The diversity and clinical implications of genetic variants influencing clopidogrel bioactivation and response in the Emirati population.

BACKGROUND: Clopidogrel is a widely prescribed prodrug that requires activation via specific pharmacogenes to exert its anti-platelet function. Genetic variations in the genes encoding its transporter, metabolizing enzymes, and target receptor lead to variability in its activation and platelet inhibition and, consequently, its efficacy. This variability increases the risk of secondary cardiovascular events, and therefore, some variations have been utilized as genetic biomarkers when prescribing clopidogrel. METHODS: Our study examined clopidogrel-related genes (CYP2C19, ABCB1, PON1, and P2Y12R) in a cohort of 298 healthy Emiratis individuals. The study used whole exome sequencing (WES) data to comprehensively analyze pertinent variations of these genes, including their minor allele frequencies, haplotype distribution, and their resulting phenotypes. RESULTS: Our data shows that approximately 37% (n = 119) of the cohort are likely to benefit from the use of alternative anti-platelet drugs due to their classification as intermediate or poor CYP2C19 metabolizers. Additionally, more than 50% of the studied cohort exhibited variants in ABCB1, PON1, and P2YR12 genes, potentially influencing clopidogrel's transport, enzymatic clearance, and receptor performance. CONCLUSIONS: Recognizing these alleles and genotype frequencies may explain the clinical differences in medication response across different ethnicities and predict adverse events. Our findings underscore the need to consider genetic variations in prescribing clopidogrel, with potential implications for implementing personalized anti-platelet therapy among Emiratis based on their genetic profiles.

Low-dose aspirin for the prevention of atherosclerotic cardiovascular disease.

During the past 30 years, several developments have occurred in the antiplatelet field, including the role of aspirin in primary prevention of atherosclerotic cardiovascular disease. There have been several attempts to develop antiplatelet drugs more effective and safer than aspirin and a shift in emphasis from efficacy to safety, advocating aspirin-free antiplatelet regimens after percutaneous coronary intervention. Evidence supporting a chemopreventive effect of low-dose aspirin against colorectal (and other digestive tract) cancer has also strengthened. The aim of this article is to revisit the role of aspirin in the prevention of atherothrombosis across the cardiovascular risk continuum, in view of developments in the antiplatelet field. The review will offer a clinical perspective on aspirin's mechanism of action, pharmacokinetics, and pharmacodynamics. This will be followed by a detailed discussion of its clinical efficacy and safety.

Aspirin-free antiplatelet strategies after percutaneous coronary interventions.

Dual antiplatelet therapy (DAPT) with aspirin and a platelet P2Y12 receptor inhibitor is the standard antithrombotic treatment after percutaneous coronary interventions (PCI). Several trials have challenged guideline-recommended DAPT after PCI by testing the relative clinical effect of an aspirin-free antiplatelet approach-consisting of P2Y12 inhibitor monotherapy after a short course (mostly 1-3 months) of DAPT-among patients undergoing PCI without a concomitant indication for oral anticoagulation (OAC). Overall, these studies have shown P2Y12 inhibitor monotherapy after short DAPT to be associated with a significant reduction in the risk of bleeding without an increase in thrombotic or ischaemic events compared with continued DAPT. Moreover, the effects of the P2Y12 inhibitor monotherapy without prior DAPT or following a very short course of DAPT after PCI are being investigated in emerging studies, of which one has recently reported unfavourable efficacy results associated with the aspirin-free approach compared with conventional DAPT. Finally, P2Y12 inhibitor alone has been compared with aspirin alone as chronic therapy after DAPT discontinuation, thus challenging the historical role of aspirin as a standard of care for secondary prevention following PCI. A thorough understanding of study designs, populations, treatments, results, and limitations of trials testing P2Y12 inhibitor monotherapy vs. DAPT or vs. aspirin is required to consider adopting this treatment in clinical practice. This review addresses the use of aspirin-free antiplatelet strategies among patients undergoing PCI without a concomitant indication for OAC, providing an overview of clinical evidence, guideline indications, practical implications, ongoing issues, and future perspectives.

Comparison of 3- to 6-Month Versus 12-Month Dual Antiplatelet Therapy After Coronary Intervention Using the Contemporary Drug-Eluting Stents With Ultrathin Struts: The HOST-IDEA Randomized Clinical Trial.

BACKGROUND: Limited data are available on short-term dual antiplatelet therapy (DAPT) after percutaneous coronary intervention using third-generation drug-eluting stents with ultrathin struts and advanced polymer technology. We investigated whether 3- to 6-month DAPT was noninferior to 12-month DAPT after implantation of drug-eluting stents with ultrathin struts and advanced polymer technology. METHODS: We performed an open-label, randomized trial at 37 centers in South Korea. We enrolled patients undergoing percutaneous coronary intervention using the Orsiro biodegradable-polymer sirolimus-eluting stents or the Coroflex ISAR polymer-free sirolimus-eluting stents. Patients with ST-segment-elevation myocardial infarction were excluded. Patients were randomly assigned to receive either 3- to 6-month or 12-month DAPT after percutaneous coronary intervention. The choice of antiplatelet medications was at the physician's discretion. The primary outcome was a net adverse clinical event, a composite of cardiac death, target vessel myocardial infarction, clinically driven target lesion revascularization, stent thrombosis, or major bleeding, defined as Bleeding Academic Research Consortium type 3 or 5 at 12 months. The major secondary outcomes were target lesion failure, a composite of cardiac death, target vessel myocardial infarction, clinically driven target lesion revascularization, and major bleeding. RESULTS: A total of 2013 patients (mean age, 65.7±10.5 years; 1487 males [73.9%]; 1110 [55.1%] presented with acute coronary syndrome) were randomly assigned to 3- to 6-month DAPT (n=1002) or 12-month DAPT (n=1011). The primary outcome occurred in 37 (3.7%) patients in the 3- to 6-month DAPT group and 41 (4.1%) in the 12-month DAPT group. The noninferiority of the 3- to 6-month DAPT group to the 12-month DAPT group was met (absolute risk difference, -0.4% [1-sided 95% CI, -∞% to 1.1%]; P<0.001 for noninferiority). There were no significant differences in target lesion failure (hazard ratio, 0.98 [95% CI, 0.56-1.71], P=0.94) or major bleeding (hazard ratio, 0.82 [95% CI, 0.41-1.61], P=0.56) between the 2 groups. Across various subgroups, the treatment effect of 3- to 6-month DAPT was consistent for net adverse clinical event. CONCLUSIONS: Among patients undergoing percutaneous coronary intervention using third-generation drug-eluting stents, 3- to 6-month DAPT was noninferior to 12-month DAPT for net adverse clinical event. Further research is needed to generalize this finding to other populations and to determine the ideal regimen for 3- to 6-month DAPT. REGISTRATION: URL: https://www. CLINICALTRIALS: gov; Unique identifier: NCT02601157.

P2Y12 adenosine receptor inhibitors preloading among patients with non-ST-elevation acute coronary syndromes; insights from a nationwide multicenter registry.

BACKGROUND: The safety and efficacy of treatment with P2Y12 adenosine-diphosphate receptor inhibitors (P2Y12-RI) before coronary angiography among patients with non-ST-segment elevation acute coronary syndromes (NSTEACS) are questionable. AIMS: To assess the pretreatment rate with P2Y12-RI and its association with ischemic and bleeding risks among patients with NSTEACS. METHODS: The study comprised patients with NSTEACS referred for coronary angiography and included in the Acute Coronary Syndrome Israeli Surveys between 2013 and 2021. Patients were divided into two groups according to the timing of P2Y12-RI loading concerning coronary angiography: pretreatment and posttreatment. The primary endpoints were 30-day major adverse cardiovascular events (MACE; composite of cardiovascular mortality, myocardial infarction, stroke, stent thrombosis, and urgent revascularization) and 1-year all-cause mortality. RESULTS: Of 3076 patients, 2423 (78.8%) received pretreatment with a P2Y12-RI, and 653 (21.2%) received P2Y12-RI posttreatment. Prasugrel and ticagrelor were used more in the posttreatment group compared to the pretreatment group (16% vs. 6% and 38% vs. 25%, respectively, p < 0.001 for both). No difference was observed in the rate of 30-day MACE comparing pretreatment and posttreatment (5.3% vs. 2.2%, respectively, p = 0.62). A sensitivity analysis of 30-day MACE among patients from the 2021 survey demonstrated similar results (2.5% in the posttreatment group vs. 8.0% in the pretreatment group, p = 0.13). There were no differences in 1-year all-cause mortality rates between the pretreatment and posttreatment groups (4.8% vs. 3.8%, p = 0.31). CONCLUSIONS: Among patients with NSTEACS referred for an invasive strategy, the P2Y12-RI posttreatment strategy was associated with similar 30-day and 1-year MACE as the pretreatment strategy. These large-scale, multicenter, real-world data provide reassurance on the safety and efficacy of delaying P2Y12-IR until after coronary stratification to improve clinical decision-making.

Electroacupuncture Alleviates Pain by Suppressing P2Y12R-Dependent Microglial Activation in Monoarthritic Rats.

Electroacupuncture (EA) effectively improves arthritis-induced hyperalgesia and allodynia by repressing spinal microglial activation, which plays a crucial role in pain hypersensitivity following tissue inflammation. However, the mechanism by which EA suppresses spinal microglial activation in monoarthritis (MA) remains unclear. In the present study, a rat model of MA was established through unilateral ankle intra-articular injection of complete Freund's adjuvant (CFA). The relationship among P2Y12 receptor (P2Y12R) expression, spinal microglial activation, and EA analgesia was investigated using quantitative real-time PCR (qRT‒PCR), western blotting, immunofluorescence (IF), and behavioral testing. The results found that EA treatment at the ipsilateral "Huantiao" (GB30) and "Yanglingquan" (GB34) acupoints markedly attenuated pain and spinal microglia M1 polarization in MA rats. In particular, P2Y12R expression was significantly increased at the mRNA and protein levels in the spinal dorsal horn in MA rats, whereas EA treatment effectively repressed the MA-induced upregulation of P2Y12R. IF analysis further revealed that most P2Y12R was expressed in microglia in the spinal dorsal horn. Pharmacological inhibition of P2Y12R by its antagonist (AR-C69931MX) decreased MA-induced spinal microglial activation and subsequent proinflammatory cytokine production. Consequently, AR-C69931MX significantly intensified the anti-pain hypersensitive function of EA in MA rats. Taken together, these results demonstrate that EA alleviates MA-induced pain by suppressing P2Y12R-dependent microglial activation.

Role and recent progress of P2Y12 receptor in cancer development.

P2Y12 receptor (P2Y12R) is an adenosine-activated G protein-coupled receptor (GPCR) that plays a central role in platelet function, hemostasis, and thrombosis. P2Y12R activation can promote platelet aggregation and adhesion to cancer cells, promote tumor angiogenesis, and affect the tumor immune microenvironment (TIME) and tumor drug resistance, which is conducive to the progression of cancers. Meanwhile, P2Y12R inhibitors can inhibit this effect, suggesting that P2Y12R may be a potential therapeutic target for cancer. P2Y12R is involved in cancer development and metastasis, while P2Y12R inhibitors are effective in inhibiting cancer. However, a new study suggests that long-term use of P2Y12R inhibitors may increase the risk of cancer and the mechanism remains to be explored. In this paper, we reviewed the structural and functional characteristics of P2Y12R and its role in cancer. We explored the role of P2Y12R inhibitors in different tumors and the latest advances by summarizing the basic and clinical studies on the effects of P2Y12R inhibitors on tumors.

Cell Surface Platelet Tissue Factor Expression: Regulation by P2Y12 and Link to Residual Platelet Reactivity.

BACKGROUND: ADP-induced platelet activation leads to cell surface expression of several proteins, including TF (tissue factor). The role of ADP receptors in platelet TF modulation is still unknown. We aimed to assess the (1) involvement of P2Y1 and P2Y12 receptors in ADP-induced TF exposure; (2) modulation of TFpos-platelets in anti-P2Y12-treated patients with coronary artery disease. Based on the obtained results, we revisited the intracellular localization of TF in platelets. METHODS: The effects of P2Y1 or P2Y12 antagonists on ADP-induced TF expression and activity were analyzed in vitro by flow cytometry and thrombin generation assay in blood from healthy subjects, P2Y12-/-, and patients with gray platelet syndrome. Ex vivo, P2Y12 inhibition of TF expression by clopidogrel/prasugrel/ticagrelor, assessed by VASP (vasodilator-stimulated phosphoprotein) platelet reactivity index, was investigated in coronary artery disease (n=238). Inhibition of open canalicular system externalization and electron microscopy (TEM) were used for TF localization. RESULTS: In blood from healthy subjects, stimulated in vitro by ADP, the percentage of TFpos-platelets (17.3±5.5%) was significantly reduced in a concentration-dependent manner by P2Y12 inhibition only (-81.7±9.5% with 100 nM AR-C69931MX). In coronary artery disease, inhibition of P2Y12 is paralleled by reduction of ADP-induced platelet TF expression (VASP platelet reactivity index: 17.9±11%, 20.9±11.3%, 40.3±13%; TFpos-platelets: 10.5±4.8%, 9.8±5.9%, 13.6±6.3%, in prasugrel/ticagrelor/clopidogrel-treated patients, respectively). Despite this, 15% of clopidogrel good responders had a level of TFpos-platelets similar to the poor-responder group. Indeed, a stronger P2Y12 inhibition (130-fold) is required to inhibit TF than VASP. Thus, a VASP platelet reactivity index <20% (as in prasugrel/ticagrelor-treated patients) identifies patients with TFpos-platelets <20% (92% sensitivity). Finally, colchicine impaired in vitro ADP-induced TF expression but not α-granule release, suggesting that TF is open canalicular system stored as confirmed by TEM and platelet analysis of patients with gray platelet syndrome. CONCLUSIONS: Data show that TF expression is regulated by P2Y12 and not P2Y1; P2Y12 antagonists downregulate the percentage of TFpos-platelets. In clopidogrel good-responder patients, assessment of TFpos-platelets highlights those with residual platelet reactivity. TF is stored in open canalicular system, and its membrane exposure upon activation is prevented by colchicine.

Impact of P2Y12 inhibitors on clinical outcomes in sepsis-3 patients receiving aspirin: a propensity score matched analysis.

BACKGROUND: Sepsis is a life-threatening disease accompanied by disorders of the coagulation and immune systems. P2Y12 inhibitors, widely used for arterial thrombosis prevention and treatment, possess recently discovered anti-inflammatory properties, raising potential for improved sepsis prognosis. METHOD: We conducted a retrospective analysis using the data from Medical Information Mart for Intensive Care-IV database. Patients were divided into an aspirin-alone group versus a combination group based on the use of a P2Y12 inhibitor or not. Differences in 30-day mortality, length of stay (LOS) in intensive care unit (ICU), LOS in hospital, bleeding events and thrombotic events were compared between the two groups. RESULT: A total of 1701 pairs of matched patients were obtained by propensity score matching. We found that no statistically significant difference in 30-day mortality in aspirin-alone group and combination group (15.3% vs. 13.7%, log-rank p = 0.154). In addition, patients received P2Y12 inhibitors had a higher incidence of gastrointestinal bleeding (0.5% vs. 1.6%, p = 0.004) and ischemic stroke (1.7% vs. 2.9%, p = 0.023), despite having a shorter LOS in hospital (11.1 vs. 10.3, days, p = 0.043). Cox regression showed that P2Y12 inhibitor was not associated with 30-day mortality (HR = 1.14, 95% CI 0.95-1.36, p = 0.154). CONCLUSION: P2Y12 inhibitors did not provide a survival benefit for patients with sepsis 3 and even led to additional adverse clinical outcomes.

Trajectories of P2Y12 inhibitor adherence in patients with acute coronary syndromes.

PURPOSE: P2Y12 inhibitors (P2Y12i) reduce cardiac events after acute coronary syndromes (ACS). However, suboptimal P2Y12i adherence persists. We aimed to examine P2Y12i non-adherence using group-based trajectory methods and to identify adherence predictors. METHODS: We conducted a population-based, retrospective cohort study using administrative data in Ontario, Canada of patients ≥65 years admitted for ACS between April 2014 and March 2018 with a P2Y12i dispensed within 7 days of discharge. We used group-based trajectory models to characterize longitudinal 1-year adherence patterns. Predictors associated with each adherence trajectory were identified by multinomial logistic regression. RESULTS: We included 11 917 patients using clopidogrel and 9763 using ticagrelor, aged [mean ± SD]: 77.33 ± 8.31/73.59 ± 6.79 years; men: 56.2%/65.4%, respectively. We identified 3 longitudinal adherence trajectories, that differed by agent: 75% of clopidogrel and 68% of ticagrelor patients showed a consistently adherent trajectory, while 13%/17% were gradually, and 12%/15% were rapidly non-adherent, respectively (p < 0.001). Differing baseline characteristics in each cohort were associated with observed adherence trajectories. Concomitant atrial fibrillation and prior bleeding history were associated with non-adherence among clopidogrel users. Among ticagrelor users, women and older persons were more likely to be rapidly non-adherent, adherence declining steeply starting 1 month post-ACS. CONCLUSIONS: We identified distinct adherence trajectories for clopidogrel and ticagrelor post-ACS, with 3 out of 4 clopidogrel patients but only 2 out of 3 ticagrelor patients in the consistently adherent trajectory. Intensive interventions targeted to the period of steep adherence decline post-ACS, particularly for women and older persons initiating ticagrelor, and patients with atrial fibrillation on clopidogrel should be considered and investigated further.

P2X7 receptor-mediated depression-like reactions arising in the mouse medial prefrontal cortex.

Major depressive disorder is a frequent and debilitating psychiatric disease. We have shown in some of the acute animal models of major depressive disorder (tail suspension test and forced swim test) that depression-like behavior can be aggravated in mice by the microinjection into the medial prefrontal cortex of the P2X7R agonistic adenosine 5'-triphosphate or its structural analog dibenzoyl-ATP, and these effects can be reversed by the P2X7R antagonistic JNJ-47965567. When measuring tail suspension test, the prolongation of immobility time by the P2YR agonist adenosine 5'-[ß-thio]diphosphate and the reduction of the adenosine 5'-(γ-thio)triphosphate effect by P2Y1R (MRS 2179) or P2Y12R (PSB 0739) antagonists, but not by JNJ-47965567, all suggest the involvement of P2YRs. In order to elucidate the localization of the modulatory P2X7Rs in the brain, we recorded current responses to dibenzoyl-ATP in layer V astrocytes and pyramidal neurons of medial prefrontal cortex brain slices by the whole-cell patch-clamp procedure; the current amplitudes were not altered in preparations taken from tail suspension test or foot shock-treated mice. The release of adenosine 5'-triphosphate was decreased by foot shock, although not by tail suspension test both in the hippocampus and PFC. In conclusion, we suggest, that in the medial prefrontal cortex, acute stressful stimuli cause supersensitivity of P2X7Rs facilitating the learned helplessness reaction.

Low-dose prasugrel versus standard-dose ticagrelor in east Asian patients with acute coronary syndrome.

Low-dose prasugrel demonstrated a similar effectiveness profile to clopidogrel in East Asian ACS patients, but its comparison with another new-generation potent P2Y12 inhibitor, ticagrelor, remains unclear. To compare the effectiveness and safety of low-dose prasugrel against those of standard-dose ticagrelor in East Asian patients with ACS. This retrospective cohort study used Taiwan's National Health and Welfare Database. This study included ACS patients who underwent percutaneous coronary intervention and, at discharge between January 1, 2018 and December 31, 2020, were prescribed with low-dose prasugrel plus aspirin or standard-dose ticagrelor plus aspirin. Stabilized inverse probability of treatment weighting was used to balance the covariates across these two groups. The primary effectiveness outcome was a composite of acute myocardial infarction, ischemic stroke, and cardiovascular death; the secondary effectiveness outcome was each of the individual components of the primary outcome, transient ischemic attack, and repeat revascularization. The primary safety outcome was a composite of intracranial hemorrhage and gastrointestinal bleeding, and the two secondary safety outcomes were intracranial hemorrhage and gastrointestinal bleeding. A total of 24,807 patients were included in this study. Among them, 1,493 were low-dose prasugrel users and 23,314 were standard-dose ticagrelor users. No significant differences were found in primary effectiveness [HR: 0.97 (0.74-1.28)] or primary safety outcomes [HR: 1.22 (0.73-2.01)] between the two study groups. For East Asian patients with ACS, low-dose prasugrel provides comparable effectiveness without increasing bleeding risk compared to standard-dose ticagrelor. Low-dose prasugrel may be an appropriate alternative for East Asian populations.

Dual antiplatelet therapy duration after percutaneous coronary intervention in high bleeding risk: a meta-analysis of randomized trials.

AIMS: The optimal duration of dual antiplatelet therapy (DAPT) after percutaneous coronary intervention (PCI) in patients at high bleeding risk (HBR) is still debated. The current study, using the totality of existing evidence, evaluated the impact of an abbreviated DAPT regimen in HBR patients. METHODS AND RESULTS: A systematic review and meta-analysis was performed to search randomized clinical trials comparing abbreviated [i.e. very-short (1 month) or short (3 months)] with standard (≥6 months) DAPT in HBR patients without indication for oral anticoagulation. A total of 11 trials, including 9006 HBR patients, were included. Abbreviated DAPT reduced major or clinically relevant non-major bleeding [risk ratio (RR): 0.76, 95% confidence interval (CI): 0.61-0.94; I2 = 28%], major bleeding (RR: 0.80, 95% CI: 0.64-0.99, I2 = 0%), and cardiovascular mortality (RR: 0.79, 95% CI: 0.65-0.95, I2 = 0%) compared with standard DAPT. No difference in all-cause mortality, major adverse cardiovascular events, myocardial infarction, or stent thrombosis was observed. Results were consistent, irrespective of HBR definition and clinical presentation. CONCLUSION: In HBR patients undergoing PCI, a 1- or 3-month abbreviated DAPT regimen was associated with lower bleeding and cardiovascular mortality, without increasing ischaemic events, compared with a ≥6-month DAPT regimen. STUDY REGISTRATION: PROSPERO registration number CRD42021284004.

Moxibustion's protective role against atherosclerosis: Inhibition of Ca2+ overload-triggered oxidative stress and inflammatory response via P2Y12/PI3K/AKT pathway.

OBJECTIVE: This study aims to investigate the protective mechanism of moxibustion in combating atherosclerosis (AS). METHODS: Apolipoprotein E (ApoE)-deficient mice, aged 8 weeks, were randomly assigned into four groups: the model group (n = 6), SC79 group (n = 6), moxibustion group (n = 6), and moxibustion+SC79 group (n = 6). All mice were fed with a high-fat diet (HFD). Concurrently, 8-week-old C57BL/6 mice of the same genetic background were utilized as the control group (n = 6) and were given a regular diet. Macrophages were isolated via flow cytometry. The intracellular Ca2+ expression in macrophages was evaluated, and aortic plaques were quantitatively assessed through en face oil red O and Masson staining. The presence of macrophages and smooth muscle cells in AS plaques was determined by MAC-3 and α-smooth muscle actin (α-SMA) immunohistochemistry. The relative fluorescence intensity of nuclear factor-κB (NF-κB) in macrophages was identified by immunofluorescence staining. The expressions of proteins related to the P2Y12/phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT) signaling pathway were examined by Western blotting. RESULTS: Moxibustion reduced free Ca2+ expression in macrophage cytoplasm, inhibiting Ca2+ influx and oxidative stress. Significant reductions in atherosclerotic plaque formation and inflammation markers, including TNF-α and IL-1ß, were noted in the moxibustion group. Moxibustion modulated the P2Y12/PI3K/AKT pathway, impacting various inflammatory and oxidative stress-related proteins. Introduction of the AKT activator SC79 counteracted moxibustion's benefits, highlighting the P2Y12/PI3K/AKT pathway's central role. CONCLUSION: Moxibustion, through the P2Y12/PI3K/AKT signaling pathway, can inhibit Ca2+ overload-induced oxidative stress and inflammatory response, decrease macrophage infiltration, and increase the content of smooth muscle cells and collagen, thereby exerting a protective effect against AS.

Initial Despair and Current Hope of Identifying a Clinically Useful Treatment of Myocardial Reperfusion Injury: Insights Derived from Studies of Platelet P2Y12 Antagonists and Interference with Inflammation and NLRP3 Assembly.

Myocardial necrosis following the successful reperfusion of a coronary artery occluded by thrombus in a patient presenting with ST-elevation myocardial infarction (STEMI) continues to be a serious problem, despite the multiple attempts to attenuate the necrosis with agents that have shown promise in pre-clinical investigations. Possible reasons include confounding clinical risk factors, the delayed application of protective agents, poorly designed pre-clinical investigations, the possible effects of routinely administered agents that might unknowingly already have protected the myocardium or that might have blocked protection, and the biological differences of the myocardium in humans and experimental animals. A better understanding of the pathobiology of myocardial infarction is needed to stem this reperfusion injury. P2Y12 receptor antagonists minimize platelet aggregation and are currently part of the standard treatment to prevent thrombus formation and propagation in STEMI protocols. Serendipitously, these P2Y12 antagonists also dramatically attenuate reperfusion injury in experimental animals and are presumed to provide a similar protection in STEMI patients. However, additional protective agents are needed to further diminish reperfusion injury. It is possible to achieve additive protection if the added intervention protects by a mechanism different from that of P2Y12 antagonists. Inflammation is now recognized to be a critical factor in the complex intracellular response to ischemia and reperfusion that leads to tissue necrosis. Interference with cardiomyocyte inflammasome assembly and activation has shown great promise in attenuating reperfusion injury in pre-clinical animal models. And the blockade of the executioner protease caspase-1, indeed, supplements the protection already seen after the administration of P2Y12 antagonists. Importantly, protective interventions must be applied in the first minutes of reperfusion, if protection is to be achieved. The promise of such a combination of protective strategies provides hope that the successful attenuation of reperfusion injury is attainable.

Microglial remodeling of actin network by Tau oligomers, via G protein-coupled purinergic receptor, P2Y12R-driven chemotaxis.

Alzheimer's disease (AD) is associated with age-related neurodegeneration, synaptic deformation and chronic inflammation mediated by microglia and infiltrated macrophages in the brain. Tau oligomers can be released from damaged neurons via various mechanisms such as exosomes, neurotransmitter, membrane leakage etc. Microglia sense the extracellular Tau through several cell-surface receptors and mediate chemotaxis and phagocytosis. The purinergic receptor P2Y12R recently gained interest in neurodegeneration for neuro-glial communication and microglial chemotaxis towards the site of plaque deposition. To understand the effect of extracellular Tau oligomers in microglial migration, the P2Y12R-mediated actin remodeling, reorientation of tubulin network and rate of migration were studied in the presence of ATP. The extracellular Tau species directly interacted with P2Y12R and also induced this purinoceptor expression in microglia. Microglial P2Y12R colocalized with remodeled membrane-associated actin network as a component of migration in response to Tau oligomers. As an inducer of P2Y12R, ATP facilitated the localization of P2Y12R in lamellipodia and filopodia during accelerated microglial migration. The direct interaction of extracellular Tau oligomers with microglial P2Y12R would facilitate the signal transduction in both way, directional chemotaxis and receptor-mediated phagocytosis. These unprecedented findings emphasize that microglia can modulate the membrane-associated actin structure and incorporate P2Y12R to perceive the axis and rate of chemotaxis in Tauopathy.

Dehydrocorydaline alleviates sleep deprivation-induced persistent postoperative pain in adolescent mice through inhibiting microglial P2Y12 receptor expression in the spinal cord.

During adolescence, a second period of central nervous system (CNS) plasticity that follows the fetal period, which involves sleep deprivation (SD), becomes apparent. SD during adolescence may result in abnormal development of neural circuits, causing imbalance in neuronal excitation and inhibition, which not only results in pain, but increases the chances of developing emotion disorders in adulthood, such as anxiety and depression. The quantity of surgeries during adolescence is also consistently on the rise, yet the impact and underlying mechanism of preoperative SD on postoperative pain remain unexplored. This study demonstrates that preoperative SD induces upregulation of the P2Y12 receptor, which is exclusively expressed on spinal microglia, and phosphorylation of its downstream signaling pathway p38Mitogen-activated protein/Nuclear transcription factor-κB (p38MAPK/NF-κB)in spinal microglia, thereby promoting microglia activation and microglial transformation into the proinflammatory M1 phenotype, resulting in increased expression of proinflammatory cytokines that exacerbate persisting postoperative incisional pain in adolescent mice. Both intrathecal minocycline (a microglia activation inhibitor) and MRS2395 (a P2Y12 receptor blocker) effectively suppressed microglial activation and proinflammatory cytokine expression. Interestingly, supplementation with dehydrocorydaline (DHC), an extract of Rhizoma Corydalis, inhibited the P2Y12/p38MAPK/NF-κB signaling pathway, microglia activation, and expression of pro-inflammatory cytokines in the model mice. Taken together, the results indicate that the P2Y12 receptor and microglial activation are important factors in persistent postoperative pain caused by preoperative SD in adolescent mice and that DHC has analgesic effects by acting on these targets.

Effects of clopidogrel bisulfate on B16-F10 cells and tumor development in a murine model of melanoma.

Metastatic melanoma is a very aggressive skin cancer. Platelets are constituents of the tumor microenvironment and, when activated, contribute to cancer progression, especially metastasis and inflammation. P2Y12 is an adenosine diphosphate receptor that triggers platelet activation. Inhibition of P2Y12 by clopidogrel bisulfate (CB) decreases platelet activation, which is also controlled by the extracellular concentration and the metabolism of purines by purinergic enzymes. We evaluated the effects of CB on the viability and proliferation of cultured B16-F10 cells. We also used a metastatic melanoma model with C57BL-6 mice to evaluate cancer development and purine metabolism modulation in platelets. B16-F10 cells were administered intraperitoneally to the mice. Two days later, the animals underwent a 12-day treatment with CB (30 mg/kg by gavage). We have found that CB reduced cell viability and proliferation in B16-F10 culture in 72 h at concentrations above 30 µm. In vivo, CB decreased tumor nodule counts and lactate dehydrogenase levels and increased platelet purine metabolism. Our results showed that CB has significant effects on melanoma progression.

GSDMD knockdown attenuates phagocytic activity of microglia and exacerbates seizure susceptibility in TLE mice.

BACKGROUND: Temporal lobe epilepsy (TLE) is often characterized pathologically by severe neuronal loss in the hippocampus. Phagocytic activity of microglia is essential for clearing apoptotic neuronal debris, allowing for repair and regeneration. Our previous research has shown that gasdermin D (GSDMD)-mediated pyroptosis is involved in the pathogenesis of TLE. However, whether GSDMD-mediated pyroptosis influences the accumulation of apoptotic neurons remains unclear. Therefore, the present study was designed to investigate whether phagocytic activity of microglia is involved in GSDMD-mediated pyroptosis and the pathogenesis of TLE. METHODS: To establish a TLE model, an intra-amygdala injection of kainic acid (KA) was performed. The Racine score and local field potential (LFP) recordings were used to assess seizure severity. Neuronal death in the bilateral hippocampus was assessed by Nissl staining and TUNEL staining. Microglial morphology and phagocytic activity were detected by immunofluorescence and verified by lipopolysaccharide (LPS) and the P2Y12R agonist 2MeSADP. RESULTS: GSDMD knockdown augmented the accumulation of apoptotic neurons and seizure susceptibility in TLE mice. Microglia activated and transition to the M1 type with increased pro-inflammatory cytokines. Furthermore, GSDMD knockdown attenuated the migration and phagocytic activity of microglia. Of note, LPS-activated microglia attenuated seizure susceptibility and the accumulation of apoptotic neurons in TLE after GSDMD knockdown. A P2Y12R selective agonist, 2MeSADP, enhanced the migration and phagocytic activity of microglia. CONCLUSIONS: Our results demonstrate that GSDMD knockdown exacerbates seizure susceptibility and the accumulation of apoptotic neurons by attenuating phagocytic activity of microglia. These findings suggest that GSDMD plays a protective role against KA-induced seizure susceptibility.

P2Y12 receptor residues crucial for thrombosis regulation.

Thrombosis, or the formation of blood clots, can lead to serious medical conditions such as stroke, heart attack, and deep vein thrombosis. The purinoreceptor P2Y12 plays a critical role in the thrombotic pathway and is targeted for therapy to prevent clot formation. However, it is essential to balance the regulation of thrombosis to avoid adverse situations. This study focuses on the P2Y12 receptor and aims to discern the protein residue network and differentiate residues based on their intramolecular interactions. The study utilized a statistical analysis to characterize the significant residues involved in ligand interaction, which helps to identify critical residues that are essential for the function of the receptor. A parametric analysis of interactions of residues in the intraprotein interaction was conducted, which revealed significant residue-based contacts that facilitate protein interactions. By examining the interactions between residues, the mechanisms underlying protein interactions were studied and the importance of specific residues in facilitating these interactions was determined. This research provides important information on P2Y12, and the findings based on the network based significance of interacting residues may contribute to the development of new therapies that target the receptor to prevent clot formation while maintaining a balance in thrombosis regulation to avoid adverse outcomes. Ultimately, this study could lead to improved treatments for thrombotic disorders and better patient outcomes.