Cytotoxic Effects of N-Hydroxyparacetamol in Suspensions of Isolated Rat Hepatocytes

Abstract: The cytotoxicity of N-hydroxyparacetamol (N-OH-pHAA), a postulated proximate metabolite of the hepatotoxic and nephrotoxic analgesic paracetamol, was studied in suspensions of hepatocytes isolated by collagen-perfusion of livers of male rats. Incubation of cells with 0.25–2.0 mM N-OH-pHAA led after 3–5 hours to increased cell permeability measured by increased trypan blue uptake, increased NADH penetration or leakage of prelabelled ⁵¹Cr. N-OH-pHAA rapidly depleted cellular glutathione, 16% of initial levels were seen after 30 min. incubation. ³H-N-OH-pHAA bound covalently to cellular proteins in a time- and concentration-dependent manner, considerably higher binding rates were seen with boiled cells compared to intact cells. Pretreatment of animals with the cytochrome P-450 inducer phenobarbital did not affect N-OH-pHAA cytotoxicity or covalent binding, whereas the cytochrome P-450 inhibitor metyrapone inhibited both cytotoxicity and binding. Lipid peroxidation in hepatocytes could be seen as a late event after a limited range of N-OH-pHAA concentrations. In contrast, lipid peroxidation was an early event in cells exposed to carbon tetrachloride. A minimal exposure time of 30 min. of the hepatocytes to N-OH-pHAA was sufficient to elicit cellular damage occurring after 3–5 hours.