The suppression mechanisms of quercetin in retinal and choroidal neovascularization through integrin signal pathway

Objective

To investigate the mechanisms of quercetin in retinal angiogenesis via in vitro and in vivo studies.

Methods

Human umbilical vein endothelial cells (HUVECs) was used in in vitro study, and oxygen-induced retinopathy (OIR) and laser-induced choroidal neovascularization (CNV) animal models were used in in vivo study.In the in vitro cell study, normal control group, VEGF-165 treatment group and VEGF-165 combined with quercetin treatment group were used.Cell counting kit-8 (CCK8) and Transwell were used to measure HUVECs cell viability, including proliferation and migration.OIR and CNV animal models were administrated through intraperitoneal injection of quercetin, and the non-perfusion area and CNV aera were evaluated.Western blot assay was used to detect the expression of integrin α5 and integrin β3 expression.In the animal study, 18 breastfeeding mice and 18 infant mice were randomly divided into normal control group, model control group and quercetin treatment group, 6 for each group.The animal feeding and use was in accordance with the standards set by the ARVO, and the experiment was approved by the Ethic Committee for Experimental Animal of Fu Xing Hospital, Capital Medical University (2016-KY-0036).

Results

In the in vitro study, VEGF-165 (20 ng/ml) promoted the proliferation and migration of HUVECs, while quercetin (50 μmol/L) inhibited HUVECs proliferation and migration significantly, comparing to the VEGF-165 treatment group (proliferation: F_groups=18.51, P=0.00; migration: F=85.74, P=0.00). In the in vivo studies, quercetin 20 mg/(kg·day) decreased the non-perfusion area and CNV area comparing to the untreated groups, with significant differences between them (t=6.02, P=0.00; t=5.79, P=0.00). Besides, quercetin down-regulated the expression of integrin α5 and integrin β3 significantly both in the in vitro and in vivo studies.Western blot test showed that integrin α5 and integrin β3 in VEGF-165+ quercetin processing 24 hours group were significantly decreased than those in the VEGF-165 group, with significant differences between them (t=4.46, P<0.05; t=5.18, P<0.01). Compared with the VEGF-165+ quercetin 24 hours group, the integrin α5 and integrin β3 in VEGF-165+ quercetin processing 48 hours group were increased, but they were still significantly increased than those in the VEGF-165 group, with significant differences between them (t=6.54, P<0.05; t=7.17, P<0.01). For the animal studies, quercetin also inhibited the levels of integrin α5 and integrin β3 in OIR and CNV models (t=5.44, 13.52; both at P=0.00).

Conclusions

Quercetin can inhibit the retinal and choroidal neovascularization through integrin pathway, which provides a new treatment strategy for clinical therapy.

Quercetin; Retinal angiogenesis; Choroidal neovascularization; Umbilical vein endothelial cells; Integrin