Abstract 14378: An Adenosine Monophosphate Activated Protein Kinase / Activating Transcription Factor-1 Cascade Initiates Commitment to Mhem Atheroprotective Macrophages in Response to Heme or Metformin
Background: Intraplaque hemorrhage (IPH) is now viewed as a key driver in atherogenesis. Mhem is a macrophage phenotype that responds to IPH and paradoxically exhibits less inflammatory activation, foam cell formation and oxidant stress than other plaque macrophages. This active protection is directed by activating transcription factor 1 (ATF1), itself activated by phosphorylation. The ATF1-Mhem pathway probably reduces the atherogenicity of IPH, Understanding the mechanisms of this protection may facilitate anti-atherosclerotic therapies.
Methods: We tested the hypothesis that heme in pathologically relevant concentration (10uM) activates the ATF1-Mhem pathway via AMP-activated protein kinase (AMPK) in human blood-derived macrophages, using standard cell and molecular biology methods.
Results: Heme (10uM) activated AMPK, and thence a downstream ATF1-HO-1/LXR network leading to the Mhem phenotype. Heme increased macrophage phospho-AMPK (20.8-fold, peak at 7-8mins), phospho-ATF1 (4-fold) and its target genes (HO-1 9.6±0.8-fold, ATF1 5±0.8-fold, transcript levels by qPCR) each prevented by the AMPK antagonist compound-C, or by AMPK-knockdown with RNAi (each p<0.05, Student’s t-test). The AMPK-activating antidiabetic drug metformin, at a clinically relevant concentration (10uM), activated ATF1, and reproduced key therapeutic features of the Mhem phenotype. The effects of metformin were inhibited by AMPK-knockdown and included suppression of macrophage oxidative stress (p<0.05, t-test, n=5 donors); increased cholesterol export (p<0.05, t-test, n=5 donors); protection from foam cell formation [*p<0.05, ANOVA, % macrophage foam cells, mean ± SE, n=5 donors, control-RNAi 19±2%, AMPK-RNAi 23±2%, control-RNAi + metformin 2±0.5 %, AMPK-RNAi + metformin 23±2 %]; and suppression of macrophage inflammatory activation (HLA-DR expression, ~1 log suppression of fluorescence, n=3 experiments).
Conclusions: Our data indicate that heme activates the atheroprotective ATF1 pathway in human macrophages via AMPK, and that a similar response occurs following treatment of cells with metformin. Our results add to the increasing evidence that metformin may have vascular protective effects beyond its role in treating hyperglycemia.
- © 2012 by American Heart Association, Inc.