Structural and functional studies of histidine biosynthesis in Acanthamoeba spp. demonstrates a novel molecular arrangement and target for antimicrobials

Acanthamoeba is normally free-living, but sometimes facultative and occasionally opportunistic parasites. Current therapies are, by necessity, arduous and yet poorly effective due to their inabilities to kill cyst stages or in some cases to actually induce encystation. Acanthamoeba can therefore survive as cysts and cause disease recurrence. Herein, in pursuit of better therapies and to understand the biochemistry of this understudied organism, we characterize its histidine biosynthesis pathway and explore the potential of targeting this with antimicrobials. We demonstrate that Acanthamoeba is a histidine autotroph, but with the ability to scavenge preformed histidine. It is able to grow in defined media lacking this amino acid, but is inhibited by 3-amino-1,2,4-triazole (3AT) that targets Imidazoleglycerol-Phosphate Dehydratase (IGPD) the rate limiting step of histidine biosynthesis. The structure of Acanthamoeba IGPD has also been determined in complex with 2-hydroxy-3-(1,2,4-triazol-1-yl) propylphosphonate [(R)-C348], a recently described novel inhibitor of Arabidopsis thaliana IGPD. This compound inhibited the growth of four Acanthamoeba species, having a 50% inhibitory concentration (IC50) ranging from 250-526 nM. This effect could be ablated by the addition of 1 mM exogenous free histidine, but importantly not by physiological concentrations found in mammalian tissues. The ability of 3AT and (R)-C348 to restrict the growth of four strains of Acanthamoeba spp. including a recently isolated clinical strain, while not inducing encystment, demonstrates the potential therapeutic utility of targeting the histidine biosynthesis pathway in Acanthamoeba

Clinical characteristics of women captured by extending the definition of severe postpartum haemorrhage with 'refractoriness to treatment': a cohort study

Background: The absence of a uniform and clinically relevant definition of severe postpartum haemorrhage hampers comparative studies and optimization of clinical management. The concept of persistent postpartum haemorrhage, based on refractoriness to initial first-line treatment, was proposed as an alternative to common definitions that are either based on estimations of blood loss or transfused units of packed red blood cells (RBC). We compared characteristics and outcomes of women with severe postpartum haemorrhage captured by these three types of definitions. Methods: In this large retrospective cohort study in 61 hospitals in the Netherlands we included 1391 consecutive women with postpartum haemorrhage who received either ≥4 units of RBC or a multicomponent transfusion. Clinical characteristics and outcomes of women with severe postpartum haemorrhage defined as persistent postpartum haemorrhage were compared to definitions based on estimated blood loss or transfused units of RBC within 24 h following birth. Adverse maternal outcome was a composite of maternal mortality, hysterectomy, arterial embolisation and intensive care unit admission. Results: One thousand two hundred sixty out of 1391 women (90.6%) with postpartum haemorrhage fulfilled the definition of persistent postpartum haemorrhage. The majority, 820/1260 (65.1%), fulfilled this definition within 1 h following birth, compared to 819/1391 (58.7%) applying the definition of ≥1 L blood loss and 37/845 (4.4%) applying the definition of ≥4 units of RBC. The definition persistent postpartum haemorrhage captured 430/471 adverse maternal outcomes (91.3%), compared to 471/471 (100%) for ≥1 L blood loss and 383/471 (81.3%) for ≥4 units of RBC. Persistent postpartum haemorrhage did not capture all adverse outcomes because of missing data on timing of initial, first-line treatment. Conclusion: The definition persistent postpartum haemo

Identifying useful human correction feedback from an on-line machine translation service

Post-editing feedback provided by users of on-line translation services offers an excellent opportunity for automatic improvement of statistical machine translation (SMT) systems. However, feedback provided by casual users is very noisy, and must be automatically filtered in order to identify the potentially useful cases. We present a study on automatic feedback filtering in a real weblog collected from Reverso.net. We extend and re-annotate a training corpus, define an extended set of simple features and approach the problem as a binary classification task, experimenting with linear and kernelbased classifiers and feature selection. Results on the feedback filtering task show a significant improvement over the majority class, but also a precision ceiling around 70-80%. This reflects the inherent difficulty of the problem and indicates that shallow features cannot fully capture the semantic nature of the problem. Despite the modest results on the filtering task, the classifiers are proven effective in an application-based evaluation. The incorporation of a filtered set of feedback instances selected from a larger corpus significantly improves the performance of a phrase-based SMT system, according to a set of standard evaluation metrics

The regulation of miRNAs by reconstituted high-density lipoproteins in diabetes-impaired angiogenesis

Diabetic vascular complications are associated with impaired ischaemia-driven angiogenesis. We recently found that reconstituted high-density lipoproteins (rHDL) rescue diabetes-impaired angiogenesis. microRNAs (miRNAs) regulate angiogenesis and are transported within HDL to sites of injury/repair. The role of miRNAs in the rescue of diabetes-impaired angiogenesis by rHDL is unknown. Using a miRNA array, we found that rHDL inhibits hsa-miR-181c-5p expression in vitro and using a hsa-miR-181c-5p mimic and antimiR identify a novel anti-angiogenic role for miR-181c-5p. miRNA expression was tracked over time post-hindlimb ischaemic induction in diabetic mice. Early post-ischaemia when angiogenesis is important, rHDL suppressed hindlimb mmu-miR-181c-5p. mmu-miR-181c-5p was not detected in the plasma or within HDL, suggesting rHDL specifically targets mmu-miR-181c-5p at the ischaemic site. Three known angiogenic miRNAs (mmu-miR-223-3p, mmu-miR-27b-3p, mmu-miR-92a-3p) were elevated in the HDL fraction of diabetic rHDL-infused mice early post-ischaemia. This was accompanied by a decrease in plasma levels. Only mmu-miR-223-3p levels were elevated in the hindlimb 3 days post-ischaemia, indicating that rHDL regulates mmu-miR-223-3p in a time-dependent and site-specific manner. The early regulation of miRNAs, particularly miR-181c-5p, may underpin the rescue of diabetes-impaired angiogenesis by rHDL and has implications for the treatment of diabetes-related vascular complications.Samuel T. Hourigan, Emma L. Solly, Victoria A. Nankivell, Anisyah Ridiandries, Benjamin M. Weimann, Rodney Henriquez, Edward R. Tepper, Jennifer Q. J. Zhang, Tania Tsatralis, Zoe E. Clayton, Laura Z. Vanags, Stacy Robertson, Stephen J. Nicholls, Martin K. C. Ng, Christina A. Bursill, Joanne T. M. Ta

VEGFR2 is activated by high-density lipoproteins and plays a key role in the proangiogenic action of HDL in ischemia

High-density lipoproteins augment hypoxia-induced angiogenesis by inducing the key angiogenic vascular endothelial growth factor A (VEGFA) and total protein levels of its receptor 2 (VEGFR2). The activation/phosphorylation of VEGFR2 is critical for mediating downstream, angiogenic signaling events. This study aimed to determine whether reconstituted high-density lipoprotein (rHDL) activates VEGFR2 phosphorylation and the downstream signaling events and the importance of VEGFR2 in the proangiogenic effects of rHDL in hypoxia. In vitro, rHDL increased VEGFR2 activation and enhanced phosphorylation of downstream, angiogenic signaling proteins ERK1/2 and p38 MAPK in hypoxia. Incubation with a VEGFR2-neutralizing antibody attenuated rHDL-induced phosphorylation of VEGFR2, ERK1/2, p38 MAPK, and tubule formation. In a murine model of ischemia-driven neovascularization, rHDL infusions enhanced blood perfusion and augmented capillary and arteriolar density. Infusion of a VEGFR2-neutralizing antibody ablated those proangiogenic effects of rHDL. Circulating Sca1+/CXCR4+ angiogenic progenitor cell levels, important for neovascularization in response to ischemia, were higher in rHDL-infused mice 3 d after ischemic induction, but that did not occur in mice that also received the VEGFR2-neutralizing antibody. In summary, VEGFR2 has a key role in the proangiogenic effects of rHDL in hypoxia/ischemia. These findings have therapeutic implications for angiogenic diseases associated with an impaired response to tissue ischemia.Carla M. Cannizzo, Aaron A. Adonopulos, Emma L. Solly, Anisyah Ridiandries, Laura Z. Vanags, Jocelyne Mulangala, Sui Ching G. Yuen, Tania Tsatralis, Rodney Henriquez, Stacy Robertson, Stephen J. Nicholls, Belinda A. Di Bartolo, Martin K.C. Ng, Yuen Ting Lam, Christina A. Bursill, and Joanne T.M. Ta