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Sami Kamjoo, Vishal R. Sharma, Jing Yang, Patricia Checa, Irene Molina-Martinez, Rocio Herrero-Vanrell, G Astrid Limb, Henry Klassen, M. C. Kenney, Baruch D. Kuppermann; Glial Cell Line-Derived Neurotrophic Factor (GDNF) Enriched Poly Lactic-co-Glycolic Acid (PLGA) Microspheres Suppress Apoptosis In Vitro. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4443.
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© ARVO (1962-2015); The Authors (2016-present)
To study effects of GDNF enriched PLGA microspheres on apoptosis induced by Benzo-e-Pyrene (BeP, a constituent of cigarette smoke) in human ARPE-19 and MIO-M1 retinal cells in vitro.
ARPE-19 and MIO-M1 cells were pre-treated with 0.5mg/ml GDNF microspheres containing 7ng/ml of GDNF for 6 hours then incubated with 400µM BeP for 24 hours. Apoptosis was studied using the JC-1 assay which measures mitochondrial membrane potential (ΔΨm), and also the YO-PRO-1 Apoptosis kit which measures YO-PRO dye fluorescence in apoptotic cells. Controls included 0.5mg/ml blank microspheres and untreated cells.
ARPE-19 cells treated with BeP alone showed reduced ΔΨm levels (1.49±0.08 versus 3.68±0.14 for untreated controls, p<0.0001). GDNF microspheres plus BeP showed increased ΔΨm levels (2.01± 0.04, p=0.002) compared to BeP alone. MIO-M1 cell ΔΨm values were 3.24±0.22 for BeP versus 5.15±0.23 for GDNF-microspheres plus BeP (p=0.001) and 7.71±0.19 for untreated controls (p=0.0001 versus BeP alone). ARPE-19 cell apoptosis expressed as % fluorescence was 908.1±42.5 for BeP versus 98.25±0.47 for untreated controls (p<0.0001), 137.0±57 for GDNF microspheres plus BeP ( p<0.0001 vs BeP alone), and 669.3±20.83 for blank microspheres plus BeP (p<0.002 versus BeP alone). MIO-M1 apoptosis % fluorescence was 248.5±5.6 for BeP versus 98.25±0.47 for untreated controls (p<0.001), 156.5±15 for GDNF microspheres plus BeP (p=0.0016 vs BeP alone), and 181.9±11 for blank microspheres (p<0.001 vs BeP alone).
GDNF microspheres partially protect ARPE-19 and MIO-M1 cells from the induction of apoptosis caused by BeP as measured by both JC-1 and YO-PRO assays. Blank microspheres also showed reduced apoptosis though to a lesser degree than GDNF microspheres. This reduced apoptosis by blank microspheres may be due to sequestering of BeP into the empty microspheres rather than any direct inhibitory effect of the microspheres.
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