Proinvasive properties of ovarian cancer ascites-derived membrane vesicles.
Malignant ovarian ascites are rich in cellular components, membrane-bound vesicles, and soluble proteins. This study focused on the structure of membrane-bound vesicles and their ability to promote invasion in cultured malignant ovarian epithelium. Membrane vesicles were derived from women with stage I-IV malignant ovarian ascites and from nonmalignant gynecologic ascites. Isolated vesicles were characterized by immunofluorescence and Western blot analysis. Using gel zymography for matrix metalloproteinase (MMP) detection and a colorimetric assay for urokinase-type plasminogen activator (uPA) analysis, we analyzed the proteinase activities of MMP-2, MMP-9, and uPA from the prepared vesicles, whole cells isolated from ascites, and the cell-free ultracentrifuged supernatant. The invasiveness of established cultured malignant ovarian epithelium on addition of ascites-derived vesicles was tested using a Matrigel-based invasion assay. Fractionation of malignant ascites revealed that extracellular matrix-degrading proteinases including MMPs and uPA are localized preferentially in membrane vesicles. All malignant vesicles tested, regardless of cancer stage, stimulated invasion. Furthermore, the combination of ovarian cancer cells and membrane vesicles resulted in greater uPA activation than that of cells or vesicles alone. Membrane vesicles from malignant ascites were also found to contain activated MMP-2, MMP-9, and uPA. Our data suggest that vesicle-stimulated proteinase activation leads to increased extracellular matrix degradation, which may facilitate tumor cell invasion and metastasis.
Department of Cellular and Molecular Biology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA.
Laura E
LE
Graves
Edgardo V
EV
Ariztia
Jason R
JR
Navari
Heather J
HJ
Matzel
M Sharon
MS
Stack
David A
DA
Fishman
D001201:Ascites / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000201:enzymology / Q000473:pathology; D004789:Enzyme Activation; D066246:ErbB Receptors / Q000378:metabolism; D005260:Female; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000378:metabolism; D009361:Neoplasm Invasiveness; D009367:Neoplasm Staging; D010051:Ovarian Neoplasms / Q000201:enzymology* / Q000378:metabolism / Q000473:pathology*; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
D019012:Integrin beta1; D066246:ErbB Receptors; D014568:Urokinase-Type Plasminogen Activator; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9