牙髓干细胞在壳聚糖-纤维蛋白原复合支架材料上的黏附与增殖  

Adhesion and proliferation of dental pulp stem cells on the chitosan-fibrin composite scaffold

作　　者：郑丽珠[1] 李小兵[2] 张苗[1] 于璐[1] 刘奕杉[1] 

机构地区：[1]新疆医科大学第一附属医院儿童口腔科-口腔预防科,新疆维吾尔自治区乌鲁木齐市830054 [2]四川大学华西口腔医院儿童口腔科,四川省成都市610041

出　　处：《中国组织工程研究》2017年第10期1552-1557,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金(81560178)~~

摘　　要：背景:随着组织工程技术的飞速发展,单一的生物支架材料很难满足组织工程的需要,通过合适的方法将两种或多种类型的材料组合起来,可以得到性能更加优良的复合支架材料。目的:检测牙髓干细胞在壳聚糖-纤维蛋白原复合支架材料上的黏附与增殖情况。方法:(1)采用改良酶消化法分离提取C57乳鼠牙髓干细胞,传代培养至第3代,体外成脂成骨诱导分化对细胞进行鉴定;(2)制备壳聚糖-纤维蛋白原复合支架材料,测定其孔径及孔隙率;(3)将壳聚糖-纤维蛋白原复合支架材料与第3代小鼠牙髓干细胞共培养,以细胞直接接种于培养板底部作为对照组,MTT法检测细胞增殖情况;(4)扫描电镜观察壳聚糖-纤维蛋白原复合支架材料的形态,细胞黏附、增殖及基质分泌情况。结果与结论:(1)成功分离培养C57小鼠牙髓干细胞,经成脂成骨诱导分化后结果呈阳性;(2)壳聚糖-纤维蛋白原复合支架材料孔径为(105.32±22.10)μm,孔隙率为(87.714±1.276)%;(3)牙髓干细胞在壳聚糖-纤维蛋白原复合支架材料上的增殖曲线与对照组基本一致,均为"S"型,培养第4-8天实验组增殖速度显著快于对照组,差异有显著性意义(P<0.05);(4)共培养第2天,细胞与壳聚糖-纤维蛋白原复合支架材料黏附紧密,生长状态良好,第4天,细胞明显开始增殖,体积增大,有丰富的细胞外基质,第6天,铺满于整个材料表面及空隙;(5)壳聚糖-纤维蛋白原复合材料是一种适宜牙髓干细胞黏附与增殖的支架材料。BACKGROUND： With the rapid development of tissue engineering, a single biological scaffold material is hard to meet the needs of tissue engineering. Therefore, composite scaffolds with excellent performance will be obtained by combining two or more kinds of materials.OBJECTIVE： To detect the adherence and proliferation of dental pulp stem cells on the Chitosan-fibrin composite scaffold.METHODS： Dental pulp stem cells were isolated and extracted from C57 neonatal rats through modified enzyme-digestion method, and subcultured to the third generation, followed by adipogenic and osteogenic induction in vitro. Then, induced cells were identified. The chitosan-fibrinogen composite scaffold was prepared, and the pore size and porosity were determined. The chitosan-fibrin composite scaffold was co-cultured with passage 3 dental pulp stem cells to observe the cell proliferation by MTT assay, and the morphology of the composite scaffold, cell adhesion,proliferation and extracellular matrix secretion were observed under scanning electron microscope. In addition, the cells were inoculated directly on the bottom of culture plate as controls.RESULTS AND CONCLUSION： The dental pulp stem cells were successfully isolated and cultivated, and positive for osteogenic and adipogenic differentiation. The pore size and porosity of the composite scaffold was （105.32±22.10） μm and （87.714±1.276）%, respectively. The ＂S＂-shaped proliferation curve in the experimental group was similar with that in the control group; the proliferation rate in the experimental group was significantly higher than that in the control group after 4-8 days of culture （P 〈 0.05）. At the 2nd day after co-culture, the cells adhered tightly and grew well onto the composite scaffold; at the 4th day, enlarged cells began to proliferate obviously with abundant extracellular matrix; the surface and pores of the scaffold were full of cells at the 6th day. These results indicate that the chitosan-fibrin composite scaffold is suitable for the a

关 键 词：生物材料 材料相容性 壳聚糖 纤维蛋白原 支架材料 年轻恒牙 牙髓干细胞 牙髓-牙本质复合体 国家自然科学基金 

分 类 号：R318[医药卫生—生物医学工程]