The inhibition of staphylococcal delta-haemolysin by human serum

It is well established that staphylococcal delta-haemolysin is inhibited by normal mammalian sera and forms a precipitin line on gel diffusion with normal sera. The purpose of this thesis was to quantitate inhibitory activity against delta-haemolysin and to identify the serum factor(s) involved. The inhibitory titre of normal human serum assayed using purified delta-haemolysin with cod erythrocytes as indicators of haemolysis was 1/500 - 1/1000. Fractionation of serum by ultrafiltration, gel filtration in Sephadex G-150 and Sepharose 6b and ammonium sulphate precipitation indicated that at least two components of molecular weights approximately 300,000 and > 106 were involved. When affinity chromatography was done using delta-haemolysin covalently coupled to Sepharose-4B the selective removal of g-lipoprotein from whole serum was shown by immunoelectrophoresis. High density (alpha-) and low density (beta-) lipoproteins were prepared by zonal ultracentrifugation of serum using potassium bromide density gradients and characterised by polyacrylamide gel electrophoresis, immunoelectrophoresis and gel diffusion using monospecific antisera. Both alpha- and beta-lipoproteins inhibited delta-haemolysin and gave a precipitin line on gel diffusion, while fractions which were shown by immunoelectrophoresis to be devoid of such lipoproteins were non-inhibitory. Proteolytic digestion of lipoproteins with trypsin or papain did not diminish their inhibitory activity, whereas this was diminished by pretreatment with Bacillus cereus phospholipase C. Delta-haemolysin was inhibited hy phospholipids and pretreatment with phospholipase decreased the inhibitory capacity of such phospholipids and also of a serum lipid extract. The results indicated that binding of delta-haemolysin to serum lipoproteins was via the phospholipid moiety and the implications of such an interaction in terms of in vivo activity of the haemolysin are discussed

Chemotherapy and immunity in murine African trypanosomiasis

The experiments described in this thesis investigated aspects of immunity and chemotherapy in mice infected with T. brucei and T. congolense. It was found that mice with infections of T. brucei of less than 7 days duration were completely cured if treated at this stage with trypanocidal drugs" However, if treatment was delayed beyond 14 days, a proportion of the mice relapsed, in some cases more than 3 months after chemotherapy. This was not due to drug resistance of the parasite, and tissue transfer experiments during the aparasitaemic period following chemotherapy indicated the presence of trypanosomes only in brain tissue. Transmission of immunity from mother to offspring was investigated in mice. Mothers with a T. brucei infection conferred immunity to homologous challenge of approximately 3 weeks duration in newborn mice. This immunity was by way of colostrum/milk after birth, since fostering experiments showed that mice born of infected mothers but suckled by an uninfected mother were susceptible to challenge. Furthermore there was an augmentative effect when chemoprophylaxis was given on the day of birth. This on its own afforded protection against infection for a similar period as passively acquired antibody, but young mice which received both drug and antibody resisted challenge for over 6 weeks. Genetic resistance to T. congolense infection was studied in C57B1 mice, which are able to repeatedly limit the numbers .of circulating parasites and survive for approximately 80 days, and CFLP mice which die within 10 days with an uncontrolled fulminating parasitaemia. No evidence was found to implicate pathophysiological factors in this resistance, but C57B1 mice were able to remove radiolabelled parasites from their circulation, and their immune response, as judged by in vitro immunological methods, was superior to the CFLP strain Attempts to enhance the response of susceptible mice by passive immunisation, priming of the immune system and activation of the mono" nuclear phagocytic system were unsuccessful, and it was concluded that the ability to produce and maintain levels of IgM in the plasma was essential for resistance. Finally, a reliable and simple technique for the in vivo labelling of trypanosomes with 75Se-methionine was developed" The fate of such trypanosomes after injection into normal and immune mice was investigated, and it was found that in immune mice the liver was the principal site of uptake. From this finding, studies were done on the mechanisms of hepatic uptake, which was found to be antibody dependent, and at low antibody titres also C3 dependents No evidence was found to suggest that intravascular lysis or activated macrophages were involved in immune clearance

Temporal Trends and Clinical Trial Characteristics Associated with the Inclusion of Women in Heart Failure Trial Steering Committees:A Systematic Review

Background: Trial steering committees (TSCs) steer the conduct of randomized controlled trials (RCTs). We examined the gender composition of TSCs in impactful heart failure RCTs and explored whether trial leadership by a woman was independently associated with the inclusion of women in TSCs. Methods: We systematically searched MEDLINE, EMBASE, and CINAHL for heart failure RCTs published in journals with impact factor ≥10 between January 2000 and May 2019. We used the Jonckheere-Terpstra test to assess temporal trends and multivariable logistic regression to explore trial characteristics associated with TSC inclusion of women. Results: Of 403 RCTs that met inclusion criteria, 127 (31.5%) reported having a TSC but 20 of these (15.7%) did not identify members. Among 107 TSCs that listed members, 56 (52.3%) included women and 6 of these (10.7%) restricted women members to the RCT leaders. Of 1213 TSC members, 11.1% (95% CI, 9.4%-13.0%) were women, with no change in temporal trends (P=0.55). Women had greater odds of TSC inclusion in RCTs led by women (adjusted odds ratio, 2.48 [95% CI, 1.05-8.72], P=0.042); this association was nonsignificant when analysis excluded TSCs that restricted women to the RCT leaders (adjusted odds ratio 1.46 [95% CI, 0.43-4.91], P=0.36). Conclusions: Women were included in 52.3% of TSCs and represented 11.1% of TSC members in 107 heart failure RCTs, with no change in trends since 2000. RCTs led by women had higher adjusted odds of including women in TSCs, partly due to the self-inclusion of RCT leaders in TSCs

Differential regulation of the alpha-globin locus by Kruppel-like factor 3 in erythroid and non-erythroid cells

Background: Krüppel-like Factor 3 (KLF3) is a broadly expressed zinc-finger transcriptional repressor with diverse biological roles. During erythropoiesis, KLF3 acts as a feedback repressor of a set of genes that are activated by Krüppel-like Factor 1 (KLF1). Noting that KLF1 binds α-globin gene regulatory sequences during erythroid maturation, we sought to determine whether KLF3 also interacts with the α-globin locus to regulate transcription. Results: We found that expression of a human transgenic α-globin reporter gene is markedly up-regulated in fetal and adult erythroid cells of Klf3−/− mice. Inspection of the mouse and human α-globin promoters revealed a number of canonical KLF-binding sites, and indeed, KLF3 was shown to bind to these regions both in vitro and in vivo. Despite these observations, we did not detect an increase in endogenous murine α-globin expression in Klf3−/− erythroid tissue. However, examination of murine embryonic fibroblasts lacking KLF3 revealed significant de-repression of α-globin gene expression. This suggests that KLF3 may contribute to the silencing of the α-globin locus in non-erythroid tissue. Moreover, ChIP-Seq analysis of murine fibroblasts demonstrated that across the locus, KLF3 does not occupy the promoter regions of the α-globin genes in these cells, but rather, binds to upstream, DNase hypersensitive regulatory regions. Conclusions: These findings reveal that the occupancy profile of KLF3 at the α-globin locus differs in erythroid and non-erythroid cells. In erythroid cells, KLF3 primarily binds to the promoters of the adult α-globin genes, but appears dispensable for normal transcriptional regulation. In non-erythroid cells, KLF3 distinctly binds to the HS-12 and HS-26 elements and plays a non-redundant, albeit modest, role in the silencing of α-globin expression. </p

Rapid Diagnosis of Tuberculosis with the Xpert MTB/RIF Assay in High Burden Countries: A Cost-Effectiveness Analysis

A cost-effectiveness study by Frank Cobelens and colleagues reveals that Xpert MTB/RIF is a cost-effective method of tuberculosis diagnosis that is suitable for use in low- and middle-income settings

Global gene expression analysis of canine osteosarcoma stem cells reveals a novel role for COX-2 in tumour initiation

Osteosarcoma is the most common primary bone tumour of both children and dogs. It is an aggressive tumour in both species with a rapid clinical course leading ultimately to metastasis. In dogs and children distant metastasis occurs in >80% of individuals treated by surgery alone. Both canine and human osteosarcoma has been shown to contain a sub-population of cancer stem cells (CSCs), which may drive tumour growth, recurrence and metastasis, suggesting that naturally occurring canine osteosarcoma could act as a preclinical model for the human disease. Here we report the successful isolation of CSCs from primary canine osteosarcoma, as well as established cell lines. We show that these cells can form tumourspheres, and demonstrate relative resistance to chemotherapy. We demonstrate similar results for the human osteosarcma cell lines, U2OS and SAOS2. Utilizing the Affymetrix canine microarray, we are able to definitively show that there are significant differences in global gene expression profiles of isolated osteosarcoma stem cells and the daughter adherent cells. We identified 13,221 significant differences (p = 0.05), and significantly, COX-2 was expressed 141-fold more in CSC spheres than daughter adherent cells. To study the role of COX-2 expression in CSCs we utilized the COX-2 inhibitors meloxicam and mavacoxib. We found that COX-2 inhibition had no effect on CSC growth, or resistance to chemotherapy. However inhibition of COX-2 in daughter cells prevented sphere formation, indicating a potential significant role for COX-2 in tumour initiation

HIV-1 Residual Viremia Correlates with Persistent T-Cell Activation in Poor Immunological Responders to Combination Antiretroviral Therapy

BACKGROUND:The clinical significance and cellular sources of residual human immunodeficiency virus type 1 (HIV-1) production despite suppressive combination antiretroviral therapy (cART) remain unclear and the effect of low-level viremia on T-cell homeostasis is still debated. METHODOLOGY/PRINCIPAL FINDINGS:We characterized the recently produced residual viruses in the plasma and short-lived blood monocytes of 23 patients with various immunological responses to sustained suppressive cART. We quantified the residual HIV-1 in the plasma below 50 copies/ml, and in the CD14(high) CD16(-) and CD16+ monocyte subsets sorted by flow cytometry, and predicted coreceptor usage by genotyping V3 env sequences. We detected residual viremia in the plasma of 8 of 10 patients with poor CD4+ T-cell reconstitution in response to cART and in only 5 of 13 patients with good CD4+ T-cell reconstitution. CXCR4-using viruses were frequent among the recently produced viruses in the plasma and in the main CD14(high) CD16(-) monocyte subset. Finally, the residual viremia was correlated with persistent CD4+ and CD8+ T-cell activation in patients with poor immune reconstitution. CONCLUSIONS:Low-level viremia could result from the release of archived viruses from cellular reservoirs and/or from ongoing virus replication in some patients. The compartmentalization of the viruses between the plasma and the blood monocytes suggests at least two origins of residual virus production during effective cART. CXCR4-using viruses might be produced preferentially in patients on cART. Our results also suggest that low-level HIV-1 production in some patients may contribute to persistent immune dysfunction despite cART

BMQ

BMQ: Boston Medical Quarterly was published from 1950-1966 by the Boston University School of Medicine and the Massachusetts Memorial Hospitals

Factors affecting general practice collaboration with voluntary and community sector organisations.

Collaborative working between general practice (GP) and voluntary and community sector (VCS) organisations is increasingly championed as a means of primary care doing more with less and of addressing patients' "wicked problems". This paper aims to add to the knowledge base around collaborative practice between GPs and VCS organisations by examining the factors that aid or inhibit such collaboration. A case study design was used to examine the lived-experience of GPs and VCS organisations working collaboratively. Four cases, each consisting of a GP and a VCS organisation with whom they work collaboratively, were identified. Interviews (n = 18) and a focus group (n = 1) were conducted with staff within each organisation. Transcribed data were analysed thematically. Whilet there are similarities across cases in their use of, for example, Health Trainers and social prescribing, the form and function of GP-VCS collaborations were unique to their local context. The identified factors affecting GP-VCS collaboration reflect those found in previous service evaluations and the broader literature on partnership working; shared understanding, time and resources, trust, strong leadership, operational systems and governance and the "negotiation" of professional boundaries. While the current political environment may represent an opportunity for collaborations to develop, there are issues yet to be resolved before collaboration-especially more holistic and integrated approaches-becomes systematically embedded into practice

α-thalassaemia

Alpha-thalassaemia is inherited as an autosomal recessive disorder characterised by a microcytic hypochromic anaemia, and a clinical phenotype varying from almost asymptomatic to a lethal haemolytic anaemia