乌司他丁调控MKP-1靶点改善脓毒症小鼠肺组织炎性损伤的机制研究

The mechanism of ulinastatin regulating MKP-1 to improve lung tissue inflammatory injury in septic mice

目的探讨乌司他丁调控MKP-1靶点改善脓毒症小鼠肺组织炎性损伤的机制。方法实验分成假手术组(Sham)、脓毒症模型组(Model)、脓毒症模型乌司他丁处理组(ulinastatin)和脓毒症模型乌司他丁+siRNA对照处理组(ulinastatin+si-NC)、脓毒症模型乌司他丁+MKP-1 siRNA处理组(ulinastatin+si-MKP-1),qRT-PCR检测MKP-1 mRNA表达,Western blot方法分析MKP-1、iNOS、p-JNK、JNK、p-p38和p38蛋白表达变化,检测肺组织湿干比重,苏木素-伊红(HE)染色对肺组织损伤程度进行评分,ELISA法检测肺组织中TNF-α、IL-6和IL-1β含量,Griess法检测肺组织中NO含量。结果与Sham组比较,Model组小鼠肺组织中MKP-1 mRNA和蛋白表达水平均降低,肺组织湿干比重和肺损伤评分均升高,TNF-α、IL-6、IL-1β和NO含量升高,iNOS蛋白表达增多,p-JNK/JNK、p-p38/p38蛋白水平升高(P<0.05)。与Model组比较,ulinastatin组小鼠肺组织中MKP-1 mRNA和蛋白表达水平均升高,肺组织湿干比重和肺损伤评分均降低,TNF-α、IL-6、IL-1β和NO含量降低,iNOS蛋白表达减少,p-JNK/JNK、p-p38/p38蛋白水平降低(P<0.05)。与ulinastatin+si-NC组比较,ulinastatin+si-MKP-1组小鼠肺组织中MKP-1 mRNA和蛋白表达水平均降低,肺组织湿干比重和肺损伤评分均升高,TNF-α、IL-6、IL-1β和NO含量升高,iNOS蛋白表达增多,p-JNK/JNK、p-p38/p38蛋白水平均升高(P<0.05)。结论乌司他丁上调MKP-1降低JNK和p38信号通路激活水平减轻脓毒症小鼠肺组织炎性损伤。

Objective To investigate the mechanism of ulinastatin regulating MKP-1 to improve lung tissue inflammatory injury in septic mice.Methods The experiment was divided into Sham,Model(sepsis model),ulinastatin(sepsis model,ulinastatin treatment),ulinastatin+si-NC(sepsis model,ulinastatin and siRNA control treatment),ulinastatin+si-MKP-1 group(sepsis model,ulinastatin and MKP-1 siRNA treatment),qRT-PCR was used to detect the expression of MKP-1 mRNA,Western blot analysis of MKP-1,iNOS,p-JNK,JNK,p-p38 and p38 proteins,detection of wet-to-dry specific gravity of lung tissue,HE staining to score lung tissue damage,ELISA method to detect TNF-α,IL-6,IL-1βcontent in lung tissue,and Griess method to detect the content of NO.Results Compared with the Sham group,the expression levels of MKP-1 mRNA and protein in the lung tissues of the Model group decreased,and the wet-to-dry ratio of lung tissues and lung injury scores were increased,TNF-α,IL-6,IL-1β,NO increased,iNOS protein expression increased,p-JNK/JNK,pp38/p38 protein levels increased(P<0.05).Compared with the Model group,the expression levels of MKP-1 mRNA and protein in the lung tissues of the ulinastatin group increased,the wet-to-dry ratio of the lung tissues and lung injury scores were reduced,and the contents of TNF-α,IL-6,IL-1β,and NO decreased,iNOS protein expression decreased,p-JNK/JNK,p-p38/p38 protein levels decreased(P<0.05).Compared with the ulinastatin+si-NC group,the expression of MKP-1 mRNA and protein in the lung tissue of the ulinastatin+si-MKP-1 group was decreased,and the wet-dry proportion of lung tissue and lung injury score was increased,the content of TNF-α,IL-6,IL-1β,NO was increased,the expression of iNOS protein was increased,and the protein levels of p-JNK/JNK and p-p38/p38 was increased(P<0.05).Conclusion Ulinastatin can up-regulate MKP-1 and reduce the activation levels of JNK and p38 signaling pathways to reduce the inflammatory injury of lung tissue in septic mice.