信号转导和转录激活蛋白STAT1/3/5在高氧性急性肺损伤中的作用及机制研究  

Role and mechanism of signal transducer and activator of transcription protein STAT1/3/5 in hyperoxia-induced acute lung injury

作　　者：梅鸿 冯帮海 刘君亚 陈淼 覃松 Mei Hong;Feng Banghai;Liu Junya;Chen Miao;Qin Song(The Second Ward,Department of Critical Care Medicine,Affiliated Hospital of Zunyi Medical University,Zunyi 563000,Guizhou,China;Department of Critical Care Medicine,Zunyi Hospital of Traditional Chinese Medicine,Zunyi 563000,Guizhou,China)

机构地区：[1]遵义医科大学附属医院重症医学科,贵州遵义563000 [2]遵义市中医院重症医学科,贵州遵义563000

出　　处：《中华危重病急救医学》2022年第6期602-607,共6页Chinese Critical Care Medicine

基　　金：国家自然科学基金(81960362,81960024)。

摘　　要：目的探讨信号转导和转录激活蛋白(STAT1/3/5)是否对高氧性急性肺损伤(HALI)具有保护作用及其机制。方法将70只C57BL/6J小鼠按随机数字表法分为常氧对照组、HALI组、STAT1/3/5抑制剂组共5组,每组14只。在90%以上高氧中暴露48 h建立HALI模型;3个STAT抑制剂组分别于制模前1周腹腔注射STAT1抑制剂40 mg/kg、STAT3抑制剂5 mg/kg、STAT5抑制剂10 mg/kg,连续预处理1周。每组随机取6只采血,采用实时荧光定量反转录-聚合酶链反应(RT-PCR)检测微小RNA-21(miR-21)表达。随后处死小鼠取肺组织,采用酶联免疫吸附试验(ELISA)检测肿瘤坏死因子-α(TNF-α)、白细胞介素(IL-6、IL-1β)、超氧化物歧化酶(SOD)、丙二醛(MDA)、基质金属蛋白酶9(MMP9)含量,计算肺组织含水量,在光镜下观察肺组织病理学变化并进行肺损伤病理评分,采用蛋白质免疫印迹试验(Western blotting)检测肺组织磷酸化STAT(p-STAT1、p-STAT3、p-STAT5)表达。采用Kaplan-Meier生存曲线分析每组剩余8只小鼠7 d累积生存率。结果光镜下可见HALI组及STAT1抑制剂组肺泡结构破坏,肺泡和肺间质大量中性粒细胞(NEU)浸润,肺间质增厚,肺损伤病理评分及肺组织含水量明显升高;STAT3抑制剂组和STAT5抑制剂组肺泡腔清晰,NEU浸润程度和肺间质厚度不及HALI组,肺损伤病理评分和肺组织含水量明显降低,STAT3抑制剂组更为明显。与常氧对照组比较,HALI组TNF-α、IL-6、IL-1β、MDA和MMP9含量及p-STAT3、p-STAT5表达水平均明显升高,而SOD和miR-21表达则明显下降。与HALI组比较,STAT3抑制剂组及STAT5抑制剂组TNF-α、IL-6、IL-1β、MDA和MMP9含量均明显下降,而SOD及miR-21表达明显升高,以STAT3抑制剂组更为明显〔TNF-α(μg/L):42.53±3.25比86.36±5.48,IL-6(ng/L):68.46±4.28比145.00±6.89,IL-1β(μg/L):28.74±3.53比68.00±5.64,MDA(μmol/g):20.33±2.74比42.58±3.45,MMP9(ng/L):128.55±6.35比325.13±6.65,SOD(kU/g):50.53±4.19比22.53±3.27,miR-21(2-ΔΔCtObjective To investigate whether signal transducer and activator of transcription(STAT1/3/5)have a protective effect on hyperoxia-induced acute lung injury(HALI)and its mechanism.Methods Seventy C57BL/6J mice were randomly divided into five groups:normoxia control group,HALI group,and STAT1/3/5 inhibitor groups,with 14 mice in each group.The HALI model was established by exposure to more than 90%hyperoxia for 48 hours;three STAT inhibitor groups were pretreated by intraperitoneal injection of STAT1 inhibitor 40 mg/kg and STAT3 inhibitor 5 mg/kg,and STAT5 inhibitor 10 mg/kg for 1 week.Six blood samples were randomly collected from each group,and microRNA-21(miR-21)expression was measured by quantitative real-time reverse transcription-polymerase chain reaction(RT-PCR).Lung tissue of the sacrificed mice was obtained,and enzyme linked immunosorbent assay(ELISA)was used to detect the contents of tumor necrosis factor-α(TNF-α),interleukins(IL-6,IL-1β),superoxide dismutase(SOD),malonic dialdehyde(MDA),and matrix metalloproteinase 9(MMP9).The water content of lung tissue was calculated.The pathological changes in lung tissue were observed under the light microscope,and the pathological score of lung injury was performed.Western blotting was used to detect the expression of phosphorylated STAT(p-STAT1,p-STAT3,p-STAT5)in lung tissue.The 7-day cumulative survival rates of the remaining 8 mice in each group were analyzed using Kaplan-Meier survival curves.Results Under the light microscope,the alveolar structures in the HALI group and the STAT1 inhibitor group were destroyed,a large number of neutrophils(NEU)infiltrated in the alveoli and lung interstitium,which were thickened.The pathological score of lung injury and the water content of the lung tissue was significantly increased.In STAT3 inhibitor and STAT5 inhibitor groups,the alveolar cavity was clear,the degree of NEU infiltration and the thickness of lung interstitium were lower than those in HALI group,the pathological score of lung injury and the water content o

关 键 词：信号转导和转录激活蛋白1/3/5 高氧性急性肺损伤 急性呼吸窘迫综合征 微小RNA 凋亡 

分 类 号：R563[医药卫生—呼吸系统]