Amplification and propagation of endothelial signaling is likely to be dependent upon the release of local paracrine factors. Reactive oxygen species, including superoxide derived from activated inflammatory cells, activate endothelial cells under inflammatory and ischemia/reperfusion conditions. Subsequently, endothelial damage may potentiate inflammation and ischemic organ injury. Determining a means to reduce endothelial damage is therefore warranted, and may lead to the development of novel therapeutic approaches to treat ischemic and inflammatory diseases. The long-term goal of the proposed research project is to understand the molecular mechanisms of superoxide- mediated endothelial injury. In this proposal, we postulate that superoxide is a signaling molecule that facilitates endoplasmic reticulum Ca2+ signaling and triggers mitochondrial and nuclear stress. The guiding hypothesis is that superoxide raises intracellular Ca2+ and leads to mitochondrial Ca2+ overload, resulting in mitochondrial signaling. Extracellular superoxide selectively induces Ca2+-dependent depolarization independent of other oxidant species. Further, superoxide-evoked signals activate the redox sensitive transcription factor NF-:B. The uncoupling of extracellular superoxide-evoked signal from the mitochondria results in mitochondrial membrane potential preservation and endothelial cell survival.
The specific aims of this project are to examine (Specific Aim 1) how superoxide triggers upstream Ca2+ signaling and selectively activates inositol 1,4,5-trisphosphate receptors (InsP3R) both in vitro and in vivo;(Specific Aim 2) the role of extracellular superoxide-mediated Ca2+ signaling in mitochondrial pathophysiology (mitochondrial redox status-NADH and glutathione levels, mitochondrial ROS generation and mitochondrial membrane potential);(Specific Aim 3) the nuclear events that activate endothelial inflammatory mechanisms by InsP3R-linked ROS signaling. The proposed experiments should provide information on: (A) the involvement of InsP3Rs and their selective role in endothelial Ca2+ signaling during oxidative stress. (B) The relationship between mitochondrial dysfunction and inflammatory signaling in endothelial dysfunction. (C) This result will provide a novel signaling link between inflammatory and endothelial cells under pathophysiological conditions and lead to the development of novel therapeutic targets.

Public Health Relevance

The pulmonary vascular endothelium plays a key role in lung heath by translating blood signals into vascular function. During septic shock, an important and under-recognized signal is leukocyte-derived superoxide, which stimulates pulmonary endothelial cells by mobilizing calcium. Understanding the mechanisms of superoxide-mediated calcium signaling and mitochondria-nuclear stress in endothelial cells are crucial to elucidating the fundamental role of superoxide in both cell survival and death during physiological and pathological processes.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL086699-05
Application #
8274848
Study Section
Vascular Cell and Molecular Biology Study Section (VCMB)
Program Officer
Charette, Marc F
Project Start
2008-09-01
Project End
2014-05-31
Budget Start
2012-06-01
Budget End
2014-05-31
Support Year
5
Fiscal Year
2012
Total Cost
$334,125
Indirect Cost
$111,375
Name
Temple University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
057123192
City
Philadelphia
State
PA
Country
United States
Zip Code
19122
Lee, Samuel K; Shanmughapriya, Santhanam; Mok, Mac C Y et al. (2016) Structural Insights into Mitochondrial Calcium Uniporter Regulation by Divalent Cations. Cell Chem Biol 23:1157-69
Bao, Lei; Chen, Shu-Jen; Conrad, Kathleen et al. (2016) Depletion of the Human Ion Channel TRPM2 in Neuroblastoma Demonstrates Its Key Role in Cell Survival through Modulation of Mitochondrial Reactive Oxygen Species and Bioenergetics. J Biol Chem 291:24449-24464
Tomar, Dhanendra; Dong, Zhiwei; Shanmughapriya, Santhanam et al. (2016) MCUR1 Is a Scaffold Factor for the MCU Complex Function and Promotes Mitochondrial Bioenergetics. Cell Rep 15:1673-85
Scheitlin, Christopher G; Julian, Justin A; Shanmughapriya, Santhanam et al. (2016) Endothelial mitochondria regulate the intracellular Ca2+ response to fluid shear stress. Am J Physiol Cell Physiol 310:C479-90
Chu, Jin; Li, Jian-Guo; Joshi, Yash B et al. (2015) Gamma secretase-activating protein is a substrate for caspase-3: implications for Alzheimer's disease. Biol Psychiatry 77:720-8
Hoffman, Nicholas E; Miller, Barbara A; Wang, JuFang et al. (2015) Ca²⁺ entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance. Am J Physiol Heart Circ Physiol 308:H637-50
Shanmughapriya, Santhanam; Rajan, Sudarsan; Hoffman, Nicholas E et al. (2015) Ca2+ signals regulate mitochondrial metabolism by stimulating CREB-mediated expression of the mitochondrial Ca2+ uniporter gene MCU. Sci Signal 8:ra23
Chu, Jin; Li, Jian-Guo; Hoffman, Nicholas E et al. (2015) Degradation of gamma secretase activating protein by the ubiquitin-proteasome pathway. J Neurochem 133:432-9
Woitek, Felix; Zentilin, Lorena; Hoffman, Nicholas E et al. (2015) Intracoronary Cytoprotective Gene Therapy: A Study of VEGF-B167 in a Pre-Clinical Animal Model of Dilated Cardiomyopathy. J Am Coll Cardiol 66:139-53
Chu, Jin; Li, Jian-Guo; Hoffman, Nicholas E et al. (2015) Regulation of gamma-secretase activating protein by the 5Lipoxygenase: in vitro and in vivo evidence. Sci Rep 5:11086

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