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FGF-2 Attenuates Neuronal Apoptosis via FGFR3/PI3k/Akt Signaling Pathway After Subarachnoid Hemorrhage

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Abstract

Neuronal apoptosis is a common and critical pathology following subarachnoid hemorrhage (SAH). We investigated the anti-apoptotic property of fibroblast growth factor (FGF)-2 after SAH in rats. A total of 289 rats underwent endovascular perforation to induce SAH or sham operation. Three dosages (3, 9, or 27 μg) of recombinant FGF-2 (rFGF-2) or vehicle was administered intranasally to rats 30 min after SAH induction. The pan-FGF receptor (FGFR) inhibitor PD173074 or vehicle was administered intracerebroventricularly (i.c.v.) 1 h before modeling, in addition to rFGF-2 treatment. Small interfering ribonucleic acid (siRNA) for FGFR1 and FGFR3 or scrambled siRNA was administered i.c.v. 48 h before SAH induction in addition to rFGF-2 treatment. Anti-FGF-2 neutralizing antibody or normal mouse immunoglobulin G (IgG) was administered i.c.v. 1 h before SAH model. Neurobehavioral tests, SAH severity, brain water content, immunofluorescence, Fluoro-Jade C, TUNEL staining, and western blot were evaluated. The expression of FGF-2, FGFR1, and FGFR3 increased after SAH. FGFR1 and FGFR3 were expressed in the neurons. Nine micrograms of FGF-2 alleviated neurological impairments, brain edema, and neuronal apoptosis following SAH. A rFGF-2 treatment improved motor skill learning and spatial memory and increased the number of surviving neurons postinjury to 28 days after SAH. PD173074 abolished the anti-apoptotic effects of rFGF-2 via suppression of the expression of PI3k, phosphorylated Akt (p-Akt), and Bcl-2 leading to enhancement of the expression of Bax. FGFR3 siRNA worsened neurobehavioral function and suppressed the expression of PI3k, p-Akt, and Bcl-2 rather than FGFR1 siRNA in SAH rats treated with rFGF-2. Anti-FGF-2 neutralizing antibody suppressed the expression of PI3k and p-Akt after SAH. FGF-2 may be a promising therapy to reduce post-SAH neuronal apoptosis via activation of the FGFR3/PI3k/Akt signaling pathway.

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Abbreviations

BWC :

brain water content

CA :

cornu ammonis

CNS :

central nervous system

DG :

dentate gyrus

DMSO :

dimethyl sulfoxide

EBI :

early brain injury

ERK :

extracellular signal-regulated kinase

FGF :

fibroblast growth factor

FGFR :

fibroblast growth factor receptor

FJC :

Fluoro-Jade C

GRB2 :

growth factor receptor-bound protein 2

i.c.v.:

intracerebroventricular

IgG :

immunoglobulin G

IHC :

immunohistochemistry

Nab :

neutralizing antibody

NeuN :

neuron-specific nuclear protein

p-Akt :

phosphorylated Akt

PBS :

phosphate-buffered saline

p-FGFR :

phosphorylated fibroblast growth factor receptor

rFGF-2 :

recombinant fibroblast growth factor-2

rpm :

revolutions per minute

SAH :

subarachnoid hemorrhage

Scr siRNA :

scrambled small interfering ribonucleic acid

siRNA :

small interfering ribonucleic acid

SOS :

son of sevenless

TUNEL :

terminal deoxynucleotidyl transferase dUTP nick end labeling

WB :

western blot

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Acknowledgments

The authors acknowledge the technical assistance provided by Desislava Doycheva, Jerry Flores, Marcin Gamdzyk, Helen Huang, Prativa Sherchan, Orhan Altay, Weilin Xu, Yuchun Zuo, Ganz Zuo, and Jun Peng.

Funding

This article is supported partially by grants from the National Institutes of Health (NS081740 and NS082184) to Dr. Zhang.

Author information

Authors and Affiliations

  1. Department of Physiology and Pharmacology, Loma Linda University, Risley Hall, Room 219, 11041 Campus St, Loma Linda, CA, 92354, USA

    Takeshi Okada, Budbazar Enkhjargal, Zachary D. Travis, Umut Ocak, Jiping Tang & John H. Zhang

  2. Department of Neurosurgery, Mie University Graduate School of Medicine, 2-174 Edobashi, Tsu, Mie, 514-8507, Japan

    Takeshi Okada & Hidenori Suzuki

  3. Department of Anesthesiology, Loma Linda University, Risley Hall, Room 219, 11041 Campus St, Loma Linda, CA, 92354, USA

    John H. Zhang

Authors
  1. Takeshi Okada
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  2. Budbazar Enkhjargal
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  3. Zachary D. Travis
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  4. Umut Ocak
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  5. Jiping Tang
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  6. Hidenori Suzuki
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  7. John H. Zhang
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Contributions

TO, BE, SH, JT, and JHZ were involved in the study design. TO, BE, and JHZ were involved in data interpretation and writing of the manuscript; TO performed the majority of the laboratory work and contributed to the analysis of data; BE was responsible for the animal model; ZDT was involved in manuscript editing. UO was responsible for the rotarod and water maze test; ZDT was involved in manuscript editing.

Corresponding author

Correspondence to John H. Zhang.

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Ethics Approval and Consent to Participate

The Institutional Animal Care and Use Committee (IACUC) of Loma Linda University approved the study protocol, and all experiments were conducted in accordance with the NIH Guidelines for Use of Animals in Neuroscience Research. All authors have read and approved the submitted manuscript.

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The authors declare that they have no competing interests.

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Okada, T., Enkhjargal, B., Travis, Z.D. et al. FGF-2 Attenuates Neuronal Apoptosis via FGFR3/PI3k/Akt Signaling Pathway After Subarachnoid Hemorrhage. Mol Neurobiol 56, 8203–8219 (2019). https://doi.org/10.1007/s12035-019-01668-9

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