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Infections are common in patients with diabetes. Moreover, increasing incidence of antibiotic resistance impedes the complete bacterial clearance and calls for alternative treatment strategies. Along with antibacterial resistance, compromised host conditions create a favorable condition for the disease progression. In particular, cell junction proteins are of major importance as they contribute to a tight cell barrier, protecting against invading pathogens. However, the impact of high glucose on cell junction proteins has received little attention in the urinary bladder but merits closer investigation. Here, we report that during diabetes the expression of cell junction protein, claudin 14 is compromised in the human urine exfoliated cells and in the urinary bladder of type 2 diabetic mouse. Further in vitro analysis confirmed a direct correlation of lower intracellular calcium levels with claudin 14 expression in high glucose-treated human uroepithelial cells. Moreover, external calcium supplementation in high glucose-treated cells significantly affected the cell migration and restored the claudin 14 expression through focal adhesion and β-1 integrins. Strengthening the epithelial barrier is essential, especially in individuals with diabetes where basal calcium levels could contribute., (© 2024. The Author(s).)

The International Mouse Phenotyping Consortium (IMPC) systematically produces and phenotypes mouse lines with presumptive null mutations to provide insight into gene function. The IMPC now uses the programmable RNA-guided nuclease Cas9 for its increased capacity and flexibility to efficiently generate null alleles in the C57BL/6N strain. In addition to being a valuable novel and accessible research resource, the production of 3313 knockout mouse lines using comparable protocols provides a rich dataset to analyze experimental and biological variables affecting in vivo gene engineering with Cas9. Mouse line production has two critical steps - generation of founders with the desired allele and germline transmission (GLT) of that allele from founders to offspring. A systematic evaluation of the variables impacting success rates identified gene essentiality as the primary factor influencing successful production of null alleles. Collectively, our findings provide best practice recommendations for using Cas9 to generate alleles in mouse essential genes, many of which are orthologs of genes linked to human disease., (© 2024. The Author(s).)

The laboratory mouse has served as the premier animal model system for both basic and preclinical investigations for over a century. However, laboratory mice capture only a subset of the genetic variation found in wild mouse populations, ultimately limiting the potential of classical inbred strains to uncover phenotype-associated variants and pathways. Wild mouse populations are reservoirs of genetic diversity that could facilitate the discovery of new functional and disease-associated alleles, but the scarcity of commercially available, well-characterized wild mouse strains limits their broader adoption in biomedical research. To overcome this barrier, we have recently developed, sequenced, and phenotyped a set of 11 inbred strains derived from wild-caught Mus musculus domesticus. Each of these "Nachman strains" immortalizes a unique wild haplotype sampled from one of five environmentally distinct locations across North and South America. Whole genome sequence analysis reveals that each strain carries between 4.73-6.54 million single nucleotide differences relative to the GRCm39 mouse reference, with 42.5% of variants in the Nachman strain genomes absent from current classical inbred mouse strain panels. We phenotyped the Nachman strains on a customized pipeline to assess the scope of disease-relevant neurobehavioral, biochemical, physiological, metabolic, and morphological trait variation. The Nachman strains exhibit significant inter-strain variation in >90% of 1119 surveyed traits and expand the range of phenotypic diversity captured in classical inbred strain panels. These novel wild-derived inbred mouse strain resources are set to empower new discoveries in both basic and preclinical research., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Dumont et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Substance use disorders are heritable disorders characterized by compulsive drug use, the biological mechanisms for which remain largely unknown. Genetic correlations reveal that predisposing drug-naïve phenotypes, including anxiety, depression, novelty preference and sensation seeking, are predictive of drug-use phenotypes, thereby implicating shared genetic mechanisms. High-throughput behavioral screening in knockout (KO) mice allows efficient discovery of the function of genes. We used this strategy in two rounds of candidate prioritization in which we identified 33 drug-use candidate genes based upon predisposing drug-naïve phenotypes and ultimately validated the perturbation of 22 genes as causal drivers of substance intake. We selected 19/221 KO strains (8.5%) that had a difference from control on at least one drug-naïve predictive behavioral phenotype and determined that 15/19 (~80%) affected the consumption or preference for alcohol, methamphetamine or both. No mutant exhibited a difference in nicotine consumption or preference which was possibly confounded with saccharin. In the second round of prioritization, we employed a multivariate approach to identify outliers and performed validation using methamphetamine two-bottle choice and ethanol drinking-in-the-dark protocols. We identified 15/401 KO strains (3.7%, which included one gene from the first cohort) that differed most from controls for the predisposing phenotypes. 8 of 15 gene deletions (53%) affected intake or preference for alcohol, methamphetamine or both. Using multivariate and bioinformatic analyses, we observed multiple relations between predisposing behaviors and drug intake, revealing many distinct biobehavioral processes underlying these relationships. The set of mouse models identified in this study can be used to characterize these addiction-related processes further., (© 2023 The Authors. Genes, Brain and Behavior published by International Behavioural and Neural Genetics Society and John Wiley & Sons Ltd.)

Behavioral and economic theory dictate that we decide between options based on their values. However, humans and animals eagerly seek information about uncertain future rewards, even when this does not provide any objective value. This implies that decisions are made by endowing information with subjective value and integrating it with the value of extrinsic rewards, but the mechanism is unknown. Here, we show that human and monkey value judgements obey strikingly conserved computational principles during multi-attribute decisions trading off information and extrinsic reward. We then identify a neural substrate in a highly conserved ancient structure, the lateral habenula (LHb). LHb neurons signal subjective value, integrating information's value with extrinsic rewards, and the LHb predicts and causally influences ongoing decisions. Neurons in key input areas to the LHb largely signal components of these computations, not integrated value signals. Thus, our data uncover neural mechanisms of conserved computations underlying decisions to seek information about the future., (© 2024. The Author(s).)

Purpose: To evaluate the safety and efficacy of performing histotripsy through overlying gas-filled bowel in an ex vivo swine model., Methods: An ex vivo model was created to simulate histotripsy treatment of solid organs through gas-filled bowel. Spherical 2.5 cm histotripsy treatments were performed in agar phantoms for each of five treatment groups: 1) control with no overlying bowel ( n  = 6), 2) bowel 0 cm above phantom ( n  = 6), 3) bowel 1 cm above phantom ( n  = 6), 4) bowel 2 cm above phantom ( n  = 6), and 5) bowel 0 cm above the phantom with increased treatment amplitude ( n  = 6). Bowel was inspected for gross and microscopic damage, and treatment zones were measured. A ray-tracing simulation estimated the percentage of therapeutic beam path blockage by bowel in each scenario., Results: All histotripsy treatments through partial blockage were successful (24/24). No visible or microscopic damage was observed to intervening bowel. Partial blockage resulted in a small increase in treatment volume compared to controls ( p  = 0.002 and p  = 0.036 for groups with bowel 0 cm above the phantom, p  > 0.3 for bowel 1 cm and 2 cm above the phantom). Gas-filled bowel was estimated to have blocked 49.6%, 35.0%, and 27.3% of the therapeutic beam at 0, 1, and 2 cm, respectively., Conclusion: Histotripsy has the potential to be applied through partial gas blockage of the therapeutic beam path, as shown by this ex vivo small bowel model. Further work in an in vivo survival model appears indicated.

Background: To avoid the overuse of antibiotics, non-steroidal anti-inflammatory drugs (NSAIDs), acting via cyclooxygenase (COX) inhibition, have been used to reduce pain and as an alternative treatment for uncomplicated urinary tract infections (UTIs). However, clinical studies evaluating NSAIDs versus antibiotics have reported an increased risk of acute pyelonephritis. Therefore, we hypothesized that COX inhibition could compromise the innate immune response and contribute to complications in patients with uncomplicated UTI., Results: We here demonstrate that in particular COX-2 inhibition led to decreased expression of the antimicrobial peptides psoriasin and human β-defensin-2 in human uroepithelial cells. Psoriasin expression was altered in neutrophils and macrophages. COX-2 inhibition also had impact on the inflammasome mediated IL-1β expression in response to uroepithelial E. coli infection. Further, COX-2 inhibition downregulated free radicals and the epithelial barrier protein claudin 1, favoring infectivity. In addition, conditioned media from COX-2 inhibited uroepithelial cells infected with E. coli failed to activate macrophages., Conclusions: Taken together, our data suggests an adverse innate immune effect of COX-2 inhibition on uroepithelial cells during UTI., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Genome editing with CRISPR-associated (Cas) proteins holds exceptional promise for "correcting" variants causing genetic disease. To realize this promise, off-target genomic changes cannot occur during the editing process. Here, we use whole genome sequencing to compare the genomes of 50 Cas9-edited founder mice to 28 untreated control mice to assess the occurrence of S. pyogenes Cas9-induced off-target mutagenesis. Computational analysis of whole-genome sequencing data detects 26 unique sequence variants at 23 predicted off-target sites for 18/163 guides used. While computationally detected variants are identified in 30% (15/50) of Cas9 gene-edited founder animals, only 38% (10/26) of the variants in 8/15 founders validate by Sanger sequencing. In vitro assays for Cas9 off-target activity identify only two unpredicted off-target sites present in genome sequencing data. In total, only 4.9% (8/163) of guides tested have detectable off-target activity, a rate of 0.2 Cas9 off-target mutations per founder analyzed. In comparison, we observe ~1,100 unique variants in each mouse regardless of genome exposure to Cas9 indicating off-target variants comprise a small fraction of genetic heterogeneity in Cas9-edited mice. These findings will inform future design and use of Cas9-edited animal models as well as provide context for evaluating off-target potential in genetically diverse patient populations., (© 2023. The Author(s).)

Reference ranges provide a powerful tool for diagnostic decision-making in clinical medicine and are enormously valuable for understanding normality in pre-clinical scientific research that uses in vivo models. As yet, there are no published reference ranges for electrocardiography (ECG) in the laboratory mouse. The first mouse-specific reference ranges for the assessment of electrical conduction are reported herein generated from an ECG dataset of unprecedented scale. International Mouse Phenotyping Consortium data from over 26,000 conscious or anesthetized C57BL/6N wildtype control mice were stratified by sex and age to develop robust ECG reference ranges. Interesting findings include that heart rate and key elements from the ECG waveform (RR-, PR-, ST-, QT-interval, QT corrected, and QRS complex) demonstrate minimal sexual dimorphism. As expected, anesthesia induces a decrease in heart rate and was shown for both inhalation (isoflurane) and injectable (tribromoethanol) anesthesia. In the absence of pharmacological, environmental, or genetic challenges, we did not observe major age-related ECG changes in C57BL/6N-inbred mice as the differences in the reference ranges of 12-week-old compared to 62-week-old mice were negligible. The generalizability of the C57BL/6N substrain reference ranges was demonstrated by comparison with ECG data from a wide range of non-IMPC studies. The close overlap in data from a wide range of mouse strains suggests that the C57BL/6N-based reference ranges can be used as a robust and comprehensive indicator of normality. We report a unique ECG reference resource of fundamental importance for any experimental study of cardiac function in mice., (© 2023. The Author(s).)

Dermal infections requiring treatment are usually treated with conventional antibiotics, but the rise of bacterial resistance to first-line antibiotics warrants alternative therapeutics. Here, we report that a backbone-cyclized antimicrobial peptide, CD4-PP, designed from the human host defense peptide LL-37, has strong direct antibacterial effects on antibiotic sensitive as well as resistant-type strains and clinical isolates of common skin pathogens in the low (<2) μM range. In addition, it influences innate immunity in keratinocytes, and treatment with CD4-PP is able to clear bacterial infections in infected keratinocytes. Additionally, CD4-PP treatment significantly reduces the wound area in a lawn of keratinocytes infected with MRSA. In conclusion, CD4-PP has the potential to serve as a future drug treating wounds infected with antibiotic-resistant bacteria.

Purpose: To compare electromagnetic navigation (EMN) with computed tomography (CT) fluoroscopy for guiding percutaneous biopsies in the abdomen and pelvis., Materials and Methods: A retrospective matched-cohort design was used to compare biopsies in the abdomen and pelvis performed with EMN (consecutive cases, n = 50; CT-Navigation; Imactis, Saint-Martin-d'Hères, France) with those performed with CT fluoroscopy (n = 100). Cases were matched 1:2 (EMN:CT fluoroscopy) for target organ and lesion size (±10 mm)., Results: The population was well-matched (age, 65 vs 65 years; target size, 2.0 vs 2.1 cm; skin-to-target distance, 11.4 vs 10.7 cm; P > .05, EMN vs CT fluoroscopy, respectively). Technical success (98% vs 100%), diagnostic yield (98% vs 95%), adverse events (2% vs 5%), and procedure time (33 minutes vs 31 minutes) were not statistically different (P > .05). Operator radiation dose was less with EMN than with CT fluoroscopy (0.04 vs 1.2 μGy; P < .001), but patient dose was greater (30.1 vs 9.6 mSv; P < .001) owing to more helical scans during EMN guidance (3.9 vs 2.1; P < .001). CT fluoroscopy was performed with a mean of 29.7 tap scans per case. In 3 (3%) cases, CT fluoroscopy was performed with gantry tilt, and the mean angle out of plane for EMN cases was 13.4°., Conclusions: Percutaneous biopsies guided by EMN and CT fluoroscopy were closely matched for technical success, diagnostic yield, procedure time, and adverse events in a matched cohort of patients. EMN cases were more likely to be performed outside of the gantry plane. Radiation dose to the operator was higher with CT fluoroscopy, and patient radiation dose was higher with EMN. Further study with a wider array of procedures and anatomic locations is warranted., (Copyright © 2023 SIR. Published by Elsevier Inc. All rights reserved.)

High static field MR scanners can produce human tissue images of astounding clarity, but rely on high frequency electromagnetic radiation that generates complicated in-tissue field patterns that are patient-specific and potentially harmful. Many such scanners use parallel transmitters to better control field patterns, but then adjust the transmitters based on general guidelines rather than optimizing for the specific patient, mostly because computing patient-specific fields was presumed far too slow. It was recently demonstrated that the combination of fast low-resolution tissue mapping and fast voxel-based field simulation can be used to perform a patient-specific MR safety check in minutes. However, the field simulation required several of those minutes, making it too slow to perform the dozens of simulations that would be needed for patient-specific optimization. In this paper we describe a compressed-perturbation-matrix technique that nearly eliminates the computational cost of including complex coils (or coils and shields) in voxel-based field simulation of tissue, thereby reducing simulation time from minutes to seconds. The approach is demonstrated on a wide variety of head+coil and head+coil+shield configurations, using the implementation in MARIE 2.0, the latest version of the open-source MR field simulator MARIE.

The aim of this study is to obtain knowledge about which cultivable bacterial species are present in indoor air in homes, and whether the concentration and diversity of airborne bacteria are associated with different factors. Measurements have been performed for one whole year inside different rooms in five homes and once in 52 homes. Within homes, a room-to-room variation for concentrations of airborne bacteria was found, but an overlap in bacterial species was found across rooms. Eleven species were found very commonly and included: Acinetobacter lowffii , Bacillus megaterium, B. pumilus , Kocuria carniphila , K. palustris , K. rhizophila, Micrococcus flavus , M. luteus, Moraxella osloensis and Paracoccus yeei . The concentrations of Gram-negative bacteria in general and the species P. yeei were significantly associated with the season with the highest concentrations in spring. The concentrations of P. yeei , K. rhizophila and B. pumilus were associated positively with relative humidity (RH), and concentrations of K. rhizophila were associated negatively with temperature and air change rate (ACR). Micrococcus flavus concentrations were associated negatively with ACR. Overall, this study identified species which are commonly present in indoor air in homes, and that the concentrations of some species were associated with the factors: season, ACR and RH., Competing Interests: Research ethics statementNot applicable to this article.Consent for publication statementThe authors declare that research participants’ informed consent to publication of findings – including photos, videos and any personal or identifiable information – was secured prior to publication.Conflicts of interest statementThe authors declare no conflicts of interest with this work.The authors declare no conflicts of interest with this work., (© 2023 The Authors.)

In human Alzheimer's disease (AD) patients and AD mouse models, both differential pre-disease brain features and differential disease-associated memory decline are observed, suggesting that certain neurological features may protect against AD-related cognitive decline. The combination of these features is known as brain reserve, and understanding the genetic underpinnings of brain reserve may advance AD treatment in genetically diverse human populations. One potential source of brain reserve is brain microstructure, which is genetically influenced and can be measured with diffusion MRI (dMRI). To investigate variation of dMRI metrics in pre-disease-onset, genetically diverse AD mouse models, we utilized a population of genetically distinct AD mice produced by crossing the 5XFAD transgenic mouse model of AD to 3 inbred strains (C57BL/6J, DBA/2J, FVB/NJ) and two wild-derived strains (CAST/EiJ, WSB/EiJ). At 3 months of age, these mice underwent diffusion magnetic resonance imaging (dMRI) to probe neural microanatomy in 83 regions of interest (ROIs). At 5 months of age, these mice underwent contextual fear conditioning (CFC). Strain had a significant effect on dMRI measures in most ROIs tested, while far fewer effects of sex, sex*strain interactions, or strain*sex*5XFAD genotype interactions were observed. A main effect of 5XFAD genotype was observed in only 1 ROI, suggesting that the 5XFAD transgene does not strongly disrupt neural development or microstructure of mice in early adulthood. Strain also explained the most variance in mouse baseline motor activity and long-term fear memory. Additionally, significant effects of sex and strain*sex interaction were observed on baseline motor activity, and significant strain*sex and sex*5XFAD genotype interactions were observed on long-term memory. We are the first to study the genetic influences of brain microanatomy in genetically diverse AD mice. Thus, we demonstrated that strain is the primary factor influencing brain microstructure in young adult AD mice and that neural development and early adult microstructure are not strongly altered by the 5XFAD transgene. We also demonstrated that strain, sex, and 5XFAD genotype interact to influence memory in genetically diverse adult mice. Our results support the usefulness of the 5XFAD mouse model and convey strong relationships between natural genetic variation, brain microstructure, and memory., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Murdy, Dunn, Singh, Telpoukhovskaia, Zhang, White, Kahn, Febo and Kaczorowski.)

Objective: We developed a hybrid volume surface integral equation (VSIE) method based on domain decomposition to perform fast and accurate magnetic resonance imaging (MRI) simulations that include both remote and local conductive elements., Methods: We separated the conductive surfaces present in MRI setups into two domains and optimized electromagnetic (EM) modeling for each case. Specifically, interactions between the body and EM waves originating from local radiofrequency (RF) coils were modeled with the precorrected fast Fourier transform, whereas the interactions with remote conductive surfaces (RF shield, scanner bore) were modeled with a novel cross tensor train-based algorithm. We compared the hybrid-VSIE with other VSIE methods for realistic MRI simulation setups., Results: The hybrid-VSIE was the only practical method for simulation using 1 mm voxel isotropic resolution (VIR). For 2 mm VIR, our method could be solved at least 23 times faster and required 760 times lower memory than traditional VSIE methods., Conclusion: The hybrid-VSIE demonstrated a marked improvement in terms of convergence times of the numerical EM simulation compared to traditional approaches in multiple realistic MRI scenarios., Significance: The efficiency of the novel hybrid-VSIE method could enable rapid simulations of complex and comprehensive MRI setups.

Careful phenotype analysis of genetically altered mouse embryos/fetuses is vital for deciphering the function of pre- and perinatally lethal genes. Usually this involves comparing the anatomy of mutants with that of wild types of identical developmental stages. Detailed three dimensional information on regular cranial nerve (CN) anatomy of prenatal mice is very scarce. We therefore set out to provide such information to be used as reference data and selected mutants to demonstrate its potential for diagnosing CN abnormalities. Digital volume data of 152 wild type mice, harvested on embryonic day (E)14.5 and of 18 mutants of the Col4a2 , Arid1b , Rpgrip1l and Cc2d2a null lines were examined. The volume data had been created with High Resolution Episcopic Microscopy (HREM) as part of the deciphering the mechanisms of developmental disorders (DMDD) program. Employing volume and surface models, oblique slicing and digital measuring tools, we provide highly detailed anatomic descriptions of the CNs and measurements of the diameter of selected segments. Specifics of the developmental stages of E14.5 mice and anatomic norm variations were acknowledged. Using the provided data as reference enabled us to objectively diagnose CN abnormalities, such as abnormal formation of CN3 ( Col4a2 ), neuroma of the motor portion of CN5 ( Arid1b ), thinning of CN7 ( Rpgrip1l ) and abnormal topology of CN12 ( Cc2d2a ). Although, in a first glimpse perceived as unspectacular, defects of the motor CN5 or CN7, like enlargement or thinning can cause death of newborns, by hindering feeding. Furthermore, abnormal topology of CN12 was recently identified as a highly reliable marker for low penetrating, but potentially lethal defects of the central nervous system., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Reissig, Geyer, Winkler, Preineder, Prin, Wilson, Galli, Tudor, White, Mohun and Weninger.)

Background: The diagnostic rate of Mendelian disorders in sequencing studies continues to increase, along with the pace of novel disease gene discovery. However, variant interpretation in novel genes not currently associated with disease is particularly challenging and strategies combining gene functional evidence with approaches that evaluate the phenotypic similarities between patients and model organisms have proven successful. A full spectrum of intolerance to loss-of-function variation has been previously described, providing evidence that gene essentiality should not be considered as a simple and fixed binary property., Methods: Here we further dissected this spectrum by assessing the embryonic stage at which homozygous loss-of-function results in lethality in mice from the International Mouse Phenotyping Consortium, classifying the set of lethal genes into one of three windows of lethality: early, mid, or late gestation lethal. We studied the correlation between these windows of lethality and various gene features including expression across development, paralogy and constraint metrics together with human disease phenotypes. We explored a gene similarity approach for novel gene discovery and investigated unsolved cases from the 100,000 Genomes Project., Results: We found that genes in the early gestation lethal category have distinct characteristics and are enriched for genes linked with recessive forms of inherited metabolic disease. We identified several genes sharing multiple features with known biallelic forms of inborn errors of the metabolism and found signs of enrichment of biallelic predicted pathogenic variants among early gestation lethal genes in patients recruited under this disease category. We highlight two novel gene candidates with phenotypic overlap between the patients and the mouse knockouts., Conclusions: Information on the developmental period at which embryonic lethality occurs in the knockout mouse may be used for novel disease gene discovery that helps to prioritise variants in unsolved rare disease cases., (© 2022. The Author(s).)

The increasing antibiotic resistance among uropathogenic bacteria warrants alternative therapeutic strategies. We demonstrate the potential of the synthetic peptide CD4-PP, designed by dimerization and backbone cyclization of the shortest antimicrobial region of human cathelicidin, LL-37. CD4-PP is active against clinical and type strains of common uropathogens Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa at concentrations substantially below cellular cytotoxic levels and induced membrane deformation and leakage in E. coli and P. aeruginosa. Furthermore, CD4-PP treatment prevented the formation of new biofilm and dissolved mature biofilm created by E. coli and P. aeruginosa and targeted curli amyloid in E. coli biofilms. In addition, CD4-PP also induced production of LL-37 by uroepithelial cells and increased the expression of tight junction proteins claudin-14 and occludin. During uroepithelial cell infection, CD4-PP significantly reduced uropathogen survival when treatment was given at the start of infection. Low micromolar of CD4-PP treatment initiated after 2 h was successful with all tested species, except P. aeruginosa where CD4-PP was unable to reduce survival, which could be attributed by early biofilm formation. Finally, we demonstrated that urinary catheter pieces coated with saline fluid supplemented with CD4-PP reduced the attachment of E. coli, giving it a potential clinical application., (© 2022. The Author(s).)

Abstract: Identifying the genetic determinants of pain is a scientific imperative given the magnitude of the global health burden that pain causes. Here, we report a genetic screen for nociception, performed under the auspices of the International Mouse Phenotyping Consortium. A biased set of 110 single-gene knockout mouse strains was screened for 1 or more nociception and hypersensitivity assays, including chemical nociception (formalin) and mechanical and thermal nociception (von Frey filaments and Hargreaves tests, respectively), with or without an inflammatory agent (complete Freund's adjuvant). We identified 13 single-gene knockout strains with altered nocifensive behavior in 1 or more assays. All these novel mouse models are openly available to the scientific community to study gene function. Two of the 13 genes (Gria1 and Htr3a) have been previously reported with nociception-related phenotypes in genetically engineered mouse strains and represent useful benchmarking standards. One of the 13 genes (Cnrip1) is known from human studies to play a role in pain modulation and the knockout mouse reported herein can be used to explore this function further. The remaining 10 genes (Abhd13, Alg6, BC048562, Cgnl1, Cp, Mmp16, Oxa1l, Tecpr2, Trim14, and Trim2) reveal novel pathways involved in nociception and may provide new knowledge to better understand genetic mechanisms of inflammatory pain and to serve as models for therapeutic target validation and drug development., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the International Association for the Study of Pain.)

Background: Mice carrying targeted mutations are important for investigating gene function and the role of genes in disease, but off-target mutagenic effects associated with the processes of generating targeted alleles, for instance using Crispr, and culturing embryonic stem cells, offer opportunities for spontaneous mutations to arise. Identifying spontaneous mutations relies on the detection of phenotypes segregating independently of targeted alleles, and having a broad estimate of the level of mutations generated by intensive breeding programmes is difficult given that many phenotypes are easy to miss if not specifically looked for. Here we present data from a large, targeted knockout programme in which mice were analysed through a phenotyping pipeline. Such spontaneous mutations segregating within mutant lines may confound phenotypic analyses, highlighting the importance of record-keeping and maintaining correct pedigrees., Results: Twenty-five lines out of 1311 displayed different deafness phenotypes that did not segregate with the targeted allele. We observed a variety of phenotypes by Auditory Brainstem Response (ABR) and behavioural assessment and isolated eight lines showing early-onset severe progressive hearing loss, later-onset progressive hearing loss, low frequency hearing loss, or complete deafness, with vestibular dysfunction. The causative mutations identified include deletions, insertions, and point mutations, some of which involve new genes not previously associated with deafness while others are new alleles of genes known to underlie hearing loss. Two of the latter show a phenotype much reduced in severity compared to other mutant alleles of the same gene. We investigated the ES cells from which these lines were derived and determined that only one of the 8 mutations could have arisen in the ES cell, and in that case, only after targeting. Instead, most of the non-segregating mutations appear to have occurred during breeding of mutant mice. In one case, the mutation arose within the wildtype colony used for expanding mutant lines., Conclusions: Our data show that spontaneous mutations with observable effects on phenotype are a common side effect of intensive breeding programmes, including those underlying targeted mutation programmes. Such spontaneous mutations segregating within mutant lines may confound phenotypic analyses, highlighting the importance of record-keeping and maintaining correct pedigrees., (© 2022. The Author(s).)

Pigeon breeding is associated with symptoms of the airways. The aim of this study is to illuminate the bacteriological and toxicological characteristics of airborne dust in pigeon coops. Airborne dust was sampled in 31 urban pigeon coops with homing and fancy pigeons, and following the dust was characterized. In total 141 different bacterial species were identified using MALDI-TOF MS, and of these 11 species are classified in risk group 2. Of the cultivable bacteria, Staphylococcus equorum was present in the highest concentration. Microorganisms in the dust were able to form biofilm, and the amount correlated positively with the number of bacteria. Next generation sequencing showed 180 genera with Acinetobacter in highest reads. On average 999 ± 225 ZOTUs were observed per sample with a Shannon-Wiener biodiversity index of 6.17 ± 0.24. Of the identified species the following have previously been suggested as causative agents of extrinsic allergic alveolitis: Alcaligenes faecalis, Bacillus subtilis, Pantoea agglomerans, Sphingobacterium spiritivorum, Thermoactinomyces sp., and Streptomyces albus. Staphylococcus was present on particles with sizes between 1.1 and > 7.0 μm with a geometric mean diameter of particles on 4.7 ± 1.1 μm. Concentrations of airborne endotoxin and dust were elevated compared to references, and the geometric mean concentrations were 102 EU/m 3 and 1.07 mg dust/m 3 , respectively. Upon exposure to the airborne dust human granulocytes produced Reactive Oxidative Species during the first 5 min, and then no further reaction was observed. The concentrations of bacteria in general, Staphylococcus spp., and endotoxin and biodiversity were associated significantly with season, temperature and/or relative humidity, but not with type or density of pigeons. The bacterial composition and biodiversity indices were not affected by type of pigeon. In conclusion, the exposure to bacteria and endotoxin in pigeon houses should not be neglected in the evaluation of causative agents of airways symptoms among pigeon breeders., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Clinical presentation of congenital heart disease is heterogeneous, making identification of the disease-causing genes and their genetic pathways and mechanisms of action challenging. By using in vivo electrocardiography, transthoracic echocardiography and microcomputed tomography imaging to screen 3,894 single-gene-null mouse lines for structural and functional cardiac abnormalities, here we identify 705 lines with cardiac arrhythmia, myocardial hypertrophy and/or ventricular dilation. Among these 705 genes, 486 have not been previously associated with cardiac dysfunction in humans, and some of them represent variants of unknown relevance (VUR). Mice with mutations in Casz1, Dnajc18, Pde4dip, Rnf38 or Tmem161b genes show developmental cardiac structural abnormalities, with their human orthologs being categorized as VUR. Using UK Biobank data, we validate the importance of the DNAJC18 gene for cardiac homeostasis by showing that its loss of function is associated with altered left ventricular systolic function. Our results identify hundreds of previously unappreciated genes with potential function in congenital heart disease and suggest causal function of five VUR in congenital heart disease., (© 2022. The Author(s).)

Purpose: To test an integrated "AC/DC" array approach at 7T, where B 0 inhomogeneity poses an obstacle for functional imaging, diffusion-weighted MRI, MR spectroscopy, and other applications., Methods: A close-fitting 7T 31-channel (31-ch) brain array was constructed and tested using combined Rx and ΔB 0 shim channels driven by a set of rapidly switchable current amplifiers. The coil was compared to a shape-matched 31-ch reference receive-only array for RF safety, signal-to-noise ratio (SNR), and inter-element noise correlation. We characterize the coil array's ability to provide global and dynamic (slice-optimized) shimming using ΔB 0 field maps and echo planar imaging (EPI) acquisitions., Results: The SNR and average noise correlation were similar to the 31-ch reference array. Global and slice-optimized shimming provide 11% and 40% improvements respectively compared to baseline second-order spherical harmonic shimming. Birdcage transmit coil efficiency was similar for the reference and AC/DC array setups., Conclusion: Adding ΔB 0 shim capability to a 31-ch 7T receive array can significantly boost 7T brain B 0 homogeneity without sacrificing the array's rdiofrequency performance, potentially improving ultra-high field neuroimaging applications that are vulnerable to off-resonance effects., (© 2021 International Society for Magnetic Resonance in Medicine.)

Approximately one-third of randomly produced knockout mouse lines produce homozygous offspring, which fail to survive the perinatal period. The majority of these die around or after embryonic day (E)14.5, presumably from cardiovascular insufficiency. For diagnosing structural abnormalities underlying death and diseases and for researching gene function, the phenotype of these individuals has to be analysed. This makes the creation of reference data, which define normal anatomy and normal variations the highest priority. While such data do exist for the heart and arteries, they are still missing for the venous system. Here we provide high-quality descriptive and metric information on the normal anatomy of the venous system of E14.5 embryos. Using high-resolution digital volume data and 3D models from 206 genetically normal embryos, bred on the C57BL/6N background, we present precise descriptive and metric information of the venous system as it presents itself in each of the six developmental stages of E14.5. The resulting data shed new light on the maturation and remodelling of the venous system at transition of embryo to foetal life and provide a reference that can be used for detecting venous abnormalities in mutants. To explore this capacity, we analysed the venous phenotype of embryos from 7 knockout lines (Atp11a, Morc2a, 1700067K01Rik, B9d2, Oaz1, Celf4 and Coro1c). Careful comparisons enabled the diagnosis of not only simple malformations, such as dual inferior vena cava, but also complex and subtle abnormalities, which would have escaped diagnosis in the absence of detailed, stage-specific referenced data., (© 2021 The Authors. Journal of Anatomy published by John Wiley & Sons Ltd on behalf of Anatomical Society.)

In this work, we propose a method for the compression of the coupling matrix in volume-surface integral equation (VSIE) formulations. VSIE methods are used for electromagnetic analysis in magnetic resonance imaging (MRI) applications, for which the coupling matrix models the interactions between the coil and the body. We showed that these effects can be represented as independent interactions between remote elements in 3D tensor formats, and subsequently decomposed with the Tucker model. Our method can work in tandem with the adaptive cross approximation technique to provide fast solutions of VSIE problems. We demonstrated that our compression approaches can enable the use of VSIE matrices of prohibitive memory requirements, by allowing the effective use of modern graphical processing units (GPUs) to accelerate the arising matrix-vector products. This is critical to enable numerical MRI simulations at clinical voxel resolutions in a feasible computation time. In this paper, we demonstrate that the VSIE matrix-vector products needed to calculate the electromagnetic field produced by an MRI coil inside a numerical body model with 1 mm 3 voxel resolution, could be performed in ~ 33 seconds in a GPU, after compressing the associated coupling matrix from ~ 80 TB to ~ 43 MB.

High resolution episcopic microscopy (HREM) produces digital volume data by physically sectioning histologically processed specimens, while capturing images of the subsequently exposed block faces. Our study aims to systematically define the spectrum of typical artefacts inherent to HREM data and to research their effect on the interpretation of the phenotype of wildtype and mutant mouse embryos. A total of 607 (198 wildtypes, 409 mutants) HREM data sets of mouse embryos harvested at embryonic day (E) 14.5 were systematically and comprehensively examined. The specimens had been processed according to essentially identical protocols. Each data set comprised 2000 to 4000 single digital images. Voxel dimensions were 3 × 3 × 3 µm 3 . Using 3D volume models and virtual resections, we identified a number of characteristic artefacts and grouped them according to their most likely causality. Furthermore, we highlight those that affect the interpretation of embryo data and provide examples for artefacts mimicking tissue defects and structural pathologies. Our results aid in optimizing specimen preparation and data generation, are vital for the correct interpretation of HREM data and allow distinguishing tissue defects and pathologies from harmless artificial alterations. In particular, they enable correct diagnosis of pathologies in mouse embryos serving as models for deciphering the mechanisms of developmental disorders.

Urinary tract infection frequently caused by E. coli is one of the most common bacterial infections. Increasing antibiotic resistance jeopardizes successful treatment and alternative treatment strategies are therefore mandatory. Metformin, an oral antidiabetic drug, has been shown to activate macrophages in the protection against certain infecting microorganisms. Since epithelial cells often form the first line of defense, we here investigated the effect on uroepithelial cells during E. coli infection. Metformin upregulated the human antimicrobial peptides cathelicidin LL-37 and RNase7 via modulation of the TRPA1 channel and AMPK pathway. Interestingly, metformin stimulation enriched both LL-37 and TRPA1 in lysosomes. In addition, metformin specifically increased nitric oxide and mitochondrial, but not cytosolic ROS. Moreover, metformin also triggered mRNA expression of the proinflammatory cytokines IL1B, CXCL8 and growth factor GDF15 in human uroepithelial cells. The GDF15 peptide stimulated macrophages increased LL-37 expression, with increased bacterial killing. In conclusion, metformin stimulation strengthened the innate immunity of uroepithelial cells inducing enhanced extracellular and intracellular bacterial killing suggesting a favorable role of metformin in the host defense., (© 2021. The Author(s).)

A Ru(ii) intercalating complex capped with a Mn(i) photoCORM allows for a new mode of DNA intercalator delivery. The steric bulk of the Mn(i) photoCORM inhibits intercalation in the dark, and visible light irradiation (470 nm) dissociates the photoCORM, allowing for DNA intercalation of the Ru(ii) complex.