间充质干细胞对巨噬细胞活化及其功能影响的实验研究

目的 探讨骨髓间充质干细胞(MSC)对脂多糖(LPS)诱导的巨噬细胞活化的影响.方法 采用贴壁筛选法分离、纯化小鼠骨髓MSC,巯基乙酸钠腹腔注射刺激后收集小鼠腹腔巨噬细胞(MPM).实验分为四组(A组:MPM;B组:MPM+LPS;C组:MPM+LPS+MSC;D组:MPM+LPS+MSC上清),建立共培养体系.在LPS(终浓度1 μg/ml)刺激巨噬细胞18 h后收集细胞培养上清,检测TNF-α、TGF-β和一氧化氮(NO)等细胞因子分泌量的变化,同时在体系中加入大肠杆菌标准菌株ATCC25922,共孵育24 h后瑞特染色检查巨噬细胞吞噬功能的变化.结果 巨噬细胞活化后培养上清中TNF-α和NO的含量明显上升分别为(147.4±37.1)pg/ml,(59.9±8.7)μmol/L];而MSC存在时,TNF-α显著减少(97.6±30.3)pg/ml,P=0.032],NO降到(50.9±29.5)μmol/L(P＞0.05);当有MSC上清存在时,TNF-α和NO进一步减少为(58.3±31.5)pg/ml,(-3.4±2.3)μmol/L(P＜0.01).MSC对巨噬细胞活化后的吞噬率及吞噬指数没有影响.结论 MSC可抑制LPS诱导的小鼠腹腔巨噬细胞的活化,而对其吞噬功能没有影响.

Experimental study on influence of bone marrow mesenchymal stem cells on activation and function of mouse peritoneal macrophages

Objective To explore the influence of bone marrow(BM)mesenchymal stem cells (MSCs)on macrophage activation after ipopolysarchride(LPS)stimulation.Methods Mouse BM MSCs were isolated and purified by adherence screening,and mouse peritoneal macrophages(MPM)were collected by sodium thioglycollate peritoneal injection,and the co-culture system was established by planting macrophages on the MSCs monolayer.The grouping of experiments:group A:MPM;group B:MPM+LPS;group C:MPM+LPS+MSC;group D:MPM+LPS+MSC supernatant.Cell culture supematants were collected to detect the changes of TNF-α/TGF-β and nitrogen monoxide(NO)after stimulating macrophages with LPS for 18 hours.At the same time Escbericbia coli standard strain(ATCC25922)was added into the culture system and incubated for another 24 hours,macrophages were stained and phagocytoses were examined.Results The concentrations of TNF-α and NO in culture supernatants were increased significantly to(147.4±37.1)Pg/ml and(59.9±8.7)μmol/L respectively after macrophage activation,however,at the present of MSC,the concentration of TNF-α was dramatically decreased(97.6 ±30.3)pg/ml,P=0.032],and the concentration of NO was decreased to(50.9±29.5)μmol/L(P＞0.05).The concentrations of TNF-α and NO were further decreased after addition of MSC supematants(58.3±31.5)pg/ml and(-3.4±2.3)μmoL/L respectively,P＜0.01].There was no change in the phagocytic rate and phagoindex of macrophages after activation.Conclusions MSCs can inhibit the activation of mouse peritoneal macrophages after stimulating with LPS but has no influence on the phagocytosis.