Dapagliflozin protects neural and vascular dysfunction of the retina in diabetes

Abstract

Introduction: Dapagliflozin, a sodium-glucose transporter inhibitor, effectively reduces blood glucose and is indicated for individuals with kidney diseases and cardiovascular disorders. In this study, we further expand the therapeutic benefit of dapagliflozin in the neural and vascular retina, with the potential to effectively manage diabetic retinopathy (DR), the most common complication of diabetes.

Research design and methods: Db/db mice, an animal model of type 2 diabetes, were treated with dapagliflozin orally, and the electroretinogram (ERG) response and acellular capillary numbers were assessed. Messenger RNA levels of inflammatory cytokines were studied using real-time quantitative (q)PCR. We assessed endothelial cell migration in a scratch wound assay and retinal glucose uptake using human retinal endothelial cells.

Results: The dapagliflozin treatment improved the ERG b-wave amplitude and decreased acellular capillary numbers. The scratch wound assay demonstrated a reduction in wound closure after dapagliflozin treatment. Retinal glucose uptake reduced after dapagliflozin treatment compared with the respective controls.

Conclusions: Our studies suggest that dapagliflozin treatment effectively corrects neural and vascular dysfunction of the retina in diabetes. This effect is mediated by a decrease in inflammation and improved glycemic control. In addition, dapagliflozin exhibits decreased wound healing and glucose uptake, which could benefit the retina. Thus, dapagliflozin could be helpful in the management of DR, with multimodal therapeutic effects.

Keywords: Diabetes Complications; Diabetes Mellitus, Type 2; Diabetic Retinopathy; Retina.

Figure 1

ERG changes in mice treated with dapagliflozin. Line graph showing different parameters of ERG under scotopic conditions: (A) b-wave amplitude, (B) b-wave implicit time, (C) a-wave amplitude, (D) a-wave implicit time, (E) oscillatory potential and photopic conditions, (F) b-wave amplitude, (G) b-wave implicit time, (H) a-wave amplitude, (I) a-wave implicit time, and (J) oscillatory potential. n=8 db/m; n=5 db/db; n=6 db/db+Dapa. Data were analyzed using two-way ANOVA, followed by Fisher’s LSD test. ANOVA, analysis of variance; Dapa, dapagliflozin; ERG, electroretinogram; LSD, least square design.

Figure 2

Decrease in acellular capillary number after dapagliflozin treatment. (A) Representative images of trypsin digested retinas showing changes in acellular capillary numbers (black arrows). (B) Bar chart showing enumeration of acellular capillaries. n=9, db/m; n=8 db/db; n=5 db/db+Dapa. One-way ANOVA, followed by Fisher’s LSD test. *P<0.05, ***P<0.001. ANOVA, analysis of variance; Dapa, dapagliflozin; LSD, least square design.

Figure 3

mRNA expression in the retina. Bar chart showing mRNA levels on (A) TNF-α, (B) IL-1β, (C) adiponectin, and (D) ACE2, after dapagliflozin treatment. n=9, db/m; n=8, db/db; n=5 db/db+Dapa. The mRNA expression was determined in triplicate for each mouse and represented as an average data point for an individual mouse. Raw data were analyzed using linear mixed model EM means, followed by comparison of a particular group with LSD. *P<0.05, **P<0.01, ***P<0.001. Dapa, dapagliflozin; EM, estimated marginal; IL-1β, interleukin 1 beta; LSD, least square design; mRNA, messenger RNA, TNF-α, tumor necrosis factor-alpha.

Figure 4

Wound healing after dapagliflozin treatment. (A) Representative photomicrographs showing scratch wound assay after dapagliflozin treatment and respective alcohol controls. (B) Bar chart showing quantification of per cent wound healing. Data were analyzed using one-way ANOVA, followed by Fisher’s LSD test. *P<0.05, ***P<0.001. n=32, medium only; n=12, control and treatment. ANOVA, analysis of variance; CTRL, vehicle control; Dapa, dapagliflozin; LSD, least square design.

Figure 5

Decrease in glucose uptake after dapagliflozin treatment. Bar chart showing 2-NBDG uptake in HRECs treated with dapagliflozin at different concentrations and respective controls. The results were tested for significance using one-way ANOVA, followed by Fisher’s LSD test. *P<0.05, ***P<0.001. n=14. 2-NBDG, 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose; ANOVA, analysis of variance; CTRL, vehicle control; Dapa, dapagliflozin; HRECs, human retinal endothelial cells; LSD, least square design.