537Microparticles and exosomes differentially impact on endothelial cell function in coronary artery disease

Background and Purpose: Microparticles (MPs) and exosomes are released by cells using different mechanisms. Thus, quantitative as well as qualitative changes of both particle populations, MPs and exosomes, in patients with coronary artery disease (CAD) might reflect an altered activation status of the endothelium, platelets and leukocytes. Moreover, they might exert differential effects on the target organs, such as the endothelium. Yet, alterations in both populations have not been studied side-by-side so far. The aim of the study was to compare the impact of MPs and exosomes from healthy subjects and CAD patients on endothelial cell (EC) functional characteristics. Methods: MPs and exosomes were isolated by stepwise filtration and ultracentrifugation from citrate-plasma and verified by electron microscopy and dynamic light scattering. MP and exosome fractions, as well as the vehicle (PBS), were added to human arterial ECs and EC apoptosis, number, size, capacity for in vitro-reendothelialisation after scratching, expression of adhesion molecules ICAM-1 and VCAM-1 were assessed. In parallel, platelet-, endothelial- and leukocyte-derived MPs were quantified. In a separate sub-study, the same parameters were assessed in plasma of CAD patients undergoing standard medical rehabilitation or an exercise-based cardiac rehabilitation programme. Results: MPs of healthy, but not of CAD patients supported in vitro re-endothelialisation, while exosomes had no influence. Exercise, but not standard rehabilitation improved CAD MP capacity to support in vitro rehabilitation. This was negatively correlated to the number of leukocyte- and endothelial-derived MPs, but not total or platelet MPs. EC number was negatively affected by exposure to CAD MPs. ANCOVA analysis identified disease, but not the particle type as influencing factor. Instead, apoptotic cell death was influenced by particle type, but not by the disease, and was not altered in rehabilitation. Similarly, ICAM-1 and VCAM-1 expression were enhanced on ECs after incubation with exosomes, but not with MPs, with no effect of disease or rehabilitation. Conclusion: MPs and exosomes differentially affect endothelial cell function and underlie differential modulation in disease and rehabilitation. Those findings might in the future help to optimize and monitor cardiovascular therap

Molecular monitoring of Plasmodium falciparum resistance to artemisinin in Tanzania

Artemisinin-based combination therapies (ACTs) are recommended for use against uncomplicated malaria in areas of multi-drug resistant malaria, such as sub-Saharan Africa. However, their long-term usefulness in these high transmission areas remains unclear. It has been suggested that documentation of the S769N PfATPase6 mutations may indicate an emergence of artemisinin resistance of Plasmodium falciparum in the field. The present study assessed PfATPase6 mutations (S769N and A623E) in 615 asymptomatic P. falciparum infections in Tanzania but no mutant genotype was detected. This observation suggests that resistance to artemisinin has not yet been selected in Tanzania, supporting the Ministry of Health's decision to adopt artemether+lumefantrine as first-line malaria treatment. The findings recommend further studies to assess PfATPase6 mutations in sentinel sites and verify their usefulness in monitoring emergency of ACT resistance

Inadequacy of zero-width approximation for a light Higgs boson signal

In the Higgs search at the LHC, a light Higgs boson (115 GeV <~ M_H <~ 130 GeV) is not excluded by experimental data. In this mass range, the width of the Standard Model Higgs boson is more than four orders of magnitude smaller than its mass. The zero-width approximation is hence expected to be an excellent approximation. We show that this is not always the case. The inclusion of off-shell contributions is essential to obtain an accurate Higgs signal normalisation at the 1% precision level. For gg (-> H) -> VV, V= W,Z, O(10%) corrections occur due to an enhanced Higgs signal in the region M_VV > 2 M_V, where also sizable Higgs-continuum interference occurs. We discuss how experimental selection cuts can be used to exclude this region in search channels where the Higgs invariant mass cannot be reconstructed. We note that the H -> VV decay modes in weak boson fusion are similarly affected.Comment: 26 pages, 18 figures, 6 tables; added references, expanded introduction, version to appear in JHE

Experimental Evolution of Resistance to Artemisinin Combination Therapy Results in Amplification of the mdr1 Gene in a Rodent Malaria Parasite

Background: Lacking suitable alternatives, the control of malaria increasingly depends upon Artemisinin Combination Treatments (ACT): resistance to these drugs would therefore be disastrous. For ACTs, the biology of resistance to the individual components has been investigated, but experimentally induced resistance to component drugs in combination has not been generated. Methodology/Principal Findings: We have used the rodent malaria parasite Plasmodium chabaudi to select in vivo resistance to the artesunate (ATN) + mefloquine (MF) version of ACT, through prolonged exposure of parasites to both drugs over many generations. The selection procedure was carried out over twenty-seven consecutive sub-inoculations under increasing ATN + MF doses, after which a genetically stable resistant parasite, AS-ATNMF1, was cloned. AS-ATNMF1 showed increased resistance to ATN + MF treatment and to artesunate or mefloquine administered separately. Investigation of candidate genes revealed an mdr1 duplication in the resistant parasites and increased levels of mdr1 transcripts and protein. There were no point mutations in the atpase6 or ubp1genes. Conclusion: Resistance to ACTs may evolve even when the two drugs within the combination are taken simultaneously and amplification of the mdr1 gene may contribute to this phenotype. However, we propose that other gene(s), as ye

Molecular markers of anti-malarial drug resistance in Central, West and East African children with severe malaria.

BACKGROUND: The Plasmodium falciparum multidrug resistance 1 (PfMDR1), P. falciparum Ca(2+)-ATPase (PfATP6) and Kelch-13 propeller domain (PfK13) loci are molecular markers of parasite susceptibility to anti-malarial drugs. Their frequency distributions were determined in the isolates collected from children with severe malaria originating from three African countries. METHODS: Samples from 287 children with severe malaria [(Gabon: n = 114); (Ghana: n = 89); (Kenya: n = 84)] were genotyped for pfmdr1, pfatp6 and pfk13 loci by DNA sequencing and assessing pfmdr1 copy number variation (CNV) by real-time PCR. RESULTS: Pfmdr1-N86Y mutation was detected in 48, 10 and 10% in Lambaréné, Kumasi and Kisumu, respectively. At codon 184, the prevalence of the mutation was 73% in Lambaréné, 63% in Kumasi and 49% Kisumu. The S1034C and N1042D variants were absent at all three sites, while the frequency of the D1246Y mutation was 1, 3 and 13% in Lambaréné, Kumasi and Kisumu, respectively. Isolates with two pfmdr1 gene copy number predominantly harboured the N86Y wild-type allele and were mostly found in Kumasi (10%) (P < 0.0001). Among the main pfmdr1 haplotypes (NFD, NYD and YFD), NYD was associated with highest parasitaemia (P = 0.04). At the pfatp6 locus, H243Y and A623E mutations were observed at very low frequency at all three sites. The prevalence of the pfatp6 E431K variant was 6, 18 and 17% in Lambaréné, Kumasi and Kisumu, respectively. The L263E and S769N mutations were absent in all isolates. The pfk13 variants associated with artemisinin resistance in Southeast Asia were not observed. Eleven novel substitutions in the pfk13 locus occurring at low frequency were observed. CONCLUSIONS: Artemisinins are still highly efficacious in large malaria-endemic regions though declining efficacy has occurred in Southeast Asia. The return of chloroquine-sensitive strains following the removal of drug pressure is observed. However, selection of wild-type alleles in the multidrug-resistance gene and the increased gene copy number is associated with reduced lumefantrine sensitivity. This study indicates a need to constantly monitor drug resistance to artemisinin in field isolates from malaria-endemic countries

Genomic identification of cryptic susceptibility to penicillins and β-lactamase inhibitors in methicillin-resistant Staphylococcus aureus.

Antibiotic resistance in bacterial pathogens threatens the future of modern medicine. One such resistant pathogen is methicillin-resistant Staphylococcus aureus (MRSA), which is resistant to nearly all β-lactam antibiotics, limiting treatment options. Here, we show that a significant proportion of MRSA isolates from different lineages, including the epidemic USA300 lineage, are susceptible to penicillins when used in combination with β-lactamase inhibitors such as clavulanic acid. Susceptibility is mediated by a combination of two different mutations in the mecA promoter region that lowers mecA-encoded penicillin-binding protein 2a (PBP2a) expression, and in the majority of isolates by either one of two substitutions in PBP2a (E246G or M122I) that increase the affinity of PBP2a for penicillin in the presence of clavulanic acid. Treatment of S. aureus infections in wax moth and mouse models shows that penicillin/β-lactamase inhibitor susceptibility can be exploited as an effective therapeutic choice for 'susceptible' MRSA infection. Finally, we show that isolates with the PBP2a E246G substitution have a growth advantage in the presence of penicillin but the absence of clavulanic acid, which suggests that penicillin/β-lactamase susceptibility is an example of collateral sensitivity (resistance to one antibiotic increases sensitivity to another). Our findings suggest that widely available and currently disregarded antibiotics could be effective in a significant proportion of MRSA infections.MRC - G1001787/1 MRC - MR/N002660/1 WT098600 HICF-T5-342 MR/S00291X/1 201344/Z/16/Z MR/P007201/

Ca2+ monitoring in Plasmodium falciparum using the yellow cameleon-Nano biosensor

Calcium (Ca2+)-mediated signaling is a conserved mechanism in eukaryotes, including the human malaria parasite, Plasmodium falciparum. Due to its small size (300?nM). We determined that the mammalian SERCA inhibitor thapsigargin and antimalarial dihydroartemisinin did not perturb SERCA activity. The change of the cytosolic Ca2+ level in P. falciparum was additionally detectable by flow cytometry. Thus, we propose that the developed YC-Nano-based system is useful to study Ca2+ signaling in P. falciparum and is applicable for drug screening.We are grateful to Japanese Red Cross Blood Society for providing human RBC and plasma. We also thank Tanaka R, Ogoshi (Sakura) M and Matsumoto N for technical assistance and Templeton TJ for critical reading. This study was conducted at the Joint Usage / Research Center on Tropical Disease, Institute of Tropical Medicine, Nagasaki University, Japan. KP was a Tokyo Biochemical Research Foundation (TBRF, http://www.tokyobrf.or.jp) post-doctoral fellow and PEF was a Japanese Society of Promotion Sciences (JSPS) post-doctoral fellow. This work was supported in part by the TBRF (K.P.), JSPS (P.E.F.), Takeda Science Foundation (K.Y.), Grants-in-Aids for Scientific Research 24590509 (K.Y.), 22390079 (O.K.), and for Scientific Research on Innovative Areas 23117008 (O.K.), MEXT, Japan. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Consistent truncations of supergravity and 1/2-BPS RG flows in 4d SCFTs

With the purpose of holographically describing flows from a large family of four dimensional ${\cal N}=1$ and ${\cal N}=2$ conformal field theories, we discuss truncations of seven dimensional supergravity to five dimensions. We write explicitly the reduced gauged supergravity and find BPS equations for simple configurations. Lifting these flows to eleven dimensions or Massive IIA supergravity, we present string duals to RG flows from strongly coupled conformal theories when deformed by marginal and/or relevant operators. We further discuss observables common to infinite families of ${\cal N}=1$ and ${\cal N}=2$ QFTs in this context.Comment: 28 pages plus appendixes. JHEP versio

Global response of Plasmodium falciparum to hyperoxia: a combined transcriptomic and proteomic approach

<p>Abstract</p> <p>Background</p> <p>Over its life cycle, the <it>Plasmodium falciparum </it>parasite is exposed to different environmental conditions, particularly to variations in O₂pressure. For example, the parasite circulates in human venous blood at 5% O₂pressure and in arterial blood, particularly in the lungs, at 13% O₂pressure. Moreover, the parasite is exposed to 21% O₂levels in the salivary glands of mosquitoes.</p> <p>Methods</p> <p>To study the metabolic adaptation of <it>P. falciparum </it>to different oxygen pressures during the intraerythrocytic cycle, a combined approach using transcriptomic and proteomic techniques was undertaken.</p> <p>Results</p> <p>Even though hyperoxia lengthens the parasitic cycle, significant transcriptional changes were detected in hyperoxic conditions in the late-ring stage. Using PS 6.0™ software (Ariadne Genomics) for microarray analysis, this study demonstrate up-expression of genes involved in antioxidant systems and down-expression of genes involved in the digestive vacuole metabolism and the glycolysis in favour of mitochondrial respiration. Proteomic analysis revealed increased levels of heat shock proteins, and decreased levels of glycolytic enzymes. Some of this regulation reflected post-transcriptional modifications during the hyperoxia response.</p> <p>Conclusions</p> <p>These results seem to indicate that hyperoxia activates antioxidant defence systems in parasites to preserve the integrity of its cellular structures. Moreover, environmental constraints seem to induce an energetic metabolism adaptation of <it>P. falciparum</it>. This study provides a better understanding of the adaptive capabilities of <it>P. falciparum </it>to environmental changes and may lead to the development of novel therapeutic targets.</p

The plant-based immunomodulator curcumin as a potential candidate for the development of an adjunctive therapy for cerebral malaria

The clinical manifestations of cerebral malaria (CM) are well correlated with underlying major pathophysiological events occurring during an acute malaria infection, the most important of which, is the adherence of parasitized erythrocytes to endothelial cells ultimately leading to sequestration and obstruction of brain capillaries. The consequent reduction in blood flow, leads to cerebral hypoxia, localized inflammation and release of neurotoxic molecules and inflammatory cytokines by the endothelium. The pharmacological regulation of these immunopathological processes by immunomodulatory molecules may potentially benefit the management of this severe complication. Adjunctive therapy of CM patients with an appropriate immunomodulatory compound possessing even moderate anti-malarial activity with the capacity to down regulate excess production of proinflammatory cytokines and expression of adhesion molecules, could potentially reverse cytoadherence, improve survival and prevent neurological sequelae. Current major drug discovery programmes are mainly focused on novel parasite targets and mechanisms of action. However, the discovery of compounds targeting the host remains a largely unexplored but attractive area of drug discovery research for the treatment of CM. This review discusses the properties of the plant immune-modifier curcumin and its potential as an adjunctive therapy for the management of this complication