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Craig Pearson, Keith R Martin, Herbert M Geller; Arylsulfatase B modifies chondroitin sulfation and enhances retinal ganglion cell axon regeneration. Invest. Ophthalmol. Vis. Sci. 2018;59(9):3723.
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© ARVO (1962-2015); The Authors (2016-present)
Strategies to enhance retinal ganglion cell (RGC) regeneration after injury may involve manipulation of factors intrinsic and/or extrinsic to RGC. Our aim was to improve RGC regeneration after optic nerve crush (ONC) in adult mice by combining an intrinsic stimulus (intravitreal zymosan) with the enzyme arylsulfatase B (ARSB), which cleaves axon-inhibiting sulfate groups from chondroitin sulfate proteoglycans (CSPGs) in the extracellular matrix.
CSPG expression was evaluated at 1, 3, 5, 7, 14, and 21 days post crush (dpc) with antibodies to CSPGs, including the 4S sulfation motif, which inhibits neurons in culture; reactive astrocytes (GFAP); and activated microglia (Iba1) (n = 3 per time point). Another group of mice received ONC followed at 3 dpc by intravitreal injection of Zymosan (12.5 mg/mL) and CPT-cAMP (50 µM) and implantation of gelfoam soaked in ARSB (1 mg/mL), chondroitinase ABC (50 U/mL), or control buffer (n = 8 per group). Optic nerves were harvested at 14 dpc and probed with GAP-43. Additional cohorts were harvested at 7 and 28 dpc to determine the therapeutic window.
CSPGs, including 4S-CSPGs, were elevated at the lesion starting at 5 dpc. Elevated levels were sustained at 21 dpc. Zymosan/CPT-cAMP induced significantly more axon regeneration than PBS controls at 14 dpc (282±83.4 vs. 42.3±11.1 axons, p < 0.05). When Zymosan/CPT-cAMP was combined with enzyme treatment, both ARSB and ChABC significantly enhanced RGC axon regeneration compared with buffer control (472±62, 535±123, and 217±53 axons, respectively, p < 0.001). Iba1 staining was higher in ChABC-treated nerves than ARSB. At 7 dpc, a significant increase in the number of axons navigating through the lesion site was already detectable in the ARSB-treated group compared with the buffer control (69.2±12.3 vs. 16.0±8.9, p < 0.05). By 28 dpc, regenerating axons were found extending as far as 4.0 mm beyond the lesion site, to the optic chiasm. ARSB significantly enhanced axon regeneration versus buffer controls (568±96.3 vs. 273±63.0, p < 0.001).
Removing 4S groups from CSPGs with ARSB enhances intrinsically-stimulated regeneration of RGC axons. Treatment can be administered 3 days after injury and exhibits a sustained effect over 4 weeks. Such combined approaches may be worth further investigation as potential treatments for human CNS injuries or degenerative conditions such as glaucoma.
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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