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目的:探讨新型糖尿病药物exendin-4(Ex-4)影响小鼠巨噬细胞一氧化氮(NO)生成和IL-10/IL-12分泌的量效-时效关系,以及是否经由PI3K/Akt/mTOR信号转导通路产生该作用。方法:体外培养小鼠RAW264.7巨噬细胞株,在有或者无细菌脂多糖(LPS)诱导下,给予不同浓度的Ex-4(1、10、50 nmol/L)刺激,并在培养4、8和24 h收集上清,分别采用Griess和ELISA方法检测上清NO和IL-10/IL-12的分泌水平。此外,分别给予mTOR抑制剂雷帕霉素或PI3K抑制剂LY294002预处理巨噬细胞1 h,再给予低、中浓度Ex-4(1、10 nmol/L)刺激,观察巨噬细胞NO的产生和IL-10/IL-12的分泌是否发生改变。结果:无论是否存在LPS,Ex-4均能显著促进小鼠RAW264.7细胞分泌IL-10,同时抑制IL-12和NO的分泌,并且星明显的时效-量效关系,中浓度Ex-4已达最大效应(与低浓度比较,P<0.05;与高浓度比较,P>0.05),该作用能被LY294002和雷帕霉素逆转(P<0.05或P<0.01),尤以雷帕霉素最为显著。结论:Ex-4可通过PI3K/Akt/mTOR信号通路调控单核细胞NO的分泌,并影响IL-10/IL-12的平衡,可能是其参与免疫调节的机制之一。
Abstract：Objective:To investigate the dose-and time-dependent effects of exendin-4 on the production of nitric oxide(NO) and IL-10/IL-12 balance in macrophage,and to ascertain whether the phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/Akt/mTOR) pathway is involved.Methods:Murine macrophage RAW264.7 cell line was treated with different doses of exendin-4(1,10,50 nmol/L) for 4,8 or 24 hours with or without lipopolysaccharide(LPS).In another set of experiment,RAW264.7 macrophage was cultured with low(1 nmol/L) or medium(10 nmol/L) dose of Ex-4 after 1-hour pretreatment with PI3K inhibitor(LY294002) or prototypic mTOR inhibitor(Rapamycin) for 4 hours.Cell-free supernatants were harvested for NO and IL-10/IL-12 assessment with Griess reagent and ELISA,respectively.Results;Compared with control group,exendin-4 could significantly increase the production of IL-10 while inhibit the secretion of NO and IL-12with or without LPS in a dose-and time-dependent manner,and a medium dose of Ex-4 exerted the most significant effect(P<0.05 compared to low dose of Ex-4 but P>0.05 compared to high-dose of Ex-4).LY294002 and Rapamycin,the two PI3K/Akt/mTOR inhibitors,had the potential to reverse the effect of exendin-4(P<0.05 or P<0.01),and between them,rapamycin was more effective.Conclusions:Exendin-4 might regulate,at least in part,the secretion of NO and IL-10/IL-12 balance in macrophage via PI3K/Akt/mTOR pathway,contributing to immune regulation.
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基本信息:
中图分类号:R459.7
引用信息:
[1]高玉雷,吕艺,柴艳芬,等.Exendin-4对小鼠巨噬细胞一氧化氮生成和促炎/抗炎反应平衡的影响[J].感染、炎症、修复,2013,14(04):201-206.
基金信息:
国家自然科学基金项且(30973120,81272089,81372054);; 国家重点基础研究发展计划项目(2012CB518102);; 军队“十二五”医药卫生科研基金重点项目(BWS12J050)
2013-12-20
2013-12-20