兔骨髓基质干细胞骨向诱导实验研究

Experimental Study on BMSC Culture and Differentiation Towards Osteoblasts

目的:研究兔骨髓基质干细胞(bone marrow stromal cells,BMSC)的体外培养以及骨向诱导分化情况,进一步探讨其作为骨组织工程种子细胞的可行性。方法:无菌条件下抽取兔的股骨骨髓,然后进行骨髓基质干细胞的分离和体外培养,并向成骨细胞定向诱导分化。结果:体外分离的骨髓基质细胞呈贴壁生长,改良MTT法测定显示骨髓基质细胞于第7.8天左右可达到增殖高峰;诱导后BMSC可向成骨细胞分化,细胞呈成骨细胞形态为梭形和多角形并可形成钙化结节。改良钙钴染色法检测诱导细胞碱性磷酸活性呈强阳性。扫描电镜观察细胞呈较典型的成骨细胞状态,并可见钙盐沉积。结论:骨髓基质细胞具有来源充足,取材方便,创伤小无明显并发症。骨髓基质干细胞分化增殖能力较强,成骨能力确定。所以采用骨髓基质细胞作为种子细胞构建组织工程骨有比较可靠的依据。

Objective： To study the culture and osteogenic differentiation of bone marrow stromal ceils （BMSC） so as to search a kind of appropriate seed cells for bone tissue engineering. Methods： We obtained BMSC from the rabbit and induced differentiation toward osteoblasts with special cultural fluid in vitro. The condition of both induced cells and normal cells were observed under an inverted phase contrast microscope, scanning electron microscope （SEM）, and ALP was tested by Gomori method. Results： BMSC growed by static adherence, were differentiated to osteoblasts and had strong capacity of proliferation. The cells proliferation reached summit at 7d or 8d by MTT. And ability of formation of calcifying nodules was detected. Alkaline phosphatase （ALP） activity of BMSC increased significandy by Gomori method. Osteoblast--like and calcium salts were tested through scanning electron microscope. Conclusions： To obtaining the BMSC from bone is convenient and cells are sufficient with a little trauma and no complication in this experiment. BMSC have a strong activity of proliferation and osteogenic capacity. So BMSC are apt to serve as seed cells for bone tissue engineering.