Primary toxicity targets of alcohol and its metabolites in the pancreas are cellular energetics and endoplasmic reticulum (ER). Therefore, the role of AMP-Activated Protein Kinase (AMPKα) in amelioration of ethanol (EtOH)-induced pancreatic acinar cell injury including ER/oxidative stress, inflammatory responses, the formation of fatty acid ethyl esters (FAEEs) and mitochondrial bioenergetics were determined in human pancreatic acinar cells (hPACs) and AR42J cells incubated with/without AMPKα activator [5-aminoimidazole-4-carboxamide ribonucleotide (AICAR)]. EtOH treated hPACs showed concentration and time-dependent increases for FAEEs and inactivation of AMPKα, along with the upregulation of ACC1 and FAS (key lipogenic proteins) and downregulation of CPT1A (involved β-oxidation of fatty acids). These cells also showed significant ER stress as evidenced by the increased expression for GRP78, IRE1α, and PERK/CHOP arm of unfolded protein response promoting apoptosis and activating p-JNK1/2 and p-ERK1/2 with increased secretion of cytokines. AR42J cells treated with EtOH showed increased oxidative stress, impaired mitochondrial biogenesis, and decreased ATP production rate. However, AMPKα activation by AICAR attenuated EtOH-induced ER/oxidative stress, lipogenesis, and inflammatory responses as well as the formation of FAEEs and restored mitochondrial function in *

The single most common cause of chronic pancreatitis (CP, a serious inflammatory disease) is chronic alcohol abuse, which impairs hepatic alcohol dehydrogenase (ADH, a major ethanol oxidizing enzyme). Previously, we found ~5 fold greater fatty acid ethyl esters (FAEEs), and injury in the pancreas of hepatic ADH deficient (ADH) vs. hepatic normal ADH (ADH) deer mice fed 3.5g% ethanol via liquid diet daily for two months. Therefore, progression of ethanol-induced pancreatic injury was determined in ADH deer mice fed ethanol for four months to delineate the mechanism and metabolic basis of alcoholic chronic pancreatitis (ACP). In addition to a substantially increased blood alcohol concentration and plasma FAEEs, significant degenerative changes, including atrophy and loss of acinar cells in some areas, ultrastructural changes evident by such features as swelling and disintegration of endoplasmic reticulum (ER) cisternae and ER stress were observed in the pancreas of ethanol-fed ADH deer mice vs. ADH deer mice. These changes are consistent with noted increases in pancreatic injury markers (plasma lipase, pancreatic trypsinogen activation peptide, FAEE synthase and cathepsin B) in ethanol-fed ADH deer mice. Most importantly, an increased levels of pancreatic glucose regulated protein (GRP) 78 (a prominent ER stress marker) were found to be closely associated with increased phosphorylated eukaryotic initiation factor (eIF) 2α signaling molecule in PKR-like ER kinase branch of unfolded protein response (UPR) as compared to X box binding protein 1S and activating transcription factor (ATF)6 - 50kDa protein of inositol requiring enzyme 1α and ATF6 branches of UPR, respectively, in ethanol-fed ADH vs. ADH deer mice. These results along with findings on plasma FAEEs, and pancreatic histology and injury markers suggest a metabolic basis of ethanol-induced pancreatic injury, and provide new avenues to understand metabolic basis and molecular mechanism of ACP.

Background Chronic alcohol abuse, a major risk factor for such diseases as hepatitis and cirrhosis, impairs hepatic alcohol dehydrogenase (ADH, key ethanol metabolizing enzyme). Therefore, differentially altered hepatic and plasma proteomes were identified in chronic ethanol feeding model of hepatic ADH-deficient (ADH−) deer mice to understand the metabolic basis of alcoholic liver disease (ALD). Methods ADH− deer mice were fed 3.5g% ethanol via Lieber-DeCarli liquid diet daily for three months, and histology of the liver assessed. Liver and plasma proteins were separated by 2-dimensional gel electrophoresis. The proteins differentially expressed were identified by MALDI-TOF/TOF mass spectrometry (MS). Results Histology of the liver showed panlobular steatosis and infiltration of T-lymphocytes. Using the criteria of ≥1.5 for fold change (p value ≤0.05) with expectation value (E ≤10−3) and protein score (≥64), eighteen proteins in the livers and five in the plasma of ethanol-fed mice were differentially expressed and identified. Prolyl 4-hydroxylase, cytochrome b-5, endo A cytokeratin, ATP synthase, heat shock 70kD proteins, enoyl CoA hydratase, stress-70 protein, peroxiredoxin 1 and ornithine carbamoyl-transferase were up regulated in the livers. However, carbonic anhydrase 3, mitochondrial ATP synthase, aldolase 2, actin γ, laminin receptor and carbamoyl-phosphate synthase were down regulated. Contrary to the increased expression of creatine kinase m-type, a decreased expression of serine protease inhibitor A3A precursor, sulfated glycoprotein-2 (clusterin) and apolipoprotein E isoforms were found in the plasma of ethanol group. Conclusions Chronic ethanol feeding in ADH− deer mice causes steatosis and infiltration of T-lymphocytes in the livers along with increased expression of proteins involved in ER stress, fibrosis, fatty acid β oxidation and biogenesis, and decreased expression of proteins involved in ATP synthesis, carbohydrate metabolism, in cell regulation and architecture. Reduced expression of various carrier proteins as found in the plasma of ethanol group has a biomarker potential.