scholarly journals Platelet Priming Following Naturally Occurring Thermal Burn Injuries And Wildfire Smoke Exposure is Associated with Intracardiac Thrombosis in Feline Survivors

Author(s):  
Ronald H. L. Li ◽  
Avalene W. K. Tan ◽  
Yu Ueda ◽  
Joshua A. Stern ◽  
Mehrab Hussain ◽  
...  

Abstract Wildfires pose a major health risk for humans, wildlife, and domestic animals. We previously discovered pathophysiologic parallels between domestic cats with naturally occurring smoke inhalation and thermal burn injuries and human beings with similar injuries; these were characterized by transient myocardial thickening, cardiac troponin I elevation and formation of intracardiac thrombosis. While the underlying mechanisms remain unclear, results from murine models suggest that platelet priming and activation may contribute to a global hypercoagulable state and thrombosis. Herein, we evaluated and compared the degree of platelet activation, platelet response to physiologic agonists and levels of platelet-derived microvesicles (PDMV) in 29 cats with naturally occurring wildfire thermal injuries, 21 clinically healthy cats with subclinical hypertrophic cardiomyopathy (HCM) and 11 healthy cats without HCM (CC). We also quantified and compared circulating PDMVs in WF cats to CC cats. In addition, we examined the association between thrombotic events, severity of burn injuries, myocardial changes, and the degree of platelet activation in cats exposed to wildfires. Flow cytometric detection of platelet surface P-selectin expression showed that cats in the wildfire group (WF) had increased platelet response to adenosine diphosphate (ADP) and thrombin compared to the 2 control groups indicating the presence of primed platelets in circulation. In addition, cats in the WF group had increased circulating levels of PDMV, characterized by increased phosphatidylserine on the external leaflet. Cats in the WF group with documented intracardiac thrombosis had elevated platelet activation and platelet priming in the presence of ADP or thrombin. While high dose arachidonic acid (AA) mostly resulted in platelet inhibition, persistent platelet activation in response to AA was noted among cats in the WF group with intracardiac thrombosis. Univariate and multiple logistic regression analyses demonstrated that increased platelet response to AA was independently associated with thrombotic events. This is the first study reporting the significant association between platelet priming and intracardiac thrombosis in domestic cats with naturally occurring wildfire-related injuries and smoke inhalation. Further studies are required to delineate additional mechanisms between inflammation and thrombosis, especially regarding platelet primers and the cyclooxygenase pathway.

1996 ◽  
Vol 16 (12) ◽  
pp. 1532-1543 ◽  
Author(s):  
Michel Humbert ◽  
Paquita Nurden ◽  
Claude Bihour ◽  
Jean-Max Pasquet ◽  
Joëlle Winckler ◽  
...  

Our study investigated the effect of the antithrombotic drug clopidogrel (75 mg/d for 7 days) on the ultrastructure of platelet aggregates induced by ADP or 2-methylthio-ADP (2-MeS-ADP) in citrated platelet-rich plasma and examined the activation state of the GP IIb/IIIa complexes. Results were compared with those obtained for patient M.L., who has a congenital disorder characterized by a reduced and reversible platelet response to ADP. When untreated normal platelets were stimulated with high-dose ADP, electron microscopy revealed large and stable aggregates often surrounded by a layer of what appeared to be degranulated platelets. The reversible aggregates of platelets from subjects receiving clopidogrel or from patient M.L. did not show this layer. Electron microscopy showed that in both situations, the aggregates were composed of loosely bound platelets with few contact points. Immunogold labeling of ultrathin sections of Lowicryl-embedded aggregates formed by ADP or 2-MeS-ADP showed a much decreased platelet surface staining by (1) a polyclonal anti-fibrinogen antibody and (2) AP-6, a murine anti–ligand-induced binding site monoclonal antibody specific for GP IIb/IIIa complexes occupied with fibrinogen. Similar findings were seen after disaggregation, when many single platelets were present that showed no signs of secretion. Flow cytometry confirmed that the number of ligand-occupied GP IIb/IIIa complexes was much lower on platelets stimulated with ADP or 2-MeS-ADP after clopidogrel treatment. As expected from previous studies, ADP-induced platelet shape change and Ca 2+ influx were unaffected by clopidogrel. These results agree with the hypothesis that platelet activation by ADP is biphasic and highlight a receptor-induced activation pathway affected by clopidogrel (or congenitally impaired in patient M.L.) that is necessary for the full activation of GP IIb/IIIa and the formation of stable macroaggregates.


1979 ◽  
Author(s):  
M.L. Tiffany ◽  
J.A. Penner

Confusing data exists in the literature on the effect of heparin on platelet aggregation. Clarification is achieved by the discovery that heparin, itself, produces some platelet aggregation with maximal effect a t 0.6 U/ml heparin with human PRP diluted 1:1 with PO4 buffered saline at pH 7.6. The extra stimulus of platelet activation by heparin (usually unnoticed) increases synergistically the platelet response to most aggregating agents with maximal effect also at 0.6 U/ml of added heparin. With collagen, as with thrombin, however, the addition of heparin decreases platelet aggrega tion due to specific binding of heparin. At high concentrations of heparin (25-50 U/ml) all platelet aggregation is inhibited. Excess heparin, over and above the amount that can bind to the platelets produces electrostatic shielding that prevents the negatively charged platelets from interacting. It is also possible that aggregation is reduced by a loss of fibrinogen and fibronectin from the platelet surface by the formation of soluble heparin complexes.


2005 ◽  
Vol 93 (06) ◽  
pp. 1137-1146 ◽  
Author(s):  
Ali Amirkhosravi ◽  
Todd Meyer ◽  
Farooq Siddiqui ◽  
Sarfraz Ahmad ◽  
Jamie Walker ◽  
...  

SummaryOur initial finding that CD40– and CD40 ligand (CD40L)-deficient mice displayed prolonged tail bleeding and platelet function analyzer (PFA-100) closure times prompted us to further investigate the role of the CD40-CD40L dyad in primary hemostasis and platelet function. Recombinant human soluble CD40L (rhs CD40L), chemical cross-linking of which suggested a trimeric structure of the protein in solution, activated platelets in a CD40-dependent manner as evidenced by increased CD62P expression. CD40 monoclonal antibody (mAb) M3, which completely blocked rhs CD40L-induced platelet activation, also prolonged PFA-100 closure times of normal human blood. In contrast, CD40 mAb G28–5 showed less potential in blocking rhs CD40L-induced CD62P expression and did not affect PFA-100 closure times. However, when added to the platelets after rhs CD40L, G28–5 significantly enhanced the platelet response by causing clustering of, and signaling through, FcγRII. Similarly, higher order multimeric immune complexes formed at a 1/3 molar ratio of M90, a CD40L mAb, to rhs CD40L induced strong FcγRII-mediated platelet activation when translocated to the platelet surface in a CD40-dependent manner, including the induction of morphological shape changes, fibrinogen binding, platelet aggregation, dense granule release, microparticle generation and monocyte-platelet-conjugate formation. The results suggest that CD40 may play a role in primary hemostasis and platelet biology by two independent mechanisms: First, by functioning as a primary signaling receptor for CD40L and, second, by serving as a docking molecule for CD40L immune complexes. The latter would also provide a potential mechanistic explanation for the unexpected high incidence of CD40L mAb-associated thrombotic events in recent human and animal studies.Parts of this work have been presented on the 46th Annual Meeting of the American Society of Hematology (San Diego, 2004).


Blood ◽  
2000 ◽  
Vol 95 (4) ◽  
pp. 1301-1308 ◽  
Author(s):  
James P. McRedmond ◽  
Patrick Harriott ◽  
Brian Walker ◽  
Desmond J. Fitzgerald

Streptokinase activates platelets, limiting its effectiveness as a thrombolytic agent. The role of antistreptokinase antibodies and proteases in streptokinase-induced platelet activation was investigated. Streptokinase induced localization of human IgG to the platelet surface, platelet aggregation, and thromboxane A2production. These effects were inhibited by a monoclonal antibody to the platelet Fc receptor, IV.3. The platelet response to streptokinase was also blocked by an antibody directed against the cleavage site of the platelet thrombin receptor, protease-activated receptor-1 (PAR-1), but not by hirudin or an active site thrombin inhibitor, Ro46-6240. In plasma depleted of plasminogen, exogenous wild-type plasminogen, but not an inactive mutant protein, S741A plasminogen, supported platelet aggregation, suggesting that the protease cleaving PAR-1 was streptokinase-plasminogen. Streptokinase-plasminogen cleaved a synthetic peptide corresponding to PAR-1, resulting in generation of PAR-1 tethered ligand sequence and selectively reduced binding of a cleavage-sensitive PAR-1 antibody in intact cells. A combination of streptokinase, plasminogen, and antistreptokinase antibodies activated human erythroleukemic cells and was inhibited by pretreatment with IV.3 or pretreating the cells with the PAR-1 agonist SFLLRN, suggesting Fc receptor and PAR-1 interactions are necessary for cell activation in this system also. Streptokinase-induced platelet activation is dependent on both antistreptokinase-Fc receptor interactions and cleavage of PAR-1.


1986 ◽  
Vol 55 (02) ◽  
pp. 268-270
Author(s):  
R J Alexander

SummaryAn attempt was made to isolate from plasma the platelet surface substrate for thrombin, glycoprotein V (GPV), because a GPV antigen was reported to be present in plasma (3). Plasma fractionation based on procedures for purification of GPV from platelets revealed a thrombin-sensitive protein with appropriate electrophoretic mobility. The protein was purified; an antiserum against it i) reacted with detergent-solubilized platelet proteins or secreted proteins in a double diffusion assay, ii) adsorbed a protein from the supernatant solution of activated platelets, and iii) inhibited thrombin-induced platelet activation, but the antiserum did not adsorb labeled GPV. The purified protein was immunochemically related to prothrombin rather than to GPV. Other antibodies against prothrombin were also able to adsorb a protein from platelets. It is concluded that 1) plasma does not contain appreciable amounts of GPV, and 2) platelets contain prothrombin or an immunochemically similar protein.


2013 ◽  
Vol 34 (9) ◽  
pp. 395-404
Author(s):  
Ramin Jamshidi ◽  
Thomas T. Sato

2013 ◽  
Vol 33 (suppl_1) ◽  
Author(s):  
Marieli Gonzalez ◽  
Fiorella Reyes ◽  
Deborah Marrero ◽  
A V Washington

Platelet activation at sites of inflammation triggers the secretion of molecules that induce the transition of atherosclerosis from fatty streak to an acute disease, featuring an increased vulnerability of the atherosclerotic lesion that results in plaque rupture and thrombosis. TLT-1 (Triggering Receptor Expressed in Myeloid cells (TREM)-like transcript-1) is a molecule exclusively found in the α-granules of megakarocytes and platelets and has a demonstrated effect in inflammatory responses. Upon platelet activation, TLT-1 is moved to the platelet surface, while its soluble form, s-TLT-1, is secreted and detected in serum. Studies using the C57Bl/6 treml1 - /- mouse demonstrated a predisposition to hemorrhage after an acute inflammatory challenge suggesting that TLT-1 may be a key regulatory molecule in the interface between hemostatic and inflammatory mechanisms. Because we have found that sTLT-1 levels are significantly elevated in apoE mice when compared to wild type, we hypothesized that TLT-1 may be playing an important role in the progression of atherosclerosis. To address this possibility, we generated apoE - /- / treml1 - /- double knockout mice [DN]. Assessment of lesions after 4 weeks high-fat diet (HFD) demonstrated that at early stages, TLT-1 deficiency accelerates fatty streak formation. After 20 weeks on HFD, lesions in both apoE - /- and [DN] mice progressed to an advance fibrous plaque stage. Although their lesion sizes were not substantially different, lesion compositions were. The mechanistic basis of these differences appears to be that the [DN] mice have significantly higher cholesterol levels when compared to apoE - /- mice. The increased cholesterol levels extend to the treml1 -/- mouse when compared to wild type mice at 4 weeks on HFD, this difference, however, gradually subsides as wild type mice cholesterol levels increase over 20 weeks. Interestingly, cholesterol levels in 50 week old mice on chow diet revealed minimal differences between test and control mice suggesting the higher cholesterol levels are related to increased dietary intake. Our work defines a surprising role for TLT-1 in the regulation of serum cholesterol levels during atherogenesis.


1999 ◽  
Vol 82 (08) ◽  
pp. 365-376 ◽  
Author(s):  
Steve Watson

IntroductionThe extracellular matrix protein, collagen, plays a primary role in hemostasis. Collagen fibers provide an important site for adhesion of platelets to the exposed subendothelium, trapping them at the site of vascular damage and enabling the formation of a monolayer of cells over the damaged area. Collagen fibers also stimulate platelet activation, leading to inside-out regulation of the integrin glycoprotein (GP) IIb-IIIa (also known as αIIbβ3), secretion from dense and α granules, generation of thromboxanes, and expression of procoagulant activity, all of which support the hemostatic process. The role of collagen in supporting platelet adhesion to the subendothelium is mediated through indirect and direct interactions. The indirect interaction is mediated through von Willebrand factor (vWF), which binds to the GP Ib-IX-V complex on the platelet surface.1-3 The interaction with vWF is critical for platelet adhesion at medium to high rates of flow because of the fast rate of association between vWF and GP Ib-IX. The importance of this interaction is demonstrated by the severe bleeding problems experienced by individuals with functional impairment of vWF (von Willebrand disease) or GP Ib-IX (Bernard-Soulier syndrome). At low rates of flow, collagen fibers are able to support adhesion in the absence of vWF through a direct interaction with a number of platelet surface glycoproteins i.e. collagen receptors,4,5 this also serves to support vWF-dependent adhesion at higher rates of flow by preventing dissociation. Crosslinking of platelet surface glycoproteins by collagen also generates intracellular signals, leading to platelet activation.The number of proteins on the platelet surface proposed to be collagen receptors is approaching double figures, but it is generally accepted that the integrin GP Ia-IIa (also known as α2β1) and glycoprotein VI (GP VI) are among the most important of these, playing critical roles in adhesion and activation, respectively6 (Fig. 1). This is illustrated by the mild bleeding problems of patients with a low level of expression or the presence of autoantibodies to GP Ia-IIa and the spontaneous, severe bleeding episodes that are occasionally seen in patients whose platelets are deficient in GP VI.6 There is evidence, however, that other collagen receptors have supporting roles in adhesion and activation. For example, GP VI supports platelet adhesion to collagen7 and GP IV, also known as CD36, may also play a similar role.8 The role of the recently cloned collagen receptor p65 in adhesion is not known. Evidence that the interaction of collagen with receptors, such as GPIV and p65, is of less importance than for interactions with GP Ia-IIa, and GP VI is provided by the absence of individuals with bleeding problems caused by deficiencies in these proteins. This is illustrated most clearly for GP IV, which is absent in 3% to 5 % of the Japanese population, and yet such individuals display no major vascular problems.Due to the large number of glycoproteins that bind collagen on the platelet surface, it has been difficult to gain a full understanding of the role of individual collagen receptors in adhesion and activation responses. This is complicated further by the interactions between vWF and GP Ib-IX-V, vWF or fibrinogen to activated GP IIb-IIIa especially as both glycoprotein receptors generate intracellular signals. The relative importance of individual collagen receptors in adhesion also varies with the rate of flow and between collagen types. A full discussion of platelet adhesion to collagen is beyond the scope of this article, and the reader is referred to a number of excellent recent reviews for further information.4-6,9,10 The present chapter focuses on the signaling events generated by the activation (or more correctly crosslinking) of platelet surface glycoproteins by collagen and the implications that this has for platelet activation under normal and diseased conditions.


2016 ◽  
Vol 115 (02) ◽  
pp. 324-332 ◽  
Author(s):  
Rabie Jouni ◽  
Heike Zöllner ◽  
Ahmad Khadour ◽  
Jan Wesche ◽  
Anne Grotevendt ◽  
...  

SummaryProtamine (PRT) is the standard drug to neutralise heparin. PRT/heparin complexes induce an immune response similar to that observed in heparin-induced thrombocytopenia (HIT). Partially desulfated heparin (ODSH) was shown to interfere with anti-platelet factor 4/heparin antibodies (Abs), which are responsible for HIT. In this study, we analyse the impact of ODSH on the interaction between anti-PRT/heparin Abs and platelets. The ability of ODSH to prevent anti-PRT/heparin Ab-induced platelet destruction in vivo was investigated using the NOD/ SCID mouse model. ODSH improved platelet survival in the presence of PRT, heparin and anti-PRT/heparin Abs (median platelet survival after 300 minutes (min) with 20 μg/ml ODSH: 75 %, range 70–81 % vs without ODSH: 49%, range 44–59%, p=0.006). Furthermore, when ODSH was applied 60 min after Ab injection platelet survival was improved (median platelet survival after 300 min with ODSH: 83 %, range 77–93 % vs without ODSH: 59 %, range 29–61 %, p=0.02). In in vitro experiments ODSH inhibited platelet activation at concentrations > 16 μg/mL (p< 0.001), as well as PRT/heparin complex binding to platelets (mean fluorescence intensity [MFI] without ODSH: 85 ± 14 vs with ODSH: 15 ± 0.6, p=0.013). ODSH also displaced pre-bound complexes from the platelet surface (MFI without ODSH: 324 ± 43 vs with 32 μg/ml ODSH: 53 ± 9, p< 0.001). While interfering with platelet activation by anti-PRT/heparin Abs, up to a concentration of 16 μg/ml, ODSH had only minimal impact on neutralisation of heparin by PRT. In conclusion, our study shows that ODSH is able to inhibit platelet activation and destruction suggesting a potential clinical use to reduce anti-PRT/heparin Ab-mediated adverse effects.


Blood ◽  
1987 ◽  
Vol 70 (2) ◽  
pp. 511-515
Author(s):  
DJ Carty ◽  
DL Freas ◽  
AR Gear

We developed a general quenched-flow approach to study platelet function as early as 0.3 seconds after stimulation. Phosphorylation of 20- and 47-kiloDalton (kD) proteins was analyzed during the first 5 seconds of platelet response to ADP from 0.5 to 10.0 mumol/L and compared with the progress of aggregation. The onset time for aggregation and phosphorylation of both proteins was less than 1 second; 20-K phosphorylation was increased greater than 200% and 47-K phosphorylation was increased 50%. The ADP sensitivity of 20-K phosphorylation was greater than that of 47-K phosphorylation (P less than .025), and of that of aggregation (P less than .01), with Ka values of 0.7, 1.0, and 1.2 mumol/L of ADP, respectively. The cyclooxygenase inhibitor indomethacin had no effect on aggregation, but inhibited both phosphorylations. Its inhibition of 20-K phosphorylation was greater than that of 47-K phosphorylation. Platelet activation by ADP thus induced biochemical changes well before 1 second. The quenched- flow approach may help to reveal relationships between phospholipase activation, calcium fluxes, and protein phosphorylation during these early periods of platelet activation.


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