渐进性糖化终产物与糖尿病肾病

高级糖化终末产物(advanced glycation end product,AGE)参与了糖尿病、动脉粥样硬化、癌症等多种疾病的发生和发展,尤其是其导致的糖尿病肾病(diabetic nephropathy,DN)是终末期肾衰竭的主要病因,因此探索以AGEs为靶点的DN治疗手段成为了国内外研究的热点。本文概述了国内外关于AGE参与DN的发病机制,靶向AGE的DN治疗策略,以及天然中药基于AGE为靶点干预DN的研究进展,初步探讨了靶向AGE的DN天然药物的筛选模型。     

Formation of advanced glycation end (AGE) products in diabetes: Prevention by pyruvate and a-keto glutarate

Glycation of proteins and their subsequent structural and functional modifications have been ascribed to play a prominent role in the pathogenesis of several secondary complications of diabetes, such as cataract and retinopathy. In addition, it plays a role in the generalized ageing process as well. Investigations have been conducted to explore the possibility of preventing the above process by use of pyruvate and a-keto glutarate as representatives of physiologically compatible keto acids. The results demonstrate that both these compounds are effective in preventing the initial glycation reaction as well as the formation of AGE products. Both these compounds also inhibit the generation of high molecular weight aggregates associated with cataract formation. Mechanistically, the preventive effects appear to be due to (1) competitive inhibition of glycation by the keto acids and (2) the antioxidant (radical scavenging) properties of these compounds. The results are hence considered usefu l from the point of view of developing these and other keto acid derivatives as pharmacological agents useful in preventing glycation related protein changes and consequent tissue pathological manifestations.     

晚期糖基化终产物在糖尿病肾病中的病理作用

高糖环境下体内积聚的晚期糖基化终产物(advanced glycation end products,AGEs)是糖尿病慢性并发症的主要致病因素.AGEs可通过对蛋白的修饰直接作用于机体或通过受体介导的作用影响机体.本文就AGEs的来源、病理生理作用,尤其是在糖尿病肾病(diabetic nephropathy,DN)发生发展中的作用及治疗干预作一综述.     

晚期糖基化终末产物、一氧化氮在糖尿病周围神经病变中的作用

目的:研究晚期糖基化终末产物(advanced glycation end products,AGE)、一氧化氮(nitric oxide,NO)在糖尿病神经病变中的作用.方法:选择69名糖尿病患者,通过是否合并周围神经病变,分为糖尿病无神经病变组39例,糖尿病合并有神经病变组30例,另外设正常对照组30例,分别测血清AGE、NO水平、胆固醇、空腹血糖、糖化血红蛋白等三组间进行比较.结果:糖尿病患者血清AGE高于正常对照组(P＜0.05),糖尿病患者血清NO低于正常对照组(P＜0.05).结论:AGE表达的上调可能与糖尿病神经病变的发生、发展关系密切;NO表达的下调可能与糖尿病神经病变的发生、发展关系密切.     

Advanced glycation end products and diabetic retinopathy

Studies have established hyperglycemia as the most important factor in the progress of vascular complications. Formation of advanced glycation end products (AGEs) correlates with glycemic control. The AGE hypothesis proposes that hyperglycemia contributes to the pathogenesis of diabetic complications including retinopathy. However, their role in diabetic retinopathy remains largely unknown. This review discusses the chemistry of AGEs formation and their patho-biochemistry particularly in relation to diabetic retinopathy. AGEs exert deleterious effects by acting directly to induce cross-linking of long-lived proteins to promote vascular stiffness, altering vascular structure and function and interacting with receptor for AGE, to induce intracellular signaling leading to enhanced oxidative stress and elaboration of key proinflammatory and prosclerotic cytokines. Novel anti-AGE strategies are being developed hoping that in next few years, some of these promising therapies will be successfully evaluated in clinical context aiming to reduce the major economical and medical burden caused by diabetic retinopathy.     

Detection and determination of glyceraldehyde-derived pyridinium-type advanced glycation end product in streptozotocin-induced diabetic rats

GLAP, glyceraldehyde-derived pyridinium-type advanced glycation end product (AGE), formed by glyceraldehyde-related glycation, was identified in the plasma protein and the tail tendon collagen of streptozotocin (STZ)-induced diabetic rats. It was detected in the plasma protein and the collagen in diabetic rats by LC-MS and LC-MS/MS analysis, but was not detected in normal rats. In addition, GLAP was formed from glyceraldehyde-3-phosphate (GA3P) with lysine as well as glyceraldehyde (GLA) with lysine in vitro. Accordingly, it is suggested that an increase in the GLAP level reflects an increase in the GLA level and the GA3P level. GLAP might be a biomarker for reduced activity of the glyceraldehyde-related enzymes in the metabolic diseases such as diabetic complications.     

Cellular signalling of the receptor for advanced glycation end products (RAGE)

The receptor for advanced glycation end-product (RAGE) is the signal transduction receptor which senses a variety of signalling molecules including advanced glycation end products (AGEs), HMGB1, S100/calgranulins, β-amyloid, phosphatidylserine, C3a and advanced oxidation protein products (AOPPs). It is usually abnormally up-regulated and plays crucial roles during the development of many human diseases such as diabetes, cardiovascular diseases, osteoarthritis and cancer. RAGE regulates a number of cell processes of pivotal importance like inflammation, apoptosis, proliferation and autophagy. Therapeutic strategies to block RAGE may represent great therapeutic potentials and therefore it has been under extensive investigation during the last decade. Accordingly, there is a growing interest of unraveling the intracellular signalling pathways by which RAGE controls these disease-related processes. Early studies are mainly focused on inflammatory pathways involving the NFκB and the MAPK pathways. Nevertheless, many novel signalling pathways implicated in other cell processes, such as autophagy, have also recently been found to be activated upon RAGE stimulation and contribute to the detrimental effects of RAGE. In this review, we aim to provide a comprehensive summary of previous and recent studies relating to the complex molecular network of RAGE signalling, with a particular emphasis on RAGE transgenic mouse models.     

Acetoacetate promotes the formation of fluorescent advanced glycation end products (AGEs)

Acetoacetate (AA) is an important ketone body, which produces reactive oxygen species (ROS). Advanced glycation end products (AGEs) are defined as final products of glycation process whose production is influenced by the levels of ROS. The accumulation of AGEs in the body contributes to pathogenesis of many diseases including complications of diabetes, and Alzheimer’s and Parkinson’s disease. Here, we evaluated the impact of AA on production of AGEs upon incubation of human serum albumin (HSA) with glucose. The effect of AA on the AGEs formation of HSA was studied under physiological conditions after incubation with glucose for 35 days. The physical techniques including circular dichroism (CD) and fluorescence spectroscopy were used to assess the impact of AA on formation and structural changes of glycated HSA (GHSA). Our results indicated that the secondary and tertiary structural changes of GHSA were increased in the presence of AA. The fluorescence intensity measurements of AGEs also showed an increase in AGEs formation. Acetoacetate has an activator effect in formation of AGEs through ROS production. The presence of AA may result in enhanced glycation in the presence of glucose and severity of complications associated with accumulation of AGEs.     

Regulation of advanced glycation end product (AGE) receptors and apoptosis by AGEs in osteoblast-like cells

It has been reported that apelin functions as an adipokine, which has been associated to obesity and insulin resistance. The objective of this study was to analyze the apelin mRNA expression in white adipose tissue (WAT) from high-fat (Cafeteria) fed rats, in order to examine potential relationships with obesity markers and other related risk factors. Animals fed on the high-fat diet during 56 days increased their body weight, total body fat and WAT depots weights when compared to controls. Apelin subcutaneous mRNA expression was higher in the Cafeteria than in the Control fed group and this increase was partially reversed by dietary vitamin C supplementation. Statistically significant associations between subcutaneous apelin gene expression and almost all the studied variables were identified, being of special interest the correlations found with serum leptin (r = 0.517), liver malondialdehyde (MDA) levels (r = 0.477), and leptin, IRS-3 and IL-1ra retroperitoneal mRNA expression (r = 0.701; r = 0.692 and r = 0.561, respectively). These associations evidence a possible role for apelin in the excessive weight gain induced by high-fat feeding and increased adiposity, insulin-resistance, liver oxidative stress and inflammation.     

Increased levels of vascular endothelial growth factor and advanced glycation end products in aqueous humor of patients with diabetic retinopathy.

Clinical studies have shown a relationship between diabetic retinopathy and vascular endothelial growth factor (VEGF) levels in ocular fluid. Advanced glycation end products (AGEs) have been implicated in diabetes complications, including diabetic retinopathy. Nepsilon-(carboxymethyl) lysine (CML) is a glycoxidation product that may be a marker of oxidative stress. In this study, we used enzyme-linked immunosorbent assays to determine the levels of VEGF, non-CML AGE and CML in the aqueous humor and serum of 82 Japanese patients with type 2 diabetes and 60 non-diabetic subjects. VEGF, non-CML AGE, and CML concentrations in aqueous humor and serum were then compared with the severity of diabetic retinopathy. Immunohistochemical detection analysis of non-CML AGE and CML was also performed using retinal tissues from patients with progressive diabetic retinopathy. Aqueous levels of VEGF, non-CML AGE and CML increased along with the progression of diabetic retinopathy compared to age-matched controls. After coagulation therapy, the VEGF, non-CML AGE, and CML levels were significantly reduced. Immunostaining showed diffuse co-localization of non-CML AGE and CML around microvessels and in the glial cells of proliferative membranes from patients with progressive diabetic retinopathy. These findings suggest that glycation and glycoxidation reactions (or oxidation, as revealed by CML) may contribute to both the onset and progression of diabetic retinopathy.     

Impact of advanced glycation end products (AGEs) signaling in coronary artery disease

Coronary artery disease remains the leading cause of mortality in adult diabetic population with however, a high predominance also in non-diabetic subjects. In search of common molecular mechanisms and metabolic by-products with potential pathogenic role, increased advanced glycation end products (AGEs) present a critical biomarker for CAD development in both cases. Interaction of AGEs with their transmembrane cell receptor, RAGE in endothelial and smooth muscle cells as well as in platelets, activates intracellular signaling that leads to endothelial injury, modulation of vascular smooth muscle cell function and altered platelet activity. Furthermore, tissue accumulation of AGEs affects current treatment approaches being involved in stent restenosis. The present review provides an update of AGE-induced molecular mechanisms involved in CAD pathophysiology while it discusses emerging therapeutic interventions targeting AGE reduction and AGE-RAGE signaling with beneficial clinical outcome.     

Solution structure of the soluble receptor for advanced glycation end products (sRAGE)

The receptor for advanced glycation end products (RAGE) is a multiligand cell surface receptor involved in various human diseases, as it binds to numerous molecules and proteins that modulate the activity of other proteins. Elucidating the three-dimensional structure of this receptor is therefore most important for understanding its function during activation and cellular signaling. The major alternative splice product of RAGE comprises its extracellular region that occurs as a soluble protein (sRAGE). Although the structures of sRAGE domains were available, their assembly into the functional full-length protein remained unknown. We observed that the protein has concentration-dependent oligomerization behavior, and this is also mediated by the presence of Ca(2+) ions. Moreover, using synchrotron small angle x-ray scattering, the solution structure of human sRAGE was determined in the monomeric and dimeric forms. The model for the monomer displays a J-like shape, whereas the dimer is formed through the association of the two N-terminal domains and has an elongated structure. These results provide insights into the assembly of the RAGE homodimer, which is essential for signal transduction, and the sRAGE:RAGE heterodimer that leads to blockage of the receptor signaling, paving the way for the design of therapeutic strategies for a large number of different pathologies.     

Purification and characterization of mouse soluble receptor for advanced glycation end products (sRAGE)

The receptor for advanced glycation end products (RAGE) is a member of the immunoglobulin superfamily of cell surface proteins that has been implicated as a progression factor in a number of pathologic conditions from chronic inflammation to cancer to Alzheimer's disease. In such conditions, RAGE acts to facilitate pathogenic processes. Its secreted isoform, soluble RAGE or sRAGE, has the ability to prevent RAGE signaling by acting as a decoy. sRAGE has been used successfully in animal models of a range of diseases to antagonize RAGE-mediated pathologic processes. In humans, sRAGE results from alternative splicing of RAGE mRNA. This study was aimed to determine whether the same holds true for mouse sRAGE and, in addition, to biochemically characterize mouse sRAGE. The biochemical characteristics examined include glycosylation and disulfide patterns. In addition, sRAGE was found to bind heparin, which may mediate its distribution in the extracellular matrix and cell surfaces of tissues. Finally, our data indicated that sRAGE in the mouse is likely produced by carboxyl-terminal truncation, in contrast to the alternative splicing mechanism reported in humans.     

CD36-mediated endocytic uptake of advanced glycation end products (AGE) in mouse 3T3-L1 and human subcutaneous adipocytes

Interaction of advanced glycation end products (AGE) with AGE receptors induces several cellular phenomena potentially relating to diabetic complications. We here show that AGE-modified bovine serum albumin (BSA) is endocytosed by adipocytes via CD36. Upon differentiation, 3T3-L1 and human subcutaneous adipose cells showed marked increases in endocytic uptake and subsequent degradation of [(125)I]AGE-BSA, which were inhibited effectively by the anti-CD36 antibody. Ligand specificity of CD36 for modified BSAs was compared with that of LOX-1 and scavenger receptor class A. Effect of fucoidan on [(125)I]AGE-BSA binding showed a sharp contrast to that on [(125)I]-oxidized low density lipoprotein. These results implicate that CD36-mediated interaction of AGE-modified proteins with adipocytes might play a pathological role in obesity or insulin-resistance.     

Detection and determination of glyceraldehyde-derived advanced glycation end product

Protein is modified by carbonyl compound in the Maillard reaction, and the irreversible structure is formed as the advanced glycation end product (AGE). We identified GLAP (glyceraldehyde-derived pyridinium compound) as an AGE formed from glyceraldehyde and lysine residue of protein. In the present study, we investigated detection and determination of GLAP from glycated protein using fluorescence HPLC method. Albumin (BSA) and carbonyls (glyceraldehyde, glycolaldehyde, methylglyoxal, glyoxal, three pentoses or three hexoses) were dissolved in phosphate buffed solution (pH 7.4), and incubated at 37 degrees C for a week. GLAP was formed only in the glyceraldehyde-modified BSA. It is suggested that GLAP was specific AGE derived from glyceraldehyde. In addition, GLAP depressed the intracellular glutathione level and induced the reactive oxygen species (ROS) in HL-60 cells. GLAP caused the oxidative stress. Therefore, GLAP will be a biomarker in the AGE related disease such as diabetic complications or chronic renal failure.     

Overexpression of glyoxalase-I reduces hyperglycemia-induced levels of advanced glycation end products and oxidative stress in diabetic rats

The reactive advanced glycation end product (AGE) precursor methylglyoxal (MGO) and MGO-derived AGEs are associated with diabetic vascular complications and also with an increase in oxidative stress. Glyoxalase-I (GLO-I) transgenic rats were used to explore whether overexpression of this MGO detoxifying enzyme reduces levels of AGEs and oxidative stress in a rat model of diabetes. Rats were made diabetic with streptozotocin, and after 12 weeks, plasma and multiple tissues were isolated for analysis of AGEs, carbonyl stress, and oxidative stress. GLO-I activity was significantly elevated in multiple tissues of all transgenic rats compared with wild-type (WT) littermates. Streptozotocin treatment resulted in a 5-fold increase in blood glucose concentrations irrespective of GLO-I overexpression. Levels of MGO, glyoxal, 3-deoxyglucosone, AGEs, and oxidative stress markers nitrotyrosine, malondialdehyde, and F2-isoprostane were elevated in the diabetic WT rats. In diabetic GLO-I rats, glyoxal and MGO composite scores were significantly decreased by 81%, and plasma AGEs and oxidative stress markers scores were significantly decreased by ～50%. Hyperglycemia induced a decrease in protein levels of the mitochondrial oxidative phosphorylation complex in the gastrocnemius muscle, which was accompanied by an increase in the lipid peroxidation product 4-hydroxy-2-nonenal, and this was counteracted by GLO-I overexpression. This study shows for the first time in an in vivo model of diabetes that GLO-I overexpression reduces hyperglycemia-induced levels of carbonyl stress, AGEs, and oxidative stress. The reduction of oxidative stress by GLO-I overexpression directly demonstrates the link between glycation and oxidative stress.     

Telmisartan inhibits advanced glycation end products (AGEs)-elicited endothelial cell injury by suppressing AGE receptor (RAGE) expression via peroxisome proliferator-activated receptor-gammaactivation

Advanced glycation end products (AGEs)-their receptor (RAGE) axis plays a central role in the pathogenesis of diabetic microangiopathy. Since the pathophysiological crosstalk between the AGEs-RAGE system and angiotensin II has also been associated with diabetic microangiopathy, we examined here whether and how telmisartan, a unique angiotensin II type 1 receptor blocker (ARB) with peroxisome proliferator-activated receptor-gamma (PPAR-gamma)-modulating activity, could inhibit the AGEs-elicited endothelial cell injury by suppressing RAGE expression in vitro. Telmisartan suppressed RAGE expression at both mRNA and protein levels in human cultured microvascular endothelial cells (ECs), which were prevented by GW9662, an inhibitor of PPAR-gamma. Further, telmisartan was found to inhibit up-regulation of mRNA levels for monocyte chemoattractant protein-1, intercellular adhesion molecule-1 and vascular endothelial growth factor in AGEs-exposed ECs. These results suggest that telmisartan inhibits the AGEs-elicited EC injury by down-regulating RAGE expression via PPAR-gamma activation. Our present study provides a unique beneficial aspect of telmisartan. Specifically, it could work as an anti-inflammatory agent against AGEs via PPAR-gamma activation and may play a protective role against diabetic microangiopathy.