血红素氧化酶1调控巨噬细胞极化对动脉粥样硬化的影响

Effect of heme oxidase 1 regulating macrophage polarization on atherosclerosis

目的探讨血红素氧化酶1(HO-1)调节巨噬细胞对动脉粥样硬化的作用。方法100 mg/L脂多糖(LPS)和20 mg/L白介素4(IL-4)分别诱导RAW264.7细胞24 h,提取信使核糖核酸(mRNA)和蛋白,实时定量聚合酶链式反应法(RT-qPCR)检测LPS和IL-4诱导后巨噬细胞中HO-1、CD163和诱导型一氧化氮合酶(iNOS)mRNA表达,蛋白质印迹法(Western blot)检测LPS和IL-4诱导巨噬细胞后HO-1蛋白表达。大体油红O染色检测载脂蛋白E敲除(ApoE-/-)小鼠喂食高脂饮食后主动脉斑块面积变化,RT-qPCR和Western blot检测主动脉中HO-1 mRNA和蛋白表达。组织免疫荧光检测HO-1在主动脉窦部斑块中与促炎型(M1)和抑炎型(M2)巨噬细胞的共定位表达。结果LPS和IL-4成功诱导巨噬细胞分化为M1型与M2型巨噬细胞;IL-4诱导巨噬细胞中HO-1表达显著高于LPS及对照组(P<0.05);动脉粥样硬化模型构建成功;HO-1在ApoE-/-小鼠中表达显著高于C57BL/6对照组(P<0.05),主动脉窦中HO-1与CD206的共定位表达显著高于HO-1与CD86。结论HO-1在M2型巨噬细胞中的表达显著高于M1型巨噬细胞,在动脉硬化发展中起保护作用。

Objective To investigate the effect of heme oxygenase-1 on the regulation polarization of macrophages during the development of atherosclerosis.Methods The mRNA and protein of RAW264.7 cells were extracted by inducing RAW264.7 cells with 100 mg/L lipopolysaccharide(LPS)and 20 mg/L interleukin-4(IL-4)for 24 h respectively.The expression of HO-1,CD163 and Inducible nitric oxide synthetase(iNOS)mRNA in macrophages induced by LPS and IL-4 was detected by real-time quantitative polymerase chain reaction(RT-qPCR),and the expression of HO-1 protein in macrophages induced by LPS and IL-4 was detected by Western blot.The changes of aortic plaque area after apoE knockout(ApoE-/-)mice were detected by gross Oil Red O staining,and HO-1 mRNA and protein expression were detected by RT-qPCR and Western blot.The colocalization of HO-1 with macrophages of pro-inflammatory type(M1)and anti-inflammatory type(M2)was detected by tissue immunofluorescence.Results LPS and IL-4 successfully induced M1 and M2 macrophages.The expression of HO-1 in macrophages induced by IL-4 was significantly higher than that in LPS and control group(P<0.05).The establishment of atherosclerosis model was successful.The expression of HO-1 in ApoE-/-mice was significantly higher than that in C57BL/6 control group(P<0.05).The colocalization expression of HO-1 and CD206 in aortic sinus was significantly higher than that in HO-1 and CD86.Conclusions HO-1 expression in M2 macrophages is significantly higher than M1 macrophages,which plays a protective role in the development of arteriosclerosis.