Effect Of Resolvin D1 On The Autophgy And NLRP3 Inflammasomes Of Experimental Acute Pancreatitis In Mice

Background Acute pancreatitis(AP)is one of the most common acute abdomen diseases,despite significant progress in medical knowledge and treatment,and its incidence and mortality remain high.Increasing studies indicated that autophagy is involved in the onset and progression of acute pancreatitis,implicated in the process of trypsinogen activation and inflammatory response.The basal autophagy is capable to maintain the stability of pancreatic acinar cells,besides,regulating autophagy pathway can reduce the severity of AP to a certain extent.Resolvin D1(Rv D1),an endogenous anti-inflammatory and pro-resolving lipid mediator derived from omega-3polyunsaturated fatty acids,has been shown to have anti-inflammatory effect in a variety of inflammatory diseases.Zhao XW et al found that administration of Resolvin D1 can reduce the level of Inflammatory mediators and finally improve the inflammatory injury in models of AP.Prieto et al previously have shown that Resolvin D1 promote activation of autophagy,autophagosome fusion with lysosomes,and process of autophagic flux in macrophage,finally improve macrophage function and Inflammatory response regression.Objective to analyze the effect of Resolvin D1 on autophagy in AP mice,and further explore the mechanism of Resolvin D1 protective effects in an experimental acute pancreatitis.Methods Adult male C57BL/6 mice were randomly assigned into control group,AP group and Rv D1 group.AP AP group was established by 7 hourly intraperitoneally cerulein,and Rv D1 group was established at 1 hour before and 4 hours during the induction of AP;meanwhile,control group given same volume normal saline.After AP induction completing,pancreatic tissue,lung tissue,and serum were rapidly obtained for analysis.Levels of amylase and lipase in serum,levels of IL-1?、IL-8、TNF-ɑ、IL-6,as well as myeloperoxidase activity in pancreatic and lung tissue were measured.Pathological changes of pancreas and lung were present by H&E staining,and autophagic vacuole were observed by transmission electron microscope.Additionally,autophagic marker were detected in pancreatic tissue using quantitative RT-PCR and Western blot methods.Results AP model mice were successfully established.The serum amylase and lipase levels,relative pancreatic and lung weight were markedly higher than those in Rv D1group(P <0.05).Compared with AP model group,the level of IL-1β,IL-8,TNF-ɑ,IL-6 and the activity of MPO in the lung and pancreatic tissue in Rv D1 group were significantly decreased(P <0.05).there is difference between control group and AP group as more and larger autophagic vacuoles were presented in AP group,contrast to AP group,the number and volume of autophagic vacuoles were cut down in Rv D1group(P <0.05).Correspond to autophagic vacuoles changes,the expression of autophagy-related markers,such as Beclin-1,p62,LC3-Ⅱ,were higher in AP group than control group and Rv D1 group,which reached statistically difference(P <0.05).Conclusion Possible mechanism of Rv D1 improvement of inflammatory injury in experimental acute pancreatitis is associated with its restoration autophgic flux,reduction impaired autophagy and inflammatory mediators.Background Acute pancreatitis(AP)is an inflammatory reaction of the pancreatic exocrine gland that begins with the injury of acinar cells,which can amplify into severe systemic inflammatory response syndrome and multiple organ failure.Inflammation plays an important role in the pathogenesis of acute pancreatitis.In the acute pancreatitis model,the levels of inflammatory mediators in the tissue and serum such as IL-1β,IL-6 and MPO were significantly increased.It’s reported that NLRP3 inflammasomes stimulate and regulate AP inflammatory responses,regulating the synthesis and release of inflammatory mediators in AP.Resolvin D1,derived from w-3polyunsaturated fatty acids,is an endogenous anti-inflammatory proresolving lipid mediator.Resolvin D1 has been shown to improve inflammatory injury in acute pancreatitis models and reduce the release of inflammatory mediators.However,the mechanism of Resolvin D1 protective role in AP is not clear.Yan et al found that omega-3 fatty acids prevent inflammation through the inhibition of NLRP3 inflammasome activation.Resolvin D1 reduces the homocysteine-induced glomerular injury through regulates the activation of NLRP3 inflammasome and inhibits the release of inflammatory mediators.Objective to investigate the effect of Rv D1 on the NLRP3 inflammasome pathway in models of AP and further explore the possible mechanisms of Rv D1 protects against AP.Methods C57BL/6 mice were randomly divided into control group,AP model group and Rv D1 group.AP models were induced by cerulein(50 μg/kg/h)intraperitoneally for seven times.Rv D1(50 ug/kg)was given intraperitoneally to the mice 1 hour before and 4 hours during the induction of AP.Control group mice received same volume of normal salineof normal saline.Colorimetry,pathology,ELISA and other tests to evaluate the role of Resolvin D1 in acute pancreatitis of inflammatory injury.EMSA assay was used to evaluate the effect of Resolvin D1 on NF-κB DNA binding activity in pancreatic tissue.RT-q PCR and Western Blot were used to detect the relative expression of NLRP3,ASC caspase-1 and pro-IL-1β and IL-1β in pancreatic tissues of acute pancreatitis.Results Compared with AP model group,serum amylase and lipase levels,relative weight of pancreas and lung,scores of pathological damage of pancreas and lung,levels of IL-1β,IL-6,IL-8,TNF-ɑ,PGE2 levels and MPO activity in tissues were decreased in Rv D1 group.Inflammatory injury in Rv D1 group was significantly reduced.The difference was statistically significant(P<0.05).Compared with AP model group,the binding activity of NF-κB DNA in pancreatic tissue in Rv D1 group were significantly decreased,with statistical significance(P<0.05).Compared with AP model group,the relative expression of NLRP3 m RNA in Rv D1 group was decreased,the difference was statistically significant(P <0.001).Besides,compared with the AP model group,the relative expression of NLRP3,ASC and caspase-1 protein of NLRP3 inflammasome decreased significantly(P <0.05).In addition,the relative expression levels of pro-IL-1β and IL-1βprotein in NLRP3 inflammasomes in the retreated D1 group decreased significantly(P <0.001).Conclusion The NLRP3 inflammatory pathway is involved in a model of acute pancreatitis,and the mechanism of Rv D1 protects against acute pancreatitis may be related to inhibition of NLRP3 inflammasome pathway acitivation.