Role of urokinase plasminogen activator receptor in thrombospondin 1-mediated tumor cell invasion.
We previously showed that thrombospondin 1 (TSP-1) upregulates the plasminogen/plasmin system and promotes breast tumor cell invasion. Preliminary data from our laboratory using neutralizing antibodies suggested that the upregulation in breast tumor cell invasion seen in response to TSP-1 involved the urokinase plasminogen activator receptor (uPAR). To confirm these findings in MDA-MB-231 breast cancer cells, we developed three other strategies to study the role of uPAR in tumor cell adhesion and TSP-1-mediated tumor cell invasion: (a) enzymatic cleavage of uPAR with glycosylphosphatidylinositol-specific phospholipase C; (b) inhibition at the mRNA level with a uPAR antisense construct (cells named LKAS-MDA); (c) inhibition of plasminogen binding with the lysine analogue epsilon-aminocaproic acid. Adhesion to laminin and type I and type IV collagen with and without the addition of epsilon-aminocaproic acid was studied. Tumor cell invasion was studied in a modified Boyden chamber collagen invasion assay. Antisense uPAR inhibition decreased uPAR expression by 48-66% and cell-associated urokinase plasminogen activator (uPA) by 30-68%. Additionally, antisense uPAR inhibition induced a 68-70% reduction in uPA and plasmin activities. Antisense uPAR transfection increased tumor cell adhesion by 46-53%. A similar effect was observed in epsilon-aminocaproic acid-treated MDA-MB-231 cells. TSP-1-mediated tumor cell invasion was almost completely inhibited by either antisense uPAR inhibition or treatment with phospholipase C or epsilon-aminocaproic acid. We conclude that uPAR plays a crucial role in the regulation of tumor cell adhesion and TSP-1-mediated tumor cell invasion.
Albo
D
D
Department of Surgery, Medical College of Pennsylvania and Hahnemann University, Philadelphia, Pennsylvania, 19103, USA.
Berger
D H
DH
Rothman
V L
VL
Tuszynski
G P
GP
D016375:Antisense Elements (Genetics) / Q000494:pharmacology; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology* / Q000503:physiopathology; D002448:Cell Adhesion / Q000187:drug effects; D005260:Female; D005341:Fibrinolysin / Q000378:metabolism / Q000502:physiology; D006801:Humans; D009361:Neoplasm Invasiveness / Q000503:physiopathology; D010958:Plasminogen / Q000502:physiology; D012333:RNA, Messenger / Q000037:antagonists & inhibitors; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D055293:Receptors, Urokinase Plasminogen Activator; D019700:Thrombospondin 1 / Q000502:physiology*; D014162:Transfection; D014407:Tumor Cells, Cultured; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
D016375:Antisense Elements (Genetics); C524898:PLAUR protein, human; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D019700:Thrombospondin 1; D010958:Plasminogen; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator