Retinal Microvascular Complications Research Articles

Is Pregnancy a Risk Factor for Progression of Diabetic Retinopathy and Nephropathy in Type 1 Diabetes? A Matched Cohort Study

Objective This study aimed to test the hypothesis that the development or deterioration of nephropathy and retinopathy over time is not affected by pregnancy in women with pregestational type 1 diabetes mellitus (T1DM). Study design Prospective, observational study of nephropathy and retinopathy follow-up during pregnancy and in a subsequent period of 2 years in a group of pregnant women with T1DM (study group) that we compared with pair-matched non-pregnant women with T1DM (control group) who underwent similar intensive follow-up. Results The rate of renal microvascular complications was similar at entry, 17.4% (4/23) in the study group and 21.7% (5/23) in the control group. At the last visit, both groups had nephropathy rates of 17.4% (4/23) and paired p-value of 1.00. Similarly, the rate of retinal microvascular complications of any grade was similar in both groups and remained so at the last follow-up examination. Conclusion Pregnancy per se does not appear to increase the risk for the development of, or the acceleration of the progression of retinopathy and nephropathy during a follow-up of at least 2 years in relatively healthy T1DM patients. This information is important for counseling young women with T1DM who are considering becoming pregnant. Key Points

TRNA-derived fragment tRF-30 propels diabetes-induced retinal microvascular complications by regulating STAT3 signaling.

Transfer RNA-derived fragments (tRFs) represent a novel class of non-coding RNA transcripts that possess specific biological functions. However, the involvement of tRFs in retinal microvascular diseases remains poorly understood. In this study, we aimed to reveal whether modulation of tRF-30 expression could attenuate pathological retinal neovascular diseases. Our findings demonstrate a significant upregulation of tRF-30 expression levels in both in vivo models of diabetic retinopathy (DR) and in vitro endothelial sprouting models. Conversely, inhibition of tRF-30 expression suppressed the formation of abnormal neovascularization in the retina in vivo, while reducing the proliferation and migration activity of retinal vascular endothelial cells in vitro. We also found that tRF-30 modulates retinal neovascularization through the tRF-30/TRIB3/signal transducer and activated transcription 3 signaling pathway. Furthermore, we validated a significant upregulation of tRF-30 expression levels in the vitreous humor of DR patients, with high levels of both validity and specificity in diagnostic testing. Collectively, our findings highlight a pro-angiogenic role for tRF-30 in DR. Intervening in the tRF-30 signaling pathway may represent a promising prevention and treatment strategy for retinal angiogenesis.

417-P: Diabetic Foot Ulcer Amputation Increases the Risk of Severe Renal and Retinal Microvascular Complications—A Korean National Health Information Database Study

417-P: Diabetic Foot Ulcer Amputation Increases the Risk of Severe Renal and Retinal Microvascular Complications—A Korean National Health Information Database Study

TRNA-derived fragment tRF-1020 ameliorates diabetes-induced retinal microvascular complications.

Transfer RNA (tRNA)‐derived fragments are the non‐coding single‐stranded RNAs involved in several physiological and pathological processes. Herein, we investigated the role of tRF‐1020, a tRNA fragment, in diabetes‐induced retinal microvascular complications. The results showed that the levels of tRF‐1020 expression were down‐regulated in diabetic retinal vessels and retinal endothelial cells following high glucose or H2O2 stress. Overexpressing tRF‐1020 led to decreased endothelial cell viability, proliferation, migration, and tube formation and alleviated retinal vascular dysfunction as shown by decreased retinal acellular capillaries, vascular leakage, and inflammation. By contrast, tRF‐1020 silencing displayed the opposite effects. tRF‐1020 regulated endothelial angiogenic functions and retinal vascular dysfunction by targeting Wnt signalling. Moreover, the levels of tRF‐1020 expression were reduced in aqueous humour and vitreous samples of the patients with diabetic retinopathy. Collectively, tRF‐1020 is a potential target for the diagnosis and treatment of diabetic retinopathy.

Comprehensive Proteomic Profiling of Aqueous Humor Proteins in Proliferative Diabetic Retinopathy.

PurposeProliferative diabetic retinopathy (PDR) is a serious ocular disease that can lead to retinal microvascular complications in patients with diabetes mellitus. To date, no studies have explored PDR development by analyzing the aqueous humor (AH). Therefore we carried out tandem mass tag (TMT) proteomic quantification to compare AH protein profiles between PDR and non-PDR subjects.MethodsWe enrolled six PDR and six control (senile cataract) subjects. AH samples were collected during surgery and stored at –80°C. Proteins were extracted, trypsin-digested, and labeled with TMTs for mass spectrometric analysis.ResultsWe found 191 proteins to be changed with |log2 (fold change)| ≥1 (P < 0.05 and identification with at least two peptides per protein). Of them, 111 were downregulated, whereas 80 were upregulated in the PDR group. Proteomic bioinformatic analysis indicated that PDR development was related to complement and coagulation cascades, platelet activation, extracellular matrix–receptor interaction, focal adhesion, protein digestion and absorption, human papillomavirus infection, PI3K-Akt signaling pathway, cholesterol metabolism, peroxisome proliferator-activated receptor signaling pathways, fat digestion and absorption, and vitamin digestion and absorption pathways.ConclusionsComprehensive proteomic profiling of the AH revealed 191 differentially expressed proteins between the two groups. Most of these proteins belong to secretory pathways, and therefore can be used as biomarkers in clinical testing and basic research.Translational RelevancePathway analysis and a review of the literature enabled us to draw a novel biological map that will support further studies on the underlying mechanisms and therapeutic control of PDR development.

Characterization of Retinal Microvascular Complications and the Effects of Endoplasmic Reticulum Stress in Mouse Models of Diabetic Atherosclerosis.

PurposeRecent evidence suggests that there is a correlation between the micro- and macrovascular complications of diabetes mellitus. The aim of this study is to investigate the molecular mechanisms by which diabetes promotes the development of microvascular disease (diabetic retinopathy [DR]) through characterization of the effects of hyperglycemia in the retina of mouse models of diabetic atherosclerosis.MethodsHyperglycemia was induced in apolipoprotein E–deficient (ApoE–/–) mice, a model of accelerated atherosclerosis, either through streptozotocin (STZ) injection or introduction of the Ins2Akita mutation (ApoE–/–Ins2+/Akita). Another subset of ApoE–/– mice was supplemented with glucosamine (GlcN). To attenuate atherosclerosis, subsets of mice from each experimental group were treated with the chemical chaperone, 4-phenylbutyric acid (4PBA). Eyes from 15-week-old mice were either trypsin digested and stained with periodic acid–Schiff (PAS) or frozen for cryostat sectioning and immunostained for endoplasmic reticulum (ER) stress markers, including C/EBP homologous protein (CHOP) and 78-kDa glucose-regulated protein (GRP78). PAS-stained retinal flatmounts were analyzed for microvessel density, acellular capillaries, and pericyte ghosts.ResultsFeatures of DR, including pericyte ghosts and reduced microvessel density, were observed in hyperglycemic and GlcN-supplemented mice. Treatment with 4PBA reduced ER stress in the retinal periphery and attenuated DR in the experimental groups.ConclusionsMouse models of diabetic atherosclerosis show characteristic pathologies of DR that correlate with atherosclerosis. The increased magnitude of these changes and responses to 4PBA in the peripheral retina suggest that future studies should be aimed at assessing regional differences in mechanisms of ER stress-related pathways in these mouse models.

Disrupted topological organization of human brain connectome in diabetic retinopathy patients.

ObjectiveThere is increasing neuroimaging evidence that type 2 diabetes patients with retinal microvascular complications show abnormal brain functional and structural architecture and are at an increased risk of cognitive decline and dementia. However, changes in the topological properties of the functional brain connectome in diabetic retinopathy (DR) patients remain unknown. The aim of this study was to explore the topological organization of the brain connectome in DR patients using graph theory approaches.MethodsThirty-five DR patients (18 males and 17 females) and 38 healthy controls (HCs) (18 males and 20 females), matched for age, sex, and education, underwent resting-state magnetic resonance imaging scans. Graph theory analysis was performed to investigate the topological properties of brain functional connectome at both global and nodal levels.ResultsBoth DR and HC groups showed high-efficiency small-world network in their brain functional networks. Notably, the DR group showed reduction in the clustering coefficient (P=0.0572) and local efficiency (P=0.0151). Furthermore, the DR group showed reduced nodal centralities in the default-mode network (DMN) and increased nodal centralities in the visual network (VN) (P<0.01, Bonferroni-corrected). The DR group also showed abnormal functional connections among the VN, DMN, salience network (SN), and sensorimotor network (SMN). Altered network metrics and nodal centralities were significantly correlated with visual acuity and fasting blood glucose level in DR patients.ConclusionDR patients showed abnormal topological organization of the human brain connectome. Specifically, the DR group showed reduction in the clustering coefficient and local efficiency, relative to HC group. Abnormal nodal centralities and functional disconnections were mainly located in the DMN, VN, SN, and SMN in DR patients. Furthermore, the disrupted topological attributes showed correlations with clinical variables. These findings offer important insight into the neural mechanism of visual loss and cognitive deficits in DR patients.

Metformin suppresses retinal angiogenesis and inflammation in vitro and in vivo.

The oral anti-diabetic drug metformin has been found to reduce cardiovascular complications independent of glycemic control in diabetic patients. However, its role in diabetic retinal microvascular complications is not clear. This study is to investigate the effects of metformin on retinal vascular endothelium and its possible mechanisms, regarding two major pathogenic features of diabetic retinopathy: angiogenesis and inflammation. In human retinal vascular endothelial cell culture, metformin inhibited various steps of angiogenesis including endothelial cell proliferation, migration, and tube formation in a dose-dependent manner. Its anti-angiogenic activity was confirmed in vivo that metformin significantly reduced spontaneous intraretinal neovascularization in a very-low-density lipoprotein receptor knockout mutant mouse (p<0.05). Several inflammatory molecules upregulated by tumor necrosis factor-α in human retinal vascular endothelial cells were markedly reduced by metformin, including nuclear factor kappa B p65 (NFκB p65), intercellular adhesion molecule-1 (ICAM-1), monocyte chemotactic protein-1 (MCP-1), and interleukin-8 (IL-8). Further, metformin significantly decreased retinal leukocyte adhesion (p<0.05) in streptozotocin-induced diabetic mice. Activation of AMP-activated protein kinase was found to play a partial role in the suppression of ICAM-1 and MCP-1 by metformin, but not in those of NFκB p65 and IL-8. Our findings support the notion that metformin has considerable anti-angiogenic and anti-inflammatory effects on retinal vasculature. Metformin could be potentially used for the purpose of treating diabetic retinopathy in addition to blood glucose control in diabetic patients.

Association of Skin Intrinsic Fluorescence with Retinal Microvascular Complications of Long Term Type 1 Diabetes in the Wisconsin Epidemiologic Study of Diabetic Retinopathy

ABSTRACTObjective: To determine the association between skin intrinsic fluorescence (SIF), a noninvasive measure of advanced glycation endproducts and oxidative stress in skin, and retinal microvascular complications of long duration type 1 diabetes, proliferative diabetic retinopathy (PDR) and macular edema.Methods: A cross-sectional cohort study of persons with type 1 diabetes in the Wisconsin Epidemiologic Study of Diabetic Retinopathy (WESDR) who participated in a 32-year follow-up examination in 2012–2014. Subjects underwent a physical examination, answered a health questionnaire, and had fundus photographs taken. SIF was measured on the underside of the left forearm near the elbow with the SCOUT DS® skin fluorescence spectrometer. Two representative SIF measures were used for these analyses: SIF01 excited by an LED centered at 375 nm with correction factors Kx = 0.6 and Km = 0.2 and SIF15 excited by an LED centered at 456 nm with correction factors Kx = 0.4 and Km = 0.9.Results: The 414 participants had mean diabetes duration of 42.2 years (standard deviation 6.8 years, range 32.9–67.9 years). PDR was statistically significantly associated (p < 0.05) with both SIF measures in multivariate models including other relevant factors (odds ratio [OR] = 1.17 for SIF01 and 1.20 for SIF15).Conclusion: Skin intrinsic fluorescence measures are independently associated with PDR in the WESDR. Incidence information is needed to evaluate whether there is a causal relationship.

Differential Modulation of Circulating Endothelial Progenitor Cells in Peripheral Blood Monocyte Population of an Early Stage of Streptozotocin-Induced Diabetic Retinopathy

Objective: To enumerate the circulating endothelial progenitor cells (EPCs) in peripheral blood mononuclear cells (PBMC) in streptozotocin (STZ) induced diabetic retinopathy (DR) rat model. Materials and Methods: Retinal microvascular complications in STZ induced diabetic Wistar rats were validated after 4 months using trypsin digestion assay of retinal vasculature, histological analysis of retina, glial fibrillary acidic protein (GFAP) immunostaining. Subsequently, circulating EPCs were quantified in PBMCs from diabetes induced rats after 4 months using markers such as CD-34, CD-31, CD-146, VE-Cadherin and Flk-1 by flow cytometry. Results: After 4 months of diabetes induction, pericyte loss, acellular capillaries, retinal layer thickness, microglial activation and increased nitric oxide (NO) levels were observed. While the EPC markers such as CD-34, CD-31, VE-Cadherin were decreased, the Flk-1 +ve cells were up-regulated in the endogenous EPC pool. Conclusion: The quantity of EPCs are modulated in diabetes which may provide an insight into the use of EPCs as a biomarker for the progression of diabetes induced microvascular complication. Key words: Diabetic retinopathy, Endothelial progenitor cells, FlK-1, GFAP, Streptozotocin.

Bloodstream-To-Eye Infections Are Facilitated by Outer Blood-Retinal Barrier Dysfunction.

The blood-retinal barrier (BRB) functions to maintain the immune privilege of the eye, which is necessary for normal vision. The outer BRB is formed by tightly-associated retinal pigment epithelial (RPE) cells which limit transport within the retinal environment, maintaining retinal function and viability. Retinal microvascular complications and RPE dysfunction resulting from diabetes and diabetic retinopathy cause permeability changes in the BRB that compromise barrier function. Diabetes is the major predisposing condition underlying endogenous bacterial endophthalmitis (EBE), a blinding intraocular infection resulting from bacterial invasion of the eye from the bloodstream. However, significant numbers of EBE cases occur in non-diabetics. In this work, we hypothesized that dysfunction of the outer BRB may be associated with EBE development. To disrupt the RPE component of the outer BRB in vivo, sodium iodate (NaIO3) was administered to C57BL/6J mice. NaIO3-treated and untreated mice were intravenously injected with 108 colony forming units (cfu) of Staphylococcus aureus or Klebsiella pneumoniae. At 4 and 6 days postinfection, EBE was observed in NaIO3-treated mice after infection with K. pneumoniae and S. aureus, although the incidence was higher following S. aureus infection. Invasion of the eye was observed in control mice following S. aureus infection, but not in control mice following K. pneumoniae infection. Immunohistochemistry and FITC-dextran conjugate transmigration assays of human RPE barriers after infection with an exoprotein-deficient agr/sar mutant of S. aureus suggested that S. aureus exoproteins may be required for the loss of the tight junction protein, ZO-1, and for permeability of this in vitro barrier. Our results support the clinical findings that for both pathogens, complications which result in BRB permeability increase the likelihood of bacterial transmigration from the bloodstream into the eye. For S. aureus, however, BRB permeability is not required for the development of EBE, but toxin production may facilitate EBE pathogenesis.

Photooxidation of glycated and non‐glycated phosphatidylethanolamines monitored by mass spectrometry

Phosphatidylethanolamines (PE) are one of the major components of cells membranes, namely in skin and in retina, that are continuously exposed to solar UV radiation being major targets of photooxidation damage. In addition, due to the presence of the free amine group, PE can also undergo glycation, in hyperglycemic conditions which may increase the susceptibility to oxidation. The aim of this study is to develop a model, based on mass spectrometry (MS) analysis, to identify photooxidative degradation of selected PE (POPE: PE 16:0/18:1, PLPE: PE 16:0/18:2, PAPE: PE 16:0/20:4) and glycated PEs due to UV irradiation. Photooxidation products were analysed by electrospray ionization MS (ESI-MS) and tandem MS (ESI-MS/MS) in positive and negative mode. Emphasis is placed in the influence of glycation in the generation of distinct photooxidation products. ESI-MS spectra of PE after UV photo-irradiation showed mainly hydroperoxy derivatives, due to oxidation of unsaturated fatty acyl chains. Glycated PE gave rise to several new photooxidation products formed due to oxidative cleavages of the glucose moiety, namely between C1 and C2, C2 and C3, and C5 and C6 of this sugar unit. These new products were identified by ESI-MS/MS in positive mode showing distinct neutral loss depending on the different structure of the polar head group. These new identified advanced glycated photooxidation products may have a deleterious role in the etiology of diabetic retinopathy and in diabetic retinal microvascular complications.

P122 * Relationship between retinal microvascular complications in metabolic syndrome (MS) and its association with serum uric acid level

P122 * Relationship between retinal microvascular complications in metabolic syndrome (MS) and its association with serum uric acid level

A PKC-β inhibitor treatment reverses cardiac microvascular barrier dysfunction in diabetic rats

The PKC-β inhibitor ruboxistaurin (RBX or LY333531) prevents diabetic renal and retinal microvascular complications. However, the effect of RBX on diabetic cardiac microvascular dysfunction is still unclear. In this study, we aimed to investigate the effects and mechanisms of RBX treatment upon cardiac endothelial barrier dysfunction in high glucose states. We demonstrated RBX treatment suppressed high glucose induced PKC-βII activation and phosphorylation of β-catenin in vivo and in vitro experiments. Meanwhile, RBX treatment protected cardiac microvascular barrier function in diabetic animals and monolayer barrier function of cultured cardiac microvascular endothelial cells (CMECs), reproducing the same effect as PKC-βII siRNA. These results provide new insight into protective properties of PKC-β inhibitor against cardiac endothelial barrier dysfunction. PKC-β inhibitor RBX prevented chronic cardiac microvascular barrier dysfunction and improved endothelial cell–cell junctional function in high glucose states.

Kidney disease in diabetology: lessons from 2008

Microand macroalbuminuria are important markers for early and progressive diabetic kidney disease. Patients with type 1 diabetes carry a 20–50% risk of developing end-stage renal disease (ESRD) requiring dialysis or renal transplantation. The primary goal of managing childhood type 1 diabetes is to prevent or delay renal and retinal microvascular complications. In contrast to patients with type 2 diabetes, the cumulative incidence of nephropathy has significantly declined in patients with type 1 diabetes over the past three decades, which was due to more intensified treatment regimens for control of hyperglycaemia, hypertension and dyslipidaemia. However, many patients with type 1 diabetes have their manifestation of disease already at very young age, when strict diabetes control is more difficult to reach compared to patients with manifestation of diabetes later in life.

Vascular Endothelial Growth Factor and Diabetic Retinopathy: Role of Oxidative Stress

Retinal neovascularization and macular edema are central features of diabetic retinopathy, a major cause of blindness in working age adults. The currently established treatment for diabetic retinopathy targets the vascular pathology by laser photocoagulation. This approach is associated with significant adverse effects due the destruction of neural tissue and is not always effective. Characterization of the molecular and cellular processes involved in vascular growth and hyperpermeability has led to the recognition that the angiogenic growth factor and vascular permeability factor VEGF (vascular endothelial growth factor) play a pivotal role in the retinal microvascular complications of diabetes. Thus, VEGF represents an important target for therapeutic intervention in diabetic retinopathy. Agents that directly inhibit the actions of VEGF and its receptors show considerable promise, but have not proven to be completely effective in blocking pathological angiogenesis. Therefore, a better understanding of the molecular events that control VEGF expression and mediate its downstream actions is important to define more precise therapeutic targets for intervention in diabetic retinopathy. This review highlights the current understanding of the process by which VEGF gene expression is regulated and how VEGF's biological effects are altered during diabetes. In particular, cellular and molecular alterations seen in diabetic models are considered in the context of high glucose-mediated oxidative stress effects on VEGF expression and action. Potential therapeutic strategies for preventing VEGF overexpression or blocking its pathological actions in the diabetic retina are considered.

Vascular endothelial growth factor and diabetic retinopathy: pathophysiological mechanisms and treatment perspectives.

Retinal neovascularization and macular edema are central features of diabetic retinopathy, the major cause of blindness in the developed world. Current treatments are limited in their efficacy and are associated with significant adverse effects. Characterization of the molecular and cellular processes involved in vascular growth and permeability has led to the recognition that the angiogenic growth factor and vascular permeability factor vascular endothelial growth factor (VEGF) plays a pivotal role in the retinal microvascular complications of diabetes. Therefore, VEGF represents an exciting target for therapeutic intervention in diabetic retinopathy. This review highlights the current understanding of the mechanisms that regulate VEGF gene expression and mediate its biological effects and how these processes may become altered during diabetes. The cellular and molecular alterations that characterize experimental models of diabetes are considered in relation to the influence of high glucose-mediated oxidative stress on VEGF expression and on the mechanisms of VEGF's actions under hyperglycemic induction. Finally, potential therapeutic strategies for preventing VEGF overexpression or blocking its pathological effects in the diabetic retina are considered.

Optic disc and retinal microvasculopathy after high-dose chemotherapy and autologous hematopoietic progenitor cell support.

The purpose of this study was to prospectively evaluate the retinal and optic nerve changes in patients undergoing high-dose chemotherapy (HDC) followed by autologous hematopoietic progenitor cell support (AHPCS). One hundred and forty patients undergoing HDC and AHPCS underwent extensive pre- and post-transplant ophthalmologic evaluations for development of retinal microvascular complications. One hundred and ten patients received high-dose cyclophosphamide, cisplatin and BCNU; thirty received identical doses of cyclophosphamide and cisplatin, but received paclitaxel instead of BCNU. Thirty-four patients (24%) had retinal findings of either cotton wool spots (CWS) (n = 20) or retinal hemorrhages (n = 18) during follow-up, which ranged from 1 to 12 months. Ten (7%) of these patients, all of whom received BCNU, showed ocular toxicity characterized by CWS 1 to 4 months post transplant (n = 10); optic disc edema (n = 3); and variable vision loss associated with the onset of BCNU-induced pulmonary toxicity. Retinal and optic disc microvascular complications may occur after high-dose chemotherapy followed by AHPCS. The association of ischemic retinal lesions and/or optic disc edema with BCNU-induced pulmonary toxicity and the lack of ocular toxicity in patients that did not receive BCNU may suggest that BCNU is the etiologic agent.

Dysregulation of the hypothalamo-pituitary-adrenal axis and duration of diabetes

We compared insulin-dependent diabetic outpatients with and without retinopathy for plasma indices of hypothalamo-pituitary-adrenal (HPA) axis activity. Diabetic patients with moderate-to-severe retinopathy had significantly higher postdexamethasone plasma levels of adrenocorticotropic hormone than patients with minimal or no retinopathy. However, when duration of diabetes was taken into account this difference was no longer significant. These data suggest that dysregulation of the HPA axis and retinal microvascular complications found in diabetic patients may both be a function of duration of diabetes.