Hepatoprotective effect of sodium hydrosulfide on hepatic encephalopathy in rats

Abstract

Hydrogen sulfide is well-known to exhibit anti-inflammatory and cytoprotective activities, and also has protective effects in the liver. This study aimed to examine the protective effect of hydrogen sulfide in rats with hepatic encephalopathy, which was induced by mild bile duct ligation. In this rat model, bile ducts were mildly ligated for 26 days. Rats were treated for the final 5 days with sodium hydrosulfide (NaHS). NaHS (25 µmol/kg), 0.5% sodium carboxymethyl cellulose, or silymarin (100 mg/kg) was administered intraperitoneally once per day for 5 consecutive days. Mild bile duct ligation caused hepatotoxicity and inflammation in rats. Intraperitoneal NaHS administration reduced levels of aspartate aminotransferase and alanine aminotransferase, which are indicators of liver disease, compared to levels in the control mild bile duct ligation group. Levels of ammonia, a major causative factor of hepatic encephalopathy, were also significantly decreased. Malondialdehyde, myeloperoxidase, catalase, and tumor necrosis factor-α levels were measured to confirm antioxidative and anti-inflammatory effects. N-Methyl-D-aspartic acid (NMDA) receptors with neurotoxic activity were assessed for subunit NMDA receptor subtype 2B. Based on these data, NaHS is suggested to exhibit hepatoprotective effects and guard against neurotoxicity through antioxidant and anti-inflammatory actions.

Keywords: Hepatic encephalopathy; Hydrogen sulfide; Inflammation.

Fig. 1. Effect of sodium hydrosulfide (NaHS) on serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST).

(A, B) Mild bile duct ligation (MBDL) surgery increased ALT and AST levels compared to those in the sham control group. NaHS administration significantly decreased serum ALT and AST levels, which were reduced to levels similar to those of the silymarin group. Sham: sham operation control rats, MBDL: MBDL rats, Silymarin: MBDL rats treated with silymarin (100 mg/kg), NaHS: MBDL rats treated with NaHS (25 µmol/kg). Data are expressed as means ± standard error of the mean (n = 6); ^**p < 0.01 compared to sham control, ^##p < 0.01 and ^###p < 0.001 compared to MBDL.

Fig. 2. Photomicrographs of liver sections stained with H&E.

Stained liver sections from the (A) sham-operation group, (B) MBDL group, (C) silymarin (100 mg/kg) treatment group, and (D) NaHS (25 µmol/kg) treatment group. Representative images of each section were captured at 20 × 10 magnification. MBDL, mild bile duct ligation; NaHS, sodium hydrosulfide.

Fig. 3. Effects of sodium hydrosulfide (NaHS) on serum ammonia levels.

Animals that underwent mild bile duct ligation (MBDL) surgery showed increased ammonia levels compared to those in the sham control group. NaHS administration significantly decreased ammonia levels to values similar to those observed in the silymarin group. Sham: sham operation control rats, MBDL: MBDL rats, silymarin: MBDL rats treated with silymarin (100 mg/kg), NaHS: MBDL rats treated with NaHS (25 µmol/kg). Data are expressed as means ± standard error of the mean (n = 6); ^***p < 0.001 compared to sham control, ^###p < 0.001 compared to MBDL.

Fig. 4. Effect of sodium hydrosulfide (NaHS) on serum malondialdehyde (MDA) and catalase (CAT) levels.

(A) Animals that underwent mild bile duct ligation (MBDL) surgery showed increased MDA levels compared to those observed in the sham control group. NaHS administration significantly decreased MDA levels to values similar to those in the silymarin group. (B) MBDL surgery significantly reduced CAT levels compared to those observed in the sham control group. NaHS administration significantly increased CAT levels to those similar to those found in the silymarin group. Sham: sham operation control rats; MBDL: MBDL rats, silymarin: MBDL rats treated with silymarin (100 mg/kg), NaHS: MBDL rats treated with NaHS (25 µmol/kg). Data are expressed as means ± standard error of the mean (n = 6); ^**p < 0.01 compared to sham control, ^#p < 0.05, ^##p < 0.01 and ^###p < 0.001 compared to MBDL.

Fig. 5. Effects of sodium hydrosulfide (NaHS) on serum myeloperoxidase (MPO) and liver tumor necrosis factor (TNF)-α levels.

(A) Mild bile duct ligation (MBDL) surgery significantly increased MPO activity compared to that observed in the sham control group. NaHS administration significantly decreased MPO activity. (B) MBDL surgery also increased TNF-α expression compared to that observed in the sham control group. NaHS administration significantly decreased serum TNF-α expression to levels similar to those found in the silymarin group. Sham: sham operation control rats, MBDL: MBDL rats, silymarin: MBDL rats treated with silymarin (100 mg/kg), NaHS: MBDL rats treated with NaHS (25 µmol/kg). Data are expressed as means ± standard error of the mean (n = 6); ^**p < 0.01 compared to sham control, ^#p < 0.05 and ^##p < 0.01 compared to MBDL.

Fig. 6. Effect of sodium hydrosulfide (NaHS) on hippocampus NR2B protein levels.

MBDL surgery increased the expression of N-Methyl-D-aspartic acid receptor subtype 2B (NR2B) protein in the hippocampus compared to that observed in the sham group. Administration of NaHS significantly reduced the expression of NR2B protein in the hippocampus. Sham: sham operation control rats, MBDL: MBDL rats, NaHS: MBDL rats treated with NaHS (25 µmol/kg). Data are expressed as means ± standard error of the mean (n = 6); ^*p < 0.05 compared to sham control, ^#p < 0.05 compared to MBDL.