Angiotensin II Induces Nuclear Factor- κ B Activation in Cultured Neonatal Rat Cardiomyocytes Through Protein Kinase C Signaling Pathway

Rat neonatal ventricular cardiomyocytes (RNVM) possess G protein-coupled AT1receptors for angiotensin II (AngII) that activate multiple intracellular pathways. To elucidate potential signaling mechanisms involved, we focussed on the nuclear transcription factor-kappa B (NF- κ B) in RNVM culture. Using specific antibody to NF-κ Bp65, immunolocalization of NF- κ B was cytoplasmic in unstimulated cardiomyocytes, whereas NF-κ B was translocated into the RNVM nucleus in response to AngII. This translocation was inhibited in the presence of calphostin C, a specific inhibitor of protein kinase C (PKC). Western blot analysis showed an increase of NF- κ B in AngII-stimulated cardiomyocyte nuclear extracts as compared to controls. Biomolecular interaction analysis (BIA analysis) of NF-κ B activation showed that only AngII-nuclear extracts bound to NF- κ B consensus sequence with a high degree of affinity. This DNA-binding capacity was completely lost in calphostin C-treated cells. At transcriptional level in RNVM, AngII mediates the upregulation of matrix gelatinase (MMP-9), which is totally inhibited by calphostin C treatment. In conclusion, cardiomyocyte nuclear NF- κ B translocation in response to Ang II via PKC pathway activates cardiomyocyte-specific transcription of MMP-9 and may activate transcription from responsive genes which are involved in cardiac hypertrophy process and/or cardiac remodeling.