TWEAK诱导骨髓间充质干细胞增殖作用的初步实验研究

Studies of tumor necrosis factor-like weak inducer of apoptosis labeling on the proliferation of rat bone marrow mesenchymal stem cells

目的探讨肿瘤坏死因子样弱凋亡因子(tumor necrosis factor-like weak inducer of apoptosis,TWEAK)基因转染骨髓间充质干细胞(bone mesenchymal stem cells,BMSCs)后对其增殖功能的影响。方法全骨髓贴壁法筛选培养幼年大鼠BMSCs,用腺病毒载体Ad5CMV-TWEAK转染TWEAK基因,通过逆转录-聚合酶链式扩增实验(RT-PCR)观察大鼠BMSCs细胞转染后TWEAK蛋白的表达情况。实验分成3组,即转染Ad5CMV-TWEAK的实验组,转染Ad5CMV-EGFP的对照组及未转染任何基因的空白对照组。用MTT法比较各组BMSCs细胞的增殖变化情况。结果全骨髓贴壁培养法能有效分离纯化大鼠BMSCs,RT-PCR可证明Ad5CMV-TWEAK成功的转染BMSCs细胞,转染后在BMSCs产生较高的表达。MTT值经χ2检验,TWEAK组与无干预组比较差异显著(P<0.05),而EGFP组与无干预组无显著差异(P>0.05)。结论TWEAK转染有刺激BMSCs的增殖作用,如进一步证明其促分化作用,其在骨修复材料的改进以及种植体涂层修饰方面将有广阔的应用前景。

Objective To approach the effect of tumor necrosis factor-like weak inducer of apoptosis （TWEAK） gene on the proliferation of rat bone mesenchymal stem cell （BMSC）. Methods The rat BMSCs were isolated and purified from bone marrow of young rats by adherent culture. TWEAK genes were transfected by AdSCMV-TWEAK, and RT-PCR was used to observe the expression of TWEAK. Then the cells were divided into 3 groups, transfected AdSCMV-TWEAK group, transfected AdSCMV- EGFP group and untransfeeted group, and the MTT assay was used to test the proliferation effect. Results The purified BMSCs were obtained by adherent culture. RT-PCR results showed high TWEAK expression after AdSCMV-TWEAK transfected. The MTF assay results demonstrated significant difference between AdSCMV-TWEAK group and untransfected group （P 〈 0.05 ） but no differencebetween the transfected AdSCMV-EGFP group and untransfected group （ P 〉 0. 05 ）. Conclusion Transfecting TWEAK gene can stimulate BMSCs proliferation, the next research will test if it can promote the differentiation of the cells to show application, prospect in bone restore material and implant coating.