N-乙酰半胱氨酸对山羊子宫内膜基质细胞增殖及相关基因表达的影响

The Effect of N-acetylcysteine on Proliferation and Related Gene Expression of Goat Endometrial Stromal Cells in vitro

旨在探究N-乙酰半胱氨酸(N-acetyl-L-cysteine,NAC)对山羊子宫内膜基质细胞(goat endometrial stromal cells,gESCs)的影响。本研究以山羊子宫内膜基质细胞为对象,体外培养基中添加浓度梯度分别为100、200、400μmol·L^(-1)的NAC,将0μmol·L^(-1)NAC视为空白组,运用CCK-8法筛选NAC的最适浓度并判断其对细胞的增殖情况,随后通过DCFH-DA探针、RT-qPCR分别检测最适浓度NAC对细胞内活性氧含量、细胞周期、增殖相关因子及山羊繁殖性能相关基因TGF-β1、TGF-β3 mRNA表达水平的影响。结果表明:200μmol·L^(-1)NAC浓度较其它梯度对山羊子宫内膜基质细胞增殖的影响更为显著,此外,200μmol·L^(-1)NAC较空白组降低了细胞ROS的含量(P<0.01),从而延缓细胞的衰老。RT-qPCR结果显示:200μmol·L^(-1)NAC添加能极显著提高细胞周期及增殖相关因子CyclinA1、CyclinE、PCNA mRNA的表达水平(P<0.01),降低细胞周期因子CyclinD2的mRNA表达量(P<0.01);并极显著降低转化生长因子家族TGF-β1基因(P<0.01)、显著降低TGF-β3基因(P<0.05)的mRNA表达水平。故本研究推测200μmol·L^(-1)NAC可能通过调控细胞中CyclinA1、CyclinE、PCNA、CyclinD2的表达水平来促进细胞的增殖并通过调控转化生长家族因子TGF-β1、TGF-β3的表达水平影响山羊的繁殖性状,为进一步研究NAC对山羊繁殖性状的影响机理提供理论基础。

The aim was to explore the effect of N-acetyl-L-cysteine(NAC)on goat endometrial stromal cells(gESCs).In this study,goat endometrial stromal cells were used as the object.Three concentration gradients of 100,200,400μmol·L^(-1)NAC were added to the culture medium.The 0μmol·L^(-1)NAC was regarded as the blank group and CCK-8 method was used to screen the optimal concentration of NAC and judge its effect on cell proliferation.Then DCFH-DA probe and RT-qPCR were used to detect the optimal concentration of NAC on the intracellular reactive oxygen content,cell cycle and proliferation related factors,and goat reproductive performance-related genes TGF-β1 and TGF-β3 mRNA expression levels.The results showed that the concentration of 200μmol·L^(-1)NAC had a more significant effect on the proliferation of goat endometrial stromal cells than other gradients.In addition,200μmol·L^(-1)NAC reduced the ROS content of cells compared with the blank group(P<0.01),thereby delaying cell senescence.RT-qPCR results showed that the addition of 200μmol·L^(-1)NAC significantly increased the expression levels of cell cycle and proliferation-related factors CyclinA1,CyclinE,PCNA mRNA(P<0.01),and reduced the mRNA expression of cell cycle factor CyclinD2(P<0.01);and extremely significantly reduced the TGF-β1 gene(P<0.01)of the transforming growth factor family and significantly reduced the mRNA expression level of the TGF-β3 gene(P<0.05).Therefore,this study speculates that 200μmol·L^(-1)NAC may promote cell proliferation by regulating the expression levels of CyclinA1,CyclinE,PCNA,and CyclinD2 in cells,and affect goat reproduction traits by regulating the expression levels of transforming growth family factors TGF-β1 and TGF-β3.This study provide a theoretical basis for further research on the mechanism of NAC’s influence on goat reproductive traits.