The Effects Of Emdogain,Simvastatin And Their Combination On The Proliferation And Differentiation Of Human Periodontal Ligament Cells

ObjectsTo observe the effects of Emdqgain,Simvastatin and their combination on the proliferation and differentiation of human periodontal ligament cells(HPLCs)Methods1,Preparation of periodontal ligament tissue.Eight periodontally healthy and non-carious premolars extracted in the course of orthodontic treatment were collected from two patients with their informed consent. Then the teeth were putted into the sterile PBS solution, and the periodontal ligament tissue which attached to the mid-third of each root was extracted within two hours to culture the periodontal ligament cells.2,Preparation and culture of HPLCsHPLCs were cultured to confluence in a 75mm~2 culture flask with Dulbecco's modified Eagle's medium (DMEM)supplemented with 15% fetal bovine serum, 100 U:ml penicillin, 100 mg:ml streptomycin and 1 mg:ml amphotericin B (medium A). When HPLCs formed a confluent monolayer, cells were harvested with 0. 25% trypsin transferred to a 150mm~2 culture flask. HPLCs in cultures at the 4th,5th,6th passage were used in the following experiments.3,Indentification for human periodontal ligament cells ①Cells morphology on phase-contrast microscope.②Mineralization nodule staining(Alizarin Bordeaux staining method),③Indentification on immunohistochemistry4, To observe the effects of Emdogain(200,100,50,25mg/L )on the proliferation anddifferentiation of human periodontal ligament cells①To assay proliferation of HPLCs by MTT.②To assay ALP activity of HPLCs by PNPP.③Type- I collagen in HPLCs were detected by assaying the level of hydroxyproline.5. To observe the effects of Simvastatins(0.125μmol/L,0.25μmol/L,0.5μmol/L,1.0μmol/L)on the proliferation and differentiation of human periodontal ligament cells①To assay proliferation of HPLCs by MTT.②To assay ALP activity of HPLCs by PNPP.③Type-I collagen in HPLCs were detected by assaying the level of hydroxyproline.6,To observe the effects of the combination of 50mg/L Emdogain and 0.125μmol/L Simvastatins on the proliferation and differentiation of human periodontal ligament cells①To assay proliferation of HPLCs by MTT.②To assay ALP activity of HPLCs by PNPP.③Type- I collagen in HPLCs were detected by assaying the level of hydroxyproline.Results1,To observe the effects of Emdogain(200 mg/L,100 mg/L,50 mg/L,25mg/L)on the proliferation and differentiation of human periodontal ligament cells. Compared with control group, Emdogain(100 mg/L,50 mg/L,25mg/L)can increase the proliferation, the synethesis of Type- I collagen of HPLCs.Compared with control group ,Emdogain(100 mg/L,50 mg/L,25mg/L )can increase the ALP activity ,but 200mg/L Emdogain can not increase the ALP activity during the experiment. No difference was found among treated groups,and the effects of 50mg/L Emdogain was the highest(P<0.05) according to the datas of this experiment.2. The effects of Simvastatins(0.125μmol/L,0. 25μmol/L,0.5μmol/L,1.0μmol/L) on the proliferation and differentiation of human periodontal ligament cells. Compared with control group, Simvastatins(0.125μmol/L,0. 25μmol/L,0.5μmol/L,1.0μmol/L )can increase the ALP activity and the synethesis of Type- I collagen of HPLCs, no differences was found among treated groups ,and the value of 0.125μmol/L Simvastatins was highest.(P<0.05) according to this experiment;compared with control group, Simvastatins(0.125μmol/L,0. 25μmol/L,0.5μmol/L,1.0μmol/L )significantly inhibited the proliferation of HPLCs.3,To observe the effects of 50mg/L Emdogain and 0.125μmol/L Simvastatins on the proliferation and differentiation of human periodontal ligament cells.Compared with control group, treated groups can increase the ALP activity and the synethesis of Type- I collagen in HPLCs, but treated groups have little effects on the proliferation of HPLCs.Conclusions1,Emdogain can promote the proliferation,ALP activity,the synthesis of Collagen type I ;2,Simvastain have the same effects on the differentiation of HPLCs,but inhibit the proliferation of HPLCs.3,Their combination have little effects on the proliferation of HPLCs,but significantly stimulate the differentiation of HPLCs.4,All above may give the experiment proofs to the treatment of periodontal disease combinated with cardiovascular disease and diabetes,but their concrete ways,sequence,time and concentration of the medicince in clinical application are worthy of further research.