可注射生物活性玻璃支架修复兔膝关节软骨缺损

Repair of rabbit articular cartilage defect by a novel injectable bioactive glass scaffoldObjective[Key Words]

目的通过比较不同方法对兔膝关节软骨缺损的修复效果,探讨一种新型生物活性玻璃支架对关节软骨的修复能力。方法应用成分为10.8%P2O5-54.2%SiO2-35%CaO(PSC)的生物活性玻璃粉末、壳聚糖溶液(chitosan solution,CS)以及半水硫酸钙(calcium sulfate hemihydrate,CSH)制得PSC/CS骨水泥,经模具定型得到支架材料。将18只新西兰大白兔随机分为对照组、生物活性玻璃(bioactiveglass, BG)组及加入TGF-β1因子的TGF-β1组。制作双膝关节软骨缺损模型,对照组不予任何处理,BG组和TGF-β1组分别放置两种支架材料。于术后6周及12周分别取材,进行大体观察、HE染色、甲苯胺蓝染色、番红O染色及Ⅱ型胶原免疫组化等的对比观察。依照国际软骨修复协会(International Cartilage Repair Society,ICRS)评分及Wakitani评分定量评价软骨修复的效果。结果术后6周,对照组与BG组的大体观察无明显差异,而TGF-β1组的大体观察优于其他两组,并且其ICRS评分(4.67±0.52)分优于BG组(2.83±0.75)分,差异有统计学意义(t=-2.817,P=0.015);组织学染色比较,三组间均差异无统计学意义;对照组Wakitani评分(13.67±0.52)分,BG组(13.83±0.41)分,TGF-β1组(13.33±1.03)分的差异均无统计学意义。术后12周对照组与BG组的大体观察依然无明显差异,而TGF-β1组的大体观察显著优于其他两组,ICRS评分(9.01±0.63)分也明显较高(对照组 vs TGF-β1组: t=-2.289,P=0.022;BG组 vs TGF-β1组:t=-2.326,P=0.020);组织学染色结果TGF-β1组的阳性染色程度同样较高,Wakitani评分(6.50±1.38)分显著低于对照组(9.83±1.33)分和BG组(9.51±1.05)分TGF-β1组,差异均有统计学意义(对照组 vs TGF-β1组:t=-2.771,P=0.007;BG组 vs TGF-β1组:t=-2.756,P=0.006),并且新生组织的染色程度与Ⅱ胶原表达程度与正常软骨相近。结论单纯PSC/CS生物活性玻璃支架不具有良好的修复软骨缺损能力,而添加TGF-β1的PSC/CS支架可促进膝关节软骨缺损的修复,新生组织接近正常软骨。

Objective By comparing different repairing effects of different methods on articular cartilage defects in rabbit model, the ability of a new bioactive glass scaffold to repair cartilage defect was studied. Methods We prepared the PSC/CS bone cement by mixing the bioactive glass (BG) powder composed of 10.8%P2O5-54.2%SiO2-35%CaO (PSC), chitosan solution (CS) and calcium sulfate hemihydrate (CSH), then the bone cement was sized by a mold to form cylindrical scaffold. In vivo experiment, 18 male rabbits were divided into three groups randomly, including blank group, BG group and TGF-β1 group, which was added TGF-β1 into BG scaffold. Both knees of each rabbit were made cartilage defect for the same group, and no intervention was applied in B group, then implanted scaffolds into defects in both experimental groups. At 6 and 12 weeks after surgery, observed the macroscopic growth, histologic staining and collagen II immunohistochemistry (IHC), and the International Cartilage Repair Society (ICRS) and Wakitani score were used to analyze the experimental results quantitatively. Results At 6 weeks after surgery, there was no obvious difference between blank group and BG groups, but the macroscopic result of TGF-β1 group was better than the other two groups and its ICRS score 4.67±0.52 points was statistically higher than BG group 2.83±0.75 points (t=-2.817, P=0.015). As to the comparison of histologic staining and Wakitani socre among three groups, no statistical difference was observed (blank group=13.67±0.52, BG group=13.83±0.41,TGF-β1 group=13.33±1.03). At 12 weeks after surgery, there was still no obvious difference between blank and BG groups, while the results of macroscopic observation and ICRS score in TGF-β1 group were significantly higher than them 9.01±0.63 points (blank group vs TGF-β1 group: t=-2.289, P=0.022;BG group vs TGF-β1 group: t=-2.326, P=0.020). More importantly, much deeper positive staining were observed in TGF-β1 group, and the Wakitani score was higher than the other two groups(blank group=9.83±1.33, BG group=9.51±1.05,TGF-β1 group=6.50±1.38, blank group vs TGF-β1 group: t=-2.771, P=0.007;BG group vs TGF-β1 group: t=-2.756, P=0.006). By comparing the degree of histologic staining and Col II expression with normal cartilage, the regenerated tissue in TGF-β1 group was similar. Conclusion Single PSC/CS scaffold doesn’t possess excellent ability to repair cartilage defect. When TGF-β1 was added into PSC/CS bioactive glass, the scaffold was able to promote cartilage defect repair, and the regenerated tissue was similar to normal cartilage.