Assessment of apoptosis and oxidative stress in cryopreserved ovary after grafting in fibrin-alginate scaffold with endothelial cells and melatonin in wistar rats

Abstract

Objective

Infertility is a significant public health concern affecting 10–15 % of couples. Young women undergoing gonadotoxic treatment are at higher risk of ovarian dysfunction and infertility. To mitigate this risk, ovarian tissue freezing and transplantation have been developed as a novel strategy. However, challenges such as follicular loss and dysfunction during the freezing process, and ovarian damage during transplantation, persist. This study aimed to investigate the potential of using appropriate antifreeze, antioxidant, wound healing, and biological hydrogels to reduce these injuries. Specifically, the effect of fibrin scaffold with endothelial cells and melatonin on apoptotic gene expression and antioxidants in cryopreserved ovaries after transplantation was examined.

Methods

A total of 36 adult female wistar rats) 6–8-week-old and weighing from 200 to 220 g) were divided equally into six groups (n = 6): 1) control group (C), 2) transplanted ovarian tissue after vitrification and thawing process (Group 1), 3) transplanted vitrified/thawed ovarian tissue while encapsulated in Fib/Alg hydrogel (Group 2), 4) transplanted vitrified/thawed ovarian tissue while encapsulated in Fib/Alg hydrogel in addition with melatonin (Group 3), 5) transplanted vitrified/thawed ovarian tissue while encapsulated in Fib/Alg hydrogel in addition with endothelial cells (Group 4) and 6) transplanted vitrified/thawed ovarian tissue while encapsulated in Fib/Alg hydrogel in addition with melatonin endothelial cells (Group 5). The ovaries were auto-transplanted in the rats' lumbar region. After 14 days, the ovaries were removed. Antioxidant levels (SOD, GPx, MDA, and TAC) were evaluated using ELISA, and apoptotic gene expressions (Bax/Bcl2 and caspase 3) were analyzed by real-time RT-PCR to determine apoptosis.

Results

In the transplanted frozen ovary group, Bax/Bcl2 and caspase 3 gene expression increased significantly (P < 0.05), while antioxidant levels (SOD, GPx, MDA, and TAC) decreased. The encapsulated frozen ovary group showed decreased gene expression and increased antioxidant levels. The ovary group encapsulated with fibrin scaffold, endothelial cells, and melatonin had the most significant decrease in gene expression and increase in antioxidant levels (P < 0.05).

Conclusion

Coordinated action of Fibrin-based scaffold with endothelial cells and melatonin could decrease apoptosis gene expression and increase antioxidant levels in cryopreserved ovaries after transplantation, providing valuable insights into preserving fertility in young women undergoing gonadotoxic treatment.