Abstract
Purpose :
To investigate the changes of RPE cell viability, proliferative potential, and wound healing activity at high glucose condition and evaluate the effects of anti-VEGF antibodies to the RPE cells under high glucose level.
Methods :
ARPE-19 cells were cultured in DMEM/Ham’s F-12 (1:1) at different glucose level (5, 25, and 75 mM). Viability was evaluated in methyl thiazolyl tetrazolium (MTT) assay at 5 days after treatment with each glucose concentration. Migration ability was measured in a wound healing assay at 3 days. Cell Death Detection Kit was used to assess apoptosis at 3 days and 14days. And proliferative potential was assessed by EdU Imaging kit at 3days. Cultured cells were treated with anti-VEGF antibodies at clinically relevant concentrations (bevacizumab 250 µg/mL, ranibizumab 125 µg/mL, aflibercept 500 µg/mL). Then, experiment was repeated at different glucose level. Significance was evaluated with an unpaired, two-tailed Student t test. A P-value < 0.05 was considered to indicate significance.
Results :
The viability and migration of ARPE-19 cells were significantly decreased in high glucose levels. (Viability; 87.6±2.6% at75mM with 5mM set as 100% p<0.05, Migration; 24.5±1.3 % at 5mM, 25.6±1.2 % at25mM, 13.7±1.3% at 75mM, p<0.05). At 14 days, the percentage of TUNEL positive cell was significantly increased in 75mM glucose level compared to 5mM (75mM :152.3±18.8% with 25mM set as 100%). Percentage of EdU positive cell was also significantly decreasd (75mM : 35.8±10.5 % with 5mM set as 100%, p<0.05). In 75mM glucose level, the groups treated with Anti-VEGF, especially showed to decrease of cell viability, proliferation and increase of apoptosis (p<0.05). However, there was no significant difference generally between the anti-VEGF groups.
Conclusions :
High glucose concentration interfered RPE cell viability, wound healing activity, and proliferation. Further, anti-VEGF antibodies aggravated those functions of RPE cells on high glucose level. It may be consider that anti-VEGF long-term use in diabetic retinopathy patients can give a negative effect on RPE function.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.