Based on the preferential elevation of MCP-1 among other CC-chemokines in alcoholic hepatitis patients4, 5 and its importance in the modulation of proinflammatory cytokines,9, 10 we hypothesized that MCP-1 contributes to chronic alcoholic liver injury and steatosis via the modulation of inflammatory cytokines. Using MCP-1-deficient mice, we sought to investigate whether MCP-1 and its receptor, chemokine (C-C motif) receptor 2 (CCR2), play a causative role in alcoholic liver injury. ACOX, acyl coenzyme A oxidase; ALD, alcoholic liver disease; ALT, alanine aminotransferase; CCL2, chemokine (C-C motif) ligand 2; CCR2, chemokine (C-C motif) receptor 2; CD, cluster of differentiation; CoA, coenzyme Pirfenidone price A; CPT-1,
carnitine palmitoyl transferase 1A; CYP2E1, cytochrome P450 2E1; ELISA, enzyme-linked immunosorbent assay; EMSA, electrophoretic mobility shift analysis; H&E, hematoxylin and eosin; ICAM-1, intercellular adhesion molecule 1; IL, interleukin; IP, intraperitoneal; KCs, Kupffer cells; KO, knockout; LCAD, long-chain acyl-CoA dehydrogenase; LPS, lipopolysaccharide; MCAD, medium-chain acyl-CoA dehydrogenase; MCP-1, monocyte chemoattractant protein-1; MIP-1, macrophage inflammatory Crizotinib datasheet protein 1; mRNA, messenger RNA; NF-κB, nuclear factor kappa light-chain enhancer of activated B cells; PPARα, peroxisome proliferator-activated receptor
alpha; PPARγ, peroxisome proliferator-activated receptor gamma; PPRE, peroxisome proliferator response element; qPCR, quantitative polymerase chain reaction; SEM, standard error of the mean; TBARS, thiobarbituric acid-reactive substances; TLR4, Toll-like receptor 4; TNFα, tumor necrosis factor alpha; VCAM-1, vascular cell adhesion molecule 1; WT, wild type. Additional descriptions enough of methods are available
in the Supporting Information. All animals received proper care in agreement with animal protocols approved by the Institutional Animal Use and Care Committee of the University of Massachusetts Medical School (Worcester, MA). Six- to eight-week-old female wild-type (WT) (C57BL/6) and MCP-1-deficient and CCR2-deficient mice (all generated on a C57BL/6 background; The Jackson Laboratory, Bar Harbor, ME) received Lieber-DeCarli diet (Bio-Serv, Frenchtown, NJ) with 5% (v/v) ethanol (36% ethanol-derived calories) for 6 weeks; pair-fed control mice received an equal amount of calories as their alcohol-fed counterparts with the alcohol-derived calories substituted with dextrin maltose. All strains of mice consumed comparable daily calories. In some cases, mice from both the alcohol-fed and pair-fed groups were administered an intraperitoneal (IP) injection of either 0.2 mL of 0.9% saline (phosphate-buffered, pH 7.4) alone as a vehicle control or 0.2 mL 0.9% saline containing 0.5 mg/kg of nonpurified lipopolysaccharide (LPS; from Escherichia coli 0111:B4; Sigma-Aldrich, St. Louis, MO) and sacrificed 2 hours later.