Abstract
The glutamate transporter 1 (GLT1) is upregulated during astrocyte development and maturation in vivo and is vital for astrocyte function. Yet it is expressed at low levels by most cultured astrocytes. We previously showed that maturation of human and mouse stem cell-derived astrocytes – including functional glutamate uptake – could be enhanced by fibroblast growth factor (FGF)1 or FGF2. Here, we examined the specificity and mechanism of action of FGF2 and other FGF family members, as well as neurotrophic and differentiation factors, on mouse embryonic stem cell-derived astrocytes. We found that some FGFs – including FGF2, strongly increased GLT1 expression and enhanced astrocyte proliferation, while others (FGF16 and FGF18) mainly affected maturation. Interestingly, BMP4 increased astrocytic GFAP expression, and BMP4-treated astrocytes failed to promote the survival of motor neurons in vitro. Whole transcriptome analysis showed that FGF2 treatment regulated multiple genes linked to cell division, and that the mRNA encoding GLT1 was one of the most strongly upregulated of all astrocyte canonical markers. Since GLT1 is expressed at reduced levels in many neurodegenerative diseases, activation of this pathway is of potential therapeutic interest. Furthermore, treatment with FGFs provides a robust means for expansion of functionally mature stem cell-derived astrocytes for preclinical investigation.
Highlights
Service, Pathology Department, Hospital and University Center of Porto, Largo Professor Abel Salazar, 4099-001, Porto, Portugal. 7Center for Motor Neuron Biology and Disease, Columbia Stem Cell Initiative, Columbia Translational Neuroscience Initiative, Columbia University, New York, NY, 10032, USA. 8Department of Pathology and Cell Biology, Neurology, and Neuroscience, College of Physicians and Surgeons, Columbia University, New York, NY, 10032, USA. 9Project A.L.S./Jenifer Estess Laboratory for Stem Cell Research, New York, NY, 10032, USA. 10Solomon H
MESC were differentiated into motor neurons (MNs) cultures for 7 days using caudalizing retinoic acid (RA) and ventralizing smoothened agonist (SAG) agents[33]
On day 21, neurospheres, which were devoid of Hb9::GFP MNs and mainly contained neural progenitors, were mechanically dissociated and progenitors were differentiated into astrocytes on adherent surfaces using medium supplemented with 10% fetal bovine serum (FBS) for 2 weeks
Summary
Pathology Department, Hospital and University Center of Porto, Largo Professor Abel Salazar, 4099-001, Porto, Portugal. 7Center for Motor Neuron Biology and Disease, Columbia Stem Cell Initiative, Columbia Translational Neuroscience Initiative, Columbia University, New York, NY, 10032, USA. 8Department of Pathology and Cell Biology, Neurology, and Neuroscience, College of Physicians and Surgeons, Columbia University, New York, NY, 10032, USA. 9Project A.L.S./Jenifer Estess Laboratory for Stem Cell Research, New York, NY, 10032, USA. 10Solomon H. Under pathological stimuli, activated astrocytes, which are characterized by the up-regulation of glial fibrillary acidic protein (GFAP) and altered morphology, can play a key role in detrimental neuro-inflammatory processes[1,6]. Another important function of astrocytes is the uptake of the main excitatory neurotransmitter glutamate[7,8], by high affinity transporters glutamate aspartate transporter (GLAST; known as EAAT1 in human and encoded by the Slc1a3 gene) and glutamate transporter 1 (GLT1; known as EAAT2 in human and encoded by the Slc1a2 gene)[9]. FGFs have various functions and they may differently regulate GLT1
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