摘要
目的 探讨软骨细胞与骨髓基质细胞 (BMSC)共培养体外构建软骨的可行性 ,以阐明软骨细胞能否诱导BMSC向软骨细胞分化并形成软骨组织。方法 分别培养猪的BMSC与耳软骨细胞 ,将 2种细胞按 8:2 (BMSC :软骨细胞 )比例混匀 ,以 5 0× 10 7/ml的终浓度接种于聚羟基乙酸 /聚乳酸支架 (PGA/PLA ,直径 9mm ,高 3mm)作为共培养组 ,相同终浓度的单纯软骨细胞和单纯BMSC分别接种于相同支架作为阳性对照及阴性对照 ,1 0× 10 7/ml(相当于共培养组中软骨细胞数 )的单纯软骨细胞接种作为低浓度软骨细胞对照。每组各接种 6例标本 ,每例接种细胞悬液 2 0 0 μl。全部标本均于体外培养 8周后取材 ,通过大体观察、湿重测定、蛋白多糖含量测定、组织学及免疫组织化学等相关检测对新生软骨进行评价。结果各组细胞均与材料支架黏附良好。共培养组及阳性对照组 (软骨细胞组 )体外培养 8周后均形成了成熟的软骨组织 ,并基本保持了复合物初始的大小和形状 ,2组新生软骨外观及组织学特征基本相同 ,免疫组织化学也显示 2组均表达软骨特异性细胞外基质Ⅱ型胶原。定量测定结果表明 ,共培养组的平均湿重和蛋白多糖含量均达到阳性对照组的 80 %以上。阴性对照组 (单纯BMSC组 )在体外培养过程中逐渐皱缩变形 ,仅在组织块边缘的局部区?
Objective Chondrogenic microenvironments play a very important role in chondrogenesis of bone marrow stromal cells (BMSC) This study explored the feasibility of in vitro chondrogenesis by co culture of BMSC and chondrocytes so as to confirm the hypothesis that chondrocytes can provide chondrogenic microenvironment to induce chondrogenic differentiation of BMSC and thus promote in vitro chondrogenesis of BMSC Methods Porcine BMSC and auricular chondrocytes were in vitro expanded respectively and then were mixed at a ratio of 8: 2 (BMSC: chondrocyte) 200 μl mixed cells(5 0× 10 7/ml) were seeded onto a polyglycolic acid/polylactic acid (PGA/PLA) scaffold,9 mm in diameter and 3 mm in thickness, as co culture group Chondrocytes and BMSC with the same cell number were seeded respectively onto the scaffolds as positive control (chondrocyte group) and negative control (BMSC group) 200 μl chondrocytes (1 0× 10 7/ml, equal to the chondrocyte number of co culture group) alone were seeded as low concentration chondrocyte group There were 6 specimens in each group All specimens were harvested after in vitro culture for 8 weeks in DMEM plus 10% FBS Gross observation, average wet weight measurement, glycosaminoglycan (GAG) quantification, histology and immunohistochemistry were used to evaluate the results Results Cells in all groups had fine adhesion to the scaffolds and could secrete extracellular matrix In both co culture group and positive control group, the cell scaffold constructs could maintain the original size and shape during in vitro culture and formed homogenous mature cartilage after 8 weeks of in vitro culture Furthermore, the neo cartilages in both groups were similar to each other in gross appearance and histological features, and abundant type II collagen was also detected by immunohistochemistry in both groups The average wet weight and GAG content of co culture group were both more than 80% of those of positive control group In negative control group, however, the constructs shrunk gradually during in vitro culture and cartilage like tissue could only be observed at the edge area of the construct In low concentration chondrocyte group, the constructs also shrunk gradually during in vitro culture and the average wet weight was below 40% of that of the positive control group although histology showed a small amount cartilage formation Conclusion Chondrocytes can provide a chondrogenic microenvironment to induce a chondrogenic differentiation of BMSC and thus promote the in vitro chondrogenesis of BMSC
出处
《中华医学杂志》
CAS
CSCD
北大核心
2004年第20期1716-1720,共5页
National Medical Journal of China
基金
国家自然科学基金资助项目 (3 0 3 0 0 3 5 3 )
国家高技术研究发展计划 (863计划)基金资助项目(2 0 0 2AA2 0 5 0 2 1)
