bluuebunny/crossref_metadata_2025 huggingface dataset:
Assessing and predicting neuropathic pain after spinal cord injury: a TRACK-SCI study
Authors: Fond Kenneth A. , Torres-Espin Abel , Chou Austin , Fernandez Xuan Duong , Moncivais Sara L. , Huie J. Russell , Hemmerle Debra D. , Keller Anastasia V. , Singh Vineeta , Pascual Lisa U. , DiGiorgio Anthony M. , Burke John F. , Talbott Jason F. , Whetstone William D. , Pan Jonathan Z. , Weinstein Phil R. , Dhall Sanjay S. , Ferguson Adam R. , Bresnahan Jacqueline C. , Beattie Michael S. , Kyritsis Nikos
Published: December 2021

Neuropathic pain is one of the most common secondary complications occurring after spinal cord injury (SCI), and often surpasses motor and sensory deficits in the patient population preferences of the most important aspects to be treated. Despite the better understanding of the molecular and physiological mechanisms of neuropathic pain, reliable treatments are still lacking and exhibit wide variations in efficiency. Previous reports have suggested that the most effective pain management is early treatment. To this end, we utilized the TRACK-SCI prospective clinical research database to assess the neuropathic pain status of all enrolled patients and identify acute care variables that can predict the development of neuropathic pain 6- and 12-months post SCI. 36 out of 61 patients of our study cohort reported neuropathic pain at the chronic stages post SCI. Using multidimensional analytics and logistic regression we discovered that (1) the number of total injuries the patient sustained, (2) the injury severity score (ISS), (3) the lower limb total motor score, and (4) the sensory pin prick total score together predict the development of chronic neuropathic pain after SCI. The balanced accuracy of the corresponding logistic regression model is 74.3%, and repeated 5-fold cross validation showed an AUC of 0.708. Our study suggests a crucial role of polytrauma in chronic pain development after SCI and offers a predictive model using variables routinely collected at every hospital setting.
Interpreting dN/dS under different selective regimes in cancer evolution
Authors: Pérez-Figueroa Andrés , Posada David
Published: December 2021

The standard relationship between the dN/dS statistic and the selection coefficient is contingent upon the computation of the rate of fixation of non-synonymous and synonymous mutations among divergent lineages (substitutions). In cancer genomics, however, dN/dS is typically calculated by including mutations that are still segregating in the cell population. The interpretation of dN/dS within sexual populations has been shown to be problematic. Here we used a simple model of somatic evolution to study the relationship between dN/dS and the selection coefficient in the presence of deleterious, neutral, and beneficial mutations in cancer. We found that dN/dS can be used to distinguish cancer genes under positive or negative selection, but it is not always informative about the magnitude of the selection coefficient. In particular, under the asexual scenario simulated, dN/dS is insensitive to negative selection strength. Furthermore, the relationship between dN/dS and the positive selection coefficient depends on the mutation detection threshold, and, in particular scenarios, it can become non-linear. Our results warn about the necessary caution when interpreting the results drawn from dN/dS estimates in cancer.
Designing anti-predator training to maximize learning and efficacy assessments
Authors: Greggor Alison L. , Masuda Bryce M. , Sabol Anne C. , Swaisgood Ronald R.
Published: December 2021

Despite the growing need to use conservation breeding and translocations in species’ recovery, many attempts to reintroduce animals to the wild fail due to predation post-release. Released animals often lack appropriate behaviours for survival, including anti-predator responses. Anti-predator training—a method for encouraging animals to exhibit wariness and defensive responses to predators—has been used to help address this challenge with varying degrees of success. The efficacy of anti-predator training hinges on animals learning to recognize and respond to predators, but learning is rarely assessed, or interventions miss key experimental controls to document learning. An accurate measure of learning serves as a diagnostic tool for improving training if it otherwise fails to reduce predation. Here we present an experimental framework for designing anti-predator training that incorporates suitable controls to infer predator-specific learning and illustrate their use with the critically endangered Hawaiian crow, ‘alalā (Corvus hawaiiensis). We conducted anti-predator training within a conservation breeding facility to increase anti-predator behaviour towards a natural predator, the Hawaiian hawk, ‘io (Buteo solitaries). In addition to running live-predator training trials, we included two control groups, aimed at determining if responses could otherwise be due to accumulated stress and agitation, or to generalized increases in fear of movement. We found that without these control groups we may have wrongly concluded that predator-specific learning occurred. Additionally, despite generations in human care that can erode anti-predator responses, ‘alalā showed unexpectedly high levels of predatory wariness during baseline assessments. We discuss the implications of a learning-focused approach to training for managing endangered species that require improved behavioural competence for dealing with predatory threats, and the importance of understanding learning mechanisms in diagnosing behavioural problems.
GBS and a newly developed mRNA-GBS approach to link population genetic and transcriptome analyses reveal pattern differences between sites and treatments in red clover (Trifolium pratenseL.)
Authors: Gemeinholzer B , Rupp O , Becker A , Strickert M. , Müller C-M
Published: December 2021

The important worldwide forage crop red clover (Trifolium pratenseL.) is widely cultivated as cattle feed and for soil improvement. Wild populations and landraces have great natural diversity that could be used to improve cultivated red clover. However, to date, there is still insufficient knowledge about the natural genetic and phenotypic diversity of the species. Here, we developed a low-cost transcriptome analysis (mRNA-GBS) with reduced complexity and compared the results with population genetic (GBS) and previously published mRNA-Seq data, to assess whether analysis of intraspecific variation within and between populations and transcriptome responses is possible simultaneously. The mRNA-GBS approach was successful. SNP analyses from the mRNA-GBS approach revealed comparable patterns to the GBS results, but it was not possible to link transcriptome analyses with reduced complexity and sequencing depth to previously published greenhouse and field expression studies. The use of short sequences upstream of the poly(A) tail of mRNA to reduce complexity are promising approaches that combine population genetics and expression profiling to analyze many individuals with trait differences simultaneously and cost-effectively, even in non-model species. Our mRNA-GBS approach revealed too many additional short mRNA sequences, hampering sequence alignment depth and SNP recovery. Optimizations are being discussed. Nevertheless, our study design across different regions in Germany was also challenging as the use of differential expression analyses with reduced complexity, in which mRNA is fragmented at specific sites rather than randomly, is most likely counteracted under natural conditions by highly complex plant reactions at low sequencing depth.
Clinical implementation of single-cell RNA sequencing using liver fine needle aspirate tissue sampling and centralized processing captures compartment specific immuno-diversity
Authors: Genshaft Alex S. , Subudhi Sonu , Keo Arlin , Vasquez Juan D. Sanchez , Conceição-Neto Nádia , Mahamed Deeqa , Boeijen Lauke L. , Alatrakchi Nadia , Oetheimer Chris , Vilme Mike , Drake Riley , Fleming Ira , Tran Nancy , Tzouanas Constantine , Joseph-Chazan Jasmin , Villanueva Martin Arreola , van de Werken Harmen J. G. , van Oord Gertine W. , Groothuismink Zwier M.A. , Beudeker Boris J. , Osmani Zgjim , Nkongolo Shirin , Mehrotra Aman , Feld Jordan , Chung Raymond T. , de Knegt Robert J. , Janssen Harry L. A. , Aerssens Jeroen , Bollekens Jacques , Hacohen Nir , Lauer Georg M. , Boonstra Andre , Shalek Alex K. , Gehring Adam
Published: December 2021

Blood samples are frequently collected in human studies of the immune system but poorly represent tissue-resident immunity. Understanding the immunopathogenesis of tissue-restricted diseases, such as chronic hepatitis B, necessitates direct investigation of local immune responses. We developed a workflow that enables frequent, minimally invasive collection of liver fine-needle aspirates in multi-site international studies and centralized single-cell RNA sequencing data generation using the Seq-Well S3 picowell-based technology. All immunological cell types were captured, including liver macrophages, and showed distinct compartmentalization and transcriptional profiles, providing a systematic assessment of the capabilities and limitations of peripheral blood samples when investigating tissue-restricted diseases. The ability to electively sample the liver of chronic viral hepatitis patients and generate high-resolution data will enable multi-site clinical studies to power fundamental and therapeutic discovery.
The influence of sponge-dwelling gobies (Elacatinus horsti) on feeding efficiency of Caribbean sponge hosts, Aplysina lacunosa and Aplysina archeri
Authors: Siemann Megan J. , Turco Aldo
Published: December 2021

Mutualistic associations between benthic marine invertebrates and reef taxa are common. Sponge-dwelling gobies benefit from protection within sponge tubes and greater food availability. Sponge-dwelling gobies are hypothesized to increase sponge pump rates by consuming polychaete parasites, but such increases have not yet been demonstrated. We investigated the association between sponge-dwelling gobies (Elacatinus horsti) and two species of tube sponge (Aplysina lacunosa and Aplysina archeri) in Bonaire, Caribbean Netherlands. We visually assessed goby presence in sponges and used in situ methods with fluorescein dye to estimate feeding rates via pump rates. Aplysina archeri were more likely to host a goby than A. lacunosa. For both sponge species, pump rates of tubes with gobies were higher on average than those of tubes without gobies. Our observations, therefore, suggest that E. horsti associations with Aplysina are likely mutualistic relationships in which sponges benefit from higher feeding rates when gobies are present.
Emotion Dynamics as Hierarchical Bayesian Inference in Time
Authors: Majumdar Gargi , Yazin Fahd , Banerjee Arpan , Roy Dipanjan
Published: December 2021

What fundamental property of our environment would be most valuable and optimal in characterizing the emotional dynamics we experience in our daily life? Empirical work has shown that an accurate estimation of uncertainty is necessary for our optimal perception, learning, and decision-making. However, the role of this uncertainty in governing our affective dynamics remains unexplored. Using Bayesian encoding, decoding and computational modelling, we show that emotional experiences naturally arise due to ongoing uncertainty estimations in a hierarchical neural architecture. This hierarchical organization involves a number of prefrontal sub-regions, with the lateral orbitofrontal cortex having the highest representational complexity of uncertainty. Crucially, this representational complexity, was sensitive to temporal fluctuations in uncertainty and was predictive of participants’ predisposition to anxiety. Furthermore, the temporal dynamics of uncertainty revealed a distinct functional double dissociation within the OFC. Specifically, the medial OFC showed higher connectivity with the DMN, while the lateral OFC with that of the FPN in response to the evolving affect. Finally, we uncovered a temporally predictive code updating individual’s beliefs swiftly in the face of fluctuating uncertainty in the lateral OFC. A biologically relevant and computationally crucial parameter in theories of brain function, we extend uncertainty to be a defining component of complex emotions.
Synthetic evolution of herbicide resistance using a T7 RNAP-based random DNA base editor
Authors: Butt Haroon , Ramirez Jose Luis Moreno , Mahfouz Magdy
Published: December 2021

Synthetic directed evolution via localized sequence diversification and the simultaneous application of selection pressure is a promising method for producing new, beneficial alleles that affect traits of interest in diverse species; however, this technique has rarely been applied in plants. Developing systems to induce localized sequence diversification at high efficiency will expand our ability to evolve traits of interest that improve global food security. In this study, we designed, built, and tested a chimeric fusion of T7 RNA Polymerase (RNAP) and deaminase to enable the localized sequence diversification of a target sequence of interest. We tested our T7 RNAP–DNA base editor in Nicotiana benthamiana transient assays to target a transgene expressing GFP under the control of the T7 promoter. More than 7% of C nucleotides were converted to T in long segments of the GFP sequence. We then targeted the T7 promoter-driven ACETOLACTATE SYNTHASE (ALS) sequence that had been stably integrated in the rice (Oryza sativa) genome and generated C-to-T and G-to-A transitions. We used herbicide treatment as selection pressure for the evolution of the ALS sequence, resulting in the enrichment of herbicide-responsive residues. We then targeted these herbicide-responsive regions in the rice genome using a CRISPR-directed evolution platform and identified herbicide-resistant ALS variants. Thus, our system could be used for the continuous synthetic evolution of gene functions to produce variants with improved herbicide resistance, as well as for other trait engineering applications.
Acceptability of Rectal Artesunate as pre-referral treatment for severe malaria in children under 5 years by health workers and caregivers in the Democratic Republic of the Congo, Nigeria and Uganda
Authors: Awor Phyllis , Kimera Joseph , Athieno Proscovia , Tumukunde Gloria , Okitawutshu Jean , Tshefu Antoinette , Omoluabi Elizabeth , Signorell Aita , Brunner Nina , Kalenga Jean-Claude , Akano Babatunde , Ayodeji Kazeem , Okon Charles , Yusuf Ocheche , Delvento Giulia , Lee Tristan , Burri Christian , Lengeler Christian , Hetzel Manuel W.
Published: December 2021

Background

In children below 6 years with suspected severe malaria who are several hours from facilities providing parenteral treatment, pre-referral rectal artesunate (RAS) is recommended by the World Health Organization to prevent death and disability. A number of African countries are in the process of rolling out quality-assured RAS for pre-referral treatment of severe malaria at community-level. The success of RAS depends, among other factors, on the acceptability of RAS in the communities where it is being rolled-out. Yet to date, there is limited literature on RAS acceptability. This study aimed to determine the acceptability of RAS by health care providers and child caregivers in communities where quality assured RAS was rolled out.

Methods

This study was nested within the comprehensive multi-country observational research project Community Access to Rectal Artesunate for Malaria (CARAMAL). The CARAMAL project was implemented in the Democratic Republic of the Congo (DRC), Nigeria, and Uganda between 2018 and 2020. Data from three different sources were analysed to understand RAS acceptability: Interviews with health workers during three healthcare provider surveys, with caregivers of children under 5 years of age during three household surveys, and with caregivers of children who were recently treated with RAS and enrolled in the CARAMAL Patient Surveillance System.

Results

RAS acceptability was high among all interviewed stakeholders in the three countries. After the roll-out of RAS, 97-100% heath care providers in DRC considered RAS medication as very good or good, as well as 98-100% in Nigeria and 93-100% in Uganda. Majority of caregivers whose children had received rectal artesunate for pre-referral management of severe malaria indicated that they would want to get the medication again, if their child had the same illness (99.8% of caregivers in DRC, 100% in Nigeria and 99.9% in Uganda). Further, using data from three household surveys, 67-80% of caregivers whose children had not received RAS considered the medication as useful.

Conclusion

RAS was well accepted by health workers and child caregivers in DRC, Nigeria and Uganda. Acceptability is unlikely to be an obstacle to the large-scale roll-out of RAS in the studied settings.
Longitudinal prediction of motor dysfunction after stroke: a disconnectome study
Authors: Dulyan Lilit , Talozzi Lia , Pacella Valentina , Corbetta Maurizio , Forkel Stephanie J , Thiebaut de Schotten Michel
Published: December 2021

Motricity is the most commonly affected ability after a stroke. While many clinical studies attempt to predict motor symptoms at different chronic time points after a stroke, longitudinal acute-to-chronic studies remain scarce. Taking advantage of recent advances in mapping brain disconnections, we predict motor outcomes in 62 patients assessed longitudinally two weeks, three months, and one year after their stroke. Results indicate that brain disconnection patterns accurately predict motor impairments. However, disconnection patterns leading to impairment differ between the three time points and between left and right motor impairments. These results were cross-validated using resampling techniques. In sum, we demonstrated that while some neuroplasticity mechanisms exist changing the structure-function relationship, disconnection patterns prevail when predicting motor impairment at different time points after stroke.
Stomata Detector: High-throughput automation of stomata counting in a population of African rice (Oryza glaberrima) using transfer learning
Authors: Cowling Sophie B. , Soltani Hamidreza , Mayes Sean , Murchie Erik H.
Published: December 2021

Stomata are dynamic structures that control the gaseous exchange of CO2 from the external to internal environment and water loss through transpiration. The density and morphology of stomata have important consequences in crop productivity and water use efficiency, both are integral considerations when breeding climate change resilient crops. The phenotyping of stomata is a slow manual process and provides a substantial bottleneck when characterising phenotypic and genetic variation for crop improvement. There are currently no open-source methods to automate stomatal counting. We used 380 human annotated micrographs of O. glaberrima and O. sativa at x20 and x40 objectives for testing and training. Training was completed using the transfer learning for deep neural networks method and R-CNN object detection model. At a x40 objective our method was able to accurately detect stomata (n = 540, r = 0.94, p<0.0001), with an overall similarity of 99% between human and automated counting methods. Our method can batch process large files of images. As proof of concept, characterised the stomatal density in a population of 155 O. glaberrima accessions, using 13,100 micrographs. Here, we present developed Stomata Detector; an open source, sophisticated piece of software for the plant science community that can accurately identify stomata in Oryza spp., and potentially other monocot species.
Possible Interference in Protein – Protein interaction as a new approach in microinhibition of respiratory pathogens on nasal– oral epithelium: An early on-screen study with reference toSARS-Cov-2–ACE2 binding interference
Authors: Datta Debatosh , Pant Suyash , Dhaked Devendra Kumar , Arumugam Somasundaram , Velayutham Ravichandiran , Datta Pallab
Published: December 2021

Upper and lower respiratory pathogens – both microbes and viruses –are responsible for very high morbidity, man-hour loss, residual long term clinical conditions and even mortality. In india only, high incidence of annual respiratory infections – both UT and LT – demands prophylactic intervention in addition to all therapeutic interventions available.The issue of respiratory infections is more pronounced now in the backdrop of nearly uncontrolled high incidences of SARS-Cov-2 affection resulting in death and damage of human lives to the extent of hundreds of millions spreading over entire world, with incidence variations from country to country. After the initial unanswered phase of spread of SARS-Cov-2 virus with attendant unseen mortalities, quickest invention of a series of unusual vaccines have stemmed the lethal progress to a very significant extent, although vaccinating each and every human subject – nearly 8 to 9 bn in supremely divided world –economically-- is an unthinkable proposition where economic disparity dictates vaccine availability and implementation.Moreover, being of highly unstable nucleic acid composition, the original virus, by now has a thick set of variants around the globe with variable clinical outcome. Given this complex background of scanty availability and inefficient implementation, there always is a need of a preventive approach which can possibly micro-fix the pathogens, including SARS-2 on nasal epithelium so as to interfere with viral [or any pathogen] entry through specified receptor gate[s] or any other ways. The present formulation is under study -- as a candidate of interference on nasal / oral mucosa for all respiratory pathogens. This brief report describes dry on-screen studies of protein – protein interaction as well as its possible interference by an amino acid Lysine. Phospholipid bilayerresponses in presence of added loads of the same essential amino acid –Lysine – showed unusual and unexplained behavior both in structural integrity as well in spatial orientation.
Dopaminergic challenge dissociates learning from primary versus secondary sources of information
Authors: Rybicki A. J. , Sowden S. L. , Schuster B. A. , Cook J. L
Published: December 2021

Some theories of human cultural evolution posit that humans have social-specific learning mechanisms that are adaptive specialisations moulded by natural selection to cope with the pressures of group living. However, the existence of neurochemical pathways that are specialised for learning from social information and from individual experience is widely debated. Cognitive neuroscientific studies present mixed evidence for social-specific learning mechanisms: some studies find dissociable neural correlates for social and individual learning whereas others find the same brain areas and, dopamine-mediated, computations involved in both. Here we demonstrate that, like individual learning, social learning is modulated by the dopamine D2 receptor antagonist haloperidol when social information is the primary learning source, but not when it comprises a secondary, additional element. Two groups (total N = 43) completed a decision-making task which required primary learning, from own experience, and secondary learning from an additional source. For one group the primary source was social, and secondary was individual; for the other group this was reversed. Haloperidol affected primary learning irrespective of social/individual nature, with no effect on learning from the secondary source. Thus, we illustrate that neurochemical mechanisms underpinning learning can be dissociated along a primary-secondary but not a social-individual axis. These results resolve conflict in the literature and support an expanding field showing that, rather than being specialised for particular inputs, neurochemical pathways in the human brain can process both social and non-social cues and arbitrate between the two depending upon which cue is primarily relevant for the task at hand.
Input-specific control of interneuron numbers in nascent striatal networks
Authors: Sreenivasan Varun , Serafeimidou-Pouliou Eleni , Exposito-Alonso David , Bercsenyi Kinga , Bernard Clémence , Bae Sung-Eun , Oozeer Fazal , Hanusz-Godoy Alicia , Edwards Robert , Marín Oscar
Published: December 2021

The assembly of functional neuronal circuits requires appropriate numbers of distinct classes of neurons, but the mechanisms through which their relative proportions are established remain poorly defined. Investigating the mouse striatum, here we found that the two most prominent subtypes of striatal interneurons, parvalbumin-expressing (PV+) GABAergic and cholinergic (ChAT+) interneurons, undergo extensive programmed cell death between the first and second postnatal weeks. Remarkably, the survival of PV+ and ChAT+ interneurons is regulated by distinct mechanisms mediated by their specific afferent connectivity. While long-range cortical inputs control PV+ interneuron survival, ChAT+ interneuron survival is regulated by local input from the medium spiny neurons. Our results identify input-specific circuit mechanisms that operate during the period of programmed cell death to establish the final number of interneurons in nascent striatal networks.
Anin vitro, in uteroandin silicoframework of oxygen diffusion in intricate vascular networks of the placenta
Authors: Bappoo Nikhilesh , Kelsey Lachlan J. , Tongpob Yutthapong , Feindel Kirk W. , Caddy Harrison , Wyrwoll Caitlin S. , Doyle Barry J.
Published: December 2021

Abstract

The placenta is a temporary and complex organ critical for fetal development through its subtle but convoluted harmonization of endocrine, vascular, haemodynamic and exchange adaptations. Yet, due to experimental, technological and ethical constraints, this unique organ remains poorly understood.In silicotools are emerging as a powerful means to overcome these challenges and have the potential to actualize novel breakthroughs. Here, we present an interdisciplinary framework combiningin vitroexperiments used to develop an elegant and scalablein silicomodel of oxygen diffusion. We then usein uteroimaging of placental perfusion and oxygenation in both control and growth-restricted rodent placentas for validation of ourin silicomodel. Our framework revealed the structure-function relationship in the feto-placental vasculature; oxygen diffusion is impaired in growth-restricted placentas, due to the diminished arborization of growth-restricted feto-placental vasculature and the lack of decelerated flow for adequate oxygen diffusion and exchange. We highlight the mechanisms of impairment in a rat model of growth restriction, underpinned by placental vascular impairment. Our framework reports and validates the prediction of blood flow deceleration impairment in growth restricted placentas with the placenta’s oxygen transfer capability being significantly impaired, both globally and locally.
A new protocol for speed vernalisation of winter cereals
Authors: Cha Jin-Kyung , O’Connor Kathryn , Alahmad Samir , Lee Jong-Hee , Dinglasan Eric , Park Hyeonjin , Lee So-Myeong , Hirsz Dominique , Kwon Soon-Wook , Kwon Youngho , Kim Kyeong-Min , Ko Jong-Min , Hickey Lee T. , Shin Dongjin , Dixon Laura E.
Published: December 2021

There are many challenges facing the development of high-yielding, nutritious crops for future environments. One limiting factor is generation time, which prolongs research and plant breeding timelines. Recent advances in speed breeding protocols have dramatically reduced generation time for many short-day and long-day species by optimising light and temperature conditions during plant growth. However, winter crops with a vernalisation requirement still require up to 6–10 weeks in low-temperature conditions before transition to reproductive development. Here, we tested a suite of environmental conditions and protocols to investigate if vernalisation can be satisfied more efficiently. We identified a vernalisation method consisting of exposing seeds at the soil surface to an extended photoperiod of 22 h day:2 h night at 10°C with transfer to speed breeding conditions that dramatically reduces generation time in both winter wheat (Triticum aestivum) and winter barley (Hordeum vulgare). Implementation of this protocol achieved up to five generations per year for winter wheat or barley, instead of the two typically obtained under standard vernalisation and plant growth conditions. The protocol has great potential to enhance training and to accelerate research, pre-breeding, and breeding outcomes focussed on winter crop improvement.
Interleukin-13 and its receptor are synaptic proteins involved in plasticity and neuroprotection
Authors: Li Shun , olde Heuvel Florian , Rehman Rida , Li Zhenghui , Aousji Oumayma , Froehlich Albrecht , Zhang Wanhong , Conquest Alison , Woelfle Sarah , Schoen Michael , O’Meara Caitlin , Reinhardt Richard Lee , Voehringer David , Kassubek Jan , Ludolph Albert , Huber-Lang Markus , Knöll Bernd , Morganti-Kossmann Maria Cristina , Boeckers Tobias , Roselli Francesco
Published: December 2021

Immune system molecules are expressed by neurons, often for unknown functions. We have identified IL-13 and its receptor IL-13Ra1 as neuronal, synaptic proteins in mouse, rat, and human brains, whose engagement upregulates the phosphorylation of NMDAR and AMPAR subunits and, in turn, increases synaptic activity and CREB-mediated transcription. We demonstrate that increased IL-13 is a hallmark of traumatic brain injury (TBI) in mice as well as in two distinct cohorts of human patients. We also provide evidence that IL-13 upregulation protects neurons from excitotoxic death. We show IL-13 upregulation occurring in several cohorts of human brain samples and in CSF. Thus, IL-13 is a previously unrecognized physiological modulator of synaptic physiology of neuronal origin, with implications for the establishment of synaptic plasticity and the survival of neurons under injury conditions. Furthermore, we suggest that the neuroprotection afforded through the upregulation of IL-13 represents a new entry point for interventions in the pathophysiology of TBI.
Never cared for what they do. High structural stability of Guanine-quadruplexes in presence of strand-break damages
Authors: Miclot Tom , Hognon Cécilia , Bignon Emmanuelle , Terenzi Alessio , Grandemange Stéphanie , Barone Giampaolo , Monari Antonio
Published: December 2021

DNA integrity is an important factor to assure genome stability and, more generally, cells and organisms’ viability. In presence of DNA damage, the normal cell cycle is perturbed while cells activate their repair processes. Although efficient, the repair system is not always able to ensure the complete restoration of gene integrity. In these cases, not only mutations may occur, but the accumulation of lesions can either lead to carcinogenesis or reach a threshold which induces apoptosis and the programmed cell death. Among the different types of DNA lesions, strand breaks produced by ionizing radiations are the most toxic, due to their inherently difficult repair, which may lead to genomic instability. In this article we show, by using classical molecular simulations techniques, that differently from the canonical double-helical B-DNA, guanine-quadruplex (G4) arrangements show a remarkable structural stability, even in presence of two strand breaks. Since G4-DNA are recognized for their regulatory roles in cell senescence and gene expression, also involving oncogene, their stability can be related to an evolutionary cellular response aimed at minimizing the effects of ionizing radiation.
A 3D printable device allowing fast and reproducible longitudinal preparation of mouse intestines
Authors: DeLucia Beckey , Samorezov Sergey , Zangara Megan T , Markley Rachel L , Osborn Lucas J , Schultz Karlee B , McDonald Christine , Claesen Jan
Published: December 2021

Accurate and reproducible analysis of mouse small and large intestinal lumen is key for research involving intestinal pathology in preclinical models. Currently, there is no easily accessible, standardized method that allows researchers of different skill levels to consistently dissect intestines in a time-efficient manner. Here, we describe the design and use of the 3D printed “Mouse Intestinal Slicing Tool” (MIST), which can be used to longitudinally prepare murine intestines for further analysis. We benchmarked the MIST against a commonly used procedure involving scissors to make a longitudinal cut along the intestines. Use of the MIST halved the time per mouse to prepare the intestines and outperformed alternative methods in smoothness of the cutting edge and general reproducibility. By sharing the plans for printing the MIST, we hope to contribute a uniformly applicable method for saving time and increasing consistency in studies of the mouse gastrointestinal tract.
Glucocorticoids Unmask Silent Non-Coding Genetic Risk Variants for Common Diseases
Authors: Le Nguyen Thanh Thanh , Gao Huanyao , Liu Duan , Ye Zhenqing , Lee Jeong-Heon , Shi Geng-xian , Copenhaver Kaleigh , Zhang Lingxin , Wei Lixuan , Yu Jia , Zhang Cheng , Li Hu , Wang Liewei , Ordog Tamas , Weinshilboum Richard M.
Published: December 2021

Understanding the function of non-coding genetic variants represents a formidable challenge for biomedicine. We previously identified genetic variants that influence gene expression only after exposure to a hormone or drug. Using glucocorticoid signaling as a model system, we have now demonstrated, in a genome-wide manner, that exposure to glucocorticoids triggered disease risk variants with previously unclear function to influence the expression of genes involved in autoimmunity, metabolic and mood disorders, osteoporosis and cancer. Integrating a series of pharmacogenomic and pharmacoepigenomic datasets, we identified the cis-regulatory elements and 3-dimensional interactions underlying the ligand-dependent associations between those genetic variants and distant risk genes. These observations increase our understanding of mechanisms of non-coding genetic variant-chemical environment interactions and advance the fine-mapping of disease risk and pharmacogenomic loci.
Thermal Analysis of Protein Stability and Ligand Binding in Complex Media
Authors: Eskew Matthew W. , Benight Albert S.
Published: December 2021

Screening of ligands that can bind to biologic products of in vitro expression systems typically requires some purification of the expressed biologic target. Such purification is often laborious and time consuming and a limiting challenge. What is required, that could represent an enormous advantage, is the ability to screen expressed proteins in the crude lysate stage without purification. For that purpose, we explore here the utility of differential scanning calorimetry (DSC) measurements for detecting the presence of specific proteins and their interactions with ligands in the complex media where they were prepared, i.e. crude lysates. Model systems were designed to mimic analogous conditions comparable to those that might be encountered in actual in vitro expression systems. Results are reported for several examples where DSC measurements distinctly showed differences in the thermal denaturation behaviors of the crude lysate alone, proteins and proteins plus binding ligands added to the crude lysate. Results were obtained for Streptavidin/Biotin binding in E. coli lysate, and binding of Angiotensin Converting Enzyme 2 (ACE2) by captopril or lisinopril in the lysate supernatant derived from cultured Human Kidney cells (HEK293). ACE2 binding by the reactive binding domain (RBC) of SARS-CoV-2 was also examined. Binding of ACE2 by RBC and lisinopril were similar and consistent with the reported ACE2 inhibitory activity of lisinopril.
Dioxin Disrupts Thyroid Hormone and Glucocorticoid Induction of klf9, a Master Regulator of Frog Metamorphosis
Authors: Han David T. , Zhao Weichen , Powell Wade H.
Published: December 2021

Frog metamorphosis, the development of an air-breathing froglet from an aquatic tadpole, is controlled by thyroid hormone (TH) and glucocorticoids (GC). Metamorphosis is susceptible to disruption by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), an aryl hydrocarbon receptor (AHR) agonist. Krüppel-Like Factor 9 (klf9), an immediate early gene in the endocrine-controlled cascade of expression changes governing metamorphosis, can be synergistically induced by both hormones. This process is mediated by an upstream enhancer cluster, the klf9 synergy module (KSM). klf9 is also an AHR target. We measured klf9 mRNA following exposures to triiodothyronine (T3), corticosterone (CORT), and TCDD in the Xenopus laevis cell line XLK-WG. klf9 was induced 6-fold by 50 nM T3, 4-fold by 100 nM CORT, and 3-fold by 175 nM TCDD. Co- treatments of CORT and TCDD or T3 and TCDD induced klf9 7- and 11-fold, respectively, while treatment with all 3 agents induced a 15-fold increase. Transactivation assays examined enhancers from the Xenopus tropicalis klf9 upstream region. KSM-containing segments mediated a strong T3 response and a larger T3/CORT response, while induction by TCDD was mediated by a region ∼1 kb farther upstream containing 5 AHR response elements (AHREs). This region also supported a CORT response in the absence of readily-identifiable glucocorticoid responsive elements, suggesting mediation by protein-protein interactions. A functional AHRE cluster is positionally conserved in the human genome, and klf9 was induced by TCDD and TH in HepG2 cells. These results indicate that AHR binding to upstream AHREs represents an early key event in TCDD’s disruption of endocrine-regulated klf9 expression and metamorphosis.
Lifelong Cytomegalovirus and Early-Life Irradiation Synergistically Potentiate Age-Related Defects in Response to Vaccination and Infection
Authors: Pugh Jason L. , Coplen Christopher P. , Sukhina Alona S. , Uhrlaub Jennifer L. , Padilla-Torres Jose , Hayashi Tomonori , Nikolich-Zugich Janko
Published: December 2021

A popular “DNA-damage theory” of aging posits that unrepaired DNA damage leads to cellular (and organismal) senescence. Indeed, some hallmarks of immune aging are more prevalent in individuals exposed to Whole-Body Irradiation (WBI). To test this hypothesis in a model relevant to human immune aging, we examined separate and joint effects of lifelong latent Murine Cytomegalovirus (MCMV) and early-life WBI (i) over the course of the lifespan; (ii) in response to a West Nile virus (WNV) live attenuated vaccine; and (iii) following lethal WNV challenge subsequent to vaccination. We recently published that a single dose of non-lethal WBI in youth, on its own, was not sufficient to accelerate aging of the murine immune system despite causing widespread DNA damage and repopulation stress in hematopoietic cells. However, 4Gy sub-lethal WBI caused manifest reactivation of MCMV. Following vaccination and challenge with WNV in the old age, MCMV-infected animals experiencing 4Gy, but not lower, dose of sub-lethal WBI in youth had reduced survival. By contrast, old irradiated mice lacking MCMV and MCMV-infected, but not irradiated, mice were both protected to the same high level as the old non-irradiated, uninfected controls. Analysis of the quality and quantity of anti-WNV immunity showed that higher mortality in MCMV-positive WBI mice correlated with increased levels of MCMV-specific immune activation during WNV challenge. Moreover, we demonstrate that infection, including that by WNV, led to MCMV reactivation. Our data suggest that MCMV reactivation may be an important determinant of increased late-life mortality following early-life irradiation and late-life acute infection.
Single-molecule biophysics experiments in silico: Towards a physical model of a replisome
Authors: Maffeo Christopher , Chou Han-Yi , Aksimentiev Aleksei
Published: December 2021

The interpretation of single-molecule experiments is frequently aided by computational modeling of biomolecular dynamics. The growth of computing power and ongoing validation of computational models suggest that it soon may be possible to replace some experiments out-right with computational mimics. Here we offer a blueprint for performing single-molecule studies in silico using a DNA binding protein as a test bed. We demonstrate how atomistic simulations, typically limited to sub-millisecond durations and zeptoliter volumes, can guide development of a coarse-grained model for use in simulations that mimic experimental assays. We show that, after initially correcting excess attraction between the DNA and protein, qualitative consistency between several experiments and their computational equivalents is achieved, while additionally providing a detailed portrait of the underlying mechanics. Finally the model is used to simulate the trombone loop of a replication fork, a large complex of proteins and DNA.
The functional evolution of termite gut microbiota
Authors: Arora Jigyasa , Kinjo Yukihiro , Šobotník Jan , Buček Aleš , Clitheroe Crystal , Stiblik Petr , Roisin Yves , Žifčáková Lucia , Park Yung Chul , Kim Ki Yoon , Sillam-Dussès David , Hervé Vincent , Lo Nathan , Tokuda Gaku , Brune Andreas , Bourguignon Thomas
Published: December 2021

Termites primarily feed on lignocellulose or soil in association with specific gut microbes. The functioning of the termite gut microbiota is partly understood in a handful of wood-feeding pest species, but remains largely unknown in other taxa. We intend to feel this gap and provide a global understanding of the functional evolution of termite gut microbiota. We sequenced the gut metagenomes of 145 samples representative of the termite diversity. We show that the prokaryotic fraction of the gut microbiota of all termites possesses similar genes for carbohydrate and nitrogen metabolisms, in proportions varying with termite phylogenetic position and diet. The presence of a conserved set of gut prokaryotic genes implies that key nutritional functions were present in the ancestor of modern termites. Furthermore, the abundance of these genes largely correlated with the host phylogeny. Finally, we found that the adaptation to a diet of soil by some termite lineages was accompanied by a change in the stoichiometry of genes involved in important nutritional functions rather than by the acquisition of new genes and pathways. Our results reveal that the composition and function of termite gut prokaryotic communities have been remarkably conserved since termites first appeared ∼150 million years ago. Therefore, the “world smallest bioreactor” has been operating as a multipartite symbiosis composed of termites, archaea, bacteria, and cellulolytic flagellates since its inception.
Global inflammatory response in in vitro organ cultured testes using single-cell RNA-sequencing
Authors: Suzuki Takahiro , Abe Takeru , Ikegaya Mika , Suzuki Kaori , Yabukami Haruka , Sato Takuya , Komeya Mitsuru , Ogawa Takehiko
Published: December 2021

In vitro functional sperm production is important for understanding spermatogenesis and for the treatment of male infertility. Here, we describe similarities and differences between testis tissues in vivo and in vitro and clarify abnormalities in the early stage of in vitro spermatogenesis at single-cell resolution. While in vitro spermatogenesis progressed similarly to in vivo spermatogenesis until the early pachytene spermatocyte stage, a noticeable acute inflammatory response occurred in immune cells and non-immune testicular somatic cells immediately after cultivation. Inhibitor treatment revealed that NLRP3 inflammasome signaling is key to the inflammation. We observed damaged/dead germ cell accumulation in cultured testis, which may be due to dysfunctional phagocytosis by Sertoli cells. Our data revealed abnormal testicular milieu of in vitro cultured testes caused by tissue-wide sterile inflammation, in which the danger-associated molecular pattern-NLRP3 inflammasome axis may be a key element.
MgrB dependent colistin resistance in Klebsiella pneumoniae is associated with an increase in host-to-host transmission
Authors: Bray Andrew S. , Smith Richard D. , Hudson Andrew W. , Hernandez Giovanna E. , Young Taylor M. , Ernst Robert K. , Zafar M. Ammar
Published: December 2021

Due to its high transmissibility, Klebsiella pneumoniae (Kpn) is one of the leading causes of nosocomial infections. Here, we studied the biological cost of colistin resistance, an antibiotic of last resort, of this opportunistic pathogen using a murine model of gut colonization and transmission. Colistin resistance in Kpn is commonly the result of inactivation of the small regulatory protein MgrB. Without a functional MgrB, the two-component system PhoPQ is constitutively active, leading to increased lipid A modifications and subsequent colistin resistance. Using an engineered MgrB mutant, we observed that MgrB-dependent colistin resistance is not associated with a fitness defect during in vitro growth conditions. However, colistin-resistant Kpn colonizes the murine gut poorly, which may be due to the decreased production of capsular polysaccharide by the mutant. The colistin-resistant mutant of Kpn had increased survival outside the host when compared to the parental colistin-sensitive strain. We attribute this enhanced survivability to dysregulation of the PhoPQ two-component system and accumulation of the master stress regulator RpoS. The enhanced survival of the colistin resistant strain may be a key factor in the observed rapid host-to-host transmission in our model. Together, our data demonstrate that colistin-resistant Kpn experiences a biological cost in gastrointestinal colonization. However, this cost is mitigated by enhanced survival outside the host, increasing the risk of transmission. Additionally, it underscores the importance of considering the entire life cycle of a pathogen to truly determine the biological cost associated with antibiotic resistance.

Importance

The biological cost associated with colistin resistance in Klebsiella pneumoniae (Kpn) was examined using a murine model of Kpn gut colonization and fecal-oral transmission. A common mutation resulting in colistin resistance in Kpn is a loss-of-function mutation of the small regulatory protein MgrB that regulates the two-component system PhoPQ. Even though colistin resistance in Kpn comes with a fitness defect in gut colonization, it increases bacterial survival outside the host enabling it to more effectively transmit to a new host. The enhanced survival is dependent upon the accumulation of RpoS and dysregulation of the PhoPQ. Hence, our study expands our understanding of the underlying molecular mechanism contributing to the transmission of colistin-resistant Kpn.
CTP synthase does not form cytoophidia in Drosophila interfollicular stalks
Authors: Wu Zheng , Liu Ji-Long
Published: December 2021

CTP synthase (CTPS) catalyzes the final step of de novo synthesis of the nucleotide CTP. In 2010, CTPS has been found to form filamentous structures termed cytoophidia in Drosophila follicle cells and germline cells. Subsequently, cytoophidia have been reported in many species across three domains of life: bacteria, eukaryotes and archaea. Forming cytoophidia appears to be a highly conserved and ancient property of CTPS. To our surprise, here we find that polar cells and stalk cells, two specialized types of cells composing Drosophila interfollicular stalks, do not possess obvious cytoophidia. Moreover, we show that Myc level is low in these two types of cells, supporting the idea that Myc regulates cytoophidium assembly. Treatment with a glutamine analog, 6-diazo-5-oxo-l-norleucine (DON), increases cytoophidium assembly in main follicle cells, but not in polar cells or stalk cells. Our findings provide an interesting paradigm for the in vivo study of cytoophidium assembly and disassembly among different populations of follicle cells.