氧化型烟酰胺腺嘌呤二核苷酸抗辐射损伤作用及其机制

目的探讨氧化型烟酰胺腺嘌呤二核苷酸(NAD+)的抗辐射损伤作用及其机制。方法将培养的人正常肝细胞株L02细胞分为3组:对照组、照射组和照射+NAD+组。对照组细胞不予照射,直接加入不含NAD+的RPMI-1640培养基培养;照射组细胞照射后加入不含NAD+的RPMI-1640培养基培养;照射+NAD+组细胞照射后加入含有NAD+(终浓度1 g.L-1)的RPMI-1640培养基培养。采用四甲基偶氮唑盐(MTT)法检测NAD+对受照射L02细胞生长活性的影响;应用流式细胞仪检测各组细胞凋亡率以及凋亡相关蛋白p53、bax、bcl-2阳性表达率和各周期细胞含量;采用Caspase-3活性检测试剂盒检测细胞Caspase-3活性。结果细胞受照射后加入NAD+能够对抗X射线照射对L02细胞的生长抑制作用,细胞生长活性较照射组细胞活性显著提高;L02细胞在受照射后p53、bax蛋白表达增加,bcl-2表达下调,Caspase-3活性增加,而且细胞周期表现为G1期阻滞;受照射后加入NAD+,则p53、bax蛋白表达下调,bcl-2表达增加,Caspase-3活性下降,G2/M期细胞比例明显增加。结论 NAD+可对抗L02细胞的X线辐射损伤,其作用机制可能通过下调p53、bax与上调bcl-2表达以及抑制Caspase-3活性,抑制受照射细胞凋亡。

Antiradiation effect and mechanisms of oxidized form of nicotinamide-adenine dinucleotide

Objective To study the antiradiation effect and mechanisms of oxidized form of nicotinamide-adenine dinucleotide(NAD+).Methods L02 human liver cells were divided into three groups:control group,irradiation group and irradiation plus NAD+ group.Cells of the irradiation plus NAD+ group and the irradiation group were respectively cultured in the RPMI-1640 medium containing NAD+(1 g·L-1)or non-NAD+ after irradiation.Cells of the control group were directly cultured in the RPMI-1640 medium.The cell growth activity was detected by MTT assay and the cell apoptosis rate determined by Annexin V/PI staining,the apoptosis-related proteins expression(bax,bcl-2 and p53)detected by flow cytometry and the Caspase-3 activity detected by Caspase-3 activity assay kit.Results NAD+ could inhibit significantly apoptosis of the L02 liver cells induced by radiation damage.As compared with the irradiation group,the expressions of p53,bax proteins up-regulated in irradiation plus NAD+ group cells,and the Caspase-3 activity significantly lower,the bcl-2 expressions significantly higher(P<0.05).Meanwhile,G2/M phase cells increased significantly in irradiation plus NAD+ group cells.Conclusion NAD+ can significantly inhibit L02 cells apoptosis induced by X-ray irradiation damage possibly by the mechanism of regulating the apoptosis-related proteins expression and Caspase-3 activity.