摘要
目的观察小鼠RAW264.7巨噬细胞不同极化状态下lincRNA-cox2表达变化，初步探讨lincRNA-cox2对巨噬细胞极化的影响。方法以IFN-γ/LPS刺激小鼠RAW264.7细胞诱导M1型极化，IL-4刺激RAW264.7细胞诱导M2型极化，采用实时荧光定量PCR（RT-PCR）检测各组细胞lincRNA-cox2的表达。设计并合成针对lincRNA-cox2基因的siRNA及无关序列，脂质体转染RAW264.7细胞，48 h后分别加入IFN-γ/LPS或IL-4继续诱导M1/M2型巨噬细胞，采用酶联免疫试验（ELISA）检测细胞因子IL-12及IL-10的分泌量；RT-PCR检测与巨噬细胞极化相关的iNOS、TNF-α、Arg-1、Fizz1的表达。将lincRNA-cox2-siRNA转染M1型巨噬细胞，检测其对M1极化表型的影响。结果RT-PCR结果显示，M1型巨噬细胞的lincRNA-cox2表达水平显著高于未极化细胞和M2型巨噬细胞。与对照组相比，lincRNA-cox2-siRNA转染RAW264.7细胞后，经IFN-γ/LPS诱导的M1型巨噬细胞中IL-12分泌降低，iNOS和TNF-α表达下降；而经IL-4诱导的巨噬细胞中IL-10产生增加，Arg-1和Fizz1表达升高。lincRNA-cox2-siRNA转染M1型巨噬细胞后，IL-12分泌降低，IL-10产生增加，其特点向M2样巨噬细胞转换。结论lincRNA-cox2参与调控小鼠巨噬细胞的极化，促进M1型巨噬细胞极化，抑制M2型巨噬细胞极化。
To investigate the effects of lincRNA-cox2 on the polarization of murine RAW264.7 macrophages by analyzing the expression of lincRNA-cox2 in RAW264.7 macrophages of M1 and M2 phenotypes.Methods Murine RAW264.7 cells were induced by IFN-γ and LPS to polarize to M1 phenotype, and were induced by IL-4 to polarize to M2 phenotype. The expression of lincRNA-cox2 in M1 and M2 macrophages were analyzed by real-time quantitative PCR （RT-PCR）. We designed and synthesized siRNA oligo for lincRNA-cox2 and unrelated sequences. Then the siRNA oligo and NC oligo were transfected into RAW264.7 cells by Lipofectmine^TM 2000. The transfected RAW264.7 cells were induced by IFN-γ and LPS or by IL-4 to polarize to M1 or M2 macrophages. Enzyme linked immunosorbent assay （ELISA） was performed to measure the secretion of IL-10 and IL-12 induced in different conditions. The expression of inducible nitric oxide synthase （iNOS）, TNF-α, arginase 1 （Arg-1） and found in inflammatory zone 1 （Fizz1） at mRNA level were detected by RT-PCR. The M1 macrophages were transfected with siRNAs to knock down the expression of lincRNA-cox2 for analyzing the biological effects of lincRNA-cox2 on the polarization of macrophages.ResultsThe relative expression of lincRNA-cox2 in M1 macrophages was significantly higher than that in RAW264.7 cells and M2 macrophages. Compared with the control group, the RAW264.7 cells transfected with lincRNA-cox2-siRNA showed decreased secretion of IL-12 and inhibited expression of iNOS and TNF-α at mRNA level after IFN-γ and LPS induction, but increased secretion of IL-10 and enhanced expression of Arg1 and Fizz1 at mRNA level after IL-4 induction. Transfecting the M1 macrophages with lincRNA-cox2-siRNA inhibited the secretion of IL-12, but promoted the secretion of IL-10.ConclusionThis study indicated that lincRNA-cox2 was involved in the regulation of macrophage phenotypes by promoting the polarization to M1 macrophages and inhibiting the polarization to M2 macrophages.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2016年第12期881-886,共6页
Chinese Journal of Microbiology and Immunology
基金
国家自然科学基金项目（81560001）
江西省青年科学基金项目（20142BAB215057）
江西省科技支撑计划项目（20151BBG70208）
江西省教育厅青年基金项目（GJJ14176）
江西省卫生和计生委科技项目（20161018）
