In vivo significance of ICAM-1--dependent leukocyte adhesion in early corneal angiogenesis. (1/340)

PURPOSE: Numerous investigations have stressed the significance of leukocytes in early angiogenesis. Leukocytes invade the cornea, and the location of their extravasation corresponds to the site of vessel ingrowth. The interactions between leukocytes and vascular endothelium are mediated by various proteins, including adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1). In this study, the role of ICAM-1 during early corneal angiogenesis was evaluated in vivo. METHODS: Corneal neovascularization was induced in New Zealand White rabbits by use of intrastromal pellets containing 750 ng vascular endothelial growth factor (VEGF). The fluorescent dye rhodamine 6G was used to stain leukocytes in vivo. Leukocyte adhesion and vessel growth were quantified in vivo by high-resolution fluorescence angiography. To inhibit ICAM-1 interactions a microemulsion containing anti-ICAM-1 antibody was applied topically. RESULTS: Limbal vessels showed increased leukocyte adhesion 24 hours after pellet implantation: The number of rolling and sticking leukocytes was significantly increased compared with the number in control animals (P < 0.01). Treatment with anti-ICAM-1 antibody resulted in reduced leukocyte sticking and increased leukocyte rolling. The area covered by new blood vessels was significantly diminished in eyes treated with anti-ICAM-1 (P < 0.05). CONCLUSIONS: The results support the hypothesis that ICAM-1-mediated leukocyte adhesion is a key event in early angiogenesis. This model may serve for investigation of the significance of adhesion molecules by in vivo observation and quantification.  (+info)

Suppression of angiogenesis and tumor growth by the inhibitor K1-5 generated by plasmin-mediated proteolysis. (2/340)

Proteolytic enzymes are involved in generation of a number of endogenous angiogenesis inhibitors. Previously, we reported that angiostatin, a potent angiogenesis inhibitor, is a proteolytic fragment containing the first four kringle modules of plasminogen. In this report, we demonstrate that urokinase-activated plasmin can process plasminogen to release an angiogenesis inhibitor, K1-5 (protease-activated kringles 1-5). K1-5 inhibits endothelial-cell proliferation with a half-maximal concentration of approximately 50 pM. This inhibitory effect is endothelial-cell-specific and appears to be at least approximately 50-fold greater than that of angiostatin. A synergistic efficacy of endothelial inhibition was observed when angiostatin and kringle 5 (K5) were coincubated with capillary endothelial cells. The synergistic effect is comparable to that produced by K1-5 alone. Systemic treatment of mice with K1-5 at a low dose significantly blocked the fibroblast growth factor-induced corneal neovascularization, whereas angiostatin had no effect at the same dose. K1-5 also suppressed angiogenesis in chicken embryos. Systemic administration of K1-5 at a low dose at which angiostatin was ineffective significantly suppressed the growth of a murine T241 fibrosarcoma in mice. The antitumor effect correlates with the reduced neovascularization. These findings suggest that the plasmin-mediated proteolysis may be involved in the negative switch of angiogenesis.  (+info)

Expression of cell adhesion molecules on limbal and neovascular endothelium in corneal inflammatory neovascularization. (3/340)

PURPOSE: To investigate the expression of cell-adhesion molecules on corneolimbal and neovascular endothelium and the associated leukocyte infiltration in an experimental model of inflammatory corneal neovascularization (NV). METHODS: Corneal NV was induced in BALB/c mice by placement of nylon sutures. Interleukin-1 receptor antagonist (IL-1ra) was used topically to determine whether suppression of IL-1 could affect adhesion molecule expression and leukocytic infiltration. At set time points, corneal samples were analyzed immunohistochemically for expression of P-selectin, E-selectin, intercellular adhesion molecule (ICAM)-1, vascular adhesion molecule (VCAM)-1, and platelet- endothelial adhesion molecule (PECAM)-1. Leukocytic infiltration at different time points was quantified histologically. In companion experiments mice deficient in ICAM-1 were investigated to determine the functional relevance of this molecule in corneal leukocyte infiltration. RESULTS: Significant enhanced expression of ICAM-1 was detected on the corneolimbal vascular endothelium as early as 8 hours and on the newly formed corneal NV by day 3, and treatment with IL-1ra led to significant suppression of this expression. IL-1ra-induced suppression of ICAM-1 expression was accompanied by a profound decrease in corneal leukocytic infiltration by 44.6% at day 1 (P < 0.003), 71.8% at day 3 (P < 0.001), 60.1% at day 7 (P < 0.001), and 63.8% at day 14 (P < 0.001), compared with control corneas. Similarly, in ICAM-1 knockout mice, the corneal leukocytic infiltration was 50.3%, 52.9%, and 36.4%, compared with wild-type control animals on day 1 (P < 0.001), day 7 (P < 0.005), and day 14 (P < 0.001), respectively. Expression of PECAM-1 was constitutively present on perilimbal vascular endothelium and had no response to IL-1ra treatment. No significant expression of P-selectin, E-selectin, or VCAM-1 was detected in this experimental model. CONCLUSIONS: These results suggest that leukocytic infiltration in this model of inflammatory corneal NV is closely associated with ICAM-1 expression, and that topical IL-1ra displays corneal anti-inflammatory effects, largely by suppressing ICAM-1 expression on vascular endothelial cells.  (+info)

VEGF contributes to postnatal neovascularization by mobilizing bone marrow-derived endothelial progenitor cells. (4/340)

Vascular endothelial growth factor (VEGF) has been shown to promote neovascularization in animal models and, more recently, in human subjects. This feature has been assumed to result exclusively from its direct effects on fully differentiated endothelial cells, i.e. angiogenesis. Given its regulatory role in both angiogenesis and vasculogenesis during fetal development, we investigated the hypothesis that VEGF may modulate endothelial progenitor cell (EPC) kinetics for postnatal neovascularization. Indeed, we observed an increase in circulating EPCs following VEGF administration in vivo. VEGF-induced mobilization of bone marrow-derived EPCs resulted in increased differentiated EPCs in vitro and augmented corneal neovascularization in vivo. These findings thus establish a novel role for VEGF in postnatal neovascularization which complements its known impact on angiogenesis.  (+info)

Inhibition of rat corneal angiogenesis by 16-kDa prolactin and by endogenous prolactin-like molecules. (5/340)

PURPOSE: The cornea is an avascular organ, where induction of new blood vessels involves the turn-on of proangiogenic factors and/or the turn-off of antiangiogenic regulators. Prolactin (PRL) fragments of 14 kDa and 16 kDa bind to endothelial cell receptors and inhibit angiogenesis. This study was designed to determine whether antiangiogenic PRL-like molecules are involved in cornea avascularity. METHODS: Sixteen-kDa PRL and basic fibroblast growth factor (bFGF) or anti-PRL antibodies were placed into rat cornea micropockets and neovascularization evaluated by the optical density associated with capillaries stained by the peroxidase reaction and by the number of vessels growing into the implants. Prolactin receptors in corneal epithelium were investigated by immunocytochemistry. RESULTS: bFGF induced a dose-dependent stimulation of corneal neovascularization. This effect was inhibited by coadministration of 16-kDa PRL, as indicated by a 65% reduction in vessel density and a 50% decrement in the incidence of angiogenic responses. Corneal angiogenic reactions of different intensities were induced by implantation of polyclonal and monoclonal anti-PRL antibodies. Corneal epithelial cells were labeled by several anti-PRL receptor monoclonal antibodies. CONCLUSIONS: These findings show that exogenous 16-kDa PRL inhibits bFGF-induced corneal neovascularization and suggest that PRL-like molecules with antiangiogenic actions function in the cornea. PRL receptors in the corneal epithelium may imply that PRL in the cornea derives from lacrimal PRL internalized through an intracellular pathway. These observations are consistent with the notion that members of the PRL family are potential regulators of corneal angiogenesis.  (+info)

Nitric oxide synthase-II is expressed in severe corneal alkali burns and inhibits neovascularization. (6/340)

PURPOSE: Inducible nitric oxide synthase (NOS-II) is expressed in many inflammatory conditions. The implication of nitric oxide (NO) in angiogenesis remains controversial. The role of NOS-II and its influence on angiogenesis in corneal neovascularization is unknown and was investigated in this study. METHODS: A mouse model of corneal neovascularization induced by chemical cauterization was used. NOS-II mRNA expression was analyzed by reverse transcriptase-polymerase chain reaction, and NOS-II protein was studied in situ by immunohistochemical analysis of the cornea. The influence of NOS-II on neovascularization was determined by comparison of vessel development in "normal" wild-type mice and mice with a targeted disruption of the NOS-II gene. RESULTS: NOS-II mRNA was induced to very high levels after corneal cauterization and remained upregulated throughout the disease. Migratory cells in the center of the cauterization area expressed NOS-II protein. The neovascular response in mice lacking the NOS-II gene was significantly stronger than in wild-type mice, and the difference increased over time. CONCLUSIONS: These data are the first evidence that NOS-II is expressed in this model of sterile corneal inflammation. NOS-II expression inhibited angiogenesis in severe corneal alkali burns.  (+info)

IFN-gamma-inducible protein-10 attenuates bleomycin-induced pulmonary fibrosis via inhibition of angiogenesis. (7/340)

Few studies have addressed the importance of vascular remodeling in the lung during the development of bleomycin-induced pulmonary fibrosis (BPF). For fibroplasia and deposition of extracellular matrix to occur, there must be a geometric increase in neovascularization. We hypothesized that net angiogenesis during the pathogenesis of fibroplasia and deposition of extracellular matrix during BPF are dependent in part on a relative deficiency of the angiostatic CXC chemokine, IFN-gamma-inducible protein-10 (IP-10). To test this hypothesis, we measured IP-10 by specific ELISA in whole lung homogenates in either bleomycin-treated or control mice and correlated these levels with lung hydroxyproline. We found that lung tissue from mice treated with bleomycin, compared with that from saline-treated controls, demonstrated a decrease in the presence of IP-10 that was correlated to a greater angiogenic response and total lung hydroxyproline content. Systemic administration of IP-10 significantly reduced BPF without any alteration in lung lymphocyte or NK cell populations. This was also paralleled by a reduction in angiogenesis. Furthermore, IP-10 had no direct effect on isolated pulmonary fibroblasts. These results demonstrate that the angiostatic CXC chemokine, IP-10, inhibits fibroplasia and deposition of extracellular matrix by regulating angiogenesis.  (+info)

Effects of 1alpha,25-dihydroxyvitamin D3 on Langerhans cell migration and corneal neovascularization in mice. (8/340)

PURPOSE: To examine the effects of 1alpha,25-dihydroxyvitamin D3 (1alpha,25[OH]2D3), a hormone that has immunosuppressive properties, on Langerhans cell (LC) migration and corneal neovascularization in mouse corneas. METHODS: Two 10-0 nylon interrupted sutures were placed in the center of 50 BALB/c mouse corneas to induce LC migration and corneal neovascularization. The mice were then randomly assigned to one of five groups. Three groups (n = 11, n = 11, n = 6) received topical 1alpha,25(OH)2D3 (at concentrations of 10(-7) M, 10(-)8 M, 10(-9) M), one group (n = 11) received vehicle only, and one group (n = 11) received no eye drops. Instillation (three times a day) began on the first day after suturing. Corneal neovascularization was assessed by slit lamp microscopy and scored according to the length of newly formed corneal vessels. Fourteen days after suturing, the number of LCs that had migrated into the central corneal epithelium was counted by an immunofluorescence assay using an anti-Ia antibody. RESULTS: The number of LCs in the central cornea was 21.9 +/- 2.8 cells/mm2 in the nontreated group and 17.8 +/- 3.9 cells/mm2 in the vehicle-only group. Significantly fewer LCs were detected in all groups that had received 1alpha,25(OH)2D3 compared with the vehicle only and nontreated groups (10(-7) M: 7.4 +/- 1.2 cells/mm2, 10(-8) M: 7.2 +/- 2.0 cells/mm2, 10(-9) M: 6.2 +/- 0.7 cells/mm2). Moderate inhibition of corneal vascularization was observed in the 10(-7) M 1alpha,25(OH)2D3 group, but not the other groups. CONCLUSIONS: Topical administration of 1alpha,25(OH)2D3 can be effective in suppressing ocular surface inflammation by inhibiting LC migration into mouse corneas.  (+info)

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