上调miR-126对LPS诱导的小鼠急性肺损伤的保护作用及机制

Protective effect and mechanism of miR-126 up-regulation against LPS-induced acute lung injury in mice

目的探讨上调微小RNA(miR)-126对小鼠急性肺损伤(acute lung injury,ALI)的影响及可能的作用机制。方法将40只雄性C57BL/6小鼠随机分为control组、ALI组、ALI+agomiR-NC和ALI+agomiR-126组,每组10只。利用腹腔注射脂多糖(LPS)诱导建立ALI小鼠模型。ALI+agomiR-NC组和ALI+agomiR-126组在建立ALI模型后,分别给予尾静脉注射agomiR-NC和agomiR-126,control组仅以等量的生理盐水进行腹腔和尾静脉注射。建模后24 h行血气分析检测小鼠动脉血氧分压(PaO 2)、二氧化碳分压(PaCO 2)和pH值。处死所有小鼠,制备支气管肺泡灌洗液(BALF),ELISA法测定BALF中IL-1β、TNF-α和IL-6的浓度;取小鼠肺组织,行HE染色观察病理改变并进行肺损伤评分,计算肺湿重/干重(W/D)比值,测定髓过氧化物酶(MPO)活性,采用qRT-PCR检测肺组织中miR-126的表达,采用Western blot法测定肺组织HMGB1蛋白的表达。结果与control组比较,ALI组小鼠动脉血pH值和PaO 2明显降低(P<0.05),BALF中IL-1β、TNF-α和IL-6的含量明显增多(P<0.05),肺损伤评分、W/D比值、MPO活性明显升高(P<0.05),肺组织miR-126表达明显降低(P<0.05),HMGB1蛋白表达明显升高(P<0.05)。与ALI组比较,ALI+agomiR-NC组各指标差异无统计学意义(P>0.05)。与ALI+agomiR-NC组比较,ALI+agomiR-126组小鼠动脉血pH值和PaO 2明显升高(P<0.05),BALF中IL-1β、TNF-α和IL-6的含量明显减少(P<0.05),肺损伤评分、W/D比值、MPO活性明显降低(P<0.05),肺组织miR-126表达明显升高(P<0.05),HMGB1蛋白表达明显降低(P<0.05)。结论miR-126在ALI小鼠肺组织中表达降低,上调miR-126能够减轻ALI小鼠肺组织的炎症反应,其机制可能与其对HMGB1的调控有关。

Objective To investigate the effect of up-regulation of microRNA(miR)-126 on acute lung injury(ALI)in mice and its possible mechanism.Methods Forty male C57BL/6 mice were randomly divided into control group,ALI group,ALI+agomiR-NC group,ALI+agomiR-126 group,10 mice in each group.The mouse model of ALI was established by intraperitoneal injection of lipopolysaccharide(LPS).In ALI+agomiR-NC group and ALI+agomiR-126 group,agomiR-NC and agomiR-126 were injected into caudal vein respectively after ALI model was established.In control group,only equal amount of normal saline was injected into abdominal cavity and caudal vein.The arterial partial pressure of oxygen(PaO 2),partial pressure of carbon dioxide(PaCO 2)and pH were measured by blood gas analysis 24 h after the establishment of the animal model.All mice were killed,and then bronchoalveolar lavage fluid(BALF)was prepared.The concentrations of IL-1β,TNF-αand IL-6 in BALF were determined by ELISA.The lung tissue of mice was stained with HE staining,the pathological changes of lung tissue were observed,and the lung injury score was evaluated.Furthermore,the ratio of wet weight to dry weight(W/D)was calculated,the activity of myeloperoxidase(MPO)was measured,the expression of miR-126 was detected by qRT-PCR,and the expression of HMGB1 protein was detected by Western blot.Results Compared with control group,pH value and PaO 2 of arterial blood were significantly reduced in ALI group(P<0.05),the contents of IL-1β,TNF-αand IL-6 in BALF were significantly increased(P<0.05),the lung injury score,W/D ratio and MPO activity were significantly increased(P<0.05),the expression of miR-126 in lung tissue was significantly decreased(P<0.05),and the expression of HMGB1 protein was significantly increased(P<0.05).There was no significant difference in each index between ALI group and ALI+agomiR-NC group(P>0.05).Compared with ALI+agomiR-NC group,pH value and PaO 2 of arterial blood were significantly increased in ALI+agomiR-126 group(P<0.05),the contents of IL-1β,TNF-αand IL-6 in BALF were significantly decreased(P<0.05),the lung injury score,W/D ratio and MPO activity were significantly reduced(P<0.05),the expression of miR-126 in lung tissue was significantly enhanced(P<0.05),and the expression of HMGB1 protein was significantly decreased(P<0.05).Conclusion The expression of miR-126 in the lung tissue of ALI mice is decreased,and the up-regulation of miR-126 can reduce the inflammatory response of ALI mice,which may be related to the regulation of HMGB1.