Extracellular matrix from different tissues is different. Since normal healthy cells create their own optimal environment, it follows that an optimal environment for cell culture can be created from the tissue whence these cells were derived. Surprisingly, no such "cultureware" is currently available to researchers interested in investigating the growth and differentiation of tissue-specific regenerative cell types (so called adult stem cells). While there are generic coatings and 3-D scaffolds, these products are not specific to the various adult mammalian stem and progenitor cells being studied by many investigators, hence they are not optimized for a given system. Several researchers have recently shown that subtle changes to extracellular matrix (ECM) components can have a dramatic effect on growth, differentiation, and tissue formation in culture. To that end, we propose to study tissue-specific ECM coatings that provide cell-substrate interactions, and the effect these interactions may have on growth and differentiation of adult stem and progenitor cells. In so doing, we will provide essential tools for the in vitro growth and differentiation of mammalian cells, specifically stem and progenitor cells, thereby accelerating the pace of development in many areas of regenerative medicine. In this study we will first aim to develop cell culture coatings to demonstrate the feasibility of our approach. We will test three cell types on three related ECM coatings as the substrate of interest: 1) skin-derived stem cells differentiated into keratinocytes (acellular dermis ECM), 2) muscle satellite cells differentiated into skeletal myoblasts (skeletal muscle ECM), and 3) amniotic fluid-derived stem cells differentiated into hepatocytes (liver ECM). This will be accomplished by using our standard protocols for tissue decellularization, by developing new protocols for acellular tissue digestion, and finally by coating these digestates onto tissue culture dishes. Cells will be tested for growth and differentiation potential on their respective substrates, as well as the other substrates, to demonstrate the specificity of the ECM-cell interactions. Characterization will include both quantitative (e.g. flow cytometry) qualitative (e.g. immunocytochemistry), and functional (i.e. metabolic) assays. We hypothesize that the cell-substrate interactions mediated by tissue-specific ECM will facilitate more efficient, complete, and rapid differentiation, thereby resulting in more functional tissue regeneration. These new biomaterials could eventually lead to more rapid development of cell and tissue-based therapies, as well as more efficient industrial scale-up and clinical translation.
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