p38丝裂原激活蛋白激酶特异性抑制剂SB203580减轻小鼠气道炎症及黏液高分泌

Airway Inflammation and Mucus Hypersecretion in Mice are Ameliorated by SB203580,A p38 Mitogen-activated Protein Kinase Specific Inhibitor

目的探讨小鼠吸入丙烯醛后是否出现气道黏液高分泌及其产生机制,观察p38丝裂原激活蛋白激酶(p38MAPK)信号通路在气道炎症损伤、黏液高分泌中的作用,以及应用p38MAPK特异性抑制剂SB203580干预后对气道炎症、MUC5AC的影响。方法64只小鼠随机分为四组丙烯醛模型组、丙烯醛+SB203580治疗组、生理盐水对照组和SB203580对照组,每组16只小鼠。于第7和21d各组随机处死8只,采集支气管肺泡灌洗液(BALF)和肺组织标本。BALF作细胞总数及分类计数,石蜡包埋肺组织切片HE染色观察形态学,AB-PAS染色了解气道杯状细胞和黏液物质,免疫组织化学及RT-PCR方法进行黏蛋白基因半定量分析,Western-Blot定量分析信号通路蛋白。结果小鼠吸入丙烯醛21d后出现明显的气道炎症和黏液高分泌,经SB203580治疗后,炎性细胞浸润减少,杯状细胞增生改善,黏液分泌明显减轻。丙烯醛模型组、丙烯醛+SB203580治疗组的AB-PAS阳性着色面积比较差异有统计学意义[(15.54±0.93)%比(9.82±0.56)%,P<0.01];丙烯醛模型组与丙烯醛+SB203580治疗组之间MUC5AC阳染积分光密度比较差异有统计学意义(0.153±0.017比0.066±0.007,P<0.01)。拮抗p38MAPK后,MUC5ACmRNA表达量下降(7d0.934±0.061,21d1.041±0.026),与模型组相应时间点比较差异有统计学意义(P<0.01)。治疗组磷酸化p38MAPK表达与模型组比较明显下调(0.844±0.014比1.004±0.039,P<0.01)。结论小鼠吸入丙烯醛模型中,p38MAPK信号通路调控MUC5AC基因转录,SB203580可减轻气道炎症和气道黏液高分泌。

Objective： To identify the role of p38 mitogen- activated protein kinase （p38 MAPK） and its specific inhibitor, SB203580, in airway inflammation and mucus hypersecretion in vivo. Methods： Sixty- four mice were randondy divided into four groups （n = 16 in each group）, ie.NS control, aerolein - only, acrolein - plus SB203580, and SB203580 - only groups.On days 7 and 21, eight mice of each group were killed so as to collect lung tissue sample and bronchoalveolar lavage fluid （BALF） .Total and differential cell counts in BALF were measured, and lung tissues were embedded in paraffin for histological analysis by HE staining. Airway goblet cells and mucus production were observed by AB - PAS slaing method. Meanwhile, the semi - quantitative analyses on mucin gene and signaling pathway protein were implemented by means of IHC, RT - PCR, and western- blot teclmiques respectively.Results： Both inflammatory cells and mucus- secreting goblet cells were strikingly increased in exposed mice treated with acrolein for 21 days, which could be apparently attenuated after administration of SB203580. In acmlein group and acrolein ＋ SB203580 group, AB/PAS - positive staining areas were. eaculated as （ 15.54±0.93 ） % and （9.82±0.56） % respectively, with a statistically significant difference （ P 〈 0.01 ） . MUC5AC protein and mRNA expression were decreased （ P 〈 0.01） by blocking the action of p38 MAPK signaling pathway.Furthermore,, Western blot analysis showed that phospho- p38 MAPK was dramatically activated in acrolein group compared with that in NS control group; there was a markedly inhibition of p38 MAPK phosphorylation via SB203580. Conclusions： p38 MAPK signaling pathway contributes to regtdating MUC5AC mucin gene trauscripafion in this murine model exposed to acrolein, in which SB203580 can ameliorate airway mucus overproduction and inflammation as well.