Vol. 29, Issue 12, 1535-1538, December 2001

In Vivo Perturbation of Rat Hepatocyte Canalicular Membrane Function by Diclofenac

Benedetta C. Sallustio and Felicity L. Holbrook

Department of Clinical Pharmacology, The Queen Elizabeth Hospital, Woodville, South Australia (B.C.S., F.L.H.); and Department of Clinical and Experimental Pharmacology, the University of Adelaide, Adelaide, South Australia (B.C.S.)

Clinical use of diclofenac is associated with a small but significant incidence of hepatotoxicity. It has been reported that in vivo diclofenac treatment results in decreased activity of the extracellular canalicular membrane protein dipeptidylpeptidase IV in rats as a consequence of protein adduct formation by its electrophilic metabolite diclofenac acyl glucuronide. The present study has investigated the effects of in vivo diclofenac treatment (15 mg/kg/day for 7 days) on the activity of an another four rat extracellular canalicular membrane proteins. Animals administered diclofenac (n = 6) had 47.9, 60.4, and 51.6% lower (p < 0.05) canalicular activities of -glutamyltransferase, Mg²⁺-ATPase, and leucine aminopeptidase, respectively, compared with controls (n = 6), but there was no difference in alkaline phosphatase activity. In general, protein adduct formation by acyl glucuronides has been associated with decreased protein function, and the lower canalicular enzyme activities in diclofenac-treated rats may suggest that -glutamyltransferase, Mg²⁺-ATPase, and leucine aminopeptidase are also targets of adduct formation by acyl glucuronide metabolites of diclofenac. However, intracellular redistribution and/or decreased synthesis of these enzymes would also be consistent with our results. The ability of diclofenac acyl glucuronide (200 µg/ml) to form covalently bound adducts with -glutamyltransferase (10 mg/ml) was demonstrated following in vitro incubations (16 h, pH 7.4, and 37°C) in which 20.7 ± 2.1 ng of diclofenac were covalently bound per milligram of protein. In these in vitro studies, the low concentration of protein adducts formed was not associated with any significant change in -glutamyltransferase activity.