Vascular Damage In Diabetes Research Articles

#2052 Reciprocal interplay between miR-155-5p and SOCS1 in the development of renal and vascular damage in experimental diabetes

Abstract Background and Aims Chronic kidney disease and atherosclerosis are common vascular complication of diabetes and a global health issue. Hyperglycemia, dyslipidemia, and cytokines activate Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathway to induce mediators of inflammation and oxidative stress, which are critical events in the progression of renal and vascular damage in diabetic patients. MicroRNAs are small, non-coding molecules and key regulators of gene expression. There is evidence of microRNA-155 (miR-155) involvement in diabetes complications, but the underlying mechanisms are unclear. In this study, gain- and loss-of-function experiments were applied to investigate the interplay between miR-155-5p and suppressor of cytokine signaling 1 (SOCS1) in the regulation of JAK/STAT pathway during renal and vascular injury in diabetes. Method Mesangial, tubuloepithelial, and vascular smooth muscle cells were transfected with miR-155-5p mimic or inhibitor, SOCS1 expression plasmid or siRNA, before stimulation with cytokines or high-glucose. Apolipoprotein E deficient mice with streptozotocin-induced diabetes were treated with either SOCS1-encoding adenovirus or miR-155-5p inhibitor. Samples were analyzed for the expression of miR-155-5p, SOCS1, and mediators of renal and vascular damage. Results In diabetic mice, the expression of miR-155-5p correlated inversely with SOCS1 levels and positively with markers of renal dysfunction, atherosclerosis, inflammation and oxidative stress. In renal cells, transfection with miR-155-5p mimic downregulated SOCS1, activated STAT1 and cytokine expression, and stimulated cell proliferation and migration. Mimic transfection also promoted the transition from contractile to synthetic phenotype in vascular smooth muscle cells. Conversely, both miR-155-5p antagonism and SOCS1 overexpression protected cells from inflammatory damage. In vivo SOCS1 gene delivery in diabetic mice decreased miR-155-5p and prevented kidney and vascular injury. Finally, therapeutic inhibition of miR-155-5p alleviates albuminuria, renal inflammation and fibrosis in diabetic mice, and also reduced atherosclerosis burden and improved plaque stability. Conclusion The modulation of miR-155/SOCS1 axis protects against diabetic renal and vascular damage in mice, thus highlighting its potential as therapeutic target for chronic complications of diabetes.

Albumin Oxidation and Albumin Glycation Discordance During Type 2 Diabetes Therapy: Biological and Clinical Implications.

Aims: Cys34 albumin redox modifications (reversible "cysteinylation" and irreversible "di/trioxidation"), besides being just oxidative stress biomarkers, may have primary pathogenetic roles to initiate and/or aggravate cell, tissue, and vascular damage in diabetes. In an exploratory "proof-of-concept" pilot study, we examined longitudinal changes in albumin oxidation during diabetes therapy. Methods: Mass spectrometric analysis was utilized to monitor changes in human serum albumin (HSA) post-translational modifications {glycation [glycated albumin (GA)], cysteinylation [cysteinylated albumin (CA) or human non-mercaptalbumin-1; reversible], di/trioxidation (di/trioxidized albumin or human non-mercaptalbumin-2; irreversible), and truncation (truncated albumin)} during ongoing therapy. Four informative groups of subjects were evaluated [type 1 diabetes (T1DM), type 2 diabetes (T2DM), prediabetes-obesity, and healthy controls] at baseline, and subjects with diabetes were followed for a period up to 280 days. Results: At baseline, T2DM was associated with relatively enhanced albumin cysteinylation (CA% total) compared with T1DM (P = 0.004), despite comparable mean hyperglycemia (P values: hemoglobin A1c = 0.09; GA = 0.09). T2DM, compared with T1DM, exhibited selectively and significantly higher elevations of all the "individual" glycated cum cysteinylated ("multimodified") albumin isoforms (P values: CysHSA+1G = 0.003; CysHSA+2G = 0.007; and CysHSA+3G = 0.001). Improvements in glycemic control and decreases in albumin glycation during diabetes therapy in T2DM were not always associated with concurrent reductions of albumin cysteinylation, and in some therapeutic situations, albumin cysteinylation worsened (glycation-cysteinylation discordance). Important differences were observed between the effects of sulfonylureas and metformin on albumin molecular modifications. Conclusions: T2DM was associated with higher oxidative (cysteinylation) and combined (cysteinylation plus glycation) albumin molecular modifications, which are not ameliorated by improved glucose control alone. Further studies are required to establish the clinical significance and optimal therapeutic strategies to address oxidative protein damage and resulting consequences in diabetes.

Traditional Chinese medicine and its active substances reduce vascular injury in diabetes via regulating autophagic activity.

Due to its high prevalence, poor prognosis, and heavy burden on healthcare costs, diabetic vascular complications have become a significant public health issue. Currently, the molecular and pathophysiological mechanisms underlying diabetes-induced vascular complications remain incompletely understood. Autophagy, a highly conserved process of lysosomal degradation, maintains intracellular homeostasis and energy balance via removing protein aggregates, damaged organelles, and exogenous pathogens. Increasing evidence suggests that dysregulated autophagy may contribute to vascular abnormalities in various types of blood vessels, including both microvessels and large vessels, under diabetic conditions. Traditional Chinese medicine (TCM) possesses the characteristics of "multiple components, multiple targets and multiple pathways," and its safety has been demonstrated, particularly with minimal toxicity in liver and kidney. Thus, TCM has gained increasing attention from researchers. Moreover, recent studies have indicated that Chinese herbal medicine and its active compounds can improve vascular damage in diabetes by regulating autophagy. Based on this background, this review summarizes the classification, occurrence process, and related molecular mechanisms of autophagy, with a focus on discussing the role of autophagy in diabetic vascular damage and the protective effects of TCM and its active compounds through the regulation of autophagy in diabetes. Moreover, we systematically elucidate the autophagic mechanisms by which TCM formulations, individual herbal extracts, and active compounds regulate diabetic vascular damage, thereby providing new candidate drugs for clinical treatment of vascular complications in diabetes. Therefore, further exploration of TCM and its active compounds with autophagy-regulating effects holds significant research value for achieving targeted therapeutic approaches for diabetic vascular complications.

Correlation of renal function indicators and vascular damage in T2DM patients with normal renal function.

This study aimed to assess the correlation between renal function-related indices and vascular damages among patients with type 2 diabetes mellitus (T2DM) and normal renal function. We screened a cohort of eligible patients with T2DM, ultimately including 826 individuals. Utilizing multifactorial logistic regression, we conducted an in-depth analysis to explore the potential associations between renal function-related indices-specifically BUN, Cr, ALB, ACR, and eGFR-and the incidence of diabetic vascular damage. Additionally, to comprehensively understand the relationships, we employed Spearman correlation analysis to assess the connections between these indicators and the occurrence of vascular damage. In this cross-sectional study of 532 patients with carotid atherosclerosis (CA), the prevalence of CA was positively correlated with Cr (53.1%, 72.3%, 68.0%, P<0.05) and negatively correlated with eGFR (71.6%, 68.5%, 53.1%, P<0.05). the higher the Cr, the higher the predominance ratio of CA (T1: reference; T2:OR. 2.166,95%CI:1.454,3.225; T3:OR:1.677, 95%CI:1.075, 2.616; P<0.05), along with an eGFR of 66.9% and 52.0% in terms of sensitivity and specificity, with a 95% CI of 0.562-0.644. Within our experimental sample, a noteworthy observation emerged: Creatinine (Cr) exhibited a positive correlation with the prevalence of individuals affected by carotid atherosclerosis (CA), underscoring a potential connection between Cr levels and CA incidence. Conversely, the estimated Glomerular Filtration Rate (eGFR) demonstrated a negative correlation with the occurrence of CA, implying that lower eGFR values might be associated with an increased likelihood of CA development.

MiR-31 ameliorates type 2 diabetic vascular damage through up-regulation of hypoxia-inducible factor-1α/vascular endothelial growth factor-A.

microRNA may be a new therapeutic direction for diabetes. As a typical tumor marker, miR-31 is involved in a variety of metabolic diseases, but the specific role is still unclear. This study aimed to investigate the effect of miR-31 on type 2 diabetes mellitus and its accompanying vascular injury, as well as on the effects of hypoxia-inducible factor-1α inhibitor (HIF1AN), hypoxia-inducible factor (HIF)-1α, and vascular endothelial growth factor (VEGF)-A expression in vitro and in vivo. In vitro, a model of high-fat and high-glucose-induced human aortic endothelial cell (HAEC) injury was established to simulate diabetes mellitus (DM). Cell functions were compared between the control group, the DM damage group, and the group transfected with miR-31 after DM damage. In vivo, overexpressing miR-31 FVB mice and FVB mice were divided into the control and induced type 2 diabetes mellitus groups. Type 2 diabetes mellitus models were induced by a high-fat diet combined with streptozotocin. The lipid metabolism levels, viscera, and vascular damage were compared between the control and type 2 diabetes mellitus groups. In vitro, miR-31 improved the proliferation ability of damaged cells by targeting HIF1AN and up-regulating the expression of HIF-1α and VEGF-A. In vivo, miR-31 ameliorated the development of type 2 diabetes mellitus, disturbance of glucose and lipid metabolism, and damage to some organs. Meanwhile, miR-31 had a protective effect on vascular damage complicated by type 2 diabetes mellitus by increasing the levels of HIF-1α and VEGF-A. Our experiments show that miR-31 can delay the progression of type 2 diabetes mellitus and ameliorate diabetic vascular injury.

A preliminary study of changes in carotid artery elasticity in type 2 diabetes mellitus.

Carotid stiffening is found to be present in patients with type 2 diabetes mellitus (T2DM) together with endothelial dysfunction and it remains unclear about the role of carotid elasticity in the development of diabetic vascular damage. The aim of the study was to investigate changes and significance of carotid artery elasticity in diabetic patients with or without microvascular complications using velocity vector imaging (VVI) analysis. Fifty participants were enrolled and divided into health Control group, the uncomplicated DM (uDM) group and the complicated DM (cDM) group. All of them underwent carotid ultrasound examinations. VVI was used to evaluate the common carotid artery (CCA) elasticity and intima-media thickness (IMT) was also measured. Flow-mediated dilation (FMD) was performed to detect the vascular endothelial function. Then differences and correlations of variables between three groups were compared and analyzed. CCA elasticity measured by VVI decreased significantly between three groups (p < 0.05), while FMD decreased significantly only in cDM group (p < 0.01) and only IMT in cDM group was significantly thicker than that of Control group (p < 0.05). Representative VVI variables were independently, negatively related to the known duration and microalbuminuria (p < 0.05). All VVI variables were significantly correlated with FMD (0.5 ≤ |r | <0.8, p < 0.001), and just a small part of VVI variables were significantly correlated with IMT (0.3 ≤ |r | <0.5, p < 0.05). Compared with FMD, CCA elasticity measured by VVI showed more obvious changes in diabetic patients with different levels of vascular damage and may be considered as an alternative indicator in evaluating arterial status of T2DM.

A Potential Target for Diabetic Vascular Damage: High Glucose-Induced Monocyte Extracellular Vesicles Impair Endothelial Cells by Delivering miR-142-5p.

Endothelial dysfunction is a key accessory to diabetic cardiovascular complications, and the regulatory role of the extracellular vesicles (EVs) from the innate immune system is growing. We tested whether EVs derived from high glucose-induced monocytes could shuttle microRNAs and impair endothelial cells. EVs from high glucose- and basal glucose-treated THP-1 cells (HG-THP-1 EVs and BG-THP-1 EVs) were isolated and identified. After coculture with THP-1 EVs, human umbilical vein endothelial cells (HUVECs) were tested by proliferation, migration, reactive oxygen species (ROS) detection assays, and western blot for Nrf2/NLRP3 signaling. MiR-142-5p was predicted by miRNAs databases and further verified by RT–qPCR and dual-luciferase reporter gene assays that inhibit Nrf2 expression. The regulation of miR-142-5p in HUVECs was further evaluated. A type 1 diabetes mellitus (T1DM) mouse model was developed for miR-142-5p inhibition. Aorta tissue was harvested for hematoxylin-eosin staining and immunohistochemistry of interleukin-1β (IL-1β). Compared to BG-THP-1 EVs, HG-THP-1 EVs significantly reduced migration and increased ROS production in HUVECs but did not affect proliferation. HG-THP-1 EVs induced suppression of Nrf2 signaling and NLRP3 signaling activation. RT–qPCR results showed that HG-THP-1 EVs overexpressed miR-142-5p in HUVECs. The transfection of miR-142-5p mimics into HUVECs exhibited consistent regulatory effects on HG-THP-1 EVs, whereas miR-142-5p inhibitors demonstrated protective effects. The miR-142-5p antagomir significantly reduced the IL-1β level in T1DM aortas despite morphological changes. To conclude, miR-142-5p transferred by high glucose-induced monocyte EVs participates in diabetic endothelial damage. The inhibition of miR-142-5p could be a potential adjuvant to diabetic cardiovascular protection.

Myoinositol Reduces Inflammation and Oxidative Stress in Human Endothelial Cells Exposed In Vivo to Chronic Hyperglycemia.

Myo-inositol (Myo) improves insulin resistance, glucose metabolism, and helps gestational diabetes (GDM) management. GDM is associated with a pro-inflammatory state and increased oxidative stress, which are both involved in vascular damage in diabetes. Our aim was to study Myo anti-inflammatory/antioxidant potential effects on an in vitro model of human umbilical vein endothelial cells (HUVECs). To this end, monocyte cell adhesion to HUVECs, adhesion molecule membrane exposure, and oxidative stress levels were determined in cells from control (C-) and GDM women treated during pregnancy either with diet only (GD-) or with diet plus Myo (GD+Myo). To deeply study the vascular effects of Myo, the same evaluations were performed in C- and GD-HUVECs following 48 h in vitro stimulation with Myo. Notably, we first observed that GD-HUVECs obtained from women assuming Myo supplementation exhibited a significantly decreased number of monocytes that adhered to endothelial cells, less adhesion molecule exposure, and lower intracellular reactive oxygen species (ROS) levels in the basal state as compared to GD-HUVECs obtained from women treated by diet only. This Myo anti-inflammatory/antioxidant effect was confirmed by 48 h in vitro stimulation of GD-HUVECs as compared to controls. Altogether, these results strongly suggest that Myo may exert protective actions against chronic inflammation induced by endothelial dysfunction in diabetes.

Diabetes and Thrombosis: A Central Role for Vascular Oxidative Stress.

Diabetes mellitus is the fifth most common cause of death worldwide. Due to its chronic nature, diabetes is a debilitating disease for the patient and a relevant cost for the national health system. Type 2 diabetes mellitus is the most common form of diabetes mellitus (90% of cases) and is characteristically multifactorial, with both genetic and environmental causes. Diabetes patients display a significant increase in the risk of developing cardiovascular disease compared to the rest of the population. This is associated with increased blood clotting, which results in circulatory complications and vascular damage. Platelets are circulating cells within the vascular system that contribute to hemostasis. Their increased tendency to activate and form thrombi has been observed in diabetes mellitus patients (i.e., platelet hyperactivity). The oxidative damage of platelets and the function of pro-oxidant enzymes such as the NADPH oxidases appear central to diabetes-dependent platelet hyperactivity. In addition to platelet hyperactivity, endothelial cell damage and alterations of the coagulation response also participate in the vascular damage associated with diabetes. Here, we present an updated interpretation of the molecular mechanisms underlying vascular damage in diabetes, including current therapeutic options for its control.

An oral absorbent, AST-120, restores vascular growth and blood flow in ischemic muscles in diabetic mice via modulation of macrophage transition

BackgroundDiabetes has a pronounced effect on the peripheral vasculature. The accumulation of advanced glycation end products (AGEs) is regarded as the crucial mechanism responsible for vascular damage in diabetes, but it is not easy to be avoided from food. In this study, we aimed to investigate the effects of an oral absorbent, AST-120, on the accumulation of AGEs and changes in blood flow recovery in diabetic mice.MethodsThe mice were divided into four groups, wild-type (WT) mice without treatment, WT mice treated with 5% AST-120 mixed into pulverized chow, streptozotocin-induced diabetes mellitus (DM) mice, and DM mice treated with 5% AST-120. Six weeks after hind-limb ischemia surgery, blood flow reperfusion, histology, plasma AGE, and cytokine were examined. Bone marrow cells were cultured and derived into macrophages to evaluate the effects of AGEs on macrophage polarization.ResultsPlasma AGEs were significantly increased in diabetic mice. AST-120 could bind to AGEs and reduced their plasma concentrations. Histological analysis revealed fewer collateral vessels with corresponding impairment of blood flow recovery in diabetic mice. In these mice, AGE-positive and AGE receptor-positive macrophages were numerous in ischemic limbs compared with non- diabetic mice. In diabetic mice, macrophages in ischemic tissues demonstrated greater M1 polarization than M2 polarization; this pattern was reversed in the AST-120 treatment group. The change in macrophage polarization was associated with the corresponding expression of pro-inflammatory cytokines in the ischemic tissues. In cell cultures, AGEs triggered the transformation of bone marrow-derived macrophages into the M1 phenotype. The alterations in the polarization of macrophages were reversed after treatment with AST-120.ConclusionsOral administration of AST-120 decreased the serum levels of AGEs in diabetic mice and improved neovascularization of ischemic limbs. This benefit may be due to, at least partially, the alterations in macrophage polarization and the associated changes in inflammatory cytokines.

Photoreceptor cells and RPE contribute to the development of diabetic retinopathy.

Diabetic retinopathy (DR) is a leading cause of blindness. It has long been regarded as vascular disease, but work in the past years has shown abnormalities also in the neural retina. Unfortunately, research on the vascular and neural abnormalities have remained largely separate, instead of being integrated into a comprehensive view of DR that includes both the neural and vascular components. Recent evidence suggests that the most predominant neural cell in the retina (photoreceptors) and the adjacent retinal pigment epithelium (RPE) play an important role in the development of vascular lesions characteristic of DR. This review summarizes evidence that the outer retina is altered in diabetes, and that photoreceptors and RPE contribute to retinal vascular alterations in the early stages of the retinopathy. The possible molecular mechanisms by which cells of the outer retina might contribute to retinal vascular damage in diabetes also are discussed. Diabetes-induced alterations in the outer retina represent a novel therapeutic target to inhibit DR.

Potential protective effect of vitamin D on aortic tissue of Streptozotocin-induced diabetic vascular damage in adult male rats

Background: Diabetes mellitus is a major risk factor of cardiovascular disease. There is evidence that vitamin D decreases type 1 diabetes mellitus risk during early adulthood and improves insulin secretion and resistance in diabetic patients. low vitamin D level was observed to increase the cardiovascular disease. Objectives: This study aimed to assess the protective effect of vitamin D on diabetic vascular damages in aorta. Materials and Methods: 40 adult male rats were randomly divided into: group I(control), group II (vitamin D), group III (diabetic) and groupVI (diabetic plus Vitamin D)(n = 10 rats /each group). Injection of streptozotocin (60 mg/kg) as a single dose intraperitoneal to induce diabetes. Vitamin D was administered orally every other day in a dose of 12.5 mg/kg. After 12weeksof treatment period, aortic samples were collected for histological examination. Results: Morphological changes of aortic tissue in diabetic untreated group in the form of reduction of tunica media thickness and areas of tunica intima detachment. The elastic lamellae became irregular, fragmented or branched. Shrunken dark or lysed nuclei of smooth muscle fibers were seen in tunica media of diabetic group. The diabetic treated group with vitamin D showed more or less normal structure of the layers of aortic tissuewith comparable thickness to the control group. Conclusion: Vitamin D may reduce the vascular complications and tissue injuries induced by diabetes in aorta. This effect has a positive influence on the function of the cardiovascular system. Keywords: Diabetes, Vitamin D, Aorta.

High glucose concentration produces a short-term increase in pERK1/2 and p85 proteins, having a direct angiogenetic effect by an action similar to VEGF.

Excessive glucose serum concentration, endothelial dysfunction and microangiopathy are key features of diabetes mellitus, being both diagnostic parameters and pathogenetic mechanisms. Vascular endothelial growth factor (VEGF) is importantly implicated in the physiology and pathology of blood vessels, including diabetic vascular damage. These factors certainly affect endothelial cells, and to evaluate mechanisms involved, we took advantage of telomerase-immortalized human microvascular endothelial (TIME) cells. TIME cells were exposed to different glucose concentrations and to VEGF treatments. Culture conditions also included the use of basement membrane extract, as an in vitro differentiation model. Cell morphology was then evaluated in the different conditions, and cellular proteins were extracted to analyze specific protein products by Western blot. High glucose concentrations and VEGF did substantially affect neither morphology nor growth of cultured TIME cells, while both considerably increased differentiation into "capillary-like" structures when cells were cultured on basement membrane extract. Under these conditions, high glucose concentration and VEGF also produced a short-term increase in pERK1/2 and p85 proteins, while total and phosphorylated AKT were not affected. These data suggest a direct angiogenetic effect of glucose, affecting intracellular transduction mechanisms with an action similar to that of VEGF. This effect on endothelial cell proliferation and differentiation could be part of pathogenetic mechanisms producing diabetic microvascular alterations.

Diabetic Vascular Damage: Review of Pathogenesis and Possible Evaluation Technologies

Diabetes mellitus (DM) is a rapidly increasing problem in health care worldwide: recent forecast indicates that the number of DM patients will rise to 640 million by 2040. Vascular damage is associated with severe complications, including cardiac neuropathy and limb amputation. Therefore, early prediction of diabetic vascular damage using advanced technologies is an important challenge because timely preventive and therapeutic measures could diminish the risk of development and burden of complications. The aim of the article is to provide a review of the initial stages of vascular damage and main mechanisms for development, as well as appropriate modern technologies for prediction and diagnosis. The manuscript provides an overview of interrelated vascular damage mechanisms influenced by diabetes, along with a review of possible technologies for early prediction and diagnosis. A comparative analysis of technologies appropriate for particular issues of prediction is summarised in the discussion.

Investigating RAGE Ectodomain Shedding (sRAGE) and Splice Variant (esRAGE) in Diabetic and Inflammatory Conditions

Obesity, diabetes and its complications represent an increasing worldwide health epidemic, however little is known about the specific cellular and molecular mechanisms that drive these complications. The defining features of obesity and diabetes are hyperlipidemia and hyperglycemia respectively, which create a cellular environment conducive to increased formation of advanced glycation end products (AGEs). AGEs bind to its chief cell surface receptor for AGE (RAGE) thereby activating downstream signaling pathways, including NF‐κB, which leads to the transcription of genes that in part create an inflammatory environment and ultimately contribute to the development of diabetic complications.RAGE has various splice variants and isoforms, a full length (fl) membrane bound RAGE (flRAGE), which can be cleaved from the cell surface by metalloprotease (MMP9) and ADAM10 to produce soluble RAGE (sRAGE). A second form of soluble RAGE is alternatively spliced and secreted by the cell known as endogenous secretory RAGE (esRAGE or RAGEv1). Both sRAGE and esRAGE act as a decoy to RAGE as they can bind to and sequester RAGE ligands, potentially preventing ligand binding to flRAGE. The objective of this study is to investigate RAGE ectodomain shedding and the splice variant of RAGE. Although studies have examined the regulation of RAGE shedding, little investigation into both sRAGE and esRAGE in cells exposed to stresses that provoke vascular damage in diabetes, such as hyperglycemia with the addition of an inflammatory setting has yet to be explored.We hypothesized that the glucose/AGE‐stimulated upregulation of inflammatory mechanisms might contribute to the generation of soluble RAGEs. Therefore, modulation of sRAGE and esRAGE levels in cells exposed to high glucose and to the prototypic inflammatory stimulus TNFα, were measured. RAGE expressing HEK293T cells exposed to diabetes‐relevant levels of high glucose, was not sufficient to induce changes in RAGE ectodomain shedding. However high glucose together with TNFα treatment resulted in a significant (p&lt;0.01) increase in RAGE ectodomain shedding as determined by ELISA. Western blot analysis under the same conditions showed that RAGE protein levels in the cell lysate remained unchanged indicating that the increase in ectodomain shedding was not due to increased cellular RAGE levels. The production of esRAGE remained unchanged when esRAGE expressing HEK293T cells were exposed to high glucose. Unlike RAGE ectodomain shedding, TNFα treatment alone resulted in a significant (p&lt;0.0001) increase in RAGE splice variant levels measured by esRAGE ELISA, which was also observed at the protein level by western blot analysis. These findings were also confirmed at the endogenous mRNA level using human monocyte‐like THP‐1 cells by qPCR. Both isoforms of soluble RAGE increased in the presence of stresses that mimic diabetic complications, with the addition of TNFα treatment being a key factor. Further mechanistic studies will elucidate the regulation of increased soluble RAGE production in these conditions with the goal of identifying novel therapeutic strategies to block adverse AGE‐RAGE consequences.Support or Funding InformationNYUAD Research Institute.Al Jalila Foundation.

Molecular Imaging of Retinal Endothelial Injury in Diabetic Animals.

Purpose:Diabetic retinopathy is a leading cause of vision loss. There is a great need for early diagnosis prior to the occurrence of irreversible structural damages. Expression of endothelial adhesion molecules is observed before the onset of diabetic vascular damage; however, to date, these molecules cannot be visualized in vivo.Methods:To quantify the expression of endothelial surface molecules, we generated imaging probes that bind to ICAM-1. The α-ICAM-1 probes were characterized via flow cytometry under microfluidic conditions. Probes were systemically injected into normal and diabetic rats, and their adhesion in the retinal microvessels was visualized via confocal scanning laser ophthalmoscopy. Histology was performed to validate in vivo imaging results. Vascular pathologies were visualized using trypsin-digested retinal preparations.Results:The α-ICAM-1 probes showed significantly higher adhesion to retinal microvessels in diabetic rats than in normal controls (P < 0.01), whereas binding of control probes did not differ between the two groups. Western blotting results showed higher ICAM-1 expression in retinas of T1D animals than in normal controls. Retinal endothelial ICAM-1 expression was observed via molecular imaging before markers of structural damage, such as pericyte ghosts and acellular capillaries.Conclusion:Results indicate that molecular imaging can be used to detect subtle changes in the diabetic retina prior to the occurrence of irreversible pathology. Thus, ICAM-1 could serve as a diagnostic target in patients with diabetes. This study provides a proof of principle for non-invasive subclinical diagnosis in experimental diabetic retinopathy. Further development of this technology could improve management of diabetic complications.

C-peptide protects against hyperglycemic memory and vascular endothelial cell apoptosis.

C-peptide exerts protective effects against diabetic complications; however, its role in inhibiting hyperglycemic memory (HGM) has not been elucidated. We investigated the beneficial effect of C-peptide on HGM-induced vascular damage in vitro and in vivo using human umbilical vein endothelial cells and diabetic mice. HGM induced apoptosis by persistent generation of intracellular ROS and sustained formation of ONOO(-) and nitrotyrosine. These HGM-induced intracellular events were normalized by treatment with C-peptide, but not insulin, in endothelial cells. C-peptide also inhibited persistent upregulation of p53 and activation of mitochondrial adaptor p66(shc) after glucose normalization. Further, C-peptide replacement therapy prevented persistent generation of ROS and ONOO(-) in the aorta of diabetic mice whose glucose levels were normalized by the administration of insulin. C-peptide, but not insulin, also prevented HGM-induced endothelial apoptosis in the murine diabetic aorta. This study highlights a promising role for C-peptide in preventing HGM-induced intracellular events and diabetic vascular damage.

The novel mitochondria-targeted hydrogen sulfide (H2S) donors AP123 and AP39 protect against hyperglycemic injury in microvascular endothelial cells in vitro

The development of diabetic vascular complications is initiated, at least in part, by mitochondrial reactive oxygen species (ROS) production in endothelial cells. Hyperglycemia induces superoxide production in the mitochondria and initiates changes in the mitochondrial membrane potential that leads to mitochondrial dysfunction. Hydrogen sulfide (H2S) supplementation has been shown to reduce the mitochondrial oxidant production and shows efficacy against diabetic vascular damage in vivo. However, the half-life of H2S is very short and it is not specific for the mitochondria. We have therefore evaluated two novel mitochondria-targeted anethole dithiolethione and hydroxythiobenzamide H2S donors (AP39 and AP123 respectively) at preventing hyperglycemia-induced oxidative stress and metabolic changes in microvascular endothelial cells in vitro. Hyperglycemia (HG) induced significant increase in the activity of the citric acid cycle and led to elevated mitochondrial membrane potential. Mitochondrial oxidant production was increased and the mitochondrial electron transport decreased in hyperglycemic cells. AP39 and AP123 (30–300nM) decreased HG-induced hyperpolarisation of the mitochondrial membrane and inhibited the mitochondrial oxidant production. Both H2S donors (30–300nM) increased the electron transport at respiratory complex III and improved the cellular metabolism. Targeting H2S to mitochondria retained the cytoprotective effect of H2S against glucose-induced damage in endothelial cells suggesting that the molecular target of H2S action is within the mitochondria. Mitochondrial targeting of H2S also induced >1000-fold increase in the potency of H2S against hyperglycemia-induced injury. The high potency and long-lasting effect elicited by these H2S donors strongly suggests that these compounds could be useful against diabetic vascular complications.

Methylglyoxal-Induced Endothelial Cell Loss and Inflammation Contribute to the Development of Diabetic Cardiomyopathy.

The mechanisms for the development of diabetic cardiomyopathy remain largely unknown. Methylglyoxal (MG) can accumulate and promote inflammation and vascular damage in diabetes. We examined if overexpression of the MG-metabolizing enzyme glyoxalase 1 (GLO1) in macrophages and the vasculature could reduce MG-induced inflammation and prevent ventricular dysfunction in diabetes. Hyperglycemia increased circulating inflammatory markers in wild-type (WT) but not in GLO1-overexpressing mice. Endothelial cell number was reduced in WT-diabetic hearts compared with nondiabetic controls, whereas GLO1 overexpression preserved capillary density. Neuregulin production, endothelial nitric oxide synthase dimerization, and Bcl-2 expression in endothelial cells was maintained in the hearts of GLO1-diabetic mice and corresponded to less myocardial cell death compared with the WT-diabetic group. Lower receptor for advanced glycation end products and tumor necrosis factor-α (TNF-α) levels were also observed in GLO1-diabetic versus WT-diabetic mice. Over a period of 8 weeks of hyperglycemia, GLO1 overexpression delayed and limited the loss of cardiac function. In vitro, MG and TNF-α were shown to synergize in promoting endothelial cell death, which was associated with increased angiopoietin 2 expression and reduced Bcl-2 expression. These results suggest that MG in diabetes increases inflammation, leading to endothelial cell loss. This contributes to the development of diabetic cardiomyopathy and identifies MG-induced endothelial inflammation as a target for therapy.

Aflibercept for the treatment of diabetic macular edema.

Diabetic macular edema (DME) is an accumulation of fluid in the central retina, secondary to vascular-leakage from diabetic vascular damage. DME and other ophthalmic sequela of diabetes are the leading cause of blindness in 20 to 74-year-olds. The development of VEGF-inhibitors (anti-VEGF) has revolutionized DME treatment improving the clinician's ability to remove excess fluid from the macula, improving visual-acuity. Aflibercept is an anti-VEGF agent made of a recombinant fusion protein (consisting of VEGF receptors 1 and 2 extracellular domains) fused with the Fc-portion of human-IgG1, which binds both VEGF isoforms A and B, and placental growth factor. Phase III clinical trials and published scientific studies have demonstrated the efficacy of intravitreal aflibercept injection in the treatment of DME.