双酚A对小鼠精原干细胞C18-4的增殖及表观遗传机制的影响

Effects of bisphenol A on cell proliferation and epigenetic mechanisms of mouse spermatogonial C18-4 cell line

目的：研究不同剂量的双酚A（BPA）暴露对小鼠精原干细胞C18-4的增殖及表观遗传机制的影响。方法：C18-4细胞暴露于不同浓度的BPA（10-9-10-4 mol/L）96 h,用CCK-8方法测定细胞的增殖;通过点杂交、Real-time PCR、Western blotting方法检测10-5 mol/L BPA、10-9 mol/L BPA对DNA甲基化水平、组蛋白甲基转移酶基因、组蛋白H3K27Me3、H3K36Me3水平的变化。结果：110-9-10-6 mol/L BPA促进C18-4细胞增殖;10-5 mol/L BPA抑制细胞增殖,促进细胞凋亡;10-4 mol/L BPA导致细胞死亡。210-5 mol/L高剂量BPA引起C18-4细胞DNA整体甲基化水平下降（P〈0.01）。3 10-5 mol/L高剂量BPA能引起H3K27Me3、H3K36Me3水平下降（P〈0.01）。结论：BPA能干扰小鼠精原干细胞C18-4的细胞增殖;高剂量BPA能引起DNA甲基化水平和组蛋白甲基化水平的下降。

Objective： To study the effects of different doses of bisphenol A （BPA） exposure on cell proliferation and epigenetic mechanisms in mouse spermatogonial stem cell C18-4. Methods： C18-4 cell proliferation was measured after exposure to different concentrations of bisphenol A （10-9 -104 mol/L） for 96 h using a CCK-8 assay. The levels of global DNA methylation, histone methyltransferase gene expression and histone H3K27Me3, H3K36Me3 were detected after the C18-4 cells exposed to two different concentrations of BPA （10-s mol/L and 10.9 mol/L）, by using the methods of dot blotting, real-time RT-PCR, Western blotting, respectively. Results： 1） 10.9-10.6 mol/L BPA exposure promoted C 18-4 cell proliferation, while 10-5 mol/L BPA inhibited cell proliferation meanwhile increased cell apoptosis rate, and 104 mol/L B PA led to cell death. 2） 10-5 mol/L BPA decreased the level of global DNA methylation ofC18-4 cells （P〈0.01）. 3） 10.5 mol/L BPA decreased the levels of H3K27Me3 and H3K36Me3 （P〈0.01）. Conclusion： BPA can disturb cell proliferation of sperrnatogonial stem cell C 18-4; 10.5 mol/L BPA exposure can decrease the levels of DNA methylation and histone methylation of C 18-4 cells.