Matrix-degrading proteinases are shed in membrane vesicles by ovarian cancer cells in vivo and in vitro

The in vitro release of matrix-degrading proteinases from breast cancer cel ls is associated in part with shed membrane vesicles. To determine whether shed vesicles might play a similar role in ovarian cancer cells, we analyze d the shedding phenomenon in vivo and in vitro as well as the enzymatic con tent of their vesicles. This is the first time that an immunoelectron micro scopical analysis revealed membrane vesicles carrying tumor-associated anti gen alpha-Folate Receptor (alpha-FR), circulating in biological fluids (asc ites and serum) of an ovarian carcinoma patient. These vesicles were trappe d in a fiber network with characteristic fibrin periodicity. An ovarian can cer cell line (CABA I) established from ascitic fluid cells of this patient , grew in Matrigel and formed tubular structures suggesting invasive capabi lity. Immunofluorescence analysis demonstrated strong cytoplasmic staining of CABA I cells with anti-matrix metalloproteinase-9 (MMP-9) and anti-uroki nase-type plasminogen activator (uPA) antibodies. CABA I cells shed membran e vesicles, which were morphologically similar to those identified in vivo, as determined by electron microscopy. Gelatin zymography of vesicles isola ted both in vivo and in vitro revealed major gelatinolytic bands of the MMP family, identified as the zymogen and active forms of gelatinase B (MMP-9) and gelatinase A (MMP-2). By casein-plasminogen zymography we observed hig h-molecular weight (HMW)-uPA and plasmin bands. Incubation of purified vesi cles from CABA I cells with Matrigel led to cleavage of Matrigel components . Taken together, our results point to a possible role of shed vesicles, bo th in vivo and in vitro, in proteolysis that mediates invasion and spread o f ovarian epithelial carcinoma cells.