Cross-talk between the transcription factor Sp1 and C/EBPβ modulates TGFβ1 production to negatively regulate the expression of chemokine RANTES

RANTES is a key chemokine for atherosclerosis, and obesity is associated with progression of atherosclerosis. Substance P (SP) increases glucose uptake and accumulation of lipids in adipocytes, and SP may upregulate RANTES expression. This study investigated the mechanism of RANTES expression by human M1 macrophages stimulated with SP. SP upregulated RANTES protein expression, whereas aprepitant (an NK1R antagonist) blunted this response. Pretreatment of macrophages with BIRB796 (a combined p38γ/p38δ inhibitor) led to a significant decrease of RANTES expression. Next, we investigated the effect of several NK1R internalization factors on RANTES expression, including GRK2, β-arrestin 2, dynamin, ROCK, and TGFβ1. Exposure of macrophages to SP upregulated TGFβ1 expression. Silencing of β-arrestin 2 or GRK2 significantly enhanced the RANTES protein level after stimulation by SP, whereas TGFβ1/2/3 siRNA or dynasore (a dynamin inhibitor) decreased RANTES and Y-27632 (a ROCK inhibitor) had no effect. Surprisingly, silencing of transcription factor specificity protein 1 (Sp1) or inhibition of Sp1 activity by mithramycin led to significant upregulation of TGFβ1 protein and corresponding enhancement of RANTES expression (by ELISA or western blotting), whereas siRNA for C/EBPβ attenuated expression of both TGFβ1 and RANTES. Next, we investigated transcriptional cross-talk among Sp1 and C/EBPβ, TIF1β, or Fli-1 in relation to RANTES expression. Compared with TIF1β or Fli-1 siRNA, C/EBPβ siRNA showed significantly stronger inhibition of RANTES production by Sp1 siRNA-transfected macrophages after stimulation with SP. In conclusion, transcription factor Sp1 engages in cross-talk with C/EBPβ and modulates TGFβ1 production to negatively regulate RANTES expression in macrophages stimulated with SP. In conclusion, cross-talk between the transcription factor Sp1 and C/EBPβ modulates TGFβ1 production to negatively regulate expression of the atherogenic chemokine RANTES in SP-stimulated macrophages, while RANTES is upregulated by SP via the p38γδMAPK/C/EBPβ/TGFβ1 signaling pathway.