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Bioengineering the neural stem cell niche to identify regulators of fate determination

Final Report Summary - BIOENGINEERED NICHES (Bioengineering the neural stem cell niche to identify regulators of fate determination)

The stem cell mediated regenerative potential of tissues, namely the intrinsic capacity to give rise to new cells upon loss, damage, death or malfunctioning, captures research interests and attention on many different levels. On the one hand, there’s interest in elucidating the mechanisms that have been selected throughout evolution to ensure tissue maintenance, on the other hand, with the increasing understanding of basic mechanisms controlling cell proliferation and differentiation within a tissue, the possibility of exploiting these features in a clinical setting has become more and more appealing and feasible. During embryogenesis and organ development, active proliferation of cells is responsible for increased organ size. This step is then followed by differentiation of cells in specific types that will perform tissue functions. In many somatic tissues, stem cells remain also after development is terminated and these are responsible for tissue turnover during adulthood. Stem cells are by definition able to self-renew and therefore generate more stem cells, and are able to generate, through activation of a differentiation program, specific cell types. While embryonic stem cells, isolated from early stages of embryonic development, are capable of generating all tissue types, stem cells isolated from adult tissues are generally only capable, under physiological conditions, to generate cells of the tissue in which they reside. Some tissues like the skin or the intestinal tracks, require constant cellular turnover, and are characterized by high levels of stem cell proliferation, others, such as the brain or the heart, are quiescent tissues and regenerative activity is limited as well as stem cell number is scarce (Li and Clevers, 2010). In all these tissues, stem cells are neighbored by adjacent cells which provide support and signals to regulate their activity, and are embedded in a three dimensional environment. These cellular and extracellular components surrounding stem cells have been named “niches” (Miller and Gauthier-Fisher, 2009).

While massive amount of research has been done to isolate and ex vivo culture stem cells, little attention has been put in trying to preserve these niche signals, despite the fact that they may be required to maintain stem cell functions (Lutolf et al., 2009). Stem cell isolation is feasible for many different tissues, however in vitro culture paradigms often consist of expansion of cells on plastic dishes, while extracellular signals are provided by addition of growth factors in solution, which very little match with the native environment. In many cases, this leads to rapid loss in culture of stem cell characteristics. Key examples are the culture of hematopoietic stem cells and muscle stem cells. By exploiting bioengineering techniques, we developed a novel cell culture platform to grow stem cells in vitro, by trying to emulate some key niche characteristics. Cells are cultured on soft hydrated gels (poly-ethylen-glycol hydrogels), largely consisting of water. By controlling the chemical composition of the gels, the stiffness of the substrate can be easily modified and the effect on stem cell behavior can be studied. The surface of these gels, contacting the cells, was modified in a way to provide signals: adhesion molecules, cell-cell contact signals and growth factors, that would mimic the native niche signals, from where the cells were extracted. This platform offered furthermore the possibility to systematically address, in a combinatorial fashion, which parameters would be more relevant in terms of achieving better proliferation or better differentiation of cells. Namely, protein concentrations and combinations could be easily modified (Gobaa et al., 2011). We combined this novel culture platform with time-lapse video microscopy, in order to obtain dynamic information on how the cell would respond to the environmental cues we provided. Monitoring cells continuously, instead of taking snap-shots in time of a culture, allows to better understand how the end point has been reached (Schroeder, 2011).