Imunofenotipagem de precursores neurais diferenciados in vitro a partir de célulastronco embrionárias de camundongos

Abstract

Embryonic stem cells has been the focus of studies because they are a promising source for cell therapy, since they proliferate without limits, respond to molecular signals and are easily accessible to genetic manipulation. In order to differentiate these cells into neural precursors and mature neurons, to obtain material for treatment of neurodegenerative diseases like Parkinson's disease, a five-step protocol was described in the literature and has been widely studied. In this study we intend to manipulate the current protocols, controlling the differentiation phenotype in vitro of a new line of embryonic stem cell (CT-4) isolated at Laboratório de Reprodução Humana Prof. Dr. Aroldo Fernando Camargos, Hospital das Clínicas, UFMG by immuno-phenotyping, to obtain a high rate of Tirosine Hidroxilase-positive neurons. The percentage of undifferentiated cells (SSEA-1+) decreased substantially along the stages and at stage 5 less than 2% of total cells were SSEA-1+. The neural precursors (nestin-positive) also decreased gradually in the stages of differentiation (between stages 3 and 5), whereas the expression of panneuronal (PGP) and dopaminergic neurons (TH) markers remained virtually stable, at 20% and 10% respectively. Our protocol has promoted terminal differentiation (neuronal) instead of proliferation of neural precursors, which may have reduced the number of dopaminergic neurons at the end.