ABSTRACT
Bactericidal/permeability-increasing protein (BPI) was originally identified as a lipopolysaccharide (LPS) binding protein with gram-negative bactericidal activity in the leukocytes. In this study, we characterized the previously unknown effects of BPI in the eye and the molecular mechanisms involved in its action. BPI mRNA was detected in bovine retina; retinal pigment epithelium; and primary cultures of bovine retinal pigment epithelial cells (RPE), pericytes (RPC), and endothelial cells (REC); while BPI protein was measured in human vitreous and plasma. BPI, but not control protein thaumatin, activated extracellular regulated kinase (ERK) and AKT, and increased DNA synthesis in RPE and RPC but not in REC. A human recombinant 21 kDa modified amino-terminal fragment of BPI (rBPI21) reduced H2O2-induced apoptosis in RPE and inhibited vascular endothelial growth factor (VEGF)-stimulated ERK phosphorylation in REC when preincubated with VEGF. Intraperitoneal (i.p.)-injected rBPI21 reduced ischemia-induced retinal neovascularization and diabetes-induced retinal permeability. Since BPI has unusual dual properties of promoting RPC and RPE growth while suppressing VEGF-induced REC growth and vascular permeability, the mechanistic understanding of BPI's action may provide novel therapeutic opportunities for diabetic retinopathy and age-related macular degeneration.
Subject(s)
Angiogenesis Inhibitors/pharmacology , Antimicrobial Cationic Peptides/pharmacology , Blood Proteins/pharmacology , Membrane Proteins/pharmacology , Retina/cytology , Retinal Vessels/drug effects , Signal Transduction , Animals , Antimicrobial Cationic Peptides/analysis , Apoptosis/drug effects , Blood Proteins/analysis , Capillary Permeability/drug effects , Cattle , Cells, Cultured , Endothelial Cells/cytology , Endothelial Cells/drug effects , Endothelial Cells/metabolism , Humans , Membrane Proteins/analysis , Neovascularization, Pathologic/drug therapy , Pericytes/cytology , Pericytes/drug effects , Pericytes/metabolism , Pigment Epithelium of Eye/cytology , Pigment Epithelium of Eye/drug effects , Pigment Epithelium of Eye/metabolism , Plasma/chemistry , Recombinant Proteins/pharmacology , Retina/drug effects , Retina/metabolism , Retinal Vessels/growth & development , Vascular Endothelial Growth Factor A/pharmacology , Vitreous Body/chemistryABSTRACT
To evaluate whether beta-catenin signaling has a role in the regulation of angiogenesis in colon cancer, a series of angiogenesis-related gene promoters was analyzed for beta-catenin/TCF binding sites. Strikingly, the gene promoter of human vascular endothelial growth factor (VEGF, or VEGF-A) contains seven consensus binding sites for beta-catenin/TCF. Analysis of laser capture microdissected human colon cancer tissue indicated a direct correlation between up-regulation of VEGF-A expression and adenomatous polyposis coli (APC) mutational status (activation of beta-catenin signaling) in primary tumors. In metastases, this correlation was not observed. Analysis by immunohistochemistry of intestinal polyps in mice heterozygous for the multiple intestinal neoplasia gene (Min/+) at 5 months revealed an increase and redistribution of VEGF-A in proximity to those cells expressing nuclear beta-catenin with a corresponding increase in vessel density. Transfection of normal colon epithelial cells with activated beta-catenin up-regulated levels of VEGF-A mRNA and protein by 250-300%. When colon cancer cells with elevated beta-catenin levels were treated with beta-catenin antisense oligodeoxynucleotides, VEGF-A expression was reduced by more than 50%. Taken together, our observations indicate a close link between beta-catenin signaling and the regulation of VEGF-A expression in colon cancer.