降钙素基因相关肽通过降低炎性小体活性抑制巨噬细胞功能

Calcitonin gene-related peptide inhibits the function of macrophage through inhibiting inflammasome activation

目的探讨降钙素基因相关肽(CGRP)抑制巨噬细胞炎性反应的机制。方法使用不同浓度CGRP处理LPS活化/未活化的小鼠巨噬细胞(RAW264.7),实时荧光定量PCR(qRT-PCR)检测巨噬细胞内促炎细胞因子白介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)和还原型辅酶Ⅱ氧化酶-活性氧簇-NOD样受体蛋白3(NLRP3)mRNA表达水平;酶联免疫吸附法(ELISA)定量检测细胞上清分泌型IL-1β和TNF-α蛋白表达量;蛋白免疫印迹法(Western blot)检测细胞内NLRP3蛋白表达水平。结果 CGRP显著降低LPS活化/未活化RAW264.7细胞内炎性因子IL-1β、TNF-α、NLRP3 m RNA水平与蛋白水平,同时,NLRP3下游调节分子Caspase-1的蛋白水平被显著下调。在mRNA水平,NLRP3随CGRP剂量变化而变化的趋势与IL-1β、TNF-α非常相似。结论CGRP抑制巨噬细胞炎性激活,其机制可能与CGRP抑制NLRP3表达与活性有关。

This study was designed to explore the calcitonin-gene-related peptide（CGRP） mediatedmechanism in repressing the secretion of inflammatory factors in mouse macrophage cells. Macrophages RAW264.7 were treated by different concentrations of CGRP, then the mRNA levels of interleukin-1β（IL-1β）, tumor necrosisfactor-α（TNF-α） and nicotinamide adenine dinucleotide phosphate oxidases-reactive oxygen species-NOD likereceptor protein 3（NLRP3） were detected by quantitative real-time PCR（qRT-PCR）, and the secretion levels ofIL-1β and TNF-α were analyzed by enzyme linked immune sorbent assay（ELISA）, while the protein level ofNLRP3 was determined by Western blotting. Data showed that CGRP dose-dependently reduced the m RNA levelsof IL-1β, TNF-α and NLRP3 in macrophages, and also reduced the protein levels of IL-1β, TNF-α and NLRP3 inthe cultural supernatants. CGRP attenuated LPS-induced expression of IL-1β and TNF-α in RAW264.7 cells.Taken together, CGRP can repress the secretion of inflammatory factors in mouse macrophage cells, and themechanism is related with the inhibition of NLRP3 expression and activity.