骨形态发生蛋白2和富血小板血浆凝胶对兔骨髓间充质干细胞体外成软骨分化的影响

Effects of BMP-2 and platelet-rich plasma gel on chondrogenic differentiation of rabbit BMSCs in vitro

目的观察富血小板血浆(PRP)凝胶及腺病毒介导的骨形态发生蛋白2(BMP-2)基因转染对兔骨髓间充质干细胞(BMSCs)体外成软骨分化的影响。方法取新西兰大白兔骨髓培养BMSCs,抽取静脉血制备PRP,以Ad-GFP-hBMP-2(BMP-2-BMSCs组)及Ad-GFP(对照组)转染的BMSCs分别与PRP凝胶复合,继续培养。用扫描电镜、MTT检测PRP凝胶的生物相容性,RT-PCR检测各组的Ⅰ型胶原、Ⅱ型胶原、Ⅹ型胶原、蛋白聚糖、SOX-9表达。结果扫描电镜及MTT检测证实PRP凝胶具有良好的生物相容性。RT-PCR分析表明,单层培养的BMP-2-BMSCs组的Ⅱ型胶原、蛋白聚糖和SOX-9的表达水平均高于对照组(P<0.05),但Ⅰ型胶原表达水平较对照组明显下降(P<0.05),同时Ⅹ型胶原表达水平与对照组相比差异无统计学意义(P>0.05);与PRP复合后,BMP-2-BMSCs组的Ⅱ型胶原、蛋白聚糖和SOX-9表达水平较对照组升高(P<0.05),而Ⅰ型胶原和Ⅹ型胶原表达水平与对照组比较差异无统计学意义(P>0.05)。结论转染BMP-2的BMSCs在PRP凝胶中生长良好,BMP-2基因转染能促进BMSCs的体外成软骨分化。

Objective To observe effects of platelet-rich plasma(PRP) gel and gene transfection of adenovirus-induced rabbit bone morphogenetic protein-2(BMP-2) on chondrogenic differentiation of bone marrow mesenchymal stem cells(BMSCs). Methods BMSCs were isolated and cultured from the bone marrow of New Zealand rabbits and PRP was acquired from the venous blood. The Ad-GFP-BMP-2(BMP-2-BMSCs group) and Ad-GFP(control group) transduced BMSCs were mixed with PRP. The scanning electron microscopy and MTT test were used to evaluate the biocompatibility of PRP. In addition,the RT-PCR was used to test the expressions of collagen Ⅰ,collagen Ⅱ,collagen Ⅹ,aggrecan and SOX-9. Results The scanning electron microscopy and MTT test indicated that PRP presented good biocompatibility. Furthermore,the RT-PCR demonstrated that the expressions of collagenⅡ,aggrecan and SOX-9 in monolayer cultured BMP-2-BMSCs group were significantly up-regulated compared with control group(P < 0.05). However,expression of collagen Ⅰ was significantly lower in the former group than in the latter group(P < 0.05). No difference in expression of collagen Ⅹ was found between the 2 groups(P > 0.05). After mixed with PRP,expressions of collagenⅡ,aggrecan and SOX-9 in BMP-2-BMSCs group were significantly higher than those in control group(P < 0.05). No statistical difference in collagenⅠ and collagen Ⅹ was found between the BMP-2 transfected group and control group(P > 0.05). Conclusion The BMP-2 transfected BMSCs could grow well in PRP,and Ad-GFP-hBMP-2 transfection can effectively facilitate the chondrogenic differentiation of BMSCs in vitro.