In vitroeffects of melatonin on colonization of neonate mouse spermatogonial stem cells

Abstract

We have recently reported that antioxidant supplements enhance the efficacy of cryopreserved spermatogonial stem cells. Melatonin is considered a free radical scavenger which has direct and indirect antioxidant effects in in vitro and in vivo microenvironments. Due to the anti-apoptotic properties of melatonin, researchers have proposed that melatonin may improve the efficiency of spermatogonial stem cell (SSC) transplantation. However, the appropriate methodology which facilitates SSC proliferation remains to be determined. Identification of a proper propagation system is essential for the future application of SSCs in the field of infertility. The aim of the present study was to investigate the effects of melatonin on the colonization of SSCs. SSCs were isolated from the testes of three to six day old mice, and their purities were assessed by cytometry using Plzf antibody. Isolated testicular cells were cultured in the absence or presence of melatonin extract for two weeks. Suppression of differentiation and maintenance of spermatogonial stem cells was confirmed by alkaline phosphatase staining and immunocytochemistry using Plzf antibody. The number and diameter of the colonies of SSCs were assessed during the 7 and 14 days of culture, and the expression of Id4, Plzf, and C-kit were evaluated using real-time PCR at the end of the culture period. The survival rate of the cultured cells in the presence of melatonin was significantly higher than the control group. The number and diameter of colonies also increased in the cells treated with melatonin. The results of our study suggest that culture of SSCs with 100 μM melatonin supplementation can increase SSCs proliferation significantly.

Natural Cryoprotective and Cytoprotective Agents in Cryopreservation: A Focus on Melatonin

“…According to these findings in goat SSCs, MLT was found to promote beneficial effects in terms of cell proliferation and survival rates in mice SSCs [ 98 ].…”

Melatonin in cryopreservation media improves transplantation efficiency of frozen–thawed spermatogonial stem cells into testes of azoospermic mice

“…In addition, it has been shown that the melatonin acts as a cryoprotectant in the freezing media containing sperm and positively affects male fertility preservation [ 20 – 22 ]. Previous research has shown that adding melatonin to the culture medium increases the efficiency and performance of SSCs isolated from mice [ 15 , 16 ]. Cryopreservation of SSCs can provide appropriate options such as SSCs transplantation and in vitro spermatogenesis for prepubertal boys and adult male after cancer treatment [ 23 ].…”

Tissue Engineering to Improve Immature Testicular Tissue and Cell Transplantation Outcomes: One Step Closer to Fertility Restoration for Prepubertal Boys Exposed to Gonadotoxic Treatments

“…This drug has antioxidant and antiapoptotic properties that rely on the radical scavenger effect delivered by its indole derivate [ 84 ]. While melatonin appeared to be beneficial in SSC transplantation experiments [ 85 , 86 ], addition of melatonin to the vitrification-warming medium did not reduce expression of apoptotic genes in frozen/thawed murine ITT [ 87 ].…”