Study Of The Expression Of HGFmRNA In Distraction Osteogenesis And Periosteal Distraction Osteogenesis

Objective:In situ hybridization method was used to observe the expression of hepatocyte growth factor(HGF) in the course of distraction osteogenesis(DO) and periosteal distraction osteogenesis(PDO) on the basis of the rabbit mandibular experiment model. To explore the role of HGF on the forming of new bone and provide theoretical basis for the exogenous application and gene therapy.Methods:Fifty-six adult New Zealand white rabbits were selected for the experiment of distraction osteogenesis and periosteal distraction osteogenesis. In the experiment of distraction osteogenesis the animals were divided into two groups as distraction group and control group. There were 20 rabbits for each group and the operations were conducted on every animal. For distraction group, the stretch rate was set as 0.5mm/12h after 7days interval and fixed for 7days. At the end of the intermittent period,distraction period and the end of 1^thw,3^thw,5^thw of fixed period, and four animals were sacrificed. The control group was free of distraction and four animals were sacrificed at the end of 1w,2w,3w,5w,7w after operations. In periosteal distraction osteogenesis experiment the device was rigidly fixed to the lateral surface of the mandible in 16 mature New Zealand white rabbits. The periosteum were fixed in situ for 7 days after operation. It was distracted by 2mm from the surface of the mandible at 8^th day postoperatively.During the next 20 days periosteum was further distracted by 1mm every 4 days.Finally, it was distracted 7mm from the surface of the mandible. The unoperated, contralateral side of the mandible served as the control.Four rabbits were sacrificed randomly at postoperative days 28, 35, 42, and 56. The specimens were studied by hematoxylin-eosin stain(HE stain) examination and in situ hybridization to observe the expression pattern of HGF.Result:In distraction osteogenesis groups the intermittent gap were filled with the fibroblasts and collagen fibers. Inflammatory cells were seen scattered among the fibers. There were new trabecular bone and osteoid formed along the direction of distraction. Larger amount of bone was formed and fibrous tissue reduced after 5weeks fixation. HGFmRNA was mainly expressed in the inflammatory cells at the end of the intermittent stage. Positive staining was seen at the end of distraction stage in the vascular endothelial cells and mesenchymal cells. The peak intensity of expression was found at the end of first week of fixation and there is little expression at the 5th week. No remarkable difference was found between the two groups at the end of the fixation.In periosteal distraction osteogenesis experiment, there were active proliferation of cells at the contact sites of periosteal and bone. The osteocytes grew and some were surrounded by bone matrix deposition. Trabecular bone was seen after 42 days fixation and new osteoporosis grew mature when it was at day 56. There was weakly positive expression of HGF mRNA in normal periosteum. In the conditions of external stretch the expression of HGF mRNA increased and was seen at the neonatal bone matrix. The expression level of HGF mRNA decreased gradually and no remarkable difference was seen at the postoperative days 56. Conclusions:There was positive expression of HGF mRNA in tissue forming of distraction osteogenesis and periosteal distraction osteogenesis. The expression in mesenchymal and vascular cells lasted longer than other cells and the positive staining was also strong. Endogenous HGF may played an important role on cell proliferation, differentiation and tissue formation. The result of this experiment provided some theoretical basis for further study of exogenous application and gene therapy with HGF.