Tumor necrosis factor α differentially modulates the cellular response of rat hepatocytes in periportal- and pericentral-equivalent cultures

Abstract

Alterations of cellular functions induced by recombinant human tumor necrosis factor α (TNFα) were compared in rat hepatocytes cultured under either periportal-equivalent (10 nM insulin; 10 nM glucagon; 13% O 2) or perivenous-equivalent conditions (10 nM insulin; 1 nM glucagon; 4% O 2). TNFα induced a time- and dose-dependent increase in nitric oxide (NO) production and an acute phase response (inhibition of albumin secretion and elevation of α 2-macroglobulin production) under both culture conditions. NO production was more pronounced in periportal cultures, while the acute phase response was stronger in pericentral cultures. This suggests that NO production and the acute phase response are controlled by different pathways. After exposure to TNFα, DNA content was measured fluorimetrically and biochemically. A marked decrease in nuclear DNA content was found exclusively in pericentral cultures after an 8-h exposure, followed by an elevation of lactic dehydrogenase (LDH) release after a 12-h exposure. Aurintricarboxylic acid (100 μM), an inhibitor of endonuclease, significantly inhibited the TNFα-induced decrease in nuclear DNA content but only partially inhibited the LDH release. This indicates that the loss of nuclear DNA content in pericentral cultures is due to an activation of endonuclease and the resulting DNA fragmentation and does not correlate with NO production. Furthermore, the release of LDH seems to be only partially associated with DNA damage. Dexamethasone (100 nM) completely inhibited both TNFα-induced DNA fragmentation and the elevation of LDH release. The results clearly indicate that the toxicity of TNFα is influenced by the metabolic state of hepatocytes. Accordingly, the preferential perivenous cell injury observed after exposure to endotoxins in vivo seems to be due to a higher sensitivity of the pericentrally localized hepatocytes towards TNFα rather than a TNFα concentration gradient.