背景:TRP蛋白对外部温度变化敏感,但其在上气道黏膜中的致病机制尚不清楚。
目的:探讨TRPV1和TRPA1调节鼻腔上皮细胞分泌炎症因子的作用机制。
方法:采用免疫荧光法检测鼻黏膜上皮细胞中TRPV1和TRPA1的表达。用TRPV1和TRPA1激动剂和拮抗剂刺激上皮细胞,并检测上皮细胞Ca2+释放和炎症因子分泌的变化。评价了用钙螯合剂EGTA刺激的TSLP分泌。免疫荧光染色观察转录因子NFAT。
结果:在鼻上皮细胞中检测到TRPV1和TRPA1的表达,和Ca2+内流在用激动剂刺激后增加。TRPV1和TRPA1激活后,TSLP基因表达,IL-25和IL-33以及TSLP和IL-33的蛋白表达水平升高,只有TSLP可以被拮抗剂和siRNA抑制。EGTA给药后,TSLP的分泌受到显著抑制,在上皮细胞中TRPV1和TRPA1蛋白激活后,观察到转录因子NFAT在细胞核中的表达。
结论:鼻腔上皮细胞TRPV1和TRPA1的激活通过Ca2+/NFAT途径刺激TSLP的生成。它还诱导IL-25和IL-33基因表达水平的上调和IL-33蛋白水平的升高,导致气道炎症的发展。
BACKGROUND: TRP protein is sensitive to external temperature changes, but its pathogenic mechanism in the upper airway mucosa is still unclear.
OBJECTIVE: To investigate the mechanism of TRPV1and TRPA1 in regulating the secretion of inflammatory factors in nasal epithelial cells.
METHODS: The expression of TRPV1 and TRPA1 in nasal mucosal epithelial cells was investigated using immunofluorescence assays. Epithelial cells were stimulated with TRPV1 and TRPA1 agonists and antagonists, and changes in Ca2+ release and inflammatory factor secretion in epithelial cells were detected. TSLP secretion stimulated with the calcium chelating agent EGTA was evaluated. The transcription factor NFAT was observed by immunofluorescence staining.
RESULTS: TRPV1 and TRPA1 expression was detected in nasal epithelial cells, and Ca2+ influx was increased after stimulation with agonists. After the activation of TRPV1 and TRPA1, the gene expression of TSLP, IL-25, and IL-33 and the protein expression levels of TSLP and IL-33 were increased, and only TSLP could be inhibited by antagonists and siRNAs. After administration of EGTA, the secretion of TSLP was inhibited significantly, and the expression of the transcription factor NFAT in the nucleus was observed after activation of the TRPV1 and TRPA1 proteins in epithelial cells.
CONCLUSIONS: Activation of TRPV1 and TRPA1 on nasal epithelial cells stimulates the generation of TSLP through the Ca2+/NFAT pathway. It also induces upregulation of IL-25 and IL-33 gene expression levels and increased levels of IL-33 protein, leading to the development of airway inflammation.