摘要
目的探究脂多糖(lipopolysaccharide,LPS)诱导人肺微血管内皮细胞(human pulmonary microvascular endothelial cells,HPMEC)模型中趋化因子受体4(chemokine receptor type 4,CXCR4)、Ras同源基因家族成员A(Ras homolog gene family,member A,RhoA)表达及其对细胞增殖、凋亡的影响。方法取对数期细胞随机分为Control组、LPS组、si-CXCR4组、si-CXCR4+ov+NC组、si-CXCR4+ov+RhoA组,LPS组细胞采用LPS处理,si-CXCR4组以CXCR4-3为干扰靶点转染CXCR4干扰质粒,si-CXCR4+ov+NC组转染CXCR4干扰质粒与空白质粒,si-CXCR4+ov+RhoA组转染RhoA过表达质粒,Control组细胞不进行LPS处理以及质粒转染,在正常条件下继续培养。分别采用CCK8(cell counting kit-8)法、TUNEL法、Western blot法、酶联免疫吸附实验、实时荧光定量PCR法测定细胞增殖、细胞凋亡、细胞调网相关标志因子、炎性因子以及CXCR4、RhoA表达水平差异。结果对比Control组,LPS组的CXCR4 mRNA及蛋白表达水平均明显升高(t值分别为5.12,6.55,P值均<0.05);对比Control组与si-CXCR4+ov+NC组,si-CXCR4+ov+RhoA组的RhoA mRNA与蛋白表达水平均明显升高(F值分别为7.54,6.75,P值均<0.05);对比Control组、si-CXCR4组,LPS组的细胞活力明显降低,对比si-CXCR4+ov+NC组,si-CXCR4+ov+RhoA组细胞活力明显降低(F=6.90,P<0.05);对比Control组、si-CXCR4组,LPS组的肿瘤坏死因子-α(tumor necrosis factor,TNF-α)明显升高,对比si-CXCR4+ov+NC组,si-CXCR4+ov+RhoA组肿瘤坏死因子(tumor necrosis factor-α,TNF-α)、白细胞介素(interleukin,IL)-1β、IL-6明显升高,对比Control组、si-CXCR4+ov+NC组,LPS组的IL-1β、IL-6均明显升高(F=9.45,P<0.05);对比Control组、si-CXCR4+ov+NC组,LPS组的细胞凋亡水平均明显升高且si-CXCR4+ov+RhoA组细胞凋亡水平更高(F=9.00,P<0.05);对比Control组、si-CXCR4组,LPS组的凋亡促进基因(Bax)、活化天冬氨酸特异性半胱氨酸蛋白酶3(cleaved caspase3)以及RhoA蛋白表达均明显升高,对比si-CXCR4+ov+NC组,si-CXCR4+ov+RhoA�
ObjectiveTo explore the induction of lipopolysaccharide(LPS)in human pulmonary microvascular endothelial cells(HPMEC),The expression of chemokine receptor 4(CXCR4)and ras homolog gene family(member A,RhoA)in HPMEC cell model and its effects on cell proliferation and apoptosis.MethodsLogarithmic HPMEC cells were randomly divided into Control group,LPS group,si-CXCR4 group,si-CXCR4+ov+NC group and si-CXCR4+ov+RhoA group,and the cells in LPS group were treated with LPS.The si-CXCR4 group transfected CXCR4 interference plasmid with CXCR4-3 as the interference target,the si-CXCR4+ov+NC group transfected CXCR4 interference plasmid and blank plasmid,the si-CXCR4+ov+RhoA group transfected RhoA overexpression plasmid,and the cells in the Control group were not treated with LPS or transfected with plasmid.Continue cultivation under normal conditions.CCK8(cell counting kit-8),TUNEL,Western blot,enzyme-linked immunosorbent assay(ELISA)and real time quantitative PCR were used to determine the differences in the expression levels of cell proliferation,apoptosis,markers of cell regulatory network,inflammatory factors and CXCR4 and RhoA.ResultsCompared with the Control group,the mrna and protein expression levels of CXCR4 in the LPS group significantly increased(t values were 5.12 and 6.55,both P values<0.05);Compared with the Control group and Si-CXCR4+OV+NC group,the expression levels of RhoA mRNA and protein in Si-CXCR4+OV+RhoA group significantly increased(F values were 7.54 and 6.75,both P values<0.05);Compared with Control group and Si-CXCR4 group,the cell viability of LPS group significantly decreased,and compared with Si-CXCR4+OV+NC group,the cell viability of Si-CXCR4+OV+RhoA group significantly decreased(F=6.90,P<0.05);Compared with Control group and Si-CXCR4 group,tumor necrosis factor-α(TNF-α)in LPS group significantly increased,compared with Si-CXCR4+OV+NC group.Tnf-α,interleukin-1β(IL-1β)and interleukin-6(IL-6)significantly increased in Si-CXCR4+OV+RhoA group.Compared with Control group and Si-CXCR4+OV+NC grou
作者
孙兵
曹家军
符宜龙
唐中建
舒艾娅
SUN Bing;CAO Jiajun;FU Yilong;TANG Zhongjian;SHU Aiya(Department of Critical Care Medicine,Chongqing University Fuling Hospital,Chongqing University,Chongqing 408000,China)
出处
《国际免疫学杂志》
CAS
2024年第5期471-478,共8页
International Journal of Immunology
基金
重庆市自然科学基金面上项目(cstc2021jcyj-msxmX0682)。
