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Yingqian Li, Kin-Sang Cho, Yanhui Yang, Hetian Lei, Dong Feng Chen; A negative regulator of neurite growth – Insulin-like growth factor binding protein 7. Invest. Ophthalmol. Vis. Sci. 2018;59(9):1861.
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Activation of axonal growth program is a critical step in successful nerve regeneration following injury. As a standard model of CNS neurons, retinal ganglion cells (RGCs) shut down the intrinsic axon growth program during the perinatal period (embryonic day 16 (E16) – postnatal day 0 (P0)) in mice. We found that the expression levels of insulin-like growth factor binding protein (IGFBPs), especially IGFBP7 changed drastically between E16 and P0, covering the time window that RGCs lose their ability to elongate and regenerate axons. This prompts us to further examine the roles of IGFBP7 in neurite growth under in vitro and in vivo conditions.
We first determined the roles of IGFBP7 in mediating neurite outgrowth in vitro in both PC12 cell and primary RGC cultures. Percentage of cell bearing neurite and cell survival and average neurite length were quantified under a masked fashion. IGFBP7 knockdown was then achieved in vitro and in vivo using the technology of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein-9 nuclease (Cas9) and adeno-associated viral gene delivery of IGFBP7 siRNA. In addition, we compared the expression level of IGFBP7 in optic nerve trauma (crush injury) and glaucoma (anterior chamber injection of microbeads) models by real time reverse transcriptase polymerase chain reaction, Western blot analysis, and immunohistochemistry.
We found that the addition of IGFBP7 to PC12 cell and RGC cultures inhibited neurite outgrowth in a dose-dependent manner without affecting cell survival. Average neurite length and percentage of cells bearing neurite were significantly reduced in cultures treated by IGFBP7 as compared to the vehicle control. The level of IGFBP7 in mouse RGCs was increased after optic nerve crush injury and in the glaucoma model, suggesting its involvement in RGC damage. Analysis of outcomes of IGFBP7 knockdown in these disease models is currently ongoing.
Our results suggest that IGFBP7 functions as a negative regulator of RGC axon growth and presents a potentially important new therapeutic target for treating optic nerve disease and injury.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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