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Rocio Herrero-Vanrell, Cristina García-Caballero, Blanca Arango-González, Irene Bravo-Osuna, Marius Ueffing; BIOACTIVITY STUDIES OF BIODEGRADABLE GDNF/VitE LOADED MICROSPHERES IN ORGANOTYPIC CULTURES OF NEONATAL RAT RETINA. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4028.
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© ARVO (1962-2015); The Authors (2016-present)
Organotypic culture models of neonatal rat retina preserve three-dimensional interactions among neural cell populations in a controlled environment and are used to explore ex vivo bioactivity of retinal neuroprotectants. The purpose of the present study was to explore this ex vivo tool to assess the effect of neuroprotective active substances released from drug delivery systems intended for retinal degeneration therapy. To this, we have focused on poly (lactic-co-glycolic acid) (PLGA) microspheres (MSs) loaded with Glial-Derived Neurotrophic Factor (GDNF) and different amounts of Vitamin E (VitE).
GDNF/VitE (20 µg GDNF and 20 µL or 40 µL of VitE) loaded MSs were prepared using a S/O/W emulsion solvent extraction-evaporation technique. Microspheres were characterized (morphology, particle size, encapsulation efficiency and in vitro release) and sterilized by gamma radiation (25 kGy) at low (−78oC) temperature. Retinal explants were harvested from 10-days-old Crl:CD(SD) wild type rats and maintained in organotypic culture during 20 days in serum-free conditions. Explants were exposed to homogeneous suspension of microspheres in culture medium (0.5 % w/v) under three different treatments [GDNF/20µLVitE, GDNF/40µLVitE MSs and unloaded MSs] or kept as untreated control (n=3). Immunohistochemistry and transferase dUTP nick end labeling (TUNEL) techniques were used for the morphologically analysis of retinal sections.
Administration of GDNF/VitE-loaded microspheres led to survival of 8.5 ± 0.77 and 8.6 ± 0.63 rows in the ONL for 20µL and 40µL vitE respectively, compared to 6.87 ± 0.83 rows in unloaded particles. TUNEL-positive cells appeared in the ONL of control groups and after exposure to unloaded MSs with higher rates (1.73 ± 0.67 % and 1.32 ± 1.11 % respectively) compared to the values in retinae treated with GDNF/VitE MSs. After 20 days in culture, we still observed a statistically significant decrease of TUNEL positive cells in the GDNF/40µL group (0.33 ± 0.11 %, P < 0.05).
GDNF/VitE-loaded PLGA microspheres increase the survival of photoreceptor cells in long term retinal explants. Organotypic cultures are a suitable tool for the screening of combined neuroprotective factors microencapsulated in PLGA MSs and for research on photoreceptor cell death.
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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