摘要
目的:探讨回阳生肌汤对体外RAW264.7巨噬细胞极化的作用及可能机制。方法:采用体外培养RAW264.7巨噬细胞,使用脂多糖(LPS)和干扰素γ(IFN-γ)将RAW264.7巨噬细胞诱导为M1型(经典激活型巨噬细胞),白介素(IL)-4使其诱导为M2型(替代激活型巨噬细胞)。采用CCK-8法检测回阳生肌汤提取物对巨噬细胞增殖作用。回阳生肌汤提取物预先作用巨噬细胞24 h,LPS+IFN-γ刺激12 h后,通过比色法检测细胞上清液中一氧化氮(NO)水平;luminex高通量检测技术检测细胞上清液中IL-1β、IL-6、IL-12、肿瘤坏死因子α(TNF-α)、IL-10、血管内皮生长因子(VEGF)的含量;流式细胞术检测RAW264.7巨噬细胞表面白细胞分化抗原86(CD86)、甘露糖受体(CD206)阳性率;Western Blot检测一氧化氮合酶(iNOS)、精氨酸酶1(Arg-1)、磷酸化信号传导和转录激活蛋白6(p-STAT6)和STAT6的表达。结果:回阳生肌汤提取物在520、260、130 mg/L时对巨噬细胞有增殖作用;与模型组比较,回阳生肌汤提取物能显著抑制巨噬细胞NO释放(P<0.01,P<0.05),抑制IL-1β、IL-6、IL-12分泌,同时促进IL-10分泌(P<0.01);提高RAW264.7巨噬细胞CD206阳性率(P<0.01);抑制iNOS蛋白表达,促进Arg-1、p-STAT6蛋白表达(P<0.01)。结论:回阳生肌汤提取物体外可抑制LPS+IFN-γ诱导的RAW264.7巨噬细胞向M1型极化,促进向M2型极化,可能通过激活STAT6信号通路发挥作用。
Objective:To investigate the polarization effect of Huiyang Shengji Decoction on macrophages and its possible mechanism.Methods:In this experiment,RAW264.7 macrophages were cultured in vitro,and lipopolysaccharide(LPS)and interferonγ(IFN-γ)were used to induce macrophages into M1 type(classical activated macrophages),interleukin(IL)-4 was used to induce M2 type(replacement of activated macrophages).The CCK-8 method was used to detect the proliferation effect of the extract of Huiyang Shengji Decoction on RAW264.7 macrophages.The extract of Huiyang Shengji Decoction was preacted for 24 h,and after 12 h stimulation with LPS+IFN-γ,the level of nitric oxide(NO)in the cell supernatant was detected by colorimetry;the IL-1β,IL-6,IL-12,tumor necrosis factorα(TNF-α),IL-10,vascular endothelial growth factor(VEGF)in the cell supernatant was detected by luminex high-throughput detection technology.The expression of mannose receptor(CD206)protein was detected by Flow cytometry.The expression of nitric oxide synthase(iNOS),arginase 1(Arg-1),phospho-signal transduction and activation of transcription protein 6(p-STAT6)and STAT6 was detected by Western Blot.Results:The extract of Huiyang Shengji Decoction at 520,260,130 mg/L had a proliferation effect on macrophages.Compared with the model group,the extract of Huiyang Shengji Decoction could significantly reduce the release of NO of macrophages(P<0.01,P<0.05);reduced the secretion of IL-1β,IL-6,and IL-12,simultaneously promoted the secretion of IL-10 and the expression of CD206(P<0.01),and reduced iNOS protein expression,but increased Arg-1 and p-STAT6 protein expression(P<0.01).Conclusion:The extract of Huiyang Shengji Decoction can inhibit LPS+IFN-γ-induced macrophages to polarize to M1 type,and promote the polarization to M2 type,which may play a role by activating the STAT6 signaling pathway.
作者
陈佳
刘青武
韩旭阳
刘欣
林燕
马慧可
何秀娟
李萍
CHEN Jia;LIU Qing-wu;HAN Xu-yang;LIU Xin;LIN Yan;MA Hui-ke;HE Xiu-juan;LI Ping(Beijing Hospital of Traditional Chinese Medicine,Capital Medical University,Beijing 100010,China;Beijing Institute of Traditional Chinese Medicine,Beijing 100010,China;Beijing University of Chinese Medicine,Beijing 100029,China)
出处
《中华中医药杂志》
CAS
CSCD
北大核心
2022年第6期3503-3509,共7页
China Journal of Traditional Chinese Medicine and Pharmacy
基金
国家自然科学基金项目(No.81774328,No.81804093)。
