Basic Study of Suitable Condition for Bone Formation by Human Osteoblasts and Clinical Application

为了在生长和改建过程中骨形成机制的基础上开发复合成骨细胞的骨替代材料，使从骨髓中获得的分化成骨细胞产生骨组织是必不可少的。本研究探讨了体外培养的人成骨细胞向成骨细胞分化的适宜条件及体内成骨能力。培养6d后，细胞碱性磷酸酶(ALP)活性呈剂量依赖性增加。此外，10^lt；-7&gt；M地塞米松处理3-9天后，ALP活性呈时间依赖性增加。地塞米松作用5d后，成骨细胞分化的生化指标，包括碱性磷酸酶活性、I型前胶原羧基末端多肽和骨钙素的分泌均显著增强。在单层培养中，成骨细胞的矿化也被…刺激。此外，人成骨细胞在含有变性I型胶原的胶原海绵中培养，加入或不加入10^&lt；-7&gt；M Dex。将这些细胞与胶原海绵一起移植到SCID小鼠体内，移植后2~8周收获。2~4周，经地塞米松处理的成骨细胞移植形成类骨样组织，数量呈时间依赖性增加至8周。这种骨样组织是由移植细胞新合成的矿化胶原基质组成的。ALP活性和骨钙素在类骨样组织周围的移植细胞中有较高的表达。在超微结构上，这种矿化的基质类似于骨。未经处理的细胞和胶原海绵单独移植无法形成骨样组织。这些研究表明，地塞米松处理的成骨细胞在体内形成了类骨组织，其形成过程与骨形成机制相似，提示从生长和改建过程中骨形成的机制出发，培养的成骨细胞作为骨替代物是有价值的。较少

For the development of the bone substitutes combined with osteblats on the basis of the mechanism of bone formation during growth and remodeling, it is indispensable to make the differentiated osteoblastic cells obtained from bone marrow produce bone tissue. In this study, we evaluated the suitable condition for the differentiation into osteoblast and the capacity for in vivo bone formation of cultured human osteoblastic cells.The alkaline phosphatase (ALP) activity of the cells cultured for 6 days in 10^<-8>-10^<-6>M Dex increased in a dose-dependent manner. Furthermore, an increase of ALP activity by 10^<-7>M Dex treatment for 3-9 days was observed in a time-dependent manner. Biochemical indicators of osteoblastic differentiation, which include ALP activity and secretion of procollagen type I carboxy-terminal peptide and osteocalcin, were significantly enhanced by Dex treatment at 10^<-7>M for 5 days. In monolayr culture, mineralization by the osteoblastic cells was also stimulated b … More y Dex treatement.Furthermore, human osteoblastic cells were cultured within collagen sponge, which consists of denatured type I collagen, in the presence or absence of 10^<-7>M Dex. These cells along with collagen sponge were transplanted into the SCID mice, and the transplants were harvested at 2-8 weeks. At 2-4 weeks, the transplants of Dex-treated osteoblastic cells formed bone-like tissue, the quantity of which increased in a time-dependent manner to 8 weeks. This bone-like tissue was composed of mineralized collagen matrix newly synthesized by the transplanted cells. A higher expression of ALP activity and osteocalcin was detected in the transplanted cells surrounding the bone-like tissue. Ultrastructurally, this mineralized matrix was similar to bone. The transplants of non-treated cells and collagen sponge alone failed to from bone-like tissue. Thses studies indicate that the bone-like tissue is formed in vivo by the Dex-treated osteoblastic cells via the process resembled mechanism of bone formation, suggesting that the cultured osteoblastic cells is useful as a bone substitutes on the basis of the mechanism of bone formation during growth and remodeling. Less