二苯乙烯苷对衰老睾丸间质细胞Ttc29基因的影响

EFFECTS OF STILBENE GLYCOSIDES ON TTC29 GENE IN AGING LEYDIG CELLS

目的:探讨二苯乙烯苷对衰老间质细胞Ttc29基因的影响。方法:原代培养大鼠睾丸间质细胞,采用自由基氧化损伤建立间质细胞衰老模型,通过β-半乳糖苷酶染色检测衰老间质细胞百分比,运用甲基化PCR法及荧光定量PCR法检测Ttc29基因甲基化程度和转录水平,染色质免疫共沉淀检测Ttc29基因启动子区H3K27me3和H3K9me3组蛋白的富集程度。结果:与正常组相比,衰老组衰老细胞阳性率明显升高(P<0.05);二苯乙烯苷可明显降低衰老细胞阳性率(P<0.05)。衰老间质细胞较正常间质细胞Ttc29基因甲基化程度显著升高,mRNA表达水平显著降低(P<0.05),且二者呈负相关关系(P<0.05)。二苯乙烯苷可显著降低衰老间质细胞Ttc29基因甲基化程度和mRNA的表达水平(P<0.05)。Ttc29基因启动子区域未检测到H3K27me3和H3K9me3的富集。结论:二苯乙烯苷可通过改变Ttc29基因的甲基化水平和mRNA的表达水平抑制间质细胞衰老。

Objective:To investigate the effects of stilbene glycosides on Ttc29 gene in aging Leydig cells.Methods:The rats’ Leydig cells were isolated and primarily cultured.The model of Leydig cells senescence was established by free radical oxidative damage and identified by detecting the cell aging marker β-galactosidase.Methylation PCR and fluorescence quantitative PCR were used to detect the methylation status and transcription level of Ttc29 gene.Chromatin immunocoprecipitation was used to detect the enrichment of H3K27me3 and H3K9me3 in the promoter region of Ttc29 gene.Results:Compared with the cells in normal group,the positive rate of senescent cells in the aging group increased significantly (P<0.05);Stilbene glycosides could significantly reduce the positive rate of senescent cells (P<0.05).Compared with the normal group,Ttc29 gene methylation was significantly increased and mRNA expression level was significantly decreased in aging Leydig cells (P<0.05);And there had a negative correlation between them (P<0.05).Stilbene glycosides could significantly reduce the methylation of Ttc29 gene and increase mRNA expression level in aging leydig cells (P<0.05).No enrichment of H3K27me3 and H3K9me3 was detected in the promoter region of Ttc29 gene.Conclusions:Stilbene glycosides can inhibit the senescence of Leydig cells by changing the methylation level and mRNA expression level of Ttc29 gene.