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Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is an important malaria virulence factor. The protein family can be divided into clinically relevant subfamilies. ICAM-1-binding group A PfEMP1 proteins also bind endothelial protein C receptor and have been associated with cerebral malaria in children. IgG to these PfEMP1 proteins is acquired later in life than that to group A PfEMP1 not binding ICAM-1., Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is an important malaria virulence factor. The protein family can be divided into clinically relevant subfamilies. ICAM-1-binding group A PfEMP1 proteins also bind endothelial protein C receptor and have been associated with cerebral malaria in children. IgG to these PfEMP1 proteins is acquired later in life than that to group A PfEMP1 not binding ICAM-1. The kinetics of acquisition of IgG to group B and C PfEMP1 proteins binding ICAM-1 is unclear and was studied here. Gene sequences encoding group B and C PfEMP1 with DBLβ domains known to bind ICAM-1 were used to identify additional binders. Levels of IgG specific for DBLβ domains from group A, B, and C PfEMP1 binding or not binding ICAM-1 were measured in plasma from Ghanaian children with or without malaria. Seven new ICAM-1-binding DBLβ domains from group B and C PfEMP1 were identified. Healthy children had higher levels of IgG specific for ICAM-1-binding DBLβ domains from group A than from groups B and C. However, the opposite pattern was found in children with malaria, particularly among young patients. Acquisition of IgG specific for DBLβ domains binding ICAM-1 differs between PfEMP1 groups.

A Labyrinthula sp. was isolated from symptomatic rough bluegrass (Poa trivialis L.) and perennial ryegrass (Lolium perenne L.) from a golf course in Arizona. Initial symptoms were a water-soaked appearance and rapid collapse of small patches of turf foliage. The affected turf died, and patches coalesced to form large dead areas after several weeks. The symptoms were those of the disease recently termed “rapid blight” for which the causal agent has not been identified (1). Rapid blight was first observed in southern California in 1995 and has become increasingly problematic in 10 other states on several cool-season turfgrasses (1). In Arizona, it is associated with high salinity irrigation water. In microscopic examinations of symptomatic P. trivialis and L. perenne leaf tissue from November 2002 to February 2003, fusiform or spindle-shaped vegetative cells (4 to 5 × 15 to 20 μm) were observed in leaf cells. These cells are consistently associated with rapid blight (1) and are typical in size and shape of those described for Labyrinthula spp. (3,4). The fusiform cells were cultured in 1% horse serum water agar medium made with irrigation water (electrical conductivity [EC] = 3.5 to 4.0 dS/m) from a golf course in central Arizona with rapid blight. The cells readily formed colonies on this medium and exhibited gliding motility along a network of hyaline slime filaments as previously described for the genus Labyrinthula (3,4). Koch's postulates were fulfilled by inoculating P. trivialis and L. perenne seedlings with Labyrinthula sp. isolated from naturally infested P. trivialis in two experiments. The grasses were started from seed and grown as a lawn in containers in the laboratory. Both experiments were repeated once. In the first experiment, infested autoclaved leaf pieces of P. trivialis were used as inoculum. Inoculated leaf pieces were placed within each of several bundles of 4 to 6 leaves and held loosely in place with a 0.5-cm wide ring of tygon tubing. Seedlings were irrigated with sterilized irrigation water from the golf course (EC = 4.0 dS/m). In the second experiment, agar discs from Labyrinthula sp. colonies on 1% horse serum agar were used as inoculum by placing the agar discs in contact with leaves. Seedlings were irrigated with sterile tap water adjusted to 4.0 dS/m using synthetic sea salt (Instant Ocean, Aquarium Systems, Inc., Mentor, OH) Leaf tissue of all inoculated seedlings became water soaked within 3 to 7 days and collapsed within 10 days in both experiments. Fusiform cells were observed in inoculated leaf tissue cells, and the Labyrinthula sp. was reisolated from 100% of selected symptomatic seedlings. Control seedlings treated with noninfested leaf pieces or sterile agar pieces did not develop symptoms, and no fusiform cells were isolated from the leaf tissue. Labyrinthula spp. are usually associated with marine systems (3). Labyrinthula zosterae D. Porter & Muehlst. has been identified as the causal agent in a marine grass wasting disease (2), but to our knowledge, no Labyrinthula spp. have been described as pathogens of terrestrial plants. References: (1) S. B. Martin et al. Phytopathology (Abstr.) 92:(suppl)S52, 2002. (2) L. K. Muehlstein et al. Mycologia 83:180, 1991. (3) K. S. Porkorny. J. Protozool. 14:697, 1967. (4) D. Porter. Handbook of Protoctista. Jones and Bartlett, Boston, MA, 1990.

Different strains of Xylella fastidiosa cause a variety of significant disease problems in agricultural and ornamental plants, including Pierce's disease in grapes, oleander leaf scorch, pecan bacterial leaf scorch, and alfalfa dwarf disease. X. fastidiosa has never been reported in New Mexico but is known to exist in surrounding states (California, Arizona, and Texas). During the summer of 2006, several chitalpa (Chitalpa tashkinensis) hybrid trees with leaf scorch symptoms and branch die back were observed in Las Cruces, NM and they tested positive for X. fastidiosa by ELISA. Additional samples from these plants and others were analyzed by ELISA, PCR (2), and cultured on XfD2 medium (1). Known positive and negative oleander samples from Arizona were included as controls. Fifteen of thirty tested chitalpa were PCR and ELISA positive, indicating that they were infected with X. fastidiosa. Bacterial colonies that were PCR positive were also recovered from 10 of the XF positive samples that were plated. DNA sequences of PCR products amplified from chitalpa and isolated bacterial colonies (GenBank Accession Nos. EF109936 and EF109937) were identical to each other, 97% similar to X. fastidiosa strain JB-USNA, and 96% similar to the Temecula 1 strain. Independent ELISA testing (Barry Hill, California Department Food and Agriculture, Sacramento, CA) confirmed our ELISA and PCR results. On the basis of these results, we conclude that X. fastidiosa is present in New Mexico and that the common landscape ornamental chitalpa is a host for X. fastidiosa. Additional work is required to determine if X. fastidiosa is pathogenic to chitalpa and to examine the relevance of this potential X. fastidiosa reservoir to agricultural production in New Mexico and other areas where chitalpa is grown. References: (1) R. P. P. Almeida et al. Curr. Microbiol. 48:368, 2004. (2) M. R. Pooler et al. Lett. Appl. Microbiol. 25:123, 1997.

Bright yellow, interveinal chlorosis was observed for the first time on leaves of the older and mid-growth of cucurbit plants in southern Arizona and Sonora (Mexico) during September and October of 2006. Some cultivars exhibited substantial yield losses of 30 to 80%. In Arizona, symptoms were in Cucumis melo (muskmelon and honeydew melon) fields in the Yuma Valley and Hyder. In Sonora, honeydew and muskmelon, Cucurbita pepo (acorn, spaghetti, and summer [yellow and zucchini] squash), and Citrullus lanatus (watermelon) were symptomatic in Hermosillo, whereas, in Caborca, honeydew and cantaloupe developed similar symptoms. Interveinal chlorosis was observed in 60 to 100% of the plants in each field. Crops planted mid-to-late season were 100% infected, whereas, the early-season fields experienced approximately 60 to 80% incidence. All symptomatic fields in the Sonoran Desert and vicinity were infested by the whitefly Bemisia tabaci (Genn.), which was identified as the ‘B biotype’ on the basis of mitochondria COI sequence analysis (data not shown). Whitefly population levels were variable and ranged from 5 to 200 per plant. Total RNA was isolated from leaf samples collected from symptomatic plants using Tri Reagent (Molecular Research Center, Cincinnati, OH). Purified RNA was used in reverse transcriptase-PCR with primers specific to the Cucurbit yellow stunting disorder virus (CYSDV) coat protein (CP) gene (RNA2-deoxyribonucleotide coordinates 4927-4950 and 5657-5679) for the suspected whitefly-transmitted bipartite CYSDV (4). PCR yielded the CYSDV CP fragment, at 753 bp (GenBank Accession Nos. EF21058 and EF21059), which was cloned into pGEM T-Easy and sequenced in both directions using universal primers. The CYSDV CP nucleotide sequences (n = 16) obtained from acorn squash, honeydew melon, muskmelon, yellow squash, and watermelon had 99 to 100% identity. The Arizona (AZ) and Sonora (SON) CYSDV CP sequences shared 99 to 100% identity with previously described CYSDV isolates from the Eastern Hemisphere (GenBank Accession Nos. DQ903105 and DQ903108) and also with two isolates of CYSDV collected during 2004 from Zacapa Valley, Guatemala (GenBank Accession Nos. EF21060 and EF21061) (J. K. Brown, unpublished data). CYSDV is a member of the genus Crinivirus, family Closteroviridae. CYSDV was first identified in cucumber and melon crops in the Middle East approximately 15 years ago and 10 years ago in Spain (1). Most recently, this virus was introduced into Texas (2), Guatemala (J. K. Brown, unpublished data), and Arizona and California (3). CYSDV has therefore emerged as an important and potentially worldwide threat to the production of cultivated cucurbits (3). The threat appears to be significant in light of the introduction or establishment of the exotic B. tabaci biotypes B and Q vectors, which also originated in the Middle Eastern-North African-Mediterranean region. To our knowledge, this is the first report of CYSDV infecting field-grown C. pepo (four types) and watermelon, reported previously only as experimental laboratory hosts, and of CYSDV in two types of melon (C. melo) in Mexico. References: (1) A. Celix et al. Phytopathology 86:1370, 1996. (2) J. Kao et al. Plant Dis. 84:101, 2000. (3) Y.-W. Kuo et al. Plant Dis. 91:330, 2007. (4) L. Rubio et al. J. Gen. Virol. 82:929, 2001.

Background Duplications of 15q11.2-q13.1 (Dup15q syndrome), including the paternally imprinted gene UBE3A and three nonimprinted gamma-aminobutyric acid type-A (GABAA) receptor genes, are highly penetrant for neurodevelopmental disorders such as autism spectrum disorder (ASD). To guide targeted treatments of Dup15q syndrome and other forms of ASD, biomarkers are needed that reflect molecular mechanisms of pathology. We recently described a beta EEG phenotype of Dup15q syndrome, but it remains unknown which specific genes drive this phenotype. Methods To test the hypothesis that UBE3A overexpression is not necessary for the beta EEG phenotype, we compared EEG from a reference cohort of children with Dup15q syndrome (n = 27) to (1) the pharmacological effects of the GABAA modulator midazolam (n = 12) on EEG from healthy adults, (2) EEG from typically developing (TD) children (n = 14), and (3) EEG from two children with duplications of paternal 15q (i.e., the UBE3A-silenced allele). Results Peak beta power was significantly increased in the reference cohort relative to TD controls. Midazolam administration recapitulated the beta EEG phenotype in healthy adults with a similar peak frequency in central channels (f = 23.0 Hz) as Dup15q syndrome (f = 23.1 Hz). Both paternal Dup15q syndrome cases displayed beta power comparable to the reference cohort. Conclusions Our results suggest a critical role for GABAergic transmission in the Dup15q syndrome beta EEG phenotype, which cannot be explained by UBE3A dysfunction alone. If this mechanism is confirmed, the phenotype may be used as a marker of GABAergic pathology in clinical trials for Dup15q syndrome. Electronic supplementary material The online version of this article (10.1186/s13229-019-0280-6) contains supplementary material, which is available to authorized users.

Following publication of the original article [1], we have been notified that the Ethics statement of the articles should be changed. The Ethics statement now reads

Northern East Asia was inhabited by modern humans as early as 40 thousand years ago (ka), as demonstrated by the Tianyuan individual. Using genome-wide data obtained from 25 individuals dated to 33.6-3.4 ka from the Amur region, we show that Tianyuan-related ancestry was widespread in northern East Asia before the Last Glacial Maximum (LGM). At the close of the LGM stadial, the earliest northern East Asian appeared in the Amur region, and this population is basal to ancient northern East Asians. Human populations in the Amur region have maintained genetic continuity from 14 ka, and these early inhabitants represent the closest East Asian source known for Ancient Paleo-Siberians. We also observed that EDAR V370A was likely to have been elevated to high frequency after the LGM, suggesting the possible timing for its selection. This study provides a deep look into the population dynamics of northern East Asia.

Ethnopharmacological relevance Previously, the phytochemical constituents of Biebersteinia heterostemon Maxim (BHM) and Arenaria kansuensis Maxim (AKM) were studied and the evaluation of anxiolytic effect based on their extracts was also investigated. The two traditional Tibetan herbs, BHM and AKM, have been widely used in Qinghai-Tibet Plateau for cardiopulmonary disorders and neuropsychiatric diseases. The anxiolytic activities of a number of agents mediated by α2/3-containing GABAA receptors (GABAARs) have been demonstrated through the genetic and pharmacological studies. Flavonoids, such as flavones and flavanols, are a class of ligands that act at GABAARs and exhibit anxiolytic effects in vivo. Here, the flavonoids are the predominant constituents isolated from BHM and AKM. And our purpose is to investigate structure-activity relationships of the flavonoid compounds with binding to BZ-S of GABAAR complexes, and to search for anxiolytic constituents that lack undesirable-effects such as sedation and myorelaxation. Materials and methods The flavonoid constituents were separated and purified through the repeatedly silica gel or/and C18 column chromatography. The affinities of the compounds for BZ-S of GABAARs were detected by the radioreceptor binding assay with bovine cerebellum membranes, in which the different recombinant subunits-containing GABAARs were expressed in HEK 293T cells. The behavior tests, including elevated plus maze, locomotor activity, holeboard, rotarod and horizontal wire, were used to determine and evaluate the anxiolytic, sedative, and myorelaxant effects of these flavonoids. Results Eleven total flavonoid compounds were obtained from the Tibetan herbs (BHM and AKM). The flavones with 6-and/or 8-OMe possessed the most potent binding affinity to GABAARs, which were based on the result of structure-activity relationships analysis. Demethoxysudachitin (DMS, Ki = 0.59 μM), a flavone that binds to recombinant α1-3/5 subunit-containing GABAARs, was isolated from BHM, and exhibited high anxiolytic activity, without inducing sedation and myorelaxation. Moreover, the anxiolytic effect of DMS was antagonized by flumazenil, suggesting that a mode of action was mediated via the BZ-S of GABAARs. Conclusions This present study indicated that the flavones, especially DMS, are novel GABAAR ligands and therapeutic potential candidates for anxiety.

In the Northern Hemisphere, the Last Glacial Maximum (LGM) is recognized as a cold and dry period that marks the maximum southward extension of the Scandinavian Inlands in Europe. In Asia, the ice sheet did not expand from the Arctic into Siberia, yet the LGM had a significant impact at high latitudes and elevations, as well as in regions with a continental climate. How much these changes affected the human occupation of Siberia and Mongolia is still a matter of debate and various models dealing with continuity, discontinuity, demographic movement and adaptation have been put forth. The present paper is a critical review of available empirical data regarding the impact of the LGM on landscapes and human settlements in Mongolia. This review underscores the caveats in the data collected and further analyses are proposed to test several basic hypotheses. The results obtained suggest that during Marine Isotope Stages (MIS) 3 and MIS 2, there were hiatuses in the human occupation of Mongolia. These gaps are potentially linked with significant changes in climate. It is recognized that one of the main breaks in the cultural sequence is associated with the LGM, suggesting that Mongolia experienced periods of depopulation associated with this dramatic climatic change.

In the last few decades, there has been a decline in average sleep duration and quality that has adversely affected general health of the population. Increased use of artificial lighting, television, noise, and light emitting gadgets in the post-industrial society has led to an environment full of factors that can adversely affect sleep. Sleep is essential for the restoration of health and well-being, and has a direct effect on the quality of life of an individual. It is thus imperative to design constructions that provide a sleep permissive environment. In addition to environmental factors, sleep and wakefulness in humans is affected by a multitude of physiological parameters as well. The mechanisms and interplay between these factors that influence sleep permissiveness or promotion have received relatively little attention. The present review takes a holistic approach in explaining the basic circadian and homeostatic processes that influence sleep and sleepiness, and then delves deeper into how the thermal rhythms of the body affect these processes. It also discusses previous research and models in construction design that influence ambient temperature, skin surface temperature, and other thermoregulatory factors that can be controlled for optimal sleep.

Purpose To describe an economic (Ec) model for estimating the impact of screening and treatment for retinopathy of prematurity (ROP). Design EcROP is a cost-effectiveness, cost-utility, and cost-benefit analysis. Methods We surveyed caregivers of 52 children at schools for the blind or pediatric eye clinics in Atlanta, Georgia and 43 in Mexico City. A decision analytic model with sensitivity analysis determined the incremental cost-effectiveness (primary outcome) and incremental monetary benefit (secondary outcome) of an ideal (100% screening) national ROP program as compared to estimates of current practice. Direct costs included screening and treatment expenditures. Indirect costs estimated lost productivity of caretaker(s) and blind individuals as determined by face-to-face surveys. Utility and effectiveness were measured in quality-adjusted life years and benefit in US dollars. EcROP includes a sensitivity analysis to assesses the incremental cost-effectiveness and societal impact of ROP screening and treatment within a country or economic region. Estimates are based on evidence-based clinical data and region-specific economic data acquired from direct field survey. Results In both Mexico and the United States, an ideal national ROP screening and treatment program was highly cost-saving. The incremental net benefit of an ideal ROP program over current practice is $5556 per child ($206 574 333 annually) and $3628 per child ($205 906 959 annually) in Mexico and the United States, respectively. Conclusion EcROP demonstrates that ROP screening and treatment is highly beneficial for quality of life, cost saving, and cost-effectiveness in the United States and Mexico. EcROP can be applied to any country or region to provide data for informed allocation of limited health care resources.

Edible films may be used in food packaging, for which they must deliver good barrier and mechanical properties. Films based on proteins have good gas barrier and mechanical properties, but poor water barrier properties. Films made from lipids have good water barrier properties, but poor mechanical properties. Protein and lipids were then combined to form composite films, and the particle size of the lipid phase was reduced to evaluate its effect on the mechanical and barrier properties. Micro and nanodroplets of Acetem and Tween 60 were added into an isolated soy protein (ISP) solution. Oil-in-water droplets were formed by direct emulsification at 1300 rpm for 30 min (microdroplets) or 5 h (nanodroplets), dispersed into the ISP solution, and cast into films. Emulsified films showed reduced strength and increased elongation, indicating a plasticizing effect of emulsions. The water barrier properties were either unchanged or slightly improved by the addition of hydrophobic compounds. Reducing droplet size improved the barrier properties as surface area of the lipid increased. Carvacrol and cinnamaldehyde were also added, and they either improved or unaffected the mechanical and barrier properties of ISP films. Addition of emulsions and reduction of droplet size generally decreased total pore volume in the films as well as water vapor permeation, indicating that the microvoid model suitably explains the water barrier properties of ISP films. This work further clarifies possible differences in barrier properties by examining water diffusivity and adsorption/desorption kinetics.

Retinal Vein Occlusions Preferred Practice Pattern® Guidelines New evidence-based Retinal Vein Occlusions Preferred Practice Pattern® (PPP) guidelines, discussing the prognosis and risk factors of retinal vein occlusions and the treatment options.

This is one of the latest volumes in the distinguished series Transformation of the Classical Heritage (Peter Brown, general editor). It is a history of early Arianism, but less of the Arianism of ...

Watermelon (Citrullus lanatus) juice is known for its refreshing flavor, but its high perishability limits its availability throughout the year. Watermelon juice concentrate has extended shelf-life and lower transportation and storage costs, but the conventional thermal evaporation process for concentrating juice degrades the nutritional components and sensory quality of the product. Thus, in this work, a large-scale, non-thermal forward osmosis (FO) process was used to concentrate fresh watermelon juice up to 65&deg, Brix. The FO concentrate was compared to thermal concentrate and fresh juices, and to commercially available refrigerated watermelon juices, in terms of lycopene and citrulline content, total soluble phenolics, antioxidant activity, and sensory properties. The FO concentrate had statistically similar (p &lt, 0.05) levels of all the nutrients of interest except antioxidant activity, when compared to the thermal concentrate. The reconstituted FO concentrate maintained the same antioxidant activity as the raw source juice, which was 45% higher than that of the reconstituted thermal concentrate. Sensory results showed that reconstituted FO concentrate resulted in highly liked juice, and it outperformed the reconstituted thermal concentrate in the sensory hedonic rating. This work demonstrates the possibility to produce a high-quality watermelon juice concentrate by forward osmosis.

We recently identified three novel thioredoxin-like genes in the genome of the protozoan parasite Plasmodium that belong to the Phosducin-like family of proteins (PhLP). PhLPs are small cytosolic proteins hypothesized to function in G-protein signaling and protein folding. Although PhLPs are highly conserved in eukaryotes from yeast to mammals, only a few representatives have been experimentally characterized to date. In addition, while PhLPs contain a thioredoxin domain, they lack a CXXC motif, a strong indicator for redox activity, and it is unclear whether members of the PhLP family are enzymatically active. Here, we describe PbPhLP-3 as the first phosducin-like protein of a protozoan organism, Plasmodium berghei. Initial transcription analysis revealed continuous low-level expression of pbphlp-3 throughout the complex Plasmodium life cycle. Attempts to knockout pbphlp-3 in P. berghei did not yield live parasites, suggesting an essential role for the gene in Plasmodium. We cloned, expressed and purified PbPhLP-3 and determined that the recombinant protein is redox active in vitro in a thioredoxin-coupled redox assay. It also has the capacity to reduce the organic compound tert-Butyl hydroperoxide (TBHP) in vitro, albeit at low efficiency. Sequence analysis, structural modeling, and site-directed mutagenesis revealed a conserved cysteine in the thioredoxin domain to be the redox active residue. Lastly, we provide evidence that recombinant human PhLP-3 exhibits redox activity similar to that of PbPhLP-3 and suggest that redox activity may be conserved in PhLP-3 homologs of other species. Our data provide new insight into the function of PhLP-3, which is hypothesized to act as co-chaperones in the folding and regulation of cytoskeletal proteins. We discuss the potential implications of PhLP-3 as a thioredoxin-target protein and possible links between the cellular redox network and the eukaryotic protein folding machinery.

There is a need for treatments that can safely promote regulatory lymphocyte responses. T cells express GABA receptors (GABAA-Rs) and GABA administration can inhibit Th1-mediated processes such as type 1 diabetes and rheumatoid arthritis in mouse models. Whether GABAA-R agonists can also inhibit Th17-driven processes such as experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis (MS), is an open question. GABA does not pass through the blood-brain barrier (BBB) making it ill-suited to inhibit the spreading of autoreactivity within the CNS. Homotaurine is a BBB-permeable amino acid that antagonizes amyloid fibril formation and was found to be safe but ineffective in long-term Alzheimer’s disease clinical trials. Homotaurine also acts as GABAA-R agonist with better pharmacokinetics than that of GABA. Working with both monophasic and relapsing-remitting mouse models of EAE, we show that oral administration of homotaurine can (1) enhance CD8+CD122+PD-1+ and CD4+Foxp3+ Treg, but not Breg, responses, (2) inhibit autoreactive Th17 and Th1 responses, and (3) effectively ameliorate ongoing disease. These observations demonstrate the potential of BBB-permeable GABAA-R agonists as a new class of treatment to enhance CD8+ and CD4+ Treg responses and limit Th17 and Th1-medaited inflammation in the CNS.

Background To describe a case of von Hippel-Lindau disease with peripheral retinal nonperfusion. Case presentation A 66-year-old female with known cerebellar and midbrain hemangioblastomas was evaluated for a retinal hemangioblastoma in the right eye. She underwent widefield fluorescein angiography, which showed hyperfluorescence localized to the hemangioblastoma surrounded by peripheral retinal nonperfusion in the same quadrant. Conclusions Further widefield imaging studies are required to determine if peripheral retinal nonperfusion is a common finding in von Hippel-Lindau disease.

Background Age-related macular degeneration (AMD) is a degenerative disorder of the central retina and the foremost cause of blindness. The retinal pigment epithelium (RPE) is a primary site of disease pathogenesis. The genetic basis of AMD is relatively well understood; however, this knowledge is yet to yield a treatment for the most prevalent non-neovascular disease forms. Therefore, tissue-specific epigenetic mechanisms of gene regulation are of considerable interest in AMD. We aimed to identify differentially methylated genes associated with AMD in the RPE and differentiate local DNA methylation aberrations from global DNA methylation changes, as local DNA methylation changes may be more amenable to therapeutic manipulation. Methods Epigenome-wide association study and targeted gene expression profiling were carried out in RPE cells from eyes of human donors. We performed genome-wide DNA methylation profiling (Illumina 450k BeadChip array) on RPE cells from 44 human donor eyes (25 AMD and 19 normal controls). We validated the findings using bisulfite pyrosequencing in 55 RPE samples (30 AMD and 25 normal controls) including technical (n = 38) and independent replicate samples (n = 17). Long interspersed nucleotide element 1 (LINE-1) analysis was then applied to assess global DNA methylation changes in the RPE. RT-qPCR on independent donor RPE samples was performed to assess gene expression changes. Results Genome-wide DNA methylation profiling identified differential methylation of multiple loci including the SKI proto-oncogene (SKI) (p = 1.18 × 10−9), general transcription factor IIH subunit H4 (GTF2H4) (p = 7.03 × 10−7), and Tenascin X (TNXB) (p = 6.30 × 10−6) genes in AMD. Bisulfite pyrosequencing validated the differentially methylated locus cg18934822 in SKI, and cg22508626 within GTF2H4, and excluded global DNA methylation changes in the RPE in AMD. We further demonstrated the differential expression of SKI, GTF2H4, and TNXB in the RPE of independent AMD donors. Conclusions We report the largest genome-wide methylation analysis of RPE in AMD along with associated gene expression changes to date, for the first-time reaching genome-wide significance, and identified novel targets for functional and future therapeutic intervention studies. The novel differentially methylated genes SKI and GTF2H4 have not been previously associated with AMD, and regulate disease pathways implicated in AMD, including TGF beta signaling (SKI) and transcription-dependent DNA repair mechanisms (GTF2H4). Electronic supplementary material The online version of this article (10.1186/s13148-019-0608-2) contains supplementary material, which is available to authorized users.

Importance Identifying and prioritizing unanswered clinical questions may help to best allocate limited resources for research associated with the treatment of age-related macular degeneration (AMD). Objective To identify and prioritize clinical questions and outcomes for research associated with the treatment of AMD through engagement with professional and patient stakeholders. Design, Setting, and Participants Multiple cross-sectional survey questions were used in a modified Delphi process for panel members of US and international organizations, the American Academy of Ophthalmology (AAO) Retina/Vitreous Panel (n=7), health care professionals from the American Society of Retinal Specialists (ASRS) (n=90), Atlantic Coast Retina Conference (ACRC) and Macula 2017 meeting (n=34); and patients from MD (Macular Degeneration) Support (n=46). Data were collected from January 20, 2015, to January 9, 2017. Main Outcomes and Measures The prioritizing of clinical questions and patient-important outcomes for AMD. Results Seventy clinical questions were derived from the AAO Preferred Practice Patterns for AMD and suggestions by the AAO Retina/Vitreous Panel. The AAO Retina/Vitreous Panel assessed all 70 clinical questions and rated 17 of 70 questions (24%) as highly important. Health care professionals assessed the 17 highly important clinical questions and rated 12 of 17 questions (71%) as high priority for research to answer; 9 of 12 high-priority clinical questions were associated with aspects of anti–vascular endothelial growth factor agents. Patients assessed the 17 highly important clinical questions and rated all as high priority. Additionally, patients identified 6 of 33 outcomes (18%) as most important to them (choroidal neovascularization, development of advanced AMD, retinal hemorrhage, gain of vision, slowing vision loss, and serious ocular events). Conclusions and Relevance Input from 4 stakeholder groups suggests good agreement on which 12 priority clinical questions can be used to underpin research related to the treatment of AMD. The 6 most important outcomes identified by patients were balanced between intended effects of AMD treatment (eg, slowing vision loss) and adverse events. Consideration of these patient-important outcomes may help to guide clinical care and future areas of research.

Purpose To describe modern multimodal imaging of the choroidal and optic disk vessels in optic disk pits. Methods Case reports of four patients with optic disk pit who underwent multimodal imaging of the optic pit and surrounding structures. Patients included in this article were found to have optic disk pits and subsequently underwent multimodal imaging. Results Cilioretinal arteries were present in two of the four cases (50%). SPECTRALIS optical coherence tomography showed intraretinal and subretinal fluid in all cases. Small vessels in the choroid and in the disk around the pit were also present in all cases through optical coherence tomographic angiography. Confocal fluorescein angiographic imaging in the first case showed leakage from the vessels adjacent to the optic disk pit. Conclusion Modern multimodal imaging shows that there are anomalous vessels in and around an optic pit. Whether these vessels affect the development of optic pit, maculopathy needs to be further evaluated.

The trabecular meshwork (TM) plays a fundamental role in intraocular pressure regulation, but its mechanotransduction pathway is poorly understood. Yarishkin et al. show that the mechanosensing channel TREK-1 regulates TM membrane potential, pressure sensitivity, calcium homeostasis, and impedance., Mechanotransduction by the trabecular meshwork (TM) is an essential component of intraocular pressure regulation in the vertebrate eye. This process is compromised in glaucoma but is poorly understood. In this study, we identify transient receptor potential vanilloid isoform 4 (TRPV4) and TWIK-related potassium channel-1 (TREK-1) as key molecular determinants of TM membrane potential, pressure sensitivity, calcium homeostasis, and transcellular permeability. We show that resting membrane potential in human TM cells is unaffected by “classical” inhibitors of voltage-activated, calcium-activated, and inwardly rectifying potassium channels but is depolarized by blockers of tandem-pore K+ channels. Using gene profiling, we reveal the presence of TREK-1, TASK-1, TWIK-2, and THIK transcripts in TM cells. Pressure stimuli, arachidonic acid, and TREK-1 activators hyperpolarize these cells, effects that are antagonized by quinine, amlodipine, spadin, and short-hairpin RNA–mediated knockdown of TREK-1 but not TASK-1. Activation and inhibition of TREK-1 modulates [Ca2+]TM and lowers the impedance of cell monolayers. Together, these results suggest that tensile homeostasis in the TM may be regulated by balanced, pressure-dependent activation of TRPV4 and TREK-1 mechanotransducers.

Our previous studies on Asterothamnus centrali-asiaticus Novopokr. (ACN) and Arenaria kansuensis Maxim. (AKM) had led to the isolation of some phytochemical constituents and evaluation of anticonvulsant effect based on their extracts. ACN and AKM have been widely used in traditional Tibetan herbs for neuropsychiatric diseases and cardiopulmonary disorders.The purpose is to investigate structure-activity relationships of flavonoids isolated from ACN and AKM, for binding to the benzodiazepine site (BZ-S) of γ-aminobutyric acid type A (GABAThe affinities of these flavonoids for the BZ-S of GABAFifteen and thirteen flavonoids were isolated from ACN and AKM, respectively. Structure-activity relationships analysis indicated that 6-and/or 8-OMe flavones exhibited the most potent binding affinity to GABAThis study suggested that these flavones, especially DMF, are new BZ receptor ligands and prospective therapeutic candidates for seizures.

Purpose We measure and compare surgical devices using an ex vivo, temperature-controlled, choroidal incision model during thermal energy transfer with a high-resolution infrared camera. Methods Ex vivo porcine choroidal tissue specimens (n = 516) were isolated and placed on a temperature-regulated (37°C) perfusion platform. We tested the pulsed electron avalanche knife (PEAK), micropulse laser (MpL), continuous laser (CL), and bipolar cautery (BpC) at three energy settings (11 [low], 45 [medium], and 134 [high] mJ/mm). Each device was clamped to a stationary mechanical arm. Movement of tissue specimens beneath the surgical device was achieved using a stepping motor-driven x-y table. An infrared video camera measured orthogonal temperature variation in the surrounding tissue. Results Increased power resulted in greater lateral thermal spread using all modalities (P < 0.001). Mean (standard deviation) lateral thermal spread at low energy was smallest for the MpL at 0.0 (0.01) mm (P < 0.001), whereas BpC had the least collateral tissue damage at medium and high energies (0.02 [0.08] and 0.34 [0.22] mm, respectively; P < 0.001). Fluidics of the ex vivo system may limit thermal spread. The PEAK had the greatest thermal spread across all energy groups (P < 0.001), with clinically relevant variation between disposable blades. Conclusions Our ex vivo model enabled direct comparison of threshold thermal tissue injury across four devices. MpL and BpC showed the least thermal damage. PEAK had a higher variation in energy delivery, but also has the advantage of more effective tissue cutting. Translational Relevance Our ex vivo surgical device analysis provides thermal tissue injury predictions for choroidal surgery.

The Tibetan Plateau is the highest and one of the most demanding environments ever inhabited by humans. We investigated the timing and mechanisms of its initial colonization at the Nwya Devu site, located nearly 4600 meters above sea level. This site, dating from 40,000 to 30,000 years ago, is the highest Paleolithic archaeological site yet identified globally. Nwya Devu has yielded an abundant blade tool assemblage, indicating hitherto-unknown capacities for the survival of modern humans who camped in this environment. This site deepens the history of the peopling of the “roof of the world” and the antiquity of human high-altitude occupations more generally.

Cerebral malaria (CM) is a potentially deadly outcome of Plasmodium falciparum malaria that is precipitated by sequestration of infected erythrocytes (IEs) in the brain. The adhesion of IEs to brain endothelial cells is mediated by a subtype of parasite-encoded erythrocyte membrane protein 1 (PfEMP1) that facilitates dual binding to host intercellular adhesion molecule 1 (ICAM-1) and endothelial protein receptor C (EPCR). The PfEMP1 subtype is characterized by the presence of a particular motif (DBLβ_motif) in the constituent ICAM-1-binding DBLβ domain. The rate of natural acquisition of DBLβ_motif-specific IgG antibodies and the ability to induce such antibodies by vaccination are unknown, and the aim of this study was to provide such data. We used an enzyme-linked immunosorbent assay (ELISA) to measure DBLβ-specific IgG in plasma from Ghanaian children with malaria. The ability of human immune plasma and DBLβ-specific rat antisera to inhibit the interaction between ICAM-1 and DBLβ was assessed using ELISA and in vitro assays of IE adhesion under flow. The acquisition of DBLβ_motif-specific IgG coincided with age-specific susceptibility to CM. Broadly cross-reactive antibodies inhibiting the interaction between ICAM-1 and DBLβ_motif domains were detectable in immune plasma and in sera of rats immunized with specific DBLβ_motif antigens. Importantly, antibodies against the DBLβ_motif inhibited ICAM-1-specific in vitro adhesion of erythrocytes infected by four of five P. falciparum isolates from cerebral malaria patients. We conclude that natural exposure to P. falciparum as well as immunization with specific DBLβ_motif antigens can induce cross-reactive antibodies that inhibit the interaction between ICAM-1 and a broad range of DBLβ_motif domains. These findings raise hope that a vaccine designed specifically to prevent CM is feasible.

GABAA receptors (GABAARs) are the main inhibitory neurotransmitter receptors and mediate rapid synaptic as well as slow extrasynaptic inhibitory neurotransmission. Structurally, GABAARs are ligand-gated ion channels formed by a total of 19 homologous subunits, each with four transmembrane domains assembled as pentamers, forming a GABA-gated Cl– channels. The major classical synaptic GABAAR subtypes are formed by 2α2β and a γ subunit, with six different possible α subunits, three different β subunits, and three γ subunits, with the most abundant subtype, α1β2γ2 receptors. More recently, highly GABA-sensitive extrasynaptic δ subunit-containing receptors that are persistently (tonically) activated by low ambient levels of GABA have entered the limelight. GABAARs are targets for sedative/hypnotic and anxiolytic drugs (e.g., benzodiazepines [BZs] and other BZ site ligands), as well as general anesthetics (e.g., etomidate, propofol, barbiturate, and neurosteroid anesthetics, and possibly volatile agents and long-chain alcohols), and also are important targets for alcohol actions.

Contradictory data have been presented regarding the implication of the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome in age-related macular degeneration (AMD), the leading cause of vision loss in the Western world. Recognizing that antibody specificity may explain this discrepancy and in line with recent National Institutes of Health (NIH) guidelines requiring authentication of key biological resources, the specificity of anti-NLRP3 antibodies was assessed to elucidate whether non-immune RPE cells express NLRP3. Using validated resources, NLRP3 was not detected in human primary or human established RPE cell lines under multiple inflammasome-priming conditions, including purported NLRP3 stimuli in RPE such as DICER1 deletion and Alu RNA transfection. Furthermore, NLRP3 was below detection limits in ex vivo macular RPE from AMD patients, as well as in human induced pluripotent stem cell (hiPSC)-derived RPE from patients with overactive NLRP3 syndrome (Chronic infantile neurologic cutaneous and articulate, CINCA syndrome). Evidence presented in this study provides new data regarding the interpretation of published results reporting NLRP3 expression and upregulation in RPE and addresses the role that this inflammasome plays in AMD pathogenesis.

A new enzymatic assay for the bacterial enzyme succinyl-diaminopimelate desuccinylase (DapE, E.C. 3.5.1.18) is described. This assay employs N6-methyl-N2-succinyl-L,L-diaminopimelic acid (N6-methyl-L,L-SDAP) as the substrate with ninhydrin used to detect cleavage of the amide bond of the modified substrate, wherein N6-methylation enables selective detection of the primary amine enzymatic product. Molecular modeling supported preparation of the mono-N6-methylated-L,L-SDAP as an alternate substrate for the assay, given binding in the active site of DapE predicted to be comparable to the endogenous substrate. The alternate substrate for the assay, N6-methyl-L,L-SDAP, was synthesized from the tert-butyl ester of Boc-L-glutamic acid employing a Horner-Wadsworth-Emmons olefination followed by an enantioselective reduction employing Rh(I)(COD)(S,S)-Et-DuPHOS as the chiral catalyst. Validation of the new ninhydrin assay was demonstrated with known inhibitors of DapE from Haemophilus influenza (HiDapE) including captopril (IC50 = 3.4 [± 0.2] μM, 3-mercaptobenzoic acid (IC50 = 21.8 [±2.2] μM, phenylboronic acid (IC50 = 316 [± 23.6] μM, and 2-thiopheneboronic acid (IC50 = 111 [± 16] μM. Based on these data, this assay is simple and robust, and should be amenable to high-throughput screening, which is an important step forward as it opens the door to medicinal chemistry efforts toward the discovery of DapE inhibitors that can function as a new class of antibiotics.

The lack of suitable animal models for the study of cytoadhesion of P. falciparum-infected erythrocytes (IEs) has necessitated in vitro studies employing a range of cell lines of either human tumour origin (e.g., BeWo and C32 cells) or non-human origin (e.g., CHO cells). Of the human cells available, many were isolated from adults, or derived from a pool of donors (e.g., HBEC-5i). Here we demonstrate, for the first time, the successful isolation of blood outgrowth endothelial cells (BOECs) from frozen stabilates of peripheral blood mononuclear cells obtained from small-volume peripheral blood samples from paediatric malaria patients. BOECs are a sub-population of human endothelial cells, found within the peripheral blood. We demonstrate that these cells express receptors such as Intercellular Adhesion Molecule 1 (ICAM-1/CD54), Endothelial Protein C Receptor (EPCR/CD201), platelet/endothelial cell adhesion molecule 1 (PECAM-1/CD31), Thrombomodulin (CD141), and support adhesion of P. falciparum IEs.

Here, we present technological, typological and morphological analyses of the Pleistocene lithic assemblages excavated from Horizons 3–2.5 of the multicomponent Chikhen-2 site located on the southern piedmont of the Gobi Altai Range, southern Mongolia. Descriptions of geomorphology, stratigraphy, and archaeological finds are given, along with an analysis of reduction and secondary trimming techniques and retouch typology. Single-platform mono-frontal flat cores, double-platform bi-longitudinal cores, orthogonal cores, and narrow-front cores dominate the reduction strategy. Microcores are also present. Levallois-like cores appear initially in Horizon 2.7. Within the tool assemblage, retouched blades, end-scrapers on blades, notched-denticulate tools and large side-scrapers (skreblos) were identified. There are also various micro-tools present, including bladelets with blunt edges, oblique points, and truncated tools. The lithic industries of Horizons 3–2.5 are classified as Early Upper Paleolithic; a conclusion supported by radiocarbon determinations (ca. 30,000 BP). The lithic complexes described here are broadly analogous with other known Early Upper Paleolithic sites both in Mongolia (e.g., Chikhen-Agui; Orkhon-1 and -7; Tolbor-4, -15, and -16), and within a larger territory including the Russian Trans-Baikal region, North China, and the Altai Mountains. Archaeological materials from the lowermost strata at Chikhen-2 are important because they illustrate the emergence of the Upper Paleolithic in southern Mongolia.

Labyrinthula terrestris, the causal agent of rapid blight disease in cool-season turfgrasses, is the first Labyrinthula species found to be associated with higher plants in terrestrial ecosystems. An initial study suggested a single common genotype associated with turfgrass in the US, but a later study revealed genetic diversity in this pathogen. In the present study, genetic diversity as well as differences in pathogenicity and morphology was determined in Labyrinthula spp. from turfgrasses in New Mexico and Arizona where rapid blight had not previously been identified. In total, 96 Labyrinthula isolates from New Mexico and four from Arizona were sequenced in 2011 and 2012. A phylogenetic approach based on 18S and ITS-LSU of ribosomal DNA was used for genotypic diversity, and pathogenicity and microscopy were used for phenotypic diversity. A maximum parsimony analysis revealed four genetic groups including two previously described Labyrinthula groups, Laby10 (L. terrestris) and Laby31. Laby10 was the most predominant with 70 % of isolates followed by Laby31 with 24 %. Two new groups, Laby879 and Laby2020, contained only five and one isolate, respectively. An association between genotypes and phenotypes was observed. Laby10 and Laby31 were highly virulent as well as larger in cell size (14.9 and 18.1 μm, respectively), while Laby879 was less virulent with an average cell size of 8.5 μm. Laby2020 was the smallest (0.8 μm) and non-pathogenic. Results showed a prevalence of pathogenic and non-pathogenic Labyrinthula spp. in turfgrasses in New Mexico and the presence of the Laby31 group in the US for the first time.

A diblock copolymer, poly(ethylene glycol)-block-poly(lactic acid) (PEG-b-PLA), modulates the crystal growth of tolazamide (TLZ), resulting in a crystal morphology change from needles to plates in aqueous media. To understand this crystal surface drug-polymer interaction, we conducted molecular dynamics simulations on crystal surfaces of TLZ in water containing PEG-b-PLA. A 130-ns simulation of the polymer in a large water box was run before initiating 50 ns simulations with each of the crystal surfaces. The simulations demonstrated differentiated drug-polymer interactions that are consistent with experimental studies. Interaction of PEG-b-PLA with the (001) face occurred more rapidly (≤10 ns) and strongly (total interaction energy of -121.1 kJ/mol/monomer) than that with the (010) face (∼35 ns, -85.4 kJ/mol/monomer). There was little interaction with the (100) face. Hydrophobic and van der Waals (VDW) interactions were the dominant forces, accounting for more than 90% of total interaction energies. It suggests that polymers capable of forming strong hydrophobic and VDW interactions might be more effective in inhibiting crystallization of poorly water-soluble and hydrophobic drugs in aqueous media (such as gastrointestinal fluid) than those with hydrogen-bonding capacities. Such in-depth analysis and understanding facilitate the rational selection of polymers in designing supersaturation-based enabling formulations.

Glutathione peroxidases (GPx) comprise an important group of redox active proteins with diverse functions, including antioxidant defense and signaling. Although the genome of the malaria parasite Plasmodium does not contain a genuine GPx gene a glutathione peroxidase-like thioredoxin peroxidase (TPx(Gl)) has recently been identified and biochemically characterized in the human malaria parasite P. falciparum. To gain more insight into the potential biological function of this enzyme we have cloned and expressed TPx(Gl) of the rodent model system P. berghei (PbTPx(Gl)). Biochemical characterization confirmed that the protein is redox active with the P. berghei thioredoxin system. We compared PbTPx(Gl) to recently characterized thioredoxin-dependent GPx-type proteins of other organisms, and generated the first hypothetical 3D model of a Plasmodium TPx(Gl), which shows the conservation of the thioredoxin-fold as well as the spatial orientation of a classic GPx catalytic tetrad. In vivo studies indicate that PbTPx(Gl) is continuously expressed in all P. berghei asexual blood stages, gametocytes and in early mosquito-stage parasites. Confocal microscopy suggest a cytoplasmic localization of PbTPx(Gl) in all investigated parasite life stages, specifically in mature ookinetes. Our data provides new insights into the structure and ubiquitous expression of Plasmodium TPx(Gl) and warrants further investigation into this potentially important redox enzyme.

In current standard MIE tests for dusts various methods are proposed for synchronizing the electric spark with the transient dust cloud, but they all have a lower spark energy limit of 1–3 mJ. Some industrial dusts have MIEs in air significantly lower than 1 mJ. There is a need, therefore, for a synchronization method that works below 1 mJ. The present paper describes an improved version of the synchronized-spark generator developed by Randeberg et al. [25] . Adequate performance of the improved generator is indicated by a comparison of the MIE obtained for propane/air with values published by other workers.

Bacteria and plants produce glycogen and starch, respectively, as their primary storage polysaccharides. The synthesis of these polymers occurs with the donation of a glucose moiety from ADP-glucose (ADP-Glc) to an existing α-1,4-polyglucan chain. It is the production of ADP-Glc, catalyzed by the enzyme ADP-glucose pyrophosphorylase (ATP:α-D-glucose-1-phosphate adenylyltransferase; EC 2.7.7.27; ADP-Glc PPase), that acts as the rate limiting step in glycogen and starch production.1 This Mg2+ dependent reaction proceeds with ATP binding first to the enzyme, followed by glucose-1-phosphate (Glc1P), with the subsequent production of ADP-Glc and pyrophosphate.2,3 All known plant and bacterial ADP-Glc PPases are derived from a common ancestor and are predicted to share considerable structural similarities.4,5 These structural features are evident in the crystallographic structures of the potato tuber6 and Agrobacterium tumefaciens7 enzymes. Notably, the enzymes are tetramers with ∼50 kDa subunits. Enteric and cyanobacterial enzymes are homotetrameric (α4), whereas those of Firmicutes, unicellular algae, and higher plants are heterotetrameric (α2β2).4,5 In the latter, the two subunit types have similar structure and share a common ancestry, although it is sometimes the case that just one subunit type is catalytic.8,9 In most organisms, this enzyme is allosterically regulated by intermediates of the main carbon assimilatory pathway that exist within that species.4,5,10 In the case of E. coli, glycolysis is the major pathway and fructose-1,6-bisphosphate (FBP) is the primary activator of the E. coli ADP-Glc PPase.4 Of the ADP-Glc PPases that have been characterized, the E. coli enzyme has some of the largest changes in basal activity brought about by the activator. For instance, it increases specific activity and ATP affinity by at least 25- and 15-fold, respectively.11,12 Previous mutagenesis studies involving ADP-Glc PPases have provided some information regarding the allosteric activation mechanism of these enzymes. For example, studies on chimeric ADP-Glc PPases have shown that the C-terminal domain is important for allosteric effector specificity.12 Some alanine mutants of N-terminal arginine residues in the A. tumefaciens enzyme had altered allosteric responses, such as R32A which had significantly reduced affinity for the activator fructose-6-P.13 Several N-terminal residues in the E. coli ADP-Glc PPase have also been associated with allosteric activation. Lys39 is part of the FBP binding site,14 and Gln74, which is universally conserved amongst ADP-Glc PPases, is needed in order for FBP to exhibit regulatory role.11 The alanine mutant of the nearly universally conserved Trp113 had a similar kinetic profile to Q74A, even though FBP still binds to these enzymes.11 W116A and Q75A mutants of the potato tuber enzyme also fail to respond to allosteric activator.15 A model of the E. coli ADP-Glc PPase that highlights these regions is shown in Figure 1. Despite all of these data, it is not known how the allosteric signal is transmitted and what specific interactions are involved. Figure 1 Model of the E. coli ADP-Glc PPase subunit, shown in cartoon representation. For reference, the Pro103–Arg115 loop is colored red, the C-terminal domain is in orange, Asn38–Ala40 are colored blue, Thr73–Gln75 are yellow, Leu25–Gly30 ... Here, we use molecular dynamics (MD) to get insight into what residues are involved in an allosteric pathway in the E. coli ADP-Glc PPase. Allosteric effectors may alter communication pathways that exist within an enzyme.16 For that reason, correlated movement analysis, which examines cooperative motions within a protein, may be a useful tool for elucidating such interactions.17 However, because this technique does not consider the physical proximity of correlated regions, further analysis may be needed to identify spatially connected communication pathways. To that end, a network pathway analysis, which accounts for the physical connectivity of correlated regions, can help pinpoint such tracts.18 These techniques, along with an examination of some hydrogen bond networks and structural comparisons with other ADP-Glc PPases, suggested that the span of residues from Pro103 to Arg115 is important in an allosteric activation pathway in the E. coli enzyme. We tested this hypothesis by performing alanine scanning mutagenesis on the residues of this loop and kinetically characterizing the mutants, several of which were distinguished by a disruption of the allosteric response. In addition to this, sequence alignments of known ADP-Glc PPases that focused on this loop region, as well as structural alignments of different bacterial NDP-glucose pyrophosphorylases (NDP-Glc PPases), provided insight into the evolution of allosteric regulation in ADP-Glc PPases.

Human perfluorooctanesulfonate (PFOS) body burdens are attributable to both direct PFOS and indirect PFOS precursor (PreFOS) exposure. The relative importance of these two pathways has been estimated, but the relative temporal trajectory of exposure to PFOS and PreFOS has not been examined. Here, two hypothesized biomarkers of PreFOS exposure, PFOS isomer profiles (quantified as percent branched PFOS, %br-PFOS) and chiral 1m-PFOS enantiomer fractions (1m-PFOS EF) were analyzed in archived human serum samples of individual American adults (1974–2010) and pooled samples of Swedish primiparous women (1996–2010). After correcting for potential confounders, significant correlations between %br-PFOS and 1m-PFOS EFs were observed in American samples and in Swedish samples for the 1996–2000 period, supporting the hypothesis that both %br-PFOS and 1m-PFOS EF are biomarkers of PreFOS exposure. Significant trends of increasing %br-PFOS, from 2000 to 2010, and increasingly non-racemic 1m-PFOS EFs, from 1996 to 2000, were detected in Swedish samples. No statistically significant trend for %br-PFOS or 1m-PFOS EF was observed in American samples, but American males had significantly higher %br-PFOS and significantly lower 1m-PFOS EF (i.e. more non-racemic) than females, and a similar significant difference was shown in the older age group, relative to the younger age group. These temporal trends in %br-PFOS and 1m-PFOS EF are not easily explained and the results highlight uncertainties about how humans are exposed to PFOS. Keywords: Perfluorooctanesulfonate (PFOS), Perfluorooctanesulfonate precursors (PreFOS), Isomers, Enantiomers, Temporal trends, Human exposure

We present a shoreline-based, millennial-scale record of lake-level changes spanning 12.8–2.3 ka for a large closed-basin lake system on the southwestern Tibetan Plateau. Fifty-three radiocarbon and eight U–Th series ages of tufa and beach cement provide age control on paleoshorelines ringing the basin, supplemented by nineteen ages from shell and aquatic plant material from natural exposures generally recording lake regressions. Our results show that paleo-Ngangla Ring Tso exceeded modern lake level (4727 m asl) continuously between ~ 12.8 and 2.3 ka. The lake was at its highstand 135 m (4862 m asl) above the modern lake from 10.3 ka to 8.6 ka. This is similar to other closed-basin lakes in western Tibet, and coincides with peak Northern Hemisphere summer insolation and peak Indian Summer Monsoon intensity. The lake experienced a series of millennial-scale oscillations centered on 11.5, 10.8, 8.3, 5.9 and 3.6 ka, consistent with weak monsoon events in proxy records of the Indian Summer Monsoon. It is unclear whether these events were forced by North Atlantic or Indian Ocean conditions, but based on the abrupt lake-level regressions recorded for Ngangla Ring Tso, they resulted in significant periodic reductions in rainfall over the western Tibetan Plateau throughout the Holocene.

This paper examines choices of earth-working tools made by Neolithic Chinese populations. In the Hemudu Culture (7000‐ 5000 B.P.), bone (scapula) digging tools were used from the earliest times, whereas peoples in surrounding areas used stone spades. A range of experiments on manufacturing costs, durability, and use efficiency under realistic conditions show that bone and stone spades are functionally equivalent when soils are soft, but that stone implements provide significant and easily perceived advantages when working harder soils. The persistence of scapular spades in the Hemudu Culture would have constrained decisions about undertaking large construction projects under normal soil conditions. Our results show that, in addition to generalized labor for construction, labor demands for producing earth-working implements for largescale prehistoric earthworks could have also been substantial. These findings not only help explain the processes of inten sifying rice-agriculture and sedentary settlements in the Lower Yangzi Basin, but also create a solid foundation for further investigation of how the recruitment of both generalized and specialized laborers, the organization of craft production, and the relevant logistics for large-scale earthworks may have paralleled concentrations of political power in prehistory. Este articulo examina las elecciones de herramientas de excavacion en poblaciones chinas del neolitico. Las herramientas de excavacion hechas de hueso (escapula) fueron utilizadas desde los inicios de la cultura Hemudu (7000‐5000 A.P), a diferencia de los habitantes de las zonas aledanas, quienes utilizaron palas liticas. Una serie de experimentos sobre costos de manufactura, durabilidad y eficiencia bajo condiciones reales demuestran que las palas liticas y de hueso son funcionalmente equivalentes en condiciones de suelo blando, sin embargo las herramientas liticas proveen importantes y evidentes ventajas en condiciones de suelo mas duro. El persistente uso de las palas de escapula en la cultura Hemudu habria limitado decisiones relacionadas

Alcohol (ethanol, EtOH) is one of the most widely abused drugs with profound effects on brain function and behavior. GABAA receptors (GABAARs) are one of the major targets for EtOH in the brain. Temporary plastic changes in GABAARs after withdrawal from a single EtOH exposure occur both in vivo and in vitro, which may be the basis for chronic EtOH addiction, tolerance and withdrawal symptoms. Extrasynaptic δ-GABAAR endocytosis is implicated in EtOH-induced GABAAR plasticity, but the mechanisms by which the relative abundance and localization of specific GABAARs are altered by EtOH exposure and withdrawal remain unclear. In this study, we investigated the mechanisms underlying rapid regulation of extrasynaptic δ-GABAAR by a single EtOH withdrawal in cultured rat hippocampal neurons. Thirty-minutes EtOH (60 mM) exposure increased extrasynaptic tonic current (Itonic) amplitude without affecting synaptic GABAAR function in neurons. In contrast, at 30 min after withdrawal, Itonic amplitude and responsiveness to acute EtOH were both reduced. Similar results occurred in neurons with okadaic acid (OA) or phorbol 12,13-dibutyrate (PDBu) exposure. Protein kinase C (PKC) inhibition prevented the reduction of Itonic amplitude and the tolerance to acute EtOH, as well as the reduction of GABAAR-δ subunit abundance induced by a single EtOH withdrawal. Moreover, EtOH withdrawal selectively increased PKCδ level, whereas PKCδ inhibition specifically rescued the EtOH-induced alterations in GABAAR-δ subunit level and δ-GABAAR function. Together, we provided strong evidence for the important roles of PKCδ in the rapid regulation of extrasynaptic δ-GABAAR induced by a single EtOH withdrawal.

Purpose To refine the Minnesota Grading System (MGS) using definitions from the Age-Related Eye Disease Studies (AREDS) into a nine-step grading scale (MGS-9). Methods A nine-step grading scale descriptive analysis using three key phenotypic features (total drusen area, increased, and decreased pigmentation) of human eyebank eyes that were graded according to definitions from the AREDS criteria in order to harmonize studies of disease progression for research involving human tissue. From 2005 through February 2017, we have analyzed 1159 human eyes, procured from two eyebanks. Each macula was imaged using high-resolution, stereoscopic color fundus photography with both direct- and transillumination. Fundus images were digitally overlaid with a grading template and triangulated for foveal centration. Results We documented and stratified risk for each globe by applying the AREDS nine-step grading scale to the key clinical features from the MGS-9. We found a good distribution within the MGS categories (1-9) with few level eight globes. Eyes were processed within 12.1 ± 6.3, hours from the time of death through imaging, dissection, and freezing or fixation. Applying the MGS-9 to 331 pairs (662 eyes were simultaneously graded), 84% were within one-grading step and 93% within two steps of the fellow eye. We also document reticular pseudodrusen, basal laminar drusen, and pattern dystrophy. Conclusions The MGS nine-step grading scale enables researchers using human tissue to refine the risk assessment of donor tissue. This analysis will harmonize results among researchers when grading human tissue using MGS criteria. Most importantly, the MGS-9 links directly to the known risk for progression from the AREDS.

gamma-Aminobutyric acid (GABA)-mediated inhibitory neurotransmission and the gene products involved were discovered during the mid-twentieth century. Historically, myriad existing nervous system drugs act as positive and negative allosteric modulators of these proteins, making GABA a major component of modern neuropharmacology, and suggesting that many potential drugs will be found that share these targets. Although some of these drugs act on proteins involved in synthesis, degradation, and membrane trasnsport of GABA, the GABA receptors Type A (GABAAR) and Type B (GABABR) are the targets of the great majority of GABAergic drugs. This discovery is due in no small part to Professor Norman Bowery. Whereas the topic of GABABR is appropriately emphasized in this special issue, Norman Bowery also made many insights into GABAAR pharmacology, the topic of this article. GABAAR are members of the ligand-gated ion channel receptor superfamily, a chloride channel family of a dozen or more heteropentameric subtypes containing 19 possible different subunits. These subtypes show different brain regional and subcellular localization, age-dependent expression, and potential for plastic changes with experience including drug exposure. Not only are GABAAR the targets of agonist depressants and antagonist convulsants, but most GABAAR drugs act at other (allosteric) binding sites on the GABAAR proteins. Some anxiolytic and sedative drugs, like benzodiazepine and related drugs, act on GABAAR subtype-dependent extracellular domain sites. General anesthetics including alcohols and neurosteroids act at GABAAR subunit-interface trans-membrane sites. Ethanol at high anesthetic doses acts on GABAAR subtype-dependent trans-membrane domain sites. Ethanol at low intoxicating doses acts at GABAAR subtype-dependent extracellular domain sites. Thus GABAAR subtypes possess pharmacologically specific receptor binding sites for a large group of different chemical classes of clinically important neuropharmacological agents.