藻酸钙复合材料构建组织工程化骨及软骨的实验

背景软骨组织工程的两个关键问题是种子细胞和支架材料,保证支架材料上有足够数量的功能细胞是体内形成骨和软骨组织的基础.目的对经体外培养的骨髓基质干细胞与藻酸钙载体及生长因子复合形成的组织工程性骨及软骨进行形态学观察.设计细胞形态对比,观察12周实验结果.单位郑州大学骨科研究所,上海第二医科大学骨科实验室.对象实验于2001-09/2002-06在上海第二医科大学骨科实验室完成.3月龄雄性新西兰大白兔9只,平均体质量3.2kg.方法复合材料分别为①藻酸钙.②藻酸钙+骨髓基质细胞.③藻酸钙+骨髓基质细胞+类胰岛素生长因子Ⅰ.④藻酸钙+骨髓基质细胞+转化生长因子β.⑤藻酸钙+骨髓基质细胞+转化生长因子β+类胰岛素生长因子Ⅰ.将5种组合注射于同一个体新西兰大白兔背部不同部位皮下组织中,每组各注射2处,共10处,每处0.2mL,分别做好标记.按组分别于术后4,8,12周取材,各时间点处死动物3只,取兔背部皮下增殖物进行骨和软骨组织生成物观察,应用苏木精-伊红、甲苯胺蓝、Masson、番红-O染色技术.主要观察指标兔背部皮下增殖物的组织学观察.结果兔背部皮下增殖物的组织学观察结果4周时可见藻酸钙+骨髓基质细胞+类胰岛素生长因子Ⅰ、藻酸钙+骨髓基质细胞+转化生长因子β、藻酸钙+骨髓基质细胞+转化生长因子β+类胰岛素生长因子Ⅰ组有岛状软骨样组织形成,软骨细胞包绕于碱性基质中,8周时复合物中形成大量的类软骨结构,并部分向骨组织转化,细胞周围有大量的基质分泌,部分软骨细胞呈簇状分布.12周时藻酸钙+骨髓基质细胞+转化生长因子β+类胰岛素生长因子Ⅰ组软骨组织基本转变为骨组织,并开始吸收.结论骨髓基质细胞-藻酸钙-生长因子复合材料能形成软骨及骨组织.藻酸钙适合骨髓基质细胞生长,是一种良好的骨及软骨组织工程载体.

Experimental study of calcium alginate compound in the construction of tissue engineered bone and cartilage

BACKGROUND: Seed cells and supporting materials are two key problems for cartilaginous tissue engineering. Enough functional cells in supporting materials is the foundation in the formation of bone and cartilaginous tissue in vivo.OBJECTIVE:To perform a morphological study on the tissue engineered bone and cartilage constructed with calcium alginate, in vitro cultured bone marrow stem cells(BMSCs) and growth factors.DESIGN:Morphological comparative study with the results observed for 12 weeksSETTING:Orthopaedic Institute of Zhengzhou University and Orthopedic Laboratory of the Second Affiliated Hospital of Shanghai Medical University.MATERIALS:This experiment was completed from September 2001 to June 2002 in the Orthopedic Laboratory of the Second Affiliated Hospital of Shanghai Medical University. Nine New Zealand male rabbits at the age of 3 months were selected, their mean body mass was 3.2 kg.METHODS: Rabbits received dorsal subcutaneous injection of five kinds of the following combinations at two points for each combination and in the Ⅰ+TGF-β. Totally ten points were marked. Three rabbits were put to death at the 4th, 8th, 12th weeks after injection respectively and dorsal subcutaneous hyperplasia tissues were collected for observation by using hematoxylin and eosin (HE), toluidine blue, Masson, tomato red-O staining techniques.MAIN OUTCOME MEASURES: Histological observation of dorsal subcutaneous hyperplasia tissues.RESULTS: At the fourth week, cartilaginous like tissues island could be observed in calcium alginate+ BMSCs + IGF-Ⅰ group, calcium alginate+BMSCs +TGF-β group and calcium alginate + BMSCs + IGF-Ⅰ +TGF-βgroup, with chondrocblasts surrounded by alkaline matrix; A great deal of cartilaginous-like tissue appeared at the eighth week with part of it transforming into bone, and cells secrete much matrix, part of the chondroblast clustered. At the 12th week, approximately all of the cartilaginous tissues turned into bone in calcium alginate+ BMSCs + IGF- Ⅰ +TGF-β group, and began to absorb.CONCLUSION: The compound composed of calcium alginate, BMSCs and growth factor might form bone and cartilaginous tissue. Calcium alginate is suitable for the growth of BMSCs and is believed to be an optimal bone and cartilaginous engineering carrier.