Abstract
Hepatocytes were cultured in the presence of recombinant tumor necrosis factor (TNF) alpha or mutated TNF alpha peptides that specifically activate either p55 or p75 TNF receptors to determine if TNF alpha can activate cytokine-inducible CCAAT/enhancer binding protein (C/EBP) isoforms by post-transcriptional mechanisms that are initiated by TNF receptors. Within 5-10 min after treatment with any of these agents, nuclear concentrations of C/EBP beta and C/EBP delta double and remain 2-4-fold greater than control cultures for 30 min (p < 0.01). Consistent with these results, gel mobility shift assays demonstrate 3-fold increased nuclear C/EBP beta- and C/EBP delta-DNA binding activity in TNF alpha-treated cells, and immunocytochemistry confirms rapid redistribution of these C/EBP isoforms into the nucleus. In contrast, mRNA and whole cell protein concentrations of C/EBP beta and delta are not altered by TNF alpha exposure, and nuclear concentrations of another C/EBP isoform, C/EBP alpha, are decreased by 80%. This novel evidence that TNF alpha initiates post-transcriptional activation of cytokine-inducible C/EBP isoforms identifies a mechanism that enables hepatocytes to respond immediately to inflammatory stress.
Highlights
Because IL-1 and IL-6 are induced after partial hepatectomy (PH) [15, 16], it is unclear if tumor necrosis factor (TNF) ␣ or other cytokines are responsible for the observed variations in CCAAT/enhancer binding protein (C/EBP) expression that occur in the regenerating liver
Recombinant TNF ␣ can down-regulate C/EBP ␣ expression in several cell lines [27], an indirect role for TNF ␣ in C/EBP  and C/EBP ␦ induction has been presumed because, in vitro, recombinant IL-1 or IL-6 can reproduce most of changes in C/EBP-regulated gene expression that occur in vivo during an acute inflammatory response
The present results demonstrate that at least in this conditionally transformed adult hepatocyte line, TNF ␣ interacts directly with its receptors to rapidly increase nuclear concentrations of these C/EBP isoforms
Summary
TNF ␣ induces Jun nuclear kinase, which phosphorylates and transcriptionally activates c-Jun, promoting increased expression of this transcription factor [2]. TNF ␣ can induce NF- activity by triggering events that favor redistribution of this transcription factor from the cytosol to the nucleus [3,4,5]. Evaluation of the regenerating liver after partial hepatectomy (PH) suggests that TNF ␣ may operate post-transcriptionally to activate members of the CCAAT/enhancer binding protein (C/EBP) family of transcription factors. Nuclear concentrations of C/EBP  and C/EBP ␦ proteins increase early during the prereplicative period after PH (6 –10) These increases are inhibited in animals pretreated with neutralizing anti-TNF antibodies, there is no difference in post-PH induction of their respective mRNAs [7].
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