Abstract
Purpose :
Diabetic retinopathy (DR) is a microvascular complication of diabetes mellites and it is the leading cause of blindness among working-age adults in the United States. Proliferative DR (PDR), the advanced stage of DR, is characterized by the aberrant formation of leaky retinal blood vessels that lead to vision loss. The enzyme histone deacetylase-3 (HDAC3) is involved in cellular proliferation and transcriptional repression. HDAC3 is highly expressed in endothelial cells and was found to regulate inflammatory cytokine response in human umbilical vein endothelial cells (HUVEC). Furthermore, HDAC3 act as a negative regulator of HUVEC angiogenesis. This study aimed to investigate the role of HDAC3 in retinal neoangiogenesis.
Methods :
Diabetes was induced in C57BL/6J mice by streptozotocin (STZ)(n=4). Retinas were collected after 8 weeks of diabetes and analyzed for HDAC3 expression by Western blotting. In vitro experiments were conducted using bovine retinal endothelial cells (BRE). BRE (P=4-8) were grown to confluence and subjected to normoxia or oxygen glucose deprivation (OGD, <1% O2) for 6 hours to mimic retinal ischemia in PDR followed by 18 hours of reperfusion (R). Cells were treated with the selective HDAC3 inhibitor, RGFP966 (2, 8 μM), or DMSO for control. Endpoints included cell viability, and assays of migration, and tube formation as important steps in new vessel formation.
Results :
Diabetic mice retinas showed upregulation of HDAC3 as compared to controls (p=0.13, 158.4±26.7% of control, mean±SEM). Treatment with RGFP966 did not affect BRE viability as measured by MTT assay and lactate dehydrogenase (LDH) release into the media. Interestingly, RGFP966 inhibited BRE cell migration under OGD/R (p=0.11, 262.6±97.6% wound remaining vs control for 2 μM concentration, and p=0.33, 618.9±200.3% of wound remaining vs control for 8 μM concentration) and tube formation (p<0.01, 61.54±6.6% of control for 2 μM, and p<0.01, 15.38±4.6% of control for 8 μM concentration).
Conclusions :
HDAC3 could be involved in retinal neoangiogenesis associated with PDR. Here, we have found that inhibiting HDAC3 decreases tube formation in BRE cells. This is in contrast with previous studies which report HDAC3 as a negative regulator of neoangiogenesis in HUVEC cells. Our results warrant further exploration of therapeutic strategies aimed at inhibiting retinal endothelial HDAC3 as a treatment of PDR.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.