T cell receptor signaling is essential for the generation and maturation of T lymphocyte precursors. Here we identify the deubiquitinating enzyme CYLD as a positive regulator of proximal T cell receptor signaling in thymocytes. CYLD physically interacted with active Lck and promoted recruitment of active Lck to its substrate, Zap70. CYLD also removed both Lys 48- and Lys 63-linked polyubiquitin chains from Lck. Because of a cell-autonomous defect in T cell development, CYLD-deficient mice had substantially fewer mature CD4(+) and CD8(+) single-positive thymocytes and peripheral T cells.

Diabetes pathology derives from the combination of hyperglycemia and hypoinsulinemia or insulin resistance leading to diabetic complications including diabetic neuropathy, nephropathy and retinopathy. Diabetic retinopathy is characterized by numerous retinal defects affecting the vasculature and the neuro-retina, but the relative contributions of the loss of retinal insulin signaling and hyperglycemia have never been directly compared. In this study we tested the hypothesis that increased retinal insulin signaling and glycemic normalization would exert differential effects on retinal cell survival and retinal physiology during diabetes. We have demonstrated in this study that both subconjunctival insulin administration and systemic glycemic reduction using the sodium-glucose linked transporter inhibitor phloridzin affected the regulation of retinal cell survival in diabetic rats. Both treatments partially restored the retinal insulin signaling without increasing plasma insulin levels. Retinal transcriptomic and histological analysis also clearly demonstrated that local administration of insulin and systemic glycemia normalization use different pathways to counteract the effects of diabetes on the retina. While local insulin primarily affected inflammation-associated pathways, systemic glycemic control affected pathways involved in the regulation of cell signaling and metabolism. These results suggest that hyperglycemia induces resistance to growth factor action in the retina and clearly demonstrate that both restoration of glycemic control and retinal insulin signaling can act through different pathways to both normalize diabetes-induced retinal abnormality and prevent vision loss.

Diabetic retinopathy is the leading cause of blindness in working age persons. Targeted studies have uncovered several components of the pathophysiology of the disease without unveiling the basic mechanisms. This study describes the use of complementary proteomic and genomic discovery methods that revealed that the proteins of the crystallin superfamily are increased dramatically in early diabetic retinopathy. Orthogonal methods confirmed that the amplitude of the up-regulation is greater than other changes described so far in diabetic retinopathy. A detailed time course study during diabetes showed differential up-regulation of the different isoforms of the crystallins superfamily. ␣-and ␤-crystallins were regulated primarily at the translation level, whereas ␥-crystallins were also regulated transcriptionally. We also demonstrated cell-specific patterns of expression of the different crystallins in normal and diabetic rat retinas. Diabetic retinopathy is the leading cause of blindness in working age persons, and despite numerous studies, the pathophysiological mechanisms, especially during the early stages of diabetes, remain to be elucidated. Diabetic retinopathy develops, to some degree, in nearly all patients with diabetes and is the most common cause of new cases of blindness among adults. The predominant causes of vision loss are clinically significant macular edema and proliferative diabetic retinopathy, but vision impairment can be prevented or minimized if the retinopathy is identified in its early stages. Diabetic retinopathy includes microvascular and neuronal, glial, and microglial cell defects early in the course of the disease before clinically visible vascular lesions. Photoreceptor and ganglion cell death occurs as early as 2-4 weeks after the onset of diabetes (1-3).Because of the difficulty of studying the mechanisms of the early stages of diabetes in humans, rodent models of type 1 and type 2 diabetes have been developed. Those models have been used to study various specific aspects of early stages of diabetic retinopathy including blood retinal barrier leakage (4), growth factor signaling such as the insulin/insulinlike growth factor receptor and vascular endothelial cell receptor pathways (5, 6), and oxidative stress mechanisms such as reactive oxygen species and advanced glycation end product production (7,8). These targeted studies provide valuable information on different aspects of the pathology and are useful in new therapeutic development, but more detailed discovery research is also needed to understand the full range of metabolic dysregulation that leads to diabetes complications. Proteome profiling using two-dimensional (2D) 1 DIGE and/or isobaric tag for relative and absolute quantitation (iTRAQ) are methods that can characterize new pathophysiological components and potential therapeutic targets.Several studies have used 2D DIGE to gain a better understanding of diabetic complications in the heart and retina (9 -12). These global approaches revealed biochemical changes in the ret...