Abstract P332: Microfluidic Model Of Late-stage Calcific Aortic Valve Disease Develops Calcium Phosphate Mineralizations

2021 ◽  
Vol 129 (Suppl_1) ◽  
Author(s):  
Melissa Mendoza ◽  
Mei-Hsiu Chen ◽  
Bruce Murray ◽  
Peter Huang ◽  
Gretchen Mahler
Keyword(s):  
Shear Stress ◽  
Calcium Phosphate ◽  
Aortic Valve ◽  
Late Stage ◽  
Aortic Valve Disease ◽  
Collagen I ◽  
Valve Disease ◽  
Calcium Deposition ◽  
Nodule Formation ◽  
Calcific Aortic Valve Disease

Introduction: Calcific aortic valve disease (CAVD) is an active pathological process leading to severe valve calcification. Late-stage CAVD is characterized by increased leaflet stiffness, disorganized collagen bundles and the deposition of glycosaminoglycans, such as chondroitin sulfate (CS), in the fibrosa layer. However, many details of the cellular pathological cascade remain unknown. Animal models such as mice, rabbits, and pigs are used in understanding human CAVD, but mice do not have similar anatomy, rabbits cannot spontaneously develop atherosclerotic lesions, and pigs require long, expensive and complex studies. Here we utilize microfluidic devices of the aortic valve fibrosa to model late-stage CAVD. Hypothesis: We assessed the hypothesis that microfluidic calcification will increase with increased shear rates and CS content. Methods: Valve-on-a-chip devices contained a hydrogel of 1.5 mg/mL collagen I-only healthy controls or 1.5 mg/mL collagen I with 1 mg/mL or 20 mg/mL CS. Porcine aortic valve interstitial cells (PAVIC) were embedded within and endothelial cells (PAVEC) were seeded onto the matrix. Steady shear stress at 1 dyne/cm 2 and 20 dyne/cm 2 were applied using a peristaltic pump for 14 days. Alizarin Red S (ARS), an assay to assess calcium deposition, was used to quantify calcific nodule formation. Scanning electron microscopy with energy dispersive x-ray (SEM/EDX) was used to further analyze sample mineralization. Results: Co-cultures in the presence of increasing shear stress and CS exhibit increased calcific nodule formation compared to static controls, both qualitatively and quantitatively (n≥3). SEM revealed the microstructure of calcified nodules and EDX confirmed calcium phosphate mineralization with physiologically-relevant calcium to phosphorous ratios (Ca/P= 0.88 - 1.4). Conclusions: These results show that in vitro calcification is driven by shear stress in the presence of PAVEC and CS. As seen in ex vivo studies of human calcification, these microfluidic-derived nodules are similarly composed of a range of naturally-occurring calcium phosphates. Given that CAVD has no targeted therapy, the creation of a physiologically relevant model of the aortic valve can provide a test bed for novel therapeutic interventions.

Abstract P098: Inhibitory Role of Notch1 in Calcific Aortic Valve Disease Is Mediated by Sox9

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Chetan P Hans ◽  
Asha Acharya ◽  
Sara N Koenig ◽  
Haley A Nichols ◽  
Cristi L Galindo ◽  
...  
Keyword(s):  
Aortic Valve ◽  
Notch Signaling ◽  
Signaling Pathway ◽  
Aortic Valve Disease ◽  
Notch Signaling Pathway ◽  
Valve Disease ◽  
Calcium Deposition ◽  
Aortic Valve Calcification ◽  
Calcific Aortic Valve Disease ◽  
Valve Calcification

Introduction: Aortic valve calcification is the most common form of valvular heart disease; however the mechanism(s) underlying calcific aortic valve disease (CAVD) are unknown. NOTCH1 mutations are associated with aortic valve malformations and adult-onset calcification in families with inherited disease. The Notch signaling pathway is critical for multiple cell differentiation processes, but its role in the development of CAVD is not well understood. Objective: To investigate the molecular changes associated with the calcification of aortic valve that occurs with inhibition of Notch signaling. Methods and Results: The expression of Notch signaling pathway members was validated in the aortic valve cusps from adult mice, and examination of diseased human aortic valves revealed decreased expression of NOTCH1 in areas of calcium deposition. To identify downstream mediators of Notch1 signaling, we examined gene expression changes that occur with chemical inhibition of Notch signaling in rat aortic valve interstitial cells (AVICs). We found significant downregulation of many cartilage-specific genes that constitute the valve extracellular matrix (ECM). Analysis of these cartilage-specific genes demonstrated that several were transcriptional targets of Sox9, a master regulator of chondrogenesis, which has been previously shown to be essential for proper valve development and maintenance. Utilizing an in vitro porcine aortic valve calcification model system, inhibition of Notch activity resulted in accelerated calcification while stimulation of Notch signaling attenuated the calcific process. Finally, utilizing transfection studies, addition of Sox9 was able to prevent the calcification of porcine AVICs that occurs with Notch inhibition. Conclusions: Loss of Notch signaling contributes to aortic valve calcification by a Sox9-dependent mechanism. Further elucidation of the Notch1-Sox9 molecular pathway and its role in the maintenance of the ECM will lead to an improved mechanistic understanding of aortic valve calcification and development of novel therapeutic strategies for CAVD.

Abstract 295: Hemodynamic Shear Stress Alterations Contribute to Calcific Aortic Valve Disease in a Frequency- and Magnitude-Dependent Manner

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Ling Sun ◽  
John LeCluyse ◽  
Brian Robillard ◽  
Philippe Sucosky
Keyword(s):  
Shear Stress ◽  
Aortic Valve ◽  
Aortic Valve Disease ◽  
Ex Vivo ◽  
Fold Increase ◽  
Endothelial Activation ◽  
Valve Disease ◽  
Dependent Manner ◽  
Molecular Signaling ◽  
Calcific Aortic Valve Disease

INTRODUCTION: Calcific aortic valve disease (CAVD) is an active process presumably triggered by interplays between atherogenic risk factors, molecular signaling networks and hemodynamic cues. While our earlier work demonstrated that progressive alterations in fluid wall-shear stress (WSS) on the fibrosa could trigger leaflet inflammation, the mechanisms of CAVD pathogenesis secondary to side-specific WSS abnormalities are poorly understood. HYPOTHESIS: Supported by our previous studies, we hypothesize that valve leaflets are sensitive to both WSS magnitude and pulsatility and that abnormalities in either promote CAVD development. OBJECTIVE: This study aims at elucidating ex vivo the contribution of isolated and combined alterations in WSS magnitude and pulsatility to valvular calcification. METHODS: The fibrosa and ventricularis of porcine leaflets were subjected simultaneously to different combinations of WSS magnitude and pulsatility (i.e., physiologic, sub- and supra-physiologic levels) for 48 hours in a double-sided shear stress bioreactor. Endothelial activation (ICAM-1, VCAM-1), paracrine expression (TGF-β and BMP-4), and proteinase/collagenase expression (MMP-2, cathepsin L) were detected by immunohistochemistry, while osteogenic differentiation (α-SMA) was assessed via western blot. RESULTS: Regardless of the magnitude or frequency, non-physiologic WSS conditions did not result in endothelial activation. Tissue exposure to either supra-physiologic WSS magnitude or pulsatility significantly upregulated paracrine (74-fold increase), proteinase (4-fold increase), collagenase (5-fold increase) and α-SMA (23-fold increase) expressions relative to the levels measured under physiologic WSS. In contrast, combined alterations in WSS magnitude and pulsatility downregulated those responses. CONCLUSION: This study demonstrates the sensitivity of aortic valve leaflets to both WSS magnitude and pulsatility and the ability of supra-physiologic WSS magnitude or pulsatility to trigger events involved in early CAVD pathogenesis. The results provide new potential insights into the mechanisms of CAVD secondary to hypertension and Paget’s disease, which are associated with abnormal blood flow and leaflet WSS.

Contribution of Hemodynamic Shear Stress Magnitude and Frequency Abnormalities to Calcific Aortic Valve Disease Pathogenesis

2013 ◽  
Author(s):  
Ling Sun ◽  
Philippe Sucosky
Keyword(s):  
Shear Stress ◽  
Aortic Valve ◽  
Aortic Valve Disease ◽  
Fluid Shear Stress ◽  
Ex Vivo ◽  
Valve Disease ◽  
Molecular Signaling ◽  
Fluid Shear ◽  
Calcific Aortic Valve Disease ◽  
Hemodynamic Shear Stress

Calcific aortic valve disease (CAVD) is an active process presumably triggered by interplays between atherogenic risk factors, molecular signaling networks and hemodynamic cues. While our earlier work demonstrated that progressive alterations in fluid shear stress (FSS) on the fibrosa could trigger valvular inflammation [1], the mechanisms of CAVD pathogenesis secondary to side-specific FSS abnormalities are poorly understood. Supported by our previous studies, we hypothesize that valve leaflets are sensitive to both WSS magnitude and pulsatility and that abnormalities in either promote CAVD development. This study aims at elucidating ex vivo the contribution of isolated and combined alterations in FSS magnitude and pulsatility to valvular calcification.

Abstract 141: Bicuspid Aortic Valve Hemodynamic Abnormalities Promote Early Development of Calcific Aortic Valve Disease

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Ling Sun ◽  
Santanu Chandra ◽  
Philippe Sucosky
Keyword(s):  
Shear Stress ◽  
Aortic Valve ◽  
Osteogenic Differentiation ◽  
Bicuspid Aortic Valve ◽  
Aortic Valve Disease ◽  
Tissue Response ◽  
Valve Disease ◽  
Type I ◽  
Calcific Aortic Valve Disease ◽  
Congenital Cardiac Anomaly

INTRODUCTION: The bicuspid aortic valve (BAV) is the most common congenital cardiac anomaly and is frequently associated with calcific aortic valve disease (CAVD). Although CAVD also develops in the normal tricuspid aortic valve (TAV), its progression in the BAV is more rapid. While the accelerated calcification of BAV leaflets has been linked to genetic and atherogenic predispositions, hemodynamic abnormalities are increasingly pointed as potential pathogenic contributors. HYPOTHESIS: Supported by our previous work, which demonstrated the sensitivity of valve leaflets to the surrounding blood flow and associated wall-shear stress (WSS), we hypothesize that the abnormal WSS experienced by BAV leaflets contribute to CAVD development by promoting valvular inflammation, remodeling and osteogenic differentiation. OBJECTIVE: This study aims at comparing ex vivo the effects of TAV and BAV leaflet WSS on valvular pathogenesis. METHODS: The native, side-specific WSS experienced by TAV and type-I (i.e., fused and non-coronary) BAV leaflets were obtained computationally using fluid-structure interaction simulations. Fresh porcine leaflets were subjected for 48 hours to each of the three WSS conditions using a novel double-sided shear stress bioreactor. Tissue response was characterized via Western blot and immunohistochemistry in terms of markers of endothelial activation (VCAM-1, ICAM-1), paracrine expression (BMP-4), TGF-β/Wnt signaling pathways (TGF-β1, β-catenin), extracellular matrix remodeling (cathepsin L, MMP-2, MMP-9) and osteogenic differentiation (α-SMA, osteocalcin). RESULTS: No significant differences in VCAM-1 and ICAM-1 expressions were detected between tissue exposed to TAV and BAV WSS. While the native WSS experienced by the TAV and non-coronary BAV leaflets maintained tissue homeostasis, tissue exposure to the fused BAV leaflet WSS resulted in a significant pathological response marked by the upregulations of BMP-4, β-catenin, MMP-2 and osteocalcin expressions. CONCLUSION: This study demonstrates the pathological nature of the native BAV hemodynamics and confirms the higher susceptibility of the fused BAV leaflet to calcify. The results provide new insights into the hemodynamic theory of BAV calcification.

scholarly journals Calcific Aortic Valve Disease: Molecular Mechanisms And Therapeutic Approaches

2015 ◽  
Vol 10 (2) ◽  
pp. 108 ◽  
Author(s):  
Daniel Alejandro Lerman ◽  
Sai Prasad ◽  
Nasri Alotti ◽  
◽  
◽  
...  
Keyword(s):  
Aortic Valve ◽  
Aortic Valve Disease ◽  
Molecular Mechanisms ◽  
Cell Model ◽  
Human Monoclonal Antibody ◽  
Valve Disease ◽  
Vascular Lesions ◽  
Calcium Deposition ◽  
Calcific Aortic Valve Disease

Calcification occurs in atherosclerotic vascular lesions and in the aortic valve. Calcific aortic valve disease (CAVD) is a slow, progressive disorder that ranges from mild valve thickening without obstruction of blood flow, termed aortic sclerosis, to severe calcification with impaired leaflet motion, termed aortic stenosis. In the past, this process was thought to be ‘degenerative’ because of time-dependent wear and tear of the leaflets, with passive calcium deposition. The presence of osteoblasts in atherosclerotic vascular lesions and in CAVD implies that calcification is an active, regulated process akin to atherosclerosis, with lipoprotein deposition and chronic inflammation. If calcification is active, via pro-osteogenic pathways, one might expect that development and progression of calcification could be inhibited. The overlap in the clinical factors associated with calcific valve disease and atherosclerosis provides further support for a shared disease mechanism. In our recent research we used an in vitro porcine valve interstitial cell model to study spontaneous calcification and potential promoters and inhibitors. Using this model, we found that denosumab, a human monoclonal antibody targeting the receptor activator of nuclear factor-κB ligand may, at a working concentration of 50 μg/mL, inhibit induced calcium deposition to basal levels.

Abstract 15192: Leptin Effects on Osteoblast Differentiation of Human Valvular Interstitial Cells: New Insight Into Calcific Aortic Valve Disease

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Mickael Rosa ◽  
Rodrigo Lorenzi ◽  
Madjid Tagzirt ◽  
Francis Juthier ◽  
Antoine Rauch ◽  
...  
Keyword(s):  
Aortic Valve ◽  
Aortic Valve Disease ◽  
Osteoblast Differentiation ◽  
Interstitial Cells ◽  
Valve Disease ◽  
Calcium Deposition ◽  
Valvular Interstitial Cells ◽  
Calcific Aortic Valve Disease ◽  
Aortic Valves ◽  
Alp Activity

Introduction: Calcific aortic valve disease (CAVD) affects 2% to 6% of the population over 65 years and results from dysregulated processes such as calcification, supported in part by the osteoblast differentiation of valvular interstitial cells (VIC), the most prevalent cell type in the human aortic valves. Leptin has recently been linked to aortic valve calcification in ApoE-/- mice. Hypothesis: Our hypothesis is that leptin could play a role in the calcifying processes implicated in CAVD via direct effects on human VIC. Methods: Patients who underwent aortic valve replacement for severe CAVD (n=43) or with coronary artery disease (CAD) but without CAVD (n=129) were included in this study. Presence of leptin was analyzed in human explanted calcified aortic valves and blood samples. Leptin receptors expression was analyzed in aortic valves and VIC isolated from aortic valves. Leptin effects on osteoblast differentiation of VIC in presence or not of Akt and ERK inhibitors were investigated by alizarin red staining, alkaline phosphatase (ALP) activity, and RT-qPCR analysis for osteopontin, ALP, bone morphogenetic protein BMP-2, and RUNX2. Results: Patients with CAVD have significant higher serum leptin concentration than CAD patients (p=0.002). The presence of leptin was observed by immunochemistry in human calcified aortic valves, with higher concentrations in calcified vs non-calcified zones (p=0.01). Both short and long leptin receptor isoforms were expressed in VIC. Chronic leptin stimulation of VIC enhanced ALP, BMP-2 and RUNX2 expression and decreased osteopontin expression. This treatment led to a higher, dose dependent, ALP activity and calcium deposition in VIC. Inhibiting Akt or ERK during leptin stimulation led to a reduced calcification by bringing the expression of calcification genes to the control levels. Conclusions: Together, these novel findings depict the potential role of leptin in the process of CAVD by triggering calcification processes in human VIC.

Structural Deformation of Native Aortic Valve Leaflet Under Hypertension: An In Vitro Study

2009 ◽  
Author(s):  
C. H. Yap ◽  
H. S. Kim ◽  
L. P. Dasi ◽  
M. J. Weiler ◽  
K. Balachandran ◽  
...  
Keyword(s):  
Shear Stress ◽  
Aortic Valve ◽  
Aortic Valve Disease ◽  
Fluid Shear Stress ◽  
Complex Structure ◽  
Valve Disease ◽  
Structural Deformation ◽  
Fluid Shear ◽  
Calcific Aortic Valve Disease ◽  
Increased Risk

The aortic valve (AV) is a complex structure that functions in a complex dynamic environment. During systole, the valve leaflets bend at the base to open and experience fluid shear stress on both ventricular and aortic sides of the leaflet. During diastole, adverse pressure gradient closes the valve causing it to structurally support the systemic afterload pressure. Ex vivo experiments has shown that isolated mechanical forces such as pressure, membrane tension, and fluid shear stress affects the remodeling activities of the valve leaflets and also elicit pathological responses [1], potentially leading to calcific aortic valve disease in the long term. Clinically, patients with hypertension have increased risk of developing calcific aortic valve disease [2], which could be a result of the increased pressure or the increased stretch on the valve leaflets.

scholarly journals Ultrastructural Pathology of Atherosclerosis, Calcific Aortic Valve Disease, and Bioprosthetic Heart Valve Degeneration: Commonalities and Differences

2020 ◽  
Vol 21 (20) ◽  
pp. 7434 ◽  
Author(s):  
Alexander Kostyunin ◽  
Rinat Mukhamadiyarov ◽  
Tatiana Glushkova ◽  
Leo Bogdanov ◽  
Daria Shishkova ◽  
...  
Keyword(s):  
Aortic Valve ◽  
Heart Valve ◽  
Aortic Valve Disease ◽  
Circulatory System ◽  
Valve Disease ◽  
Calcium Deposition ◽  
Bioprosthetic Heart Valve ◽  
Calcific Aortic Valve Disease ◽  
Ultrastructural Pathology ◽  
Structural Valve Deterioration

Atherosclerosis, calcific aortic valve disease (CAVD), and bioprosthetic heart valve degeneration (alternatively termed structural valve deterioration, SVD) represent three diseases affecting distinct components of the circulatory system and their substitutes, yet sharing multiple risk factors and commonly leading to the extraskeletal calcification. Whereas the histopathology of the mentioned disorders is well-described, their ultrastructural pathology is largely obscure due to the lack of appropriate investigation techniques. Employing an original method for sample preparation and the electron microscopy visualisation of calcified cardiovascular tissues, here we revisited the ultrastructural features of lipid retention, macrophage infiltration, intraplaque/intraleaflet haemorrhage, and calcification which are common or unique for the indicated types of cardiovascular disease. Atherosclerotic plaques were notable for the massive accumulation of lipids in the extracellular matrix (ECM), abundant macrophage content, and pronounced neovascularisation associated with blood leakage and calcium deposition. In contrast, CAVD and SVD generally did not require vasculo- or angiogenesis to occur, instead relying on fatigue-induced ECM degradation and the concurrent migration of immune cells. Unlike native tissues, bioprosthetic heart valves contained numerous specialised macrophages and were not capable of the regeneration that underscores ECM integrity as a pivotal factor for SVD prevention. While atherosclerosis, CAVD, and SVD show similar pathogenesis patterns, these disorders demonstrate considerable ultrastructural differences.

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