低温冻存同种异体软骨细胞在喉功能重建中的应用

Application of cryogenic allogeneic chondrocytes in the reconstruction of laryng eal function

目的:通过对深低温冻存的软骨细胞作为种子细胞构建组织工程化软骨修复喉软骨缺损的能力研究,寻找一种活性软骨细胞的保存方法。方法:取3周龄新西兰兔关节软骨,经深低温冻存,复苏冻存不同时间的软骨细胞,体外培养,培养细胞呈单层细胞铺满培养瓶底后收集细胞,制成细胞悬液,接种于聚羟基乙酸(polyglycolicacid,PGA)三维支架材料上,复合物体外培养1周,接种于同种异体兔甲状软骨缺损处,术后2,4,8周取材,行大体及组织学观察。结果:经深低温冻存的软骨细胞与新鲜软骨细胞修复区愈合良好,无瘢痕及坏死现象,组织学观察均有软骨生成及基质分泌;冻存不同时间的软骨细胞组之间无明显差异。结论:经深低温冻存的软骨细胞保持了分裂增殖及合成基质的能力,可用于组织工程修复软骨缺损,冻存时间对其功能无明显的影响。

AIM:To study the ability of cryogenically frozen chondrocytes as seeding cells to construct tissue engineered chondrocytes and repair the defect of laryngeal cartilages so as to find a preserving method for active chondrocytes. METHODS:The articular cartilages of New Zealand rabbits aged three weeks were taken,and cryogenically frozen.The chondrocytes which frozen at different time w ere thawed and cultured in vitro,the cells were collected after the cultured cel ls were put everywhere on the bottom of the culture bottle as monolayer cells,an d they were made into cell suspension which was inoculated to three dimensional polyglycolicacid(PGA)scaffolds,and the compound was cultured in vitro for one w eek,and were inoculated to the injured area of thyroid cartilage in allogeneic r abbits.The material was taken at 2,4 and 8 weeks after operation for the gross a nd histological observations. RESULTS:The repaired areas of cryogenically frozen chondrocytes and fresh chon drocytes were of good healing,and no symptoms of scar and necrosis were found.Th e cartilage and excretion of matrix were observed histologically.There was no ob vious difference among the chondrocytes frozen at different time. CONCLUSION:The cryogenically frozen chondrocytes has the ability of split,prol iferation and synthesis of matrix.It can be used in tissue engineering for repai ring the defect of cartilage and the frozen time has no obvious effect on its fu nction.