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Researches showed that mitochondria and oxidative stress play a crucial role in retinal photochemical injury,but the relationship between the damage of human retinal pigment epithelium (RPE) cell-induced by blue light and light-irradiated time is less studied.
The aim of this study was to research the possible mechanism of RPE oxidative damage induced by blue light in vitro.
Human RPE cells were isolated from healthy donors and cultured.The cells were divided into the normal control group and the light exposure group.The cells of light exposure group were irradiated using the blue light of (4.0±0.5) mW/cm2 for 0.5,1,2,3,4,5,6,12 and 24 hours,respectively,and the cells of the normal control group were cultured in dark environment.Cellular viability was detected by MTT method,and the ultrastructure change of subcellular organelles in RPE cells was examined under the transmission electron microscope(TEM).The content of reactive oxygen species (ROS) was assayed by flow cytometry for the assessment of oxidative stress reaction.The relative expressions of nicotinamide adenine dinucleotide phosphate (NADPH) mRNA and cyclooxygenase 1 (COX1) mRNA in the cells were detected by real-time fluorescence quantitative PCR to evaluate the mitochondria function.
The percentages of cellular viability were (100.00±20.00)%,(95.73±0.89)%,(94.67±2.56)%,(84.23±0.16)%,(78.57±3.09)%,(75.43±2.18)%,(66.13±1.42)%,(53.43±1.91)% and (47.97±1.36)% in the normal control group and light exposure for 1-hour,2-hour,3-hour,4-hour,5-hour,6-hour,12-hour and 24-hour groups,respectively,showing a significant difference among the groups (F=172.270,P=0.000),and the percentages of light exposure for the more than 3 hours groups were significantly lower than those of the normal control group (all at P<0.05).The vacuoles-like degeneration,mitochondrial swelling,decreased microvilli were seen under the TEM.The contents of ROS in RPE cells were (14.75±2.49)%,(19.04±1.02)%,(22.81±3.20)%,(28.75±2.15)%,(33.06±0.96)%,(40.64±2.11)%,(48.25±2.50)% and (60.44±2.68)% in the normal control group and light exposure for 0.5-hour,1-hour,2-hour,3-hour,4-hour,5-hour,6-hour groups,and with significant increases in ROS contents in various light exposure groups compared with the normal control group (all at P<0.05).The relative expression levels of NAPDH mRNA in the cells were gradually elevated 3 hours after light exposure with the increase of time in comparison with the normal control group (all at P<0.05),and the relative expression levels of COX1 mRNA in the cells were higher in the light exposure for 2-hour,3-hour,4-hour and 5-hour group compared with the normal control group (all at P<0.05),and after that the COX1 mRNA levels were gradually declined and were close to the normal level.
Blue light irradiation for more than 3 hours causes oxidative stress damage of mitochondria in RPE in vitro, and the damage was more obvious after irradiation for 5-6 hours.