摘要
目的 :以胶原蛋白和人血纤维蛋白混合物为载体在体外进行关节软骨细胞三维立体培养 ,构建人工软骨组织 ,研究载体对软骨细胞在其中的表达及合成 型胶原的影响。方法 :取 2周龄的新生兔关节软骨 ,经消化 ,将获得的软骨细胞与牛 型胶原、人血冻干纤维蛋白原、凝血酶按一定比例混合 ,制成软骨培养物并在体外培养。培养第 3周时 ,取材进行 Masson染色、 型胶原寡核苷酸探针原位杂交和 型胶原的免疫组化观察。结果 :体外培养 3周 ,培养物内细胞大部分存活 ,Masson染色形成软骨陷窝 ,同源性细胞簇出现 ,细胞周围有蓝色物质环绕存在 ,原位杂交证实其软骨细胞表达 型胶原 m RNA,免疫组化证实软骨细胞分泌 型胶原。结论 :用胶原蛋白和人血纤维蛋白为载体支架体外培养软骨细胞 ,可促进关节软骨细胞合成分泌 型胶原 , 型胶原是软骨组织最主要的细胞外基质成分。
Objective To detect collagen Ⅱ in constructed articular cartilage by three-dimensional culturing system using collagen and human fibrin as supporter. Methods Chondrocytes were isolated from articular cartilage of 2-week-old rabbit, digested by collagenase Ⅱ, and mixed with bovine collagen I, human blood freeze-dry fibrinogen and thrombosin according to a certain proportion,and were cultured in vitro. Biopsies was analyzed by Masson stain, hybridization in situ using oligonucleotied probe of collagen Ⅱ and immunohistochemistry test of collagen Ⅱ after 3 weeks. Results After 3 weeks, all cells were viable, formed cartilage lacunae, appeared homologous cell cluster and secreted extracellular matrix that could be stained with Masson stain. In situ hybridization assay showed that chondrocytes expressed of collagen Ⅱ mRNA. It was demonstrated that chondrocytes could secrete collagen Ⅱ with immunohistochemistry method. Conclusion Chondrocytes still secrete collagen Ⅱin three-dimensional culturing system using collagen and human fibrin as supporter. Collagen Ⅱ is one of the most important component of cartilage tissue.
出处
《吉林大学学报（医学版）》
CAS
CSCD
北大核心
2004年第4期524-526,F002,共4页
Journal of Jilin University：Medicine Edition
基金
国家自然科学基金资助课题 (39870 781
30 2 71319)
