Synergistic induction of interleukin-6 expression by endothelin-1 and cyclic AMP in adipocytes

Abstract

We have demonstrated previously that endothelin-1 (ET-1) may stimulate interleukin-6 (IL-6) release from 3T3-L1 adipocytes. In this study, we further examined the combined effect of ET-1 and cyclic adenosine monophosphate (cAMP) on IL-6 release. IL-6 release was measured by enzyme-linked immuosorbent assay. Reverse transcriptase-PCR and real-time PCR analyses were used to determine cellular mRNA levels. A luciferase reporter driven by promoter (-1310/+198) of mouse IL-6 gene was transfected into 3T3-L1 adipocytes to monitor IL-6 transcription. ET-1 and cAMP induced IL-6 release in a synergistic manner that can be attributed to their synergistic induction of IL-6 gene expression, as evidenced by IL-6 mRNA analysis and the IL-6 promoter reporter assay. Both ET(A) and ET(B) receptors seem to be involved. In addition, enhanced IL-6 promoter activity can be similarly induced by ET-1 and catecholamines (epinephrine and norepinephrine). The cooperative interaction between ET-1 and cAMP on IL-6 expression seems distinctive, as no other proinflammatory cytokines, such as tumor necrosis factor-α (TNF-α) and IL-1β, are similarly affected. In fact, cAMP inhibited ET-1-stimulated TNF-α and IL-1β expressions in adipocytes. Furthermore, injection of mice with epinephrine and ET-1 induced a tremendously synergistic increase in serum IL-6 levels. Nevertheless, whereas cAMP induced IL-6 expression in RAW264.7 mouse macrophages, ET-1 had no effect on either the basal or the cAMP-induced IL-6 expression. ET-1 and epinephrine may boost plasma IL-6 levels in mice in a synergistic manner, probably through their synergistic induction of IL-6 expression in adipocytes. This study should provide a new perspective for treating IL-6-related diseases, especially those accompanied with elevated ET-1 and catecholamine levels.