目的 探讨小鼠骨髓间充质干细胞（Bone marrow mesenchymal stem cells, BMSCs）移植缓解1型糖尿病小鼠（T1DM）发病及机制。 方法 BMSCs分离培养及鉴定；第3代培养的上清作为条件培养基，与2周龄NOD小鼠脾细胞共培养，FACS检测CD4+和CD8+T细胞增殖及活化；2周龄雌性NOD小鼠随机分为BMSCs组、PBS组、对照组，每组6只，对照组2周处死；BMSCs组和PBS组12周分别腹腔注射100 μl（2×107细胞）BMSCs或等体积PBS，监测小鼠血糖和体重至26周；HE、免疫组化、免疫荧光染色观察胰腺炎性细胞浸润；FACS检测CD4+和CD8+T细胞增殖及活化。 结果 成功分离培养BMSCs；BMSCs条件培养基抑制CD4+及CD8+T细胞的增殖及活化（P<0.05）；BMSCs组小鼠体重高于PBS组（P<0.05），糖尿病发病率降低，炎性浸润减少（P<0.05），脾脏CD4+和CD8+T细胞增殖及活化降低（P<0.05）。结论 BMSCs移植可能通过T细胞免疫抑制缓解T1DM小鼠发病。
Abstract
Objective To investigate the effects and mechanism of bone marrow mesenchymal stem cells (BMSCs) transplantation on Type 1 diabetes mellitus (T1DM) mice. Methods Isolation, culture and identification of BMSCs were performed. BMSCs supernatant was used to be the conditioned medium after third passage (P3), co-cultured with 2 weeks NOD splenocytes, and the proliferation and activation of CD4+ and CD8+T cells were detected by FACS. Two weeks female NOD mice were randomly divided into three groups: a BMSCs group, a PBS group and a control group (6 mice in each group). Mice in the control group were killed at 2 weeks. 100 μl (2×107 cells) BMSCs or equal volume PBS was injected intraperitoneally at 12 weeks in the BMSCs group and the PBS group respectively, then the mice were monitored for blood glucose and body weight till 26 weeks. Inflammatory cells infiltration of pancreases was detected by H&E, immunohistochemical and immunofluorescent staining. The splenocytes were extracted to observe the proliferation and activation of CD4+ and CD8+T cells by FACS. Results BMSCs were successfully isolated and cultured. BMSCs conditioned medium inhibited the proliferation and activation of CD4+ and CD8+T cells (P<0.05). Mice body weight of BMSCs group was higher than that of PBS group (P<0.05), the incidence rate of diabetes and inflammatory infiltration decreased (P<0.05) as well as the proliferation and activation of CD4+ and CD8+ T cells in the spleen (P<0.05). Conclusions BMSCs transplantation may attenuate T1DM by immune suppressing T cells.
关键词
骨髓间充质干细胞 /
NOD/ShiLtJ小鼠 /
1型糖尿病 /
免疫抑制 /
T细胞
Key words
BMSCs /
NOD/ShiLtJ mice /
T1DM Immunosuppression /
T cells
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参考文献
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基金
贵州省科学技术基金项目[黔科合基础（2016）1125号] ；贵州省科技支撑计划（黔科合支撑 [2017]2970号）；贵州省普通高校科技拔尖人才支持计划（黔教合KY字[2017]071）；贵州省科技合作计划项目（黔科合LH字[2016]7363）；国家自然科学基金（NSFC 81660033）
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