应用富血小板血浆和骨髓基质细胞构建细胞-膜片及其异位成骨的实验研究  

Cell-sheet constructed by combined marrow stromal cells and platelet-rich plasma and its ectopic bone formation

作　　者：张森林[1] 毛天球[2] 

机构地区：[1]南京军区南京总医院口腔科,江苏南京210002 [2]第四军医大学口腔医学院口腔颌面外科,陕西西安710032

出　　处：《临床口腔医学杂志》2009年第3期147-150,共4页Journal of Clinical Stomatology

基　　金：南京军区医学科学技术研究"十一五"计划课题(06MA128);南京军区医学科技创新资助课题(07M087);南京军区南京总医院重点科研课题(Z2008044)

摘　　要：目的:探讨应用富血小板血浆(platelet rich plasma,PRP)和骨髓基质细胞(marrow stromal cells,MSCs)构建细胞-膜片的方法并评价膜片包裹珊瑚支架后的异位成骨能力。方法:将体外培养扩增的兔MSCs悬液与PRP混合,滴加到6孔培养板上,再滴加牛凝血酶溶液,形成凝胶样MSCs/PRP混合物,置于孵箱内培养。第1周用DMEM完全培养液,后2周改用DMEM诱导培养液。用得到的细胞-膜片包裹珊瑚圆片后,植入裸鼠背部皮下,以单纯珊瑚植入作对照。术后4周和8周取材,通过组织学观察,评价其异位成骨情况。结果:细胞-膜片具有一定的韧性和可操作性。扫描电镜观察显示:膜片由大量的纺锤样成骨细胞有规律地密集在一起而形成。组织学观察显示:术后4周和8周,膜-支架复合体珊瑚支架的表面及其孔洞内有大量的软骨或骨形成,以软骨内成骨的方式成骨;单纯珊瑚植入组,珊瑚表面及其孔洞内有大量纤维结缔组织长入,未见软骨或骨形成。结论:用PRP和MSCs构建细胞-膜片的方法简便,膜片包裹珊瑚支架形成的膜-支架复合体具有良好异位成骨能力。Objective： To investigate method of growing cell-sheet by combined marrow stromal cells （MSCs） and platelet rich plasma （PRP） and its ectopic bone formation. Method： A suspension of 5 ×10^6 MSCs in 500 μl medium were dissolved in 500 μl of PRP. The mixture of MSCs and PRP was plated in one well of 6-well plate and cultured in a humidi- fied incubator for 3 weeks. The culture media was standard DMEM for first week and then was changed to osteogenic DMEM for next two weeks. Cell-sheet was formed and wrapped around the coral scaffold,and the sheet-scaffold construct was implanted into the dorsal subcutaneous area of athymic mice. Coral scaffolds acted as control. The ectopic bone forma- tion was investigated by histological observation 4 and 8 weeks after operation. Result： The sheet comprising multilayered spindle-like osteoblasts possessed good mechanical properties. New bone and/or cartilage formation could be observed both inside and outside the scaffold in specimens from the sheet-scaffold construct group in ectopic sites 4 and 8 weeks after im- plantation, and osteogenesis followed the pattern of endochondral bone formation. In contrast, no bone or cartilage formation could be observed in coral group. Conclusion： The method of growing cell-sheet by combined MSCs and PRP was convenient and simple, and bone graft could be engineered through combination of the sheet and coral scaffold.

关 键 词：骨组织工程 富血小板血浆 骨髓基质细胞 珊瑚 细胞-膜片技术 

分 类 号：R318.08[医药卫生—生物医学工程]