Patrolling Monocytes Are Recruited and Activated by Diabetes to Protect Retinal Microvessels.

In diabetes there is a long latency between the onset of hyperglycemia and the appearance of structural microangiopathy. Because Ly6Clow patrolling monocytes (PMo) behave as housekeepers of the vasculature, we tested whether PMo protect microvessels against diabetes. We found that in wild-type mice, diabetes reduced PMo in the general circulation but increased by fourfold the absolute number of PMo adherent to retinal vessels (leukostasis). Conversely, in diabetic NR4A1-/- mice, a model of absence of PMo, there was no increase in leukostasis, and at 6 months of diabetes, the number of retinal acellular capillaries almost doubled compared with diabetic wild-type mice. Circulating PMo showed gene expression changes indicative of enhanced migratory, vasculoprotective, and housekeeping activities, as well as profound suppression of genes related to inflammation and apoptosis. Promigratory CXCR4 was no longer upregulated at longer duration when retinal acellular capillaries begin to increase. Thus, after a short diabetes duration, PMo are the cells preferentially recruited to the retinal vessels and protect vessels from diabetic damage. These observations support the need for reinterpretation of the functional meaning of leukostasis in diabetes and document within the natural history of diabetic retinopathy processes of protection and repair that can provide novel paradigms for prevention.

Endothelial progenitor cells and response to ranibizumab in age-related macular degeneration.

BACKGROUND: Choroidal neovascularization (CNV) is the main cause of vision loss in age-related macular degeneration (AMD). In experimental CNV, endothelial progenitor cells (EPCs) contribute to the formation of new vessels. The aim of this study was to investigate whether the behavior of EPCs in patients with AMD supports a role for EPCs in human CNV. METHODS: The number of circulating EPCs that are considered pure endothelial precursors and EPCs with monocytic characteristics, and the plasma levels of regulatory cytokines were evaluated in 23 patients with AMD with active CNV and 20 matched controls. In the patients, this profile was re-evaluated after ranibizumab. RESULTS: When compared with controls, the patients with AMD showed a lower number of both EPC types (P = 0.03) and higher plasma levels (P = 0.03) of stromal cell-derived factor 1. Three monthly injections of ranibizumab returned to control levels the number of circulating EPCs considered pure endothelial precursors and of stromal cell-derived factor 1, but not of monocytic EPCs. CONCLUSION: The observations indicate responsiveness of circulating EPCs to the CNV process in AMD. They suggest the hypothesis that increased stromal cell-derived factor 1 production at the CNV site (reflected in higher plasma levels) recruits EPCs from the circulation, and that antivascular endothelial growth factor therapy selectively decreases the recruitment of cells to be incorporated into new vessels.

Macular micropseudocysts in early stages of diabetic retinopathy.

PURPOSE: To identify by noninvasive means early retinal abnormalities that may predict diabetic macular edema. METHODS: The authors analyzed retrospectively data from consecutive patients with Type 1 (n = 16) or Type 2 (n = 23) diabetes who presented for routine follow-up of early retinopathy, had no clinical signs or symptoms of diabetic macular edema, and were evaluated with spectral-domain optical coherence tomography. Age- and gender-matched nondiabetic subjects provided normative data. RESULTS: Spectral-domain optical coherence tomography revealed in the macular region of diabetic patients small hyporeflective areas (median diameter, 55 µm) contained within discrete retinal layers that we named micropseudocysts (MPCs). Micropseudocysts are associated with vascular leakage. The patients showing MPCs had more frequently systemic hypertension and increased central foveal thickness than those without MPCs. The association with increased central foveal thickness was only in the patients with Type 2 diabetes. CONCLUSION: Macular MPCs in patients with mild diabetic retinopathy appear to reflect leakage and can precede macular thickening. The association of MPCs with increased central foveal thickness in patients with Type 2 diabetes, but not in patients with Type 1 diabetes, points to a greater tendency to retinal fluid accumulation in patients with Type 2 diabetes. Studies in larger cohorts will determine the usefulness of MPCs in strategies to abort diabetic macular edema.

Early complement activation and decreased levels of glycosylphosphatidylinositol-anchored complement inhibitors in human and experimental diabetic retinopathy.

Diabetic retinal microangiopathy is characterized by increased permeability, leukostasis, microthrombosis, and apoptosis of capillary cells, all of which could be caused or compounded by activation of complement. In this study, we observed deposition of C5b-9, the terminal product of complement activation, in the wall of retinal vessels of human eye donors with 9 +/- 3 years of type 2 diabetes, but not in the vessels of age-matched nondiabetic donors. C5b-9 often colocalized with von Willebrand factor in luminal endothelium. C1q and C4, the complement components unique to the classical pathway, were not detected in the diabetic retinas, suggesting that C5b-9 was generated via the alternative pathway, the spontaneous activation of which is regulated by complement inhibitors. The diabetic donors showed a prominent reduction in the retinal levels of CD55 and CD59, the two complement inhibitors linked to the plasma membrane by glycosylphosphatidylinositol anchors, but not in the levels of transmembrane CD46. Similar complement activation in retinal vessels and selective reduction in the levels of retinal CD55 and CD59 were observed in rats with a 10-week duration of streptozotocin-induced diabetes. Thus, diabetes causes defective regulation of complement inhibitors and complement activation that precede most other manifestations of diabetic retinal microangiopathy. These are novel clues for probing how diabetes affects and damages vascular cells.

Activation of nuclear factor-kappaB induced by diabetes and high glucose regulates a proapoptotic program in retinal pericytes.

To reconstruct the events that may contribute to the accelerated death of retinal vascular cells in diabetes, we investigated in situ and in vitro the activation of nuclear factor-kappaB (NF-kappaB), which is triggered by cellular stress and controls several programs of gene expression. The retinal capillaries of diabetic eye donors showed an increased number of pericyte nuclei positive for NF-kappaB, when compared with nondiabetic donors, whereas endothelial cells were negative. Microvascular cell apoptosis and acellular capillaries were increased only in the diabetic donors with numerous NF-kappaB-positive pericytes. Likewise, high glucose in vitro activated NF-kappaB in retinal pericytes but not in endothelial cells, and increased apoptosis only in pericytes. Studies with NF-kappaB inhibitors suggested that in pericytes, basal NF-kappaB has prosurvival functions, whereas NF-kappaB activation induced by high glucose is proapoptotic. Pericytes exposed to high glucose showed increased expression of Bax and of tumor necrosis factor-alpha, which were prevented by the NF-kappaB inhibitors and mimicked by transfection with the p65 subunit of NF-kappaB, and failed to increase the levels of the NF-kappaB-dependent inhibitors of apoptosis. Colocalization of activated NF-kappaB and Bax overexpression was observed in the retinal pericytes of diabetic donors. A proapoptotic program triggered by NF-kappaB selectively in retinal pericytes in response to hyperglycemia is a possible mechanism for the early demise of pericytes in diabetic retinopathy.