Characterization of adult neural stem cells from the SVZ after different cultivation time: differentiating role of TGF-beta.

PIANTANIDA, ANA PAULA, MATHIEU, PATRÍCIA AND PITOSSI, FERNANDO

Cordoba, Argentina

Congreso; First Joint Meeting of the Argentine Society for Neurosciences (SAN) and the Argentine Workshop in Neurosciences (TAN); 2009

Parkinson´s disease is a neurodegenerative disorder characterized by a progressive loss of dopaminergic neurons of the substancia nigra pars compacta. In the search of new sources of cells for regenerative therapies, the adult neural stem cells (aNSC) are an alternative because these cells are able to differentiate into neurons, astrocytes and oligodendrocytes in vitro and in vivo. Our group observed that the transforming growth factor beta (TGF-beta) is pro-neurogenic on hippocampus NSC in vivo and in vitro. The objective of our work is to study the use in regenerative and protective therapies of aNSC obtained from one of main areas in the brain where neurons are generated during adulthood, the subventricular layer in the lateral ventricles (SVZ). In order to prove the therapeutic potential of these cells is important the standardization of the isolation protocol and the characterization of the cells in culture.             We performed primary cultures dissecting the SVZ from adult Wistar rats (8-10 weeks old). These cultures were characterized after two, four and six weeks in culture, by Nestin, astrocyte marker (GFAP), and beta III tubulin (Tuj) immunocytochemistry. The effect of TGF-beta on aNSC was analyzed by treatment of cultures with this cytokine for a five days period, after what there were characterized by immunocytochemistry. Preliminary results showed that the percentage of Nestin and Tuj positive cells in the cultures increases from week 2 to 6 (42-65% and 38-60% for Nestin and Tuj markers respectively). In contrast, the fraction of GFAP-positive cells remains relatively constant over 6 weeks (3-4%). On the other hand, the treatment with TGF-beta increases 10 % the Tuj positive cells.. This approach for isolation and culture of aNSC from the SVZ shows that the culture is mainly form by Tuj positive cells. This result indicate that these cells are already committed toward a neural phenotype, so it could become an important source for cell based therapies for neurodegenerative disease. In addition TGF-beta might be a promising tool to increase neural differentiation.