人参皂甙Rg3下调树突状细胞IDO杀伤小鼠肾癌细胞的实验研究

Ginsenoside Rg3 Down-Regulates IDO in Dendritic Cells to Kill Mouse Renal Cancer Cells

目的探讨人参皂甙Rg3促进树突状细胞(DC)的免疫增强作用,比较Rg3 DC疫苗与常规DC疫苗的抗肿瘤作用及机制。方法分离小鼠骨髓细胞制备DC,以普通DC诱导培养体系为Control组(C组),G10、G20、G50组在DC培养液中分别加入10、20、50μg·mL^(-1)人参皂甙Rg3,观察4组DC形态变化,诱导6 d后流式细胞术(FCM)检测CD11c、CD80和CD86、MHC-Ⅱ的表达;以RAG肿瘤抗原致敏后成熟DC将其中吲哚胺-2,3-双加氧酶(IDO)沉默,用qPCR方法检测IDO沉默率,然后与同种异体T细胞混合培养,采用CCK-8法检测淋巴细胞增殖功能,ELISA法检测共培养上清IL-12、IFN-γ水平;经激活的细胞毒性T淋巴细胞(CTL)与肿瘤细胞共培养,采用CCK-8法检测CTL对靶细胞的杀伤能力。结果G20组人参皂甙Rg3可以明显下调DC中IDO的表达,且诱导CTL杀伤肿瘤的能力明显强于其他组(P<0.05),IL-12、IFN-γ的表达水平随着Rg3浓度的升高而升高(P<0.05)。结论一定浓度范围内的Rg3可在体外下调DC的IDO表达,促进DC成熟,进而促进其抗原递呈能力,有效地活化和增强CTL的抗肿瘤作用。

Objective To investigate the immunological enhancement effect of ginsenoside Rg3 on dendritic cells(DCs),and to compare the antitumor effect and mechanism of Rg3 DC vaccine and conventional DC vaccine.Methods DCs were isolated from mouse bone marrow cells.The cells in the control group were cultured in the common induction system.The cells in G10,G20 and G50 groups were additionally given 10,20 and 50μg·mL^(-1) Rg3,respectively.The morphological changes of DCs were observed in all the four groups.The expression of CD11c,CD80,CD86 and MHC-Ⅱwas detected by flow cytometry after 6 days of induction.Indolylamine 2,3-dioxygenase(IDO)was silenced in mature DCs after RAG tumor antigen sensitization.After detecting the silencing rate by qPCR,DCs were co-cultured with allogeneic T cells.The proliferation of lymphocytes was determined by CCK-8 assay,and levels of IL-12 and IFN-γin the supernatant were measured by ELISA.The activated cytotoxic T lymphocytes(CTLs)were co-cultured with tumor cells,and the cytotoxicity of CTLs to target cells was detected by CCK-8 method.Results Treatment with 20μg·mL^(-1) Rg3 down-regulated IDO expression in DCs(P<0.05).Furthermore,the ability of inducing CTLs to kill tumor in G20 group was better than that in other groups(P<0.05).In addition,the expression of IL-12 and IFN-γincreased with the Rg3 concentrations(P<0.05).Conclusion In a certain concentration range,Rg3 can down-regulate IDO expression,promote maturation,and thus motivate antigen presentation ability in DCs.Therefore,it effectively activates and enhances the antitumor effect of CTLs.