Nicole Feric

Biomaterials in myocardial tissue engineering.

Cardiovascular disease is the leading cause of death in the developed world, and as such there is a pressing need for treatment options. Cardiac tissue engineering emerged from the need to develop alternative sources and methods of replacing tissue damaged by cardiovascular diseases, as the ultimate treatment option for many who suffer from end‐stage heart failure is a heart transplant. In this review we focus on biomaterial approaches to augmenting injured or impaired myocardium, with specific emphasis on: the design criteria for these biomaterials; the types of scaffolds – composed of natural or synthetic biomaterials or decellularized extracellular matrix – that have been used to develop cardiac patches and tissue models; methods to vascularize scaffolds and engineered tissue; and finally, injectable biomaterials (hydrogels) designed for endogenous repair, exogenous repair or as bulking agents to maintain ventricular geometry post‐infarct. The challenges facing the field and obstacles that must be overcome to develop truly clinically viable cardiac therapies are also discussed. Copyright

Maturing human pluripotent stem cell-derived cardiomyocytes in human engineered cardiac tissues.

Engineering functional human cardiac tissue that mimics the native adult morphological and functional phenotype has been a long held objective. In the last 5 years, the field of cardiac tissue engineering has transitioned from cardiac tissues derived from various animal species to the production of the first generation of human engineered cardiac tissues (hECTs), due to recent advances in human stem cell biology. Despite this progress, the hECTs generated to date remain immature relative to the native adult myocardium. In this review, we focus on the maturation challenge in the context of hECTs, the present state of the art, and future perspectives in terms of regenerative medicine, drug discovery, preclinical safety testing and pathophysiological studies.

The Role of Tissue Engineering and Biomaterials in Cardiac Regenerative Medicine

In recent years, the development of 3-dimensional engineered heart tissue (EHT) has made large strides forward because of advances in stem cell biology, materials science, prevascularization strategies, and nanotechnology. As a result, the role of tissue engineering in cardiac regenerative medicine has become multifaceted as new applications become feasible. Cardiac tissue engineering has long been established to have the potential to partially or fully restore cardiac function after cardiac injury. However, EHTs may also serve as surrogate human cardiac tissue for drug-related toxicity screening. Cardiotoxicity remains a major cause of drug withdrawal in the pharmaceutical industry. Unsafe drugs reach the market because preclinical evaluation is insufficient to weed out cardiotoxic drugs in all their forms. Bioengineering methods could provide functional and mature human myocardial tissues, ie, physiologically relevant platforms, for screening the cardiotoxic effects of pharmaceutical agents and facilitate the discovery of new therapeutic agents. Finally, advances in induced pluripotent stem cells have made patient-specific EHTs possible, which opens up the possibility of personalized medicine. Herein, we give an overview of the present state of the art in cardiac tissue engineering, the challenges to the field, and future perspectives.

Hydrogel substrate stiffness and topography interact to induce contact guidance in cardiac fibroblasts.

Previous studies demonstrated the importance of substrate stiffness and topography on the phenotype of many different cell types including fibroblasts. Yet the interaction of these two physical parameters remains insufficiently characterized, in particular for cardiac fibroblasts. Most studies focusing on contact guidance use rigid patterned substrates. It is not known how the ability of cardiac fibroblasts to follow grooves and ridges changes as the substrate stiffness is decreased to match the range of stiffness found in native heart tissues. This report demonstrates a significant interactive effect of substrate stiffness and topography on cardiac fibroblast elongation and orientation using polyacrylamide substrates of different stiffness and topography.

Strategies and Challenges to Myocardial Replacement Therapy

Cardiovascular diseases account for the majority of deaths globally and are a significant drain on economic resources. Although heart transplants and left‐ventricle assist devices are the solution for some, the best chance for many patients who suffer because of a myocardial infarction, heart failure, or a congenital heart disease may be cell‐based regenerative therapies. Such therapies can be divided into two categories: the application of a cell suspension and the implantation of an in vitro engineered tissue construct to the damaged area of the heart. Both strategies have their advantages and challenges, and in this review, we discuss the current state of the art in myocardial regeneration, the challenges to success, and the future direction of the field.

Diabetic wound regeneration using peptide-modified hydrogels to target re-epithelialization

Significance Current treatments for diabetic chronic wounds fail to achieve effective therapeutic outcomes. The majority of these treatments focus on angiogenesis, but diabetes often involves endothelial dysfunction. A hallmark of regenerative wound healing is rapid, effective re-epithelialization. In this study, we present QHREDGS (glutamine-histidine-arginine-glutamic acid-aspartic acid-glycine-serine), a prosurvival peptide derived from angiopoietin-1, as a therapeutic candidate that targets re-epithelialization. Immobilized QHREDGS peptide promoted cell survival against hydrogen peroxide stress and collective cell migration of both normal and diabetic human keratinocytes in vitro. The clinical relevance was demonstrated further in type 2 diabetic mice: A single treatment with a low QHREDGS dose immobilized in chitosan–collagen was effective in promoting wound healing, and a single high-dose peptide treatment outperformed a clinically approved porous collagen dressing. There is a clinical need for new, more effective treatments for chronic wounds in diabetic patients. Lack of epithelial cell migration is a hallmark of nonhealing wounds, and diabetes often involves endothelial dysfunction. Therefore, targeting re-epithelialization, which mainly involves keratinocytes, may improve therapeutic outcomes of current treatments. In this study, we present an integrin-binding prosurvival peptide derived from angiopoietin-1, QHREDGS (glutamine-histidine-arginine-glutamic acid-aspartic acid-glycine-serine), as a therapeutic candidate for diabetic wound treatments by demonstrating its efficacy in promoting the attachment, survival, and collective migration of human primary keratinocytes and the activation of protein kinase B Akt and MAPKp42/44. The QHREDGS peptide, both as a soluble supplement and when immobilized in a substrate, protected keratinocytes against hydrogen peroxide stress in a dose-dependent manner. Collective migration of both normal and diabetic human keratinocytes was promoted on chitosan–collagen films with the immobilized QHREDGS peptide. The clinical relevance was demonstrated further by assessing the chitosan–collagen hydrogel with immobilized QHREDGS in full-thickness excisional wounds in a db/db diabetic mouse model; QHREDGS showed significantly accelerated and enhanced wound closure compared with a clinically approved collagen wound dressing, peptide-free hydrogel, or blank wound controls. The accelerated wound closure resulted primarily from faster re-epithelialization and increased formation of granulation tissue. There were no observable differences in blood vessel density or size within the wound; however, the total number of blood vessels was greater in the peptide-hydrogel–treated wounds. Together, these findings indicate that QHREDGS is a promising candidate for wound-healing interventions that enhance re-epithelialization and the formation of granulation tissue.

Hydrogels With Integrin-Binding Angiopoietin-1-Derived Peptide, QHREDGS, for Treatment of Acute Myocardial Infarction

Background—Hydrogels are being actively investigated for direct delivery of cells or bioactive molecules to the heart after myocardial infarction (MI) to prevent cardiac functional loss. We postulate that immobilization of the prosurvival angiopoietin-1–derived peptide, QHREDGS, to a chitosan-collagen hydrogel could produce a clinically translatable thermoresponsive hydrogel to attenuate post-MI cardiac remodeling. Methods and Results—In a rat MI model, QHREDGS-conjugated hydrogel (QHG213H), control gel, or PBS was injected into the peri-infarct/MI zone. By in vivo tracking and chitosan staining, the hydrogel was demonstrated to remain in situ for 2 weeks and was cleared in ≈3 weeks. By echocardiography and pressure–volume analysis, the QHG213H hydrogel significantly improved cardiac function compared with the controls. Scar thickness and scar area fraction were also significantly improved with QHG213H gel injection compared with the controls. There were significantly more cardiomyocytes, determined by cardiac troponin-T staining, in the MI zone of the QHG213H hydrogel group; and hydrogel injection did not induce a significant inflammatory response as assessed by polymerase chain reaction and an inflammatory cytokine assay. The interaction of cardiomyocytes and cardiac fibroblasts with QHREDGS was found to be mediated by &bgr;1-integrins. Conclusions—We demonstrated for the first time that the QHG213H peptide–modified hydrogel can be injected in the beating heart where it remains localized for a clinically effective period. Moreover, the QHG213H hydrogel induced significant cardiac functional and morphological improvements after MI relative to the controls.

Angiopoietin-1 peptide QHREDGS promotes osteoblast differentiation, bone matrix deposition and mineralization on biomedical materials.

Bone loss occurs as a consequence of a variety of diseases as well as from traumatic injuries, and often requires therapeutic intervention. Strategies for repairing and replacing damaged and/or lost bone tissue include the use of biomaterials and medical implant devices with and without osteoinductive coatings. The soluble growth factor angiopoietin-1 (Ang-1) has been found to promote cell adhesion and survival in a range of cell types including cardiac myocytes, endothelial cells and fibroblasts through an integrin-dependent mechanism. Furthermore, the short sequence QHREDGS has been identified as the integrin-binding sequence of Ang-1 and as a synthetic peptide has been found to possess similar integrin-dependent effects as Ang-1 in the aforementioned cell types. Integrins have been implicated in osteoblast differentiation and bone mineralization, processes critical to bone regeneration. By binding integrins on the osteoblast surface, QHREDGS could promote cell survival and adhesion, as well as conceivably osteoblast differentiation and bone mineralization. Here we immobilized QHREDGS onto polyacrylate (PA)-coated titanium (Ti) plates and polyethylene glycol (PEG) hydrogels. The osteoblast differentiation marker, alkaline phosphatase, peaked in activity 4-12 days earlier on the QHREDGS-immobilized PA-coated Ti plates than on the unimmobilized, DGQESHR (scrambled)- and RGDS-immobilized surfaces. Significantly more bone matrix was deposited on the QHREDGS-immobilized Ti surface than on the other surfaces as determined by atomic force microscopy. The QHREDGS-immobilized hydrogels also had a significantly higher mineral-to-matrix (M/M) ratio determined by Fourier transform infrared spectroscopy. Alizarin Red S and von Kossa staining and quantification, and environmental scanning electron microscopy showed that while both the QHREDGS- and RGDS-immobilized surfaces had extensive mineralization relative to the unimmobilized and DGQESHR-immobilized surfaces, the mineralization was more considerable on the QHREDGS-immobilized surface, both with and without the induction of osteoblast differentiation. Finally, treatment of cell monolayers with soluble QHREDGS was demonstrated to upregulate osteogenic gene expression. Taken together, these results demonstrate that the QHREDGS peptide is osteoinductive, inducing osteoblast differentiation, bone matrix deposition and mineralization.

The role of Wnt regulation in heart development, cardiac repair and disease: A tissue engineering perspective

Wingless-related integration site (Wnt) signaling has proven to be a fundamental mechanism in cardiovascular development as well as disease. Understanding its particular role in heart formation has helped to develop pluripotent stem cell differentiation protocols that produce relatively pure cardiomyocyte populations. The resultant cardiomyocytes have been used to generate heart tissue for pharmaceutical testing, and to study physiological and disease states. Such protocols in combination with induced pluripotent stem cell technology have yielded patient-derived cardiomyocytes that exhibit some of the hallmarks of cardiovascular disease and are therefore being used to model disease states. While FDA approval of new treatments typically requires animal experiments, the burgeoning field of tissue engineering could act as a replacement. This would necessitate the generation of reproducible three-dimensional cardiac tissues in a well-controlled environment, which exhibit native heart properties, such as cellular density, composition, extracellular matrix composition, and structure-function. Such tissues could also enable the further study of Wnt signaling. Furthermore, as Wnt signaling has been found to have a mechanistic role in cardiac pathophysiology, e.g. heart attack, hypertrophy, atherosclerosis, and aortic stenosis, its strategic manipulation could provide a means of generating reproducible and specific, physiological and pathological cardiac models.

Human pluripotent stem cell-derived cardiomyocyte based models for cardiotoxicity and drug discovery.

The drug development process, from discovery to safety studies, aims to bring efficacious and safe drugs to market. Unfortunately, this ambition is often thwarted by unanticipated adverse effects that are not detected by the current drug-testing paradigm. Cardiotoxicity ranks as one of the primary causes of drug recalls, an extraordinarily costly event in terms of both the manufacturer’s bottom line and the negative patient outcomes [1]. As such, developing more predictive cardiotoxicity assessments is a main focus of research. Currently, preclinical cardiac assessments mainly consist of animal studies and heterologous expression systems of human cardiac ion channel in noncardiac cell lines. Animal models provide valuable in vivo pharmacodynamic and pharmacokinetic data, and their genes can be precisely manipulated for mechanistic investigations. However, species–species differences hinder their applicability as a model of human patients. For example, rodents have heart rates 3–10 times faster than humans [2], virtually no delayed rectifier potassium IKr (human ether-a-gogo-related gene [hERG]) current, and animal ion channel expression and action potential profiles are generally distinct from those of humans [3]. Human-specific drug screening data have been obtained using primary heart cells and tissue, tumor-derived immortalized mammalian cardiac cell lines, and heterologous expression systems in which noncardiac cells (Chinese Hamster Ovary or Human Embryonic Kidney) are transfected to express an individual human ion channel, most commonly the hERG channel. Most of the cardiotoxicity/arrhythmia safety screens performed to date have employed the latter as a means of assessing the potential for acute drug-induced electrocardiogram (ECG) abnormalities. These human-specific models are limited by genetic instability and the absence of important cardiac cell characteristics. Taken as a whole, the preclinical paradigm is able to predict 70–90% [1] of human patient drug effects, yet many promising drugs are triaged early in development and some cardiotoxic drugs still enter the market. To minimize the use of animal models and to improve the precision of risk prediction, various alternative platforms have been developed for drug safety evaluation. Here, we discuss the current challenges to the cardiotoxicity and drug discovery fields, the state of the art and we envision a more predictive human cardiac model.