Neuroprotective effects of brimonidine on retinal ganglion cells induced by oxidative stress mitochondrial dysfunction

Authors: Wang Lu,  Lin Binwu,  Wang Yan,  Pang Long

DOI: 10.3760/cma.j.issn.2095-0160.2018.01.003
Published 2018-01-10
Cite as Chin J Exp Ophthalmol, 2018,36(1): 12-15.

Abstract

Objective

To explore whether brimonidine has a protective effect on retinal ganglion cells (RGCs) through improving mitochondrial function under the oxidative stress.

Methods

Mouse RGC-5 cells were cultured in DMEM medium containing low concentration of glucose (1 g/L), 10% fetal bovine serum and 100 U/ml penicillin-streptomycin solution.The cells were divided into normal control group, H2O2-treated group and brimonidine+ H2O2 group.H2O2 at the concentration of 800 μmol/L was added into the medium in the H2O2-treated group, and 1 μmol/L brimonidine was added into the medium for 2 hours prior to the addition of H2O2 in the brimonidine+ H2O2 group.The cells were sequently cultured for 24 hours.The morphology of the cell nucleus was examined by Hoechst fluorscence staining.The expressions of apoptosis-related protein in the cells were detected by Western blot assay.Mitochondrial membrane potential was assessed by JC-1 staining.

Results

The cell nuclei showed round or oval in shape with consistent size in the normal control group.The pycnosis and karyorrhexis of the cell nuclei were seen in the H2O2-treated group, and less abnormal nuclei were found in the brimonidine+ H2O2 group.The relative expression level of bcl-2 protein in the cells was 0.76±0.15, 0.50±0.13 and 0.75±0.17 in the normal control group, H2O2-treated group and brimonidine+ H2O2 group, respectively, and the expression of bcl-2 protein in the H2O2-treated group was significantly lower than that in the normal control group and brimonidine+ H2O2 group (both at P<0.05). The relative expression level of bax protein in the cells was 0.65±0.13, 0.83±0.07 and 0.70±0.10 in the normal control group, H2O2-treated group and brimonidine+ H2O2 group, respectively, and the expression of bax protein in the H2O2-treated group was significantly higher than that in the normal control group and brimonidine+ H2O2 group (both at P<0.05). A strong orange fluorescence was seen in the mitochondrial membrane of RGC-5 in the normal control group with a co-expression with the green fluorescence of cell membrane.In the H2O2-treated group, the orange fluorescence intensity in the cells was evidently weakened, and the number of JC-1 responsed cells was considerably increased and the orange fluorescence intensity was enhanced in the brimonidine+ H2O2 group.

Conclusions

Brimonidine can prevent RGCs from oxidative-stress damage by improving the mitochondrial function and therefore play a potential neuroprotective effect on optic nerve.

Key words:

Neuroprotective agents/pharmacology; Cell hypoxia/drug effects; Oxidative stress; Retinal ganglion cells; Apoptosis; Mitochondria/pathology; Brimonidine; Cell line

Contributor Information

Wang Lu
Department of Ophthalmology, Traditional Chinese Medical Hospital of Guangdong Province, Guangzhou 510120, China
Lin Binwu
Wang Yan
Pang Long
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Updated: September 4, 2019 — 7:28 am