摘要
目的观察TLR4/NF-κB信号通路在黄芪多糖抑制慢性阻塞性肺疾病(COPD)大鼠炎性反应中的作用,以及炎性因子与平均血小板体积(MPV)的关系。方法2019年12月—2020年2月于河北北方学院附属第一医院进行实验,60只SPF级雄性SD大鼠随机分为6组,空白对照组,模型组,黄芪多糖400、600、800 mg/kg组和泼尼松(6 mg/kg)组,各10只。除空白对照组外,其余各组给予大鼠气管灌注脂多糖(LPS)与吸烟建立COPD模型。黄芪多糖组与泼尼松组于灌注LPS或吸烟前灌胃给药。第29天后全自动血液分析仪测定大鼠血浆MPV水平;取大鼠肺组织,ELISA法测定IL-1β、IL-8、TNF-α水平,Western-blot测定TLR4、NF-κB蛋白水平,实时荧光定量PCR测定IL-1β、IL-8、TNF-αmRNA表达水平;股骨取骨髓巨核细胞瑞氏染色,观察细胞形态并计算全片巨核细胞数和产板巨核细胞数。人巨核细胞株Meg-01细胞分为6组:对照组、黄芪多糖组、IL-1β组、IL-8组、TNF-α组、TPO组,各组在相应条件下孵育24 h后,流式细胞仪测定各组细胞CD41、GPIba的表达。结果与空白对照组比较,模型组IL-1β、IL-8、TNF-α、TLR4、NF-κB蛋白水平显著提高,IL-1β、IL-8、TNF-αmRNA水平表达显著提高,产板巨核细胞数量、MPV显著降低(P均<0.05)。与模型组比较,黄芪多糖各组和泼尼松组IL-1β、IL-8、TNF-α、TLR4、NF-κB蛋白水平显著降低,IL-1β、IL-8、TNF-αmRNA水平表达显著降低,产板巨核细胞数显著增加,MPV显著升高(P均<0.05)。当黄芪多糖为800 mg/kg时各指标改善程度与泼尼松组无显著差异(P>0.05)。与对照组比较,IL-1β组、IL-8组、TNF-α组Mge-01细胞CD41、GPIba的表达显著下调,黄芪多糖组CD41、GPIba的表达无明显差异(P>0.05)。结论黄芪多糖通过抑制TLR4/NF-κB通路减轻COPD大鼠炎性反应并使MPV升高,COPD随炎性反应强度的增加巨核细胞成熟与分化能力降低,导致MPV下降,检测MPV水平可有效评估COPD炎性反应的严
Objective To explore the role of TLR4/NF-κB signaling pathway in astragalus polysaccharide(APS)inhibition of inflammatory response in COPD rats,and the relationship between inflammatory factors with mean platelet volume.Methods 60 SPF male SD rats were randomly divided into 6 groups,namely,normal control group,model group,APS 400,600,800 mg/kg group and prednisone(6 mg/kg)group.In addition to the blank control group,other groups were given LPS and cigarette smoke to establish COPD model.APS groups and prednisone group were administrated before given LPS or smoke.At the 29th day,plasma MPV of rats was analyzed by automatic blood analyzer;lung tissues of rats were taken,and the expression levels of IL-1β、IL-8、TNF-αwere determined by ELISA;TLR4 and NF-κB protein were determined by Western blot;the mRNA expression levels of IL-1β、IL-8、TNF-αwere determined by real-time quantitative PCR.In addition,bone marrow megakaryocytes were collected from femur,Wright's staining was performed to observe the cell morphology and calculate the number of megakaryocyte and the number of thromocytogenic megakaryocyte.Human megakaryocyte Meg-01 cells were divided into 6 groups:control group,APS group,IL-1βgroup,IL-8 group,TNF-αgroup,TPO group.After being incubated for 24h in each culture media,the expressions of CD41 and GPIba were measured by flow cytometry.Results Compared with blank control group,the protein levels of IL-1β,IL-8,TNF-α,TLR4,NF-κB in the model group were significantly increased,and the mRNA levels of IL-1β,IL-8,TNF-αwere significantly increased.The number of bone marrow thromocytogenic megakaryocytes and MPV decreased significantly.Compared with model group,the protein levels of IL-1β,IL-8,TNF-α,TLR4,NF-κB in APS groups and prednisone group were significantly decreased,and the mRNA levels of IL-1β,IL-8,TNF-αwere significantly decreased.The number of bone marrow thromocytogenic megakaryocytes and MPV significantly increased.When the level of APS went up to 800 mg/kg,there was no significant dif
作者
曹亮
刘建华
冯平
赵建清
Cao Liang;Liu Jianhua;Feng Ping;Zhao Jianqing(Department of Respiratory,the First Affiliated Hospital to Hebei North College,Hebei Province,Zhangjiakou 075061,China)
出处
《疑难病杂志》
CAS
2020年第7期723-729,共7页
Chinese Journal of Difficult and Complicated Cases
基金
河北省医学科学研究重点课题计划项目(20160035)。
