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.

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.

Utility of VerifyNow to assess the immediate pharmacodynamic response of chewed and swallowed aspirin: comparison with aggregometry.

VerifyNow (VN) test is a less laborious method to assess pharmacodynamics (PD) compared to light transmittance aggregometry (LTA). VN assay has not been used to study the immediate PD effects of acetylsalicylic acid (ASA). Ten healthy volunteers were randomly assigned to a single 162 or 650 mg dose of chewed and swallowed ASA. Pharmacodynamic and pharmacokinetic measurements were performed at baseline and serially up to 60 min after ASA administration. Onset by VN was 20 ± 7 min with 162 mg and 13 ± 7 min with 650 mg ASA (p = .07). Onset by 1 mM AA-induced PA was 13 ± 12 min with 162 mg and 7 ± 3 min with 650 mg ASA (p=NS). VN correlated with AA-induced PA (r = 0.80, p < .001) and serum TxB2 levels (r = 0.76, p < .001). 95% inhibition of serum TxB2 was achieved at 38 ± 22 min and 22 ± 8 min with the 162 and 650 mg ASA, respectively (p = .08). The onset and extent of the antiplatelet effect of 650 mg ASA is numerically faster and greater than the 162 mg dose. VN identifies the onset, extent, and dose response to ASA therapy. The ease of using VN should facilitate multicenter PD investigations of ASA.

Aspirin (acetylsalicylic acid) is an important drug widely used to prevent adverse ischemic events in patients with cardiovascular disease. Platelet aggregation and thromboxane B₂ levels in blood samples by complex laboratory methods are used to assess platelet response to aspirin. VerifyNow assay is a simple laboratory test that has not been used to assess the immediate effect of aspirin. In this study, conducted in 10 healthy volunteers, we compared the immediate platelet response to aspirin by serially assessing platelet aggregation by aggregometry and VerifyNow assay, and thromboxane B₂ levels. We also measured plasma levels of acetylsalicylic acid and salicylic acid. Our study demonstrated that the VerifyNow Aspirin test identifies the onset, extent, and dose-response to aspirin therapy. The ease of using the VerifyNow test should facilitate multicenter pharmacodynamic investigations of aspirin.

[Clinical study on the effect of acupuncture on pregnancy outcome in patients with prethrombotic infertility after repeated implantation failure].

OBJECTIVE: To observe the effect of acupuncture combined with medication on the pregnancy outcome of kidney deficiency and blood stasis type recurrent implantation failure (RIF) in prethrombotic infertility patients. METHODS: A total of 70 RIF patients of kidney deficiency and blood stasis type who were to undergo freeze-thaw embryo transplantation were randomly divided into control and treatment groups, with 35 cases in each group. Patients in the control group were given the basic treatment of artificial periodic freeze-thaw embryo transfer and oral aspirin enteric-coated tablet, 25 mg each time, twice a day, for 3 consecutive menstrual cycles. Patients in the treatment group were treated with acupuncture on the basis of the control group, 30 min each time, 3 times a week, for 3 consecutive menstrual cycles. The clinical pregnancy rate, embryo implantation rate and live birth rate of the two groups were compared. Before treatment and 1 day before transplantation, the scores of kidney deficiency and blood stasis symptom were compared. The blood flow pulse index (PI) and resistance index (RI) of the uterus spiral artery were detected by Doppler ultrasound before treatment and 1 day before transplantation. The endometrial thickness was detected 1 day before transplantation. The contents of plasma D-dimer, serum homocysteine (Hcy) and serum thromboxane B2 (TXB2) were detected. RESULTS: The clinical pregnancy rate, embryo implantation rate and live birth rate in the treatment group were higher than those in the control group (P<0.05). After treatment, the scores of kidney deficiency and blood stasis symptom, the levels of plasma D-dimer, serum Hcy and TXB2, the PI and RI value in both groups were decreased (P<0.05) compared with those before treatment, and the indexes in the treatment group were decreased (P<0.05) more than those in the control group. There was no significant difference in endometrial thickness between the two groups. In the course of treatments, 7 patients in the control group underwent gastric distension pain, poor appetite, constipation, nausea and other gastrointestinal reactions, while only 2 patients in the treatment group had the above discomfort reactions. CONCLUSION: On the basis of medication, acupuncture can reduce the serum TXB2 content in RIF patients in prethrombotic state, improve vascular endothelial function, enhance endometrial tolerance, alleviate the symptoms of kidney deficiency and blood stasis, reduce drug adverse reactions, and ultimately improve the pregnancy outcome and increase the rate of embryo implantation.

Agomelatine inhibits platelet aggregation through melatonin receptor-dependent and independent mechanisms.

AIMS: Melatonin is known to inhibit platelet aggregation induced by arachidonic acid (AA). In the present study we investigated whether agomelatine (Ago), an antidepressant with agonist activity at melatonin receptor 1 (MT1) and MT2 could reduce platelets aggregation and adhesion. MAIN METHODS: Human platelets from healthy donors were used to test the in vitro effects of Ago in the presence of different platelet activators. We performed aggregation and adhesion assays, thromboxane B2 (TxB2), cAMP and cGMP measurements, intra-platelet calcium registration and flow cytometry assays. KEY FINDINGS: Our data revealed that different concentrations of Ago reduced AA- and collagen-induced human platelet aggregation in vitro. Ago also reduced AA-induced increase in thromboxane B2 (TxB2) production, intracellular calcium levels and P-selectin expression at plasma membrane. The effects of Ago in AA-activated platelets were likely dependent on MT1 as they were blocked by luzindole (a MT1/MT2 antagonist) and mimicked by the MT1 agonist UCM871 in a luzindole-sensitive manner. The MT2 agonist UCM924 was also able to inhibit platelet aggregation, but this response was not affected by luzindole. On the other hand, although UCM871 and UCM924 reduced collagen-induced platelet aggregation and adhesion, inhibition of collagen-induced platelet aggregation by Ago was not mediated by melatonin receptors because it was not affected by luzindole. SIGNIFICANCE: The present data show that Ago suppresses human platelet aggregation and suggest that this antidepressant may have the potential to prevent atherothrombotic ischemic events by reducing thrombus formation and vessel occlusion.

Establishment of 11-dehydro-thromboxane B2 time-resolved immunoassay and application in membranous nephropathy.

BACKGROUND: 11-Dehydro-thromboxane B2 (11-dehydro-TXB2) is the final stable metabolite of thromboxane A2 (TXA2) and is involved in thrombus formation. Patients with membranous nephropathy (MN) are prone to thromboembolism events. METHODS: Time-resolved fluorescence immunoassay (TRFIA) for 11-dehydro-TXB2 was established by indirect competitive method. The coated 11-dehydro-TXB2-BSA conjugate was used to bind the 11-dehydro-TXB2 antibody competitively to the 11-dehydro-TXB2 antigen in the samples, followed by Eu3+-labeled goat anti-mouse IgG antibody, to detect 11-dehydro-TXB2. This study measured 11-dehydro-TXB2 concentrations in serum samples from healthy individuals and patients with MN. RESULTS: The linear range of TRFIA was 16.38-2000 pg/mL, the sensitivity was 4.70 pg/mL, the average coefficients of variation from intra-assay and inter-assay were 3.50% and 4.95%, respectively, and the recovery was 99.38%. The serum level of 11-dehydro-TXB2 in patients with MN was significantly higher than that in healthy subjects (P < 0.05). The serum 11-dehydro-TXB2 concentration detected by TRFIA was highly consistent with that by ELISA (ρ = 0.900). DISCUSSION: This study successfully established a new highly sensitive method for the detection of 11-dehydro-TXB2 in serum. 11-Dehydro-TXB2 has great potential in evaluating the risk of thromboembolic events in patients with MN and is expected to be applied to other thromboembolic-related diseases.

Effect of low-dose aspirin on urinary 11-dehydro-thromboxane B2 in the ASCEND (A Study of Cardiovascular Events iN Diabetes) randomized controlled trial.

BACKGROUND: Aspirin is widely used for cardioprotection with its antiplatelet effects due to the blocking of thromboxane A2 production. However, it has been suggested that platelet abnormalities in those with diabetes prevent adequate suppression with once daily aspirin. METHODS: In the ASCEND randomized double-blind trial of aspirin 100 mg once daily versus placebo in participants with diabetes but no history of cardiovascular disease, suppression was assessed by measuring 11-dehydro-thromboxane B2 excretion in urine (U-TXM) in a randomly selected sample of 152 participants (76 aspirin arm, 74 placebo arm), plus 198 (93 aspirin arm, 105 placebo arm) adherent to study drugs and selected to maximize the numbers ingesting their last tablet 12-24 h before urine sampling. U-TXM was assayed using a competitive ELISA assay in samples mailed a mean of 2 years after randomization, with time since taking last aspirin/placebo tablet recorded at the time of sample provision. Effective suppression (U-TXM < 1500 pg/mg creatinine) and percentage reductions in U-TXM by aspirin allocation were compared. RESULTS: In the random sample, U-TXM was 71% (95% CI 64-76%) lower among aspirin vs placebo-allocated participants. Among adherent participants in the aspirin arm, U-TXM was 72% (95% CI 69-75%) lower than in the placebo arm and 77% achieved effective suppression overall. Suppression was similar among those who ingested their last tablet more than 12 h before urine sampling with levels in the aspirin arm 72% (95% CI 67-77%) lower than in the placebo arm and 70% achieving effective suppression. CONCLUSIONS: Daily aspirin significantly reduces U-TXM in participants with diabetes, including at 12-24 h after ingestion. TRIAL REGISTRATION: ISRCTN ISRCTN60635500. Registered on 1 Sept 2005; ClinicalTrials.gov NCT00135226. Registered on 24 Aug 2005.

Myeloproliferative Neoplasms and Aspirin: Does Increased Platelet Turnover Matter?

Objective: Platelet aggregation tests and the analysis of thromboxane A2 metabolites [serum thromboxane B2 (TXB2) and urine 11-dehydro TXB2] are used to evaluate the efficacy of aspirin. In myeloproliferative neoplasms (MPNs), the immature platelet fraction (IPF) rises due to enhanced platelet turnover, and this has been thought to reduce the efficacy of aspirin. This phenomenon is overcome by the recommendation of aspirin intake in divided doses. We aimed to evaluate aspirin efficacy in patients who were receiving aspirin treatment of 100 mg/day. Materials and Methods: Thirty-eight MPN patients and 30 control patients (non-MPN patients who received a single daily dose of aspirin at 100 mg for nonhematological conditions) were enrolled. IPF, serum TXB2, and urine 11-dehydro TXB2 levels were measured and aggregation tests with arachidonic acid and adenosine diphosphate were performed by light transmission aggregometry (LTA). Results: Mean IPF and TXB2 levels were higher in the MPN group (p=0.008 and p=0.003, respectively). IPF levels were lower in patients on cytoreductive therapy in the MPN group (p=0.001), but these values were similar between patients on hydroxyurea and the non-MPN group (p=0.72). TXB2 levels did not differ according to hydroxyurea treatment status but were higher in the MPN group compared to non-MPN patients (23.63 ng/mL and 19.78 ng/mL, respectively; p=0.04). TXB2 values were higher in patients with essential thrombocythemia and a history of thrombotic events (p=0.031). No difference was observed in LTA between the MPN and non-MPN patient groups (p=0.513). Conclusion: Higher levels of IPF and TXB2 in the MPN patient group indicated platelets that could not be inhibited by aspirin. It was observed that patients under cytoreductive therapy had lower IPF values, but the expected decrease in TXB2 levels was not observed. These findings suggest that a lack of response to aspirin may be due to additional intrinsic factors rather than increased platelet turnover.

Adherence to the Mediterranean Diet Association with Serum Levels of Nitric Oxide, Prostacyclin, and Thromboxane B2 among Prinzmetal Angina Patients and Healthy Persons.

This study aimed to assess the association between adherence to the Mediterranean diet with serum Nitric oxide, Prostacyclin, and Thromboxane B2 among Prinzmetal angina patients and healthy persons. This case-control study was conducted among 100 Prinzmetal angina patients and 100 healthy persons referred to the Ardabil Imam Khomeini hospital between 2021 and 2022. Blood samples were obtained from all study participants for measurement of serum Nitric oxide, Prostacyclin, and Thromboxane B2. To calculate adherence to the Mediterranean diet, the ten-item screener was used. The serum Nitric oxide in patients who adhered more to the Mediterranean diet was higher than patients with less adherence (coeff. = 0.41 p = 0.04). The serum Prostacyclin level in patients with greater adherence to the Mediterranean diet was 0.34 units higher than patients with less adherence (coeff. = 0.34 p = 0.02). The level of serum Thromboxane B2 had a negative association with adherence to the Mediterranean diet (coeff. = -0.48 p = 0.04). The amount of consumption of olive oil, fruits, vegetables, and legumes in healthy people was more than Prinzmetal angina patients. In Prinzmetal angina patients, more adherence to the Mediterranean diet can decrease the serum Thromboxane B2 and increase the serum Nitric oxide and Prostacyclin.

Toll-Like Receptor 4-Dependent Platelet-Related Thrombosis in SARS-CoV-2 Infection.

BACKGROUND: SARS-CoV-2 is associated with an increased risk of venous and arterial thrombosis, but the underlying mechanism is still unclear. METHODS: We performed a cross-sectional analysis of platelet function in 25 SARS-CoV-2 and 10 healthy subjects by measuring Nox2 (NADPH oxidase 2)-derived oxidative stress and thromboxane B2, and investigated if administration of monoclonal antibodies against the S protein (Spike protein) of SARS-CoV-2 affects platelet activation. Furthermore, we investigated in vitro if the S protein of SARS-CoV-2 or plasma from SARS-CoV-2 enhanced platelet activation. RESULTS: Ex vivo studies showed enhanced platelet Nox2-derived oxidative stress and thromboxane B2 biosynthesis and under laminar flow platelet-dependent thrombus growth in SARS-CoV-2 compared with controls; both effects were lowered by Nox2 and TLR4 (Toll-like receptor 4) inhibitors. Two hours after administration of monoclonal antibodies, a significant inhibition of platelet activation was observed in patients with SARS-CoV-2 compared with untreated ones. In vitro study showed that S protein per se did not elicit platelet activation but amplified the platelet response to subthreshold concentrations of agonists and functionally interacted with platelet TLR4. A docking simulation analysis suggested that TLR4 binds to S protein via three receptor-binding domains; furthermore, immunoprecipitation and immunofluorescence showed S protein-TLR4 colocalization in platelets from SARS-CoV-2. Plasma from patients with SARS-CoV-2 enhanced platelet activation and Nox2-related oxidative stress, an effect blunted by TNF (tumor necrosis factor) α inhibitor; this effect was recapitulated by an in vitro study documenting that TNFα alone promoted platelet activation and amplified the platelet response to S protein via p47phox (phagocyte oxidase) upregulation. CONCLUSIONS: The study identifies 2 TLR4-dependent and independent pathways promoting platelet-dependent thrombus growth and suggests inhibition of TLR4. or p47phox as a tool to counteract thrombosis in SARS-CoV-2.

Personalized allocation of acetylsalicylic acid therapy for secondary prevention of coronary artery disease.

Background: A single-daily dose of 75 mg of acetylsalicylic acid inhibits 100% of thromboxane-B2 synthesis within 30-60 min. Thromboxane-B2 synthesis then recovers slowly as new platelets are released from the bone marrow. Normally, only 10% of the platelets are replaced daily by new platelets entering circulation. Hence, 24 h after a dose of acetylsalicylic acid, thromboxane-B2 synthesis is still suppressed by more than 90%. Hence, there is an adequate anti-platelet effect even after 24 h of acetylsalicylic acid intake. However, some patients treated with once-daily acetylsalicylic acid may have an incomplete 24-h suppression of thromboxane-B2 synthesis due to increased platelet turnover. The response could be improved in such patients by twice-daily acetylsalicylic acid administration. This study aimed to identify such a group of patients who would benefit from a twice-daily dose of acetylsalicylic acid. Materials and methods: Serum thromboxane-B2 levels were measured in 79 patients with coronary artery disease receiving 75 mg of acetylsalicylic acid for secondary prophylaxis. Serum levels of thromboxane-B2 were measured after 4 and 24 h of acetylsalicylic acid intake. Patients were then classified into three groups: steady suppression group (serum thromboxane B2 is adequately suppressed at 4 and 24 h), i.e., adequate response to acetylsalicylic acid; fast recovery group (more than 10% rise in serum thromboxane-B2 levels at 24-h when compared to at 4-h) and non-responders (serum thromboxane-B2 levels of >3,100 pg/ml after 4 h of acetylsalicylic acid intake). Patients in the fast recovery group were given twice-daily acetylsalicylic acid and thromboxane-B2 levels were re-measured. Results: A total of 20 patients (24.3%) had steady suppression of thromboxane-B2 and 11 patients (13.9%) belonged to the fast recovery group, i.e., thromboxane-B2 levels were adequately suppressed at 4 h but had recovered by more than 10% at 24 h; which was adequately suppressed by twice-daily acetylsalicylic acid (p 0.004). A total of 48 patients (60.8%) were non-responders. Conclusion: Twice-daily acetylsalicylic acid may be beneficial if serum thromboxane-B2 levels at 4 h are <3,100 and >3,100 pg/ml at 24 h. If thromboxane-B2 levels at 4 and 24 h is <3100 pg/ml but if there is a >10% rise in serum thromboxane B2 at 24 h as compared to that at 4 h, then twice-daily acetylsalicylic acid should be considered. However, if thromboxane-B2 at 4 and 24 h is >3,100 pg/ml consider switching over to a P2Y12 inhibitor.

Factors Associated with Aspirin Resistance in Hong Kong Chinese Patients with Stable Coronary Heart Disease Using the Multiplate® Analyzer and Serum Thromboxane B2.

Introduction: Aspirin resistance may be associated with various conditions. We measured serum thromboxane B2 (TXB2) and platelet function using the Multiplate® Analyzer with arachidonic acid (ASPI) in patients on long term aspirin therapy to identify aspirin resistance and associated factors. Materials and Methods: Chinese patients with stable coronary heart disease had samples for serum TXB2 and ASPI measurement taken before and 1 h after taking a morning dose of 80 mg aspirin. Results: In 266 patients with mean age 66.6 ± 10.7 years, 17% were female and 55% were current or previous smokers. TXB2 and ASPI measurements were significantly higher before the dose than at 1 h post dose, with 46% of subjects having high ASPI values (AUC > 300 AU*min) pre dose compared with 27% at 1 h post dose. TXB2 and ASPI measures of platelet aggregation showed weak correlations, which were only significant before the dose (r = 0.219, p = 0.001). Increased ASPI measurements were associated with white blood cell (WBC) count, haematocrit, platelet count and heart rate at 24 h post dose but only with WBC count, smoking history and heart rate at 1 h post dose. Diabetes was not associated with reduced platelet response to aspirin. The WBC count associated with aspirin resistance was over 6.55 × 109/L by receiver operating characteristic analysis. Conclusions: The antiplatelet response to aspirin was reduced in a large proportion of patients. Patients with higher WBC count within the normal range appear to be at increased risk of aspirin resistance. Higher or more frequent doses of aspirin may be needed in many patients.

The impact of enteric coating of aspirin on aspirin responsiveness in patients with suspected or newly diagnosed ischemic stroke: prospective cohort study: results from the (ECASIS) study.

BACKGROUND AND PURPOSE: Uncertainty remains regarding the impact of enteric-coated aspirin (EC-ASA) on secondary prevention of ischemic stroke compared to plain aspirin (P-ASA). Hence, this study was designed to investigate the effect of EC formulation on ASA response via evaluating thromboxane B2 (TXB2) levels in patients with suspected or newly diagnosed stroke. METHODS: A prospective cohort study on suspected or newly diagnosed ischemic stroke patients who are aspirin-naive was conducted. Patients were received either EC aspirin or plain aspirin for at least 3 days. The primary outcome was the proportion of aspirin non-responsiveness between two groups (level of residual serum TXB2 associated with elevated thrombotic risk (< 99.0% inhibition or TXB2 > 3.1 ng/ml) within 72 h after three daily aspirin doses, while secondary outcomes were the incidence of early gastrointestinal tract (GIT) bleeding with the various aspirin preparations. (Trial registration: Clinicaltrials.gov NCT04330872 registered on 02 April 2020). RESULTS: Of 42 patients, ischemic strokes were confirmed in both P-ASA (81%) and EC-ASA (67%) arms. ASA non-responsiveness showed no significant difference between the two formulations (P-ASA vs. EC-ASA; 28.6% vs 23.8%; P = 0.726). Univariate and multivariate logistic regression analysis showed that patients treated with EC-ASA were more likely to have a lower rate of non-responders compared to P-ASA (unadjusted OR 0.78; 95% CI 0.20, 3.11); with the risk highest in type 2 diabetic patients with HBA1c > 6.5% (adjusted OR 6; 95% CI 1.02, 35.27; P = 0.047). No incidence of GIT bleeding observed throughout the study. CONCLUSION: A significant proportion of ASA non-responsiveness was recorded regardless of ASA formulation administered. The increased risk of ASA non-responsiveness in diabetic patients needs further exploration by larger prospective studies.

Cardioprotective effects of sinomenine in myocardial ischemia/reperfusion injury in a rat model.

Background: Ischemia reperfusion (I/R) play an imperative role in the expansion of cardiovascular disease. Sinomenine (SM) has been exhibited to possess antioxidant, anticancer, anti-inflammatory, antiviral and anticarcinogenic properties. The aim of the study was scrutinized the cardioprotective effect of SM against I/R injury in rat. Methods: Rat were randomly divided into normal control (NC), I/R control and I/R + SM (5, 10 and 20 mg/kg), respectively. Ventricular arrhythmias, body weight and heart weight were estimated. Antioxidant, inflammatory cytokines, inflammatory mediators and plasmin system indicator were accessed. Results: Pre-treated SM group rats exhibited the reduction in the duration and incidence of ventricular fibrillation, ventricular ectopic beat (VEB) and ventricular tachycardia along with suppression of arrhythmia score during the ischemia (30 and 120 min). SM treated rats significantly (P < 0.001) altered the level of antioxidant parameters. SM treatment significantly (P < 0.001) repressed the level of creatine kinase MB (CK-MB), creatine kinase (CK) and troponin I (Tnl). SM treated rats significantly (P < 0.001) repressed the tissue factor (TF), thromboxane B2 (TXB2), plasminogen activator inhibitor 1 (PAI-1) and plasma fibrinogen (Fbg) and inflammatory cytokines and inflammatory mediators. Conclusion: Our result clearly indicated that SM plays anti-arrhythmia effect in I/R injury in the rats via alteration of oxidative stress and inflammatory reaction.

Urine 11-Dehydro-Thromboxane B2 in Aspirin-Naive Males with Metabolic Syndrome.

Urine 11-dehydro-thromboxane B2 (11-dehydro-TXB2), an indirect measure of platelet activity, is elevated in cardiovascular diseases and diabetes. The purpose of our study was to determine whether urine 11-dehydro-TXB2 is elevated in aspirin-naive males with metabolic syndrome (MS) and to determine predictors of 11-dehydro-TXB2 levels. The secondary aim was to evaluate whether these MS patients could be potential candidates for the aspirin-mediated prevention of atherosclerotic cardiovascular diseases (ASCVDs). In 82 males with MS (76 hypertensive), anthropometric measures, urine 11-dehydro-TXB2, platelet count, creatinine, glucose, insulin, estimated insulin resistance, lipid parameters, high-sensitivity C-reactive protein (hs-CRP), adiponectin, homocysteine, and ten-year risk of fatal cardiovascular disease (SCORE) were assessed. Urine 11-dehydro-TXB2 levels were elevated (≥2500 pg/mg creatinine) in two-thirds of patients, including 11 high-risk patients (SCORE ≥ 5%). Homocysteine, adiponectin, hs-CRP, waist-to-hip ratio, and total cholesterol were found to be predictors of urine 11-dehydro-TXB2. In conclusion, there is a high incidence of elevated urine 11-dehydro-TXB2 in males with MS, including in some patients who are at a high or very high risk of ASCVDs. 11-dehydro-TXB2 levels are associated with hyperhomocysteinemia, inflammation, fat distribution, hypercholesterolemia, and adiponectin concentrations. Elevated 11-dehydro-TXB2 levels may support the use of personalised aspirin ASCVD prevention in high-risk males with MS. Giuseppe Patti.

Concentrations of PGE2 and TXB2 in the Eyes of Mice with Disseminated Acanthamoebiasis.

Previous studies have shown that Acanthamoeba spp. may invade the eyes by migrating along the optic nerve to the eyes from the brain. This study aimed to confirm the presence of inflammation in the eyes of mice with disseminated acanthamoebiasis by examining prostaglandin E2 (PGE2) and thromboxane B2 (TXB2) concentrations in the eyes of immunocompetent and immunocompromised mice intranasally inoculated with Acanthamoeba spp. The PGE2 concentration was statistically significantly lower in the immunocompromised amoebae-infected mice on 8 dpi compared with the noninfected group of animals, and it was higher in the eyes of immunosuppressed amoebae-infected mice on 16 dpi than in the control group of animals. There was a statistically significant lower TXB2 concentration in the eyes of immunocompetent infected mice compared with the noninfected group on 8 dpi. However, on 24 dpi, we noted statistically significant higher TXB2 levels in the immunocompetent infected mice than in the control group. In immunocompromised mice, there was a lower TXB2 level on 8 dpi than in control mice. This study confirmed the existence of an inflammatory process in the eyes of immunocompetent and immunocompromised mice infected with Acanthamoeba spp. without damaged corneas.

Flavonoids: Antiplatelet Effect as Inhibitors of COX-1.

Flavonoids are compounds with a benzopyranic structure that exhibits multiple pharmacological activities. They are known for their venotonic activity, but their mechanism of action remains unclear. It is thought that, as this mechanism is mediated by prostaglandins, these compounds may interfere with the arachidonic acid (AA) cascade. These assays are designed to measure the antiplatelet aggregation capacity of quercetin, rutin, diosmetin, diosmin, and hidrosmin, as well as to evaluate a potential structure-activity ratio. In this paper, several studies on platelet aggregation at different concentrations (from 0.33 mM to 1.5 mM) of different flavone compounds are conducted, measuring platelet aggregation by impedance aggregometry, and the cyclooxygenase (COX) activity by metabolites generated, including the activity of the pure recombinant enzyme in the presence of these polyphenols. The results obtained showed that quercetin and diosmetin aglycones have a greater antiplatelet effect and inhibit the COX enzyme activity to a greater extent than their heterosides; however, the fact that greater inhibition of the pure recombinant enzyme was achieved by heterosides suggests that these compounds may have difficulty in crossing biological membranes. In any case, in view of the results obtained, it can be concluded that flavonoids could be useful as coadjuvants in the treatment of cardiovascular pathologies.

Serum thromboxane B2 but not soluble P-selectin levels identify ischemic stroke patients with persistent platelet reactivity while on aspirin therapy.

BACKGROUND: Aspirin non-response due to persistent platelet reactivity has been associated with adverse vascular events. Light transmission aggregometry (LTA), the 'gold standard' for measuring the platelet response to aspirin therapy, is a cumbersome procedure and a simple and reliable alternative is required. Our aim was to explore whether serum thromboxane B2 (sTXB2) and soluble P-selectin can be used to identify patients who are at risk of increased platelet reactivity while on aspirin. METHODS AND RESULTS: We recruited 293 ischemic stroke patients, taking aspirin for more than seven days, and performed LTA to classify them. Based on therapeutic serum salicylate levels, 63 patients were excluded due to suspected non-compliance, followed by ELISA measurement of TXB2 and P-selectin in serum. Accordingly, patients were classified into 'Responders' (n = 122, 53%), 'Semi-responders' (n = 76, 33%) and 'Non-responders' (n=32, 14%) by LTA. Patients who had platelet aggregation of ≥70% with 10µM ADP and ≥20% with 0.5mM AA were defined as 'Non-responders'. In comparison with 'Responders', 'Non-responders' had 8.63-fold increased risk of secondary vascular events (p = 0.008). ROC curve analysis revealed that sTXB2, at a cut-off level of >4.15 ng/mL, could distinguish the patient group with elevated platelet reactivity with a sensitivity of 84.3% (AUC = 0.84), and was in fair agreement with the LTA-based classification of patients. Soluble P-selectin levels, on the other hand, had no discriminatory ability. CONCLUSION: We suggest sTXB2 measurement as an alternative to the LTA approach for identifying aspirin-treated ischemic stroke patients who are at risk of enhanced platelet reactivity and subsequent vascular events.

First Experience Addressing the Prognostic Utility of Novel Urinary Biomarkers in Patients With COVID-19.

Urine 11-dehydro-thromboxane B2 (u11-dh-TxB2), 8-hydroxy-2'-deoxyguanosine (8-OHdG), and liver-type fatty acid binding protein levels (L-FABP) at the time of hospitalization were higher in coronavirus disease 2019 (COVID-19) patients with adverse events vs without events. Higher u11-dh-TxB2 and L-FABP levels were associated with longer hospitalization, more thrombotic events, and greater mortality, providing evidence for potential utility as early prognostic biomarkers for COVID-19.