miR-21在脓毒症大鼠肺损伤中的作用:与TRPM2表达的关系

Role of microRNA-21 in sepsis-induced lung injury in rats:relationship with TRPM2 expression

目的评价miR-21在脓毒症大鼠肺损伤中的作用及其与瞬时受体电位M2(TRPM2)表达的关系。方法清洁级健康Wistar大鼠48只,雌雄各半,体重250~300 g,采用随机数字表法分为4组(n=12):假手术组(SH组)、脓毒症组(S组)、miR-21抑制剂组(MI组)和miR-21抑制剂+TRPM2阻断剂Gd3+组(MIG组)。采用盲肠结扎穿孔(CLP)法制备大鼠脓毒症模型。于CLP前12 h,MI组和MIG组分别尾静脉注射miR-21抑制剂和miR-21抑制剂+TRPM2阻断剂组Gd3+。于CLP后24 h时取颈动脉血样,行血气分析,记录PaO2,计算氧合指数;随后处死大鼠取肺组织,光镜下行肺损伤评分,确定湿重/干重(W/D)比值,采用qRT-PCR法检测miR-21和TRPM2的mRNA表达,分光光度计比色法检测MDA和SOD水平,ELISA法检测血清IL-6、IL-8和TNF-α的浓度。结果与SH组比较,其余3组氧合指数和肺组织SOD活性降低,肺组织W/D比值、肺损伤评分、MDA含量、血清IL-6、IL-8和TNF-α浓度升高,S组肺组织miR-21 mRNA表达上调,TRPM2 mRNA表达下调(P<0.05);与S组比较,MI组氧合指数、肺组织SOD活性和血清IL-6、IL-8和TNF-α浓度升高,肺组织W/D比值、肺损伤评分、MDA含量降低,MI组和MIG组肺组织miR-21 mRNA表达下调,TRPM2 mRNA表达上调(P<0.05);与MI组比较,MIG组氧合指数和肺组织SOD活性降低,肺组织W/D比值、肺损伤评分、MDA含量和血清IL-6、IL-8和TNF-α浓度升高,肺组织TRPM2 mRNA表达下调(P<0.05)。结论miR-21表达上调,TRPM2表达下调参与了脓毒症大鼠肺损伤的过程。

Objective To evaluate the role of microRNA-21(miR-21)in sepsis-induced lung injury and the relationship with transient receptor potential melastatin 2(TRPM2)expression in rats.Methods Forty-eight clean-grade Wistar rats,half male and half female,weighing 250-300 g,were divided into 4 groups(n=12 each)by a random number table method:sham operation group(SH group),sepsis group(S group),miR-21 inhibitor group(group MI),and miR-21 inhibitor plus TRPM2 blocker Gd3+group.Sepsis was induced by cecal ligation and puncture(CLP)in anesthetized rats.In MI group and MIG group,miR-21 inhibitor and miR-21 inhibitor plus TRPM2 blocker Gd3+were injected through the tail vein,respectively,at 12 h before CLP.Carotid artery blood samples were collected at 24 h after CLP for blood gas analysis,PaO2 was recorded,and oxygenation index was calculated.Animals were sacrificed,and lung tissues were removed for microscopic examination of the pathological changes which were scored and for determination of wet to dry weight ratio(W/D ratio),expression of miR-21 and TRPM2 mRNA(quantitative real-time polymerase chain reaction),levels of malondialdehyde(MDA)and superoxide dismutase(SOD)(by spectrophotometer colorimetry)and levels of interleukin-6(IL-6),IL-8 and tumor necrosis factor-alpha(TNF-α)in serum(by enzyme-linked immunosorbent assay).Results Compared with group SH,the oxygenation index and SOD activity were significantly decreased,and W/D ratio,lung injury score,MDA content,serum IL-6,IL-8 and TNF-αconcentrations were increased in the other three groups,and the expression of miR-21 mRNA was up-regulated,and the expression of TRPM2 mRNA was down-regulated in group S(P<0.05).Compared with group S,the oxygenation index,SOD activity and serum IL-6,IL-8 and TNF-αconcentrations were significantly increased,and W/D ratio,lung injury score and MDA content were decreased in group MI,and the expression of miR-21 mRNA was down-regulated,and the expression of TRPM2 mRNA was up-regulated in MI and MIG groups(P<0.05).Compared with group MI,the oxygenation index and SOD activity were significantly decreased,W/D ratio,lung injury score,MDA content,serum IL-6,IL-8 and TNF-αconcentrations were increased,and the expression of TRPM2 mRNA was down-regulated in group MIG(P<0.05).Conclusion Up-regulated expression of miR-21 and down-regulated expression of TRPM2 are involved in the process of sepsis-induced lung injury in rats.