Cell Journal (Apr 2023)

Comparative Epigenetic Analysis of Imprinting Genes Involved in Fertility, in Cryopreserved Human Sperms with Rapid Freezing versus Vitrification Methods

  • Nahid Khosronezhad,
  • Vahideh Hassanzadeh,
  • Maryam Hezavehei,
  • Abdolhossein Shahverdi,
  • Maryam Shahhoseini

DOI
https://doi.org/10.22074/cellj.2023.1974291.1171
Journal volume & issue
Vol. 25, no. 4
pp. 238 – 246

Abstract

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Objective: Choosing the optimal method for human sperm cryopreservation seems necessary to reduce cryoinjury.The aim of this study is to compare two cryopreservation methods including rapid-freezing and vitrification, in termsof cellular parameters, epigenetic patterns and expression of paternally imprinted genes (PAX8, PEG3 and RTL1) inhuman sperm which play a role in male fertility.Materials and Methods: In this experimental study, semen samples were collected from 20 normozoospermic men.After washing the sperms, cellular parameters were investigated. DNA methylation and expression of genes wereinvestigated using methylation-specific polymerase chain reaction (PCR) and real-time PCR methods, respectively.Results: The results showed a significant decrease in sperm motility and viability, while a significant increase wasobserved in DNA fragmentation index of cryopreserved groups in comparison with the fresh group. Moreover, a significantreduction in sperm total motility (TM, P<0.01) and viability (P<0.01) was determined, whereas a significant increasewas observed in DNA fragmentation index (P<0.05) of the vitrification group compared to the rapid-freezing group.Our results also showed a significant decrease in expression of PAX8, PEG3 and RTL1 genes in the cryopreservedgroups compared to the fresh group. However, expression of PEG3 (P<0.01) and RTL1 (P<0.05) genes were reducedin the vitrification compared to the rapid-freezing group. Moreover, a significant increase in the percentage of PAX8,PEG3 and RTL1 methylation was detected in the rapid-freezing group (P<0.01, P<0.0001 and P<0.001, respectively)and vitrification group (P<0.01, P<0.0001 and P<0.0001, respectively) compared to the fresh group. Additionally,percentage of PEG3 and RTL1 methylation in the vitrification group was significantly increased (P<0.05 and P<0.05,respectively) compared to the rapid-freezing group.Conclusion: Our findings showed that rapid-freezing is more suitable method for maintaining sperm cell quality. Inaddition, due to the role of these genes in fertility, changes in their expression and epigenetic modification may affectfertility.

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