Characterization of 4-hydroxy-2-nonenal metabolism in stellate cell lines derived from normal and cirrhotic rat liver

During oxidative stress, reactive aldehydes, including trans-4-hydroxy-2-no nenal (4-HNE), are generated by peroxidation of membrane lipids and purport edly stimulate hepatic stellate cells to produce excessive extracellular ma trix, including type I collagen. An important question concerning the abili ty of 4-HNE to modulate collagen production by stellate cells is the potent ial of these specialized cells to detoxify 4-HNE. The objective of the pres ent study was to characterize the ability of stellate cell lines, derived f rom normal (NFSC) and cirrhotic (CFSC) rat livers, to metabolize 4-HNE by o xidative, reductive and conjugative pathways. These two stellate cell lines were noted to have differing susceptibilities to the cytotoxic effect of 4 -HNE. Treatment of both stellate cell lines with a range of 4-HNE doses dem onstrated that the concentration which was cytotoxic to 50% of CFSC (TD50) was 25% greater than that for NFSC (967.57 +/- 9.26 nmol/10(6) cells vs. 76 9.90 +/- 5.32 nmol/10(6) cells respectively). The capacity of these cell li nes to metabolizes 4-HNE was determined by incubating them in suspension wi th 50 mu M 4-HNE (10 nmol/10(6) cell); 4-HNE elimination and metabolite for mation were quantified over a 20 min time course. Both stellate cell lines rapidly metabolized 4-HNE, with the CFSC line eliminating 4-HNE at a rate t hat was approx. 2-fold greater than the NFSC line. The rate of 4-HNE metabo lism attributable to glutathione S-transferase (GST) was similar in both ce ll lines, though differential cell specific expressions of GST isoforms GST P1-1 and GSTA4-4 were observed. The greater rate of 4-HNE elimination by CF SC was attributable to its aldehyde dehydrogenase (ALDH) activity which acc ounted for approx. 50% of 4-HNE metabolism in CFSC but was insignificant in NFSC. Neither cell line had detectable alcohol dehydrogenase activity or p rotein levels. Measurement of cellular GSH concentrations revealed that NFS C contain approx. 2-fold greater concentrations of GSH when compared to CFS C and that following 4-HNE treatment, GSH levels were rapidly depleted from both cell lines. Concomitant with 4-HNE mediated GSH depletion, a correspo nding increase in the 4-HNE-glutathione adduct formation was observed with the NFSC line forming greater amounts of the glutathione adduct than did th e CFSC line. Taken together, these data demonstrate that both stellate cell lines have the capacity to metabolize 4-HNE but that CFSC have a greater r ate of metabolism which is attributable to their greater ALDH activity, sug gesting that the stellate cells isolated from cirrhotic Liver may be differ entially responsive to the biologic effects of 4-HNE. (C) 2000 Elsevier Sci ence B.V. All rights reserved.