表面活性蛋白A在星形胶质细胞和小胶质细胞中的表达及炎症调节作用  

Surfactant protein A expression in astrocytes and microglia and its regulatory effects on inflammation

作　　者：赵正[1] 王晓凤[1] 李茂新 杨雪[1] ZHAO Zheng;WANG Xiaofeng;LI Maoxin;YANG Xue(Department of Emergency,Shengjing Hospital of China Medical University,Shenyang 110004,China)

机构地区：[1]中国医科大学附属盛京医院急诊科,沈阳110004

出　　处：《中国医科大学学报》2021年第5期418-424,共7页Journal of China Medical University

基　　金：辽宁省博士科研启动基金(2019-BS-278);中国医科大学青年骨干支持计划(QGZ2018056)。

摘　　要：目的探讨表面活性蛋白A(SPA)在星形胶质细胞和小胶质细胞中的表达及其炎症调节作用。方法分别采用免疫荧光双染和免疫组织化学染色法检测SPA在健康大鼠脑组织的星形胶质细胞、体外培养的星形胶质细胞和小胶质细胞中的表达情况。采用不同浓度(1、5、10μg/mL)脂多糖(LPS)培养液培养人星形胶质细胞及小胶质细胞24 h,10μg/mL LPS培养液培养人星形胶质细胞及小胶质细胞2、4、8、16、24 h,采用Western blotting检测细胞中SPA的表达情况。将人星形胶质细胞及小胶质细胞随机各分为LPS组(含有5μg/mL LPS的培养液)、LPS+SPA组(含有5μg/mL LPS和0.5μg/mL SPA的培养液)、SPA组(含有0.5μg/mL SPA的培养液)和对照组(未加入任何处理因素的培养基)。各组细胞培养8 h,采用Western blotting检测各组细胞中TLR4和NF-κB p65蛋白表达;ELISA法检测各组培养液中肿瘤坏死因子-α(TNF-α)和白细胞介素-1β(IL-1β)水平。结果SPA在大鼠脑组织的星形胶质细胞中表达,在体外培养的人星形胶质细胞和小胶质细胞中均有表达。LPS诱导人星形胶质细胞和小胶质细胞中SPA表达水平升高,且与LPS浓度呈剂量相关(P<0.05),与LPS处理时间不相关(P>0.05)。与LPS组比较,LPS+SPA组人星形胶质细胞和小胶质细胞中Toll样受体4(TLR4)和核转录因子κB(NF-κB)p65相对表达量显著下降(均P<0.05),培养液中TNF-α和IL-1β水平亦均显著下降(均P<0.05)。结论SPA在星形胶质细胞和小胶质细胞中表达,并可能通过抑制TLR4/NFκ-B信号通路来抑制TNF-α和IL-1β炎性细胞因子的产生,进而参与中枢神经系统免疫炎症反应的调控。Objective To characterize surfactant protein A(SPA)expression in astrocytes and microglia,and explore its regulatory effects on inflammation.Methods Immunofluorescence double staining and immunohistochemical staining were used to detect SPA protein expression in healthy rat brain tissues,and in astrocytes and microglia in vitro.Human astrocytes and microglia were each exposed to a range of lipopolysaccharide(LPS)concentrations(0,1,5,and 10μg/mL)for 24 h,or 10μg/mL LPS for different durations(2,4,8,16,and 24 h).Human astrocytes and microglial cells were each randomly divided into four groups:an LPS group(incubated with 5μg/mL LPS for 8 h),an LPS+SPA group(incubated with 5μg/mL LPS and 0.5μg/mL SPA for 8 h),an SPA group(incubated with 0.5μg/mL SPA for 8 h),and a control group(incubated with normal culture medium).Western blotting and ELISA were used to detect expression of TLR4 and NF-κB p65 proteins,and levels of TNF-αand IL-1βin the culture medium.Results SPA was expressed in astrocytes in rat brain tissue,and in human astrocyte and microglia cultures in vitro.LPS induced a significant increase in SPA expression in two glial cell types(P<0.05).LPS’effect on SPA expression was dose-dependent(P<0.05),but not clearly related to LPS exposure time(P>0.05).Relative expression of TLR4 and NF-κB p65 protein in the LPS+SPA treated human astrocytes and microglia was significantly lower than that in the LPS group(P<0.05).The levels of TNF-αand IL-1βin the culture medium of the LPS+SPA group were significantly lower than those in the LPS group(both P<0.05).Conclusion SPA is expressed in astrocytes and microglia,and participates in immune inflammatory regulation of the central nervous system,possibly by regulating TNF-αand IL-1βinflammatory factor secretion by inhibiting the TLR4/NF-κB signaling pathway.

关 键 词：表面活性蛋白A 星型胶质细胞 小胶质细胞 炎症调节 

分 类 号：R392.9[医药卫生—免疫学]