Human Valve Research Articles

Patient Prosthesis Mismatch

AbstractConcept of patient prosthesis mismatch came into existence from 1978 onward when Rahimtoola first defined it as “mismatch can be considered to be present when the effective prosthetic valve area, after insertion into the patient, is less than that of a normal human valve.” Patient prosthesis mismatch produces higher than expected pressure gradient through normally functioning valve. Since insertion of first ball caged mechanical valve in descending aorta by Dr. Charles Hufnagel in 1952, prosthetic valve had undergone tremendous improvement in terms of valve design, hemodynamics, durability, and thrombogenicity. Despite these marked changes in valve design, prosthetic valves are still subjected to inherent complications.

Abstract 10223: Tissue-Entrapped Extracellular Vesicles Modulate Divergent Mechanisms of Cardiovascular Calcification

Introduction: Fewer than 50% of patients develop vascular and valvular calcification, implying differential pathogenesis. Tissue-entrapped extracellular vesicles (EVs) are implicated in mineralization but their contents and functions are unstudied. We investigated entrapped EV cargoes in human cardiovascular disease. Methods: Human carotid endarterectomy specimens and stenotic aortic valves were obtained from 53 patients. Disease stage-specific proteomics was performed on whole tissue (non-diseased/fibrotic/calcified areas). Tissue EVs were enriched by gradient fractionation then underwent proteomics and miRNA-seq. miR targets were predicted by TargetScan, pathway analyses utilized BioCarta/KEGG/Reactome, and protein-protein interaction networks employed STRING. Results: Disease progression drove significant convergence (p<0.0001) of atherosclerotic plaque and valve proteomes (2,318 proteins). 548 and 158 proteins were exclusively altered (q<0.05) by disease in plaques or valves, respectively. Vesicular GO terms increased 2.2x (p<0.01) amongst proteins altered by disease in both tissues (202). Proteomics found 24 EV markers in the low-density fractions of plaques and valves, confirmed by electron microscopy and nanoparticle tracking. EV omics quantified 1,104 proteins and 123 miR cargoes. Networks of proteins and miR targets shared by plaque and valve EVs revealed common regulation of Rho GTPase and MAPK signaling. 179 proteins and 5 miRs were altered between plaque and valve EVs (q<0.05); multi-omics integration found that EVs modulated cellular contraction and p53-mediated transcriptional regulation in plaques and valves, respectively. Conclusions: This first comparative proteomics study of human valves and arteries finds shared EV functionality in both diseases. Using novel means to examine tissue EV molecular cargoes, we also reveal critical divergent tissue-specific roles for EVs in mediating cardiovascular disease.

Abstract 13243: Integrin Linked Kinase Expression in Human Valvular Endothelial Cells Plays a Protective Role in Calcific Aortic Valve Disease

Introduction: Calcific aortic valve disease (CAVD) is the most common valvular heart disease in the aging population. CAVD is a highly active ossification process originated in valvular interstitial cells which differentiate towards an osteoblastic phenotype. However, the role of valvular endothelial cells in CAVD has not been fully explored. We previously demonstrated that ILK expression in vascular endothelium plays an essential cardioprotective role. Hypothesis: Endothelial ILK plays a protective role in human valvular endothelial cells osteogenesis. Methods and Results: Human aortic valves were obtained from 52 patients with calcific aortic stenosis compared with 16 non-calcified aortic valves and ILK levels were analyzed. Our study showed a significant relationship between the degree of AV calcification and decreased ILK expression detected by IHQ and qPCR (p< 0,013). Moreover, decreased ILK expression correlated with BMP2 and Runx2, two markers of valvular calcification (p<0.01). Human valvular endothelial cells (hVECs) isolated from non-calcified controls display the ability to calcify when cultured in pro-calcifiying conditions and ILK levels were unaffected by the pro-calcifiying conditions. However, silencing ILK expression in hVECs resulted in decreased NO production, increased expression of calcific disease markers Runx2, BMP2, and osteopontin. ILK silencing in hVECs induced calcium deposition, determined by alizarin red staining, when cultured in pro-calcifiying conditions. Thus, our results suggest that decreased ILK expression could promote endothelial to osteogenic phenotype transition. Restoring nitric oxide levels by DETA-NONOate treatment in ILK silenced hVEC was able to reverse those effects partially. Conclusions: Collectively, our results point to a crucial role of ILK in maintaining human valve endothelial cell function preventing valvular calcification.

Abstract 11048: Serotonin Transporter Deficiency Leads to a Specific Fibrotic Pattern in Valvular and Myocardial Tissue

Introduction: Increased serotonin (5HT) levels can lead to heart valve disease. This mechanism is observed in patients with 5HT-secreting carcinoid tumors and after treatment with the diet drug Dexfenfluoramine, which acts as a 5HT receptor (HTR)-2B agonist and as an inhibitor of the 5HT transporter (SERT). SERT internalizes 5HT, limiting HTR signaling (Figure 1A). We showed that SERT reduction on human mitral valve (MV) contributes to the progression of MV regurgitation. We investigated the impact of SERT deficiency on cardiac substructures in a SERT-/- murine model. Methods: SERT-/- and wild type (WT) C57BL/6J mice (n≥8/group) were sacrificed at 16 weeks of age, after echocardiography (Fig. 1B). Cardiac substructures (left ventricle (LV) apex, MV leaflets, aortic valve (AV) leaflets and aortic-mitral (AM curtain) were isolated in three animals/group (Fig. 1E). Results: Ventricular dimensions were significantly larger in SERT-/- mice vs. WT at end-systole and end-diastole (Fig. 1C). Fractional shortening was significantly reduced in SERT-/- mice vs. WT. Pricosirius staining of collagen showed that SERT-/- mice had increased MV leaflet fibrosis, MV and AV sub-valvular fibrosis at leaflet attachment points (AM curtain, Fig. 1D), and epicardial fibrosis. PCR (Fig. 1F) showed significantly increased COL1A1 expression in the LV myocardium, MV, AV and AM curtain in SERT-/- mice vs. WT. Fibrosis marker alpha smooth muscle actin (αSMA) was significantly increased in the myocardium and AM curtain of SERT-/- mice, but not in the valve leaflets. HTR2B expression was significantly increased in the MV and AM curtain of SERT-/- mice. Conclusions: SERT-/- mice have decreased cardiac function, and cardiac fibrosis that is associated with HTR2B upregulation in the MV leaflets and valve leaflet attachment points. Our results support the hypothesis that decreased 5HT clearance by SERT downregulation may increase HTR2B signaling in the MV leading to remodeling and regurgitation.

Differential proteome profile, biological pathways, and network relationships of osteogenic proteins in calcified human aortic valves.

Calcific aortic valve disease (CAVD) is the most common heart valve disease requiring intervention. Most research on CAVD has focused on inflammation, ossification, and cellular phenotype transformation. To gain a broader picture into the wide range of cellular and molecular mechanisms involved in this disease, we compared the total protein profiles between calcified and non-calcified areas from 5 human valves resected during surgery. The 1413 positively identified proteins were filtered down to 248 proteins present in both calcified and non-calcified segments of at least 3 of the 5 valves, which were then analyzed using Ingenuity Pathway Analysis. Concurrently, the top 40 differentially abundant proteins were grouped according to their biological functions and shown in interactive networks. Finally, the abundance of selected osteogenic proteins (osteopontin, osteonectin, osteocalcin, osteoprotegerin, and RANK) was quantified using ELISA and/or immunohistochemistry. The top pathways identified were complement system, acute phase response signaling, metabolism, LXR/RXR and FXR/RXR activation, actin cytoskeleton, mineral binding, nucleic acid interaction, structural extracellular matrix (ECM), and angiogenesis. There was a greater abundance of osteopontin, osteonectin, osteocalcin, osteoprotegerin, and RANK in the calcified regions than the non-calcified ones. The osteogenic proteins also formed key connections between the biological signaling pathways in the network model. In conclusion, this proteomic analysis demonstrated the involvement of multiple signaling pathways in CAVD. The interconnectedness of these pathways provides new insights for the treatment of this disease.

A cadaveric study on chordae tendinae of human tricuspid valve

Aim of the present study was to observe the types of chordae tendinae present in tricuspid valve of human heart. Morphology of chordae tendinae in tricuspid valve gains utmost importance in cardiac surgeries in recent times because advent in modern technologies in treatment of tricuspid valve diseases.This study was carried out on 96 normal formalin fixed human post-mortem heart specimens. Types of chordae tendinae observed on the basis of their attachments.Chordae tendinae were observed in all specimens. Five types of chordae tendinae were identified namely rough zone, free edge, fan shaped, deep and basal chordae. Anterior papillary muscle was seen providing attachment to 2 to 9; Posterior papillary muscles were seen with 1 to 6 and Septal papillary muscles provided attachment to 1 to 4 chordae tendinae.We hope this study will serve to understand the tricuspid valve complex and types of different chordae tendinae better and it will help in various surgical procedures done on tricuspid valve.

Palmdelphin Regulates Nuclear Resilience to Mechanical Stress in the Endothelium.

PALMD (palmdelphin) belongs to the family of paralemmin proteins implicated in cytoskeletal regulation. Single nucleotide polymorphisms in the PALMD locus that result in reduced expression are strong risk factors for development of calcific aortic valve stenosis and predict severity of the disease. Immunodetection and public database screening showed dominant expression of PALMD in endothelial cells (ECs) in brain and cardiovascular tissues including aortic valves. Mass spectrometry, coimmunoprecipitation, and immunofluorescent staining allowed identification of PALMD partners. The consequence of loss of PALMD expression was assessed in small interferring RNA-treated EC cultures, knockout mice, and human valve samples. RNA sequencing of ECs and transcript arrays on valve samples from an aortic valve study cohort including patients with the single nucleotide polymorphism rs7543130 informed about gene regulatory changes. ECs express the cytosolic PALMD-KKVI splice variant, which associated with RANGAP1 (RAN GTP hydrolyase activating protein 1). RANGAP1 regulates the activity of the GTPase RAN and thereby nucleocytoplasmic shuttling via XPO1 (Exportin1). Reduced PALMD expression resulted in subcellular relocalization of RANGAP1 and XPO1, and nuclear arrest of the XPO1 cargoes p53 and p21. This indicates an important role for PALMD in nucleocytoplasmic transport and consequently in gene regulation because of the effect on localization of transcriptional regulators. Changes in EC responsiveness on loss of PALMD expression included failure to form a perinuclear actin cap when exposed to flow, indicating lack of protection against mechanical stress. Loss of the actin cap correlated with misalignment of the nuclear long axis relative to the cell body, observed in PALMD-deficient ECs, Palmd-/- mouse aorta, and human aortic valve samples derived from patients with calcific aortic valve stenosis. In agreement with these changes in EC behavior, gene ontology analysis showed enrichment of nuclear- and cytoskeleton-related terms in PALMD-silenced ECs. We identify RANGAP1 as a PALMD partner in ECs. Disrupting the PALMD/RANGAP1 complex alters the subcellular localization of RANGAP1 and XPO1, and leads to nuclear arrest of the XPO1 cargoes p53 and p21, accompanied by gene regulatory changes and loss of actin-dependent nuclear resilience. Combined, these consequences of reduced PALMD expression provide a mechanistic underpinning for PALMD's contribution to calcific aortic valve stenosis pathology.

Hypoxic Culture Maintains Cell Growth of the Primary Human Valve Interstitial Cells with Stemness.

The characterization of aortic valve interstitial cells (VICs) cultured under optimal conditions is essential for understanding the molecular mechanisms underlying aortic valve stenosis. Here, we propose 2% hypoxia as an optimum VIC culture condition. Leaflets harvested from patients with aortic valve regurgitation were digested using collagenase and VICs were cultured under the 2% hypoxic condition. A significant increase in VIC growth was observed in 2% hypoxia (hypo-VICs), compared to normoxia (normo-VICs). RNA-sequencing revealed that downregulation of oxidative stress-marker genes (such as superoxide dismutase) and upregulation of cell cycle accelerators (such as cyclins) occurred in hypo-VICs. Accumulation of reactive oxygen species was observed in normo-VICs, indicating that low oxygen tension can avoid oxidative stress with cell-cycle arrest. Further mRNA quantifications revealed significant upregulation of several mesenchymal and hematopoietic progenitor markers, including CD34, in hypo-VICs. The stemness of hypo-VICs was confirmed using osteoblast differentiation assays, indicating that hypoxic culture is beneficial for maintaining growth and stemness, as well as for avoiding senescence via oxidative stress. The availability of hypoxic culture was also demonstrated in the molecular screening using proteomics. Therefore, hypoxic culture can be helpful for the identification of therapeutic targets and the evaluation of VIC molecular functions in vitro.

Mutations in EPHB4 cause human venous valve aplasia.

Venous valve (VV) failure causes chronic venous insufficiency, but the molecular regulation of valve development is poorly understood. A primary lymphatic anomaly, caused by mutations in the receptor tyrosine kinase EPHB4, was recently described, with these patients also presenting with venous insufficiency. Whether the venous anomalies are the result of an effect on VVs is not known. VV formation requires complex “organization” of valve-forming endothelial cells, including their reorientation perpendicular to the direction of blood flow. Using quantitative ultrasound, we identified substantial VV aplasia and deep venous reflux in patients with mutations in EPHB4. We used a GFP reporter in mice to study expression of its ligand, ephrinB2, and analyzed developmental phenotypes after conditional deletion of floxed Ephb4 and Efnb2 alleles. EphB4 and ephrinB2 expression patterns were dynamically regulated around organizing valve-forming cells. Efnb2 deletion disrupted the normal endothelial expression patterns of the gap junction proteins connexin37 and connexin43 (both required for normal valve development) around reorientating valve-forming cells and produced deficient valve-forming cell elongation, reorientation, polarity, and proliferation. Ephb4 was also required for valve-forming cell organization and subsequent growth of the valve leaflets. These results uncover a potentially novel cause of primary human VV aplasia.

Abstract MP08: Telomerase Reverse Transcriptase Mediates Osteogenesis In Calcific Aortic Valve Disease

Calcific aortic valve disease (CAVD) is the leading heart valve disorder in the US. It is characterized by an active accumulation of calcium nodules on the aortic valve leaflets which lead to stiffening and remodeling of the valve leaflets causing valve dysfunction, cardiac failure and increased stroke risk. Inflammation and mechanical stresses contribute to CAVD pathogenesis. However, the mechanisms driving the fibrocalcific remodeling of the aortic valve are currently ill-defined. Multiple studies have revealed that the catalytic subunit of telomerase reverse transcriptase (TERT) can induce gene transcription and its overexpression primes mesenchymal stem cells to differentiate into osteoblasts, suggesting that TERT has a role in the activation of osteogenic transcriptional programs. We hypothesized that TERT contributes to early events leading to calcification of the valve leaflet. In human calcified valve tissue, we found that TERT protein is highly expressed in areas of calcification compared to control valve tissue, with no effect on telomere length. Alpha-SMA, a VIC activation marker, and RUNX2, a key transcription factor involved in the osteogenic differentiation of osteoblast, were also elevated in CAVD tissue. Under osteogenic differentiation conditions, human valve interstitial cells (VICs) upregulated TERT, RUNX2, and alpha-SMA protein levels and calcified, while CAVD VICs calcified de novo. Inflammatory stimuli intensified in vitro calcification, and induced TERT, RUNX2, and alpha-SMA protein expression. PLA and ChIP analysis showed that TERT interacts with interacted with Signal Transducer and Activator of Transcription 5A/B (STAT5) and together bind to RUNX2 promoter, respectively. shRNA-mediated TERT downregulation reduced expression of RUNX2 and alpha-SMA and genetic deletion of Tert in murine mesenchymal stem cells and vascular smooth muscle cells prevented calcification. These data provide evidence that TERT is required for calcification, regulates the transition of quiescent VICs into calcifying VICs, and that STAT5 functions as a TERT-interacting partner for DNA binding.

Abstract P113: Multi Omics Analysis Identified Fndc1 And Mxra5 As Novel Extracellular Matrix Proteins In Aortic Stenosis

Calcified aortic valve disease (CAVD) affects over six million Americans and is associated with changes in valve leaflets' mechanical properties, resulting in impaired valvular blood flow. Currently, there is no viable pharmacological treatment to stop the disease's progression or activate mineral regression. The only effective therapy to treat CAVD is aortic valve replacement (AVR) or transcatherization (TAVR). Therefore, it is imperative to understand the molecular mechanisms leading to aortic valve mineralization to identify new pharmacological targets. In this work, we examined the expression of extracellular matrix proteins (ECM) during valve calcification. Method: We used both RNAseq (n=18) and Proteomics (n=15) on explanted human valves of two different human cohorts to study the modulation of ECM proteins. Proteins were isolated from the fibrotic and the calcific side of each valve tissue. We next confirm our data in isolated human valve cells. Cells were treated with calcifying medium for one week and secreted ECM proteins were analyzed at different time points: Baseline, 3 days, and 7 days. Results: our network analysis revealed an interaction between ECM, metabolic, complement, and lipids transporters proteins. We identified SPP1, VTN, THSB2, MATN2, FNDC1, and MXRA5 as the common modulated ECM protein in both RNAseq and proteomics data set. Conclusion: This study yields new insight into ECM proteins expression during the aortic valve calcification process, and identified two new ECM proteins, MXRA5 and FNDC1, in aortic stenosis. The disruption of ECM proteins was more significant in the calcified stage and it was accompanied by a down-regulation of many metabolic enzymes.

High fructose induced osteogenic differentiation of human valve interstitial cells via activating PI3K/AKT/mitochondria signaling

BackgroundAortic valve stenosis (AS) is a common, lethal cardiovascular disease. There is no cure except the valve replacement at last stage. Therefore, an understanding of the detail mechanism is imperative to prevent and intervene AS. Metabolic syndrome (MetS) is one of the major risk factors of AS whereas fructose overconsuming tops the list of MetS risk factors. However, whether the fructose under physiological level induces AS is currently unknown. MethodsThe human valve interstitial cells (hVICs), a crucial source to develop calcification, were co-incubated with fructose at 2 or 20 mM to mimic the serum fructose at fasting or post-fructose consumption, respectively, for 24 h. The cell proliferation was evaluated by WST-1 assays. The expressions of osteogenic and fibrotic proteins, PI3K/AKT signaling, insulin receptor substrate 1 and mitochondrial dynamic proteins were detected by Western blot analyses. The mitochondrial oxidative phosphorylation (OXPHOS) was examined by Seahorse analyzer. ResultshVICs proliferation was significantly suppressed by 20 mM fructose. The expressions of alkaline phosphatase (ALP) and osteocalcin were enhanced concurrent with the upregulated PI3K p85, AKT, phospho(p)S473-AKT, and pS636-insulin receptor substrate 1 (p-IRS-1) by high fructose. Moreover, ATP production capacity and maximal respiratory capacity were enhanced in the high fructose groups. Synchronically, the expressions of mitochondrial fission 1 and optic atrophy type 1 were increased. ConclusionsThese results suggested that high fructose stimulated the osteogenic differentiation of hVICs via the activation of PI3K/AKT/mitochondria signaling at the early stage. These results implied that high fructose at physiological level might have a direct, hazard effect on the progression of AS.

Clinical and Technical Challenges of Prosthesis–Patient Mismatch After Transcatheter Aortic Valve Implantation

Prosthesis–patient mismatch (PPM) is present when the effective area of a prosthetic valve inserted into a patient is inferior to that of a normal human valve; the hemodynamic consequence of a valve too small compared with the size of the patient's body is the generation of higher than expected transprosthetic gradients. Despite evidence of increased risk of short- and long-term mortality and of structural valve degeneration in patients with PPM after surgical aortic valve replacement, its clinical impact in patients subject to transcatheter aortic valve implantation (TAVI) is yet unclear. We aim to review and update on the definition and incidence of PPM after TAVI, and its prognostic implications in the overall population and in higher-risk subgroups, such as small aortic annuli or valve-in-valve procedures. Last, we will focus on the armamentarium available in order to reduce risk of PPM when planning a TAVI procedure.

Influence of Annular Dynamics and Material Behavior in Finite Element Analysis of Barlow\u2019s Mitral Valve Disease

Barlow’s disease affects the entire mitral valve apparatus, by altering several of the fundamental mechanisms in the mitral valve which ensures unidirectional blood flow between the left atrium and the left ventricle. In this paper, a finite element model of a patient diagnosed with Barlow’s disease with patient-specific geometry and boundary conditions is presented. The geometry and boundary conditions are extracted from the echocardiographic assessment of the patient prior to surgery. Material properties representing myxomatous, healthy human and animal mitral valves are implemented and computed response are compared with each other and the echocardiographic images of the patient. This study shows that the annular dilation observed in Barlow’s patients controls several aspects of the mitral valve behavior during ventricular systole. The coaptation of the leaflets is observed to be highly dependent on annular dilation, and the coaptation area reduces rapidly at the onset of mitral regurgitation. Furthermore, the leaflet material implementation is important to predict lack of closure in the FE model correctly. It was observed that using healthy human material parameters in the Barlow’s diseased FE geometry gave severe lack of closure from the onset of mitral regurgitation, while myxomatous material properties showed a more physiological leakage.

Isolation of Human Primary Valve Cells for In vitro Disease Modeling

Isolation of Human Primary Valve Cells for In vitro Disease Modeling

Isolation of Human Primary Valve Cells for In vitro Disease Modeling.

Calcific aortic valve disease (CAVD) is present in nearly a third of the elderly population. Thickening, stiffening, and calcification of the aortic valve causes aortic stenosis and contributes to heart failure and stroke. Disease pathogenesis is multifactorial, and stresses such as inflammation, extracellular matrix remodeling, turbulent flow, and mechanical stress and strain contribute to the osteogenic differentiation of valve endothelial and valve interstitial cells. However, the precise initiating factors that drive the osteogenic transition of a healthy cell into a calcifying cell are not fully defined. Further, the only current therapy for CAVD-induced aortic stenosis is aortic valve replacement, whereby the native valve is removed (surgical aortic valve replacement, SAVR) or a fully collapsible replacement valve is inserted via a catheter (transcatheter aortic valve replacement, TAVR). These surgical procedures come at a high cost and with serious risks; thus, identifying novel therapeutic targets for drug discovery is imperative. To that end, the present study develops a workflow where surgically removed tissues from patients and donor cadaver tissues are used to create patient-specific primary lines of valvular cells for in vitro disease modeling. This protocol introduces the utilization of a cold storage solution, commonly utilized in organ transplant, to reduce the damage caused by the often-lengthy procurement time between tissue excision and laboratory processing with the benefit of greatly stabilizing cells of the excised tissue. The results of the present study demonstrate that isolated valve cells retain their proliferative capacity and endothelial and interstitial phenotypes in culture upwards of several days after valve removal from the donor. Using these materials allows for the collection of control and CAVD cells, from which both control and disease cell lines are established.

Anatomical Similarities Between Mitral and Tricuspid Valves in Sheep, Ovis aries, Hearts

With the recent rapid growth in transcatheter mitral and tricuspid valve interventions, it has become increasingly important to understand detailed anatomy of the valves. In this study, we investigated the similarities, differences, and associations between the dimensions of the atrioventricular valves in a sheep model, as sheep heart valves have a similar morphology to human valves. A systematic dissection of twenty-five sheep hearts was performed, with annular circumference measurement, and sub-valvular anatomy documentation. There was a significant association (r=0.865; p=0.007) between the circumference of the mitral and tricuspid annuli. Authors also identified significantly more chordae tendinea in the subvalvular mitral apparatus compared to the tricuspid valve (15.8±1.2 vs. 13.9±1.5; p<0.001). In conclusion, there is a significant association between the size of mitral and tricuspid valve annuli, and the morphology of leaflets and subvalvular apparatus is different between the two valves. These findings could have important implications in transcatheter device design, sizing, and optimal intervention timing.

Engineered Hydrogels as Valve Tissue Mimetics That Recapitulate Sexual Dimorphisms Observed in Calcification and Osteopontin Activity

Objective: Recent clinical reports indicate that patients with calcific aortic valve stenosis (AVS) have sexually dimorphic disease phenotypes, where female patients have reduced valve calcification compared to males. Yet, the cellular mechanisms contributing to this dimorphism remain unknown. Our results reveal that diseased valve leaflets from a female tend to have reduced and smaller-scale microcalcifications compared to males. Additionally, osteopontin (OPN) expression, a pro-fibrotic but anti-calcific bone sialoprotein, is elevated in human female valve tissue sections. We hypothesize that OPN may play a role in reducing calcification in female aortic valves. Methods: To this end, we use a hydrogel system with cellularly-degradable crosslinkers and adhesive peptides to culture VICs isolated from male and female pigs in 3D. We further introduced a secondary interpenetrating network of collagen type I and studied differences in the cellular response to osteogenic stimuli. Results: The hydrogel matrix supported VIC survival, matrix remodeling, and recapitulated the mineralization differences that are observed in human diseased valve tissue. Specifically, female valve tissue and female VICs cultured in pro-calcifying hydrogels have more punctate calcification (< 3000 μm2) relative to male samples (> 3000 μm2) (Figure 1). Further, OPN localization shifted from the hydrogel extracellular matrix to the nucleus in female cells in response to exogenously delivered calcification stimuli. Conclusions: OPN localization corresponded to sites of VIC-mediated matrix calcification, suggesting OPN may have a role in suppressing mineralization growth in AVS progression. This 3D hydrogel culture system should prove useful to test additional hypotheses related to sex-specific AVS calcification and potentially guide the development of sex-specific treatments. Figure 1. Von Kossa stained histological sections display reduced calcification of female samples (left) relative to male (right), in both tissue sections of human diseased aortic valve (top) and within in vitro 3D hydrogel valve mimetics (bottom). Scale bar = 100 μm

Obesity Promotes Extracellular Matrix and Metabolic Proteins Network in Aortic Stenosis

Objective: Obesity is one the main risk factors promoting the progression of aortic stenosis. Today, there is a critical need to identify new therapeutic targets to reverse or stop the progression of aortic stenosis. This study aimed to evaluate the modulation of extracellular matrix proteins (ECM) and its impact on metabolic enzymes during aortic valve mineralization. Methods: We analyzed ECM markers expression by mass spectrometry and RNA sequencing on explanted human aortic valves and isolated human valve cells, in 2 independent human cohorts (n= 15 vs n=17). Results: We identified a complex linkage between calcified and fibrotic phases, with an elaborate interaction between ECM, metabolic, complement and lipid transporter proteins. Aldehyde Dehydrogenase 1 Family Member A1 (ALDH1), a major enzyme in alcohol metabolism, was down regulated in both fibrotic and calcified stages. We identified Fibronectin Type III Domain Containing 1 (FNDC1) and Matrix-remodeling Associated 5 (MXRA5) as the common modulated ECM protein in calcified valve tissue in both proteomic and RNAseq data sets. Conclusion: This work highlights the interactome between metabolic and ECM markers, and identified FNDC1 and MXRA5 as novel ECM markers in calcified valves, electing them as potential targets in the development and the progression of AS.

Pre-surgical Prediction of Ischemic Mitral Regurgitation Recurrence Using In Vivo Mitral Valve Leaflet Strains.

Ischemic mitral regurgitation (IMR) is a prevalent cardiac disease associated with substantial morbidity and mortality. Contemporary surgical treatments continue to have limited long-term success, in part due to the complex and multi-factorial nature of IMR. There is thus a need to better understand IMR etiology to guide optimal patient specific treatments. Herein, we applied our finite element-based shape-matching technique to non-invasively estimate peak systolic leaflet strains in human mitral valves (MVs) from in-vivo 3D echocardiographic images taken immediately prior to and post-annuloplasty repair. From a total of 21 MVs, we found statistically significant differences in pre-surgical MV size, shape, and deformation patterns between the with and without IMR recurrence patient groups at 6 months post-surgery. Recurrent MVs had significantly less compressive circumferential strains in the anterior commissure region compared to the recurrent MVs (p=0.0223) and were significantly larger. A logistic regression analysis revealed that average pre-surgical circumferential leaflet strain in the Carpentier A1 region independently predicted 6-month recurrence of IMR (optimal cutoff value -18%, p=0.0362). Collectively, these results suggest greater disease progression in the recurrent group and underscore the highly patient-specific nature of IMR. Importantly, the ability to identify such factors pre-surgically could be used to guide optimal treatment methods to reduce post-surgical IMR recurrence.