Cardiac tissue engineering enables the generation of functional human cardiac tissue using cells in combination with biocompatible materials. Human pluripotent stem cell (hPSC)-derived cardiomyocytes provide a cell source for cardiac tissue engineering; however, their immaturity limits their potential applications. Here we sought to study the effect of mechanical conditioning and electrical pacing on the maturation of hPSC-derived cardiac tissues. In the first part of the study, cardiomyocytes derived from human induced pluripotent stem cells (hIPSCs) were used to generate collagen-based bioengineered human cardiac tissue. Engineered tissue constructs were subject to different stress and electrical pacing conditions. This engineered human myocardium exhibits Frank-Starling curve-type force-length relationships. After 2 weeks of static stress conditioning, the engineered myocardium demonstrated at least 10-fold increase in contractility and tensile stiffness, greater cell alignment, and a 1.5-fold increase in cell size and cell volume fraction within the constructs. Stress conditioning also increased sarco-endoplasmic reticulum calcium transport ATPase 2 (SERCA2) expression. When electrical pacing was combined with static stress conditioning, the tissues showed an additional 2-fold increase in force production, tensile stiffness, and contractility, with no change in cell alignment or cell size, suggesting maturation of excitation-contraction coupling. Supporting this notion, we found expression of RYR2 and SERCA2 further increased by combined static stress and electrical stimulation. These studies demonstrate that electrical pacing and mechanical stimulation promote both the structural and functional maturation of hiPSC-derived cardiac tissues. In the second part of the study, cardiovascular progenitor (CVP) cells derived from hPSC were used as the input cell population to generate engineered tissues. The effects of a 3-D microenvironment and mechanical stress on differentiation and maturation of human cardiovascular progenitors into myocardial tissue were evaluated. Compared to 2-D culture, the unstressed 3-D environment increased cardiomyocyte numbers and decreased smooth muscle numbers. Additionally, 3-D culture suppressed smooth muscle cell maturation. Mechanical stress conditioning further improved cardiomyocyte maturation. Cyclic stress-conditioning increased expression of several cardiac markers, like beta-myosin and cTnT, and the tissue showed enhanced force production. This 3-D system has facilitated understanding of the effect of mechanical stress on the differentiation and morphogenesis of distinct cardiovascular cell populations into organized, functional human cardiovascular tissues. In conclusion, we were able to create a complex engineered human cardiac tissue with both stem cell-derived cardiomyocytes and CVP cells. We showed that how environmental stimulations like mechanical stress, electrical pacing, and 3-D culturing can affect the maturation and specification of cells within the engineered cardiac tissues. The study paves our way to further apply these engineered cardiac tissues to other in vitro and in vivo usages like drug testing, clinical translation, and disease modeling.

Research centering on blood flow in the heart continues to hold an important position, especially since a better understanding of the subject may help reduce the incidence of coronary arterial disease and heart attacks. This book summarizes recent advances in the field; it is the product of fruitful cooperation among international scientists who met in Japan in May, 1990 to discuss the regulation of coronary blood flow.

Ischemic heart disease is a major cause of human death worldwide owing to the heart minimal ability to repair following injury. Other than heart transplantation, there are currently no effective or long-lasting therapies for end-stage heart failure. Therefore, it is crucial to develop not only alternative therapies that potentiate heart regeneration or repair, but also new tools to study human cardiac physiology and pathophysiology in vitro. In this context, cardiac tissue engineering arises a promising strategy, as it is aimed at generating cardiac tissue analogues that would act as in vitro models of human cardiac tissue or as surrogates for heart repair. Thus, having 3D human cardiac tissue constructs resembling human myocardium could revolutionize drug discovery and toxicity testing, cardiac disease modelling and regenerative medicine. An strategy to obtain reliable cardiac tissue constructs is to mimic the native cardiac environment. The classical approach is based on seeding cardiomyocytes in biocompatible 3D scaffolds, and then culturing the construct in a biomimetic signaling system, usually a bioreactor. Although major advances have been made, the generation of thick and mature tissue constructs from human induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CM) is still a challenge. Therefore, the hypothesis of our study is that the combination of hiPSC-CM with 3D scaffolds and appropriate regulatory signals may lead to the generation of mature human cardiac tissue constructs resembling human myocardium, both functionally and structurally. To address this, we have characterized a collagen-based 3D scaffold and established an efficient method for cell seeding into the scaffold. We have also developed a parallelized perfusion bioreactor system, which ensures an effective mass transport between cells and culture medium and allows culturing multiple replicas of tissue constructs. In addition, we have designed and fabricated a perfusion chamber including electrodes to electrically stimulate cells during culture, as well as to monitor tissue function. With this advanced 3D culture system, we have been able to generate thick 3D human cardiac constructs with tissue-like functionality. Our results indicate that perfusion of culture medium combined with electrical stimulation and collagen-based scaffold improve the structural and functional maturation of hiPSC-CM. In general terms, electrical stimulation has improved the structural organization, alignment and coupling of cardiomyocytes in our cardiac tissue constructs. Moreover, electrical stimulation has promoted the formation of synchronous contractile constructs at the macroscale with improved electrophysiological functions. Through the development of a new electrophysiological recording system, we report for the first time to our knowledge a technique that provides information about the electrical activity of intact cardiac tissue constructs in real time. Specifically, the combination of action potentials generated by hiPSC-CM composing cardiac constructs produces ECG-like signals, which could be monitored online. Finally, we have demonstrated the ability of stimulated human cardiac tissue constructs to detect drug-induced cardiotoxicity, as typical features of arrhythmias (e.g. prolongation of RR intervals and regular blockades) could be observed upon treatment with sotalol. Taken together, results indicate that macroscopic human cardiac tissue constructs with tissue-like functionality can be obtained through the use of our advanced 3D culture system. We have studied the effects of electrical stimulation on cardiomyocytes at multiple levels: molecular (presence, distribution and expression of cardiac proteins), ultrastructural (sarcomere width and presence of specialized cellular junctions), cellular (morphology and alignment), and functional (amplitude, directionality and strain of contractions, and electrophysiological recordings). Findings validate our in vitro approach as a valuable system to obtain 3D cardiac patches with an improved maturity and functionality. Importantly, the online monitoring system developed in this study can provide essential electrophysiological information of intact cardiac tissue constructs, which opens up myriad possibilities in the field of cardiovascular research.

Cellular and Molecular Pathobiology of Cardiovascular Disease focuses on the pathophysiology of common cardiovascular disease in the context of its underlying mechanisms and molecular biology. This book has been developed from the editors' experiences teaching an advanced cardiovascular pathology course for PhD trainees in the biomedical sciences, and trainees in cardiology, pathology, public health, and veterinary medicine. No other single text-reference combines clinical cardiology and cardiovascular pathology with enough molecular content for graduate students in both biomedical research and clinical departments. The text is complemented and supported by a rich variety of photomicrographs, diagrams of molecular relationships, and tables. It is uniquely useful to a wide audience of graduate students and post-doctoral fellows in areas from pathology to physiology, genetics, pharmacology, and more, as well as medical residents in pathology, laboratory medicine, internal medicine, cardiovascular surgery, and cardiology. - Explains how to identify cardiovascular pathologies and compare with normal physiology to aid research - Gives concise explanations of key issues and background reading suggestions - Covers molecular bases of diseases for better understanding of molecular events that precede or accompany the development of pathology

Cardiac Tissue Engineering: Methods and Protocols presents a collection of protocols on cardiac tissue engineering from pioneering and leading researchers around the globe. These include methods and protocols for cell preparation, biomaterial preparation, cell seeding, and cultivation in various systems. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Cardiac Tissue Engineering: Methods and Protocols highlights the major techniques, both experimental and computational, for the study of cardiovascular tissue engineering.