Therapeutic immunoglobulin reduces Ca2+ mobilization and von Willebrand factor secretion, and increases nitric oxide release in human endothelial cells.
Intravenous gamma-immunoglobulin (i.v.Ig) is commonly used in the treatment of autoimmune and inflammatory vascular disorders to prevent thrombotic complications. The mechanism of action of i.v.Ig is, however, not yet elucidated. In view of this, we investigated the ability of i.v.Ig to modulate i) Ca(2+) signals of fura-2 loaded endothelial cells, and ii) the associated release of nitric oxide (NO) and von Willebrand factor (vWf). NO was measured either indirectly by radioimmunoassay of cGMP in unstimulated cells or directly by electrochemistry at the surface of stimulated endothelial cells from human umbilical cord veins (HUVEC). Short-term treatment of unstimulated HUVEC with intact i.v.Ig decreased the basal cytosolic Ca(2+) concentration by 20% while it activated the NO/cGMP synthesis. Following i.v.Ig treatment of HUVEC, the Ca(2+) liberation from internal stores and the vWf secretion induced by ATP, thrombin or histamine were significantly reduced by 38 and 60%, respectively. The effects on Ca(2+) signals were observed with intact i.v.Ig as well as with the F(ab')2 or the Fc fragments indicating that both portions are involved in the mechanism of action. The i.v.Ig treatment of HUVECs had no effect on the NO release induced by thrombin or histamine. By contrast, the i.v.Ig treatment increased the ATP-activated NO release by amplifying the Ser1177-eNOS phosphorylation. The i.v.Ig also activated the NO-dependent cGMP release in resting and collagen-stimulated platelets. Since NO is a potent inhibitor of platelet activation and vWF is a platelet adhesion cofactor, the beneficial effects of therapeutic i.v.Ig may lie in the inhibition of platelet adhesion to damaged endothelium.
David-Dufilho
Monique
M
Research unit Cellular signalling and early arthersclerosis, CNRS UMR 7131, Hôpital Broussais, 102 rue Didot, 75764 Paris Cedex 14, France. monique.dufilho@brs.ap-hop-paris.fr
Schussler
Olivier
O
Pernollet
Marie-Gabrielle
MG
Brunet
Annie
A
Millanvoye-Van Brussel
Elisabeth
E
Le Quan Sang
Kim Hahn
KH
Rendu
Francine
F
D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects*; D006152:Cyclic GMP / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D016756:Immunoglobulins, Intravenous / Q000494:pharmacology*; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III; D010973:Platelet Adhesiveness / Q000187:drug effects; D014471:Umbilical Veins; D014841:von Willebrand Factor / Q000187:drug effects / Q000378:metabolism*
D016756:Immunoglobulins, Intravenous; D014841:von Willebrand Factor; D009569:Nitric Oxide; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; D006152:Cyclic GMP