藻酸钙复合自体软骨细胞修复羊膝关节负重区软骨缺损的实验研究

Calcium alginate combined with autologous chondrocytes for repairing of full-thickness defects of femoral weight-bearing articular cartilage in goat

[目的]以中国山羊为动物模型,观察藻酸钙复合自体软骨细胞修复膝关节负重区软骨缺损的可行性。[方法]取羊肩关节软骨,分离、培养软骨细胞,蕃红"O"、 Giemsa及Ⅱ型胶原免疫组织化学染色对其进行鉴定。将自体软骨细胞与藻酸钙凝胶复合,修复山羊股骨髁负重区全层软骨缺损(直径6 mm),实验分为四组:(1)缺损旷置组:缺损内未植入任何组织;(2)骨膜覆盖组:自体骨膜覆盖缺损区;(3)藻酸钙+骨膜组:凝胶植入软骨缺损区,并用自体骨膜覆盖;(4)藻酸钙+细胞+骨膜组:藻酸钙复合自体软骨细胞植入软骨缺损区,自体骨膜覆盖;分别于手术后3、6个月取材,通过大体观察及组织学评分检测修复效果。[结果]软骨细胞复合物蕃红"O"、 Giemsa染色及Ⅱ型胶原免疫组化染色结果均为阳性,将藻酸钙凝胶-软骨细胞复合物用于羊负重区关节面软骨缺损修复,从大体观察和组织学评分进行比较,发现各组均有不同程度的组织修复,藻酸钙+细胞+骨膜组效果最好,与其他组差异有统计学意义(P<0.05)。[结论]藻酸钙凝胶-软骨细胞复合物结合自体骨膜覆盖,可较好修复山羊膝关节负重区软骨缺损。

[Objective] To investigate the feasibility of calcium alginate combined with autologous chondrocytes for repairing of full-thickness defects of femoral weight-bearing articular cartilage in goat. [Methods]Chondrocytes were isolated from China goat shoulder joint, and determined the phenotype with Safranin "O" and Giemsa staining, as well as immunohistochemical staining for collagen type Ⅱ. The full-thickness articular-cartilage defects of 6 mm in diameter were created in the weightbearing area of femoral condyle in goats, which underwent following four treatments: the blank defect without any implant, the defect covered with periosteum, the defect filled with calcium alginate and covered with periosteum, and the defect filled with cell-hydrogel and covered with periosteum. The outcomes of defect repair were assessed by gross and histological observation after 3 and 6 months. [Results]The chondrocytes, isolated and cultured in calcium alginate, preserved the chondrocyte phenotype, with positive staining of Safranin "O" and Giemsa, as well as immunohistochemical staining for collagen type II.The regeneration tissue in the cartilage defects of the animal models consisted of fibrocartilage and hyaline cartilage in each group in different extent. In term of gross and histological scores, the defect filled with cell-hydrogel and covered with periosteum group was statistically superior to the other3 groups(P<0.05). [Conclusion]The calcium alginate-autologous chondrocytes constructs combined with periosteum does improve repairing of the full-thickness cartilage defects in the femoral condyle of goats.