| dc.description.abstract | The reprogramming of different specialized cell types into others has been widely studied
in recent years. More specifically, the generation of induced neurons (iNs) from other
differentiated cells is applied to study the molecular mechanisms of neuronal
differentiation, to generate humanized models of neurological and psychiatric diseases,
as well to obtain iNs that can be used in cell therapies for neurodegenerative diseases or
acute lesions in the central nervous system. The first type of non-progenitor cell converted
to iNs through expression of an exogenous gene was astroglia. Then, non-neural cells,
such as fibroblasts and hepatocytes, were also reprogrammed into iNs by genetic
manipulation. More recently, the use of small molecules, capable of interfering with
specific intracellular signaling cascades, has been proposed as an alternative for the
reprogramming of differentiated cells in iNs without direct gene manipulation. However, it
is still unclear what is the best combination of small molecules to reprogram astroglial cells
isolated from the postnatal brain. In this work, we evaluated the possibility of
reprogramming astrocytes in iNs, using a combination of small molecules previously used
to reprogram embryonic fibroblasts. For this, astrocytes isolated from the neocortex and
cerebellum of postnatal mice were cultured and exposed to small molecules for 8-20 days.
After this period, the culture cell phenotype was evaluated by immunocytochemistry, RTqPCR and time-lapse video-microscopy. We evaluated aspects such as the expression of
mRNA and specific proteins of neurons and astrocytes, morphology, cell survival and
proliferation. Our results indicate that only a fraction of the cultured astrocytes acquire
typically neuronal characteristics when exposed to small molecules. In addition, some of
these cells maintain astrocytic properties, indicating an incomplete reprogramming state.
Time-lapse video-microscopy analyzes also indicate that treatment with small molecules
leads to increased cell death, which could contribute to the low conversion rate in iNs
observed. Alternatively, the combination of molecules used may not be the most suitable
to reprogram astrocytes into iNs, indicating the need for different combinations for this
purpose. | |
