Circulating microvesicles are less procoagulant and carry different miRNA cargo in myelodysplasia.

BACKGROUND AND AIMS: Myelodysplasia (MDS) is characterised by abnormal haematopoiesis and increased risk of bleeding. Microvesicles (MV) play a key role in coagulation and their impact in MDS is unknown. METHODS: Platelet free plasma from 35 red-cell transfusion-dependent MDS patients and 15 controls were analysed. Pro-coagulant function was assessed by the XaCT assay and by thrombin generation (ETP). Total MV were enumerated by nano-tracking analysis. MV subsets were quantified by flow cytometry after staining with specific antibodies for various endovascular cell types. Small RNA was quantitated and sequenced. The MV measurements were correlated with MDS clinical risk scores and level of transfusion dependence. RESULTS: The pro-coagulant function of MV was significantly lower in MDS. All the MV subtypes, as measured by flow cytometric markers, were also significantly lower. The small RNA and miRNA cargo were significantly higher in MDS. The miRNA profile showed that mir-28 and mir-LETD7 were under expressed whilst mir-584J and mir-4485 were over expressed in MV from MDS. CONCLUSIONS: Circulating MV in MDS show reduced pro-coagulant functional activity, reduced subtypes by flow cytometry and significantly different miRNA content. However, the levels or subtypes of MV did not predict the clinical phenotype or level of transfusion dependence.

All-trans-retinoic acid, idarubicin, and IV arsenic trioxide as initial therapy in acute promyelocytic leukemia (APML4).

The treatment of acute promyelocytic leukemia has improved considerably after recognition of the effectiveness of all-trans-retinoic acid (ATRA), anthracycline-based chemotherapy, and arsenic trioxide (ATO). Here we report the use of all 3 agents in combination in an APML4 phase 2 protocol. For induction, ATO was superimposed on an ATRA and idarubicin backbone, with scheduling designed to exploit antileukemic synergy while minimizing cardiotoxicity and the severity of differentiation syndrome. Consolidation comprised 2 cycles of ATRA and ATO without chemotherapy, followed by 2 years of maintenance with ATRA, oral methotrexate, and 6-mercaptopurine. Of 124 evaluable patients, there were 4 (3.2%) early deaths, 118 (95%) hematologic complete remissions, and all 112 patients who commenced consolidation attained molecular complete remission. The 2-year rate for freedom from relapse is 97.5%, failure-free survival 88.1%, and overall survival 93.2%. These outcomes were not influenced by FLT3 mutation status, whereas failure-free survival was correlated with Sanz risk stratification (P[trend] = .03). Compared with our previously reported ATRA/idarubicin-based protocol (APML3), APML4 patients had statistically significantly improved freedom from relapse (P = .006) and failure-free survival (P = .01). In conclusion, the use of ATO in both induction and consolidation achieved excellent outcomes despite a substantial reduction in anthracycline exposure. This trial was registered at the Australian New Zealand Clinical Trials Registry (www.anzctr.org.au) as ACTRN12605000070639.

Thrombotic microangiopathies, thrombotic thrombocytopenic purpura, and ADAMTS-13.

Thrombotic microangiopathy (TMA) is a term used to describe a group of disorders characterized by hemolytic anemia (with prominent red blood cell fragmentation), thrombocytopenia, and thrombosis in the microvasculature. It may be used when describing patients with thrombotic thrombocytopenic purpura (TTP), hemolytic uremic syndrome, atypical hemolytic uremic syndrome, as well as a myriad of other disorders in which the TMA may be secondary to another disease or disorder. While limited information exists as to the exact cause of microthrombosis in many TMA, recent advances have been made in the understanding of TTP and its pathophysiology. This progress can be attributed to discovery of the von Willebrand factor cleaving protease ADAMTS-13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13), whose absence in TTP has given the disorder a distinct molecular identity. The discovery of this metalloprotease has prompted a significant amount of research relating to its role in TTP as well as its general function in hemostasis. The exact mechanisms by which this metalloprotease achieves its role are slowly being understood and these now provide other avenues by which TMA may occur.

Implications of a needs assessment intervention for people with progressive cancer: impact on clinical assessment, response and service utilisation.

OBJECTIVE: To assess the impact of the systematic use of the Palliative Care Needs Assessment Guidelines and Needs Assessment Tool: Progressive Disease-Cancer (NAT: PD-C) on clinical assessment, response and service utilisation. STUDY SETTING: Three major oncology treatment centres in NSW, Australia. STUDY DESIGN: Between March 2007 and December 2009, 219 people with advanced cancer were recruited to complete bi-monthly telephone interviews. The intervention, introduced after at least two baseline interviews, involved training health professionals to complete the NAT: PD-C with patients approximately monthly. DATA COLLECTION: Rates of service use and referrals were compared pre- and post-introduction of the NAT: PD-C. Rates of completion of the tool; its impact on consultation length; and the types of needs and follow-up care to address these were also assessed. PRINCIPAL FINDINGS: The NAT: PD-C had a high rate of completion; identified needs consistent with those self-reported by patients in interviews; and did not alter consultation length. No changes in the number of health professionals seen by patients were found pre- and post-intervention. CONCLUSION: The NAT: PD-C is an efficient and acceptable strategy for supporting needs-based cancer care that can potentially be incorporated into standard routine care without increasing the burden on care providers.

The risk of coronary thrombosis with cyclo-oxygenase-2 inhibitors does not vary with polymorphisms in two regions of the cyclo-oxygenase-2 gene.

AIMS: To investigate whether polymorphisms of the cyclo-oxygenase-2 (COX-2) gene modify the adverse cardiovascular effects of COX-2 inhibitors. METHODS: A case control study was conducted in the Hunter Region of New South Wales, Australia. Cases (n= 460) were hospitalized with acute coronary syndrome (ACS). Controls (n= 640) were recruited from the electoral rolls. Structured interviews gathered information on variables including recent ingestion of non-steroidal anti-inflammatory drugs (NSAIDs). Targeted genotyping of rs 20417(G > C) and rs5275 (T > C) polymorphisms was performed by real-time polymerase chain reaction using allele-specific probes. RESULTS: Ingestion of any NSAID in the week prior to interview was associated with an elevated risk for ACS: adjusted odds ratio 1.8 (1.2, 2.5). The rs 20417 and rs 5275 polymorphisms were not singly associated with risk for ACS: adjusted odds ratios 1.1 (0.80, 1.5) and 1.2 (0.88, 1.5), respectively. Individually, the polymorphisms did not modify the risk of ACS with the drugs. When analyses were conducted by haplotype, the adjusted odds ratio with celecoxib or rofecoxib in individuals who had one or two copies of the 'low risk' haplotype (no GT) was 1.2 (0.29, 5.0), compared with 2.1 (1.1, 4.0) with the 'high risk' haplotype (one or two copies of GT). CONCLUSIONS: We found little evidence of a gene/drug interaction. We found a statistically non-significant trend toward a lower risk of coronary events with NSAIDs in the presence of the 'low risk' haplotype. Even if confirmed, the clinical utility of the finding would be limited as this haplotype is carried by a minority of the population.

Thrombin generation as a predictor of radiotherapy induced skin erythema.

BACKGROUND AND PURPOSE: Biological mechanisms underlying radiation induced erythema remain largely unknown, with no simple way to accurately predict or prevent extreme cases. Based on the recent findings in patients suffering from chronic urticaria, we sought to determine if similar mechanisms of hypercoagulation contributed to comparable skin reactions during radiotherapy. MATERIALS AND METHODS: Plasma levels of prothrombin factor 1+2 (F1+2), D-dimers and plasminogen activator inhibitor-1 (Pai-1) were tested in 32 women undergoing irradiation following breast conserving surgery for early breast cancer. Reflectance spectrophotometry was used to objectively assess erythema throughout the treatment by measuring the amount of light reflected from the skin surface as a function of wavelength. Correlations between peak levels of erythema and plasma biomarkers were then assessed. RESULTS: Individual peak reflectance readings generally occurred between day 29 of treatment and 2 weeks post radiotherapy, and represented a median increase of 66% (range: 11-146%; p<0.001) from baseline. Peak reflectance correlated with F1+2 and Pai-1 levels measured both at baseline and day 29 of treatment, and multivariate analysis indicated that these two baseline measurements were the best predictors of peak reflectance, accounting for 59% of the variability in erythema (p=0.000004). CONCLUSIONS: Patients with signs of intravascular thrombin generation are at higher risk of radiotherapy-induced skin reactions, providing a new therapeutic avenue for possibly predicting and preventing this side effect of cancer treatment.

Circulating microvesicles in snakebite patients with microangiopathy.

BACKGROUND: Venom-induced consumption coagulopathy is a common consequence of snake envenoming that can lead to life-threatening hemorrhage, and is associated with microangiopathic hemolytic anemia (MAHA), acute kidney injury and thrombocytopenia. The role of microvesicles (MV) in snakebite patients has not been previously investigated. OBJECTIVE: To compare changes in subsets of circulating MV levels in snakebite patients with venom induced consumption coagulopathy and with or without microangiopathic hemolysis to those of healthy controls. METHODS: This study used samples from patients recruited to the Australian Snakebite Project (ASP) with snake envenoming, including bites by brown snakes, tiger snakes, and taipans. Citrated blood from envenomed patients was collected, processed, and stored according to a national standardized protocol. Full blood count and coagulation parameters were measured as per routine clinical care and blood films were examined for evidence of hemolysis. Baseline coagulation parameters were measured on a Behring Coagulation System. Flow cytometry was performed to detect CD41a (platelet), CD62e (endothelial), and glycophorin (red cell) MV. The results were analyzed using BD software and appropriate statistical tools. RESULTS AND CONCLUSIONS: The red cell MV in snakebite patients with MAHA (n = 13) were significantly higher than those without MAHA (n = 17) while there was no significant difference in platelet MV levels between the snakebite patients with and without MAHA. Interestingly, the endothelial MV were reduced in all snakebite patient samples compared to the control samples. Measuring red cell MV at presentation could be useful as a predictive marker for MAHA in patients with snakebites.

Microparticles in health and disease.

Microparticles (MPs) are small fragments of membrane-bound cytoplasm that are shed from the surface of an activated or apoptotic cell. Recently, their function as vectors of transcellular exchange of biologic information, in addition to better described forms of intercellular communication such as growth factors, cytokines, and chemokines, has become well recognized. Circulating levels of MPs are thought to reflect a balance between cell stimulation, proliferation, and death. The production of MPs is thought to predominately occur by vesiculation or blebbing of the cell membrane. The mechanisms governing the remodeling of the plasma membrane are complex, involving cytoskeletal changes and a shift from normal phospholipid asymmetry. Increased intracellular calcium subsequent to cell activation leads to intracellular increases in several proteins including gelsolin and calpain, as well as the activity of enzymes such as translocase, floppase, and scramblase, which play important roles in the homeostasis of the cell membrane. The membrane vesiculation and phospholipids asymmetry leading to the production of MPs occurs by the complex interplay of the proteins involved. There are several clinical conditions associated with elevated MPs, and most are also associated with an increased risk of thrombosis. Apart from cardiovascular disease and venous thromboembolism, MPs are also elevated in solid tumors with metastatic disease. The measurement of MPs is being regarded as a potential biomarker, given the range of conditions in which they are elevated and the association with prothrombotic states. The utility of measuring MPs as a diagnostic and prognostic marker is currently a subject of intense investigation. The further development of the various methods for the detection and measurement of MPs and prospective clinical trials establishing the utility of such tests will be critical prior to the routine measurement of MPs in the diagnostic laboratory.

Polymorphisms of the tissue factor pathway inhibitor gene are associated with venous thromboembolism in the antiphospholipid syndrome and carriers of factor V Leiden.

Polymorphisms within the tissue factor pathway inhibitor (TFPI) gene may determine TFPI expression and increase the risk of venous thromboembolism (VTE) in predisposed individuals. We tested this hypothesis by comparing TFPI activity and the frequency of common TFPI polymorphisms, -33T->C, -399C->T and -287T->C, in patients with antiphospholipid syndrome (APS) (n = 24) or factor V Leiden (n = 44) who had a history of VTE (n = 26), compared with those without VTE (n = 42) and also with normal control individuals (n = 56). TFPI activity was measured using a modified amidolytic assay and genotypes were determined by polymerase chain reaction and restriction fragment length polymorphism. We found that only APS patients with a history of venous thrombosis had TFPI activity levels significantly different from control individuals (1.77 +/- 0.60 vs 0.77 +/- 0.19 U/ml; P = 0.0001), and this was associated with inheritance of the TFPI -33C allele (1.70 +/- 0.72 U/ml for TC/CC genotypes vs 0.97 +/- 0.56 U/ml for TT; P = 0.01). Multivariate analysis of APS and factor V Leiden patients revealed that the greatest independent contributor to VTE was TFPI activity (adjusted odds ratio = 16.84; 95% confidence interval = 2.47-114.36, P = 0.004), while inheritance of either the TFPI -33C or -399T alleles each increased the odds of VTE by nearly 13 times (95% confidence interval = 2.39-69.91, P = 0.003; and 95% confidence interval = 2.25-71.23, P = 0.004, respectively). These results indicate that the TFPI -33T->C and -399C->T polymorphisms are significantly associated with venous thrombosis in the presence of other risk factors, especially APS, and may be clinically relevant in patients who are prone to hypercoagulability.

Circulating microvesicle number, function and small RNA content vary with age, gender, smoking status, lipid and hormone profiles.

BACKGROUND: Characterization of circulating microvesicles (MV) in healthy subjects in relation to various biological factors is not well studied. OBJECTIVES: We evaluated the influence of age, gender, smoking status, lipid and hormone profiles on circulating MV in healthy subjects. METHODS: Platelet free plasma from 143 volunteer blood donors (males=80, females=63) was evaluated by standardized flow cytometry for MV expressing CD41 (platelet-derived), CD105 (endothelial-derived), CD235 (red cell-derived), TF (tissue factor) and phosphatidylserine (PS) MV. Procoagulant function was measured by the Xa based assay (XaCT) and endogenous thrombin potential (ETP) using thrombin generation assay. RESULTS: Those ≤29years and ≥60years had higher levels of MV subsets (CD41, CD235, TF and PS) compared to those aged 30-59years. The median CD41, CD105, CD235, TF and PS expressing MV by flow cytometry were similar or lower in females, whilst procoagulant activity by the XaCT assay was higher (p=0.002). In smokers (n=21), certain MV subsets (CD41, TF and PS) and functional activity (ETP) was lower (p<0.05). Regression analysis showed that MV parameters of CD41, CD105, TF and ETP could be predicted independently by age, whilst smoking predicted for CD105, CD235, TF, PS and ETP. Certain MV parameters also correlated with BMI, lipid and hormone levels. The small RNA and miRNA levels did not differ by age group, smoking status or gender. CONCLUSIONS: It is important to recognize that differences may arise depending on age, gender, BMI, lipid, hormone levels and smoking status in apparently healthy subjects when evaluating MV for pathogenic potential.

Correlative analysis of nanoparticle tracking, flow cytometric and functional measurements for circulating microvesicles in normal subjects.

INTRODUCTION: Circulating microvesicles (MV) can be analysed using a number of different techniques. The aim of this study was to evaluate the correlation between functional procoagulant based assays including thrombin generation, factor Xa activation test (XaCT), and phosphatidylserine factor Xa-activity by ELISA with optical MV enumeration by flow cytometry and nanoparticle tracking analysis. METHODS: Citrated blood samples were collected from 60 healthy volunteer blood donors after informed consent. Platelet free plasma was prepared using a standardized published protocol. MV subsets were enumerated by flow cytometry (BDFACS Canto) after staining with specific antibodies for platelets (CD41), endothelial cells (CD105), red cells (CD235) monocytes (CD14), tissue factor (CD142) and for phosphatidylserine expression by binding to annexin V. A standardized protocol using counting beads was employed. Nanotracking analysis was performed on both scatter and fluorescent settings after MV staining with quantum dot stain, Qdot 655. Procoagulant function was assessed by the XaCT assay on an automated coagulation analyser and by thrombin generation assay measuring endogenous thrombin potential (ETP), lagtime, peak (PEAK) and time to peak (ttPEAK) using a Calibrated Automated Thrombogram (CAT). The statistical analysis was carried out with Statistica 12 software using non-parametric tests (Spearman rank order correlations, with significance set at p<0.05). RESULTS: In normal healthy subjects, thrombin generation parameters correlated with levels of MV measured by flow cytometry. ETP, lagtime, ttPEAK and PEAK correlated with MV expressing phosphatidylserine (rs, Spearman rank order correlation was 0.29, 0.40, 0.31 and 0.34 respectively, p<0.05), and MV expressing tissue factor (rs was 0.29, 0.40, 0.31 and 0.34 respectively, p<0.05), whilst red cell derived MV correlated with lagtime, ttPEAK and PEAK (rs, was 0.35,0.30 and 0.3, respectively, p<0.05). Lagtime and ttPEAK negatively correlated with the clot based XaCT test (rs, was -0.34 and -0.30 respectively, p<0.05) and positively correlated with the ELISA MP-activity assay (rs=0.42 for both, p<0.05). In addition, endothelial MV levels weakly correlated with white cell counts (rs = 0.27, p<0.05). CONCLUSIONS: Thrombin generation and flow cytometry for phosphatidylserine or tissue factor expressing MV correlate well as markers for procoagulant activity. A combination of optical or non-optical enumeration as well as functional methods may be required for a complete profiling of circulating MV.

An imatinib-only window followed by imatinib and chemotherapy for Philadelphia chromosome-positive acute leukemia: long-term results of the CMLALL1 trial.

We report long-term results in 40 patients with Philadlephia chromosome-positive (Ph+) acute leukemia who received an imatinib monotherapy window to evaluate in vivo effects on BCR-ABL signaling prior to induction chemotherapy. The first 25 patients (cohort 1) received the LALA-94 protocol without further imatinib (newly diagnosed Ph+ acute lymphoblastic leukemia [ALL]) or induction chemotherapy followed by single-agent imatinib. Subsequent patients (cohort 2) continued imatinib concurrently with either LALA-94 (newly diagnosed Ph + ALL) or other intensive chemotherapy regimens. Cohort 2 had a complete response (CR) rate of 93% and 5-year survival of 69%. For newly diagnosed Ph+ ALL, survival was superior in cohort 2 compared with cohort 1. Toxicity was similar to that expected for chemotherapy alone. Among 10 evaluable patients, rapid loss of phospho-CRKL occurred during the imatinib window in seven patients (all achieved CR) and incomplete inhibition in three patients (none with CR). In summary, a pharmacodynamic window design permitted biomarker assessment of BCR-ABL targeting without compromising clinical outcomes.

Use of arsenic trioxide in remission induction and consolidation therapy for acute promyelocytic leukaemia in the Australasian Leukaemia and Lymphoma Group (ALLG) APML4 study: a non-randomised phase 2 trial.

BACKGROUND: Initial treatment of acute promyelocytic leukaemia traditionally involves tretinoin (all-trans retinoic acid) combined with anthracycline-based risk-adapted chemotherapy, with arsenic trioxide being the treatment of choice at relapse. To try to reduce the relapse rate, we combined arsenic trioxide with tretinoin and idarubicin in induction therapy, and used arsenic trioxide with tretinoin as consolidation therapy. METHODS: Patients with previously untreated genetically confirmed acute promyelocytic leukaemia were eligible for this study. Eligibilty also required Eastern Cooperative Oncology Group performance status 0-3, age older than 1 year, normal left ventricular ejection fraction, Q-Tc interval less than 500 ms, absence of serious comorbidity, and written informed consent. Patients with genetic variants of acute promyelocytic leukaemia (fusion of genes other than PML with RARA) were ineligible. Induction comprised 45 mg/m(2) oral tretinoin in four divided doses daily on days 1-36, 6-12 mg/m(2) intravenous idarubicin on days 2, 4, 6, and 8, adjusted for age, and 0·15 mg/kg intravenous arsenic trioxide once daily on days 9-36. Supportive therapy included blood products for protocol-specified haemostatic targets, and 1 mg/kg prednisone daily as prophylaxis against differentiation syndrome. Two consolidation cycles with tretinoin and arsenic trioxide were followed by maintenance therapy with oral tretinoin, 6-mercaptopurine, and methotrexate for 2 years. The primary endpoints of the study were freedom from relapse and early death (within 36 days of treatment start) and we assessed improvement compared with the 2 year interim results. To assess durability of remission we compared the primary endpoints and disease-free and overall survival at 5 years in APML4 with the 2 year interim APML4 data and the APML3 treatment protocol that excluded arsenic trioxide. This study is registered with the Australian New Zealand Clinical Trials Registry, number ACTRN12605000070639. FINDINGS: 124 patients were enrolled between Nov 10, 2004, and Sept 23, 2009, with data cutoff of March 15, 2012. Four (3%) patients died early. After a median follow-up of 4·2 years (IQR, 3·2-5·2), the 5 year freedom from relapse was 95% (95% CI 89-98), disease-free survival was 95% (89-98), event-free survival was 90% (83-94), and overall survival was 94% (89-97). The comparison with APML3 data showed that hazard ratios were 0·23 (95% CI 0·08-0·64, p=0·002) for freedom from relapse, 0·21 (0·07-0·59, p=0·001) for disease-free survival, 0·34 (0·16-0·69, p=0·002) for event-free survival, and 0·35 (0·14-0·91, p=0·02) for overall survival. INTERPRETATION: Incorporation of arsenic trioxide in initial therapy induction and consolidation for acute promyelocytic leukaemia reduced the risk of relapse when compared with historical controls. This improvement, together with a non-significant reduction in early deaths and absence of deaths in remission, translated into better event-free and overall survival. FUNDING: Phebra.

Fatigue during breast radiotherapy and its relationship to biological factors.

INTRODUCTION: Fatigue is commonly reported in patients receiving radiotherapy for breast conservation, but the underlying mechanisms remain unclear. METHODS: Patients with early breast cancer participating in a prospective study of the impact of inflammatory processes on early and delayed breast morbidity were assessed for fatigue levels using the Functional Assessment of Cancer Therapy (FACT) fatigue subscale before and at intervals during and after radiotherapy. Blood for analysis of a variety of circulating cytokines, coagulation factors, peripheral blood indices and biochemical factors was collected at the same time points. RESULTS: Fifty-two eligible patients were assessed. Twenty-one patients (43%) developed significant fatigue during radiotherapy, whereas 28 (54%) developed minimal or no fatigue. Fatigue appeared to plateau between week 4 of treatment and 2 weeks after treatment. The fatigue was beginning to settle by 6 weeks after treatment. Significant fatigue was predicted by a higher baseline fatigue score, red cell count, neutrophil count, and D-dimer level. Baseline fatigue correlated with higher body mass index, C-reactive protein, soluble thrombomodulin, tissue plasminogen activator, von Willebrand factor antigen, interleukin-6, ICAM-1, hemoglobin and red cell, monocyte, and neutrophil counts. There were also significant correlations between body mass index and tissue plasminogen activator, C-reactive protein, interleukin-6, ICAM-1, and red cell count. After these factors were controlled for, baseline fatigue was seen to be associated with higher body mass index, soluble thrombomodulin, tissue plasminogen activator, von Willebrand factor antigen, monocyte count, and neutrophil count. Multiple logistic regression procedures indicated that the most predictive factors for fatigue during radiotherapy were higher baseline fatigue level and higher baseline neutrophil and red cell counts. CONCLUSION: This study has shown that significant fatigue is common in patients receiving breast irradiation and is precipitated during radiotherapy in some patients but not others. The factors shown to be associated with fatigue in this study will be helpful in shaping future studies.

Induction with oral chemotherapy (CID) followed by early autologous stem cell transplantation for de novo multiple myeloma patients.

In an effort to minimise induction therapy-related toxicity, avoid unnecessary hospitalisation and facilitate early ASCT, we have prospectively evaluated an outpatient-based oral chemotherapeutic regimen, cyclophosphamide, idarubicin, dexamethasone (CID) in patients with previously untreated multiple myeloma (mm). Patients subsequently underwent ASCT conditioned with melphalan 200 g/m(2). A total of 36 newly diagnosed MM patients were enrolled between February 1997 and March 2000. In all 136 cycles of CID were administered requiring only four unplanned hospital admissions. Grade 3 or 4 haematological toxicities were documented following 14 cycles (10%). There were no treatment-related deaths. Response rates (PR + CR) based on an intention-to-treat basis were 66% (23 of 35, 9% CR) post-CID and 80% (28 of 35, 34% CR) post-ASCT. In all, 28 of the 35 patients remain alive with a median follow-up for surviving patients of 40 months (range, 30-67 months). Progression-free survival from the time of diagnosis was 32 months (range, 3-55+ months). Median overall survival from diagnosis has not been reached with an estimated overall survival at 4 years of 77%. CID in combination with early ASCT is an effective and well-tolerated antimyelomatous regimen and is a practical alternative to more toxic and invasive therapeutic approaches.

Reduction of prothrombin and Factor V levels following supplementation with omega-3 fatty acids is sex dependent: a randomised controlled study.

BACKGROUND: LCn-3PUFA comprised of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) offer cardioprotection involving a decrease in coagulant activity; however, the evidence is equivocal. We have previously demonstrated that the acute (24 h) effects and chronic (4 weeks) effects of LCn-3PUFA supplementation on platelet aggregation in human subjects are sex specific. This study investigated the mechanisms of the sex-dependent effects of LCn-3PUFA with 4 weeks supplementation of EPA-rich vs. DHA-rich oils on procoagulant and platelet activity in healthy subjects. DESIGN: A double-blinded, placebo-controlled randomised trial was conducted in 94 healthy adults: male (n=41) and female (n=53). Platelet coagulation parameters including factors I, II, V, VII, VIII, IX, X, vWF:Ag and endogenous thrombin potential were measured at baseline and 4 weeks postsupplementation with EPA-rich or DHA-rich oil capsules. RESULTS: We have previously reported that platelet aggregation is specifically reduced by supplementation with EPA in males and DHA in females. This sex-specific effect was also observed for decreases in plasma levels of Factor II (-7.9 ± 3.8%, P=.026), Factor V (-6.5 ± 4.5%, P=.022) and vWF:Ag (-7.3 ± 2.1%, P=.034) and was most pronounced in males supplemented with EPA. In contrast, DHA-mediated reduction in platelet aggregation in females was not accompanied by any significant changes in the coagulation parameters tested. CONCLUSION: Significant interactions between sex and specific LCn-3PUFA exist to reduce procoagulant activity differentially in males vs. females and could have profound effects on managing risk of thrombotic disease.

Comparative sensitivity of commercially available aPTT reagents to mulga snake (Pseudechis australis) venom.

This study aimed to determine the relative sensitivity of activated partial thromboplastin time (aPTT) reagents to the anticoagulant effects of phospholipases in mulga snake (Pseudechis australis) venom.Twenty-one haematology laboratories participating in the Royal College of Pathologists of Australasia Quality Assurance Programs were sent human plasma samples spiked with mulga venom (n=25 total results). Results for 17 patients with mulga snake envenoming were available through the Australian Snakebite Project.Only 12 of 25 venom spiked samples returned an abnormally prolonged aPTT. Tests performed with Dade Actin FS (n=7) did not identify any of the spiked samples as abnormal. Although clotting times were significantly prolonged using the lupus anticoagulant sensitive Actin FSL (n=5, p=0.043), only one was reported as abnormal. Only laboratories using TriniCLOT aPTT S (n=6), HemosIL APTT SP (n=2) and Stago PTT-A (n=1) consistently recorded the spiked sample as being above the upper normal reference interval. Abnormally prolonged aPTTs were recorded for four of eight patients whose tests were performed with Actin FSL, five of eight patients with TriniCLOT aPTT HS, and three of three patients using TriniCLOT aPTT S.We conclude that some reagents used for routine aPTT testing are relatively insensitive to the anticoagulant effects of mulga snake venom. Tests performed with these reagents should be interpreted with caution.