The effect of Bruton's tyrosine kinase inhibitor ibrutinib on atherothrombus formation under stenotic flow conditions.

BACKGROUND: Bruton's kinase (Btk) is critical for collagen-triggered platelet signal transduction. The Btk inhibitor ibrutinib has been shown to selectively block platelet adhesion to atherosclerotic plaque material under laminar arterial flow. However, this has not been studied under a shear gradient, which is characteristic for atherothrombosis. OBJECTIVE: To determine the effect of ibrutinib treatment on in vitro thrombus formation on collagen and atherosclerotic plaque material in the absence or presence of a shear gradient. METHODS: Blood was obtained from patients with chronic lymphocytic leukemia, mantle-cell lymphoma and Waldenström macroglobulinemia with and without ibrutinib treatment and perfused through a microfluidic channel with(out) 60% stenosis over Horm type I collagen or human atherosclerotic plaque homogenate. RESULTS: At a constant shear rate of 1500 s-1, platelet deposition was significantly decreased in blood from haematological malignancy patients treated with ibrutinib as compared to untreated patients, on atherosclerotic plaque material but not on collagen. However, thrombus size, stability, and height, were reduced on both plaque material and collagen. An increase in shear rate up to 3900 s-1, as induced by 60% stenosis, resulted in decreased platelet deposition and thrombus parameters on plaque material but not on collagen when compared to a laminar shear of 1500 s-1. Ibrutinib treatment decreased platelet deposition and thrombus parameters even further around the stenosis. CONCLUSION: Treatment of patients with haematological disorders with the Btk inhibitor ibrutinib reduces in vitro platelet deposition, thrombus size and contraction on human atherosclerotic plaque around a stenosis when compared to patients not receiving ibrutinib.

Finding the "switch" in platelet activation: prediction of key mediators involved in reversal of platelet activation using a novel network biology approach.

The cAMP-protein kinase A (PKA) pathway in platelets is important for both platelet activation and inactivation. We hypothesize that proteins/processes downstream of the cAMP-PKA pathway that are regulated after platelet activation ánd subsequent inactivation can serve as a "switch" in platelet activation and inhibition. We used a STRING-based protein-protein interaction network from proteins of interest distilled from publicly available quantitative platelet proteome datasets. The protein network was integrated with biological pathway information by functional enrichment analysis, phosphorylation by PKA, and drug-target information. Functional enrichment analysis revealed biological processes related to vesicle secretion and cytoskeletal reorganization to be overrepresented among these 30 proteins coinciding with topological clusters in the network. Our method identified proteins/processes with functions related to vesicle transport, cyclin-dependent protein kinases, tight junctions, and small GTPases as potential switches in platelet activation and inhibition. Next to established enzymes in cAMP-PKA signaling, such as PDE3A, proteins with an unknown/less well-known role in platelet biology, such as Stonin-2 and ABLIM-3, emerged from our analysis as interesting candidates for reversal of platelet activation. Our method can be used to repurpose existing datasets and provide a coherent overview of mechanisms involved to predict novel connections, by visually integrating multiple datasets. SIGNIFICANCE: This article presents a novel approach of visually incorporating multiple existing tools and proteomics datasets and in doing so provides novel insight into the complex molecular mechanisms involved in platelet activation. Using our approach, we also highlight several interesting candidates for future research into pathologies with high platelet reactivity.

Characterization of Atherosclerotic Plaque Coating for Thrombosis Microfluidics Assays.

INTRODUCTION: Studying arterial thrombus formation by in vitro flow assays is a widely used approach. Incorporating human atherosclerotic plaque material as a thrombogenic surface in these assays represents a method to model the pathophysiological environment of thrombus formation upon plaque disruption. Up until now, achieving a homogeneous coating of plaque material and subsequent reproducible platelet adhesion has been challenging. Here, we characterized a novel method for coating of plaque material on glass coverslips for use in thrombosis microfluidic assays. METHODS: A homogenate of human atherosclerotic plaques was coated on glass coverslips by conventional manual droplet coating or by spin coating. Prior to coating, a subset of coverslips was plasma treated. Water contact angle measurements were performed as an indicator for the hydrophilicity of the coverslips. Homogeneity of plaque coatings was determined using profilometric analysis and scanning electron microscopy. Thrombogenicity of the plaque material was assessed in real time by microscopic imaging while perfusing whole blood at a shear rate of 1500 s-1 over the plaque material. RESULTS: Plasma treatment of glass coverslips, prior to spin coating with plaque material, increased the hydrophilicity of the coverslip compared to no plasma treatment. The most homogeneous plaque coating and highest platelet adhesion was obtained upon plasma treatment followed by spin coating of the plaque material. Manual plaque coating on non-plasma treated coverslips yielded lowest coating homogeneity and platelet adhesion and activation. CONCLUSION: Spin coating of atherosclerotic plaque material on plasma treated coverslips leads to a more homogenous coating and improved platelet adhesion to the plaque when compared to conventional droplet coating on non-plasma treated coverslips. These properties are beneficial in ensuring the quality and reproducibility of flow experiments. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12195-021-00713-9.

Ethics review for sale? Conflict of interest and commercial research review boards.

Research review boards, established to protect the rights and welfare of human research subjects, have to ensure that conflicts of interest do not interfere with the ethical conduct of medical research. Private, commercial review boards, which increasingly review research protocols, are themselves affected by a structural conflict of interest. Within the regulatory setting, procedural conflict-of-interest rules are essential because of the absence of clear substantive rules in research review and the reliance on the fairness and good judgment of institutional review board members. Current guidelines and regulations lack adequate conflict-of-interest rules and provide insufficient details on the substantive rules. Because commercial review boards are similar to administrative courts and tribunals, rules of administrative law on bias are applied to determine when a conflict of interest jeopardizes the purposes of research review; administrative law has always judged financial conflicts of interest severely. The structure of private review tends to breach a core principle of administrative law and procedural justice. Reform of the research review system will reinforce public trust in the process.

Selective justice, genetic discrimination, and insurance: should we single out genes in our laws?

This article discusses the desirability of legislation focusing on genetic discrimination, in particular in the context of insurance. Many American states and some European countries as well as the Council of Europe have introduced protective measures against discrimination on the basis of genetic susceptibility. The author questions their effectiveness and queries whether they may be inequitable, because they fail to address more fundamental underlying issues related to the nature of insurance, access to health care, and unequal distribution of wealth. There is also a problem of definition in these statutes. They fail to capture what constitutes genetic information. Nonetheless, the author argues it is important to consider the social consequences of genetic testing. Michael Walzer's theory of justice is used to examine the role of insurance and health care. Using this approach, the author finds the American system of distribution for health care to be problematic. This is then used to inform the author's discussion of the future of health care in Canada. Anti-discrimination provisions could be used in a way that is consistent with Walzer's theory of justice. They would encompass both genetic and non-genetic health factors. These can be modelled on current anti-discrimination statutes in Canada. The author then proposes administrative committee structures to regulate the use of genetic data in Canada.

Primary prevention of arterial thromboembolism in non-rheumatic atrial fibrillation in primary care: randomised controlled trial comparing two intensities of coumarin with aspirin.

OBJECTIVE: To investigate the effectiveness of aspirin and coumarin in preventing thromboembolism in patients with non-rheumatic atrial fibrillation in general practice. DESIGN: Randomised controlled trial. PARTICIPANTS: 729 patients aged >/=60 years with atrial fibrillation, recruited in general practice, who had no established indication for coumarin. Mean age was 75 years and mean follow up 2. 7 years. SETTING: Primary care in the Netherlands. INTERVENTIONS: Patients eligible for standard intensity coumarin (international normalised ratio 2.5-3.5) were randomly assigned to standard anticoagulation, very low intensity coumarin (international normalised ratio 1.1-1.6), or aspirin (150 mg/day) (stratum 1). Patients ineligible for standard anticoagulation were randomly assigned to low anticoagulation or aspirin (stratum 2). MAIN OUTCOME MEASURES: Stroke, systemic embolism, major haemorrhage, and vascular death. RESULTS: 108 primary events occurred (annual event rate 5.5%), including 13 major haemorrhages (0.7% a year). The hazard ratio was 0.91 (0.61 to 1.36) for low anticoagulation versus aspirin and 0.78 (0.34 to 1.81) for standard anticoagulation versus aspirin. Non-vascular death was less common in the low anticoagulation group than in the aspirin group (0.41, 0.20 to 0.82). There was no significant difference between the treatment groups in bleeding incidence. High systolic and low diastolic blood pressure and age were independent prognostic factors. CONCLUSION: In a general practice population (without established indications for coumarin) neither low nor standard intensity anticoagulation is better than aspirin in preventing primary outcome events. Aspirin may therefore be the first choice in patients with atrial fibrillation in general practice.

Primary prevention of arterial thromboembolism in nonrheumatic atrial fibrillation: the PATAF trial study design.

Patients with nonrheumatic atrial fibrillation (NRAF) have a higher risk of thromboembolism than patients in sinus rhythm. Several trials have been conducted to establish the best preventive regimen in patients with NRAF, but not in the primary-care setting. The Primary Prevention of Arterial Thromboembolism in Nonrheumatic Atrial Fibrillation (PATAF) study, a primary-care-based trial, was set up to compare the preventive efficacy of low-intensity anticoagulation (AC), target range International Normalized Ratio (INR) 1.1 < INR < 1.6 and regular-intensity AC (2.5 < INR < 3.5) therapies with that of aspirin 150 mg/d for the occurrence of thromboembolism in NRAF patients. Patients eligible for regular-intensity AC were randomly assigned to aspirin at 150 mg/d, low-intensity AC, or regular AC in group I. In cases of noneligibility for regular AC, the trial randomized patients between aspirin and low-intensity AC (assigned to group II). Primary outcome events were stroke (including intracranial hemorrhage), systemic embolism, major hemorrhage, or vascular death. Analysis of the data was based on Cox regression to compute the hazard ratio (HR) with a 95% confidence interval, using the likelihood ratio test. The trial randomized 729 patients. Patient enrollment and follow-up has been stopped, and the final analysis is now complete. We shall publish the main results as soon as possible.

Bioethics for clinicians: 17. Conflict of interest in research, education and patient care.

A conflict of interest occurs in a situation in which professional judgement regarding a primary interest, such as research, education or patient care, may be unduly influenced by a secondary interest, such as financial gain or personal prestige. Conflicts of interest exist in every walk of life, including medicine and science. There is nothing inherently unethical in finding oneself in a conflict of interest. Rather, the key questions are whether one recognizes the conflict and how one deals with it. Strategies include disclosing the conflict, establishing a system of review and authorization, and prohibiting the activities that lead to the conflict.

Towards the right to be killed? Treatment refusal, assisted suicide and euthanasia in the United States and Canada.

This chapter describes some dominant trends of American and Canadian law in relation to treatment refusal, physician-assisted suicide and euthanasia. Although common law in both countries recognizes the right of patients to refuse treatment, problems have arisen, especially in the US, over treatment refusal on behalf of incompetent patients. One response has been to enact advance-directive legislation, promoting the use of living wills and proxy appointments. Courts have also specified criteria for withholding and withdrawing treatment from incompetent patients. The notion of a "right to die', developed in court cases on treatment refusal, is now being invoked to support the legalization of assisted suicide. Courts are generally reluctant to recognize an extention of this right. Debates and court cases following the recent initiative to legalize assisted suicide in Oregon and the Sue Rodriguez case in Canada's Supreme Court, which resulted in a special report of a Canadian Senate Committee, are of major importance for the development of law in this area.