Construction Of NICD Overexpression Vector And Its Effect On Proliferation Of Human Periodontal Ligament Stem Cells

Objective: To construct periodontal ligament stem cells(PDLSCs)stably overexpressing Notch1 intracellular domain(NICD)and investigate the effect of overexpressing NICD gene on the proliferation of human periodontal ligament stem cells(PDLSCs).Methods: the recovery of ourprevious cryopreserved human periodontal ligament stem cells were subcultured for 3~5 generation experiment.The retroviral particles containing NICD gene overexpression were constructed and transfected into PDLSCs.Puromycin screening was used to construct PDLSCs/NICD cell line overexpressing NICD.PDLSCs/NICD were used as experimental group,PDLSCs/wt as normal cell group and PDLSCs/vector empty virus transfection group as control group.Quantitative Real time-polymerase chain reaction(qRT-PCR)and Western blot were used to detect the expression of NICD gene and protein;CCK-8 method was used to detect the proliferation of cells;Transwell test was used to detect the migration ability of cells.Results: Western blot and qRT-PCR results showed that compared with PDLSCs/wt group and PDLSCs/vectors group,the expression level of NICD protein and its mRNA in PDLSCs/NICD group were significantly higher(P<0.05);CCK-8 results indicated that compared with PDLSCs/vectors group,the proliferation rate of PDLSCs/NICD group was faster,and Transwell results showed that the cell migration rate in PDLSCs/NICD group was significantly higher(40.20±1.74)than that in PDLSCs/vectors group(21.20±1.18).Conclusion: The proliferation and migration of PDLSCs overexpressed by NICD gene are enhanced to some extent.