IL-27通过信号通路p38 MAPK和PI3K途径调节中性粒细胞Mac-1、fMLP-R及IL-1β表达

IL-27 Regulates the Expression of Mac-1,fMLP-R and IL-1β in Human Neutrophils through p38 MAPK and PI3K Signal Pathways

本研究探讨IL-27调节人外周血中性粒细胞中黏附分子Mac-1、趋化因子受体fMLP-R和促炎因子IL-1β表达的信号通路。采用Ficoll密度梯度离心及低渗裂解残存红细胞方法分离高纯度的外周血中性粒细胞。用RT-PCR方法检测中性粒细胞IL-27受体(WSX-1/TCCR和gp130)mRNA表达。分别应用IL-27及信号通路p38 MAPK、PI3K和ERK抑制剂SB203580、LY294002、U0126处理中性粒细胞,用实时定量RT-PCR检测作用前后fMLP-R和IL-1βmRNA水平变化;应用放射免疫法检测作用前后培养上清液中IL-1β的水平变化;流式细胞术检测作用前后Mac-1表达水平变化。结果表明:人外周血中性粒细胞协同表达IL-27受体的两个亚单位WSX-1/TCCR和gp130,IL-27下调中性粒细胞表面黏附分子Mac-1表达(p<0.05),SB203580阻断了IL-27对Mac-1的下调作用(p<0.05),而LY294002及U0126无阻断作用(p>0.05)。相反,IL-27上调中性粒细胞中fMLP-R和IL-1βmRNA的表达并能增加IL-1β释放(p<0.05),LY294002阻断了IL-27对fMLP-R和IL-1β的上调作用(p<0.05),而SB203580及U0126无阻断作用(p>0.05)。结论:IL-27与IL-27R相互作用,可能通过p38 MAPK和PI3K信号途径调节人外周血中性粒细胞中Mac-1、fMLP-R及IL-1β表达。

The present study was aimed to investigate the pathways, by which IL-27 regulates the expression of adherent molecule Mac-l, chemotactic factor receptor fMLP-R and pro-inflammatory cytokine IL-1β in human neutrophils. Highly purified human neutrophils were isolated from peripheral blood using Ficoll-Hypaque gradients centrifugation and erythrocyte lysis. The mRNA expression of IL-27 receptor components （WSX-1/TCCR and gp130 ） in human neutrophils was detected by reverse transcription polymerase chain reaction （RT-PCR）. After incubation with IL-27 and specific inhibitors （p38 MAPK inhibitor SB203580, PI3K inhibitor LY294002 and ERK inhibitor U0126 ）, the mRNA levels of fMLP-R and IL-1β were determined by real time RT-PCR, and the adherent molecule Mac-1 expression in human neutrophils was determined by flow cytometry. The IL-1β level in culture supematant of human neutrophils was assayed by radioimmunoassay. The results showed that IL-27 receptor components （WSX-1/TCCR and gpl30） were constitutively expressed in human neutrophils. IL-27 down-regulated Mac-1 expression in human neutrophils （p 〈 0.05）. After incubation with specific inhibitors, SB203580, not LY294002 and U0126, inhibited the down-regulation of Mac-1 expression by IL-27. However, 1L-27 up-regulated the mRNA expression of fMLP-R and IL-1 β, and increased the release of IL-1β （p 〈 0.05 ）. Interestingly, LY294002, not SB203580 and U0126, inhibited the up- regulation of fMLP-R and IL-1β by IL-27. It is concluded that the IL-27 may regulate the expression of Mac-1, fMLP-R and IL-1β in human neutrophils through p38 MAPK and PI3K signal pathways.