RESUMEN
Increasing evidence is implicating roles for platelets in the development and progression of ovarian cancer, a highly lethal disease that can arise from the fallopian tubes, and has no current method of early detection or prevention. Thrombosis is a major cause of mortality of ovarian cancer patients suggesting that the cancer alters platelet behavior. The objective of this study was to develop a cell culture model of the pathological interactions of human platelets and ovarian cancer cells, using normal FT epithelial cells as a healthy control, and to test effects of the anti-platelet dihomo-gamma-linolenic acid (DGLA) in the model. Both healthy and cancer cells caused platelet aggregation, however platelets only affected spheroid formation by cancer cells and had no effect on healthy cell spheroid formation. When naturally-formed spheroids of epithelial cells were exposed to platelets in transwell inserts that did not allow direct interactions of the two cell types, platelets caused increased size of the spheroids formed by cancer cells, but not healthy cells. When cancer cell spheroids formed using magnetic nanoshuttle technology were put in direct physical contact with platelets, the platelets caused spheroid condensation. In ovarian cancer cells, DGLA promoted epithelial-to-mesenchymal (EMT) transition at doses as low as 100 µM, and inhibited metabolic viability and induced apoptosis at doses ≥150 µM. DGLA doses ≤150 µM used to avoid direct DGLA effects on cancer cells, had no effect on the pathological interactions of platelets and ovarian cancer cells in our models. These results demonstrate that the pathological interactions of platelets with ovarian cancer cells can be modeled in cell culture, and that DGLA has no effect on these interactions, suggesting that targeting platelets is a rational approach for reducing cancer aggressiveness and thrombosis risk in ovarian cancer patients, however DGLA is not an appropriate candidate for this strategy.
Asunto(s)
Ácido 8,11,14-Eicosatrienoico , Plaquetas , Células Epiteliales , Neoplasias Ováricas , Esferoides Celulares , Humanos , Plaquetas/efectos de los fármacos , Femenino , Células Epiteliales/efectos de los fármacos , Neoplasias Ováricas/patología , Ácido 8,11,14-Eicosatrienoico/farmacología , Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Esferoides Celulares/efectos de los fármacos , Línea Celular Tumoral , Agregación Plaquetaria/efectos de los fármacos , Técnicas de Cultivo de Célula/métodos , Comunicación Celular/efectos de los fármacosRESUMEN
BACKGROUND: This study aimed to explore the effect of a P2Y12 inhibitor regimen on the occurrence of postoperative atrial fibrillation (POAF) after off-pump coronary artery bypass graft surgery in carriers with the cytochrome P450 family 2 subfamily C member19 loss-of-function allele. METHODS AND RESULTS: From May 2019 to November 2023, patients containing the cytochrome P450 family 2 subfamily C member19*2 or *3 allele undergoing elective first-time off-pump coronary artery bypass graft surgery including aspirin 100 mg/d and ticagrelor 180 mg/d (AT group; n=95) versus clopidogrel 75 mg/d (aspirin and clopidogrel group; n=95) were prospectively followed. The primary end point was the cumulative incidence of POAF in a week. The secondary end points were POAF burden, platelet aggregability, systemic immune-inflammation index and heart rate variability. The incidence of POAF was 21.1% in the AT group versus 41.1% in the aspirin and clopidogrel group (hazard ratio, 0.46 [95% CI, 0.27-0.76]; P=0.003). POAF burden, ADP-induced platelet aggregation and systemic immune-inflammation index was notably lower in the AT group than the aspirin and clopidogrel group. Heart rate variability data showed an increase in both high-frequency and SD of normal-to-normal RR intervals in the AT group with a decreased low-frequency/high-frequency ratio, suggesting that the sympathetic/parasympathetic activation was balanced. CONCLUSIONS: In patients carrying the cytochrome P450 family 2 subfamily C member19 loss-of-function allele, an AT regimen after off-pump coronary artery bypass grafting was associated with a lower incidence of POAF, paralleled by lower atrial fibrillation burden, ADP-induced platelet aggregation, lower systemic immune-inflammation index reaction, and a balanced automatic nerve system compared with an aspirin and clopidogrel regimen. Inhibiting the systemic immune-inflammation response and sustaining automatic nerve balance may underlie the therapeutic effect of POAF by a potent antiplatelet combination.
Asunto(s)
Fibrilación Atrial , Clopidogrel , Puente de Arteria Coronaria Off-Pump , Inhibidores de Agregación Plaquetaria , Ticagrelor , Humanos , Clopidogrel/uso terapéutico , Clopidogrel/efectos adversos , Ticagrelor/uso terapéutico , Fibrilación Atrial/etiología , Fibrilación Atrial/epidemiología , Fibrilación Atrial/prevención & control , Fibrilación Atrial/diagnóstico , Masculino , Femenino , Puente de Arteria Coronaria Off-Pump/efectos adversos , Persona de Mediana Edad , Anciano , Inhibidores de Agregación Plaquetaria/uso terapéutico , Inhibidores de Agregación Plaquetaria/efectos adversos , Complicaciones Posoperatorias/epidemiología , Complicaciones Posoperatorias/prevención & control , Incidencia , Citocromo P-450 CYP2C19/genética , Citocromo P-450 CYP2C19/metabolismo , Estudios Prospectivos , Agregación Plaquetaria/efectos de los fármacos , Antagonistas del Receptor Purinérgico P2Y/uso terapéutico , Antagonistas del Receptor Purinérgico P2Y/efectos adversos , Enfermedad de la Arteria Coronaria/cirugía , Resultado del TratamientoRESUMEN
Thrombosis is the primary cause of death in patients with cancer. Resveratrol inhibits platelet activation, a crucial pathophysiological basis of thrombosis, in healthy individuals. However, its effects and mechanisms of action in patients with colon cancer remain unknown. Here, we investigated the effect of resveratrol on platelet adhesion and aggregation in patients with colon cancer. Through numerous in vitro and in vivo analyses, including flow cytometry, western blotting, ELISA, and immunofluorescence and colon cancer rat models, we demonstrated that resveratrol reduced thrombosis in patients with colon cancer by inhibiting the phosphorylation of the MAPK and activating the cyclic-GMP/vasodilator-stimulated phosphoprotein pathway. These findings demonstrate the potential of resveratrol in reducing thrombosis in patients with colon cancer and could be used to develop novel therapeutic strategies for this condition.
Asunto(s)
Moléculas de Adhesión Celular , Neoplasias del Colon , GMP Cíclico , Fosfoproteínas , Activación Plaquetaria , Resveratrol , Trombosis , Resveratrol/farmacología , Resveratrol/uso terapéutico , Trombosis/tratamiento farmacológico , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/complicaciones , Humanos , Animales , Activación Plaquetaria/efectos de los fármacos , Masculino , Ratas , GMP Cíclico/metabolismo , Moléculas de Adhesión Celular/metabolismo , Fosfoproteínas/metabolismo , Femenino , Estilbenos/farmacología , Estilbenos/uso terapéutico , Ratas Sprague-Dawley , Persona de Mediana Edad , Transducción de Señal/efectos de los fármacos , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Agregación Plaquetaria/efectos de los fármacos , Proteínas de MicrofilamentosRESUMEN
The study aimed to evaluate the antithrombotic action of Acrocomia aculeata pulp oil (AAPO) in natura, in an in vitro experimental model. AAPO was obtained by solvent extraction, and its chemical characterization was performed by gas chromatography coupled to a mass spectrometer (GC-MS). In vitro toxicity was evaluated with the Trypan Blue exclusion test and in vivo by the Galleria mellonella model. ADP/epinephrine-induced platelet aggregation after treatment with AAPO (50, 100, 200, 400, and 800 µg/mL) was evaluated by turbidimetry, and coagulation was determined by prothrombin activity time (PT) and activated partial thromboplastin time (aPTT). Platelet activation was measured by expression of P-selectin on the platelet surface by flow cytometry and intraplatelet content of reactive oxygen species (ROS) by fluorimetry. The results showed that AAPO has as major components such as oleic acid, palmitic acid, lauric acid, caprylic acid, and squalene. AAPO showed no toxicity in vitro or in vivo. Platelet aggregation decreased against agonists using treatment with different concentrations of AAPO. Oil did not interfere in PT and aPTT. Moreover, it expressively decreased ROS-induced platelet activation and P-selectin expression. Therefore, AAPO showed antiplatelet action since it decreased platelet activation verified by the decrease in P-selectin expression as well as in ROS production.
Asunto(s)
Fibrinolíticos , Selectina-P , Aceites de Plantas , Agregación Plaquetaria , Especies Reactivas de Oxígeno , Animales , Agregación Plaquetaria/efectos de los fármacos , Selectina-P/metabolismo , Humanos , Aceites de Plantas/farmacología , Aceites de Plantas/química , Especies Reactivas de Oxígeno/metabolismo , Fibrinolíticos/farmacología , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Coagulación Sanguínea/efectos de los fármacos , Activación Plaquetaria/efectos de los fármacosRESUMEN
ABSTRACT: Monoclonal antibodies (mAbs) have provided valuable information regarding the structure and function of platelet αIIbß3. Protein disulfide isomerase (PDI) has been implicated in αIIbß3 activation and binds to thrombin-activated αIIbß3. Using human platelets as the immunogen, we identified a new mAb (R21D10) that inhibits the binding of PDI to platelets activated with thrombin receptor-activating peptide (T6). R21D10 also partially inhibited T6-induced fibrinogen and PAC-1 binding to platelets, as well as T6- and adenosine 5'-diphosphate-induced platelet aggregation. Mutual competition experiments showed that R21D10 does not inhibit the binding of mAbs 10E5 (anti-αIIb cap domain) or 7E3 (anti-ß3 ß-I domain), and immunoblot studies indicated that R21D10 binds to ß3. The dissociation of αIIbß3 by EDTA had a minimal effect on R21D10 binding. Cryogenic electron microscopy of the αIIbß3-R21D10 Fab complex revealed that R21D10 binds to the ß3 integrin-epidermal growth factor 1 (I-EGF1) domain and traps an intermediate conformation of αIIbß3 with semiextended leg domains. The binding of R21D10 produces a major structural change in the ß3 I-EGF2 domain associated with a new interaction between the ß3 I-EGF2 and αIIb thigh domains, which may prevent the swing-out motion of the ß3 hybrid domain required for high-affinity ligand binding and protect αIIbß3 from EDTA-induced dissociation. R21D10 partially reversed the ligand binding priming effect of eptifibatide, suggesting that it could convert the swung-out conformation into a semiextended conformation. We concluded that R21D10 inhibits ligand binding to αIIbß3 via a unique allosteric mechanism, which may or may not be related to its inhibition of PDI binding.
Asunto(s)
Anticuerpos Monoclonales , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria , Unión Proteica , Humanos , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/metabolismo , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/inmunología , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/química , Anticuerpos Monoclonales/química , Ligandos , Plaquetas/metabolismo , Agregación Plaquetaria/efectos de los fármacos , Conformación Proteica , Regulación Alostérica , Proteína Disulfuro Isomerasas/metabolismo , Proteína Disulfuro Isomerasas/química , Proteína Disulfuro Isomerasas/antagonistas & inhibidoresRESUMEN
BACKGROUND: Leukocyte-platelet aggregates comprise a pathogenic link between hemostasis and immunity, but the prerequisites and mechanisms of their formation remain not understood. AIMS: To quantify the formation, composition, and morphology of leukocyte-platelet aggregates in vitro under the influence of various cellular activators. METHODS: Phorbol-12-myristate-13-acetate (PMA), lipopolysaccharide (LPS), thrombin receptor-activating peptide (TRAP-6), and adenosine diphosphate (ADP) were used as cellular activators. Flow cytometry was utilized to identify and quantify aggregates in whole human blood and platelet-rich plasma. Cell types and cellular aggregates were identified using fluorescently labeled antibodies against the appropriate cellular markers, and cell activation was assessed by the expression of appropriate surface markers. For confocal fluorescent microscopy, cell membranes and nuclei were labeled. Neutrophil-platelet aggregates were studied using scanning electron microscopy. RESULTS: In the presence of PMA, ADP or TRAP-6, about 17-38 % of neutrophils and 61-77 % of monocytes formed aggregates with platelets in whole blood, whereas LPS did not induce platelet aggregation with either neutrophils or monocytes due the inability to activate platelets. Similar results were obtained when isolated neutrophils were added to platelet-rich plasma. All the cell types involved in the heterotypic aggregation expressed molecular markers of activation. Fluorescent and electron microscopy of the aggregates showed that the predominant platelet/leukocyte ratios were 1:1 and 2:1. CONCLUSIONS: Formation of leukocyte-platelet aggregates depends on the nature of the cellular activator and the spectrum of its cell-activating ability. An indispensable condition for formation of leukocyte-platelet aggregates is activation of all cell types including platelets, which is the restrictive step.
Asunto(s)
Plaquetas , Leucocitos , Lipopolisacáridos , Agregación Plaquetaria , Acetato de Tetradecanoilforbol , Humanos , Plaquetas/metabolismo , Plaquetas/efectos de los fármacos , Acetato de Tetradecanoilforbol/farmacología , Agregación Plaquetaria/efectos de los fármacos , Leucocitos/efectos de los fármacos , Leucocitos/metabolismo , Lipopolisacáridos/farmacología , Adenosina Difosfato/farmacología , Adenosina Difosfato/metabolismo , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Fragmentos de Péptidos/farmacología , Citometría de Flujo , Monocitos/efectos de los fármacos , Monocitos/metabolismoRESUMEN
Antiplatelet therapy is an important factor influencing the postterm patency rate of carotid artery stenting (CAS). Clopidogrel is a platelet aggregation inhibitor mediated by the adenosine diphosphate receptor and is affected by CYP2C19 gene polymorphisms in vivo. When the CYP2C19 gene has a nonfunctional mutation, the activity of the encoded enzyme will be weakened or lost, which directly affects the metabolism of clopidogrel and ultimately weakens its antiplatelet aggregation ability. Therefore, based on network pharmacology, analyzing the influence of CYP2C19 gene polymorphisms on the antiplatelet therapeutic effect of clopidogrel after CAS is highly important for the formulation of individualized clinical drug regimens. The effect of the CYP2C19 gene polymorphism on the antiplatelet aggregation of clopidogrel after CAS was analyzed based on network pharmacology. A total of 100 patients with ischemic cerebrovascular disease who were confirmed by the neurology department and required CAS treatment were studied. CYP2C19 genotyping was performed on all patients via a gene chip. All patients were classified into the wild-type (WT) group (*1/*1), heterozygous mutation (HTM) group (CYP2C19*1/*2, CYP2C19*1/*3), and homozygous mutation (HMM) group (CYP2C19*2/*2, CYP2C19*2/*3, and CYP2C19*3/*3). High-performance liquid chromatography (HPLC) with tandem mass spectrometry (MS/MS) was used to detect the blood concentration of clopidogrel and the plasma clopidogrel clearance (CL) rate in different groups of patients before and after clopidogrel treatment. The platelet aggregation rate of patients with different genotypes was measured by turbidimetry. The incidences of clopidogrel resistance (CR) and stent thrombosis in different groups after three months of treatment were analyzed. The results showed that among the different CYP2C19 genotypes, patients from the HTM group accounted for the most patients, while patients from the HTM group accounted for the least patients. Similarly, the clopidogrel CL of patients in the HMM group was lower than that of patients in the WT group and HTM group (P < 0.01). The platelet inhibition rate of patients in the HMM group was evidently inferior to that of patients in the WT group and HTM group (P < 0.01). The incidence of CR and stent thrombosis in the WT group was notably lower than that in the HTM and HMM groups (P < 0.01). These results indicate that the CYP2C19 gene can affect CR occurrence and stent thrombosis after CAS by influencing clopidogrel metabolism and platelet count.
Asunto(s)
Clopidogrel , Citocromo P-450 CYP2C19 , Inhibidores de Agregación Plaquetaria , Agregación Plaquetaria , Stents , Humanos , Citocromo P-450 CYP2C19/genética , Clopidogrel/uso terapéutico , Clopidogrel/farmacología , Clopidogrel/farmacocinética , Inhibidores de Agregación Plaquetaria/uso terapéutico , Inhibidores de Agregación Plaquetaria/farmacología , Inhibidores de Agregación Plaquetaria/farmacocinética , Masculino , Femenino , Agregación Plaquetaria/efectos de los fármacos , Anciano , Persona de Mediana Edad , Polimorfismo Genético , Ticlopidina/análogos & derivados , Ticlopidina/uso terapéutico , Ticlopidina/farmacología , Genotipo , Arterias Carótidas/efectos de los fármacos , Arterias Carótidas/cirugíaRESUMEN
Platelets play essential roles in the formation of blood clots by clumping with coagulation factors at the site of vascular injury to stop bleeding; therefore, a reduction in the platelet number or disorder in their function causes bleeding risk. In our research, we developed a method to assess platelet aggregation using an optical approach within a microfluidic chip's channel by evaluating the size of laser speckles. These speckles, associated with slowed blood flow in the microfluidic channel, had a baseline size of 28.54 ± 0.72 µm in whole blood. Removing platelets from the sample led to a notable decrease in speckle size to 27.04 ± 1.23 µm. Moreover, the addition of an ADP-containing agonist, which activates platelets, resulted in an increased speckle size of 32.89 ± 1.69 µm. This finding may provide a simple optical method via microfluidics that could be utilized to assess platelet functionality in diagnosing bleeding disorders and potentially in monitoring therapies that target platelets.
Asunto(s)
Plaquetas , Agregación Plaquetaria , Plaquetas/efectos de los fármacos , Humanos , Agregación Plaquetaria/efectos de los fármacos , Pruebas de Función Plaquetaria/métodos , Pruebas de Función Plaquetaria/instrumentación , Dispositivos Laboratorio en un Chip , Técnicas Analíticas Microfluídicas/instrumentación , Técnicas Analíticas Microfluídicas/métodos , Microfluídica/métodos , Adenosina Difosfato/farmacologíaRESUMEN
In in vitro model of short-term therapeutic inhalation of Xe/O2 mixture, xenon in millimolar concentrations led to a pronounced decrease in induced platelet aggregation in the platelet-enriched blood plasma. The maximum and statistically significant decrease occurred in response to induction by collagen (by ≈30%, p≤0.01) and ADP (by ≈25%, p≤0.01). A slightly weaker but statistically significant reduction in aggregation appeared in response to ristocetin (by ≈12%, p≤0.01) and epinephrine (by ≈9%, p≤0.01). It should be noted that the spontaneous aggregation exceeded the reference values in the control group. Nevertheless, even at minimal absolute values, spontaneous platelet aggregation decreased by 2 times in response to xenon (p≤0.01). The reasons for the decrease of spontaneous and induced aggregation are xenon accumulation in the lipid bilayer of the membrane with subsequent nonspecific (mechanical) disassociation of membrane platelet structures and specific block of its distinct from neuronal NMDA receptor.
Asunto(s)
Agregación Plaquetaria , Xenón , Xenón/farmacología , Agregación Plaquetaria/efectos de los fármacos , Humanos , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Adenosina Difosfato/farmacología , Inhibidores de Agregación Plaquetaria/farmacología , Plasma Rico en Plaquetas/metabolismo , Epinefrina/farmacología , Epinefrina/sangre , Colágeno/metabolismoRESUMEN
BACKGROUND: Clopidogrel has been the primary choice of antiplatelet in ischemic stroke that inhibits adenosine diphosphate (ADP)-induced platelet aggregation. P-glycoprotein (P-gp) multidrug resistance-1 (MDR1) is a transmembrane efflux transporter in intestinal cells that plays a significant role in clopidogrel absorption, therefore may affect platelet aggregation. P-gp is encoded by the ABCB1 gene. This study aims to evaluate the effect of ABCB1 polymorphism on clopidogrel response variability in ischemic stroke patients and its genotype frequency. METHODS: A cross-sectional study was conducted in ischemic stroke patients who received clopidogrel between 2020 and 2023 in RSUI/RSCM. All subjects were assessed for ABCB1 polymorphisms C3435T and C1236T. Platelet aggregation were measured using VerifyNow PRU. Clopidogrel response variability was classified into unresponsive (> 208 PRU), responsive (95-208 PRU), and bleeding risk (< 95 PRU). RESULTS: 124 subjects enrolled in this study, with 12,9% of subjects classified as non-responsive/resistant, 49,5% as responsive, and 41,9% as bleeding risk. ABCB1 C1236T homozygote wildtype (CC) was associated with 3,76 times higher bleeding risk than other variants (p = 0,008; 95%CI 1,41 - 10,07). Genotype frequency of ABCB1 C3435T homozygote wildtype, heterozygote, and homozygote variants were 35,9%, 43,5% and 16,9%, respectively; while the genotype frequency of ABCB1 C1236T were 17,8%, 39,5%, and 42,7%, respectively. CONCLUSION: ABCB1 C1236T homozygote wildtype was associated with 3,76 times higher bleeding risk than other variants. The most common genotype frequency of ABCB1 C1236T was homozygote variant; while for ABCB1 C3435T was heterozygote.
Asunto(s)
Subfamilia B de Transportador de Casetes de Unión a ATP , Clopidogrel , Accidente Cerebrovascular Isquémico , Inhibidores de Agregación Plaquetaria , Humanos , Clopidogrel/uso terapéutico , Clopidogrel/administración & dosificación , Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Estudios Transversales , Masculino , Femenino , Persona de Mediana Edad , Accidente Cerebrovascular Isquémico/tratamiento farmacológico , Accidente Cerebrovascular Isquémico/genética , Anciano , Inhibidores de Agregación Plaquetaria/uso terapéutico , Polimorfismo de Nucleótido Simple/genética , Genotipo , Agregación Plaquetaria/efectos de los fármacos , Agregación Plaquetaria/genéticaRESUMEN
ABSTRACT: Glycoprotein VI (GPVI) plays a key role in collagen-induced platelet aggregation. Affimers are engineered binding protein alternatives to antibodies. We screened and characterized GPVI-binding Affimers as novel tools to probe GPVI function. Among the positive clones, M17, D22, and D18 bound GPVI with the highest affinities (dissociation constant (KD) in the nanomolar range). These Affimers inhibited GPVI-collagen-related peptide (CRP)-XL/collagen interactions, CRP-XL/collagen-induced platelet aggregation, and D22 also inhibited in vitro thrombus formation on a collagen surface under flow. D18 bound GPVI dimer but not monomer. GPVI binding was increased for D18 but not M17/D22 upon platelet activation by CRP-XL and adenosine 5'-diphosphate. D22 but not M17/D18 displaced nanobody 2 (Nb2) binding to GPVI, indicating similar epitopes for D22 with Nb2 but not for M17/D18. Mapping of binding sites revealed that D22 binds a site that overlaps with Nb2 on the D1 domain, whereas M17 targets a site on the D2 domain, overlapping in part with the glenzocimab binding site, a humanized GPVI antibody fragment antigen-binding fragment. D18 targets a new region on the D2 domain. We found that D18 is a stable noncovalent dimer and forms a stable complex with dimeric GPVI with 1:1 stoichiometry. Taken together, our data demonstrate that Affimers modulate GPVI-ligand interactions and bind different sites on GPVI D1/D2 domains. D18 is dimer-specific and could be used as a tool to detect GPVI dimerization or clustering in platelets. A dimeric epitope regulating ligand binding was identified on the GPVI D2 domain, which could be used for the development of novel bivalent antithrombotic agents selectively targeting GPVI dimer on platelets.
Asunto(s)
Plaquetas , Agregación Plaquetaria , Glicoproteínas de Membrana Plaquetaria , Unión Proteica , Multimerización de Proteína , Glicoproteínas de Membrana Plaquetaria/metabolismo , Glicoproteínas de Membrana Plaquetaria/química , Humanos , Plaquetas/metabolismo , Ligandos , Agregación Plaquetaria/efectos de los fármacos , Sitios de Unión , Colágeno/metabolismo , Colágeno/química , Proteínas Portadoras , PéptidosRESUMEN
This cohort study aims to assess the connection between cytochrome P450 family 2 subfamily C member 19 (CYP2C19) genotyping, platelet aggregability following oral clopidogrel administration, and the occurrence of postoperative atrial fibrillation (POAF) after off-pump coronary artery bypass graft (CABG) surgery. From May 2017 to November 2022, a total of 258 patients undergoing elective first-time CABG surgery, receiving 100 mg/day oral aspirin and 75 mg/day oral clopidogrel postoperatively, was included for analysis. These patients were categorized based on CYP2C19 genotyping. Platelet aggregability was assessed serially using multiple-electrode aggregometry before CABG, 1 and 5 days after the procedure, and before discharge. The incidences of POAF were compared using the log-rank test for cumulative risk. CYP2C19 genotyping led to categorization into CYP2C19*1*1 (WT group, n = 123) and CYP2C19*2 or *3 (LOF group, n = 135). Baseline characteristics and operative data showed no significant differences between the two groups. The incidence of POAF after CABG was 42.2% in the LOF group, contrasting with 22.8% in the WT group (hazard risk [HR]: 2.061; 95% confidence interval [CI]: 1.347, 3.153; p = 0.0013). Adenosine diphosphate-stimulated platelet aggregation was notably higher in the LOF group compared to the WT group 5 days after CABG (30.4% ± 6.5% vs. 17.9% ± 4.1%, p < 0.001), remaining a similar higher level at hospital discharge (25.6% ± 6.1% vs. 12.2% ± 3.5%, p < 0.001). The presence of CYP2C19 LOF was linked to a higher incidence of POAF and relatively elevated platelet aggregation after CABG surgery under the same oral clopidogrel regimen.
Asunto(s)
Fibrilación Atrial , Clopidogrel , Puente de Arteria Coronaria , Citocromo P-450 CYP2C19 , Genotipo , Inhibidores de Agregación Plaquetaria , Agregación Plaquetaria , Complicaciones Posoperatorias , Humanos , Citocromo P-450 CYP2C19/genética , Citocromo P-450 CYP2C19/metabolismo , Fibrilación Atrial/etiología , Fibrilación Atrial/genética , Fibrilación Atrial/epidemiología , Masculino , Femenino , Anciano , Puente de Arteria Coronaria/efectos adversos , Persona de Mediana Edad , Clopidogrel/administración & dosificación , Clopidogrel/efectos adversos , Complicaciones Posoperatorias/etiología , Complicaciones Posoperatorias/epidemiología , Inhibidores de Agregación Plaquetaria/administración & dosificación , Inhibidores de Agregación Plaquetaria/efectos adversos , Agregación Plaquetaria/efectos de los fármacos , Incidencia , Aspirina/administración & dosificación , Aspirina/efectos adversosRESUMEN
AIMS: Dysregulated platelet aggregation is a fatal condition in many bacterial- and virus-induced diseases. However, classical antithrombotics cannot completely prevent immunothrombosis, due to the unaddressed mechanisms towards inflammation. Thus, targeting platelet hyperactivation together with inflammation might provide new treatment options in diseases, characterized by immunothrombosis, such as COVID-19 and sepsis. The aim of this study was to investigate the antiaggregatory effect and mode of action of 1.8-cineole, a monoterpene derived from the essential oil of eucalyptus leaves, known for its anti-inflammatory proprieties. MAIN METHODS: Platelet activity was monitored by measuring the expression and release of platelet activation markers, i.e., P-selectin, CD63 and CCL5, as well as platelet aggregation, upon treatment with 1.8-cineole and stimulation with several classical stimuli and bacteria. A kinase activity assay was used to elucidate the mode of action, followed by a detailed analysis of the involvement of the adenylyl-cyclase (AC)-cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) pathway by Western blot and ELISA. KEY FINDINGS: 1.8-cineole prevented the expression and release of platelet activation markers, as well as platelet aggregation, upon induction of aggregation with classical stimuli and immunological agonists. Mechanistically, 1.8- cineole influences the activation of the AC-cAMP-PKA pathway, leading to higher cAMP levels and vasodilator-stimulated phosphoprotein (VASP) phosphorylation. Finally, blocking the adenosine A2A receptor reversed the antithrombotic effect of 1.8-cineole. SIGNIFICANCE: Given the recognized anti-inflammatory attributes of 1.8-cineole, coupled with our findings, 1.8-cineole might emerge as a promising candidate for treating conditions marked by platelet activation and abnormal inflammation.
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AMP Cíclico , Eucaliptol , Activación Plaquetaria , Agregación Plaquetaria , Receptor de Adenosina A2A , Eucaliptol/farmacología , Receptor de Adenosina A2A/metabolismo , Activación Plaquetaria/efectos de los fármacos , Agregación Plaquetaria/efectos de los fármacos , Humanos , AMP Cíclico/metabolismo , Plaquetas/metabolismo , Plaquetas/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Selectina-P/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Inhibidores de Agregación Plaquetaria/farmacología , Antiinflamatorios/farmacología , COVID-19/metabolismoRESUMEN
Sweet potatoes are rich in cardioprotective phytochemicals with potential anti-platelet aggregation activity, although this benefit may vary among cultivars/genotypes. The phenolic profile [HPLC-ESI(-)-qTOF-MS2], cheminformatics (ADMET properties, affinity toward platelet proteins) and anti-PA activity of phenolic-rich hydroalcoholic extracts obtained from orange (OSP) and purple (PSP) sweet potato storage roots, was evaluated. The phenolic richness [Hydroxycinnamic acids> flavonoids> benzoic acids] was PSP > OSP. Their main chlorogenic acids could interact with platelet proteins (integrins/adhesins, kinases/metalloenzymes) but their bioavailability could be poor. Just OSP exhibited a dose-dependent anti-platelet aggregation activity [inductor (IC50, mg.ml-1): thrombin receptor activator peptide-6 (0.55) > Adenosine-5'-diphosphate (1.02) > collagen (1.56)] and reduced P-selectin expression (0.75-1.0 mg.ml-1) but not glycoprotein IIb/IIIa secretion. The explored anti-PA activity of OSP/PSP seems to be inversely related to their phenolic richness. The poor first-pass bioavailability of its chlorogenic acids (documented in silico) may represent a further obstacle for their anti-PA in vivo.
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Ipomoea batatas , Fenoles , Extractos Vegetales , Raíces de Plantas , Inhibidores de Agregación Plaquetaria , Agregación Plaquetaria , Ipomoea batatas/química , Fenoles/química , Agregación Plaquetaria/efectos de los fármacos , Extractos Vegetales/química , Extractos Vegetales/farmacología , Inhibidores de Agregación Plaquetaria/química , Inhibidores de Agregación Plaquetaria/farmacología , Raíces de Plantas/química , Humanos , Quimioinformática , Animales , Plaquetas/metabolismo , Plaquetas/efectos de los fármacosRESUMEN
INTRODUCTION: Previous literature suggests that sphingolipids may impact systemic coagulation and platelet aggregation, thus modulating the risks of thrombotic events. The goal of this investigation was to evaluate the role of serum sphingolipids on intrinsic platelet function to assess whether pharmacologic manipulation of sphingolipid metabolites would impact platelet aggregability. METHODS: C57BL/6J mice were injected with either normal saline, 1 mg/kg FTY720 (synthetic sphingosine-1-phosphate [S1P] receptor analog), or 5 mg/kg SLM6031434 (sphingosine kinase two inhibitor). Mice were sacrificed at 6 h and whole blood (WB) was collected for impedance aggregometry assessing platelet responsiveness to arachidonic acid or adenosine diphosphate. Ex vivo studies utilized WB or platelet-rich plasma that was pretreated with S1P, FTY720, amitriptyline, or d-sphingosine then analyzed by aggregability and flow cytometry for platelet and platelet-derived microvesicle characteristics. RESULTS: FTY720 and SLM6031434 pretreated induced similar arachidonic acid and adenosine diphosphate-mediated platelet aggregation as controls. Ex vivo WB and platelet-rich plasma treatment with S1P, FTY720, amitriptyline and d-sphingosine did not impact platelet aggregation. The percentages of CD41+, CD62P+ and CD41+/ceramide+, CD62P+/ceramide + platelets, and platelet-derived microvesicle were not significantly different between amitriptyline-treated and normal saline-treated cohorts. CONCLUSIONS: Sphingolipid modulating agents, such as FTY720, SLM6031434, S1P, amitriptyline, ceramide, and d-sphingosine do not appear to independently impact platelet aggregation in murine models.
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Plaquetas , Clorhidrato de Fingolimod , Ratones Endogámicos C57BL , Agregación Plaquetaria , Esfingolípidos , Esfingosina , Animales , Agregación Plaquetaria/efectos de los fármacos , Clorhidrato de Fingolimod/farmacología , Esfingosina/análogos & derivados , Esfingosina/sangre , Ratones , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Esfingolípidos/sangre , Esfingolípidos/metabolismo , Masculino , Lisofosfolípidos/farmacología , Lisofosfolípidos/sangre , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/antagonistas & inhibidores , Ácido Araquidónico/farmacología , Amitriptilina/farmacología , Adenosina Difosfato/farmacologíaRESUMEN
BACKGROUND: ADP and ATP are importantly involved in vascular and thrombotic homeostasis, via multiple receptor pathways. Blockade of ADP P2Y12 receptors inhibits platelet aggregation and represents an effective cardiovascular disease prevention strategy. AZD3366 (APT102), a long-acting recombinant form of an optimized CD39L3 human apyrase, has effectively reduced ATP, ADP, and platelet aggregation and provided tissue protection in preclinical models, features that could be very beneficial in treating patients with cardiovascular disease. METHODS AND RESULTS: We conducted this phase 1, first-in-human study of single ascending doses of intravenous AZD3366 or placebo, including doses added to dual antiplatelet therapy with ticagrelor and acetylsalicylic acid. The primary objective was safety and tolerability; secondary and exploratory objectives included pharmacokinetics, pharmacodynamics (measured as inhibition of platelet aggregation), adenosine diphosphatase (ADPase) activity, and ATP/ADP metabolism. In total, 104 participants were randomized. AZD3366 was generally well tolerated, with no major safety concerns observed. ADPase activity increased in a dose-dependent manner with a strong correlation to AZD3366 exposure. Inhibition of ADP-stimulated platelet aggregation was immediate, substantial, and durable. In addition, there was a prompt decrease in systemic ATP concentration and an increase in adenosine monophosphate concentrations, whereas ADP concentration appeared generally unaltered. At higher doses, there was a prolongation of capillary bleeding time without detectable changes in the ex vivo thromboelastometric parameters. CONCLUSIONS: AZD3366 was well tolerated in healthy participants and demonstrated substantial and durable inhibition of platelet aggregation after single dosing. Higher doses prolonged capillary bleeding time without detectable changes in ex vivo thromboelastometric parameters. REGISTRATION: URL: https://www.clinicaltrials.gov; Unique Identifier: NCT04588727.
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Apirasa , Aspirina , Inhibidores de Agregación Plaquetaria , Agregación Plaquetaria , Ticagrelor , Humanos , Masculino , Ticagrelor/farmacocinética , Ticagrelor/administración & dosificación , Ticagrelor/efectos adversos , Femenino , Apirasa/metabolismo , Apirasa/administración & dosificación , Agregación Plaquetaria/efectos de los fármacos , Aspirina/administración & dosificación , Aspirina/farmacocinética , Aspirina/efectos adversos , Inhibidores de Agregación Plaquetaria/farmacocinética , Inhibidores de Agregación Plaquetaria/administración & dosificación , Inhibidores de Agregación Plaquetaria/efectos adversos , Persona de Mediana Edad , Adulto , Método Doble Ciego , Terapia Antiplaquetaria Doble , Quimioterapia Combinada , Adulto Joven , Adenosina Difosfato , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Relación Dosis-Respuesta a Droga , Resultado del Tratamiento , Proteínas Recombinantes/administración & dosificación , Proteínas Recombinantes/farmacocinética , Antagonistas del Receptor Purinérgico P2Y/farmacocinética , Antagonistas del Receptor Purinérgico P2Y/administración & dosificación , Antagonistas del Receptor Purinérgico P2Y/efectos adversos , Antagonistas del Receptor Purinérgico P2Y/farmacologíaRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Green tea (Camellia sinensis) is a popular beverage consumed all over the world due to its health benefits. Many of these beneficial effects of green tea are attributed to polyphenols, particularly catechins. AIM OF THE STUDY: The present study focuses on underlying anti-platelet aggregation, anti-thrombotic, and anti-lipidemic molecular mechanisms of green tea in South Indian smokers. MATERIALS AND METHODS: We selected 120 South Indian male volunteers for this study to collect the blood and categorised them into four groups; control group individuals (Controls), smokers, healthy control individuals consuming green tea, and smokers consuming green tea. Smokers group subjects have been smoking an average 16-18 cigarettes per day for the last 7 years or more. The subjects (green tea consumed groups) consumed 100 mL of green tea each time, thrice a day for a one-year period. RESULTS: LC-MS analysis revealed the presence of multiple phytocompounds along with catechins in green tea extract. Increased plasma lipid peroxidation (LPO), protein carbonyls, cholesterol, triglycerides, and LDL-cholesterol with decreased HDL-cholesterol levels were observed in smokers compared to the control group and the consumption of green tea showed beneficial effect. Furthermore, docking studies revealed that natural compounds of green tea had high binding capacity with 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG-CoA) when compared to their positive controls, whereas (-) epigallocatechin-3-gallate (EGCG) and (-) epicatechin-gallate (ECG) had high binding capacity with sterol regulatory element-binding transcription factor 1 (SREBP1c). Further, our ex vivo studies showed that green tea extract (GTE) significantly inhibited platelet aggregation and increased thrombolytic activity in a dose dependent manner. CONCLUSION: In conclusion, in smokers, catechins synergistically lowered oxidative stress, platelet aggregation and modified the aberrant lipid profile. Furthermore, molecular docking studies supported green tea catechins' antihyperlipidemic efficacy through strong inhibitory activity on HMG-CoA reductase and SREBP1c. The mitigating effects of green tea on cardiovascular disease risk factors in smokers that have been reported can be attributed majorly to catechins or to their synergistic effects.
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Camellia sinensis , Simulación del Acoplamiento Molecular , Extractos Vegetales , Té , Humanos , Masculino , India , Adulto , Camellia sinensis/química , Té/química , Extractos Vegetales/farmacología , Extractos Vegetales/uso terapéutico , Extractos Vegetales/química , Fumar , Persona de Mediana Edad , Aterosclerosis/tratamiento farmacológico , Aterosclerosis/prevención & control , Fumadores , Catequina/farmacología , Catequina/análogos & derivados , Lípidos/sangre , Antioxidantes/farmacología , Inhibidores de Agregación Plaquetaria/farmacología , Agregación Plaquetaria/efectos de los fármacosRESUMEN
BACKGROUND: Plaque erosion, a type of coronary atherothrombosis, involves superficial injury to smooth muscle cell (SMC)-rich plaques. Elevated levels of coagulation factor VIII (FVIII) correlate with an increased ischemic heart disease risk. FVIII may contribute to thrombus formation on eroded plaques. AIMS: We aimed to elucidate the role of elevated FVIII in arterial thrombus formation within SMC-rich neointima in rabbits. METHODS AND RESULTS: We assessed the effect of recombinant human FVIII (rFVIII) on blood coagulation in vitro and platelet aggregation ex vivo. An SMC-rich neointima was induced through balloon injury to the unilateral femoral artery. Three weeks after the first balloon injury, superficial erosive injury and thrombus formation were initiated with a second balloon injury of the bilateral femoral arteries 45 min after the administration of rFVIII (100 IU/kg) or saline. The thrombus area and contents were histologically measured 15 min after the second balloon injury. rFVIII administration reduced the activated partial thromboplastin time and augmented botrocetin-induced, but not collagen- or adenosine 5'-diphosphate-induced, platelet aggregation. While rFVIII did not influence platelet-thrombus formation in normal intima, it increased thrombus formation on SMC-rich neointima post-superficial erosive injury. Enhanced immunopositivity for glycoprotein IIb/IIIa and fibrin was observed in rFVIII-administered SMC-rich neointima. Neutrophil count in the arterial thrombus on the SMC-rich neointima correlated positively with thrombus size in the control group, unlike the rFVIII group. CONCLUSIONS: Increased FVIII contributes to thrombus propagation within erosive SMC-rich neointima, highlighting FVIII's potential role in plaque erosion-related atherothrombosis.
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Factor VIII , Miocitos del Músculo Liso , Neointima , Trombosis , Conejos , Animales , Neointima/patología , Neointima/sangre , Trombosis/sangre , Trombosis/patología , Masculino , Miocitos del Músculo Liso/patología , Miocitos del Músculo Liso/efectos de los fármacos , Túnica Íntima/patología , Túnica Íntima/efectos de los fármacos , Humanos , Agregación Plaquetaria/efectos de los fármacos , Arteria Femoral/patología , Arteria Femoral/lesionesRESUMEN
The toxicity of inhaled particulate air pollution perseveres even at lower concentrations than those of the existing air quality limit. Therefore, the identification of safe and effective measures against pollutant particles-induced vascular toxicity is warranted. Carnosol is a bioactive phenolic diterpene found in rosemary herb, with anti-inflammatory and antioxidant actions. However, its possible protective effect on the thrombotic and vascular injury induced by diesel exhaust particles (DEP) has not been studied before. We assessed here the potential alleviating effect of carnosol (20 mg/kg) administered intraperitoneally 1 h before intratracheal (i.t.) instillation of DEP (20 µg/mouse). Twenty-four hours after the administration of DEP, various parameters were assessed. Carnosol administration prevented the increase in the plasma concentrations of C-reactive protein, fibrinogen, and tissue factor induced by DEP exposure. Carnosol inhibited DEP-induced prothrombotic effects in pial microvessels in vivo and platelet aggregation in vitro. The shortening of activated partial thromboplastin time and prothrombin time induced by DEP was abated by carnosol administration. Carnosol inhibited the increase in pro-inflammatory cytokines (interleukin-6 and tumor necrosis factor α) and adhesion molecules (intercellular adhesion molecule-1, vascular cell adhesion molecule-1, E-selectin, and P-selectin) in aortic tissue. Moreover, it averted the effects of DEP-induced increase of thiobarbituric acid reactive substances, depletion of antioxidants and DNA damage in the aortic tissue. Likewise, carnosol prevented the decrease in the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) caused by DEP. We conclude that carnosol alleviates DEP-induced thrombogenicity and vascular inflammation, oxidative damage, and DNA injury through Nrf2 and HO-1 activation.
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Abietanos , Trombosis , Emisiones de Vehículos , Animales , Abietanos/farmacología , Ratones , Masculino , Emisiones de Vehículos/toxicidad , Trombosis/prevención & control , Trombosis/tratamiento farmacológico , Trombosis/inducido químicamente , Pulmón/efectos de los fármacos , Pulmón/patología , Pulmón/metabolismo , Lesiones del Sistema Vascular/tratamiento farmacológico , Antioxidantes/farmacología , Material Particulado/toxicidad , Material Particulado/efectos adversos , Factor 2 Relacionado con NF-E2/metabolismo , Contaminantes Atmosféricos/toxicidad , Estrés Oxidativo/efectos de los fármacos , Agregación Plaquetaria/efectos de los fármacosRESUMEN
Platelet functional activity was assessed in healthy volunteers (HV, n=92), patients with stable angina pectoris (SA, n=42) and acute coronary syndrome (ACS, n=73), treated with acetylsalicylic acid (ASA) + clopidogrel and ASA + ticagrelor, respectively. In all HV and patients we have compared parameters of platelet aggregation (maximum light transmission and velocity, Tmax and Vmax) and parameters, characterizing exposure of platelet activation markers, evaluated by flow cytometry. HV platelets were activated by 10 µM, 1 µM TRAP, and 20 µM, 5 µM, 2.5 µM ADP; patient platelets were activated by 10 µM TRAP and by 20 µM and 5 µM ADP. Strong and significant correlations between the aggregation and flow cytometry parameters (the r correlation coefficient from 0.4 up to >0.6) most frequently were registered in HV platelet during activation by 1 µM TRAP and in SA patients during platelet activation by 20 µM and 5 µM ADP. However, in many other cases these correlations were rather weak (r < 0.3) and sometimes statistically insignificant. In HV the differences in PAC-1 binding parameters between platelets activated by 10 µM TRAP (the strongest agonist) and all ADP concentrations were negligible (≤ 10%), while CD62P binding (at all ADP concentrations) and LTA parameters for (5 µM and 2.5 µM ADP) were significantly lower (by 40-60%). Antiplatelet therapy in patients decreased all parameters as compared to HV, but to varying extents. For 10 µM TRAP the MFI index for PAC-1 binding (40-50% decrease) and for both ADP concentrations the Tmax values (60-85% decrease) appeared to be the most sensitive in comparison with the other parameters that decreased to a lesser extent. The data obtained indicate a possibility of inconsistency between different LTA and flow cytometry parameters in assessing platelet activity and efficacy of antiplatelet drugs.