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8. BIOREACTOR CULTIVATION 8.1. Hydrodynamic Environment Convective mixing improves the kinetic rate, ef ciency, and spatial uniformity of cell seeding on three-dimensional polymer scaffolds and improves tissue structure and composition by enhancing mass transport within the culture medium and at the tissue surfaces [Vunjak-Novakovic et al., 1998; Radisic et al., 2003]. Hydrodynamic factors present during culture can modulate cell function and tissue development in at least two ways: via associated effects on mass transport between the developing tissue and culture medium (e.g., oxygen, nutrients, growth factors) and by physical stimulation of the cells (e.g., shear, pressure). The composition, morphology, and mechanical properties of engineered tissues grown in hydrodynamically active environments were generally better than in static environments. For cultivations in vessels described in Fig. 6.5, construct compositions and mechanical properties were better in stirred asks and rotating vessels than in static asks. Direct perfusion through cultured tissue constructs stimulated chondrogenesis, presumably because of the combined effects of enhanced mass transport, pH regulation, and uid shear in the cell microenvironment [Sittinger et al., 1994; Dunkelman et al., 1995; Pazzano et al., 2000], in particular at physiological interstitial ow velocities [Maroudas, 1979; Mow et al., 1980, 1991; Vunjak-Novakovic et al., 1996]. However, physiologically thick, functional cartilage could be engineered with diffusional transport within the cultured tissue constructs (e.g., in rotating bioreactors, [Vunjak-Novakovic et al., 1999; Pei et al., 2002a]). In contrast, the same culture conditions yielded engineered cardiac tissue in which the functional layer was only approximately 0.1 mm thick [Carrier et al., 1999]. This nding could be attributed to diffusional limitations of oxygen transport to the cells, as supported by calculations that showed that the pO2 would decrease to zero at a depth of approximately 0.1 mm [Carrier et al., 2002b]. The engineering of a thick layer of functional cardiac tissue requires the perfusion of oxygen-rich culture medium directly through the growing construct [Radisic et al., 2004a]. The combination of rapid cell inoculation and immediate establishment of medium perfusion enabled physiological densities of viable cells in engineered cardiac constructs, because of the maintenance of ef cient oxygen supply to the cells at all times of cultivation. 8.2. Growth Factors Growth factors (e.g., FGF-2, TGF- 1) are generally required to engineer cartilaginous tissues starting from bone marrow-derived mesenchymal stem cells
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