microRNA-1285-3p在脓毒症急性肺损伤中的表达及其与炎症因子水平的相关性

The expression of microRNA-1285-3p in sepsis-induced acute lung injury and its correlation with inflammatory factors

目的探讨microRNA(miRNA)-1285-3p在脓毒症急性肺损伤(ALI)中的表达及其与炎症因子、肿瘤坏死因子(TNF)-α、白介素(IL)-6水平的相关性。方法对大鼠进行腹腔注射内毒素脂多糖(LPS)诱导急性肺损伤大鼠模型,选取相同数量的大鼠作为对照组;各模型组大鼠注射LPS 4 h、8 h、12 h以及24 h后,观察各组大鼠肺组织病理学变化,并检测肺组织中microRNA-1285-3p、TNF-α与IL-6的含量变化。利用LPS刺激大鼠肺泡巨噬细胞(NR8383细胞),在4 h、8 h、12 h以及24 h不同时间点检测细胞中microRNA-1285-3p、TNF-α与IL-6的含量变化,并分析细胞中microRNA-1285-3p的表达与TNF-α、IL-6表达的相关性。以生物信息学方法对microRNA-1285-3p及其靶基因进行预测,上调或下调NR8383细胞microRNA-1285-3p表达,以LPS刺激后,12 h后观察microRNA-1285-3p的表达变化,利用蛋白免疫印迹试验(Western blot)检测24 h后细胞中microRNA-1285-3p靶蛋白与NF-κB的表达变化。结果LPS 4 h组、LPS 8 h组、LPS 12 h组、LPS 24 h组肺组织及NR8383细胞系中的microRNA-1285-3p表达量均分别低于对照组(P<0.05),而TNF-α、IL-6水平均高于对照组(P<0.05);microRNA-1285-3p表达与TNF-α、IL-6均呈负相关关系(均P<0.05);microRNA-1285-3p mimic组Notch1蛋白(Notch信号通路受体蛋白之一)的表达低于对照组(P<0.05);microRNA-1285-3p抑制剂(inhibitor)组Notch1蛋白的表达水平高于对照组(P<0.05)。结论microRNA-1285-3p可能通过调控脓毒症急性肺损伤大鼠的Notch1蛋白的表达,影响炎症因子TNF-α与IL-6的表达,参与其免疫炎症调控过程。

The purpose of this study is to investigate the expression of microrna-1285-3 p in acute lung injury(ALI)of sepsis and its correlation with the levels of TNF-a and IL-6.Firstly,rats were injected with LPS to induce acute lung injury,and the same number of rats were selected as the control group.After injection of LPS for 4 h,8 h,12 h and 24 h,the pathological changes of lung tissue were observed,and the contents of microrna-1285-3 p,TNF-αand IL-6 were detected.The contents of microrna-1285-3 p,TNF-αand IL-6 in nr8383 cells stimulated by LPS were measured at 4 h,8 h,12 h and 24 h,and the correlation between the expression of microrna-1285-3 p and the expression of TNF-αand IL-6 were analyzed.Meanwhile,microrna-1285-3 p and its target gene were predicted by bioinformatics method.The nr8383 cells,in which microrna-1285-3 p expression was up-regulated or down regulated,were stimulated with LPS,and then the expression of microrna-1285-3 p was observed 12 hours later,and the expression of microrna-1285-3 p target protein and NF-κB were detected 24 hours later by Western blotting.Data showed that the expression of microrna-1285-3 p in lung tissue and nr8383 cell line of LPS 4 h group,LPS 8 h group,LPS 12 h group,LPS 24 h group were all significantly lower than that of the control group(P<0.05),while the levels of TNF-αand IL-6 were significantly higher than those in the control group(P<0.05);the expression of microrna-1285-3 p was negatively correlated with TNF-αand IL-6(P<0.05);the expression of Notch 1 protein in microrna-1285-3 p mimic group was lower than that in the control group(P<0.05),while the expression of Notch 1 protein in microrna-1285-3 p inhibitor group was higher than that in the control group(P<0.05).All these result suggests that microrna-1285-3 p may play an important role in the regulation of immune inflammation by regulating the expression of Notch-1 protein and the expression of TNF-αand IL-6.