二妙散对巨噬细胞分化的调控作用研究
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摘要
目的研究二妙散对脂多糖(lipopolysaccharide,LPS)诱导巨噬细胞分化的影响。方法噻唑蓝(methylthiazolyl tetrazolium,MTT)法测定二妙散水煎液对细胞活力的影响;将LPS(100ng·mL~(-1))及不同浓度(0.1,1.0,10mg·mL~(-1))的二妙散水煎液共同作用RAW264.7细胞后,酶联免疫吸附法(enzyme-linked immune sorbent assay,ELISA)检测上清液中肿瘤坏死因子-α(tumornecrosisfactor-α,TNF-α)、白介素-10(interleukin10,IL-10)的含量,实时荧光定量PCR(Real-time fluorescent quantitative PCR,qPCR)检测RAW264.7细胞中白介素-1β(Interleukin 1β,IL-1β),IL-6的mRNA水平,Western blot检测RAW264.7细胞NOS2的蛋白表达情况。结果二妙散水煎液在0.01～100mg·mL~(-1)内对RAW264.7细胞无明显毒性。与LPS诱导的模型组相比,二妙散水煎液能明显降低细胞上清液中TNF-α的含量(P<0.05)、升高IL-10的含量(P<0.05),且可显著抑制RAW264.7细胞中M1分化标记物IL-1β、IL-6的mRNA水平(P<0.05)及NOS2的蛋白表达(P<0.05)。结论二妙散可通过抑制巨噬细胞向M1促炎方向分化,从而发挥抗炎作用。
OBJECTIVE To investigate the effects of Ermiao Recipe on macrophage polarization induced by LPS.METHODS The cell viability was detected by MTT assay, and the contents of TNF-α and IL-10 in supernatant were determined by ELISA, the mRNA expression of IL-1β and IL-6 in RAW 264.7 were detected by qPCR and the protein expression of NOS2 in RAW 264.7 was detected by Western blot. RESULTS Ermiao Recipe water decoction significantly decreased the content of proinflammatory factor TNF-α and increased the content of anti-inflammatory factor IL-10 in cell supernatant, and also inhibited the protein expression of NOS2 and the mRNA level of IL-1β, IL-6, which were M1 polarization markers in RAW264.7 cells. CONCLUSION Ermiao Recipe can inhibit the polarization of macrophage into proinflammatory prescription and thus exert an anti-inflammatory effect.
OBJECTIVE To investigate the effects of Ermiao Recipe on macrophage polarization induced by LPS.METHODS The cell viability was detected by MTT assay, and the contents of TNF-α and IL-10 in supernatant were determined by ELISA, the mRNA expression of IL-1β and IL-6 in RAW 264.7 were detected by qPCR and the protein expression of NOS2 in RAW 264.7 was detected by Western blot. RESULTS Ermiao Recipe water decoction significantly decreased the content of proinflammatory factor TNF-α and increased the content of anti-inflammatory factor IL-10 in cell supernatant, and also inhibited the protein expression of NOS2 and the mRNA level of IL-1β, IL-6, which were M1 polarization markers in RAW264.7 cells. CONCLUSION Ermiao Recipe can inhibit the polarization of macrophage into proinflammatory prescription and thus exert an anti-inflammatory effect.
引文
[1]王勇.二妙散对类风湿关节炎大鼠抗炎免疫效果研究[J].中国民族民间医药,2010,19(9B):24.
[2]考希良,董嘉琪.二妙散不同配伍对大鼠佐剂性关节炎的抗炎效应[J].中医药学报,2013,41(3):107-109.
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[6]HAMILTON J A.Colony-stimulating factors in inflammation and autoimmunity[J].Nat Rev Immunol,2008,8(7):533-544.
[2]考希良,董嘉琪.二妙散不同配伍对大鼠佐剂性关节炎的抗炎效应[J].中医药学报,2013,41(3):107-109.
[3]BRENNAN F M,MCINNES I B.Evidence that cytokines play a role in rheumatoid arthritis[J].J Clin Invest,2008,118(11):3537-3545.
[4]FELDMANN M,BRENNAN F M,MAINI R N.Role of cytokines in rheumatoid arthritis[J].Ann Rev Immunol,1996,14(1):397-440.
[5]MANTOVANI A,SICA A,SOZZANI S,et al.The chemokine system in diverse forms of macrophage activation and polarization[J].Trends Immunol,2004,25(12):677-686.
[6]HAMILTON J A.Colony-stimulating factors in inflammation and autoimmunity[J].Nat Rev Immunol,2008,8(7):533-544.