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目的 探讨转染IL 6基因的骨髓基质细胞系对同基因骨髓移植 (BMT)后小鼠免疫功能重建的促进作用。方法 将IL 6cDNA片段连接到pcDNA3真核表达载体上。用脂质体将pcD NA3IL 6转入骨髓基质细胞系QXMSC1,ELISA法测定转染IL 6基因骨髓基质细胞QXMSC1IL 6培养上清中IL 6的含量 ,有限稀释挑选多个细胞克隆 ,选择表达量最高的转基因细胞系QXMSC1IL 6用于动物实验。BABL c小鼠经γ射线致死量照射后 ,输入同基因骨髓细胞 (10 7 只 )同时输入骨髓基质细胞QXMSC1IL 6 (5× 10 5 只 )。在骨髓移植后 30d、6 0d检测BMT小鼠淋巴细胞对LPS ,ConA增殖反应 ,T辅助细胞前体 (helpTlymphocyteprecursor,HTLp) ,杀伤性T细胞前体 (cytotoxicTlymphocyteprecursor,CTLp) ,迟发型超敏反应 (delayed typehypersensitivity ,DTH)及空斑形成细胞数 (plaqueformingcell,PFC) ,反映骨髓移植后小鼠免疫功能。结果 成功构建pcDNA3IL 6重组体。该细胞体外培养 2 4h分泌IL 6的含量为 11.15 (± 2 .4 1) μg 10 6 。QXMSC1IL 6细胞系能明显增强BMT后淋巴细胞对LPS、ConA反应性 ,小鼠对异基因小鼠脾细胞DTH反应增强 ,脾脏中HTLp ,CTLp及PFC数明显增加。转入外源IL 6cDNA基因的骨髓基质细胞系QXMSC1IL 6在体内能明显促进BMT后小鼠T、B淋巴?
Objective To investigate the effects of transfection of bone marrow stromal cells transfected with IL-6 gene on the immune function of mice after homologous bone marrow transplantation (BMT). Methods IL 6 cDNA fragments were ligated into pcDNA3 eukaryotic expression vector. PcD NA3IL 6 was transfected into bone marrow stromal cell line QXMSC1 by lipofectamine. The content of IL 6 in the supernatant of QXMSC1IL 6 transfected with IL 6 gene was assayed by ELISA. Multiple cell clones were selected by limiting dilution, and the highest expression level The transgenic cell line QXMSC1IL 6 was used in animal experiments. BABL c mice were irradiated with γ-rays and then injected with the same gene myeloid cells (10 7) into the bone marrow stromal cells QXMSC1IL 6 (5 × 10 5). The proliferation of BMT mouse lymphocytes on LPS, ConA, T helper lymphocyte (HTLp), cytotoxic T lymphocyte precursor (CTLp), delayed type hypersensitivity delayed type hypersensitivity (DTH) and plaqueformingcell (PFC), reflecting the immune function of mice after bone marrow transplantation. Results The pcDNA3IL 6 recombinant was successfully constructed. The amount of IL-6 secreted by the cells cultured 24 h in vitro was 11.15 (± 2.41) μg 10 6. QXMSC1IL 6 cell line can significantly enhance the lymphocyte response to LPS and ConA after BMT, increase the DTH response of allogeneic mouse spleen cells in mice, and obviously increase the number of HTLp, CTLp and PFC in the spleen. Bone marrow stromal cell line QXMSC1IL6 transfected with exogenous IL6 cDNA can significantly promote T, B lymphocytes in mice after BMT.