Increased thrombin generation in a mouse model of cancer cachexia is partially interleukin-6 dependent.

Essentials Cancer cachexia and cancer-associated thrombosis have not previously been mechanistically linked. We assessed thrombin generation and coagulation parameters in cachectic C26 tumor-bearing mice. C26 mice are hypercoagulable, partially corrected by blocking tumor derived interleukin-6. Coagulability and anti-inflammatory interventions may be clinically important in cancer cachexia. SUMMARY: Background Cancer cachexia and cancer-associated thrombosis are potentially fatal outcomes of advanced cancer, which have not previously been mechanistically linked. The colon 26 (C26) carcinoma is a well-established mouse model of complications of advanced cancer cachexia, partially dependent on high levels of interleukin-6 (IL-6) produced by the tumor. Objectives To assess if cancer cachexia altered the coagulation state and if this was attributable to tumor IL-6 production. Methods In male BALB/c*DBA2 (F1 hybrid) mice with a C26 tumor we used modified calibrated automated thrombogram and fibrin generation (based on overall hemostatic potential) assays to assess the functional coagulation state, and also examined fibrinogen, erythrocyte sedimentation rate (ESR), platelet count, tissue factor pathway inhibitor (TFPI) and hepatic expression of coagulation factors by microarray. C26 mice were compared with non-cachectic NC26, pair-fed and sham control mice. IL-6 expression in C26 cells was knocked down by lentiviral shRNA constructs. Results C26 mice with significant weight loss and highly elevated IL-6 had elevated thrombin generation, fibrinogen, ESR, platelets and TFPI compared with all control groups. Fibrin generation was elevated compared with pair-fed and sham controls but not compared with NC26 tumor mice. Hepatic expression of coagulation factors and fibrinolytic inhibitors was increased. Silencing IL-6 in the tumor significantly, but incompletely, attenuated the increased thrombin generation, fibrinogen and TFPI. Conclusions Cachectic C26 tumor-bearing mice are in a hypercoagulable state, which is partly attributable to IL-6 release by the tumor. The findings support the importance of the coagulation state in cancer cachexia and the clinical utility of anti-inflammatory interventions.

New oral anticoagulants: a practical guide on prescription, laboratory testing and peri-procedural/bleeding management. Australasian Society of Thrombosis and Haemostasis.

New oral anticoagulants (NOAC) are becoming available as alternatives to warfarin to prevent systemic embolism in patients with non-valvular atrial fibrillation and for the treatment and prevention of venous thromboembolism. An in-depth understanding of their pharmacology is invaluable for appropriate prescription and optimal management of patients receiving these drugs should unexpected complications (such as bleeding) occur, or the patient requires urgent surgery. The Australasian Society of Thrombosis and Haemostasis has set out to inform physicians on the use of the different NOAC based on current available evidence focusing on: (i) selection of the most suitable patient groups to receive NOAC, (ii) laboratory measurements of NOAC in appropriate circumstances and (iii) management of patients taking NOAC in the perioperative period, and strategies to manage bleeding complications or 'reverse' the anticoagulant effects for urgent invasive procedures.

Acute pulmonary embolism during warfarin therapy and long-term risk of recurrent fatal pulmonary embolism.

The clinical characteristics and long-term outcomes of patients presenting with acute pulmonary embolism (PE) during treatment with warfarin have not been described. Clinical details of all patients admitted to a tertiary institution from 2000-2007 with acute PE were retrieved retrospectively, baseline warfarin status and the international normalised ratio (INR) were recorded, and their outcomes tracked using a statewide death registry. Of 923 patients with clearly documented warfarin status included in this study, 83 (9%) were taking warfarin. Mean (± standard deviation) day-1 INR of those taking warfarin was 2.3 ± 0.9, with 67% of patients therapeutically anti-coagulated (INR ≥2.0) at presentation (49 patients with INR <2.5 and 34 with INR ≥2.5). Patients taking warfarin on admission were more likely to have heart failure, atrial fibrillation and valvular heart disease, with similar prevalence of malignancy and ischaemic heart disease, compared to patients not on warfarin. Total mortality of the cohort (mean follow-up 4.0 ± 2.5 years) was 31.6% (in-hospital mortality 1.5%), and was similar between warfarin and no warfarin groups. There was however a greater than four-fold increased risk of post-discharge death due to recurrent PE for the patients taking warfarin on admission (hazard ratio [HR] 4.43, 95% confidence interval [CI] 1.36-14.42, p=0.01). Among patients taking warfarin on admission, day-1 INR <2.5 significantly increased long-term all-cause mortality compared to INR ≥2.5 (adjusted HR 2.51, 95% CI 1.08-5.86, p=0.03). In conclusion, patients presenting with PE during treatment with warfarin have an increased risk of death from recurrent PE. Admission INR appears to have independent long-term prognostic importance in these patients.

The use of FEIBA® in the correction of coagulation abnormalities induced by dabigatran.

INTRODUCTION: Studies have shown dabigatran to be an effective anticoagulant with an acceptable bleeding profile. None the less, these patients do suffer from bleeding complications. Unfortunately, there are currently no direct reversal agents to dabigatran or established guidelines on the management of bleeding in these circumstances. METHODS: We examined the effects on thrombin generation parameters, after ex-vivo spiking the plasma of patients on dabigatran (n = 8) with FEIBA(®). These parameters were measured using the calibrated automated thrombography (CAT) machine. RESULTS: In our study, we showed the ability of FEIBA(®) to improve the abnormal thrombin generation parameters caused by dabigatran in these patients. CONCLUSION: This provides evidence, lacking in the literature, that this agent may be able to provide haemostatic support in situations where dabigatran induced coagulopathy exists.

Reduced fibrinolysis and increased fibrin generation can be detected in hypercoagulable patients using the overall hemostatic potential assay.

BACKGROUND: Routinely available coagulation assays are not capable of detecting clinically defined hypercoagulable states. A number of global coagulation assays have been developed with the potential to evaluate hypercoagulability, which predisposes to the common clinical events of arterial and venous thromboembolism (VTE). OBJECTIVES: We hypothesized that the overall hemostatic potential (OHP) assay would show abnormal fibrin generation and lysis in patients with clinically defined hypercoagulable states. METHODS: We used the OHP assay as described by Blombäck and colleagues [1,2] in 161 clinically hypercoagulable patients with arterial or VTE, pregnancy complications or autoimmune disease. Eighty patients had associated antiphospholipid antibodies (APLA). Ninety-eight normal plasma donors were tested for comparison. RESULTS: We derived three new assay parameters for correlation with hypercoagulable states: the maximum optical density, maximum slope, and delay in onset of fibrin generation. We found significantly different assay results for all patients' parameters examined when compared with controls, indicating both increased fibrin generation and reduced fibrinolysis in hypercoagulable patients. The findings were similar whether samples were collected in association with an acute thrombotic event or not. Estimated assay sensitivity for detection of a clinically defined hypercoagulable state was 96%. CONCLUSIONS: The OHP assay is a simple, inexpensive global test that is useful for assessing patients with hypercoagulable states including APLA. OHP results are significantly abnormal in hypercoagulable groups compared with controls, indicating that both increased fibrin generation and reduced fibrinolysis contribute to hypercoagulable states. The assay may ultimately assist in tailoring clinical management to patients' individual requirements.

Vaginal algometer: development and application of a device to monitor vaginal wall pressure pain threshold.

We describe a device (which we have named a vaginal algometer) which can measure the pressure pain threshold (PPT) on the lateral walls of the vagina. The device was assessed in 63 healthy women and a normal range for this measurement was established. Each woman had her vaginal wall PPT measured and was asked about the acceptability of the procedure. We demonstrate that the vaginal algometer can provide a quantitative assessment of vaginal PPT and that the procedure is acceptable to most women.

The relation between Ashworth scale scores and the excitability of the alpha motor neurones in patients with post-stroke muscle spasticity.

BACKGROUND: The modified Ashworth scale (MAS) is the most widely used method for assessing muscle spasticity in clinical practice and research. However, the validity of this scale has been challenged. OBJECTIVES: To compare the MAS with objective neurophysiological tests of spasticity. METHODS: The MAS was recorded in patients with post-stroke lower limb muscle spasticity and correlated with the excitability of the alpha motor neurones. The latter was evaluated by measuring the latency of the Hoffmann reflex (H reflex) and the ratio of the amplitude of the maximum H reflex (H(max)) to that of the compound action motor potential of the soleus muscle (M(max)). RESULTS: Data on 24 randomly recruited patients were analysed. Patients were divided into two groups according to their MAS score: 14 had a MAS score of 1 (group A) and 10 scored 2 (group B). The two groups were comparable with respect to age and sex, but in group A there was a longer period since the stroke. The H reflex latency was reduced and the H(max):M(max) ratio was increased in both groups. The H(max):M(max) ratio values were higher for group B but the differences were not statistically significant. CONCLUSIONS: There is a relation between the MAS scores and alpha motor neurone excitability, although it is not linear. This suggests that the MAS measures muscle hypertonia rather than spasticity.

Presentation by myoblasts of an epitope from endogenous acetylcholine receptor indicates a potential role in the spreading of the immune response.

It is generally considered that myoblasts are unable to prime naive T cell responses without help from professional antigen-presenting cells (APC). However, their ability to present endogenous antigens to previously primed T lymphocytes in the secondary phase of a T cell response has not been well studied. We show here that primary human myoblasts, when stimulated with IFNgamma to express class II MHC, can present an endogenous epitope, probably an acetylcholine receptor (AChR) peptide, to a CD4(+) AChR-specific T helper lymphocyte clone. Presentation leads to secretion of IFNgamma by the T cell clone and, in addition, killing of the myoblast. Our results suggest that, during the effector phase of the immune response, myoblasts could enhance the inflammatory response by presenting endogenous antigen, and thereby become targets for CD4(+) T lymphocyte-induced cytotoxicity; subsequent release of myoblast antigens could then lead to inter- and intra-molecular determinant spreading.

Restricted IgG1 subclass of anti-Yo antibodies in paraneoplastic cerebellar degeneration.

Paraneoplastic cerebellar degeneration (PCD) occurs as a non-metastatic manifestation of cancer in a small proportion of patients with certain breast or gynaecological tumours, and is characterised by widespread Purkinje cell loss. Antibodies against a Purkinje cell cytoplasmic antigen, called Yo, that is expressed by the tumours, are present in the majority of these patients, but the pathogenic role of the antibodies is not clear. To characterise further the immune response in these cases, 13 anti-Yo positive sera were tested for IgG subclasses by immunohistochemistry and western blotting and, in four cases, PHA-stimulated cytokine secretion by peripheral blood lymphocytes was measured. Surprisingly, anti-Yo antibodies were entirely restricted to the IgG1 subclass, whereas antibodies against the small cell cancer-associated antigen, Hu, were found in all four IgG subclasses. There was a trend towards raised IgG1 levels in the total IgG of the anti-Yo positive patients and, in two, PHA-stimulated peripheral blood lymphocytes secreted raised levels of IFN-gamma. By contrast, in the other two cases tested, raised levels of IL-4 were secreted. Patients with PCD associated with anti-Yo antibodies appear to have strong immune responses that are polarised with respect to the IgG subclass and Th cytokine profiles.

Receiver operating characteristic analysis for intelligent medical systems--a new approach for finding confidence intervals.

Intelligent systems are increasingly being deployed in medicine and healthcare, but there is a need for a robust and objective methodology for evaluating such systems. Potentially, receiver operating characteristic (ROC) analysis could form a basis for the objective evaluation of intelligent medical systems. However, it has several weaknesses when applied to the types of data used to evaluate intelligent medical systems. First, small data sets are often used, which are unsatisfactory with existing methods. Second, many existing ROC methods use parametric assumptions which may not always be valid for the test cases selected. Third, system evaluations are often more concerned with particular, clinically meaningful, points on the curve, rather than on global indexes such as the more commonly used area under the curve. A novel, robust and accurate method is proposed, derived from first principles, which calculates the probability density function (pdf) for each point on a ROC curve for any given sample size. Confidence intervals are produced as contours on the pdf. The theoretical work has been validated by Monte Carlo simulations. It has also been applied to two real-world examples of ROC analysis, taken from the literature (classification of mammograms and differential diagnosis of pancreatic diseases), to investigate the confidence surfaces produced for real cases, and to illustrate how analysis of system performance can be enhanced. We illustrate the impact of sample size on system performance from analysis of ROC pdf's and 95% confidence boundaries. This work establishes an important new method for generating pdf's, and provides an accurate and robust method of producing confidence intervals for ROC curves for the small sample sizes typical of intelligent medical systems. It is conjectured that, potentially, the method could be extended to determine risks associated with the deployment of intelligent medical systems in clinical practice.

Environmental risk factors for sporadic Escherichia coli O157 infection in Scotland: results of a descriptive epidemiology study.

The objective of the study was to obtain detailed descriptive epidemiological information on sporadic verocytotoxin-producing Escherichia coli O157 infection in Scotland in relation to transmission routes and host-related risk factors. Using a standardized questionnaire, the study was carried out throughout Scotland over an 18-month period from July 1992 and co-ordinated at the Communicable Diseases and Environmental Health (Scotland) Unit, Glasgow (CDEH(S)U). The subjects were laboratory-confirmed cases of Escherichia coli O157 infection, of whom 138 met the criteria for inclusion in the study. The most important findings were the high proportion of cases who had been exposed to environmental factors such as farm animals and/or their by-products; or who had participated in gardening or garden-play; or who had suspected or confirmed household water supply problems, prior to the onset of illness. The frequency and relative importance of environmental risk factors requires further quantification and study in order to assess where control measures can be directed most effectively. The implications for the NHS in preventing this crippling, life-threatening infection are considerable, not least in relation to hospitalization, dialysis and renal transplantation costs.

Determinant spreading and immune responses to acetylcholine receptors in myasthenia gravis.

In myasthenia gravis (MG), antibodies to the muscle acetylcholine receptor (AChR) cause muscle weakness. Experimental autoimmune myasthenia gravis (EAMG) can be induced by immunisation against purified AChR; the main immunogenic region (MIR) is a conformation-dependent site that includes alpha 67-76. EAMG can also occur after immunisation against extracellular AChR sequences, but this probably involves intramolecular determinant spreading. In MG patients, thymic hyperplasia and germinal centres are found in about 50%, and thymoma in 10-15%. The heterogeneous, high affinity, IgG anti-AChR antibodies appear to be end-products of germinal centre responses, and react mainly with the MIR or a site on fetal AChR; the latter contains a gamma subunit and is mainly expressed on myoid cells in the thymic medulla. T cells cloned against recombinant AChR subunits recognise principally two naturally processed epitopes: epsilon 201-219 derived from adult AChR which is expressed in muscle, and sometimes in thymic epithelium, and alpha 146-160, common to fetal and adult AChR. Since AChR is not normally co-expressed with class II, it is unclear how CD4+ responses to AChR alpha and epsilon subunits are initiated, and how and where these spread to induce antibodies against fetal AChR. Various possibilities, including upregulation of class II on muscle/myoid cells and involvement of CD8+ responses to AChR and other muscle antigens, are discussed.

A pathogenetic role for the thymoma in myasthenia gravis. Autosensitization of IL-4- producing T cell clones recognizing extracellular acetylcholine receptor epitopes presented by minority class II isotypes.

Myasthenia gravis (MG) is caused by helper T cell-dependent autoantibodies against the muscle acetylcholine receptor (AChR). Thymic epithelial tumors (thymomas) occur in 10% of MG patients, but their autoimmunizing potential is unclear. They express mRNAs encoding AChR alpha and epsilon subunits, and might aberrantly select or sensitize developing thymocytes or recirculating peripheral T cells against AChR epitopes. Alternatively, there could be defective self-tolerance induction in the abundant maturing thymocytes that they usually generate. For the first time, we have isolated and characterized AChR-specific T cell clones from two MG thymomas. They recognize extracellular epitopes (alpha75-90 and alpha149-158) which are processed very efficiently from muscle AChR. Both clones express CD4 and CD8alpha, and have a Th-0 cytokine profile, producing IL-4 as well as IFN-gamma. They are restricted to HLA-DP14 and DR52a; expression of these minority isotypes was strong on professional antigen-presenting cells in the donors' tumors, although it is generally weak in the periphery. The two clones' T cell receptor beta chains are different, but their alpha chain sequences are very similar. These resemblances, and the striking contrasts with T cells previously cloned from non-thymoma patients, show that thymomas generate and actively induce specific T cells rather than merely failing to tolerize them against self antigens.

Epidemiology of infection due to Escherichia coli O157: a 3-year prospective study.

A 3-year study of Escherichia coli infections in Grampian Region was conducted to ascertain the incidence, document clinical sequelae and identify at-risk groups. Approximately 30,000 stools from patients with acute diarrhoea were screened for E. coli O157, and an epidemiological questionnaire filled in for each patient whose stool was positive. Eighty-three patients were studied. The annual incidence was 6 per 100, 000. Proportionately more infections occurred in people involved in agriculture. Evidence was seen of case-to-case transmission, and contamination of a water supply. Eight cases developed haemolytic uraemic syndrome (HUS). There were 2 deaths due to HUS and 2 due to haemorrhagic colitis (HC). Symptomatic E. coli infection is relatively common in the Grampian Region, more common in the agricultural community, and is the main cause of HUS in this Region.

Scottish national survey of tuberculosis notifications 1993 with special reference to the prevalence of HIV seropositivity.

BACKGROUND: The study sought to determine the contribution of HIV seropositivity to the arrest of decline in tuberculosis notifications in Scotland. METHODS: Survey forms relating to each tuberculosis notification in 1993 were completed by the notifying consultant. Voluntary anonymous HIV testing of tuberculosis cases aged under 65 was requested. Age, sex, ethnic status, country of birth, employment status, occupation, previous tuberculosis, contact status, risk factors for HIV infection, HIV serostatus of cases aged under 65, site, radiological extent, and bacteriological status of tuberculous disease were determined. RESULTS: Five hundred and seventy four cases of tuberculosis were originally notified, of which 77 (14%) subsequently proved to be non-tuberculous and were therefore denotified. Of the 497 cases 423 (85%) were white and 58 (12%) were from the Indian subcontinent. Eighty five per cent of patients from the Indian subcontinent were aged < 55 years whereas 64% of white patients were aged > 55 years. Pulmonary disease was found in 74%, non-pulmonary in 22%, and combined disease in 4% of patients. Of 242 HIV tests performed, three were positive and five other HIV positive patients were known, giving an HIV positivity rate of 1.6% of all tuberculosis notifications in 1993. Annual notification rates for Scotland were 9.7 per 10(5) before and 8.7 per 10(5) after exclusion of previously treated cases; rates were 8.4 per 10(5) for the white population and 179 per 10(5) for those from the Indian subcontinent. CONCLUSIONS: The study documents the distribution of tuberculous disease in Scotland by age, sex, site, and ethnic group for the first time. Notification practices, with respect to denotification, need to be improved. Infection with HIV is presently uncommon in cases of tuberculosis in Scotland but continued vigilance is essential.

T cell development in TCR-alpha beta transgenic mice. Analysis using V(D)J recombination substrates.

The major pathway of intrathymic T cell differentiation leads CD4-8- (DN) T lineage-committed precursors to TCR-alpha beta+ CD4+8- or CD4-84+ (SP) T lymphocytes. The expression of functionally rearranged TCR-alpha beta transgenes (Tg-TCR) may influence thymocyte development by affecting the various selection events that control T cell differentiation. To gain insights into these processes, we have produced double transgenic animals carrying V(D)J recombination substrates in addition to the MHC class I (H-2Kb) allospecific KB5C20 Tg-TCR. We have analyzed substrate rearrangements in purified populations of Tg-TCR+ thymocytes in the situation of positive or negative selection. The profile of rearrangements found in SP thymocytes, positively selected for the Tg-TCR, suggests that expression of the KB5C20 Tg-TCR has only a minimal influence on substrate V(D)J recombination in cells differentiating along the major alpha beta T cell developmental pathway. In contrast, Tg-TCR+ DN thymocytes, in both positively and negatively selecting haplotypes, presented a profile that implies premature cessation of substrate rearrangements. This profile was maintained in peripheral Tg-TCR+ DN cells and was distinct from the one found in CD25+, alpha beta+, or gamma delta+ DN cells purified from mice transgenic for the recombination substrates only. These results are discussed with respect to the possible origin and differentiation pathway of Tg-TCR+ DN and SP cells.

Four channel foetal ECG data collection system.

Recently there has been an increased interest in the development of improved techniques for the diagnosis of foetal distress during labour. Many of the techniques have been based on extracting extra information from the foetal ECG obtained from a scalp electrode. To fully develop and test the prototypes of these systems requires recorded data from patients. However due to the poor level of prediction of these cases at present, it is very difficult to collect the data using simple single channel data collection systems. The system described here will automatically collect and document data from up to four deliveries at the same time and does not add to the work load of the clinical staff.