Publications by authors named "Shabana Aafaqi"

6 Publications

  • Page 1 of 1

Ankyrin-B dysfunction predisposes to arrhythmogenic cardiomyopathy and is amenable to therapy.

J Clin Invest 2019 07 2;129(8):3171-3184. Epub 2019 Jul 2.

Division of Cardiology, Department of Medicine, Johns Hopkins University, Baltimore, Maryland, USA.

Arrhythmogenic cardiomyopathy (ACM) is an inherited arrhythmia syndrome characterized by severe structural and electrical cardiac phenotypes, including myocardial fibrofatty replacement and sudden cardiac death. Clinical management of ACM is largely palliative, owing to an absence of therapies that target its underlying pathophysiology, which stems partially from our limited insight into the condition. Following identification of deceased ACM probands possessing ANK2 rare variants and evidence of ankyrin-B loss of function on cardiac tissue analysis, an ANK2 mouse model was found to develop dramatic structural abnormalities reflective of human ACM, including biventricular dilation, reduced ejection fraction, cardiac fibrosis, and premature death. Desmosomal structure and function appeared preserved in diseased human and murine specimens in the presence of markedly abnormal β-catenin expression and patterning, leading to identification of a previously unknown interaction between ankyrin-B and β-catenin. A pharmacological activator of the WNT/β-catenin pathway, SB-216763, successfully prevented and partially reversed the murine ACM phenotypes. Our findings introduce what we believe to be a new pathway for ACM, a role of ankyrin-B in cardiac structure and signaling, a molecular link between ankyrin-B and β-catenin, and evidence for targeted activation of the WNT/β-catenin pathway as a potential treatment for this disease.
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http://dx.doi.org/10.1172/JCI125538DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6668697PMC
July 2019

TMEM43 mutation p.S358L alters intercalated disc protein expression and reduces conduction velocity in arrhythmogenic right ventricular cardiomyopathy.

PLoS One 2014 24;9(10):e109128. Epub 2014 Oct 24.

Physiology and Experimental Medicine, The Hospital for Sick Children and Research Institute, Toronto, Ontario, Canada.

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a myocardial disease characterized by fibro-fatty replacement of myocardium in the right ventricular free wall and frequently results in life-threatening ventricular arrhythmias and sudden cardiac death. A heterozygous missense mutation in the transmembrane protein 43 (TMEM43) gene, p.S358L, has been genetically identified to cause autosomal dominant ARVC type 5 in a founder population from the island of Newfoundland, Canada. Little is known about the function of the TMEM43 protein or how it leads to the pathogenesis of ARVC. We sought to determine the distribution of TMEM43 and the effect of the p.S358L mutation on the expression and distribution of various intercalated (IC) disc proteins as well as functional effects on IC disc gap junction dye transfer and conduction velocity in cell culture. Through Western blot analysis, transmission electron microscopy (TEM), immunofluorescence (IF), and electrophysiological analysis, our results showed that the stable expression of p.S358L mutation in the HL-1 cardiac cell line resulted in decreased Zonula Occludens (ZO-1) expression and the loss of ZO-1 localization to cell-cell junctions. Junctional Plakoglobin (JUP) and α-catenin proteins were redistributed to the cytoplasm with decreased localization to cell-cell junctions. Connexin-43 (Cx43) phosphorylation was altered, and there was reduced gap junction dye transfer and conduction velocity in mutant TMEM43-transfected cells. These observations suggest that expression of the p.S358L mutant of TMEM43 found in ARVC type 5 may affect localization of proteins involved in conduction, alter gap junction function and reduce conduction velocity in cardiac tissue.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0109128PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4208740PMC
June 2015

Integrin-linked kinase mediates force transduction in cardiomyocytes by modulating SERCA2a/PLN function.

Nat Commun 2014 Sep 11;5:4533. Epub 2014 Sep 11.

Cardiology Division, Department of Paediatrics, Hospital for Sick Children, Toronto, Ontario, Canada M5G 1X8.

Human dilated cardiomyopathy (DCM) manifests as a profound reduction in biventricular cardiac function that typically progresses to death or cardiac transplantation. There is no effective mechanism-based therapy currently available for DCM, in part because the transduction of mechanical load into dynamic changes in cardiac contractility (termed mechanotransduction) remains an incompletely understood process during both normal cardiac function and in disease states. Here we show that the mechanoreceptor protein integrin-linked kinase (ILK) mediates cardiomyocyte force transduction through regulation of the key calcium regulatory protein sarcoplasmic/endoplasmic reticulum Ca(2+)ATPase isoform 2a (SERCA-2a) and phosphorylation of phospholamban (PLN) in the human heart. A non-oncogenic ILK mutation with a synthetic point mutation in the pleckstrin homology-like domain (ILK(R211A)) is shown to enhance global cardiac function through SERCA-2a/PLN. Thus, ILK serves to link mechanoreception to the dynamic modulation of cardiac contractility through a previously undiscovered interaction with the functional SERCA-2a/PLN module that can be exploited to rescue impaired mechanotransduction in DCM.
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http://dx.doi.org/10.1038/ncomms5533DOI Listing
September 2014

Mutation in integrin-linked kinase (ILK(R211A)) and heat-shock protein 70 comprise a broadly cardioprotective complex.

PLoS One 2013 18;8(11):e77331. Epub 2013 Nov 18.

Division of Cardiovascular Research, Hospital for Sick Children, Toronto, Canada.

Rationale: Integrin-linked kinase (ILK) has been proposed as a novel molecular target that has translational potential in diverse cardiac diseases, since its upregulation promotes a broadly cardioprotective phenotype. However, ILK has been implicated as both a cardioprotective and oncogenic target, which imposes therapeutic constraints that are generally relevant to the translational potential of many kinases.

Objective: To study the cardioprotective properties of the activation-resistant, non-oncogenic, mutation of ILK (ILK(R211A)) against experimental MI in vivo and Doxorubicin induced apoptosis in vitro and it's relationships to stress induced heat shock proteins.

Methods/results: The transgenic mouse heart over-expressing a point mutation in the ILK pleckstrin homology (PH) domain (Tg(R211A)) exhibits a highly cardioprotective phenotype based on LAD-ligation-induced MI reduction in vivo, and on protection against doxorubicin (DOX)-induced cardiomyocyte apoptosis when overexpressed in human induced pluripotent stem cell (iPS)-derived cardiomyocytes in vitro. Intriguingly, the degree of cardioprotection seen with the ILK(R211A) mutation exceeded that with the ILK(S343D) mutation. Microarray and immunoprecipitation analyses revealed upregulation of expression levels and specific binding of ILK(WT), ILK(S343D) and ILK(R211A) to both constitutively active heat-shock protein 70 (Hsc70) and inducible Hsp70 in response to MI, and to acute ILK overexpression in iPSC-cardiomyocytes. ILK-mediated cardioprotection was shown to depend upon Hsp70 ATPase activity.

Conclusions: These findings indicate that wild type ILK and the non-oncogenic ILK(R211A) mutation comprise a cardioprotective module with Hsp/c70. These results advance a novel target discovery theme in which kinase mutations can be safely engineered to enhance cardioprotective effects.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0077331PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3832499PMC
July 2014

Pulmonary vein stenosis and the pathophysiology of "upstream" pulmonary veins.

J Thorac Cardiovasc Surg 2014 Jul 29;148(1):245-53. Epub 2013 Sep 29.

Division of Cardiovascular Surgery, Hospital for Sick Children, Labatt Family Heart Center and University of Toronto, Toronto, Ontario, Canada. Electronic address:

Background: Surgical and catheter-based interventions on pulmonary veins are associated with pulmonary vein stenosis (PVS), which can progress diffusely through the "upstream" pulmonary veins. The mechanism has been rarely studied. We used a porcine model of PVS to assess disease progression with emphasis on the potential role of endothelial-mesenchymal transition (EndMT).

Methods: Neonatal piglets underwent bilateral pulmonary vein banding (banded, n = 6) or sham operations (sham, n = 6). Additional piglets underwent identical banding and stent implantation in a single-banded pulmonary vein 3 weeks postbanding (stented, n = 6). At 7 weeks postbanding, hemodynamics and upstream PV pathology were assessed.

Results: Banded piglets developed pulmonary hypertension. The upstream pulmonary veins exhibited intimal thickening associated with features of EndMT, including increased transforming growth factor (TGF)-β1 and Smad expression, loss of endothelial and gain of mesenchymal marker expression, and coexpression of endothelial and mesenchymal markers in banded pulmonary vein intimal cells. These immunopathologic changes and a prominent myofibroblast phenotype in the remodeled pulmonary veins were consistently identified in specimens from patients with PVS, in vitro TGF-β1-stimulated cells isolated from piglet and human pulmonary veins, and human umbilical vein endothelial cells. After stent implantation, decompression of a pulmonary vein was associated with reappearance of endothelial marker expression, suggesting the potential for plasticity in the observed pathologic changes, followed by rapid in-stent restenosis.

Conclusions: Neonatal pulmonary vein banding in piglets recapitulates critical aspects of clinical PVS and highlights a pathologic profile consistent with EndMT, supporting the rationale for evaluating therapeutic strategies designed to exploit reversibility of upstream pulmonary vein pathology.
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http://dx.doi.org/10.1016/j.jtcvs.2013.08.046DOI Listing
July 2014

ILK induces cardiomyogenesis in the human heart.

PLoS One 2012 29;7(5):e37802. Epub 2012 May 29.

Division of Cardiovascular Research, Hospital for Sick Children, Toronto, Canada.

Background: Integrin-linked kinase (ILK) is a widely conserved serine/threonine kinase that regulates diverse signal transduction pathways implicated in cardiac hypertrophy and contractility. In this study we explored whether experimental overexpression of ILK would up-regulate morphogenesis in the human fetal heart.

Methodology/principal Findings: Primary cultures of human fetal myocardial cells (19-22 weeks gestation) yielded scattered aggregates of cardioblasts positive for the early cardiac lineage marker nk × 2.5 and containing nascent sarcomeres. Cardiac cells in colonies uniformly expressed the gap junction protein connexin 43 (C × 43) and displayed a spectrum of differentiation with only a subset of cells exhibiting the late cardiomyogenic marker troponin T (cTnT) and evidence of electrical excitability. Adenovirus-mediated overexpression of ILK potently increased the number of new aggregates of primitive cardioblasts (p<0.001). The number of cardioblast colonies was significantly decreased (p<0.05) when ILK expression was knocked down with ILK targeted siRNA. Interestingly, overexpression of the activation resistant ILK mutant (ILK(R211A)) resulted in much greater increase in the number of new cell aggregates as compared to overexpression of wild-type ILK (ILK(WT)). The cardiomyogenic effects of ILK(R211A) and ILK(WT) were accompanied by concurrent activation of β-catenin (p<0.001) and increase expression of progenitor cell marker islet-1, which was also observed in lysates of transgenic mice with cardiac-specific over-expression of ILK(R211A) and ILK(WT). Finally, endogenous ILK expression was shown to increase in concert with those of cardiomyogenic markers during directed cardiomyogenic differentiation in human embryonic stem cells (hESCs).

Conclusions/significance: In the human fetal heart ILK activation is instructive to the specification of mesodermal precursor cells towards a cardiomyogenic lineage. Induction of cardiomyogenesis by ILK overexpression bypasses the requirement of proximal PI3K activation for transduction of growth factor- and β1-integrin-mediated differentiation signals. Altogether, our data indicate that ILK represents a novel regulatory checkpoint during human cardiomyogenesis.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0037802PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3362604PMC
October 2012
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