Effect Of Dimethyloxalylglycine On Osteogenic Differentiation And Angiogenesis Of Human Periodontal Ligament Stem Cells In Vitro

Objective To investigate the effect of DMOG on osteogenic differentiation and angiogenesis of hPDLSCs in vitro.Methods HPDLSCs were isolated and cultured by tissue block method,and the cell were identified by flow cytometry using surface and immunohistochemistry.HPDLSCs were cultured in basic medium containing 0,0.1,1,10 and 100 μmol/L,MTT assay was used to analyze the effect of DMOG on proliferation and cell viability of hPDLSCs.At the 24th,48th and 72th hours,the total cellular proteins of each group were respectively extracted,the expressions of HIF-la and VEGF protein were detected by Western blot;The mRNA was extracted on day 1,day 3,day 7 and day 14,and the expressions of RUNX2,ALP,OCN and VEGF were detected by RT-PCR.HPDLSCs were cultured in osteogenic medium containing 0,0.1,1,10 and 100 μmol/L DMOG for 14 days,the bone formation in vitro was observed by alkaline phosphatase(ALP)staining and alizarin red staining.The test data were expressed as mean ± standard deviation,using SPSS 19.0 statistical software to analyze the data,P<0.05 was considered statistically significant.Results The hPDLSCs were isolated and cultured by tissue block method.About 7-15 days,cells were observed to crawl out from the tissue.Most of the cells were long fusiform and radially arranged.Flow cytometry showed that hPDLSCs surface antigens such as CD44、CD90、CD 105、CD 146 positive,CD34、CD45 negative,immunohistochemical staining showed that vimentin was positive and keratin was negative,the above result ssuggesting that the cultured cells are mesenchymal stem cells.The results of MTT showed that DMOG could inhibit the proliferation of hPDLSCs in a dose-dependent manner(P<0.05).DMOG could increase the viability of hPDLSCs under the condition of lack of serum(P<0.05).Western blot results showed that the expression of HIF-1α and VEGF protein increased with the increase of DMOG concentration.RT-PCR results showed that DMOG at 10 μmol/L significantly up-regulated RUNX2,ALP and OCN mRNA expression(P<0.05).Alkaline phosphatase and alizarin red staining showed that 10 mol/L DMOG could significantly promote the expression of ALP and the formation of calcium nodules in hPDLSCs osteogenesis.Conclusion DMOG through the upregulation of the expression of HIF-1 alpha,promote the osteogenic differentiation and angiogenesis of hPDLSCs.