Effect of Alcohol on Pancreas

Pooja Goswami

AI.I.M.S. New Delhi

Point to be covered

• Pancreas normal physiology and anatomy

• Exocrine & endocrine function of pancreas

• Epidemiology

• Different hypothesis for pancreatitis

• Effect of alcohol on pancreas

• Effect of alcohol on PSCs

• Role of PSCs in fibrosis or cancer progression

Pancreas• Anatomy

– Endocrine• Pancreatic islets produce insulin and glucagon

– Exocrine• Acini produce digestive enzymes

– Regions: Head, body, tail

Normal Pancreas

• The pancreatic gland contains three major types of cells.

• The ductular cells make up about 10% of the pancreas and secrete solutions rich in bicarbonate.

• The acinar cells comprise over 80% of the pancreas and they synthesize and secrete pancreatic enzymes.

• The islet cells make up about 10% of the pancreas and form the endocrine portion of the pancreas. These cells secrete the hormones insulin, glucagon, somatostatin, and pancreatic polypeptide.

Islet cells

– Alpha cells secrete glucagon- elevates blood glucose concentrations

– Beta cells secrete insulin- reduces blood glucose concentrations

– Delta cells secrete somatostatin- slows the rate of food absorption and digestive enzyme secretion

Pancreatic enzyme

1. Occurs in the islets of Langerhans2. Beta cells secrete insulin3. Alpha cells secrete glucagon4. Delta cells secrete somatostatin

1. The acinar cells secrete amylase, proteases, and lipases, enzymes responsible for the digestion of 3 food types: carbohydrate, protein, and fat.

2. Trypsin is the most abundant enzyme3. Stored in its inactive form, trypsinogen;

activated by enterokinase

Exocrine Endocrine

Exocrine & Endocrine function of Pancreas

Alcohol & Pancreatitis

• Alcohol abuse accounts for 38-94% CP• Daily 80g/day for 13-21 years to develop

alcohol induced pancreatitis• Only 10 % of heavy drinker develop

pancreatic inflammation• Black ppl are 3-4 times more chances to

develop pancreatitis• Patient get diagnosed at age of 35-40 years of

age

Ductal obstruction hypothesis

• Chronic alcohol use • acinar and ductal cell

• protein rich pancreatic juice, low in volume and HCO3 • formation of protein precipitates – plug

• calcification of ppt – ductal stone formation• ductule obstruction

• parenchymal damage

• Pancreatic ductal stone are seen in alcoholic, tropical, hereditary, idiopathic

• Histologic changes of CP may be seen with out ductal obstruction

Toxic metabolic hypothesis

• (alcohol) Direct injurious effect on acinar and ductal cells• Increased membrane lipid peroxidation (oxidative stress),

free radical production• Increase acinar cell sensitivity to pathogenic stimuli• Activation of pancreatic stellate cells (alcohol, cytokines) –

produce proteins of extracellular matrix

Necrosis fibrosis hypothesis

• Repeated episodes of acute pancreatitis with cellular necrosis or apoptosis, healing replaces necrotic tissue with fibrosis

• Evidence from natural history studies - more severe and frequent attacks

• More evidence from hereditary pancreatitis and animal models

• But some have evidence of chronic pancreatitis at time of first clinical acute attack

Genetics Basis of CP

Autosomal dominant disorder Cationic Trypsinogen gene mutation….PRSS1(R122H)

Autosomal Recessive….CFTR mutation

Susceptabilty gene….SPINK1

Course Modifiers…..various polymorphisms(ADH2,ADH3,ALDH2,CYP2E1 ETC.)

34

35

36

37

38

39

40

41

Allele Type

Mea

n A

ge

(in

yea

rs)

2-1 2-1 2-1 2-2 2-2 2-2

40.76 ± 8

38.5 ± 3

34.71 ± 10

It appears that When genotype 2-2/2-2 is conferring risk ( double dose of

Lys; allele 2-2 *) onset of pancreatitis occurs at younger age

But when genotype 2-1/2-2 (heterozygous) is conferring risk (single dose of Lys; allele 2-2 *) onset of pancreatitis is slightly delayed

When genotype 2-1/2-1 is present (absence of allele 2-2 *) onset of pancreatitis is significantly delayed

Distribution of ALDH genotypes with age at onset of Alcoholic Pancreatitis

Distribution of ALDH genotypes with age at onset of Alcoholic Pancreatitis

Alcohol and Pancreas

• Incompletely understood and intensely studied.

• Why 10% heavy alcoholics develop chronic pancreatitis and the rest not, or limited to asymptomatic pancreatic fibrosis

How alcohol affect pancreas

• Earlier theories suggested pancreatic duct is the central organ to develop alcohol induced pancreatitis

• New theory evolved, alcohol have direct effect on acinar cell as well as PSCs

Mechanism of ethanol induced Pancreas dysfunction

• Ethanol & other factors combined affect – Pancreatic blood flow

– Pancreatic exocrine secretion

– Pancreatic duct permeability

– Zymogen activation

– Intracellular signaling

– Oxidative stress generation

– Interaction of ethanol and its metabolite

Spasm in sphincter of oddi causes

• The sphincter of Oddi separates the common bile duct & pancreatic duct from the small intestine. So by relaxing it, bile and pancreatic juices can be dumped into the small intestine.

• Digestive enzyme of pancreas instead of entering to intestine to digest food, “ digest” pancreatic cell

• Backflow of bile or duodenum content back into the pancreatic duct lead

Effect of alcohol on small duct

• Small pancreatic duct begins at the acini and drain into large pancreatic duct

• Small duct blocked by protein plug formation

• Protein plug get enlarge and calcify, which may be a cause or effect of disease

Freedman at al 1993

Protein plug: pancreatic digestive enzyme and lithostathine & GP2

• Pancreatic Lithostathine– Form 5-10% protein of pancreatic secretion

– Inhibit the deposition of ca+ from pancreatic juice, therfore its ↓ level promote calcification of protein (Bernard et al 1992)

– Convert lithostathine into lithostathine S1( to initiate plug formation

– Long term alcohol consumption leads to ↑ conc. of lithostathine in juice, which promotes protein deposition in ducts (Apte et al 1996)

• Pancreatic GP2– Help in protein precipitation from pancreatic juice

– Alcohol consumption lead to ↑ conc. of GP2, favor plug formation

Effect of protein plug formation on pancreas

• Protein plug formation or stone in small duct lead to ulceration, scarring further obstruction & finally atrophy and fibrosis via– Reduces pancreatic secretion

– Increased viscosity of secretion

– Decrease citrate conc. In pancreatic juice, a predisposing factor for crystal formation

– Producing protein to increase stone formation

Protein plug known to be a player in progression of disease if not initiation

Direct effect of alcohol on acinar cell• A single acinar cell can

synthesize 10 million enzyme mol/day

• They protect themselves by synthesizing most digestive enzymes (trypsin) as inactive precursor i.e. zymogen granule

• Any disruption could lead to premature activation of zymogen & causes auto-digestion of pancreatic acinar cells whitcomb et al 1996

Effect of alcohol on fragility of zymogen & lysosome

• Long term alcohol consumption premature activation of zymogen enzyme

• Alcohol ↑ the synthesis of digestive enzyme

• Alcohol consumption lead to↑ in fragility of lysosome & zymogen granule allowing zymogen leakage to the cell

Wilson et al 1992, Apte et al 1995

How fragility start auto-digestion of pancreas

• Alcoholic metabolites makes fragile zymogen granule & lysosomal membrane

• Trypsin can not be degrade by protective enzyme of acinar cell

• So due to fragile membrane lysosomal enzyme (cathepsin B) will active trypsin to trypsinogen which further lead to activate other enzyme & start the cascade of event or autodigestion of pancrease

ZL

TrypsinTrypsinogen

Cathepsin B

Activate other enzyme and strat autodigestion of pancreas

Ethanol Metabolism

Ethanol Metabolism

Non-oxidative PathwayOxidative Pathway

Rate is 21 fold higher

Acetaldehyde FAEE

Alcohol dehydrogenaseCytochrome P-4502E1 (CYP2E1)

Catalse FAEE synthases

ROS

Induces morphological Alteration in pancreas

Harmful to cell membrane, intracellular protein & DNA

lysosomal fragility

Alcohol induced oxidative stress in pancreas• The cell is normally protected from the disruptive effect of free

radicals by antioxidant system, which releases during normal metabolism of alcohol via CYP2E1

• Oxidative stress is the imbalance between production of ROS & defense mechanism (Antioxidant glutathione, peroxidase, superoxide mutase & catalase)

• This imbalance may be a due to – ROS release during ethanol oxidation via CYP2E1

– Depletion of ROS scavenger gluthathione

• As a result oxidative stress destabilizes zymogen & lysosome granules lead to auto digestion of pancreas via acinar cell activation

•

Altomare et al 1996

Effect of alcohol metabolism on pancreas

• Acetaldehyde induces the stellate cells & lead to fibrosis

• FAEE induces the acinar cell & lead to necrosis and continuous insult lead to fibrosis

Alcoholic Chronic Pancreatitis

ALCOHOL

Oxidative pathway

Acetaldehyde

Non-oxidative pathway

Fatty acid ethyl esters (FAEEs)

Pancreatic Stellate Cell Activation

Pancreatic stellate cells (PSCs)

• PSCs are 4% cell of total pancreatic cell

• Vit A containing lipid droplets

• On activation loose vitamin A – Maintained matrix turn over

– Protective immune function as phagocytic cell

– Work as progenitor cell (in acute injury secrte insulin after differentiation)

– CCK (Cholecystokinin ) induced pancreatic exocrine function

Stellate cells

Quiscent PSc Active PSc

Vitamin A lipid droplets Present Absent

a Smooth muscle actin Absent Present

Proliferation Limited Increased

Migration Limited Increased

ECM Limited Increased

MMPs and TIMPs Maintain normal ECM turnover

Change in types of MMPs and TIMPs to facilitate ECM deposition

Production of cytokines Limited Increased ((PDGF, TGFb, CTGF, IL1, IL6, IL15)

Capacity for phagocytosis Absent Present

ActivePSc

PDGF induced Proliferation mediated by ERK & JAK/STAT

•PDGF induced migration mediated by PI3K•Migration also mediated by hedgehog pathway

Quiscent PSc

TGFB mediated

LPS receptor (TLR 2 &4)

Stellate cells formation, proliferation & migration

SMA positve

PSCs Activation• PSCs are activated via paracrine

pathways by exogenous factors such as cytokines, oxidant stress, ethanol and its metabolites

• Activated PSCs secrete cytokines which act on PSC in autocrine way

• These remain active PSCs even in the absence if the initial trigger factors, leading to excessive ECM production and eventually causing pancreatic fibrosis

ActivePSC

Pancreatic fibrosis

Quiescent PSC

Cytokine autocrine effect

ActivePSC

Quiescent PSC

Cell proliferationSMA expression

ECM protein synthesis Matrix degradation

via

Vit. A loss (Altered retinol metabolism)Imbalance MMP/TIMP

Cell migration Contractility

Pancreatic fibrosis

Ethanol insult & Endotoxin

Mainly acetaldehydeLPS ↑ in blood of alcoholics

Activated PSCs leads to fibrosis

PSC in Chronic Pancreatitis: Animal studies

• Fibrosis has been produced in rat model via (1) trinitrobenzene sulfonic acid (TNBS) injection into the pancreatic

duct (Haber et al., 1999)

(2) intravenous injection of an organotin compound dubutyltin chloride

(DBTC) (Emmrich et al., 2000)

(3) spontaneous chronic pancreatitis in WBN/Kob rats (Ohashi et al., 1990)

(4) severe hyperstimula- tion obstructive pancreatitis (SHOP),

involving intraperitoneal (IP) injections of supramaximal doses of caerulein (a synthetic analogue of CCK, a major pancreatic

secretagogue) + bile- pancreatic duct ligation (Murayama et al., 1999)

(5) repeated IP injections of a superoxide dismutase inhibitor (Matsumura et al., 2001)

Continued..• (6) intragastric high dose alcohol administration + repeated

caerulein injections (Tsukamoto et al., 1988; Uesugi et al., 2004)

• (7) chronic alcohol administration (liquid diet) with repeated cyclosporin and caerulein injections (Gukovsky et al., 2008)

• (8) chronic alcohol administration with repeated endo- toxin LPS, injections (Vonlaufen et al., 2007b).

•Rat model produced by chronic alcohol administration and repeated endotoxin exposuren that is based on a well recognized clinical phenomenon, namely endo- toxinaemia (secondary to increased gut mucosal permeability) in alcoholics (Bode et al., 1993; Parlesak, 2005). •Thus, the alcohol feeding, LPS challenge model possibly represents the most physiologically relevant model of chronic alcoholic pancreatitis described to date.

Pancreatic fibrosis

Lost vitamin A

Inflammation towards PDAC

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