生物衍生骨复合成骨诱导骨髓间充质干细胞构建组织工程化骨修复兔桡骨-骨膜缺损的血管化进程

Vascularization of tissue engineered bone constructed by bio-derived bone co-cultured with induced bone marrow mesenchymal stem cells to repair segmental defects of rabbit radius-periosteum

背景:移植骨能否血管化是移植成活的关键。目的:验证生物衍生骨复合骨髓间充质干细胞构建组织化工程骨修复兔桡骨-骨膜缺损的血管化进程。设计、时间及地点:随机分组设计的动物对照实验,2007-01/2008-01在南华大学生命科学院实验室完成。材料:健康成年新西兰大白兔25只,双侧桡骨制备成中段1.5cm的桡骨-骨膜缺损。方法:经脱脂、脱蛋白、部分脱钙和冻干等一系列的物理、化学方法处理后制备生物衍生骨支架材料。与自体骨髓间充质干细胞复合制备组织工程骨支架材料。按随机数字表法,20只兔右侧桡骨为实验组植入组织工程骨,左侧为对照组植入单纯生物衍生骨,不做内外固定;另5只兔双侧造成骨缺损后不植入任何材料作为空白组。主要观察指标:植入后2,4,8,12周观察组织工程骨、生物衍生骨与断端的愈合情况;组织学观察骨组织中血管形成情况,并进行血管面积图像分析。结果:25只兔无脱落。大体观察显示实验组植入后8周两断端已融合为一体;植入后12周,已经塑型,外观接近正常。对照组各时间点愈合程度不如实验组。组织学观察显示,实验组和对照组植入后4周均有大量血管生成,植入后8周,实验组修复区的血管网较对照组开始排列规律;植入后12周,实验组血管已达成熟阶段,血管排列方向比对照组均匀、有序,形态结构接近正常皮质骨的血管形态。血管面积图像法测定的血管面积结果表明,植入后2周实验组和对照组血管面积低于正常骨组织,植入后4,8周实验组和对照组血管面积高于正常骨组织,差异均有显著性意义(P<0.05)。植入后4周实验组血管面积低于对照组,差异有显著性意义(P<0.05)。结论:生物衍生骨复合成骨诱导的骨髓间充质干细胞可加快骨修复后的血管化进程。

BACKGROUND： It is a key for the survival of a transplanted bone to vascularize. OBJECTIVE： To study the vascularization of repairing segmental defects of rabbit radius-periosteum by tissue engineering bone constructed by the co-culture of bio-derived bone with induced bone marrow mesenchymal stem cells （BMSCs）. DESIGN, TIME AND SETTING： Randomization design animal controlled trials were performed at the laboratory of Life Science College of Nanhua University from January 2007 to January 2008. MATERIALS： Twenty-five healthy adult New Zealand rabbits were used in this study to establish a 15-mm length of radius-periosteum defects on both sides of radial bone. METHODS： Bio-derived bone scaffold was prepared with a serial of physical and chemical processes, including degreasing, deproteinization, partial decalcification and freeze drying. Bio-derived bone scaffold was then co-cultured with induced BMSCs to construct tissue engineering bones, which were implanted into the segmental bone defects of 20 rabbits in right radius, taken as the experimental group; Bio-derived bone scaffold was implanted into the left radius, serving as the controls. No implant was given into the other 5 rabbits as the blank group. MAIN OUTCOME MEASURES： The healing on the broken ends of fractured bone was observed after tissue engineering bone and bio-derived bone were implanted for 2, 4, 8 and 12 weeks. Histological examination was performed to observe the angiopoiesis after operation, we also did the image analysis of blood vessels. RESULTS： All the 25 rabbits come to result analysis. The broken ends of fractured bone healed at 8 weeks postoperation in the experimental group, and there were moldings at 12 weeks, the appearance was close to the normal. In the control group, the healing was not as good as that of the experimental group at every time. Histological examination showed that, a lot of blood vessels were observed in both experimental and control groups at 4 weeks postoperation. In the experimental, group, the vascular net began to arrange regularly at 8 weeks, and the blood vessels were mature at 12 weeks, with the orientating of blood vessels more well-distributed and well-ordered than that of control group, the shape and structure approached normal blood vessel in cortex bone. Blood vessel image analysis showed that, the areas of blood vessel in both experimental and control groups were significantly less at 2 weeks after operation, but larger than the normal one at 4 and 8 weeks （P 〈 0.05）. At 4 weeks posttransplant, the area of blood vessels in the experimental group was significantly lower than that in the control group （P 〈 0.05）. CONCLUSION： The tissue engineering bone constructed by bio-derived bone co-cultured with induced BMSCs has good vascularization in repairing segmental defects of rabbit radial bone.