Submit manuscript

Abstract

Purpose

To evaluate the efficiency of ovarian tissue treatment with Z-VAD-FMK, a broad-spectrum caspase inhibitor, to prevent follicle loss induced by ischemia/reperfusion injury after transplantation.

Methods

In vitro, granulosa cells were exposed to hypoxic conditions, reproducing early ischemia after ovarian tissue transplantation, and treated with Z-VAD-FMK (50 μM). In vivo, cryopreserved human ovarian fragments (n = 39) were embedded in a collagen matrix containing or not Z-VAD-FMK (50 μM) and xenotransplanted on SCID mice ovaries for 3 days or 3 weeks.

Results

In vitro, Z-VAD-FMK maintained the metabolic activity of granulosa cells, reduced HGL5 cell death, and decreased PARP cleavage. In vivo, no improvement of follicular pool and global tissue preservation was observed with Z-VAD-FMK in ovarian tissue recovered 3-days post-grafting. Conversely, after 3 weeks of transplantation, the primary follicular density was higher in fragments treated with Z-VAD-FMK. This improvement was associated with a decreased percentage of apoptosis in the tissue.

Conclusions

In situ administration of Z-VAD-FMK slightly improves primary follicular preservation and reduces global apoptosis after 3 weeks of transplantation. Data presented herein will help to guide further researches towards a combined approach targeting multiple cell death pathways, angiogenesis stimulation, and follicular recruitment inhibition.

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References

  1. Oktay K, Sonmezer M. Chemotherapy and amenorrhea: risks and treatment options. Curr Opin Obstet Gynecol. 2008;20(4):408–15. https://doi.org/10.1097/GCO.0b013e328307ebad.

    Article  PubMed  Google Scholar 

  2. Demeestere I, Simon P, Emiliani S, Delbaere A, Englert Y. Orthotopic and heterotopic ovarian tissue transplantation. Hum Reprod Update. 2009;15(6):649–65. https://doi.org/10.1093/humupd/dmp021.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  3. Demeestere I, Simon P, Dedeken L, Moffa F, Tsepelidis S, Brachet C, et al. Live birth after autograft of ovarian tissue cryopreserved during childhood. Hum Reprod. 2015;30(9):2107–9. https://doi.org/10.1093/humrep/dev128.

    Article  PubMed  Google Scholar 

  4. Jensen AK, Macklon KT, Fedder J, Ernst E, Humaidan P, Andersen CY. 86 successful births and 9 ongoing pregnancies worldwide in women transplanted with frozen-thawed ovarian tissue: focus on birth and perinatal outcome in 40 of these children. J Assist Reprod Genet. 2016;34:325–36. https://doi.org/10.1007/s10815-016-0843-9.

    Article  PubMed  PubMed Central  Google Scholar 

  5. Donnez J, Dolmans MM. Fertility preservation in women. N Engl J Med. 2017;377(17):1657–65. https://doi.org/10.1056/NEJMra1614676.

    Article  PubMed  Google Scholar 

  6. Van Eyck AS, Jordan BF, Gallez B, Heilier JF, Van Langendonckt A, Donnez J. Electron paramagnetic resonance as a tool to evaluate human ovarian tissue reoxygenation after xenografting. Fertil Steril. 2009;92(1):374–81.

    Article  PubMed  CAS  Google Scholar 

  7. Nugent D, Newton H, Gallivan L, Gosden RG. Protective effect of vitamin E on ischaemia-reperfusion injury in ovarian grafts. J Reprod Fertil. 1998;114(2):341–6.

    Article  PubMed  CAS  Google Scholar 

  8. Nisolle M, Casanas-Roux F, Qu J, Motta P, Donnez J. Histologic and ultrastructural evaluation of fresh and frozen-thawed human ovarian xenografts in nude mice. Fertil Steril. 2000;74(1):122–9.

    Article  PubMed  CAS  Google Scholar 

  9. Camboni A, Martinez-Madrid B, Dolmans MM, Nottola S, Van Langendonckt A, Donnez J. Autotransplantation of frozen-thawed ovarian tissue in a young woman: ultrastructure and viability of grafted tissue. Fertil Steril. 2008;90(4):1215–8. https://doi.org/10.1016/j.fertnstert.2007.08.084.

    Article  PubMed  Google Scholar 

  10. Israely T, Nevo N, Harmelin A, Neeman M, Tsafriri A. Reducing ischaemic damage in rodent ovarian xenografts transplanted into granulation tissue. Hum Reprod. 2006;21(6):1368–79.

    Article  PubMed  Google Scholar 

  11. Abir R, Fisch B, Jessel S, Felz C, Ben Haroush A, Orvieto R. Improving posttransplantation survival of human ovarian tissue by treating the host and graft. Fertil Steril. 2011;95(4):1205–10.

    Article  PubMed  Google Scholar 

  12. Shikanov A, Zhang Z, Xu M, Smith RM, Rajan A, Woodruff TK, et al. Fibrin encapsulation and vascular endothelial growth factor delivery promotes ovarian graft survival in mice. Tissue Eng Part A. 2011;17(23–24):3095–104. https://doi.org/10.1089/ten.TEA.2011.0204.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  13. Soleimani R, Heytens E, Oktay K. Enhancement of neoangiogenesis and follicle survival by sphingosine-1-phosphate in human ovarian tissue xenotransplants. PLoS One. 2011;6(4):e19475.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  14. Labied S, Delforge Y, Munaut C, Blacher S, Colige A, Delcombel R, et al. Isoform 111 of vascular endothelial growth factor (VEGF111) improves angiogenesis of ovarian tissue xenotransplantation. Transplantation. 2013;95(3):426–33. https://doi.org/10.1097/TP.0b013e318279965c.

    Article  PubMed  CAS  Google Scholar 

  15. Commin L, Buff S, Rosset E, Galet C, Allard A, Bruyere P, et al. Follicle development in cryopreserved bitch ovarian tissue grafted to immunodeficient mouse. Reprod Fertil Dev. 2012;24(3):461–71. https://doi.org/10.1071/RD11166.

    Article  PubMed  CAS  Google Scholar 

  16. Wang L, Ying YF, Ouyang YL, Wang JF, Xu J. VEGF and bFGF increase survival of xenografted human ovarian tissue in an experimental rabbit model. J Assist Reprod Genet. 2013;30(10):1301–11. https://doi.org/10.1007/s10815-013-0043-9.

    Article  PubMed  PubMed Central  Google Scholar 

  17. Henry L, Labied S, Fransolet M, Kirschvink N, Blacher S, Noel A, et al. Isoform 165 of vascular endothelial growth factor in collagen matrix improves ovine cryopreserved ovarian tissue revascularisation after xenotransplantation in mice. Reprod Biol Endocrinol. 2015;13:12. https://doi.org/10.1186/s12958-015-0015-2.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  18. Langbeen A, Ginneken CV, Fransen E, Bosmans E, Leroy JL, Bols PE. Morphometrical analysis of preantral follicular survival of VEGF-treated bovine ovarian cortex tissue following xenotransplantation in an immune deficient mouse model. Anim Reprod Sci. 2016;168:73–85. https://doi.org/10.1016/j.anireprosci.2016.02.029.

    Article  PubMed  CAS  Google Scholar 

  19. Kalogeris T, Baines CP, Krenz M, Korthuis RJ. Cell biology of ischemia/reperfusion injury. Int Rev Cell Mol Biol. 2012;298:229–317. https://doi.org/10.1016/B978-0-12-394309-5.00006-7.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  20. Liu J, Van der Elst J, Van den Broecke R, Dhont M. Early massive follicle loss and apoptosis in heterotopically grafted newborn mouse ovaries. Hum Reprod. 2002;17(3):605–11.

    Article  PubMed  Google Scholar 

  21. Yang H, Lee HH, Lee HC, Ko DS, Kim SS. Assessment of vascular endothelial growth factor expression and apoptosis in the ovarian graft: can exogenous gonadotropin promote angiogenesis after ovarian transplantation? Fertil Steril. 2008;90(4 Suppl):1550–8. https://doi.org/10.1016/j.fertnstert.2007.08.086.

    Article  PubMed  CAS  Google Scholar 

  22. Goldar S, Khaniani MS, Derakhshan SM, Baradaran B. Molecular mechanisms of apoptosis and roles in cancer development and treatment. Asian Pac J Cancer Prev. 2015;16(6):2129–44.

    Article  PubMed  Google Scholar 

  23. Yaoita H, Ogawa K, Maehara K, Maruyama Y. Attenuation of ischemia/reperfusion injury in rats by a caspase inhibitor. Circulation. 1998;97(3):276–81.

    Article  PubMed  CAS  Google Scholar 

  24. Himi T, Ishizaki Y, Murota S. A caspase inhibitor blocks ischaemia-induced delayed neuronal death in the gerbil. Eur J Neurosci. 1998;10(2):777–81.

    Article  PubMed  CAS  Google Scholar 

  25. Cursio R, Gugenheim J, Ricci JE, Crenesse D, Rostagno P, Maulon L, et al. Caspase inhibition protects from liver injury following ischemia and reperfusion in rats. Transpl Int. 2000;13(Suppl 1):S568–72.

    Article  PubMed  Google Scholar 

  26. Iwata A, Harlan JM, Vedder NB, Winn RK. The caspase inhibitor z-VAD is more effective than CD18 adhesion blockade in reducing muscle ischemia-reperfusion injury: implication for clinical trials. Blood. 2002;100(6):2077–80. https://doi.org/10.1182/blood-2002-03-0752.

    Article  PubMed  CAS  Google Scholar 

  27. Azuara D, Sola A, Hotter G, Calatayud L, de Oca J. Apoptosis inhibition plays a greater role than necrosis inhibition in decreasing bacterial translocation in experimental intestinal transplantation. Surgery. 2005;137(1):85–91. https://doi.org/10.1016/j.surg.2004.06.008.

    Article  PubMed  Google Scholar 

  28. Emamaullee JA, Stanton L, Schur C, Shapiro AM. Caspase inhibitor therapy enhances marginal mass islet graft survival and preserves long-term function in islet transplantation. Diabetes. 2007;56(5):1289–98. https://doi.org/10.2337/db06-1653.

    Article  PubMed  CAS  Google Scholar 

  29. Slee EA, Zhu H, Chow SC, MacFarlane M, Nicholson DW, Cohen GM. Benzyloxycarbonyl-Val-Ala-Asp (OMe) fluoromethylketone (Z-VAD.FMK) inhibits apoptosis by blocking the processing of CPP32. Biochem J. 1996;315(Pt 1):21–4.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  30. Henry L, Fransolet M, Labied S, Blacher S, Masereel MC, Foidart JM, et al. Supplementation of transport and freezing media with anti-apoptotic drugs improves ovarian cortex survival. J Ovarian Res. 2016;9(1):4. https://doi.org/10.1186/s13048-016-0216-0.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  31. Zhang JM, Li LX, Yang YX, Liu XL, Wan XP. Is caspase inhibition a valid therapeutic strategy in cryopreservation of ovarian tissue? J Assist Reprod Genet. 2009;26(7):415–20. https://doi.org/10.1007/s10815-009-9331-9.

    Article  PubMed  PubMed Central  Google Scholar 

  32. Shi FT, Cheung AP, Leung PC. Growth differentiation factor 9 enhances activin a-induced inhibin B production in human granulosa cells. Endocrinology. 2009;150(8):3540–6. https://doi.org/10.1210/en.2009-0267.

    Article  PubMed  CAS  Google Scholar 

  33. Fransolet M, Henry L, Labied S, Noël A, Nisolle M, Munaut C. In vitro evaluation of the anti-apoptotic drug Z-VAD-FMK on human ovarian granulosa cell lines for further use in ovarian tissue transplantation. J Assist Reprod Genet. 2015;32(10):1551–9. https://doi.org/10.1007/s10815-015-0536-9.

    Article  PubMed  PubMed Central  Google Scholar 

  34. Gosden RG, Baird DT, Wade JC, Webb R. Restoration of fertility to oophorectomized sheep by ovarian autografts stored at −196 degrees C. Hum Reprod. 1994;9(4):597–603.

    Article  PubMed  CAS  Google Scholar 

  35. Fransolet M, Henry L, Labied S, Masereel MC, Blacher S, Noel A, et al. Influence of mouse strain on ovarian tissue recovery after engraftment with angiogenic factor. J Ovarian Res. 2015;8(1):14. https://doi.org/10.1186/s13048-015-0142-6.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  36. Aubard Y, Piver P, Cogni Y, Fermeaux V, Poulin N, Driancourt MA. Orthotopic and heterotopic autografts of frozen-thawed ovarian cortex in sheep. Hum Reprod. 1999;14(8):2149–54.

    Article  PubMed  CAS  Google Scholar 

  37. Tilly JL, Kolesnick RN. Sphingolipids, apoptosis, cancer treatments and the ovary: investigating a crime against female fertility. Biochim Biophys Acta. 2002;1585(2–3):135–8.

    Article  PubMed  CAS  Google Scholar 

  38. Baird DT, Webb R, Campbell BK, Harkness LM, Gosden RG. Long-term ovarian function in sheep after ovariectomy and transplantation of autografts stored at −196 C. Endocrinology. 1999;140(1):462–71. https://doi.org/10.1210/en.140.1.462.

    Article  PubMed  CAS  Google Scholar 

  39. Rimon E, Cohen T, Dantes A, Hirsh L, Amit A, Lessing JB, et al. Apoptosis in cryopreserved human ovarian tissue obtained from cancer patients: a tool for evaluating cryopreservation utility. Int J Oncol. 2005;27(2):345–53.

    PubMed  CAS  Google Scholar 

  40. Fauque P, Ben Amor A, Joanne C, Agnani G, Bresson JL, Roux C. Use of trypan blue staining to assess the quality of ovarian cryopreservation. Fertil Steril. 2007;87(5):1200–7. https://doi.org/10.1016/j.fertnstert.2006.08.115.

    Article  PubMed  Google Scholar 

  41. Xiao Z, Wang Y, Li L, Li SW. Cryopreservation of the human ovarian tissue induces the expression of Fas system in morphologically normal primordial follicles. Cryo Letters. 2010;31(2):112–9.

    PubMed  Google Scholar 

  42. Hussein MR. Apoptosis in the ovary: molecular mechanisms. Hum Reprod Update. 2005;11(2):162–77. https://doi.org/10.1093/humupd/dmi001.

    Article  PubMed  CAS  Google Scholar 

  43. Isachenko V, Mallmann P, Petrunkina AM, Rahimi G, Nawroth F, Hancke K, et al. Comparison of in vitro- and chorioallantoic membrane (CAM)-culture systems for cryopreserved medulla-contained human ovarian tissue. PLoS One. 2012;7(3):e32549. https://doi.org/10.1371/journal.pone.0032549.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  44. Stroh C, Cassens U, Samraj AK, Sibrowski W, Schulze-Osthoff K, Los M. The role of caspases in cryoinjury: caspase inhibition strongly improves the recovery of cryopreserved hematopoietic and other cells. FASEB J. 2002;16(12):1651–3. https://doi.org/10.1096/fj.02-0034fje.

    Article  PubMed  CAS  Google Scholar 

  45. Gururajan R, Lahti JM, Kidd VJ. Molecular analysis of programmed cell death in mammals. In: Adolph KW, editor. Human genome methods. New York, CRC Press; 1998. p. 135–157.

  46. Roness H, Gavish Z, Cohen Y, Meirow D. Ovarian follicle burnout: a universal phenomenon? Cell Cycle. 2013;12(20):3245–6. https://doi.org/10.4161/cc.26358.

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  47. Silber S. Ovarian tissue cryopreservation and transplantation: scientific implications. J Assist Reprod Genet. 2016;33:1595–603. https://doi.org/10.1007/s10815-016-0814-1.

    Article  PubMed  PubMed Central  Google Scholar 

  48. Gavish Z, Spector I, Peer G, Schlatt S, Wistuba J, Roness H, et al. Follicle activation is a significant and immediate cause of follicle loss after ovarian tissue transplantation. J Assist Reprod Genet. 2017;35:61–9. https://doi.org/10.1007/s10815-017-1079-z.

    Article  PubMed  PubMed Central  Google Scholar 

  49. Kong HS, Kim SK, Lee J, Youm HW, Lee JR, Suh CS, et al. Effect of exogenous anti-Mullerian hormone treatment on cryopreserved and transplanted mouse ovaries. Reprod Sci. 2016;23(1):51–60. https://doi.org/10.1177/1933719115594021.

    Article  PubMed  CAS  Google Scholar 

  50. Ayuandari S, Winkler-Crepaz K, Paulitsch M, Wagner C, Zavadil C, Manzl C, et al. Follicular growth after xenotransplantation of cryopreserved/thawed human ovarian tissue in SCID mice: dynamics and molecular aspects. J Assist Reprod Genet. 2016;33:1585–93. https://doi.org/10.1007/s10815-016-0769-2.

    Article  PubMed  PubMed Central  Google Scholar 

  51. Hancke K, Walker E, Strauch O, Gobel H, Hanjalic-Beck A, Denschlag D. Ovarian transplantation for fertility preservation in a sheep model: can follicle loss be prevented by antiapoptotic sphingosine-1-phosphate administration? Gynecol Endocrinol. 2009;25(12):839–43. https://doi.org/10.3109/09513590903159524.

    Article  PubMed  CAS  Google Scholar 

  52. Martinez-Madrid B, Donnez J, Van Eyck AS, Veiga-Lopez A, Dolmans MM, Van Langendonckt A. Chick embryo chorioallantoic membrane (CAM) model: a useful tool to study short-term transplantation of cryopreserved human ovarian tissue. Fertil Steril. 2009;91(1):285–92. https://doi.org/10.1016/j.fertnstert.2007.11.026.

    Article  PubMed  Google Scholar 

  53. Lee JR, Youm HW, Kim SK, Jee BC, Suh CS, Kim SH. Effect of necrostatin on mouse ovarian cryopreservation and transplantation. Eur J Obstet Gynecol Reprod Biol. 2014;178:16–20. https://doi.org/10.1016/j.ejogrb.2014.04.040.

    Article  PubMed  CAS  Google Scholar 

  54. Gong JS, Kim GJ. The role of autophagy in the placenta as a regulator of cell death. Clin Exp Reprod Med. 2014;41(3):97–107. https://doi.org/10.5653/cerm.2014.41.3.97.

    Article  PubMed  PubMed Central  Google Scholar 

  55. Adhikari D, Liu K. mTOR signaling in the control of activation of primordial follicles. Cell Cycle. 2010;9(9):1673–4. https://doi.org/10.4161/cc.9.9.11626.

    Article  PubMed  CAS  Google Scholar 

  56. McLaughlin M, Patrizio P, Kayisli U, Luk J, Thomson TC, Anderson RA, et al. mTOR kinase inhibition results in oocyte loss characterized by empty follicles in human ovarian cortical strips cultured in vitro. Fertil Steril. 2011;96(5):1154–9 e1. https://doi.org/10.1016/j.fertnstert.2011.08.040.

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

The authors thank the CPMA-CHR Citadelle Liège for their contribution to follicular fluids collection. The authors are very grateful to M. C. Masereel and S. Schoenen for their help for follicular quantification. The authors acknowledge M. Dehuy, E. Konradowski, E. Feyereisen, and I. Dasoul for their excellent technical assistance.

The authors thank the Fonds de la Recherche Scientifique-FNRS (F.R.S.-FNRS, Télévie, Belgium; 7.4597.12-7.4622.14-7.4590.16), the Fondation contre le Cancer (foundation of public interest, Belgium), the Fonds spéciaux de la Recherche (University of Liège), the Centre Anticancéreux près l’Université de Liège, the Fonds Léon Fredericq (University of Liège), the Direction Générale Opérationnelle de l’Economie, de l’Emploi et de la Recherche from the Service Public de Wallonie (SPW, Belgium), the Interuniversity Attraction Poles Programme-Belgian Science Policy (Brussels, Belgium), the Plan National Cancer (Service Public Fédéral), and the Actions de Recherche Concertées (University of Liege, Belgium). The authors also thank the GIGA (Groupe Interdisciplinaire de Génoprotéomique Appliquée, University of Liege, Belgium) for the access to the GIGA-Imaging and Flow Cytometry platform and Dr. S. Ormenese and R. Stephan for their support with FACS analyses.

Author’s roles

MF performed experiments, interpreted data, and wrote the manuscript. LN contributed to in vitro experiments. LH and SL interpreted data and revised the manuscript. SB performed the computer images analysis. MN conceived and designed the study and corrected the manuscript. CM conceived and designed the study, interpreted data, corrected the manuscript, and substantially contributed to critical revisions. All authors read and approved the final manuscript.

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Authors and Affiliations

  1. Laboratory of Tumor and Developmental Biology, GIGA-R, University of Liège, Tour de Pathologie (B23), Avenue Hippocrate, 13, Sart Tilman, B-4000, Liège, Belgium

    Maïté Fransolet, Laure Noël, Silvia Blacher, Michelle Nisolle & Carine Munaut

  2. Department of Gynecology and Obstetrics, CPMA, University of Liège, CHR de la Citadelle, Boulevard du 12ième de Ligne, 1, B-4000, Liège, Belgium

    Laure Noël, Laurie Henry, Soraya Labied & Michelle Nisolle

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  1. Maïté Fransolet
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  2. Laure Noël
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  7. Carine Munaut
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Correspondence to Carine Munaut.

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Ethical approval

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted.

The use of human follicular fluid and ovarian tissue was approved by the Ethics Committee of the CHR Citadelle, University of Liège (CE412/1508 and CE412/1448, respectively). The use of the xenograft model was approved by the Animal Ethics Committee of the University of Liège (Reference 1304, October 2012).

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Informed consent was obtained from all individual participants included in the study.

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The authors declare that they have no conflict of interest.

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Fransolet, M., Noël, L., Henry, L. et al. Evaluation of Z-VAD-FMK as an anti-apoptotic drug to prevent granulosa cell apoptosis and follicular death after human ovarian tissue transplantation. J Assist Reprod Genet 36, 349–359 (2019). https://doi.org/10.1007/s10815-018-1353-8

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