Serial hepatic gene expression profiling in Angus steers during feed restriction and realimentation

Growing ruminants maintained under dietary restriction for extended periods will exhibit compensatory growth when reverted to ad libitum feeding. This period of compensatory growth is associated with increased feed efficiency, lower basal energy requirements, and changes in circulating concentrations of metabolic hormones. To identify genetic mechanisms contributing to these physiological changes, 8 month-old steers were fed either ad libitum (control; n = 6) or 60-70% of intake of control animals (feed-restricted; n=6) for a period of 12 weeks. All steers were then fed ad libitum for the remaining 8 weeks of the experiment (realimentation period). Liver was biopsied from each animal at days -14, +1 and +14 relative to realimentation for RNA extraction and gene expression analysis by microarray hybridization. Overall design: Steers were assigned randomly to one of two treatment groups, control or feed-restricted, and housed indoors in individual pens. Steers were acclimated to their pens for 5 d prior to starting the experimental treatments. Feed was offered once daily between 0630 and 0930 and orts from the previous day's feeding were collected and weighed to estimate actual intake. Control animals were fed ad libitum throughout the 20-wk experimental period. Feed-restricted steers were offered 60-70% of intake of control animals for 12 wks to target a limited rate of gain of approximately 0.5 kg/d. Restricted steers were then fed ad libitum for the remaining 8 wks of the experiment (realimentation period). During the first 3 d of realimentation, feed offered to both treatment groups was divided into two equal rations to gradually adjust restricted animals to full intake. Water was offered ad libitum throughout the experimental period. Approximately 200 mg of liver tissue was collected from each steer by needle biopsy using a Tru-Cut biopsy needle at -14, +1, +14 d relative to realimentation. Liver samples were immediately frozen in liquid nitrogen and stored at -80C until RNA isolation. Total RNA was isolated from 36 liver samples using TRIZOL Reagent (Invitrogen Corp., Carlsbad, CA). Samples were DNase-treated using the TURBO DNA-free kit (Ambion, Inc., Austin, TX) according to manufacturer’s instructions, followed by column purification using the RNeasy Mini Kit (Qiagen, Valencia, CA). Quality and concentration of RNA were assessed using a 2100 Bioanlayzer (Agilent Technologies, Palo Alto, CA) and ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, DE). Probe labeling, hybridizations of probes to the oligo microarray, and array scanning were performed by the Roche NimbleGen Systems, Inc. Microarray Core Facility in Reykjavik, Iceland according to standard procedures (Madison, WI; http://www.nimblegen.com).