Ravi K. Birla

Self-organization of rat cardiac cells into contractile 3-D cardiac tissue

The mammalian heart is not known to regenerate following injury. Therefore, there is great interest in developing viable tissue‐based models for cardiac assist. Recent years have brought numerous advances in the development of scaffold‐based models of cardiac tissue, but a self‐organizing model has yet to be described. Here, we report the development of an in vitro cardiac tissue without scaffolding materials in the contractile region. Using an optimal concentration of the adhesion molecule laminin, a confluent layer of neonatal rat cardiomyogenic cells can be induced to self‐organize into a cylindrical construct, resembling a papillary muscle, which we have termed a cardioid. Like endogenous heart tissue, cardioids contract spontaneously and can be electrically paced between 1 and 5 Hz indefinitely without fatigue. These engineered cardiac tissues also show an increased rate of spontaneous contraction (chronotropy), increased rate of relaxation (lusitropy), and increased force production (inotropy) in response to epinephrine. Cardioids have a developmental protein phenotype that expresses both α‐ and β‐tropomyosin, very low levels of SERCA2a, and very little of the mature isoform of cardiac troponin T.

Microfeature guided skeletal muscle tissue engineering for highly organized 3-dimensional free-standing constructs.

Engineering tissue similar in structure to their natural equivalents is a major challenge and crucial to function. Despite attempts to engineer skeletal muscle, it is still difficult to effectively mimic tissue architecture. Rigid scaffolds can guide cell alignment but have the critical drawback of hindering mechanical function of the resultant tissue. We present a method for creating highly ordered tissue-only constructs by using rigid microtopographically patterned surfaces to first guide myoblast alignment, followed by transfer of aligned myotubes into a degradable hydrogel and self-organization of the ordered cells into a functional, 3-dimensional, free-standing construct independent of the initial template substrate. Histology revealed an intracellular organization resembling that of native muscle. Aligned cell constructs exhibited a 2-fold increase in peak force production compared to controls. Effective specific force, or force normalized over cross-sectional area, was increased by 23%. This template, transfer, and self-organization strategy is envisioned to be broadly useful in improving construct function and clinical applicability for highly ordered tissues like muscle.

Poly(glycerol-dodecanoate), a biodegradable polyester for medical devices and tissue engineering scaffolds

In this paper we describe the mechanical and biological features of a thermosetting polyester synthesized from glycerol and dodecanedioic acid named Poly-Glycerol-Dodecanoate (PGD). This polymer shows a glass transition temperature (T(g)) around 32 degrees C, and this accounts for its mechanical properties. At room temperature (21 degrees ) PGD behaves like a stiff elastic-plastic material, while at body temperature (37 degrees C), it shows a compliant non-linear elastic behavior. Together with biodegradability and biocompatibility PGD has distinct shape memory features. After the polymer is cured, no matter what the final configuration is, we can recover the original shape by heating PGD to temperatures of 32 degrees C and higher. The mechanical properties together with biocompatibility/biodegradability and shape memory features make PGD an attractive polymer for biomedical applications.

Engineering the heart piece by piece: state of the art in cardiac tissue engineering

According to the National Transplant Society, more than 7000 Americans in need of organs die every year owing to a lack of lifesaving organs. Bioengineering 3D organs in vitro for subsequent implantation may provide a solution to this problem. The field of tissue engineering in its most rudimentary form is focused on the developed of transplantable organ substitutes in the laboratory. The objective of this article is to introduce important technological hurdles in the field of cardiac tissue engineering. This review starts with an overview of tissue engineering, followed by an introduction to the field of cardiovascular tissue engineering and finally summarizes some of the key advances in cardiac tissue engineering; specific topics discussed in this article include cell sourcing and biomaterials, in vitro models of cardiac muscle and bioreactors. The article concludes with thoughts on the utility of tissue-engineering models in basic research as well as critical technological hurdles that need to be addressed in the future.

Tissue-engineered heart valve prostheses: 'state of the heart'

In this article, we will review the current state of the art in heart valve tissue engineering. We provide an overview of mechanical and biological replacement options, outlining advantages and limitations of each option. Tissue engineering, as a field, is introduced, and specific aspects of valve tissue engineering are discussed (e.g., biomaterials, cells and bioreactors). Technological hurdles, which need to be overcome for advancement of the field, are also discussed.

Cell-based cardiac pumps and tissue-engineered ventricles

Mortalities resulting from cardiovascular disorders remain high, with an urgent need to develop novel treatment modalities. Tissue-engineering therapies aim to provide cell-based alternatives to conventional options. Significant technological advancements have occurred during the last decade towards the fabrication of functional 3D heart muscle in vitro. More recent research has focused on the development of cell-based cardiac pumps and tissue-engineered ventricles. The global objective of this collective work is to simulate the functional performance of the left ventricle, utilizing completely cell-based options. Current prototypes have shown several physiological performance metrics, including the ability of these devices to generate intraluminal pressure upon electrical stimulation. This review will highlight the transition from tissue engineering 3D heart muscle to cell-based cardiac pumps/ventricles.

Functional evaluation of isolated zebrafish hearts

Traditional working heart preparations, based on the original Langendorff setup, are widely used experimental models that have tremendously advanced the cardiovascular field. However, these systems can be deceivingly complex, requiring the maintenance of pH with CO(2), the delivery of oxygenated perfusate, and the need for extensive laboratory equipment. We have examined the feasibility of using isolated zebrafish (Danio rerio) hearts as an experimental model system, in which experimental procedures can be performed in the absence of the traditional requirements and sophisticated setup equipment. Isolated zebrafish hearts exhibited spontaneous contractile activity, could be electrically paced, and were responsive to pharmacologic stimulation with isoproterenol for 1.5 h after in vivo removal. Isolated zebrafish hearts offer a time- and cost-effective alternative to traditional Langendorff/working heart preparation models, and could be used to investigate cardiac function and repair.

Fabrication of Functional Cardiac, Skeletal, and Smooth Muscle Pumps In Vitro

Cardiovascular disease is one of the leading causes of death in the United States, and new treatments need to be developed in order to provide novel therapies. Tissue engineering aims to develop biologic substitutes that restore tissue function. The purpose of the current study was to construct cell-based pumps, which can be viewed as biologic left ventricular assist devices. The pumps were fabricated by culturing cardiac, skeletal, and smooth muscle cells within a fibrin gel and then each 3-D tissue construct was wrapped around a decellularized rodent aorta. We described the methodology for pump fabrication along with functional performance metric, determined by the intra-luminal pressure. In addition, histologic evaluation showed a concentric organization of components, with the muscle cells positioned on the outermost surface, followed by the fibrin gel and the decellularized aorta formed the innermost layer. Though early in development, cell-based muscle pumps have tremendous potential to be used for basic and applied research, and with further development, can be used clinically as cell-based left ventricular assist devices.

Getting to the Heart of Tissue Engineering

Cardiovascular disease affects 80 million people in the USA and is the leading cause of death. Significant limitations of current treatments necessitate the development of novel strategies. Cardiovascular tissue engineering is an emerging field focused on the development of biological substitutes to restore, maintain, or improve tissue function. In this article, we present an overview of trends in the field and scientific milestones achieved during the last decade. Various 3D bioengineered models of functional cardiovascular structures, including cell-based cardiac pumps, ventricles, patches, vessels, and valves, are described. We discuss critical technological hurdles that must be addressed for continued progress and an outlook for the future of cardiovascular tissue engineering.

Development of a microperfusion system for the culture of bioengineered heart muscle.

Tissue engineering strategies are being used to develop functional 3D heart muscle in vitro. Work within our own group has been focused on the development of bioengineered heart muscle using fibrin gel as a support matrix. As tissue engineering models of heart muscle are developed in the laboratory, a critical technologic challenge remains the ability to delivery nutrients to the entire tissue construct. To address this specific need, we have developed a novel perfusion system for cardiac tissue engineering applications. The system consists of a custom microincubator, designed to house ten 35-mm tissue culture plates on independent platforms for controlled fluid delivery and aspiration. Temperature, pH, and media flow rate and oxygenation are all regulated. In the current study, we describe the compatibility of this microperfusion system with bioengineered heart muscles. We demonstrate that the perfusion system is capable of supporting construct viability (mitochondrial activity, total protein, and total RNA) and maintaining contractile properties (twitch force, specific force, and electrical pacing).