组蛋白H4对小鼠急性呼吸窘迫综合征肺泡巨噬细胞极化的影响  

Effect of histone H4 on polarization of alveolar macrophages in mice with acute respiratory distress syndrome

作　　者：张雁林[1] 陈明[1] 赵怡然 郭利霞[1] 李晓 郑亦沐 关里[1] 李树强[1] ZHANG Yanlin;CHEN Ming;ZHAO Yiran;GUO Lixia;LI Xiao;ZHENG Yimu;GUAN Li;LI Shuqiang(Department of Occupational Medicine,Peking University Third Hospital,Beijing 100191,China)

机构地区：[1]北京大学第三医院职业病科,北京100191

出　　处：《中国职业医学》2023年第3期248-254,共7页China Occupational Medicine

基　　金：国家自然科学基金面上项目(81773374);北京市自然科学基金面上项目(7182179);北京大学第三医院学术带头人后备人才基金(Y7551308)。

摘　　要：目的探讨组蛋白H4在脂多糖诱导的小鼠急性呼吸窘迫综合征(ARDS)中对肺泡巨噬细胞(AM)极化的作用。方法(1)将无特定病原体级C57BL/6雄性小鼠随机分为对照组和2、4、6、8 mg/kg脂多糖组,每组6只。采用一次性气管滴注法,后4组小鼠予相应剂量脂多糖染毒,对照组小鼠予等体积的0.9%氯化钠溶液。12 h后,采集各组小鼠动脉血进行血气分析,取肺组织观察肺水肿和病理组织学改变情况,采用酶联免疫吸附实验检测小鼠支气管肺泡灌洗液(BALF)中组蛋白H4水平,并采用贴壁法分离5组小鼠AM。(2)采用贴壁法分离正常小鼠AM,随机分为对照组、组蛋白H4损伤组、BALF损伤组和抗组蛋白H4中和抗体(anti-H4)干预组。组蛋白H4损伤组AM中加入终质量浓度为20 mg/L的组蛋白H4;BALF损伤组和anti-H4干预组AM中均加入经剂量为6 mg/kg体质量脂多糖染毒小鼠的BALF上清200μL,后者同时加入终质量浓度为25 mg/L的anti-H4抗体;对照组AM中加入等体积的磷酸盐缓冲液溶液。刺激12 h后,收集AM。(3)采用实时定量聚合酶链式反应法检测AM中肿瘤坏死因子-α(Tnfa)、白细胞介素-1β(Il1b)、分化抗原206(Cd206)和精氨酸酶1(Arg1)的mRNA相对表达水平。结果(1)与对照组比较,4个脂多糖组小鼠均出现呼吸急促,肺组织出现炎症反应和肺水肿,均呈剂量依赖性加重。随着脂多糖染毒剂量的增加,小鼠动脉血氧分压与吸入气中氧浓度分数比值下降(P<0.05),肺湿/干质量比值、BALF中组蛋白H4水平和AM的Tnfa、Il1b mRNA相对表达水平均增加(P值均<0.05)。其中,6和8 mg/kg脂多糖组小鼠均出现ARDS。6和8 mg/kg脂多糖组小鼠BALF中组蛋白H4水平和AM的Tnfa、Il1b mRNA相对表达水平均高于其余3组(P值均<0.05)。(2)与对照组比较,组蛋白H4损伤组、BALF损伤组AM中Tnfa和Il1b mRNA相对表达水平均升高(P值均<0.05),Cd206和Arg1 mRNA相对表达水平均下降(P值均<0.05)。与BALF损伤组比较,antObjective To investigate the role of histone H4 in the polarization of alveolar macrophages(AM)in lipopolysaccharide(LPS)-induced acute respiratory distress syndrome(ARDS)in mice.Methods i)The specific pathogen free male C57BL/6 mice were randomly divided into control group and 2,4,6 and 8 mg/kg LPS groups,with six mice in each group.The mice in the LPS groups were intratracheally administered LPS according to their respective doses,while the mice in the control group received an equivalent volume of 0.9%saline.After 12 hours,the arterial blood gas was analyzed,and the pulmonary edema and histopathological changes in lung tissues of mice in each group were observed.The level of histone H4 in bronchoalveolar lavage fluid(BALF)of mice was detected using enzyme-linked immunosorbent assay,and mice AMs of the five group were isolated using adherent method.ii)AMs from normal mice were isolated using adherent method and randomly divided into control group,histone H4 injury group,BALF injury group and anti-histone H4 antibody(anti-H4)intervention group.In the histone H4 injury group,AMs were treated with histone H4 at a final concentration of 20 mg/L.In the BALF injury group and anti-H4 intervention group,AMs were treated with 200μL BALF supernatant from mice intratracheally administered 6 mg/kg body weight LPS,with the latter group treated with 25 mg/L anti-H4 antibody.The control group AMs were treated with phosphate-buffered saline.iii)After 12 hours of stimulation,the cells were collected,and the relative expression of tumor necrosis factor-α(Tnfa),interleukin-1β(Il1b),differentiation antigen 206(Cd206)and arginase 1(Arg1)in AMs was detected using real-time quantitative polymerase chain reaction.Results i)Compared with the control group,mice in all four LPS groups exhibited rapid breathing,inflammatory reaction and lung edema in lung tissues,which were aggravated in a dose-dependent manner.The ratio of partial pressure of arterial oxygen to fraction of inspired oxygen in mice decreased with the increase of LPS dos

关 键 词：急性呼吸窘迫综合征 组蛋白H4 肺泡巨噬细胞 脂多糖 肿瘤坏死因子-α 白细胞介素-1β 剂量-效应关系 小鼠 

分 类 号：R135.1[医药卫生—劳动卫生]