Over 15 million people in the United States have a history of myocardial infarction (heart attack), which causes irreversible tissue damage and accounts for approximately 1000 deaths daily. Improved understanding of cardiac physiology and pathology is crucial for the development of new therapeutic and diagnostic strategies. This thesis was motivated by the need to create a functional substitute for cardiac tissue that could serve as an in vitro model for scientific research and could eventually be used to replace irreversibly damaged cardiac tissue in vivo.

Experimental cardiovascular research in vitro often involves the use of two-dimensional (2-D) cell cultures (monolayers). However, it is known that cardiac myocytes grown in monolayers express a dedifferentiated phenotype. Hence, the main objective of this thesis was to engineer 3-D cardiac tissue that would better mimic the structure and function of native tissue and could serve as a novel in vitro model system for electrophysiological studies of cardiac muscle. The general hypothesis was that myocytes cultivated in a 3-D environment would yield cardiac tissue that, compared to 2-D cultures, was more similar to native tissue. Additionally, it was hypothesized that tissue properties could be modulated by changes in the biochemical and physical regulatory signals in the cell microenvironment. These hypotheses were tested by changing the conditions (e.g. cellular composition, cell culture substrate, environmental factors) and duration of cultivation of engineered tissue, and by quantifying the relevant structural, biochemical, and functional properties on both the cellular and tissue scales. Measured properties were compared with those of native cardiac tissue and conventional 2-D monolayer cultures, and mechanisms for some of the observed differences were explored.

3-D engineered cardiac tissue provided an environment for cell development and interaction that promoted retention of more differentiated cell phenotype compared to 2-D cultures. This environment yielded an in vitro cardiac tissue substitute that exhibited tissue- and cell-scale electrophysiological properties similar to those observed in vivo. Thus, 3-D engineered cardiac tissue could be used as a complementary model system to conventional monolayer cultures of cardiac myocytes to study the development, pharmacology, and physiology of cardiac muscle in vitro.