New species of carex from Mexico and Guatemala

C.subbracteata var.tolucensis (sect.Ovales) andC. distentiformis (sect.Extensae) from Mexico,C. guatemalensis (sect.Longicaules) andC. caxinensis (sect.Viridiflorae) from Guatemala, andC. volcanica (sect.Ovales) from Mexico and Guatemala are described as new.     

The loss of health status in rheumatoid arthritis and the effect of biologic therapy: a longitudinal observational study

Introduction  

The long-term course of rheumatoid arthritis (RA) in terms of health status is not well understood, nor is the degree of effectiveness of biologic therapy in the community. We modeled the progression of loss of health status, and measured incremental costs and effectiveness of biologic therapy in the community.     

Cretaceous Therian Tarsals and the Metatherian-Eutherian Dichotomy

Diverse metatherian and eutherian tarsal remains from the Late Cretaceous (middle-late Turonian) Bissekty Formation, Kyzylkum Desert, Uzbekistan (ca 90 MYA) are described. Their functional and taxonomic properties, along with those of other tarsal evidence, led to a reassessment of polarity hypotheses of therian, metatherian, and eutherian cruropedal attributes, and the consequences of this for phylogeny of taxa. There are calcaneal remains of several types of marsupials, and a single astragalus that probably belongs to one of these. This represents greater taxonomic diversity than the dental record suggests. Exceptionally large and distally extending peroneal processes, and small and steeply angled calcaneocuboid articulations facing mediodistally, as seen in the Early Cretaceous Sinodelphys and other Cretaceous and Paleogene taxa, attest not only to the metatherian status of these specimens, but also to the retention of many ancestral therian features, even more so than in both the Tiupampa and Itaboraí marsupials of the South American Paleocene (both the calcanea and the astragalus suggest therian traits that are decidedly unlike those of symmetrodonts). Calcanea allocated to the deltatherioid species at Bissekty testify unequivocally to their metatherian affinity. The morphology of the best represented sample of eutherian calcanea from Bissekty, presumably of a number of zhelestid species, appears to be more derived than that of the Late Cretaceous/Paleocene Protungulatum in having a much more reduced peroneal process and a calcaneocuboid articulation that faces distally, oriented nearly at a 90° angle to the long axis of the calcaneus. In fact, this distally facing facet, common in later eutherians (except for lineages in the Paleogene record, and various Carnivora), may not be diagnostic of either the protoeutherian, or even of the protoplacentalian, in spite of its presence in Eomaia. Many putatively “basal” lineages have derived characters, hence such outgroups should not be considered the unequivocal repositories of only ancestral character states.     

Improvement of thermostable aldehyde dehydrogenase by directed evolution for application in Synthetic Cascade Biomanufacturing

The aldehyde dehydrogenase from Thermoplasma acidophilum, which was previously implemented as a key enzyme in a synthetic cell-free reaction cascade for the production of alcohols, was optimized by directed evolution. Improvements have been made to enhance reaction velocity and solubility. Using a random approach followed by site-directed and saturation mutagenesis, three beneficial amino acid mutations were found after screening of ca. 20,000 variants. Mutation Y399C enhanced the protein solubility after recombinant expression in Escherichia coli 6-fold. Two further mutations, F34M and S405N, enhanced enzyme activity with the cofactor NAD⁺ by a factor of eight. Impacts on enzyme stability and substrate specificity were negligible.     

Macrophages Create an Acidic Extracellular Hydrolytic Compartment to Digest Aggregated Lipoproteins

A critical event in atherogenesis is the interaction of macrophages with subendothelial lipoproteins. Although most studies model this interaction by incubating macrophages with monomeric lipoproteins, macrophages in vivo encounter lipoproteins that are aggregated. The physical features of the lipoproteins require distinctive mechanisms for their uptake. We show that macrophages create an extracellular, acidic, hydrolytic compartment to carry out digestion of aggregated low-density lipoproteins. We demonstrate delivery of lysosomal contents to these specialized compartments and their acidification by vacuolar ATPase, enabling aggregate catabolism by lysosomal acid hydrolases. We observe transient sealing of portions of the compartments, allowing formation of an “extracellular” proton gradient. An increase in free cholesterol is observed in aggregates contained in these compartments. Thus, cholesteryl ester hydrolysis can occur extracellularly in a specialized compartment, a lysosomal synapse, during the interaction of macrophages with aggregated low-density lipoprotein. A detailed understanding of these processes is essential for developing strategies to prevent atherosclerosis.     

RNAi-Mediated Gene Suppression in a GCAP1(L151F) Cone-Rod Dystrophy Mouse Model

Dominant mutations occurring in the high-affinity Ca²⁺-binding sites (EF-hands) of the GUCA1A gene encoding guanylate cyclase-activating protein 1 (GCAP1) cause slowly progressing cone-rod dystrophy (CORD) in a dozen families worldwide. We developed a nonallele-specific adeno-associated virus (AAV)-based RNAi knockdown strategy to rescue the retina degeneration caused by GCAP1 mutations. We generated three genomic transgenic mouse lines expressing wildtype (WT) and L151F mutant mouse GCAP1 with or without a C-terminal GFP fusion. Under control of endogenous regulatory elements, the transgenes were expressed specifically in mouse photoreceptors. GCAP1(L151F) and GCAP1(L151F)-GFP transgenic mice presented with a late onset and slowly progressive photoreceptor degeneration, similar to that observed in human GCAP1-CORD patients. Transgenic expression of WT GCAP1-EGFP in photoreceptors had no adverse effect. Toward therapy development, a highly effective anti-mGCAP1 shRNA, mG1hp4, was selected from four candidate shRNAs using an in-vitro screening assay. Subsequently a self-complementary (sc) AAV serotype 2/8 expressing mG1hp4 was delivered subretinally to GCAP1(L151F)-GFP transgenic mice. Knockdown of the GCAP1(L151F)-GFP transgene product was visualized by fluorescence live imaging in the scAAV2/8-mG1hp4-treated retinas. Concomitant with the mutant GCAP1-GFP fusion protein, endogenous GCAP1 decreased as well in treated retinas. We propose nonallele-specific RNAi knockdown of GCAP1 as a general therapeutic strategy to rescue any GCAP1-based dominant cone-rod dystrophy in human patients.     

Protonation of the Human PIEZO1 Ion Channel Stabilizes Inactivation

PIEZO1 is a recently cloned eukaryotic cation-selective channel that opens with mechanical force. We found that extracellular protonation inhibits channel activation by ≈90% by increased occupancy in the closed or the inactivated state. Titration between pH 6.3 and 8.3 exhibited a pK of ≈6.9. The steepness of the titration data suggests positive cooperativity, implying the involvement of at least two protonation sites. Whole-cell recordings yielded results similar to patches, and pH 6.5 reduced whole-cell currents by >80%. The effects were reversible. To assess whether pH acts on the open or the inactivated state, we tested a double-mutant PIEZO1 that does not inactivate. Cell-attached patches and whole-cell currents from this mutant channel were pH-insensitive. Thus, protonation appears to be associated with domain(s) of the channel involved with inactivation. pH also did not affect mutant channels with point mutations at position 2456 that are known to exhibit slow inactivation. To determine whether the physical properties of the membrane are altered by pH and thereby affect channel gating, we measured patch capacitance during mechanical stimuli at pH 6.5 and 7.3. The rate constants for changes in patch capacitance were independent of pH, suggesting that bilayer mechanics are not involved. In summary, low pH stabilizes the inactivated state. This effect may be important when channels are activated under pathological conditions in which the pH is reduced, such as during ischemia.     

ARGoS: a modular, parallel, multi-engine simulator for multi-robot systems

We present a novel multi-robot simulator named ARGoS. ARGoS is designed to simulate complex experiments involving large swarms of robots of different types. ARGoS is the first multi-robot simulator that is at the same time both efficient (fast performance with many robots) and flexible (highly customizable for specific experiments). Novel design choices in ARGoS have enabled this breakthrough. First, in ARGoS, it is possible to partition the simulated space into multiple sub-spaces, managed by different physics engines running in parallel. Second, ARGoS?? architecture is multi-threaded, thus designed to optimize the usage of modern multi-core CPUs. Finally, the architecture of ARGoS is highly modular, enabling easy addition of custom features and appropriate allocation of computational resources. We assess the efficiency of ARGoS and showcase its flexibility with targeted experiments. Experimental results demonstrate that simulation run-time increases linearly with the number of robots. A 2D-dynamics simulation of 10,000 e-puck robots can be performed in 60?% of the time taken by the corresponding real-world experiment. We show how ARGoS can be extended to suit the needs of an experiment in which custom functionality is necessary to achieve sufficient simulation accuracy. ARGoS is open source software licensed under GPL3 and is downloadable free of charge.