Endothelial cell-cardiomyocyte cross-talk: understanding bidirectional paracrine signaling in cardiovascular homeostasis and disease.

To maintain homeostasis in the heart, endothelial cells and cardiomyocytes engage in dynamic cross-talk through paracrine signals that regulate both cardiac development and function. Here, we review the paracrine signals that endothelial cells release to regulate cardiomyocyte growth, hypertrophy and contractility, and the factors that cardiomyocytes release to influence angiogenesis and vascular tone. Dysregulated communication between these cell types can drive pathophysiology of disease, as seen in ischemia-reperfusion injury, diabetes, maladaptive hypertrophy, and chemotherapy-induced cardiotoxicity.

The use of a multi-metric readout screen to identify EHMT2/G9a-inhibition as a modulator of cancer-associated fibroblast activation state.

Cancer-associated fibroblasts (CAFs) play a pivotal role in cancer progression, including mediating tumour cell invasion via their pro-invasive secretory profile and ability to remodel the extracellular matrix (ECM). Given that reduced CAF abundance in tumours correlates with improved outcomes in various cancers, we set out to identify epigenetic targets involved in CAF activation in regions of tumour-stromal mixing with the goal of reducing tumour aggressiveness. Using the GLAnCE (Gels for Live Analysis of Compartmentalized Environments) platform, we performed an image-based, phenotypic screen that enabled us to identify modulators of CAF abundance and the capacity of CAFs to induce tumour cell invasion.

Characterization of pediatric porcine pulmonary valves as a model for tissue engineered heart valves.

Heart valve tissue engineering holds the potential to transform the surgical management of congenital heart defects affecting the pediatric pulmonary valve (PV) by offering a viable valve replacement. While aiming to recapitulate the native valve, the minimum requirement for tissue engineered heart valves (TEHVs) has historically been adequate mechanical function at implantation. However, long-term in situ functionality of TEHVs remains elusive, suggesting that a closer approximation of the native valve is required.

Sensitivity and Validation of Porous Membrane Electrical Cell Substrate Impedance Spectroscopy (PM-ECIS) for Measuring Endothelial Barrier Properties.

Measurement of endothelial and epithelial barrier integrity is important for a variety of in vitro models, including Transwell assays, cocultures, and organ-on-chip platforms. Barrier resistance is typically measured by trans-endothelial electrical resistance (TEER), but TEER is invasive and cannot accurately measure isolated monolayer resistance in coculture or most organ-on-chip devices. These limitations are addressed by porous membrane electrical cell-substrate impedance sensing (PM-ECIS), which measures barrier integrity in cell monolayers grown directly on permeable membranes patterned with electrodes.

Identification of congenital aortic valve malformations in juvenile natriuretic peptide receptor 2-deficient mice using high-frequency ultrasound.

Mouse models of congenital aortic valve malformations are useful for studying disease pathobiology, but most models have incomplete penetrance [e.g., ∼2 to 77% prevalence of bicuspid aortic valves (BAVs) across multiple models].

Bridging barriers: advances and challenges in modeling biological barriers and measuring barrier integrity in organ-on-chip systems.

Biological barriers such as the blood-brain barrier, skin, and intestinal mucosal barrier play key roles in homeostasis, disease physiology, and drug delivery - as such, it is important to create representative models to improve understanding of barrier biology and serve as tools for therapeutic development. Microfluidic cell culture and organ-on-a-chip (OOC) systems enable barrier modelling with greater physiological fidelity than conventional platforms by mimicking key environmental aspects such as fluid shear, accurate microscale dimensions, mechanical cues, extracellular matrix, and geometrically defined co-culture. As the prevalence of barrier-on-chip models increases, so does the importance of tools that can accurately assess barrier integrity and function without disturbing the carefully engineered microenvironment.

Myosin inhibitor reverses hypertrophic cardiomyopathy in genotypically diverse pediatric iPSC-cardiomyocytes to mirror variant correction.

Pathogenic variants in MYH7 and MYBPC3 account for the majority of hypertrophic cardiomyopathy (HCM). Targeted drugs like myosin ATPase inhibitors have not been evaluated in children. We generate patient and variant-corrected iPSC-cardiomyocytes (CMs) from pediatric HCM patients harboring single variants in MYH7 (V606M; R453C), MYBPC3 (G148R) or digenic variants (MYBPC3 P955fs, TNNI3 A157V).

Interrogating Matrix Stiffness and Metabolomics in Pancreatic Ductal Carcinoma Using an Openable Microfluidic Tumor-on-a-Chip.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a dense fibrotic stroma that contributes to aggressive tumor biology and therapeutic resistance. Current in vitro PDAC models lack sufficient optical and physical access for fibrous network visualization, in situ mechanical stiffness measurement, and metabolomic profiling. Here, we describe an openable multilayer microfluidic PDAC-on-a-chip platform that consists of pancreatic tumor cells (PTCs) and pancreatic stellate cells (PSCs) embedded in a 3D collagen matrix that mimics the stroma.

Porous Membrane Electrical Cell-Substrate Impedance Spectroscopy for Versatile Assessment of Biological Barriers In Vitro.

Cell culture models of endothelial and epithelial barriers typically use porous membrane inserts (e.g., Transwell inserts) as a permeable substrate on which barrier cells are grown, often in coculture with other cell types on the opposite side of the membrane.

Biomechanical properties of the aortic root are distinct from those of the ascending aorta in both normal and aneurysmal states.

Background: Although aneurysms of the ascending aorta and the aortic root are treated similarly in clinical guidelines, how biomechanical properties differ between these 2 segments of aorta is poorly defined.

Methods: Biomechanical testing was performed on tissue collected from the aortic root (normal = 11, aneurysm = 51) and the ascending aorta (normal = 21, aneurysm = 76). Energy loss, tangent modulus of elasticity, and delamination strength were evaluated.

Noninvasive Quantification of Contractile Dynamics in Cardiac Cells, Spheroids, and Organs-on-a-Chip Using High-Frequency Ultrasound.

Cell-based models that mimic in vivo heart physiology are poised to make significant advances in cardiac disease modeling and drug discovery. In these systems, cardiomyocyte (CM) contractility is an important functional metric, but current measurement methods are inaccurate and low-throughput or require complex setups. To address this need, we developed a standalone noninvasive, label-free ultrasound technique operating at 40-200 MHz to measure the contractile kinetics of cardiac models, ranging from single adult CMs to 3D microtissue constructs in standard cell culture formats.

Bioinspired Oligo-Urethane Nanoparticles for Delivering Exogenous C-Type Natriuretic Peptide: Synthetic Biomaterial Nanocarrier Complexes and Their Interactions with Cardiac Myofibroblasts.

In a healthy heart, cells naturally secrete C-type natriuretic peptide (CNP), a cytokine that protects against myofibroblast differentiation of cardiac fibroblasts and extracellular matrix deposition leading to fibrosis. CNP availability during myocardial remodeling is important to prevent cardiac fibrosis, but CNP is limited after an injury because of the loss of cardiomyocytes and the activation of cardiac fibroblasts to myofibroblasts. We hypothesized that the sustained release of exogenous CNP from oligo-urethane nanoparticles (NPs) would reduce differentiation of human cardiac fibroblasts toward a myofibrogenic phenotype.

Serum- and xeno-free culture of human umbilical cord perivascular cells for pediatric heart valve tissue engineering.

Background: Constructs currently used to repair or replace congenitally diseased pediatric heart valves lack a viable cell population capable of functional adaptation in situ, necessitating repeated surgical intervention. Heart valve tissue engineering (HVTE) can address these limitations by producing functional living tissue in vitro that holds the potential for somatic growth and remodelling upon implantation. However, clinical translation of HVTE strategies requires an appropriate source of autologous cells that can be non-invasively harvested from mesenchymal stem cell (MSC)-rich tissues and cultured under serum- and xeno-free conditions.

Ascending aortic geometry and its relationship to the biomechanical properties of aortic tissue.

Objective: The objective of this study was to evaluate the relationship between ascending aortic geometry and biomechanical properties.

Methods: Preoperative computed tomography scans from ascending aortic aneurysm patients were analyzed using a center line technique (n = 68). Aortic length was measured from annulus to innominate artery, and maximal diameter from this segment was recorded.

Assessing engineered tissues and biomaterials using ultrasound imaging: In vitro and in vivo applications.

Quantitative assessment of the structural, functional, and mechanical properties of engineered tissues and biomaterials is fundamental to their development for regenerative medicine applications. Ultrasound (US) imaging is a non-invasive, non-destructive, and cost-effective technique capable of longitudinal and quantitative monitoring of tissue structure and function across centimeter to sub-micron length scales. Here we present the fundamentals of US to contextualize its application for the assessment of biomaterials and engineered tissues, both in vivo and in vitro.

High-frequency quantitative ultrasound for the assessment of the acoustic properties of engineered tissues in vitro.

Acoustic properties of biomaterials and engineered tissues reflect their structure and cellularity. High-frequency ultrasound (US) can non-invasively characterize and monitor these properties with sub-millimetre resolution. We present an approach to estimate the speed of sound, acoustic impedance, and acoustic attenuation of cell-laden hydrogels that accounts for frequency-dependent effects of attenuation in coupling media, hydrogel thickness, and interfacial transmission/reflection coefficients of US waves, all of which can bias attenuation estimates.

Tmem65 is critical for the structure and function of the intercalated discs in mouse hearts.

The intercalated disc (ICD) is a unique membrane structure that is indispensable to normal heart function, yet its structural organization is not completely understood. Previously, we showed that the ICD-bound transmembrane protein 65 (Tmem65) was required for connexin43 (Cx43) localization and function in cultured mouse neonatal cardiomyocytes. Here, we investigate the functional and cellular effects of Tmem65 reductions on the myocardium in a mouse model by injecting CD1 mouse pups (3-7 days after birth) with recombinant adeno-associated virus 9 (rAAV9) harboring Tmem65 shRNA, which reduces Tmem65 expression by 90% in mouse ventricles compared to scrambled shRNA injection.

Harnessing conserved signaling and metabolic pathways to enhance the maturation of functional engineered tissues.

The development of induced-pluripotent stem cell (iPSC)-derived cell types offers promise for basic science, drug testing, disease modeling, personalized medicine, and translatable cell therapies across many tissue types. However, in practice many iPSC-derived cells have presented as immature in physiological function, and despite efforts to recapitulate adult maturity, most have yet to meet the necessary benchmarks for the intended tissues. Here, we summarize the available state of knowledge surrounding the physiological mechanisms underlying cell maturation in several key tissues.

Immunomagnetic Isolation and Enrichment of Microvascular Endothelial Cells from Human Adipose Tissue.

Human adipose tissue-resident microvascular endothelial cells are not only garnering attention for their emergent role in the pathogenesis of obesity-related metabolic disorders, but are also of considerable interest for vascular tissue engineering due, in part, to the abundant, accessible, and uniquely dispensable nature of the tissue. Here, we delineate a protocol for the acquisition of microvascular endothelial cells from human fat. A cheaper, smaller, and simpler alternative to fluorescence-assisted cell sorting for the immunoselection of cells, our protocol adapts magnet-assisted cell sorting for the isolation of endothelial cells from enzymatically digested adipose tissue and the subsequent enrichment of their primary cultures.

Hearts by design.

Scalable biofabrication of heart helical tissue pattern augments pumping function.

A Carbon-Based Biosensing Platform for Simultaneously Measuring the Contraction and Electrophysiology of iPSC-Cardiomyocyte Monolayers.

Heart beating is triggered by the generation and propagation of action potentials through the myocardium, resulting in the synchronous contraction of cardiomyocytes. This process highlights the importance of electrical and mechanical coordination in organ function. Investigating the pathogenesis of heart diseases and potential therapeutic actions requires biosensing technologies which allow for long-term and simultaneous measurement of the contractility and electrophysiology of cardiomyocytes.

An SCPPPQ1/LAM332 protein complex enhances the adhesion and migration of oral epithelial cells: Implications for dentogingival regeneration.

Common periodontal disease treatment procedures often fail to restore the structural integrity of the junctional epithelium (JE), the epithelial attachment of the gum to the tooth, leaving the tooth-gum interface prone to bacterial colonization. To address this issue, we introduced a novel bio-inspired protein complex comprised of a proline-rich enamel protein, SCPPPQ1, and laminin 332 (LAM332) to enhance the JE attachment. Using quartz crystal microbalance with dissipation monitoring (QCM-D), we showed that SCPPPQ1 and LAM332 interacted and assembled into a protein complex with high-affinity adsorption of 5.

Design of a Mechanobioreactor to Apply Anisotropic, Biaxial Strain to Large Thin Biomaterials for Tissue Engineered Heart Valve Applications.

Repair and replacement solutions for congenitally diseased heart valves capable of post-surgery growth and adaptation have remained elusive. Tissue engineered heart valves (TEHVs) offer a potential biological solution that addresses the drawbacks of existing valve replacements. Typically, TEHVs are made from thin, fibrous biomaterials that either become cell populated in vitro or in situ.