302 Inhibitors of the lipoxygenase arachidonic acid pathway impair glycocholate efflux in isolated rat hepatocytes Jorge Quiroga, Jose L. Rodriguez-SanromBn, Francisco Guamer, Carlos Rodriguez Ortigosa, Josh M. Art?jola and Jesds Prieto Dqm”P”l o,l”w”al hhi,Ci”P, ““ivmiry Chic o,i+l”Orro, F%wlp,ono. spoin (Received 19December 1989) Lipoxygennre arachidonic acid metabolires mediate secretory processes in several tissues. but their possible involvement in liver transport functions is still unknown. Thrs study evaluated the influence of the lipoxygenase inhibitor nordihydro- guayaretic acid (NDGA), the cyclooxygenase inhibitor indomethacin (INDO), and the dual cycle and lipoxygenase inhibi- tors 3-amino-l-[m-(tri~uoramethyl)-phenyl]-2-pyr~oline (BW 755~) and eicosatetraynoic acid (ETYA) on the handling of glycocholic acid (GC) by isolated rat hepatocytes. No drug modified cell viability or oxygen consumption in hepato- cytes. In 30.min incubations with 50pM GC the initial rate of GC uptake zyxwvutsrqponmlkjihgfedcba (Vo) in control hepatocytes was 1.15 f 0.03 nmal.mg proteinv’,mbi’. The cellular GC content remained eonstent from 10 to 30 min (steady-state phase), the 30-min value being 6.63 i: 0.35 nmol.mg protein-‘. NDGA (lo-50/rM), BW 755~ (25~200/rM) and ETYA (5-100gM). pre- vented Ihc steady-state phase occurring, thus determining a progressive accumulation of GC in cells with time. As cotn- pared to controls, 50pM NDGA (+37%, p < O.Ol), 2OOpM BW 755~ (+3Y%..u < 0.01) and 5pM ETYA (+19%,p < 0.05) i&red the higheq increases in the amount of GC in cells at 30 min, in all cases zyxwvutsrqponmlkjihgfedcba V, being unchanged. Concentrations of BW 155~ and EPA above those indicated also decreased zyxwvutsrqponmlkjihgfedcbaZYXWVUT V,. Both V, and tbz amount of cellular GC in the steady-state phase were proportionally decreased by high INDG concentrations (2.-1OOpM) which did not modify the morphology of the uptake curve. Since experiments with dual and lipoxygenase inhibirors suggested an impairment of GC &flux, the ini- tial rate of GC efflux (V,,) was measured in hepatocytes prelaaded with 50yM GC and transferred to a GC-free medium. In controls, V,, was 1.12 + 0.12 nmolmg protein-‘.min-‘. BW 755~ (ZOOycM) and NDGA (SOgM) reduced Voet by 45 and 38%. respectively. The kinetic analysis of the effect of 2CQpM BW 755~ on the efflux process using hepatocytes preloaded with GC from 5 to 200pM disciosed a non-competitive inhibition. V,,,, wasreducedfrom1.37i0.15to0.89+0.10@< O.Ol), wherzxs K,,, was unchanged (3.79 + 0.33 K 4.25 f 0.54, N.S.). In summary, inhibitors of the IipoxySenase arachi- donic acid pathway impaired the &flux of CC from isolated rat hepatocytes. The hypothesis is raised that oxidized metabo- lites of arachidonic acid may participate on the secretion of bile salts in these cells. Lipoxygenases (LO) wch as S-LO. 12.LO and IS-LO, tives leukotriene B, and the cysteinyl leukotricnes C,, D, catalyre the oxidation of arachidonic acid at a different and E, which are synthesized by hepatic cells (2,3) and and specific positwn to generate the initial compounds of the LO arachidonic acid oathwav (1). Recent studies demonstrated rcievant r&tionships between the liver and these substances,in particular with the S-LO drriva- play an important role as mediators of liver injury (4). Furthermore, specific recemnrs for Icukotrimr C, have been found in hepatocytes &), cell:; which are able t, take up cysteinylleukotrienes from systemic blood, and mera-