Effects Of IL-17 And Human Periodontal Ligament Fibroblasts On The Production Of Osteoblast-like Cells In Vitro

Ojective:Root resorption is a common complication in orthodontic treatment,whereas IL-17 and periodontal ligament fibroblasts may induce osteoporosis differentiation and activation by inducing the formation of osteoclasts.In this study,we investigated the effect of IL-17 and periodontal ligament fibroblasts on the biological behavior of peripheral blood mononuclear cells.The effects of different concentrations of IL-17 on periodontal ligament fibroblast conditioned medium and RANKL were studied at different time And the expression of CTSK,MMP-9 and CA II mRNA in PBMC,and the morphological and characteristic markers of osteoclast-like cells were observed under co-culture system.The effects of IL-17 and periodontal ligament fibroblasts on orthodontic root resorption Possible role and its mechanism of action.Methods:The human peripheral blood mononuclear cells were cultured by density gradient centrifugation method.The cells were grown in 48-well plates and the growth state was observed by inverted microscope.The periodontal ligament tissue was cultured by tissue culture.The indirect co-culture system of HPDLF and PBMC was established.2.Take the well-grown HPDLF from the 4-8 generation,add the prepared DMEM medium with IL-17 concentration of 0 or 20ng/ml for 42h,and use the DMEM medium containing 10%FBS for 12 hours,Collecting HPDLF cell supernatant,adding 50ng/ml RANKL as conditioned medium;The expression of CTSK,MMP-9 and CA ? mRNA in osteoblasts induced by peripheral blood mononuclear cells were detected by RT-qPCR in 3 days,5 days,7 days and 10 days respectively with different HPDLF conditioned medium.Expression level;4.Different conditions of culture medium on the role of peripheral blood mononuclear cells,indirect co-culture induced osteoclast-like cells for 14d,phalloidin staining and count osteoclast-like cell cytoskeleton of fibrous actin ring morphology,number.Results:1.Cultured peripheral blood mononuclear cells are round,loose adherent growth,as osteoclast precursor cells.Periodontal ligament fibroblasts are long spindle,radial or swirl-like growth,stable growth state.2.Compared with the control group,the expression of CTSK,MMP-9 and CA II mRNA was up-regulated by IL-17(+)conditioned medium and RANKL in PBMC under RT-qPCR detection.(P<0.05).The expression of IL-17(+),RANKL(+)and 10d was the most significant difference between the two groups(P<0.05).3.IL-17(+)and RANKL(+)and IL-17(-),RANKL(-),and IL-17(-)were treated with monoclonal antibody(P<0.01),and the number of osteoclast-like cells with fibrous actin ring morphology was statistically significant(P<0.01).Conclusion:1.The indirect co-culture system of human peripheral blood mononuclear cells and human periodontal ligament fibroblasts was successfully established by density gradient centrifugation' and tissue culture.The biological characteristics were stable.The appropriate concentration of IL-17(+)Conditioned medium and RANKL could induce PBMC to produce osteoclast-like cells expressing CTSK,MMP-9 and CA ? mRNA.2.IL-17 can promote the expression of CTSK,MMP-9 and CA ? mRNA in periodontal ligament fibroblasts conditioned medium and RANKL.3.Human peripheral blood mononuclear cells can differentiate into osteoclast-like cells with fibrous actin ring morphology under the direct co-culture of human periodontal ligament fibroblasts.4.IL-17 and human periodontal ligament fibroblasts promote the formation of osteoclast-like cells,the mechanism may be through human periodontal ligament fibroblasts expressing RANKL in human peripheral blood mononuclear cells.