The P2X1 receptor and platelet function

Extracellular nucleotides are ubiquitous signalling molecules, acting via the P2 class of surface receptors. Platelets express three P2 receptor subtypes, ADP-dependent P2Y1 and P2Y12 G-protein-coupled receptors and the ATP-gated P2X1 non-selective cation channel. Platelet P2X1 receptors can generate significant increases in intracellular Ca²⁺, leading to shape change, movement of secretory granules and low levels of α_IIbβ₃ integrin activation. P2X1 can also synergise with several other receptors to amplify signalling and functional events in the platelet. In particular, activation of P2X1 receptors by ATP released from dense granules amplifies the aggregation responses to low levels of the major agonists, collagen and thrombin. In vivo studies using transgenic murine models show that P2X1 receptors amplify localised thrombosis following damage of small arteries and arterioles and also contribute to thromboembolism induced by intravenous co-injection of collagen and adrenaline. In vitro, under flow conditions, P2X1 receptors contribute more to aggregate formation on collagen-coated surfaces as the shear rate is increased, which may explain their greater contribution to localised thrombosis in arterioles compared to venules within in vivo models. Since shear increases substantially near sites of stenosis, anti-P2X1 therapy represents a potential means of reducing thrombotic events at atherosclerotic plaques.     

A protective effect of the synthetic coumarine derivative Cloricromene against DNB-colitis in the rat

Biologic therapies, namely antibodies against tumor necrosis factor-alpha (TNF- alpha) or its receptors, have been recently introduced for the treatment of patients with inflammatory bowel disease (IBD). In the present study the effects of cloricromene, an agent with known antithrombotic actions and with demonstrated anti-TNF- alpha activity were investigated in a rat model of experimental colitis induced with dinitrobenzenesulphonic acid (DNB)/ethanol. We investigated three experimental groups: (i) sham-colitis with vehicle-treatment (controls, n = 6), (ii) colitis with vehicle-treatment (saline, 0.1 ml s.c., daily) (DNB-V, n = 7), (iii) colitis with cloricromene-treatment (10 mg/kg/day s.c.; DNB-C, n = 8). After 7 days, the weight gain, colon wet weight, macroscopic damage score, coagulation parameters, colon mucosal myeloperoxidase activity (MPO), and tissue concentrations of TNF- alpha and of macrophage inhibitory peptide-2 (MIP-2) were assessed. The macroscopic damage scores, colon wet weights, and tissue MIP-2 levels were significantly increased in untreated and in cloricromene-treated rats compared with controls. Cloricromene treatment was associated with a minor body weight loss (p < 0.025) and significantly reduced tissue concentrations of MPO and TNF-alpha (p < 0.02, both). Blood coagulation parameters were not affected by treatment. In the DNB-model treatment with cloricromene effectively reduces tissue levels of TNF- alpha and of myeloperoxidase, whereas MIP-2 concentrations were not influenced. Blood coagulation parameters remained unchanged indicating safety of treatment. Since biological therapies frequently fail to improve disease course of IBD, other therapies with similar targets should be further investigated.     

Optimization of ketone-based P2Y12 receptor antagonists as antithrombotic agents: Pharmacodynamics and receptor kinetics considerations

Modification of a series of P2Y₁₂ receptor antagonists by replacement of the ester functionality was aimed at minimizing the risk of in vivo metabolic instability and pharmacokinetic variability. The resulting ketones were then optimized for their P2Y₁₂ antagonistic and anticoagulation effects in combination with their physicochemical and absorption profiles. The most promising compound showed very potent antiplatelet action in vivo. However, pharmacodynamic–pharmacokinetic analysis did not reveal a significant separation between its anti-platelet and bleeding effects. The relevance of receptor binding kinetics to the in vivo profile is described.     

Low-dose intravenous nitrite improves hemodynamics in a canine model of acute pulmonary thromboembolism

Acute pulmonary thomboembolism (APT)-induced pulmonary hypertension can be counteracted by activating the nitric oxide (NO)-cGMP pathway. Recent studies have demonstrated that the naturally occurring anion nitrite (NO₂^-) is a bioactive storage reservoir for NO, and is reduced to NO under conditions of hypoxia and acidosis. We hypothesized that nitrite infused intravenously could attenuate the hemodynamic changes associated with APT. APT was induced with autologous blood clots injected into the right atrium in mongrel dogs. After APT (or saline), the dogs received an intravenous nitrite (or saline) infusion (6.75 μmol/kg over 15 min and then 0.28 μmol/kg/min) and hemodynamic evaluations were carried out for 2 h. Plasma nitrite concentrations were measured using ozone-based reductive chemiluminescence methodologies. APT decreased cardiac index (CI) and increased pulmonary vascular resistance index (PVRI); these effects were improved during infusions of sodium nitrite. Accordingly, nitrite infusion increased cardiac index by 28%, reduced the PVRI by 48%, and the systemic vascular resistance index (SVRI) by 21% in embolized dogs, suggesting a greater effect on the ischemic embolized vascular system than the systemic circulation following embolization. Interestingly, in nonembolized control dogs the same nitrite infusion decreased MAP and CI (all P < 0.05). The nitrite infusion increased plasma nitrite concentrations by approximately 2 μM, and produced dose-dependent effects on PVRI, MAP, and SVRI. Remarkably, blood levels of nitrite as low as 500 nM decreased PVRI and SVRI in this model, suggesting a potential role of nitrite in physiological blood flow regulation. These results suggest that a low-dose nitrite infusion produces beneficial hemodynamic effects in a dog model of APT. These findings suggest a new therapeutic application for nitrite and support emerging evidence for a surprisingly potent and potentially physiological vasoactivity of nitrite.     

肾病综合征患者血清同型半胱氨酸、D-二聚体和B型钠尿肽水平表达及与血栓栓塞的关系分析

摘要 目的：探讨肾病综合征（NS）患者血清同型半胱氨酸（Hcy）、D-二聚体（D-D）和B型钠尿肽（BNP）水平及与发生血栓栓塞（TE）的关系。方法：选取2017年1月~2019年1月我院收治的241例NS患者,根据是否并发TE分为TE组56例,非TE组185例,另选取65名体检健康者为对照组,对比各组血清Hcy、D-D和BNP水平,多因素Logistics回归分析NS患者并发TE的影响因素,ROC曲线判断血清Hcy、D-D和BNP水平对NS患者并发TE的诊断价值。结果：三组血清Hcy、D-D和BNP水平比较差异有统计学意义（P＜0.05）。TE组血清Hcy、D-D和BNP水平明显高于非TE组,非TE组血清Hcy、D-D和BNP水平明显高于对照组,差异均有统计学意义（P＜0.05）。TE组高血压史、利尿剂使用史、糖皮质激使用史比例和BUN、24hUPQ水平明显高于非TE组,差异均有统计学意义（P＜0.05）。多因素Logistics回归分析显示,Hcy、D-D、BNP、24hUPQ为NS患者并发TE独立影响因素（P＜0.05）。ROC曲线显示,血清Hcy+D-D+BNP水平诊断NS患者并发TE的AUC明显大于血清Hcy、D-D和BNP单独诊断,敏感度和特异度也高于各指标单独诊断。结论：NS患者血清Hcy、D-D和BNP水平明显提升,合并TE后进一步提高,且血清Hcy、D-D和BNP水平为NS患者发生TE的危险因素,联合测定可提高诊断价值。     

Antithrombotic therapy in heart failure

Patients with congestive heart failure have a significant risk of stroke due to thromboembolism from the dilated left ventricle. Two relatively small trials suggest that oral anticoagulation with vitamin-K antagonists may reduce this risk when compared with placebo, aspirin or clopidogrel. However, more studies are eagerly awaited. So far, physicians seeing patients with heart failure should decide who needs antithrombotic prophylaxis on a case-by-case basis, especially since most heart failure patients have significant comorbidity precluding the use of oral anticoagulant.     

Discovery of 9-(1-phenoxyethyl)-2-morpholino-4-oxo-pyrido[1,2-a]pyrimidine-7-carboxamides as oral PI3Kβ inhibitors,useful as antiplatelet agents

Optimization of AZD6482 (2), the first antiplatelet PI3Kβ inhibitor evaluated in man, focused on improving the pharmacokinetic profile to a level compatible with once daily oral dosing as well as achieving adequate selectivity towards PI3Kα to minimize the risk for insulin resistance. Structure-based design and optimization of DMPK properties resulted in (R)-16, a novel, orally bioavailable PI3Kβ inhibitor with potent in vivo anti-thrombotic effect with excellent separation to bleeding risk and insulin resistance.     

The association between two genetic polymorphisms in ITGB3 and increase risk of venous thromboembolism in cancer patients in Eastern Province of Saudi Arabia

Venous thromboembolism (VTE) is one of the major complications in most cancer patients leading to poor prognosis and short survival. Several common clinical risk factors coexist in cancer patients are used as risk predictive biomarkers to help in the management and prevention of VTE. These include cancer site and stage, chemotherapy regimen and elevated biological markers. However, Genetic polymorphisms in genes controlling coagulation and fibrinolysis are significantly associated with VTE if detected, then they might be more sensitive individual predictive biomarkers for VTE risk assessment. This study was conducted to evaluate the association between ITGB3 rs3809865 and rs5918 with VTE risk as well as monitor the effect of VTE on overall survival of these cancer patients. In this retrospective case-control study, 195 cancer patients’ formalin-fixed paraffin embedded tissue (FFPE) samples were collected (controls n = 157, case n = 38) using the stored data through Jan 2010 to Sep 2018 from King Fahad Specialist Hospital in Dammam. Samples were genotyped using TaqMan genotyping assay, then logistic regression analysis and Chi-square were used to predict the association between risk factors and VTE. Survival Comparison was tested by the log-rank test. Genetic polymorphisms in ITGB3 (rs3809865 and rs5918) found not to be associated with VTE increasing risk in cancer patients (p>0.05). While the advanced stage was potentially increasing the risk of VTE events (OR 5.1 CI 2.01–12.9p = 0.001). Patients with VTE showed a poor overall survival reflected by the median survival rate of only three years compared to seven years for cancer patients without VTE. This study highlighted the potential influence of VTE on prognosis and survival of cancer patients and raised the importance of exploring risk predictive biomarkers in our population. This will improve the risk prediction biomarkers leading to implementing safe and effective thrombosis prophylaxis strategies.     

心房颤动并发血栓栓塞患者脑钠肽与D-二聚体的表达及相关性分析

目的:观察房颤及房颤并发血栓栓塞患者血浆内脑钠肽(brain natriuretic peptide,BNP)和D-二聚体(D-dimer)的表达水平;探讨两者表达水平的关联性以及两者对房颤血栓栓塞的预测价值。方法:回顾分析2010年5月-2012年12月上海市第一人民医院心内科住院病人;根据入组及排除标准将符合条件的研究对象74例分为对照组、单纯房颤组与房颤血栓组;对所有对象进行数据采集,包括年龄、性别、血脂情况、高血压病史、血糖等情况;对所有对象进行D-dimer及BNP水平的数据采集。结果:(1)房颤血栓组的年龄明显高于对照组(P0.01)和单纯房颤组(P0.001);(2)房颤血栓组的D-dimer和BNP水平高于单纯房颤组(P0.05)和对照组(P0.001);(3)单纯房颤组BNP水平与D-dimer水平呈正相关性(r=0.507,P=0.004),房颤血栓组BNP水平与D-dimer水平呈正相关性(r=0.680,P0.001)。结论:(1)心房颤动患者随着年龄的增加并发血栓栓塞风险也增加,指导我们在临床治疗时需要重视年龄因素。(2)患者血浆中D-dimer和BNP水平的增高是心房颤动并发血栓栓塞患者的危险信号。(3)D-dimer和BNP检测在预防心房颤动并发血栓栓塞中有重要的临床意义。