Ethylene glycol monomethyl ether (EGME) induces testicular lesion in rats and human.  To investigate miRNAs expression in EGME testicular lesion, miRNA array assay and real-time RT-PCR analysis were conducted by using testis in rats treated with 50 and 2,000 mg/kg EGME for 6 and 24 hr. The expression of corresponding target gene for miRNAs was also examined. At 50 mg/kg, there were no changes in the gene expression and histopathological examination. At 2,000 mg/kg, slight decrease of phacytene spermatocytes with cell shrinkage and nucleus pyknosis at 6 hr and remarkable decrease (or cell death) of phacytene spermatocytes with Sertoli cell vacuolation at 24 hr were observed. After 24 hr, miR-449a and miR-92a decreased obviously and, miR-320, miR-134 and miR-188 increased, while only miR-760-5p increased after 6 hr. Above these miRNAs are reported to have an important role for spermatogenesis. The gene expression of Bcl-2, target for miR-449a, increased and therefore it is considered anti-apoptotic reaction has started in this stage. The expression of high mobility group AT-hook 2 (target of miR-92a) which regulates histone structure, was increased. Furthermore, histone deacethylase 4, targets for miR-320, was also affected. Above prohibiting apoptosis or activating epigenetic genes might be protective reaction to spermatocytes death under the miRNAs regulation in EGME testicular lesion.