The central hypothesis is that autophagy is downregulated in the settings of advanced age and metabolic syndrome, leading to attenuation ofthe endogenous cardioprotective response and increased inflammation, which in turn results in pathologic remodeling of the heart in response to ischemic injury. In the heart, the predominant targets of autophagy are mitochondria. Impaired mitophagy results in inefficient ATP production and excessive production of reactive oxygen species, leading to cellular dysfunction and inflammatory signaling.
Specific Aim One will examine the molecular basis of mitochondrial turnover in the heart. We have shown that Parkin mediates mitophagy and is required for ischemic preconditioning (IPC), and we now examine the effect of statins to trigger mitophagy and Parkin-dependent cardioprotection. We hypothesize that Parkin-dependent mitophagy is a prerequisite for biogenesis. We will examine the role of the ATP- sensitive potassium channel (mitoK[ATp]) and the permeability transition pore in Parkin translocation.
Specific Aim Two will use organelle flow cytometry to isolate mitochondrial subpopulations based on the fluorescent protein. Timer, targeted to the mitochondrial matrix (MitoTimer), as well as on mitochondrial DNA synthesis (BrdU or EdU incorporation), membrane potential (Rhodamine123), and protein import (Tat-Ndi1-TexasRed). The sorted mitochondrial subpopulations will be used for proteomic studies of post-translational modifications (notably protein carbonylation and thiol modifications, phosphorylation, and acetylation), as well as protein composition. Impairing mitophagy may increase accumulation of posttranslational modifications arising from oxidative stress, while stimulating mitophagy will lead to replacement of dysfunctional mitochondria that have a low threshold for opening of the mitochondrial permeability transition pore. The relative abundance of these molecular signatures of mitochondrial turnover identified in the mouse models will be used to quantify turnover in non-transgenic animals.
Specific Aim Three will assess the effect of metabolic syndrome and advanced age on mitophagy and biogenesis, and will assess functional consequences of impaired mitochondrial turnover with respect to cardioprotection. We will also assess the effect of pharmacologic stimulation of autophagy (and mitophagy) with respect to mitochondrial function and cardioprotection. Flow cytometric analysis of autophagy in lymphocytes will be used to develop a surrogate marker of autophagy suitable for noninvasive studies in humans. These studies will identify molecular signatures of mitochondrial turnover in heart tissue. An understanding of the relationship between mitochondrial turnover and the heart's response to ischemic stress in the contest of metabolic syndrome and aging may lead to new therapeutic approaches for heart disease in patients with comorbid conditions.

Public Health Relevance

These studies will help us understand why cardiovascular outcomes are worse in the elderly or those with obesity and features of metabolic syndrome. We will use novel methodology to monitor mitochondrial destruction and replacement in mice, which will be applied in large animals. These biochemical signatures may be used to indicate which drugs are likely to improve outcome in patients with heart disease.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Program Projects (P01)
Project #
7P01HL112730-02
Application #
8683227
Study Section
Heart, Lung, and Blood Initial Review Group (HLBP)
Project Start
Project End
Budget Start
2014-07-21
Budget End
2015-06-30
Support Year
2
Fiscal Year
2014
Total Cost
$465,715
Indirect Cost
$191,765
Name
Cedars-Sinai Medical Center
Department
Type
DUNS #
075307785
City
Los Angeles
State
CA
Country
United States
Zip Code
90048
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Stotland, Aleksandr; Gottlieb, Roberta A (2016) α-MHC MitoTimer mouse: In vivo mitochondrial turnover model reveals remarkable mitochondrial heterogeneity in the heart. J Mol Cell Cardiol 90:53-8
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