September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Insulin-like growth factor 1 rescues R28 retinal neurons from apoptotic death through ERK-mediated BimEL phosphorylation independent of Akt
Author Affiliations & Notes
  • Dejuan Kong
    Ophthalmology and Visual Sciences, University of Michigan Kellogg Eye Center, Ann Arbor, Michigan, United States
  • Sumathi Shanmugam
    Ophthalmology and Visual Sciences, University of Michigan Kellogg Eye Center, Ann Arbor, Michigan, United States
  • Lijie Gong
    Ophthalmology and Visual Sciences, University of Michigan Kellogg Eye Center, Ann Arbor, Michigan, United States
  • Patrice E Fort
    Ophthalmology and Visual Sciences, University of Michigan Kellogg Eye Center, Ann Arbor, Michigan, United States
  • Thomas W Gardner
    Ophthalmology and Visual Sciences, University of Michigan Kellogg Eye Center, Ann Arbor, Michigan, United States
  • Steven F Abcouwer
    Ophthalmology and Visual Sciences, University of Michigan Kellogg Eye Center, Ann Arbor, Michigan, United States
  • Footnotes
    Commercial Relationships   Dejuan Kong, None; Sumathi Shanmugam, None; Lijie Gong, None; Patrice Fort, None; Thomas Gardner, None; Steven Abcouwer, None
  • Footnotes
    Support  Supported by NIH R01EY020582 (SFA, TWG), NIH R01EY007739 (SFA) and NIH P30EY007003 (Vision Core)
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 2741. doi:
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      Dejuan Kong, Sumathi Shanmugam, Lijie Gong, Patrice E Fort, Thomas W Gardner, Steven F Abcouwer; Insulin-like growth factor 1 rescues R28 retinal neurons from apoptotic death through ERK-mediated BimEL phosphorylation independent of Akt. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2741.

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      © 2017 Association for Research in Vision and Ophthalmology.

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Abstract

Purpose : IGF-1 is thought to promote neuronal cell survival largely through activation of Akt and deactivation of FoxO proteins. We employed serum deprivation (SD) of R28 cells to explore the mechanisms by which insulin-like growth factor 1 (IGF-1) protects retinal neurons from apoptotic death and to determine the contribution of the Akt/FoxO and MEK/ERK/BimEL pathways.

Methods : R28 cells were induced to undergo apoptosis by SD for up to 48 h. IGF-1 was added at concentrations of 10 to 100 ng/mL. Apoptotic cell death was evaluated by western blotting for caspase-9 and caspase-3 cleavage, caspase-3/7 activity, and lactate dehydrogenase (LDH) activity release. Cell signaling was evaluated by western blotting with phospho-specific antibodies to Akt, GSK3, FoxO1, FoxO3, ERK1/2, and Bim isoforms. FoxO protein translocation was assayed by immunofluorescence and subcellular fractionation. Transfection with siRNA was used to silence expression of Rictor, FoxO1, FoxO3 and Bim.

Results : SD caused a progressive increase in cell death. IGF-1 as low as 10 ng/mL afforded significant protection. IGF-1 caused a rapid activation of Akt, as indicated by phosphorylation of Akt-T308, Akt-S473, PRAS40-T246, and FoxO1/3-T32/T24. This led to prevention of FoxO1/3 nuclear translocation and FoxO-mediated Bim mRNA upregulation in response to SD. IGF-1 also caused MAPK/MEK pathway activation as indicated by ERK1/2-T202/Y204 and Bim-S65 phosphorylation. Surprisingly, inhibition of Akt activation with LY294002 or by Rictor knockdown did not block the protective effect of IGF-1. In contrast, inhibition of MEK activity with PD98059 prevented Bim phosphorylation and blocked IGF-1 protection. In addition, knockdown of Bim expression was protective while co-silencing of FoxO1/3 expression had no effect.

Conclusions : Although IGF-1 may provide long-term neurotrophic support by activation of Akt, inhibition of FoxO activity and prevention of Bim gene transcription, this pathway was not essential for protection from SD-induced apoptosis by IGF-1. Instead, IGF-1 protection was dependent on activation of the MEK/ERK pathway leading to BimEL phosphorylation, which is known to prevent Bax/Bak oligomerization and activation of the intrinsic mitochondrial apoptosis pathway. Targeting this pathway may lead to novel neuroprotective strategies to prevent the progression of sight-threatening diseases.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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