干扰S100A4在屋尘螨诱导的哮喘气道上皮屏障功能的作用

Role of Interfering S100A4 in HDM Induced Airway Epithelial Barrier Dysfunction

目的:探讨干扰16HBE细胞中S100A4基因表达后,HDM(屋尘螨)诱导的气道上皮屏障功能和信号蛋白的变化。方法:合成靶向S100A4基因的siRNA,采用脂质体转染正常16HBE细胞,PCR和Western blot检测干扰效果,并使用HDM刺激正常和干扰后的16HBE细胞,检测跨细胞电阻、FITC通透性,以及屏障蛋白(E-cadherin、β-catenin)、下游信号分子VEGFR2/p-VEGFR2表达的变化。结果:siRNA干扰后成功抑制16HBE细胞的S100A4基因和蛋白表达。HDM引起正常16HBE细胞TEER减少和FITC通透性增加,干扰S100A4可抑制HDM对跨细胞电阻和FITC通透性的损害。HDM刺激导致E-cadherin、β-catenin表达下调,干扰S100A4可减轻HDM对16HBE细胞屏障蛋白的破坏。HDM刺激后引起培养液中S100A4释放增加,细胞p-VEGFR2表达显著增加,而siRNA干扰后的16HBE细胞p-VEGFR2的表达仅轻度增加。结论:S100A4在HDM诱导的气道上皮细胞屏障损害中具有重要作用,可为哮喘的防治提供重要科学依据。

Objective:To investigate the changes of airway epithelial barrier function and signaling proteins in asthma induced by HDM after RNAi with S100A4 gene expression in 16HBE cells.Method:16HBE were transfected by S100A4 siRNA,the interference effect was detected by PCR and Western blot.16HBE were stimulated by HDM.The changes of TEER,FITC permeability,barrier proteins and VEGFR2/p-VEGFR2 were detected.Result:S100A4 gene and protein in 16HBE cells was inhibited successfully by siRNA interference.HDM stimulation resulted in decrease of TEER and increase of FITC permeability in normal 16HBE.S100A4 RNAi ameliorated the impairment of HDM on TEER and FITC permeability.The expression of E-cadherin and β-catenin was downregulated by HDM stimulation,while interference with S100A4 could alleviate the damage of barrier proteins induced by HDM.HDM promoted the release of S100A4 in culture medium and profoundly elevated the expression of p-VEGFR2 in normal cells,while the expression of p-VEGFR2 in RNAi 16HBE slightly increased.Conclusion:S100A4 plays an important role in HDM-induced airway epithelial barrier dysfunction,and these findings can provide scientific basis for the prevention and treatment of asthma.