Folic acid reduces methomyl insecticide damage to testicular cells by altering the DNA methylation environment

Abstract Methomyl is a carbamate insecticide with confirmed testicular toxicity. This study intends to observe the damage of methomyl on testicular cells and the protective effect of folic acid through in vitro experiments. We treated the GC-1 spermatogonia, TM4 Sertoli cells, and TM3 Leydig cells with Methomyl (0, 250, 500, and 1000 μM) with or not folic acid (0, 10, 100, and 1000 nM) for 24h. It was found that folic acid attenuated the testicular cytotoxicity of methomyl in a dose-dependent manner. In spermatogonia, methomyl significantly inhibited the expression of proliferation genes Ki67 and PCNA at 1000 μM, and increased the expression of apoptosis genes Caspase3 and Bax at each dose, while folic acid could reverse the change caused by methomyl. In Sertoli cells, methomyl dose-dependently inhibited the expression of blood-testis barrier function genes TJP1, Cx43, and N-cadherin, but did not affect Occludin and E-cadherin, while folic acid could significantly up-regulate the expression of TJP1, Cx43, and Occludin. In Leydig cells, methomyl dose-dependently inhibited the expression of steroid synthase P450scc, StAR, Hsd3b1 and down-regulated the level of testosterone, but did not affect Cyp17a1 and Hsd17b1, while folic acid could significantly up-regulate the expression of P450scc, StAR, Hsd3b, Cyp17a1, and Hsd17b1, and reverse testosterone level downtrend. Simultaneously, methomyl did not affect the expression of Dnmt1, Dnmt3A, Dnmt3B genes in testicular cells, while folic acid dose-dependently up-regulated the expression of Dnmt1, Dnmt3A, and Dnmt3B in spermatogonia, Dnmt1 and Dnmt3B in Sertoli cells, and Dnmt1 and Dnmt3A in Leydig cells. In conclusion, folic acid attenuated the damage of methomyl on testicular spermatogonia, Sertoli cells, and Leydig cells by altering the DNA methylation environment.