Reduction of advanced glycation end-product (AGE) levels in nervous tissue proteins of diabetic Lewis rats following islet transplants is related to different durations of poor metabolic control

Advanced glycation end-products (AGEs) are irreversible compounds which, by abnormally accumulating over proteins as a consequence of diabetic hyperglycaemia, can damage tissues and thus contribute to the pathogenesis of diabetic complications. This study was performed to evaluate whether restoration of euglycaemia by islet transplantation modifies AGE accumulation in central and peripheral nervous tissue proteins and, as a comparison, in proteins from a non-nervous tissue. Two groups of streptozotocin diabetic inbred Lewis rats with 4 (T1) or 8 (T2) months disease duration were grafted into the liver via the portal vein with 1200-1500 islets freshly isolated from normal Lewis rats. Transplanted rats, age-matched control and diabetic rats studied in parallel, were followed for a further 4-month period. At study conclusion, glycaemia, glycated haemoglobin and body weight were measured in all animals, and an oral glucose tolerance test (OGTT) performed in transplanted rats. AGE levels in cerebral cortex, spinal cord, sciatic nerve proteins and tail tendon collagen were measured by enzyme-linked immunosorbent assay (ELISA). Transplanted animal OGTTs were within normal limits, as were glycaemia and glycated haemoglobin. Diabetic animal AGEs were significantly higher than those of control animals. Protein AGE values were reduced in many transplanted animals compared to diabetic animals, reaching statistical significance in spinal cord (P < 0.05), sciatic nerve (P < 0.02) and tail tendon collagen (P < 0.05) of T1 animals. Thus, return to euglycaemia following islet transplantation after 4 months of diabetes with poor metabolic control reduces AGE accumulation rate in the protein fractions of the mixed and purely peripheral nervous tissues (spinal cord and sciatic nerve, respectively). However, after a double duration of bad metabolic control, a statistically significant AGE reduction has not been achieved in any of the tissues, suggesting the importance of an early therapeutic intervention to prevent the possibly pathological accumulation of AGEs in nervous and other proteins.     

Advanced glycation end products in vitreous: Structural and functional implications for diabetic vitreopathy

PURPOSE: Advanced glycation end products (AGEs) form irreversible cross-links with many macromolecules and have been shown to accumulate in tissues at an accelerated rate in diabetes. In the present study, AGE formation in vitreous was examined in patients of various ages and in patients with diabetes. Ex vivo investigations were performed on bovine vitreous incubated in glucose to determine AGE formation and cross-linking of vitreous collagen. METHODS: By means of an AGE-specific enzyme-linked immunosorbent assay (ELISA), AGE formation was investigated in vitreous samples obtained after pars plana vitrectomy in patients with and without diabetes. In addition, vitreous AGEs were investigated in bovine vitreous collagen after incubation in high glucose, high glucose with aminoguanidine, or normal saline for as long as 8 weeks. AGEs and AGE cross-linking was subsequently determined by quantitative and qualitative assays. RESULTS: There was a significant correlation between AGEs and increasing age in patients without diabetes (r = 0.74). Furthermore, a comparison between age-matched diabetic and nondiabetic vitreous showed a significantly higher level of AGEs in the patients with diabetes (P < 0.005). Collagen purified from bovine vitreous incubated in 0.5 M glucose showed an increase in AGE formation when observed in dot blot analysis, immunogold labeling, and AGE ELISA. Furthermore, there was increased cross-linking of collagen in the glucose-incubated vitreous, when observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and protein separation. This cross-linking was effectively inhibited by coincubation with 10 mM aminoguanidine. CONCLUSIONS: This study suggests that AGEs may form in vitreous with increasing age. This process seems to be accelerated in the presence of diabetes and as a consequence of exposure to high glucose. Advanced glycation and AGE cross-linking of the vitreous collagen network may help to explain the vitreous abnormalities characteristic of diabetes.     

Nonenzymatic glycation of type IV collagen and matrix metalloproteinase susceptibility

The glomerular basement membrane (GBM) is damaged in diabetes through complex mechanisms that are not fully understood. Prominent among them is nonenzymatic protein glycation leading to the formation of so-called advanced glycation end products (AGEs). We examined the effects of in vitro glycation of intact collagen type IV in bovine lens capsule (LBM) and kidney glomerular (GBM) basement membranes on their susceptibility to matrix metalloproteinases, using stromelysin 1 (MMP-3) and gelatinase B (MMP-9). Sites of cleavage of unmodified LBM collagen were located in the triple helical region. In vitro glycation by glucose severely inhibited the release of soluble collagen cleavage peptides by MMP-3 and MMP-9. The distribution of AGEs within the three domains of collagen IV (7S, triple helical, and noncollagenous NC1) were compared for LBM glycation using AGE fluorescence, pentosidine quantitation, and immunoreactivity towards anti-AGE antibodies that recognize the AGE carboxymethyllysine (CML). Marked asymmetry was observed, with the flexible triple helical domain having the most pentosidine and fluorescent AGEs but the least CML. The in vivo relevance of these findings is supported by preliminary studies of AGE distribution in renal basement membrane (RBM) collagen IV domains from human kidneys of two insulin-dependent diabetics and one normal subject. Pentosidine and fluorescent AGE distributions of diabetic RBM were similar to LBM, but the CML AGE in diabetic kidney was less in the triple helical domain than in NC1. Our results support the hypothesis that nonenzymatic glycation of collagen IV contributes to the thickening of basement membranes, a hallmark of diabetic nephropathy.     

Congenital malformations in experimental diabetic pregnancy: aetiology and antioxidative treatment. Minireview based on a doctoral thesis

Diabetes mellitus in pregnancy causes congenital malformations in the offspring. The aim of this work was to characterize biochemical and morphologic anomalies in the conceptus of an animal model of diabetic pregnancy. In addition, a preventive treatment against diabetes-induced dysmorphogenesis was developed. Congenital cataract was often found in the offspring of diabetic rats. The fetal lenses had increased water accumulation, sorbitol concentration and aldose reductase activity compared to control lenses. The results suggest that the cataracts form via osmotic attraction of water due to sorbitol accumulation in the fetal lens. Another set of malformations, with possible neural crest cell origin, occurred frequently in offspring of diabetic rats. These included low set ears, micrognathia, hypoplasia of the thymus, thyroid and parathyroid glands, as well as anomalies of the heart and great vessels. Furthermore, diabetes caused intrauterine death and resorptions more frequently in the late part of gestation. When the pregnant diabetic rats were treated with the antioxidants butylated hydroxytoluene, vitamin E or vitamin C, the occurrence of gross malformations was reduced from approximately 25% to less than 8%, and late resorptions from 17% to 7%. This suggests that an abnormal handling of reactive oxygen species (ROS) is involved in diabetes-induced dysmorphogenesis in vivo. Indeed, an increased concentration of lipid peroxides, indicating damage caused by ROS, was found in fetuses of diabetes rats. In addition, embryos of diabetic rats had low concentrations of the antioxidant vitamin E compared to control embryos. These biochemical alterations were normalized by vitamin E treatment of the pregnant diabetic rats. The antioxidants are likely to have prevented ROS injury in the embryos of the diabetic rats, in particular in the neural crest cells, thereby normalizing embryonic development. These results provide a rationale for developing new anti-teratogenic treatments for pregnant women with diabetes mellitus.     

The effects of streptozocin-induced diabetes on renal plasma flow

To evaluate the significance of collagen levels, nonenzymatic glycation, and effective renal plasma flow in diabetes, we studied 52 Streptozocin induced-diabetic rats. After 10 weeks of diabetes, rats were injected i.v. 0.5 microCi/g 99mTechnetium Mercaptoacetyl triglycine and effective renal plasma flow was calculated from the renograms obtained. The collagen content and hydroxymethyl furfural levels of kidney and tail tissue as well as renal plasma flow increased significantly in diabetic rats (p < 0.05). The increase in renal plasma flow indicates that hyperperfusion may play a role in diabetic nephropathy mechanism. The correlation between renal plasma flow and glycation was not as high as the correlations between blood glucose concentrations and hydroxymethyl furfural and collagen levels, suggesting that factors other than glycation are important in increasing the renal plasma flow.     

Protein glycation in the aging male Sprague-Dawley rat: effects of antiaging diet restrictions

Protein glycation and accumulation of advanced glycosylated end-products (AGEs) are supposed to play an important role in the process of aging. Dietary restriction increases life span and delays the onset of most age-associated diseases. Age-dependent changes in glucose homeostasis and glycated plasma proteins and hemoglobin were determined, and AGEs formation was measured as fluorescence in skin and aortic collagens in male Sprague-Dawley rats fed ad libitum or subjected to every-other-day feeding or 40% food restriction. In aging control rats, skin and aortic collagen-linked fluorescence increased with a similar exponential curve (aortic value being always higher), whereas glycated plasma protein and hemoglobin decreased slightly. Dietary restrictions decreased glycated plasma proteins and fluorescent products in skin collagen of younger but not older rats, and did not affect glycated hemoglobin or aortic collagen fluorescence. In conclusion, our data indicate that age-related changes in glucose homeostasis do not play a substantial role in aging; and collagen-linked fluorescence increases significantly during aging, but it may not be sensitive to dietary intervention.     

Depletion of reactive advanced glycation endproducts from diabetic uremic sera using a lysozyme-linked matrix

Diabetic uremic sera contain excessive amounts of reactive advanced glycation endproducts (AGEs), which accelerate the vasculopathy of diabetes and end-stage renal disease. To capture in vivo-derived toxic AGEs, high affinity AGE-binding protein lysozyme (LZ) was linked to a Sepharose 4B matrix. Initial studies showed that > 80% of 125I-AGE-BSA was retained by the LZ matrix, compared with < 10% retained by a control matrix. More than 60% of AGE-lysine was captured by the LZ matrix, and the LZ-bound fraction retained immunoreactivity and cross-linking activity, but had little intrinsic fluorescence (370/440 nm). After passage through the LZ matrix, AGE levels in diabetic sera (0.37+/-0.04 U/mg) were significantly reduced to a level (0.09+/-0.01 U/mg; n = 10; P < 0. 0001) comparable with the level of normal human serum, whereas total protein absorption was < 3%. The AGE-enriched serum fraction exhibited cross-linking activity, which was completely prevented by aminoguanidine. Among numerous LZ-bound proteins in diabetic uremic sera, three major proteins "susceptible" to AGE modification were identified: the immunoglobulin G light chain, apolipoprotein J (clusterin/SP-40,40), and the complement 3b beta chain. These findings indicate that the LZ-linked AGE affinity column may serve as an efficient method for the depletion of toxic AGEs from sera, including specific AGE-modified proteins that may be linked to altered immunity, lipoprotein metabolism, and accelerated vasculopathy in renal failure patients with or without diabetes.     

Accumulation of pyrraline-modified albumin in phagocytes due to reduced degradation by lysosomal enzymes

Previous studies suggested that the interaction between proteins modified by advanced glycation end products (AGEs) and cells, such as macrophages, may be involved in diabetic angiopathy. Pyrraline is one of the AGEs and known to be elevated in plasma of diabetic rats and humans, and is present in vascular lesions of diabetic and elderly subjects. We examined whether modification of albumin by pyrraline influences its degradation by macrophage-like cell line, P388D1 cells. Degradation of pyrraline-modified albumin by these cells was diminished, causing accumulation of the albumin in these cells. The susceptibility of pyrraline-modified albumin to lysosomal proteolytic enzymes was reduced by approximately 40% in vitro, while lysosomal activity in the cells per se was not affected. This phenomenon was also observed when human monocytes were used instead of P388D1 cells. Our results suggest that accumulation of pyrraline-modified albumin in P388D1 cells is due to the reduced susceptibility of the protein to lysosomal enzymatic degradation. Such alterations in the interaction between AGEs-modified protein and phagocytes may contribute to angiopathy in elderly subjects and patients with diabetes.     

Potential benefit of inhibitors of advanced glycation end products in the progression of type II diabetes: a study with aminoguanidine in C57/BLKsJ diabetic mice

Prolonged hyperglycemia in type II diabetic patients is linked both with diabetic complications and with further impairment of glucose homeostasis, possibly due to glucose toxicity of the beta cell. While the connection between the accumulation of extracellular advanced glycation end products (AGEs) and the development of complications is well established, it has only recently been suggested that intracellular glycation may be equally adverse and could be involved in the pathogenesis of glucose toxicity in vitro. Aminoguanidine is a recognized inhibitor of the formation of both extracellular and intracellular AGEs. In this study, we show that the development of diabetes, measured by increased water intake and concomitant midday blood glucose levels in type II genetically diabetic mice, is reduced by treatment with aminoguanidine at a dosage of 500 mg/kg/d for 12 weeks in the diet. In addition, at the end of the study, aminoguanidine reduced the decline in serum and pancreatic insulin levels and the degree of pancreatic islet morphological degeneration, all of which are associated with pancreatic insufficiency following prolonged hyperglycemia in this animal model. These results suggest that AGEs may be involved in the aggravation of type II diabetes in vivo and aminoguanidine may be beneficial in its treatment.     

Tissue ascorbic acid and polyol pathway metabolism in experimental diabetes

Previous studies demonstrating reduced plasma concentrations of ascorbic acid (AA) in diabetes and interactions between this vitamin and biochemical mechanisms such as synthesis of structural proteins, oxidative stress, polyol pathway and nonenzymatic glycation of proteins suggest that disturbed AA metabolism may be important in the pathogenesis of diabetic microangiopathy. However, limited information is available on the concentration of AA in tissues which develop diabetic complications. This study demonstrates reduced renal but not sciatic nerve or plasma AA concentration in two animal models of insulin-dependent diabetes mellitus, namely the STZ-diabetic rat and the spontaneously diabetic BB rat. Decreased lens AA concentration was also observed in STZ-diabetic rats. Improvement of glycaemic control by insulin treatment (albeit insufficient to achieve normoglycaemia) partially corrected lens and renal AA concentration in STZ-diabetic rats. AA treatment increased kidney and lens AA concentrations of STZ-diabetic and non-diabetic rats and corrected the abnormalities observed for untreated diabetic rats. Sciatic nerve AA concentration was not increased by AA treatment in any group. Tissue ratios of dehydroascorbic acid (DHAA)/AA, one index of oxidative stress, were not different between the diabetic and non-diabetic groups and were unaltered by AA supplementation. AA treatment of STZ-diabetic rats had no effect on elevated tissue concentrations of glucose, sorbitol and fructose or reduced myo-inositol concentration. The effect of reduced tissue AA levels in diabetes on either collagen synthesis or ability to combat increased free radical production is not known. However, correction of abnormal kidney and lens AA concentrations in experimental diabetes by AA supplementation suggests that if AA does have a role in the development or progression of the renal and ocular complications of diabetes, this treatment could be beneficial.     

Specific fluorescence assay for advanced glycation end products in blood and urine of diabetic patients

Late rearrangement products that accumulate by glycation of proteins, known as advanced glycation end products (AGEs), have been implicated in the pathogenesis of complications related to diabetes. Circulating AGEs, especially in the form of a small peptide (AGE-peptide) of less than 10 kd, increase in the blood of diabetic patients with end-stage renal disease (ESRD). The aim of the study was to evaluate AGE-peptide levels by measuring AGE-specific fluorescence (excitation at 370 nm and emission at 440 nm) and to examine the relationship between AGE-peptide and diabetic nephropathy. AGE-specific fluorescence in serum and urine were examined in diabetic subjects with various levels of renal complications of varying severity: normoalbuminuria (N), microalbuminuria (Mi), macroalbuminuria (Ma), chronic renal failure (C), and hemodialysis (HD). We also assessed correlations among the AGE-peptide level and age, duration of diabetes, hemoglobin A1c (HbA1c), serum creatinine, and creatinine clearance. Serum and urine AGE-peptide levels in C and HD were significantly higher than in N, Mi, and Ma. Serum AGE-peptide levels were significantly correlated with serum creatinine (r=.866, P < .0001) and creatinine clearance (r=-.720, P < .0001) but not with duration of diabetes or age. There was a significant correlation between AGE-peptide levels measured by enzyme-linked immunosorbent assay (ELISA) and levels determined from the specific fluorescence intensity (r=.688, P < .0001). These findings suggest that renal function may play a greater role in the accumulation of AGEs than persistent hyperglycemia in diabetic patients. Measurement of AGE-specific fluorescence (ie, AGE-peptide) may serve as a simple and useful test to assess circulating AGE levels and monitor AGE excretion.     

Vitamin E decreases the occurrence of malformations in the offspring of diabetic rats

An association between excess oxygen radical activity and disturbed embryogenesis in diabetic pregnancy has been suggested. In the present study, the protective capacity of vitamin E with different treatment regimens was investigated in early and late pregnancy of streptozotocin-induced diabetic rats. Daily gavaging of 0.2 g/kg or 0.8 g/kg of vitamin E exerted moderate protective effects. In contrast, treatment with a diet enriched with 2% (wt/wt) of vitamin E, yielding an approximate daily dosage of 2 g/kg of vitamin E, clearly restored both embryonic and fetal morphology. High-performance liquid chromatography measurement showed that maternal diabetes decreased embryonic content of vitamin E. When pregnant diabetic animals were supplemented with vitamin E, increased concentrations of the vitamin were found in maternal, embryonic, and fetal tissues. Thus, despite marked accumulation of vitamin E in maternal tissues, the compound apparently reached the conceptus. Thiobarbituric acid reactive substances (TBARS) were estimated as a measure of lipid peroxidation, and no changes were observed in maternal tissue, embryonic tissue, placenta, and fetal brain in the untreated diabetic group. In contrast, a fivefold increase of TBARS was found in fetal liver, a rise that was reduced with vitamin E treatment of the diabetic pregnant rats and completely normalized with 2% vitamin E in the diet. Congenital malformations caused by experimental diabetes can be prevented by antioxidants in vivo. These findings further corroborate the notion that an imbalance in the metabolism of free oxygen radicals is involved in the embryonic maldevelopment of diabetic pregnancy, and suggest a direction for prophylactic treatment in the future.     

Effect of glycated collagen on proliferation of human smooth muscle cells in vitro

While non-enzymatic glycation of long-lived tissue proteins such as collagen has been implicated in chronic complications of diabetes mellitus, its role in the aetiology of diabetic macroangiopathy has not been elucidated. To test the hypothesis that glycation of collagen abolishes the inhibitory effect of native collagen on the proliferation of human smooth muscle cells, we obtained smooth muscle cells from human gastric arteries and cultured them on dishes coated with glycated or non-glycated collagen. The proliferation of human smooth muscle cells in the presence of 10% fetal calf serum or platelet derived growth factor-BB (10 ng/ml) was inhibited by type 1 collagen coated on the dishes. Glycation of collagen with glucose 6-phosphate for 7 days abolished the growth-inhibitory effect of native collagen. Succinylation of collagen, which like glycation blocked the lysyl residues in collagen, also abolished the growth-inhibitory effect. Adhesion of human smooth muscle cells to collagen-coated dishes was not affected by glycation of collagen. Addition of glycated albumin to the medium did not affect the growth of human smooth muscle cells on plastic dishes. The inhibition of human smooth muscle cell proliferation by collagen was not reversed by the glycation of collagen in the presence of aminoguanidine. Results suggest that early glycation abolishes the inhibitory effect of collagen on human smooth muscle cell proliferation and may thus participate in the progression of macro-angiopathy in diabetes.     

A murine model of accelerated periodontal disease in diabetes

Diabetes is a risk factor for periodontal disease in humans. In hyperglycemia, glycoxidation of proteins and lipids results in the formation of advanced glycation endproducts, or AGEs. The accumulation of AGEs in the plasma and tissues, and their interaction with their cellular receptor for AGE (RAGE), has been implicated in diabetic complications. In order to establish a model with which to delineate the specific host response factors that underlie the development of periodontal disease in diabetes, male C57BL/6J mice were rendered diabetic with streptozotocin. One month after documentation of diabetes or control state, mice were inoculated with the human periodontal pathogen Porphyromonas gingivalis, strain 381 (P. gingivalis) or treated with vehicle. Infection with P. gingivalis was achieved, as demonstrated by infiltration of gingival tissue with granulocytes, presence of DNA specific for P. gingivalis as well as increased serum antibody titer to P. gingivalis. At 2 and 3 months after infection, increased alveolar bone loss was demonstrated in P. gingivalis-inoculated diabetic vs. non-diabetic mice, along with enhanced tissue-destructive capacity, as demonstrated by increased collagenolytic activity in gingival extracts. Consistent with an important role for AGE-RAGE interaction, increased AGE deposition and expression of vascular and monocyte RAGE were demonstrated in diabetic gingiva compared with non-diabetic controls. Taken together, these data indicate that we have established a murine model of enhanced periodontal disease in diabetes. This model will serve to delineate molecular mechanisms which account for the increased susceptibility of diabetic patients to periodontal disease.     

Influence of dietary curcumin and cholesterol on the progression of experimentally induced diabetes in albino rat

Effect of feeding 0.5% curcumin diet or 1% cholesterol diet was examined in albino rats rendered diabetic with streptozotocin injection. Diabetic rats maintained on curcumin diet for 8 weeks excreted comparatively less amounts of albumin, urea, creatinine and inorganic phosphorus. Urinary excretion of the electrolytes sodium and potassium were also significantly lowered under curcumin treatment. Dietary curcumin also partially reversed the abnormalities in plasma albumin, urea, creatine and inorganic phosphorus in diabetic animals. On the other hand, glucose excretion or the fasting sugar level was unaffected by dietary curcumin and so also the body weights were not improved to any significant extent. Diabetic rats fed curcumin diet had a lowered relative liver weight at the end of the study compared to other diabetic groups. Diabetic rats fed a curcumin diet also showed lowered lipid peroxidation in plasma and urine when compared to other diabetic groups. The extent of lipid peroxidation on the other hand, was still higher in cholesterol fed diabetic groups compared to diabetic rats fed with control diet. Thus, the study reveals that curcumin feeding improves the metabolic status in diabetic conditions, despite no effect on hyperglycemic status or the body weights. The mechanism by which curcumin improves this situation is probably by virtue of its hypocholesterolemic influence, antioxidant nature and free radical scavenging property.     

Deposition of advanced glycation end products (AGE) and expression of the receptor for AGE in cardiovascular tissue of the diabetic rat

Advanced glycation end products (AGE) in tissues are important for the central pathological features of diabetic complication. Although AGE bind to several cell-surface sites, resulting in altered cellular functions, receptor for AGE (RAGE) appears to have a central role. We examined AGE accumulation and RAGE expression in the aorta and heart of rats with streptozotocin (STZ)-induced diabetes, 0, 4, 8, 12, 16 and 24 weeks after STZ administration. Early atherosclerotic findings in the intima and medial thinning were observed in the aorta after 16 weeks of STZ-Induced diabetes. Immunohistochemistry and microscope spectrophotometry showed that AGE deposition increased significantly in the aorta and vessels of the myocardium, depending on the period of hyperglycaemia. RAGE was expressed in the endothelial cells and vascular smooth muscle cells of all animals. The number of smooth muscle cells with RAGE immunoreactivity increased until 12 weeks after STZ injection, and then decreased in rats with diabetes between 16 and 24 weeks. On the other hand, total RAGE mRNA levels in the aorta and heart continued to increase with the duration of hyperglycaemia. Furthermore, AGE-BSA induced RAGE mRNA expression of human umbilical vein endothelial cells in vitro. Taken together, the AGE accumulation might initiate diabetic macroangiopathy through RAGE, and the increase of RAGE expression by endothelial cells could be a reason that diabetes mellitus accelerates atherosclerosis rapidly.     

Age-dependent increase in ortho-tyrosine and methionine sulfoxide in human skin collagen is not accelerated in diabetes. Evidence against a generalized increase in oxidative stress in diabetes

The glycoxidation products Nepsilon-(carboxymethyl)lysine and pentosidine increase in skin collagen with age and at an accelerated rate in diabetes. Their age-adjusted concentrations in skin collagen are correlated with the severity of diabetic complications. To determine the relative roles of increased glycation and/or oxidation in the accelerated formation of glycoxidation products in diabetes, we measured levels of amino acid oxidation products, distinct from glycoxidative modifications of amino acids, as independent indicators of oxidative stress and damage to collagen in aging and diabetes. We show that ortho-tyrosine and methionine sulfoxide are formed in concert with Nepsilon-(carboxymethyl)lysine and pentosidine during glycoxidation of collagen in vitro, and that they also increase with age in human skin collagen. The age-adjusted levels of these oxidized amino acids in collagen was the same in diabetic and nondiabetic subjects, arguing that diabetes per se does not cause an increase in oxidative stress or damage to extracellular matrix proteins. These results provide evidence for an age-dependent increase in oxidative damage to collagen and support previous conclusions that the increase in glycoxidation products in skin collagen in diabetes can be explained by the increase in glycemia alone, without invoking a generalized, diabetes-dependent increase in oxidative stress.     

Immunohistochemical localization of advanced glycosylation end products in coronary atheroma and cardiac tissue in diabetes mellitus

Advanced glycosylation end products (AGEs) accumulate on long-lived extracellular matrix proteins and have been implicated in the micro- and macrovascular complications of diabetes mellitus. Within the arterial wall, AGE-modified proteins increase vascular permeability, inactivate nitric oxide activity, and induce the release of growth-promoting cytokines. Recently developed anti-AGE antibodies were used in an immunohistochemical analysis of coronary arteries obtained from type II diabetic and nondiabetic patients. High levels of AGE reactivity were observed within the atherosclerotic plaque present in vessels from selected patients with diabetes. Considered together with the pathological effects of AGEs on vascular wall homeostasis, these data support the role of advanced glycosylation in the rapidly progressive atherosclerosis associated with diabetes mellitus.     

The platysma myocutaneous flap. Indications and caveats

Arterial hypertension has been identified as a major secondary risk factor for diabetic retinopathy. However, the mechanisms by which hypertension worsens retinopathy are unknown. Inhibition of advanced glycation product formation prevents the development of experimental diabetic retinopathy in normotensive diabetic rats. In this study the effect of hypertension on the rate of diabetic retinopathy development and the formation of arteriolar thrombosis was evaluated. We also evaluated the effect of aminoguanidine, an inhibitor of advanced glycation and product formation on retinal pathology of diabetic hypertensive rats. After 26 weeks of diabetes, hypertension accelerated the development of retinopathy despite a lower mean blood glucose level than in the non-hypertensive group (diabetic spontaneous hypertensive rats (SHR) 16.00 +/- 6.83 mmol/l; diabetic normotensive Wistar Kyoto rats (WKY) 34.9 +/- 3.64 mmol/l; p < 0.0001). Diabetic SHR had nearly twice as many acellular capillaries as diabetic WKY (SHR diabetic: 91.9 +/- 7.5 acellular capillaries per mm2 of retinal area vs WKY diabetic: 53.7 +/- 8.5 acellular capillaries per mm2 of retinal area), and a 3.8-fold increase in the number of arteriolar microthromboses (SHR diabetic 23,504 +/- 5523 microns2 vs SHR non-diabetic 6228 +/- 2707 microns2). Aminoguanidine treatment of SHR diabetic rats reduced the number of acellular capillaries by 50%, and completely prevented both arteriolar deposition of PAS-positive material and abnormal microthrombus formation. These data suggest that hypertension-induced deposition of glycated proteins in the retinal vasculature plays a central role in the acceleration of diabetic retinopathy by hypertension.     

Identification of potential ferric binding residues in the iron-binding protein of pathogenic Neisseria meningitidis through structure-based multiple sequence alignments

Toxic effects of hyperglycemia-induced advanced glycosylated end products (AGEs) may explain some vasculopathic complications of diabetes. Aminoguanidine, a known inhibitor of AGE formation, was administered by gavage to Sprague-Dawley streptozotocin-induced diabetic rats made azotemic by surgical reduction of renal mass. All rats became hyperglycemic. Renal ablation caused renal insufficiency, as evidenced by markedly reduced endogenous creatinine clearances at days 7 and 14. Aminoguanidine-treated rats had significantly (P < 0.04) superior survival to that of untreated azotemic diabetic rats. We infer from the extended life in a rat model of uremia in diabetic nephropathy that aminoguanidine may prove beneficial in human diabetes.