高迁移率族蛋白A2加重脂多糖诱导的内皮细胞损伤

High mobility group A2 aggravates lipopolysaccharide-induced endothelial cell injury

目的研究高迁移率族蛋白A2(HMGA2)在脂多糖诱导的人脐静脉内皮细胞(HUVEC)损伤中的作用及其机制。方法探究脂多糖刺激下HMGA2表达变化,将细胞分为2组,空白模型组,空白对照组。探究HMGA2过表达对内皮细胞损伤的影响,将细胞分为4组,对照组,HMGA2组,模型组,实验组。采用脂多糖(100 ng/ml)刺激12 h诱导HUVEC损伤。采用Western blot检测蛋白表达以及筛选可能的信号通路;采用PCR检测炎性细胞因子的转录水平;TNUEL染色检测细胞凋亡;荧光探针二乙酰二氯氢化荧光素检测细胞活性氧水平;硝酸酶还原法检测细胞内NO水平。结果Western blot检测结果显示,与空白对照组比较,空白模型组HMGA2表达明显增高,差异有统计学意义(0.40±0.03 vs 0.14±0.01,P<0.05)。与模型组比较,实验组白细胞介素1、白细胞介素6和TNF-αmRNA表达明显上调;凋亡阳性细胞数目增多;活性氧荧光强度增强;NO水平明显升高;磷酸化IκB、磷酸化p65和诱导型一氧化氮合酶(iNOS)表达明显增高,差异有统计学意义(P<0.05)。结论脂多糖刺激可以诱导HUVEC的HMGA2表达上调,并且HMGA2通过激活NF-κB/iNOS/NO信号通路促进内皮细胞损伤。

Objective To study the role of HMGA2 in lipopolysaccharide(LPS)-induced injury of HUVEC and its mechasnism.Methods The HUVEC were divided into blank model group and blank control group,and into control group 1,HMGA2 group,model group,experimental group.The injury of HUVEC was induced by LPS(100 ng/ml)stimulation for 12 h.The expressions of HMGA2,p-IκB,p-p65,iNOS and their signaling pathways were detected by Western blot.The transcription of inflammatory cytokines was detected by PCR.The apoptosis of HUVEC was assayed with TNUEL staining.The cellular reactive oxygen species was measured with fluorescent probe and the serum NO level was measured using nitrate reduction method.Results The expression level of HMGA2 was significantly higher in blank model group than in blank control group(0.40±0.03 vs 0.14±0.01,P<0.05).The expression levels of IL-1,IL-6 and TNF-αmRNA were significantly higher,the number of apoptotic positive HUVEC was significantly larger,the fluorescence intensity of reactive oxygen species,serum NO level and expression levels of p-IκB,p-p65 and iNOS were significantly higher in experimental group than in model group(P<0.05).Conclusion LPS can upregulate the expression of HMGA2 in HUVEC.HMGA2 promotes the endothelial cell injury by activating the NF-κB,iNOS and NO signaling pathways.