Adhesion molecules expressed on platelets play important roles for in the interaction between platelets and vascular endothelial cells.
However, little is known whether megakaryocytes bear the same adhesion molecules as those expressed on platelets.
In this study, we have examined the expression of adhesion molecules, integrin family, immunoglobulin superfamily, selectin family and CD44 family on the purified megakaryocytes, in comparison with those on platelets.
Megakaryocytes were purified by Percoll density centrifugation followed by albumin gravity sedimentation. Megakaryocytes doubly stained by anti-GP IIb/IIIa (Plt-1) and anti-adhesion molecule antibodies were analyzed by flowcytometer. All the adhesion molecules expressed on platelets were also expressed on megakaryocytes. In addition, among tested adhesion molecules, endothelial-leukocyte adhesion molecule-1 (ELAM-1) was detected on megakaryocytes whereas not on platelets. This study may help for understanding the expression of adhesion molecules during megakaryocytic maturation.

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We describe here a simple enzyme-linked immunosorbent assay (ELISA) for the quantiation of GPIb of freshly-prepared washed platelets. This assay used rabbit antihuman platelet antibody as a platelet-capturing antibody and an anti-GPIb monoclonal antibody (OP-F1) conjugated with horseraddish peroxidase as a second antibody. The lowest concentration of GPIb antigen measurable by this ELISA was 0.01% of normal platelets when determined using a platelet concentration of 10⁵/μl. The average value (mean±2SD) of platelet GPIb antigen from 30 normal individuals was 93.1±20.3%. Two unrelative patients with Bernard-Soulier syndrome showed less than 0.1% and 4.5%, respectively of normal platelet GPIb. In 30 patients with chronic renal failure (CRF), the GPIb antigen levels (43.1±19.9%) were also significantly reduced (p<0.001). These results also suggest that hemorrhagic tendency in patients with CRF is due to, in part, the decreased levels of GPIb.

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A novel antiplatelet agent, satigrel (E5510), was administered to 9 patients with arteriosclerosis obliterans (ASO) and 6 patients with thromboangiitis obliterans (TAO) at a dosage of 4mg/day for 2 weeks followed by 8mg/day for another 2 weeks and its effect on platelet function and coagulation-fibrinolytic system as well as hemodynamics were evaluated. Satigrel significantly inhibited platelet aggregation induced by collagen (p<0.001), lowered plasma thromboxane B₂ (TXB₂) (p<0.01) and 11-dehydro TXB₂ (p<0.1), and increased plasma antithrombin III (AT-III) (p<0.001). Hopwever, there were no significant change in plasma levels of β-thromboglobulin (β-TG), platelet factor 4 (PF4), 6-keto prostaglandin F₁α (6-KF), thrombin-antithrombin III complex (TAT), fibrinopeptide A (FPA) and plasmin-α₂ plasmin inhibitor complex (PIC). Plasma D-Dimer and fibrinopeptide Bβ_15-42 (FPBβ_15-42) levels were higher in ASO patients than in TAO patients before treatment and tended to increase in both groups following the administration of satigrel. Furthermore, satigrel improved tissue blood flow in the affected feet (p<0.05) even though clinical improvement and increase in ankle pressure index were not significant. These results suggest that satigrel is beneficial to a failed circulation in these arterial occlusive diseases.

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This paper describes a quantitative assay for the binding ability of plasma factor VII to human tissue factor (TF) on microplate. Plasma samples from eight patients with congenital factor VII abnormality were measured by this method. The procedure is simple for using recombinant TF from commercial prothrombin time reagent, and non isotopic for using anti-factor VII IgG labeled biotin. Its sensitivity to detect factor VII binding ability to tissue factor was about 3% of normal subjects. The factor VII-TF binding ability was inhibited by monoclonal anti-TF antibody that is characterized by its inhibition of interaction between factor VII and TF. Being partially detected immediately after the addition of the plasma sample on the wells of micro-plate coated with TF, the factor VII binding to TF had completely achieved in 2hr at 37°C or 8hr at 4°C. Measured factor VII binding ability to TF in normal subjects was 101.17±18.90% (X±SD), and the value correlated well both with factor VII activity (r=0.788) and antigen (r=0.662). Two homozygous subjects with the mutation of Arg 79 to Gln in factor VII (Human Molecular Genetics, 2: 1335, 1994) had a half level of a normal subject for factor VII-TF binding. Two heterozygous subjects who are sons of the homozygous subjects had lower levels in the normal range as for factor VII-TF binding. One of 4 patients with congenital factor VII abnormality whose mutation site had not been detected yet, had 26% of normal subjects. Three of the 4 patients had the normal range as for factor VII-TF binding.

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A technique is described for the induction and registration of changes in vasomotor tone in mesenteric arterioles of the guinea pig. Through a platinum electrode, insulated except at the tip and positioned at a fixed distance from the selected arteriolar segment, a negative direct current (1μA) applied for 20 seconds induced a vasoconstriction followed by relaxation within three minutes. If experimental conditions remained unchanged, arteriolar constriction could be induced every 4 minutes, repeated 10 times in succession, and with almost identical changes in vasomotor tone. Quantification was performed on video recordings of the investigated arteriolar segment by measuring the intraluminal diameter directly from the image on the monitor screen. Visual inspection, confirmed by statistical analysis, demonstrated that the constriction-relaxation phenomena of the arteriole were fully reproducible at the site directly opposite the electrode. In the applied experimental protocol, the arteriole served as its own control for evaluation of the effect of drug administration. Phentolamine as well as carbacyclin, although affecting vasomotricity through different mechanisms, both significantly decreased the standardised constrictive responses. Our findings clearly indicate that other drugs and/or mediators deserve investigation by this method in order to unravel their vasoactive properties in in vivo conditions.

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Thrombomodulin (TM) is a surface glycoprotein that forms a stoichiometric complex with thrombin thereby serving as a natural anticoagulant on endothelium of arteries, veins, capillaries, and lymphatics. Recently, it was shown that monocytes and macrophages express TM in vitro (McCachren et al, 1991). We here studied immunohistochemical expression of TM on inflammatory exudating cells in granulomatous inflammation. Tissue sections from sarcoidosis were reacted with anti-human recombinant TM polyclonal rabbit antiserum, and stained with avidin-biotin peroxidase complex immunohistochemistry or indirect immunofluorescence techniques. Results clearly showed that a subpopulation of CD11b-positive monocytic histiocytes infiltrating into the tissue is TM-positive. The TM-positive inflammatory cells are round, cuboidal and dendritic in cell morphology and they surround the granulomas but not observed in the center of organized granulomas where lysozyme-positive and CD68-positive epithelioid and multinucleated cells were accumulated. The results indicate that TM is potentially a unique and useful immunomarker of monocytic histiocytes particularly in the study of chronic granulomatous inflammation.

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