Materials and methodsThe study protocol conforms with the United Kingdom Animals (Scientific Procedures) Act of 1986, the Home Office (UK) and was approved by the local institutional review board.Sixty 10 to 14 week old, male Sprague-Dawley rats weighing selleck inhibitor 340 to 390 g were used in this study. Animals were acclimatized to laboratory conditions for three days before experimental use, housed at 21��C with a 12-hour light-dark cycle, and allowed free access to tap water and standard rodent chow. On the day of study, the rats were weighed and anesthetized with an intraperitoneal injection of pentobarbital sodium 50 mg/kg repeated twice (every three hours). The internal jugular vein was cannulated to draw blood samples and for the sedative infusion. The rats were then randomized to saline infusion (C group), midazolam infusion at 0.

6 mg/kg/hr (M group) or dexmedetomidine infusion at 5 ��g/kg/hr (D group) [22] for eight hours (n = 20 per group) with equal volume infusion rate at 1 ml/hr in each group. Drug doses were calculated from human doses scaled for body surface area using the Meeh-Rubner formula. The dexmedetomidine dose had previously been applied in rats [22] and the midazolam dose was the calculated equivalent of the dexmedetomidine dose for the rat (scaled from human dosing). All animals appeared sedated and did not need further sedation to maintain immobility. All groups were administered intravenous fluids at 1 ml/hr (thus ensuring the same volume of fluid resuscitation). After this procedure, the animals were rested for 30 minutes followed by a baseline venous blood sample (time = 0 hours).

Body temperature was maintained at 37 �� 0.2��C with the aid of a heating pad.Cecal ligation and double intestinal punctureAfter cannulation and the start of the sedative infusions cecal ligation and double intestinal puncture (CLIP) was performed as previously described [33,34] under additional pentobarbital anesthesia. The procedure was performed under sterile conditions with the abdominal skin disinfected with 70% alcohol. Laparotomy was conducted through a 2 cm lower-midline incision. The cecum was exposed and ligated immediately distal to the ileocecal valve to avoid intestinal obstruction and then punctured twice with an 18-gauge needle, squeezed gently to force out a small amount of feces, and then returned to the abdominal cavity.

The abdomen is closed with 3-0 silk sutures in two layers. Following completion of CLIP the sedative (or saline) infusions were continued without bolus administration.Plasma cytokine measurementVenous Cilengitide blood samples (1 ml) were drawn for the measurement of plasma cytokine (TNF-�� and IL-6) concentrations at two, four, and six hours after CLIP (n = four to six per group). Double the volume of saline was injected to replace blood lost after each sampling.