Long-term pharmacodynamic and clinical effects of twice- versus once-daily low-dose aspirin in essential thrombocythemia: The ARES trial.

Patients with essential thrombocythemia (ET) are treated with once-daily low-dose aspirin to prevent thrombosis, but their accelerated platelet turnover shortens the antiplatelet effect. The short-term Aspirin Regimens in EsSential Thrombocythemia trial showed that twice-daily aspirin dosing restores persistent platelet thromboxane (TX) inhibition. However, the long-term pharmacodynamic efficacy, safety and tolerability of twice-daily aspirin remain untested. We performed a multicenter, randomized, open-label, blinded-endpoint, phase-2 trial in which 242 patients with ET were randomized to 100 mg aspirin twice- or once-daily and followed for 20 months. The primary endpoint was the persistence of low serum TXB2, a surrogate biomarker of antithrombotic efficacy. Secondary endpoints were major and clinically relevant non-major bleedings, serious vascular events, symptom burden assessed by validated questionnaires, and in vivo platelet activation. Serum TXB2 was consistently lower in the twice-daily versus once-daily regimen on 10 study visits over 20 months: median 3.9 ng/mL versus 19.2 ng/mL, respectively; p < .001; 80% median reduction; 95% CI, 74%-85%. No major bleeding occurred. Clinically relevant non-major bleedings were non-significantly higher (6.6% vs. 1.7%), and major thromboses lower (0.8% vs. 2.5%) in the twice-daily versus once-daily group. Patients on the twice-daily regimen had significantly lower frequencies of disease-specific symptoms and severe hand and foot microvascular pain. Upper gastrointestinal pain was comparable in the two arms. In vivo platelet activation was significantly reduced by the twice-daily regimen. In patients with ET, twice-daily was persistently superior to once-daily low-dose aspirin in suppressing thromboxane biosynthesis and reducing symptom burden, with no detectable excess of bleeding and gastrointestinal discomfort.

Reference interval establishment and correlation research of urine thromboxane metabolites: Unveiling new possibilities in platelet activation.

BACKGROUND: Thromboxane metabolites could indirectly reflect platelet activation, among which 11-dehydro-thromboxane B2 (11dhTxB2) and 11-dehydro-2, 3-dinor thromboxane B2 (11dh23dinorTxB2) are two stable metabolites that are abundant in urine, and both are closely related to disease progression and drug use. However, most clinical application studies have focused on the single indicator of 11dhTxB2. We propose an LC-MS/MS method suitable for routine clinical screening with simultaneous determination of both metabolites and conduct preliminary studies in different populations. METHODS AND RESULTS: The thromboxane metabolites were extracted by liquid-liquid extraction and determined by LC-MS/MS. Reference intervals (RI) were established in 333 healthy adults and validated in 25 patients with coronary atherosclerosis (CA). This LC-MS/MS method was over a wide quantitative range (0.1-10 µmol/L), the imprecision and accuracy were 5.2 %-11 % and 89.3 %-106.5 %, and was suitable for clinical routine quantitative screening. The 95th percentile RI of unire 11dhTxB2 was 1220 (95 % CI: 1048, 1376) pg mg Cr -1, for 11dh23dinorTxB2, RI was 908 (95 % CI: 821, 1102) pg mg Cr -1. For the first time, we found a significant correlation between 11dhTxB2 and 11dh23dinorTxB2 in both healthy adults (r = 0.67, P < 0.001) and CA patients (r = 0.77, P < 0.001). CONCLUSION: The establishment of RI provides a reference for diseases related to platelet activation and the use of drugs, and the first discovery of the correlation between 11dhTxB2 and 11dh23dinorTxB2 in urine provides a new possibilitie for the diagnostic and prognostic of cardiovascular diseases.

Plasma Instead of Serum Avoids Critical Confounding of Clinical Metabolomics Studies by Platelets.

Metabolomics is an emerging and powerful bioanalytical method supporting clinical investigations. Serum and plasma are commonly used without rational prioritization. Serum is collected after blood coagulation, a complex biochemical process involving active platelet metabolism. This may affect the metabolome and increase the variance, as platelet counts and function may vary substantially in individuals. A multiomics approach systematically investigating the suitability of serum and plasma for clinical studies demonstrated that metabolites correlated well (n = 461, R2 = 0.991), whereas lipid mediators (n = 83, R2 = 0.906) and proteins (n = 322, R2 = 0.860) differed substantially between specimen. Independently, analysis of platelet releasates identified most biomolecules significantly enriched in serum compared to plasma. A prospective, randomized, controlled parallel group metabolomics trial with acetylsalicylic acid administered for 7 days demonstrated that the apparent drug effects significantly differ depending on the analyzed specimen. Only serum analyses of healthy individuals suggested a significant downregulation of TXB2 and 12-HETE, which were specifically formed during coagulation in vitro. Plasma analyses reliably identified acetylsalicylic acid effects on metabolites and lipids occurring in vivo such as an increase in serotonin, 15-deoxy-PGJ2 and sphingosine-1-phosphate and a decrease in polyunsaturated fatty acids. The present data suggest that plasma should be preferred above serum for clinical metabolomics studies as the serum metabolome may be substantially confounded by platelets.

Adjustment for Renal Function Improves the Prognostic Performance of Urinary Thromboxane Metabolites.

BACKGROUND: Systemic thromboxane A2 generation, assessed by quantifying the concentration of stable thromboxane B2 metabolites (TXB2-M) in the urine adjusted for urinary creatinine, is strongly associated with mortality risk. We sought to define optimal TXB2-M cutpoints for aspirin users and nonusers and determine if adjusting TXB2-M for estimated glomerular filtration rate (eGFR) in addition to urinary creatinine improved mortality risk assessment. METHODS: Urinary TXB2-M were measured by competitive ELISA in 1363 aspirin users and 1681 nonusers participating in the Framingham Heart Study. Cutpoints were determined for TXB2-M and TXB2-M/eGFR using log-rank statistics and used to assess mortality risk by Cox proportional hazard modeling and restricted mean survival time. Multivariable models were compared using the Akaike Information Criterion (AIC). A cohort of 105 aspirin users with heart failure was used for external validation. RESULTS: Optimized cutpoints of TXB2-M were 1291 and 5609 pg/mg creatinine and of TXB2-M/eGFR were 16.6 and 62.1 filtered prostanoid units (defined as pg·min/creatinine·mL·1.73 m2), for aspirin users and nonusers, respectively. TXB2-M/eGFR cutpoints provided more robust all-cause mortality risk discrimination than TXB2-M cutpoints, with a larger unadjusted hazard ratio (2.88 vs 2.16, AIC P < 0.0001) and greater differences in restricted mean survival time between exposure groups (1.46 vs 1.10 years), findings that were confirmed in the external validation cohort of aspirin users. TXB2-M/eGFR cutpoints also provided better cardiovascular/stroke mortality risk discrimination than TXB2-M cutpoints (unadjusted hazard ratio 3.31 vs 2.13, AIC P < 0.0001). CONCLUSION: Adjustment for eGFR strengthens the association of urinary TXB2-M with long-term mortality risk irrespective of aspirin use.

Thromboxane biosynthesis and future events in diabetes: the ASCEND trial.

BACKGROUND AND AIMS: Thromboxane (TX) A2, released by activated platelets, plays an important role in atherothrombosis. Urinary 11-dehydro-TXB2 (U-TXM), a stable metabolite reflecting the whole-body TXA2 biosynthesis, is reduced by ∼70% by daily low-dose aspirin. The U-TXM represents a non-invasive biomarker of in vivo platelet activation and is enhanced in patients with diabetes. This study assessed whether U-TXM is associated with the risk of future serious vascular events or revascularizations (SVE-R), major bleeding, or cancer in patients with diabetes. METHODS: The U-TXM was measured pre-randomization to aspirin or placebo in 5948 people with type 1 or 2 diabetes and no cardiovascular disease, in the ASCEND trial. Associations between log U-TXM and SVE-R (n = 618), major bleed (n = 206), and cancer (n = 700) during 6.6 years of follow-up were investigated by Cox regression; comparisons of these associations with the effects of randomization to aspirin were made. RESULTS: Higher U-TXM was associated with older age, female sex, current smoking, type 2 diabetes, higher body size, urinary albumin/creatinine ratio of ≥3 mg/mmol, and higher estimated glomerular filtration rate. After adjustment for these, U-TXM was marginally statistically significantly associated with SVE-R and major bleed but not cancer [hazard ratios per 1 SD higher log U-TXM (95% confidence interval): 1.09 (1.00-1.18), 1.16 (1.01-1.34), and 1.06 (0.98-1.14)]. The hazard ratio was similar to that implied by the clinical effects of randomization to aspirin for SVE-R but not for major bleed. CONCLUSIONS: The U-TXM was log-linearly independently associated with SVE-R in diabetes. This is consistent with the involvement of platelet TXA2 in diabetic atherothrombosis.

Dietary inflammation index association with serum levels of nitric oxide, prostacyclin, and thromboxane B2 among prinzmetal angina patients and healthy persons.

BACKGROUND AND AIM: This study aimed to assess the association between dietary inflammation index with serum Nitric oxide, Prostacyclin, and Thromboxane B2 among Prinzmetal angina patients and healthy persons. METHODS AND RESULTS: This case-control study was conducted among 120 Prinzmetal angina patients and 120 healthy persons referred to the Ardabil Imam Khomeini Hospital between 2021 and 2022. Blood samples were gained from all study participants for measurement of serum Nitric oxide, Prostacyclin, and Thromboxane B2. The serum Nitric oxide in patients who had higher DII was less than in patients with less dietary inflammation index (ß = -0.75 p = 0.02). The serum Prostacyclin level in patients with greater dietary inflammation index was 0.68 ng/ml less than in patients with less dietary inflammation index (ß = -0.68 p = 0.04). The level of serum Thromboxane B2 had a positive association with dietary inflammation index (ß = 0.81 p = 0.04). CONCLUSION: In Prinzmetal angina patients, more dietary inflammation index can increase the serum Thromboxane B2 and decrease the serum Nitric oxide and Prostacyclin. More clinical trial study is needed to confirm these results.

La combinación estradiol-acetano de noretisterona sobre los niveles del tromboxano B2 en mujeres posmenopáusicas / Estradiol norethisterone acetate combination on thromboxane B2 levels in postmenopausal women

Investigar los niveles plasmáticos de tromboxano B2 en las mujeres posmenopáusica antes y después de la administración de estradiol-acetato de noretisterona y compararlos con los niveles de mujeres premenopáusicas. Se seleccionaron 22 mujeres premenopáusicas. Las mujeres posmenopáusicas fueron tratadas con 2mg de estradiol y 1 mg de acetato de noretisterona diario por 6 meses, y fue en la fase folicular del ciclo artificial de la terapia hormonal de reemplazo al momento de la toma de la muestra. El tromboxano B2 (producto final del tromboxano A2) se analizó usando una prueba de radioinmunoensayo. Los niveles plasmáticos de tromboxano B2 fueron los siguientes: 198ñ83,2 pg/mL en las mujeres premenopáusicas 1 215ñ153,8 pg/mL en las mujeres posmenopáusicas antes del tratamientoy 711ñ99,1 pg/mL en las posmenopáusicas después del tratamiento. El tratamiento por 6 meses con estradiol-acetato de noretisterona produjo una disminución estadísticamente significativa en el tromboxano B2 en mujeres posmenopáusicas (p<0.01). La diferencia fue estadísticamente significativa cuando se comparó a las mujeres posmenopáusicas tanto antes como después del tratamiento. Existe un aumento en la producción de tromboxano B2 en mujeres posmenopáusicas, lo cual probablemente refleja el incremeto del estrés visto en la posmenopáusia. Después de 6 meses de tratamiento diario con la combinación estradiol-acetato de noretisterona disminuyen los niveles de tromboxano B2 plamático.

Tromboxano B2 y factor de crecimiento derivado de las plaquetas en la trombocitemia esencial tratada con anagrelide / Thromboxane B2 and platelet derived growth factor in essential thrombocythemia treated with anagrelide

Presentamos los resultados obtenidos en 17 pacientes con trombocitemia esencial antes del tratamiento con anagrelide y durante la remisión hematológica y clínica inducida por esta droga. Diez pacientes tenían manifestaciones clínicas relacionadas con la trombocitemia, en 8 de ellos al momento de iniciar el tratamiento. Se midieron los niveles plasmáticos del TXB2 y del PDGF por técnica de ELISA. Antes del tratamiento los valores plasmáticos del PDGF corregidos por el recuento de plaquetas eran significativamente menores que en los controles (p = 0.02), y no difirieron de los resultados obtenidos durante la remisión. En cambio, los valores plasmáticos del TXB2, corregidos de acuerdo al recuento de plaquetas, estaban más elevados que en los controles (p = 0.04), y disminuyeron significativamente durante la remisión (p = 0.04), aunque todavía estaban por sobre los valores normales (p = 0.008). Los síntomas desaparecieron en todos los pacientes cuando el recuento de plaquetas se normalizó. Estos resultados muestran que los pacientes con trombocitemia esencial tratados con anagrelide y en remisión hematológica y clínica tienen tendencia a normalizar los niveles del TXB2.

Variations in white blood count, thromboxane B2 levels and hematocrit in chronic venous hypertension

Objective: To analyze variations in leukocyte count and thromboxane B2 production in the femoral vein of patients with chronic venous hypertension (CVH). Design: Prospective clinical study, controlled, non randomized and open. Location: Hospital das Cl[inicas, Faculdade de Medicina da Universidade de Sao Paulo, referral center, university hospital. Participants: 15 patients with recurring stasis ulcer were analyzed, selected randomly from the venous diseases outpatient center, and 4 without lower limb venous alterations were also analyzed. Intervention: Blood samples from the femoral and brachial veins were drawn following supine and 45º reverse Trendelenburg. Main outcomes measures: Direct leukocyte count and analysis of the thomboxane B2 with enzyme linked immunosorbent assay test. Results: After 30 minutes in reverse Trendelenburg, patients with CVH showed a leukocyte count reduced by +27 per cent (p=0.02) and thromboxane B2 levels increased by +158 per cent (p=0.02). Conclusions: We suggest that future studies of medications for stasis ulcers include their effects on leukocyte entrapment and thromboxane B2 production in the lower limb venous system.

The effects of melatonin in human platelets

La melatonina pineal ha sido implicada en el control de varios procesos fisiológicos, como la ritmicidad circadiana y la reproducción estacional en mamíferos. Sin embargo la función de la melatonina en el hombre permanece aún indefinida. La melatonina actúa en el SNC para modificar distintos ritmos biológicos; además se han descrito varios efectos periféricos de la hormona pineal entre ellos su actividad plaquetaria. En este trabajo se analizan los datos experimentales que indican que las plaquetas son células sensibles a la melatonina de posible utilidad clínica para elucidar la función de la melatonina y la glándula pineal en el hombre. La melatonina inhibe distintos procesos fisiológicos plaquetarios, como la agregación plaquetaria, la producción de tromboxanos y la liberación de ATP y serotonina, ambos índices de secreción plaquetaria. En general, el efecto de la melatonina es mayor en horas de la tarde y comienzo de la noche, coincidiendo con los datos in vivo que indican un mayor efecto de la melatonina administrada sistématicamente. El máximo en actividad in vitro de la melatonina en plaquetas precede al máximo en niveles circulantes de la hormona, indicando la existencia de disociación entre niveles circulantes y sensibilidad tisular a la hormona. En estudios de unión de radiologando in vitro se detectaron sitios aceptores para la 3H-melatonina en membranas de plaquetas humanas, con Kd de rango nanomolar. Estos resultados sugieren en conjunto que las plaquetas pueden utilizar-se clínicamente como "ventanas" circulantes de los procesos modificados por la melatonina en el SNC