Modulation of procoagulant and fibrinolytic properties of endothelial cells mediated by adhesion with activated T lymphocytes was studied by focusing on the CD40-CD40 Ligand (CD40L) interaction. Cultured human umbilical vein endothelial cells (HUVECs) were treated with interferon-γ to increase the expression of CD40, and co-cultured with T lymphocytes that had been isolated from peripheral blood, activated with PMA and ionomycin to induce the expression of CD40L and fixed with 1% paraformaldehyde. This co-culture gave rise to the generation of tissue factor (TF) on the surface of HUVECs, but co-culture of the HUVECs with non-activated T lymphocytes failed to do so. TF expressed on the HUVECs was specifically but only partially inhibited by an anti-CD40L monoclonal antibody. By contrast, co-culture of HUVECs with the activated and fixed T lymphocytes resulted in both down-regulation of thrombomodulin in HUVECs and decrease of release of tissue factor pathway inhibitor from HUVECs. Futhermore, the co-culture enhanced the release of tissue-type plasminogen activator (t-PA) from the HUVECs without affecting the release of plasminogen activator inhibitor-1, which was still in great excess of t-PA. These results altogether suggest that the CD40-CD40L-involved adhesion of activated T lymphocytes to endothelial cells that could occur at the inflammatory sites is likely to shift the nature of endothelial cell surface toward the thrombogenic state.

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A 9 years old Japanese non-albino girl has presented with mild to moderate nosebleed. Routine studies revealed normal counts and sizes of the patient's platelets and normal coagulation tests but prolonged bleeding times (over 12min). The platelets showed normal shape change in response to ADP. Aggregations of the platelets by ADP and collagen were significantly delayed in onset and weaker than controls, while aggregation/agglutination by epinephrine, thrombin, A23187 or ristocetin were normal, suggesting that the patient's platelets are abnormal only in ADP-induced activation. The platelet ATP secretion by 20μg/ml collagen was about half of controls by a Lumi-aggregometer. ³H-ADP binding to the patient's platelets as well as Ca²⁺ influx and mobilization induced by ADP were normal, suggesting a normal quantity of three ADP receptors and a normal function of P2X₁, a component of the receptors. Immunoblots revealed that Giα subunit, a G-protein associated with P2Yac that is the second component of the receptors, was decreased in the patient's platelets than controls, but Gqα subunit, a G-protein associated with P2Y₁ that is the third component of the receptors, was normal in the patient's platelets. Stimulation of PGE₁-pretreated platelets by 10μM ADP or 10μM epinephrine reduced the cAMP levels in the patient's platelets to the same extent as in control platelets, indicating a normal function of P2Yac in spite of the decreased Giα subunit. The patient's platelet abnormality characterized with defective activation induced by ADP appears to originate from defective coordination between decreased Giαsubunit and other ADP receptors.

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