甲酰肽受体在缺氧时对血管平滑肌细胞IL-6和IL-8产生的作用

The role of Formyl peptide receptor in the production of IL-6 and IL-8 from vascular smooth muscle cells during hypoxia

目的探讨甲酰肽受体(formyl peptide receptor,FPR)在缺氧时对血管平滑肌细胞(vascular smooth muscle cells,VSMCs)炎症因子产生的作用。方法按照缺氧时间将大鼠血管平滑肌细胞随机分为0 h、12 h、24 h组,将各组细胞放入缺氧培养箱(1%O2、5%CO2、99%N2)培养相应时间后,用Western blot检测平滑肌细胞中IL-6、IL-8的蛋白水平,用RT-PCR检测其m RNA水平。同时使用FPR拮抗剂t BOC预处理细胞并缺氧不同时间(0 h、12 h、24 h、)后,检测IL-6、IL-8的表达水平。另外,为了进一步明确FPR与炎症因子表达的关系,用FPR激动剂f MLP直接处理细胞之后,检测IL-6、IL-8的表达水平。然后使用细胞在缺氧条件下培养后的上清液作用于未缺氧的细胞,检测IL-6、IL-8的表达水平变化,并通过观察FPR拮抗剂处理后的变化,明确FPR在其中的作用。用f MLP刺激p38抑制剂预处理过的细胞,检测IL-6、IL-8的表达水平变化,以观察FPR激活后引起炎症因子表达对p38途径的依赖性。结果缺氧后血管平滑肌细胞的IL-6、IL-8 m RNA和蛋白含量均显著高于0 h组;当t BOC处理细胞后显著削弱了这一效应(P<0.05)。而且FPR激动剂能直接显著提高未缺氧的细胞的IL-6、IL-8的表达水平(P<0.05)。使用缺氧细胞的上清作用于常氧细胞后IL-6、IL-8的表达水平显著高于常氧细胞上清的作用效果;FPR拮抗剂处理显著削弱了这一效应(P<0.05)。用p38抑制剂预处理过的细胞加入f MLP与单纯加入f MLP的细胞相比,血管平滑肌细胞IL-6、IL-8表达水平显著低于单纯加入f MLP组(P<0.05)。结论缺氧引起血管平滑肌细胞IL-6、IL-8的表达,FPR在这一过程中发挥了重要的介导作用,而FPR是被缺氧细胞释放出来的激动剂激活。激活后的FPR引起IL-6、IL-8的表达依赖于p38信号途径。

This study was performed to investigate the role of Formyl peptide receptor （FPR） in the production of inflammatory cytokines from vascular smooth muscle cells （VSMCs） during hypoxia. The mRNA levels of IL-6 and IL-8 in VSMCs of hypoxia rats were examined with RT-PCR, while the protein levels of IL-6 and IL-8 in VSMCs of hypoxia rats were detected by Western blotting. The mRNA and protein levels of IL-6 and IL-8 in vascular smooth muscle cells of hypoxia group were significantly higher than those in control group, and this effect was significantly attenuated by tBOC treatment （P〈O.05）. Moreover, FPR agonist directly and significantly upregulated the expression of IL-6 and IL-8 in non-hypoxic cells （P〈0.05）. The expression levels of IL-6 and IL-8 in the cells stimulated by hypoxic supernatant was significantly higher than that in the cells stimulated by normoxic supematant, while FPR antagonist treatment could significantly reduced the difference （P〈0.05） The fMLP-induced IL-6 and IL-8 expression in VSMCs were significantly down-regulated by p38 inhibitor （P〈0.05）. In conclusion, hypoxia induces the expression of IL-6 and IL-8 in vascular smooth muscle cells. FPR plays an important mediating role in this process. FPRs can be activated by some agonists released by hypoxic cells. And the upregulation of IL-6 and IL-8 expression by activated FPRs is dependent on the p38 signaling pathway.