免疫抑制小鼠急性重症肺部感染中巨噬细胞亚型变化与乌司他丁调控效应  

Alterations of macrophage subsets and the regulation of Ulinastatin in the immunosuppressed mouse with severe pulmonary infection

作　　者：秦科[1] 邝晓聪[2] 孙煦勇[1] 董建辉[1] 蓝柳根[1] 黄莹[1] 曹嵩[1] 李美思[1] 李壮江[1] 李海滨[1] 苏庆东[1] 周洁惠[1] 

机构地区：[1]广西南宁中国人民解放军第303医院移植医学研究院,广州军区器官移植中心,广西移植医学重点实验室,广西移植医学工程技术研究中心,广西南宁530021 [2]广西医科大学基础医学院,广西南宁530021

出　　处：《实用器官移植电子杂志》2016年第1期46-51,共6页Practical Journal of Organ Transplantation(Electronic Version)

基　　金：桂科自2013GXNSFAA019253,桂科自2013GXNSFAA019244;广西科学研究与技术开发计划项目(桂科攻14124003-8);广西壮族自治区卫生与计生委员会课题(Z2015293)

摘　　要：目的探讨免疫抑制状态下重症肺部感染小鼠肺巨噬细胞亚型变化情况,观察乌司他丁(UTI)干预的巨噬细胞调控效应,初步明确其拮抗重症肺部炎症的作用机制。方法选取60只Balb/c小鼠,分为对照组、模型组和乌司他丁组,每组20只小鼠;其中模型组采用腹腔注射甲泼尼龙(30 mg/kg),气管内滴注内毒素(10 mg/kg)制备免疫低下急性重症肺部感染小鼠模型;乌司他丁组则在建模前后腹腔注射乌司他丁(1×10~5 U/kg)干预;主要观察小鼠呼吸情况、肺泡灌洗液炎性细胞构成、肺组织病理改变与评分以及M1和M2巨噬细胞等指标,采用免疫组化法检测肺组织M2巨噬细胞的CD163表达,采用实时定量PCR检测肺泡灌洗液细胞中M1巨噬细胞分泌分子肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)、诱导型一氧化氮合成酶(i NOS)和M2巨噬细胞标志与分泌分子CD206、Arg-1、IL-10的m RNA表达水平。结果模型组和乌司他丁组小鼠一般情况较差,部分小鼠出现呼吸窘迫现象,其中模型组小鼠肺泡腔与间质炎性细胞浸润,肺间质增宽,乌司他丁组病变减轻,与模型组相比,肺损伤评分显著降低〔(肺损伤评分(分):6.77±0.85比8.21±0.92,P<0.05〕,乌司他丁组与模型组肺泡灌洗液中中性粒细胞比例显著增高,但两组差异不大。除i NOS外,M1和M2巨噬细胞相关分子均高于对照组,与模型组相比,乌司他丁组M1巨噬细胞相关分子TNF-α、IL-6以及M2巨噬细胞相关分子IL-10、CD206均显著降低〔TNF(2^(-ΔΔCt)):0.42±0.18比0.76±0.12,IL-6(2^(-ΔΔCt)):0.72±0.26比1.22±0.32,IL-10(2^(-ΔΔCt)):3.91±1.35比6.84±2.13,CD206(2^(-ΔΔCt)):0.61±0.91比0.92±0.31,均P<0.05〕,而i NOS和Arg-1则与模型组无显著差别。此外,免疫组化染色检测乌司他丁组肺组织M2巨噬细胞的CD163表达,其结果明显低于模型组〔CD163(A)值:31 430.0±2 417.97比69 855.0±6 489.5,P<0.05〕,但高于对照组。结论在辅助T细胞Th1单向偏移Th2Objective To observe the alterations of macrophage M1 and M2 subsets and the regulation of Ulinastatin（UTI）and to explore the mechanisms of UTI anti-inflammatory in the immunocompromised mouse with severe pulmonary infection. Methods A total number of 60 Balb/c mice were randomly divided into control group, model group and UTI-group, with 20 mice in each group. The mice in model group were prepared with methylprednisolone by intraperitoneal injection and dripping LPS by trachea. The mice in UTI-group were treated by intervention with UTI（1×105 U/kg）, before or after LPS Intervention. At 6 hours after the intratracheal administration of LPS, the indexes were detected as fellow： the cell types and ratios of cells in bron choalveolar lavage fluid（BALF）, pathological changes expression of CD163 in the lung of mice were detected by hematoxylin-eosin（HE） staining and immunohistochemistry staining, respectively. The m RNA expression of M1-type markers（TNF-α, IL-6, i NOS） and M2-type markers（CD206, Arg-1, IL-10） were analyzed by real-time quantitative PCR in BALF. Results There was acute respiratory distress phenomenon in the model group. The pathological changes were wider alveolar walls and predominant polymorphonuclear neutrophils（PMN） and mononuclear inflammatory infiltrates. In UTI group, the pulmonary injury were alleviated and lung injury score were significantly lower than that of model group 〔lung injury score（score）：6.77±0.85 vs. 8.21±0.92,P 〈0.05〕. The ratios of PMN and macrophage were significantly increased in in UTI group and model group, but there were no significant difference between the two groups. Except i NOS, the markers of M1 and M2 macrophages were higher than those of control group. In UTI group, M1-type markers（TNF-α, IL-6） and M2-type markers（CD206, IL-10） were significantly lower than that in model group 〔TNF（2^（-ΔΔCt））：0.42±0.18 vs. 0.76±0.17,IL-6（2^（-ΔΔCt））：0.72±0.26 vs. 1.22±0.32,IL-10（2^（-ΔΔCt））�

关 键 词：免疫抑制 小鼠 重症肺部感染 巨噬细胞 UTI 

分 类 号：R617[医药卫生—外科学]