Investigation Into Mechanisms of Prothrombinase Complex Inhibition by a Covalent Antithrombin-Heparin Complex.
Notice bibliographique
Résumé
Abstract Abstract 2136 Poster Board II-113 Introduction: Formation of the prothrombinase complex results in efficient generation of thrombin (IIa) by a vast enhancement of the factor Xa (Xa) reaction with prothrombin (II). However, the integration of Xa into the complex significantly decreases its availability for inhibition by antithrombin+heparin (AT+H). Our laboratory has developed a novel anticoagulant by covalently linking antithrombin to heparin (ATH). We have shown that ATH, compared to AT+H, significantly increased the rate of inhibition of clot-bound thrombin as well as several other coagulation factors in isolation. The present study was performed to extend understanding of the anticoagulant mechanisms of ATH by determining its inhibition of Xa within the critical prothrombinase system. Methods: Pseudo first-order kinetic experiments for the inhibition of Xa in the prothrombinase complex were performed by reacting AT+H or ATH with CaCl2, phospholipid vesicles (PC/PS), factor Va (Va), II, specific IIa inhibitor Pefabloc and Xa in buffer. At specific time intervals, the reactions were neutralized with Na2EDTA, polybrene and Xa substrate (S-2222) in buffer, followed by measurement of residual enzyme activity. Second-order rate constants (k2) were calculated from semi-logarithmic plots of residual Xa activity remaining versus time. To investigate the roles of individual components of the prothrombinase complex on the anticoagulant effects of ATH, additional experiments were performed where components of the complex (II, PC/PS or Va) were removed prior to reaction with AT+H or ATH. Results: The k2value (× 108 M−1min−1) for the inhibition of Xa alone by AT+H was 1.47 ± 0.08. Incorporation of Xa into the prothrombinase complex significantly decreased this value to 0.67 ± 0.05, p<0.001. In comparison, ATH inhibited free-Xa and prothrombinase-complexed Xa at significantly faster rates than AT+H (2.83 ± 0.15 and 2.11 ± 0.38 (p<0.01), respectively). Relative to intact prothrombinase, rates for inhibition by AT+H of Xa in complexes devoid of PC/PS, Va or II significantly increased to 1.49 ± 0.08, 1.44 ± 0.08 and 1.24 ± 0.04, p<0.001, respectively. In contrast, removal of II from prothrombinase significantly decreased the k2 for Xa inhibition by ATH to 1.57 ± 0.04, p<0.05. Although removal of PC/PS from the system resulted in a higher k2 (2.79 ± 0.14, p<0.01), removal of Va resulted in no significant change (2.09 ± 0.09, p=0.79) for ATH reactions. Conclusion: Based on these results, we conclude that the prothrombinase complex hinders the inhibitory action of AT+H on Xa, whereas ATH is less affected. In fact, ATH inhibition of the prothrombinase complex was significantly higher than the inhibition rate for either free or prothrominase-bound Xa by AT+H. We speculate that, similar to inhibition of fibrin-bound thrombin, H in the non-covalent AT+H dissociate and bind to components within the prothrombinase that repel incoming AT+H and preventing Xa inhibition. These complexes cannot occur when AT is covalently linked to H, thus allowing for enhanced inhibition of prothrominase-bound Xa by ATH. Absence of II from the prothrombinase complex removes interference of AT+H reactions by any H complexes within prothrombinase, but may antagonize inhibition by the conjugate through unproductive interactions of ATH with uncovered sites close to Xa at the phospholipid surface. These findings reveal important features involved in the enhanced anticoagulant action of ATH against prothrombinase surface-bound enzymes. Disclosures: No relevant conflicts of interest to declare.
Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.
Comment cette classification a été obtenuedéplier
Prédiction machine sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Le volet Gemma est une étiquette directe du modèle pour chaque travail de la base, lue sur la notice réduite au titre. Le volet Codex est un classifieur appris des 10 348 étiquettes directes de Codex et calibré sur les taux pondérés de l'échantillon; les champs sans appui suffisant ne portent aucun appel Codex. Le mode candidate est l'union des deux volets; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont pas des étiquettes humaines.
Scores du classifieur distillé par catégorie (deux têtes)
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,000 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,000 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,002 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule source (Gemma direct ou Codex distillé), pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».