髓源性抑制细胞(MDSC)相关趋化因子MIP-1γ及其受体CCR1在肝癌荷瘤小鼠脾脏的表达及意义

Expression and significance of myeloid-derived suppressor cells-associated chemokine MIP-1γ and its receptor CCR1 in the spleen of hepatoma H22-bearing mice

目的检测髓源性抑制细胞(MDSC)相关趋化因子在肝癌小鼠脾脏中的表达及其意义。方法用流式细胞术检测脾脏MDSC的比例,并采用流式细胞术分选出脾脏MDSC,与活化的脾细胞共培养,ELISA检测共培养上清中IFN-γ的表达水平。制备小鼠原位种植性肝癌模型,用蛋白芯片检测正常小鼠和荷瘤小鼠脾脏的细胞因子差异表达,并用ELISA对脾脏MDSC相关趋化因子进行验证。最后用流式细胞术检测脾脏MDSC表面的趋化因子受体的表达。结果荷瘤小鼠脾脏MDSC的比例显著升高,并且具有免疫抑制功能。蛋白芯片筛选出10个上调的细胞因子和9个下调的细胞因子。在上调的细胞因子中,5个趋化因子是B淋巴细胞趋化因子(BLC)、CXC趋化因子配体16(CXCL16)、巨噬细胞/单核细胞趋化蛋白5(MCP-5)、巨噬细胞炎性蛋白1γ(MIP-1γ)和巨噬细胞炎性蛋白2(MIP-2)。脾脏MDSC相关趋化因子MIP-1γ在荷瘤小鼠脾脏中显著升高,并且脾脏MDSC表面有其受体CC趋化因子受体1(CCR1)的表达。结论用蛋白芯片筛选出脾脏MDSC相关趋化因子MIP-1γ及其受体CCR1,可能与荷瘤小鼠脾脏MDSC聚集有关。

Objective To study the expression and significance of myeloid-derived suppressor cell(MDSC)-associated chemokines in the spleen of hepatoma H22-bearing mice.Methods The percentage of splenic MDSCs was examined by flow cytometry.Splenic MDSCs were sorted using flow cytometry and co-cultured with activated splenocytes,and then the level of IFN-γin the supernatant of co-cultured cells was assayed by ELISA.The murine orthotopic hepatoma model was established.The differential cytokine expression in the spleens of normal and tumor-bearing mice was assayed by protein chip,and splenic MDSC-associated chemokines were verified using ELISA.The chemokine receptors on splenic MDSCs were also detected by flow cytometry.Results The percentage of splenic MDSCs was markedly raised in the tumor-bearing mice,and splenic MDSCs exhibited immune-inhibitory function.Ten up-regulated cytokines and nine down-regulated ones were found out using protein chip.Among the up-regulated cytokines,5 cytokines were chemokines,namely B lymphocyte chemoattractant(BLC),chemokine(C-X-C motif)ligand 16(CXCL16),macrophage/monocyte chemotactic protein-5(MCP-5),macrophage inflammatory protein 1γ(MIP-1γ)and MIP-2.The level of splenic MDSC-associated chemokine MIP-1γsignificantly increased in the spleen of tumor-bearing mice,and its receptor CCR1 was also expressed on the cell surface of splenic MDSCs.Conclusion Splenic MDSC-associated chemokine MIP-1γand its receptor CCR1 were obtained by protein chip,and they might be associated with the accumulation of splenic MDSCs in hepatoma H22-bearing mice.