An In Vitro Model of Human Acute Ethanol Exposure That Incorporates CXCR3- and CXCR4-Dependent Recruitment of Immune Cells

Abstract

Alcoholic liver disease (ALD) is one of the commonest causes of cirrhosis and liver failure in the developed world. Hepatic inflammation is the critical stage in progression of both ALD and non-ALD, but it remains difficult to study the underlying mechanisms in a human system, and current animal models do not fully recapitulate human liver disease. We developed a human tissue-based system to study lymphocyte recruitment in response to ethanol challenge. Precision-cut liver slices (PCLS) from human livers were incubated in culture, and hepatic function was determined by albumin production, 3-(4,5-dimethylthiazol)-2,5-diphenyl tetrazolium bromide assay, glucose uptake responses, and morphometric assessment. Responses of tissue and lymphocytes to ethanol exposure were determined by PCR, flow cytometry, histology, and lymphocyte infiltration assays. Human PCLS demonstrated appropriate upregulation of CYP2E1, ADH1α, and ADH3 in response to ethanol treatment. Ethanol also induced expression of endothelial VCAM-1 and ICAM-1, production of sICAM-1 and CXCL8, and the chemokine receptors CXCR3 and CXCR4 on CD4 and CD8 lymphocytes. CXCR3- and CXCR4-dependent migration of lymphocytes into the tissue increased significantly in response to treatment with ethanol. We have demonstrated that ethanol increases chemokine receptor expression and lymphocyte recruitment into human liver tissue, suggesting that it may operate directly to promote hepatitis in ALD. The physiological and pathophysiological responses of the PCLS to ethanol in vitro highlight the potential of this assay for dissecting the molecular mechanisms underlying human liver inflammation and as a screening tool for novel therapeutics.

Translational Research in Pharmacology and Toxicology Using Precision-Cut Tissue Slices

“…In human precision-cut liver slices, exposure of ethanol increased the chemokine receptor expression and lymphocyte recruitment into the liver tissue. 35 All these results were in line with in vivo results, demonstrating the added value of the slices in the early onset of alcoholic liver disease. In animal models, CCl 4 and thioacetamide (TAA) are the most commonly used hepatotoxins to induce brosis.…”

“…34 In addition, co-cultures of liver slices and lymphocytes showed increased inltration of lymphocytes in the liver slices upon exposure to ethanol. 35 However the applications of co-incubation of PCTS from different human organs are hampered by the requirement that all the tissues should be available on the same day, and that the opportunity to obtain samples from several different organs from one donor is very limited. Therefore extension of the preservation possibilities, either by developing better cold preservation solutions, cryopreservation methods or long-term culturing at 37 C, will be instrumental in the future application of multi-organ perifusion or co-incubation systems for human PCTS.…”

Altered Distribution of Peripheral Blood Maturation-Associated B-Cell Subsets in Chronic Alcoholism

“…Such apparent discrepancy between the 2 patient groups could potentially be explained, at least in part, because of the potential migration of plasmablasts into the liver in AH, as previously reported also for autoimmune hepatitis (Brandão et al, 2010). In this regard, EtOH has been recently found to increase chemokine receptor-dependent migration and lymphocyte recruitment into the human liver, suggesting that it may directly promote ALD (Karim et al, 2013), but the specific liver infiltrating Blymphocyte subsets were not analyzed. Nevertheless, both in ALD and non-ALD patients, accumulation of intrahepatic memory B cells associated with decreased numbers of PB memory B lymphocytes have been reported (Dhirapong et al, 2011).…”

Profile analysis of hepatic porcine and murine brain tissue slices obtained with a vibratome

“…Unlike classical sectioning procedures based on the use of microtomes, vibratome slicing does not require any tissue fixation, dehydration and embedding thus cell viability and native tissue structure are conserved. Fresh tissue slices are suitable candidates for in-vitro tissue models (Parrish, Gandolfi & Brendel, 1995;Parrish et al, 2002;van de Bovenkamp et al, 2005;Groothuis & de Graaf, 2013) as well as for structural and morphometric analysis (Karim et al, 2013;Eide et al, 2013). For instance, precision-cut liver slices are powerful tools for the in-vitro study of pharmacological metabolism, toxicology and efficacy of novel substances under standardized conditions (van de Bovenkamp et al, 2005(van de Bovenkamp et al, , 2006Van de Bovenkamp et al, 2007;Karim et al, 2013;Eide et al, 2013).…”

“…Fresh tissue slices are suitable candidates for in-vitro tissue models (Parrish, Gandolfi & Brendel, 1995;Parrish et al, 2002;van de Bovenkamp et al, 2005;Groothuis & de Graaf, 2013) as well as for structural and morphometric analysis (Karim et al, 2013;Eide et al, 2013). For instance, precision-cut liver slices are powerful tools for the in-vitro study of pharmacological metabolism, toxicology and efficacy of novel substances under standardized conditions (van de Bovenkamp et al, 2005(van de Bovenkamp et al, , 2006Van de Bovenkamp et al, 2007;Karim et al, 2013;Eide et al, 2013). They have been used extensively for rank-ordering the toxicity of chemicals and examining the mechanisms of liver injury as well as for investigating the induction of cytochrome P-450 enzymes and the expression of stress proteins or peroxisomal enzymes, thus offering a valuable bridge between in-vivo and cell culture systems (Gandolfi, Wijeweera & Brendel, 1996;Olinga & Schuppan, 2013).…”