RESUMO
Peroxisome proliferator-activated receptors (PPARs), retinoid X receptors (RXRs), CCAAT/enhancer binding proteins (C/EBPs) and beta-catenin are transcription factors involved in cell differentiation. The aim of this work was to investigate the occurrence and variations of these proteins during astrocyte differentiation. Primary cultures of mouse cortical astrocytes were characterized using nestin, A2B5 and glial fibrillary acidic protein (GFAP) as differentiation markers, during a period of 21 days in vitro (DIV). Glycogen and triglyceride accumulation were also studied. At 3 DIV the cultures were mainly constituted by neural progenitor cells, as assessed by their immunofluorescent pattern. At this time PPARs and beta-catenin were localized to the cytoplasm. Interestingly, some cells contained Oil Red O-positive lipid droplets. Between 7 and 21 DIV, nestin decreased, while GFAP increased, indicating ongoing astroglial differentiation. beta-catenin, predominantly nuclear at 7 DIV, later localized to membranes. Redistribution of all three PPAR isotypes from the cytoplasm to the nucleus was observed starting from 7 DIV. Between 7 and 14 DIV, C/EBPalpha, PPARalpha, RXRalpha and glycogen content increased. Between 14 and 21 DIV, PPARbeta/delta decreased, while PPARgamma, C/EBPbeta and delta and lipid droplet-containing cells increased. At 21 DIV both A2B5-/GFAP+ and A2B5+/GFAP+ cells were predominantly observed, indicating differentiation toward type-1 and type-2 astrocytes, although the presence of GFAP- cells demonstrates the persistence of neural precursors in the culture even at this time point. In conclusion, our results, reporting modifications of PPARs, RXRs, C/EBPs and beta-catenin during culture time, strongly suggest the involvement of these transcription factors in astrocyte differentiation. Specifically, beta-catenin translocation from the nucleus to plasma membrane, together with PPARbeta/delta decrease and C/EBPalpha increase, could be related to decreased proliferation at confluence, while PPARalpha and gamma and all C/EBPs could participate in differentiation processes, such as glycogenesis and lipidogenesis.