genego.com) and Ingenuity Pathway Analysis (IPA; Ingenuity Systems; www.ingenuity.com). Consensus findings

from these tools were used to interpret and understand the biological mechanisms behind the data. Microarray data have been deposited to the NCBI’s Gene Expression Omnibus repository (accession no.: GSE38872). Mice were fasted for 4 hours and were intraperitoneally (IP) injected Tyrosine Kinase Inhibitor Library with 10 µCi of [1-14C]-sodium acetate (PerkinElmer, Waltham, MA). Mice were sacrificed 30 minutes after injection, and ∼250 mg of liver were rinsed in ice-cold 1× phosphate-buffered saline. Liver tissue was saponified in a 2.2-mL mixture of 50% KOH/95% ethanol (1:10, v/v) at 70°C overnight. 3H-cholesterol (1 µCi) was added to the same tube as a recovery control. Sterols were extracted in 3 mL of hexane, dried, and redissolved AZD4547 mouse in a 300-μL mixture of acetone/ethanol (1:1, v/v). Sterols were then precipitated with 1 mL of digitonin (0.5% in 95% ethanol) overnight at room temperature. Saponified fatty acids were acidified and extracted with petroleum ether. WT and Cyp7a1-tg mice were IP injected with a single dose of glucose (8 g/kg) containing 5 µCi of [1-14C] glucose (PerkinElmer) as an isotope tracer. Mice were allowed free access to standard chow and water, and feces were collected for 3 consecutive days. Fecal samples were then homogenized in a 5-mL mixture of 50% KOH/95% ethanol (1:10, v/v) at 70°C overnight.

3H-cholesterol (1 µCi) was added to the same tube as a recovery control. Neutral sterols were separated from bile acids by extraction in 6 mL of hexane, dried, and redissolved in a 1-mL mixture of acetone/ethanol (1:1, v/v) and were then precipitated with 3 mL of digitonin (0.5% in 95% ethanol) overnight at room temperature. Adenovirus-expressing miR-33a was purchased from Applied Biological Materials, Inc. (Richmond, British Columbia, Canada). Adeno-null, which does not express a gene product,

was purchased from Vector Biolabs (Philadelphia, PA). Adenovirus was administered find more at 2 × 109 pfu/mouse through the tail vein. Experiments were carried out 7 days postinjection. Results were expressed as mean ± standard error (SE), unless noted. Statistical analysis was performed by the Student t test; P < 0.05 indicates statistical significance. To obtain molecular insight into the role of bile acid synthesis in maintaining hepatic lipid homeostasis, we used microarray gene profiling to identify differentially expressed genes in livers of chow-fed and Western high-fat-diet (HFD)-fed Cyp7a1-tg and WT mice. Under chow-fed conditions, 77 genes were identified as differentially expressed with a 2-fold induction or a 50% inhibition, whereas under WD-fed conditions, 144 genes were differentially expressed (Supporting Fig 1A). There were 52 differentially expressed genes that were identified by comparison under both chow-fed and WD-fed conditions.