Studies on effect of long-term lutein treatment on early diabetic retinopathy using the Ins2Akita/+ mouse model

Abstract

Diabetic retinopathy (DR) is a sight-threatening complication of diabetes. It remains a main cause of visual impairment worldwide. Management of DR remains challenging. Current treatments directly target sight-threatening vascular lesions in the relative late stage of disease. However, increasing evidence showed that prior to the presence of retinal vascular signs, diabetes-induced changes such as inflammation and neurodegeneration already exist in the diabetic retina, thus may lead to irreversible retinal damages. Therefore, novel early interventions with high safety profile are urgently needed. Lutein, a naturally occurring carotenoid with potent antioxidant capacity, is a promising candidate. Studies have shown evident anti-inflammatory and neuroprotective effects of lutein in many eye diseases. The aim of this study is to investigate therapeutic potential of long-term lutein treatment in DR in the Ins2Akita/+ mouse. This mouse model develops stable diabetes and exhibits an early onset of retinal complications mimicking early DR in diabetic patients. The effect of lutein in this model has not been reported. Heterozygous male Ins2Akita/+ and age-matched wild-type mice were used. Hyperglycemic phenotype was confirmed by blood glucose >250 mg/dL. Lutein (4.2 and 8.4 mg/kg/d) was given to animals daily in drinking water from 6 weeks till 4.5, 6.5 or 9 months of age. Microglial reactivity was examined in retinal flat-mounts, by immunolabeling with specific microglial markers. Vascular endothelial growth factor (VEGF) expression in the mouse retina was evaluated via enzyme-linked immunosorbent assay (ELISA). Retinal vascular permeability was assessed in retinal flat-mounts in 9-month-old mice, following perfusion with retro-orbitally injected fluorescein isothiocyanate (FITC)-dextran. Tight junction protein occludin expression level in the mouse retina was evaluated via Western blot. Retinal function was assessed utilizing the electroretinogram (ERG) test. Immunofluorescence staining showed that microglial reactivity in the Ins2Akita/+ mouse retina was increased when compared with wild-type control at 4.5 months of age. With lutein administration, microglial reactivity in the diabetic retina was significantly decreased. The ELISA results showed increased VEGF expression in the Ins2Akita/+ mouse retina at 6.5 and 9 months of age, which was decreased by lutein treatment. Following FITC-dextran perfusion, extravasation of FITC-dextran was detected in the retinas of the Ins2Akita/+ mice. With lutein treatment, fluorescein dye leakage in the diabetic retina was alleviated. Western blot results showed decreases of retinal occludin expression in the Ins2Akita/+ mice. With lutein treatment, the decrease in occludin expression was ameliorated. ERG recordings showed diminished scotopic a- and b-wave amplitudes in the Ins2Akita/+ mice. The functional deficits detected with ERG were significantly ameliorated in the lutein-treated Ins2Akita/+ mice. Taken together, my study demonstrated significant therapeutic efficacy of lutein on diabetes-induced retinal inflammation (indicated by microglial activation) and retinal neurodegeneration (indicated by impaired ERG responses), suggesting that lutein may serve as an inflammation/neurodegeneration targeting intervention therapy for early DR. Moreover, lutein administration showed significant efficacy in alleviating microvascular injuries, suggesting that long-term lutein supplementation may exert beneficial effects on diabetes-induced vasculopathy in DR patients. (476 words)