angiogenin and Corneal-Neovascularization

Angiogenin (ANG), a component of tears, is involved in the innate immune system and is related with inflammatory disease. We investigated whether ANG has an immune modulatory function in human corneal fibroblasts (HCFs).. HCFs were cultured from excised corneal tissues. The gene or protein expression levels of interleukin (IL)-1beta (β), IL-4, IL-6, IL-8, IL-10, complements, toll-like receptor (TLR)4, myeloid differentiation primary response gene (MYD)88, TANK-binding kinase (TBK)1, IkappaB kinase-epsilon (IKK-ε) and nuclear factor-kappaB (NF-κB) were analyzed with or without ANG treatment in tumor necrosis factor-alpha (TNF-α)- or lipopolysaccharide (LPS)-induced inflammatory HCFs by real-time polymerase chain reaction (PCR), Western blotting and immunocytochemistry. Inflammatory cytokine profiles with or without ANG were evaluated through immunodot blot analysis in inflammatory HCFs. Corneal neovascularization and opacity in a rat model of corneal alkali burn were evaluated after application of ANG eye drops.. ANG decreased the mRNA levels of IL-1β, IL-6, IL-8, TNF-α receptor (TNFR)1, 2, TLR4, MYD88, and complement components except for C1r and C1s and elevated the mRNA expression of IL-4 and IL-10. Increased signal intensity of IL-6, IL-8 and monocyte chemotactic protein (MCP)-1 and MCP-2 induced by TNF-α or LPS was weakened by ANG treatment. ANG reduced the protein levels of IKK-ε by either TNF-α and LPS, and decreased TBK1 production induced by TNF-α, but not induced by LPS. The expression of NF-κB in the nuclei was decreased after ANG treatment. ANG application lowered corneal neovascularization and opacity in rats compared to controls.. These results demonstrate that ANG reduces the inflammatory response induced by TNF-α or LPS in HCFs through common suppression of IKK-ε-mediated activation of NF-κB. This may support the targeting of immune-mediated corneal inflammation by using ANG.

Topics: Angiogenesis Inducing Agents; Animals; Blotting, Western; Burns, Chemical; Chemokines; Cornea; Corneal Neovascularization; Corneal Opacity; Cytokines; Disease Models, Animal; Eye Burns; Fibroblasts; Humans; Immunity, Innate; Immunohistochemistry; Interleukins; Male; Polymerase Chain Reaction; Rats; Rats, Sprague-Dawley; Ribonuclease, Pancreatic; RNA, Messenger

S179D prolactin (PRL) is an experimentally useful mimic of naturally phosphorylated human prolactin. S179D PRL, but not unmodified PRL, was found to be anti-angiogenic in both the chorioallantoic membrane and corneal assays. Further investigation using human endothelial in vitro models showed reduced cell number, reduced tubule formation in Matrigel, and reduced migration and invasion, as a function of treatment with S179D PRL. Analysis of growth factors in human endothelial cells in response to S179D PRL showed: a decreased expression or release of endogenous PRL, heme-oxygenase-1, basic fibroblast growth factor (bFGF), angiogenin, epidermal growth factor and vascular endothelial growth factor; and an increased expression of inhibitors of matrix metalloproteases. S179D PRL also blocked signaling from bFGF in these cells. We conclude that this molecular mimic of a pituitary hormone is a potent anti-angiogenic protein, partly as a result of its ability to reduce utilization of several well-established endothelial autocrine growth loops, partly by its ability to block signaling from bFGF and partly because of its ability to decrease endothelial migration. These findings suggest that circulating levels of phosphorylated PRL may influence the progression of cancer and, furthermore, that S179D PRL may be a useful anti-angiogenic therapeutic.

Topics: Angiogenesis Inhibitors; Animals; Cell Movement; Chickens; Chorioallantoic Membrane; Collagen; Corneal Neovascularization; Drug Combinations; Endothelium, Vascular; Epidermal Growth Factor; Fibroblast Growth Factor 2; Gene Expression; Heme Oxygenase-1; Humans; Laminin; Matrix Metalloproteinases; Mice; Molecular Mimicry; Phosphorylation; Prolactin; Proteoglycans; Rats; Rats, Sprague-Dawley; Ribonuclease, Pancreatic; Umbilical Veins; Vascular Endothelial Growth Factor A

Angiostatin is a potent angiogenesis inhibitor that has been identified as a cryptic fragment of plasminogen molecule containing the first four kringle domain. Angiogenin, a 14-kDa monomeric protein, a potent blood vessel inducer, is expressed in tumors and present in mammalian plasma. The purpose of this study was to determine whether recombinant kringle 1-3 (rKI-3) of human plasminogen could interfere with angiogenesis induced by angiogenin and to evaluate the role of angiogenin in corneal angiogenesis in rabbit.. A Hydron polymer pellet containing 2.0 microg of angiogenin was implanted intrastromally into the superior cornea of each of 44 rabbit eyes. All eyes received an intrastromal pellet and were randomized into either one group treated with 12.5 microg of rKI-3 (n = 25) or the other group treated with phosphate-buffered saline (PBS; n = 19). Both pellets were positioned in parallel at the site 1.2 mm from the superior limbus. Two masked observers kept the angiogenesis score daily, based on the number and the length of new vessels. The corneas with induced angiogenesis also were examined histologically.. On the third day of the angiogenin pellets implantation, the eye treated with rKI-3 had less angiogenesis (mean score, 4.2 +/- 6.6) than eye treated with PBS (mean score, 16.1 +/- 17.1; p < 0.05, Mann-Whitney U test). The cornea treated with PBS also showed much more leukocyte adhesion than the cornea treated with rKI-3.. Recombinant kringle 1-3 appears to inhibit angiogenin-induced angiogenesis in a rabbit corneal pocket assay. Recombinant kringle 1-3 may have therapeutic potential as an antiangiogenic agent.

Topics: Angiogenesis Inducing Agents; Animals; Antineoplastic Agents; Cell Adhesion; Cornea; Corneal Neovascularization; Disease Models, Animal; Drug Implants; Leukocytes; Male; Peptide Fragments; Plasminogen; Rabbits; Random Allocation; Recombinant Proteins; Ribonuclease, Pancreatic