黄芪甲苷对马兜铃酸诱导的巨噬细胞M1型极化的作用及机制研究  

Effect of Astragaloside Ⅳ on Aristolochic Acid-induced M1-type Polarization of Macrophages and Its Mechanism Study

作　　者：徐维佳 陈揭剑[1,2] 严丽群 张勇 王丽萍 庄永泽[1,2] XU Wei-jia;CHEN Jie-jian;YAN Li-qun;ZHANG Yong;WANG Li-ping;ZHUANG Yong-ze(Department of Nephrology,900 Hospital of PLA Joint Service Support Force,Fuzhou,Fujian,350025,China;Fuzhou General Hospital of Clinical Medicine,Fujian Medical University,Fuzhou,Fujian,350004,China)

机构地区：[1]中国人民解放军联勤保障部队第九〇〇医院肾脏病科,福建福州350025 [2]福建医科大学福总临床医学院,福建福州350004

出　　处：《现代生物医学进展》2021年第6期1040-1045,共6页Progress in Modern Biomedicine

基　　金：福建省自然科学基金面上项目(2017J01324);国家自然科学基金面上项目(81373837);福建省临床重点专科建设项目(闽卫医政函[2017]739号)。

摘　　要：目的:探讨黄芪甲苷对马兜铃酸诱导的RAW264.7细胞向M1型极化的影响,并初步探索其可能的作用机制。方法:分别采用马兜铃酸和脂多糖(LPS)刺激RAW264.7细胞24 h,伴或不伴黄芪甲苷进行药物干预处理。采用细胞计数检测试剂盒-8(CCK 8)检测细胞活性变化,流式细胞仪检测巨噬细胞分型,酶联免疫吸附试验(ELISA)检测细胞上清液中白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)的分泌量。反转录实时定量PCR(RT-qPCR)技术检测RAW264.7细胞IL-6、TNF-αmRNA表达。蛋白免疫印迹法(Western blot)检测RAW264.7细胞p-p38和p38 MAPK蛋白表达水平。结果:CCK8结果提示黄芪甲苷在5~50μg/mL浓度范围对RAW264.7巨噬细胞无明显毒性,本研究选取10μg/mL作为实验干预浓度。黄芪甲苷能够显著改善马兜铃酸诱导的巨噬细胞活性(P<0.05),同时减少IL-6和TNF-α的分泌水平和mRNA表达水平(均P<0.05),抑制马兜铃酸和LPS诱导的M1/M2巨噬细胞比例(P<0.05)。黄芪甲苷可部分抑制马兜铃酸诱导的巨噬细胞p38 MAPK磷酸化水平(P<0.05)。结论:黄芪甲苷可减少巨噬细胞M1型极化,降低炎症因子IL-6和TNF-α水平,减少巨噬细胞的活性,从而起到减缓马兜铃酸肾损害的作用,其作用机制可能与部分抑制p38 MAPK信号活性有关。Objective: To investigate the effect of astragaloside Ⅳ on the polarization of RAW264.7 cells induced by aristolochia acid to M1-type, and to explore its possible mechanism. Methods: Aristolocholic acid and Lipopolysaccharide(LPS) were used to stimulate RAW264.7 cells for 24 h successively, with or without astragaloside Ⅳ for drug intervention. Cell counting kit-8(CCK-8) was used to detect the change of cell activity, flow cytometry was used to detect the type of macrophages, and enzyme-linked immunosorbent assay(ELISA) was used to detect the secretion of interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α) in cell supernatant. The expression of IL-6 and TNF-α m RNA in RAW264.7 cells were detected by real-time quantitative reverse transcription PCR(RT-qPCR). The protein expression levels of p-p38 and p38 MAPK in RAW264.7 cells were detected by Western blot. Results: The results of CCK8 indicated that astragaloside Ⅳ had no obvious toxicity to RAW264.7 macrophages in the concentration range of 5-50 μg/mL, and 10 μg/m L was selected as the experimental intervention concentration in this study. Astragaloside IV could significantly improve the increase of macrophage activity induced by aristolochic acid(P<0.05), and decreased the secretion and expression levels of IL-6, TNF-α and m RNA expression(all P<0.05), inhibit the increase of M1/M2 macrophage ratio induced by aristolochic acid and LPS(P<0.05). Astragaloside IV could partially inhibited the increased phosphorylation of p38 MAPK in macrophages induced by aristolochic acid(P<0.05). Conclusions: Astragaloside Ⅳ can reduce the M1-type polarization of macrophages, reduce the levels of inflammatory cytokines IL-6 and TNF-α, and reduce the activity of macrophages, thus playing a role in slowing the renal damage of aristolochian acid. The mechanism of action may be related to the partial inhibition of p38 MAPK signaling activity.

关 键 词：黄芪甲苷 马兜铃酸 巨噬细胞 M1型极化 p38 MAPK信号通路 

分 类 号：R-33[医药卫生] R692