中文摘要：

目的探讨miR-148b/DUSP1信号通路调节巨噬细胞分泌细胞因子CD206的表达对肝癌发生的影响。方法利用全自动磁珠提取纯化系统提取外周血单核细胞并培养,用粒细胞-巨噬细胞集落刺激因子（granulocyte-macrophage colony stimulating factor,GM-CSF）和巨噬细胞集落刺激因子（macrophage colony stimulating factor,M-CSF）分别诱导生成M1型和M2型巨噬细胞,CD68、CD206进行表型鉴定,ELISA检测M1和M2型巨噬细胞分泌的细胞因子CD206的表达,CCK-8和Transwell实验检测巨噬细胞分泌细胞因子CD206对肝癌细胞（Hep G2和Huh7细胞）增殖、侵袭、转移的影响,双荧光素酶报告基因系统验证miR-148b与DUSP1的靶向结合。结果初步分离并鉴定M1和M2型巨噬细胞,M2型巨噬细胞分泌的细胞因子CD206促进肝癌细胞的生长和侵袭、转移,双荧光素酶报告基因证实DUSP1为miR-148b的靶基因,miR-148b/DUSP1信号通路促进肝癌的发生、发展,巨噬细胞标志物与肝癌患者的临床病理特征相关。结论 miR-148b/DUSP1信号通路影响巨噬细胞分泌的细胞因子促进肝癌发生、发展。

英文摘要：

Objective To determine the effect of miR-148 b /DUSP1 signaling pathway on the level of cytokine CD206 produced by macrophages and on the growth of hepatocarcinoma cells. Methods Peripheral blood mononuclear cells（ PBMC） were screened with CD14 microbeads,and then induced to M1 and M2 macrophages with granulocyte-macrophage colony stimulating factor（ GM-CSF） and macrophage colony stimulating factor（ M-CSF） respectively. Phenotypes and functions of the obtained polarized monocyte-derived macrophages were identified by CD68 and CD206. ELISA was used to determine the secretion of CD206 by M1 and M2 macrophages. CCK-8 and Transwell assays were used to verify the growth,invasion and migration of hepatoma cells（ Hep G2 and Huh7 cell lines） after the treatment of the secreted growth factors. Dual luciferase reporter system was used to determine whether DUSP1 was a bona fide target of miR-148 b. Results Monocyte-derived M1 and M2 macrophage cultures were established successfully. The presence of growth factor CD206 produced by M2 macrophages promoted the proliferation,invasion and migration of hepatoma cell lines. Dual luciferase reporter system indicated that miR-148 b functioned to directly suppress DUSP1 gene expression through the miR-148b-binding sequence located at the 3＇-UTR of the DUSP1 mRNA. miR-148 b /DUSP1 signaling pathway promoted the occurrence and development of hepatoma,and macrophage markers were associated with clinical and pathological features of liver cancer. Conclusion miR-148 b/DUSP1 signaling pathway affects macrophage-secreted cytokines in promotion of occurrence and development of hepatocarcinoma,and may provide new evidence for targeted therapy of liver cancer.