昆明犬胎儿成纤维细胞系的建立及生物学特性分析

本研究旨在探索和建立昆明犬胎儿成纤维细胞体外分离、培养的技术方法并观察其生物学特性.试验利用剖腹手术法获得发育30 d的昆明犬胎儿12个,性别鉴定结果为7公5母.公、母胎儿体尺和体重分别为:体长1.90 cm±0.14 cm和2.00 cm±0.05 cm,体宽0.90 cm±0.13 cm和0.90 cm±0.15 cm;体重0.68 g±0.17 g和0.66 g±0.06 g,差异均不显著(P>0.05).以昆明犬胎儿组织为材料,采用胶原酶消化法和细胞冷冻保存技术建立了昆明犬胎儿成纤维细胞系,并对所构建的细胞系进行生物学特性研究.结果表明,分离的胎儿成纤维细胞呈典型的成纤维细胞形态,生长曲线呈S型,倍增时间约为36 h;冻存前和复苏后的存活率分别为96.1%和94.6%;染色体核型分析显示2n=78(XY),并在体外培养多代后仍能保持正常核型.本研究建立的昆明犬胎儿成纤维细胞系可在体外快速贴壁生长、增殖,进行稳定培养,这不仅使昆明犬遗传资源在细胞水平得以保存,且所获成纤维细胞能作为相关研究中所需细胞的来源.

Establishment and Biological Characteristics Analysis of Fetal Fibroblast Cell Lines in Kunming Dog

The objective of this study was to explore the method of isolation and cultivation of fibroblasts of Kunming dog in vitro. 12 fetus at 30th day in gestation of Kunming dog were obtained by laparotomy and the sex indentification of fetus were 7 males and 5 females. The body size and weight of fetus were measured. The results showed that male and female fetus of body length were 1.90 cm ± 0.14 cm and 2.00 cm ± 0.05 cm, body width were 0.90 cm ± 0.13 cm and 0.90 cm ± 0.15 cm, body weight were 0.68 g ± 0.17 g and 0.66 g ± 0.06 g, the difference was not significant (P>0.05). Additionally, the fetal fibroblast cell lines were successfully established by the method of collagenase digestion and cryopreservation. The research were designed to examine the biological characteristics of fibroblasts. The cells showed typical morphologic characteristics of fibrous and long spindle appearance. Analysis of the growth of the fibroblasts revealed a sigmoid curve with the population doubling time (PDT) of 36 h. Moreover, the average viability of the fetal fibroblasts was 96.1% before freezing and 94.6% after thawing. Karyotyping indicated the chromosome number of Kunming dog was 2n=78 (XY). In conclusion, the fetal fibroblast cell lines in Kunming dog were successfully established in vitro. It will not only preserve the genetic resources of Kunming dog at the cellular level but also provide valuable materials for further research.