Cardiac tissue engineering: Cell seeding, cultivation parameters, and tissue construct characterization

TLDR

Cardiac tissue engineering seeks to generate functional tissue equivalents for research and repair. The study aimed to determine how cell source, seeding density, seeding vessel, and culture vessel influence the structure and composition of engineered cardiac muscle. The authors employed isolated cells, 3‑D polymer scaffolds, and bioreactors to produce and culture the constructs. High‑density seeding and laminar‑flow bioreactors produced constructs with maintained structural integrity, higher seeding efficiency, reduced cell damage, greater cellularity, and more physiological metabolism, and after 1–2 weeks the constructs expressed cardiac proteins, exhibited lower cellularity yet retained metabolic activity comparable to native tissue.

Abstract

Cardiac tissue engineering has been motivated by the need to create functional tissue equivalents for scientific studies and cardiac tissue repair. We previously demonstrated that contractile cardiac cell-polymer constructs can be cultivated using isolated cells, 3-dimensional scaffolds, and bioreactors. In the present work, we examined the effects of (1) cell source (neonatal rat or embryonic chick), (2) initial cell seeding density, (3) cell seeding vessel, and (4) tissue culture vessel on the structure and composition of engineered cardiac muscle. Constructs seeded under well-mixed conditions with rat heart cells at a high initial density ((6-8) x 10(6) cells/polymer scaffold) maintained structural integrity and contained macroscopic contractile areas (approximately 20 mm(2)). Seeding in rotating vessels (laminar flow) rather than mixed flasks (turbulent flow) resulted in 23% higher seeding efficiency and 20% less cell damage as assessed by medium lactate dehydrogenase levels (p < 0.05). Advantages of culturing constructs under mixed rather than static conditions included the maintenance of metabolic parameters in physiological ranges, 2-4 times higher construct cellularity (p &le 0.0001), more aerobic cell metabolism, and a more physiological, elongated cell shape. Cultivations in rotating bioreactors, in which flow patterns are laminar and dynamic, yielded constructs with a more active, aerobic metabolism as compared to constructs cultured in mixed or static flasks. After 1-2 weeks of cultivation, tissue constructs expressed cardiac specific proteins and ultrastructural features and had approximately 2-6 times lower cellularity (p < 0.05) but similar metabolic activity per unit cell when compared to native cardiac tissue.