Wild Mouse Gut Microbiota Promotes Host Fitness and Improves Disease Resistance.

Laboratory mice, while paramount for understanding basic biological phenomena, are limited in modeling complex diseases of humans and other free-living mammals. Because the microbiome is a major factor in mammalian physiology, we aimed to identify a naturally evolved reference microbiome to better recapitulate physiological phenomena relevant in the natural world outside the laboratory. Among 21 distinct mouse populations worldwide, we identified a closely related wild relative to standard laboratory mouse strains. Its bacterial gut microbiome differed significantly from its laboratory mouse counterpart and was transferred to and maintained in laboratory mice over several generations. Laboratory mice reconstituted with natural microbiota exhibited reduced inflammation and increased survival following influenza virus infection and improved resistance against mutagen/inflammation-induced colorectal tumorigenesis. By demonstrating the host fitness-promoting traits of natural microbiota, our findings should enable the discovery of protective mechanisms relevant in the natural world and improve the modeling of complex diseases of free-living mammals. VIDEO ABSTRACT.

Targeting myostatin/activin A protects against skeletal muscle and bone loss during spaceflight.

Among the physiological consequences of extended spaceflight are loss of skeletal muscle and bone mass. One signaling pathway that plays an important role in maintaining muscle and bone homeostasis is that regulated by the secreted signaling proteins, myostatin (MSTN) and activin A. Here, we used both genetic and pharmacological approaches to investigate the effect of targeting MSTN/activin A signaling in mice that were sent to the International Space Station. Wild type mice lost significant muscle and bone mass during the 33 d spent in microgravity. Muscle weights of Mstn-/- mice, which are about twice those of wild type mice, were largely maintained during spaceflight. Systemic inhibition of MSTN/activin A signaling using a soluble form of the activin type IIB receptor (ACVR2B), which can bind each of these ligands, led to dramatic increases in both muscle and bone mass, with effects being comparable in ground and flight mice. Exposure to microgravity and treatment with the soluble receptor each led to alterations in numerous signaling pathways, which were reflected in changes in levels of key signaling components in the blood as well as their RNA expression levels in muscle and bone. These findings have implications for therapeutic strategies to combat the concomitant muscle and bone loss occurring in people afflicted with disuse atrophy on Earth as well as in astronauts in space, especially during prolonged missions.

De Novo Mutation Rate Variation and Its Determinants in Chlamydomonas.

De novo mutations are central for evolution, since they provide the raw material for natural selection by regenerating genetic variation. However, studying de novo mutations is challenging and is generally restricted to model species, so we have a limited understanding of the evolution of the mutation rate and spectrum between closely related species. Here, we present a mutation accumulation (MA) experiment to study de novo mutation in the unicellular green alga Chlamydomonas incerta and perform comparative analyses with its closest known relative, Chlamydomonas reinhardtii. Using whole-genome sequencing data, we estimate that the median single nucleotide mutation (SNM) rate in C. incerta is µ = 7.6 × 10-10, and is highly variable between MA lines, ranging from µ = 0.35 × 10-10 to µ = 131.7 × 10-10. The SNM rate is strongly positively correlated with the mutation rate for insertions and deletions between lines (r > 0.97). We infer that the genomic factors associated with variation in the mutation rate are similar to those in C. reinhardtii, allowing for cross-prediction between species. Among these genomic factors, sequence context and complexity are more important than GC content. With the exception of a remarkably high C→T bias, the SNM spectrum differs markedly between the two Chlamydomonas species. Our results suggest that similar genomic and biological characteristics may result in a similar mutation rate in the two species, whereas the SNM spectrum has more freedom to diverge.

Investigating cell autonomy in microorganisms.

Cell-cell signaling in microorganisms is still poorly characterized. In this Methods paper, we describe a genetic procedure for detecting cell-nonautonomous genetic effects, and in particular cell-cell signaling, termed the chimeric colony assay (CCA). The CCA measures the effect of a gene on a biological response in a neighboring cell. This assay can measure cell autonomy for range of biological activities including transcript or protein accumulation, subcellular localization, and cell differentiation. To date, the CCA has been used exclusively to investigate colony patterning in the budding yeast Saccharomyces cerevisiae. To demonstrate the wider potential of the assay, we applied this assay to two other systems: the effect of Grr1 on glucose repression of GAL1 transcription in yeast and the effect of rpsL on stop-codon translational readthrough in Escherichia coli. We also describe variations of the standard CCA that address specific aspects of cell-cell signaling, and we delineate essential controls for this assay. Finally, we discuss complementary approaches to the CCA. Taken together, this Methods paper demonstrates how genetic assays can reveal and explore the roles of cell-cell signaling in microbial processes.

Observations of phase changes in monoolein during high viscous injection.

Serial crystallography of membrane proteins often employs high-viscosity injectors (HVIs) to deliver micrometre-sized crystals to the X-ray beam. Typically, the carrier medium is a lipidic cubic phase (LCP) media, which can also be used to nucleate and grow the crystals. However, despite the fact that the LCP is widely used with HVIs, the potential impact of the injection process on the LCP structure has not been reported and hence is not yet well understood. The self-assembled structure of the LCP can be affected by pressure, dehydration and temperature changes, all of which occur during continuous flow injection. These changes to the LCP structure may in turn impact the results of X-ray diffraction measurements from membrane protein crystals. To investigate the influence of HVIs on the structure of the LCP we conducted a study of the phase changes in monoolein/water and monoolein/buffer mixtures during continuous flow injection, at both atmospheric pressure and under vacuum. The reservoir pressure in the HVI was tracked to determine if there is any correlation with the phase behaviour of the LCP. The results indicated that, even though the reservoir pressure underwent (at times) significant variation, this did not appear to correlate with observed phase changes in the sample stream or correspond to shifts in the LCP lattice parameter. During vacuum injection, there was a three-way coexistence of the gyroid cubic phase, diamond cubic phase and lamellar phase. During injection at atmospheric pressure, the coexistence of a cubic phase and lamellar phase in the monoolein/water mixtures was also observed. The degree to which the lamellar phase is formed was found to be strongly dependent on the co-flowing gas conditions used to stabilize the LCP stream. A combination of laboratory-based optical polarization microscopy and simulation studies was used to investigate these observations.

Population structure and inbreeding in wild house mice (Mus musculus) at different geographic scales.

House mice (Mus musculus) have spread globally as a result of their commensal relationship with humans. In the form of laboratory strains, both inbred and outbred, they are also among the most widely used model organisms in biomedical research. Although the general outlines of house mouse dispersal and population structure are well known, details have been obscured by either limited sample size or small numbers of markers. Here we examine ancestry, population structure, and inbreeding using SNP microarray genotypes in a cohort of 814 wild mice spanning five continents and all major subspecies of Mus, with a focus on M. m. domesticus. We find that the major axis of genetic variation in M. m. domesticus is a south-to-north gradient within Europe and the Mediterranean. The dominant ancestry component in North America, Australia, New Zealand, and various small offshore islands are of northern European origin. Next we show that inbreeding is surprisingly pervasive and highly variable, even between nearby populations. By inspecting the length distribution of homozygous segments in individual genomes, we find that inbreeding in commensal populations is mostly due to consanguinity. Our results offer new insight into the natural history of an important model organism for medicine and evolutionary biology.

'I Don't Like Uncertainty, I Like to Know': How and why uveal melanoma patients consent to life expectancy prognostication.

BACKGROUND: Technological advances have led to cancer prognostication that is increasingly accurate but often unalterable. However, a reliable prognosis of limited life expectancy can cause psychological distress. People should carefully consider offers of prognostication, but little is known about how and why they decide on prognostication. Using uveal melanoma (UM) patients, we aimed to identify (i) how and why do people with UM decide to accept prognostication and (ii) alignment and divergence of their decision-making from conceptualizations of a 'well-considered' decision. METHODS: UM provides a paradigm to elucidate clinical and ethical perspectives on prognostication, because prognostication is reliable but prognoses are largely nonameliorable. We used qualitative methods to examine how and why 20 UM people with UM chose prognostication. We compared findings to a template of 'well-considered' decision-making, where 'well-considered' decisions involve consideration of all likely outcomes. RESULTS: Participants wanted prognostication to reduce future worry about uncertain life expectancy. They spontaneously spoke of hoping for a good prognosis when making their decisions, but largely did not consider the 50% possibility of a poor prognosis. When pressed, they argued that a poor outcome at least brings certainty. CONCLUSIONS: While respecting decisions as valid expressions of participants' wishes, we are concerned that they did not explicitly consider the realistic possibility of a poor outcome and how this would affect them. Thus, it is difficult to see their decisions as 'well-considered'. We propose that nondirective preference exploration techniques could help people to consider the possibility of a poor outcome. PATIENT OR PUBLIC CONTRIBUTION: This paper is a direct response to a patient-identified and defined problem that arose in therapeutic and conversational discourse. The research was informed by the responses of patient participants, as we used the material from interviews to dynamically shape the interview guide. Thus, participants' ideas drove the analysis and shaped the interviews to come.

NHS frontline staff experiences of an in-house psychological support service during the COVID-19 pandemic.

The COVID-19 pandemic has led to enormous practical and emotional challenges for healthcare workers globally, including NHS staff. Psychological support provisions have been established by an NHS healthcare trust in the North of England, including 1:1 psychological support provided by the in-house psychology team. This study sought to understand how staff experienced the service, what worked well and what could be improved. Five participants who had accessed the staff support service took part in semi-structured interviews. Data were analysed using Interpretative Phenomenological Analysis (IPA). Three main themes emerged from the data: 'The need for a flexible, responsive approach', 'Individual and group benefits' and 'The future of staff support: "we need to invest in staff mentally"'. Accessing the service was seen as personally and professionally rewarding for the NHS staff interviewed. These findings are discussed in relation to the relevant literature. Clinical implications, methodological limitations and directions for future research are discussed.

Distribution and Temporal Dynamics of Plasmodium falciparum Chloroquine Resistance Transporter Mutations Associated With Piperaquine Resistance in Northern Cambodia.

BACKGROUND: Newly emerged mutations within the Plasmodium falciparum chloroquine resistance transporter (PfCRT) can confer piperaquine resistance in the absence of amplified plasmepsin II (pfpm2). In this study, we estimated the prevalence of co-circulating piperaquine resistance mutations in P. falciparum isolates collected in northern Cambodia from 2009 to 2017. METHODS: The sequence of pfcrt was determined for 410 P. falciparum isolates using PacBio amplicon sequencing or whole genome sequencing. Quantitative polymerase chain reaction was used to estimate pfpm2 and pfmdr1 copy number. RESULTS: Newly emerged PfCRT mutations increased in prevalence after the change to dihydroartemisinin-piperaquine in 2010, with >98% of parasites harboring these mutations by 2017. After 2014, the prevalence of PfCRT F145I declined, being outcompeted by parasites with less resistant, but more fit PfCRT alleles. After the change to artesunate-mefloquine, the prevalence of parasites with amplified pfpm2 decreased, with nearly half of piperaquine-resistant PfCRT mutants having single-copy pfpm2. CONCLUSIONS: The large proportion of PfCRT mutants that lack pfpm2 amplification emphasizes the importance of including PfCRT mutations as part of molecular surveillance for piperaquine resistance in this region. Likewise, it is critical to monitor for amplified pfmdr1 in these PfCRT mutants, as increased mefloquine pressure could lead to mutants resistant to both drugs.

LayNii: A software suite for layer-fMRI.

High-resolution fMRI in the sub-millimeter regime allows researchers to resolve brain activity across cortical layers and columns non-invasively. While these high-resolution data make it possible to address novel questions of directional information flow within and across brain circuits, the corresponding data analyses are challenged by MRI artifacts, including image blurring, image distortions, low SNR, and restricted coverage. These challenges often result in insufficient spatial accuracy of conventional analysis pipelines. Here we introduce a new software suite that is specifically designed for layer-specific functional MRI: LayNii. This toolbox is a collection of command-line executable programs written in C/C++ and is distributed opensource and as pre-compiled binaries for Linux, Windows, and macOS. LayNii is designed for layer-fMRI data that suffer from SNR and coverage constraints and thus cannot be straightforwardly analyzed in alternative software packages. Some of the most popular programs of LayNii contain 'layerification' and columnarization in the native voxel space of functional data as well as many other layer-fMRI specific analysis tasks: layer-specific smoothing, model-based vein mitigation of GE-BOLD data, quality assessment of artifact dominated sub-millimeter fMRI, as well as analyses of VASO data.

Prdm9 Intersubspecific Interactions in Hybrid Male Sterility of House Mouse.

The classical definition posits hybrid sterility as a phenomenon when two parental taxa each of which is fertile produce a hybrid that is sterile. The first hybrid sterility gene in vertebrates, Prdm9, coding for a histone methyltransferase, was identified in crosses between two laboratory mouse strains derived from Mus mus musculus and M. m. domesticus subspecies. The unique function of PRDM9 protein in the initiation of meiotic recombination led to the discovery of the basic molecular mechanism of hybrid sterility in laboratory crosses. However, the role of this protein as a component of reproductive barrier outside the laboratory model remained unclear. Here, we show that the Prdm9 allelic incompatibilities represent the primary cause of reduced fertility in intersubspecific hybrids between M. m. musculus and M. m. domesticus including 16 musculus and domesticus wild-derived strains. Disruption of fertility phenotypes correlated with the rate of failure of synapsis between homologous chromosomes in meiosis I and with early meiotic arrest. All phenotypes were restored to normal when the domesticus Prdm9dom2 allele was substituted with the Prdm9dom2H humanized variant. To conclude, our data show for the first time the male infertility of wild-derived musculus and domesticus subspecies F1 hybrids controlled by Prdm9 as the major hybrid sterility gene. The impairment of fertility surrogates, testes weight and sperm count, correlated with increasing difficulties of meiotic synapsis of homologous chromosomes and with meiotic arrest, which we suppose reflect the increasing asymmetry of PRDM9-dependent DNA double-strand breaks.

A self-supervised deep neural network for image completion resembles early visual cortex fMRI activity patterns for occluded scenes.

The promise of artificial intelligence in understanding biological vision relies on the comparison of computational models with brain data with the goal of capturing functional principles of visual information processing. Convolutional neural networks (CNN) have successfully matched the transformations in hierarchical processing occurring along the brain's feedforward visual pathway, extending into ventral temporal cortex. However, we are still to learn if CNNs can successfully describe feedback processes in early visual cortex. Here, we investigated similarities between human early visual cortex and a CNN with encoder/decoder architecture, trained with self-supervised learning to fill occlusions and reconstruct an unseen image. Using representational similarity analysis (RSA), we compared 3T functional magnetic resonance imaging (fMRI) data from a nonstimulated patch of early visual cortex in human participants viewing partially occluded images, with the different CNN layer activations from the same images. Results show that our self-supervised image-completion network outperforms a classical object-recognition supervised network (VGG16) in terms of similarity to fMRI data. This work provides additional evidence that optimal models of the visual system might come from less feedforward architectures trained with less supervision. We also find that CNN decoder pathway activations are more similar to brain processing compared to encoder activations, suggesting an integration of mid- and low/middle-level features in early visual cortex. Challenging an artificial intelligence model to learn natural image representations via self-supervised learning and comparing them with brain data can help us to constrain our understanding of information processing, such as neuronal predictive coding.

Scene Representations Conveyed by Cortical Feedback to Early Visual Cortex Can Be Described by Line Drawings.

Human behavior is dependent on the ability of neuronal circuits to predict the outside world. Neuronal circuits in early visual areas make these predictions based on internal models that are delivered via non-feedforward connections. Despite our extensive knowledge of the feedforward sensory features that drive cortical neurons, we have a limited grasp on the structure of the brain's internal models. Progress in neuroscience therefore depends on our ability to replicate the models that the brain creates internally. Here we record human fMRI data while presenting partially occluded visual scenes. Visual occlusion allows us to experimentally control sensory input to subregions of visual cortex while internal models continue to influence activity in these regions. Because the observed activity is dependent on internal models, but not on sensory input, we have the opportunity to map visual features conveyed by the brain's internal models. Our results show that activity related to internal models in early visual cortex are more related to scene-specific features than to categorical or depth features. We further demonstrate that behavioral line drawings provide a good description of internal model structure representing scene-specific features. These findings extend our understanding of internal models, showing that line drawings provide a window into our brains' internal models of vision.SIGNIFICANCE STATEMENT We find that fMRI activity patterns corresponding to occluded visual information in early visual cortex fill in scene-specific features. Line drawings of the missing scene information correlate with our recorded activity patterns, and thus to internal models. Despite our extensive knowledge of the sensory features that drive cortical neurons, we have a limited grasp on the structure of our brains' internal models. These results therefore constitute an advance to the field of neuroscience by extending our knowledge about the models that our brains construct to efficiently represent and predict the world. Moreover, they link a behavioral measure to these internal models, which play an active role in many components of human behavior, including visual predictions, action planning, and decision making.

Multi-site harmonization of 7 tesla MRI neuroimaging protocols.

Increasing numbers of 7 T (7 T) magnetic resonance imaging (MRI) scanners are in research and clinical use. 7 T MRI can increase the scanning speed, spatial resolution and contrast-to-noise-ratio of many neuroimaging protocols, but technical challenges in implementation have been addressed in a variety of ways across sites. In order to facilitate multi-centre studies and ensure consistency of findings across sites, it is desirable that 7 T MRI sites implement common high-quality neuroimaging protocols that can accommodate different scanner models and software versions. With the installation of several new 7 T MRI scanners in the United Kingdom, the UK7T Network was established with an aim to create a set of harmonized structural and functional neuroimaging sequences and protocols. The Network currently includes five sites, which use three different scanner platforms, provided by two different vendors. Here we describe the harmonization of functional and anatomical imaging protocols across the three different scanner models, detailing the necessary changes to pulse sequences and reconstruction methods. The harmonized sequences are fully described, along with implementation details. Example datasets acquired from the same subject on all Network scanners are made available. Based on these data, an evaluation of the harmonization is provided. In addition, the implementation and validation of a common system calibration process is described.

Multi-centre, multi-vendor reproducibility of 7T QSM and R2* in the human brain: Results from the UK7T study.

INTRODUCTION: We present the reliability of ultra-high field T2* MRI at 7T, as part of the UK7T Network's "Travelling Heads" study. T2*-weighted MRI images can be processed to produce quantitative susceptibility maps (QSM) and R2* maps. These reflect iron and myelin concentrations, which are altered in many pathophysiological processes. The relaxation parameters of human brain tissue are such that R2* mapping and QSM show particularly strong gains in contrast-to-noise ratio at ultra-high field (7T) vs clinical field strengths (1.5-3T). We aimed to determine the inter-subject and inter-site reproducibility of QSM and R2* mapping at 7T, in readiness for future multi-site clinical studies. METHODS: Ten healthy volunteers were scanned with harmonised single- and multi-echo T2*-weighted gradient echo pulse sequences. Participants were scanned five times at each "home" site and once at each of four other sites. The five sites had 1× Philips, 2× Siemens Magnetom, and 2× Siemens Terra scanners. QSM and R2* maps were computed with the Multi-Scale Dipole Inversion (MSDI) algorithm (https://github.com/fil-physics/Publication-Code). Results were assessed in relevant subcortical and cortical regions of interest (ROIs) defined manually or by the MNI152 standard space. RESULTS AND DISCUSSION: Mean susceptibility (χ) and R2* values agreed broadly with literature values in all ROIs. The inter-site within-subject standard deviation was 0.001-0.005 ppm (χ) and 0.0005-0.001 ms-1 (R2*). For χ this is 2.1-4.8 fold better than 3T reports, and 1.1-3.4 fold better for R2*. The median ICC from within- and cross-site R2* data was 0.98 and 0.91, respectively. Multi-echo QSM had greater variability vs single-echo QSM especially in areas with large B0 inhomogeneity such as the inferior frontal cortex. Across sites, R2* values were more consistent than QSM in subcortical structures due to differences in B0-shimming. On a between-subject level, our measured χ and R2* cross-site variance is comparable to within-site variance in the literature, suggesting that it is reasonable to pool data across sites using our harmonised protocol. CONCLUSION: The harmonized UK7T protocol and pipeline delivers on average a 3-fold improvement in the coefficient of reproducibility for QSM and R2* at 7T compared to previous reports of multi-site reproducibility at 3T. These protocols are ready for use in multi-site clinical studies at 7T.

Falciparum malaria from coastal Tanzania and Zanzibar remains highly connected despite effective control efforts on the archipelago.

BACKGROUND: Tanzania's Zanzibar archipelago has made significant gains in malaria control over the last decade and is a target for malaria elimination. Despite consistent implementation of effective tools since 2002, elimination has not been achieved. Importation of parasites from outside of the archipelago is thought to be an important cause of malaria's persistence, but this paradigm has not been studied using modern genetic tools. METHODS: Whole-genome sequencing (WGS) was used to investigate the impact of importation, employing population genetic analyses of Plasmodium falciparum isolates from both the archipelago and mainland Tanzania. Ancestry, levels of genetic diversity and differentiation, patterns of relatedness, and patterns of selection between these two populations were assessed by leveraging recent advances in deconvolution of genomes from polyclonal malaria infections. RESULTS: Significant decreases in the effective population sizes were inferred in both populations that coincide with a period of decreasing malaria transmission in Tanzania. Identity by descent analysis showed that parasites in the two populations shared long segments of their genomes, on the order of 5 cM, suggesting shared ancestry within the last 10 generations. Even with limited sampling, two of isolates between the mainland and Zanzibar were identified that are related at the expected level of half-siblings, consistent with recent importation. CONCLUSIONS: These findings suggest that importation plays an important role for malaria incidence on Zanzibar and demonstrate the value of genomic approaches for identifying corridors of parasite movement to the island.

Locally Transplanted Adipose Stem Cells Reduce Anastomotic Leaks in Ischemic Colorectal Anastomoses: A Rat Model.

BACKGROUND: Anastomotic leakage remains a dreaded complication after colorectal surgery. Stem-cell-based therapies have been shown to increase angiogenesis and cell proliferation. OBJECTIVE: The purpose of this research was to investigate the use of adipose-derived stem cells on the healing of ischemic colonic anastomoses in a rat model. DESIGN: This is an animal research study using xenotransplantation. SETTINGS: Male Wistar rats (300-400 g, n = 48) were purchased from a licensed breeder. PATIENTS: Adipose stem cells were isolated from the subcutaneous fat of healthy human donors. INTERVENTIONS: The rats underwent laparotomy with creation of an ischemic colorectal anastomosis created by ligation of mesenteric vessels. The animals were divided into 3 groups: control group with an ischemic anastomosis, vehicle-only group in which the ischemic anastomosis was treated with an absorbable gelatin sponge, and a treatment group in which the ischemic anastomosis was treated with an absorbable gelatin sponge plus adipose stem cells. Animals were killed at postoperative days 3 and 7. MAIN OUTCOME MEASURES: Anastomotic leakage was defined as the finding of feculent peritonitis or perianastomotic abscess on necropsy. Rat mRNA expression was measured using real-time polymerase chain reaction. RESULTS: Adipose-derived stem cells significantly decreased anastomotic leakage when compared with control at both postoperative days 3 (25.0% vs 87.5%; p = 0.02) and 7 (25.0% vs 87.5%; p = 0.02). The use of an absorbable gelatin sponge alone had no effect on anastomotic leakage when compared with control and postoperative days 3 or 7. We found that stem cell-treated animals had a 5.9-fold and 7.4-fold increase in the expression of vascular endothelial growth factor when compared with control at 3 and 7 days; however, this difference was not statistically significant when compared with the absorbable gelatin sponge group. LIMITATIONS: This is a preclinical animal research study using xenotransplantation of cultured stem cells. CONCLUSIONS: Locally transplanted adipose stem cells enhance the healing of ischemic colorectal anastomoses and may be a novel strategy for reducing the risk of anastomotic leakage in colorectal surgery. See Video Abstract at http://links.lww.com/DCR/B203. EL TRANSPLANTE LOCAL DE CÉLULAS MADRE ADIPOSAS REDUCE LA FUGA ANASTOMÓTICA EN LAS SUTURAS COLORRECTALES ISQUÉMICAS: MODELO EN RATAS: Las fugas anastomóticas son una complicación pusilánime después de toda cirugía colorrectal. Se ha demostrado que el tratamiento con células madre aumenta la angiogénesis y la proliferación celular.Investigar el uso de células madre derivadas de tejido adiposo en la cicatrización de una anastomosis colónica isquémica basada en ratas como modelo.Estudio de investigación en animales utilizando xenotrasplantes.Adquisición de típicas ratas de laboratorio raza Wistar, todas machos (300-400 g, n = 48) de un criadero autorizado.Aislamiento de células madre de tipo adiposo del tejido celular subcutáneo en donantes humanos sanos.Las ratas se sometieron a laparotomía con la creación de una anastomosis colorrectal isquémica obtenida mediante ligadura controlada de los vasos mesentéricos correspondientes. Los animales se dividieron en tres grupos: grupo de control con anastomosis isquémica, grupo de vehículo único en el que la anastomosis isquémica se trató con una esponja de gelatina absorbible, y un grupo de tratamiento en el que la anastomosis isquémica se trató con una esponja de gelatina absorbible asociada a un vástago adiposo de células madre. Los animales fueron sacrificados el POD3 y el POD7.La fuga anastomótica fué definida como el hallazgo de peritonitis fecaloidea o absceso perianastomótico a la necropsia. La expresión de RNAm de las ratas se midió usando PCR en tiempo real.Las células madre derivadas de tejido adiposo disminuyeron significativamente la fuga anastomótica en comparación con el grupo control tanto en el POD3 (25% frente a 87.5%, p = 0.02) como en el POD7 (25% frente a 87.5%, p = 0.02). El uso de una esponja de gelatina absorbible sola, no tuvo efecto sobre la fuga anastomótica en comparación con los controles el POD3 o el POD7. Descubrimos que los animales tratados con células madre adiposas tenían un aumento de 5,9 y 7,4 veces en la expresión de VEGF en comparación con el control a los 3 y 7 días, respectivamente; sin embargo, esta diferencia no fue estadísticamente significativa en comparación con el grupo de esponja de gelatina absorbible.Este es un estudio preclínico de investigación en animales que utiliza xenotrasplantes de células madre adiposas cultivadas.Las células madre de tipo adiposo trasplantadas localmente mejoran la cicatrisación en casos de anastomosis colorrectales isquémicas, y podrían convertirse en una nueva estrategia para reducir el riesgo de fugas anastomóticas en casos de cirugía colorrectal. Consulte Video Resumen en http://links.lww.com/DCR/B203. (Traducción-Dr Xavier Delgadillo).

X-ray diffractive imaging of controlled gas-phase molecules: Toward imaging of dynamics in the molecular frame.

We report experimental results on the diffractive imaging of three-dimensionally aligned 2,5-diiodothiophene molecules. The molecules were aligned by chirped near-infrared laser pulses, and their structure was probed at a photon energy of 9.5 keV (λ ≈ 130 pm) provided by the Linac Coherent Light Source. Diffracted photons were recorded on the Cornell-SLAC pixel array detector, and a two-dimensional diffraction pattern of the equilibrium structure of 2,5-diiodothiophene was recorded. The retrieved distance between the two iodine atoms agrees with the quantum-chemically calculated molecular structure to be within 5%. The experimental approach allows for the imaging of intrinsic molecular dynamics in the molecular frame, albeit this requires more experimental data, which should be readily available at upcoming high-repetition-rate facilities.

Low-signal limit of X-ray single particle diffractive imaging.

An outstanding question in X-ray single particle imaging experiments has been the feasibility of imaging sub 10-nm-sized biomolecules under realistic experimental conditions where very few photons are expected to be measured in a single snapshot and instrument background may be significant relative to particle scattering. While analyses of simulated data have shown that the determination of an average image should be feasible using Bayesian methods such as the EMC algorithm, this has yet to be demonstrated using experimental data containing realistic non-isotropic instrument background, sample variability and other experimental factors. In this work, we show that the orientation and phase retrieval steps work at photon counts diluted to the signal levels one expects from smaller molecules or with weaker pulses, using data from experimental measurements of 60-nm PR772 viruses. Even when the signal is reduced to a fraction as little as 1/256, the virus electron density determined using ab initio phasing is of almost the same quality as the high-signal data. However, we are still limited by the total number of patterns collected, which may soon be mitigated by the advent of high repetition-rate sources like the European XFEL and LCLS-II.

Multilayer Laue lenses at high X-ray energies: performance and applications.

X-ray microscopy at photon energies above 15 keV is very attractive for the investigation of atomic and nanoscale properties of technologically relevant structural and bio materials. This method is limited by the quality of X-ray optics. Multilayer Laue lenses (MLLs) have the potential to make a major impact in this field because, as compared to other X-ray optics, they become more efficient and effective with increasing photon energy. In this work, MLLs were utilized with hard X-rays at photon energies up to 34.5 keV. The design, fabrication, and performance of these lenses are presented, and their application in several imaging configurations is described. In particular, two "full field" modes of imaging were explored, which provide various contrast modalities that are useful for materials characterisation. These include point projection imaging (or Gabor holography) for phase contrast imaging and direct imaging with both bright-field and dark-field illumination. With high-efficiency MLLs, such modes offer rapid data collection as compared with scanning methods as well as a large field of views.