血红素氧化酶-1基因过表达对RAW264.7细胞增殖和凋亡的影响  

Effect of heme oxygenase-1 over expression on proliferation and apoptosis of RAW264.7 macrophages

作　　者：周茂彬[1,2] 李鑫[3] 李琦[3] 姬文婕[3] 焉力方 闫睿[4] 周欣[1,2] 李玉明[1,2] 

机构地区：[1]武警后勤学院附属医院心血管病研究所,天津300162 [2]天津市心血管重塑与靶器官损伤重点实验室,天津300162 [3]武警后勤学院附属医院呼吸与重症医学科,天津300162 [4]武警后勤学院附属医院胸外科,天津300162

出　　处：《武警后勤学院学报（医学版）》2017年第4期277-280,共4页Journal of Logistics University of PAP（Medical Sciences）

基　　金：国家自然科学基金资助项目(81570335);武警后勤学院附属医院种子基金项目(FYM201538);武警后勤学院中心实验室开放基金项目(2015ZXKF11)

摘　　要：【目的】建立血红素氧化酶-1(heme oxygenase-1,HO-1)稳定过表达细胞株RAW264.7,探讨HO-1过表达对其增殖活性和细胞凋亡的作用。【方法】用携带小鼠HO-1基因的重组慢病毒感染小鼠RAW264.7细胞,经嘌呤霉素筛选获得HO-1稳定过表达细胞株。细胞分为野生型组(WT)、阴性对照组(NC)和过表达组(Lenti-HO-1)。用Real-Time PCR检测细胞中HO-1mRNA的表达。用Western blotting检测细胞中HO-1蛋白的表达。用CCK-8法检测细胞的增殖活性。用流式细胞术检测细胞的凋亡水平。【结果】HO-1慢病毒能促进RAW264.7细胞HO-1 mRNA和蛋白的稳定过表达。Lenti-HO-1组细胞的增殖活性显著低于WT组和NC组(P<0.05)。Lenti-HO-1组凋亡细胞的比例显著高于NC组(P<0.05)。【结论】本研究成功构建了HO-1稳定过表达的RAW264.7细胞株,HO-1过表达能够降低RAW264.7细胞的增殖活性,增加凋亡细胞的比例,其为进一步探讨HO-1在巨噬细胞所参与病理生理机制中的作用奠定了基础。【Objective】To establish a cell line of stable over-expressing of heme oxygenase-1（HO-1） in mouse RAW264.7 macrophages, and to investigate the effect of HO-1 over-expression on the proliferation and apoptosis in this cell line.【Methods】The lentiviral vector of HO-1 gene（Lenti-HO-1） was transfected into mouse RAW264.7 macrophages, followed by puromycin selection to obtain the cell line with stably over-expressed HO-1. The cells were then divided into wild type group（WT）, negative control group（NC） and over expression group（Lenti-HO-1）. The HO-1 mRNA expression level was measured by real-time PCR, and HO-1 protein expression level was determined by Western blot. The cell proliferative activity was analyzed by CCK-8 assay. The cell apoptosis was detected by flow cytometry.【Results】The Lenti-HO-1 tranfection led to a stable over expression of HO-1 mRNA, as well as in protein level in RAW264.7 cells. The HO-1 over-expressed cells showed lower proliferative activity than WT and NC cells（all P〈0.05）. In addition,the proportion of apoptotic cells in Lenti-HO-1 group was significantly higher than that in group WT and NC（all P〈0.05）.【Conclusion】Using lentiviral vector, we established HO-1 overexpression cell line of mouse RAW264.7 macrophage background, which showed reduced proliferative activity and enhanced apoptotic response. This cell line may be useful for examining the role of HO-1 mediated, macrophage participated physiology and pathological processes.

关 键 词：血红素氧化酶-1 RAW264.7巨噬细胞 细胞增殖 细胞凋亡 慢病毒 

分 类 号：R34[医药卫生—基础医学]