DNA Cell Biol. 2023 Jun 08.
Folate, as the initial substrate in one-carbon metabolism, is involved in the synthesis of important substances such as DNA, RNA, and protein. Folate deficiency (FD) is associated with male subfertility and impaired spermatogenesis, yet the underlying mechanisms are poorly understood. In the present study, we established an animal model of FD to investigate the effect of FD on spermatogenesis. GC-1 spermatogonia were used as a model to investigate the effect of FD on proliferation, viability, and chromosomal instability (CIN). Furthermore, we explored the expression of core genes and proteins of spindle assembly checkpoint (SAC), a signaling cascade ensuring accurate chromosome segregation and preventing CIN during mitosis. Cells were maintained in medium containing 0, 20, 200, or 2000 nM folate for 14 days. CIN was measured by using a cytokinesis-blocked micronucleus cytome assay. We found that sperm counts decreased significantly (p < 0.001) and the rate of sperm with defects in the head increased significantly (p < 0.05) in FD diet mice. We also found, relative to the folate-sufficient conditions (2000 nM), cells cultured with 0, 20, or 200 nM folate exhibited delayed growth and increased apoptosis in an inverse dose-dependent manner. FD (0, 20, or 200 nM) significantly induced CIN (p < 0.001, p < 0.001, and p < 0.05, respectively). Moreover, FD significantly and inverse dose dependently increased the mRNA and protein expression of several key SAC-related genes. The results indicate that FD impairs SAC activity, which contributes to mitotic aberrations and CIN. These findings establish a novel association between FD and SAC dysfunction. Thus, FD-impaired spermatogenesis may be partly due to genomic instability and proliferation inhibition of spermatogonia.
Keywords: chromosomal instability; folate deficiency; male subfertility; spermatogonia; spindle assembly checkpoint