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目的探讨基质细胞衍生因子-1α(SDF-1α)受体CXCR4、CXCR7在骨髓间充质干细胞(BMSCs)中蛋白和mRNA的表达;及SDF-1α/CXCR4/CXCR7轴对BMSCs迁移作用的可能机制。方法体外培养大鼠BMSCs,流式细胞术鉴定细胞表面抗原CD29、CD44和CD34。应用CXCR4特异性拮抗剂AMD3100及CXCR7中和抗体分别阻断CXCR4及CXCR7,通过Western blotting和RT-PCR分别检测BMSCs蛋白和mRNA的表达变化;Transwell法检测细胞迁移能力。本次实验分为单纯BMSCs组(A)、AMD3100预处理BMSCs组(B)、CXCR7中和抗体预处理BMSCs组(C)及AMD3100+CXCR7中和抗体预处理BMSCs组(D)。结果经鉴定第3代大鼠BMSCs中CD29和CD44均呈阳性表达,而CD34表达阴性。BMSCs中CXCR4、CXCR7蛋白和mRNA均有表达。与A组相比,B组及D组CXCR4及CXCR7蛋白表达明显受到抑制(P<0.05),C组只有CXCR7蛋白表达降低(P<0.05);各组CXCR4 mRNA和CXCR7 mRNA的表达差异均无显著性。SDF-1α可以诱导BMSCs迁移,与0μg/L组相比,10μg/L组和100μg/L组穿膜细胞数均显著增多(P<0.01),与10μg/L组相比,100μg/L组穿膜细胞数亦明显增多(P<0.01);AMD3100和CXCR7中和抗体均能抑制BMSCs的迁移作用(P<0.05),当两者同时作用时,抑制效应更为显著(P<0.05)。结论 BMSCs共表达CXCR4、CXCR7蛋白及mRNA;BMSCs的迁移具有SDF-1α浓度依赖性;SDF-1α/CXCR4/CXCR7轴介导BMSCs的迁移作用,CXCR4受体和CXCR7受体对BMSCs的迁移可能具有协同促进作用。
Objective To investigate the expression of protein and mRNA of stromal cell-derived factor-1α (SDF-1α) CXCR4 and CXCR7 in bone marrow mesenchymal stem cells (BMSCs) and the possible mechanism of SDF-1α / CXCR4 / CXCR7 axis migration on BMSCs . Methods Rat BMSCs were cultured in vitro, and cell surface antigen CD29, CD44 and CD34 were identified by flow cytometry. The CXCR4-specific antagonist AMD3100 and CXCR7 neutralizing antibody were used to block CXCR4 and CXCR7 respectively. Western blotting and RT-PCR were used to detect the protein and mRNA expression of BMSCs respectively. Transwell assay was used to detect cell migration. This experiment was divided into BMSCs group (A), BMSCs group (B), AMD3100 pretreatment group (B), CXCR7 neutralizing antibody pretreatment BMSCs group (C) and AMD3100 + CXCR7 neutralizing antibody pretreatment BMSCs group (D). Results The third generation of BMSCs were identified as positive for CD29 and CD44 but negative for CD34. BMSCs CXCR4, CXCR7 protein and mRNA are expressed. Compared with group A, the expressions of CXCR4 and CXCR7 in group B and group D were significantly inhibited (P <0.05), and only CXCR7 protein in group C was decreased (P <0.05). There was no difference in the expression of CXCR4 and CXCR7 between group B and group D Significance. SDF-1α could induce the migration of BMSCs. Compared with 0μg / L group, the number of transmembrane cells in 10μg / L group and 100μg / L group were significantly increased (P <0.01). Compared with 10μg / L group, (P <0.01). Neutralizing antibody of both AMD3100 and CXCR7 inhibited the migration of BMSCs (P <0.05). When both of them were treated simultaneously, the inhibitory effect was more significant (P <0.05). CONCLUSIONS: BMSCs co-express CXCR4, CXCR7 protein and mRNA. The migration of BMSCs is dependent on SDF-1α concentration. SDF-1α / CXCR4 / CXCR7 axis mediates the migration of BMSCs. Migration of CXCR4 and CXCR7 receptors may have Synergistic effect.