Successful establishment of a durable endothelial cell (EC) monolayer inside a ventricular blood sac requires homogeneous coverage of the entire luminal surface with attached cells. For this purpose, a new device was developed that slowly rotates a fully assembled cardiac prosthesis with three degrees of freedom. We seeded ECs derived from human adipose tissue at a density of ≈3.5 × 104 cells/cm2 onto the surfaces of polyurethane-made blood sacs and “ersatz” bladders (consisting of T-25 tissue culture flasks). The kinetics of cell attachment, spreading, and proliferation were determined using video microscopy combined with image analysis and cell viability assays. After 60 min of seeding at 5–10 rotations/hr, the plating efficiency inside the blood sacs was 35.7 ± 11%, with cell viability remaining ≈ 90 ± 5%. After 3 hr, when the plating efficiency reached a plateau (≈ 70%), the rotation was stopped and the ECs were allowed to spread and proliferate under static conditions. Within 48 hr, the entire luminal surface was evenly covered by a confluent EC monolayer. Our long-term studies show that with a proper feeding schedule, such an EC monolayer can be maintained intact in vitro for more than 2 weeks.
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