Heart valves are dynamic structures that, in the average human, open and close over 100,000 times per day, and 3 × 109 times per lifetime to maintain unidirectional blood flow. Efficient, coordinated movement of the valve structures during the cardiac cycle is mediated by the intricate and sophisticated network of extracellular matrix (ECM) components that provide the necessary biomechanical properties to meet these mechanical demands. Organized in layers that accommodate passive functional movements of the valve leaflets, heart valve ECM is synthesized during embryonic development, and remodeled and maintained by resident cells throughout life. The failure of ECM organization compromises biomechanical function, and may lead to obstruction or leaking, which if left untreated can lead to heart failure. At present, effective treatment for heart valve dysfunction is limited and frequently ends with surgical repair or replacement, which comes with insuperable complications for many high-risk patients including aged and pediatric populations. Therefore, there is a critical need to fully appreciate the pathobiology of biomechanical valve failure in order to develop better, alternative therapies. To date, the majority of studies have focused on delineating valve disease mechanisms at the cellular level, namely the interstitial and endothelial lineages. However, less focus has been on the ECM, shown previously in other systems, to be a promising mechanism-inspired therapeutic target. Here, we highlight and review the biology and biomechanical contributions of key components of the heart valve ECM. Furthermore, we discuss how human diseases, including connective tissue disorders lead to aberrations in the abundance, organization and quality of these matrix proteins, resulting in instability of the valve infrastructure and gross functional impairment.

Human genetic studies identified a strong association between loss of function mutations in RBFOX2 and hypoplastic left heart syndrome (HLHS). There are currently no Rbfox2 mouse models that recapitulate HLHS. Therefore, it is still unknown how RBFOX2 as an RNA binding protein contributes to heart development. To address this, we conditionally deleted Rbfox2 in embryonic mouse hearts and found profound defects in cardiac chamber and yolk sac vasculature formation. Importantly, our Rbfox2 conditional knockout mouse model recapitulated several molecular and phenotypic features of HLHS. To determine the molecular drivers of these cardiac defects, we performed RNA-sequencing in Rbfox2 mutant hearts and identified dysregulated alternative splicing (AS) networks that affect cell adhesion to extracellular matrix (ECM) mediated by Rho GTPases. We identified two Rho GTPase cycling genes as targets of RBFOX2. Modulating AS of these two genes using antisense oligos led to cell cycle and cell-ECM adhesion defects. Consistently, Rbfox2 mutant hearts displayed cell cycle defects and inability to undergo endocardial-mesenchymal transition, processes dependent on cell-ECM adhesion and that are seen in HLHS. Overall, our work not only revealed that loss of Rbfox2 leads to heart development defects resembling HLHS, but also identified RBFOX2-regulated AS networks that influence cell-ECM communication vital for heart development.

Rationale: Calcific Aortic Valve Disease (CAVD) affects more than 5.2 million people in the US. The only effective treatment is surgery and this comes with complications and no guarantee of long-term success. Objective: Outcomes from pharmacological initiatives remain unsubstantiated and therefore the aim of this study is to determine if repurposing a selective XPO1 inhibitor drug (KPT-330) is beneficial in the treatment of CAVD. Methods and Results: We show that KPT-330 prevents, attenuates and mitigates calcific nodule formation in heart valve interstitial cells (VICs) in vitro, and prevents CAVD in Klotho -/- mice. Using RNA-sequencing and Mass Spectrometry we show that KPT-330's beneficial effect is mediated by inhibiting nuclear export of the transcription factor C/EBPβ in VICs, leading to repression of canonical Wnt signaling, in part through activation of the Wnt antagonist Axin1, and a subsequent decrease in pro-osteogenic markers and cell viability. Conclusions: Our findings have met a critical need to discover alterative, pharmacological-based therapies in the treatment of CAVD.

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While current clinical utilization of large vascular grafts for vascular transplantation is encouraging, tissue engineering of small grafts still faces numerous challenges. This study aims to investigate the feasibility of constructing a small vascular graft from the decellularized amniotic membranes (DAMs). DAMs were rolled around a catheter and each of the resulting grafts was crosslinked with a) 0.1% glutaraldehyde; b) EDC (20mM)-NHS (10mM); c) 0.5% genipin; and d) no-crosslinking, respectively. Our results demonstrated the feasibility of using a rolling technique followed by lyophilization to transform DAM into a vessel-like structure. The genipin-crosslinked DAM graft showed an improved integrated structure, prolonged stability, proper mechanical property, and superior biocompatibility. After transplantation in rat abdominal aorta, the genipin-crosslinked DAM graft remained patent up to 16-month, with both endothelial and smooth muscle cell regeneration, which suggests that the genipin-crosslinked DAM graft has great potential to be implemented as a small tissue engineered graft for future vascular transplantation.

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