不同培养基对骨髓间充质干细胞免疫学特性的影响

Influences of Different Culture Media on the Immunological Characteristics of Bone Marrow Mesenchymal Stem Cells

目的比较ScienCell(SCI)干细胞培养基和低糖(LG)完全培养基培养下骨髓间充质干细胞(BMSC)的免疫学特性。方法利用流式细胞分析BMSC表面标志物的表达;通过体外成骨和成脂诱导,鉴定BMSC的多向分化能力;应用混合淋巴细胞反应实验,比较SCI组和LG组BMSC的免疫原性及免疫调节功能;采用流式细胞仪分析IFN-γ刺激前后BMSC表面免疫分子的表达情况。结果混合淋巴细胞反应结果显示,两组培养基培养的BMSC均具有低免疫原性和免疫抑制功能,但SCI组单向混合淋巴细胞增殖率显著高于LG组,而两组双向混合淋巴细胞增殖率未见显著差异。流式细胞仪分析结果显示,SCI组BMSC中免疫共刺激因子CD40阳性的细胞比例显著高于LG组,而HLA-DR以及免疫共抑制因子B7-H1和B7-DC阳性细胞比例均显著低于LG组。IFN-γ刺激后可显著上调两组BMSC中HLADR的阳性细胞比例至90%以上。低浓度的IFN-γ(10 U/mL)仅轻微上调LG组BMSCs的CD40的表达水平;而高浓度IFN-γ(1 000 U/mL)可显著提高两组BMSC中CD40和B7-H1的阳性细胞比例,以及SCI组BMSC中B7-DC的阳性细胞比例。结论在体外,含有不同成分的干细胞培养基可改变BMSC免疫分子的表达水平;SCI组BMSC的免疫原性显著高于LG组,但免疫抑制功能两组无明显差异。

Objective To compare the immunological characteristics of BMSCs cultured in ScienCell (SCI) stem cell growth medium and low glucose (LG) complete medium. Methods The expressions of cell surface markers on BMSC were characterized by flow cytometry analysis and the multipotency of BMSC was confirmed by functional osteogenic and adipogenic differentiation. The immunogenicity and immunomodulatory properties of BMSC in SCI and LG groups were assessed and compared by mix lymphocyte reaction (MLR). The expression profiles of immune molecules on BMSC with and without IFN-γ stimulation were characterized by flow cytometry analysis. Results The MLR results showed that BMSCs in both SCI group and LG group exhibited low immunogenicity and immunosuppressive abilities. One-way lymphocyte proliferation index was significantly higher in SCI group than in LG group while no significant difference was observed for the two-way mix lymphocyte proliferation index between SCI group and LG group. Compared with BMSC in LG group, BMSCs in SCI group exhibited significantly higher percentage of cells that positive for co-stimulatory molecule CD40 but lower percentage of cells expressing HLA-DR and co-inhibitory molecules B7-H1 and B7-DC. The percentages of HLA-DR+cells of BMSC in both SCI group and LG group were significantly increased to more than 90% in response to IFN-γ stimuli. Treatment with low concentration of IFN-γ (10 U/mL) only slightly increased the percentage of CD40+cells in BMSCs of LG group, while treatment with high concentration of IFN-γ (1 000 U/mL) significantly increased the expression of CD40 and B7-H1 of BMSCs in both groups, and B7-DC in BMSCs in LG group. Conclusion The expression of immune relevant molecules on BMSC cultured in SCI medium in vitro was changed. The immunogenicity of BMSC in SCI group was significantly higher than in LG group while no significant difference was observed for their immunosuppressive properties.