Does Hyperglycemia Cause Oxidative Stress in the Diabetic Rat Retina?

Mohammad Shamsul Ola

doi:10.3390/cells10040794

Abstract

Diabetes, being a metabolic disease dysregulates a large number of metabolites and factors. However, among those altered metabolites, hyperglycemia is considered as the major factor to cause an increase in oxidative stress that initiates the pathophysiology of retinal damage leading to diabetic retinopathy. Diabetes-induced oxidative stress in the diabetic retina and its damaging effects are well known, but still, the exact source and the mechanism of hyperglycemia-induced reactive oxygen species (ROS) generation especially through mitochondria remains uncertain. In this study, we analyzed precisely the generation of ROS and the antioxidant capacity of enzymes in a real-time situation under ex vivo and in vivo conditions in the control and streptozotocin-induced diabetic rat retinas. We also measured the rate of flux through the citric acid cycle by determining the oxidation of glucose to CO2 and glutamate, under ex vivo conditions in the control and diabetic retinas. Measurements of H2O2 clearance from the ex vivo control and diabetic retinas indicated that activities of mitochondrial antioxidant enzymes are intact in the diabetic retina. Short-term hyperglycemia seems to influence a decrease in ROS generation in the diabetic retina compared to controls, which is also correlated with a decreased oxidation rate of glucose in the diabetic retina. However, an increase in the formation of ROS was observed in the diabetic retinas compared to controls under in vivo conditions. Thus, our results suggest of diabetes/hyperglycemia-induced non-mitochondrial sources may serve as major sources of ROS generation in the diabetic retina as opposed to widely believed hyperglycemia-induced mitochondrial sources of excess ROS. Therefore, hyperglycemia per se may not cause an increase in oxidative stress, especially through mitochondria to damage the retina as in the case of diabetic retinopathy.

Highlights

Diabetes is an endocrinological disorder that dysregulates several metabolic processes and so forth alters the levels of a multitude of metabolites and signaling molecules, either due to lack of insulin or insulin signaling
We employed in vivo techniques to analyze free radical generation in both control and diabetic rats by intravitreal injection of fluorogenic cell-permeant marker CM-H2DCFDA, as the oxidized fluorescent product of the dye gives a measure of intracellular level of reactive oxygen species (ROS) generation in the retina
Our previous metabolic studies in the ex vivo rat retinas, using unique radio-isotopic techniques, indicated a decreased flux of glycolytic and citric acid cycle intermediates in diabetic retinas, which did not support an increase in ROS by mitochondria under hyperglycemic conditions [24]

Summary

Introduction

Diabetes is an endocrinological disorder that dysregulates several metabolic processes and so forth alters the levels of a multitude of metabolites and signaling molecules, either due to lack of insulin or insulin signaling. Many investigators reported that diabetesinduced hyperglycemia stimulates glycolysis and tricarboxylic acid cycle fluxes that increase NADH/NAD+ ratios both in the cytosol and mitochondria of the cells [8,9,16,17,18,19]. This in turn increases electron disposal at the electron transport chain which, thereby produces superoxide radicals by partial reduction of oxygen [20,21]. Proper techniques and physiological conditions are warranted to measure the exact level of oxygen-free radicals in diabetic retinas

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