ABSTRACT
The aspartyl-protease cathepsin D (cath-D) is overexpressed and hypersecreted by epithelial breast cancer cells and stimulates their proliferation. As tumor epithelial-fibroblast cell interactions are important events in cancer progression, we investigated whether cath-D overexpression affects also fibroblast behavior. We demonstrate a requirement of cath-D for fibroblast invasive growth using a three-dimensional (3D) coculture assay with cancer cells secreting or not pro-cath-D. Ectopic expression of cath-D in cath-D-deficient fibroblasts stimulates 3D outgrowth that is associated with a significant increase in fibroblast proliferation, survival, motility, and invasive capacity, accompanied by activation of the ras-MAPK pathway. Interestingly, all these stimulatory effects on fibroblasts are independent of cath-D proteolytic activity. Finally, we show that pro-cath-D secreted by cancer cells is captured by fibroblasts and partially mimics effects of transfected cath-D. We conclude that cath-D is crucial for fibroblast invasive outgrowth and could act as a key paracrine communicator between cancer and stromal cells, independently of its catalytic activity.
Subject(s)
Cathepsin D/physiology , Cell Movement/physiology , Fibroblasts/cytology , Animals , Apoptosis/physiology , Butadienes/pharmacology , Cathepsin D/genetics , Cathepsin D/metabolism , Cell Enlargement/drug effects , Cell Growth Processes/drug effects , Cell Growth Processes/physiology , Cell Line , Cell Line, Tumor , Cell Movement/drug effects , Cell Proliferation/drug effects , Cell Survival/physiology , Coculture Techniques , Culture Media, Conditioned/pharmacology , Endocytosis/drug effects , Enzyme Inhibitors/pharmacology , Enzyme Precursors/metabolism , Enzyme Precursors/physiology , Fibroblasts/drug effects , Fibroblasts/ultrastructure , Humans , Mannosephosphates/pharmacology , Mice , Microscopy, Electron , Mitogen-Activated Protein Kinase 1/metabolism , Mitogen-Activated Protein Kinase 3/metabolism , Neoplasm Invasiveness , Neoplasms, Glandular and Epithelial/enzymology , Neoplasms, Glandular and Epithelial/metabolism , Neoplasms, Glandular and Epithelial/pathology , Nitriles/pharmacology , Paracrine Communication/physiology , Phosphorylation/drug effects , RNA, Small Interfering/genetics , Transfection , Wound HealingABSTRACT
PURPOSE: To evaluate the presence and potential involvement of members of the plasminogen/plasminogen activator (Plg/PA) system in the exudative form of age-related macular degeneration (AMD). METHODS: The expression of PA members mRNA was evaluated in human and experimental choroidal neovascularization (CNV) by RT-PCR. The presence and activity of PA was studied by immunofluorescence and in situ zymography. The influence of endogenous plasminogen (Plg), urokinase (uPA), tissue type plasminogen activator (tPA), and uPA receptor (uPAR) was explored in single-gene-deficient mice in a model of laser-induced CNV. RESULTS: Members of the Plg/PA system were present both in human and murine CNV. The absence of Plg, uPA, or tPA significantly decreased the development of experimental CNV compared with wild-type or uPAR-deficient mice. This effect could be attributable, partly to a modulation of matrix metalloproteinase activity, but also to an accumulation of fibrinogen-fibrin in the laser-induced wounds. CONCLUSIONS: Together with previous work done by the authors, this study indicates that choroidal neovascularization is extremely sensitive to the modulation of Plg/PA system activity. This may provide a new strategy for the treatment of exudative AMD.
