LA RIPROGRAMMAZIONE DI ASTROCITI UMANI IN CELLULE NEURO-STAMINALI E NEURONI COME POSSIBILE STRUMENTO TERAPEUTICO PER LE PATOLOGIE NEUROLOGICHE
Tesi di Dottorato
Data di Pubblicazione:
2012
Citazione:
LA RIPROGRAMMAZIONE DI ASTROCITI UMANI IN CELLULE NEURO-STAMINALI E NEURONI COME POSSIBILE STRUMENTO TERAPEUTICO PER LE PATOLOGIE NEUROLOGICHE / M. Falcone ; tutore: G. Comi ; coordinatore: P. Corti. Universita' degli Studi di Milano, 2012 Feb 08. 24. ciclo, Anno Accademico 2011. [10.13130/falcone-marianna_phd2012-02-08].
Abstract:
Generating neural stem cells and neurons from reprogrammed human astrocytes
is a potential strategy for repair in neurological diseases.
It has been recently showed that astrocytes from murine cerebral cortex can be
differentiated into neurons by the forced expression of a single transcription factor
(Heinrich et al., 2010). While these studies have evaluated astrocyte conversion in
the murine context, a similar possibility has yet to be demonstrated in human cells.
An essential element for developing such applications with therapeutic value is a
thorough comprehension of the mechanisms that regulate reprogramming of adult
cells into induced pluripotent stem cells (iPSCs) (Hanna et al., 2010) or directly into
another committed lineage, such as fibroblasts converted into neurons and also
specific neuronal subpopulations like dopaminergic neurons (Pang et al., 2011,
Caiazzo et al., 2011).
Here we demonstrate the possibility to obtain progenitors and mature cells of the
neural fate directly from human cortical astrocytes with a dedifferentiation into
neural stem/progenitor phenotype.
Even if for the purpose of autologous cell transplantation in neurological disorders,
fibroblasts from patients resemble a much more suitable source of neurons than
astrocytes from patients, nevertheless, shading light in the mechanisms that make
possible to reprogram astrocytes into NSCs is useful for the final goal of using
these cells as endogenous cell source for in situ neural repair in the CNS without
any invasive cell graft. Human astrocytes can be reprogrammed into iPSCs, with
similar efficiencies to other cells, using the viral expression of four reprogramming
factors (Oct4, Sox2, Klf4, and cMyc) (Riuz et al., 2008). Remarkably,
overexpression of a single factor like OCT4 in adult cells can induce full
reprogramming, as when it is expressed in NSCs (human and mouse) (Kim et al.,
2009 a, b) or promote the formation of another phenotype, such as the generation
of blood cells with its expression in human fibroblasts (Szabo et al., 2010). These
data suggest that the effect of these stem reprogramming factors changes in
relationship to the lineage and the differentiation stage of the cells expressing
them. In the current work, using the individual expression of OCT4, SOX2, or
NANOG, we demonstrated and characterized the direct neural fate conversion of
human astrocytes into multipotent neural progenitors, in vitro and in vivo. These
cells were generated in a manner that is independent of iPSC production.
Individual ectopic expression of the reprogramming factors OCT4 or SOX2 or
NANOG into astrocytes, together with specific cytokine/culture conditions,
activated the neural stem gene program and induced the generation of cells
expressing neural stem/precursors markers. This change of lineage commitment
was obtained also in pure CD44+ mature astrocytes and did not require passing
through a pluripotent state. These unique astrocytes-derived neural stem cells
gave rise to neurons, astrocytes and oligodendrocytes, and showed in vivo
engraftment properties. ASCL1 expression further promotes the acquisition of a
neuronal phenotype in vitro and in vivo. ASCL1 expression further promotes the
acquisition of a neuronal phenotype in vitro and in vivo (Kim et al., 2009). To
develop a broader understanding of astrocytes reprogramming we performed a methylation analysis demonstrating that epigenetic modifications underlie this
process.
These observations indicated that the sites of epigenetic and gene expression
changes during reprogramming of astrocytes to NSCs are tightly linked to genes
that are functionally important for pluripotency. These data demonstrate restoration
of multipotency from human astrocytes, and poi
Tipologia IRIS:
Tesi di dottorato
Elenco autori:
M. Falcone
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