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ABSTRACT: AIMSAortic valve sclerosis (AVSc) is a hallmark of several cardiovascular conditions ranging from chronic heart failure and myocardial infarction to calcific aortic valve stenosis (AVS). AVSc, present in 25%-30% of patients over 65 years of age, is characterized by thickening of the leaflets with marginal effects on the mechanical proprieties of the valve making its presentation asymptomatic. Despite its clinical prevalence, few studies have investigated the pathogenesis of this disease using human AVSc specimens. Here we investigate in vitro and ex vivo BMP4-mediated transdifferentiation of human valve interstitial cells (VIC) toward an osteogenic-like phenotype in AVSc.METHODS AND RESULTSHuman specimens from 60 patients were collected at the time of aortic valve replacement (AVS) or through the heart transplant program (Controls and AVSc). We show that non-calcified leaflets from AVSc patients can be induced to express markers of osteogenic transdifferentiation and biomineralization through the combinatory effect of BMP4 and mechanical stimulation. We show that BMP4 antagonist Noggin attenuates VIC activation and biomineralization. Additionally, patient-derived VICs were induced to transdifferentiate using either cell culture or a Tissue Engineering Aortic Valve model. We determine that while BMP4 alone is not sufficient to induce osteogenic transdifferentiation of AVSc-derived cells, the combinatory effect of BMP4 and mechanical stretch induces VIC activation towards a phenotype typical of late calcified stage of the disease.CONCLUSION
This work demonstrates, for the first time using AVSc specimens, that human sclerotic aortic valves can be induced to express marker of osteogenic-like phenotype typical of advanced severe aortic stenosis.
Cardiovascular research 03/2013; · 5.80 Impact Factor
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The Journal of heart valve disease 03/2013; 22(2):156-165. · 0.81 Impact Factor
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Emanuela Branchetti,
Rachana Sainger,
Paolo Poggio,
Juan B Grau,
Jeffrey Patterson-Fortin,
Joseph E Bavaria,
Michael Chorny,
Eric Lai,
Robert C Gorman,
Robert J Levy,
Giovanni Ferrari
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ABSTRACT: OBJECTIVE: Accumulation of reactive oxygen species (ROS) and remodeling of the microstructure of the cusp characterize aortic valve sclerosis, the early phase of calcific aortic valve disease. These events are associated with activation of valvular interstitial cells (VICs) toward an osteogenic-like phenotype. Because ROS cause DNA damage and transcriptional activation we investigated the relationship between ROS, DNA damage response, and transdifferentiation of VICs. METHODS AND RESULTS: Human aortic valve cusps and patient-matched VICs were collected from 39 patients both with and without calcific aortic valve disease. VICs were exposed to hydrogen peroxide (0.1-1 mmol/L) after cell transduction with extracellular superoxide dismutase/catalase adenoviruses and characterized for DNA-damage response, osteogenic transdifferentiation, and calcification. ROS induce relocalization of phosphorylated γH2AX, MRE11, and XRCC1 proteins with expression of osteogenic signaling molecule RUNX2 via AKT. We report a sustained activation of γH2AX in aortic valve sclerosis-derived VICs suggesting their impaired ability to repair DNA damage. Adenovirus superoxide dismutase/catalase transduction decreases ROS-induced DNA damage and VIC transdifferentiation in aortic valve sclerosis-derived cells. Finally, adenoviral transduction with catalase reverts ROS-mediated calcification and cellular transdifferentiation. CONCLUSIONS: We conclude that the ROS-induced DNA damage response is dysfunctional in early asymptomatic stages of calcific aortic valve disease. We unveiled an association among ROS, DNA-damage response, and cellular transdifferentiation, reversible by antioxidant enzymes delivery.
Arteriosclerosis Thrombosis and Vascular Biology 12/2012; · 6.37 Impact Factor
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ABSTRACT: Calcific Aortic Valve Disease (CAVD) is an active pathological process leading to biomineralization of the aortic cusps. We characterized circulating and tissue Osteopontin (OPN) as a biomarker for CAVD.
Here we investigate the post-translational modifications of circulating OPN and correlate the phosphorylation status with the ability to prevent calcification.
Circulating OPN levels were estimated in CAVD patients (n = 51) and controls (n = 56). In a subgroup of 27 subjects, OPN was purified and the phosphorylation status analyzed.
Plasma OPN levels were significantly elevated in CAVD patients as compared to the controls and correlates with the aortic valve calcium score. Our study demonstrates that phospho-threonine levels of OPN purified from controls were higher when compared to CAVD subjects, whereas phospho-serine and phospho-tyrosine levels were comparable between the two groups.
The dephosphorylation of circulating OPN correlates with severe valvular calcification in patients with CAVD.
Biomarkers 12/2011; 17(2):111-8. · 2.21 Impact Factor
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Rachana Sainger,
Juan B Grau, Emanuela Branchetti,
Paolo Poggio,
William F Seefried,
Benjamin C Field,
Michael A Acker,
Robert C Gorman,
Joseph H Gorman,
Clark W Hargrove,
Joseph E Bavaria,
Giovanni Ferrari
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ABSTRACT: Myxomatous mitral valve prolapse (MVP) is the most common cardiac valvular abnormality in industrialized countries and a leading cause of mitral valve surgery for isolated mitral regurgitation. The key role of valvular interstitial cells (VICs) during mitral valve development and homeostasis has been recently suggested, however little is known about the molecular pathways leading to MVP. We aim to characterize bone morphogenetic protein 4 (BMP4) as a cellular regulator of mitral VIC activation towards a pathologic synthetic phenotype and to analyze the cellular phenotypic changes and extracellular matrix (ECM) reorganization associated with the development of myxomatous MVP. Microarray analysis showed significant up regulation of BMP4-mediated signaling molecules in myxomatous MVP when compared to controls. Histological analysis and cellular characterization suggest that during myxomatous MVP development, healthy quiescent mitral VICs undergo a phenotypic activation via up regulation of BMP4-mediated pathway. In vitro hBMP4 treatment of isolated human mitral VICs mimics the cellular activation and ECM remodeling as seen in MVP tissues. The present study characterizes the cell biology of mitral VICs in physiological and pathological conditions and provides insights into the molecular and cellular mechanisms mediated by BMP4 during MVP. The ability to test and control the plasticity of VICs using different molecules may help in developing new diagnostic and therapeutic strategies for myxomatous MVP.
Journal of Cellular Physiology 11/2011; 227(6):2595-604. · 3.87 Impact Factor
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ABSTRACT: Calcific aortic valve disease (CAVD) is the most common cause of acquired valve disease. Initial phases of CAVD include thickening of the cusps, whereas advanced stages are associated with biomineralization and reduction of the aortic valve area. These conditions are known as aortic valve sclerosis (AVSc) and aortic valve stenosis (AVS), respectively. Because of its asymptomatic presentation, little is known about the molecular determinants of AVSc. The aim of this study was to correlate plasma and tissue osteopontin (OPN) levels with echocardiographic evaluation for the identification of asymptomatic patients at risk for CAVD. In addition, our aim was to analyze the differential expression and biological function of OPN splicing variants as biomarkers of early and late stages of CAVD.
From January 2010 to February 2011, 310 patients were enrolled in the study. Patients were divided into 3 groups based on transesophageal echocardiographic (TEE) evaluation: controls (56 patients), AVSc (90 patients), and AVS (164 patients). Plasma and tissue OPN levels were measured by immunohistochemical evaluation, enzyme-linked immunosorbent assay (ELISA), and real-time quantitative polymerase chain reaction (qPCR).
Patients with AVSc and AVS have higher OPN levels compared with controls. OPN levels are elevated in asymptomatic patients with AVSc with no appearance of calcification during TEE evaluation. OPN splicing variants OPN-a, OPN-b, and OPN-c are differentially expressed during CAVD progression and are able to inhibit biomineralization in a cell-based biomineralization assay.
The analysis of the differential expression of OPN splicing variants during CAVD may help in developing diagnostic and risk stratification tools to follow the progression of asymptomatic aortic valve degeneration.
The Annals of thoracic surgery 11/2011; 93(1):79-86. · 3.74 Impact Factor
