镉诱导LLC-PK_1细胞凋亡与bcl-2、p53蛋白表达关系的研究

目的　探讨镉诱导LLC PK1 细胞凋亡及与bcl 2、p5 3(mtp5 3)蛋白表达的相互关系。方法　采用透射电镜观察凋亡小体、流式细胞仪分析凋亡率、琼脂糖凝胶DNA电泳方法确定镉对LLC PK1 细胞诱导的凋亡作用 ,以及流式细胞仪分别测定bcl 2和p5 3基因表达产物bcl 2蛋白、mtp5 3蛋白。结果　透射电镜观察发现 4 0 μmol LCdCl2 作用LLC PK1 细胞 12h后 ,出现典型的凋亡小体 ;流式细胞仪分析其凋亡率为 32 6 1% ,并高于对照组 (1 0 8% ) (P <0 0 1) ;琼脂糖凝胶DNA电泳呈明显梯形条带。 0、10、2 0、4 0 μmol LCdCl2 作用LLC PK1 细胞 4h、8h ,8h后 ,bcl 2基因表达逐渐下降 ,并呈良好的剂量 -反应关系 (r=- 0 910 ,P <0 0 5 ) ;作用 4h、8h后mtp5 3蛋白表达均明显下降 ,并有剂量 -反应关系 (r值分别为 - 0 716、- 0 972 ,P值均 <0 0 5 )。结论　镉诱导LLC PK1 细胞凋亡可能与镉抑制bcl 2、mtp5 3蛋白表达有关

Apoptosis induced by cadmium and the expression of bcl-2 and p53 genes in LLC-PK1 cells

OBJECTIVE: To study the effect of cadmium on the apoptosis of porcine renal proximal tubule epithelial cells (LLC-PK1) and the relationships between the apoptosis and gene expression of the bcl-2 and mutant p53 (mtp53) protein. METHODS: Apoptotic bodies were examined by electron microscopy, the rates of apoptosis were measured by flow cytometry (FCM) and DNA ladder was detected by DNA gel electrophoresis. The expressions of bcl-2 and mtp53 were measured respectively by FCM. RESULTS: The apoptotic bodies of LLC-PK1 cells treated with CdCl2 at dose of 40 micromol/L for 12 h were observed by electron microscopy and the apoptosis rates of LLC-PK1 cells were 32.61%, which were higher than those of the control group (P < 0.01). Apparent DNA ladder was showed by DNA gel electrophoresis. LLC-PK1 cells were incubated with CdCl2 at doses of 0, 10, 20, 40 micromol/L for 4 h or 8 h, it was found that expressions of bcl-2 and mtp53 were decreased and where a significant dose-response was found (r = -0.910 for bcl-2 8 h, r = -0.716, -0.972 for mtp53 4 h, 8 h, respectively). CONCLUSION: The results were suggested that the apoptosis of LLC-PK1 cells induced by cadmium might be associated with the inhibitions of the expression of bcl-2 and p53.