Reprogramming cardiomyocyte mechanosensing by crosstalk between integrins and hyaluronic acid receptors
ABSTRACT The elastic modulus of bioengineered materials has a strong influence on the phenotype of many cells including cardiomyocytes. On polyacrylamide (PAA) gels that are laminated with ligands for integrins, cardiac myocytes develop well organized sarcomeres only when cultured on substrates with elastic moduli in the range 10 kPa-30 kPa, near those of the healthy tissue. On stiffer substrates (>60 kPa) approximating the damaged heart, myocytes form stress fiber-like filament bundles but lack organized sarcomeres or an elongated shape. On soft (<1 kPa) PAA gels myocytes exhibit disorganized actin networks and sarcomeres. However, when the polyacrylamide matrix is replaced by hyaluronic acid (HA) as the gel network to which integrin ligands are attached, robust development of functional neonatal rat ventricular myocytes occurs on gels with elastic moduli of 200 Pa, a stiffness far below that of the neonatal heart and on which myocytes would be amorphous and dysfunctional when cultured on polyacrylamide-based gels. The HA matrix by itself is not adhesive for myocytes, and the myocyte phenotype depends on the type of integrin ligand that is incorporated within the HA gel, with fibronectin, gelatin, or fibrinogen being more effective than collagen I. These results show that HA alters the integrin-dependent stiffness response of cells in vitro and suggests that expression of HA within the extracellular matrix (ECM) in vivo might similarly alter the response of cells that bind the ECM through integrins. The integration of HA with integrin-specific ECM signaling proteins provides a rationale for engineering a new class of soft hybrid hydrogels that can be used in therapeutic strategies to reverse the remodeling of the injured myocardium.
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ABSTRACT: The extracellular matrix is no longer considered a static support structure for cells, but a dynamic signaling network with the power to influence cell, tissue and whole organ physiology. In the myocardium, cardiac fibroblasts are the primary cell type responsible for the synthesis, deposition and degradation of matrix proteins and they therefore play a critical role in the development and maintenance of functional heart tissue. This review will summarize the extensive research conducted in vivo and in vitro demonstrating the influence of both physical and chemical stimuli on cardiac fibroblasts and how these interactions impact both cardiomyocytes and the extracellular matrix. This work is of considerable significance given that cardiovascular diseases are marked by extensive remodeling of the extracellular matrix, which ultimately impairs the functional capacity of the heart. We seek to summarize the unique role of cardiac fibroblasts in normal cardiac development and the most prevalent cardiac pathologies including congenital heart defects, hypertension, hypertrophy, and the remodeled heart following myocardial infarction. We will conclude by identifying existing holes in the research that, if answered, have the potential to dramatically improve current therapeutic strategies for the repair and regeneration of damaged myocardium via mechanotransductive signaling.Journal of Biomechanical Engineering 05/2013; DOI:10.1115/1.4024349 · 1.75 Impact Factor
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ABSTRACT: The current practice of cell therapy, in which multipotent or terminally differentiated cells are injected into tissues or intravenously, is inefficient. Few therapeutic cells are retained at the site of administration and engraftment is low. An injectable and biologically appropriate vehicle for delivery, retention, growth and differentiation of therapeutic cells is needed to improve the efficacy of cell therapy. We focus on a hyaluronan-based semi-synthetic extracellular matrix (sECM), HyStem®, which is a manufacturable, approvable and affordable clinical product. The composition of this sECM can be customized for use with mesenchymal stem cells as well as cells derived from embryonic or induced pluripotent sources. In addition, it can support therapeutic uses of progenitor and mature cell populations obtained from skin, fat, liver, heart, muscle, bone, cartilage, nerves and other tissues. This overview presents four pre-clinical uses of HyStem® for cell therapy to repair injured vocal folds, improve post-myocardial infarct heart function, regenerate damaged liver tissue and restore brain function following ischemic stroke. Finally, we address the real-world limitations - manufacture, regulation, market acceptance and financing - surrounding cell therapy and the development of clinical combination products.Acta biomaterialia 07/2012; 8(12). DOI:10.1016/j.actbio.2012.06.043 · 5.68 Impact Factor
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ABSTRACT: INTRODUCTION: The mechanical environment is a key regulator of function in cardiomyocytes. We studied the role of substrate stiffness on the organization of sarcomeres and costameres in adult rat cardiomyocytes and further examined the resulting changes in cell shortening and calcium dynamics. METHODS: Cardiomyocytes isolated from adult rats were plated on laminin-coated polydimethylsiloxane substrates of defined stiffness (255 kPa, 117 kPa, 27 kPa, and 7 kPa) for 48 h. Levels of α-actinin and β1 integrins were determined by immunofluoresence imaging and immunoblotting, both in the absence and presence of the phosphatase inhibitor calyculin A. Quantitative reverse transcriptase polymerase chain reaction was used to measure message levels of key structural proteins (α-actinin, α7 integrin, β1 integrin, vinculin). Sarcomere shortening and calcium dynamics were measured at 2, 24, and 48 h. RESULTS: Overall cardiomyocyte morphology was similar on all substrates. However, well organized sarcomere structures were observed on only the stiffest (255 kPa) and the most compliant (7 kPa) substrates. Levels of α-actinin in cells were the same on all substrates, while message levels of structural proteins were up-regulated on substrates of intermediate stiffness. Inhibition of phosphatase activity blocked the degradation of contractile structures, but altered overall cardiomyocyte morphology. Shortening and calcium dynamics also were dependent on substrate stiffness; however, there was no clear causative relationship between the phenomena. CONCLUSIONS: Extracellular matrix stiffness can affect structural remodeling by adult cardiomyocytes, and the resulting contractile activity. These findings illuminate changes in cardiomyocyte function in cardiac fibrosis, and may suggest cardiac-specific phosphatases as a target for therapeutic intervention.Cardiovascular pathology: the official journal of the Society for Cardiovascular Pathology 12/2012; 22(3). DOI:10.1016/j.carpath.2012.10.003 · 2.34 Impact Factor