Department of Reproduction and Genetics, Heping Hospital Affiliated to Changzhi Medical College, Changzhi, Shanxi 046000, People's Republic of China.
Drug Des Devel Ther. 2020 Nov 10;14:4845-4854. doi: 10.2147/DDDT.S269236. eCollection 2020.
MicroRNAs play important roles in testicular development and spermatogenesis. Previous research has indicated that the level of miR-200a was significantly upregulated in patients with different spermatogenic impairments. However, the mechanism by which miR-200a regulated spermatogenic impairments remains unclear.
Leydig cells were treated with triptolide (TP) to mimic spermatogenic impairments. CCK-8 and flow cytometry were used to detect the proliferation and apoptosis in Leydig cells, respectively. In addition, Western blot assay was used to examine ATG7, ATG5, p62 protein levels in MLTC-1 cells.
TP dose-dependently upregulated the expression of miR-200a in MLTC-1 cells. In addition, TP inhibited the proliferation of MLTC-1 cells via inducing apoptosis and oxidative stress; however, these phenomena were notably reversed by miR-200a antagomir. Furthermore, luciferase reporter assay identified that ATG7 was the direct binding target of miR-200a. TP treatment markedly inhibited the activation of autophagy in MLTC-1 cells via inhibition of ATG7. Conversely, downregulation of miR-200a significantly induced autophagy in TP-treated MLTC-1 cells by activation of ATG7. Meanwhile, the cell protective effects of miR-200a against TP were reversed by autophagy inhibitor 3MA, indicating that autophagy plays an important role.
These results indicated that downregulation of miR-200a could protect MLTC-1 cells against TP by inducing autophagy. Therefore, miR-200a might serve as a new therapeutic target for the treatment of male hypogonadism.
MicroRNAs 在睾丸发育和精子发生中发挥重要作用。先前的研究表明，miR-200a 的水平在不同生精障碍患者中显著上调。然而，miR-200a 调节生精障碍的机制尚不清楚。
用雷公藤内酯醇 (TP) 处理 Leydig 细胞模拟生精障碍。分别用 CCK-8 和流式细胞术检测 Leydig 细胞的增殖和凋亡。此外，Western blot 检测 MLTC-1 细胞中 ATG7、ATG5、p62 蛋白水平。
TP 剂量依赖性地上调 MLTC-1 细胞中 miR-200a 的表达。此外，TP 通过诱导凋亡和氧化应激抑制 MLTC-1 细胞的增殖，而这些现象明显被 miR-200a 拮抗剂逆转。此外，荧光素酶报告基因实验鉴定出 ATG7 是 miR-200a 的直接靶基因。TP 处理通过抑制 ATG7 显著抑制 MLTC-1 细胞中的自噬激活。相反，下调 miR-200a 通过激活 ATG7 显著诱导 TP 处理的 MLTC-1 细胞中的自噬。同时，自噬抑制剂 3MA 逆转了 miR-200a 对 TP 的细胞保护作用，表明自噬起重要作用。
这些结果表明下调 miR-200a 通过诱导自噬可保护 MLTC-1 细胞免受 TP 的影响。因此，miR-200a 可能成为治疗男性性腺功能减退症的新治疗靶点。
