Formation Of Acellular Capillaries Research Articles

Role of Interleukin-1β in the Development of Retinopathy in Rats: Effect of Antioxidants

Diabetic retinopathy is shown to share many similarities with chronic inflammatory disease, and in diabetes, accelerated apoptosis of retinal capillary cells is evident before histopathology can be seen. The purpose of this study was to examine the effect of interleukin (IL)-1beta on capillary cell apoptosis in rat retina and to determine the effect of antioxidants on diabetes-induced changes in retinal IL-1beta. The effect of injection of IL-1beta into the vitreous (5 ng/5 microL) of normal rats on capillary cell apoptosis (detected by terminal transferase dUTP nick-end labeling [TUNEL]) and formation of acellular capillaries was investigated in the trypsin digested retinal microvessels. The levels of IL-1beta were quantified (by ELISA and Western blot) in the retina of rats diabetic for 2 months, and the effect of administration of antioxidants on diabetes-induced changes in retinal IL-1beta was determined. The number of TUNEL-positive capillary cells in the retinal microvessels obtained from normal rats that received intravitreal injection of IL-1beta was increased by more than threefold and that of acellular capillaries by more than twofold, compared with the microvessels obtained from rats that received an intravitreal injection of PBS (5 microL) or BSA (5 ng/5 microL). IL-1beta also increased the levels of 8-hydroxy-2'-deoxyguanosine (an indicator of oxidative stress) and nitric oxide by more than 40% and activated NF-kappaB by 35% to 55%. Two months of diabetes in rats increased retinal IL-1beta levels by more than twofold, and antioxidants inhibited such increases. IL-1beta, by activation of NF-kappaB and an increase in oxidative stress, plays an important role in the retinal microvascular disease that is characteristic of diabetic retinopathy. Antioxidants inhibit diabetes-induced increases in retinal IL-1beta. These studies offer a possible rationale to test IL-1beta-targeted therapies to inhibit the development of retinopathy in diabetes.

A central role for inflammation in the pathogenesis of diabetic retinopathy.

Diabetic retinopathy is a leading cause of adult vision loss and blindness. Much of the retinal damage that characterizes the disease results from retinal vascular leakage and nonperfusion. Diabetic retinal vascular leakage, capillary nonperfusion, and endothelial cell damage are temporary and spatially associated with retinal leukocyte stasis in early experimental diabetes. Retinal leukostasis increases within days of developing diabetes and correlates with the increased expression of retinal intercellular adhesion molecule-1 (ICAM-1) and CD18. Mice deficient in the genes encoding for the leukocyte adhesion molecules CD18 and ICAM-1 were studied in two models of diabetic retinopathy with respect to the long-term development of retinal vascular lesions. CD18-/- and ICAM-1-/- mice demonstrate significantly fewer adherent leukocytes in the retinal vasculature at 11 and 15 months after induction of diabetes with STZ. This condition is associated with fewer damaged endothelial cells and lesser vascular leakage. Galactosemia of up to 24 months causes pericyte and endothelial cell loss and formation of acellular capillaries. These changes are significantly reduced in CD18- and ICAM-1-deficient mice. Basement membrane thickening of the retinal vessels is increased in long-term galactosemic animals independent of the genetic strain. Here we show that chronic, low-grade subclinical inflammation is responsible for many of the signature vascular lesions of diabetic retinopathy. These data highlight the central and causal role of adherent leukocytes in the pathogenesis of diabetic retinopathy. They also underscore the potential utility of anti-inflammatory treatment in diabetic retinopathy.

Pathogenetic mechanisms of diabetic microangiopathy

Hyperglycaemia has been shown to play a central role in diabetic microangiopathy, together with the individual background. The pathogenetic sequence is initiated by a series of interrelated biochemical abnormalities associated with hyperglycaemia, including increased flux through the polyol and hexosamine pathways, oxidative stress, AGE formation and protein kinase C (PKC) activation. These abnormalities are capable of modifying the production pattern of several autocrine/paracrine factors by resident and nonresident cells of the tissues. These changes impair the vessel wall turnover, thus leading to an abnormal vascular remodelling, characterised by altered cell and matrix turnover and contacts, vascular tone and permeability and coagulation pattern, with distinct features depending on the target tissue. The hallmark of nephropathy is an abnormal extracellular matrix (ECM) accumulation within the mesangium, sustained by an upregulation of TGF-β, possibly triggered by a local activation of the renin–angiotensin system, whereas the characteristic alteration of retinopathy is retinal ischaemia due to the formation of acellular capillaries and triggering of a process of vascular endothelial growth factor (VEGF)-dependent neovascularization. This response is very active, but leads to the formation of noncompetent vessels, at variance with macrovascular disease, characterised by an impaired angiogenic response, with generation of competent vessels, potentially compensating for reduced flow.

15th Golgi lecture: from hyperglycaemia to the dysregulation of vascular remodelling in diabetes.

Hyperglycaemia has been shown to play a central part in diabetic vascular disease, which is also influenced by individual background. Hyperglycaemia initiates the pathogenetic sequence through a series of interrelated biochemical abnormalities, including increased flux through the polyol and hexosamine pathways, oxidative stress, AGE formation and protein kinase C activation. These abnormalities are capable of modifying the function of resident and non-resident vascular cells by changing their production pattern of several autocrine and paracrine factors, including growth, vasoactive and coagulation factors and adhesion molecules. These mediators profoundly impair the physiologic turnover of the vessel wall, thus leading to an abnormal process of vascular remodelling, with alterations in cell and matrix turnover and contacts, vascular tone and permeability and coagulation pattern. This process has distinct features depending on the target tissue. The hallmark of nephropathy is an abnormal accumulation of extracellular matrix within the mesangium, sustained by an upregulation of TGF-beta, possibly triggered by a local activation of the renin-angiotensin system. The central pathological lesion in retinopathy is retinal ischaemia due to the formation of acellular capillaries. The resulting vascular endothelial growth factor-dependent neovascularization is a detrimental phenomenon leading to the formation of noncompetent vessels. Conversely, in macrovascular disease, arterial occlusion resulting from plaque formation with superimposed thrombosis elicits an angiogenic response which is impaired, but generates competent vessels, potentially compensating for reduced flow. Thus, upstream interventions interrupting the pathogenetic sequence at the level of hyperglycaemia (and related biochemical events) are the most effective, whereas downstream interventions should be targeted to the tissue affected.

Antioxidant treatment of experimental diabetic retinopathy in rats with nicanartine.

In order to study the contribution of oxidant stress to the pathogenesis of experimental diabetic retinopathy, male streptozotocin diabetic Lewis rats were treated with the antioxidant and lipid-lowering compound nicanartine (80 mg/kg; n = 8, blood glucose level 16.7 +/- 3.9 mmol/l; HbA1 11.8 +/- 1.6%) by oral supplementation for 6 months and compared with untreated diabetic (n = 6; blood glucose 18.1 +/- 1.4 mmol/l; HbA1 11.5 +/- 1.5%) and untreated, non-diabetic rats (n = 8; blood glucose 4.0 +/- 0.6 mmol/l; HbA1 4.8 +/- 0.9%). Diabetic retinopathy was evaluated by computer-assisted quantitative morphometry including measurement of autofluorescent advanced glycated end-products and immunohistochemistry for heme oxygenase I. Antioxidant treatment did not inhibit the 3.1-fold increase of retinal advanced glycation end products, while the expression of heme oxygenase I in both vascular and glial structures was markedly reduced. Chronic hyperglycaemia led to a 37.3% increase in endothelial cells (p < 0.001 vs normal controls) and a 26.6% reduction in pericyte numbers (p < 0.001 vs controls). Both abnormalities were significantly ameliorated by nicanartine (p < 0.001 vs diabetic controls). No effect was observed on the formation of acellular capillaries or microaneurysms. These data indicate that antioxidant therapy with nicanartine is of limited benefit in diabetic retinopathy, at least in the rodent model of streptozotocin-induced diabetes.

Acceleration of experimental diabetic retinopathy in the rat by omega-3 fatty acids

Omega-3 fatty acids exert several important biological effects on factors that may predispose to diabetic retinopathy. Potential pathogenetic mechanisms include platelet dysfunction, altered eicosanoid production, increased blood viscosity in association with impaired cell deformability and pathologic leucocyte/endothelium interaction. Therefore, we tested whether a 6-month administration of fish oil (750 mg Maxepa, 5 times per week), containing 14% eicosapentaenoic acid (EPA) and 10% docosahexaenic acid, could inhibit the development of experimental retinopathy of the streptozotocin-diabetic rat. The efficiency of fish oil supplementation was evaluated by measuring EPA concentrations in total, plasma and membrane fatty acids and by measuring the generation of lipid mediators (leukotrienes and thromboxanes). Retinal digest preparations were quantitatively analysed for pericyte loss, and the formation of acellular capillaries. Omega-3 fatty acid administration to diabetic rats resulted in a twofold increase of EPA 20:5 in total fatty acids, and a reduction of the thromboxane ratio from 600 (untreated diabetic rats) to 50 (treated diabetic rats). Despite these biochemical changes, diabetes-associated pericyte loss remained unaffected and the formation of acellular, occluded capillaries was increased by 75% in the fish oil treated diabetic group (115.1 +/- 26.8; untreated diabetic 65.2 +/- 15.0 acellular capillary segments/mm2 of retinal area). We conclude from this study that dietary fish oil supplementation may be harmful for the diabetic microvasculature in the retina.

Effect of aldose reductase inhibitors on the progression of retinopathy in galactose-fed dogs

The onset and progression of diabetic-like retinopathy has been investigated in age-matched male beagle dogs fed a 30% galactose diet. Examination of the intact retinal vessels, isolated by gentle trypsin digestion, reveals that the destruction of pericytes to form pericyte ghosts is the earliest observable retinal vessel change. This results in an irregular distribution of endothelial cell nuclei near pericyte ghosts and the subsequent formation of acellular capillaries containing neither endothelial cells or pericytes. This was followed by the histological appearance of microaneurysms and later, by the funduscopic appearance of intraretinal hemorrhages. Studies in similar galactose-fed dogs treated with aldose reductase inhibitors indicate that all of these changes are linked to the initial aldose reductase associated destruction of pericytes.