同型半胱氨酸对NIT-1细胞凋亡及细胞内SOD活性的影响

目的探讨同型半胱氨酸（Hcy）对克隆的胰岛β细胞NIT-1细胞株的存活率和凋亡的影响，以及对细胞内超氧化物歧化酶（SOD）的影响。方法将不同浓度的Hcy作用于NIT-1细胞后，用MTT的方法检测细胞生存率；用流式细胞仪Annexin V/PI双染色法和琼脂糖凝胶电泳检测不同浓度的Hcy作用不同时间对NIT-1细胞凋亡的影响；用黄嘌呤氧化酶法和WST结合的方法检测Hcy作用后的NIT-1细胞内SOD的活性。结果Hcy以时间、剂量依赖性的方式抑制NIT-1细胞的存活率（P〈0.01）。Hcy可诱导NIT-1细胞的凋亡，随作用时间的延长和Hcy浓度增加NIT-1细胞的凋亡率逐渐增加，浓度为100μmol／L的Hcy作用24h细胞凋亡明显增加，凋亡率为7、21％（P〈0．01），250μmol／L的Hcy作用12h后细胞凋亡率达8．91％（P〈0．01）。100μmol／L的Hcy作用24h后NIT-1细胞内SOD活性较正常组细胞下降20．2％（P〈0．01）。结论Hcy可抑制NIT-1细胞的存活率，并以时间和剂量依赖性的方式诱导细胞凋亡；这些有害作用可能是通过抑制细胞内的SOD的活性而发挥作用，为研究保护胰岛细胞功能提供一种新的思路。

Effects of homocysteine on apoptosis and SOD activity of NIT-1 cells

Objective To study the possible effect of homocysteine on apoptosis and superoxide dismutase （SOD） activity in cells of NIT-1 cell line, which is a mouse pancreatic beta-cell line derived from a transgenic NOD/Lt mouse. Methods NIT-1 cells were cultured in the culture medium containing 5.6mmol/L glucose. Homocysteine at various concentrations was added to the culture medium according to a time schedule. The cell viability was determined by MTT colorimetric assay. The apoptosis of NIT-1 cells was evaluated by flow cytometry Annexin V/PI double staining method and electrophoresis on agarose gel. The SOD activity in NIT-1 cells was also assayed. Results In comparison with that in the control group, the cell viability was inhibited by homocysteine in a time- and concentration-dependent manner. Both Annexin V/PI FACS and electrophoresis analysis on agarose gel indicated that cell apoptosis could be induced by homocysteine in a time- and concentration-dependent manner. In comparison with the control group, intra-cellular SOD activity significantly declined as the concentration of homocysteine increased to 100 μmol/L （P 〈 0.01）. Conclusion Homocysteine can inhibit the cell viability and induce cell apoptosis of NIT-1 cells in a time- and dose-dependent manner. These detrimental effects may be effectuated throngh inhibiting SOD activity in the cells. This provides a new thought for studying protection of pancreatic islet cell function.