Duclienne muscular dystrophy is a common genetic disorder resulting from mutations within the dystrophin gene. It is likely that the dystrophic phenotype does not result directly from alteration to the myofibrillar Structures, rather it is a disruption of sarcolemmal membrane integrity that nomnally confers tight control of intracellular Ca homeostasis, which leads to elevated intracellular Ca and eventual muscle degeneration. The pathophysiological mechanism responsible for elevation of intracellular Ca levels is not dear, however our recent findings suggest that store-operated Ca entry (SOCE) linked to uncontrolled Ca spark activity may contribute to the aberrant Ca influx observed in dystrophic muscle. While the mechanism of SOCE activation is still a matter of intensive study, it has been recently detemnined that the Ca insensitive phospholipase A2 (iPLA2) is an important mediator of SOCE. The focus of this project is to test the hypothesis that aben-ant Ca spark activity acts as a trigger for SOCE and thus induces a dystrophic cascade in mammalian skeletal muscle through a pathway that involves iPLA2 mediated signaling. We will test this with three specific aims:
Aim 1; To establish induced Ca sparks as a trigger for SOCE in healthy and dystrophic muscle. We will utilize multiple methods for measurement of SOCE to examine alteration to Ca entry in dystrophic fiber. Patch-clamp measurement and modulation of Oa influx will provide insight into SR Ca release and the activation of SOCE in dystrophic fibers.
Aim 2 : Detemiine the contribution of iPLA2 activity to Ca influx in muscular dystrophy. We will examine the altered characteristics of iPLA2 function in dystrophic muscle using various in vitro molecular and pharmacological methods.
Aim 3 : To elucidate if SOCE facilitates the dystrophic cascade in skeletal muscle. Due to the inability to assess changes in the dystrophic phenotype in skeletal muscle we wili modulate SOCE in dystrophic animals and assay changes in dystrophic phenotypes.

Public Health Relevance

This project will contribute to our understanding of how muscle damage occurs in cases of Duchenne muscular dystrophy (DMD), the most common genetic disease in the United States. Our experiments will establish that defects in how calcium enters muscle cells result in the development of DMD. Such knowledge will suggest new methods to treat DMD, some of which will be tested on experimental animals in this study.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Transition Award (R00)
Project #
4R00AR054793-03
Application #
8073247
Study Section
Special Emphasis Panel (NSS)
Program Officer
Boyce, Amanda T
Project Start
2011-02-18
Project End
2014-01-31
Budget Start
2011-02-18
Budget End
2012-01-31
Support Year
3
Fiscal Year
2011
Total Cost
$248,999
Indirect Cost
Name
University of Medicine & Dentistry of NJ
Department
Physiology
Type
Schools of Medicine
DUNS #
617022384
City
Piscataway
State
NJ
Country
United States
Zip Code
08854
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Ferrante, Christopher; Szappanos, Henrietta; Csernoch, László et al. (2013) Analysis of osmotic stress induced Ca2+ spark termination in mammalian skeletal muscle. Indian J Biochem Biophys 50:411-8
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Weisleder, Noah; Zhou, Jingsong; Ma, Jianjie (2012) Detection of calcium sparks in intact and permeabilized skeletal muscle fibers. Methods Mol Biol 798:395-410
Zhao, Xiaoli; Moloughney, Joseph G; Zhang, Sai et al. (2012) Orai1 mediates exacerbated Ca(2+) entry in dystrophic skeletal muscle. PLoS One 7:e49862
Thornton, Angela M; Zhao, Xiaoli; Weisleder, Noah et al. (2011) Store-operated Ca(2+) entry (SOCE) contributes to normal skeletal muscle contractility in young but not in aged skeletal muscle. Aging (Albany NY) 3:621-34
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Weisleder, Noah; Lin, Peihui; Zhao, Xiaoli et al. (2011) Visualization of MG53-mediated cell membrane repair using in vivo and in vitro systems. J Vis Exp :
Zhu, Hua; Lin, Peihui; De, Gejing et al. (2011) Polymerase transcriptase release factor (PTRF) anchors MG53 protein to cell injury site for initiation of membrane repair. J Biol Chem 286:12820-4

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