Bioreactor systems for cardiovascular tissue engineering

Abstract

A successful cardiovascular tissue engineering construct should consist of functional cardiac cells and cardiac extracellular matrix (ECM) that can provide structural and biochemical support. Embryonic stem cells derived cardiomyocytes (ESC-CMs) represent one promising source; however, differentiated ESC-CMs often possess an immature phenotype compared to adult CMs. Fibroblasts have been commonly used as an alternative cell source for in vitro generation of cardiac ECM, nevertheless, the formation of one of the most important structures in the cardiac ECM, elastic fibers, remains a challenge. Mechanical forces, such as pulsatile shear stress and regular contraction, are crucial for heart development and growth. Therefore, it was hypothesized that the application of physiologically relevant mechanical stimuli to ESC-CMs using bioreactor systems can potentially induce ESC-CM maturation and elastogenesis by fibroblasts. A flow/stretch bioreactor system was designed, validated and utilized to apply pulsatile flow and cyclic strain to murine and human ESC-CMs. The dynamic stimulation of ESC-CMs over extended culture time resulted in an increased expression of cardiac-associated proteins and ion channel-related genes, as well as an improved Ca2+ handling property, compared to static controls. In addition, using Raman spectroscopy, it was shown that dynamically cultured ESC-CMs display a comparable Raman fingerprint to primary isolated CMs. This data highlights the potency of combined pulsatile flow, cyclic strain and extended culture in the maturation process of murine and human ESC-CMs. Subsequently, another custom fluid flow bioreactor system was used to promote elastogenesis in human fibroblasts. Fibroblasts seeded in hybrid electrospun scaffolds showed an increase in the elastin and elastin-associated proteins, followed by elastin fiber deposition after 6-days of culture under pulsatile shear stress conditions, compared to static controls. Furthermore, to visualize the activity of cells in a non-invasive fashion, a flow bioreactor was designed and validated to be compatible with high-resolution imaging systems. This newly designed bioreactor offers significant advantages compared to traditional bioreactor systems, where the culture needs to be sacrificed at regular intervals to analyze samples using invasive methods. The possible applications of the designed bioreactor include in vitro ESC differentiation, ECM synthesis and cell-scaffold interactions.