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目的探讨体外骨髓间充质干细胞(MSCs)联合雷帕霉素对BALB/c鼠T、B淋巴细胞的免疫调节作用及可能机制。方法从5～6周龄BALB/c鼠骨髓中分离培养MSCs并鉴定其纯度。用EZ-SepTM Mouse1X分离异体BALB/c鼠脾脏淋巴细胞,分别在刀豆蛋白A(ConA)、脂多糖(LPS)刺激下,用MSCs和/或雷帕霉素处理,MTT法检测淋巴细胞的增殖,流式细胞术检测T、B淋巴细胞CD69、CD28和CD86的表达和T淋巴细胞凋亡情况,Real-time PCR测定T淋巴细胞IL-10 mRNA、IFN-γ mRNA的表达水平。结果MSCs明显抑制T、B淋巴细胞增殖,联合雷帕霉素组抑制作用更加显著(P<0.01)。MSCs可抑制T淋巴细胞的凋亡,联合雷帕霉素组较单独MSCs组抑制作用更加明显(P<0.01)。MSCs联合雷帕霉素具有协同促进IFN-γ mRNA表达和协同抑制IL-10 mRNA表达的作用。单独MSCs或MSCs联合雷帕霉素对T、B淋巴细胞CD69、CD28和CD86的表达无显著影响。结论MSCs联合雷帕霉素对BALB/c鼠T、B淋巴细胞有免疫负调节作用,可能与协同抑制淋巴细胞增殖、T淋巴细胞凋亡和干预IFN-γ mRNA、IL-10 mRNA表达有关。
Objective To investigate the immunomodulatory effects of mesenchymal stem cells (MSCs) combined with rapamycin on T and B lymphocytes of BALB / c mice and its possible mechanism. Methods MSCs were isolated from bone marrow of 5-6 weeks old BALB / c mice and their purity was identified. The spleen lymphocytes of allogeneic BALB / c mice were isolated by EZ-SepTM Mouse1X and were treated with MSCs and / or rapamycin under the stimulation of ConA and LPS respectively. The levels of lymphocytes The expression of CD69, CD28 and CD86 and the apoptosis of T lymphocytes in T, B lymphocytes were detected by flow cytometry. The expression of IL-10 mRNA and IFN-γ mRNA in T lymphocytes were detected by Real-time PCR. Results MSCs significantly inhibited the proliferation of T and B lymphocytes, and the inhibitory effect was more significant when combined with rapamycin (P <0.01). MSCs can inhibit the apoptosis of T lymphocytes, and the combination of rapamycin and MSCs alone inhibited more obviously (P <0.01). MSCs combined with rapamycin have the synergistic effect of promoting the expression of IFN-γ mRNA and synergistically inhibiting the expression of IL-10 mRNA. MSCs alone or MSCs combined with rapamycin had no significant effect on the expression of CD69, CD28 and CD86 on T and B lymphocytes. Conclusion MSCs combined with rapamycin have immunoregulatory effects on T and B lymphocytes of BALB / c mice, which may be related to the synergistic inhibition of lymphocyte proliferation, T lymphocyte apoptosis and IFN-γ mRNA and IL-10 mRNA expression.