Study On The Mechanism Of DNA Polymerase ? Regulating Rheumatoid Arthritis By Inhibiting Macrophage Pyroptosis

Rheumatoid arthritis(RA)is a chronic and progressive autoimmune disease,which is characterized by erosive synovitis and joint lesions,and is known as"immortal cancer".The main manifestations of RA patients are joint pain,swelling and irreversible joint deformity,which seriously affect the quality of life of patients.However,the pathogenesis of RA is complex and has not been fully elucidated.Therefore,it is of great scientific value and clinical significance to study the new mechanism of the occurrence and development of RA and provide a new theoretical basis for the treatment of RA and the development of a new generation of anti RA drugs.Inflammatory response is an important feature of RA,and it is also an important factor of bone destruction in RA.Macrophages play an important role in the inflammatory response of RA.There are a lot of macrophages in cartilage and synovial tissue of RA patients;At the same time,the number of macrophages has a certain correlation with the degree of joint injury and clinical symptoms.Macrophages can recruit and activate other immune cells,and can produce TNF,IL-1,IL-6 and other pro inflammatory cytokines to further initiate inflammatory response.The activation of inflammatory response is closely related to the inflammatory bodies of nod like receptor protein 3(NLRP3)in macrophages.NLRP3 inflammasome has the biological function of recognizing pathogens,can sense the changes of the internal environment,and activate a unique programmed cell death pathway-pyroptosis.Its continuous activation can increase the expression level of pro-inflammatory factors,lead to the occurrence of chronic inflammatory diseases,and play an induction or promotion role in the pathogenesis of RA.Base excision repair(BER)is an important DNA damage repair pathway,which is mainly responsible for the repair of genomic DNA base damage and single strand breaks.DNA polymerase?(Pol?)is a key enzyme in the base excision repair pathway.It is widely present in the nucleus of mammalian cells and participates in the repair of DNA oxidative damage and maintains genome stability.The loss of Pol?can lead to accumulation of DNA damage and genome instability,thereby inducing tissue inflammation and autoimmune diseases.However,the relationship between the functional defects of Pol?and RA is still unclear.In this work,we explored the regulatory effects of Pol?on the development and treatment of RA in vitro and in vivo.To assess whether Pol?is involved in RA formation,we compared the expression of Pol?in CD4~+T cells from active RA patients and healthy donors using the GEO database(GSE56649).The levels of Pol?were decreased in CD4~+T cells from active RA patients compared with those in healthy donors.At the same time,based on the successful construction of a mouse model of collagen-induced arthritis induced by chicken type II collagen(CIA mouse model),it was showed that the expression of Pol?were significantly decreased in CIA mice compared with normal control mice measured by RT-q PCR and Western blot.Subsequently,through animal experiments,it was found that knocking down the Pol?gene in mice or mutating the active site of the Pol?gene increased the incidence of CIA mice.Histopathological analysis also showed that inflammatory cell infiltration and bone destruction were more serious in the ankle joints of Pol?-deficient mice.In vitro cell experiments found that knocking down Pol?can exacerbate the pyroptosis macrophages induced by LPS combined with ATP,while overexpression of Pol?can inhibit this process.NLRP3-Caspase-1-IL-1?is an important molecular axis in cell pyroptosis and plays an important role in inflammatory response.Through RT-q PCR,Western blot and immunofluorescence experiments,it was found that the expression of pyroptosis-related proteins were up-regulated in Pol?knockdown cells;overexpression of Pol?can effectively reverse this phenomenon.At the same time,our experiments showed that the DNA repair ability of macrophages was reduced after knocking down Pol?,resulting in the accumulation of DNA damage intermediates,and ds DNA leaks from the nucleus into the cytoplasm,activating the c GAS-STING-NF-?B signaling pathway.In the complex inflammatory network mechanism,activated NF-?B can up-regulate the expression of NLRP3,promote the activation of NLRP3 inflammasomes and cause a strong immune response.In summary,this study clarified the influence of Pol?on the formation of RA and the molecular mechanism by which Pol?regulates the formation of RA via inhibiting macrophage pyroptosis.