Alcohol-induced etiology represents the most common cause of acute pancreatitis in most countries [14
]. While the pathomechanisms of alcohol-induced pancreatitis are yet to be fully understood, there is substantial evidence for both direct acinar cell damage mediated by alcohol and its metabolites as well as inflammatory cell recruitment and activation as pivotal components of alcoholic pancreatitis [14
]. In addition, regulation of inflammatory cell responses by alcohol may play a role in the development and progression of pancreatitis. In this study, we investigated the effect of acute alcohol treatment on inflammatory cell function and activation in response to a single or multiple inflammatory stimuli. We found that acute alcohol can differently regulate monocyte TNFα production depending on the complexity of the co-stimulatory signals (fig. ). In vivo alcohol intake or acute alcohol treatment in vitro had anti-inflammatory effects on TLR4-induced cell activation [22
]. In contrast, even acute alcohol could augment inflammation when monocytes were stimulated with both TLR2 and TLR4 ligands, suggesting that alcohol regulates monocytes differently in the presence of more complex pro-inflammatory or danger signals [33
]. It is tempting to speculate that some of these mechanisms may play a role in acute pancreatitis related to acute alcohol intake or binge drinking where local tissue damage could produce multiple different ligands for inflammatory cell activation. For example, acinar cell death could be triggered as a result of direct effects of alcohol and these apoptotic and necrotic cells induce pro-inflammatory activation of monocytes [14
]. It has been demonstrated that chronic alcohol exposure increases the susceptibility of the pancreas to LPS-induced necrosis [34
]. While LPS induces acinar cell apoptosis, alcohol-exposed acinar cells have increased susceptibility to LPS-induced necrosis. It is conceivable that LPS-induced TLR4 stimulation occurs if excessive amounts of acute or binge drinking cause an increase in gut permeability and result in elevations in serum LPS [35
]. Elevated levels of serum endotoxin have been reported after chronic alcohol intake both in humans and in animal models [35
]. Furthermore, proteins derived from dying cells such as acinar cells are recognized as ‘danger’ signals by inflammatory cells [10
]. For example, toll-like receptor 2 (TLR2) is activated by heat shock protein 70 and cellular components of dying cells activate damage-associated pattern recognition receptors [10
]. Thus, under conditions where multiple cellular stimuli occur even acute alcohol could augment pro-inflammatory cell activation. Our results demonstrated that increased TNFα production in the presence of acute alcohol and dual stimulation with TLR2 and TLR4 ligands was mediated by an increase in NF-κB activation and DNA binding [33
]. We found that TLR-induced IRAK-1 phosphorylation was attenuated by acute alcohol after TLR4 but it was upregulated after TLR2 plus TLR4 stimulation [33
]. In addition, we have recently reported that differential activation of MAP kinases by acute alcohol in the presence of a single or multiple TLR ligands correlates with these effects [33
]. Specifically, acute alcohol selectively inhibits Erk phosphorylation in LPS stimulated cells while JNK phosphorylation is increased by acute alcohol in the presence of dual TLR2 plus TLR4 stimulation. Taken together, we conclude that acute alcohol may attenuate or augment TNFα production and pro-inflammatory signaling pathways depending on the presence of co-stimulatory signals from the environment (fig. ).
Regulation of monocyte TNF-α production is dependent on the TLR costimulatory signals.
Studies in acute non-alcoholic experimental pancreatitis attempting to delineate the effect of TLR-4 signaling resulted in conflicting results. While in the report by Pastor et al. the absence of TLR-4 expression had no impact on local or systemic injury in caerulein-induced pancreatitis [36
], Saluja et al. [pers. commun.] found ameliorated pancreatitis in TLR4-deficient mice. In another study, administration of anti-TLR-4 blocking antibody led to increased local pancreatic and systemic injury [37
]. Interestingly, MD-2, an essential co-receptor of TLR-4, is proteolytically inactivated by trypsin abundantly released in acute pancreatitis, favoring the notion of attenuated pancreatitis in the TLR4-deficient mice.
Inflammation is regulated at multiple levels and under normal conditions monocytes also produce anti-inflammatory cytokines that can keep the initial pro-inflammatory activation under control [39
]. Interleukin-10 (IL-10) produced by monocytes and T cells is an anti-inflammatory cytokine that also has immunosuppressive effects. We found that the anti-inflammatory and immunosuppressive potential of acute alcohol is partially linked to increased IL-10 induction in LPS-stimulated monocytes [5
]. Conversely, IL-10 production was attenuated by acute alcohol in combination with TLR2 or TLR2 plus TLR4 stimulation [33
]. While the mechanisms for this difference in the effects of alcohol on TLR2 and TLR4-induced IL-10 production are yet to be delineated, it is striking that the overall inflammatory cell regulation by alcohol appear to show a coordinated fashion. When acute alcohol inhibited LPS-induced NF-κB and TNFα production, IL-10 was concomitantly induced. In contrast, augmentation of TLR2 plus TLR4-induced TNFα and NF-κB activation by acute alcohol occurred in the presence of reduced monocyte IL-10 production. The role of IL-10 is yet to be explored in alcoholic pancreatitis; however, monocytes recruited to the local tissue damage site are likely to respond in a fashion similar to our in vitro observations.
In alcoholic liver disease, where alcoholic hepatitis is induced by chronic and usually excessive alcohol intake, increased serum levels of pro-inflammatory cytokines correlate with disease outcome [26
]. While IL-8, IL-1, IL-6 and TNFα levels are all elevated in alcoholic hepatitis, increased serum TNFα is a predictor of poor survival [28
]. Elevations in pro-inflammatory cytokines have also been found in chronic pancreatitis, Circulating levels of IL-6 and MCP-1 were increased by acute alcohol intake in chronic pancreatitis [40
]. In in vitro experiments we demonstrated that prolonged alcohol exposure resulted in sensitization of monocytes to LPS-induced TNFα production. We also showed that monocytes exposed to prolonged alcohol treatment showed an increase in NF-κB activation suggesting that this is a likely mechanism involved in increased TNFα induction. Importantly, increased NF-κB activation was associated with pancreatitis induced by hormones (CCK and caerulein) or taurocholate [41
] and ethanol potentiated the cholecystokinine-induced NF-κB activation in pancreatic acinar cells [43
]. Thus, it appears that NF-κB activation is a major indicator and mediator of cellular activation both in acinar cells and in monocytes and it is likely to play a regulatory role in alcoholic pancreatitis.
The clinical course of chronic alcoholic pancreatitis often evolves into pancreatic fibrosis due to pancreatic stellate cell activation which is similar to the outcome of chronic alcoholic hepatitis leading to liver fibrosis and cirrhosis. The mechanisms of stellate cell activation both in the pancreas and in the liver likely involve direct activation by alcohol and its metabolites and pro-inflammatory mediators. In addition to the pro-inflammatory mediators, transforming factor-β (TGFβ) is a major regulator of collagen production in stellate cells [21
]. We have previously reported that alcohol treatment of monocytes results in both induction and augmentation of TGFβ production suggesting an inflammatory cell-mediated mechanisms for stellate cell activation [44
]. Thus, it is conceivable that monocyte-derived pro-inflammatory and pro-fibrinogenic cytokines contribute not only to the inflammation but also to fibrosis in chronic alcoholic pancreatitis.
In summary, we propose that alcohol may amplify inflammatory cell activation which is a potential mechanism contributing to alcoholic pancreatitis. In addition to the components of alcohol-induced pancreatitis elegantly summarized by Apte et al. [17
], we speculate that inflammatory cell activation can be induced by necrotic acinar cells and LPS from increased gut permeability after alcohol use (fig. ). These stimuli in combination with acute or chronic alcohol intake can increase inflammatory responses. Inflammatory cell recruitment and activation can induce a vicious cycle in the presence of ongoing alcohol-induced insult to stimulate local production of pro-inflammatory cytokines that may worsen pancreatic necrosis. In addition, inflammatory cell-derived pro-inflammatory cytokines and TGFβ will result in stellate cell activation leading to fibrosis in the pancreas.
The role of inflammatory responses in alcohol-induced pancreatitis.