Effects Of Total Glucosides Of Paeony On Proliferation And Wnt Signaling Pathways In Human Fibroblast-like Synoviocyte-osteoarthritis

Objective:To observe the growth and characteristics of human fibroblast-like synoviocytes-osteoarthritis（HFLS-OA）in vitro,and compare the results between tissue culture and enzyme digestion culture,cell identification at last;To investigate the effects of different concentrations of total glucosides of paeony（TGP）on the proliferation of HFLS-OA;To analyze the regulation of TGP to the Wnt signaling pathway in HFLS-OA.Methods:Fresh synovial tissues in patients with osteoarthritis（OA）were cultured by using tissue culture and enzyme digestion culture to obtain HFLS-OA in primary culture;Primary culture cells were subcultured to P₃,then purified FLS marked by Vimentin and CD68 were detected by flow cytometry;HFLS-OA were treated by different concentrations of TGP（312.5ug/ml;625ug/ml;1250ug/ml;2500ug/ml;5000ug/ml）for 24h,then CCK8 was used to investigate the growth of HFLS-OA;After HFLS-OA were treated by different concentrations of TGP（625ug/ml;1250ug/ml;2500ug/ml;5000ug/ml）for 24h,the expression levels of DKK-1、sFRP-1、β-catenin were compared by Western blot.Results:Compared with enzyme digestion culture,FLS with tissue culture grew well and more stable;Primary FLS showed different forms and coexisted with some other cells,howerver,purified FLS were cultured at the generation of P₃ with the progress of culture;Flow cytometry showed that more than 95%P₃ cells were Vimentin positive and CD68 negative,which was in line with the characteristics of FLS;With increasing concentrations of TGP（625ug/ml;1250ug/ml;2500ug/ml;5 000ug/ml）,OD values decreased with statistical significance（p=0.016,p<0.001,p<0.001,p<0.001）,at the lower concentration of TGP（312.5ug/ml）,the OD values had no significant difference compared with the control group;With the change of the concentrations of TGP（625ug/ml;1250ug/ml;2500ug/ml;5000ug/ml）,the expression ofβ-catenin decreased（p<0.001）and the portain expression of DKK-1,SFRP-1 increased（p<0.05）.Conclusions:Tissue culture is a more reliable method to obtain a sufficient amount of purified FLS compared to enzyme digestion culture;TGP can inhibit the proliferation of HFLS-OA with a dose-dependent manner;TGP can regulate the activation of Wnt signaling pathway in HFLS-OA within a certain concentration range.