不同来源间充质干细胞成骨分化时序性过程的比较研究

Comparative study of osteogenesis differentiation performance based on time-sequence of mesenchymal stem cells derived from different source

目的:比较骨髓间充质干细胞(BMSCs)和脂肪间充质干细胞(ADSCs)成骨分化时序性特点及其潜能的差异。方法:分离提取同一SD大鼠的BMSCs和ADSCs两种细胞进行培养,通过免疫荧光染色和Von Kossa染色分别观察两种细胞经成骨诱导培养后的成骨分化蛋白I型胶原(Col1α1)的分泌和矿化结节沉积。利用实时荧光定量PCR(RT-q PCR)方法检测两种细胞成骨分化相关基因的表达情况。结果:免疫荧光染色结果显示第7 d时ADSCs的Col1α1表达强度较BMSCs弱,但在14 d和21 d时两者差异并不明显。Von Kossa染色显示21 d时两者均呈现出较好的矿化能力。RT-q PCR结果显示ADSCs成骨相关基因表达水平在早中期(7、14 d)低于BMSCs(P<0.05),而在晚期(21、28 d)无显著性差异(P>0.05)。结论:ADSCs在早期成骨分化水平低于BMSCs,但在晚期接近BMSCs,显示出良好的成骨分化潜能,在骨组织工程中将有广泛的应用前景。

Objective：The purpose of this study was to elucidate the differences of osteogenic potential between bone marrow derived mesenchymal stem cells（BMSCs） and adipose-derived mesenchymal stem cells（ADSCs） by exploring the time course differences of ECM formed in vitro. Method：ADSCs and BMSCs were isolated from the same SD rat and cultured in osteogenic medium for 7,14 and 21 days,followed by immunofluorescence staining of the collagen1α1（Col1α1）.Von Kossa staining was used for observation mineralized nodules. To further explore the difference in organic components,their underlying genotypes,including collagen1α1（Col1α1）,osteopontin（OPN）and osteocalcin（OCN）,were analyzed by RT-qPCR. Result：Immunofluorescence staining showed fluorescence intensity of Col1α1.At the 7th day,ADSCs was weaker than BMSCs,while got close at the 14 th and 21 th day. Von Kossa staining displayed that both ADSCs and BMSCs presented good mineralization ability. RT-qPCR showed that higher expression levels of osteogenic markers including RUNX2,Col1α1,OCN and OPN BMSCs during the initial 7 and 14 days when compared with ADSCs,while no statistic differences were found between the two cells at the 21 th and 28 th day. Conclusion：These results exhibited that osteogenic differentiation level of ADSCs was lower than BMSCs at the early stage,but almost the same at the later stage of cultivation,showed good osteogenetic differentiation potential. Our reach could help in choosing seed cells in bone regenerative medicine.