硒对氟诱导NRK-52E细胞凋亡干预作用机制

目的 探讨硒对氟诱导NRK-52E细胞凋亡的拮抗作用。方法 实验设对照组、染氟组、染硒组、硒干预组，培养NRK-52E细胞，染毒72 h，采用流式细胞仪检测细胞凋亡，Western blot检测AMPK及线粒体通路相关蛋白表达水平。结果 与对照组[（5.97±0.09）%]比较，5、20 mg/L NaF组细胞凋亡率[分别为（7.92±0.24）%、（11.06±0.17）%]明显升高（P<0.05）；与20 mg/L NaF组比较，17.1、34.2 μg/L硒干预组细胞凋亡率[分别为（8.46±0.09）%、（9.88±0.08）%]明显降低（P<0.05）；与对照组比较，20 mg/L NaF组细胞AMPK磷酸化蛋白、bax、Cyt-C蛋白表达[分别为（0.73±0.16）、（0.99±0.16）、（0.73±0.08）]、活性caspase-9和caspase-3蛋白表达[分别为（1.17±0.17）、（1.51±0.42）]均升高（P<0.05）；与20 mg/L NaF组比较，17.1、34.2 g/L硒干预组AMPK磷酸化蛋白表达量[分别为（0.46±0.12）、（0.48±0.15）]、bax、Cyt-C蛋白表达量[分别为（0.55±0.09）、（0.61±0.16）与（0.30±0.06）、（0.34±0.05）]及活性caspase-9、caspase-3蛋白表达量[分别为（0.76±0.11）、（0.40±0.12）与（0.35±0.12）、（0.27±0.04）]均降低（P<0.05）。结论 AMPK介导的线粒体通路参与了硒拮抗氟诱导NRK-52E细胞凋亡过程。

Mechanism of interference effect of selenium on sodium fluoride-induced apoptosis in NRK-52E cells

Objective To study antagonistic effect of selenium on sodium fluoride-induced apoptosis of NRK-52E cells.Methods NRK-52E cells were cultured and divided into following groups:a control group,two sodium fluoride (NaF)-treated (5,20 mg/L) groups,two sodium selenite (Na₂SeO₃)-treated (17.1,34.2 μg/L) groups,and four NaF+Na₂SeO₃-treated (5 or 20 mg/L NaF+17.1 or 34.2 μg/L Na₂SeO₃) groups.Cell apoptosis was detected by flow cytometry 72 hours after the treatments.Protein expressions of adenosine monophosphate-activated protein kinase (AMPK) and related mitochondria pathway proteins were determined using Western blot.Results Compared to the control group(5.97±0.09%),the cell apoptosis rate of the 5 and 20 mg/L NaF-treated groups (7.92±0.24% and 11.06±0.17%) were remarkably increased (P<0.05),while the rates of 20 mg/L NaF+17.1 and 34.2 g/L Na₂SeO₃-treated groups (8.46±0.09% and 9.88±0.08%) were statistically significantly decreased compared to that of the 20 mg/L NaF-treated groups (both P<0.05).Compared to those of the control group,the expression levels of phosphorylated-AMPK (0.73±0.16),bax (0.99±0.16),cytochrome C (Cyt-C)(0.73±0.08),caspase-9 (1.17±0.17),and caspase-3 (1.51±0.42) were obviously up-regulated in 20 mg/L NaF-treated group (P<0.05).Compared to those of the 20 mg/L NaF-treated group,the expression levels of phosphorylated -AMPK(046±0.12,0.48±0.15),bax(0.55±0.09,0.61±0.16),Cyt-C (0.30±0.06,0.34±0.05),caspase-9 (0.76±0.11,0.40±0.12),caspase-3 (0.35±0.1,0.27±0.04) were down-regulated in 20 mg/L NaF+17.1 and 34.2 g/L Na₂SeO₃-treated groups (all P<0.05).Conclusion Fluoride can induce apoptosis in NRK-52E cells and the effect can be ameliorated by selenium by regulation of AMPK mediated mitochondrial pathway.