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Nuclear matrix metalloproteinase-2 and investigation of its potential targets in myocardial ischemia-reperfusion injury Open Access


Other title
Nuclear MMP-2
Nuclear matrix metalloproteinase-2
myocardial ischemia-reperfusion injury
Type of item
Degree grantor
University of Alberta
Author or creator
Baghirova, Sabina
Supervisor and department
Schulz, Richard (Pharmacology & Paediatrics)
Examining committee member and department
Schulz, Richard (Pharmacology & Paediatrics)
Bourque, Stephane (Pharmacology)
Plane, Frances (Pharmacology)
Hendzel, Michael (Oncology)
Department of Pharmacology

Date accepted
Graduation date
Master of Science
Degree level
Matrix metalloproteinases (MMPs) are zinc-dependent proteases involved in intra- and extra-cellular matrix remodeling. MMP-2 was the first to be localized to the nucleus; however the biological functions and substrates of nuclear MMP-2 are mostly unknown. We hypothesized that MMP‐2 is present in the nucleus under normal physiological conditions but increases during myocardial ischemia‐reperfusion (I/R) injury induced oxidative stress, proteolyzing nuclear structural proteins (lamins). Lamin A/C, a putative nuclear MMP‐2 target, is an intermediate filament protein that provides structural support to the nucleus. Immunofluorescent confocal microscopy and subcellular fractionation showed the presence of MMP-2 in cytoplasm and nuclei of neonatal rat ventricular myocytes. The distribution of MMP-2 in cytoplasm and nuclei was verified by immunofluorescent confocal microscopy in the human fibrosarcoma HT 1080 cells. Further analysis by flow cytometry determined that 88.6% of a sample of ~10,000 HT 1080 cells had MMP-2 concentrated to the nucleus. Rat hearts were isolated and perfused by the Langendorff method aerobically, or subjected to global, no-flow ischemia followed by aerobic reperfusion in the presence or absence of an MMP inhibitor (100 µM o-phenanthroline). Nuclear fractions extracted from the rat hearts showed increased MMP-2 activity, but not protein level in hearts subjected to I/R injury. To identify possible targets, an in vitro proteolysis assay was performed with lamin A or B incubated with MMP-2. Lamin A, but not lamin B, was proteolysed by MMP-2 in to a putative 50 kDa fragment, which was also predicted by in silico cleavage site analysis. Protein levels of troponin I, a known sarcomeric target of MMP-2 showed a trend for decrease in I/R hearts and was normalized by o-phenanthroline, demonstrating efficacy of the MMP inhibitor. However, lamin A and lamin C protein levels remained unchanged in I/R hearts. PARP-1, a nuclear DNA repair protein, previously shown to be proteolysed by MMP-2 in vitro was measured in I/R hearts. It was observed that PARP-1 protein levels were decreased in I/R hearts treated with o-phenanthroline compared to I/R hearts without o-phenanthroline. Nuclear MMP-2 is present in cardiomyocytes under normal physiological conditions, and is increased as a result of I/R injury. This increase of MMP-2 activity in extracts from I/R hearts leads to proteolysis of troponin I, but not, PARP-1, lamin A or C. The activation of genes in myocardial I/R injury suggests that other nuclear functions of MMP-2 are likely.
This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for the purpose of private, scholarly or scientific research. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.
Citation for previous publication
Baghirova, S., B.G. Hughes, M.J. Hendzel and R. Schulz. 2015. Sequential fractionation and isolation of subcellular proteins from tissue or cultured cells. MethodsX 2: 440-445. doi:10.1016/j.mex.2015.11.001.

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