纤维连接素对髁突软骨细胞在Cytodex-3微载体表面贴附、铺展及增殖的影响

目的:研究纤维连接素(FN)对下颌骨髁突软骨(MCC)细胞在Cytodex-3微载体表面生长的影响.方法:采用机械分离及酶消化法获得MCC细胞,将细胞以1×108/L密度分别接种于含有空白及FN(20mg∥L)包埋的微载体的自制转瓶细胞培养系统,利用环境扫描电镜对培养MCC细胞在两组微载体表面的生长方式进行动态观察.结果:MCC细胞可快速贴附于FN包埋的Cytodex-3微载体表面,铺展速度快于空白微载体组,细胞形态不规则,更为扁平,有较多伪足.细胞在包埋微载体表面生长速度显著快于空白微载体组,细胞指数生长期提前,细胞倍增时间缩短,最终生长密度高于对照组.结论:细胞在微载体表面的生长有赖于适当的细胞外基质成分,FN可促进MCC细胞在微载体表面的贴附及增殖,可用于MCC细胞大规模扩增的微载体表面处理.

Fibronectin in Adhesion, Spreading and Proliferation of Mandibular Condylar Cartilage Cells on Cytodex-3 Microcarrier

Objective:The purpose of this study was to investigate the effects of exogenous fibronectin on the attachment and growth of mandibular condylar cartilage (MCC) cells on DEAE dextran micro carrier.Methods: MCC cells were harvested from newborn New Zealand white rabbits by sequential digestion with trypsin and collagenase. They were grown on the 20 mg/L fibronectin coated and uncoated cytodex 3 micro carrier. Samples were collected after 1,3,5,7,9,11 days. The kinetics of adhesion and growth were observed with phase contrast microscope and environmental scanning electronic microscope (ESEM) and quantified by 0.1% crystal violet nuclear extrusion method.Results: MCC cells could rapidly attach and spread onto the fibronectin coated cytodex 3, and nearly 60 percent cells attached to the micro carrier within 2 hours. Spreading cells were flat and lowly refractile observed under phase contrast microscope. While the attachment and spreading of the cells in the untreated group were slow, and only 36 percent cells attached to the uncoated micro carrier. However, most cells attached to the micro carrier in both groups after 24 hours. The MCC cells attaching to FN coated cytodex 3 observed under ESEM were flatter and adhered tightly with more processes (pseudopodia) stretching out from cytoplasm. In contrast, cells on untreated cytodex 3 remained regularly round even 24 hours after being plated. Meanwhile, there was an accelerated growth rate of the MCC cells on the fibronectin treated cytodex 3. The cell density in the FN treated group was higher than that of the untreated group at the end of the culture. Conclusion: Fibronectin, one major glycoprotein of extracellular matrix, plays an important role in cell attachment and spreading.\ The better understanding of the extracellular matrix will be helpful in selecting the optimal substrate for cell growth in vitro.