Abstract 19391: Activation of AMP-activated Protein Kinase Displays a Novel Strategy Against Reperfusion-Induced Barrier Failure in Endothelial Cells
Background: Capillary leakage and edema formation is a complication of ischemia compromising the outcome of reperfusion and the recovery of heart function. Recently it became apparent that the AMP-activated protein kinase (AMPK) not only plays a crucial role as a fuel sensor during hypoxia but also is involved in stabilizing endothelial barrier function. Here we tested the hypothesis that the imminent barrier failure induced by ischemia/reperfusion can be prevented by pharmacological activation of AMPK at the onset of reperfusion.
Methods and Results: Cultured human umbilical vein endothelial cells (HUVEC) were exposed to ischemia (40 min, Po2<5 mmHg; pH 6.4) and reperfusion (40 min, Po2=140 mm Hg; pH 7.4) while control cells were exposed to normoxia. Ischemia induced endothelial barrier failure (interendothelial gap formation (GF), video-imaging technique), activation of the contractile machinery (phosphorylation of myosin light chain (MLC) and MYPT1, actin stress fibre formation), and loss of VE-cadherin/β-catenin at cell adhesions. All these parameters were further aggravated during reperfusion. While AMPK activation (AMPK∼P, phosphorylation) was detected during ischemia, AMPK∼P declined to basal levels rapidly after onset of reperfusion. However, application of the AMPK activators AICAR (5-aminoimidizole-4-carboxamide riboside, 1 mM) or Metformin (1 mM) at the onset of reperfusion preserved AMPK activation, abolished interendothelial GF, MLC and MYPT1 phosphorylation, and induced rearrangement of cortical actin at the cell borders, as well as re-established VE-cadherin/β-catenin at cell adhesions. This protective effect of AICAR and Metformin on all parameters was abolished in presence of the AMPK inhibitor adenine 9-beta-d-arabinofuranoside (1mM) or silencing of AMPK by siRNA.
Conclusions: AMPK is involved in the maintenance of endothelial barrier. Targeted activation of AMPK at the onset of reperfusion abolishes reperfusion-induced hyperpermeability. Therefore, activation of AMPK may be a promising new therapeutic option to prevent reperfusion-induced endothelial barrier failure.
- © 2010 by American Heart Association, Inc.