Study of bone-forming factors and their clinical evaluation

1.建立了以20日龄胎鼠顶骨为材料的无血清成骨器官培养体系。在有氧条件下，将骨在网格上培养并放置在摇床上。这种条件增加了培养的大鼠顶骨的成骨活性。测定培养液中钙含量、骨干重和骨组织产生的乳酸，以研究生命骨的精确成骨过程。2.利用改进的成骨器官培养体系，研究胰岛素对骨形成的影响。胰岛素可增加骨钙含量和骨干重。组织形态计量学分析显示，胰岛素作用10℃时，矿化骨和骨基质面积增大，成骨细胞较多，破骨细胞较少，胰岛素抑制钙向培养液中的释放。这些结果表明，胰岛素对骨形成有促进作用，促进骨矿化和骨基质形成。3.采用聚合酶链式反应方法克隆骨形态发生蛋白(BMP)基因。4.用秋水仙碱诱导骨形成和骨吸收的大鼠胫骨骨髓细胞进行原位杂交，证实骨桥蛋白在骨吸收过程中有表达。5.对儿童骨折病例进行临床统计和长期随访。

1.Serum-free bone-forming organ culture system using twenty-day fetal rat parietal bones was established. Bones were cultured on grids and placed on a rocking platform under the aerobic condition. This condition increased the bone-forming activities of cultured rat parietal bones. The calcium content, dry weight of bone, and lactate produced by bone tissue in the medium were measured to study the precise bone-forming process of vital bones.2.Using this improved bone-forming organ culture system, the effects of insulin on bone formation were studied. Insulin increased calcium content and dry weight of bone. These effects were maximum in bones treated with insulin at 10^<-6>M.Histomorphometrical analysis showed that the areas of mineralized bone and bone matrix increased, and that many osteoblasts and few osteoclasts appeared in bones treated with insulin at 10^<-6>Mor higher.Insulin inhibited the release of ^<45>Ca into the medium. From these results, it was suggested that insulin has a stimulatory effect on bone formation enhancing both bone mineralization and bone matrix formation.3.Cloning of bone morphogenetic protein (BMP) cDNA by PCR method was performed. Ectopic bone formation induced by BMP,was investigated, and also the molecular cloning of BMP type I receptor was done.4.In situ hybridization of the rat tibial bone marrow cvity, in which bone formation and resorption were indluced by colchicine, demonstrated that osteopontin mRNA was expressed during bone resorption process.5.Clinicostatistical and long-term follow-up studies of the pediatric fracture cases were performed.