Membrane type-1 matrix metalloproteinase stimulates tumour cell-induced platelet aggregation: role of receptor glycoproteins.
1. Matrix metalloproteinase-2 (MMP-2) plays a role in agonist- and tumour cell-induced platelet aggregation (TCIPA). 2. MMP-2 is synthesized as a proenzyme and is activated at the cell surface by membrane type-1 matrix metalloproteinase (MT1-MMP, MMP-14). 3. The significance of tumour cell-associated MT1-MMP for TCIPA was investigated using human breast carcinoma MCF7 cells stably coexpressing the integrin alphavbeta3 with MT1-MMP, cells expressing alphavbeta3 alone and mock-transfected cells. 4. Western blot and zymography confirmed that alphavbeta3/MT1-MMP cells expressed MT1-MMP and efficiently processed proMMP-2 to MMP-2. 5. Aggregometry, phase-contrast and transmission electron microscopy and flow cytometry were used to characterize TCIPA induced by MCF7 cell lines. 6. The aggregating potency of cells was: alphavbeta3/MT1-MMP >alphavbeta3=mock cells, as shown by aggregometry and phase-contrast microscopy. 7. Electron microscopy revealed close, membrane-membrane interactions between activated platelets and alphavbeta3/MT1-MMP cells during TCIPA. 8. Inhibition of MMP-2 with the neutralizing anti-MMP-2 antibody (5 microg ml(-1)) and o-phenanthroline (100 microm) reduced aggregation induced by alphavbeta3/MT1-MMP cells. 9. TCIPA induced by alphavbeta3/MT1-MMP cells was also reduced by inhibiting the generation and actions of ADP with apyrase (250 microg ml(-1)) and 2-methylthio-AMP (2-MeSAMP) (30 microm), but not N(6)-methyl-2'-deoxyadenosine-3',5'-bisphosphate (MRS2179) (30 microm). 10. Flow cytometry demonstrated that TCIPA enhanced expression of glycoprotein (GP) Ib and IIb/IIIa receptors not only on platelets but also on breast cancer cells. 11. Thus, (a) human breast carcinoma cell surface-associated MT1-MMP, via activating proMMP-2, stimulates TCIPA; (b) ADP amplifies the effects of MMPs via stimulation of P2Y(12) receptors and (c) both tumour- and platelet-derived GPIb and GPIIb/IIIa are involved in the aggregatory effects of MT1-MMP.
Alonso-Escolano
David
D
Department of Integrative Biology and Pharmacology, University of Texas, Houston, TX 77030, USA.
Strongin
Alex Y
AY
Chung
Ada W
AW
Deryugina
Elena I
EI
Radomski
Marek W
MW
D001943:Breast Neoplasms / Q000097:blood / Q000201:enzymology / Q000235:genetics; D006801:Humans; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000096:biosynthesis* / Q000235:genetics; D010974:Platelet Aggregation / Q000502:physiology*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000502:physiology*; D014162:Transfection / Q000379:methods; D014407:Tumor Cells, Cultured
D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases