Elsevier

Neuroscience

Volume 131, Issue 3, 2005, Pages 577-587
Neuroscience

Peroxisome Proliferator-Activated Receptors (PPARs) and related transcription factors in differentiating astrocyte cultures

https://doi.org/10.1016/j.neuroscience.2004.11.008Get rights and content

Abstract

Peroxisome proliferator-activated receptors (PPARs), retinoid X receptors (RXRs), CCAAT/enhancer binding proteins (C/EBPs) and β-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 β-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. β-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/EBPα, PPARα, RXRα and glycogen content increased. Between 14 and 21 DIV, PPARβ/δ decreased, while PPARγ, C/EBPβ and δ 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 β-catenin during culture time, strongly suggest the involvement of these transcription factors in astrocyte differentiation. Specifically, β-catenin translocation from the nucleus to plasma membrane, together with PPARβ/δ decrease and C/EBPα increase, could be related to decreased proliferation at confluence, while PPARα and γ and all C/EBPs could participate in differentiation processes, such as glycogenesis and lipidogenesis.

Section snippets

Cell cultures

Primary cultures of cerebral cortical astrocytes were prepared from 1- or 2-day-old CD1 mice (Charles River Italia, Lecco, Italy) according to McCarthy and de Vellis (1980). Experiments were carried out in accordance with the European Communities Council Directive 86/609/EEC. Formal approval for the described experiments was obtained by the Italian Ministry of Health (D. L.vo 116/92) and all efforts were made to minimize the number of animals used and their suffering. Animals were rapidly

Results

The following markers were utilized to study the composition and differentiation of primary astrocyte cultures: nestin, expressed in neuroepithelial stem cells (Lendahl et al., 1990); A2B5, marker of neural precursor cells (Raff et al., 1979); GFAP, marker of terminally differentiated type 2 and type 1 astrocytes (Raff et al., 1984); O4, oligodendrocyte marker (Sommer and Schachner, 1981); β-tubulin III, neuronal marker (Sullivan, 1988).

Discussion

In this paper we studied the possible involvement of PPARs, RXRs and C/EBPs in astrocyte differentiation. To this purpose, the presence and variations of these transcription factors in 3–21 DIV primary cultures from neonatal mouse cerebral cortex were examined and related to the expression of markers of different neural cell lineages.

At the earliest time point, all viable cells were nestin positive and many of them also expressed A2B5; GFAP, O4 and β-tubulin III were absent. This indicates that

Acknowledgments

The authors gratefully thank Dr. Pierpaolo Aimola (Department of Basic and Applied Biology, University of l’Aquila, Italy) for skillful technical assistance. This work was supported by a grant from the University of L’Aquila (PRIA 2001).

References (54)

  • D.J. Mangelsdorf et al.

    The RXR heterodimers and orphan receptors

    Cell

    (1995)
  • S. Moreno et al.

    Immunolocalization of peroxisome proliferator-activated receptors and retinoid X receptors in the adult rat CNS

    Neuroscience

    (2004)
  • S.R. Nahorski et al.

    An enzymatic fluorometric micro-method for determination of glycogen

    Anal Biochem

    (1972)
  • M.C. Raff et al.

    Two glial cell lineages diverge prenatally in rat optic nerve

    Dev Biol

    (1984)
  • M.C. Raff et al.

    Cell-type-specific markers for distinguishing and studying neurons and the major classes of glial cells in culture

    Brain Res

    (1979)
  • I. Sommer et al.

    Monoclonal antibodies (O1 to O4) to oligodendrocyte cell surfacesan immunocytological study in the central nervous system

    Dev Biol

    (1981)
  • O. Sorg et al.

    Characterization of the glycogenolysis elicited by vasoactive intestinal peptide, noradrenaline and adenosine in primary cultures of mouse cerebral cortical astrocytes

    Brain Res

    (1991)
  • P. Thuillier et al.

    Cytosolic and nuclear distribution of PPARgamma2 in differentiating 3T3-L1 preadipocytes

    J Lipid Res

    (1998)
  • N.A.C.S. Wong et al.

    β-Catenin a linchpin in colorectal carcinogenesis?

    Am J Pathol

    (2002)
  • J.W. Woods et al.

    Localization of PPARδ in murine central nervous systemexpression in oligodendrocytes and neurons

    Brain Res

    (2003)
  • S. Yano et al.

    Regulation of CCAAT/Enhancer-binding protein family members by stimulation of glutamate receptors in cultured rat cortical astrocytes

    J Biol Chem

    (1996)
  • I. Allaman et al.

    Glucocorticoids modulate neurotrasmitter-induced glycogen metabolism in cultured cortical astrocytes

    J Neurochem

    (2004)
  • J.-R. Cardinaux et al.

    Vasoactive intestinal peptide, pituitary adenylate cyclase-activating peptide and noradrenaline induce the transcription factors CCAAT/Enhancer binding protein (C/EBP)-β and C/EBPδ in mouse cortical astrocytesinvolvement in cAMP-regulated glycogen metabolism

    J Neurosci

    (1996)
  • L. Cristiano et al.

    Peroxisome proliferator-activated receptors (PPARs) and peroxisomes in rat cortical and cerebellar astrocytes

    J Neurocytol

    (2001)
  • B. Desvergne et al.

    Peroxisome proliferator-activated receptorsnuclear control of metabolism

    Endocr Rev

    (1999)
  • D.D. Diascro et al.

    High fatty acid content in rabbit serum is responsible for the differentiation of osteoblasts into adipocyte-like cells

    J Bone Miner Res

    (1998)
  • F. Doetsch

    The glial identity of neural stem cells

    Nat Neurosci

    (2003)

    • Cited by (0)

    View full text
    Copyright © 2005 IBRO. Published by Elsevier Ltd. All rights reserved.