The In Vivo And In Vitro Study Of SCAPs/PGA Combined With CGF Repairing Periodontal Tissue Defect

Objectives:To observe the basic biological characteristics of SCAPs and compare the potential of autologous SCAPs/PGA combined with CGF to promote acute one-wall defects of periodontal regeneration in beagle dogs.Method:1.In vitro study: SCAPs and PDLSCs were isolated and cultured by enzyme digestion;SCAPs proliferation rate was detected by CCK8 and monoclonal formation rate was calculated;SCAPs surface markers were identified by flow cytometry: Stro-1,CD146 and CD45;differentiation potential of SCAPs into osteogenesis?adipogenic and cartilage were tested.The cells were inoculated and observed in linearly aligned polyglycolic acid PGA scaffold material under static tension for 2 weeks.2.In vivo study: SCAPs/PGA,PDLSCs/PGA(positive control)combined with or without CGF cultured for two weeks under static tension in vitro,and then cell constructs were implanted into one-wall intrabony defect model.After 8 weeks,the specimens were fixed,decalcified,and dehydrated.After paraffin sectioning,HE staining and Azan staining were performed to observe the histologic healing of periodontal tissues.Results:1.Isolation,culture and identification of SCAPs: It can be observed that the cells are clonal-like adherent growth,and the colony formation rate was(8.99±2.44)%.The growth curve was consistent with the logarithmic growth curve.Flow cytometry identification of SCAPs: Stro-1 was(11.12±3.09)%,CD146 was(10.57±2.22)%,hematopoietic stem cell marker CD45 is not expressed;multi-directional differentiation ability detection proved that SCAPs can form calcifying nodules and cartilage matrix,and lipid droplets can be observed.When SCAPs were inoculated on the PGA scaffold for two weeks,under the inverted phase contrast microscope,most of the SCAPs were observed to adhere to the aligned PGA fibers and the bottom of the dish,and grew along the fiber arrangement,secreting a large amount of extracellular matrix attached the PGA.Thus a tissue engineered ligament-like structure was obtained.2.After 8 weeks of surgery,histological observation of the wound area formed a good connective tissue adhesion,the experimental group defect can be seen new bone formation above the notch,The control group has less new bone mass.Undegraded materials can still be seen in most of the periodontal wound healing area,and there are dense fibers around the bone.The inner surface of the experimental group can be seen abundant connecting fibers extending into the connective tissue.Rich collagen fiber formation can be seen between acellular cementum and new alveolar bone.Histological measurements showed that the amount of connective tissue adhesion in SCAPs/PGA/CGF,SCAPs/PGA,PDLSCs/PGA/CGF,PDLSCs/PGA groups is less than that of control group(p<0.05);new bone area: SCAPs/PGA/CGF and control group were also significantly different;new bone height: SCAPs/PGA/CGF,SCAPs/PGA,PDLSCs/PGA/CGF,PDLSCs/PGA were slightly higher than the control group,but p >0.05.Conclusion:The SCAPs/PGA complex can form an engineered ligament structure in vitro,it has good biocompatibility and/or CGF and has the potential to promote periodontal tissue regeneration.