Abstract
BackgroundGlucose is the most important metabolic substrate of the retina and maintenance of normoglycemia is an essential challenge for diabetic patients. Glycemic excursions could lead to cardiovascular disease, nephropathy, neuropathy and retinopathy. A vast body of literature exists on hyperglycemia namely in the field of diabetic retinopathy, but very little is known about the deleterious effect of hypoglycemia. Therefore, we decided to study the role of acute hypoglycemia in mouse retina.Methodology/Principal FindingsTo test effects of hypoglycemia, we performed a 5-hour hyperinsulinemic/hypoglycemic clamp; to exclude an effect of insulin, we made a hyperinsulinemic/euglycemic clamp as control. We then isolated retinas from each group at different time-points after the clamp to analyze cells apoptosis and genes regulation. In parallel, we used 661W photoreceptor cells to confirm in vivo results. We showed herein that hypoglycemia induced retinal cell death in mouse via caspase 3 activation. We then tested the mRNA expression of glutathione transferase omega 1 (Gsto1) and glutathione peroxidase 3 (Gpx3), two genes involved in glutathione (GSH) homeostasis. The expression of both genes was up-regulated by low glucose, leading to a decrease of reduced glutathione (GSH). In vitro experiments confirmed the low-glucose induction of 661W cell death via superoxide production and activation of caspase 3, which was concomitant with a decrease of GSH content. Moreover, decrease of GSH content by inhibition with buthionine sulphoximine (BSO) at high glucose induced apoptosis, while complementation with extracellular glutathione ethyl ester (GSHee) at low glucose restored GSH level and reduced apoptosis.Conclusions/SignificanceWe showed, for the first time, that acute insulin-induced hypoglycemia leads to caspase 3-dependant retinal cell death with a predominant role of GSH content.
Highlights
Neural tissue, including retina, is totally dependent on glucose for normal metabolic activity
We quantified apoptotic cells from three flat-mount retinas isolated from each group and obtained 312656 Terminal dUTP Nick End-Labeling (TUNEL)-positive cells after acute hypoglycemia (Fig. 2B) while few positive cells (1566 for Eugly and 664 for Ctl) were detected in the two control groups
The majority of TUNEL positive cells were localized in the outer nuclear layer (ONL) and ganglion cell layers (GCL), as observed by confocal microscopy
Summary
Neural tissue, including retina, is totally dependent on glucose for normal metabolic activity. Since the level of glucose storage is negligible compared with the eye glucose demand, this tissue is dependent on glucose delivery by circulating blood. In both type I and II diabetes, normalization of blood glucose concentration is an important issue to avoid secondary long-term microvascular complications, including nephropathy, cardiovascular disease, neuropathy and retinopathy [1]. The role of hyperglycemia in the retina via pericyte apoptosis and in vascular complications has been extensively studied in a large number of in vivo and/or ex vivo models [5,6]. We decided to study the role of acute hypoglycemia in mouse retina
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