The purpose of these experiments was to determine the specific role of reactive oxygen species (ROS) in the blood-retinal barrier (BRB) breakdown that characterizes the early stages of vascular dysfunction in diabetes...
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The purpose of these experiments was to determine the specific role of reactive oxygen species (ROS) in the blood-retinal barrier (BRB) breakdown that characterizes the early stages of vascular dysfunction in diabetes. Based on our data showing that high glucose increases nitric oxide, superoxide, and nitrotyrosine formation in retinal endothelial cells, we hypothesized that excess formation of ROS causes BRB breakdown in diabetes. Because ROS are known to induce increases in expression of the well-known endothelial mitogen and permeability factor vascular endothelial growth factor (VEGF) we also examined their influence on the expression of VEGF and its downstream target urokinase plasminogen activator receptor (uPAR). After 2 weeks of streptozotocin-induced diabetes, analysis of albumin leakage confirmed a prominent breakdown of the BRB. This permeability defect was correlated with significant increases in the formation of nitric oxide, lipid peroxides, and the peroxynitrite biomarker nitrotyrosine as weft as with increases in the expression of VEGF and uPAR. Treatment with a nitric oxide synthase inhibitor (N-omega-nitro-L-arginine methyl ester, 50 mg/kg/day) or peroxynitrite scavenger (uric acid, 160 mg/kg/day) blocked the breakdown in the BRB and prevented the increases in formation of lipid peroxides and tyrosine nitration as well as the increases in expression of VEGF and uPAR. Taken together, these data indicate that early diabetes causes breakdown of the BRB by a mechanism involving the action of reactive nitrogen species in promoting expression of VEGF and uPAR. (Am J Pathol 2003, 162:1995-2004)
1. Pericytes, which are contractile cells located on the outer wall of microvessels, are thought to be particularly important in the retina where the ratio of these cells to vascular endothelial cells is the highest o...
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1. Pericytes, which are contractile cells located on the outer wall of microvessels, are thought to be particularly important in the retina where the ratio of these cells to vascular endothelial cells is the highest of any tissue. Retinal pericytes are of interest since they may regulate capillary blood flow and because their selective loss is an early event in diabetic retinopathy, which is a common sight-threatening disorder associated with dysfunction of the blood-retinal barrier. 2. Although a breakdown in the vascular endothelial barrier is a frequent pathophysiological event, knowledge of the effects of blood-derived molecules on pericyte function is limited. Based on the premise that ion channels play a vital role in cellular function, we examined the effect of serum on the ionic currents of retinal pericytes. To do this, we used the perforated patch configuration of the patch-clamp technique to monitor the whole-cell currents of pericytes located on freshly isolated rat retinal microvessels. 3. Exposure to serum reversibly activated inward and outward currents in virtually all of the sampled retinal pericytes. Two types of sustained conductances were induced by serum. These were a calcium-permeable non-specific cation (NSC) current and a voltage-dependent potassium current. In addition, exposure to serum increased the activity of chloride channels which caused transient depolarizing currents. 4. Associated with the activation of these conductances, the membrane potential showed a sustained decrease of 10 +/- 2 mV from -56 mV to -46 mV and, also, transient depolarizations to near -30 mV. The serum-induced depolarizations can activate the voltage-gated calcium channels expressed by the retinal pericytes. 5. Calcium-permeable NSC channels appear to play a critical role in the response of pericytes to serum-derived molecules. Consistent with this, activation of the chloride and potassium channels was sensitive to SK&F 98365, which is a blocker of NSC channels. I
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