: Neural stem cells (NSCs) can be propagated in vitro for extensive periods of time while retaining the ability to differentiate into neurons, astrocytes and oligodendrocytes. However, NSCs are limited in their potential to yield specific neuron types. Na?ve NSCs expanded in vitro give rise primarily to GABAergic interneurons and to a lesser extent glutamate neurons. This suggests that NSCs, while multipotent, may not have access to the full spectrum of neuron types. In contrast human embryonic stem cells (hESCs) differentiated towards early neural fates can be readily biased towards various region-specific neuron types such as midbrain dopamine neurons or spinal motoneurons. We have recently reported that hESC derived neural progeny responsive to such regional patterning cues are organized into columnar neuroepithelial structures termed neural rosettes (R- NSCs) and characterized R-NSCs in considerable detail1. Our study has identified R-NSCs as novel, unique NSC stage based on marker expression, clonal stem cell properties, neural differentiation potential, and genetic identity1, 2. The most intriguing finding of these studies was the broad patterning potential of R-NSCs compared to other currently available NSC types. R-NSCs are capable of comprehensive differentiation towards CNS1 and PNS3 derivatives and capable of in vivo engraftment. Despite these exciting preliminary data our studies also revealed major gaps in our current understanding of RNSC biology. Here we would like to address some of these limitations by defining heterogeneity within RNSCs and develop genetic strategies for the prospective isolation of fully patternable R-NSCs based on BAC transgenesis. BAC transgenic hESC reporter lines have been recently pioneered in the Studer lab and will serve as reliable readout of R-NSC stage, identity and function. BAC transgenic reporters will also be critical for probing function of extrinsic and intrinsic factors affecting R-NSC identity. These studies should provide fundamental insights into the genetic and epigenetic mechanisms of neural patterning and ultimately result in novel conditions for the continued in vitro expansion of fully patternable R-NSC - a key step towards establishing a stable expandable universal NSC population.

Public Health Relevance

Defining fate potential in hESC derived neural stem cells Project Narrative This proposal defines the molecular factors affecting the fate potential in human ESC derived neural stem cells. The overarching goal is to establish a truly unlimited source of neural stem cells that retains the full capacity of generating any type of neuron in the central and peripheral nervous system. The availability of such highly potent human ESC derived neural stem cell populations should have a major impact for the development of cell therapies in the nervous system and for generating large number of neural cells suitable for drug screening. Our study builds on a set of novel genetic tools developed in the Studer lab BAC transgenesis to precisely define the molecular factors controlling the potency of human neural stem cells in culture. This study will also address whether neural stem cell potency can be understood by the epigenetic profiles of a given neural stem cell stage. Finally we will manipulate the potency of neural stem cells by applying novel culture conditions and by using viral vectors that inducing gain or loss-of function for specific candidate genes involved in the control of neural stem cell potency. We hope that these studies will ultimately allow us to maintain a universal neural stem cell without any loss of fate potential. Such an ESC- equivalent of the nervous system would retain the potency of the earliest neural plate stage cells in vitro, similar to the in vitro maintenance of inner cell mass potential in ESCs.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Neurogenesis and Cell Fate Study Section (NCF)
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Owens, David F
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Sloan-Kettering Institute for Cancer Research
New York
United States
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Chambers, Stuart M; Mica, Yvonne; Lee, Gabsang et al. (2016) Dual-SMAD Inhibition/WNT Activation-Based Methods to Induce Neural Crest and Derivatives from Human Pluripotent Stem Cells. Methods Mol Biol 1307:329-43
Oricchio, Elisa; Papapetrou, Eirini P; Lafaille, Fabien et al. (2014) A cell engineering strategy to enhance the safety of stem cell therapies. Cell Rep 8:1677-1685
Maroof, Asif M; Keros, Sotirios; Tyson, Jennifer A et al. (2013) Directed differentiation and functional maturation of cortical interneurons from human embryonic stem cells. Cell Stem Cell 12:559-72
Mica, Yvonne; Lee, Gabsang; Chambers, Stuart M et al. (2013) Modeling neural crest induction, melanocyte specification, and disease-related pigmentation defects in hESCs and patient-specific iPSCs. Cell Rep 3:1140-52
Lafaille, Fabien G; Pessach, Itai M; Zhang, Shen-Ying et al. (2012) Impaired intrinsic immunity to HSV-1 in human iPSC-derived TLR3-deficient CNS cells. Nature 491:769-73
Chambers, Stuart M; Qi, Yuchen; Mica, Yvonne et al. (2012) Combined small-molecule inhibition accelerates developmental timing and converts human pluripotent stem cells into nociceptors. Nat Biotechnol 30:715-20
Ming, Guo-Li; Brustle, Oliver; Muotri, Alysson et al. (2011) Cellular reprogramming: recent advances in modeling neurological diseases. J Neurosci 31:16070-5