To construct recombinant eukaryotic expression plasmid vector of human IL-34 gene, and to study the effects of IL-34 expressed by human bone marrow-derived mesenchymal stem cells (hBM-MSCs) on THP-1 cells. Full-length IL-34 encoding sequence was amplified by PCR. And this fragment was cloned into the plasmid pIRES2-EGFP. Western blotting and ELISA were used to analyze the expression of IL-34 in hBM-MSCs. THP-1 cells were cultured with hBM-MSCs medium containing IL-34 protein. Real-time PCR detected the effects of IL-34 on the expression of IL-10 and TNFα in THP-1 cells. Restrictive enzyme analysis and sequencing demonstrated that IL-34 eukaryotic expression vector was successfully constructed. IL-34 protein expressed by hBM-MSCs could promote IL-10 and TNFα expression in THP-1 cells. Those results show that IL-34 expressed by hBM-MSCs has regulating effect on THP-1 cells.
构建人IL-34 真核表达载体并将其转染到人骨髓间充质干细胞，观察高表达IL-34 的骨髓间充质干细胞对THP-1 细胞的影响。PCR 扩增IL-34 DNA，并将其克隆到真核表达载体pIRES2-EGFP；将构建成功的重组体转染到骨髓间充质干细胞，Western blotting 和ELISA 分析IL-34 在细胞中的表达；用高表达IL-34 的骨髓间充质干细胞培养上清液来培养THP-1 细胞，Real-time PCR 分析THP-1 细胞中IL-10 和TNFα 的表达变化。经双酶切和测序鉴定，成功构建了pIRES2-EGFP-IL-34 重组体；转染至骨髓间充质干细胞的IL-34 可以促进THP-1 细胞表达IL-10 和TNFα。结果表明，骨髓间充质干细胞表达分泌的IL-34 对THP-1 有调节作用。.
Keywords: IL-10; IL-34; THP-1 cells; TNFα; human bone marrow derived mesen chymal stem cells.