Five female domestic piglets (Sus scrofa, average body weight 26.9 kg) were used in this study. Piglets were immobilized by anesthesia and mechanically ventilated via a tracheotomy for a period of five days. During this study period, the animals were sedated using isoflurane inhalation (Abbott Laboratories, Chicago, Il, USA, 0.8 – 1.3% end-tidal concentration) supplemented by intravenous bolus doses of morphine and ketamine as needed. Core body temperature (blood) was maintained in the range of 38.5 – 40°C by a servo controlled heating pad. The animals received intravenous crystalloid fluid (Ringeracetat) to maintain stable blood pressure and urinary output and a glucose infusion (Rehydrex, Fresenius Kabi, Stockholm, Sweden, 25 mg glucose /mL) in the range of 0.5 – 1.5 mg/kg/minute to decrease the effects of catabolism. Each animal received prophylactic streptomycin 750 mg/d and bensylpenicillin 600 mg/d (Streptocillin Vet, Boeringer-Ingelheim, Hellerup, Denmark). Arterial blood gas analysis as well as electrolytes and blood glucose levels were monitored regularly and kept in the normal range throughout the study period. A neuromuscular blocking agent (NMBA) was administered as a continuous infusion of pancuronium bromide 0.1mg/kg/h (Pavolun; Organon, Boxtel, The Netherlands) for 5 days while a corticosteroid (CS) was given as bolus doses of betamethasone 0.1 mg/kg (Betapred; GSK, Slona, Sweden) twice daily for 5 days. Endotoxemia was induced by a continuous infusion of Escherichia coli endotoxin, serotype O26:B6 (Sigma Labkemi, Stockholm, Sweden) at 36 µg/kg/h for 1h.
Growth protocol
All piglets originated from the same farm (Vallrums Lantbruk, Ransta, Sweden).
Extracted molecule
total RNA
Extraction protocol
Total RNA was extracted according to the standard Affymetrix protocol including Trizol extraction and Rneasy column cleanup.
Label
biotin
Label protocol
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 3 ug total RNA (Expression Analysis Technical Manual, 2007, Affymetrix).
Hybridization protocol
Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on the Porcine Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.
Scan protocol
GeneChips were scanned using the GeneChip Scanner 3000.
