Inter-species comparisons of hepatic cytochrome P450 enzyme levels in male ruminants

Abstract

Our current knowledge about the biotransformation enzymes in wild ruminants is limited. The present study aimed to compare basic levels and specific activities of cytochrome P450 isoforms (CYP1A, 2A, 2B, 2C, 2D, 2E, 3A, 4A) in males of red deer ( Cervus elaphus), fallow deer ( Dama dama), roe deer ( Capreolus capreolus) and mouflon ( Ovis musimon). The proteins from the major cytochrome P450 (CYP) subfamilies were detected in all ruminant species by Western blotting, using polyclonal antibodies raised against rat or human CYP enzymes. The immunochemical data seem to suggest that humans and wild ruminants share some similar hepatic CYP enzymes corresponding to members of subfamilies 2 and 3; ruminant liver samples also contained two proteins cross-reacting with anti-rat CYP1A antibodies. High activities of CYP1A enzymes found in liver microsomes of male fallow deer and roe deer are indicative of increased susceptibility of these species towards promutagens that are metabolically activated by these CYPs. On the other hand, low activities of CYP1A-dependent alkoxyresorufin O-dealkylase activities were detected in male mouflons. Oxidative metabolism of testosterone was significantly higher in wild ruminants than the values previously reported from bulls. Androstene-3,17-dione and 6beta-hydroxytestosterone were the most important products of testosterone oxidation in liver microsomes of all the ruminant species under study. The highest CYP3A-dependent testosterone 6beta-hydroxylase activity was found in mouflons and fallow deer. A different pattern of CYP activities towards testosterone was found in roe deer, which showed high activities of testosterone 2beta-hydroxylase and lower production of androstene-3,17-dione. An increased activity of CYP4A-dependent laurate 12-hydroxylase found in roe deer and mouflons might indicate a higher metabolic turnover of fatty acids. The data on CYP activities indicated that high metabolic rates of steroids, fatty acids, and xenobiotics may occur in male wild ruminants. The highest hepatic activities specific for CYP3A, CYP2C, CYP2D, and CYP2E enzymes were found in mouflon, suggesting that this species has the highest biotransformation capacity.

Effect of breed and gender on bovine liver cytochrome P450 3A (CYP3A) expression and inter-species comparison with other domestic ruminants

“…In our study, the CYP3A-dependent N-demethylations of erythromycin and ethylmorphine were significantly higher in small ruminants (GT > OV) than in CA, confirming previously published results [7,47,48,51,58]. However, no species-differences were observed in the hydroxylation of testosterone at the 6ß-and 2ß-positions, two reactions catalysed by CYP3A also in ruminants [30]. In a recent paper, a relatively high 6ß-OHT, compared to GT or CA, was noticed in OV [55].…”

“…Based on our results and bibliographic evidence, no clear-cut conclusions can therefore be made. On a literature basis, it is known that among the most investigated constitutional factors are the species-differences in P450-dependent DME activities [35]; this is clearly evident for ruminants [10,30,46,51,53,55]. In the present study, no difference was noticed in P450 concentrations; by contrast, the amount of CYP3A apoprotein was significantly higher in OV as compared to cattle (CA); the order of magnitude was CA < GT < OV and these results confirm the presence of species-differences in the expression of some P450 isoforms and their lower expression, including CYP3A, in cattle vs. small ruminants [55,58].…”

Modulation of porcine cytochrome P450 enzyme activities by surgical castration and immunocastration

“…The activities of CYP1A1 and CYP1A2 are usually measured as a rate of the O-dealkylation of ethoxy-and methoxyresorufin, respectively (Burke et al, 1994;Machala et al, 2003), though certain non-specificities of resorufin substrates were found in different species. CYP2B activities can be measured as a rate of the O-dealkylation of pentoxyresorufin (Burke et al, 1994;Machala et al, 2003). In pigs, CYP2E1 and CYP2A are both involved in the metabolism of skatole, a major cause of faecal-like odour in meat from entire male pigs (Diaz and Squires, 2000).…”

Effect of Breed upon Cytochromes P450 and Phase II Enzyme Expression in Cattle Liver

“…A few studies, mostly addressed to investigate the post-translational effects of species, age, and gender (some of the constitutional factors influencing the overall biotransformation capability; Nebbia, 2001), upon the cattle liver DMEs have been published in the past decade (Machala et al, 2003;Nebbia et al, 2003;Sivapathasundaram et al, 2003b;Szotáková et al, 2004;Gusson et al, 2006). In contrast, only recently, DME expression and regulation have begun to be studied at the gene expression level (Greger et al, 2006).…”

“…Likewise, commercially available antibodies, raised toward human and rat antigens, were used for CYP1A, 2B, 2C, 2E, and 3A immunoblotting; in fact, to the best of the authors' knowledge, mono-or polyclonal antibodies directed to cattle antigens have not yet produced so far. In effect, such an approach has been already adopted, at least partly, in prior studies where cattle liver drug metabolism was compared with that of other ruminants, other farm animals, or reference species (i.e., the rat) as well (Sivapathasundaram et al, 2001(Sivapathasundaram et al, , 2003aMachala et al, 2003;Nebbia et al, 2003Nebbia et al, , 2004Szotáková et al, 2004;Dacasto et al, 2005;Gusson et al, 2006;Ioannides, 2006). On the contrary, bovinespecific primer pairs were designed for P450s and phase II enzyme isoform mRNA to be used in the relative quantification by means of quantitative real-time polymerase chain reaction (Q RT-PCR).…”