The total reduced glutathione content was determined in both
The total reduced glutathione content was determined in both n-acetyl-l-cysteine to examine whether retinol was decreasing glutathione concentrations and thus decreasing the conjugation of NAPQI. Total hepatic (3.9±0.3 vs 4.9±1.0 μmol/g, retinol vs untreated, respectively) and renal (0.4±0.1 vs 0.2±0.1 μmol/g, retinol vs untreated, respectively) reduced glutathione were not significantly different from control when analysed 24 h after the last dose of retinol (Fig. 4).
Discussion We have previously demonstrated that retinol, when administered at 75 mg/kg/day for 3–7 days, did not cause hepatotoxicity as assessed by ALT activity and histopathology (Rosengren et al., 1995, Inder et al., 1999). Both of these studies were conducted in male Swiss Webster mice. Our present investigation was conducted in male BALB/c mice and the results are similar to those we have previously published. Four days of retinol treatment potentiated paracetamol-induced hepatotoxicity in this mouse strain, which indicates that the effect of retinol was not strain specific. We were also interested in determining interactions between retinol and paracetamol in the kidney. While murine nephrotoxicity induced by paracetamol has been reported at doses of 600 mg/kg (Emeigh et al., 1994, Hoivik et al., 1995), we observed no indications of nephrotoxicity following paracetamol dosed at 400 mg/kg. Interestingly, retinol potentiated paracetamol-induced hepatotoxicity but the same combination of treatments continued to show no indication of nephrotoxicity. Therefore, the combination of retinol and paracetamol produced an organ-specific response. To determine whether this organ-specific response was caused by a differential induction of CYP450 isozymes, we investigated the activity of several hepatic and renal CYP450 isoforms following retinol administration. No studies have been published on the effect of retinol on CYP450 isoforms in the mouse kidney; however, a number of studies have previously demonstrated a variable response between either retinoid deficient (Miranda et al., 1979, Siddik et al., 1980, Grover et al., 1985, Ushio et al., 1995, Westin et al., 1997) or supplemented (Badger et al., 1996, Inder et al., 1999, McCarthy et al., 1987, Miranda and Chhabra, 1981, Murray et al., 1991, Ushio et al., 1995, Westin et al., 1997) animals and hepatic CYP450 activity. Results from both hepatic and renal microsomal catalytic activity and Western immunoblotting showed no significant difference between retinol and the control groups for CYP2E1. The liver displayed approximately double the CYP2E1 catalytic activity when compared to the kidney. Differential CYP2E1 activity has been demonstrated in mice, rats, rabbits and hamsters (Arubc et al., 1991, Guengerich and Mason, 1979, Liu et al., 1993, Ueng et al., 1991, Villard et al., 1998). The lack of induction of hepatic CYP2E1 by retinol is supported by previous work from our laboratory that showed no change in CYP2E1 catalytic activity or polypeptide levels in male Swiss Webster mice following 3 days of retinol treatment (75 mg/kg/day) (Inder et al., 1999). Therefore, it appears that this response is not altered by different durations of retinol or different strains of mice. However, in other species retinol and derivatives have been shown to induce CYP2E1. Miranda and Chhabra (1981) showed an increase in hepatic aniline hydroxylation in guinea pigs fed a retinol-supplemented (500 IU retinyl palmitate/g) diet. Furthermore, retinol (75 mg/kg) induced hepatic CYP2E1 polypeptide levels and catalytic activities in the rat (Badger et al., 1996). Interestingly, in this study 7 days of retinol treatment resulted in less CYP2E1 induction than 1 day of retinol. Therefore, any relevance of this finding to our design, utilising a 4-day treatment regimen and a different species, is unlikely. In the mouse, renal CYP2E1 is induced by inhalation of cigarette smoke, which indicates that xenobiotics modulate the activity of this enzyme in the kidney (Seree et al., 1996, Villard et al., 1998). However, our results indicate that retinol does not display this activity in the mouse.