Differential effects of bevacizumab, ranibizumab and aflibercept on cell viability, phagocytosis and mitochondrial bioenergetics of retinal pigment epithelial cell.

PURPOSE: To evaluate the short- and long-term effects of most clinically used anti-vascular endothelial growth factor agents, including bevacizumab, ranibizumab or aflibercept, on cell viability, phagocytosis, mitochondrial bioenergetics and the oxidant acrolein-induced oxidative stress of human adult retinal pigment epithelial (ARPE)-19 cells. METHODS: In cultured ARPE-19 cells, cell viability was measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, phagocytotic activity and intracellular reactive oxygen species (ROS) level were determined by flow cytometry, mitochondrial bioenergetics was assessed using a Seahorse XF24 Extracellular Flux Analyzer, and protein expression was measured by Western blotting. RESULTS: Long-term exposure to all three agents had no effect on cell viability; but rescued the ARPE-19 cells from acrolein-induced decrease in cell viability. Bevacizumab, but not ranibizumab or aflibercept, suppressed the phagocytotic activity of ARPE-19 cells and exerted significantly less protection against acrolein-induced inhibition of phagocytosis. Both ranibizumab and aflibercept increased basal respiratory rate and maximal mitochondrial respiratory capacity after 1-hr exposure; but returned to baseline following 24- or 72-hr exposure. In contrast, both responses were reduced on short-term exposure, but augmented after long-term exposure to bevacizumab. Long-term pretreatment with all three agents reversed acrolein-induced impairment of mitochondrial bioenergetics, overproduction of ROS and phosphorylation of the mitogen-activated protein kinases in ARPE-19 cells. CONCLUSION: Bevacizumab might affect mitochondrial bioenergetics differently from that by ranibizumab and aflibercept. Ranibizumab and aflibercept at their therapeutic dose protect against acrolein-induced oxidative cytotoxicity in human ARPE-19 cells via an increase in mitochondrial bioenergetics. An early protective action on mitochondrial bioenergetic capacity might be used to predict possible long-term antioxidative effects of the agents in the eye.

Resveratrol stimulates mitochondrial bioenergetics to protect retinal pigment epithelial cells from oxidative damage.

PURPOSE: Resveratrol (RSV) alleviates oxidative damage in human adult retinal pigment epithelial (ARPE) cells. Mitochondrial bioenergetics is associated with oxidative stress. The purpose of this study was to examine the role of mitochondrial bioenergetics in the cytoprotective effect of RSV. Its role in protection against the adverse effects of cigarette smoke (CS) in experimental choroidal neovascularization (CNV) was also examined. METHODS: Cultured ARPE-19 cells were treated with acrolein alone or acrolein with added RSV. Temporal changes in cell viability, expression of the antioxidant protein, and mitochondrial bioenergetics were evaluated. In an animal study, CNV lesions were created in Brown Norway rats by laser-induced photocoagulation. Effects of CS alone or with additional RSV treatment on CNV lesions were quantified by fundus fluorescein angiography. RESULTS: In ARPE-19 cells, RSV rescued acrolein-induced cell death, alongside reversal of acrolein-induced superoxide dismutase expression. Resveratrol increased the mitochondrial bioenergetics, including basal respiratory rate, adenosine triphosphate synthesis via oxidative phosphorylation, and maximal mitochondrial capacity. In animal experiments, CS induced a significant increase in CNV following laser injury, and this increase in CNV was appreciably prevented following peripheral infusion of RSV. CONCLUSIONS: Our results indicate that RSV, a major polyphenol found in red wine, exerts protection against acrolein-induced cytotoxicity in human ARPE-19 cells via increases in the mitochondrial bioenergetics. In addition, the antioxidant effect of RSV may contribute to protection against laser-induced CNV in animals exposed to CS. Therefore, RSV might be beneficial for treatment of acrolein-induced or CS-evoked RPE degeneration.

High HbA1c level was the most important factor associated with prevalence of diabetic retinopathy in Taiwanese type II diabetic patients with a fixed duration.

BACKGROUND: To identify the prevalence and related risk factors for diabetic retinopathy (DR) in non-insulin dependent diabetes in Taiwan. METHODS: A retrospective review of type II diabetic patients in the Diabetes Shared Care System database of our Hospital enrolled from 2002 to 2009. A retinopathy severity score was assigned according to fundus examination by indirect ophthalmoscopy or binocular biomicroscopy. RESULTS: Data was collected on 901 subjects, 497 males and 404 females. Of these, 230 (25.53 %) had DR at enrolment. Compared with patients without DR, those with DR were more likely to be female (p = 0.03) or have higher HbA1c (p < 0.001), longer duration of diabetes (p < 0.001), hypertension (p < 0.001), higher systolic blood pressure (p < 0.001), higher diastolic blood pressure (p = 0.05), as well as impaired renal function (p = 0.001). In subgroup analysis stratified by diabetes duration, HbA1c was the most consistent independent risk factor associated to the prevalence of DR. Higher systolic blood pressure and female sex were significantly independent risk factors only in patients with a duration of diabetes < 4 years. On the contrary, old onset age showed a protective effect against DR only in those with a disease duration > 8 years. CONCLUSIONS: High HbA1c level was the most important factor associated with prevalence of DR in Taiwanese type II DM patients with a fixed duration.

Resveratrol protects human retinal pigment epithelial cells from acrolein-induced damage.

PURPOSE: Although the exact pathogenesis of age-related macular degeneration (AMD) is not clear, most studies indicate a role for retinal pigment epithelial (RPE) cell damage and death caused by oxidative stress. The purpose of this study was to examine the potential protective effects of lutein, zeaxanthin, meclofenamic acid, and resveratrol on the acrolein-induced oxidative stress in human RPE cells. METHODS: Cultured human RPE R-50 cells were treated with acrolein at different concentrations and treatment times. The protective effects of lutein (100 microM), zeaxanthin (100 microM), meclofenamic acid (30 microM), and resveratrol (10 microM) were investigated by pretreatment with the above agents before toxicant exposure in acute toxicity models and cotreatment with the toxicant in chronic toxicity models. The synergistic effects of acrolein and hydrogen peroxide exposure were also studied. Fluorescent latex beads were used to assess the phagocytic function of the cells. RESULTS: Acrolein inhibited the phagocytic function of human RPE R-50 cells, and the inhibitory effects were time dependent. Pretreatment with lutein, zeaxanthin, meclofenamic acid, or resveratrol alleviated the inhibition of phagocytosis in the acute acrolein and combined acrolein/hydrogen peroxide toxicity models. Synergistic effects were seen between zeaxanthin and resveratrol or meclofenamic acid. Cotreatment with lutein, zeaxanthin, meclofenamic acid, or resveratrol showed a protective effect against the damage caused by 7-day acrolein exposure followed by hydrogen peroxide treatment. CONCLUSIONS: Our results indicated an inhibitory effect of compounds found in cigarette smoke on human RPE phagocytosis, and lutein, zeaxanthin, meclofenamic acid, and resveratrol each offered protection against this inhibition. Therefore, red wine polyphenol, resveratrol, might ameliorate acrolein-induced or age-related RPE degeneration, such as AMD.