Transcription factor NF-κB is necessary for up-regulation of type 1 angiotensin II receptor mRNA in rat cardiac fibroblasts treated with tumor necrosis factor-α or interleukin-1β

Abstract

Tumor necrosis factor-α (TNF-α) and interleukin-1β up-regulate type 1 angiotensin II receptor (AT1) mRNA and protein in cultured neonatal rat cardiac fibroblasts. The use of pharmacologic inhibitors and a degradation-resistant mutant IκB-α demonstrated that the transcription factor nuclear factor-κB (NF-κB) is necessary for cytokine-induced AT1, up-regulation. The increase in AT1 mRNA with TNF-α treatment is slow, reaching significance by 6-12 h and peaking by 24-48 h. Electrophoretic mobility shift assays revealed that NF-κB nuclear translocation was maintained for ≥24 h with a single dose of TNF-α. Since prolonged NF-κB activation appeared necessary to maximize AT1 up-regulation, the mechanism of persistent NF-κB activation was studied further. Stimulation with TNF-α induced a >10x increase in IκB kinase (IKK) activity that quickly diminished by 20 min. IκB-α and IκB-β proteins were degraded during this time, and IκB-α was resynthesized subsequently by NF-κB-dependent transcription. However, IκB isoforms and IKK activity did not return completely to unstimulated values during a 12-h time course. These results suggest that low but persistent IKK activity and IκB degradation lead to prolonged NF-κB nuclear translocation and maximal AT1 up-regulation in the continued presence of TNF-α.