Distinct spatio-temporal Ca2+ signaling elicited by integrin alpha2beta1 and glycoprotein VI under flow.
We studied how integrin alpha2beta1 and glycoprotein VI (GPVI) contribute to collagen-induced platelet activation under flow conditions by evaluating stable adhesion and intracellular Ca(2+) concentration ([Ca(2+)](i)) of FLUO 3-AM-labeled platelets perfused over acid-soluble type I or microfibrillar type VI collagen. Adhering platelets displayed 2 kinds of [Ca(2+)](i) oscillations. Rapid alpha-like peaks were unaffected by the membrane-impermeable Ca(2+) chelator ethyleneglycoltetraacetic acid but abolished by membrane-permeable BAPTA-AM. Longer-lasting gamma-like peaks were always preceded by at least one alpha-like peak and abolished by intracellular or extracellular Ca(2+) chelation. Inhibition of phosphatidylinositol 3-kinase or phospholipase C and modulation of cyclic nucleotides, but not blockage of adenosine diphosphate receptors, prevented both Ca(2+) responses. Human or mouse platelets lacking GPVI function exhibited alpha-like but not gamma-like Ca(2+) peaks, whereas those lacking alpha2beta1 showed markedly reduced to absent alpha-like and no gamma-like Ca(2+) peaks. Specific alpha2beta1 ligation induced alpha-like but not gamma-like peaks. Thus, alpha2beta1 may generate Ca(2+) signals that are reinforced by GPVI and required for subsequent longer-lasting Ca(2+) oscillation mediated by GPVI through transmembrane ion flux. Our results delineate a GPVI-independent signaling role of alpha2beta1 in response to collagen stimulation.
Mazzucato
Mario
M
Department of Laboratory Medicine, Centro di Riferimento Oncologico (CRO) National Cancer Institute, Istituti di Ricovero e Cura a Carattere Scientifico (IRCCS), Aviano (PN), Italy.
Cozzi
Maria Rita
MR
Battiston
Monica
M
Jandrot-Perrus
Martine
M
Mongiat
Maurizio
M
Marchese
Patrizia
P
Kunicki
Thomas J
TJ
Ruggeri
Zaverio M
ZM
De Marco
Luigi
L
D000818:Animals; D001775:Blood Circulation / Q000502:physiology*; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D020013:Calcium Signaling / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D024042:Collagen Type I / Q000378:metabolism / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D038982:Integrin alpha2beta1 / Q000235:genetics / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009025:Morpholines / Q000494:pharmacology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010973:Platelet Adhesiveness / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D010980:Platelet Membrane Glycoproteins / Q000235:genetics / Q000502:physiology*; D013997:Time Factors
D002867:Chromones; D024042:Collagen Type I; D004791:Enzyme Inhibitors; D038982:Integrin alpha2beta1; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010980:Platelet Membrane Glycoproteins; C098341:platelet membrane glycoprotein VI; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one