多孔磷酸钙/骨基质明胶复合骨水泥修复兔腰椎骨缺损的实验研究

Experiment of porous calcium phosphate/bone matrix gelatin composite cement for repairing lumbar vertebral bone defect in rabbit

目的探讨多孔磷酸钙/骨基质明胶复合骨水泥(以下简称多孔复合骨水泥)修复兔腰椎骨缺损的效果。方法采用Urist等的方法制备成年新西兰兔骨基质明胶(bone matrix gelatin,BMG),参考W/O/W复乳法制作聚乳酸-羟基乙酸共聚物[poly(lactic-co-glycolic)acid,PLGA]空白微球,将PLGA空白微球和BMG与磷酸钙骨水泥(calcium phosphate cement,CPC)复合,构建多孔复合骨水泥。通过体外抗溃散观察、孔隙率测定及生物力学试验,观察其理化特性;并以CPC作对照。取30只2月龄新西兰兔,制备L_3椎体大小为4 mm×3 mm×3 mm的骨缺损后,分别填入多孔复合骨水泥(实验组,n=15)和CPC(对照组,n=15)。术后第4、8、12周,X线片观察骨融合情况,micro-CT分析骨密度(bone mineral density,BMD)、骨体积分数(bone volume fraction,BVF)、骨小梁厚度(trabecular thickness,Tb.Th.)、骨小梁数量(trabecular number,Tb.N.)和骨小梁分离度(trabecular spacing,Tb.Sp.),组织学观察新生骨形成情况。结果两种骨水泥均有较好抗溃散性,多孔复合骨水泥孔隙率为55.06%±1.18%,抗压强度为(51.63±6.73)MPa,与CPC的孔隙率(49.38%±1.75%)以及抗压强度(63.34±3.27)MPa相比,差异有统计学意义(t=4.254,P=0.006;t=2.476,P=0.034)。X线片观察示,随时间延长,两组材料与宿主之间的边界逐步模糊,且实验组第12周时边界已消失、对照组仍可见。micro-CT检测各时间点实验组BMD、BVF、Tb.Th.、Tb.N.以及Tb.Sp.与对照组相比,差异均有统计学意义(P<0.05)。组织学观察示,随时间延长,两组植入材料中均见新骨生长,其中实验组新生骨与宿主骨形成骨性连接,优于对照组。结论多孔复合骨水泥具有骨诱导活性和骨传导双重作用,可以有效促进兔腰椎骨缺损修复重建。

Objective To investigate the effect of a porous calcium phosphate/bone matrix gelatin （BMG） composite cement （hereinafter referred to as the ＂porous composite cement＂） for repairing lumbar vertebral bone defect in a rabbit model. Methods BMG was extracted from adult New Zealand rabbits according to the Urist＇s method. Poly （lactic-co-glycolic） acid （PLGA） microsphere was prepared by W/O/W double emulsion method. The porous composite cement was developed by using calcium phosphate cement （CPC） composited with BMG and PLGA microsphere. The physicochemical characterizations of the porous composite cement were assessed by anti-washout property, porosity, and biomechanical experiment, also compared with the CPC. Thirty 2-month-old New Zealand rabbits were used to construct vertebral bone defect at L3 in size of 4 mmx3 mmx3 mm. Then, the bone defect was repaired with porous composite cement （experimental group, n=15） or CPC （control group, n=15）. At 4, 8, and 12 weeks after implantation, each bone specimen was assessed by X-ray films for bone fusion, micro-CT for bone mineral density-（BMD）, bone volume fraction （BVF）, trabecular thickness （Tb. Th.）, trabecular number （Tb.N.）, and trabecular spacing （Tb. Sp.）, and histological section with toluidine blue staining for new-born bone formation. Results The study demonstrated well anti-washout property in 2 groups. The porous composite cement has 55.06%±1.18% of porosity and （51.63±6.73） MPa of compressive strength. The CPC has 49.38%±1.75% of porosity and （63.34±3.27） MPa of compressive strength. There were significant differences in porosity and compressive strength between different cements （t=4.254, P=0.006; t=2.476, P=0.034）. X-ray films revealed that the zone between the cement and host bone gradually blurred with the time extending. At 12 weeks after implantation, the zone was disappeared in the experimental group, but clear in the control group. There were significant differences in BMD, BVF, Tb. Th., Tb. N., and Tb. Sp. between 2 groups at each time point （P〈0.05）. Histological observation revealed that there was new-born bone in the cement with the time extending in 2 groups. Among them, bony connection was observed between the new-born bone and the host in the experimental group, which was prior to the control group. Conclusion The porous composite cement has dual bioactivity of osteoinductivity and osteoconductivity, which are effective to promote bone defect healing and reconstruction.