Abstract
Purpose :
Proliferative diabetic retinopathy (PDR) is the leading cause of blindness among working-age adults. One of the hallmark characteristics of PDR is increased vascular endothelial growth factor (VEGF). Furthermore, HIF-1a, a heterodimeric transcription factor, is known to mediate cellular responses such as increased VEGF synthesis under hypoxic conditions. Here, we explore the effects of distinct glucose concentrations and hypoxia on VEGF secretion in 661W mouse cone photoreceptor cells and attempt to further understand the mechanisms that lead to PDR.
Methods :
661W cells were cultured in P100 dishes until 80% confluent. Hypoxia was chemically induced using a pre-made cobalt (II) chloride hexahydrate solution. Cells were treated with culture medium containing either normal glucose (NG; 5.5mM), NG with hypoxia, high glucose (HG; 30mM), or HG with hypoxia for 24 hours. ELISA was used to measure VEGF in conditioned media and HIF-1a in cell lysates.
Results :
Total VEGF secreted into the conditioned media 24 hours after the treatment was 14, 29, 15, and 24 pg/mL in NG, NG with hypoxia, HG, and HG with hypoxia treatment groups, respectively. There was a significant between-group difference in VEGF levels secreted into conditioned media (p<0.001). Correspondingly, HIF-1a protein in the cell lysates of these treatment groups was 22, 55, 25 and 51 pg/mL, respectively. Similarly, there was a significant between-group difference in HIF-1a levels in cell lysate (p=0.001).
Conclusions :
Our results indicate that hypoxia is associated with both elevated VEGF secretion and HIF-1a. Thus, it is possible that hypoxia acts upon cone photoreceptor cells via HIF-1a to increase VEGF secretion. Further experiments are needed to show that HIF-1a mediates VEGF output in cone photoreceptor cells and serves as a key factor in the development and progression of PDR.
This is a 2020 ARVO Annual Meeting abstract.