Abstract
Granulocyte-colony stimulating factor (G-CSF), a growth factor, has known neuroprotective effects in a variety of experimental brain injury models. Herein we show that G-CSF administration attenuates neuronal apoptosis after neonatal hypoxia-ischemia (HI) via glycogen synthase kinase-3β (GSK-3β) inhibition. Ten day old Sprague-Dawley rat pups (n = 157) were subjected to unilateral carotid artery ligation followed by 2.5. h of hypoxia or sham surgery. HI animals received control siRNA, GSK-3β siRNA (4. μL/pup), G-CSF (50. μg/kg), G-CSF combined with 0.1 or 0.4. nM G-CSF receptor (G-CSFR) siRNA, phosphatidylinositol 3-kinase (PI3K) inhibitor Wortmannin (86. ng/pup), or DMSO (vehicle for Wortmannin). Pups were euthanized 48. h post-HI to quantify brain infarct volume. G-CSFR, activated Akt (p-Akt), activated GSK-3β (p-GSK-3β), Cleaved Caspase-3 (CC3), Bcl-2, and Bax were quantified using Western blot analysis and the localizations of each was visualized via immunofluorescence staining. Neuronal cell death was determined using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL). Our results showed p-GSK-3β increased after HI until its peak at 48. h post-ictus, and both GSK-3β siRNA and G-CSF administration reduced p-GSK-3β expression, as well as infarct volume. p-GSK-3β and CC3 were generally co-localized in neurons. Furthermore, G-CSF increased p-Akt expression and the Bcl-2/Bax ratio and also decreased p-GSK-3β and CC3 expression levels in the ipsilateral hemisphere, which were all reversed by G-CSFR siRNA, Wortmannin, and GSK-3β siRNA. In conclusion, G-CSF attenuated caspase activation and reduced brain injury by inhibiting GSK-3β activity after experimental HI in rat pups. This neuroprotective effect was abolished by both G-CSFR siRNA and Wortmannin.
Original language | English |
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Pages (from-to) | 141-149 |
Number of pages | 9 |
Journal | Experimental Neurology |
Volume | 263 |
DOIs | |
State | Published - Jan 1 2015 |
ASJC Scopus Subject Areas
- Neurology
- Developmental Neuroscience
Keywords
- Apoptosis
- GSK-3β
- Granulocyte-colony stimulating factor (G-CSF)
- Hypoxia-ischemia (HI)
- Neonatal
- PI3K
- Animals, Newborn
- In Situ Nick-End Labeling
- Apoptosis/drug effects
- Rats
- Glycogen Synthase Kinase 3 beta
- Neurons/drug effects
- Neuroprotective Agents/pharmacology
- Glycogen Synthase Kinase 3/metabolism
- Rats, Sprague-Dawley
- Blotting, Western
- Animals
- Transfection
- Fluorescent Antibody Technique
- Hypoxia-Ischemia, Brain/metabolism
- RNA, Small Interfering
- Granulocyte Colony-Stimulating Factor/pharmacology
- Disease Models, Animal