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Christopher L Passaglia, Mustafa M Siddiq, Yanna Zorina, Sarah Davis, Ehud Kaplan, Robert Blitzer, Ravi Iyengar; A four-drug combination promotes functional axonal regeneration in the rat optic nerve crush model. Invest. Ophthalmol. Vis. Sci. 2018;59(9):2510. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Injured axons in the mammalian optic nerve do not regenerate due to a lack of intrinsic capacity and the inhibitory environment at the injury site. Using a systems pharmacology approach, we identified a four-drug combination that allowed severed neurites of rat cortical neurons to regrow in inhibitory culture media. This study aimed to use the rat optic nerve crush model to test whether the drug combination can spur axon regrowth and perhaps restore visual signal transmission to the brain.
Adult male Sprague-Dawley or Long Evans rats (250–280g, 8-10 weeks) were anesthetized and the right optic nerve was crushed with fine forceps for 10s. Animals then received a 2.5μl intravitreal injection of 1% DMSO (n=9) or a combination of IL-6 [5μg/ml] and HU-210 [300μM] (n=32), and the injury site was covered with gelfoam soaked in APC [4.1mg/ml] and taxol [5μM]. A second intravitreal injection of the drug combination was given 3 days later. Individual controls were performed for all drugs (n=5 each). In 19 experiments, animals were euthanized 21 days after crush and the optic nerves and chiasm were extracted, fixed, and chemically cleared. Regenerating axons were traced by pre-labeling cells prior to euthanasia with a 5μl intravitreal injection of cholera toxin B [1mg/ml] coupled to Alexa-488. In 13 experiments, flash ERGs (fERG), pattern ERGs (pERG), and visual evoked potentials (VEP) were recorded from both eyes 21-28 days after crush. Flashes were delivered by a LED housed in an opaque tube that confined light to the stimulated eye. Patterns were displayed on a computer monitor.
Axon regeneration was not observed in any animal that received no drug treatment or that was treated solely with IL-6 or HU-210. APC and taxol alone promoted some regrowth (<1mm). The four-drug combination produced marked regrowth, reaching the chiasm (>7.5 mm) in over 25% of animals. fERGs from the injured and non-injured eyes were similar for all animals, while pERGs from the injured eye were abnormal for vehicle-control animals and significantly restored for animals that received the four-drug combination. Flashes to the injured eye also elicited a detectable VEP in nearly 25% of four-drug treated animals. The signal was much smaller and slower than for the non-injured eye.
We conclude that the four-drug treatment can promote functional axonal regeneration after optic nerve injury in rats.
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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