Direct conversion of mouse fibroblasts to self-renewing, tripotent neural precursor cells

Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 02/2012; 109(7):2527-32. DOI: 10.1073/pnas.1121003109
Source: PubMed


We recently showed that defined sets of transcription factors are sufficient to convert mouse and human fibroblasts directly into cells resembling functional neurons, referred to as "induced neuronal" (iN) cells. For some applications however, it would be desirable to convert fibroblasts into proliferative neural precursor cells (NPCs) instead of neurons. We hypothesized that NPC-like cells may be induced using the same principal approach used for generating iN cells. Toward this goal, we infected mouse embryonic fibroblasts derived from Sox2-EGFP mice with a set of 11 transcription factors highly expressed in NPCs. Twenty-four days after transgene induction, Sox2-EGFP(+) colonies emerged that expressed NPC-specific genes and differentiated into neuronal and astrocytic cells. Using stepwise elimination, we found that Sox2 and FoxG1 are capable of generating clonal self-renewing, bipotent induced NPCs that gave rise to astrocytes and functional neurons. When we added the Pou and Homeobox domain-containing transcription factor Brn2 to Sox2 and FoxG1, we were able to induce tripotent NPCs that could be differentiated not only into neurons and astrocytes but also into oligodendrocytes. The transcription factors FoxG1 and Brn2 alone also were capable of inducing NPC-like cells; however, these cells generated less mature neurons, although they did produce astrocytes and even oligodendrocytes capable of integration into dysmyelinated Shiverer brain. Our data demonstrate that direct lineage reprogramming using target cell-type-specific transcription factors can be used to induce NPC-like cells that potentially could be used for autologous cell transplantation-based therapies in the brain or spinal cord.

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Available from: Henrik Ahlenius
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    • "For instance, patient-derived iN cells could be used to investigate pathogenetic mechanisms and reveal cellular phenotypes that could be used as proxy for disease expression and as assay for testing therapeutic interventions such as candidate or novel small molecule drugs (Ming et al., 2011). iN cells or other induced neural cell types that are of more proliferative capacity such as induced neural progenitor cells (iNPCs) or induced oligodendrocyte precursor cells (iOPCs) could also be used as cellular grafts with therapeutic intention, such as for Parkinson's disease or myelin diseases (Han et al., 2012; Lujan et al., 2012; Thier et al., 2012; Yang et al., 2013). On the other hand, direct reprogramming could be envisioned for in situ conversion of non-neuronal cells into neurons. "

    Full-text · Article · Oct 2015
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    • "Recently, fibroblasts have been reprogrammed into induced neural stem cells (iNSC) by transcription factors [1] [2] [3] [4] [5], which makes the neural stem cell (NSC) therapy for neurodegenerative disease feasible. However, clinical utilization of patient-specific NSC for the treatment of human diseases remains elusive, mainly due to the risks associated with viral transduction vectors used for induction. "
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    ABSTRACT: Although it is possible to generate neural stem cells (NSC) from somatic cells by reprogramming technologies with transcription factors, clinical utilization of patient-specific NSC for the treatment of human diseases remains elusive. The risk hurdles are associated with viral transduction vectors induced mutagenesis, tumor formation from undifferentiated stem cells, and transcription factors-induced genomic instability. Here we describe a viral vector-free and more efficient method to induce mouse fibroblasts into NSC using small molecules. The small molecule-induced neural stem (SMINS) cells closely resemble NSC in morphology, gene expression patterns, self-renewal, excitability, and multipotency. Furthermore, the SMINS cells are able to differentiate into astrocytes, functional neurons, and oligodendrocytes in vitro and in vivo . Thus, we have established a novel way to efficiently induce neural stem cells (iNSC) from fibroblasts using only small molecules without altering the genome. Such chemical induction removes the risks associated with current techniques such as the use of viral vectors or the induction of oncogenic factors. This technique may, therefore, enable NSC to be utilized in various applications within clinical medicine.
    Full-text · Article · Aug 2015 · International Journal of Stem Cells
    • "Reprogramming adult cells directly into a neural cell identity is now possible. This occurs very rapidly, within 72 hours, and can be used to generate a variety of committed cells or even NSPCs (Vierbuchen et al., 2010; Lujan et al., 2012). These virtues of reprogramming patient-specifi c cells into a desired phenotype, rapidly and without viruses, suggests that the clinical application of cell transplantation strategies in treating SCI may soon advance exponentially. "
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    ABSTRACT: Stem cell transplantation offers an attractive potential therapy for neurological and neurodegenerative disorders such as spinal cord injury (SCI). Given the demand and expectations for the success of regenerative medicines, newly developed cellular therapeutics must be carefully designed and executed, informed by strong preclinical rationale available. This chapter will address the current state of preclinical scientific research for translation into clinical use of stem cell therapy. Focus will be given to current advances in cell transplantation strategies, with specific attention paid to critical assessment of mechanism and efficacy. Specific cell types will be discussed with respect to pathophysiological processes of SCI and their proposed targets. These therapeutics targets include: 1) trophic support and reducing cell loss, 2) remyelination and neuroprotection, 3) tissue modification, and 4) regeneration through neuroplasticity. Combinatorial strategies consisting of co-administering cell types, neurotrophins and tissue modification will also be addressed. Pressing considerations and challenges of translating preclinical findings into clinical therapy exist, and ought to be critically assessed with respect to the best current animal model data. Clinical trials of cell transplantation in human participants with spinal injuries are of significant importance, and will be critically appraised. Taken as a whole—while expectations must be measured—the current status of knowledge on stem cell transplantation for SCI has evolved substantially over the past decade. While translational knowledge gaps continue to exist with regard to cervical and chronic SCI, carefully conducted early phase clinical trials with cellular therapies are warranted, but must be complemented by a robust preclinical research strategy.
    No preview · Chapter · Jan 2015
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