Study On Mechanism Of JAK2/STAT3 Pathway In Hypertriglyceridemia Induced Acute Pancreatitis

Part OneHigh-Dose Linoleic Acid Activated JAK2-STAT3 Signaling Pathway inPancreatic Exocrine CellsObjectivesIn China,with the improvement of living standards and changes in diet structures,the incidence of hypertriglyceridemia induced acute pancreatitis(HTGP)was increasing.This project explored the role of JAK2/STAT3 signaling pathway in the pathogenesis of HTGP from cell level.Impact of free fatty acid(FFA)on JAK2/STAT3 pathway and cytokines' expression are also explored when AG490 are applied preliminarily.MethodsTaking rats pancreatic exocrine cells of AR42 J as the research object,the study explored the different concentrations(0mmol/L,0.04mmol/L,0.4mmol/L,4mmol/L)of linoleic acid(LA)on the proliferation and cytotoxicity of AR42 J cells.AR42 J cells were treated with LA in 0h,2h,4h,8h,12 h and isolated cytoplasmic and nuclear protein,then detected the expression of JAK2,p-JAK2,STAT3,p-STAT3 protein at different time points by Western blot(WB)and tested concentrations of p-STAT3 protein in the cytoplasm and nucleus in AR42 J cells.AR42 J cells were divided into five groups: control group(no drug),AG490 group(80 mol/L AG490 5H),AG490+LA group(80?mol/L AG490 pretreatment 2H adding 0.4mmol/L LA 3H),LA group(0.4mmol/L LA 3H),LA+AG490 group(0.4mmol/L LA 1H then adding 80?mol/L AG490 2H).Total protein were collected from the AR42 J cells and detected by WB.Detection of inflammatory cytokines(IL-6,TNF-?)was based on ELISA method,which is to explore the effect of LA on JAK2/STAT3 signaling pathway in AR42 J cells after AG490 inhibited JAK2 inhibition.ResultsThe cell viability of AR42 J rat pancreatic exocrine cells was inhibited,and cytotoxicity was increased by high-dose linoleic acid(0.4mmol/L).JAK2 and STAT3 proteins in pancreatic exocrine cells were activated by high-dose linoleic acid via phosphorylation and nuclear localization of phosphorylated STAT3.When combined the LA and AG490,the expression of p-JAK2 was depended on the different adding sequence.When the AG490 inhibited the phosphorylation of JAK2 in advance,the expression of p-JAK2 was down-regulated regardless of whether applying LA or not.Once the expression of p-JAK2 induced by LA,the concentrations of p-JAK2 increased regardless of whether treated with the AG490 or not.But the expression of p-STAT3 protein in two separate applications were significantly increased.Moreover,the expression of downstream proteins in JAK2-STAT3 pathway(IL-6 and TNF-?)was up-regulated by high-dose linoleic acid.The increased levels of IL-6 and TNF-? caused by high-dose linoleic acid were attenuated by JAK2 inhibitor AG490.ConclusionWe concluded that the cytotoxicity was increased and JAK2-STAT3 signaling pathway was activated by high-dose LA including cytokine production in rat pancreatic exocrine cells.This effect can be partly reversed by AG490.Part Two Circulating Levels of Molecules Associated with JAK2/STAT3 Signaling Pathway in Patients with Acute PancreatitisObjective To investigate the differences and correlation of serum free fatty acid(FFA),leptin,p-JAK2 and p-STAT3 in patients with acute pancreatitis(AP)and illustrate the possible pathogenesis of this disease.Methods A total of 34 adult patients with AP in Shanghai General Hospital from July 2015 to March 2016 were enrolled.10 volunteers formed normal control(NC)group.Serum leptin,p-JAK2 and p-STAT3 concentration were determined by enzyme-linked immunosorbent assay(ELISA)and FFA by microcolorimetry.Differences of serum FFA,leptin,p-JAK2 and p-STAT3 concentration were analysed between two groups.According to scores of the modified CT severity index(MCTSI),there were 18 nonsevere AP and 16 severe AP.The differences of serum FFA,p-JAK2 and p-STAT3 concentration in the subgroups with MCTSI scores <4 and ?4.The relevant factors affecting the serum p-STAT3 level were also evaluated.ResultsThe level of leptin,FFA,p-JAK2 and p-STAT3 was significantly increased in AP group compared to NC group(P<0.05).There were no difference of serum FFA,leptin,p-JAK2 and p-STAT3 between two subgroups.On univariate analysis,FFA and p-JAK2 level were signifcantly associated with the concentration of p-STAT3.However,on multivariate analysis,the FFA and p-JAK2 level were found to be the independent correlation factors for the level of p-STAT3(R2= 0.6831,P<0.001).ConclusionsThe JAK2/STAT3 Signal Pathway is activated in patients with AP,which is possible related to the effect of FFA and leptin and has no correlation with the severity of AP.