Experimental uveitis induced by products of activated lymphocytes: Intraocular effects of rhodopsin-induced lymphokines

Immunization of strain 13 guinea pigs by footpad injection of bovine rhodopsin in complete Freund's adjuvant produces experimental autoimmune uveitis (retinopathy) with retinal pathology characterized by destruction of the retinal photoreceptor cell layer, as we reported previously. Since rhodopsin-induced EAU can be passively transferred by immune cells but not antirhodopsin antibody, the present studies were conducted to determine if soluble products of activated lymphocytes (PAL) could cause the retinal pathology seen in experimental uveitis. These studies, reported here, show that intravitreal injection of supernatant factors generated by guinea pig lymph node cells specifically activated in vitro with rhodopsin or purified protein derivatives of tuberculin or nonspecifically activated with the mitogen concanavalin A produce uveitis in the normal guinea pig eye. The PAL produced mononuclear cell infiltration in the vitreous and a retinopathy with loss of retinal photoreceptor cells. Inflammatory cell infiltrates were found in the retina but not found in the anterior segment of the eye. Control lymphocyte supernatants did not produce retinal pathology. In guinea pigs sensitized to rhodopsin-complete Freund's adjuvant, intravitreal injection of the PAL produced a mononuclear vitreal infiltrate, disruption of the photoreceptor cell layer, necrosis of the inner retina, and a granulomatous infiltrate in the choroid with damage to the retinal pigment epithelium. Injection of control supernatants into the rhodopsin-complete Freund's adjuvant sensitized guinea pig eye did not produce inflammation and the retinal photoreceptor cell layer remained intact. Results from our studies indicate a role for the PAL along with other, yet undefined, factors in the initiation of autoimmune uveitis and in the autoimmune pathology of the retina.     

Inhibition of experimental autoimmune uveitis by retinal photoreceptor antigens coupled to spleen cells.

Experimental autoimmune uveitis (EAU) and experimental autoimmune pinealitis (EAP) are CD4+ T cell-mediated inflammatory diseases of the uveal tract and retina of the eye and of the pineal gland. EAU and EAP can be induced by several retinal autoantigens including S-antigen (S-Ag) and interphotoreceptor retinoid binding protein (IRBP). In this study we investigated the effect of intravenous administration of S-Ag and IRBP coupled to syngeneic spleen cells on the development of EAU and EAP. Injection of S-Ag or IRBP coupled to spleen cells 5 days prior to immunization with native S-Ag or IRBP, respectively, was effective in preventing the induction of EAU and EAP in LEW rats. Conversely, LEW rats receiving S-Ag-coupled spleen cells and challenged with IRBP or LEW rats receiving IRBP-coupled spleen cells and challenged with S-Ag developed a severe EAU within 10 days to 2 weeks following immunization, as did all control animals receiving sham-coupled spleen cells and challenged with the two retinal antigens. The results show that the administration of retinal autoantigens coupled to spleen cells effectively protects against the development of EAU when animals are subsequently challenged with the tolerizing antigen but not when challenged with another unrelated pathogenic retinal autoantigen.     

Immunoregulatory role of ocular macrophages: the macrophages produce RANTES to suppress experimental autoimmune uveitis

Murine experimental autoimmune uveitis (EAU) is a model of human uveitis. Ocular-infiltrating macrophages play a crucial role in the generation of tissue damage in EAU. In fact, several chemokines are actually produced in the inflamed eye. The aim of this study was to elucidate the role of ocular macrophage-derived chemokines in EAU. C57BL/6 mice were immunized with human interphotoreceptor retinoid binding protein peptide 1-20, and the EAU severity was scored at multiple time points based on microscopic fundus observations (retinal vascular dilatation and exudates) and histological examinations. The peak inflammatory response was observed 1 wk (day 16) after the beginning of macrophage infiltration to the eye (day 9). Ocular-infiltrating cells were enriched or depleted of macrophages by magnetic beads and analyzed by real-time RT-PCR for chemokine mRNA production. We found that only the macrophage-enriched cells from the eye produced RANTES, and thus proposed that macrophage-derived RANTES facilitated the ocular inflammations. In contrast to our postulate, neutralization of RANTES by specific Ab in vivo on days 9 and 13 exacerbated EAU. We also found that the ratio of ocular CD4/CD8 T cells was markedly increased after treatment. As a result, RANTES neutralization might exacerbate EAU by modulating the type of T cell subsets recruited to the eye. In conclusion, our data provide insight into the immunoregulatory role of macrophages and RANTES in the pathogenesis of ocular inflammation. Not all macrophage-derived chemokines cause local inflammation, since RANTES produced by ocular macrophages appears to suppress EAU.     

A new model of autoimmune disease. Experimental autoimmune uveoretinitis induced in mice with two different retinal antigens

Experimental autoimmune uveoretinitis (EAU) is an organ-specific, T lymphocyte-mediated autoimmune disease, which serves as a model for several human ocular inflammations of an apparently autoimmune nature. EAU pathology in some rodents and in monkeys can readily be induced by immunization with several different retinal proteins; however, advancing research into the cellular mechanisms of this disease has raised the need for an EAU model in an immunologically and genetically well defined species. We report here the induction of EAU in the mouse, which has hitherto been considered a species refractory to EAU, with two retinal Ag, the retinal soluble Ag and the interphotoreceptor retinoid-binding protein. Although all the mouse strains tested exhibited lymphocyte responses and antibody titers to both retinal Ag, EAU was inducible in only some of the strains, and the uveitogenic responses to retinal soluble Ag and interphotoreceptor retinoid-binding protein appeared to be mutually exclusive. The EAU model in mice was found to differ in several respects from the EAU model in other rodent species. Induction of the disease was achieved with a relatively high dose of Ag and an intensified immunization protocol, and the onset of disease was later, the duration was longer, and the course was less acute. Anterior segment involvement was slight or nonexistent, and damage to the retina and uvea was of a focal rather than of a diffuse nature. Murine EAU appeared to approximate some types of human uveitis more closely than the EAU models described in other rodent species with respect to its pathologic manifestations as well as its more chronic course. The relatively longer duration of the active stage of disease in murine EAU should facilitate therapeutic intervention in established disease, which was not feasible in the more acute models of EAU. The extensive knowledge of the immunologic parameters of the mouse and the availability of genetically defined strains should be of great value in the study of cellular mechanisms and immunogenetics of ocular autoimmune disease.     

Autophagy-induced regression of hyaloid vessels in early ocular development

The hyaloid vessel is a transient intraocular circulatory system that undergoes a complete regression as the retina becomes matured with retinal vascularization. If the complete involution of the hyaloid vessels failes, the pathological persistence of these vessels results in persistent hyperplastic primary vitreous (PHPV) associated with severe ocular pathologies. Unfortunately despite its clinical significance, cellular and molecular processes involved in hyaloid regression remain to be elucidated. Herein, we for the first time demonstrated that autophagy could contribute to the regression of hyaloid vessels in early developing retina. In developing retina, hyaloid vessel regression coincided with retinal vascular development; this occurred simultaneous with apoptotic and autophagic processes. Moreover, in vascular endothelial cells under hypoxic conditions, LC3-II conversion was detected along with caspase-3 activation. The autophagy inducer rapamycin induced autophagy-mediated cell death of vascular endothelial cells in a dose-dependent manner. Moreover, rapamycin significantly enhanced the involution of hyaloid vessels in the early developing eye. Therefore, our results suggest that the autophagy pathway would be involved in hyaloid regression that occurs during early ocular development. Furthermore, activation of the autophagy pathway could be considered for a therapeutic approach to PHPV.     

IL-1β is upregulated in the diabetic retina and retinal vessels: cell-specific effect of high glucose and IL-1β autostimulation

Many molecular and cellular abnormalities detected in the diabetic retina support a role for IL-1β-driven neuroinflammation in the pathogenesis of diabetic retinopathy. IL-1β is well known for its role in the induction and, through autostimulation, amplification of neuroinflammation. Upregulation of IL-1β has been consistently detected in the diabetic retina; however, the mechanisms and cellular source of IL-1β overexpression are poorly understood. The aim of this study was to investigate the effect of high glucose and IL-1β itself on IL-1β expression in microglial, macroglial (astrocytes and Müller cells) and retinal vascular endothelial cells; and to study the effect of diabetes on the expression of IL-1β in isolated retinal vessels and on the temporal pattern of IL-1β upregulation and glial reactivity in the retina of streptozotocin-diabetic rats. IL-1β was quantified by RealTime RT-PCR and ELISA, glial fibrillar acidic protein, α2-macroglobulin, and ceruloplasmin by immunoblotting. We found that high glucose induced a 3-fold increase of IL-1β expression in retinal endothelial cells but not in macroglia and microglia. IL-1β induced its own synthesis in endothelial and macroglial cells but not in microglia. In retinal endothelial cells, the high glucose-induced IL-1β overexpression was prevented by calphostin C, a protein kinase C inhibitor. The retinal vessels of diabetic rats showed increased IL-1β expression as compared to non-diabetic rats. Retinal expression of IL-1β increased early after the induction of diabetes, continued to increase with progression of the disease, and was temporally associated with upregulation of markers of glial activation. These findings point to hyperglycemia as the trigger and to the endothelium as the origin of the initial retinal upregulation of IL-1β in diabetes; and to IL-1β itself, via autostimulation in endothelial and macroglial cells, as the mechanism of sustained IL-1β overexpression. Interrupting the vicious circle triggered by IL-1β autostimulation could limit the progression of diabetic retinopathy.     

Hyperoxia causes decreased expression of vascular endothelial growth factor and endothelial cell apoptosis in adult retina

Mice or humans with photoreceptor degenerations experience permeability and dropout of retinal capillaries. Loss of photoreceptors results in decreased oxygen usage and thinning of the retina with increased oxygen delivery to the inner retina. To investigate the possibility that increased tissue oxygen plays a role in the vascular damage, we exposed adult mice to hyperoxia, which also increases oxygen in the retina. After 1, 2, or 3 weeks of hyperoxia, there was a statistically significant decrease in retinal vascular density that was not reversible, and endothelial cell apoptosis was demonstrated by TUNEL staining. Mice exposed to hyperoxia and mice with photoreceptor degeneration both showed decreased expression of VEGF in the retina. After complete or near-complete degeneration of photoreceptors, there was increased expression of VEGF in RPE cells, which may explain the association of photoreceptor degeneration and neovascularization in or around the RPE. Increased expression of VEGF in photoreceptors of transgenic mice failed to prevent hyperoxia-induced retinal capillary dropout. These data suggest that increased oxygen in the retina, either by increased inspired oxygen or by photoreceptor degeneration, results in endothelial cell death and dropout of capillaries. Decreased expression of VEGF may be a contributing factor, but the situation may be more complicated for mature retinal vessels than it is for immature vessels, because VEGF replacement does not rescue mature retinal vessels, suggesting that other factors may also be involved.     

Identification of a uveitopathogenic and lymphocyte proliferation site in bovine S-antigen

S-antigen is a well-characterized retinal protein that is highly pathogenic for the induction of experimental autoimmune uveitis (EAU), a severe inflammatory disease of the eye and the pineal gland. EAU was observed following the immunization of Lewis rats with various doses (50 to 200 micrograms) of a small synthetic peptide, peptide N (22 amino acids in length), which corresponds to amino acid positions 281 to 302 in bovine S-antigen. Peptide N consistently induced an EAU that was identical to the disease caused by native S-antigen. Clinically, the disease that developed in the eye was characterized by iris and pericorneal hyperemia, followed by inflammatory exudates in the anterior chamber and vitreous. Histopathologically, a severe inflammatory response was observed that resulted in the complete destruction of the photoreceptor cell layer of the retina. In addition, animals with ocular inflammatory disease had an associated pinealitis characterized by a lymphocytic infiltration of the pineal gland. Furthermore, draining lymph node cells of rats immunized with peptide N showed strong in vitro proliferative responses toward peptide N as measured by [3H]thymidine uptake. Our results indicate that several synthetic peptides, which correspond to the amino acid sequence of bovine S-antigen, are capable of inducing an EAU and, as such, suggest that multiple uveitopathogenic sites may be present in the molecule.     

Acquired and congenital ocular toxoplasmosis experimentally induced in Calomys callosus (Rodentia, Cricetidae)

An experimental model for acquired and congenital ocular toxoplasmosis as well as a model to induce experimental autoimmune uveitis (EAU) was investigated in Calomys callosus. Toxoplasma gondii, ME-49 strain, was used to infect males and pregnant- and not pregnant-females while S-antigen, a major glycoprotein of the retinal photoreceptor cell, was used to induce EAU. The ocular lesions elicited by T. gondii were characterized by the presence of cysts, free tachyzoites and inflammatory cells in the retina or related tissues. In the congenital form, 40% of the fetus presented ocular lesions, i.e., presence of cysts in the retina, vitreous, and extra-retinal tissues. In the acquired form, 75% of the females and 50% of the males presented unilateral ocular cysts both at 21 and 47 days post-infection. It was also demonstrated that S-antigen was not uveitogenic in the C. callosus model. No lesion was observed in the animals exclusively immunized with this retinal component, even when jacalin was used as additional adjuvant for polyclonal response to the retinal antigen. It can be concluded that C. callosus may constitute in a promising model for study both acquired and congenital ocular toxoplasmosis, particularly when it is important to make sure that a non autoimmune process is involved in the genesis of the ocular infection.     

The immunopathology of experimental allergic encephalomyelitis. II. Endothelial cell Ia increases prior to inflammatory cell infiltration

Experimental allergic encephalomyelitis (EAE) is a T cell-mediated neuroimmunologic disease model characterized by meningeal and parenchymal mononuclear cell infiltrates (see preceding companion paper). Here we report enhanced staining for Ia in the central nervous system (CNS) microvasculature endothelium in acute EAE in adult strain 13 guinea pigs (GP) sensitized with GP spinal cord homogenate (SC) or with GP myelin basic protein (MBP) in complete Freund's adjuvant (CFA). Cryostat sections of CNS and other tissues were stained with two monoclonal antibodies, 5S2 and 22C4, to GP Ia determinants, and with polyclonal antibody to factor VIII-related antigen (VIII-RA) as an endothelial cell marker. Morphometric techniques were employed on immunoperoxidase counterstained and coded sections to determine the frequency of Ia+ vessels and cells. Rare (approximately 10% of VIII-RA+) vascular endothelial cells were Ia+ in the CNS of normal and CFA-sensitized controls. SC- or MBP-sensitized strain 13 GP sacrificed on day 7, before the onset of neurologic signs (pre-clinical), had no detectable CNS mononuclear cell infiltrates, but had increased (approximately 30% of VIII-RA+) endothelial cell Ia staining over controls (p less than 0.001). The endothelial Ia staining persisted (approximately 35% of VIII-RA+) in vessels as the animals developed paralysis. There were no differences in endothelial cell Ia between SC- and MBP-induced disease. EAE-resistant strain 2 GP sensitized with SC/CFA had no neurologic signs, and had fewer inflammatory foci than strain 13 GP with EAE, but had similar numbers of Ia+ endothelial cells. No differences in endothelial cell Ia staining were found in non-CNS tissues among any GP groups. In EAE, increased endothelial cell Ia is a pre-inflammatory, target organ-specific alteration that persists during inflammation. The findings suggest that in vivo modulation of endothelial cell Ia may be part of the local immune response. Endothelial cells may play a significant role, in antigen presentation or in promoting T cell migration, in the in situ immune response in the CNS.     

Autoimmune uveitis elicited with antigen-pulsed dendritic cells has a distinct clinical signature and is driven by unique effector mechanisms: initial encounter with autoantigen defines disease phenotype

Human autoimmune uveitis is a heterogeneous group of potentially blinding ocular diseases in which most patients who exhibit immunity recognize the same retinal Ag. It is represented by the model of experimental autoimmune uveitis (EAU) induced in mice by immunization with retinal Ag in CFA. Murine EAU is characterized by a Th1/Th17 response pattern, which may not represent all types of human uveitis. We report in this study a new model of EAU induced by injection of matured dendritic cells loaded with a uveitogenic retinal peptide. Dendritic cell-induced EAU demonstrated unique characteristics compared with traditional EAU in terms of clinical manifestations, the nature of the inflammatory infiltrating cells, the cytokine response profile, and a strict requirement for IFN-gamma, whereas IL-17 appeared to play a minor role. Disease was self-limiting, but could be reinduced with the same Ag in CFA, albeit with reduced severity, suggesting post-recovery resistance. Our study demonstrates in a disease setting that the context in which the same autoantigen is initially presented to the immune system precipitates distinct forms of pathology via a distinct pathogenic pathway on the same genetic background. These findings may shed new light on the complex biology and the heterogeneous nature of human uveitis, and provide an alternative model for uveitic diseases of immune origin.     

Antigen-directed retention of an autoimmune T-cell line

We have used the T-cell-mediated, organ-specific autoimmune disease model of experimental autoimmune uveoretinitis (EAU) in the Lewis rat to study antigen-directed retention of autoimmune T helper cells in the target organ. We have compared the migration into the eye of two T-helper-cell lines: ThS, specific for retinal S antigen (S-Ag), that is uveitogenic to normal syngenic recipients, and ThP, specific to purified protein derivative of tuberculin (PPD), that is non-uveitogenic. The retention of adoptively transferred 51Cr-labeled ThS and ThP was studied up to the stage of disease induction in unprimed animals, during the acute stage of EAU induced by active immunization with S-Ag, and during the acute stage of a uveitis induced by a nonocular antigen (bovine serum albumin, BSA). Low numbers of cells from the two lymphocyte lines were detected in the eyes of unprimed animals, with no obvious increase of ThS over ThP, despite induction of EAU in the recipient animals by the injected ThS cells. In S-Ag-induced EAU many more ThS accumulated in the eye than ThP. In BSA uveitis both T-cell lines accumulated in the eye to the same extent, but more than in control noninflamed eyes. These results demonstrate the presence of increased antigen-specific retention of circulating autoimmune T helper lymphocytes during the acute stage of an ocular antigen-specific, but not ocular antigen nonspecific, inflammation. Since detectable accumulation of ThS cells in the eye was not a prerequisite for the induction of EAU, this phenomenon appears to be the result, rather than the cause, of the autoimmune process.     

A novel bispecific molecule delivered by recombinant AAV2 suppresses ocular inflammation and choroidal neovascularization

Elevated vascular endothelial growth factor (VEGF) and complement activation are implicated in the pathogenesis of different ocular diseases. The objective of this study was to investigate the hypothesis that dual inhibition of both VEGF and complement activation would confer better protection against ocular inflammation and neovascularization. In this study, we engineered a secreted chimeric VEGF inhibitor domain (VID), a complement inhibitor domain (CID) and a dual inhibitor (ACVP1). Vectors expressing these three inhibitors were constructed and packaged into AAV2 (sextY‐F) particles. The expression and secretion of the proteins were validated by Western blot. The effects of these inhibitors expressed from AAV2 vectors were examined in endotoxin‐induced uveitis (EIU), experimental autoimmune uveoretinitis (EAU) and choroidal neovascularization (CNV) mouse models. The AAV2 vectors expressing the CID‐ and ACVP1‐attenuated inflammation in EIU and EAU model, whereas the vector expressing VID showed improved retinal structure damaged by EAU, but not affect the infiltration of inflammatory cells in EAU or EIU eyes. Both VID and CID vectors improved laser‐induced retinal and choroid/RPE injuries and CNV, whereas ACVP1 vector provided significantly better protection. Our results suggest that gene therapy targeting VEGF and complement components could provide an innovative and long‐term strategy for ocular inflammatory and neovascular diseases.     

Suppression of protein kinase C-ζ attenuates vascular leakage via prevention of tight junction protein decrease in diabetic retinopathy

To investigate the effect of protein kinase C (PKC)-ζ inhibition on vascular leakage in diabetic retinopathy, streptozotocin-induced diabetic mice were intravitreously injected with siPKC-ζ. According to the fluorescein angiography of the retinal vessels, suppression of PKC-ζ effectively attenuated vascular leakage in diabetic retina. Further evaluation on the retina with western blot analysis and immunohistochemistry revealed accompanying restoration of tight junction proteins on retinal vessels. As two major contributors to vascular leakage in diabetic retinopathy, vascular endothelial growth factor (VEGF) and advanced glycation end products (AGEs) were investigated on the tight junction protein expression in endothelial cells. Inhibition of PKC-ζ attenuated VEGF-induced decrease of tight junction proteins and accompanying hyperpermeability in human retinal microvascular endothelial cells (HRMECs). PKC-ζ inhibition also attenuated AGE-induced decrease of tight junction proteins in HRMECs. Our findings suggest that inhibition of PKC-ζ could be an alternative treatment option for compromised blood-retinal barrier in diabetic retinopathy.     

Integrin-dependent and -independent functions of astrocytic fibronectin in retinal angiogenesis

Fibronectin (FN) is a major component of the extracellular matrix and functions in cell adhesion, cell spreading and cell migration. In the retina, FN is transiently expressed and assembled on astrocytes (ACs), which guide sprouting tip cells and deposit a provisional matrix for sprouting angiogenesis. The precise function of FN in retinal angiogenesis is largely unknown. Using genetic tools, we show that astrocytes are the major source of cellular FN during angiogenesis in the mouse retina. Deletion of astrocytic FN reduces radial endothelial migration during vascular plexus formation in a gene dose-dependent manner. This effect correlates with reduced VEGF receptor 2 and PI3K/AKT signalling, and can be mimicked by selectively inhibiting VEGF-A binding to FN through intraocular injection of blocking peptides. By contrast, AC-specific replacement of the integrin-binding RGD sequence with FN-RGE or endothelial deletion of itga5 shows little effect on migration and PI3K/AKT signalling, but impairs filopodial alignment along AC processes, suggesting that FN-integrin α5β1 interaction is involved in filopodial adhesion to the astrocytic matrix. AC FN shares its VEGF-binding function and cell-surface distribution with heparan-sulfate (HS), and genetic deletion of both FN and HS together greatly enhances the migration defect, indicating a synergistic function of FN and HS in VEGF binding. We propose that in vivo the VEGF-binding properties of FN and HS promote directional tip cell migration, whereas FN integrin-binding functions to support filopodia adhesion to the astrocytic migration template.     

Suppression of Autoimmune Retinal Inflammation by an Antiangiogenic Drug

Chronic and recurrent uveitis account for approximately 10% of legal blindness in the western world. Autoimmune uveitis is driven by activated CD4⁺ T cells that differentiate into effector T helper cells (Th1, Th2, and Th17) which release proinflammatory cytokines that damage the retina. In this study we investigated the effect of the methionine aminopeptidase 2 (MetAP2) inhibitor, Lodamin, on T cell activation and differentiation. MetAp2 is an enzyme which regulates cellular protein synthesis and is highly expressed in T cells. Lodamin was found to suppress T cell receptor (TCR) mediated T cell proliferation and reduced the production of Th1 and Th17 cells. Further, Lodamin suppressed overall inflammation in the mouse model of experimental autoimmune uveitis (EAU) by a six fold. This effect was attributed in part to a reduction in retinal proinflammatory cytokines, down regulation of MetAP2 expression in purified lymph node CD4⁺ T cells, and a general normalization of the systemic immune reaction.     

Intraocular Ia expression in vivo induced in the absence of immune T cells

The expression of Ia antigen on the ciliary body, retinal pigment epithelium, and retinal vascular endothelium was investigated using two models of ocular inflammation. Active systemic immunization with bovine serum albumin with subsequent ocular challenge resulted in increased Ia expression in the ciliary body and retinal pigment epithelium. This was compared with passive transfer of hyperimmune serum to bovine serum albumin followed by ocular challenge when Ia expression was found to occur only in the ciliary body. An intraocular T-cell infiltrate occurred only in the actively immunized animals and was not present after passive transfer, suggesting that the Ia induction in this latter situation occurred in the absence of T cells.     

Perturbation of Mouse Retinal Vascular Morphogenesis by Anthrax Lethal Toxin

Lethal factor, the enzymatic moiety of anthrax lethal toxin (LeTx) is a protease that inactivates mitogen activated protein kinase kinases (MEK or MKK). In vitro and in vivo studies demonstrate LeTx targets endothelial cells. However, the effects of LeTx on endothelial cells are incompletely characterized. To gain insight into this process we used a developmental model of vascularization in the murine retina. We hypothesized that application of LeTx would disrupt normal retinal vascularization, specifically during the angiogenic phase of vascular development. By immunoblotting and immunofluorescence microscopy we observed that MAPK activation occurs in a spatially and temporally regulated manner during retinal vascular development. Intravitreal administration of LeTx caused an early delay (4 d post injection) in retinal vascular development that was marked by reduced penetration of vessels into distal regions of the retina as well as failure of sprouting vessels to form the deep and intermediate plexuses within the inner retina. In contrast, later stages (8 d post injection) were characterized by the formation of abnormal vascular tufts that co-stained with phosphorylated MAPK in the outer retinal region. We also observed a significant increase in the levels of secreted VEGF in the vitreous 4 d and 8 d after LeTx injection. In contrast, the levels of over 50 cytokines other cytokines, including bFGF, EGF, MCP-1, and MMP-9, remained unchanged. Finally, co-injection of VEGF-neutralizing antibodies significantly decreased LeTx-induced neovascular growth. Our studies not only reveal that MAPK signaling plays a key role in retinal angiogenesis but also that perturbation of MAPK signaling by LeTx can profoundly alter vascular morphogenesis.     

Distribution of melatonin MT1 receptor immunoreactivity in human retina.

Melatonin is synthesized in the pineal gland and retina during the night. Retinal melatonin is believed to be involved in local cellular modulation and in regulation of light-induced entrainment of circadian rhythms. The present study provides the first immunohistochemical evidence for the localization of melatonin 1a-receptor (MT1) in human retina of aged subjects. Ganglion, amacrine, and photoreceptor cells expressed MT1. In addition, MT1 immunoreactivity was localized to cell processes in the inner plexiform layer and to central vessels of the retina, as well as to retinal vessels but not to ciliary or choroidal vessels. These results support a variety of cellular and vascular effects of melatonin in the human retina. Preliminary evidence from patients with Alzheimer's disease (AD) revealed increased MT1 immunoreactivity in ganglion and amacrine cells, as well as in vessels. In AD cases photoreceptor cells were degenerated and showed low MT1 expression.     

视网膜微血管内皮细胞在糖尿病视网膜病变中的研究现状

糖尿病视网膜疾病是导致成年人失明的主要因素,是糖尿病的一种令人恐惧的并发症,高血糖被认为是促进其发展的主要原因。高血糖不断地破坏视网膜的微血管系统最终导致视网膜的许多代谢,结构和功能的紊乱。视网膜微血管内皮细胞在微脉管系统中形成树枝状供应视网膜神经,这些内皮细胞的解剖和生理符合重要视觉保护的营养需求[1]。一方面,内皮组织务必确保氧的供应和代谢活跃的视网膜营养供应;另一方面,内皮细胞有助于血-视网膜屏障将循环产生的毒素分子,白细胞促炎性物质排出体外来保护视网膜,这种特性也可能会引起疾病,比如:视网膜血管的渗漏和新生血管,炎性物质转移,因此,视网膜内皮细胞在视网膜缺血性病变,血管炎中起到重要作用,包括糖尿病视网膜病变和视网膜炎症或感染尤其是后葡萄膜炎。使用基因表达和蛋白质组学分析等研究方法,有助于了解这些疾病的发病机制。为了进一步开展对糖尿病视网膜疾病的研究,有必要就目前有关糖尿病视网膜病变患者微血管内皮细胞的研究进展予以综述,旨在为糖尿病视网膜病变的深入研究提供参考依据。