Abstract P287: Therapeutic Hypothermia Cardioprotection During Cardiac Arrest Alters Reperfusion-Associated mTORC1 and AMPK Signaling
Introduction: Therapeutic hypothermia (TH) is highly cardioprotective in cellular and animal models of ischemia/reperfusion (I/R) and cardiac arrest and is dependent upon Akt. The mammalian target of rapamycin complex 1 (mTORC1) is a downstream target of Akt and an important regulator of cellular energy utilization and survival but its relationship to TH is not clear. We hypothesized that TH attenuates mTORC1 activation while simultaneously activating the cell survival kinases AMP activated protein kinase (AMPK) and Stat3. We further hypothesized that inhibitors of mTORC1 or activators of AMPK would replicate and/or enhance TH cardioprotection.
Methods: Cardiomyocytes isolated from 1-2-day old C57BL6/J mice, were exposed to simulated I (90 min)/R (3 h). For TH, cells were cooled from 37C to 32C during ischemia and the first hour of reperfusion. Heart samples were also obtained from C57BL/6 mice that underwent an 8-min cardiac arrest. Following 6 min, the mice were randomized to normothermia (NT, 37C) or TH (30C) extended during CPR and for 1 h after resuscitation. Protein lysates were collected at serial time points for Western blot analyses.
Results: Cardiomyocyte death during ischemia was minimal, but accelerated during reperfusion (4% vs. 46%,p<.05 n=5). TH attenuated this cell death (24% p<.05 n=5) and attenuated reperfusion-induced mTOR activation seen within 30 min as measured by differential phosphorylation of the mTOR targets 70S6K and eEF2. Compared to NT, TH-treated cardiomyocytes and hearts demonstrated increased p-AMPK after resuscitation, consistent with mTOR inhibition. Stat3 also showed enhanced phosphorylation on tyrosine 705. The mTOR inhibitor rapamycin (100nM) given during I/R similar to TH affected 70S6K and eEF2 and significantly reduced cell death (28% vs. 42%, p<.05 n=5). The AMPK activator metformin (100uM) also significantly reduced cell death (22%, p<.05 n=4).
Conclusion: TH cardioprotection is associated with altered signaling events suggesting mTORC1 inhibition and activation of the cell survival kinases AMPK and Stat3. Pharmacologic strategies that mimic these TH effects are available and could be useful as TH adjuncts or mimetics that improve survival after cardiac arrest.
- © 2011 by American Heart Association, Inc.