Medium perfusion enables engineering of compact and contractile cardiac tissue

American Journal of Physiology - Heart and Circulatory Physiology - Tập 286 Số 2 - Trang H507-H516 - 2004
Milica Radisic1, Liming Yang2, Jan Boublik2, Richard J. Cohen2, Róbert Langer2, Lisa E. Freed2, Gordana Vunjak‐Novakovic2
1Dept. of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, 02139, USA.
2Department of Chemical Engineering and Harvard-Massachusetts Institute of Technology Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139

Tóm tắt

We hypothesized that functional constructs with physiological cell densities can be engineered in vitro by mimicking convective-diffusive oxygen transport normally present in vivo. To test this hypothesis, we designed an in vitro culture system that maintains efficient oxygen supply to the cells at all times during cell seeding and construct cultivation and characterized in detail construct metabolism, structure, and function. Neonatal rat cardiomyocytes suspended in Matrigel were cultured on collagen sponges at a high initial density (1.35 × 108cells/cm3) for 7 days with interstitial flow of medium; constructs cultured in orbitally mixed dishes, neonatal rat ventricles, and freshly isolated cardiomyocytes served as controls. Constructs were assessed at timed intervals with respect to cell number, distribution, viability, metabolic activity, cell cycle, presence of contractile proteins (sarcomeric α-actin, troponin I, and tropomyosin), and contractile function in response to electrical stimulation [excitation threshold (ET), maximum capture rate (MCR), response to a gap junctional blocker]. Interstitial flow of culture medium through the central 5-mm-diameter × 1.5-mm-thick region resulted in a physiological density of viable and differentiated, aerobically metabolizing cells, whereas dish culture resulted in constructs with only a 100- to 200-μm-thick surface layer containing viable and differentiated but anaerobically metabolizing cells around an acellular interior. Perfusion resulted in significantly higher numbers of live cells, higher cell viability, and significantly more cells in the S phase compared with dish-grown constructs. In response to electrical stimulation, perfused constructs contracted synchronously, had lower ETs, and recovered their baseline function levels of ET and MCR after treatment with a gap junctional blocker; dish-grown constructs exhibited arrhythmic contractile patterns and failed to recover their baseline MCR levels.

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American Journal of Physiology - Heart and Circulatory Physiology

Tập 286 Số 2

H507-H516

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