摘要
目的探讨白藜芦醇对脂多糖(LPS)诱导的小鼠巨噬细胞系RAW264.7细胞极化表型改变的影响。方法体外培养的RAW264.7小鼠巨噬细胞在六孔板孵育12h,分为磷酸盐缓冲液(PBS)对照组、LPS(100ng/mL)组、LPS(100ng/mL)+白藜芦醇(30μmol/L)组,LPS+白藜芦醇组在加入LPS前预先加入白藜芦醇孵育12h;上述细胞在LPS刺激12h后分别收集细胞和培养上清液;实时定量PCR(RT-qPCR)检测经典的炎症活化型巨噬细胞(M1)型相关基因iNOS、TNF-αmRNA表达,以及替代活化巨噬细胞(M2)型相关基因IL-10、PPARγ和Arg-1mRNA表达,Western blot检测细胞iNOS、Arg-1蛋白表达,ELISA检测培养上清液中炎性因子IL-12p40、IL-10和TNF-α的水平。结果 RT-qPCR检测结果显示,与LPS组相比,M1型相关基因iNOS、TNF-αmRNA较LPS+白藜芦醇组显著上升(P<0.05),而M2型相关基因IL-10、PPARγ、Arg-1mRNA表达显著下调(P<0.05)。Western blot检测发现,LPS组iNOS蛋白水平显著高于LPS+白藜芦醇组(P<0.05),而Arg-1蛋白水平显著低于LPS+白藜芦醇组(P<0.05)。ELISA检测发现,LPS组培养上清液中炎性因子IL-12p40、TNF-α水平显著高于LPS+白藜芦醇组(P<0.05),而IL-10和水平显著低于LPS+白藜芦醇组(P<0.05)。结论白藜芦醇可能促进了LPS刺激的RAW264.7巨噬细胞向M2型极化改变。
Objective To investigate the effect of resveratrol on murine macrophage cell line （RAW264.7 ceils） polarizing phenotype induced by lipopolysaccharide. Methods RAW264.7 mouse macrophages seeded in a 6 well plate,then randomly divided into phosphate buffer saline（PBS） control group, LPS （100 ng/mL） group, and LPS （100 ng/mL）＋ resveratrol （30 μmol/L） group. In the LPS＋ resveratrol group,LPS was added after incubation with resveratrol for 12 h. Cells were harvested and superna- rant were collected after incubation with LPS for 12 h. Both the mRNA expression levels of M1 associated markers iNOS and TNF- and M2 associated markers IL-10, PPARγ and Arg-1 were measured by real time quantitative PCR. Expression of iNOS, Arg-1 protein were detected by Western blot,inflammatory factor IL-12 p40,IL-10 and TNF-α protein in the supernatant of were assayed by ELISA. Results PCR detection showed that the mRNA expression levels of M1 associated markers iNOS and TNF-α in the LPS group were significantly higher than that of LPS＋ resveratrol group（P〈0.05） ,but the mRNA expression levels of M2 associated markers IL-10,PPARγ, and Arg-1 were significantly lower than that of LPS＋ resveratrol group（P〈0.05）. Compared with LPS＋ resveratrol group,western blot assay showed that iNOS protein level in LPS group was significantly higher than it （P〈0.05）, but Arg-1 protein level was significantly lower than it（P〈0.05）. The levels of IL-12 p40 and TNF-α in LPS group were significantly higher than that in LPS＋ resveratrol group（P〈0.05） ,but the levels of IL-10 was significantly lower than it（P〈0.05）. Conclusion Resveratrol may promote LPS stimulated RAW264.7 macrophage polarization to M2 phenotype.
出处
《重庆医学》
CAS
北大核心
2015年第19期2593-2595,2599,共4页
Chongqing medicine
基金
国家自然科学基金资助项目(81160233)
关键词
巨噬细胞
白藜芦醇
极化
macrophages
resveratrol
polarization
