The most striking features of diabetic retinopathy are the vascular abnormalities that are apparent by fundus examination. There is also strong evidence that diabetes causes apoptosis of neural and vascular cells in the retina. Thus, there is good reason to define diabetic retinopathy as a form of chronic neurovascular degeneration. In keeping with the gradual onset of retinopathy in humans, the rate of cell loss in the animal models is insidious, even in uncontrolled diabetes. This is not surprising given that a sustained high rate of cell loss without regeneration would soon lead to catastrophic tissue destruction. The consequences of ongoing cell death are difficult to detect, and even the quantification of cumulative cell loss requires painstaking histology and microscopy. This subtle cell loss raises the issue of the relevance of the phenomenon to the progression of diabetic retinopathy and the ultimate loss of vision. Neuronal function may be compromised in advance of apoptosis, contributing to an early deterioration of vision. Here we review some of the evidence supporting apoptotic cell death as a contributing mechanism of diabetic retinopathy, explore some of the potential causes, and discuss the potential links between apoptosis and loss of visual function in diabetic retinopathy.

OBJECTIVE: Tumor necrosis factor-α (TNF-α) and interleukin-1 beta (IL-1β) are elevated in the vitreous of diabetic patients and in retinas of diabetic rats associated with increased retinal vascular permeability. However, the molecular mechanisms underlying retinal vascular permeability induced by these cytokines are poorly understood. In this study, the effects of IL-1β and TNF-α on retinal endothelial cell permeability were compared and the molecular mechanisms by which TNF-α increases cell permeability were elucidated. RESEARCH DESIGN AND METHODS: Cytokine-induced retinal vascular permeability was measured in bovine retinal endothelial cells (BRECs) and rat retinas. Western blotting, quantitative real-time PCR, and immunocytochemistry were performed to determine tight junction protein expression and localization. RESULTS: IL-1β and TNF-α increased BREC permeability, and TNF-α was more potent. TNF-α decreased the protein and mRNA content of the tight junction proteins ZO-1 and claudin-5 and altered the cellular localization of these tight junction proteins. Dexamethasone prevented TNF-α–induced cell permeability through glucocorticoid receptor transactivation and nuclear factor-kappaB (NF-κB) transrepression. Preventing NF-κB activation with an inhibitor κB kinase (IKK) chemical inhibitor or adenoviral overexpression of inhibitor κB alpha (IκBα) reduced TNF-α–stimulated permeability. Finally, inhibiting protein kinase C zeta (PKCζ) using both a peptide and a novel chemical inhibitor reduced NF-κB activation and completely prevented the alterations in the tight junction complex and cell permeability induced by TNF-α in cell culture and rat retinas. CONCLUSIONS: These results suggest that PKCζ may provide a specific therapeutic target for the prevention of vascular permeability in retinal diseases characterized by elevated TNF-α, including diabetic retinopathy.

Mammalian cells respond to endoplasmic reticulum (ER) stress by attenuation of protein translation mediated through the PERK-eIF2α pathway and transcriptional activation of genes such as Grp78/BiP encoding ER chaperone proteins. The disruption of PERK function or the blocking of eIF2α Ser⁵¹ phosphorylation fails to attenuate translation after ER stress and also results in substantial impairment of Grp78/BiP induction by ER stress. While the activation of the Grp78 promoter by the ATF6 pathway through the endoplasmic reticulum stress elements (ERSEs) is well documented, the molecular mechanism linking PERK activation to Grp78 stress induction is unknown. We report here that ATF4, a transcription factor whose translation is up-regulated by the PERK-eIF2α pathway, can activate the Grp78 promoter independent of the ERSE. The ATF4-activating site is localized to an ATF/CRE sequence upstream of the ERSEs and is distinct from the C/EBP-ATF composite site previously identified as the ATF4 binding site in the ER stress-inducible chop promoter. In vitro translated ATF4 binding to the ATF/CRE site requires other nuclear co-factors from non-stressed cells, forming a complex that exhibits identical electrophoretic mobility as a thapsigargin-stress induced complex. Here we have identified the closely related ATF1 and CREB1 as nuclear co-factors that form in vivo complexes with endogenous ATF4. ER stress induces CREB1 phosphorylation and ATF1/CREB1 binding to the Grp78 promoter. Through the use of adenoviral vector expression systems, we provide evidence that when ATF4 function is suppressed and its binding partners are not able to compensate for its function, Grp78 induction by Tg and Tu is partially inhibited. Our studies resolve a mechanism responsible for inhibition of Grp78 mRNA induction by ER stress in cells that are functionally null for PERK or devoid of eIF2α phosphorylation.

Diabetic retinopathy (DR) impairs vision of patients with type 1 and type 2 diabetes, associated with vascular dysfunction and occlusion, retinal edema, hemorrhage, and inappropriate growth of new blood vessels. The recent success of biologic treatments targeting vascular endothelial growth factor (VEGF) demonstrates that treating the vascular aspects in the later stages of the disease can preserve vision in many patients. It would also be highly desirable to prevent the onset of the disease or arrest its progression at a stage preceding the appearance of overt microvascular pathologies. The progression of DR is not necessarily linear but may follow a series of steps that evolve over the course of multiple years. Abundant data suggest that diabetes affects the entire neurovascular unit of the retina, with an early loss of neurovascular coupling, gradual neurodegeneration, gliosis, and neuroinflammation occurring before observable vascular pathologies. In this article, we consider the pathology of DR from the point of view that diabetes causes measurable dysfunctions in the complex integral network of cell types that produce and maintain human vision.