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  • Natural and synthetic glucocorticoids perform their biologic

    2019-08-17

    Natural and synthetic glucocorticoids perform their biological action in the organism through receptor-dependent mechanisms (e.g., by binding to the glucocorticoid receptors). It has been determined that glucocorticoid receptors cross-talk with other nuclear receptors, including AhR (Celander et al., 1996b, Dvorak et al., 2008, Vrzal et al., 2009). Dexamethasone is a typical inducer of mammalian CYP3A (Donato et al., 1995, McCune et al., 2000, Pascussi et al., 2000a, Pascussi et al., 2000b) and CYP2E1 activity (Tamasi et al., 2001). In fish, DEX increased protein expression and CYP1A catalytic activity in hepatocellular carcinoma Tubastatin A from Poeciliopsis lucida (Celander et al., 1996a). In this study, we investigated the in vivo effect of DEX on the catalytic activity of selected hepatic CYP450s in rainbow trout. Because DEX concentrations vary greatly in environmental waters, two environmentally relevant DEX concentrations (3 and 30ngL−1) and two higher concentrations (300 and 3000ngL−1) were included. Activities of the following CYP450s were measured: CYP1A (7-ethoxyresorufin O-deethylase [EROD]), CYP2E1-like (p-nitrophenol hydroxylase [PNPH]) and CYP3A-like (7-benzyloxy-4-trifluoromethylcoumarin O-debenzylase [BFCOD] and 7-benzyloxyquinoline O-debenzylase [BQOD]). Moreover, total CYP450 content as well as CYP1A and CYP3A-like proteins were determined.
    Materials and methods
    Results
    Discussion The induction of CYP450 in fish has been evaluated as a sensitive early warning signal of organic xenobiotics in the aquatic environment (van der Oost et al., 2003). In this sub-chronic in vivo experiment, we observed that DEX at concentrations ranging from 3 to 300ngL−1 can modify CYP450 activity when expressed as amount of product formed per min per nmol total CYP450, and it can affect total CYP450 content. Several recent studies focused on the modulation of CYP450 activity by numerous pharmaceuticals including DEX (Smith and Wilson, 2010, Wassmur et al., 2010). In general, no consistent response of fish CYP450 to DEX was found across published studies. In one study, increased metabolism of a number of CYP450 substrates was found after treating rainbow trout with DEX at a dose of 2mg/kg body weight (Haasch et al., 1994). Rainbow trout injected with DEX at a dose of 100mg/kg body weight showed significant induction in 3-cyano-7-ethoxycoumarin metabolism (mainly metabolised by CYP1A enzymes), while EROD and 7-methoxyresorufin O-demethylation activities were not altered (Smith and Wilson, 2010). Similarly, injection of DEX at 0.03–3μg/kg into male H. malabaricus did not alter EROD activity (Guiloski et al., 2015). CYP3A activity was induced in grass carp after DEX treatment (Li et al., 2008). This dissimilarity in responses can be associated with many factors, such as the age of the fish, type of exposure and differences in experimental concepts. There is evidence that the response of CYP450 activity to DEX is species-specific. CYP1A and CYP3A expressions were strongly induced by DEX in porcine hepatocytes (Rasmussen et al., 2014) and in rat and human, but not minipig or beagle dog hepatocytes (Lu and Li, 2001). Within fish, there are also species-specific differences. Thus, in the same study, the effects of DEX were observed in rainbow trout but not in killifish (Smith and Wilson, 2010). Additionally, DEX induced CYP3A activity (induction of aminopyrine N-demethylase and erythromycin N-demethylase) in two grass carp cell lines (Li et al., 2008). Given that DEX is a well-known inducer of mammalian CYP3A, we included two catalyzed reactions (BFCOD and BQOD) to evaluate CYP3A activity. Both affected rainbow trout in similar ways, and induction followed 21d exposure. In mammals, DEX can initiate transcription of CYP2E1 (Sampol et al., 1997). The CYP2E1-like protein activity has also been identified in fish (Kaplan et al., 1999, Wall and Crivello, 1999, Zamaratskaia and Zlabek, 2011). To date, there are no reports on the in vivo effect of DEX on PNPH activity. In vitro, DEX did not modify PNPH activity in rainbow trout (Burkina et al., 2013); however, we found that PNPH activity was induced by DEX after 21d exposure. Compared to mammals, the regulation of CYP2E1 in fish is less investigated and requires further study. The molecular mechanisms by which DEX regulates PNPH activity appear to be similar to those of other studied CYP450-mediated reactions in fish liver.