ASC,

apoptosis-associated speck-like CARD-domain containing protein; ATP, adenosine triphosphate; DAMP, danger-associated molecular pattern; FA, fatty acid; FFA, free fatty acid; HCV, hepatitis C virus; HFD, high-fat diet; IL, interleukin; LDH, lactate dehydrogenase; LMNC, liver mononuclear cell; LPS, lipopolysaccharide; MCD, methionine choline–deficient; MCS, methionine choline–supplemented; mRNA, messenger RNA; NAFLD, nonalcoholic fatty liver disease; NALP, NACHT, LRR, and PYD domains–containing protein; NASH, nonalcoholic steatohepatitis; NLR, NOD-like receptor; PA, palmitic acid; qPCR, quantitative polymerase chain reaction; TLR, toll-like GSK2126458 solubility dmso receptor; TNF-α, tumor necrosis factor α; ZVAD, carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone. This study was approved by the Institutional Animal Use and Care Committee of the University of Massachusetts Medical School. Female C57Bl/6 wild-type mice that were 6 to 8 weeks old (n = 6-8 per group) were fed either an MCD diet for 5 weeks or an HFD for 4 weeks or 9 months. Control mice received either the same MCD diet supplemented with DL-methionine Angiogenesis inhibitor (3 g/kg) and choline bitartrate (2 g/kg; Dyets, Inc., Bethlehem, PA) or a regular rodent chow diet. We also used 9-week-old female mice that were leptin-deficient (i.e., ob/ob mice; B6.V-Lep ob/J, Jackson Laboratories) and an age-

and sex-matched control group (C57Bl/6J). The presence of steatohepatitis was proven histologically in the MCD diet–fed mice, whereas fat deposition was proven with a liver triglyceride assay in the HFD-fed mice and the ob/ob mice. The TLR4 ligand

LPS (Sigma, St. Louis, MO) was injected intraperitoneally [0.5 mg/kg of body weight for methionine choline–supplemented (MCS) mice and MCD mice and 12 μg for ob/ob mice]. Serum this website alanine aminotransferase levels were determined with a kinetic method (D-TEK, Bensalem, PA), and liver triglyceride levels were assessed with an L-type triglyceride H kit (Wako Chemicals USA, Inc., Richmond, VA). Serum tumor necrosis factor α (TNF-α) and IL-1β levels were determined with a BD cytometric bead array (BD Biosciences, Sparks, MD). Sections of formalin-fixed livers were stained with hematoxylin-eosin, whereas optimal cutting temperature frozen samples were stained with Oil Red O; all slides were analyzed by microscopy. ImageJ and Microsuite (Olympus Soft Imaging Solutions GmbH, Münster, Germany) were used for imaging analysis at indicated magnifications on 20 high-power fields. RNA was purified with the RNeasy kit (Qiagen Sciences, Germantown, MD) and with on-column DNA digestion. Complementary DNA was transcribed with a reverse-transcription system (Promega Corp., Madison, WI). Real-time quantitative polymerase chain reaction (qPCR) was performed with the iCycler (Bio-Rad Laboratories, Inc.