载脂蛋白M通过内皮型一氧化氮合酶发挥抗炎作用

Apolipoprotein M exhibits anti-inflammatory property by endothelial nitric oxide synthase

目的探讨载脂蛋白M(apoM)的抗炎作用与内皮型一氧化氮合酶(eNOS)表达的关系。方法用携带apoM基因的慢病毒(apoM^(Tg))和空载慢病毒(apoM^(TgN))分别转染人脐静脉细胞融合细胞株EA.hy926,随后分为apoM^(Tg)+TNF-α组、apoM^(TgN)+TNF-α组、apoM^(Tg)组和apoM^(TgN)组,采用RT-PCR检测细胞中apoM和eNOS mRNA表达,Western blot法检测apoM和eNOS蛋白表达以及eNOS磷酸化水平。选取16只apoM基因野生型(apoM^(+/+))小鼠随机均分为apoM^(+/+)+脂多糖(LPS)组和apoM^(+/+)组,16只apoM基因敲除型(apoM^(-/-))小鼠随机均分为apoM^(-/-)+LPS组和apoM^(-/-)组,采用RT-PCR检测小鼠肾脏组织中apoM和eNOS mRNA表达。结果与转染apoM^(TgN)的细胞相比,转染apoM^(Tg)的细胞中apoM mRNA和蛋白表达、eNOS mRNA表达和磷酸化水平增加(P<0.05或P<0.01),而eNOS蛋白表达差异无统计学意义(P>0.05)。apoM^(Tg)+TNF-α组的apoM和eNOS mRNA表达高于apoM^(Tg)组和apoM^(TgN)+TNF-α组(P<0.01);而apoM^(TgN)+TNF-α组与apoM^(TgN)组间apoM和eNOS mRNA表达差异无统计学意义(P>0.05)。apoM^(+/+)组、apoM^(-/-)+LPS组与apoM^(-/-)组的eNOS mRNA表达差异均无统计学意义(P>0.05);而apoM^(+/+)+LPS组的eNOS mRNA表达高于apoM^(+/+)组和apoM^(-/-)+LPS组(P<0.05)。结论apoM的抗炎作用可能与其促进eNOS表达和磷酸化水平有关。

Objective To investigate the relationship between the effect of anti-inflammation of apolipoprotein M(apoM)and the expression of endothelial nitric oxide synthase(eNOS).MethodsThe human umbilical vein cell fusion cell line EA.hy926 was transfected by lentivirus carrying apoM gene(apoM^(Tg))or empty vector(apoM^(TgN)),and then divided into four groups of apoM^(Tg)+TNF-α,apoM^(TgN)+TNF-α,apoM^(Tg)and apoM^(TgN).The mRNA expressions of apoM and eNOS in EA.hy926 cells were determined by RT-PCR,and the protein expressions of apoM and eNOS and phosphorylation level of eNOS were detected by Western blot.Sixteen mice with apoM wild-type(apoM^(+/+))were equally randomized into two groups of apoM^(+/+)+LPS and apoM^(+/+),meanwhile 16 mice with apoM knock-out(apoM^(-/-))were equally randomized into two groups of apoM^(-/-)+LPS and apoM^(-/-),respectively.The mRNA expressions of apoM and eNOS in the kidney tissues of mice were measured by RT-PCR.Results Compared with the cells transfected with apoM^(TgN),the mRNA and protein expressions of apoM,and mRNA expression and phosphorylation level of eNOS were increased(P<0.05 or P<0.01),while the protein expression of eNOS was not significantly changed in the cells transfected with apoM^(Tg)(P>0.05).The mRNA expressions of apoM and eNOS in apoM^(Tg)+TNF-αgroup were higher than those in apoM^(Tg)group and apoM^(TgN)+TNF-αgroup(P<0.01).There was no statistical difference in the mRNA expressions of apoM and eNOS between groups of apoM^(TgN)+TNF-αand apoM^(TgN)(P>0.05).The mRNA expressions of eNOS in groups of apoM^(+/+)and apoM^(-/-)+LPS were similar to those in apoM^(-/-)group(P>0.05).While the mRNA expression of eNOS in apoM^(+/+)+LPS group was higher than that in groups of apoM^(+/+)and apoM^(-/-)+LPS(P<0.05).Conclusion apoM has the anti-inflammation effect probably via enhancing the expression and phosphorylation level of eNOS.