Authors: Z. PAPPALARDO1,2, L. CASSEREAU2, *B. A. ADAMS2, B. DOWNEY2, J. LIM2;
1San Francisco State Univ., San Francisco, CA; 2Xcell Biosci. Inc., San Francisco, CA
Induced pluripotent stem cells (iPSCs) can be used for autologous regenerative medicine to treat conditions such as spinal cord injuries and neuropathology-associated genetic disease. iPSCs can differentiate to neural progenitor cells (NPCs), a multipotent population that can give rise to all lineages of adult neural cells. However, a confounding limitation of neurons derived from iPSC-derived NPCs is that they are not genetically and functionally equivalent to adult neurons in vivo, rendering in vitro systems as sub-optimal surrogates. Furthermore, maturation of iPSC-derived neurons for regenerative medicine may improve clinical efficacy of neural cell transplants. Recent studies highlight the significance of micro-environmental factors such as hypoxia and mechanical force / pressure on stem cell maintenance and directed-differentiation to specific cell lineages, yet none have evaluated the combined contribution of these factors towards differentiation of iPSCs to NPCs and subsequent maturation of differentiated neurons. We demonstrate the biological impact of oxygen and atmospheric pressure on differentiation to NPCs from human iPSCs. Also, we explore how atmospheric pressure-mediated force can act with soluble factors to promote differentiation of NPCs to mature neurons. We used the AVATAR™, which allows tunable control of the microenvironment ex vivo, to allow precise control of physiologically-relevant levels of oxygen and pressure simultaneously. We demonstrate that combinatorial oxygen and pressure are significant drivers of NPC differentiation from iPSCs. We used settings for oxygen concentration (5% vs normoxia) and atmospheric pressure (2 PSI + atmospheric) during re-programming of fibroblasts to iPSCs and identified pressure-dependent increases in genes involved in re-modeling of the extra-cellular matrix and neural differentiation. We further show that in the absence of soluble differentiating factors, oxygen and pressure are sufficient to fully differentiate iPSCs to NPCs expressing PAX6, NES, and SOX2. When evaluating the effects of oxygen/pressure plus soluble factors for neural induction of iPSCs we see an increase in efficiency relative to a standard CO2 incubator workflow. We now are leveraging the differentiation potential of combinatorial oxygen and atmospheric pressure towards maturation of CNS neurons, astrocytes, and motor neurons. Our findings suggest that oxygen and pressure are important drivers of neural differentiation from human iPSCs, and that these factors have the potential to induce maturation of neurons such that they are better suited for translational studies in vitro and in the clinic.
Z. Pappalardo: A. Employment/Salary (full or part-time):; Xcell Biosciences Inc. L. Cassereau: A. Employment/Salary (full or part-time):; Xcell Biosciences Inc. B.A. Adams: A. Employment/Salary (full or part-time):; Xcell Biosciences Inc. E. Ownership Interest (stock, stock options, royalty, receipt of intellectual property rights/patent holder, excluding diversified mutual funds); Xcell Biosciences Inc. B. Downey: A. Employment/Salary (full or part-time):; Xcell Biosciences Inc. J. Lim: A. Employment/Salary (full or part-time):; Xcell Biosciences Inc. E. Ownership Interest (stock, stock options, royalty, receipt of intellectual property rights/patent holder, excluding diversified mutual funds); Xcell Biosciences Inc.
LINK: Society for Neuroscience
EDUC2-08391 CIRM bridges 2.0 to ZP