Volume 5, Issue 1 (Feb 2017)                   Res Mol Med (RMM) 2017, 5(1): 27-33 | Back to browse issues page

XML Print

Download citation:
BibTeX | RIS | EndNote | Medlars | ProCite | Reference Manager | RefWorks
Send citation to:

Shahedi A, Hosseini A, Ali Khalili M, Yeganeh F. Vitrification Affects Nuclear Maturation and Gene Expression of Immature Human Oocytes. Res Mol Med (RMM) 2017; 5 (1) :27-33
URL: http://rmm.mazums.ac.ir/article-1-225-en.html
1- Department of Biology and Anatomical Sciences, Shahid Sadoughi University of Medical Sciences , Yazd, Iran
2- Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
3- Research and Clinical Center for Infertility, Shahid Sadoughi University of Medical Sciences, Yazd, Iran
4- Department of Immunology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran. , fyeganeh@sbmu.ac.ir
Abstract:   (6245 Views)

Background: Vitrification of oocytes is a fast-freezing technique, which may affect the quality of the human oocyte, and consequently affects the embryo development, pregnancy and birth. The aim of the current study was to investigate the consequence of in-vitro vitrification on maturation status of immature human oocytes, additionally, expression levels of stress, and apoptosis related genes.

Materials and Methods: The total of 213 human immature oocytes which routinely discarded from assisted reproduction clinics were collected and divided into two groups including: (I) fresh germinal vesicle (GV) oocytes (n=106) (matured in-vitro  (fIVM) , and  (II) GV oocytes (n=107) that initially vitrified, then matured in  in-vitro (vIVM). After 36 hours of incubation, the oocytes were evaluated for nuclear maturation and expression level of DNA methyltransferase (DNMT1), stress related genes (Sod1 and Hsp70), and apoptotic related genes (Bax and Bcl-2) by quantitative Real-Time PCR.

Results: Oocyte maturation rates were reduced in vIVM compared to fIVM oocytes (P=0.001). The expression of stress (Sod1 and Hsp70), and apoptotic-related genes (Bax and Bcl-2) in vIVM were significantly higher compared to the fIVM group. Additionally, pro-apoptotic gene up-regulated 4.3 times more than anti-apoptotic gene in vIVM oocyte. However, DNMT1 gene expression was reduced in vIVM oocyte (P = 0.047).

Conclusions: The low survival rate of vitrified In-vitro matured GV oocytes could definitely be explained by the alterations of their gene expression profile. 

Full-Text [PDF 276 kb]   (2278 Downloads)    
Type of Study: Research | Subject: Molecular biology
Published: 2017/05/3

1. Wang H, Racowsky C, Combelles CMH. Is it best to cryopreserve human cumulus-free immature oocytes before or after in vitro maturation? Cryobiology. 2012; 65(2):79-87. PMID: 22691237 [DOI:10.1016/j.cryobiol.2012.06.001]
2. González C, Devesa M, Boada M, Coroleu B, Veiga A, Barri PN. Combined strategy for fertility preservation in an oncologic patient: vitrification of in vitro matured oocytes and ovarian tissue freezing. J Assist Reprod Genet. 2011; 28(12):1147-9. PMID: 21887506 [DOI:10.1007/s10815-011-9628-3]
3. Borini A, Bianchi V. Cryopreservation of mature and immature oocytes. Clin Obstet Gynecol. 2010; 53(4):763-74. [DOI:10.1097/GRF.0b013e3181f96f01]
4. Walls M, Junk S, Ryan JP, Hart R. IVF versus ICSI for the fertilization of in-vitro matured human oocytes. Reprod Biomed Online. 2012; 25(6):603-7. PMID: 23063820 [DOI:10.1016/j.rbmo.2012.08.001]
5. Azari M, Kafi M, Ebrahimi B, Fatehi R, Jamalzadeh M. Oocyte maturation, embryo development and gene expression following two different methods of bovine cumulus-oocyte complexes vitrification. Vet Res Commun. 2017; 41(1):49-56. PMID: 27943152 [DOI:10.1007/s11259-016-9671-8]
6. Men H, Monson RL, Parrish JJ, Rutledge JJ. Degeneration of cryopreserved bovine oocytes via apoptosis during subsequent culture. Cryobiology. 2003; 47(1):73-81. PMID: 12963414 [DOI:10.1016/S0011-2240(03)00070-1]
7. Uysal F, Akkoyunlu G, Ozturk S. Dynamic expression of DNA methyltransferases (DNMTs) in oocytes and early embryos. Biochimie. 2015; 116:103-13. PMID: 26143007 [DOI:10.1016/j.biochi.2015.06.019]
8. Kurihara Y, Kawamura Y, Uchijima Y, Amamo T, Kobayashi H, Asano T, et al. Maintenance of genomic methylation patterns during preimplantation development requires the somatic form of DNA methyltransferase 1. Dev Biol. 2008; 313(1):335-46. PMID: 26143007 [DOI:10.1016/j.ydbio.2007.10.033]
9. Petrussa L, Van de Velde H, De Rycke M. Dynamic regulation of DNA methyltransferases in human oocytes and preimplantation embryos after assisted reproductive technologies. Mol Hum Reprod. 2014; 20(9):861-74. PMID: 24994815 [DOI:10.1093/molehr/gau049]
10. Shi W, Haaf T. Aberrant methylation patterns at the two-cell stage as an indicator of early developmental failure. Mol Reprod Dev. 2002; 63(3):329-34. PMID: 12237948 [DOI:10.1002/mrd.90016]
11. El Mouatassim S, Guérin P, Ménézo Y. Expression of genes encoding antioxidant enzymes in human and mouse oocytes during the final stages of maturation. Mol Hum Reprod. 1999; 5(8):720-5. PMID: 10421798 [DOI:10.1093/molehr/5.8.720]
12. Blomberg LA, Long EL, Sonstegard TS, Van Tassell CP, Dobrinsky JR, Zuelke KA. Serial analysis of gene expression during elongation of the peri-implantation porcine trophectoderm (conceptus). Physiol Genomics. 2005; 20(2):188-94. PMID: 15536174 [DOI:10.1152/physiolgenomics.00157.2004]
13. Dai J, Wu C, Muneri CW, Niu Y, Zhang S, Rui R, et al. Changes in mitochondrial function in porcine vitrified MII-stage oocytes and their impacts on apoptosis and developmental ability. Cryobiology. 2015; 71(2):291-8. PMID: 26247316 [DOI:10.1016/j.cryobiol.2015.08.002]
14. Shahedi A, Hosseini A, Khalili MA, Norouzian M, Salehi M, Piriaei A, et al. The effect of vitrification on ultrastructure of human in vitro matured germinal vesicle oocytes. Eur J Obstet Gynecol Reprod Biol. 2013; 167(1):69-75. PMID: 23260597 [DOI:10.1016/j.ejogrb.2012.11.006]
15. Fasano G, Demeestere I, Englert Y. In-vitro maturation of human oocytes: before or after vitrification? J Assist Reprod Genet. 2012; 29(6):507-12. PMID: 22476503 [DOI:10.1007/s10815-012-9751-9]
16. Khalili MA, Maione M, Palmerini MG, Bianchi S, Macchiarelli G, Nottola SA. Ultrastructure of human mature oocytes after vitrification. Eur J Histochem. 2012; 56(3):e38. PMID: 23027354 [DOI:10.4081/ejh.2012.e38]
17. Vajta G. Vitrification of the oocytes and embryos of domestic animals. Anim Reprod Sci. 2000; 60-61:357-64. PMID: 10844207 [DOI:10.1016/S0378-4320(00)00097-X]
18. Hyttel P, Vajta G, Callesen H. Vitrification of bovine oocytes with the open pulled straw method: ultrastructural consequences. Mol Reprod Dev. 2000; 56(1):80-8. PMID: 10737970 https://doi.org/10.1002/(SICI)1098-2795(200005)56:1<80::AID-MRD10>3.0.CO;2-U [DOI:10.1002/(SICI)1098-2795(200005)56:13.0.CO;2-U]
19. Moussa M, Shu J, Zhang X, Zeng F. Cryopreservation of mammalian oocytes and embryos: current problems and future perspectives. Sci China Life Sci. 2014; 57(9):903-14. PMID: 25104318 [DOI:10.1007/s11427-014-4689-z]
20. Cao Y, Xing Q, Zhang Z-G, Wei Z-L, Zhou P, Cong L. Cryopreservation of immature and in-vitro matured human oocytes by vitrification. Reprod Biomed Online. 2009; 19(3):369-73. PMID: 19778481 [DOI:10.1016/S1472-6483(10)60170-8]
21. Son WY, Park SE, Lee KA, Lee WS, Ko JJ, Yoon TK, et al. Effects of 1,2-propanediol and freezing-thawing on the in vitro developmental capacity of human immature oocytes. Fertil Steril. 1996; 66(6):995-9. PMID: 8941068 [DOI:10.1016/S0015-0282(16)58696-8]
22. Combelles CMH, Cekleniak NA, Racowsky C, Albertini DF. Assessment of nuclear and cytoplasmic maturation in in-vitro matured human oocytes. Hum Reprod. 2002; 17(4):1006-16. PMID: 11925398 [DOI:10.1093/humrep/17.4.1006]
23. Baust JM, Van Buskirk, Baust JG. Cell viability improves following inhibition of cryopreservation-induced apoptosis. In Vitro Cell Dev Biol Anim. 2000; 36(4):262-70. PMID: 10852352 https://doi.org/10.1290/1071-2690(2000)036<0262:CVIFIO>2.0.CO;2 https://doi.org/10.1290/1071-2690(2000)036<0262:CVIFIO>2.3.CO;2 [DOI:10.1290/1071-2690(2000)0362.0.CO;2]
24. Dhali A, Anchamparuthy VM, Butler SP, Pearson RE, Mullarky IK, Gwazdauskas FC. Gene expression and development of mouse zygotes following droplet vitrification. Theriogenology. 2007; 68(9):1292-8. PMID: 17915304 [DOI:10.1016/j.theriogenology.2007.08.030]
25. Anchamparuthy VM, Pearson RE, Gwazdauskas FC. Expression pattern of apoptotic genes in vitrified-thawed bovine oocytes. Reprod Domest Anim. 2010; 45(5):e83-90. PMID: 19821945
26. Tatemoto H, Sakurai N, Muto N. Protection of porcine oocytes against apoptotic cell death caused by oxidative stress during In vitro maturation: role of cumulus cells. Biol Reprod. 2000; 63(3):805-10. PMID: 10952924 [DOI:10.1095/biolreprod63.3.805]
27. Turathum B, Saikhun K, Sangsuwan P, Kitiyanant Y. Effects of vitrification on nuclear maturation, ultrastructural changes and gene expression of canine oocytes. Reprod Biol Endocrinol. 2010; 8:70. PMID: 2056598 [DOI:10.1186/1477-7827-8-70]
28. Habibi A, Farrokhi N, Moreira da Silva F, Bettencourt BF, Bruges-Armas J, Amidi F, et al. The effects of vitrification on gene expression in mature mouse oocytes by nested quantitative PCR. J Assist Reprod Genet. 2010; 27(11):599-604. PMID: 20714800 [DOI:10.1007/s10815-010-9453-0]
29. Vallorani C, Spinaci M, Bucci D, Porcu E, Tamanini C, Galeati G. Pig oocyte vitrification by Cryotop method and the activation of the apoptotic cascade. Anim Reprod Sci. 2012; 135(1-4):68-74. PMID: 22974705 [DOI:10.1016/j.anireprosci.2012.08.020]
30. Zhao X-M, Ren J-J, Du W-H, Hao H-S, Wang D, Qin T, et al. Effect of vitrification on promoter CpG island methylation patterns and expression levels of DNA methyltransferase 1o, histone acetyltransferase 1, and deacetylase 1 in metaphase II mouse oocytes. Fertil Steril. 2013; 100(1):256-61. PMID: 23548937 [DOI:10.1016/j.fertnstert.2013.03.009]
31. Cheng K-R, Fu X-W, Zhang R-N, Jia G-X, Hou Y-P, Zhu S-E. Effect of oocyte vitrification on deoxyribonucleic acid methylation of H19, Peg3, and Snrpn differentially methylated regions in mouse blastocysts. Fertil Steril. 2014; 102(4):1183-90.e3. PMID: 25064401 [DOI:10.1016/j.fertnstert.2014.06.037]

Add your comments about this article : Your username or Email:

Rights and permissions
Creative Commons License This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.


Designed & Developed by : Yektaweb