Activated Notch1 is a stronger astrocytic stimulus than leukemia inhibitory factor for rat neural stem cells.
Neural stem cells (NSC) self-renew and generate specialized cell types. There are reports indicating that Notch and Leukemia Inhibitory Factor (LIF) signaling are involved in cell determination of NSC, either preventing differentiation or promoting astrocytic fate. In this work, we aimed to compare the astrocytogenic effect of activated Notch with that induced by LIF. To this end, rat cerebral cortex neural progenitors/NSC were transduced with retroviral vectors in order to express green fluorescent protein (GFP), or a fusion protein of GFP with the active Notch1 intracellular domain (NICD). In parallel, other cultures were treated with increasing concentrations of LIF. We confirmed, in proliferating NSC, that LIF activated intracellular effectors by measuring STAT3 phosphorylation and Socs3 transcription. In NICD-expressing cells, Hes5 mRNA was induced, an effect not present in GFP-transduced NSC. We quantified the proportion of cells expressing Nestin in the presence of Fibroblast Growth Factor-2 (FGF-2) with LIF or NICD treatments. LIF significantly increased the proportion of cells co-expresssing Nestin and Glial Fibrillary Acidic Protein (GFAP), an effect absent in cells with activated Notch. After FGF2 withdrawal to promote differentiation, Nestin was markedly down-regulated, and neuronal and glial markers appeared in control cultures. LIF treatment caused a significant increase in the proportion of GFAP-positive cells, but cells expressing NICD showed a significantly higher percentage of astrocytes than control and LIF-treated cultures. These experiments show that cells stimulated with NICD differentiate more readily to astrocytes than LIF-treated NSC.
Departamento de Neurociencias, Instituto de Fisiologia Celular, Universidad Nacional Autonoma de Mexico, Mexico.
Nidia S
NS
Rodríguez-Rivera
Anayansi
A
Molina-Hernández
Erika
E
Sánchez-Cruz
Diana
D
Escalante-Alcalde
Iván
I
Velasco
D000818:Animals; D001253:Astrocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation; D002478:Cells, Cultured; D020934:Ciliary Neurotrophic Factor / Q000494:pharmacology; D017931:DNA Primers / Q000235:genetics; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007381:Intermediate Filament Proteins / Q000378:metabolism; D053605:Leukemia Inhibitory Factor / Q000494:pharmacology*; D053680:Leukemia Inhibitory Factor Receptor alpha Subunit / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D064231:Nestin; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D020801:Receptor, Ciliary Neurotrophic Factor / Q000378:metabolism; D051881:Receptor, Notch1 / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism; D014161:Transduction, Genetic
D051792:Basic Helix-Loop-Helix Proteins; D020934:Ciliary Neurotrophic Factor; D017931:DNA Primers; D005904:Glial Fibrillary Acidic Protein; D049452:Green Fluorescent Proteins; D007381:Intermediate Filament Proteins; D053605:Leukemia Inhibitory Factor; D053680:Leukemia Inhibitory Factor Receptor alpha Subunit; D009419:Nerve Tissue Proteins; D064231:Nestin; D020801:Receptor, Ciliary Neurotrophic Factor; D051881:Receptor, Notch1; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C489686:Hes5 protein, rat; C507842:Lifr protein, rat; C577486:Nes protein, rat; C497763:Notch1 protein, rat