可注射生物降解型复合淀粉水凝胶修复关节骨-软骨缺损的应用研究

Preparation of an injectable and degradable composite starch hydrogel and its application in osteochondral defect repair

目的开发一种可注射和生物降解型的水凝胶,并研究该水凝胶对软骨细胞增殖和功能表达的影响,以及在大鼠膝关节骨-软骨缺损模型中的修复效果。方法以硅酸钙溶液和淀粉为主要原料加入添加剂后制备复合淀粉水凝胶,通过流变学测试、注射器推出实验和体外浸泡实验对其流变学性能、可注射性和降解性能进行表征。分离提取幼年大鼠膝关节软骨细胞并与材料浸提液进行共培养,通过CCK-8细胞增殖测试和细胞Live/Dead染色方法测定细胞毒性和活性,并通过PCR定量测试成软骨相关基因的表达量。构建大鼠膝关节骨-软骨缺损模型,通过局部注射复合淀粉水凝胶进行干预,在2周、4周和6周取大鼠膝关节标本行宏观、组织学染色评估和分析。结果复合淀粉水凝胶具有介于固态和液态之间的流变学特征,在15 N外力下可从22 G针头中稳定推出,在PBS溶液中浸泡10 d后质量损失率为38wt.%。复合淀粉水凝胶浸提液对软骨细胞增殖无抑制作用,并提高了其Ⅱ型胶原基因和SOX-9基因的表达量。动物实验显示治疗4周和6周后复合淀粉水凝胶组的组织学评分高于未治疗组,组织学染色也显示该水凝胶可促进软骨细胞外基质合成。结论复合淀粉水凝胶具有良好的可注射性、可降解性和生物相容性,可以提升软骨细胞Ⅱ型胶原基因和SOX-9基因的表达量,并可在体内促进骨-软骨缺损修复。

Objective To design an injectable and degradable hydrogel for non-invasive osteochondral defect repair by intra-articular injection.The effect of the hydrogel on the proliferation and function of chondrocytes and the repair of rat knee osteochondral defect were studied.Methods A composite starch hydrogel was prepared by adding calcium silicate and starch as main raw materials.The rheological properties,injectability and degradability were characterized by rheological test,syringe push out test and in vitro immersion test.Chondrocytes were isolated from knee joint of young rats and co-cultured with material extract.CCK-8 cell proliferation test and live/dead staining were used to determine cytotoxicity of material extract,and PCR was used to detect the expression of cartilage formation related genes.Composite starch hydrogel was injected in the rat knee joint with osteochondral defect.The rat knee joint specimens were collected at 2,4 and 6 weeks formacroscopic evaluation and histological staining analysis.Results The composite starch hydrogel had rheological properties between solid state and liquid state and can be injected from 22 G needle under 15 N external force.The mass loss rate was 38%after soaking in PBS solution for 10 days.The composite starch hydrogel had no inhibitory effect on the proliferation of chondrocytes,and promoted the gene expression level of typeⅡcollagen and SOX-9.Animal experiments showed that the macroscopic and histological scores of the composite starch hydrogel group were higher than other groups at 4 and 6 weeks.Histological staining also showed that the hydrogel could promote the formation of cartilage extracellular matrix.Conclusion The composite starch hydrogel is injectable,degradable,and biocompatible,and can increase the secretion of typeⅡcollagen and glycoprotein of the extracellular matrix of cartilage,thereby promoting the repair of articular osteochondral defect in vivo.