Advances in stem cell biology have afforded promising results for the generation of various cell types for therapies against devastating diseases. However, a prerequisite for realizing the therapeutic potential of stem cells is the development of bioprocesses for the production of stem cell progeny in quantities that satisfy clinical demands. Recent reports on the expansion and directed differentiation of human embryonic stem cells (hESCs) in scalable stirred-suspension bioreactors (SSBs) demonstrated that large-scale production of therapeutically useful hESC progeny is feasible with current state-of-the-art culture technologies. Stem cells have been cultured in SSBs as aggregates, in microcarrier suspension and after encapsulation. The various modes in which SSBs can be employed for the cultivation of hESCs and human induced pluripotent stem cells (hiPSCs) are described. To that end, this is the first account of hiPSC cultivation in a microcarrier stirred-suspension system. Given that cultured stem cells and their differentiated progeny are the actual products used in tissue engineering and cell therapies, the impact of bioreactor's operating conditions on stem cell self-renewal and commitment should be considered. The effects of variables specific to SSB operation on stem cell physiology are discussed. Finally, major challenges are presented which remain to be addressed before the mainstream use of SSBs for the large-scale culture of hESCs and hiPSCs.

Heart diseases are major causes of morbidity and mortality linked to extensive loss of cardiac cells. Embryonic stem cells (ESCs) give rise to cardiomyocyte-like cells, which may be used in heart cell replacement therapies. Most cardiogenic differentiation protocols involve the culture of ESCs as embryoid bodies (EBs). Stirred-suspension bioreactor cultures of ESC aggregates may be employed for scaling up the production of cardiomyocyte progeny but the wide range of EB sizes and the unknown effects of the hydrodynamic environment on differentiating EBs are some of the major challenges in tightly controlling the differentiation outcome. Here, we explored the cardiogenic potential of mouse ESCs (mESCs) and human ESCs (hESCs) encapsulated in poly-L-lysine (pLL)-coated alginate capsules. Liquefaction of the capsule core led to the formation of single ESC aggregates within each bead and their average size depended on the concentration of seeded ESCs. Encapsulated mESCs were directed along cardiomyogenic lineages in dishes under serum-free conditions with the addition of bone morphogenetic protein 4 (BMP4). Human ESCs in pLL-layered liquid core (LC) alginate beads were also differentiated towards heart cells in serum-containing media. Besides the robust cell proliferation, higher fractions of cells expressing cardiac markers were detected in ESCs encapsulated in LC than in solid beads. Furthermore, we demonstrated for the first time that ESCs encapsulated in pLL-layered LC alginate beads can be coaxed towards heart cells in stirred-suspension bioreactors. Encapsulated ESCs yielded higher fractions of Nkx2.5- and GATA4-positive cells in the biore-actor compared to dish cultures. Differentiated cells formed beating foci that responded to chronotropic agents in an organotypic manner. Our findings warrant further development and implementation of microencapsulation technologies in conjunction with bioreactor cultivation to enable the production of stem cell-derived cardiac cells appropriate for clinical therapies and applications.

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Myocardial infarction–induced heart failure is a prevailing cause of death in the United States and most developed countries. The cardiac tissue has extremely limited regenerative potential, and heart transplantation for reconstituting the function of damaged heart is severely hindered mainly due to the scarcity of donor organs. To that end, stem cells with their extensive proliferative capacity and their ability to differentiate toward functional cardiomyocytes may serve as a renewable cellular source for repairing the damaged myocardium. Here, we review recent studies regarding the cardiogenic potential of adult progenitor cells and embryonic stem cells. Although large strides have been made toward the engineering of cardiac tissues using stem cells, important issues remain to be addressed to enable the translation of such technologies to the clinical setting.

Regenerating islet (Reg) proteins are involved in the proliferation and differentiation of diverse cell types. However, whether embryonic stem cells (ESCs) express Reg genes and their corresponding proteins remains unknown. In this study, we probed the expression of Reg family members by mouse ESCs (mESCs). Mouse Reg1 and Reg3γ were detected in undifferentiated stem cells. Furthermore, we tested if gastrin-an inducer of Reg1 expression in committed cells-up-regulates the Reg1 gene in mESCs. Gastrin did not affect the expression of Reg1 either in self-renewing mESCs or under conditions permitting their differentiation. Moreover, overexpression of Reg genes found in various forms of cancer has been linked to dysregulated activation of the canonical Wnt/β-catenin cascade. Given the important roles of Wnt signaling in stem cells, we investigated if activation of Wnt alters the expression of Reg genes in mESCs. Wnt activation led to an increase in Reg1 gene expression with a concomitant increase in the amount of secreted Reg1 protein. Finally, the expression pattern of genes indicative of differentiation was examined in mESCs that were either exposed to soluble Reg1 or overexpressed the Reg1 gene. This is the fi rst account of expression of Reg family members by ESCs. Our results show that the canonical Wnt cascade affects Reg expression and warrants further studies into the potential roles of Reg proteins in stem cell physiology.