The Mechanism Research Of Macrophage-like Synovial Cells Activating KOA Hyperalgesia And The Treatment Effects Of "Yiceng" External Therapy

ObjectiveTo find out whether there is macrophage pyroptosis in the synovium of KOA,and to find its internal relationship with synovium inflammation,cartilage degeneration,and pain sensitivity.Based on macrophage pyroptosis/TRPs signal pathway,to study the therapeutic mechanism of"easy layer" application to effectively inhibit KOA pain.Methods(1)Establishing the KOA rat model,HE staining,immunofluorescence,Western blot and other experimental methods were used to determine whether there is macrophage pyroptosis and the relationship with synovial inflammation and cartilage degeneration;(2)Establishing the KOA rat model,the effects of synovial macrophage pyroptosis on TRPs protein expression and pain sensitivity were determined by detecting the parameters of cold and mechanical pain sensitivity,the expression of TRPs channel protein and gene in DRG;(3)We constructed the model of macrophage scorching in vitro to investigate the effect of macrophage pyroptosis on the expression and function of TRPs channel protein;(4)Establishing the KOA rat model,the therapeutic mechanism of "easy layer" application was explored by detecting the parameters of cold and mechanical pain sensitivity,the expression of TRPs channel protein and gene in DRG;(5)Establishing the KOA rat model,HE staining,immunofluorescence,Western blot and other experimental methods were used to observe the effect of "easy layer" application on the synovial macrophage pyroptosis.Results(1)In the KOA model group,typical synovial inflammation,cartilage degeneration,and high expression of macrophage pyroptosis related protein were found in synovium;(2)In KOA model group,the rats showed typical cold and mechanical pain sensitive state;after the intervention of macrophage specific caspase-1 low expression adeno-associated virus,the pain sensitive state of the rats was significantly improved,the latency of cold stimulation foot contraction was longer,and the threshold of mechanical stimulation foot contraction was significantly higher than that of the model group;the protein and gene expression of TRPA1,TRPM8,TRPV4 in dorsal root ganglion tissue were down regulated compared with the model group;(3)LPS/ATP was used to interfere with macrophages in vitro,and the model of pyroptosis was successfully constructed.The medium of macrophages was used to interfere with neuron cells,and the effect of LPS/ATP on TRPs channel of neuron cell membrane was observed.It was found that macrophage pyrolytic products could significantly up regulate the expression of TRPs protein and gene,and this effect was eliminated after the inhibition of pyrolytic;the intracellular calcium concentration was observed by neuronal calcium imaging technology,and it was found that macrophage pyrolytic products could significantly increase the intracellular calcium concentration,and inhibition of pyrolytic could reduce the intracellular calcium concentration;(4)The results showed that the application of "easy layer" could improve the cold and mechanical pain sensitivity of KOA rats,prolong the latent period of cold stimulation and increase the threshold of mechanical stimulation;(5)"Easy layer" application can reduce the high expression of macrophage pyroptosis related protein in the synovium of KOA rats,improve synovium inflammation and reduce the level of pro-inflammatory factors in serumConclusions(1)There was macrophage pyroptosis in the synovium of KOA,which promoted synovium inflammation and cartilage destruction;(2)KOA synovium macrophage pyroptosis,releases pro-inflammatory factors,activates TRPs channels,and constructs cold and mechanical pain sensitive states;(3)The application of "easy layer" can effectively improve the cold and mechanical pain sensitivity of KOA;(4)The application of "easy layer" exerts the therapeutic effect of inhibiting pain by intervening macrophage pyroptosis/TRPs signal pathway.