Investigation Of PDLCs' Biology Function By EMPs And BMP-2

The enamel matrix proteins (EMPs) is the protein composition in unmineralized enamel in enamel developed process, the research discovered that it can compound by Hertwig's Epithelial Root Sheath(HERS), and participate in the forming of the enamel. In recent years, EMPs have been introduced into the therapy field of periodontology. Compared with other method of periodontal treatment, the effect after using EMPs similar to the physiological status of periodontal attachment apparatus. But, the underlying biological mechanism is not understood.This is also the reason why people study it eagerly, cementoblastThe BMP-2 is a important kind of growth factor, it can induce undifferentiated mesenchymal cell differentiation into osteoblast unreversibleand,and the new bone formed. The BMP-2 was also apply in the therapy of the periodontal diseases,it can induce undifferentiated mesenchymal cell differentiation into cementoblast and osteoblast. The BMP-2 can be found in periodontal; it can also promote PDLCs proliferation, composing DNA and the vivid of Alkaline Phosphatase. With proper carrier, The BMP-2 can stimulate the new-born bone obviously, the area of defect will quickly fulled cartilage, subsequent formation new bone and new cement. This process is similar to the physiological status of periodontal attachment apparatus.The most ideal process of periodontal regeneration include alveolar bone,cement and periodontal ligament. periodontal ligament Cell is the main cell component ofmetabolize and physiological rebuild, among them, periodontal ligament fibreblast Cell form the crassitude fiber, osteoblast form alveolar bone and cementoblast form cement., these cells composition all renewal depend on mesenchymal cell.In commonly consider, periodontal ligament Cell is is the base and main source of periodontal regeneration.But the sources of PDLCs are very limited in the periodontal defects, so that it is difficult to obtain the satisfactory regeneration. And then tissue engineering is supposed to be able to solve the problem. Tissue engineering is the emerging field of science aimed at developing techniques for the fabrication of new tissues to replace damaged tissues. To engineer living tissues, cultured cells are coaxed to grow on bioactive degradable scaffolds that provide the physical and chemical cues to guide their differentiation and assembly into three-dimensional tissues.Therefore, the study the effect of EMPs and BMP-2 acting on PDLCs will contribute to understanding the mechanism of periodontal regeneration, periodontal regeneration.This study has included the three parts:1. The study of the influences on the proliferation of human PDLCs by different induce conditionThe influences on the proliferation of human PDLCs by BMP-2 or/and EMP were evaluated by MTT colorimetric assay, in order to analyze the possible mechanism of promoting the proliferation of PDLCs by BMP-2 or/and EMPs. The results have indicated that within 3 day, within 7 day and within 7 day,the result are likely resemble:BMP-2 and EMPs were both able to enhance the proliferation of PDLCs, and BMP-2 and EMPs had synergistic action. BMP-2 and EMPs enhanced the proliferative responses of PDLCs in a dose-dependent manner, and the effect of BMP-2 was more obvious than EMPs. The most ideal dose of EMPs is100mg/L,while The most ideal dose of BMP-2 is lOOug/L.2.the study of the influences on the activity of Alkaline Phosphatase and forming mineralize tuber of human PDLCs by different induce condition.The influences on the activity of Alkaline Phosphatase of human PDLCs by BMP-2(100ug/L) or EMPs(100mg/L) were evaluated by activity of Alkaline Phosphatase assay, compared with traditional mineralizatino inducing in order to analyze the possible mechanism of promoting the mineralize ability of PDLCs by BMP-2 or EMPs. The results have indicated that within 3 day, BMP-2 and EMPs were both able to enhance the mineralize ability of PDLCs,within 7 day and within 3-7 day,BMP-2 were able to enhance the mineralize ability of PDLCs .EMPs were not able to enhance the mineralize ability of PDLCs, and the effect of BMP-2 was more obvious than EMPs. That indicate EMPs is not an osteoinduction factor but a Osteopromotion factor.The influences on the forming mineralize tuber of human PDLCs by BMP-2(100ug/L) or EMPs(100mg/L) were also evaluated by Von-Kossa. The result have indicated BMP-2 or EMP can accelerate the forming mineralize tuber of human PDLCs, the effect of BMP-2 was more obvious than EMPs. This not only owe to the ability of enhance the mineralize ability of PDLCs by BMP-2 or EMPs. Although PDLCs possess heterogeneity,the PDLCs that didn't use induce only appear to layer growth, instead of the mineral tuber.This indicate that layer growth is not unique decision factor that the mineral,it need join the mineral induce liquid or the growth factors,and then it can be can carry out.3.1nfluences on Synthesis of noncollagen matrix proteins of Human Periodontal Ligament Cells by different induce condition..The noncollagen matrix proteins is include Osteopontin, Bone Sialoprotein,osteonectin and osteocalcin. In the hard organizations such as bone andalveolar bone, calcium and phosphoric acid ions are organize by the noncollagen matrix proteinsand collagen plank,and then mineralize formed.Because they are nearly relation with the metabolism and rebuilt of the bone,they also called Osteorelation protein.CAP is a particularity protein relating to forming cement,it can pick-up from cement. The influences on synthesis of noncollagen matrix proteinsand CAP of human PDLCs by BMP-2(100ug/L) or EMPs(100mg/L) were evaluated by immunity organization chemistry method and the picture analysis methods. The results have indicated: the PDLCs can express the OPN and BSP with the low level and can hardly express the ON under the condition of normal development,that means PDLCs express the characteristic that become the an ossification cell cultured in vitro. Our results showed that the factors could promote PDLCs' synthesis of OPN or BSP or ON in different time, especially the traditional mineral induce,the BMP-2 is senior and the EMPs is subordination, which indicates them could exert their action in promote periodontal hard tissues regeneration through promotion PDLCs' osteoblast differentiation.On the other hand , after BMP-2 act 7 day and EMPs act 4day and 7 day, part of PDLC can express CAP.This indicate that EMPs and BMP-2 promote PDLCs' cementoblast differentiation. EMPs was more obvious than BMP-2.