Plasminogen activation by t-PA on the surface of human melanoma cells in the presence of alpha 2-macroglobulin secretion.
Several human melanoma cell lines produced tissue-type plasminogen activator (t-PA), as detected by zymography and immunocapture assay of culture media and cell lysates. Urokinase (u-PA) was found at only less than or equal to 1% the level of t-PA. Acid eluates of the cell surface indicated that the melanoma cells had t-PA bound on their surface, but no u-PA, and also had a very low capacity to bind exogenous u-PA. After incubation of the melanoma cells with 10% plasminogen-depleted fetal calf serum and human plasminogen, bound plasmin activity could be eluted from the cell surface with tranexamic acid, an analogue of lysine. This indicated that plasminogen was activated on the cell surface. The cell-surface plasmin formation was inhibited by an anti-catalytic monoclonal antibody to human t-PA, and not by an anti-catalytic antibody to u-PA. The melanoma cells also synthesized and secreted alpha 2-macroglobulin (alpha 2M), as shown by alpha 2M-specific mRNA in Northern blotting and detection of alpha 2M protein in conditioned cell culture media. The media were found to inhibit u-PA but not t-PA. This inhibition was related to their alpha 2M content, and immunoabsorption of alpha 2M removed the inhibitory activity. These studies suggest that t-PA can bind to the surface of melanoma cells and generate surface-bound plasmin. Because t-PA and cell-bound plasmin are unaffected by alpha 2M, t-PA may, in the case of melanoma cells, serve an analogous function to u-PA in supporting tumor cell invasion.
Bizik
J
J
Department of Virology, University of Helsinki, Finland.
Lizonová
A
A
Stephens
R W
RW
Grófová
M
M
Vaheri
A
A
D000911:Antibodies, Monoclonal / Q000276:immunology; D007611:Aprotinin / Q000494:pharmacology; D015152:Blotting, Northern; D002462:Cell Membrane / Q000378:metabolism; D003470:Culture Media; D015342:DNA Probes; D005341:Fibrinolysin / Q000378:metabolism; D006801:Humans; D008545:Melanoma / Q000378:metabolism*; D010958:Plasminogen / Q000378:metabolism*; D010960:Plasminogen Activators / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D010959:Tissue Plasminogen Activator / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism; D000511:alpha-Macroglobulins / Q000378:metabolism* / Q000502:physiology
D000911:Antibodies, Monoclonal; D003470:Culture Media; D015342:DNA Probes; D012333:RNA, Messenger; D000511:alpha-Macroglobulins; D010958:Plasminogen; D007611:Aprotinin; D010960:Plasminogen Activators; D010959:Tissue Plasminogen Activator; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator