Brazilian green propolis suppresses acetaminophen-induced hepatocellular necrosis by modulating inflammation-related factors in rats

Abstract

Propolis is a resin-like material produced by honey bees from bud exudates and sap of plants and their own secretions. An ethanol extract of Brazilian green propolis (EEBGP) contains prenylated phenylpropanoids and flavonoids and has antioxidative and anti-inflammatory effects. Acetaminophen (N-acetyl-p-aminophenol; APAP) is a typical hepatotoxic drug, and APAP-treated rats are widely used as a model of drug-induced liver injury. Oxidative stress and inflammatory reactions cause APAP-induced hepatocellular necrosis and are also related to expansion of the lesion. In the present study, we investigated the preventive effects of EEBGP on APAP-induced hepatocellular necrosis in rats and the protective mechanism including the expression of antioxidative enzyme genes and inflammation-related genes. A histological analysis revealed that administration 0.3% EEBGP in the diet for seven days reduced centrilobular hepatocellular necrosis with inflammatory cell infiltration induced by oral administration of APAP (800 mg/kg) and significantly reduced the area of necrosis. EEBGP administration did not significantly change the mRNA expression levels of antioxidant enzyme genes in the liver of APAP-treated rats but decreased the mRNA expression of cytokines including Il10 and Il1b, with a significant difference in Il10 expression. In addition, the decrease in the mRNA levels of the Il1b and Il10 genes significantly correlated with the decrease in the percentage of hepatocellular necrosis. These findings suggest that EEBGP could suppress APAP-induced hepatocellular necrosis by modulating cytokine expression.

Keywords: Brazilian green propolis; acetaminophen; antioxidative enzyme; chemokine; cytokine; hepatocellular necrosis.

Fig. 1.

Effect of EEBGP administration on hepatocellular necrosis. A: Representative photographs of the liver after 24 h of APAP 800 mg/kg administration. A-(a): Control group. Centrilobular hepatocellular necrosis with inflammatory cell infiltration is observed. A-(b): EEBGP group. Hepatocellular necrosis with inflammatory cell infiltration is limited to the region around the central veins. CV, central vein; PT, portal triad. Tissue specimens were stained with hematoxylin and eosin (100×). B: Percentage of necrosis area in the liver of the control and EEBGP groups. The vertical bars represent the mean values and standard deviation for each group. *p<0.05 versus the control group, assessed by Student’s t-test. C: The serum ALT and AST levels in the EEBGP and control groups. The vertical bars represent the mean values and standard deviation for each group. EEBGP, ethanol extract of Brazilian green propolis; APAP, N-acetyl-p-aminophenol; ALT, alanine aminotransferase; AST, aspartate aminotransferase

Fig. 2.

mRNA levels of the antioxidative enzymes in the liver of the control and EEBGP groups. The mRNA levels are normalized to that of Gapdh expression. The vertical bars represent the mean values and standard deviation for each group. EEBGP, ethanol extract of Brazilian green propolis

Fig. 3.

mRNA levels of the inflammation-related genes in the liver of the control and EEBGP groups. A: Cytokines. B: chemokines and nitric oxide synthase 2. The mRNA levels are normalized to that of Gapdh expression. The vertical bars represent the mean values and standard deviation for each group. *p<0.05 versus the control group, assessed by Student’s t-test. EEBGP, ethanol extract of Brazilian green propolis

Fig. 4.

Correlation between the percentage of necrosis area and mRNA level of inflammation-related genes in the liver of the EEBGP and control groups. A: Il1b, B: Il6, C: Il10, and D: Cxcl2 gene. The mRNA levels are normalized to that of Gapdh expression. The vertical axis represents the individual mRNA levels, and the horizontal axis represents the individual necrosis area. The R-value (correlation coefficient) and the p-value were assessed by Spearman’s rank correlation coefficient. EEBGP, ethanol extract of Brazilian green propolis