【摘要】 目的:观察兔骨髓源成骨细胞与人部分脱钙骨基质构建组织工程骨的可行性及其骨缺损修复作用。方法:实验于2004-03/2005-01在哈尔滨医科大学第一临床医学院动物实验中心完成。①选取日本大耳白兔36只,雌雄不限,随机分成两组,即实验组和对照组各18只,每组再随机分成3小组即4,8和12周组各6只,另选3只日本大耳白兔作为空白组。②将实验组所有动物于胫骨结节处用骨穿针刺入骨髓腔抽取骨髓2.0mL,进行分离、培养和诱导,适时将功能状态良好的兔骨髓源成骨细胞以一定细胞浓度与人部分脱钙骨基质支架材料复合构建组织工程骨,并于复合后24,48和72h用扫描电镜观察细胞在支架材料孔隙内贴附、生长、增殖和功能情况。③所有动物于右侧桡骨中段制作1.5cm骨膜骨缺损模型,实验组骨缺损处植入组织工程骨,对照组骨缺损处植入人部分脱钙骨基质支架。于术后4,8,12周行X射线检查、大体观察、组织学检查,对比研究其成骨和血管化过程及其骨缺损修复能力。结果:①兔骨髓源成骨细胞与人部分脱钙骨基质复合后24h在材料孔隙内部分贴附良好,部分堆积成团;48h基本全部贴壁并有胶原分泌;72h全部贴壁,细胞完全伸展,相邻细胞间有突起相互连接,细胞分布均匀,数量增多,并有大量胶原分泌。②术后12周,实验组X线有明显阻射,骨缺损部位塑型基本完成,骨髓腔已开始再通;对照组支架材料X线阻射进一步增强,两端已愈合,已开始塑型。空白组术后无X射线阻射,无成骨。③实验组和对照组随着时间的推移其成骨量和新生血管数量逐渐增多,实验组4,8,12周组织学血管面积测定值均高于对照组[(2.068±0.216)%比(0.532±0.102)%,(2.976±0.304)%(1.302±0.354)%,(3.708±0.382)%比(2.246±0.402)%,P<0.05]。结论:兔骨髓源成骨细胞与人部分脱钙骨基质生物相容性良好,支架材料为细胞生长提供了良好的三维空间,用兔骨髓源成骨细胞与人部分脱钙骨基质构建组织工程骨是可行的,构建的组织工程骨具有良好的骨修复作用。更多还原

【Abstract】 AIM: To observe feasibility of tissue-engineered bone constructed by bone marrow derived osteoblast of rabbits and demineralized bone matrix (DBM) and bone defect repair. METHODS: The experiment was conducted at the Animal Laboratory Center, First Clinical Medicine College, Harbin Medical University between March 2004 and January 2005. ①Thirty-six Japanese flap-eared rabbits (no matter male or female) were divided into 2 groups randomly: experimental group and control group with 18 rabbits in each group divided randomly into 3 secondary groups with 6 rabbits in each subgroup for 4 weeks group, 8 weeks group and 12 weeks group. Three other rarebits were selected as blank control group. ②2.0 ml bone marrow were extracted at tibial tubercle, isolated, cultured, and induced to cell differentiation. Bone marrow derived from osteoblast grew well and DBM were selected to construct tissue-engineered bone at the right moment functional status. Cell attach, growth and multiplication in bracket pore were observed by scanning electron microscope after 24 hours, 48 hours and 72 hours. ③Animal experimental models were made with periosteal bone defect of right radial intermediate piece for 1.5 cm, and tissue-engineered bone was replanted in the experimental group and cradle of DBM in the control group. The capabilities of bone defect repair, vascularization, and ossification were studied with X-ray, in general observation and histology at 4, 8 and 12 weeks. RESULTS: ①Cells attached to the wall well in bracket pore after 24 hours when bone marrow derived from osteoblast of rabbits and DBM combined. A few of the cells piled up the conglobation; all cells almost attached to the wall and excreted collagen almost after 48 hours; all of the cells attached to the wall after 72 hours and completely expanded, connected by prominence presented in flanking cells, disposited uniformity, increased on number, excreted much collagen. ②After 12-week operation, X-ray was stopped significantly in the experimental group. The part of bone defect was made a statue completely. Medullary cavity of bone had been recanalization. The X-ray was stopped by cradle resistance stronger, and the two sides had been healing, and began to make a statue in the control group. There was no X-ray resistance and no osteoblast in the blank control group after operation. ③Ossification quantity and numbers of new vessels increased gradually along with time changed in the experimental group and control group, but those increased vale in the experimental group at 4, 8 and 12 weeks were higher than that in the control group (2.068±0.216)% vs (0.532±0.102)%,(2.976±0.304)%(1.302±0.354)%,(3.708±0.382)% vs (2.246±0.402)%,P < 0.05. CONCLUSION: There is well biocompatibility between bone marrow derived osteoblast of rabbits and DBM of human. Bracket provides good three-dimension for cell growth. The tissue-engineered bone construction with bone marrow derived osteoblast of rabbits and DBM of human is feasibility and has repaired the bone defect well.更多还原