Protective effects of garlic oil on hepatocarcinoma induced by N-nitrosodiethylamine in rats

Abstract

To investigate the protective effects and the possible mechanisms of garlic oil (GO) against N-nitrosodiethylamine (NDEA)-induced hepatocarcinoma in rats, Wistar rats were gavaged with GO (20 or 40 mg/kg) for 1 week, and then were gavaged with GO and NDEA (10 mg/kg) for the next 20 weeks. The changes of morphology, histology, the biochemical indices of serum, and DNA oxidative damage of liver were examined to assess the protective effects. Lipid peroxidation (LPO), antioxidant defense system, and apoptosis-related proteins were measured to investigate potential mechanisms. At the end of the study (21 weeks), GO administration significantly inhibited the increase of the nodule incidence and average nodule number per nodule-bearing liver induced by NDEA, improved hepatocellular architecture, and dramatically inhibited NDEA-induced elevation of serum biochemical indices (alanine aminotransferase , aspartate aminotransferase, alkaline phosphatase and gamma-glutamyl transpeptidase) and hepatic 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels in a dose-dependent manner. The mechanistic studies demonstrated that GO counteracted NDEA-induced oxidative stress in rats illustrated by the restoration of glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx), glutathione-S-transferase (GST) levels, and the reduction of the malondialdehyde (MDA) levels in liver. Furthermore, the mRNA and protein levels of Bcl-2, Bcl-xl, andβ-arrestin-2 were significantly decreased whereas those of Bax and caspase-3 were significantly increased. These data suggest that GO exhibited significant protection against NDEA-induced hepatocarcinogenesis, which might be related with the enhancement of the antioxidant activity and the induction of apoptosis.

Keywords: Antioxidant; Apoptosis; Garlic oil; Hepatocarcinoma; Nitrosodiethylamine.

Figure 1

Changes of rat body weight. The data of body weight were presented as mean ± S.D. Statistical difference was shown as: *P < 0.05, **P < 0.01, with respect to control; ^# P < 0.05, ^##P < 0.01, with respect to NDEA group.

Figure 2

Effect of GO on hepatic morphology and histology in NDEA-intoxicated rats. A: The gross appearance of liver. B: Representative photomicrographs of hepatic histology changes in 4 groups (HE staining, ×200). C: Changes of nodule incidence and average number of nodules per nodule-bearing liver in rats.

Figure 3

The content of 8-OHdG in liver DNA. The data was presented as mean ± S.D. *P<0.05, **P<0.01, compared with control group; ^#P<0.05, ^##P<0.01, compared with NDEA group.

Figure 4

Effect of GO and NDEA on the mRNA level of Bcl-2, Bcl-XL, Bax and Caspase-3. The mRNA levels were quantified with GAPDH as an internal control. The control group was defined as 100%. *P<0.05, **P<0.01, compared with control group; ^#P<0.05, ^##P<0.01, compared with NDEA group.

Figure 5

Effect of GO and NDEA on the protein level of Bcl-2, Bcl-XL, Bax and Caspase-3. A: A representative immunoblot. B: Data presented the expression of Bcl-2, Bcl-XL, Bax and Caspase-3 as percentage of control group (mean ± SD) in triplicate. The protein levels were quantified with β-actin as an internal control. *P<0.05, **P<0.01, compared with control group; ^#P<0.05, ^##P<0.01, compared with NDEA group. (lane 1) control; (lane 2) NDEA+GO (low); (lane 3) NDEA+GO (high); (lane 4) NDEA.

Figure 6

Effect of GO and NDEA on the mRNA and protein levels of β-arrestin-2. The protein levels were quantified with β-actin as an internal control. The mRNA levels were quantified with GAPDH as an internal control. *P<0.05, **P<0.01, compared with control group; ^#P<0.05, ^##P<0.01, compared with NDEA group.