Study On The Clinicopathological Features And Pathogenesis Of Alcoholic Chronic Pancreatitis

objectives Alcoholic chronic pancreatitis(ACP)ranks 3rd among all types of pancreatitis in China and is the most prevalent of all types of pancreatitis in Western countries.The incidence of ACP has increased significantly compared to previous years,and ACP is one of the risk factors for the development of pancreatic cancer,which is a serious health risk to people.However,in some cases,the clinical presentation and imaging are not easily distinguishable from pancreatic cancer,and the diagnosis can only be made after surgery and pathological examination.The current treatment for ACP is limited to symptomatic treatment and management of complications,which is unsatisfactory and prone to recurrence.This study aims to address the following key scientific questions:(1)to investigate the correlation between clinically relevant risk factors and the severity of ACP fibrosis through retrospective analysis,and to compare the incidence of ACP comorbidities and the incidence of abnormalities in commonly used clinical assessment indicators between different degrees of fibrosis;(2)to explore the changes in NDRG1 after ethanol activation of PSC and its role in the formation of ACP fibrosis;and(3)To elucidate the upstream and downstream signalling pathways of NDRG1 in the development of pancreatic fibrosis.The resolution of the above scientific questions could provide an important theoretical basis for the clinical translational application of targeting NDRG1,and provide effective therapeutic targets and new strategies for clinical mitigation and reversal of ACP fibrosis.Methods1.Collected cases of chronic pancreatitis clearly diagnosed by pathology at the First Hospital of Jilin University from January 2010 to December 2020,screened cases that met the criteria of alcoholic chronic pancreatitis,and recorded the clinical and pathological data of these patients in detail.The clinical data included patients’ basic information,clinical manifestations,alcohol consumption,duration of alcohol consumption,blood calcium,blood amylase and lipase,etc.The pathological data included the degree of atrophy of the glandular alveoli,the degree of interstitial fibrosis,whether there was pancreatic duct dilatation,pseudocysts and calcification,etc.The data were collated and pooled,and the correlation between the different degrees of fibrosis in ACP and the patients’ age,BMI and history of alcohol consumption was calculated using SPSS analysis software,and the Kruskal-Wallis test for independent samples was used for comparison between groups.In addition,the chi-square(2)test and Fisher’s exact test were used for group comparisons to compare the variability of pancreatitis comorbidities and biochemical tests between the different fibrosis grades.2.h PSC cells were cultured in the laboratory,and the stable cell line h PSC was stimulated with alcohol(50m M)for 1d-6d as the experimental group(activated state),and the control group(resting state)was set up.PCR and Deep Sequencing were used to cluster and analyse the NDRG1 m RNA expression level.3.The stable cell line h PSC was stimulated with alcohol(50m M)for 1d-6d as the experimental group(activated state),and the control group(resting state)was set up;the differences in the expression of NDRG1 total protein levels and phosphorylated forms(Ser330,Thr346),and the expression of h PSC activation indexes(α-SMA,TGF-β)were detected by Western blot.4.A control group(control si RNA)was set up,and changes in the expression of MAPK pathway indicators(ERK,JNK),EMT-related indicators(Snail,Slug,Vimentin),and cell activation-related indicators(α-SMA,TGF-β)were detected by Western blot after knockdown of NDRG1 using si RNA.5.hPSC cells were treated with specific inhibitors of JNK and ERK in the presence/absence of alcohol stimulation,and NDRG1 and EMT-related proteins were detected by Western blot to explore the effects of ERK and JNK pathways on NDRG1 and EMT-related proteins.6.Immunohistochemical staining of pancreatic tissues from ACP patients was performed,and the slices were read to observe whether there were differences in the expression of NDRG1 and its phosphorylated forms(Ser330 and Thr346)between normal tissues and fibrotic areas.Results1.The statistical results of the clinical data showed statistically significant differences(P < 0.05)in the average daily alcohol consumption,duration of alcohol consumption,total alcohol consumption,incidence of pancreatic duct dilatation,decreased blood calcium and elevated blood amylase between the different fibrosis grades of pancreatic tissue,while the differences in age,BMI,calcification,incidence of pseudocysts and elevated lipase were not statistically significant(P > 0.05).2.NDRG1 m RNA expression was progressively lower in hPSC receiving alcohol(50m M)stimulation for 48 h and 72 h compared to controls;consistent results were obtained by Real-time PCR with statistically significant differences(P<0.001).3.Western blot showed that the expression levels of both NDRG1 total protein and its phosphorylated form(Ser330,Thr346)were down-regulated,while the expression of h PSC activation indicators(α-SMA,TGF-β)was up-regulated after h PSC were stimulated by ethanol(50m M)for 1d-6d.4.The expression of MAPK pathway indicators(ERK,JNK),EMT-related indicators(Snail,Slug,Vimentin)and cell activation-related indicators(α-SMA,TGF-β)were all up-regulated after si RNA knockdown of NDRG1 compared to the control group by Western blot.5.In the presence/absence of alcohol stimulation,the ERK inhibitor UO126 and the JNK inhibitor SP600125 did not regulate NDRG1 expression,but down-regulated the EMT-related proteins Snail,Slug and Vimentin.6.Immunohistochemical staining showed that NDRG1 and its phosphorylated forms(Ser330 and Thr346)were reduced and diminished in expression in fibrotic areas compared to normal pancreatic tissue.Conclusions1.Long-term chronic intake of alcohol largely increases the risk of ACP,and the duration of alcohol consumption and total alcohol intake are positively correlated with the degree of fibrosis in chronic pancreatitis.2.During the formation of fibrosis in ACP pancreatic tissue,NDRG1 is negatively correlated with the activation of h PSC and inhibits the EMT process through the MAPK pathway,thereby inhibiting the formation of fibrosis.