摘要
目的研究不同血液透析液对U937细胞蛋白激酶C-δ(PKCδ)表达及细胞凋亡的影响。方法在对数生长期人单核细胞株U937细胞培养液中添加不同血液透析液进行干预,分为A液+B液组(血液透析液A液+B液组合)、A液+B液+PKCδ特异性抑制剂(rottlerin)组、A液+B粉组(血液透析液A液+B粉组合)、A液+B粉+rottlerin组,同时设立空白对照和正常对照组。分组干预后24h和48h分别收集细胞,Hoechst33258染色后荧光显微镜观察细胞形态学改变;Annexin V-FITC/PI双标记流式细胞术检测细胞凋亡;RT-PCR和Western blotting检测PKCδ mRNA和蛋白表达。结果A液+B粉组凋亡细胞明显增多,呈凋亡典型形态学改变,分组干预24h和48h的细胞凋亡率明显高于相应时间点空白对照组、正常对照组和A液+B粉+rottlerin组(P<0.05);同时与两对照组和A液+B粉+rottlerin组比较,A液+B粉组U937细胞PKCδ mRNA和蛋白表达均明显上调(P<0.05)。A液+B液组细胞凋亡率及PKCδ mRNA和蛋白表达与两对照组和A液+B液+rottlerin组比较,差异均无统计学意义(P>0.05)。结论血液透析液A液+B粉组合对U937细胞有明显促凋亡作用,其机制可能与上调细胞PKCδ表达有关;与A液+B粉组合比较,选择A液+B液组合作为血液透析液可降低患者外周血单核细胞的细胞凋亡率。
Objective To investigate the effects of different dialysates on expression of protein kinase C-δ （PKCδ） and apoptosis of U937 cell line. Methods Different dialysates were added into culture fluid with U937 cell line at exponential phase of growth, and groups were divided： fluid A＋fluid B group （dialysate A＋dialysate B）, fluid A＋fluid B＋rottlerin （PKCδ specific inhibitor）group, fluid A＋powder B group （dialysate A＋powder B） and fluid A＋powder B ＋ rottlerin group. Besides, blank control group and normal control group were established. Cells were harvested 24 h and 48 h after treatment, morphological changes were observed by Hoechst33258 fluorescence staining, cell apoptosis was measured by Annexin-V-FITC/PI double staining, and expression of PKCδ mRNA and protein was detected by RT-PCR and Western blotting, respectively. Results Cell apoptosis significantly increased in fluid A＋powder B group, with typical morphology of apoptosis. After treatment for 24 h and 48 h, cell apoptosis rates in fluid A＋powder B group were significantly higher than those at corresponding time points in blank control group, normal control group and fluid A＋powder B＋rottlerin group （P〈0.05）. Compared with normal control group, blank control group and fluid A＋powder B＋rottlerin group, the expression of PKCδ mRNA and protein of U937 cells in fluid A＋powder B group were significantly increased （P〈0.05）. There was no significant difference in cell apoptosis rates and expression of PKCδ mRNA and protein between fluid A＋fluid B group and blank control group, normal control group and fluid A＋fluid B＋rottlerin group （P〉0.05）. Conclusion Fluid A＋powder B can significantly increase apoptosis of U937 cell line, the mechanism of which may be associated with the up-regulation of expression of PKCδ. Compared with fluid A＋powder B, fluid A＋fluid B is superior in reducing apoptosis of peripheral blood monouclear cells.
出处
《上海交通大学学报（医学版）》
CAS
CSCD
北大核心
2009年第12期1434-1438,共5页
Journal of Shanghai Jiao tong University:Medical Science
基金
上海市卫生局基金(2007129)
上海市宝山区科委基金(08-E-28)~~
