Suppression of beta-naphthoflavone induced CYP1A expression and lipid-peroxidation by berberine.

Impacts of berberine, a major isoquinoline alkaloid in herbal plants, on beta-naphthoflavone (BNF)-induced CYP1A expression were determined both in primary mouse hepatocytes and in vivo. Berberine concentration-dependently suppressed BNF-induced CYP1A expression in mouse hepatocyte and it significantly down-regulated BNF-induced CYP1A in mouse liver via suppression of mRNA and protein expression, and decreases of EROD and MROD activities. Moreover, berberine showed significant potential on suppression of BNF-induced lipid peroxidation in mouse hepatic microsome. Therefore, using berberine as a health supplement or an alternative medication might provide extra-benefit due to its inhibitory regulation on CYP1A expression and anti-lipid peroxidation activity.

Induction of EROD activity in European eel (Anguilla anguilla) experimentally exposed to benzo[a]pyrene and beta-naphthoflavone.

The induction of liver ethoxyresorufin O-deethylase (EROD) activity was investigated in the European eel, Anguilla anguilla, collected from a Mediterranean brackish environment and experimentally exposed to benzo[a]pyrene (B[a]P) and beta-naphthoflavone (BNF). Eels were injected intraperitoneally at increasing doses (0.1, 1, 10, and 50 mg/kg wet body weight) using corn oil as a carrier and sacrificed after 7 days. The main objectives of the present study are: (1). to assess of the sensitivity of EROD induction as a biomarker to polycyclic aromatic hydrocarbon (PAH) exposure; (2). to determine an EROD dose-response relationship of the contaminants used; and (3). to compare the efficiency of B[a]P and BNF as inducers of EROD activity. Results showed that both chemicals resulted in a dose-dependent EROD induction, but increases were not linear. EROD activity seemed to reach a plateau at the exposure of 10 mg/kg in both treatment groups; B[a]P was a more potent inducer than BNF was at the higher doses (10 and 50 mg/kg), while the opposite result was observed at the lower ones (0.1 and 1 mg/kg). The greatest induction occurred in eels treated with 10 mg/kg B[a]P, in which a 261-fold increase in EROD activity was observed. Results showed that EROD activity in A. anguilla is significantly induced by B[a]P and BNF exposure, responding to a wide range of concentrations of these contaminants. We infer that this tool may be suited as a diagnostic biomarker for biomonitoring PAHs pollution in Mediterranean brackish environments and further field research is suggested.

Anguilla anguilla L. liver ethoxyresorufin O-deethylation, glutathione S-transferase, erythrocytic nuclear abnormalities, and endocrine responses to naphthalene and beta-naphthoflavone.

The effects of naphthalene (NAP) and beta-naphthoflavone (BNF) on phase I biotransformation and genotoxicity in Anguilla anguilla L. were evaluated. Phase II biotransformation and cortisol levels were also assessed in NAP-treated fish. Two groups of eels were exposed to either a NAP or a BNF concentration range (0.1-2.7 microM) for different exposure periods (2-72 h). An early significant ethoxyresorufin O-deethylation (EROD) activity inhibition was observed, especially for the highest NAP concentrations at 2-6 h exposure and for BNF at 2h exposure. However, a significant EROD activity increase was detected from 16 to 72 h exposure for NAP and from 4 to 72 h exposure for BNF. The cytochrome P450 (P450) content was not dose related. However, with regard to BNF exposure, P450 was the first biomarker to respond. Liver alanine transaminase (ALT) activity was measured as an indicator of hepatic health condition. ALT results demonstrated that the EROD activity decrease, previously described for NAP, was not related to tissue damage. Nevertheless, the highest BNF concentrations were demonstrated to induce liver damage and to impair the EROD activity response. An increased genotoxic response, measured as erythrocytic nuclear abnormalities (ENA), was observed during the first 8h NAP exposure. However, for exposures longer than 8 h, ENA frequency returned to the control levels. This response profile may reflect a considerable DNA repair capacity and/or a metabolic adaptation providing an efficient NAP biotransformation and consequent detoxification. BNF revealed no ENA alterations for all concentrations and exposure lengths. In the NAP experiment a causal relationship between immature erythrocytes (IE) and ENA frequency disappearance was not found. BNF results with regard to IE frequency revealed an ability to alter the balance between erythropoiesis and removal of erythrocytes. Liver glutathione S-transferase activity was significantly induced after 2 and 48 h NAP exposure. A cortisol-impaired response seems to occur from 4 to 24 h NAP exposure, demonstrating an endocrine disruption. However, an adaptation process seems to occur after 48 h, since the plasma cortisol had a tendency to increase. The present findings confirm the usefulness of the adopted biomarkers. The ecological risk associated with aquatic contamination by NAP was also confirmed by the present data.

Induction of DNA strand breakage and apoptosis in the eel Anguilla anguilla.

The ability of benzo[a]pyrene, Aroclor 1254, 2-3-7-8-tetrachlorodibenzo-p-dioxin and beta-naphthoflavone to induce DNA strand breaks (SB) and apoptosis in erythrocytes of the European eel (Anguilla anguilla) was investigated following by in vivo exposure. DNA damage was evaluated by the Comet assay, while the diffusion assay was used to investigate the induction of apoptosis 7 days after a single intraperitoneal administration. 2-3-7-8-Tetrachlorodibenzo-p-dioxin induced the highest genotoxic effect, followed by benzo[a]pyrene, while the other two substances had limited effects. A significant induction of apoptosis was observed at the highest doses after exposure to benzo[a]pyrene, when DNA damage was also elevated. The occurrence of apoptotic cells after exposure to Aroclor, 2-3-7-8-tetrachlorodibenzo-p-dioxin and beta-naphthoflavone was quite variable and did not show clear dose-related responses. The role of oxidative stress in mediating DNA damage was also discussed.

Naphthalene and beta-naphthoflavone effects on Anguilla anguilla L. hepatic metabolism and erythrocytic nuclear abnormalities.

The effects of a polycyclic aromatic hydrocarbon (PAH) such as naphthalene (NAP)--an environmental contaminant--and beta-naphthoflavone (BNF)--a model substance (PAH-like compound)--were investigated in European eel (Anguilla anguilla L.) over 3-, 6-, and 9-day exposure (0.1-2.7 microM). Both xenobiotics revealed to be strong biotransformation (phase I) inducers. After 3-day exposure, liver ethoxyresorufin O-deethylase (EROD) activity was significantly increased by all NAP and BNF tested concentrations. At 6 and 9 days, liver EROD activity was significantly induced mainly by the highest NAP and BNF concentrations. Liver cytochrome P450 content was significantly induced after 3-day exposure to 0.9 and 2.7 microM BNF and 9-day exposure to 0.1, 0.3 and 0.9 microM NAP. Liver alanine transaminase (ALT) activity was measured as an indicator of hepatic health condition, revealing a significant decrease after 6-day exposure to 0.9 microM BNF. Genotoxicity measured as erythrocytic nuclear abnormalities (ENA) was detected in all BNF treated fish on day 6, whereas on day 9, ENA frequencies returned to control levels, significantly decreasing at 0.9 microM BNF exposure. Immature erythrocytes (IE) frequency demonstrated a decreasing tendency along the BNF experiment and concomitantly with the above ENA response. The present experimental results elect EROD activity in A. anguilla as a useful short- to medium-term biomarker of exposure to both PAH and PAH-like compounds. However, some problems can emerge in the presence of high xenobiotic concentrations. Concerning genotoxicity, it is hypothesized that ENA response depends on different factors such as the exhaustion of the detoxification process, the balance erythropoiesis/erythrocytic catabolism and the DNA repairing capacity.

Liver phase I and phase II enzymatic induction and genotoxic responses of beta-naphthoflavone water-exposed sea bass.

Sea bass were exposed to 0 (control), 0.1, 0.3, 0.9, and 2.7 microM beta-naphthoflavone (BNF) for 0, 2, 8, and 16 h in order to assess the chronological and concentration relationships between BNF phase I and II biotransformation responses, such as liver cytochrome P450 (P450) content, ethoxyresorufin-O-deethylase (EROD), uridine diphosphate-glucuronosyl transferase (UDP-GT), and the genotoxic effects, measured either by erythrocytic micronuclei (EMN) or erythrocytic nuclear abnormalities (ENA) tests. Liver alanine aminotransferase (ALT) activity and liver somatic index (LSI) were also measured. A significant liver EROD activity was found at 8 h exposure, respectively, to 0.1, 0.3, 0.9, and 2.7 microM BNF. Maximal liver EROD activity increase was observed at 16 h exposure to 0.9 microM BNF, whereas the highest liver P450 was reached at 8 h exposure to 2.7 microM BNF. Liver UDP-GT activity was significantly increased at 2 h exposure to 0.1 and 0.3 microM BNF and at 8 h exposure to 0.1, 0.3, and 0.9 microM BNF, decreasing at 16 h, for every exposure concentration. Significant ENA increase was observed at 2h exposure, respectively, to 0.3, 0.9, and 2.7 microM BNF. Maximal ENA increase was observed at 16 h exposure to 0.9 microM BNF. The MN was significantly increased at 8 and 16 h exposure, respectively, to 2.7 and 0.9 microM BNF. Liver ALT activity significantly increases at 8 h exposure to 0.1 and 0.3 microM BNF, whereas liver somatic index was significantly increased from 2 to 16 h exposure for every BNF concentration. A slight liver EROD activity increase with a concomitant lack of liver UDP-GT activity is able to induce significant erythrocytic genotoxic effects. Liver UDP-GT high levels are important in sea bass BNF detoxification. However, high liver UDP-GT activity is not enough to prevent the BNF metabolite genotoxic effects on sea bass erythrocytes when liver EROD activity is induced at 2 and 8 h exposure to 0.3 and 0.9 microM BNF. The genotoxic effects measured as EMN and ENA suggest that the balance between the rates of liver BNF reactive and conjugated metabolites seems to be critical.

Beta-naphthoflavone liver EROD and erythrocytic nuclear abnormality induction in juvenile Dicentrarchus labrax L.

Juvenile Dicentrarchus labrax L. (sea bass) was exposed to five different beta-naphthoflavone (BNF) concentrations-0, 0.1, 0.3, 0.9 and 2.7 microM-for 0, 2, 4, 6, 8, 16, 24, 48, 72, 144, and 216 h. A battery of biomarkers was investigated, such as liver ethoxyresorufin-O-deethylase (EROD), liver cytochrome P450 (P450 content), liver aminotransferase (ALT activity), liver somatic index (LSI), micronuclei (MN), and erythrocytic nuclear abnormality (ENA) frequencies. Juvenile D. labrax L. liver EROD induction started at 2 h exposure to 2.7 microM BNF and 6 h exposure to 0.1, 0.3, and 0.9 microM BNF, respectively. A significant liver EROD decrease was observed between 8 and 16 h for all BNF concentrations, followed by a slight increase after 48 h exposure to 0.9 and 2.7 microM BNF and a definitive decrease from 72 h exposure onward. Liver P450 content significantly increased at 2, 6, and 8 h exposure, respectively, to 2.7 microM, 0.9, 0.3, and 0.1 microM BNF. However, liver P450 content remained significantly higher than that of the control from 72 to 216 h in the sea bass exposed to 2.7 microM BNF. Sea bass ENA induction started at 4h exposure to 0.9 and 2.7 microM BNF, and significantly increased to 16 and 24 h exposure, whereas for 0.3 microM BNF a significant increase started after 8 h exposure. A significant ENA frequency increase was still observed at 144 and 216 h exposure to 0.9 and 2.7 microM BNF. The micronuclei induction was observed at 4, 6, and 8 h, respectively, after 2.7, 0.9, and 0.3 microM BNF exposure. However, there was a micronuclei frequency decrease for 0.3, 0.9, and 2.7 microM BNF exposure concentrations between 8 and 16 h, followed by a slight increase after 48, 72, and 144 h exposure, respectively, to 2.7, 0.9, and 0.3 microM BNF. Liver somatic index significantly increased after 216 h, whereas ALT activity significantly decreased at 144 and 216 h 2.7 microM BNF exposure.