Abstract 18953: Phosphoproteomic Analysis in the Endothelial Cells Damaged by Hyperphosphatemia
Recent epidemiological analyses have demonstrated that hyperphosphatemia is a risk factor for cardiovascular disease (CVD) in healthy population as well as chronic kidney disease patients. Hyperphosphatemia also account for a premature aging-like phenotype including shortened lifespan, muscle and skin atrophy, osteopenia, and CVD in Klotho/fibroblast growth factor 23 (FGF23)-deficient mice. However the exact mechanism linking hyperphosphatemia and CVD has not been clarified. We have recently reported that hyperphosphatemia causes not only vascular calcification but also endothelial dysfunction. Several studies including ours demonstrated that phosphate (Pi) influx through sodium-dependent Pi transporter is important signal to mediate both endothelial dysfunction and calcification of vascular smooth muscle cells. In this study, we investigated the signal transduction pathway following Pi influx in bovine and rat aortic endothelial cells by using phosphoproteomics analysis technology. The cells were incubated with control (0.9mM) or high Pi (HP: 3mM) medium. After the incubation, protein extracts were prepared and subjected to phosphoproteomics analysis using anti-phospho-signal transduction molecules antibody. Incubation with HP medium decreased Akt/PKB expression and phosphorylation of p70S6 kinase, Mnk1 at 2 and 7 hr after the incubation. In addition, we found decreases in phosphorylation of AMP-dependent kinase (AMPK), acetyl-CoA carboxylase (ACC) and eNOS at ser1177 at 15 min after HP medium incubation. Phosphate is a substrate for ATP synthase, therefore increase in phosphate influx can stimulate ATP production, resulting that inhibited AMPK, and also inhibited phosphorylation of downstream molecules such as eNOS, ACC. In the endothelium, AMPK plays a central role in regulating mitochondrial function determining sensitivity to oxidative stress. Our results suggest that phosphate influx would modulate AMPK and/or Akt/PKB pathways, and inhibit eNOS activity and mitochondrial function, thus resulting endothelial dysfunction. The metabolic effect of AMPK may be also an important reason why hyperphosphatemia can be a risk factor for CVD and premature aging-like phenotype in Klotho/FGF23 mice.
- © 2010 by American Heart Association, Inc.