Effect Of Platelet-rich Fibrin Extract On The Proliferation Of Gingival Fibroblasts

Objective The platelet-rich fibrin was transformed into platelet-rich fibrin extract to study the optimal concentration of platelet-rich fibrin extract to promote the proliferation and differentiation of gingival fibroblasts,and to observe the effect of PDGF growth factor released by platelet-rich fibrin extract on the proliferation activity of gingival fibroblasts,so as to provide theoretical basis for its application in assisting the enhancement of gingival soft tissue..Methods From November 2017 to June 2018,patients in oral and maxillofacial surgery of Anhui Stomatological Hospital were selected.Some gingival soft tissues needed to be removed for extraction of impacted teeth.Routine examinations were performed before removal.After screening for inclusion and exclusion criteria,a small amount of gingival soft tissue removed during surgery was immediately placed in 4 C culture medium.Then gingival fibroblasts were isolated and cultured by tissue culture method.Cell morphology was observed under inverted microscope and identified by immunofluorescence cytochemistry.Then platelet-rich fibrin(PRF)was prepared and extracted,and platelet-rich fibrin(PRF)was transformed into platelet-rich fibrin extract(PRFe).The three-dimensional structure of PRF was observed by electron microscopy and the content of PDGF in PRF was quantitatively determined by ELISA.Then the PRFe ratio was 2.5% PRFe,5% PRFe,7.5% PRFe,10% PRFe,12.5% PRFe and 15%PRFe,respectively,and the stimulation of each group was inoculated into gingival fibroblasts.On the first,third and fifth day after inoculation,the effects of PRFe at different concentrations on the proliferation of gingival fibroblasts were detected by CCK8 method to determine the optimal PRFe concentration.Then the blank control group,the best PRFe concentration group and the best PRFe concentration group were added to the platelet-derived growth factor antibody group.CCK8 was detected on the first day,the third day and the fifth day,respectively.The effects of PDGF growth factor released by PRFE and PFRE on the proliferation activity of gingival fibroblasts were observed.Results 1.The morphology of cultured gingival fibroblasts was spindle-shaped and the border was clear.Immunofluorescence histochemical results showed that the anti-Vimentin staining was positive in all 8 groups.Microscopically,the oval nuclei of spindle cells were homogeneous blue stained,while the cytoplasm was red-brown.The anti-Cytokeratin staining was negative in all 8 groups.Microscopically,the nuclei were blue and the cytoplasm was opaque.2.The main body of PRF was pale yellow fibrin gel and a few red cell layers below it.There were a lot of growth factors at the junction of red and yellow.Scanning electron microscopy showed that platelet rich fibrin arranged in a rattan like three dimensional network structure.The results of ELISA detection showed that the release of PDGF was detected at different time points within 11 days,and PRF was higher in the first hour after preparation,and had a burst effect.It should decrease gradually after the first day,then increase slowly to reach the peak value on the 7th day,close to the release value of the first hour.3.The proliferative activity of HGFs was tested with different concentrations of PRFe.The results showed that HGFs had a concentration dependence on PRFe on the first day,the third day and the fifth day after inoculation with 2.5% PRFe,5% PRFe,7.5% PRFe,10% PRFe,12.5% PRFe and 15% PRFe,but the effect was the best at the concentration of 5% PRFe(P < 0.05).There was no significant difference in the effect of subsequent concentration increase on the proliferation of HGFs(P > 0.05).PDGF neutralization and experimental results showed that HGFs had a concentration dependence on PRFe.The proliferation of gingival fibroblasts was significantly inhibited by PDGF(P < 0.05).Conclusion PRF is a three-dimensional reticular structure,which contains a large number of growth factors.PDGF release has a sudden release effect,reaching its maximum at the 7th day,while PRFe at 5% concentration has the best effect in promoting gingival fibroblasts proliferation in vitro.PDGF released from PRF plays an important role in promoting gingival fibroblasts proliferation,which provides a mechanism for PRF to assist in promoting gingival soft tissue regeneration.