摘要
目的 观察绿原酸对脂多糖(LPS)致巨噬细胞激活的影响,并探讨骨髓细胞2表达的触发受体(TREM2)蛋白在其中的作用。方法 为筛选LPS造模浓度,分别以1、10、100 ng/mL的LPS培养细胞24 h,检测细胞培养上清液中白细胞介素6(IL-6)水平和细胞中诱导型一氧化氮合酶(i NOS)蛋白表达水平。为筛选绿原酸给药浓度,将细胞分为Control组、LPS处理组和3个不同浓度绿原酸(0.01、0.1、1μmol/L)干预组,检测细胞培养上清液中肿瘤坏死因子α(TNF-α)、IL-1β水平和细胞中iNOS、TREM2蛋白表达水平以及细胞活力。为观察TREM2在绿原酸抑制巨噬细胞激活中的作用,采用TREM2小干扰RNA(TREM2-siRNA)干扰细胞TREM2蛋白表达,将细胞分为Control组、LPS处理组、绿原酸+LPS组、TREM2-siRNA+绿原酸+LPS组和乱序-siRNA(SCsiRNA)+绿原酸+LPS组,在含有上述药物的培养基/空白培养基中孵育24 h后,检测细胞培养上清液中TNF-α和IL-1β水平以及细胞中TREM2、iNOS和核因子κB p65(NF-κB p65)的蛋白表达水平。结果 10 ng/mL LPS可显著促进细胞释放IL-6,并增加细胞中iNOS蛋白表达水平,故选择该质量浓度LPS进行后续实验。与LPS处理组相比,0.1μmol/L绿原酸干预组细胞培养上清液中TNF-α、IL-1β水平和细胞中iNOS蛋白表达水平均显著降低(P<0.05),细胞中TREM2蛋白表达水平显著升高(P<0.05),细胞活力未受影响,故选择该浓度的绿原酸进行后续实验。在观察TREM2蛋白在绿原酸抑制巨噬细胞激活中的作用中发现,与Control组比较,LPS处理组细胞中iNOS和NF-κB p65蛋白表达水平均显著升高(P<0.05);与LPS处理组比较,绿原酸+LPS组细胞中iNOS和NF-κB p65蛋白表达水平均显著降低、TREM2蛋白表达水平显著升高(P<0.05);与绿原酸+LPS组比较,TREM2-siRNA+绿原酸+LPS组细胞中iNOS和NF-κB p65蛋白表达水平均显著升高、TREM2蛋白表达水平显著降低(P<0.05),TREM2-siRNA明显逆转了绿原酸的上述作用;而SC-siRNA
OBJECTIVE To observe the effects of chlorogenic acid on the activation of macrophage induced by lipopolysaccharide(LPS),and to explore the role of triggering receptors expressed on myeloid cells-2(TREM2) in the action.METHODS To find a suitable LPS concentration,the cells were cultured with 1,10 and 100 ng/mL LPS for 24 h.The level of interleukin 6(IL-6) in the cell culture supernatant and protein expression of inducible nitric oxide synthase(iNOS) in the cells were detected.To search for a suitable chlorogenic acid concentration,the cells were divided into control group,LPS group and three chlorogenic acid(0.01,0.1 and 1 μmol/L)+LPS groups.The levels of tumor necrosis factor α(TNF-α) and IL-1β in the cell culture supernatant,the protein expressions of iNOS and TREM2 in the cells and cell viability were detected.To observe the effects of TREM2 in chlorogenic acid alleviating macrophage activation,TREM2-small interfering RNA(TREM2-siRNA) was taken to intervene in TREM2 protein expression.The cells were divided into control group,LPS group,chlorogenic acid+LPS group,TREM2-siRNA+chlorogenic acid+LPS group and SC-siRNA+chlorogenic acid+LPS group.After 24 h incubation,the levels of TNF-α and IL-1β in the cell culture supernatant and protein expressions of TREM2,iNOS and nuclear factor κB p65(NF-κB p65) in the cells were detected.RESULTS 10 ng/m L LPS promoted IL-6 release and increased i NOS protein expression,and 10 ng/m L LPS was taken in the next experiments.Compared with the LPS group,0.1 μmol/L chlorogenic acid decreased TNF-α and IL-1β levels,and down-regulated i NOS expression,meanwhile increased TREM2 expression without effect on cell viability,and 0.1 μmol/L chlorogenic acid was taken in the next experiments.Compared with the control group,the protein expressions of i NOS and NF-κB p65 in the LPS group were significantly increased(P<0.05);compared with the LPS group,the protein expressions of i NOS and NF-κB p65 in the chlorogenic acid+LPS group were significantly decreased,the protein expressio
作者
郑薇
郎静
黄西凤
肖锐
白荷
贾济
ZHENG Wei;LANG Jing;HUANG Xifeng;XIAO Rui;BAI He;JIA Ji(Dept.of Anesthesiology,General Hospital of Southern Theatre Command,Guangzhou 510501,China;Dept.of Orthopedics,General Hospital of Western Theatre Command,Chengdu 610083,China)
出处
《中国药房》
CAS
北大核心
2023年第21期2601-2607,共7页
China Pharmacy
基金
广东省自然科学基金项目(No.2017A030313810)。
关键词
绿原酸
巨噬细胞
脂多糖
炎症
骨髓细胞2表达的触发受体
chlorogenic acid
macrophage
lipopolysaccharide
inflammation
triggering receptors expressed on myeloid cells-2
