摘要
背景:目前常用的动物肌腱研究模型(比如兔,罗曼鸡等)的培养方法存在周期长,细胞获取量少等不足,往往成为制约肌腱组织工程实验研究进程的瓶颈。目的:希望建立SD大鼠鼠尾肌腱细胞培养流程,以期用较短时间获取大量种子细胞,为更高的工程化肌腱构造研究创造模型构建条件。设计、时间及地点:对比观察,于2006-02/11在四川大学生物治疗国家重点实验室干细胞与组织工程研究室完成。材料:7～10dSD大鼠2只用于获取鼠尾腱细胞;第4代人包皮成纤维细胞由四川大学华西医院干细胞与组织工程研究室提供。方法:选用SD乳鼠,无菌条件下抽取尾腱,体积分数为10%的小牛血清+DF培养基悬浮组织块培养法获得鼠尾肌腱原代细胞,将第3代细胞与人包皮成纤维细胞为对照,行Ⅰ,Ⅲ型胶原免疫细胞化学染色,染色结果用image-proplus 5.02行吸光度测量,并做统计分析。主要观察指标:SD大鼠尾腱细胞免疫细胞化学染色结果及染色吸光度测量结果。结果:第2代SD大鼠尾腱细胞Ⅰ型胶原染色呈阳性,Ⅲ型胶原染色呈阴性;人成纤维细胞Ⅰ、Ⅲ型胶原免疫组织化学皆呈阳性。尾腱细胞和人成纤维细胞Ⅰ型胶原表达吸光度值均明显高于Ⅲ型胶原(P<0.05)。两种细胞Ⅲ型胶原表达吸光度值与空白对照之间差异无显著性意义(P>0.05)。结论:SD乳鼠尾腱源细胞符合肌腱细胞的生物学特性。采用组织块悬浮培养可以在短期内大量获取原代或传代尾腱细胞。
BACKGROUND： The low output of seed cells and long cycle of traditional coils culture methods in tendon animal models （rabbits and chicken） restrict the researches of tendon tissue engineering study. OBJECTIVE： To establish an ideal culture protocol of tail tendon in SD rats, to get more seed cells within less time for subsequent engineered tendon construction research. DESIGN, TIME AND SETTING： Controlled observation was performed in the National Key Laboratory of Biotherapy, Department of Stern Cells and Tissue Engineering, Sichuan University between February and November in 2006. MATERIALS： Rat tail tendon cells were harvested from 2 SD rats, aged 7-10 days; human prepuce fibroblasts were offered by National Key Laboratory of Biotherapy, Department of Stem Cells and Tissue Engineering, Sichuan University. METHODS： Tail tendon of SD rats was draw off and cut into pieces, which were then cultured in 10% fetal bovine serum ＋ DF culture medium for getting primary tendoncyte by using suspension tissue culture method. The third generation cells were processed into immuocytochemistry stain with collagen type Ⅰ and Ⅲ, while human prepuce fibroblasts served as controls. Absorbanco of stain result was measured by image-pro plus 5.02 for statistical analysis. MAIN OUTCOME MEASURES： Immuocytochemistry stain and absorbanco measurement of SD rat tail tendon cells. RESULTS： The second generation of SD rat tail tendon cells were positive for type Ⅰ collagen stain, and negative for type Ⅲ collagen stain; human fibroblast were positive for both Ⅰ and Ⅲ collagen, tn the rat tail tendon cells and human fibroblasts, the absorbance value of type Ⅰ collagen expression was dramatically higher than of type Ⅲ collagen （P 〈 0.05）. There was no significant differences addressing the absorbance of type Ⅲcollagen expression between type Ⅰand Ⅲ collagen of SD rat tail tendon cells and blank control group （P 〉 0.05）. CONCLUSION： Cells cultured from SD rat tail tendon have biological characteristic of tendon calls. Tissue piece suspension culture can obtain a auantitv of nrimarv or subcultured cells of rat tail tendon within a short time.
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2009年第46期9011-9014,共4页
Journal of Clinical Rehabilitative Tissue Engineering Research
基金
国家自然科学基金资助项目(30570469)
四川省青年科技基金资助项目(2001-19-0132)~~
