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Piera Versura, Carmen Ciavarella, Marina Buzzi, Chiara Coslovi, Elisa Bergantin, Giuseppe Giannaccare, Silvia Bisti, Emilio C Campos; NEUROPROTECTIVE EFFECTS OF CORD BLOOD SERUM (CB-S) IN RAT MULLER CELLS UNDER OXIDATIVE STRESS. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4387.
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© ARVO (1962-2015); The Authors (2016-present)
Cord blood (CB) is rich of neurotrophins such as BDNF (Brain Derived Neurotrophic Factor), NGF (Nerve Growth Factor), GDNF (Glial Derived Neurotrophic Factor), TGF (Transforming Growth Factor)-α, and EGF (Epidermal Growth factor). The aim of this work was to evaluate the protective effect of CB Serum (CB-S), against the hydrogen peroxide (H2O2)-induced damage on rat retinal Muller cells.
Experiments were performed on immortalized rat Retinal Müller Cells (rMC-1, T0576, ABM, Richmond, BC, Canada). Oxidative stress was induced by increasing concentrations of H2O2 (50-500 μM) for 3, 6 and 24 hrs. Standard 5% FBS and 5% CB-S cultured rMC-1 were compared in terms of cell viability (crystal violet staining test), proliferation rate (bromodeoxyuridine –BrdU- incorporation rate), metabolic activity (MTT assay), GFAP (Glial Fibrillary Acidic Protein) expression (Immunofluorescence IF and RT-PCR) and TNF (Tumour Necrosis Factor)–α transcription (RT-PCR) after H2O2 exposure. CB-S was characterized with respect to the BDNF and NGF content by Human Magnetic Luminex Screening Assay. Aliquots used in the experiments contained BDNF 1.5 ng/ml, NGF 3.0 pg/ml, GDNF 1.5 pg/ml, TGF-α 36.3 pg/ml, and EGF 820 pg/ml as a median. Data were expressed as relative to untreated rMC-1 grown with FBS 5%. Each experiment was executed in triplicates and statistical analysis was performed (significance p<0.05).
Exposure to 100 and 200 μM H2O2 for 3 and 6 hrs was set as optimal experimental conditions for testing CB-S effectiveness in protecting rMC-1 from oxidative stress. Crystal violet and MTT assays revealed increased cell survival in CB-S rMC-1 exposed to H2O2 at 100 and 200 μM for 3 and 6 hrs, in comparison to FBS rMC-1 (p always<0.01). BrdU incorporation rate did not endure significant changes. Expression of GFAP and TNF-α transcripts in CB-S rMC-1 after 3- and 6-hour treatment with 100 and 200 uM H2O2 was reduced by the half, along with a decrease of GFAP protein, as shown by IF.
The supplementation of neurotrophins through CB-S protected rMC-1 from oxidative stress by keeping cell viability and metabolic activity. Moreover, CB-S was effective in reducing the expression of GFAP and TNF-α, markers of cell damage and inflammation. The application of CB-S could be promising in neurodegenerative diseases and further studies are necessary to confirm this issue.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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