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C.R. Schlieve, B.L. Nilsson, R.T. Raines, L.A. Levin; Synthesis and Characterization of a Novel Class of Thiol Reducing Agents Which Are Highly Neuroprotective for Retinal Ganglion Cells . Invest. Ophthalmol. Vis. Sci. 2005;46(13):188.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: We previously demonstrated that shifting the retinal ganglion cell redox state towards mild reduction and preventing thiol oxidation is neuroprotective in dissociated mixed retinal cultures in vitro and optic nerve crush in vivo. We synthesized novel modifications of the thiol reductant tris(2–carboxyethyl)phosphine (TCEP) with increased cell permeability and improved chemical stability, and tested their ability to increase RGC survival at varying concentrations. Methods: Retinal ganglion cells of postnatal day 2–4 Long Evans rats were retrogradely labeled with 4',6–diamidino–2–phenylindole (DAPI). At postnatal days 11–13 the animals were sacrificed by decapitation, the retinas enzymatically dissociated, and plated on poly–L–lysine–coated 96–well flat–bottomed tissue culture plates for 3 and 6 days in Neurobasal–A, B27 supplement lacking antioxidants, and one of 3 TCEP compounds (TCEP, TCEP–BX1, and TCEP–BX2) or 3 phosphino compounds. Viable DAPI–positive RGCs were identified by calcein–AM staining. Results: At 72 hours, TCEP–BX1 was effective at rescuing acutely axotomized RGCs at concentrations from 1 nM – 100 µM. RGC survival with 1 nM TCEP–BX1 was 173 ± 12% of control (p=0.002). At 144 hours, TCEP–BX1 rescued RGCs at concentrations from 10 pM – 1 µM. Another compound, TCEP–BX2 rescued RGCs at 10 pM (177 ± 20%; p=0.006) and 10 nM (251 ± 34%; p=0.004) at 3 days. Conclusions: Modified cell permeable thiol reductants are effective neuroprotectants for retinal ganglion cells at picomolar concentrations.
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