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Permanent link (DOI): https://doi.org/10.7939/R3NK8N

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Novel intracellular role of matrix metalloproteinase-2 in cardiac cell injury Open Access

Descriptions

Other title
Subject/Keyword
cytosolic
matrix metalloproteinase-2
cell death
ischemia/reperfusion
titin
calpain inhibitors
Type of item
Thesis
Degree grantor
University of Alberta
Author or creator
Ali, Mohammad M. A.
Supervisor and department
Schulz, Richard (Pediatrics & Pharmacology)
Examining committee member and department
Hollenberg, Morley (Pharmacology & Physiology, University of Calgary)
Sipione, Simonetta (Pharmacology)
Clanachan, Alexander (Pharmacology)
Plane, Frances (Pharmacology)
Schulz, Richard (Pharmacology)
Simmen, Thomas (Cell Biology)
Department
Department of Pharmacology
Specialization

Date accepted
2012-06-20T11:11:25Z
Graduation date
2012-11
Degree
Doctor of Philosophy
Degree level
Doctoral
Abstract
Despite originally being described as a secreted protease, matrix metalloproteinase‐2 (MMP‐2) was recently revealed to have targets within cardiomyocytes. The biological mechanism(s) that describes the intracellular localization of MMP‐2 is unknown and is thus the subject of this thesis. Additionally, activation and inhibition of MMP-2 as well as novel intracellular target(s) of it that are involved in cardiomyocyte injury were investigated. The cytosolic targeting of MMP-2 was examined and it was found that the signal sequence of MMP-2 led to its incomplete targeting to the endoplasmic reticulum for secretion. Moreover, an MMP-2 splice variant which lacks the signal sequence and is enriched in the cytosol was discovered. Thus, intracellular MMP-2 is explained by the expression of a splice variant and by the inefficient targeting of canonical MMP-2 to the secretory pathway. Intracellular MMP-2 was found to play a pathological role in myocardial ischemia/reperfusion injury by proteolyzing the giant sarcomeric protein, titin. Discrete co-localization between MMP-2 and titin was found at the sarcomeric Z-disc region. Isolated rat hearts subjected to ischemia/reperfusion injury showed cleavage of titin, whereas inhibition of MMP-2 prevented titin proteolysis and improved the contractile function. To explore whether oxidative stress affects intracellular MMP-2 activity, isolated cardiomyocytes were treated with various concentrations of hydrogen peroxide. Treatment with 200 μM hydrogen peroxide led to elevated MMP-2 level/activity in cardiomyocyte lysates. Hydrogen peroxide primarily caused necrotic and not apoptotic cell death, however, pretreatment with selective MMP inhibitors did not protect against necrosis. In myocardial ischemia/reperfusion injury, MMP-2 or calpain inhibitors were shown to improve the myocardial contractile function. In order to investigate whether calpain inhibitors target intracellular MMP-2, the inhibitory effect of some calpain inhibitors on MMP-2 activity was tested. The IC50 values of calpain inhibitors, PD- 150606 and ALLN, against MMP-2 were determined to be 9.3 and 21.9 μM, respectively, revealing that some calpain inhibitors have significant pharmacological activity as inhibitors of MMP-2. In summary, these studies describe a set of mechanisms that cells utilize to equilibrate MMP-2 in both extracellular and intracellular locations. These results suggest that MMP-2 inhibitors should be rigorously tested as a therapeutic strategy to alleviate myocardial ischemia/reperfusion injury.
Language
English
DOI
doi:10.7939/R3NK8N
Rights
Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of these terms. The author reserves all other publication and other rights in association with the copyright in the thesis and, except as herein before provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.
Citation for previous publication
Titin is a target of matrix metalloproteinase-2: implications in myocardial ischemia/reperfusion injury. Ali MA, Cho WJ, Hudson B, Kassiri Z, Granzier H, Schulz R. Circulation. 2010 Nov 16;122(20):2039-47. Epub 2010 Nov 1.Mechanisms of cytosolic targeting of matrix metalloproteinase-2. Ali MA, Chow AK, Kandasamy AD, Fan X, West LJ, Crawford BD, Simmen T, Schulz R. J Cell Physiol. 2011 Dec 29. doi: 10.1002/jcp.24040. [Epub ahead of print]Calpain inhibitors exhibit matrix metalloproteinase-2 inhibitory activity. Ali MA, Stepanko A, Fan X, Holt A, Schulz R. Biochem Biophys Res Commun. 2012 May 7. [Epub ahead of print]

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