Animal Research
Mitochondria play multiple roles in the cell, including maintaining energy homeostasis in response to environmental and cellular stress. Therefore, a key aspect of normal cellular function is the ability of the cell to respond to stress by stimulating mitochondrial biogenesis and turnover. Impairment of this ability can lead to mitochondrial dysfunction and disruption of cell energetics. Our research focuses on understanding the regulation of mitochondrial metabolism and how this changes with age. We have three related and overlapping research interests. They are to understand
1) the coupling of oxidative phosphorylation
2) how oxidative stress affects in vivo mitochondrial function
3) how the ability of the cell to respond to metabolic stress changes with age.
APPROACH
We believe that in order to understand how the body regulates mitochondrial function in response to stress it is necessary to integrate research on molecular, biochemical, and organismal levels. However, due to the lack of necessary tools to measure mitochondrial function in vivo, approaches have typically focused on in vitro measurements, particularly of the electron transport chain (ETC). However, mitochondria are sensitive to many systemic and cellular factors making it difficult to extrapolate results from isolated mitochondria to function in the intact organism. To bridge this gap we have developed state of the art molecular imaging/spectroscopy tools to study oxidative phosphorylation in skeletal muscle of intact organisms. We use optical and magnetic resonance spectroscopies to provide independent measures of O2 and ATP fluxes in vivo. By independently measuring these fluxes we determine several parameters of mitochondrial energetics in intact skeletal muscle, including the coupling of oxidative phosphorylation (P/O), phosphorylation capacity, and the sensitivity of respiration to oxygen content. Our research program combines these in vivo tools with transgenic models and molecular analysis to investigate the cellular and molecular mechanisms responsible for changes in the regulation of energy metabolism with age.
PREVIOUS RESULTS
Our work has led to significant insights into mitochondrial function in both normal and aged skeletal muscle. First, we determined that oxygen does not significantly regulate mitochondrial function in mouse skeletal muscle under normal physiological conditions (1). This finding is significant because it allowed us to separate the effects of mitochondrial dysfunction and oxygen limitation in aging and disease. A second paper demonstrated that our measurements of P/O in intact mouse hindlimb agree well with the theoretical values (2). This finding demonstrated the validity of the method, but more importantly established the baseline from which to analyze how mitochondrial function changes in response to environmental and disease stressors. Most recently we found significant mitochondrial uncoupling in resting skeletal muscle of aged mice (3). Our findings indicating significant mitochondrial uncoupling in aged skeletal muscle were recently extended to human muscle in collaboration with colleagues in the Metabolic Spectroscopy Laboratory (4). This uncoupling was associated with greater physiological stress (reduced ATP concentration) at rest and may be a key factor sensitizing the cell to apoptosis.
RESEARCH PLAN
The focus of our current research is to understand how changes in the ability of the cell to stimulate mitochondrial biogenesis in response to cellular stress affect in vivo mitochondrial function. My research addresses these issues at multiple levels of biological organization. Our strategy pairs in vivo experiments with studies of isolated muscles using pharmacological and transgenic manipulations that affect oxidative metabolism. We can then assess the change in mitochondrial function both in vivo and in vitro. This functional analysis is paired with a molecular analysis of the signaling pathways regulating the response. The long-term goal is to apply a gene expression analysis to identify and gene therapy approach to manipulate key steps signaling nodes responsible for the activation and inhibition of mitochondrial biogenesis. In this way we address how the regulation of mitochondrial function and biogenesis at the tissue, cellular, and molecular levels varies with environment and age.
1. Marcinek DJ, Ciesielski WA, Conley KE, and Schenkman KA. Oxygen regulation and limitation to cellular respiration in mouse skeletal muscle in vivo.Am J Physiol Heart Circ Physiol 285: H1900-1908, 2003.
2. Marcinek DJ, Schenkman KA, Ciesielski WA, and Conley KE. Mitochondrial coupling in vivo in mouse skeletal muscle. Am J Physiol Cell Physiol 286: 457-463, 2004.
3. Marcinek DJ, Schenkman KA, Ciesielski WA, Lee D, and Conley KE. Reduced mitochondrial coupling in vivo alters cellular energetics in aged mouse skeletal muscle. J Physiol 569: 467-473, 2005.
4. Amara CE, Shankland EG, Jubrias SA, Marcinek DJ, Kushmerick MJ, and Conley KE. Mild mitochondrial uncoupling impacts cellular aging in human muscles in vivo. Proceedings of the National Academy of Sciences of the United States of America 104: 1057-1062, 2007.