不同方法低温保存的组织工程骨修复骨缺损的实验研究

Repairing bone defects with tissue engineered bone cryopreserved with or without cryoprotectant

目的探讨不同方法低温保存的组织工程骨修复节段性骨缺损的差异。方法以骨髓基质干细胞（BMSCs）复合部分脱蛋白骨培养制备组织工程骨，组织工程骨分别用添加或不添加冻存保护剂的保存液于-196℃的液氮中冻存3个月，植入实验前复温冻存的组织工程骨。选择60只新西兰白兔，制作15mm长的桡骨节段性骨缺损模型，根据骨缺损区植入不同的材料分为A、B、C和D组，每组15只，A组：植入添加冻存剂保存的组织工程骨；B组：植入未添加冻存剂保存的组织工程骨；C组：植入未行低温保存的组织工程骨；D组：植入部分脱蛋白骨。术后4、8、16周取材，行X线检查、组织学观察、计算机图像分析和生物力学测定。结果术后4、8、16周各组骨缺损区均有新骨生成，成骨量随时间的推移而增加。经X线、组织学和生物力学评估，A组与C组比较差异无统计学意义（P〉0．05），A组或C组分别与B、D组比较，成骨能力依次为：A组或C组〉B组〉D组，差异有统计学意义（P〈0．05），其中A组或C组术后16周骨缺损完全修复，骨髓腔再通，生物力学性能接近正常骨。结论选择适宜的冻存保护剂对组织工程骨的生物活性有一定的保护作用。

Objective To study the differences of tissue engineered bone eryopreserved with or without cryoprotectant in repair of segmental bone defects. Methods BMSCs were cocuhured with partially deproteinized bone to produce tissue engineered bone. After the bone was stored in liquid nitrogen at - 196 ℃ with or without cryoprotectant for 3 months, it was thawed. Experimental models of 15 mm radial segmental defect were created in 60 New Zealand white rabbits which were divided into 4 groups according to transplant materials. The defects were repaired with tissue engineered bone cryopreserved with cryoprotectant in Group A, without cryoprotectant in Group B, with tissue engineered bone without cryopreservation in Group C, and with partially deproteinized bone in Group D. The samples were harvested at 4, 8, 16 weeks postoperatively. A series of examinations were carried out to observe the repair effects, including the roentgenography, histomorphology, biomechanics and computerized graphical analysis. Results New bone formation, increasing with time, was observed at 4, 8, 16 weeks postoperatively in all the defects treated with implants. There was no difference between Groups A and C（ P 〉 0. 05）, according to radiologieal, histomorphologieal and biomechanical evaluations. The comparison study showed that capability of new bone formation in the 4 groups was ranked as follows： A or C 〉 B 〉 D（ P 〈 0. 05） . At 16 weeks the defects in Groups A and C were bridged with marrow cavity, and the biomechanical property of implants approximated to that of normal bone. Conclusion Proper cryoprotectant may optimize the bioactivity of tissue engineered bone.