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1.
Methods Mol Biol ; 2663: 417-428, 2023.
Article in English | MEDLINE | ID: mdl-37204727

ABSTRACT

Immune-mediated heparin-induced thrombocytopenia (HIT) occurs when heparin-dependent IgG antibodies bind to heparin/platelet factor 4 (H/PF4) complexes and activate platelets. There is a vast panoply of assays to investigate HIT which can be divided into two groups, antigen-based immunoassays that detect all antibodies against H/PF4 and are used as a first diagnostic step and functional assays that will identify only the antibodies capable of activating platelets and are mandatory to confirm a diagnosis of pathological HIT. The serotonin-release assay, known as SRA, has been the gold standard for decades, but in the last 10 years, other easier alternatives have been described. The current chapter will focus on whole blood multiple electrode aggregometry, a validated method for the functional diagnosis of HIT.


Subject(s)
Platelet Function Tests , Thrombocytopenia , Humans , Electric Impedance , Platelet Function Tests/methods , Thrombocytopenia/chemically induced , Thrombocytopenia/diagnosis , Heparin/adverse effects , Immunoglobulin G , Platelet Factor 4/adverse effects , Anticoagulants/adverse effects
3.
Aust Crit Care ; 36(5): 762-768, 2023 09.
Article in English | MEDLINE | ID: mdl-36371291

ABSTRACT

BACKGROUND: The hospitalisation of a patient in intensive care impacts the psychological health of family members, with a high prevalence of anxiety, depression, and post-traumatic stress symptoms reported among families of critically ill patients. Understanding of the behavioural and physiological impact is limited and presents a new area of focus. OBJECTIVES: The objective of this study was to evaluate behavioural and physiological stress responses of visiting family members following hospitalisation of their adult relative. METHODS: Prospective longitudinal evaluation included 40 family members of adult patients with admission to intensive or coronary care in a large tertiary care metropolitan hospital. Assessments were conducted at three timepoints: in-hospital within 1 week of admission and 2 weeks and 3 months post discharge. Assessments included duration and quality of sleep (self-reported and actigraphy measured), physical activity, dietary and alcohol patterns, resting heart rate and blood pressure, and morning blood cortisol and lipid levels. Assessment of a reference group of 40 non-hospital-exposed control participants was also conducted. RESULTS: At the in-hospital assessment, study participants reported lower sleep time, altered 24-h physical activity patterns, reduced dietary and alcohol intake, and higher systolic and diastolic blood pressure than a nonhospitalised reference group. Compared to in-hospital assessment, these altered behavioural and physiological responses improved over time except for systolic blood pressures which remained unchanged at 3 months post family member discharge. CONCLUSION: Hospitalisation is associated with altered behavioural and physiological responses in family members. These findings contribute to understanding of the impact of unexpected hospitalisation on family members' cardiovascular risk factors and provide insights into potential mechanisms for the proposed increased risk during this time. Elevated systolic blood pressure at 3 months post discharge suggests a prolonged cardiovascular stress response in many family members of critical care patients that requires further study, with a focus on contributing and potential modifiable factors.


Subject(s)
Aftercare , Cardiovascular Diseases , Adult , Humans , Prospective Studies , Cardiovascular Diseases/epidemiology , Patient Discharge , Risk Factors , Family/psychology , Hospitalization , Anxiety/psychology , Stress, Physiological , Heart Disease Risk Factors , Intensive Care Units
4.
Eur J Hum Genet ; 30(10): 1121-1131, 2022 10.
Article in English | MEDLINE | ID: mdl-35970915

ABSTRACT

Whole genome sequencing (WGS) improves Mendelian disorder diagnosis over whole exome sequencing (WES); however, additional diagnostic yields and costs remain undefined. We investigated differences between diagnostic and cost outcomes of WGS and WES in a cohort with suspected Mendelian disorders. WGS was performed in 38 WES-negative families derived from a 64 family Mendelian cohort that previously underwent WES. For new WGS diagnoses, contemporary WES reanalysis determined whether variants were diagnosable by original WES or unique to WGS. Diagnostic rates were estimated for WES and WGS to simulate outcomes if both had been applied to the 64 families. Diagnostic costs were calculated for various genomic testing scenarios. WGS diagnosed 34% (13/38) of WES-negative families. However, contemporary WES reanalysis on average 2 years later would have diagnosed 18% (7/38 families) resulting in a WGS-specific diagnostic yield of 19% (6/31 remaining families). In WES-negative families, the incremental cost per additional diagnosis using WGS following WES reanalysis was AU$36,710 (£19,407;US$23,727) and WGS alone was AU$41,916 (£22,159;US$27,093) compared to WES-reanalysis. When we simulated the use of WGS alone as an initial genomic test, the incremental cost for each additional diagnosis was AU$29,708 (£15,705;US$19,201) whereas contemporary WES followed by WGS was AU$36,710 (£19,407;US$23,727) compared to contemporary WES. Our findings confirm that WGS is the optimal genomic test choice for maximal diagnosis in Mendelian disorders. However, accepting a small reduction in diagnostic yield, WES with subsequent reanalysis confers the lowest costs. Whether WES or WGS is utilised will depend on clinical scenario and local resourcing and availability.


Subject(s)
Exome , Base Sequence , Chromosome Mapping , Humans , Exome Sequencing , Whole Genome Sequencing
5.
Pathology ; 54(6): 746-754, 2022 Oct.
Article in English | MEDLINE | ID: mdl-35750510

ABSTRACT

Identification of disordered platelet function is important to guide peri-operative bleeding management as well as long term treatment and prognostic strategies in individuals with platelet bleeding disorders. Light transmission aggregometry (LTA), the current gold standard diagnostic test of platelet function is a time consuming technique almost exclusively performed in specialised laboratories and almost universally unavailable in regional centres in Australia, where there is an unmet need for access to specialised platelet function diagnostic services. 96-well plate-based aggregometry (Optimul, UK), has been utilised in research laboratories as a novel platform to investigate platelet function. We evaluated the Optimul assay at two centres in Australia, one regional and one tertiary metropolitan, to assess its feasibility as a screening test applicable to remote regional centres. Concentration-response curves were established from 45 healthy volunteers at the participating regional hospital and from 31 healthy volunteers at the tertiary institution. Optimul successfully detected anti-platelet effects in individuals taking aspirin (n=4), NSAID (n=2), clopidogrel (n=2) and dual therapy with aspirin and clopidogrel (n=1). When tested in parallel to LTA in individuals referred for the evaluation of abnormal bleeding symptoms there was overall a very good level of agreement between Optimul and LTA [Cohen's kappa (k2)=0.84], supporting its role as a useful screening tool in the assessment of platelet function. Optimul assay performance was quick and the methodology simple, requiring no specialised training or resources to be implemented at either the regional or metropolitan laboratory. Widespread implementation, particularly in regional laboratories within Australia where specialised platelet function testing is unavailable, has the potential to drastically improve the inequity of access to such services.


Subject(s)
Blood Platelet Disorders , Platelet Aggregation , Anti-Inflammatory Agents, Non-Steroidal , Aspirin/pharmacology , Blood Platelet Disorders/diagnosis , Clopidogrel/pharmacology , Humans , Pilot Projects , Platelet Function Tests/methods
6.
Platelets ; 33(8): 1139-1145, 2022 Nov 17.
Article in English | MEDLINE | ID: mdl-35316151

ABSTRACT

The use of mean platelet diameter (MPD) to classify inherited thrombocytopenia (IT) has been demonstrated in several studies. Alternatively, the mean platelet volume (MPV) may be used, but in macrothrombocytopenia this may not be available. We hypothesized that platelet forward scatter (FSC) measurements using flow cytometry may be used for the size-based classification of IT. The study aimed to assess the ability of platelet FSC to measure platelet size and whether it could be used as an alternative to the MPD or MPV.Blood samples were obtained from individuals undergoing investigation for inherited platelet function disorders (IPFD, n = 40) or platelet number disorders (IPND, n = 46). A hematology analyzer was used to obtain MPV and platelet counts, flow cytometry to measure platelet FSC and ImageJ software to measure MPD from stained blood smears. The International Society of Thrombosis and Hemostasis (ISTH) Bleeding Assessment Tool (BAT) was used to calculate bleeding scores.Twenty-nine(63%) of IPND patients had an MPV that could not be reported. A significant correlation to platelet FSC was found to the MPD (p < .0001) and MPV (p < .0001) and an inverse correlation with platelet count (p < .0001). No significant correlation was found between FSC and bleeding history. In conclusion, platelet FSC is an alternative to MPV and may be used in macrothrombocytopenia where the MPV is not recorded.


Subject(s)
Blood Platelet Disorders , Thrombocytopenia , Blood Platelets , Flow Cytometry , Hemorrhage , Humans , Mean Platelet Volume , Platelet Count , Thrombocytopenia/diagnosis
7.
Haematologica ; 107(1): 260-267, 2022 01 01.
Article in English | MEDLINE | ID: mdl-33472357

ABSTRACT

GFI1B is a transcription factor essential for the regulation of erythropoiesis and megakaryopoiesis, and pathogenic variants have been associated with thrombocytopenia and bleeding. Analysing thrombocytopenic families by whole exome sequencing, we identified a novel GFI1B variant (c.648+5G>A), which causes exon 9 skipping and overexpression of a shorter p32 isoform. We report the clinical data of our patients and critically review the phenotype observed in individuals with different GFI1B variants leading to the same effect on the p32 expression. Since p32 is increased in acute and chronic leukemia cells, we tested the expression level of genes playing a role in various type of cancers, including hematological tumors and found that they are significantly dysregulated, suggesting a potential role for GFI1B in carcinogenesis regulation. Increasing the detection of individuals with GFI1B variants will allow us to better characterize this rare disease and determine whether it is associated with an increased risk of developing malignancies.


Subject(s)
Germ-Line Mutation , Thrombocytopenia , Carcinogenesis , Humans , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , Repressor Proteins/genetics , Thrombocytopenia/genetics
8.
J Thromb Haemost ; 20(1): 252-259, 2022 01.
Article in English | MEDLINE | ID: mdl-34657390

ABSTRACT

BACKGROUND: Functional tests for the diagnosis of heparin-induced thrombocytopenia (HIT) exhibit variable performance. OBJECTIVES: We evaluated in a multicenter study whether 5B9, a monoclonal anti-PF4/heparin IgG mimicking human HIT antibodies, could be used as an internal quality control. METHODS: 5B9 was sent to 11 laboratories in seven countries, and six initial concentrations ranging from 10 to 400 µg/mL were tested by heparin-induced platelet activation assay (HIPA), serotonin release assay (SRA), platelet aggregation test (PAT), flow cytometry (FC), or heparin-induced multiple-electrode aggregometry (HIMEA). Each method was evaluated in three different laboratories using experimental procedures identical to those usually applied for the diagnosis of HIT by testing platelets from 10 different healthy donors. RESULTS: The procedures used varied among the laboratories, particularly when platelet-rich plasma and whole blood were used. Nevertheless, positive results were obtained with at least 100 µg/ml of 5B9 for most donors tested by all centers (except one) performing HIPA, SRA, or HIMEA. FC and PAT results were more heterogeneous. FC results from one center that used washed platelets preincubated with PF4 were positive with all donors at 50 µg/ml 5B9, but at least 200 µg/ml of 5B9 were required to activate cells with most donors tested using PAT. CONCLUSION: This study confirms that HIT functional tests are not well standardized and exhibit variable sensitivity for the detection of platelet-activating antibodies. However, 5B9 is a potentially useful tool to standardize functional tests, to select responding platelet donors, and consequently to improve the performance of these assays and comparability between laboratories.


Subject(s)
Platelet Factor 4 , Thrombocytopenia , Antibodies, Monoclonal , Anticoagulants/adverse effects , Blood Platelets , Communication , Heparin/adverse effects , Humans , Immunoglobulin G , Platelet Activation , Quality Control , Thrombocytopenia/chemically induced , Thrombocytopenia/diagnosis
9.
Platelets ; 33(3): 432-442, 2022 Apr 03.
Article in English | MEDLINE | ID: mdl-34223798

ABSTRACT

Variants of the Diaphanous-Related Formin 1 (DIAPH-1) gene have recently been reported causing inherited macrothrombocytopenia. The essential/"diagnostic" characteristics associated with the disorder are emerging; however, robust and complete criteria are not established. Here, we report the first cases of DIAPH1-related disorder in Australia caused by the autosomal dominant gain-of-function DIAPH1 R1213X variant formed by truncation of the protein within the diaphanous auto-regulatory domain (DAD) with loss of regulatory motifs responsible for autoinhibitory interactions within the DIAPH1 protein. We affirm phenotypic changes induced by the DIAPH1 R1213X variant to include macrothrombocytopenia, early-onset progressive sensorineural hearing loss, and mild asymptomatic neutropenia. High-resolution microscopy confirms perturbations of cytoskeletal dynamics caused by the DIAPH1 variant and we extend the repertoire of changes generated by this variant to include alteration of procoagulant platelet formation and possible dental anomalies.


Subject(s)
Blood Platelets/metabolism , Deafness/genetics , Formins/adverse effects , High-Throughput Nucleotide Sequencing/methods , Deafness/pathology , Humans , Phenotype
10.
J Am Heart Assoc ; 10(20): e020604, 2021 10 19.
Article in English | MEDLINE | ID: mdl-34622670

ABSTRACT

Background Although the association between dysregulated coagulation and atherosclerosis is well recognized, individual assays have been of minimal value in understanding disease susceptibility. Here we investigated the association of global coagulation profiles with coronary artery disease with consideration of sex differences. Methods and Results The study included patients from the BioHEART-CT (The BioHEART Study: Assessing Patients With Suspected Cardiovascular Disease for New Disease Markers and Risk Factors) biobank who had computed tomography coronary angiograms scored for coronary artery calcium score (CACS) and Gensini score. The cohort included 206 adult patients who were referred for clinically indicated computed tomography coronary angiography and had a median of 2 major cardiac risk factors; 50% were women and the average age was 62.6 years (±9.9 years). The overall hemostatic potential (OHP) and calibrated automated thrombography generation assays were performed on platelet-poor plasma. CACS and Gensini score in men were significantly correlated in bivariate analysis with measures from the OHP assay, and regression models predicting disease severity by CACS or Gensini score were improved by adding the OHP assay variables in men but not in women. The calibrated automated thrombography generation assay demonstrated a more hypercoagulable profile in women than in men. The OHP assay showed hypercoagulable profiles in women with hyperlipidemia and men with obesity. Conclusions The OHP assay identified hypercoagulable profiles associated with different risk factors for each sex and was associated with CACS and Gensini score severity in men, emphasizing the associations between increased fibrin generation and reduced fibrinolysis with cardiac risk factors and early atherosclerosis. Registration Information www.anzctr.org.au. Identifier: ACTRN12618001322224.


Subject(s)
Atherosclerosis , Cardiovascular Diseases , Coronary Artery Disease , Thrombophilia , Vascular Calcification , Adult , Atherosclerosis/diagnostic imaging , Atherosclerosis/epidemiology , Coronary Angiography , Coronary Artery Disease/diagnostic imaging , Coronary Artery Disease/epidemiology , Female , Heart Disease Risk Factors , Humans , Male , Middle Aged , Risk Factors , Tomography, X-Ray Computed
11.
J Thromb Haemost ; 19(10): 2612-2617, 2021 10.
Article in English | MEDLINE | ID: mdl-34355501

ABSTRACT

The implementation of high-throughput sequencing (HTS) technologies in research and diagnostic laboratories has linked many new genes to rare bleeding, thrombotic, and platelet disorders (BTPD), and revealed multiple genetic variants linked to those disorders, many of them being of uncertain pathogenicity when considering the accepted evidence (variant consequence, frequency in control datasets, number of reported patients, prediction models, and functional assays). The sequencing effort has also resulted in resources for gathering disease-causing variants associated with specific genes, but for BTPD, such well-curated databases exist only for a few genes. On the other hand, submissions by individuals or diagnostic laboratories to the variant database ClinVar are hampered by the lack of a submission process tailored to capture the specific features of hemostatic diseases. As we move toward the implementation of HTS in the diagnosis of BTPD, the Scientific and Standardization Committee for Genetics in Thrombosis and Haemostasis has developed and tested a REDCap-based interface, aimed at the community, to submit curated genetic variants for diagnostic-grade BTPD genes. Here, we describe the use of the interface and the initial submission of 821 variants from 30 different centers covering 14 countries. This open-access variant resource will be shared with the community to improve variant classification and regular bulk data transfer to ClinVar.


Subject(s)
Blood Platelet Disorders , Thrombosis , Blood Platelet Disorders/diagnosis , Blood Platelet Disorders/genetics , Communication , Genomics , Hemostasis/genetics , Humans , Thrombosis/diagnosis , Thrombosis/genetics
12.
Platelets ; 32(8): 998-1008, 2021 Nov 17.
Article in English | MEDLINE | ID: mdl-33284715

ABSTRACT

Ionotropic glutamate receptors include α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPAR), kainate receptors (KAR), and N-methyl-D-aspartate receptors (NMDAR). All function as cation channels; AMPAR and KAR are more permeable to sodium and NMDAR to calcium ions. Compared to the brain, receptor assemblies in platelets are unusual, suggesting distinctive functionalities.There is convincing evidence that AMPAR and KAR amplify platelet function and thrombus formation in vitro and in vivo. Transgenic mice lacking GluA1 and GluK2 (AMPAR and KAR subunits, respectively) have longer bleeding times and prolonged time to thrombosis in an arterial model. In humans, rs465566 KAR gene polymorphism associates with altered in vitro platelet responses suggesting enhanced aspirin effect. The NMDAR contribution to platelet function is less well defined. NMDA at low concentrations (≤10 µM) inhibits platelet aggregation and high concentrations (≥100 µM) have no effect. However, open NMDAR channel blockers interfere with platelet activation and aggregation induced by other agonists in vitro; anti-GluN1 antibodies interfere with thrombus formation under high shear rates ex vivo; and rats vaccinated with GluN1 develop iron deficiency anemia suggestive of mild chronic bleeding. In this review, we summarize data on glutamate receptors in platelets and propose a unifying model that reconciles some of the opposing effects observed.


Subject(s)
Blood Platelets/metabolism , Receptors, Ionotropic Glutamate/metabolism , Animals , Humans , Male , Rats
13.
Int J Mol Sci ; 21(21)2020 Nov 09.
Article in English | MEDLINE | ID: mdl-33182365

ABSTRACT

Ischaemic brain damage induces autoimmune responses, including the production of autoantibodies with potential neuroprotective effects. Platelets share unexplained similarities with neurons, and the formation of anti-platelet antibodies has been documented in neurological disorders. The aim of this study was to investigate the presence of anti-platelet antibodies in the peripheral blood of patients after ischaemic stroke and determine any clinical correlations. Using a flow cytometry-based platelet immunofluorescence method, we detected platelet-reactive antibodies in 15 of 48 (31%) stroke patients and two of 50 (4%) controls (p < 0.001). Western blotting revealed heterogeneous reactivities with platelet proteins, some of which overlapped with brain proteins. Stroke patients who carried anti-platelet antibodies presented with larger infarcts and more severe neurological dysfunction, which manifested as higher scores on the National Institutes of Health Stroke Scale (NIHSS; p = 0.009), but they had a greater recovery in the NIHSS by the time of hospital discharge (day 7 ± 2) compared with antibody-negative patients (p = 0.043). Antibodies from stroke sera reacted more strongly with activated platelets (p = 0.031) and inhibited platelet aggregation by up to 30.1 ± 2.8% (p < 0.001), suggesting the potential to interfere with thrombus formation. In conclusion, platelet-reactive antibodies can be found in patients soon after ischaemic stroke and correlate with better short-term outcomes, suggesting a potential novel mechanism limiting thrombosis.


Subject(s)
Autoantibodies/immunology , Blood Platelets/immunology , Brain Ischemia/immunology , Ischemic Stroke/immunology , Aged , Autoimmunity/immunology , Blood Coagulation/immunology , Female , Humans , Male , Platelet Aggregation/immunology , Platelet Count/methods , Thrombosis/immunology
14.
Res Pract Thromb Haemost ; 4(1): 106-110, 2020 Jan.
Article in English | MEDLINE | ID: mdl-31989091

ABSTRACT

Germline mutations of runt-related transcription factor-1 (RUNX1) cause familial platelet disorder with predisposition to myeloid malignancy (FPDMM), most commonly associated with thrombocytopenia and propensity to develop myeloid neoplasms. A key clinical question is which patients with a family history of thrombocytopenia should undergo genetic testing for RUNX1 mutations. Typically, molecular diagnosis by genetic sequencing is performed when the clinical phenotype is suggestive of this diagnosis; however, our understanding of the spectrum of associated features suggestive of this diagnosis continues to evolve. Herein, we report a case series of 3 unrelated families with RUNX1-associated FPDMM and clinical phenotypes not typically reported with this condition. These cases expand our understanding of FPDMM and highlight the complexity of transcriptional regulation of hematopoiesis and its potentially diverse phenotypes. We describe our approach to diagnosis and management of these individuals and the importance of long-term surveillance in these cases.

15.
Pathology ; 52(2): 243-255, 2020 Feb.
Article in English | MEDLINE | ID: mdl-31932033

ABSTRACT

Inherited disorders of platelet function (IPFD) and/or number (IPND) are heterogeneous conditions that result in variable mucocutaneous bleeding symptoms as a result of deranged primary haemostasis caused by platelet dysfunction or thrombocytopenia. Diagnosis is important to guide post-operative bleeding prophylactic strategies, to avoid treatment with inappropriate medications, and inform prognosis. Achieving an accurate diagnosis has traditionally been hampered by the requirement of multiple, often complex, laboratory tests that are not always available at single centres. To improve the diagnosis of these disorders a research collaborative was established, the Sydney Platelet Group, that explored an integrated approach combining traditional and contemporary platelet phenotypic and genetic diagnostic platforms available at four Sydney tertiary hospitals. Herein we report the outcomes of the first 50 patients evaluated using this approach. The cohort included 22 individuals with suspected IPFD and 28 with thrombocytopenia. Bleeding scores were higher in individuals with IPFD (mean 5.75; SD 4.83) than those with IPNDs (mean 2.14; SD 2.45). In cases with suspected IPFD, diagnosis to the level of the defective pathway was achieved in 71% and four individuals were found not to have a definitive platelet function defect. Dense granule secretion disorders were the most common platelet pathway abnormality detected (n=5). Mean bleeding scores in these individuals were not significantly different to individuals with defects in other commonly detected platelet pathways (dense granules, signal transduction and 'undetermined'). A molecular diagnosis was achieved in 52% of individuals with IPNDs and 5% with IPFD. Likely pathogenic and pathogenic variants detected included variants associated with extra-haematological complications (DIAPH1, MYH9) and potential for malignancy (ANKRD26 and RUNX1). The level of platelet investigation undertaken by this initiative is currently not available elsewhere in Australia and initial results confirm the utility of this integrated phenotypic-genetic approach.


Subject(s)
Blood Platelet Disorders/diagnosis , Platelet Function Tests/methods , Adolescent , Adult , Aged , Australia , Blood Platelet Disorders/genetics , Child , Child, Preschool , Female , Humans , Male , Middle Aged , Young Adult
16.
Am Heart J ; 220: 264-272, 2020 02.
Article in English | MEDLINE | ID: mdl-31923768

ABSTRACT

BACKGROUND: Bereavement is associated with an increased risk of cardiovascular disease; however, no reports exist of interventions to reduce risk. In a randomized, double-blind, placebo-controlled trial of 85 recently bereaved participants, we determined whether ß-blocker (metoprolol 25 mg) and aspirin (100 mg) reduce cardiovascular risk markers and anxiety, without adversely affecting bereavement intensity. METHODS: Participants were spouses (n = 73) or parents (n = 12) of deceased from 5 hospitals in Sydney, Australia, 55 females, 30 males, aged 66.1 ±â€¯9.4 years. After assessment within 2 weeks of bereavement, subjects were randomized to 6 weeks of daily treatment or placebo, and the effect evaluated using ANCOVA, adjusted for baseline values (primary analysis). RESULTS: Participants on metoprolol and aspirin had lower levels of home systolic pressure (P = .03), 24-hour average heart rate (P < .001) and anxiety (P = .01) platelet response to arachidonic acid (P < .001) and depression symptoms (P = .046) than placebo with no difference in standard deviation of NN intervals index (SDNNi), von Willebrand Factor antigen, platelet-granulocyte aggregates or bereavement intensity. No significant adverse safety impact was observed. CONCLUSIONS: In early bereavement, low dose metoprolol and aspirin for 6 weeks reduces physiological and psychological surrogate measures of cardiovascular risk. Although further research is needed, results suggest a potential preventive benefit of this approach during heightened cardiovascular risk associated with early bereavement.


Subject(s)
Adrenergic beta-1 Receptor Antagonists/therapeutic use , Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Aspirin/therapeutic use , Bereavement , Cardiovascular Diseases/prevention & control , Metoprolol/therapeutic use , Adult , Aged , Aged, 80 and over , Anxiety/drug therapy , Arachidonic Acid/pharmacology , Blood Platelets/drug effects , Depression/drug therapy , Double-Blind Method , Female , Heart Rate/drug effects , Humans , Male , Medication Therapy Management , Middle Aged , Placebos , Prospective Studies , Systole/drug effects
17.
Med J Aust ; 210(11): 509-516, 2019 06.
Article in English | MEDLINE | ID: mdl-31155728

ABSTRACT

INTRODUCTION: Heparin-induced thrombocytopenia (HIT) is a prothrombotic disorder that occurs following the administration of heparin and is caused by antibodies to platelet factor 4 and heparin. Diagnosis of HIT is essential to guide treatment strategies using non-heparin anticoagulants and to avoid unwanted and potential fatal thromboembolic complications. This consensus statement, formulated by members of the Thrombosis and Haemostasis Society of Australia and New Zealand, provides an update on HIT pathogenesis and guidance on the diagnosis and management of patients with suspected or confirmed HIT. MAIN RECOMMENDATIONS: A 4Ts score is recommended for all patients with suspected HIT prior to laboratory testing. Further laboratory testing with a screening immunoassay or confirmatory functional assay is not recommended in individuals with a low 4Ts score. However, if there are missing or unreliable clinical data, then laboratory testing should be performed. A positive functional assay result confirms the diagnosis of HIT and should be performed to confirm a positive immunoassay result. Heparin exposure must be ceased in patients with suspected or confirmed HIT and initial treatment with a non-heparin alternative instituted. Non-heparin anticoagulants (danaparoid, argatroban, fondaparinux and bivalirudin) used to treat HIT should be given in therapeutic rather than prophylactic doses. Direct oral anticoagulants may be used in place of warfarin after patients with HIT have responded to alternative parenteral anticoagulants with platelet count recovery. CHANGES IN MANAGEMENT AS A RESULT OF THIS STATEMENT: These are the first Australasian recommendations for diagnosis and management of HIT, with a focus on locally available diagnostic assays and therapeutic options. The importance of examining both clinical and laboratory data in considering a diagnosis of HIT cannot be overstated.


Subject(s)
Anticoagulants/adverse effects , Heparin/adverse effects , Thrombocytopenia/diagnosis , Thrombocytopenia/drug therapy , Anticoagulants/therapeutic use , Australia , Consensus , Hemorrhage/chemically induced , Heparin/immunology , Humans , New Zealand , Platelet Factor 4/immunology , Receptors, IgG/immunology , Thrombocytopenia/chemically induced , Thrombosis/etiology
18.
Res Pract Thromb Haemost ; 2(1): 125-138, 2018 Jan.
Article in English | MEDLINE | ID: mdl-30046713

ABSTRACT

BACKGROUND: N-methyl-d-aspartate receptors (NMDARs) contribute calcium influx in megakaryocytic cells but their roles remain unclear; both pro- and anti-differentiating effects have been shown in different contexts. OBJECTIVES: The aim of this study was to clarify NMDAR contribution to megakaryocytic differentiation in both normal and leukemic cells. METHODS: Meg-01, Set-2, and K-562 leukemic cell lines were differentiated using phorbol-12-myristate-13-acetate (PMA, 10 nmol L-1) or valproic acid (VPA, 500 µmol L-1). Normal megakaryocytes were grown from mouse marrow-derived hematopoietic progenitors (lineage-negative and CD41a-enriched) in the presence of thrombopoietin (30-40 nmol L-1). Marrow explants were used to monitor proplatelet formation in the native bone marrow milieu. In all culture systems, NMDARs were inhibited using memantine and MK-801 (100 µmol L-1); their effects compared against appropriate controls. RESULTS: The most striking observation from our studies was that NMDAR antagonists markedly inhibited proplatelet formation in all primary cultures employed. Proplatelets were either absent (in the presence of memantine) or short, broad and intertwined (with MK-801). Earlier steps of megakaryocytic differentiation (acquisition of CD41a and nuclear ploidy) were maintained, albeit reduced. In contrast, in leukemic Meg-01 cells, NMDAR antagonists inhibited differentiation in the presence of PMA and VPA but induced differentiation when applied by themselves. CONCLUSIONS: NMDAR-mediated calcium influx is required for normal megakaryocytic differentiation, in particular proplatelet formation. However, in leukemic cells, the main NMDAR role is to inhibit differentiation, suggesting diversion of NMDAR activity to support leukemia growth. Further elucidation of the NMDAR and calcium pathways in megakaryocytic cells may suggest novel ways to modulate abnormal megakaryopoiesis.

20.
Platelets ; 29(8): 793-800, 2018 Dec.
Article in English | MEDLINE | ID: mdl-29090586

ABSTRACT

MYH9-related disorders (MYH9-RDs) caused by mutation of the MYH9 gene which encodes non-muscle myosin heavy-chain-IIA (NMMHC-IIA), an important motor protein in hemopoietic cells, are the most commonly encountered cause of inherited macrothrombocytopenia. Despite distinguishing features including an autosomal dominant mode of inheritance, giant platelets on the peripheral blood film accompanied by leucocytes with cytoplasmic inclusion bodies (döhle-like bodies), these disorders remain generally under-recognized and often misdiagnosed as immune thrombocytopenia (ITP). This may result in inappropriate treatment with corticosteroids, immunosupressants and in some cases, splenectomy. We explored the efficacy of next generation sequencing (NGS) with a candidate gene panel to establish the aetiology of thrombocytopenia for individuals who had been referred to our center from hematologists in the Australasian region in whom the cause of thrombocytopenia was suspected to be secondary to an inherited condition but which remained uncharacterized despite phenotypic investigations. Pathogenic MYH9 variants were detected in 15 (15/121, 12.4%) individuals and the pathogenecity of a novel variant of uncertain significance was confirmed in a further two related individuals following immunofluorescence (IF) staining performed in our laboratory. Concerningly, only one (1/17) individual diagnosed with MYH9-RD had been referred with this as a presumptive diagnosis, in all other cases (16/17, 94.1%), a diagnosis was not suspected by referring clinicians, indicating a lack of awareness or a failing of our diagnostic approach to these conditions. We examined the mean platelet diameter (MPD) measurements as a means to better identify and quantify platelet size. MPDs in cases with MYH9-RDs were significantly larger than controls (p < 0.001) and in 91% were greater than a previously suggested threshold for platelets in cases of ITP. In addition, we undertook IF staining in a proportion of cases and confirm that this test and/or NGS are satisfactory diagnostic tests. We propose that fewer cases of MYH9-RDs would be missed if diagnostic algorithms prioritized IF and/or NGS in cases of thrombocytopenia associated with giant platelets, even if döhle-like bodies are not appreciated on the peripheral blood film. Finally, our report describes the long-term use of a thrombopoietin agonist in a case of MYH9-RD that had previously been diagnosed as ITP, and demonstrates that treatment with these agents may be possible, and is well tolerated, in this group of patients.


Subject(s)
Blood Platelets/metabolism , Hearing Loss, Sensorineural/genetics , Mutation , Myosin Heavy Chains/genetics , Purpura, Thrombocytopenic, Idiopathic/genetics , Receptors, Fc/therapeutic use , Recombinant Fusion Proteins/therapeutic use , Thrombocytopenia/congenital , Thrombopoietin/therapeutic use , Adult , Australasia , Blood Platelets/drug effects , Blood Platelets/pathology , Cell Size , Cohort Studies , Diagnosis, Differential , Female , Gene Expression , Gene Expression Profiling , Genes, Dominant , Hearing Loss, Sensorineural/blood , Hearing Loss, Sensorineural/diagnosis , Hearing Loss, Sensorineural/drug therapy , High-Throughput Nucleotide Sequencing , Humans , Inclusion Bodies/drug effects , Inclusion Bodies/metabolism , Inclusion Bodies/pathology , Male , Mean Platelet Volume , Middle Aged , Myosin Heavy Chains/blood , Purpura, Thrombocytopenic, Idiopathic/blood , Purpura, Thrombocytopenic, Idiopathic/diagnosis , Purpura, Thrombocytopenic, Idiopathic/drug therapy , Thrombocytopenia/blood , Thrombocytopenia/diagnosis , Thrombocytopenia/drug therapy , Thrombocytopenia/genetics
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