The Role Of P2X7R/NLRP3 Signaling Pathway In Alcoholic Liver Injury

Alcoholic liver disease is due to long-term,heavy drinking caused by liver damage,the development process for alcoholic fatty liver,alcoholic hepatitis,alcoholic cirrhosis.The incidence of ALD in western countries is on the rise,and the incidence of alcoholic liver disease in China is also increasing,and it has become the second biggest cause after viral hepatitis.ALD has become an important public issue in the world today.Therefore,alcoholic liver injury has attracted more and more attention.Alcohol is mainly absorbed in the upper part of the stomach and small intestine,most of which are metabolized in the liver.Drinking a large amount of alcohol in a short time causes it to be unable to be metabolized in time in the body.It may lead to acute alcoholic liver injury.Recent studies have shown that the main pathogenesis of ALD includes acetaldehyde toxicity,endotoxin injury,oxidative stress injury and inflammatory reaction injury and malnutrition,and the inflammatory reaction plays a key role in the development of ALD.Stimulated by alcohol,activated macrophages in the liver can produce a large number of inflammatory factors,including tumor necrosis factor(TNF-α),interleukin-1 beta(IL-1 beta),and interleukin-18(IL-18),and may be involved in the pathogenesis of ALD.As the pathogenesis of ALD is complex and there is no effective treatment at present,the specific pathogenesis and prevention and cure of ALD have become one of the urgent problems to be solved in the field of liver disease.Purine receptor P2X7 is an ion-type ligand-gated channel receptor,activated by the ligand ATP,can open the ion channel,promote Ca2+、Na+、influx and K+ efflux,and then mediated a series of cell signal transduction,such as NLRP3(an important member of NLR inflammatory family),NF-ΚB、transforming growth factor-beta(TGF-β)and other pathways,induced maturation and release of mature cytokines such as IL-1β,IL-18 and TNF-α,In the inflammatory response and immune regulation plays a very important role.Studies have shown that ATP-P2X7 R is an important signaling pathway to activate NLRP3,and NLRP3 inflammatory corpuscle belongs to the NOD like receptor(nucleotide binding oligomerization domain-like receptors,NLRs),and NLRs is divided into four subtypes.NLRP3 inflammatory bodies are mainly composed of NLRP3,apoptosis related speckle like protein(ASC)and cysteine protein kinase-1(caspase-1).It plays an important role in the development of various inflammatory related liver diseases.Studies have shown that activation of P2X7 receptor is a key factor inducing NLRP3 inflammatory body.Activation of NLRP3 mainly plays a downstream role by secreting IL-1 beta and IL-18.It is also reported that the expression of P2X7 receptor in liver injury tissues is up-regulated,which suggests that P2X7 R may have a close relationship with liver injury,and P2X7R-NLRP3 signaling pathway may also be involved.This experiment uses acute alcoholic liver model to explore the role of P2X7R-NLRP3 signaling pathway in alcoholic liver injury.The main contents are as follows:1.Establishment of liver-induced liver injury modelThe model of acute alcoholic liver injury in mice was established by NIAAA method.30 male C57BL/6 mice were randomly divided into 3 groups(n=10): control group,model group and P2X7 specific blocker A438079 intervention group.The following groups were treated in the last week,the control group and the model group were given equal dose of saline intraperitoneal injection(about 0.2ml / only once a day;group A438079: according to the body weight of mice,the intraperitoneal injection of A438079(A438079 with 7mg/ml by 7mg/ml)was intraperitoneally injected at 200 mu mol/kg,once a day.The last day was given a single 31.5% alcohol solution in the morning,with a dose of 0.2ml/20 g.After 9 h fasting,6 mice in each group were randomly selected to take blood and take the liver by caesarean section.The serum was used to detect the level of ALT,AST,TCHO and triglyceride(TG).The liver tissue was used for histopathological observation and immunohistochemical analysis.2.Expression of P2X7 receptor in alcoholic liver injury modelImmunohistochemical method was used to detect the expression level of P2X7 in liver tissue.The results showed that the expression of P2X7 in normal liver tissues was very low.Compared with the normal control group,the expression of P2X7 protein in the liver tissue of the model group was significantly increased,and the expression of P2X7 protein in the liver tissue of the A438079 group was significantly lower than that in the model group.The results showed that the expression of P2X7 in the alcoholic liver injury model group was significantly higher than that in the A438079 group.3.Role of purine receptor P2X7-mediated NLRP3 signaling pathway in alcoholic liver injury modelThe expressions of P2X7,NLRP3,ASC,TNF-α,IL-1β and IL-18 in liver tissue were detected by Western blot.The results showed that the expression of P2X7,NLRP3,ASC,TNF-α,IL-1β and IL-18 in the model group were significantly higher than those in the control group;Compared with the model group,the expression levels of P2X7,NLRP3,ASC,TNF-α,IL-1β and IL-18 in A438079 group were significantly decreased.It is suggested that P2X7 may be through the activation of NLRP3 inflammatory body,NLRP3 inflammatory body by affecting the secretion of TNF-α,IL-1β,IL-18 play a downstream effect.4.Conclusion In summary,in the model of alcoholic liver injury,the expression of P2X7 and NLRP3 in the model group was significantly enhanced,The expression of A438079,NLRP3,TNF-α,IL-1β and IL-18 were significantly decreased by P2X7receptor-specific inhibitors,can effectively inhibit alcoholic liver injury in inflammatory response,Suggesting that alcohol-induced liver injury may be associated with P2X7R-NLRP3 signaling pathway.