冻存复苏培养的人鼻中隔软骨细胞生物学特性观察

Biological characteristics of human nasoseptal chondrocytes cultured in vitro after cryopreservation and resuscitation

目的:观察冻存复苏过程对人鼻中隔软骨细胞生物学特性的影响。方法:体外培养人鼻中隔软骨细胞,对冻存1周组、1个月组、3个月组和新鲜制备组的软骨细胞进行倒置相差显微镜动态观察、甲苯胺蓝异染和Ⅱ型胶原的免疫组化染色,并测定35S-Na2SO4掺入量以观察对软骨细胞蛋白多糖合成量的影响,在细胞表型和功能方面进行比较。结果:各组间软骨细胞的生长过程、特殊染色无明显差别,蛋白多糖合成量(35S-Na2SO4掺入量)测定值为(3.2±0.24)×103dpm/μgDNA,统计分析其差异无显著性意义(F=0.6,P>0.05)。各组间细胞存活率差异没有显著性意义(F=2.1,P>0.05)。结论:冻存复苏的人鼻中隔软骨细胞保持了其生物特性,且冻存时间长短对冻存的软骨细胞存活率无显著影响。

AIM:To study the effects of culture in vitro after cryopreservation and resuscitation on the biological characteristics of human nasoseptal chondrocytes. METHODS:The human nasoseptal chondrocytes were cultured in vitro for the dynamic observation in 1 week group,1 month group,3 month group(cryopreservation for 1 week,1 and 3 months respectively) and control group(fresh chondrocytes) through inverted phase contrast microscope.The biological synthesis of the chondrocytes was determined by toluidine blue and type Ⅱ collagen immunohistochemistry reaction.The proteoglycan synthesis efficiencies of cryopreservation on human nasoseptal chondrocytes cultured in vitro were measured by using 35S Na2SO4 incorporation.The comparison was done between cultured chondrocytes and those after cryopreservation in cytomorphology and functions. RESULTS:The growth and special staining of the chondrocyts was insignificantly different among the four groups.There was no significant difference in the efficiency of proteoglycan synthesis(incorporation efficiency of 35S Na2SO4)[(3.2±0.24)×103 dpm/μg DNA](F=0.6,P >0.05).No significant difference was found in the survival rate among all the groups(F=2.1,P >0.05). CONCLUSION:Human nasoseptal chondrocytes after cryopreservation and resuscitation maintain their biological characteristics of chondrocytes.Duration of cryopreservation does not influence the survival rate of chondrocytes.