interleukin-8 and Retinal-Neovascularization

To investigate the mechanism of intraocular neovascularization, we studied how vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8) are expressed in the ocular tissues under hypoxic conditions. Prior to proliferation of vascular endothelial cells resulting in neovascularization, the retinal tissues such as pericytes, retinal glial cells, ganglion cells, and ciliary epithelium react directly to hypoxia expressing VEGF and/or IL-8 and stimulate endothelial cell proliferation in a paracrine manner. We demonstrated that transcription factor activator protein-1 (AP-1) is activated for expression of VEGF messenger ribonuculeic acid (mRNA) and in a similar way nuclear factor kappa B (NF-kappa B) is activated for expression of IL-8 mRNA. However, hypoxia-induced expression of VEGF and/ or IL-8 is only one aspect of the complicated processes in intraocular neovascularization. We hope that further detailed analysis of the mechanism will make it possible to inhibit and treat clinically intraocular neovascularization in the near future.

Topics: Adult; Animals; Endothelial Growth Factors; Humans; Interleukin-8; Lymphokines; Rats; Retinal Neovascularization; RNA, Messenger; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Vascular adhesion molecules play an important role in various immunological disorders, particularly in cancers. However, little is known regarding the role of these adhesion molecules in proliferative retinopathies. We observed that IL-33 regulates VCAM-1 expression in human retinal endothelial cells and that genetic deletion of IL-33 reduces hypoxia-induced VCAM-1 expression and retinal neovascularization in C57BL/6 mice. We found that VCAM-1 via JunB regulates IL-8 promoter activity and expression in human retinal endothelial cells. In addition, our study outlines the regulatory role of VCAM-1-JunB-IL-8 signaling on retinal endothelial cell sprouting and angiogenesis. Our RNA sequencing results show an induced expression of CXCL1 (a murine functional homolog of IL-8) in the hypoxic retina, and intravitreal injection of VCAM-1 siRNA not only decreases hypoxia-induced VCAM-1-JunB-CXCL1 signaling but also reduces OIR-induced sprouting and retinal neovascularization. These findings suggest that VCAM-1-JunB-IL-8 signaling plays a crucial role in retinal neovascularization, and its antagonism might provide an advanced treatment option for proliferative retinopathies.

Topics: Animals; Chemokine CXCL1; Endothelial Cells; Humans; Hypoxia; Interleukin-33; Interleukin-8; Mice; Mice, Inbred C57BL; Neovascularization, Pathologic; Retinal Neovascularization; Transcription Factors; Vascular Cell Adhesion Molecule-1

Age-related macular degeneration (AMD), the leading cause of blindness in the elderly, targets the retinal pigment epithelium (RPE), a monolayer of cells at the back of the eye. As AMD progresses, it can develop into two distinct forms of late AMD: "dry," atrophic AMD, characterized by RPE senescence and geographic RPE loss, and "wet," neovascular AMD, characterized by RPE activation with abnormal growth of choroidal vessels. The genetic and molecular pathways that lead to these diverse phenotypes are currently under investigation. We have found that bone morphogenetic protein-4 (BMP4) is differentially expressed in atrophic and neovascular AMD. In atrophic AMD, BMP4 is highly expressed in RPE, and mediates oxidative stress induced RPE senescencein vitro via Smad and p38 pathways. In contrast, in neovascular AMD lesions, BMP4 expression in RPE is low, possibly a result of local expression of pro-inflammatory mediators. Thus, BMP4 may be involved in the molecular switch determining which phenotypic pathway is taken in the progression of AMD.

Topics: Aging; Bone Morphogenetic Protein 4; Cellular Senescence; Cyclin-Dependent Kinase Inhibitor p21; Humans; Hydrogen Peroxide; Interleukin-8; Macular Degeneration; Oxidative Stress; p38 Mitogen-Activated Protein Kinases; Retinal Neovascularization; Retinal Pigment Epithelium; Smad Proteins

To determine the levels of interleukin 8 (IL-8) in the vitreous of patients with proliferative diabetic retinopathy (PDR).. Observational case-control study.. Vitreous fluid samples were obtained by vitreoretinal surgery from 71 eyes of patients with diabetes mellitus type 2 with PDR and from 17 eyes of nondiabetic patients with a macular hole. PDR was classified as active and inactive and subdivided according to the extent of large-vessel gliotic obliteration. The cytokine levels were measured by cytometric bead array method.. The vitreous levels of IL-8 were significantly higher in patients with PDR in comparison with the control subjects (P < .001) and in patients with higher extent of large vessel gliotic obliteration (P < .001). A vitreous level of IL-8 was not associated with the presence of active PDR.. Increased levels of IL-8 in PDR were associated with a higher extent of large-vessel gliotic obliteration.

Topics: Aged; Case-Control Studies; Diabetes Mellitus, Type 2; Diabetic Retinopathy; Gliosis; Humans; Interleukin-8; Middle Aged; Retinal Neovascularization; Vitreous Body

Recent studies showed that interleukin-8 (IL-8) played an important role in retinal neovascularization. In this study, the effects of genistein on the expression of IL-8 in the arising retinal pigment epithelia-19 cells were studied. The levels of IL-8 protein expression in supernatants were punctually detected by ELISA. When the cells were treated with hypoxia (5% CO2, 95% N2), IL-8 secretion increased from 0.29 +/- 0.04 to 2.59 +/- 0.42 ng/ml. To study calcium-dependent IL-8 expression, cells were treated with KCl at 25 mM, norepinephrine (NE) at 10 nM, and glutamate (Glu) at 1 microM for 8 h. As a result, the levels of IL-8 protein in supernatants were significantly increased compared with that in the controls. When the cells are treated with genistein (50, 100, 200 microM) for 30 min before hypoxia or stimulations by KCl, NE, and Glu, the elevated expression of IL-8 protein was all suppressed in a concentration-dependent manner. These results suggested that suppression of IL-8 expression in retinal pigment epithelial cells might partly account for the inhibitive effect of genistein on retinal neovascularization.

Topics: Cell Hypoxia; Cells, Cultured; Enzyme Inhibitors; Genistein; Humans; Interleukin-8; Pigment Epithelium of Eye; Retinal Neovascularization

The purpose of this study was to determine the retinal expression of angiogenic chemokines/cytokines in a mouse model of oxygen-induced retinopathy.. C57BL/6 (B6) mice were exposed to 75% oxygen from postnatal day 7 (P7) to P12 and then recovered in room air. Reverse transcription-polymerase chain reaction (RT-PCR) was used to determine relative mRNA levels of KC, macrophage inflammatory protein-2 (MIP-2), interleukin-1alpha (IL-1alpha), and interferon gamma (IFN-gamma). Immunohistochemistry was used to localize KC in the retina. IL-1alpha was also injected into the vitreous of mouse eyes, and KC expression was examined by RT-PCR, enzyme-linked immunosorbent assay (ELISA), and immunohistochemistry.. KC expression at both the mRNA and protein levels was increased in P14, P17, and P21 of hyperoxia-injured eyes. KC immunoreactivity was localized along the nerve fiber layer and in radial Müller cell processes. IL-1alpha mRNA was modestly increased in hyperoxia-injured eyes on P14 and P17. INF-gamma mRNA was not detected in the retina. Adult mouse eyes injected with IL-1alpha demonstrated increased levels of both KC mRNA and protein, with KC immunoreactivity localized to Müller cell processes.. Oxygen-induced injury to the developing retina results in the induction of the CXC chemokine KC at both the mRNA and protein levels during the peak time points of neovascularization, suggesting a possible role in the pathogenesis of retinopathy of prematurity.

Topics: Animals; Animals, Newborn; Chemokine CXCL1; Chemokine CXCL2; Chemokines; Chemokines, CXC; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Gene Expression; Hyperoxia; Immunohistochemistry; Intercellular Signaling Peptides and Proteins; Interferon-gamma; Interleukin-1; Interleukin-8; Mice; Mice, Inbred C57BL; Oxygen; Retinal Neovascularization; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

To investigate the role of chemokines in the pathogenesis of proliferative diabetic retinopathy (PDR).. In total, 41 eyes of 38 patients undergoing vitrectomy were divided into two groups; PDR and non-PDR. The PDR group was comprised of 30 eyes, and the non-PDR group of 11 eyes. Vitreous specimens obtained at vitrectomy were centrifuged and separated into supernatants and cellular components. Concentrations of vascular endothelial growth factor(VEGF), interleukin-8 (IL-8), monocyte chemotactic protein-1 (MCP-1), and regulated upon activation, normal T cell expressed and secreted(RANTES) in the supernatants were determined by enzyme-linked immunosorbent assay(ELISA) or chemiluminescence enzyme immunoassay(CLEIA). Expression of VEGF in the cellular components was determined by immunohistochemistry.. Vitreous levels of VEGF(p < 0.05), IL8 (p < 0.0001) and MCP-1 (p < 0.05) in the PDR group were significantly higher than in the non-PDR group. However, there was no significant difference in RANTES between the two groups. There was a significant correlation (p < 0.0001, r = 0.84) between vitreous IL-8 and MCP-1 levels in the PDR group. After immunohistochemical staining with antiVEGF monoclonal antibody, VEGF positivity was localized in polymorphonuclear leukocytes and monocytes of the cellular components of PDR vitreous specimens.. These results indicate that chemokines are possibly involved in the recruitment of neutrophils and monocytes into the vitreous and that they play a role in the intraocular neovascularization characteristic of PDR.

Topics: Chemokine CCL2; Chemokine CCL5; Chemokines; Diabetic Retinopathy; Endothelial Growth Factors; Enzyme-Linked Immunosorbent Assay; Female; Humans; Interleukin-8; Luminescent Measurements; Male; Retinal Neovascularization; Vitrectomy; Vitreous Body

We have demonstrated that macrophages/microglia were activated during post-ischemic inflammation in a mouse model of ischemic retinal neovascularization, and that the angiogenesis induced by tumor necrosis factor-alpha (TNF-alpha) appeared to be modulated through such angiogenic factors as interleukin-8 (IL-8), vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (bFGF) in microvascular endothelial cells. We have extended these studies, and investigated whether TNF-alpha is localized in macrophages/microglia in the mouse model of retinal neovascularization, and whether TNF-alpha can induce angiogenic factors in retinal glial cells.. C57BL/6 J pups were placed in a 75% oxygen environment on postnatal day 7 (P7) for 5 days and then returned to room air. The co-localization of TNF-alpha with macrophages/microglia in the ischemic retina was examined by fluorescent immunohistochemistry. Bovine retinal glial cells were isolated for Northern blot analysis to quantify the expression levels of monocyte chemotactic protein-1 (MCP-1), IL-8, bFGF, and VEGF.. Double staining of retinas revealed that the TNF-alpha expression level was enhanced in macrophages/microglia 4 days after the hypoxia. Cellular mRNA levels of MCP-1, IL-8, and bFGF, but not VEGF, were increased after treating retinal glial cells with TNF-alpha (100 U/ml).. The results indicate that TNF-alpha is produced by activated macrophages/microglia and may participate in retinal neovascularization during post-ischemic inflammation through the induction of potent angiogenic factors in an autocrine or paracrine manner.

Topics: Animals; Blotting, Northern; Chemokine CCL2; Fibroblast Growth Factor 2; Fluorescent Antibody Technique, Indirect; Interleukin-8; Macrophage Activation; Mice; Mice, Inbred C57BL; Microglia; Polymerase Chain Reaction; Reperfusion Injury; Retina; Retinal Neovascularization; RNA, Messenger; Tumor Necrosis Factor-alpha

To investigate the role of interleukin (IL)-8 in intraocular neovascularization and the mechanism of its production.. Interleukin-8 was measured with enzyme-linked immunosorbent assays in vitreous and aqueous fluid obtained from patients with neovascular diseases. Localization of IL-8 was examined by immunohistochemistry. An in vitro angiogenesis assay was performed on collagen gels, by using bovine aortic endothelial cells to determine the effect of the vitreous fluid. In bovine retinal glial cells under hypoxia, NF-kappaB activation was evaluated by immunoblot analysis and by electrophoretic mobility shift assay, and IL-8 and vascular endothelial growth factor (VEGF) mRNA expression was determined by semiquantitative reverse transcription-polymerase chain reaction.. The concentration of IL-8 in vitreous fluid of patients with retinal neovascularization was significantly higher than that of patients without neovascular disease. Interleukin-8 immunostaining was detected in vascular endothelial cells and glial cells in the retinas with neovascularization. Vitreous fluid with high concentrations of IL-8 induced tubular morphogenesis in endothelial cells, and this effect was inhibited to a similar extent by neutralizing antibodies to IL-8 or to VEGF. In glial cells, in vitro, hypoxia induced NF-kappaB activation and increased IL-8 and VEGF mRNA. Furthermore, pyrrolidine dithiocarbamate, a specific inhibitor of NF-kappaB activation, prevented the induction of the IL-8 gene, but not that of the VEGF gene.. These results suggest that IL-8 induced by hypoxia and mediated by NF-kappaB may contribute to the pathogenesis of intraocular neovascularization.

Topics: Animals; Aqueous Humor; Cattle; Cells, Cultured; DNA Primers; Endothelial Growth Factors; Endothelium, Vascular; Enzyme-Linked Immunosorbent Assay; Humans; Hypoxia; Immunoenzyme Techniques; Interleukin-8; Lymphokines; Neuroglia; NF-kappa B; Polymerase Chain Reaction; Pyrrolidines; Retinal Neovascularization; RNA, Messenger; Thiocarbamates; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors; Vitreous Body