异亮氨酸缓解猪睾丸支持细胞氧化应激作用

Effects of L-isoleucine on alleviating oxidative stress in porcine testicular sertoli cells

目的:研究异亮氨酸缓解过氧化氢诱导的猪睾丸支持细胞氧化应激的保护作用,并确定不同浓度异亮氨酸对缓解氧化应激导致的睾丸支持细胞活力下降的最佳浓度,及最佳异亮氨酸浓度对猪睾丸支持细胞的增殖、凋亡和抗氧化水平的影响。方法:利用CCK-8法检测细胞活力,筛选在氧化应激模型下异亮氨酸保护猪睾丸支持细胞的最适浓度。应用流式细胞术检测最适异亮氨酸浓度下支持细胞的周期和凋亡情况。应用试剂盒检测总超氧化物歧化酶(Superoxide Dismutase,T-SOD)、总抗氧化能力(Total Antioxidant Capacity,T-AOC)、谷胱甘肽过氧化物酶(Glutathione Peroxidase,GSH-Px)和丙二醛(Malondialdehyde,MDA)的表达量。结果:与氧化应激丙氨酸等氮对照组和其他浓度氧化应激异亮氨酸组相比,32 mmol/L氧化应激异亮氨酸组细胞增殖活力显著升高。与氧化应激不加药组和氧化应激丙氨酸等氮对照组相比,氧化应激异亮氨酸组细胞分化G_(0)/G_(1)期比例相对较低,S期和G_(2)/M期比例无明显差异。与氧化应激不加药组相比,氧化应激异亮氨酸组细胞凋亡比例呈显著降低,与氧化应激等氮对照组相比,氧化应激异亮氨酸组细胞凋亡比例无明显差异。与无氧化应激丙氨酸等氮对照组和氧化应激丙氨酸对照组相比,异亮氨酸极显著增强T-SOD的活性,也极显著增加氧化应激异亮氨酸组T-AOC,而对GSH-Px和MDA没有显著影响。结论:在本试验条件下32 mmol/L异亮氨酸效果最佳,有效缓解氧化应激导致的细胞活力下降、细胞周期紊乱和细胞凋亡。

Objective:To investigate the protective role of L-isoleucine in alleviating oxidative stress induced by H 2 O 2 in porcine sertoli cells,to determine the optimal concentration of L-isoleucine for mitigating the decline in cell viability due to oxidative stress,and to assess,the effects on cell proliferation,apoptosis,and antioxidant levels in porcine Sertoli cells at the optinal concentranon.Methods:Cell viability was examined by using the CCK-8 assay to select the most suitable concentration of L-isoleucine for protecting porcine Sertoli cells under oxidative stress conditions.Flow cytometry was utilized to detect the cell cycle and apoptosis of Sertoli cells at the optimal L-isoleucine concentration.The levels of Superoxide dismutase(T-SOD),Total antioxidant capacity(T-AOC),Glutathione peroxidase(GSH-Px),and Malondialdehyde(MDA)were measured by using specific assay kits.Results:Compared to the oxidative stress alanine nitrogen control group and other concentrations of oxidative stress isoleucine groups,cell proliferation vitality was significantly elevated in the 32 mmol/L oxidative stress isoleucine group.Relative to the oxidative stress untreated group and the oxidative stress alanine nitrogen control group,the oxidative stress isoleucine group exhibited a comparatively lower ratio of cells in the G_(0)/G_(1)phase,with no notable differences in the proportions of the S and G_(2)/M phases.In comparison to the oxidative stress untreated group,the proportion of cell apoptosis was significantly reduced in the oxidative stress isoleucine-treated group,with no significant difference when compared to the oxidative stress nitrogen control group.Isoleucine markedly increased T-SOD activity in comparison to both the non-oxidative stress alanine nitrogen control group and the oxidative stress alanine control group.It also significantly enhanced T-AOC levels in the oxidative stress isoleucine group,though it did not have a significant effect on GSH-Px and MDA.Conclusion:Under the experimental conditions of this study,32 mmol/L of isoleucine was found to be the most effective concentration,successfully mitigating the decline in cell viability,cell cycle dysregulation,and cell apoptosis caused by oxidative stress.