腺病毒介导HO-1基因转染肾小管上皮细胞及对PBMC增殖影响的实验研究  

Transfection of Ad-HO-1 on renal tubular epithelial cell HK-2 and its effect on PBMC proliferation

作　　者：戴卫华[1] 吴雄飞[1] 金锡御[1] 

机构地区：[1]第三军医大学西南医院肾科,重庆400038

出　　处：《第三军医大学学报》2006年第17期1756-1758,共3页Journal of Third Military Medical University

基　　金：国家自然科学基金资助项目(30170436)~~

摘　　要：目的观察Ad-HO-1感染对HK-2细胞存活率的影响,探讨表达目的蛋白血红素氧化酶-1(HO-1)的HK-2细胞能否有效降低外周血单个核细胞(peripheral blood monoeuclear cell,PBMC)增殖反应活性。方法 Ad-HO-1按感染强度(multiplicity of infection,MOI)100、200和400分别转染人肾小管上皮细胞株(HK-2)3 h,转染2 d后观察绿色荧光蛋白的表达,转染2、4 d后进行活细胞计数(台盼蓝染色法)。激光共聚焦法检测HO-1和GFP蛋白在HK-2中的表达,WesternBlot检测HK-2细胞HO-1蛋白表达,3H-TdR法检测HK-2转染后抑制PBMC增殖能力。结果重组腺病毒Ad-HO-1(MOI200)感染HK-2细胞2 d后90%的细胞表达绿色荧光,2、4 d时活细胞百分率与对照组无显著差异。激光共聚焦检测表达GFP的细胞均表达HO-1蛋白,位置分布基本一致。Western blot检测显示HK-2感染Ad-HO-1后有能与HO-1单抗特异性结合的蛋白免疫印迹条带,对PBMC增殖能力有显著抑制作用。结论 Ad-HO-1能介导HO-1目的蛋白的表达,具有抑制PBMC增殖的生物学活性。Objective To observe the cytotoxicity of Ad-HO-1 and its capacity of mediating the expression of HO-1 in cultured HK-2 cells. Methods The HK-2 cells were infected by Ad-HO-1 with 100, 200 and 400 MOI for 3 h, followed by the green fluorescence observation 2 days later. The live cells ratio was detected 2 and 4 days later. The expression of HO-1 and GFP in HK-2 were detected under laser scan confocal microscope. The expression of HO-1 was detected by Western blot analysis. 3H-TdR was used to assay the ability of HO-2 cells infected with Ad-HO-1 to inhibit the PBMC proliferation. Results Over 90% HK-2 cells expressed green fluorescence after infected by Ad-HO-1 （ MOI 200） for 2 days. The live cell ratio at 2 and 4 days had not any difference from those of the control group. The expressions of HO-1 and GFP in the Ad-HO-1 transfected HK-2 cells were determined under laser scan confocal microscope. The locations of these two proteins were the same. HK-2 cells infected by Ad-HO-1 had protein immune blot strap combined with HO-1 antibody. When the MOI of Adv-HO-1 was 0.01, 0.1, 1 and 10, the proliferative capacity of PBMC was inhibited significantly. Conclusion The stable expression of HO-1 mediated by Ad-HO-1 in cultured tubular epithelial cells can inhibit the cell proliferation of PBMC.

关 键 词：血红素氧化酶-1 基因转染 

分 类 号：R322.61[医药卫生—人体解剖和组织胚胎学] R392.12[医药卫生—基础医学]