兔骨髓间充质干细胞复合纳米羟基磷灰石/左旋聚乳酸修复兔软骨缺损的实验研究

Experiemntal Research of Repairing Osteochondral Defects of Rabbit Knees by Using Nano-HA/PLLA Composite with Rabbit Bone Marrow Mesenchymal Stem Cells

目的评价兔骨髓间充质干细胞复合纳米羟基磷灰石/左旋聚乳酸(Nano-HA/PLLA)修复兔膝关节全层软骨缺损的效果。方法体外分离培养兔骨髓间充质干细胞(rBMSCs),取第3代将其与Nano-HA/PLLA在体外构建组织工程软骨(tissue engineering cartilage,TEC)。将18只新西兰大白兔股骨内髁关节面造一直径4.5 mm、深度5 mm的软骨缺损模型,随机分成3组,即实验组、支架组和空白对照组,每组6只,分别予复合细胞的Nano-HA/PLLA、Nano-HA/PLLA及空白填充。术后12、24周取材行大体标本、组织学、免疫组织化学观察及Wakitani组织学评分。结果实验组显示缺损有骨软骨组织形成,软骨下骨基本达到生理整合,组织学及免疫组化显示有大量细胞外基质形成;支架组及对照组显示出有限的再生重建。以上三组12周Wakitani组织学评分分别为5.5±1.401,9.3±1.304和10.2±1.052,24周时为2.2±0.837,7.4±1.144和8.2±1.225,两个时间点实验组均优于支架组及空白组,差异有统计学意义(P<0.05);且实验组24周时优于12周(P<0.05)。结论复合骨髓间充质干细胞的多孔Nano-HA/PLLA能提高成年兔膝关节承重部的骨软骨缺损的修复。

Abstract： Objective To evaluate bone marrow mesenchymal stem cells composite with nano- hydroxyapatite/PLLA in repairing full-thickness cartilage defects of the knees in rabbits. Methods Rabbit bone marrow mesenchymal stem cells （rBMSCs） were cultured in vitro, 3rd generation nano hydroxyapatite/L- polylactic acid （Nano-HA/PLLA） was used in preparing tissue engineering cartilage （tissue engineering carti- lage, TEC）. Diameter of 4.5mm x 5ram cartilage defects in femoral condyle articular surface were made in 18 New Zealand white rabbits. These 18 rabbits were divided into three groups （TEC, Nano-HA/PLLA, and blank group）. Gross histology, immunohistochemistry, biomechanics observation and semi-quantitative improved Wa- kitani score were studied after 12 weeks and 24 weeks, respectively. Results In TEC group,bone and carti- lage tissue formation was found in defect site , subchondral bone basically reached physiological integration,histology and immunohistochemistry showed the extracellular matrix formation increased. There was significant difference in comparing with the Nano-HA/PLLA group, or the blank group （P 〈 0.05 ）. Moreover, there was a positive correlation between the effect of cartilage repaired and time （P 〈 0.05）. Conclusion The porous composite bone marrow mesenchymal stem cells Nano-HA/PLLA can improve cartilage regeneration in adult rabbits after cartilage defects.