中文摘要：

目的：为克服目前组织工程中细胞和材料的复合方法中存在的缺陷，利用细胞装配系统将细胞和材料的共混物按照预先设定的结构直接打印成三维复合支架，并通过体外培养检测支架里细胞的形态和活性。方法：实验于2006—06／2007—03在清华大学生物膜与膜生物工程国家重点实验室和清华大学机械系组织制造中心完成。①选用海藻酸盐和明胶的混合水凝胶作为支架材料，配制终浓度为7％且藻酸盐和明胶比例为3：4的混合物，用CaCl2溶液交联后，冷冻干燥，用扫描电镜观察共混材料的内部结构。②取出生3-5d的新西兰乳兔的关节软骨组织，用Ⅱ型胶原酶消化获得原代软骨细胞并体外扩增至2或3代后备用。③将软骨细胞悬液与复合水凝胶混合均匀后，再利用快速成形技术组装的细胞装配系统将此共混物直接打印成组织工程三维复合支架，并体外培养此复合支架。④在培养的1，7，14和21d采用MTT的方法检测软骨细胞在支架中的增殖；培养7d后用苏木精-伊红染色的组织学方法观测细胞的形态；培养14d后采用免疫组化检测支架中软骨细胞的Ⅱ型胶原表达。结果：①扫描电镜观察的结果表明共混物的内部是交错的网格结构，有着互相连通的孔。②在细胞装配系统上按照预先设计的参数将细胞和材料直接组装为网格状的细胞，凝胶复合支架，尺寸为10mm×10mm×6mm，锥虫蓝染色表明成形后支架里细胞的存活率大于90％。③MTT实验表明软骨细胞在细胞，凝胶的支架里增殖很快，21d的吸光度是1d的3．5倍。④苏木精-伊红染色显示体外培养7d后，复合支架里的软骨细胞都是圆形的，是生理状态下软骨细胞的正常状态，而且还可以看到正在分裂的细胞。⑤免疫组化的结果表明培养14d后，培养在支架里的软骨细胞仍保持着分泌Ⅱ型胶原的活性。结论：通过细胞?

英文摘要：

AIM： To overcome the shortcomings of fabricating cell and material composites in tissue engineering, this study is designed to form three-dimension （3D） chondrocytes and materials compound scaffolds with predesigned parameters by using cell controlled assembling system, and evaluate the morphology and proliferation of chondrocytes in the formed cell/gel scaffolds cultured in vitro. METHODS： The experiments were completed in the State Key Laboratory of Biomembrane and Membrane Biotechnology, Tsinghua University and Center of Organism Manufacturing, Department of Mechanical Engineering, Tsinghua University from June 2006 to March 2007.（1）Alginate and gelatin were selected to form mixed hydrogel, which was used as biomaterial of scaffold. The total concentration of hydrogel was 7% with the alginate/gelatin ratio of 3：4. After crosslinked with CaCl2 solution and cryodesiccation, the inner structure of mixed materials was observed by scanning electron microscope （SEM）.（2）Articular cartilage was harvested from young （3-5 days old） New Zealand white rabbits, and then was digested by collagen type Ⅱ to release chondrocytes. Then chondrocytes were cultured in vitro to expand for use. （3）The chondrocyte suspension was added into the mixed hydrogel. Then this mixture of hydrogel and cell was used to form 3D composite scaffold by cell assembling system with rapid prototyping, and the composite scaffold was cultured in vitro.（4）After cell culturing for 1, 7, 14 and 21 days, MTT assay was used to analyse proliferation of chondrocytes in gel. At 7 days of culture, the morphology of cells was observed by histological analysis, in which the cell/gel scaffold was stained with hematoxylin-eosin （HE）. At 14 days of culture, immunohistochemical analysis was used to detect the expression of type Ⅱ collagen for chondrocytes in gel. RESULTS： （1）The result of SEM observation revealed that, the inner structure of mixed hydrogel was reseau with interstice.（2）The porous grid-like cell/gel