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Journal articles on the topic 'Pro-hemostatic agents'
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Laurenti, Juliana Bergamasco, Gabriel Zazeri, Ana Paula Ribeiro Povinelli, Moacir Fernandes de Godoy, Domingo Marcolino Braile, Tânia R. Flores da Rocha, Élbio Antônio D' Amico, and José Geraldo Nery. "Enhanced pro-coagulant hemostatic agents based on nanometric zeolites." Microporous and Mesoporous Materials 239 (February 2017): 263–71. http://dx.doi.org/10.1016/j.micromeso.2016.10.020.
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Budko, Elena V., Daria A. Chernikova, Leonid M. Yampolsky, and Valentina Y. Yatsyuk. "Local hemostatic agents and ways of their improvement." I.P. Pavlov Russian Medical Biological Herald 27, no. 2 (July 2, 2019): 274–85. http://dx.doi.org/10.23888/pavlovj2019272274-285.
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Recently, local hemostatic agents (LHA) have become increasingly popular abroad and in our country. They act in a targeted way and can be used both in damage to large vessels and in diffuse bleeding. In the article, chemical nature, physical and chemical characteristics of materials and mechanisms of LHA activity are considered, directions of their improvement are shown. LHA are mostly classified by mechanism of action.
To date, the popular groups of hemostatic agents are «mucoadhesive agents» (chitosan, amylopectin) and «coagulation factors concentrators» (zeolites, kaolin). Other authors distinguish the group of «aggregation and adhesion stimulants» (collagen, cellulose). Here, representatives of these groups have common characteristics – very high porosity and hydration ability. Another group includes substances that «promote protein denaturation» (inorganic salts of metals, as well as salts of acrylic acid and its derivatives). Polyacrylates are the basis of adhesives with hemostatic activity. However, most modern LHA are complex drugs and it is just this group that is most promising. All means, from hemostatic sponges produced by Zelyonaya Dubrava (Russia) and Nycomed, Takeda (Austria, Norway), and to hemostatic materials of MedTrade manufacture (Great Britain), Etiguette and Z-Medica (USA), combine sorption and, actually, thrombotic properties. The trademarks often imply original compositions and, especially, technologies: Quick Relief, BioSeal, BallistiClot, Hemaderm, CELOX Gauze PRO, OMNI-STAT Hemostatic Gauze for minor external bleeding. The most effective LHA are those based on chitosan and kaolin in the form of dressings with embedded clot-forming substance, for example, with artificial platelets or other coagulation factors.
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Otrocka-Domagała, I., P. Jastrzębski, Z. Adamiak, K. Paździor-Czapula, M. Gesek, M. Mikiewicz, and T. Rotkiewicz. "Safety of the long-term application of QuikClot Combat Gauze, ChitoGauze PRO and Celox Gauze in a femoral artery injury model in swine – a preliminary study." Polish Journal of Veterinary Sciences 19, no. 2 (June 1, 2016): 337–43. http://dx.doi.org/10.1515/pjvs-2016-0041.
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AbstractThe purpose of this study was to examine the safety of the long-term application of QuikClot Combat Gauze, ChitoGauze PRO and Celox Gauze using a swine model. The study was conducted on nine pigs weighing approximately 30 kg, which were randomly divided into three groups. Under deep anesthesia, the pigs underwent complete transverse cutting of the femoral artery in the groin region. Hemostatic dressings were left in the wound for 24 hours. The animals were euthanized 24 hours after dressing application. In each group, macroscopic and microscopic severe changes and shock symptoms were observed in the lungs, liver, kidneys and heart. Fibrino-gaseous embolic material was found in the pulmonary artery of each group and in the lung vessels of the animals from the ChitoGauze PRO and Celox Gauze groups. In conclusion, the long-term application of the evaluated hemostatic dressings has the risk of coagulopathy and reaching the progressive stage of shock. The residues from the hemostatic dressings can ingress into the systemic circulation, thereby increasing the risk of embolus formation. Because of these harmful effects, the evaluated hemostatic dressings are not appropriate for long-term use. Future studies are needed on the consequences of the long-term application of these hemostatic agents.
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Schoenwaelder, Simone M., Kate E. Jarman, Elizabeth E. Gardiner, My Hua, Jianlin Qiao, Michael J. White, Emma C. Josefsson, et al. "Bcl-xL–inhibitory BH3 mimetics can induce a transient thrombocytopathy that undermines the hemostatic function of platelets." Blood 118, no. 6 (August 11, 2011): 1663–74. http://dx.doi.org/10.1182/blood-2011-04-347849.
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Abstract BH3 mimetics are a new class of proapo-ptotic anticancer agents that have shown considerable promise in preclinical animal models and early-stage human trials. These agents act by inhibiting the pro-survival function of one or more Bcl-2–related proteins. Agents that inhibit Bcl-xL induce rapid platelet death that leads to thrombocytopenia; however, their impact on the function of residual circulating platelets remains unclear. In this study, we demonstrate that the BH3 mimetics, ABT-737 or ABT-263, induce a time- and dose-dependent decrease in platelet adhesive function that correlates with ectodomain shedding of the major platelet adhesion receptors, glycoprotein Ibα and glycoprotein VI, and functional down-regulation of integrin αIIbβ3. Analysis of platelets from mice treated with higher doses of BH3 mimetics revealed the presence of a subpopulation of circulating platelets undergoing cell death that have impaired activation responses to soluble agonists. Functional analysis of platelets by intravital microscopy revealed a time-dependent defect in platelet aggregation at sites of vascular injury that correlated with an increase in tail bleeding time. Overall, these studies demonstrate that Bcl-xL–inhibitory BH3 mimetics not only induce thrombocytopenia but also a transient thrombocytopathy that can undermine the hemostatic function of platelets.
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Agraharkar, M., and M. A. Martinez. "Spontaneous Re-Canalization of Thrombosed Polytetrafluoroethylene (PTFE) Grafts." Journal of Vascular Access 2, no. 3 (July 2001): 114–18. http://dx.doi.org/10.1177/112972980100200306.
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Fibrinolysis is an important hemostatic process initiated either by tissue plasminogen activator (tPA) or pro-urokinase (pro-UK) released from endothelial cells. These agents act preferentially on plasminogen by converting it to the active molecule plasmin. This initiates the clot lysis process, which may take several days for completion. Most patients with end-stage renal disease (ESRD) have PTFE grafts for chronic hemodialysis. When these grafts are thrombosed, they are either surgically revised or percutaneously thrombolysed. When these measures fail another access is created without removing the clotted graft. However, it is possible that de-clotting of these thrombosed PTFE grafts can occur spontaneously. Once the graft develops an endothelial lining, these cells can contribute to the fibrinolytic process by secreting tPA or pro-UK. Because this endothelial lining may be less developed than that of a normal vessel, the fibrinolytic process may occur at a slower rate or not at all. Frequent cannulation during hemodialysis can denude existing endothelium, further contributing to the inadequacy of the graft to initiate thrombolysis. In practice, once the PTFE is clotted, the graft is ignored if it is not considered for declotting. In such circumstances the re-canalization process could be overlooked, resulting in the unnecessary placement of additional accesses. Presented here are three patients with clotted grafts in which re-canalization occurred without intervention.
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Sivakumar, Walavan, Jian Guan, Jean-Philippe Langevin, Neil A. Martin, Garni Barkhoudarian, Daniel F. Kelly, John Franklin Berry, and Aditya K. Iyer. "873 Code Brain Ultra-Early Clinical Pathway for Patient Identification and Management in Spontaneous Non-traumatic Intracranial Hemorrhage Single-Institution Study." Neurosurgery 70, Supplement_1 (April 2024): 173. http://dx.doi.org/10.1227/neu.0000000000002809_873.
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INTRODUCTION: Spontaneous non-traumatic intracranial hemorrhage causes high morbidity and mortality. Clinical pathways for the management of acute ischemic stroke are well established but fall short in identifying intracranial hemorrhage, preventing early therapy. METHODS: Using Class 1 recommendations from the AHA/ASA Spontaneous Intracranial Hemorrhage Guidelines, our multi-disciplinary stroke team developed a hyper-acute clinical pathway. Once the hemorrhagic stroke patient was identified, protocolized measures were immediately initiated to lower systolic blood pressure to 140 mmHg, institute factor replacement therapies in coagulopathic patients and anti-platelet reversal when appropriate. Key performance indicators and quality metrics were established to compare post-intervention groups to the baseline cohort. RESULTS: Of 180 patients, 113 and 67 patients were treated pre- and post-protocol, respectively. From pre- to post-protocol, diagnosis of hemorrhagic stroke using the alert increased from 59% to 81%. Anti-hypertensive initiation within 60 minutes increased from 79.5% to 88.7%, with the average time from CT result decreasing from 32.7 to 21.7 minutes. Time to reversal agent decreased from 53 to 23 minutes for pro-coagulation agents and 106 to 55 minutes for DDAVP. Reversal agent initiation within 60 minutes of CT result increased from 66.6% to 100% for pro-coagulation agents and 28.5% to 84.6% for DDAVP. CONCLUSIONS: The initiation of this management protocol for hemorrhagic stroke resulted in a higher proportion of patients being rapidly identified and earlier initiation of anti-hypertensive and hemostatic therapies. These processes are the key to important interventions designed to reduce injury extension associated with intracranial hemorrhage, like “time to thrombolysis” measures for acute ischemic stroke.
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Thalji, Nabil K., Lacramioara Ivanciu, Reema Jasuja, Sunita Patel-Hett, Joachim Fruebis, Debra Pittman, and Rodney M. Camire. "Zymogen-like FXa Is a Potent Bypassing Agent for Reversal of Direct FXa Inhibitors in Vivo." Blood 124, no. 21 (December 6, 2014): 582. http://dx.doi.org/10.1182/blood.v124.21.582.582.
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Abstract The pharmacokinetic challenges of warfarin therapy have led to the development of new oral anticoagulants (NOACs) that directly inhibit coagulation factor Xa (FXa). While these new drugs (including rivaroxaban and apixaban) have many benefits over warfarin, no approved strategy exists to reverse their anticoagulant effects in the event of life-threatening bleeding or emergent need for surgery. The most promising reversal strategy in clinical development is a catalytically inactive form of FXa (Gla-domainless FXaS195A, GD-FXaS195A) that binds the NOAC with high affinity to relieve inhibition of endogenous FXa. In principle, removing the inhibitor through molecular engagement is attractive, especially when administered before bleeding begins (e.g. pre-surgery), and there is in vivo evidence to support the efficacy of GD-FXaS195A as a pre-injury antidote for direct FXa inhibitors. However, since GD-FXaS195A is a scavenger of the inhibitor and not a pro-hemostatic agent, GD-FXaS195A may not be effective if given after a bleeding episode has begun. Moreover, its mechanism of action necessitates a 1:1 ratio of GD-FXaS195A to neutralize the inhibitor. Thus, high doses (hundreds of milligrams of protein) will likely be required in humans. We hypothesized that a pro-hemostatic bypassing agent might be able to reverse the effects of direct FXa inhibitors no matter when it is administered, and with high potency. To evaluate this, we used a variant of FXa (FXaI16L) that is more zymogen-like than wild-type (wt)-FXa. This "zymogen-like" variant has lower catalytic activity in in vitro assays compared to wt-FXa due to impaired active site maturation. It is also resistant to active site inhibitors including plasma protease inhibitors, resulting in an extension of its plasma half-life (>30 minutes vs ~1 minute for wt-FXa). Importantly, its activity is rescued upon binding to its cofactor FVa in vivo, and we have previously shown that FXaI16L bypasses the intrinsic pathway defect in hemophilic mice. Here we evaluated whether FXaI16L might also be able to bypass direct FXa inhibitors using in vitro thrombin generation assays (TGAs) and two in vivo injury models in mice, and directly compared FXaI16L to GD-FXaS195A. In TGA experiments, 500 nM rivaroxaban decreased peak thrombin generation to 23% of that observed in normal human plasma (NHP). This decreased thrombin generation could be reversed by the addition of 3 nM human (h)FXaI16L which normalized peak thrombin generation. In comparative studies, hFXaI16L was 300-fold more potent than GD-FXaS195A in TGA assays. These data highlight that differences in mechanism of action (pro-hemostatic vs. scavenger) will have a huge impact on amounts of protein needed to revive thrombin generation. This was further reflected in in vivo studies. In invivo experiments with wild-type mice using 7.5% FeCl3 to induce carotid artery thrombosis, 1 mg/kg rivaroxaban prevented formation of occlusive thrombi. 30 minutes after the initial FeCl3 injury to rivaroxaban-treated mice, infusion of 0.25 mg/kg murine (m)FXaI16L normalized the phenotype, causing rapid occlusion at the injury site within 5 minutes. In contrast, we did not observe carotid artery occlusion with administration of up to 25 mg/kg GD-FXaS195A 30 minutes after the injury to rivaroxaban-treated mice (Fig. 1). When both rivaroxaban and the reversal agent were given prior to the injury, both mFXaI16L (0.5 mg/kg) and GD-FXaS195A (25 mg/kg) were effective at reversing the inhibition. Furthermore, using intravital microscopy to examine thrombus formation at the site of laser injury to mouse cremasteric arterioles, we found that 1 mg/kg rivaroxaban completely abrogated fibrin deposition and substantially reduced platelet accumulation. Treatment with1 mg/kg mFXaI16L substantially increased platelet and fibrin deposition at the site of laser injury. Figure 1 Carotid occlusion time when the reversal agent was infused 30 minutes after the FeCl3 injury. Figure 1. Carotid occlusion time when the reversal agent was infused 30 minutes after the FeCl3 injury. Taken together, these data provide strong support for the in vivo efficacy and potency of FXaI16L as a potential pro-hemostatic bypass agent to reverse direct FXa inhibitors. The data also illustrate that antidotes like GD-FXaS195A may not be as effective as pro-hemostatic agents like FXaI16L following an injury, and we predict that effective reversal of NOACs will require a nuanced approach that uses an antidote or a bypassing agent depending on the clinical scenario. Disclosures Jasuja: Pfizer: Employment. Patel-Hett:Pfizer: Employment. Fruebis:Pfizer: Employment. Pittman:Pfizer: Employment. Camire:Pfizer: Consultancy, Patents & Royalties, Research Funding.
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Elbaz, Carolyne, Katerina Pavenski, Hina Chaudhry, Jerome M. Teitel, and Michelle Sholzberg. "The Frequency and Effect of Baseline Cross-Reacting and De Novo Inhibitors to Recombinant Porcine FVIII in Patients with Congenital and Acquired Hemophilia a." Blood 134, Supplement_1 (November 13, 2019): 1128. http://dx.doi.org/10.1182/blood-2019-122260.
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Background Patients with severe congenital hemophilia A (CHA) have a 25-40% lifetime risk of alloantibody (inhibitor) development to FVIII. Patients with acquired hemophilia A (AHA) spontaneously develop neutralizing autoantibodies to factor VIII. In both cases, patients require pro-hemostatic therapy with bypassing agents: recombinant factor VIIa (rFVIIa), activated prothrombin complex concentrate (aPCC) and more recently recombinant porcine factor VIII (rpFVIII). Anti-human FVIII (hFVIII) inhibitors typically bind to the A2 and C2 domains of the FVIII molecule. RpFVIII is an effective pro-hemostatic treatment for AHA and CHA given the immunologic difference in the A2 and C2 domains of the rpFVIII while maintaining sufficient hFVIII homology to act as an effective cofactor to human FIX in the intrinsic tenase. However, some anti-hFVIII antibodies cross-react with rpFVIII and may interfere with its hemostatic function. Cross-reacting antibodies were reported in 35% of subjects in a phase II/III trial prior to initiation of rpFVIII. Moreover, de novo rpFVIII inhibitors may develop during or after the treatment with rpFVIII and may affect its hemostatic function. Here we describe the largest case series to date on baseline cross-reactivity of rpFVIII inhibitors and post-treatment de novo inhibitor development in patients with CHA and AHA to address the paucity of published literature in this area. Aim First, we describe the frequency of baseline cross-reacting rpFVIII inhibitors in patients with AHA and CHA (with inhibitors) at our institution. Second, we describe the effect of baseline rpFVIII antibodies on FVIII recovery after treatment with rpFVIII. We also describe the frequency and timing of de novo rpFVIII inhibitor development after exposure to rpFVIII. Methods Institutional research ethics board approval was obtained. Electronic charts of patients admitted to our institution with AHA or CHA who underwent testing for rpFVIII inhibitors were reviewed retrospectively. RpFVIII inhibitor assay is performed in the special coagulation laboratory using the Nijmegen modified Bethesda assay. The patient sample is initially heat-treated at 57 Results Twenty-seven patients (7 CHA, 20 AHA) underwent testing for porcine inhibitors since assay availability in 2016. 61% (5/7 CHA, 11/20 AHA) of patients had a detectable rpFVIII inhibitor prior to exposure to rpFVIII; median titer 1.6 BU/ml (range 0.6-192). Eight patients with AHA with baseline cross-reacting inhibitors received rpFVIII. Of those, three achieved an initial FVIII recovery beyond 100% (132%, 148% and 177%) after approximately 100U/kg of rpFVIII and all three had very low anti-rpFVIII Bethesda titers (0.70, 0.85 and 0.9 BU/ml). Five patients did not achieve a FVIII recovery above 50% (46%, 46%, 40%, 36% and 0%) despite approximately 100U/kg of rpFVIII. Most patients who received rpFVIII were tested weekly for the duration of their treatment or hospital stay. Upon discharge, patients who were seen in clinic for follow up were tested for anti-hFVIII and anti-rpFVIII. Two AHA patients without a baseline inhibitor who received rpFVIII treatment developed a de novo inhibitor after 20 days (1 BU/ml) and 133 days (12 BU/ml), respectively. One AHA patient had a rise in baseline anti-rpFVIII titer after exposure to rpFVIII. Conclusion In conclusion, we found that 61% of patients with AHA and CHA tested for rpFVIII inhibitors had a detectable baseline cross-reacting inhibitor which is higher than previously described. Of those patients with a baseline inhibitor treated with rpFVIII, only 37.5% of patients had an appropriate rise in FVIII. Finally, 13% of patients without baseline inhibitors developed a de novo inhibitor after exposure to rpFVIII, an incidence comparable to previously published findings. Disclosures Pavenski: Bioverativ: Research Funding; Alexion: Honoraria, Research Funding; Octapharma: Research Funding; Shire: Honoraria; Ablynx: Honoraria, Research Funding. Teitel:BioMarin: Consultancy; CSL Behring: Consultancy; Octapharma: Consultancy; Novo Nordisk: Consultancy; Shire: Consultancy; Pfizer: Consultancy, Research Funding; Bayer: Consultancy, Research Funding. Sholzberg:Takeda: Honoraria, Research Funding; Baxter: Honoraria, Research Funding; Baxalta: Honoraria, Research Funding. OffLabel Disclosure: Recombinant porcine factor VIII is used to treated patients with congenital hemophilia A with allo inhibitors
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Hartmann, Rudolf, Tjerk Feenstra, Sabine Knappe, Michael Dockal, and Friedrich Scheiflinger. "Elucidating the Excessive Pro-Coagulant Effect of a Sequence Identical Analogue to ACE910 in Combination with Bypassing Agents." Blood 130, Suppl_1 (December 7, 2017): 90. http://dx.doi.org/10.1182/blood.v130.suppl_1.90.90.
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Abstract Introduction: Emicizumab (ACE910), an antibody to FIX(a) and FX(a), is currently under investigation for treatment of hemophilia with inhibitors. In a phase III trial, two thromboembolic complications and three cases of microangiopathy were reported in patients on ACE910 prophylaxis [Oldenburg et al. NEJM 2017], whose breakthrough bleeding was treated with activated prothrombin complex concentrate aPCC (FEIBA) or aPCC and rFVIIa. We generated a sequence identical analogue (SIA) to ACE910 and analyzed its synergistic interplay with bypassing agents. Aims: To monitor in vitro the pro-coagulant activity of SIA ACE910 in the presence of FEIBA and rFVIIa, and detect the source of excessive coagulation induced by SIA ACE910 combined with FEIBA. Methods: A sequence identical analogue (SIA) to ACE910 was expressed in HEK293 cells, purified as previously described [Sampei et al. PLoS One 2013], and analyzed in several global hemostatic assays at different concentrations and test conditions using plasma and whole blood assays. In thrombin generation (TG) experiments, platelet-poor plasma (PPP) from hemophilia A inhibitor patients and hemophilia A plasma reconstituted with platelets from 3 healthy donors (PRP) was used. A normal TG range was established in healthy donor plasma. Therapeutic concentrations of SIA ACE910 (20-600 nM) were tested alone and with FEIBA (0.05-1 U/mL) or rFVIIa (0.88-5.25 µg/mL). To measure FEIBA components' contribution to the synergistic effect with SIA ACE910, PPP was spiked with select FEIBA components at concentrations corresponding to 0.5 U/mL FEIBA in combination with the antibody. Thrombus formation was analyzed in FVIII-inhibited blood using rotational thromboelastometry (ROTEM) and Total Thrombus-formation Analysis System (T-TAS). Results: Normal peak thrombin was 47-144 nM for PPP and 88-231 nM for PRP. rFVIIa and FEIBA had an additive effect on TG in combination with SIA ACE910 in both plasma types. Combined with rFVIIa (0.88 µg/mL) or FEIBA (0.5 U/mL), SIA ACE910 (600 nM) induced a ~2- and ~16-fold increase over SIA ACE910 alone. SIA ACE910+rFVIIa did not reach the normal range, while SIA ACE910+FEIBA far exceeded it. Adding individual FEIBA components to PPP showed that FIX was, with a half-maximal effect, the main driver for enhanced TG, followed by FIXa. formation in FVIII-inhibited whole blood using ROTEM and T-TAS confirmed the excessive effect of SIA ACE910+FEIBA. In ROTEM, FEIBA and rFVIIa reduced clotting time to shorter than normal, whereas SIA ACE910 had only little effect. Moreover, adding SIA ACE910 to rFVIIa exerted no effect over rFVIIa alone. Conclusion: Combining SIA ACE910 at plasma concentrations observed in patients [Oldenburg et al. NEJM 2017] with FEIBA induced excessive thrombin generation and faster clot formation. In vitro, this effect is mainly mediated by FEIBA component FIX. ACE910 binds to FIX and FIXa to the same extent, and displays its pro-coagulant effect via an unregulated mechanism. Therefore, careful judgement is needed in treating breakthrough bleeds with FEIBA. Disclosures Hartmann: Shire: Employment. Feenstra: Shire: Employment. Knappe: Shire: Employment. Dockal: Baxalta: Patents & Royalties; Shire: Employment, Equity Ownership; Baxter: Equity Ownership, Patents & Royalties. Scheiflinger: Baxter: Equity Ownership; Shire: Employment, Equity Ownership.
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Jubelirer, Steven J. "Venous Thromboembolism and Malignant Brain Tumors: A Review." Clinical and Applied Thrombosis/Hemostasis 2, no. 2 (April 1996): 130–36. http://dx.doi.org/10.1177/107602969600200208.
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The incidence of venous thromboembolism (VTE) in patients with primary brain tumors varies be tween 1 and 60%. This variability in incidence is due to study differences in (a) methods of diagnosis of VTE— i.e., diagnosis at autopsy or clinical diagnosis; (b) amount of time from surgery to VTE diagnosis; (c) proportion of patients receiving deep venous thrombosis (DVT) pro phylaxis ; (d) clinical risk factors associated with VTE, such as paresis, prior thrombotic disease, and chemother apy; and (e) tumor location and histology. The etiology of VTE in patients with primary brain tumors is unknown. The preoperative hemostatic abnormalities noted in clin ical studies have been most consistent with compensated disseminated intravascular coagulation (DIC). These ab normalities, however, appear to be of little predictive value for the subsequent development of VTE. Studies involving brain tumor tissue or cell cultures have impli cated factors released by the tumor or surrounding neural tissue that activate the coagulation system or inhibit fi brinolysis. Recommendations for VTE prophylaxis in clude (a) earliest possible ambulation; (b) intermittent pneumatic compression in all nonambulatory patients preoperatively and postoperatively; and (c) s.c. heparin in high-risk patients. The role of low-molecular-weight heparin in VTE prophylaxis has not been established. Patients with malignant brain tumors can be safely anti coagulated with heparin and warfarin if these agents are carefully monitored. Of 197 patients in seven series who received anticoagulants, only 5 (2.5%) had intracranial bleeding. Vena caval filters and thrombectomy are rarely required. Thrombolytic therapy is contraindicated. Key Words: Venous thromboembolism—Deep venous throm bosis—Malignant brain tumors.
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Tucker, Erik I., Ulla M. Marzec, Sawan Hurst, András Gruber, and Stephen R. Hanson. "Moderate Pharmacological Platelet Count Reduction Limits Thrombus Formation without Hemostatic Impairment in Primates." Blood 112, no. 11 (November 16, 2008): 1033. http://dx.doi.org/10.1182/blood.v112.11.1033.1033.
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Abstract Despite the established contribution of platelets to thrombotic cardiovascular disorders, documented in part by the effectiveness of platelet function inhibitors and the increased risk of thrombosis associated with high normal and supranormal platelet counts, relationships between circulating platelet count and thrombotic events remain largely undefined. Since the initiation and propagation of arterial, platelet-dependent thrombus must depend upon platelet count, albeit in a manner that could be nonlinear, we hypothesized that reducing platelet count within the normal range would produce an anti-thrombotic benefit with minimal effects on hemostasis. To test this hypothesis, we reduced the platelet count in baboons (n=4) by targeting the megakaryocyte growth and development factor thrombopoietin (TPO). A polyclonal anti-TPO autoantibody (anti-TPOab) was purified from the serum of a baboon that developed thrombocytopenia following recombinant TPO injections. The IC50 of the purified IgG fraction was found to be 0.76 μg/ml, determined using a proliferation assay with a TPO-dependent cell line. An i.v. bolus of the anti-TPO antiserum, 30–35 ml infused into baboons, resulted in a transient, >60% decrease in the circulating platelet count after 2–3 weeks. Other blood cell counts were unaffected vs. baseline values. The effect of platelet count reduction on thrombogenesis was evaluated using an established baboon arteriovenous (AV) shunt thrombosis model. Accumulation of 111-Indium-labeled platelets and 125-Iodine-labeled fibrinogen were measured within a 4 mm i.d. thrombogenic vascular graft segment that was deployed into a chronic AV shunt for 60 min. Blood flow was maintained at 100 ml/min, producing an arterial wall shear rate of 265 sec−1. Standard template bleeding times (BTs) were used to assess hemostatic impairment at various platelet counts. Platelet count reductions, ranging from 46–61% (normal levels averaging 352,000 ± 61,000 platelets/μl), reduced platelet deposition onto the graft surface by 46–68% (vs. control values of 4.1 ± 0.9 x 109 platelets deposited, n=9). Similarly, thrombus fibrin accumulation was reduced by 14–39% (vs. control values of 2.2 ± 0.4 mgs of deposited fibrin). Thrombus formation was not affected acutely by anti-TPOab administration, but correlated directly with circulating platelet numbers. As expected, BTs were not significantly prolonged until platelet counts fell below ~100,000 cells/μl. In contrast, single dose aspirin (32 mg/kg) at normal platelet counts did not significantly reduce graft associated platelet deposition in this model but doubled the BTs to 6.8 ± 2.6 min (vs. control values of 3.4 ± 0.9 min). With further reduction in platelet counts to 90,000 ± 30,000 platelets/μL, BTs were only slightly prolonged (5.6 ± 1.7 min, n=5). When platelet counts averaged 74,000 ± 20,000 platelets/μl in animals given ASA, BTs averaged only 9.4 ± 2.7 min (n=5). Thus ASA produced a hemostatic impairment that was approximately fixed (i.e., a BT prolongation of 3–4 min) and not disproportionately prolonged at reduced platelet counts. Thus specific lowering of the platelet count by pharmacologic inhibition of megakaryocytopoiesis may be an effective anti-thrombotic strategy in populations currently treated with conventional anti-platelet agents. Since direct inhibitors of platelet function produce a significant risk of bleeding, inhibition of platelet production may represent a safer approach for reducing the pro-thrombotic capacity of the circulating platelet pool.
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McDonald, Anna G., Maureane Hoffman, Ulla Hedner, Harold R. Roberts, and Dougald M. Monroe. "Restoring Initial Thrombin Generation Does Not Normalize Cutaneous Wound Healing in Hemophilia B." Blood 108, no. 11 (November 16, 2006): 1030. http://dx.doi.org/10.1182/blood.v108.11.1030.1030.
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Abstract We recently described abnormal wound healing in a mouse model of hemophilia B (Hoffman et al, Blood 2006; DOI 10.1182/blood-2006-05-020495). Specifically: epithelial closure was delayed in hemophilia B mice; hemophilic animals, unlike wild type, developed subcutaneous hematomas; macrophage influx was delayed compared to wild-type mice; and, surprisingly, angiogenesis was enhanced in the hemophilia B mice. We hypothesized that restoring the initial hemostatic burst of thrombin generation following wounding by administration of a single dose of factor IX (FIX) replacement or factor VIIa (FVIIa) bypassing therapy would not only prevent bleeding, but correct the subsequent wound healing process. One dose of therapy was given thirty minutes prior to placement of a single three mm punch biopsy wound on the dorsal skin of each wild type, untreated hemophilia B, and treated hemophilia B mouse. The size of the wounds was measured daily until full epithelial closure. The time course of epithelial closure in treated hemophilia B was intermediate between wild type and untreated hemophilia B. FVIIa-treated hemophilia B began to heal earlier than FIX-treated. Skin from the wound site was collected at different days and examined histologically. Macrophage influx was earlier in treated hemophilia B mice compared to untreated hemophilia B, likely a due to the increased thrombin and fibrin acting as chemotactic agents. The macrophage influx in FVIIa-treated HB was significantly greater at certain time points than in FIX-treated mice, possibly reflecting some signaling effect of TF/FVIIa in addition to its effects on thrombin generation. With the earlier influx in macrophages, hemoglobin was degraded to storage iron at earlier time points. However, tissue iron continued to persist in treated hemophilia B mice similar to untreated hemophilia B, suggesting continued rebleeding. FIX treatment led to significantly more angiogenesis than FVIIa. The reasons for this difference remain to be determined. Untreated and some treated hemophilia B mice developed subcutaneous hematomas both before and after wound closure. The early hematomas are likely caused by a combination of the initial wounding trauma and vulnerability to bleeding related to the high level of vascularity within the granulation tissue. We propose the late hematomas are due to a cycle of bleeding, leading to more inflammation with production of more pro-angiogenic cytokines, leading to greater angiogenesis with its attendant risk of bleeding. In conclusion, restoring initial hemostatic thrombin generation did not normalize cutaneous wound healing in a hemophilia B mouse model. While the time frame of healing may not be the same in human and murine hemophiliacs, our findings suggest that this model could be helpful in rational determination of treatment schedules for replacement or bypassing therapy following injury or surgery.
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Li, Xena X., Ivan Stevic, Frank M. H. Lee, Keith K. Lau, Anthony K. C. Chan, and Howard H. W. Chan. "Reversal Of Dabigatran By Recombinant Factor VIIa Is Dependent On The Amount Of Tissue-Factor." Blood 122, no. 21 (November 15, 2013): 1149. http://dx.doi.org/10.1182/blood.v122.21.1149.1149.
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Abstract Background Dabigatran is a new oral anticoagulant that specifically and reversibly inhibits thrombin. Its predictable pharmacokinetics and pharmacodynamics allow for minimal monitoring. However, there is currently no specific antidote to reverse its anticoagulant effects. Instead, activated prothrombin complex concentrate (aPCC) or recombinant Factor VIIa (rFVIIa) has been used to stop bleeding complications in patients on dabigatran. Both were originally used to treat hemophilia patients with inhibitors. Currently, Factor Eight Inhibitor Bypass Activity (FEIBA) is the only clinically approved aPCC, which contains Factors II, IX, X, and VII in both active and non-active forms. In contrast, rFVIIa is a human recombinant protein which can initiate clotting via the tissue factor (TF) pathway. It has been controversial which of these hemostatic agents is more efficient at reversing the effect of dabigatran, as animal models have failed to yield consistent results. We hereby utilized a modified Hemoclot turbdity assay to determine the equivalent concentrations of FEIBA and rFVIIa that can reverse the anticoagulant effect of dabigatran. Methods A mixture was prepared by incubating normal pooled plasma (NPP) with 382 nM dabigatran and varying concentration of hemostatic agents (FEIBA or rFVIIa), in the absence or presence of TF (Thrombosel®). The mixture was diluted 1:8 in TSP buffer, from which a 50 µL-aliquot was then incubated with 100 µL of NPP at 37°C for 5 min. Clotting was initiated with 100 µL of another mixture containing 10 mM Ca2+ and 2.5 nM thrombin in TSP. Turbidity was measured at 350 nm using a SpectraMax Pro spectrophotometer at 37°C for 2 hr. Clotting time (CT) was defined as the time to reach half of the maximum turbidity. Results In this modified Hemoclot turbidity assay, the CT without dabigatran was 101 s and it linearly increased dependent on the dabigatran concentration up to 1500 nM. Dabigatran at 382 nM, the therapeutic plasma concentration, prolonged CT 5-fold to 505 s. Addition of 1 U/mL FEIBA reduced the CT approximately 35% to 328 s. The reversal effect of FEIBA plateaued at 2.5 U/mL to 5 U/mL because there was minimal further reduction in the CT even with 10 U/mL FEIBA. In contrast, rFVIIa at a therapeutic concentration of 50 nM barely reduced the CT by <10% to 469 s. The reversal effects of rFVIIa were drastically enhanced by the addition of extrinsic tissue factor. With TF at 1.5 pM, the rFVIIa reduced the CT to a level similarly achieved by 1 U/mL of FEIBA, but the CT could not be reduced further despite the concentration of rFVIIa increased 10-fold to 400 nM. Finally, although both FEIBA and rFVIIa/TF reached an eventual plateau in the reversal of dabigatran, none of them could lower the CT to the baseline level. Summary/Conclusions Our in vitro study shows that reversal of dabigatran by rFVIIa is dependent on the concentration of TF. Higher levels of TF augment the reversal effects of rFVIIa to the extents similarly achieved with FEIBA. The data may, in part, explain the inconsistency of results obtained from in vivo studies using various animal models to compare rFVIIa and FEIBA for the reversal of dabigatran. The circulating TF levels in vivo are much lower than the amount of TF used in this study, thus its availability in the microenvironment for hemostasis varies depending on the methods used to induce bleeding. The data in this study also explain the variable clinical efficacy of rFVIIa reported in the literature when it is used to reverse the bleeding complications in patients on dabigatran. Disclosures: No relevant conflicts of interest to declare.
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Gabriel, Don A., Brett E. Skolnick, Stephanie Seremetis, Philip Leese, and David Mathews. "Effect of Recombinant Activated Factor VII (rFVIIa) on Platelet and Clotting Systems in Healthy Volunteers." Blood 106, no. 11 (November 16, 2005): 4053. http://dx.doi.org/10.1182/blood.v106.11.4053.4053.
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Abstract The generation of thrombin is a critical step in clot formation and securing hemostasis. Inadequate thrombin generation may result in excessive bleeding, while uncontrolled thrombin production has the potential to induce thromboses. Recombinant FVIIa has been shown to control bleeding in hemophilia patients with inhibitors to coagulation factors VIII and IX, and several other clinical conditions in which life- and limb-threatening bleeding may occur. However, for non-hemophilia patients the dose and dosing regimen still require refinement. The use of ex-vivo assessment systems (Thromboelastograph assay, Hemodyne Hemostasis Analyzer) to generate an in vitro model of hemostasis may be useful to determine the effect of rFVIIa on the dynamics of clot formation, particularly in a patient population where clinical trials are difficult. A series of pilot studies were performed to establish an optimal punch biopsy location and biopsy size to create an effective bleeding site for further study. The current study (48 subjects consented for an IRB approved protocol) examined, in an ascending dose-escalation manner, the effect of an individual dose of rFVIIa on bleeding time, blood loss volume, coagulation parameters and coagulation status in healthy volunteers. All patients underwent three punch biopsies: the first (baseline biopsy) with no treatment, the second and third biopsy with either a low or high rFVIIa dose administered prior to biopsy. The treatment pair sequences were placebo/10, 10/20, 20/40, 40/80, 80/120, and 120/160 μg/kg. Blood samples were drawn 15 minutes pre-, 15 minutes post-, 1 hour and 5 hours post-biopsy to conduct ex-vivo assessments of hemostasis. The results indicate that overall there was a trend towards an increase in platelet contractile force and clot elastic modulus with an apparent maximal effect at 1 hour post-biopsy. At the higher doses this effect, although reduced, does not return to pre-treatment values by 5 hours post-biopsy. For the lower doses these values decreased to levels comparable to those at pre-treatment. These observations are consistent with the reports of a dose-related half-life of rFVIIa. These data imply that the administration of rFVIIa may have an effect in non-coagulopathic individuals. Although the heterogenity observed in this sample of patients may limit interpretations, it is proposed that this ex-vivo model with further refinements may be useful for evaluating the effects of future pro-hemostatic agents. Figure Figure
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Shukla, Shilpa, Jason R. Fritz, Crystal A. Kyaw, Sara P. Valentino, Catherine W. Imossi, Susmita N. Sarangi, Kevin J. Tracey, Christopher J. Czura, and Jared M. Huston. "Cholinergic Stimulation Improves Hemostasis in a Hemophilia Mouse Model." Blood 126, no. 23 (December 3, 2015): 3528. http://dx.doi.org/10.1182/blood.v126.23.3528.3528.
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Abstract Background: Management of bleeding in hemophilia patients that develop inhibitors remains a challenging clinical problem. Current bypassing agents are expensive, not universally effective and have pro-thrombotic potential. New approaches, including bispecific antibodies and others have shown potential but still require additional testing in clinical trials. As such, there is a clear need for alternative therapeutic options. The inflammatory reflex is an endogenous neuro-immune pathway that monitors and regulates systemic inflammation via the vagus nerve. The afferent vagus nerve can inform the brain of increased peripheral inflammation. The brain then down-regulates systemic inflammation via increased efferent vagus nerve signaling to the spleen, a mechanism referred to as the cholinergic anti-inflammatory pathway (KJ Tracey, Nat Rev Imm 9(6), 2009). Our previous work has demonstrated that activation of the cholinergic anti-inflammatory pathway via direct electrical vagus nerve stimulation (VNS) or by systemic administration of a pharmacological cholinergic agonist inhibits proinflammatory cytokine production and protects against lethal systemic inflammation (JR Fritz, Bioelectron Med 1(1), 2014). While studying the cholinergic anti-inflammatory pathway in mice, we observed that cholinergic stimulation reduces traumatic hemorrhage. To explore this observation further, we developed a porcine model of soft tissue trauma and hemorrhage. Electrical stimulation of the cervical vagus nerve significantly reduces total blood loss and bleeding time in pigs. Electrical vagus nerve stimulation also significantly increases local thrombin generation at the site of injury, whereas systemic thrombin concentrations remain unchanged (CJ Czura, Shock 33(6), 2010). Aim: To evaluate the beneficial hemostatic effects of cholinergic stimulation on tail hemorrhage in a mouse model of hemophilia A. Methods: Male 8-12-week-old, factor VIII knockout mice (B6;129S-F8<tm1Kaz>/J, Jackson Labs) receive either electrical cervical vagus nerve stimulation (1 V, 30 Hz, 2 ms pulse for 5 minutes) or sham stimulation under anesthesia (ketamine/xylazine 140/16 mg/kg, ip) five minutes before 2 mm distal tail transection. In separate experiments, nicotine (2 mg/kg, ip) or saline is administered 17 minutes before injury. Mouse tails are placed into a 37¡ÆC normal saline bath for five minutes prior to injury. After injury, tails are placed into 50 mL conical tubes filled with 37¡ÆC normal saline and allowed to bleed freely for a total of ten minutes. Total shed blood volume and active bleeding time are recorded, and systemic and local thrombin generation (thrombin-antithrombin complex) is measured via ELISA. Complete blood counts are measured via standard clinical assay. Results: Vagus nerve stimulation significantly reduces total blood loss and bleeding time by 75% and 18%, respectively. Administration of nicotine significantly reduces total blood loss and bleeding time by 79% and 41%, respectively (Fig. 1). Nicotine significantly increases local thrombin generation compared with vehicle controls, whereas systemic thrombin generation is unchanged. There are no differences in circulating platelet counts or hematocrit levels. Conclusions: Cholinergic stimulation results in rapid and specific improvements in hemostasis during traumatic hemorrhage in a mouse model of hemophilia A. The molecular mechanisms of localized thrombin generation remain an active area of research. Future clinical trials are necessary to determine if cholinergic stimulation is an efficacious, safe and cost-effective therapy for hemophilia A patients. Figure 1. Vagus nerve stimulation (A) and nicotine administration (B) significantly reduce blood loss following traumatic hemorrhage in hemophilia A mice. Figure 1. Vagus nerve stimulation (A) and nicotine administration (B) significantly reduce blood loss following traumatic hemorrhage in hemophilia A mice. Figure 2. Nicotine administation significantly increases thrombin generation specifically at the site of injury and not systemically. Figure 2. Nicotine administation significantly increases thrombin generation specifically at the site of injury and not systemically. Disclosures No relevant conflicts of interest to declare.
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Keshava-Prasad, Holavanahalli, Krishna Oza, Girindra Raval, and Tamim Antakli. "Management of Intractable Bleeding after Cardiac Surgery with Recombinant Activated Factor VII." Blood 112, no. 11 (November 16, 2008): 4526. http://dx.doi.org/10.1182/blood.v112.11.4526.4526.
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Abstract Background Intractable hemorrhage is a dreaded complication after cardiovascular surgery often requiring re-exploration and the administration of large quantities of blood products. In view of problems with aprotinin, a new safer effective agent is needed. Recombinant activated FVII is approved for use in patients with hemophilia A and B who have inhibitors to factors VIII and IX, and has shown promise in off-label use for the management of life-threatening hemorrhage in several clinical scenarios including cardiac surgery. It may help control bleeding, reduce blood product usage, and avoid potential morbidity. Its exact place in the management of bleeding during and after cardiac surgery is not yet fully known. Methods. We performed a retrospective review of patients who were given recombinant factor VIIa (rFVIIa; Novoseven, NovoNordisk, Copenhagen, Denmark) to control bleeding after major cardiovascular surgery requiring cardiopulmonary bypass (CPB) at our institution. The decision to administer rFVIIa was made empirically based on the observation by the surgeons of refractory bleeding that appeared unresponsive to conventional hemostasis agents including the requirement of large volumes of blood components, and was at least severe enough to prevent chest closure. We compared blood loss and blood component usage in patients before and after rFVIIa. We also performed a detailed review of the English literature to determine the role of rFVIIa in the treatment of bleeding after cardiac surgery. Results. Between August 2002 to February 2006, 1295 patients underwent open heart surgery at our institution; of these, 28 were given Novoseven either to control intractable bleeding, or to prevent major bleeding. Table 1 shows the patient characteristics. Satisfactory hemostasis was achieved in all but 3 patients after a single 90 μg/kg intravenous dose of rFVIIa. In all patients, there was a dramatic reduction in the amount of blood components (PRBCs, Platelets and FFP) used after rFVIIa infusion (Table 2). Cryoprecipitate was administered routinely with rFVIIa and its usage did not change significantly (Table 2). No thromboembolic or other complications directly related to rVIIa occurred. Conclusions. We have demonstrated that intravenous rFVIIa is effective, safe, and valuable in the management of intractable bleeding after complicated cardiac surgeries. There are several reports and reviews in the literature which corroborate our experience and indicate that recombinant factor VIIa is a potent pro-hemostatic agent which has a role in the treatment of life-threatening refractory hemorrhage associated with cardiac surgery. Earlier preemptive administration of rFVIIa during or before surgery may be of value in patients at high risk of intractable bleeding in order to limit blood loss, and to avoid potential morbidity from large volume blood component transfusions. Randomized, controlled trials are warranted to assess the efficacy, safety, and cost-benefit of this intervention in cardiac surgical patients. TABLE 1. Characteristics and operative course of the 28 patients Mean age 60 yrs (range 22–85) Male, M 24(85%); F 4(15%) Total number of surgical procedures performed: 34 Aortic valve: 7; Bentall or modified Bentall: 9 (3 emergent) Mitral Valve Replacement: 4; CABG: 10; Redo 2 Left pneumonectomy/resection of L Atrial cuff & pericardium: 1 Removal of Inferior vena cava tumor (Renal cell ca): Re-exploration: 6; Delayed closure: 5; Both re-exploration and delayed closure: Median bypass time: 214 min (65–358) Timing of Novoseven: intra op: 21 including elective use in 2 pts; post op: 7 Dose of Novoseven: 90mcg/kg in 22; 45 mcg/kg 2 patients Responders 25(89%) Outcome: Deaths 11(38%) Autopsies: 2; no evidence of systemic thrombosis Table 2. Details of the blood products administered both before and after rFVIIa infusion. Componen Mean units Before rVIIa Mean units After rVIIa Difference; p value PRBC usage 15.9 5.033333 0.045 Platelet usage 4.448276 1.37931 0.005 FFP Usage 9.931034 5.793103 0.042 Cryoppt 21.71429 12.54167 0.091
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"Glanzmann’s Thrombasthenia in the Parturient Undergoing Cesarean Delivery- A Case Report and Discussion of Clinical Consideration." Anesthesia and Perioperative Medicine, May 17, 2021, 1–3. http://dx.doi.org/10.36879/apm.2021.000103.
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Glanzmann thrombasthenia (GT) is a rare hemorrhagic disorder characterized by the absence of IIb-IIIa platelet glycoprotein (GPIIbIIIa) on the platelet surface. Many GT patients experience recurrent bleeding and require frequent platelet transfusion. Therefore, they are at risk for alloimmunization against platelet glycoprotein or Human leucocyte antigen (HLA) epitopes resulting in platelet refractoriness and therapeutic failure. We report a unique case of GT in pregnancy at risk for severe postpartum hemorrhage due to anti-GP IIb/IIIa and anti-HLA antibodies and the use of pro-hemostatic agents to optimize outcomes. A multidisciplinary approach is essential to achieving the best possible outcomes for these patients.
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Salter, Brittany, and Mark Crowther. "A Historical Perspective on the Reversal of Anticoagulants." Seminars in Thrombosis and Hemostasis, September 2, 2022. http://dx.doi.org/10.1055/s-0042-1753485.
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AbstractThere has been a landmark shift in the last several decades in the management and prevention of thromboembolic events. From the discovery of parenteral and oral agents requiring frequent monitoring as early as 1914, to the development of direct oral anticoagulants (DOACs) that do not require monitoring or dose adjustment in the late 20th century, great advances have been achieved. Despite the advent of these newer agents, bleeding continues to be a key complication, affecting 2 to 4% of DOAC-treated patients per year. Bleeding is associated with substantial morbidity and mortality. Although specific reversal agents for DOACs have lagged the release of these agents, idarucizumab and andexanet alfa are now available as antagonists. However, the efficacy of these reversal agents is uncertain, and complications, including thrombosis, have not been adequately explored. As such, guidelines continue to advise the use of nonspecific prohemostatic agents for patients requiring reversal of the anticoagulant effect of these drugs. As the indications for DOACs and the overall prevalence of their use expand, there is an unmet need for further studies to determine the efficacy of specific compared with nonspecific pro-hemostatic reversal agents. In this review, we will discuss the evidence behind specific and nonspecific reversal agents for both parenteral and oral anticoagulants.
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Kottana, Regina Komal, and Anne-Laure Papa. "Abstract 611: Effect Of Tumor-derived Extracellular Vesicles On Platelet Function." Arteriosclerosis, Thrombosis, and Vascular Biology 43, Suppl_1 (May 2023). http://dx.doi.org/10.1161/atvb.43.suppl_1.611.
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Tumor-derived extracellular vesicles (EVs) have been described as pro-coagulant particles causing cancer-associated thrombosis. Tumor-derived EVs circulating in the bloodstream may interact with platelets or trigger the coagulation cascade, leading to venous and arterial thrombosis. In order to better understand the interaction of EVs with the hemostatic components, we aim to determine the mechanistic aspects of their interaction with platelets and their potential role in initiating coagulation. Given the wide variation of protein and integrin expression between different cancer cell types and the expectation of varying mechanistic effects, we have selected to study lung (A549), breast (MDA-MB-231) and glioblastoma (LN18) cell lines. We have characterized the effect of EVs derived from these cell lines on platelet function of aggregation and activation. Our data shows that both A549 and MDA-MB-231 EVs at 50 ug/mL significantly increase platelet aggregation as compared to control when incubated with the platelets for 10 min and in the presence of an agonist (ADP 20 uM) (Fig. 1). We confirmed the expression of phosphatidylserine on the surface of EVs using Annexin V and flow cytometry, indicating that they are pro-coagulant in nature. Furthermore, we will be investigating interaction of EVs with platelet receptor GP IIb-IIIa, agonist release, and presence of proteins such as Tissue Factor, as possible mechanisms for EV-induced platelet aggregation. Thus, the understanding of the mechanistic aspects behind the interaction between EVs and the hemostatic components could prove beneficial in the design of next generation agents for cancer-associated thrombosis. Figure 1. Platelet aggregation measured by light transmission aggregometry with ADP 20uM in the presence of A549 and MDA-MB-231 EVs (n=8 to 11)
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He, Shu, Honglie Cao, Charlotte Thålin, Jan Svensson, Margareta Blombäck, and Håkan Wallén. "The Clotting Trigger Is an Important Determinant for the Coagulation Pathway In Vivo or In Vitro—Inference from Data Review." Seminars in Thrombosis and Hemostasis, December 21, 2020. http://dx.doi.org/10.1055/s-0040-1718888.
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AbstractBlood coagulation comprises a series of enzymatic reactions leading to thrombin generation and fibrin formation. This process is commonly illustrated in a waterfall-like manner, referred to as the coagulation cascade. In vivo, this “cascade” is initiated through the tissue factor (TF) pathway, once subendothelial TF is exposed and bound to coagulation factor VII (FVII) in blood. In vitro, a diminutive concentration of recombinant TF (rTF) is used as a clotting trigger in various global hemostasis assays such as the calibrated automated thrombogram, methods that assess fibrin turbidity and fibrin viscoelasticity tests such as rotational thromboelastometry. These assays aim to mimic in vivo global coagulation, and are useful in assessing hyper-/hypocoagulable disorders or monitoring therapies with hemostatic agents. An excess of rTF, a sufficient amount of negatively charged surfaces, various concentrations of exogenous thrombin, recombinant activated FVII, or recombinant activated FIXa are also used to initiate activation of specific sub-processes of the coagulation cascade in vitro. These approaches offer important information on certain specific coagulation pathways, while alterations in pro-/anticoagulants not participating in these pathways remain undetectable by these methods. Reviewing available data, we sought to enhance our knowledge of how choice of clotting trigger affects the outcome of hemostasis assays, and address the call for further investigations on this topic.
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Ginovyan, Mikayel, Hayarpi Javrushyan, Hasmik Karapetyan, Izabela Koss‐Mikołajczyk, Barbara Kusznierewicz, Anna Grigoryan, Alina Maloyan, Agnieszka Bartoszek, and Nikolay Avtandilyan. "Hypericum alpestre extract exhibits in vitro and in vivo anticancer properties by regulating the cellular antioxidant system and metabolic pathway of L‐arginine." Cell Biochemistry and Function 42, no. 1 (January 2024). http://dx.doi.org/10.1002/cbf.3914.
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AbstractConventional treatment methods are not effective enough to fight the rapid increase in cancer cases. The interest is increasing in the investigation of herbal sources for the development of new anticancer therapeutics. This study aims to investigate the antitumor capacity of Hypericum alpestre (H. alpestre) extract in vitro and in vivo, either alone or in combination with the inhibitors of the l‐arginine/polyamine/nitric oxide (NO) pathway, and to characterize its active phytochemicals using advanced chromatographic techniques. Our previous reports suggest beneficial effects of the arginase inhibitor NG‐hydroxy‐nor‐ l‐arginine and NO inhibitor NG‐nitro‐Larginine methyl ester in the treatment of breast cancer via downregulation of polyamine and NO synthesis. Here, the antitumor properties of H. alpestre and its combinations were explored in vivo, in a rat model of mammary gland carcinogenesis induced by subcutaneous injection of 7,12‐dimethylbenz[a]anthracene. The study revealed strong antiradical activity of H. alpestre aerial part extract in chemical (DPPH/ABTS) tests. In the in vitro antioxidant activity test, the H. alpestre extract demonstrated pro‐oxidant characteristics in human colorectal (HT29) cells, which were contingent upon the hemostatic condition of the cells. The H. alpestre extract expressed a cytotoxic effect on HT29 and breast cancer (MCF‐7) cells measured by the MTT test. According to comet assay results, H. alpestre extract did not exhibit genotoxic activity nor possessed antigenotoxic properties in HT29 cells. Overall, 233 substances have been identified and annotated in H. alpestre extract using the LC‐Q‐Orbitrap HRMS system. In vivo experiments using rat breast cancer models revealed that the H. alpestre extract activated the antioxidant enzymes in the liver, brain, and tumors. H. alpestre combined with chemotherapeutic agents attenuated cancer‐like histological alterations and showed significant reductions in tumor blood vessel area. Thus, either alone or in combination with Nω‐OH‐nor‐ l‐arginine and Nω‐nitro‐ l‐arginine methyl ester, H. alpestre extract exhibits pro‐ and antioxidant, antiangiogenic, and cytotoxic effects.
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Stalker, Timothy J., Elizabeth A. Traxler, Samantha Cermignano, Roman Voronov, Scott L. Diamond, and Lawrence F. Brass. "Abstract 55: Spatiotemporal Heterogeneity of Platelet Activation and Thrombus Architecture Regulates Plasma Protein Accessibility During Thrombus Formation in Vivo." Arteriosclerosis, Thrombosis, and Vascular Biology 32, suppl_1 (May 2012). http://dx.doi.org/10.1161/atvb.32.suppl_1.a55.
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While often described as a linear process in which platelets react in a uniform manner, recent in vivo studies demonstrate that the platelet response to vascular injury is quite heterogeneous. Here, we utilized confocal fluorescence intravital microscopy to provide a detailed description of the heterogeneous nature of thrombi formed in vivo while asking the following questions: 1) does platelet accumulation and activation during thrombus formation follow specific spatio-temporal patterns that result in subpopulations of platelets with different activation states; 2) does the architecture of the hemostatic mass depend on the mechanism of injury; and 3) does heterogeneity within the platelet mass result in discrete regions of a thrombus that are more or less accessible to plasma and its constituents? We found that regardless of the mode of injury, stable thrombi were composed of two distinct and contiguous populations of platelets defined both by their activation state and their packing density. Closest to the site of injury was a population of fully activated, stably adherent, degranulated platelets, which was covered by multiple layers of loosely adherent platelets that had not undergone granule release. Increased packing density of platelets within the thrombus core was associated with reduced plasma volume in this region as well as limited accessibility of large plasma borne tracer molecules and proteins. Taken together, these findings clearly demonstrate the heterogeneous nature of thrombus formation at the molecular and structural levels. Further, the finding of reduced plasma volume and protein accessibility to the thrombus core has implications for delivery of pro- and anti-thrombotic blood factors and therapeutic agents to this region of a thrombus.