Dopamine 2 receptor activation suppresses VEGF-induced proliferation of primary cultured human retinal microvascular endothelial cells.

Abstract

Purpose : Diabetic retinopathy (DR), a leading cause of blindness, results in part from chronically increased expression of vascular endothelial growth factor (VEGF) in the eye in the diabetic state. VEGF increases proliferation of endothelial cells (ECs), a critical step in abnormal angiogenesis that contributes to the pathophysiology of DR. COX-2 signaling has been shown to be required for VEGF-induced proliferation in certain ECs. Other recent evidence suggests that activation of dopamine receptors, specifically the D2-type receptor (D2R), inhibits VEGF signaling on some types of ECs. Here, we determined whether D2R activation or COX-2 inhibition of human retinal microvascular endothelial cells (HRMECs) suppresses VEGF-induced proliferation in primary cell culture.

Methods : HRMECs obtained from CSC, Inc. (Kirkland, WA), were plated in 10% serum with additional defined growth factors in 96-well plates. After overnight starvation in 0.5% serum, wells were pretreated with the D2R agonist quinpirole (QNP; 50 nM and 10 uM), the COX-2 inhibitor NS-398 (NS; 2 nM in 0.026% DMSO), or the two in combination. Only 0.026% DMSO was added to control wells. After 15 min, VEGF was added. Proliferation as a percent of control was determined using a WST assay after 48 hours. In other experiments, expression of mRNA for the dopamine receptor genes (DRD1-5) was assayed from cultures grown in 10% serum plus defined growth factors to near confluence using a Biomark PCR system (Fluidigm).

Results : VEGF induced proliferation of HRMECs. This proliferation was blocked by either Qnp or NS treatment. Treatment with either Qnp or NS without VEGF had no effect on proliferation. DRD1, 2, 3 and 5, but not DRD4, dopamine receptor gene expression was detected in HRMECs.

Conclusions : The D2R agonist quinpirole and COX-2 inhibition suppressed VEGF-induced HRMEC proliferation to control levels. These results suggest that specific dopamine receptor activation may be a useful adjunct treatment for blocking VEGF-induced EC activation in DR.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.