3. Using gene expression as a method to quantify follicle survival in ovarian tissue cryopreservation

Nguyen Thi Thu Lan, Trương Nguyen Thai Ha, Ma Pham Que Mai, Nguyen Cao Tri, Nguyen Thi Ngoc Diem, Tran Bich Thu, Truong Dinh Kiet, Dang Quang Vinh, Nguyen Van Tien, Ho Manh Tuong

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Abstract

Ovarian tissue cryopreservation (OTC) is a promising technique for reproductive preservation in women with cancer. Although this technique was applied successfully in many Invitro Fertilization (IVF) centers worldwide, OTC has not been developed recently, and there is no publication of OTC in Vietnam. Using the expression of GDF-9 and caspase-3 as a biomarker to evaluate the survival of follicles in OTC in IVFMD at My Duc Hospital (NCT04666376) is the first study conducted in Vietnam. Ovarian tissues from each donor were divided into 3 groups, which group 1 was fresh tissues as a control group, group 2 was vitrified tissues by Ova kit Type M (Kitazato, Japan) and group 3 was vitrified tissues by IVFMD medium. Then, mRNA was extracted and transcripted reversely to cDNA before realtime PCR was performed. The findings showed there was no statistical difference between 3 groups about the expression of GDF-9 and caspase-3, respectively, [0.66, 1.38 in group 1 vs group 2] and [0.71, 1.08 in group 1 vs group 3], p > 0.05. The results aligned with previous findings because of the negative effects of cryopreservation. Moreover, it was illustrated that there was no change in tissue quality between the commercial medium and IVFMD’s medium in OTC.

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References

1. Kristensen SG, Rasmussen A, Byskov AG, et al. Isolation of pre-antral follicles from human ovarian medulla tissue. Hum Reprod. 2011;26:157-66.
2. Fabbri R, Vicenti R, Magnani V, et al. Ovarian tissue cryopreservation and transplantation: 20 years experience in Bologna University. Front. Endocrinol. (Lausanne). 2022;13:1035109. 10.3389/fendo. 2022.1035109
3. Arapaki A, Christopoulos P, Kalampokas E, et al. Ovarian tissue cryopreservation in children and adolescents. Child. (Basel) 9. 2022;1256. 10.3390/children9081256
4. Kagawa N, Silber S, Kuwayama M. Successful vitrification of bovine and human ovarian tissue. RBMOnline. 2009;18(4):568-577.
5. Wang T, Yan J, Lu C, et al. Human single follicle growth in invitro from cryopreserved ovarian tissue after slow freezing or vitrification. Hum Reprod. 2016;31(4):763-773.
6. Liu L, Liu B, Liu K, et al. Identification of biomarkers for predicting ovarian reserve of primordial follicle via transcriptomic analysis. Original research. 2022;13:879974. doi: 10. 3389/fgene.2022.879974
7. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PRC and the 2-∆∆Ct method. Methods. 2001;25(4):402-408.
8. Wang N, Li C-Y, Zhu H-B, et al. Effect of vitrification on the mRNA transcriptome of bovine oocytes. Reprod. Domest. Anim.. 2017;52:531-541. 
9. Chuaqui RF, Bonner RF, Best CJ, et al. Post-analysis follow-up of a microarray experiment. Nat Genet. 2002;32:509-514.
10. Zhang JM, Li LX, Yang YX, et al. Is caspase inhibition a valid therapeutic strategy in cryopreservation of ovarian tissue? J Assit Reprod Genet. 2009;26:415-420. doi: 10.1007/s10815-009-9331-9.
11. Men H, Agca T, Riley LK, et al. Improves survival of vitrified porcine embryos after partial delipidation through chemically stimulated lipolysis and inhibition of apoptosis. Theriogenology. 2006;2008-2016.
12. Shin SY, Lee JY, Lee EY. Protective effect of vascular endothelial growth factor (VEGF) in frozen-thawed granulosa cells is mediated by inhibition of apoptosis. Eur J Obstet Gynecol Reprod Biol. 2006;125:233-238.
13. Amorim CA, Dolmas MM, David A, et al. Vitrification and xenografting of human ovarian tissue. Fertil and Steril. 2012;98(5):1291-1298.
14. QIAGEN. AllPrep DNA/RNA/Protein Mini handbook. 2020 Nov. https://www.qiagen.com/us/resources/resourcedetail?id=e00044e1-a80 b-4c54-8989-957207ac8be2&lang=en
15. Glamoclija V, Vilovic K, Saraga-Babic M, et al. Apoptosis and active caspase-3 expression in hunam granulosa cells. Fertil and Steril. 2005;83(2):426-431. doi:10.1016/j.fertns tert.2004.06.075.
16. Hurst PR, Mora JM, Fenwick MA. Caspase-3, TUNEL and ultrastructural studies of small follicles in adult human ovarian biopsies. Hum Rerod. 2006;21(8):1974-1980. doi:10.1093/humrep/del109
17. Hariya M, Suzuki H. Incidence of apoptosis cells after vitrification in canine ovarian tissues. J. of Mammalian Ova Research. 2016;33(1):69-75. https://doi.org/10.1274/jmor. 33.69
18. Silver N, Cotroneo E, Proctor G, et al. Selection of housekeeping genes for gene expression studies in the adult rat submandibular gland under normal, inflamed, atrophic and regenerative states. BMC Mol Biol. 2008;9:64-72. doi: 10.1186/1471-2199-9-64
19. Butterfield DA, Hardass SS, Lange ML.  Oxidatively modified glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and Alzheimer’s disease: many pathways to neurodegeneration. J Alzheimers Dis. 2010;20(2):369-393. doi:10.3233/JAD-2010-13 75.
20. Fu J, Bia L, Zhao L, et al. Identification of genes for normalization of quantitative realtime PCR data in ovarian tissue. ABBS. 2010;42:568-574. https://doi.org/10.1093/abbs/gmq062
21. Kolkova Z, Arakelyan A, Casslen B, et al. Normalizing to GADPH jeopardies correct quantification of gene expression in ovarian tumours - IPO8 and RPL4 are reliable reference genes. J Ovarian Res. 2013;6(60):1-10. http://www.ovarianresearch.com/content/6/1/60
22. Kristensen SG, Kumar A, Mamsen LS, et al. Intrafollicular concentration of oocyte-secreted factors GDF-9 and BMP15 vary inversely in polycystic ovaries. J Clin Endocrinol Metab. 2022;107(8):e3374-e3383. doi:10.1210/clinem/dgac272
23. Aaltonen J, Laitinen MP, Vuojolainen RJ, et al. Human Growth Differentiation Factor 9 (GDF-9) and Its novel homolog GDF-9B are expressed in Oocytes during early folliculogenesis. The Journal of Clinical Endocrinology & Metabolism. 1999;84(8):2744-2750.
24. Van Nassauw L, Tao L, Harrisson F. Distribution of apoptosis-related proteins in the quail ovary during folliculogenesis: BCL-2, BAX and CPP32. Acta Histochem. 1999;101:103-12.
25. Abdollahi M, Salehnia M, Salehpour S, et al. Human ovarian tissue vitrification/ warming has minor effect on the expression of apoptosis-related genes. Iran Biomed J. 2013;17(4):179-186.
26. Wang Y, Xiao Z, Li L, et al. Novel needle immersed vitrification: a practical and convenient method with potential advantages in mouse and human ovarian tissue cryopreservation. Hum Reprod. 2008;23(10):2256-65.
27. Cheng J, Ruan X, Zhou Q, et al. Long-time low-temperature transportation of human ovarian tissue before cryopreservation. RBMO. 2021;43 2): 72-183. https://doi.org/10.1016/j.rb mo.2021.05.006