摘要
目的:探讨棕榈酸(PA)对小鼠骨髓来源巨噬细胞(BMDM)极化的影响及机制。方法:提取BALB/c小鼠骨髓细胞,于含有10%胎牛血清和20 ng/ml GM-CSF的DMEM低糖培养基中诱导7 d后得到BMDM,给予不同浓度PA,CCK8检测巨噬细胞活性变化,确定最佳诱导浓度。倒置显微镜观察诱导后巨噬细胞形态。RT-PCR检测TNF-α、IL-6、IL-12、诱导型一氧化氮合酶(iNOS)、转化生长因子-β(TGF-β)、IL-10、Fizz1及YM-1 mRNA表达,流式细胞术检测NOS2、CD206表达,Western blot检测MAPK信号通路相关蛋白表达。结果:PA诱导巨噬细胞的最佳浓度为400μmol/ml,巨噬细胞呈现M1型极化样的长梭形,与未处理组相比,PA诱导组炎症细胞因子TNF-α、IL-6、IL-12及iNOS mRNA水平在诱导后12~72 h表达升高,而抑炎细胞因子TGF-β、IL-10、Fizz1及YM-1的转录水平受到抑制,48 h时,巨噬细胞高表达NOS2,而CD206表达一直处于较低水平。PA诱导巨噬细胞MAPK信号通路相关蛋白p-ERK、p-JNK及p-p38高表达,p-65入核。结论:PA可诱导小鼠BMDM M1型极化,可能通过MAPK信号通路传导信号给NF-κB影响巨噬细胞极化。
Objective:To investigate effect of palmitic acid(PA)on polarization of mouse bone marrow-derived macrophages(BMDM)and its mechanism.Methods:Mouse BMDM were harvested and cultured in DMEM low glucose medium containing 10%fetal bovine serum with 20 ng/ml GM-CSF for 7 d to obtain BMDM.BMDMs were treated with different concentrations of PA and CCK8 was used to detect cell viability of macrophages and determined optimal concentration for induction.Morphology of macrophages was observed after induction by inverted microscope.Relative mRNA expression levels of TNF-α,IL-6,IL-12,inducible nitric oxide synthase(iNOS),transforming growth factor-β(TGF-β),IL-10,Fizz1,YM-1 were determined by RT-PCR.Expression levels of NOS2 and CD206 were determined by flow cytometry.MAPK signaling pathway-related proteins were detected by Western blot.Results:Optimal concentration of PA-induced macrophages was 400μmol/ml.Macrophages showed a long fusiform shape like M1 phenotype.Compared with untreated group,PA induced groups mRNA expression levels of flammatory cytokines TNF-α,IL-6,IL-12 and iNOS were increased at 12-72 h,while transcription levels of anti-inflammatory cytokines TGF-β,IL-10,Fizz1,YM-1 were inhibited;macrophage incubated with PA expressed NOS2 at 48 h,while expression of CD206 was always at a low level.PA induced high expressions of p-ERK,p-JNK,p-p38 that related to JNK/P38 MAPK signaling pathway,and p-65 was transferred to nucleus.Conclusion:PA induces expressions of macrophage inflammatory cytokines and signals to NF-κB via MAPK signaling pathway,which induces M1 polarization of BMDM.
作者
张焱皓
刘芊伊
冯继红
李茂
刘利萍
秦欢
罗军敏(指导)
ZHANG Yan-Hao;LIU Qian-Yi;FENG Ji-Hong;LI Mao;LIU Li-Ping;QIN Huan;LUO Jun-Min(Department of Immunology,Zunyi Medical University,Immune Molecules Application Research Center of Guizhou Province,Zunyi 563003,China)
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2020年第20期2433-2437,共5页
Chinese Journal of Immunology
基金
国家自然科学基金地区项目(No.81860291)
浙江省中医药管理局课题项目(2020ZB292)。
关键词
巨噬细胞
棕榈酸
极化
MAPK信号通路
Macrophages
Palmitic acid
Polarization
MAPK signaling pathway
