Potentiation of gentamicin nephrotoxicity in the rat by infusion of aprotinin

The present study was undertaken to examine a possible effect of aprotinin, a 6.5-kDa polypeptide with an inhibitory effect on proteolysis, on aminoglycoside nephrotoxicity. Experimental animals (female Sprague-Dawley rats, 175-200 g body wt) were treated for 4 days with 40 mg/kg gentamicin given ip at 12-hr intervals. Aprotinin (40,000 kIU per animal) was infused i.v. over a period of 8 days, using subcutaneously implanted miniosmotic pumps. In protocol A, infusion pumps were placed 4 days before starting gentamicin treatment. In protocol B, pumps were implanted 15-18 hr prior to first gentamicin administration. In addition to rats exposed to both gentamicin and aprotinin (GAP), animals were treated with gentamicin ip+saline i.v. (G), saline ip+aprotinin i.v. (AP), or received only saline by both routes of administration (C). All rats were terminated 4 days after the end of gentamicin dosing. One hour before sacrifice, 200 microCi of [3H]thymidine was given ip to each animal in order to monitor cell turnover in renal tissue. The kidneys were analyzed with respect to (i) histopathological alterations and renal dysfunction, (ii) aminoglycoside tissue accumulation, and (iii) tubular regeneration (measurement of cell proliferation). In animals receiving aprotinin alone, histological examination of renal cortex on paraffin sections disclosed mild tubular injury with focal cell necrosis. In plastic-embedded tissue, proximal tubule epithelium was characterized by the presence of numerous inclusions densely stained with toluidine blue. At the ultrastructural level, these inclusions appeared filled with amorphous electron-dense material. In gentamicin-treated animals, cortical drug accumulation reached values higher than 0.3 mg/g renal tissue, but a significant 30-40% decrease of gentamicin accumulation was noted in GAP groups, compared to G groups. Histological examination of renal cortex (paraffin sections) revealed the development of acute tubular necrosis in both G and GAP groups. Tubular injury was accompanied by mild renal dysfunction, as shown by the level of serum creatinine which was increased almost 3-fold in the G group, compared to C and AP groups. Aprotinin infusion produced a further increase of serum creatinine, particularly in protocol A where it was 72% higher for the GAP group than for the G group. In both G and GAP groups, postnecrotic tubular regeneration was evidenced by determining the rate of DNA synthesis and the frequency of S-phase cells in renal cortex. Both methods gave consistent results and showed a 8- to 13-fold increase of cell proliferation in groups receiving gentamicin alone, compared to C groups.