Mechanical load influences embryonic ventricular growth, morphogenesis, and function. To date, little is known regarding how the embryonic left ventricular (LV) myocardium acquires a three-dimensional (3D) fiber architecture distribution or how altered mechanical load influences local myofiber architecture. We tested the hypothesis that altered mechanical load changes the maturation process of local 3D fiber architecture of the developing embryonic LV compact myocardium. We measured transmural myofiber angle distribution in the LV compact myocardium in Hamburger-Hamilton stages 21, 27, 31, and 36 chick embryos during normal development or following either left atrial ligation (LAL; LV hypoplasia model) or conotruncal banding (CTB; LV hyperplasia model). The embryonic LV was stained with f-actin and then z-serial optical sectioning was performed using a laser confocal scanning microscope. We reconstructed local 3D myofiber images and computed local transmural myofiber angle distribution. Transmural myofiber angles in compact myocardium (in LV sagittal sections) were oriented in a circumferential direction until stage 27 (Ϫ10 to 10°). Myofibers in the outer side of compact myocardium shifted to a more longitudinal direction by stage 36 (10 to 40°), producing a transmural gradient in myofiber orientation. Developmental changes in transmural myofiber angle distribution were significantly delayed following LAL, while the changes in angle distribution were accelerated following CTB. Results suggest that mechanical load modulates the maturation process of myofiber architecture distribution in the developing LV compact myocardium.

Embryonic myocardium has a high rate of cell proliferation and regulates cellular proliferation, contractile function, and myocardial architecture in response to changes in external mechanical loads. However, the small and complex three-dimensional (3D) structure of the embryonic myocardium limits our ability to directly investigate detailed relationships between mechanical load, contractile function, and cardiomyocyte proliferation. We developed a novel 3D engineered early embryonic cardiac tissue (EEECT) from early embryonic ventricular cells to test the hypothesis that EEECT retains the proliferative and contractile properties of embryonic myocardium. We combined freshly isolated White Leghorn chicken embryonic ventricular cells at Hamburger-Hamilton (HH) stage 31 (day 7 of a 46-stage, 21-day incubation period), collagen type I, and matrix factors to construct cylindrical-shaped EEECTs. We studied tissue architecture, cell proliferation patterns, and contractile function. We then generated engineered fetal cardiac tissue (EFCT) from HH stage 40 (day 14) fetal ventricular cells for direct comparison with EEECT. Tissue architecture was similar in EEECT and EFCT. EEECT maintained high cell proliferation patterns by culture day 12, whereas EFCT decreased cell proliferation rate by culture day 9 (P < 0.05). EEECT increased active contractile force from culture day 7 to day 12. The culture day 12 EEECT contractile response to the beta-adrenergic stimulation was less than culture day 9 EFCT (P < 0.05). Cyclic mechanical stretch stimulation induced myocardial hyperplasia in EEECT. Results indicate that EEECT retains the proliferative and contractile properties of developing embryonic myocardium and shows potential as a robust in vitro model of developing embryonic myocardium.

A number of factors contribute to the control of stem cell fate. In particular, the evidence for how physical forces control the stem cell differentiation program is now accruing. In this review, we discuss the types of physical forces: mechanical forces, cell shape, extracellular matrix geometry/properties, and cell-cell contacts and morphogenic factors, which evidence suggests play a role in influencing stem cell differentiation.