Introduction: The Chromogranin A (CgA)-derived peptide Catestatin (CST) is thought as a novel regulator of cardiac function and blood pressure, being able to induce anti-adrenergic, vasodilatory and cardioprotective effects, which are mainly due to nitric oxide (NO) release from endothelial cells. The cellular processes upstream eNOS activation exerted by CST are still unknown, mainly because typical high-affinity CST receptors have not yet been identified. We thus hypothesized that, on the basis of its cationic and amphipathic properties, CST acts as a cell penetrating peptide, binding to heparan sulfate proteoglycans (HSPGs) and activating eNOS phosphorylation through a PI3K-dependent, endocytosis-coupled mechanism.

Methods: In bovine aortic endothelial (BAE-1) cells, endocytotic vesicles trafficking was quantified by confocal microscopy with the membrane dye FM1-43; Caveolin (Cav)-1 shift from plasma membrane was studied by immunofluorescence staining and transfection with GFP-Cav; eNOS phosphorylation was assessed by immunofluorescence and immunoblot analysis.

Results and Conclusions: CST markedly increased both caveolae-dependent endocytosis and eNOS phosphorylation in BAE-1 cells. These effects were reduced by both heparinase III and Wortmannin.

Our results suggest a novel signal transduction pathway involved in the action of CST on endothelial cells: HSPGs interaction and caveolae endocytosis, coupled with a PI3K-dependent eNOS phosphorylation.