晚期糖基化终末产物受体促进甲苯二异氰酸脂哮喘小鼠MUC5AC表达及气道黏液高分泌

目的 探讨晚期糖基化终末产物受体(RAGE)信号对甲苯二异氰酸脂(TDI)哮喘小鼠黏蛋白MUC5AC表达及气道黏液分泌的影响.方法 在体内水平上建立TDI小鼠哮喘模型,实验分4组:对照组、TDI溶剂(AOO)致敏激发组、TDI致敏激发组、RAGE抑制剂处理组.采用糖原染色评估各组间气道黏液分泌情况,Western blotting及免疫组化法检测MUC5AC表达水平.同时通过Western blotting检测各组间细胞外调节蛋白激酶(ERK)通路磷酸化水平.在体外水平配制TDI-人血清白蛋白(TDI-HSA)复合物,刺激人支气管上皮细胞(16HBE)及经慢病毒转染空载体和shRNA-RAGE载体的人支气管上皮细胞,检测各组MUC5AC及ERK通路分子表达情况,加入ERK抑制剂U0126后,进一步评估MUC5AC mRNA表达情况.结果 与对照组相比,TDI哮喘组小鼠糖原染色阳性率及MUC5AC表达升高(P<0.05),p-ERK表达增多(P<0.05),RAGE抑制剂处理组糖原染色阳性率及MUC5AC表达较TDI组减少(P<0.05).同时,p-ERK表达下降(P<0.05).体外水平,与对照组相比,TDI-HSA处理组MUC5AC mRNA及p-ERK表达增多(P<0.05),sh RNA-RAGE组MUC5AC mRNA及p-ERK表达下调(P<0.05),ERK抑制剂预处理组,MUC5AC mRNA表达下调(P<0.05).结论 TDI小鼠哮喘模型下RAGE可能通过激活ERK通路促进黏蛋白MUC5AC的表达及黏液高分泌的发生.

Receptor for advanced glycation end products upregulates MUC5AC expression and promotes mucus overproduction in mice with toluene diisocyanate-induced asthma

Objective To explore the role of the receptor for advanced glycation end products (RAGE) in regulating the expression of MUC5AC and mucus production in a mouse model of toluene diisocyanate (TDI)-induced asthma. Methods BALB/c mice were randomly divided into control group, vehicle (AOO) group, TDI-induced asthma group and RAGE inhibitor (FPS-ZM1) group. PAS staining, Western blotting, and immunohistochemistry were used to analyze the changes in mucus production and MUC5AC expression in the airway of the mice, and the expression of p-ERK was detected with Western blotting. In vitro cultured human bronchial epithelial cell line 16HBE was transfected with lentiviral vector carrying short hairpin RNA targeting RAGE (shRNA-RAGE) and subsequently challenged with a TDI-human serum albumin (TDI-HSA)conjugate,and the changes in cellular MUC5AC mRNA expression as detected using RT-PCR;the protein expressions of ERK and p-ERK in the cells were examined with Western blotting. The effect of ERK inhibitor U0126 pretreatment on MUC5AC mRNA expression was also analyzed in the cells.Results Compared with the control mice,TDI-induced asthmatic mice showed significantly higher rates of PAS positivity and increased MUC5AC and p-ERK expressions in the airway (P<0.05). Treatment with FPS-ZM1 significantly decreased PAS positivity and lowered MUC5AC and p-ERK expressions in the airway of the asthmatic mice(P<0.05).Exposure of 16HBE cells to TDI-HSA caused a significant increase in MUC5AC mRNA expression and p-ERK protein expression (P<0.05), while RAGE knockdown obviously suppressed TDI-HSA-induced upregulation of p-ERK and MUC5AC mRNA(P<0.05). Treatment with the ERK inhibitor U0126 also lowered TDI-HSA-induced up-regulation of MUC5AC mRNA in the cells (P<0.05). Conclusion RAGE signaling induces MUC5AC expression via extracellular signal-regulated kinase pathway to promote mucus overproduction in mice with TDI-induced asthma.