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Properties and regional expression of a CYP3A-like protein in channel catfish intestine
James, M.O.; Lou, Z.; Rowland-Faux, L.; Celander, M.C. (2005). Properties and regional expression of a CYP3A-like protein in channel catfish intestine. Aquat. Toxicol. 72(4): 361-371
In: Aquatic Toxicology. Elsevier Science: Tokyo; New York; London; Amsterdam. ISSN 0166-445X, more
Peer reviewed article  

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Keywords
    Cytochromes; Toxicity; Brackish water; Fresh water

Authors  Top 
  • James, M.O.
  • Lou, Z.
  • Rowland-Faux, L.
  • Celander, M.C.

Abstract
    Biotransformation in the intestine may influence the bioavailability and toxicity of ingested xenobiotics. The objective of this study was to examine the expression and catalytic properties of a constitutive cytochrome P450 (CYP) 3A-like protein along the intestine of channel catfish, Ictalurus punctatus. Fish were maintained on commercial chow or nutritionally complete semi-purified diets. Polyclonal antibodies generated against rainbow trout CYP3A proteins reacted strongly with catfish washed intestinal microsomes on Western blots showing a major protein band with MW of 59kDa. In catfish maintained on a standard chow diet, the expression of this protein was higher in the proximal segment (0.101 ± 0.031units/mg protein, mean ± S.D., n=4) than in the distal part (0.032 ± 0.023units/mg protein). Microsomal testosterone 6beta-hydroxylation activity was monitored as the catalytic indicator of CYP3A, and was higher in proximal than distal catfish intestine (263 ± 80.3 and 88.6 ± 15.6pmol/min/mg protein for proximal and distal, respectively, mean ± S.D., n=4). CYP3A protein levels and testosterone 6beta-hydroxylation activities were lower in microsomes from the proximal segment of intestine from catfish maintained on a semi-purified diet, compared with commercial chow, but again the proximal intestine had higher CYP3A and 6beta-hydroxylase activities than distal intestine. Testosterone 6beta-hydroxylase activities in all samples correlated with the CYP3A protein levels, r2=0.8. Testosterone 6beta-hydroxylation was inhibited by specific CYP3A inhibitors, ketoconazole (IC50=0.02 µM) and erythromycin (IC50=41 µM), as well as general CYP inhibitors, metyrapone (IC50=2.8 µM) and SKF-525A (IC50=25 µM). There was evidence for the involvement of CYP3A in the mono-oxygenation of benzo(a)pyrene and of (-)-benzo(a)pyrene-7,8-dihydrodiol in intestinal microsomes from catfish maintained on the semi-purified diet. Mono-oxygenation of both substrates was increased in a concentration-dependent manner by in vitro addition of alpha-naphthoflavone. Benzo(a)pyrene hydroxylase activities were higher in proximal than in distal intestine; 3.72 ± 0.77pmol/min/mg protein, mean ± S.D., n=5 and 1.45 ± 0.42 in these respective segments. The results of this study strongly suggest that CYP3A is important in the first pass metabolism of dietary xenobiotics in untreated fish.

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