G. de Gaetano

Retraction of reptilase-clots in the presence of agents inducing or inhibiting the platelet adhesion-aggregation reaction

Abstract Human platelet-rich plasma clotted by reptilase does not retract. This seems to be due to the lack of activation of platelets exposed to this enzyme rather than to the incomplete hydrolysis of fibrinogen. Indeed, when aggregating agents such as ADP, collagen, adrenaline or Thrombofax are preincubated with unstirred PRP before adding reptilase, strong clot retraction occurs. A number of antiaggregating compounds has been evaluated in this test system. Only specific inhibitors of the release reaction were unable to influence ADP-reptilase or collagen-reptilase clot retraction. This procedure allows simple and sensitive evaluation of certain agents inhibiting the platelet adhesion-aggregation reaction. A scheme linking the platelet adhesion-aggregation reaction and clot retraction and indicating the level of inhibition of several compounds is proposed.

Plasmatic regulation of vascular prostacyclin in pregnancy.

Activity of prostacyclin-stimulating factor was measured in six normal, non-pregnant women, six women in early normal pregnancy, six in late normal pregnancy, and six in late pregnancy complicated by severe pre-eclampsia. The activity was lower in the women in late pregnancy than in those in early pregnancy and the controls but was about normal in those with severe pre-eclampsia. These results may be relevant to the physiology of pregnancy and the pathogenesis of pre-eclampsia.

Bleeding in renal failure: is von Willebrand factor implicated?

Normal or increased concentrations of factor VIII procoagulant activity (VIIIC), factor VIII-related antigen (VIIIRA), and factor VIII-von Willebrand activity (VIIIVWF) were found in the predialysis plasma of 10 out of 11 patients with chronic renal failure (CRF). All patients had a bleeding time longer than 15 minutes and platelet retention to glass beads lower than 40%. The only patient who had reduced concentrations of all three factor VIII complex components was subsequently shown to have von Willebrands disease. In four patients with CRF, very low platelet retention, and slightly prolonged bleeding time none of the three factor VIII COMPLEX COMPONENTS WERE SELECTIVely modified in predialysis samples. These findings suggest that the bleeding tendency common in CRF is not necessarily linked to defective plasma factor VIII-related activities.

Effect of platelets on clot structuration. A thrombelastographic study

Abstract Reptilase induces in the thrombelastograph (TEG) plasma clots which are different from those formed by thrombin. Platelets promote the structuration of both reptilase - and thrombin-fibrin. ADP or other aggregating substances, although not primarily involved in the interaction of platelets with reptilase-fibrin, considerably increase such interaction; indeed, in their presence, the clot formation time (k) is shortened and the maximal amplitude of TEG strongly increased. The promoting effect of aggregating agents (including thrombin) on the structuration of fibrin is inhibited by several aggregation inhibitors (Na2-EDTA, pyrimido-pyrimidine compounds, PGE1). In contrast, both aspirin and indomethacin are ineffective. It is suggested that the role of platelets in clot structuration is, at least, twofold: 1) adherence of polymerising fibrin to platelets orients the fibrin fibres; 2) activation of the platelet contractile system provides further orientation of the fibrin fibres and increases their tensile strength. In all the systems studied, the maximal amplitude of TEG was linearly related to the degree of clot retraction; this confirms that the maximal amplitude of TEG is a measure of clot retraction. Provided conditions for the basic fibrin structure (qualitatively and quantitatively) are equal.

Indomethacin and Platelet Aggregation in Chronic Glomerulonephritis: Existence of Non-responders

Indomethacin given to adults with glomerulonephritis usually reduces their urinary excretion of protein and fibrinogen/fibrin-related (F.R.) antigen. Nevertheless, non-responders exist. Platelet aggregation is significantly more strongly inhibited by indomethacin in good responders than in nonresponders. This supports the hypothesis that the reduction of urinary excretion of F.R. antigen during indomethacin administration is related, at least in part, to inhibition of intrarenal platelet aggregation and of the subsequent fibrin deposition.

Dissociation of Clot Retraction from Platelet Granule Fusion and Degranulation: an Ultrastructural Study of Reptilase—Human Platelet-rich Plasma Clots

Summary. Human platelet‐rich plasma clots, formed with Reptilase, do not retract. Electron microscopy of such a clot reveals platelets of normal shape and ultrastructural features surrounded by fibrin. When ADP is added before Reptilase, the clot strongly retracts. A sequential ultrastructural study of this process shows that, before visible fibrin formation, the platelets form small clusters and show shape change, granule centralization and pseudopod formation. Upon immobilization of the platelets by fibrin, the pseudopods develop into large cytoplasmatic protrusions, which allow cell contact. During the retraction process, the platelet granules remain intact; some granule fusion only becomes apparent in fully retracted clots. When Thrombofax is added to platelet‐rich plasma before Reptilase, the clots formed also retract. With this aggregation inducer, granule fusion occurs earlier and in more platelets. Platelet pseudopod enlargement and fibrin concentration around the platelet mass arc similar to that in ADP‐trcated samples. Inhibitors of granule fusion and secretion (suprofen, indomethacin) do not modify ADP‐Rcptilase nor Thrombofax‐Reptilase clot retraction but reduce the incidence of granule fusion in Thrombofax‐Reptilase clots. Retraction of Thrombofax‐Reptilase clots is unaffected by concentrations of apyrase which completely block ADP‐Reptilase clot retraction. Prostaglandin E1; papaverine and amitryptiline inhibit both ADP‐and Thrombofax‐Reptilase clot retraction, platelet pseudopod formation and cell‐to‐cell adhesion. These findings suggest that interaction of fibrin with the mass of fused granules (granulomere) or platelet secretion are not responsible for clot retraction. Rather, contraction around adhesion sites formed by cytoplasmic protrusions from adjacent platelets would seem to be involved.

Platelet Factor 3 — Properties and Clinical Significance

Bounameaux (2) suggested that the periphery of the platelet has a “plasmatic atmosphere” containing many plasma substances, all identified on the basis of their activity. Several so-called “platelet factors” have since then been described (Table I) (20); for most of them, however, it is not possible to state at present if they are specific constituents of platelets or if they come from the plasma; in contrast, some other factors such as phospholipid-like activity (platelet factor-3, PF-3) and antiheparin activity (platelet factor-4) seem to be specific platelet constituents.

Factor VIII--related protein on vascular intima of patients with chronic renal failure and prolonged bleeding times.

To determine whether the prolonged bleeding time so common to chronic renal failure (CRF) was due to defective factor VIII-related activities, as in von Willebrands disease, vascular-factor VIII-related protein was measured in patients with CRF. Factor VIII-related protein was detected by immunofluorescence on the vascular intima of all 13 patients with CRF and greatly prolonged bleeding times. This protein was also present on the vascular intima of a patient with CRF and moderate von Willebrands disease. These findings support a previous suggestion that the disturbed haemostasis in patients with CRF is not linked to defective factor VIII-related activities.

Etude de l'agrégation plaquettaire par le Thrombofax

Thrombofax Ortho is a stable, reproducible, ready-to-use reagent for the evaluation of the platelet release reaction.Thrombofax Ortho is a stable, reproducible, ready-to-use reagent for the evaluation of the platelet release reaction.

Platelet Factor-4. Methods of Study

“Platelet Factor-4” (PF-4) indicates one or more factors exhibiting several biological functions (for review, see ref.3), the best known of which is the ability of neutralizing the anti- clotting activity of heparin; PF-4 is also able to neutralise the anticlotting activity of some fibrinogen degradation products and to induce non-enzymatic polymerization of fibrinogen and its derivatives, a phenomenon known as “paracoagulation”.