【摘要】 目的:基于高迁移率族蛋白B1(HMGB1)信号通路探讨乌司他丁对早期放射性肺损伤的保护机制。方法:根据干预方式的不同将36只SD大鼠分为正常对照组、放射性肺损伤组及乌司他丁组,每组12只。收集各组大鼠肺支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)和肺组织,对肺组织进行苏木精-伊红染色,观察肺组织形态学变化,采用瑞式-姬姆萨染色法测定BALF中各白细胞种类计数,采用蛋白免疫印迹法测定肺组织中HMGB1蛋白及下游基因蛋白表达水平,采用实时荧光定量聚合酶链式反应测定肺组织中HMGB1及下游基因mRNA表达水平。结果:正常对照组大鼠肺组织支气管、黏膜下及肺间质中无炎症细胞浸润,放射性肺损伤组大鼠肺组织支气管、黏膜下及肺间质中有大量炎症细胞浸润,肺间质增厚,乌司他丁组大鼠肺组织支气管、黏膜下及肺间质中炎症细胞较放射性肺损伤组明显减少,肺间质水肿明显缓解;与正常对照组比较,放射性肺损伤组和乌司他丁组大鼠BALF中嗜酸性粒细胞、中性粒细胞、淋巴细胞及单核细胞比例升高,HMGB1、晚期糖基化终末产物受体(RAGE)、核转录因子κB(NF-κB)、肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)及白细胞介素1β(IL-1β)蛋白表达水平明显升高,HMGB1、RAGE、NF-κB、TNF-α、IL-6及IL-1βmRNA表达水平明显升高,差异均有统计学意义(P<0. 05);与放射性肺损伤组比较,乌司他丁组大鼠BALF中嗜酸性粒细胞、中性粒细胞、淋巴细胞及单核细胞比例降低,HMGB1、RAGE、NF-κB、TNF-α、IL-6及IL-1β蛋白表达水平明显降低,肺组织中HMGB1、RAGE、NF-κB、TNF-α、IL-6及IL-1βmRNA表达水平明显降低,差异均有统计学意义(P<0. 05)。结论:乌司他丁可通过降低HMGB1及其下游蛋白表达,中断HMGB1信号通路,减轻早期放射性肺损伤大鼠肺部炎症细胞浸润,有可能成为预防和治疗早期放射性肺损伤的药物。更多还原

【Abstract】 OBJECTIVE: To probe into the protective mechanism of ulinastatin on early radiation-induced pulmonary injury based on the signal pathway of high mobility group box-1 protein( HMGB1). METHODS: 36 SD rats were divided into normal control group,radiation-induced pulmonary injury group and ulinastatin group according to different intervention methods,with 12 cases in each group. The bronchoalveolar lavage fluid( BALF) and lung tissue of rats in each group were collected, the lung tissue was stained with hematoxylin-eosin so as to observe its morphological change. Ray-giemsa staining method was adopted to measure various white blood cells counts in BALF,Western blotting method was adopted to measure the expression levels of HMGB1 protein and downstream gene protein in lung tissue,real-time fluorescence quantification polymerase chain reaction was adopted to measure the expression levels of HMGB1 and downstream gene mRNA in lung tissue. RESULTS: There was no inflammatory cell infiltration in the bronchus,submucosa and pulmonary interstitium of rats in normal control group; while there was a large number of inflammatory cells infiltrated in the bronchus,submucosa and pulmonary interstitium of rats in the radiation-induced pulmonary injury group, with pulmonary interstitial thickening; the number of inflammatory cells in bronchi,submucosa and pulmonary interstitium of rats in the ulinastatin group was significantly less than that of the radiationinduced lung injury group,and pulmonary interstitial edema was significantly alleviated; compared with normal control group,the proportions of eosinophilic granulocyte,neutrophile granulocyte,lymphocyte and monocyte in the BALF of rats in radiation-induced pulmonary injury group and ulinastatin group were increased,the protein expression levels of HMGB1,receptor for advanced glycation end products( RAGE),nuclear transcription factor-κB( NF-κB),tumor necrosis factor α( TNF-α),interleukin-6( IL-6) and interleukin-1β( IL-1β) were increased,the mRNA expression levels of HMGB1,RAGE,NF-κB,TNF-α,IL-6 and IL-1β were increased,with statistically significant differences( P<0. 05); compared with radiation-induced pulmonary injury group,the proportions of eosinophilic granulocyte,neutrophile granulocyte,lymphocyte and monocyte in the BALF of rats in ulinastatin group were decreased,the protein expression levels of HMGB1,RAGE,NF-κB,TNF-α,IL-6 and IL-1β were decreased,the mRNA expression levels of HMGB1,RAGE,NF-κB,TNF-α,IL-6 and IL-1β were decreased,with statistically significant differences( P <0. 05). CONCLUSIONS: Ulinastatin can relieve pulmonary inflammatory infiltration in rats with early radiation-induced pulmonary injury by interrupting HMGB1 signaling pathway by reducing the expression of HMGB1 and its downstream protein,which is likely to be a drug to prevent and treat early radiation-induced pulmonary injury.更多还原