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Analysis of Osteopontin Levels for the Identification of Asymptomatic Patients With Calcific Aortic Valve Disease  Juan B. Grau, MD, Paolo Poggio, MS,

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Presentation on theme: "Analysis of Osteopontin Levels for the Identification of Asymptomatic Patients With Calcific Aortic Valve Disease  Juan B. Grau, MD, Paolo Poggio, MS,"— Presentation transcript:

1 Analysis of Osteopontin Levels for the Identification of Asymptomatic Patients With Calcific Aortic Valve Disease  Juan B. Grau, MD, Paolo Poggio, MS, Rachana Sainger, PhD, William J. Vernick, MD, William F. Seefried, BS, Emanuela Branchetti, PhD, Benjamin C. Field, BS, Joseph E. Bavaria, MD, Michael A. Acker, MS, Giovanni Ferrari, PhD  The Annals of Thoracic Surgery  Volume 93, Issue 1, Pages (January 2012) DOI: /j.athoracsur Copyright © 2012 The Society of Thoracic Surgeons Terms and Conditions

2 Fig 1 Calcific aortic valve degeneration.(A) Videoscopic-assisted intraoperative pictures of healthy aortic valve (Control), sclerotic aortic valve displaying thickened leaflets and fibrotic plaques (Aortic Sclerosis), and severe aortic calcification (Aortic Stenosis). (B) Histologic analysis of aortic valve leaflets. Composite photomicrographs of longitudinal section of human aortic valve cusps. Controls and diseased valves are indicated. High-definition images (50+) per slide have been assembled to rebuild the leaflet structures (×10 magnification). Hematoxylin and eosin (H&E) and Movat (MOVAT) pentachrome staining are indicated. (C) Examples of transesophageal echocardiographic evaluation (long axis view) for control, AVSc, and AVS. The Annals of Thoracic Surgery  , 79-86DOI: ( /j.athoracsur ) Copyright © 2012 The Society of Thoracic Surgeons Terms and Conditions

3 Fig 2 Osteopontin (OPN) level correlates with the progression of calcific aortic valve disease. (A) Composite photomicrographs of section of human aortic valve cusps (longitudinal sections) stained with alizarin red (Alz. Red) to visualize calcium content and with OPN antibody (×10 magnification). (B) Real-time quantitative polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay (ELISA) for the evaluation of OPN levels in patients with aortic sclerosis, stenosis, and in controls in tissue and plasma, respectively. For qPCR analysis, OPN levels were normalized against the control level. Absolute OPN levels (in nanograms per milliliter of plasma) are indicated in the ELISA. The Annals of Thoracic Surgery  , 79-86DOI: ( /j.athoracsur ) Copyright © 2012 The Society of Thoracic Surgeons Terms and Conditions

4 Fig 3 Comparison between echocardiographic evaluation and osteopontin (OPN) level in patients with calcific aortic valve disease (CAVD). (A) Absolute OPN levels (in nanograms per milliliter of plasma), measured by ELISA, in the plasma of patients with different calcium scores (x axis). (B) Comparison between OPN concentration in the plasma of control group (calcium score 1) and patients (calcium score 1 to 4). (C = control; ASc = aortic sclerosis; AS = aortic stenosis.) The Annals of Thoracic Surgery  , 79-86DOI: ( /j.athoracsur ) Copyright © 2012 The Society of Thoracic Surgeons Terms and Conditions

5 Fig 4 Differential expression of osteopontin (OPN) splicing variant in calcific aortic valve disease. (A) Graphic representation of OPN splicing variants. (B) Relative quantification of OPN-a, OPN-b, OPN-c in patients with aortic sclerosis, aortic stenosis, and control tissues assessed by quantitative polymerase chain reaction. (C) Tissue distribution of OPN isoforms in control and diseased tissues detected by in situ hybridization (×100 magnification). The Annals of Thoracic Surgery  , 79-86DOI: ( /j.athoracsur ) Copyright © 2012 The Society of Thoracic Surgeons Terms and Conditions

6 Fig 5 Osteopontin (OPN) splicing variants and biomineralization. (A) OPN splicing variant production and detection with Commassie blue staining. (B) OPN-a, OPN-b, and OPN-c expression detected by Western blotting using anti-FLAG and anti-OPN antibodies. (C) Bar graph representative of calcium formed on vascular smooth muscle cells (VSMC). (D) Calcification assay performed using OPN isoforms: (a, b) calcium accumulation on VSMC cultured in normal media and in calcification media (negative and positive control respectively); (c, d, e) calcium deposition on valvular smooth muscle cells treated with calcification media in the presence of OPN isoforms nonphosphorylated; (f, g, h) calcium deposition on valvular smooth muscle cells treated with calcification media in the presence of phosphorylated OPN isoforms. Calcium accumulation was quantified and normalized against protein content. The Annals of Thoracic Surgery  , 79-86DOI: ( /j.athoracsur ) Copyright © 2012 The Society of Thoracic Surgeons Terms and Conditions

7 Fig 6 Characterization of osteopontin as an early marker for the identification of asymptomatic patients with calcific aortic valve disease. (TEE = transesophageal echocardiography.) The Annals of Thoracic Surgery  , 79-86DOI: ( /j.athoracsur ) Copyright © 2012 The Society of Thoracic Surgeons Terms and Conditions

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