Ketamine-induced hepatoprotection: the role of heme oxygenase-1

Abstract

Lipopolysaccharide (LPS) causes hepatic injury that is mediated, in part, by upregulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Ketamine has been shown to prevent these effects. Because upregulation of heme oxygenase-1 (HO-1) has hepatoprotective effects, as does carbon monoxide (CO), an end product of the HO-1 catalytic reaction, we examined the effects of HO-1 inhibition on ketamine-induced hepatoprotection and assessed whether CO could attenuate LPS-induced hepatic injury. One group of rats received ketamine (70 mg/kg ip) or saline concurrently with either the HO-1 inhibitor tin protoporphyrin IX (50 micromol/kg ip) or saline. Another group of rats received inhalational CO (250 ppm over 1 h) or room air. All rats were given LPS (20 mg/kg ip) or saline 1 h later and euthanized 5 h after LPS or saline. Liver was collected for iNOS, COX-2, and HO-1 (Western blot), NF-kappaB and PPAR-gamma analysis (EMSA), and iNOS and COX-2 mRNA analysis (RT-PCR). Serum was collected to measure alanine aminotransferase as an index of hepatocellular injury. HO-1 inhibition attenuated ketamine-induced hepatoprotection and downregulation of iNOS and COX-2 protein. CO prevented LPS-induced hepatic injury and upregulation of iNOS and COX-2 proteins. Although CO abolished the ability of LPS to diminish PPAR-gamma activity, it enhanced NF-kappaB activity. These data suggest that the hepatoprotective effects of ketamine are mediated primarily by HO-1 and its end product CO.

Fig. 1.

Effects of intraperitoneal tin protoporphyrin IX (SnPPN; 50 μmol/kg) or saline and intraperitoneal ketamine (70 mg/kg) or saline given 1 h prior to intraperitoneal saline or lipopolysaccharide (LPS; 20 mg/kg) for 5 h on LPS-induced liver injury as measured by alanine aminotransferase (ALT) release. Data are means ± SE; n ≥ 5 per group. *P < 0.05 vs. saline counterpart, **P < 0.05 vs. Saline-Saline-LPS.

Fig. 2.

Effects of intraperitoneal SnPPN (50 μmol/kg) or saline and intraperitoneal ketamine (70 mg/kg) or saline given 1 h prior to intraperitoneal saline or LPS (20 mg/kg) for 5 h on LPS-induced inducible nitric oxide synthase (iNOS). A: densitometric analysis; data are means ± SE; n ≥ 5 per group. *P < 0.05 vs. saline counterpart, **P < 0.05 vs. Saline-Saline-LPS, ***P < 0.05 vs. Saline-Ketamine/LPS. B: representative Western immunoblot.

Fig. 3.

Effects of intraperitoneal SnPPN (50 μmol/kg) or saline and intraperitoneal ketamine (70 mg/kg) or saline given 1 h prior to intraperitoneal saline or LPS (20 mg/kg) for 5 h on LPS-induced cyclooxygenase-2 (COX-2). A: densitometric analysis; data are means ± SE; n ≥ 5 per group. *P < 0.05 vs. saline counterpart, **P < 0.05 vs. Saline-Saline-LPS, ***P < 0.05 vs. Saline-Ketamine/LPS. B: representative Western immunoblot.

Fig. 4.

Effects of inhaled carbon monoxide (CO; 250 ppm) over 1 h or room air, 1 h prior to intraperitoneal saline or LPS (20 mg/kg), on nuclear factor-κB (NF-κB). A: densitometric analysis; data are means ± SE; n ≥ 5 per group. *P < 0.02 vs. Air-Saline, **P < 0.0001 vs. Air-LPS. B: representative electrophoretic mobility shift assay.

Fig. 5.

Effects of inhaled CO (250 ppm) over 1 h or room air, 1 h prior to intraperitoneal saline or LPS (20 mg/kg), on peroxisome proliferator-activated receptor-γ (PPAR-γ). A: densitometric analysis; data are means ± SE; n ≥ 5 per group. *P < 0.02 vs. Air-Saline, **P < 0.0005 vs. Air-LPS. B: representative electrophoretic mobility shift assay.

Fig. 6.

Effects of inhaled CO (250 ppm) over 1 h or room air, 1 h prior to intraperitoneal saline or LPS (20 mg/kg), on LPS-induced inducible nitric oxide synthase (iNOS). A: densitometric analysis; data are means ± SE; n ≥ 5 per group. *P < 0.05 vs. Air-Saline, **P < 0.005 vs. Air-LPS. B: representative Western immunoblot.

Fig. 7.

Effects of inhaled CO (250 ppm) over 1 h or room air, 1 h prior to intraperitoneal saline or LPS (20 mg/kg), on LPS-induced cyclooxygenase-2 (COX-2) expression. A: densitometric analysis; data are means ± SE; n ≥ 5 per group. *P < 0.01 vs. Air-Saline, **P < 0.005 vs. Air-LPS. B: representative Western immunoblot.