Activation of Kupffer cells (KCs) by gut-derived lipopolysaccharide (LPS) and Toll-Like Receptors 4 (TLR4)-LPS-mediatedincrease in TNF␣ production has a central role in the pathogenesis of alcoholic liver disease. Micro-RNA (miR)-125b, miR-146a, and miR-155 can regulate inflammatory responses to LPS. Here we evaluated the involvement of miRs in alcoholinduced macrophage activation. Chronic alcohol treatment in vitro resulted in a time-dependent increase in miR-155 but not miR-125b or miR-146a levels in RAW 264.7 macrophages. Furthermore, alcohol pretreatment augmented LPS-induced miR-155 expression in macrophages. We found a linear correlation between alcohol-induced increase in miR-155 and TNF␣ induction. In a mouse model of alcoholic liver disease, we found a significant increase in both miR-155 levels and TNF␣ production in isolated KCs when compared with pairfed controls. The mechanistic role of miR-155 in TNF␣ regulation was indicated by decreased TNF␣ levels in alcohol-treated macrophages after inhibition of miR-155 and by increased TNF␣ production after miR-155 overexpression, respectively. We found that miR-155 affected TNF␣ mRNA stability because miR-155 inhibition decreased whereas miR-155 overexpression increased TNF␣ mRNA half-life. Using the NF-B inhibitors, MG-132 or Bay11-7082, we demonstrated that NF-B activation mediated the up-regulation of miR-155 by alcohol in KCs. In conclusion, our novel data demonstrate that chronic alcohol consumption increases miR-155 in macrophages via NF-B and the increased miR-155 contributes to alcohol-induced elevation in TNF␣ production via increased mRNA stability.

Summary The pathogenesis of alcoholic liver injury involves interactions of several intracellular signalling pathways in different cell types of the liver. Alcohol-induced sensitization of liver macrophages to portal endotoxin/lipopolysaccharide (LPS) is considered a hallmark of alcoholic liver disease (ALD). Intracellular mechanisms associated with LPS-induced signalling play a crucial role in the initiation and progression of alcoholic liver injury, and are being extensively explored. LPS recognition by Toll-like receptor 4 (TLR4) on macrophages and other cell types in the liver, activation of downstream signalling pathways culminating in activation of transcription factors such as NFκB, AP-1 leads to increased inflammatory cytokine production in ALD. In addition, LPS-induced MAPK such as ERK and p38 also contribute to liver injury. The importance of alcohol-induced reactive oxygen species and interactions with TLR pathways in macrophages leading to inflammation is becoming increasingly evident. Collectively, these signalling pathways induce pro- and anti-inflammatory cytokines that play an important role in ALD. In this review we describe the key signalling intermediates leading to alcohol-induced inflammation in alcoholic liver disease.

The Toll-like receptor 4 (TLR4) that recognizes endotoxin, a trigger of inflammation in alcoholic liver disease (ALD), activates two signaling pathways utilizing different adapter molecules: the common TLR adapter, myeloid differentiation factor 88 (MyD88), or Toll/interleukin immuneresponse-domain-containing adaptor inducing interferon (IFN)-␤. The MyD88 pathway induces proinflammatory cytokine activation, a critical mediator of ALD. Here we evaluated the role of MyD88 in alcohol-induced liver injury in wild-type, TLR2-deficient, TLR4-deficient, or MyD88-deficient (knockout [KO]) mice after administration of the Lieber-De-Carli diet (4.5% volume/volume ethanol) or an isocaloric liquid control diet for 5 weeks. Alcohol feeding resulted in a significant increase in serum alanine aminotransferase levels, liver steatosis and triglyceride levels suggesting liver damage in WT, TLR2-KO, and MyD88-KO but not in TLR4-KO mice. Expression of inflammatory mediators (tumor necrosis factor-␣ and interleukin-6) and TLR4 coreceptors (CD14 and MD2) was significantly higher in livers of alcohol-fed WT, TLR2-KO, or MyD88-KO, but not in TLR4-KO mice, compared to controls. Reactive oxygen radicals produced by cytochrome P450 and the nicotinamide adenine dinucleotide phosphate complexes contribute to alcoholic liver damage. Alcohol feeding-induced expression and activation of cytochrome P450 and the nicotinamide adenine dinucleotide phosphate complex were prevented by TLR4-deficiency but not by MyD88-deficiency. Liver expression of interferon regulatory factor 3 (IRF3), a MyD88-independent signaling molecule, was not affected by chronic alcohol treatment in whole livers of WT mice or in any of the KO mice. However, the induction of IRF7, an IRF3-inducible gene, was found in Kupffer cells of alcohol-fed WT mice. Alcohol feeding also induced nuclear factor-B activation in a TLR4-dependent MyD88-independent manner. Conclusion: While TLR4 deficiency was protective, MyD88 deficiency failed to prevent alcohol-induced liver damage and inflammation. These results suggest that the common TLR adapter, MyD88, is dispensable in TLR4-mediated liver injury in ALD. (HEPATOLOGY 2008;48:1224-1231 A lcoholic liver disease (ALD) is characterized by a spectrum of liver pathology ranging from fatty liver, steatohepatitis, to cirrhosis, and it represents the second leading cause for liver transplantation in the United States. 1 Gut-derived lipopolysaccharide (LPS), a component of the gram-negative bacterial wall, has been proposed as a key player in the pathogenesis of ALD. [2][3][4] Exposure to LPS during chronic alcohol consumption results in increased production of inflammatory mediators, as well as in induction of reactive oxygen species (ROS), leading to progression of liver injury. 5,6 Indeed, mice deficient in tumor necrosis factor-alpha (TNF␣) type I receptor were protected from alcohol-induced liver injury. 7 Of the different sources of ROS, the role of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation was suggested by atten...

Overview Multiple line of clinical and experimental evidence demonstrates that both acute, moderate and chronic, excessive alcohol use result in various abnormalities in the functions of the immune system. Altered inflammatory cell and adaptive immune responses in turn result in increased incidence and poor outcome of infections and other organ effects after alcohol use. This review article summarizes recent findings relevant to immunomodulation by alcohol and its consequences on host defense against microbial pathogens and tissue injury.

Cytokines are multifunctional proteins that play a critical role in cellular communication and activation. Cytokines have been classified as being proinflammatory (T helper 1, Th1) or anti-inflammatory (T helper 2, Th2) depending on their effects on the immune system. However, cytokines impact a variety of tissues in a complex manner that regulates inflammation, cell death, and cell proliferation and migration as well as healing mechanisms. Ethanol (alcohol) is known to alter cytokine levels in a variety of tissues including plasma, lung, liver, and brain. Studies on human monocyte responses to pathogens reveal ethanol disruption of cytokine production depending upon the pathogen and duration of alcohol consumption, with multiple pathogens and chronic ethanol promoting inflammatory cytokine production. In lung, cytokine production is disrupted by ethanol exacerbating respiratory distress syndrome with greatly increased expression of transforming growth factor beta (TGFbeta). Alcoholic liver disease involves an inflammatory hepatitis and an exaggerated Th1 response with increases in tumor necrosis factor alpha (TNFalpha). Recent studies suggest that the transition from Th1 to Th2 cytokines contribute to hepatic fibrosis and cirrhosis. Cytokines affect the brain and likely contribute to changes in the central nervous system that contribute to long-term changes in behavior and neurodegeneration. Together these studies suggest that ethanol disruption of cytokines and inflammation contribute in multiple ways to a diversity of alcoholic pathologies.