基质细胞衍生因子-1对骨髓源间充质干细胞迁移的影响

目的:利用Boyden chamber体外迁移体系初步探索基质细胞衍生因子-1(SDF-1)在骨髓源间充质干细胞(MSC)迁移中的作用及其信号转导机制。方法:采用经典的全骨髓贴壁法培养MSC,通过成骨、成脂肪等多向诱导分化以及流式细胞分析其表面标记(CD133、CD34、CD90、CD105)等鉴定MSC特征;以P3-MSC为实验材料,利用Boyden chamber体外迁移体系,先观察不同浓度的hSDF-1α对MSC迁移的影响,随后以50 M wortman-nin、10 uM LY294002、50 uM PD98059、10 uM U73122、0.1 mg/ml AMD3100等不同处理P3-MSC,观察最适宜浓度的hSDF-1α对MSC迁移影响的信号转导机制。结果:培养的MSC呈现出CD90、CD105强阳性,具有成骨、成脂肪等多向分化能力;MSC体外迁移能力随着hSDF-1α浓度(1 ng/ml、10 ng/ml、100 ng/ml、500 ng/ml)的递增而逐渐增强,并且hSDF-1α浓度在100 ng/ml时,MSC迁移到滤膜上的细胞数接近于锋值;Wortmannin、LY294002、PD98059、U70312、AMD3100对MSC迁移均有影响,其中U73122、AMD3100对MSC迁移阻断的效应最显著。结论:SDF-1/CXCR4所介导的MSC迁移与丝裂酶原活化蛋白激酶(MAPK)、磷脂酰肌醇特异性磷脂酶C(PI-PLC)和蛋白激酶C(PKC)等信号途径有关,且PKC途径可能处于中心环节。

The Effect of Human Stromal Derived Factor-1α on the Migration of Bone Marrow Mesenchymal Stem Cells

Objective To explore the signal transduction mechanism of stromal derived factor-1(SDF-1) on migration of mesenchymal stem cell(MSC).Methods MSC culture was performed with the classic adhering method;characteristic of MSC was identified through multipotent differentiation and surface marker assay;by using Boyden chamber in vitro migration assay system,the SDF-1 concentration-dependent manner of MSCs migration and the signal transduction pathways related with MSCs migration were observed.Results Cultured MSCs had the ability of multipotent differentiation and highly expressed CD105 and CD90;the efficiency of MSC migration into the filter increased gradually with the concentration increase of hSDF-1(1 ng/ml,10 ng/ml,100 ng/ml,500 ng/ml),and MSCs number in the filter reached the peak with hSDF-1 in 100 ng/ml.After MSCs intervention with 50M wortmannin,10 uM LY294002,50 uM PD98059,10 uM U73122 and 0.1 mg/ml AMD3100,respectively,MSCs number in the filter all decreased.Importantly,the ability of MSCs migration obviously decreased after U73122,AMD3100 administration.Conlusion SDF-1/CXCR4-mediated MSCs migration is related with MAPK,PI-PLC and PKC signal pathways,PKC signal pathway may play the central role for MSCs migration.