The Effect Of IL-1? In LPS-Induced Inflammation In Goat Temporomandibular Joint Disc Cells

Objective: Lipopolysaccharide(LPS)was applied to induce inflammation and simulate the inflammatory environment on the goat temporomandibular joint(TMJ)disc cells.We could explore the effects of inflammatory environment on goat TMJ disc cells.The role of Interleukin-1?(IL-1?)in the inflammatory response,and the effect of IL-1? on goat TMJ articular cells and extracellular matrix.Methods:First,the LPS concentration screening experiment: Goat TMJ disc tissue was separated in vitro and goat TMJ disc cells were cultured.MTT was used to detect the proliferation of goat TMJ disc cells with different concentrations of LPS(0,0.1,1,10,100?g/mL)The survival state of goat TMJ cells after dosing culture was observed by trypan blue staining.The expression level of IL-1? in each concentration group was detected by qRT-PCR.ELISA was used to detect the secretion levels of IL-6 and TNF-?.After staining with toluidine blue and Sirius red,the expression of glycosaminoglycan and total collagen in the extracellular matrix of goat TMJ joint disc was observed.Second,the effect of IL-1? on goat TMJ disc cells and cell-extracellular matrix after LPS inflammation induction: According to the experimental requirements,the experimental components were: blank control group,LPS culture group,IL-1? culture group,LPS + IL-1? culture group.MTT assay is for the proliferation ability of goat TMJ disc cells.The secretion levels of IL-6 and TNF-? in each experimental group were measured by ELISA.Immunohistochemical staining of toluidine blue,Sirius red and type I collagen was used to observe the expression of glycosaminoglycan,total collagen and type I collagen in the extracellular matrix of goat TMJ disc cells in each group.The expression of related gene mRNA in each group was detected by qRT-PCR.Results: Microscopically,the morphology of goat TMJ articular disc was long fusiform and polygonal.Immunohistochemical staining of type I collagen and toluidine blue staining showed positive results.The MTT results showed that LPS(10?g/mL)showed a greater decrease in cell viability than the control group after 24h(p<0.05).The results of trypan blue staining showed that the cell membrane of LPS(10?g/mL)group partially translucent,the membrane continuity decreased,and most of the cells survived.Small concentration of LPS(0,5,10,15?g/mL)MTT assay showed that LPS inhibited the proliferation of goat TMJ joint disc cells after 24 h.qRT-PCR showed that the expression level of IL-1? mRNA was up-regulated compared with the control group(p<0.05).The ELISA showed a significant increase in the secretion of IL-6 and TNF-? in the LPS(10?g/mL)group compared with the control group(p<0.05).Toluidine blue and Sirius red staining of the LPS(10?g/mL)showed significant positive results compared with the control group.In summary,LPS(10?g/mL)was selected for 24 h as a suitable experimental concentration and time.LPS and IL-1? acted on goat TMJ disc cells and the cells viability of the LPS+IL-1? group was significantly lower than that of the control group(p<0.05).ELISA showed that the levels of IL-6 and TNF-? in the LPS+IL-1? group was significantly higher than those in the control group(p<0.05).Toluidine blue staining,Sirius red staining,and immunohistochemical staining of type I collagen showed that the positive expression of staining was decreased in the LPS+IL-1? group compared with the control group.qRT-PCR showed a decrease in the expression level of type I collagen in the LPS+IL-1? group compared with the control group.The expression of Aggrecan protein in the LPS+IL-1? group was decreased compared with the control group.Compared with the control group,the expression levels of MMP-1,MMP-3 and MMP-13 in LPS+IL-1? group were significantly up-regulated(p<0.05).Conclusions: 1.LPS induced the production of IL-1? and other inflammatory factors in the inflammation,and the inflammatory response did have effect on the morphology and proliferation of goat TMJ disc cells.2.LPS-induced inflammation addition of IL-1? could promote the inflammatory reaction,inhibit the expression of collagen and proteoglycan in the extracellular matrix of the goat TMJ disc cells,and promote the expression of MMP-1,MMP-3 and MMP-13 in the inflammatory reaction process.