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DYT1 dystonia is a debilitating neurological movement disorder arising from mutation in the AAA+ ATPase TorsinA. The hallmark of Torsin dysfunction is nuclear envelope blebbing resulting from defects in nuclear pore complex biogenesis. Whether blebs actively contribute to disease manifestation is unknown. We report that FG-nucleoporins in the bleb lumen form aberrant condensates and contribute to DYT1 dystonia by provoking two proteotoxic insults. Short-lived ubiquitylated proteins that are normally rapidly degraded partition into the bleb lumen and become stabilized. In addition, blebs selectively sequester a specific HSP40-HSP70 chaperone network that is modulated by the bleb component MLF2. MLF2 suppresses the ectopic accumulation of FG-nucleoporins and modulates the selective properties and size of condensates in vitro. Our study identifies dual mechanisms of proteotoxicity in the context of condensate formation and establishes FG-nucleoporin-directed activities for a nuclear chaperone network.

This article aims to show the usefulness of the pneumo-computed tomography gastric distention technique in the detection and morphological characterization of subepithelial gastric lesions. We correlate the pneumo-computed tomography and pathology findings in lesions studied at our institution and review the relevant literature.Pneumo-computed tomography, combined with multiplanar reconstructions, three-dimensional reconstructions, and virtual endoscopy, is useful for delineating the morphological details of subepithelial gastric lesions, thanks to the additional gastric distention. This technique better delimits and characterizes the upper and lower margins of the lesions. Pneumo-computed tomography can be considered a useful noninvasive imaging techniques for characterizing these lesions.

Inflammation upon infectious lung injury is a double-edged sword: while tissue-infiltrating immune cells and cytokines are necessary to control infection, these same factors often aggravate injury. Full appreciation of both the sources and targets of inflammatory mediators is required to facilitate strategies to maintain antimicrobial effects while minimizing off-target epithelial and endothelial damage. Recognizing that the vasculature is centrally involved in tissue responses to injury and infection, we observed that pulmonary capillary endothelial cells (ECs) exhibit dramatic transcriptomic changes upon influenza injury punctuated by profound upregulation ofSparcl1. Endothelial deletion and overexpression of SPARCL1 implicated this secreted matricellular protein in driving key pathophysiologic symptoms of pneumonia, which we demonstrate result from its effects on macrophage polarization. SPARCL1 induces a shift to a pro-inflammatory “M1-like” phenotype (CD86+CD206-), thereby increasing associated cytokine levels. Mechanistically, SPARCL1 acts directly on macrophagesin vitroto induce the pro-inflammatory phenotype via activation of TLR4, and TLR4 inhibitionin vivoameliorates inflammatory exacerbations caused by endothelialSparcl1overexpression. Finally, we confirmed significant elevation of SPARCL1 in COVID-19 lung ECs in comparison with those from healthy donors. Survival analysis demonstrated that patients with fatal COVID-19 had higher levels of circulating SPARCL1 protein compared to those who recovered, indicating the potential of SPARCL1 as a biomarker for prognosis of pneumonia and suggesting that personalized medicine approaches might be harnessed to block SPARCL1 and improve outcomes in high-expressing patients.

Antisense oligonucleotides (ASOs) dosed into cerebrospinal fluid (CSF) distribute broadly throughout the central nervous system (CNS). By modulating RNA, they hold the promise of targeting root molecular causes of disease and hold potential to treat myriad CNS disorders. Realization of this potential requires that ASOs must be active in the disease-relevant cells, and ideally, that monitorable biomarkers also reflect ASO activity in these cells. The biodistribution and activity of such centrally delivered ASOs have been deeply characterized in rodent and non-human primate (NHP) models, but usually only in bulk tissue, limiting our understanding of the distribution of ASO activity across individual cells and across diverse CNS cell types. Moreover, in human clinical trials, target engagement is usually monitorable only in a single compartment, CSF. We sought a deeper understanding of how individual cells and cell types contribute to bulk tissue signal in the CNS, and how these are linked to CSF biomarker outcomes. We employed single nucleus transcriptomics on tissue from mice treated with RNase H1 ASOs against Prnp and Malat1 and NHPs treated with an ASO against PRNP. Pharmacologic activity was observed in every cell type, though sometimes with substantial differences in magnitude. Single cell RNA count distributions implied target RNA suppression in every single sequenced cell, rather than intense knockdown in only some cells. Duration of action up to 12 weeks post-dose differed across cell types, being shorter in microglia than in neurons. Suppression in neurons was generally similar to, or more robust than, the bulk tissue. In macaques, PrP in CSF was lowered 40% in conjunction with PRNP knockdown across all cell types including neurons, arguing that a CSF biomarker readout is likely to reflect ASO pharmacodynamic effect in disease-relevant cells in a neuronal disorder. Our results provide a reference dataset for ASO activity distribution in the CNS and establish single nucleus sequencing as a method for evaluating cell type specificity of oligonucleotide therapeutics and other modalities.

Resumen: El propósito de este ensayo es retomar el tema de los Incas en el Collasuyu, sector sudeste del Tahuantinsuyu, mediante aproximaciones al estudio de la capacocha, ceremonia fundacional de una alianza validada por un oráculo que podía ser consultado desde el Cusco. Veremos a través de cuatro casos dos de sus tipos: acuerdo entre el Inca y sus aliados, y acuerdo entre jefes regionales aliados entre sí.La Arqueología mostró la forma de estos acuerdos, y la Etnohistoria aportó microsecuencias de la realización de la ceremonia, rangos de sus participantes, los marcadores de la filiación incaica, etcétera.Abstract: The purpose of this essay is to return to the subject of the Incas in Collasuyu, in the southeast sector of Tahuantinsuyu, through approaches to the study of the capacocha, the founding ceremony of an alliance validated by an oracle that could be consulted from Cusco. We will see through four cases two of their types: agreement between the Inca and his allies, and agreement between regional leaders allied with each other.Archeology showed the form of these agreements, and Ethnohistory provided microsequences of the performance of the ceremony, ranks of its participants, the markers of Inca filiation, etc.Palabras clave: capacocha; Doncellas; Rachaite; Aconcagua; Llullaillaco; Tahuantinsuyu; Collasuyu; Incas; alianzas; atuendos; colores. Keywords: capacocha; Doncellas; Rachaite; Aconcagua; Llullaillaco; Tahuantinsuyu; Collasuyu; Incas; alliances; garments; colours.

Enteric helminths form intimate physical connections with the intestinal epithelium, yet their ability to directly alter epithelial stem cell fate has not been resolved. Here we demonstrate that infection of mice with the parasite Heligmosomoides polygyrus bakeri (Hpb) reprograms the intestinal epithelium into a fetal-like state marked by the emergence of Clusterin-expressing revival stem cells (revSCs). Organoid-based studies using parasite-derived excretory-secretory products reveal that Hpb-mediated revSC generation occurs independently of host-derived immune signals and inhibits type 2 cytokine–driven differentiation of secretory epithelial lineages that promote their expulsion. Reciprocally, type 2 cytokine signals limit revSC differentiation and, consequently, Hpb fitness, indicating that helminths compete with their host for control of the intestinal stem cell compartment to promote continuation of their life cycle.

The lung exhibits a robust, multifaceted regenerative response to severe injuries such as influenza infection, during which quiescent lung-resident epithelial progenitors participate in two distinct reparative pathways: functionally beneficial regeneration via alveolar type 2 (AT2) cell proliferation and differentiation, and dysplastic tissue remodeling via intrapulmonary airway-resident basal p63

SummaryUnderstanding the molecular anatomy and neural connectivity of the brain requires imaging technologies that can map the 3D nanoscale distribution of specific proteins in the context of brain ultrastructure. Light and electron microscopy (EM) enable visualization of either specific labels or anatomical ultrastructure, but combining molecular specificity with anatomical context is challenging. Here, we present pan-Expansion Microscopy of tissue (pan-ExM-t), an all-optical mouse brain imaging method that combines ∼24-fold linear expansion of biological samples with fluorescent pan-staining of protein densities (providing EM-like ultrastructural context), and immunolabeling of protein targets (for molecular imaging). We demonstrate the versatility of this approach by imaging the established synaptic markers Homer1, Bassoon, PSD-95, Synaptophysin, the astrocytic protein GFAP, myelin basic protein (MBP), and anti-GFP antibodies in dissociated neuron cultures and mouse brain tissue sections. pan-ExM-t reveals these markers in the context of ultrastructural features such as pre and postsynaptic densities, 3D nanoarchitecture of neuropil, and the fine structures of cellular organelles. pan-ExM-t is adoptable in any neurobiological laboratory with access to a confocal microscope and has therefore broad applicability in the research community.Highlightspan-ExM-t visualizes proteins in the context of synaptic ultrastructureLipid labeling in pan-ExM-t reveals organellar and cellular membranesAll-optical, easily accessible alternative to correlative light/electron microscopyHigh potential for high throughput connectomics studies

Zika virus (ZIKV) is a mosquito-borne pathogen that recently caused a series of increasingly severe outbreaks. We previously demonstrated that, compared with a pre-epidemic isolate (ZIKVCDN), a Brazilian ZIKV isolate (ZIKVBR) possesses a novel capacity to suppress host immunity, resulting in delayed viral clearance. However, whether ZIKVBR modulates CD4 T cell responses remains unknown. In this study, we show that, in comparison with ZIKVCDN infection, CD4 T cells are less polarized to the Th1 subtype following ZIKVBR challenge in mice. In contrast, we observed an enhanced accumulation of T follicular helper cells 10, 14, and 21 d postinfection with ZIKVBR. This response correlated with an enhanced germinal center B cell response and robust production of higher avidity-neutralizing Abs following ZIKVBR infection. Taken together, our data suggest that contemporary ZIKV strains have evolved to differentially induce CD4 T cell, B cell, and Ab responses and this could provide a model to further define the signals required for T follicular helper cell development.

Introduction: We have previously shown that alterations to the gut microbiota are an important mediator of diet-induced cardiometabolic dysfunction. One potential mechanism by which these microbial alterations may affect cardiometabolic function is via lipopolysaccharide (LPS), a component of gram-negative bacteria that can enter circulation following dysbiosis-related intestinal permeability. Once in circulation, LPS binds to toll-like receptor 4 (TLR4) in various tissues and induces a robust inflammatory response. TLR4 null mice are partially protected from obesity-related metabolic dysfunction, but it’s unclear if this mutation offers protection from vascular dysfunction. Methods: 24 male mice lacking the TLR4 receptor (KO) and 24 C57BL/6 wild type male mice (WT) were fed either a control diet (CON) containing 4% fat or a high fat diet containing 60% fat (HFD) for 6 months, yielding the following four experimental group: WT+CON; WT+HFD; KO+CON; KO+HFD. After 6 months, mice underwent measurement arterial stiffness (via pulse wave velocity), and glucose tolerance (via ip glucose tolerance test). The gut microbiota was analyzed by 16S gene sequencing. Results: Mice fed a high fat diet increased body weight, although the magnitude of weight gain was attenuated in KO+HFD vs WT+HFD (p Conclusions: These data indicate that deletion of TLR4 in male mice partially protects from arterial stiffness. These data support the supposition that LPS signaling downstream of gut dysbiosis may be an important factor in diet-induced vascular dysfunction.

Prion protein (PrP) concentration controls the kinetics of prion replication and is a genetically and pharmacologically validated therapeutic target for prion disease. In order to evaluate PrP concentration as a pharmacodynamic biomarker and assess its contribution to known prion disease risk factors, we developed and validated a plate-based immunoassay reactive for PrP across 6 species of interest and applicable to brain and cerebrospinal fluid (CSF). PrP concentration varied dramatically across different brain regions in mice, cynomolgus macaques, and humans. PrP expression did not appear to contribute to the known risk factors of age, sex, or common PRNP genetic variants. CSF PrP was lowered in the presence of rare pathogenic PRNP variants, with heterozygous carriers of P102L displaying 55%, and D178N just 31%, of the CSF PrP concentration of mutation-negative controls. In rodents, pharmacologic reduction of brain Prnp RNA was reflected in brain parenchyma PrP and, in turn in CSF PrP, validating CSF as a sampling compartment for the effect of PrP-lowering therapy. Our findings support the use of CSF PrP as a pharmacodynamic biomarker for PrP-lowering drugs and suggest that relative reduction from individual baseline CSF PrP concentration may be an appropriate marker for target engagement.

Prion protein (PrP) concentration controls the kinetics of prion replication and is a genetically and pharmacologically validated therapeutic target for prion disease. In order to evaluate PrP concentration as a pharmacodynamic biomarker and assess its contribution to known prion disease risk factors, we developed and validated a plate-based immunoassay reactive for PrP across six species of interest and applicable to brain and cerebrospinal fluid (CSF). PrP concentration varies dramatically between different brain regions in mice, cynomolgus macaques, and humans. PrP expression does not appear to contribute to the known risk factors of age, sex, or common PRNP genetic variants. CSF PrP is lowered in the presence of rare pathogenic PRNP variants, with heterozygous carriers of P102L displaying 55% and of D178N just 31% the CSF PrP concentration of mutation-negative controls. In rodents, pharmacologic reduction of brain Prnp RNA is reflected in brain parenchyma PrP, and in turn in CSF PrP, validating CSF as a sampling compartment for the effect of PrP-lowering therapy. Our findings support the use of CSF PrP as a pharmacodynamic biomarker for PrP-lowering drugs, and suggest that relative reduction from individual baseline CSF PrP concentration may be an appropriate marker for target engagement.

DYT1 dystonia is a debilitating neurological movement disorder arising from mutation in the AAA+ ATPase TorsinA. The hallmark of Torsin dysfunction is nuclear envelope blebbing resulting from defects in nuclear pore complex biogenesis. Whether blebs actively contribute to disease manifestation is unknown. We report that FG-nucleoporins in the bleb lumen form aberrant condensates and contribute to DYT1 dystonia by provoking two proteotoxic insults. Short-lived ubiquitylated proteins that are normally rapidly degraded partition into the bleb lumen and become stabilized. In addition, blebs selectively sequester a specific HSP40-HSP70 chaperone network that is modulated by the bleb component MLF2. MLF2 suppresses the ectopic accumulation of FG-nucleoporins and modulates the selective properties and size of condensates in vitro. Our study identifies dual mechanisms of proteotoxicity in the context of condensate formation and establishes FG-nucleoporin-directed activities for a nuclear chaperone network.

Enteric helminths form intimate physical connections with the intestinal epithelium, yet their ability to directly alter epithelial stem cell fate has not been resolved. Here we demonstrate that infection of mice with the symbiotic parasite Heligmosomoides polygyrus bakeri (Hbp), reprograms the intestinal epithelium into a fetal-like state marked by the emergence of Clusterin-expressing revival stem cells (revSCs). Organoid-based studies using parasite-derived excretory/secretory products reveal that Hpb-mediated revSC generation occurs independent of host-derived immune signals and inhibits type 2 cytokine-driven differentiation of secretory epithelial lineages that promote their expulsion. Reciprocally, type 2 cytokine signals limit revSC differentiation and, consequently, Hpb fitness indicating that helminths compete with their host for control of the intestinal stem cell niche to promote continuation of their life cycle.

Parasitic helminths cause significant damage as they migrate through host tissues to complete their life cycle. While chronic helminth infections are characterized by a well-described Type 2 immune response, the early, tissue-invasive stages are not well understood. Here we investigate the immune pathways activated during the early stages of Heligmosomoides polygyrus bakeri (Hpb), a natural parasitic roundworm of mice. In contrast to the Type 2 immune response present at later stages of infection, a robust Type 1 immune signature including IFNg production was dominant at the time of parasite invasion and granuloma formation. This early response was associated with an accumulation of activated Natural Killer (NK) cells, with no increase of other innate lymphoid cell populations. Parabiosis and confocal microscopy studies indicated that NK cells were recruited from circulation to the small intestine, where they surrounded parasitic larvae. NK cell recruitment required IFNγ receptor signaling, but was independent of CXCR3 expression. The depletion of tissue-infiltrating NK cells altered neither worm burden nor parasite fitness, but increased vascular injury, suggesting a role for NK cells in mediating tissue protection. Together, these data identify an unexpected role for NK cells in promoting disease tolerance during the invasive stage of an enteric helminth infection.

Sea level rise threatens the coastal landscape, including coastal wetlands, which provide a unique natural habitat to a variety of animal and plant species as well as an array of ecosystem service flows of value to people. The economic valuation of potential changes in coastal wetland areas, while challenging, allows for a comparison with other types of economic impacts from climate change and enhances our understanding of the potential benefits of greenhouse gas mitigation. In this study, we estimate an ensemble of future changes in coastal wetland areas considering both sea level rise, future greenhouse gas emissions, and accretion rate uncertainty, using outputs from the National Ocean and Atmospheric (NOAA) marsh migration model. By the end of the century, total wetland losses range from 2.0 to 10.7 million acres across sea level rise scenarios. For Representative Concentration Pathway (RCP) 4.5 and RCP8.5, respectively, cummulative net wetland area loss is 1.8 and 2.4 million acres by 2050 and 3.5 and 5.2 million acres by 2100. We then estimate economic impacts with two distinct approaches: restoration cost and ecosystem services. The ecosystem services considered are limited by what can be reliably quantified-namely, coastal property protection from coastal flooding and carbon sequestration, the latter using a social cost of carbon approach. By the end of the century, annual restoration costs reach $1.5 and $3.1 billion for RCP 4.5 and RCP8.5, respectively. The lost ecosystem services, together, reach annual economic impacts that are much higher, reaching $2.5 billion for RCP4.5 and $6.1 billion for RCP8.5.

The theme of this essay is the bullroarers and the sibilant ropes that accompanied the ceremony of founding the capacocha alliance and oracle to and on the top of the Llullaillaco volcano (6739 masl, province of Salta, Argentina).Crossing data of Inca archaeology, colonial documentation and ethnographic record, I return to the theme of boas as one of the Inca emblems because a garment of the women's dress of the entourage – the tubular tissue to which the bullroarers are attached – represents another ophidian from the same family: the machacuay.Their whistles were reproduced by the men, making the ropes that they carried with them and with which they had already participated in another ceremony in Cusco. The human voice had to add the stories guided by these sounds. With plates, tubular fabric, ropes and stories updated, sound and visually, the History of the means used by the Inca to put under the Cusco government the other non-Cusqueño without confronting war. El tema de este ensayo son las placas zumbadoras y las sogas sibilantes que acompasaron la ceremonia de fundación de la alianza y oráculo capacocha hacia y en la cima del volcán Llullaillaco (6739 msnm, provincia de Salta, República Argentina).Cruzando datos de arqueología incaica, documentación colonial y registro etnográfico, regreso al tema del amaru como uno de los emblemas incaicos porque una prenda del vestuario de las mujeres de la comitiva –el tejido tubular al que están sujetas las placas– representa otro ofidio de la familia de las boas: el machacuay. Sus silbos los reproducían los hombres haciendo girar rápidamente las sogas que llevaban consigo y con las que ya habían participado de otra ceremonia en el Cusco. La voz humana debió agregar los relatos pautados por estos sonidos.Con placas, tejido tubular, sogas y relatos se actualizaban, sonora y visualmente, la Historia de los medios empleados por el Inca para poner bajo el gobierno del Cusco al otro no-cusqueño sin confrontar bélicamente.

This article brings to the Andean studies of convergence of pre-Hispanic and European beliefs the study and analysis of two cases, their continuity and adaptation in space and time. The central theme is the eastern border of the Andes during Tahuantinsuyu, represented by the boas that populated that region. The constant attempts to alleviate the cataclysms, which were believed to have produced with their movements, date back several centuries before Pachacutec re-founded Cusco and the success of the technologies implemented each time (Chavin, Tiwanaku, Inka and its epigones) left their mark on the Pre-Hispanic time line. Este artículo aporta a los estudios andinos de convergencia de creencias prehispánicas y europeas el estudio y análisis de dos casos, su continuidad y adaptación en espacio y tiempo. El tema central es la frontera Este de los Andes durante el Tahuantinsuyu, graficada mediante las boas que poblaban esa región. Los constantes intentos de paliar los cataclismos –que se creía producían con sus movimientos– datan de varios siglos antes que Pachacutec refundase el Cusco y el éxito de las tecnologías implementadas cada vez (Chavín, Tiwanaku, Inka y sus epígonos) dejaron su marca en la línea del tiempo prehispánico.

The development impact fee is one growth management tool that is often adopted to reduce externalities associated with development on the urban fringe. But it is also used as a revenue generator that offsets property taxes. While graduated impact fees are a potential means to reduce sprawling development, it is unclear which public constituencies favor their adoption. Using an adjacent category logit model, there is limited evidence for exclusion based on race or class and, surprisingly, homeownership is not a major determinant of support. The model results indicate differences in policy preferences among longtime Phoenix residents, newcomers, city dwellers, and sub/exurbanites, which may suggest a desire to maintain the status quo and shift the burden of new development to developers and homebuyers. This article contributes to local government literature through an empirical examination of how sociodemographic factors drive public support for graduated development impact fees.

Interaction between the motor and nociceptive systems seems to play an important role in chronic pain. In this pilot study we used a combination of functional near-infrared spectroscopy (FNIRS) and laserevoked potentials (LEPs) during concurrent finger tapping task and noxious laser stimulation in fibromyalgia (FM) patients and controls. The study included 9 healthy subjects and 15 FM patients. During concurrent FNIRS and LEP recording, participants were required either to remain in resting relaxed condition or to execute a finger tapping task with the right hand. In the control group, the left motor cortex showed increased oxyhaemoglobin levels, while the early N1 LEP component was reduced, during the finger tapping task. In FM patients, motor cortex oxyhaemoglobin concentrations were lower during movement, which did not reduce LEPs. The left motor cortex oxyhaemoglobin concentrations had 79.2% diagnostic accuracy. The interplay between motor and pain-related circuits seems to be dysfunctional in FM patients. These results may support a role for motor cortex modulation in the treatment of this disabling disease.