ferase reporter gene construct under the control of a PCK In cultured rat hepatocytes, the gluconeogenic key enzyme, phosphoenolpyruvate carboxykinase (PCK), is induced by gene promoter fragment (base 0979 to base /32). Luciferase activity was determined after stimulation of the cells with glucagon via elevation of cyclic 3,5 adenosine monophosphate (cAMP). The proinflammatory cytokine, interleukin-6 CPT-cAMP in the absence or presence of IL-6. CPT-cAMP increased luciferase activity by 1.7-fold, which was inhibited (IL-6), which in the liver together with IL-1b and tumor necrosis factor a triggers the acute-phase response, had been shown in the presence of IL-6. It is concluded that IL-6 had a dual inhibitory effect on the stimulation of PCK gene expression to attenuate the glucagon-induced increase in PCK gene transcription, messenger (mRNA) levels, and enzyme activity. The by glucagon. It inhibited the increase in cAMP at a site before cAMP formation by adenylate cyclase and at a site after cAMP molecular mechanism of this inhibition was investigated in the present study. Glucagon increased cyclic cAMP and PCK formation, the activation of the PCK gene promoter by cAMP.
(HEPATOLOGY 1997;26:73-80.) mRNA levels to a transient maximum twofold and fivefold, respectively. The increases were attenuated by IL-6. Forskolin, which stimulates adenylate cyclase activity, increased
In the liver, the proinflammatory cytokine, interleukin-6 cAMP and PCK mRNA levels 1.6-fold and fivefold, respec-(IL-6), triggers the acute-phase response, which is mainly tively. However, IL-6 attenuated the forskolin-stimulated incharacterized by the increased synthesis and secretion of poscrease in PCK mRNA but not the increase in cAMP. This itive acute-phase plasma proteins and the decreased synthesis showed that IL-6 inhibited PCK mRNA increase in part by and secretion of negative acute-phase plasma proteins (for the attenuation of cAMP increase, but also beyond cAMP review, see Heinrich et al. 1 and Akira et al. 2 ). IL-6 binds to formation. This was confirmed in experiments in which PCK its 80-kd receptor at the cell surface, which associates with mRNA levels were increased by the nonhydrolyzable cAMPa dimer of the signal-transducing 130-kd glycoprotein, analogue, chlorophenylthio (CPT)-cAMP. The increase in PCK gp130. 3,4 After the binding of IL-6 to its receptor, signal mRNA was again attenuated by IL-6. In pertussis toxin-and transduction is accomplished by two different routes in the in isobutylmethylxanthine-treated hepatocytes, IL-6 still incell. Phosphorylation on threonine of the basic leucine zipper hibited the glucagon-stimulated increase in cAMP, indicating transcription factor, C/EBPb (CAAT-box enhancer binding that IL-6 did not activate an inhibitory G-protein or phosphoprotein b) (also termed NF-IL-6, IL-6-DBP, or LAP), leads diesterase, which could cause the impairment of cAMP into transcriptional activation of C/EBPb, which binds to C/ crease. To demonstrate whether the inhibition of PCK gene EBPb-binding sites in the promoter of IL-6-responsive expression by IL-6 beyond cAMP might be caused by the genes, and thereby stimulates the expression of acute-phase inhibition of the activation of the PCK gene promoter by proteins. 5 The second signal-transduction pathway is medi-cAMP, cultured rat hepatocytes were transfected with a luciated by the gp130-associated transcription factor, STAT3
(signal transducers and activators of transcription; also termed APRF, acute-phase response factor), which, after tyrosine phosphorylation by gp130-associated JAK/TYK tyro-Abbreviations: IL-6, interleukin-6; C/EBPb, CAAT-box enhancer binding protein sine kinases translocates to the nucleus, binds to DNA b; PCK, phosphoenolpyruvate carboxykinase; cAMP, cyclic 3,5 adenosine monophossequences in the promoter of IL-6-responsive genes of acutephate; CREB, cAMP response element-binding protein; mRNA, messenger RNA; CPT, chlorophenylthio; RLU, relative light units.
phase proteins, and thereby stimulates transcription of these
From the Institut fu ¨r Biochemie und Molekulare Zellbiologie, Georg-August-Unigenes. 6,7 versita ¨t, Go ¨ttingen, Germany.
The expression of the gene of the hepatic key gluconeo-