羊软骨细胞在生物反应器中的培养和扩增  

A quick way in vitro for amplification of goat chondrocytes by RCSS

作　　者：张莉[1] 郭全义[1] 眭翔[1] 许文静[1] 袁玫[1] 黄靖香[1] 卢世璧[1] 

机构地区：[1]解放军总医院骨科研究所,北京市海淀区复兴路28号100853

出　　处：《中国矫形外科杂志》2006年第6期446-448,共3页Orthopedic Journal of China

基　　金：国家863计划资助项目(2002AA205021)

摘　　要：[目的]探索在旋转生物反应器内,应用微载体技术快速扩增分化良好的羊软骨细胞的方法。[方法]将培养的羊软骨细胞应用Cytodex3微载体在旋转生物反应器(RCSS)内,进行动态培养,应用倒置显微镜对微载体表面的软骨细胞进行动态观察,并对收获的软骨细胞进行Ⅰ、Ⅱ型胶原的细胞免疫化学染色分析。[结果]关节软骨细胞于1d内贴附于Cytodex3微载体表面,细胞初期为圆球形、半球形凸起,逐渐向周围伸展,随时间的延长,贴附于微载体的细胞逐渐增多,到培养后期,细胞密度可达最初接种的15～17倍,在微载体上收获的软骨细胞经Ⅰ型胶原的免疫细胞化学染色呈阴性,Ⅱ型胶原染色则呈强阳性。[结论]利用微载体细胞培养技术可简便快速地在体外扩增羊软骨细胞,可为构建组织工程化人工软骨提供大量活性、分化良好的软骨细胞。[ Objective]To establish a method for obtaining well-differentiated goat ehondroeytes quickly in large scale by RCSS using mieroearriers technique. [ Method ] Articular ehondroeytes were harvested from goat by sequential digestion with trypsin and eollagenase , and then grown in RCSS bioreaetor euhure system which contained the Cytodex-3 mieroearriers in the euhure medium （DMEM） . Growth of chondrocytes on Cytodex-3 mieroearriers was observed dynamically under phase contrast microscope . Immunoeytoehemieal analysis was performed for type Ⅰ collagen and type Ⅱ collagen, [ Result ] The articular ehondroeytes attached rapidly to the surface of Cytodex-3 mieroearriers. Quick growth of these cells was observed after they fully spread onto the mieroearriers. The density of ehondroeytes increased by 15 - 17 times in later stage of euhure as compared with the initial density, The harvested ehondroeytes had no detectable staining for collagen type Ⅰ , but stained intensively for collagen type Ⅱ. [ Conclusion] Mieroearrier culture of ehondroeytes can yield a large quantity of goat cells within a short time ,which will be of benefit for banking cartilage cells for reconstruction of impaired cartilage by way of tissue engineering.

关 键 词：生物反应器 微载体 软骨细胞培养 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]