Competitive endothelial adhesion between Plasmodium falciparum isolates under physiological flow conditions

<p>Abstract</p> <p>Background</p> <p>Sequestration of parasitized red blood cells in the microvasculature of major organs involves a sequence of events that is believed to contribute to the pathogenesis of severe falciparum malaria. <it>Plasmodium falciparum </it>infections are commonly composed of multiple subpopulations of parasites with varied adhesive properties. A key question is: do these subpopulations compete for adhesion to endothelium? This study investigated whether, in a laboratory model of cytoadherence, there is competition in binding to endothelium between pRBC infected with <it>P. falciparum </it>of variant adhesive phenotypes, particularly under flow conditions.</p> <p>Methods</p> <p>Four different <it>P. falciparum </it>isolates, of known adherence phenotypes, were matched in pairs, mixed in different proportions and allowed to bind to cultured human endothelium. Using <it>in vitro </it>competitive static and flow-based adhesion assays, that allow simultaneous testing of the adhesive properties of two different parasite lines, adherence levels of paired <it>P. falciparum </it>isolates were quantified and analysed using either non-parametric Wilcoxon's paired signed rank test or Student paired test.</p> <p>Results</p> <p>Study findings show that <it>P. falciparum </it>parasite lines show marked differences in the efficiency of adhesion to endothelium.</p> <p>Conclusion</p> <p><it>Plasmodium falciparum </it>variants will compete for adhesion to endothelia and variants can be ranked by their efficiency of binding. These findings suggest that variants from a mixed infection will not show uniform cytoadherence and so may vary in their ability to cause disease.</p

Extending the M_(bh)-sigma diagram with dense nuclear star clusters

Abridged: Four new nuclear star cluster masses, M_nc, plus seven upper limits, are provided for galaxies with previously determined black hole masses, M_bh. Together with a sample of 64 galaxies with direct M_bh measurements, 13 of which additionally now have M_nc measurements rather than only upper limits, plus an additional 29 dwarf galaxies with available M_nc measurements and velocity dispersions sigma, an (M_bh + M_nc)-sigma diagram is constructed. Given that major dry galaxy merger events preserve the M_bh/L ratio, and given that L ~ sigma^5 for luminous galaxies, it is first noted that the observation M_bh ~ sigma^5 is consistent with expectations. For the fainter elliptical galaxies it is known that L ~ sigma^2, and assuming a constant M_nc/L ratio (Ferrarese et al.), the expectation that M_nc ~ sigma^2 is in broad agreement with our new observational result that M_nc ~ sigma^{1.57\pm0.24}. This exponent is however in contrast to the value of ~4 which has been reported previously and interpreted in terms of a regulating feedback mechanism from stellar winds.Comment: 6 pages, 2 figures. Submitted 08/08/2011 to MNRAS, first referee report received 19/01/2012, accepted 10/02/201

Specific Receptor Usage in Plasmodium falciparum Cytoadherence Is Associated with Disease Outcome

Our understanding of the basis of severe disease in malaria is incomplete. It is clear that pathology is in part related to the pro-inflammatory nature of the host response but a number of other factors are also thought to be involved, including the interaction between infected erythrocytes and endothelium. This is a complex system involving several host receptors and a major parasite-derived variant antigen (PfEMP1) expressed on the surface of the infected erythrocyte membrane. Previous studies have suggested a role for ICAM-1 in the pathology of cerebral malaria, although these have been inconclusive. In this study we have examined the cytoadherence patterns of 101 patient isolates from varying clinical syndromes to CD36 and ICAM-1, and have used variant ICAM-1 proteins to further characterise this adhesive phenotype. Our results show that increased binding to CD36 is associated with uncomplicated malaria while ICAM-1 adhesion is raised in parasites from cerebral malaria cases

The Effect of Anti-Rosetting Agents against Malaria Parasites under Physiological Flow Conditions

Rosetting remains the dominant malaria parasite adhesion phenotype associated with severe disease and pathogenicity in Africa. The formation of rosettes, whereby a Plasmodium falciparum infected erythrocyte (IE) adheres to two or more non-IEs, is thought to facilitate the occlusion of microvascular blood vessels by adhering to host endothelial cells and other bound IEs. Current methods of determining the rosette-disrupting capabilities of antibodies/drugs have focused on static assays. As IEs in vivo are exposed to shear stresses within the microvasculature, the effect of flow conditions on rosetting requires further examination. This study establishes a new rosetting flow assay using a closed perfusion system together with inverted fluorescence microscopy and image analysis, and confirms previous reports that rosettes exist under shear stresses equivalent to those present in the microvasculature (0.5-1.0 dyn/cm(2)). Furthermore, we tested the effectiveness of rosette-disrupting PfEMP1 antibodies, heparin and fucoidan over a range of concentrations on two P. falciparum strains, and found no statistically significant differences between the results of static and flow assays. The new flow assay is a valuable addition to the tools available to study rosetting. However, the static assay has good predictive value and remains useful as the standard screening test for rosette-disrupting interventions

Multiple novel prostate cancer susceptibility signals identified by fine-mapping of known risk loci among Europeans

Genome-wide association studies (GWAS) have identified numerous common prostate cancer (PrCa) susceptibility loci. We have fine-mapped 64 GWAS regions known at the conclusion of the iCOGS study using large-scale genotyping and imputation in 25 723 PrCa cases and 26 274 controls of European ancestry. We detected evidence for multiple independent signals at 16 regions, 12 of which contained additional newly identified significant associations. A single signal comprising a spectrum of correlated variation was observed at 39 regions; 35 of which are now described by a novel more significantly associated lead SNP, while the originally reported variant remained as the lead SNP only in 4 regions. We also confirmed two association signals in Europeans that had been previously reported only in East-Asian GWAS. Based on statistical evidence and linkage disequilibrium (LD) structure, we have curated and narrowed down the list of the most likely candidate causal variants for each region. Functional annotation using data from ENCODE filtered for PrCa cell lines and eQTL analysis demonstrated significant enrichment for overlap with bio-features within this set. By incorporating the novel risk variants identified here alongside the refined data for existing association signals, we estimate that these loci now explain ∼38.9% of the familial relative risk of PrCa, an 8.9% improvement over the previously reported GWAS tag SNPs. This suggests that a significant fraction of the heritability of PrCa may have been hidden during the discovery phase of GWAS, in particular due to the presence of multiple independent signals within the same regio

Effectiveness of a national quality improvement programme to improve survival after emergency abdominal surgery (EPOCH): a stepped-wedge cluster-randomised trial

Background: Emergency abdominal surgery is associated with poor patient outcomes. We studied the effectiveness of a national quality improvement (QI) programme to implement a care pathway to improve survival for these patients. Methods: We did a stepped-wedge cluster-randomised trial of patients aged 40 years or older undergoing emergency open major abdominal surgery. Eligible UK National Health Service (NHS) hospitals (those that had an emergency general surgical service, a substantial volume of emergency abdominal surgery cases, and contributed data to the National Emergency Laparotomy Audit) were organised into 15 geographical clusters and commenced the QI programme in a random order, based on a computer-generated random sequence, over an 85-week period with one geographical cluster commencing the intervention every 5 weeks from the second to the 16th time period. Patients were masked to the study group, but it was not possible to mask hospital staff or investigators. The primary outcome measure was mortality within 90 days of surgery. Analyses were done on an intention-to-treat basis. This study is registered with the ISRCTN registry, number ISRCTN80682973. Findings: Treatment took place between March 3, 2014, and Oct 19, 2015. 22 754 patients were assessed for elegibility. Of 15 873 eligible patients from 93 NHS hospitals, primary outcome data were analysed for 8482 patients in the usual care group and 7374 in the QI group. Eight patients in the usual care group and nine patients in the QI group were not included in the analysis because of missing primary outcome data. The primary outcome of 90-day mortality occurred in 1210 (16%) patients in the QI group compared with 1393 (16%) patients in the usual care group (HR 1·11, 0·96–1·28). Interpretation: No survival benefit was observed from this QI programme to implement a care pathway for patients undergoing emergency abdominal surgery. Future QI programmes should ensure that teams have both the time and resources needed to improve patient care. Funding: National Institute for Health Research Health Services and Delivery Research Programme

Effectiveness of a national quality improvement programme to improve survival after emergency abdominal surgery (EPOCH): a stepped-wedge cluster-randomised trial

BACKGROUND: Emergency abdominal surgery is associated with poor patient outcomes. We studied the effectiveness of a national quality improvement (QI) programme to implement a care pathway to improve survival for these patients. METHODS: We did a stepped-wedge cluster-randomised trial of patients aged 40 years or older undergoing emergency open major abdominal surgery. Eligible UK National Health Service (NHS) hospitals (those that had an emergency general surgical service, a substantial volume of emergency abdominal surgery cases, and contributed data to the National Emergency Laparotomy Audit) were organised into 15 geographical clusters and commenced the QI programme in a random order, based on a computer-generated random sequence, over an 85-week period with one geographical cluster commencing the intervention every 5 weeks from the second to the 16th time period. Patients were masked to the study group, but it was not possible to mask hospital staff or investigators. The primary outcome measure was mortality within 90 days of surgery. Analyses were done on an intention-to-treat basis. This study is registered with the ISRCTN registry, number ISRCTN80682973. FINDINGS: Treatment took place between March 3, 2014, and Oct 19, 2015. 22 754 patients were assessed for elegibility. Of 15 873 eligible patients from 93 NHS hospitals, primary outcome data were analysed for 8482 patients in the usual care group and 7374 in the QI group. Eight patients in the usual care group and nine patients in the QI group were not included in the analysis because of missing primary outcome data. The primary outcome of 90-day mortality occurred in 1210 (16%) patients in the QI group compared with 1393 (16%) patients in the usual care group (HR 1·11, 0·96-1·28). INTERPRETATION: No survival benefit was observed from this QI programme to implement a care pathway for patients undergoing emergency abdominal surgery. Future QI programmes should ensure that teams have both the time and resources needed to improve patient care. FUNDING: National Institute for Health Research Health Services and Delivery Research Programme

Continued cytoadherence of Plasmodium falciparum infected red blood cells after antimalarial treatment

Development of severe disease in Plasmodium falciparum malaria infection is thought to be, at least in part, due to the sequestration of trophozoite-stage infected red blood cells in the microvasculature. The process of cytoadherence is mediated by binding of the parasite protein PfEMP-1 on the surface of infected red blood cells to endothelial cell receptors. Although antimalarial treatments rapidly kill parasites, significant mortality is still seen in severe malaria, particularly within 24 h of hospital admission. We find that cytoadherence of infected red blood cells continues for several hours after killing of the parasite by antimalarials; after 24 h treatment using a range of antimalarials binding is approximately one-third the level of untreated parasite cultures. This is consistent with the maintained presence of PfEMP-1 on the surface of drug-treated infected red blood cells. A specific advantage of artesunate over other treatments tested is seen on addition of this drug to younger ring stage parasites, which do not mature to the cytoadherent trophozoite-stage. These findings show that cytoadherence, a potential pathogenic property of P. falciparum infected red blood cells, continues long after the parasite has been killed. These data support the development of adjunctive therapies to reverse the pathophysiological consequences of cytoadherence

Exported Proteins Required for Virulence and Rigidity of Plasmodium falciparum-Infected Human Erythrocytes

A major part of virulence for Plasmodium falciparum malaria infection, the most lethal parasitic disease of humans, results from increased rigidity and adhesiveness of infected host red cells. These changes are caused by parasite proteins exported to the erythrocyte using novel trafficking machinery assembled in the host cell. To understand these unique modifications, we used a large-scale gene knockout strategy combined with functional screens to identify proteins exported into parasite-infected erythrocytes and involved in remodeling these cells. Eight genes were identified encoding proteins required for export of the parasite adhesin PfEMP1 and assembly of knobs that function as physical platforms to anchor the adhesin. Additionally, we show that multiple proteins play a role in generating increased rigidity of infected erythrocytes. Collectively these proteins function as a pathogen secretion system, similar to bacteria and may provide targets for antivirulence based therapies to a disease responsible for millions of deaths annually