Low frequency of septic arthritis after arthrocentesis and intra-articular glucocorticoid injection.

Objective: The aim of this study was to evaluate the risk of septic arthritis (SA) in patients who received an intra-articular (IA) glucocorticoid (GC) injection and to describe the characteristics of these patients. Methods: All patients undergoing IA procedures at the orthopaedic and rheumatological departments on the Danish island of Funen from January 2006 to December 2013 were identified in the central database and included by register extraction. Patients who developed a clinically inflamed joint and positive synovial fluid culture within 14 days after IA GC injection were considered as having SA. Retrospectively, data on age, gender, affected joint location, bacterial agent, pre-existing inflammatory disorder, and death within 30 days were extracted from the patient files. According to local recommendations, a non-touch sterile technique was used for IA procedures. Patients were informed about the risk of SA and advised to seek medical attention on suspicion of infection or lack of improvement. Results: In total, 22 370 IA procedures were performed. Among these, 14 118 GC injections and 8252 arthrocenteses were undertaken. Only 11 patients were diagnosed with SA (0.08%, 95% confidence interval 0.03-0.12). Risk factors for SA were male gender, age, and pre-existing joint disease. Conclusion: We found a low frequency of SA subsequent to IA GC injections. Older patients with pre-existing joint disease are at higher risk of developing SA.

Prospective role of indigenous Exiguobacterium profundum PT2 in arsenic biotransformation and biosorption by planktonic cultures and biofilms.

AIMS: The aim of this study was to analyse arsenic (As) transformation and biosorption by indigenous As-resistant bacteria both in planktonic and biofilm modes of growth. METHODS AND RESULTS: As-resistant bacteria were isolated from industrial waste water and strain PT2, and identified as Exiguobacterium profundum through 16S rRNA gene sequencing was selected for further study. As transformation and biosorption by E. profundumPT2 was determined by HPLC-ICP-MS analysis. Planktonic cultures reduced 3·73 mmol l-1 As5+ into As3+ from artificial waste water effluent after 48-h incubation. In case of biosorption, planktonic cultures and biofilms exhibited 25·2 and 29·4 mg g-1 biomass biosorption, respectively. As biosorption kinetics followed Freundlich isotherm and pseudo second-order model. Biofilm formation peaked after 3 days of incubation, and in the presence of As stress, biofilm formation was significantly affected in contrast to control (P < 0·05). Homogeneous nature of mature biofilms with an increased demand of nutrients was revealed by minimum roughness and maximum surface to biovolume ratio measured through CLSM analysis. CONCLUSION: Indigenous As-resistant E. profundumPT2 was found capable of As transformation and biosorption both in the form of planktonic cultures and biofilms. SIGNIFICANCE AND IMPACT OF THE STUDY: Indigenous biofilm forming E. profundum PT2 revealing As biosorption and biotransformation potential is presented an eco-friendly and cost-effective source for As remediation that can be implemented for waste water treatment.

Prognostic factors associated with mortality in patients with septic arthritis: a descriptive cohort study.

OBJECTIVES: To evaluate the 30-day mortality rate of septic arthritis (SA) in adults in Funen, central Denmark, and to explore whether, at the time of SA presentation, risk factors for the 30-day mortality rate could be revealed. Our secondary objective was to describe the microbiological aetiologies, systemic signs of inflammation, and co-morbidity. METHOD: A descriptive study identifying patients with SA from central Denmark, during the period 2006-2013, by the use of joint fluid culture data retrieved from the electronic database at the Department of Clinical Microbiology, Odense University Hospital. Patients with a positive joint fluid culture were considered eligible and their medical records were examined. RESULTS: We identified 215 patients with SA, mean age 64.8 years. At presentation, mean C-reactive protein (CRP) was 204 mg/L, mean white blood cell count (WBC) 11.9 × 109/L, and mean body temperature 37.6°C. A total of 101 patients (47%) had a prosthetic joint, 46 (21%) had an inflammatory joint disease, and 24 (11%) had diabetes mellitus (DM). Staphylococcus aureus was the most common pathogen (104 patients, 48.4%). The 30-day mortality rate was 9.3% and the significant risk factor for death was liver disease at time of presentation [odds ratio (OR) 40.40, 95% confidence interval (CI) 5.38-303]. The other factors tested such as age > 65 years, elevated temperature, rheumatoid arthritis (RA), prostheses, and diabetes mellitus (DM) did not reach statistical significance. CONCLUSIONS: In our sample of patients with SA, we found a 30-day mortality rate in almost one in 10 adults. Among possible explanations, our study indicates that liver disease is a clinically relevant risk factor.

Web-based cognitive training for breast cancer survivors with cognitive complaints-a randomized controlled trial.

BACKGROUND: Cognitive complaints are common amongst breast cancer survivors, and no standard treatment exists. The present study evaluates whether web-based cognitive training can alleviate subjectively reported and objectively assessed cognitive complaints in a sample of breast cancer survivors. The primary and secondary outcomes were an objective measure of working memory and a measure of perceived cognitive functioning. Additional outcomes were neuropsychological tests of memory, executive function, working memory and questionnaire-based assessment of anxiety, depression and somatization. METHODS: A total of 157 female breast cancer survivors were recruited from an existing cohort and through announcements in open access cancer-related Internet fora and randomly allocated to either web-based cognitive training (eCogT) with telephone support (n = 94) or a waitlist control (WLC) condition (n = 63). eCogT encompassed 30 training sessions over 6 weeks. Neuropsychological assessments were undertaken over the telephone, and questionnaire data was collected online. Data was collected at baseline, post-intervention and at 5-month follow-up. RESULTS: Mixed linear models revealed no statistically significant change in primary or secondary outcome at follow-up in either group. Statistically significant improvements (p 0.040-0.043) were found in the eCogT group for verbal learning and on a working memory test. CONCLUSIONS: Web-based cognitive training did not result in improvements of the primary or secondary outcome. Improved performance was observed on verbal learning and working memory. These effects were observed at 5-month follow-up, indicating long-term effects of training. The intervention may be applied in a clinical setting at low cost and without risk of adverse effects.© 2016 The Authors Psycho-Oncology Published by John Wiley & Sons Ltd.

Level of Adherence to Prophylactic Osteoporosis Medication amongst Patients with Polymyalgia Rheumatica and Giant Cell Arteritis: A Cross-Sectional Study.

Objective. To estimate level of adherence to oral calcium and vitamin D supplementation as well as bisphosphonate amongst patients with PMR and GCA treated with glucocorticoids. Method. A total of 138 patients with the diagnosis of PMR and/or GCA registered in our department in December 2013. In this cross-sectional study we interviewed all the patients to measure level of adherence to calcium and vitamin D, as well as bisphosphonates. Results. Out of the 118 included patients, 88.9% of them were adherent to their prescription. Only 2 patients (1.7%) did not take calcium and vitamin D at all and 10 patients (8.5%) took their medication infrequently, 9 and 1 out of 10 patients took the medication 50-100% of the time and less than 50% of the prescribed dose, respectively. Sixty-one patients received additional treatment with bisphosphonate and 96.6% were adherent to this therapy. The remaining 3.4% of the patients did not take the medication at all. Forgetfulness, adverse side effects, and lack of understanding of treatment benefits were the most significant causes for nonadherence to calcium and vitamin D. Conclusions. Contrary to what we expected this study found that adherence to osteoporosis preventive medication in patients with PMR and GCA was high.

Recombinant activated factor VII (rFVIIa) acutely normalizes prothrombin time in patients with cirrhosis during bleeding from oesophageal varices.

BACKGROUND: Patients with cirrhosis have low levels of coagulation factors, the most pronounced deficiency being that of FVII. This may compromise haemostasis during bleeding from ruptured oesophageal varices. The objective of this trial was to evaluate the effect of rFVIIa on prothrombin time in cirrhosis patients with ongoing variceal bleeding. Safety, including signs of DIC, was monitored. METHODS: The study is a single centre, open-label trial. Ten consecutive patients with known alcoholic cirrhosis and oesophageal variceal bleeding were included. The patients received routine treatment, including Terlipressin. Each patient received one i.v. injection of rFVIIa (80 microg/kg bw). The study observation time was 12 h per patient. RESULTS: The mean age of the patients was 48 years (8 men and 2 women). The cirrhosis was classified as Child B in 5 patients and Child C in 5. At baseline, all patients had prothrombin time levels above the normal range, and all but one had FVII coagulation activity (FVII:C) levels below the normal range. rFVIIa normalized the prothrombin time in all patients within 30 min. The effect lasted for more than 4 h in 7 patients, and for about 2 h in the remaining 3 patients. Immediate bleeding control was obtained in all patients, and no patient died within the study time. There was no sign of DIC. CONCLUSIONS: rFVIIa is effective in transiently reversing the prolonged prothrombin time in cirrhosis patients with haematemesis from varices. This indicates a potential of improving haemostasis and survival in patients with compromised coagulation due to liver disease.

Potential role of NovoSeven in the prevention of rebleeding following aneurysmal subarachnoid haemorrhage.

Rebleeding following aneurysmal subarachnoid haemorrhage is a major factor contributing to unfavourable outcome. Antifibrinolytic agents reduce the rate of rebleeding but increase the risk of cerebral ischaemia and infarction and hence provide no overall benefit. To address the theoretical concern that recombinant activated factor VII (NovoSeven, Novo Nordisk A/S, Bagsvaerd, Denmark) might increase the risk of cerebral ischaemia while stabilizing the clot at the site of aneurysmal rupture, an open-label, dose-escalation safety study has been developed in collaboration with the UK Spontaneous Intracranial Haemorrhage Group. The trial design includes the recruitment of 15 patients (aged 18 years or over) in good grade with subarachnoid haemorrhage verified by computerized tomography scan or lumbar puncture. Safety evaluation includes clinical observation, monitoring of laboratory variables, positron emission tomography (PET) scanning (rCBF, rOEF, rCMRO2) and transcranial Doppler ultrasound. To date, ten patients have been recruited [NovoSeven 80 microg/kg single bolus (n = 2), NovoSeven 80 microg/kg single bolus followed by continuous infusion at 3.5 microg/kg per h (n = 2) or 7 microg/kg per h (n = 1), or control (n = 5)]. Clinical observation, transcranial Doppler ultrasound and PET studies revealed no evidence of cerebral ischaemia in the first nine patients treated with NovoSeven. The last patient developed middle cerebral artery branch thrombosis contralateral to the aneurysm. The study is currently suspended pending further investigation.

Effect of the administration of recombinant activated factor VII (rFVIIa; NovoSeven) in the management of severe uncontrolled bleeding in patients undergoing heart valve replacement surgery.

Recombinant activated factor VII (rFVIIa; NovoSeven, Novo Nordisk A/S, Bagsvaerd, Denmark) is being increasingly used to secure haemostasis in difficult clinical situations. The role of rFVIIa in the treatment of patients undergoing open-heart surgery for valvular heart disease was evaluated in an open pilot study. Study objectives included evaluation of blood loss, haemostatic effect and safety and laboratory parameters following rFVIIa administration. To date, we have treated five patients (one child aged 2.5 years and four adults) undergoing surgical procedures including arterial switch, closure of atrial septal defect and De Vega's procedure (mitral valve replacement with tricuspid valve repair). Four patients received rFVIIa intraoperatively, while the fifth received it postoperatively. Satisfactory haemostasis was achieved with a single dose (30 microg/kg) of rFVIIa. Four hours after treatment mean blood loss was 262.5 ml for adults (220-334 ml) and 85 ml for the child. No significant adverse events were reported. Laboratory parameters indicated a mean 18.5-fold (range 3.7-42) increase in FVII levels at 30 min postinjection and a mean reduction of 12 s (range 3-39 s) in prothrombin time. In conclusion, rFVIIa represents an effective and well-tolerated treatment for serious bleeding episodes both during cardiac surgery and postoperatively.

Influence of exogenous factor VIIa on thrombin generation in cord plasma of full-term and pre-term newborns.

Factor (F) VIIa has been used to treat adults and children with a variety of bleeding disorders. The results from these studies cannot be extrapolated to newborns because their hemostatic system differs significantly from adults, which may influence the effects of FVIIa on thrombin (IIa) generation. We compared the effects of FVIIa concentrates on IIa generation in plasmas from adults, full-term newborns and pre-term newborns. Defibrinated plasma (using arvin) from adults, or umbilical cords from full-term or pre-term deliveries was supplemented with FVIIa (Novo Nordisk, Bagsvaerd, Denmark), mixed with dilute thromboplastin reagent, and the resultant reaction mixture subsampled periodically into ethylenediamine tetraacetic acid, followed by measurement of total IIa activity (S-2238). Thrombin-alpha2 macroglobulin complexes, determined as residual activity after neutralization with heparin and antithrombin, were subtracted from total IIa to give free IIa. Prothrombin (FII) and inhibitor complexes were measured by enzyme-linked immunosorbent assays. Addition of FVIIa caused a reduction in the lag phase for the appearance of free IIa and consumption of FII, which was more pronounced in newborn plasma. There was no increase in peak IIa levels regardless of the amount of FVIIa added. Final inhibitor complex concentrations were increased in plasmas from adults compared with newborns, likely reflecting higher plasma concentrations of FII in adults. Generation of IIa was more rapid in pre-term plasma compared with that in adult and full-term cord plasmas due to increased endogenous tissue factor (TF). In summary, FVIIa enhanced IIa generation in plasma from different age groups, with the effect being more pronounced in plasma from pre-term newborns, possibly due to increased levels of plasma TF.

Decline of factor VIII and factor IX inhibitors during long-term treatment with NovoSeven.

Recombinant factor VIIa (rFVIIa) (NovoSeveng) is used to treat bleeding episodes in hemophilia A and B patients with inhibitor antibodies against factor VIII (FVIII) and factor IX. rFVIIIa has been studied in home treatment of mild-to-moderate joint, muscle, and mucocutaneous bleeds to assess safety and efficacy. Treatment with other factor concentrates was allowed according to treating physician's judgment. Blood samples were drawn before study start and after 6 and 12 months. It has thus been possible to follow the inhibitor titres during this period. Analyses of 53 patients (49 hemophilia A, four hemophilia B) showed inhibitor levels up to 1,208 BU/ml before study start. Based on the first analysis, hemophilia A patients were divided into high responders (> 5 BU/ml; 28 patients), low responders (> 1 and < 5 BU/ml; 15 patients) and very low responders (< or = 1 BU/ml; six patients). In high responders receiving rFVIIa as only treatment, FVIII inhibitor titre decreased to one-third of the initial level. For high responders receiving other factor treatments such as FVIII or prothrombin complex concentrates, inhibitor titre remained unchanged. Titres for low responders and very low responders remained unchanged independent of treatment. Thus, when rFVIIa is used as the only coagulation factor to treat hemophilia A/B high-responder inhibitor patients, inhibitor level declines significantly.

An approach for characterization and purification of a human monoclonal hybridoma antibody.

An IgG human-human monoclonal hybridoma antibody, AML-19, reactive with human myeloid cells of non-malignant and malignant origin has been produced by fusion of blood mononuclear cells from a patient with acute myeloid leukemia (AML) and the human B-lymphoma cell line RH-L4. The monoclonal antibody (MAb) AML-19 was purified from hybridoma supernatant by primarily anion-exchange chromatography, in order to separate the AML-19 MAb from contaminating immunoglobulin (Ig), e.g. bovine Ig and MAb derived from the parental fusion partner, and followed by immunoaffinity chromatography. This purification method gave the highest yield and purity of the AML-19 MAb. The isoelectric point (pI) of the MAb was estimated to be 5. Inhibition assays indicate an apparent dissociation constant (Kd) corresponding to 4 x 10(-9) M and an affinity constant (Ka) to 2.5 x 10(8)M-1 to K562 erythroleukemia cells. Scatchard plot demonstrated a linear slope as a manifestation of monoclonality and a low number of AML-19 specific epitopes, estimated to 1500 per cell.

Identification and characterization of an antigen specific for normal erythroid precursor cells and its application in diagnosis of erythroleukemia.

A monoclonal antibody (designated K:1-6F) generated by hybridization of mouse myeloma cells with spleen cells from mice immunized with the erythroleukemic cell line K562 was found by fluorescence-activated cell sorter analysis, dot-blot assay and electroimmunoblotting to bind to a majority of cells in the K562 and HEL erythroleukemic cell lines, to a subset of cells of the erythroid lineage from normal bone marrow, to a subset of cells in all analysed cases (total 10) of erythroleukemia, and weakly to cells from patients with myeloid leukemia. The antibody did not bind to normal erythrocytes, monocytes, T- and B lymphocytes or granulocytes, as well as a panel of human malignant cell lines of hemopoietic origin (HL60, U937, Daudi, Molt-3, RH-L4 and U266). Biochemical characterization of the antigen defined by the antibody suggests that eht epitope is defined by a carbohydrate structure alone or in combination with proteins. Four molecules with Mr 100 kD, 65 kD, 45 kD and 18 kD respectively were immunoprecipitated from Triton X-100 extract of K562 erythroleukemia cells. Neuraminidase did not affect the binding of the antibody, whereas tunicamycin reduced the K:1-6F expression. The K:1-6F Mab was in normal bone marrow found to be specific for erythroid precursor cells and may therefore be useful in examination of normal and leukemic erythropoiesis.

Specificity and diagnostic implications of the reactivity pattern of a panel of monoclonal antibodies against myeloid leukemia cells.

The immunological phenotypes of leukemia cell samples from 60 patients, of whom 54 had acute myeloid leukemia (AML), were assessed with a panel of monoclonal antibodies (Mabs) with specificity for the following epitopes: Epitopes associated with myeloid leukemia cells, Epitopes expressed only on immature myeloid cells (or subsets) and on monocytes, Epitopes only expressed on granulocytes or on granulocytes and mature myeloid cells (promyelocytes, myelocytes and monocytes), Epitopes on HLA-class II (DR) and HLA-class I molecules and on insulin receptors. This panel of Mabs proved useful to identify leukemia cells in blood and to assess their myeloid origin. The panel of Mabs was found also to be useful for immunophenotyping of leukemia cells. Furthermore, the analysis revealed considerable variations in the immunological phenotype of AML cells, reflecting antigenic heterogeneity within the individual leukemia cell population as well as abnormal or no expression of histocompatibility antigens and insulin receptors in some samples. Some of the Mabs bound preferentially to subgroups in the French-American-British (FAB) classification.

Antibody-producing human-human hybridomas. III. Derivation and characterization of two antibodies with specificity for human myeloid cells.

Peripheral blood mononuclear cells from a patient with acute myeloid leukemia (AML) and spleen cells from a patient with chronic myeloid leukemia (CML) were fused with HAT-sensitive human B lymphoma cells (RH-L4) in attempts to generate human monoclonal antibodies (Mab) against antigens with high specificity for myeloid leukemia cells. Forty-seven of 246 hybridomas secreted Ig that bound to AML cell surface constituents, as determined by FACS analysis of viable cells that were FITC-stained with the human Mab as the first-step reagent and FITC-conjugated rabbit anti-human Ig as second-step. Two of the 47 human Mab (one from each patient and designated AML-19 and CML-20, respectively) bound to both autologous and allogeneic myeloid leukemia cells. No significant binding was observed to cell surface constituents on human bone marrow cells, granulocytes, lymphocytes, erythrocytes, thymocytes, monocytes, lymphoblastic leukemia cells, fibroblasts, malignant B and T lymphocytic cell lines, and murine bone marrow cells. Both human Mab were IgG and were cytotoxic to myeloid leukemia cells in the presence of complement. About 70% of peripheral blood cell samples from 46 AML patients contained AML-19- and CML-20-positive cells, but the reactivity pattern had no correlation to the morphologic FAB classification of the samples. The promyelocytic HL60 cell line and the K562 cell line reacted with the two antibodies. Dot blot analysis of binding of AML-19 and CML-20 to cellular extracts immobilized on nitrocellulose paper showed that both human Mab in this assay also reacted with normal bone marrow cells. This was supported by microscopic immunofluorescence because both human Mab stained intracytoplasmatic structures in normal bone marrow cells, but both intracytoplasmatic and cell surface components stained in myeloid leukemia cells. Moreover, immunoblotting demonstrated that both human Mab in leukemia cells reacted with two cellular proteins with Mr approximately 14,500 and 18,000, and in normal bone marrow cells with a molecule with Mr approximately 20,000. Immunoprecipitation of cell membrane molecules with both the AML-19 and CML-20 antibody precipitated from leukemic cells only the molecule with Mr approximately 18,000 and no components from normal bone marrow cells. It is concluded that myeloid leukemogenesis may result in generation of cell surface expression of either new or abnormally processed molecules that are immunogenic in the autochthonous host. These molecules may also be useful as markers in diagnosis of myeloid leukemia.

Immune phenotype heterogeneity in AML.

Blood cells from 46 patients with acute myeloid leukaemia were studied for expression of various surface markers, defined by a panel of 12 monoclonal antibodies and the expression of Fc gamma receptors. Corresponding studies were done on normal bone marrow cells. Antibodies which bound to leukaemic cells in high frequencies were those which most frequently also bound to normal bone marrow cells. Immunophenotypic analysis revealed a marked antigenic heterogeneity in AML, also evident within single FAB subclasses. However, leukaemic cells of FAB subclass M1 significantly more often expressed HLA class I antigen than those of FAB subclass M5a, whereas Fc gamma receptors which were expressed only on a few cells in M5a, were increasingly frequent on leukaemic cells of M1-M2, M4, and M5b leukaemias. The frequency of cells reacting with the monoclonal antibody T50/12,11,2 was related to the complete remission rate of the patients. Patients with high frequencies of cells reacting with this antibody had a better complete remission rate than patients with fewer cells binding to this antibody. The immunophenotypic heterogeneity an AML may reflect a great biological variability of this disease. This variability may be of importance for the classification and treatment of AML.

A monoclonal antibody (NAT-9 II:3F-6F) that identifies a differentiation antigen on human myeloid cells.

A monoclonal antibody, designated NAT-9 II:3F-6F (IgM), was generated by hybridization of mouse myeloma cells with spleen cell from mice immunized with normal human bone marrow cells. The antibody reacted with 40-60% of bone marrow cells as analysed on samples from 40 normal individuals and only with a subpopulation of human acute myeloid leukemia (AML) cells of the M2 class (20/20 tested) and M4 class (12/12 tested) (subclasses of the French-American-British (FAB) classification), but not with leukemic cells of the M1 (0/12 tested) and M5 (0/12 tested) FAB subclasses. This is in contrast to many other myeloid-specific monoclonal antibodies. Fluorescence-activated cell sorter (FACS) analyses and morphological examination of cells stained with peroxidase as based on the NAT-9 II:3F-6F monoclonal antibody showed that this antibody reacted with a distant differentiation antigen which is absent on myeloblasts, but expressed on promyelocytes, myelocytes, metamyelocytes, band neutrophils, and on a minority of mature granulocytes. NAT-9 II:3F-6F did not bind to circulating monocytes, T and B cells, erythrocytes and a variety of different human cell culture lines. Immunoblotting demonstrated that the antibody bind to a cellular component with a Mr approximately 97.400 dalton. The antibody may be useful in immunological subclassification of non-lymphoid leukemias and in studies on hematopoiesis.