Endothelium-derived agents in pericyte function/dysfunction

The major components of blood vessels are the vascular endothelium and its supporting smooth muscle. Significant strides have been made in the understanding of the cellular and molecular biology of these two cell types and in particular their interactions have been the subject of much interest and debate over the past two decades. The vascular endothelium is now known to profoundly influence the synthetic and motor functions of the underlying smooth muscle and participate in the pathogenesis of all the major vascular disorders. Similarly, the vascular smooth muscle has important effects on the overlying endothelium, and any disruption in the cellular physiology of either cell type can result in dysfunction with important effects on blood flow and vascular permeability The majority of this accumulated knowledge relates to the vascular cells of the macrocirculation. Pericytes are the supporting cells of the microvasculature and a body of evidence is now available to show that similar regulatory mechanisms and vessel-wall cross-talk exists between these cells and the microvascular endothelium. Nowhere are these interactions more important than in the retinal microcirculation where autoregulation is vital for the maintenance of smooth and uninterrrupted blood flow. This review focuses on the interactions between retinal microvascular endothelial cells and their associated pericytes and examines the role of the endothelial cell and the pericyte in the pathogenesis of disease.

Retinal ultrastructural findings in cone degeneration

We studied an eye from a 73-year-old man with a sporadic type of retinal cone degeneration and choroidal melanoma. Histologic and ultrastructural studies of the nasal retina unaffected by the choroidal melanoma showed alterations at the outer retina predominantly involving the photoreceptors and retinal pigment epithelium. A wide spectrum of pathologic changes were observed, ranging from near normal retina showing only photoreceptor outer segment disease (distortion and kinking) to grossly pathologic regions where photoreceptor cell bodies were sparse and their outer segments absent. The retinal pigment epithelium in minimally affected regions of the retina showed an increased proportion of the melanin complement of the cell within complex granules. In severe disease, many cells showed only giant complex granules with no free melanin. Retinal pigment epithelial cell migration and relocation around blood vessels was also noted in severe disease.

Endocytosis in the retinal and choroidal microcirculation

The endocytosis of horseradish peroxidase (HRP) by the vascular cells of retinal and choroidal blood vessels was compared in immersion and perfusion fixed eyes from individual rats. The mechanisms of endocytosis of HRP appeared identical in both retinal and choroidal vessels. The bulk of internalised tracer occurred in macropinosomes 300-400 nm in diameter. Tracer was localised to a 20-30 nm layer on the internal aspect of the limiting membrane. This layer was coincident with the glycocalyx of the luminal plasma membrane as revealed by ruthenium redosmium tetroxide staining. Horseradish peroxidase was also internalised by a small scattered population of vesicles (100-130 nm in diameter). The size of these vesicles suggested that they may have arisen from clathrin coated regions of the plasma membrane. It is suggested that the endocytosis of HRP in retinal and choroidal vascular endothelium occurs as a function of plasma membrane recycling. Horseradish peroxidase may also be internalised as a 'contaminant' of the glycocalyx in coated pits involved in receptor mediated endocytosis. The smooth 80 nm plasmalemmal caveolae of the retinal and choroidal vascular endothelial cells did not appear to participate either in absorptive endocytosis or vesicular transport.

Acridine orange leukocyte fluorography in mice

Simultaneous non-invasive visualization of blood vessels and nerves in patients can be obtained in the eye. The retinal vasculature is a target of many retinopathies. Inflammation, readily manifest by leukocyte adhesion to the endothelial lining, is a key pathophysiological mechanism of many retinopathies, making it a valuable and ubiquitous target for disease research. Leukocyte fluorography has been extensively used in the past twenty years; however, fluorescent markers, visualization techniques, and recording methods have differed between studies. The lack of detailed protocol papers regarding leukocyte fluorography, coupled with lack of uniformity between studies, has led to a paucity of standards for leukocyte transit (velocity, adherence, extravasation) in the retina. Here, we give a detailed description of a convenient method using acridine orange (AO) and a commercially available scanning laser ophthalmoscope (SLO, HRA-OCT Spectralis) to view leukocyte behavior in the mouse retina. Normal mice are compared to mice with acute and chronic inflammation. This method can be readily adopted in many research labs.

Pathophysiology and Pathogenesis of Diabetic Retinopathy

Diabetic retinopathy is traditionally viewed as a disease of the retinal blood vessels, although there is increasing evidence that retinal neurons and glial cells are also affected. This article describes the changes in the diabetic retina that precede the development of clinical diabetic retinopathy, including changes in the rate of retinal blood flow, alterations in the electroretinogram and breakdown of the integrity of the blood-retinal barrier. The long term lesions of diabetic retinopathy are characterised by a complex array of vasodegenerative changes that lead directly to areas of retinal ischaemia. This frequently triggers the onset of macular oedema and/or the proliferative stages of diabetic retinopathy with risk of visual impairment and blindness. Neurodegeneration has also been reported in the retina during both human and experimental diabetic retinopathy, although presently it remains unclear to what extent such changes contribute to visual loss in diabetic retinopathy.

Calcification and ossification of the convexity of the falx cerebri and related subdural space in human cadavers

OBJECTIVE: The present work was planned to report the incidence of calcification and ossification of an isolated cranial dural fold. The form, degree of severity and range of extension of such changes will be described. Involvement of the neighboring brain tissue and blood vessels, whether meningeal or cerebral, will also be determined. The results of this study might highlight the occasional incidence of intracranial calcification and ossification in images of the head and their interpretation, by radiologists and neurologists, to be of dural or vascular origin.

METHODS: Two human formalin-fixed cadavers, one middle-aged female another older male, were investigated at the Anatomy Laboratory, College of Medicine, King Faisal University, Dammam, Kingdom of Saudi Arabia during the period from 2000 to 2003. In each cadaver, the skullcap was removed and the convexity of the cranial dura mater, as well as the individual dural folds, were carefully examined for any calcification or ossification. The meningeal and cerebral blood vessels together with the underlying brain were grossly inspected for such structural changes. Calcified or ossified tissues, when identified, were subjected to histological examination to confirm their construction.

RESULTS: The female cadaver showed a calcified parietal emissary vein piercing the skullcap and projecting into the scalp. The latter looked paler and deficient in hair on its right side. The base of the stump was surrounded by a granular patch of calcification. The upper convex border of the falx cerebri was hardened and it presented granules, plaques and a cauliflower mass, which all proved to be osseous in structure. The meningeal and right cerebral vessels were mottled with calcium granules. The underlying temporal and parietal lobes of the right cerebral hemisphere were degenerated. The male cadaver also revealed a calcified upper border of the falx cerebri and superior sagittal sinus. Osseous granules and plaques, similar to those of the first specimen, were also identified but without gross changes in the underlying brain.

CONCLUSION: Calcification or ossification of an isolated site of the cranial dura mater and the intracranial blood vessels might occur. These changes should be kept in mind while interpreting images of the skull and brain. Clinical assessment and laboratory investigations are required to determine whether these changes are idiopathic, traumatic, or as a manifestation of a generalized disease such as hyperparathyroidism, vitamin D-intoxication, or chronic renal failure.

Optical coherence tomography for the diagnosis, monitoring and guiding of treatment for neovascular age-related macular degeneration:a systematic review and economic evaluation

BACKGROUND: Age-related macular degeneration is the most common cause of sight impairment in the UK. In neovascular age-related macular degeneration (nAMD), vision worsens rapidly (over weeks) due to abnormal blood vessels developing that leak fluid and blood at the macula.

OBJECTIVES: To determine the optimal role of optical coherence tomography (OCT) in diagnosing people newly presenting with suspected nAMD and monitoring those previously diagnosed with the disease.

DATA SOURCES: Databases searched: MEDLINE (1946 to March 2013), MEDLINE In-Process & Other Non-Indexed Citations (March 2013), EMBASE (1988 to March 2013), Biosciences Information Service (1995 to March 2013), Science Citation Index (1995 to March 2013), The Cochrane Library (Issue 2 2013), Database of Abstracts of Reviews of Effects (inception to March 2013), Medion (inception to March 2013), Health Technology Assessment database (inception to March 2013).

REVIEW METHODS: Types of studies: direct/indirect studies reporting diagnostic outcomes.

INDEX TEST: time domain optical coherence tomography (TD-OCT) or spectral domain optical coherence tomography (SD-OCT).

COMPARATORS: clinical evaluation, visual acuity, Amsler grid, colour fundus photographs, infrared reflectance, red-free images/blue reflectance, fundus autofluorescence imaging, indocyanine green angiography, preferential hyperacuity perimetry, microperimetry. Reference standard: fundus fluorescein angiography (FFA). Risk of bias was assessed using quality assessment of diagnostic accuracy studies, version 2. Meta-analysis models were fitted using hierarchical summary receiver operating characteristic curves. A Markov model was developed (65-year-old cohort, nAMD prevalence 70%), with nine strategies for diagnosis and/or monitoring, and cost-utility analysis conducted. NHS and Personal Social Services perspective was adopted. Costs (2011/12 prices) and quality-adjusted life-years (QALYs) were discounted (3.5%). Deterministic and probabilistic sensitivity analyses were performed.

RESULTS: In pooled estimates of diagnostic studies (all TD-OCT), sensitivity and specificity [95% confidence interval (CI)] was 88% (46% to 98%) and 78% (64% to 88%) respectively. For monitoring, the pooled sensitivity and specificity (95% CI) was 85% (72% to 93%) and 48% (30% to 67%) respectively. The FFA for diagnosis and nurse-technician-led monitoring strategy had the lowest cost (£39,769; QALYs 10.473) and dominated all others except FFA for diagnosis and ophthalmologist-led monitoring (£44,649; QALYs 10.575; incremental cost-effectiveness ratio £47,768). The least costly strategy had a 46.4% probability of being cost-effective at £30,000 willingness-to-pay threshold.

LIMITATIONS: Very few studies provided sufficient information for inclusion in meta-analyses. Only a few studies reported other tests; for some tests no studies were identified. The modelling was hampered by a lack of data on the diagnostic accuracy of strategies involving several tests.

CONCLUSIONS: Based on a small body of evidence of variable quality, OCT had high sensitivity and moderate specificity for diagnosis, and relatively high sensitivity but low specificity for monitoring. Strategies involving OCT alone for diagnosis and/or monitoring were unlikely to be cost-effective. Further research is required on (i) the performance of SD-OCT compared with FFA, especially for monitoring but also for diagnosis; (ii) the performance of strategies involving combinations/sequences of tests, for diagnosis and monitoring; (iii) the likelihood of active and inactive nAMD becoming inactive or active respectively; and (iv) assessment of treatment-associated utility weights (e.g. decrements), through a preference-based study.

STUDY REGISTRATION: This study is registered as PROSPERO CRD42012001930.

FUNDING: The National Institute for Health Research Health Technology Assessment programme.

Discrete Conditional Phase-type Model Utilising a Multiclass Support Vector Machine for the Prediction of Retinopathy of Prematurity

Retinopathy of prematurity (ROP) is a rare disease in which retinal blood vessels of premature infants fail to develop normally, and is one of the major causes of childhood blindness throughout the world. The Discrete Conditional Phase-type (DC-Ph) model consists of two components, the conditional component measuring the inter-relationships between covariates and the survival component which models the survival distribution using a Coxian phase-type distribution. This paper expands the DC-Ph models by introducing a support vector machine (SVM), in the role of the conditional component. The SVM is capable of classifying multiple outcomes and is used to identify the infant's risk of developing ROP. Class imbalance makes predicting rare events difficult. A new class decomposition technique, which deals with the problem of multiclass imbalance, is introduced. Based on the SVM classification, the length of stay in the neonatal ward is modelled using a 5, 8 or 9 phase Coxian distribution.

Single nucleotide polymorphisms in the APCS gene influence geographic atrophy secondary to VEGF inhibition therapy in neovascular macular degeneration.

Introduction: Age-related macular degeneration (AMD) is a leading cause of vision loss in the elderly mostly due to the development of neovascular AMD (nAMD) or geographic atrophy (GA). Intravitreal injections of anti-vascular endothelial growth factor (VEGF) agents are an effective therapeutic option for nAMD. Following anti-VEGF treatments, increased atrophy of the retinal pigment epithelium (RPE) and choriocapillaries that resembles GA has been reported. We sought to evaluate the underlying genetic influences that may contribute to this process. Methods: We selected 68 single nucleotide polymorphisms (SNPs) from genes previously identified as susceptibility factors in AMD, along with 43 SNPs from genes encoding the VEGF protein and its cognate receptors as this pathway is targeted by treatment. We enrolled 467 consecutive patients (Feb 2009 to October 2011) with nAMD who received anti-VEGF therapy. The acutely presenting eye was designated as the study eye and retinal tomograms graded for macular atrophy at study exit. Statistical analysis was performed using PLINK to identify SNPs with a P value < 0.01. Logistic regression models with macular atrophy as dependent variable were fitted with age, gender, smoking status, common genetic risk factors and the identified SNPs as explanatory variables. Results: Grading for macular atrophy was available in 304 study eyes and 70% (214) were classified as showing macular atrophy. In the unadjusted analysis we observed significant associations between macular atrophy and two independent SNPs in the APCS gene: rs6695377: odds ratio (OR) = 1.98; 95% confidence intervals (CI): 1.23, 3.19; P = 0.004; rs1446965: OR = 2.49, CI: 1.29, 4.82; P = 0.006 and these associations remained significant after adjustment for covariates. Conclusions: VEGF is a mitogen and growth factor for choroidal blood vessels and the RPE and its inhibition could lead to atrophy of these key tissues. Anti-VEGF treatment can interfere with ocular vascular maintenance and may be associated with RPE and choroidal atrophy. As such, these medications, which block the effects of VEGF, may influence the development of GA. The top associated SNPs are found in the APCS gene, a highly conserved glycoprotein that encodes Serum amyloid P (SAP) which opsonizes apoptotic cells. SAP can bind to and activate complement components via binding to C1q, a mechanism by which SAP may remove cellular debris, affecting regulation of the three complement pathways.

Preoperative radial artery volume flow is predictive of arteriovenous fistula outcomes

Objective: Guidelines recommend the creation of a wrist radiocephalic arteriovenous fistula (RAVF) as initial hemodialysis vascular access. This study explored the potential of preoperative ultrasound vessel measurements to predict AVF failure to mature (FTM) in a cohort of patients with end-stage renal disease in Northern Ireland

.Methods: A retrospective analysis was performed of all patients who had preoperative ultrasound mapping of upper limb blood vessels carried out from August 2011 to December 2014 and whose AVF reached a functional outcome by March 2015.

Results: There were 152 patients (97% white) who had ultrasound mapping andan AVF functional outcome recorded; 80 (54%) had an upper arm AVF created, and 69 (46%) had a RAVF formed. Logistic regression revealed that female gender (odds ratio [OR], 2.5; 95% confidence interval [CI], 1.12-5.55; P = .025), minimum venous diameter (OR, 0.6; 95% CI, 0.39-0.95; P = .029), and RAVF (OR, 0.4; 95% CI, 0.18-0.89; P = .026) were associated with FTM. On subgroup analysis of the RAVF group, RAVFs with an arterial volume flow <50 mL/min were seven times as likely to fail as RAVFs with higher volume flows (OR, 7.0; 95% CI, 2.35-20.87; P < .001).

Conclusions: In this cohort, a radial artery flow rate <50 mL/min was associated with a sevenfold increased risk of FTM in RAVF, which to our knowledge has not been previously reported in the literature. Preoperative ultrasound mapping adds objective assessment in the clinical prediction of AVF FTM.

Vasoactive Intestinal Polypeptide (VIP) in Human Dental Pulps

Pulpal innervation is not exclusively sensory and there are potential roles for other neuropeptides such as vasoactive intestinal polypeptide (VIP) in pulpal health and disease. In the systemic circulation VIP relaxes vascular smooth muscles leading to vasodilatation. It has been shown that VIP fibres are associated with pulpal blood vessels and therefore VIP may mediate vasoactivity in the dental pulp. A growing body of evidence has now demonstrated that an additional major physiological role of VIP is to act as a survival factor. In order to gain a better understanding of the role of neuropeptides in the caries process it is of interest to specifically examine a role for VIP. Objectives: The aim of the present study was to determine the levels of VIP in carious (moderately carious and grossly carious) compared with non-carious teeth. Methods: A total of 68 teeth were included in the study (22 non-carious, 20 moderately carious and 26 grossly carious). VIP was measured in all samples using a sensitive and specific radioimmunoassay. Results: The mean concentration of VIP in the pulps of non-carious teeth was 7.69 ng/g (9.41 SD) compared to 14.93 ng/g (15.58 SD) in carious teeth. Pair-wise comparisons of VIP levels using Tukey’s test showed statistically significant differences in VIP expression between non-carious and moderately carious teeth (p=0.002) and between moderately and grossly carious teeth, (p=0.002). Conclusion: The significantly increased levels of VIP in moderately carious pulps compared with either non-carious or grossly carious pulps may suggest a role for VIP as a protective or survival factor.

Endothelial Function in Hypertension: Victim or Culprit?

Far from simply lining the inner surface of blood vessels, the cellular monolayer that comprises the endothelium is a highly active organ that regulates vascular tone. In health, the endothelium maintains the balance between opposing dilator and constrictor influences, while in disease, it is the common ground on which cardiovascular risk factors act to initiate the atherosclerotic process. As such, it is the site at which cardiovascular disease begins and consequently acts as a barometer of an individual's likely future cardiovascular health. The vascular endothelium is a very active organ responsible for the regulation of vascular tone through the effects of locally synthesized mediators, predominantly nitric oxide (NO), endothelial NO synthase (eNOS), and superoxide. NO is abundantly evident in normally functioning vasculature where it acts as a vasodilator, inhibits inflammation, and has an antiaggregant effect on platelets. Its depletion is both a sign and cause of endothelial dysfunction resulting from reduced activity of eNOS and amplified production of nicotinamide adenine dinucleotide oxidase, which, in turn, results in raised levels of reactive oxygen species. This cascade is the basis for reduced vascular compliance through an imbalanced regulation of tone with a predominance of vasoconstrictive elements. Further, structural changes in the microvasculature are a critical early step in the loss of normal function. This microvascular dysfunction is known to be highly predictive of future macrovascular events and is consequently a very attractive target for intervention in the hypertensive population in order to prevent cardiovascular events.

Gold nanoparticle induced vasculature damage in radiotherapy: Comparing protons, megavoltage photons, and kilovoltage photons

Purpose: The purpose of this work is to investigate the radiosensitizing effect of gold nanoparticle (GNP) induced vasculature damage for proton, megavoltage (MV) photon, and kilovoltage (kV) photon irradiation. Methods: Monte Carlo simulations were carried out using tool for particle simulation (TOPAS) to obtain the spatial dose distribution in close proximity up to 20 µm from the GNPs. The spatial dose distribution from GNPs was used as an input to calculate the dose deposited to the blood vessels. GNP induced vasculature damage was evaluated for three particle sources (a clinical spread out Bragg peak proton beam, a 6 MV photon beam, and two kV photon beams). For each particle source, various depths in tissue, GNP sizes (2, 10, and 20 nm diameter), and vessel diameters (8, 14, and 20 µm) were investigated. Two GNP distributions in lumen were considered, either homogeneously distributed in the vessel or attached to the inner wall of the vessel. Doses of 30 Gy and 2 Gy were considered, representing typical in vivo enhancement studies and conventional clinical fractionation, respectively. Results: These simulations showed that for 20 Au-mg/g GNP blood concentration homogeneously distributed in the vessel, the additional dose at the inner vascular wall encircling the lumen was 43% of the prescribed dose at the depth of treatment for the 250 kVp photon source, 1% for the 6 MV photon source, and 0.1% for the proton beam. For kV photons, GNPs caused 15% more dose in the vascular wall for 150 kVp source than for 250 kVp. For 6 MV photons, GNPs caused 0.2% more dose in the vascular wall at 20 cm depth in water as compared to at depth of maximum dose (Dmax). For proton therapy, GNPs caused the same dose in the vascular wall for all depths across the spread out Bragg peak with 12.7 cm range and 7 cm modulation. For the same weight of GNPs in the vessel, 2 nm diameter GNPs caused three times more damage to the vessel than 20 nm diameter GNPs. When the GNPs were attached to the inner vascular wall, the damage to the inner vascular wall can be up to 207% of the prescribed dose for the 250 kVp photon source, 4% for the 6 MV photon source, and 2% for the proton beam. Even though the average dose increase from the proton beam and MV photon beam was not large, there were high dose spikes that elevate the local dose of the parts of the blood vessel to be higher than 15 Gy even for 2 Gy prescribed dose, especially when the GNPs can be actively targeted to the endothelial cells. Conclusions: GNPs can potentially be used to enhance radiation therapy by causing vasculature damage through high dose spikes caused by the addition of GNPs especially for hypofractionated treatment. If GNPs are designed to actively accumulate at the tumor vasculature walls, vasculature damage can be increased significantly. The largest enhancement is seen using kilovoltage photons due to the photoelectric effect. Although no significant average dose enhancement was observed for the whole vasculature structure for both MV photons and protons, they can cause high local dose escalation (>15 Gy) to areas of the blood vessel that can potentially contribute to the disruption of the functionality of the blood vessels in the tumor.

Tight junctional abnormality in multiple sclerosis white matter affects all calibres of vessel and is associated with blood-brain barrier leakage and active demyelination

Blood-brain barrier (BBB) hyperpermeability in multiple sclerosis (MS) is associated with lesion pathogenesis and has been linked to pathology in microvascular tight junctions (TJs). This study quantifies the uneven distribution of TJ pathology and its association with BBB leakage. Frozen sections from plaque and normal-appearing white matter (NAWM) in 14 cases were studied together with white matter from six neurological and five normal controls. Using single and double immunofluorescence and confocal microscopy, the TJ-associated protein zonula occludens-1 (ZO-1) was examined across lesion types and tissue categories, and in relation to fibrinogen leakage. Confocal image data sets were analysed for 2198 MS and 1062 control vessels. Significant differences in the incidence of TJ abnormalities were detected between the different lesion types in MS and between MS and control white matter. These were frequent in oil-red O (ORO)+ active plaques, affecting 42% of vessel segments, but less frequent in ORO- inactive plaques (23%), NAWM (13%), and normal (3.7%) and neurological controls (8%). A similar pattern was found irrespective of the vessel size, supporting a causal role for diffusible inflammatory mediators. In both NAWM and inactive lesions, dual labelling showed that vessels with the most TJ abnormality also showed most fibrinogen leakage. This was even more pronounced in active lesions, where 41% of vessels in the highest grade for TJ alteration showed severe leakage. It is concluded that disruption of TJs in MS, affecting both paracellular and transcellular paths, contributes to BBB leakage. TJ abnormality and BBB leakage in inactive lesions suggests either failure of TJ repair or a continuing pathological process. In NAWM, it suggests either pre-lesional change or secondary damage. Clinically inapparent TJ pathology has prognostic implications and should be considered when planning disease-modifying therapy

Inhibition of Advanced Glycation and Absence of Galectin-3 Prevent Blood-Retinal Barrier Dysfunction during Short-Term Diabetes

Breakdown of the inner blood-retinal barrier (iBRB) occurs early in diabetes and is central to the development of sight-threatening diabetic macular edema (DME) as retinopathy progresses. In the current study, we examined how advanced glycation end products (AGEs) forming early in diabetes could modulate vasopermeability factor expression in the diabetic retina and alter inter-endothelial cell tight junction (TJ) integrity leading to iBRB dysfunction. We also investigated the potential for an AGE inhibitor to prevent this acute pathology and examined a role of the AGE-binding protein galectin-3 (Gal-3) in AGE-mediated cell retinal pathophysiology. Diabetes was induced in C57/BL6 wild-type (WT) mice and in Gal-3(-/-) transgenic mice. Blood glucose was monitored and AGE levels were quantified by ELISA and immunohistochemistry. The diabetic groups were subdivided, and one group was treated with the AGE-inhibitor pyridoxamine (PM) while separate groups of WT and Gal-3(-/-) mice were maintained as nondiabetic controls. iBRB integrity was assessed by Evans blue assay alongside visualisation of TJ protein complexes via occludin-1 immunolocalization in retinal flat mounts. Retinal expression levels of the vasopermeability factor VEGF were quantified using real-time RT-PCR and ELISA. WT diabetic mice showed significant AGE -immunoreactivity in the retinal microvasculature and also showed significant iBRB breakdown (P < .005). These diabetics had higher VEGF mRNA and protein expression in comparison to controls (P < .01). PM-treated diabetics had normal iBRB function and significantly reduced diabetes-mediated VEGF expression. Diabetic retinal vessels showed disrupted TJ integrity when compared to controls, while PM-treated diabetics demonstrated near-normal configuration. Gal-3(-/-) mice showed significantly less diabetes-mediated iBRB dysfunction, junctional disruption, and VEGF expression changes than their WT counterparts. The data suggests an AGE-mediated disruption of iBRB via upregulation of VEGF in the diabetic retina, possibly modulating disruption of TJ integrity, even after acute diabetes. Prevention of AGE formation or genetic deletion of Gal-3 can effectively prevent these acute diabetic retinopathy changes.