bluuebunny/crossref_metadata_2025 huggingface dataset:
Computational modeling of macrophage iron sequestration during host defense againstAspergillus
Authors: Adhikari Bandita , Scindia Yogesh , Vieira Luis Sordo , de Assis Lopes Ribeiro Henrique , Masison Joseph , Yang Ning , Fonseca Luis L. , Wheeler Matthew , Knapp Adam C. , Mei Yu , Helba Brian , Atkinson Carl , Schroeder Will , Mehrad Borna , Laubenbacher Reinhard
Published: January 2022

Iron is essential to the virulence ofAspergillusspecies, and restricting iron availability is a critical mechanism of antimicrobial host defense. Macrophages recruited to the site of infection are at the crux of this process, employing multiple intersecting mechanisms to orchestrate iron sequestration from pathogens. To gain an integrated understanding of how this is achieved in invasive aspergillosis, we generated a transcriptomic time-series of the response of human monocyte-derived macrophages toAspergillusand used this and the available literature to construct a mechanistic computational model of iron handling of macrophages during this infection. We found an overwhelming macrophage response beginning 2-4 hours after exposure to the fungus, which included upregulated transcription of iron import proteins transferrin receptor-1, divalent metal transporter-1, and ZIP family transporters, and downregulated transcription of the iron exporter ferroportin. The computational model, based on a discrete dynamical systems framework, consisted of 21 3-state nodes, and was validated with additional experimental data that were not used in model generation. The model accurately captures the steady state and the trajectories of most of the quantitatively measured nodes. In the experimental data, we surprisingly found that transferrin receptor-1 upregulation preceded the induction of inflammatory cytokines, a feature that deviated from model predictions. Model simulations suggested that direct induction of TfR1 after fungal recognition, independent of the Iron Regulatory Protein - Labile Iron Pool system, explains this finding. We anticipate that this model will contribute to a quantitative understanding of iron regulation as a fundamental host defense mechanism during aspergillosis.

Importance

Invasive pulmonary aspergillosis is a major cause of death among immunosuppressed individuals despite the best available therapy. Depriving the pathogen of iron is an essential component of host defense in this infection, but the mechanisms by which the host achieves this are complex. To understand how recruited macrophages mediate iron deprivation during the infection, we developed and validated a mechanistic computational model that integrates the available information in the field. The insights provided by this approach can help in designing iron modulation therapies as anti-fungal treatments.
Foodborne Outbreaks, Product Recalls, and Firm Learning
Authors: Akhundjanov Sherzod B. , Pozo Veronica F. , Thomas Briana
Published: January 2022

Firms in the food industry may experience more than one contamination incident over time. In the context of food safety, increasing the interval between foodborne outbreaks represents a key objective for both the food industry and public health officials. We demonstrate a systematic approach to analyzing repeated recalls, specifically to evaluate factors influencing the time until the next recall and, more importantly, to identify the extent of organizational learning from inter-event times. An analysis of meat and poultry recalls issued by publicly traded firms in the United States between 1994-2015 indicates that more diversified firms face a lower risk of repeat recalls as firm size expands, compared to firms primarily producing meat and poultry products. The hazard of a recall incident decreases with the severity of the previous recall. Some evidence of firm learning is found, but there is no definitive evidence indicating that a firm’s ability to prevent recalls improves with the number of foodborne outbreaks it has experienced.
Brain perivascular space imaging across the human lifespan
Authors: Lynch Kirsten M. , Sepehrband Farshid , Toga Arthur W. , Choupan Jeiran
Published: January 2022

Enlarged perivascular spaces (PVS) are considered a biomarker for vascular pathology and are observed in normal aging and neurological conditions; however, research on the role of PVS in health and disease are hindered by the lack of knowledge regarding the normative time course of PVS alterations with age. To this end, we characterized the influence of age, sex and cognitive performance on PVS anatomical characteristics in a large cross-sectional cohort (∼1400) of healthy subjects between 8 and 90 years of age using multimodal structural MRI data. Our results show age is associated with wider and more numerous MRI-visible PVS over the course of the lifetime with spatially-varying patterns of PVS enlargement trajectories. In particular, regions with low PVS volume fraction in childhood are associated with rapid age-related PVS enlargement (e.g., temporal regions), while regions with high PVS volume fraction in childhood are associated with minimal age-related PVS alterations (e.g., limbic regions). PVS burden was significantly elevated in males compared to females with differing morphological time courses with age. Together, these findings contribute to our understanding of perivascular physiology across the healthy lifespan and provide a normative reference for the spatial distribution of PVS enlargement patterns to which pathological alterations can be compared.
Meta-omics-aided isolation of an elusive anaerobic arsenic-methylating soil bacterium
Authors: Viacava Karen , Qiao Jiangtao , Janowczyk Andrew , Poudel Suresh , Jacquemin Nicolas , Meibom Karin Lederballe , Shrestha Him K. , Reid Matthew C. , Hettich Robert L. , Bernier-Latmani Rizlan
Published: January 2022

Soil microbiomes harbor unparalleled functional and phylogenetic diversity and are sources of novel metabolisms. However, extracting isolates with a targeted function from complex microbiomes is not straightforward, particularly if the associated phenotype does not lend itself to high-throughput screening. Here, we tackle the methylation of arsenic (As) in anoxic soils. By analogy to mercury, As methylation was proposed to be catalyzed by sulfate-reducing bacteria. However, to date, there are no anaerobic isolates capable of As methylation, whether sulfate-reducing or otherwise. The isolation of such a microorganism has been thwarted by the fact that the anaerobic bacteria harboring a functional arsenite S-adenosylmethionine methyltransferase (ArsM) tested to date did not methylate As in pure culture. Additionally, fortuitous As methylation can result from the release of non-specific methyltransferases upon lysis. Thus, we combined metagenomics, metatranscriptomics, and metaproteomics to identify the microorganisms actively methylating As in anoxic soil-derived microbial cultures. Based on the metagenome-assembled genomes of microorganisms expressing ArsM, we isolated Paraclostridium sp. strain EML, which was confirmed to actively methylate As anaerobically. This work is an example of the application of meta-omics to the isolation of elusive microorganisms.
KLF5-regulated extracellular matrix remodeling secures biliary epithelial tissue integrity against cholestatic liver injury
Authors: Yamada Minami , Okada Hajime , Ema Masatsugu , Kikkawa Yamato , Miyajima Atsushi , Itoh Tohru
Published: January 2022

Abstract / Introductory paragraph

Tubular epithelial tissues in the body play fundamental roles as infrastructure constituting conduits to transport various types of biological fluids, for which contiguous and integrated epithelial tissue structures should be maintained continuously and even under stressed conditions. Compared to tissue morphological processes that take place during ontogeny, the mechanisms whereby tubular epithelial tissues maintain their structural integrity in adulthood remains largely unclear. Here, we show that the transcription factor Klf5 is crucial for maintaining the biliary epithelial integrity in tissue remodeling processes induced under cholestatic injury conditions in the adult liver. Loss of Klf5 in the biliary epithelia led to tissue collapse in vivo in injured mouse livers, as well as in vitro in bile ductular organoids in a tissue-autonomous manner and independent of cell proliferation. Klf5 regulated cell junction organization and cell adhesion, along with extracellular matrix remodeling around the expanding biliary epithelia through deposition of Lamb3-containing laminin complexes. Targeting the Lamb3 expression in biliary epithelia in mice recapitulated the tissue collapse phenotype. Together, our results highlight a novel mechanism whereby the epithelial tissue maintains its integrity while undergoing unstable structural transformation.
Impact of secondary TCR engagement on the heterogeneity of pathogen-specific CD8+T cell response during acute and chronic toxoplasmosis
Authors: Shallberg Lindsey A , Phan Anthony T , Christian David A , Perry Joseph A , Haskins Breanne E , Beiting Daniel P , Harris Tajie H , Koshy Anita A , Hunter Christopher A
Published: January 2022

Initial TCR engagement of CD8+T cells results in T cell expansion, and these early events influence the generation of diverse effector and memory populations. During infection, some activated T cells re-encounter cognate antigen, but how these events influence local effector responses or formation of memory populations is unclear. To address this issue, OT-I T cells which express the Nur77-GFP reporter of TCR activation were paired withT. gondiithat express OVA to assess the impact of TCR activation on CD8+T cell responses. During acute infection, TCR stimulation in affected tissues correlated with parasite burden and was associated with markers of effector cells while Nur77-GFP-OT-I showed signs of effector memory potential. However, adoptive transfer of Nur77-GFP negative or positive OT-I from infected mice into naive recipients resulted in formation of similar memory populations. During the chronic stage of infection in the CNS, TCR activation was associated with large scale transcriptional changes and the acquisition of an effector T cell phenotype as well as the generation of a population of CD103+CD69+Trm like cells. However, while inhibition of parasite replication resulted in reduced effector responses it did not alter the Trm population. These data sets highlight the contribution of recent TCR activation on the phenotypic heterogeneity of the CD8+T cell response but suggest that this process has a limited impact on memory populations at acute and chronic stages of infection.

Author Summary

CD8+T cells are important to control many acute and chronic infections, however the role that recent T cell receptor stimulation plays in the formation of ongoing T cell responses is unclear. Here, we utilize a genetic reporter of TCR stimulation and high parameter flow cytometry to characterize TCR-driven phenotypes of pathogen specific T cell responses to the parasiteToxoplasma gondiiduring acute and chronic infection in the periphery and central nervous system. This work demonstrates the importance of recent TCR stimulation in driving local effector CD8+T cell responses in peripheral tissues during infection, as well as the plasticity of the formation of memory T cells. Additionally, we utilize static and live imaging to investigate howToxoplasma gondiilife cycle impacts the ability to present antigen to CD8+T cells. These studies aid in our understanding of how effector and memory CD8+T cell responses are generated and maintained during an infection.
Weighting of celestial and terrestrial cues in the monarch butterfly central complex
Authors: Nguyen Tu Anh Thi , Beetz M. Jerome , Merlin Christine , Pfeiffer Keram , Jundi Basil el
Published: January 2022

Monarch butterflies rely on external cues for orientation during their annual long-distance migration from Northern US and Canada to Central Mexico. These external cues can be celestial cues, such as the sun or polarized light, which are processed in the internal compass of the brain, termed the central complex (CX). Previous research typically focused on how individual simulated celestial cues are encoded in the butterfly’s CX. However, in nature, the butterflies perceive several celestial cues at the same time and need to integrate them to effectively use the compound of all cues for orientation. In addition, a recent behavioral study revealed that monarch butterflies can rely on terrestrial cues, such as the panoramic skyline, for orientation and use them in combination with the sun to maintain a directed flight course. How the CX encodes a combination of celestial and terrestrial cues and how they are weighted in the butterfly’s CX is still unknown. Here, we examined how input neurons of the CX, termed TL neurons, combine celestial and terrestrial information. While recording intracellularly from the neurons, we presented a sun stimulus and polarized light to the butterflies as well as a simulated sun and a panoramic scene simultaneously. Our results show that celestial cues are integrated linearly in these cells, while the combination of the sun and a panoramic skyline did not always follow a linear integration of action potential rates. Interestingly, while the weighting between the sun and polarized light was invariant between individual input neurons, it varied strongly when the sun stimulus and the panoramic skyline were presented simultaneously. Taken together, this dynamic weighting between celestial and terrestrial cues may allow the butterflies to flexibly set their cue preference during navigation.
The interferon-inducible GTPase MxB promotes capsid disassembly and genome release of herpesviruses
Authors: Serrero Manutea C. , Girault Virginie , Weigang Sebastian , Greco Todd M. , Ramos-Nascimento Ana , Anderson Fenja , Piras Antonio , Martinez Ana Hickford , Hertzog Jonny , Binz Anne , Pohlmann Anja , Prank Ute , Rehwinkel Jan , Bauerfeind Rudolf , Cristea Ileana M. , Pichlmair Andreas , Kochs Georg , Sodeik Beate
Published: January 2022

Host proteins sense viral products and induce defence mechanisms, particularly in immune cells. Using cell-free assays and quantitative mass spectrometry, we determined the interactome of capsid- host protein complexes of herpes simplex virus and identified the large dynamin-like GTPase myxovirus resistance protein B (MxB) as an interferon-inducible protein interacting with capsids. Electron microscopy analyses showed that cytosols containing MxB had the remarkable capability to disassemble the icosahedral capsids of herpes simplex viruses and varicella zoster virus into flat sheets of connected triangular faces. In contrast, capsids remained intact in cytosols with MxB mutants unable to hydrolyse GTP or to dimerize. Our data suggest that MxB senses herpesviral capsids, mediates their disassembly, and thereby restricts the efficiency of nuclear targeting of incoming capsids and/or the assembly of progeny capsids. The resulting premature release of viral genomes from capsids may enhance the activation of DNA sensors, and thereby amplify the innate immune responses.
Divergent Golgi Trafficking Limits B cell-Mediated IgG Sialylation
Authors: Glendenning Leandre M. , Zhou Julie Y. , Reynero Kalob M. , Cobb Brian A.
Published: January 2022

The degree of α2,6-linked sialylation on IgG glycans is associated with a variety of inflammatory conditions and is thought to drive IgG anti-inflammatory activity. Previous findings revealed that ablation of β-galactoside α2,6-sialyltransferase 1 (ST6Gal1) in B cells failed to alter IgG sialylation in vivo, yet resulted in the loss of B cell surface α2,6 sialylation, suggesting divergent pathways for IgG and cell surface glycoprotein glycosylation and trafficking. Employing both B cell hybridomas and ex vivo murine B cells, we discovered that IgG was poorly sialylated by ST6Gal1 and highly core fucosylated by α1,6-fucosyltransferase 8 (Fut8) in cell culture. In contrast, IgG-producing cells showed the opposite pattern by flow cytometry, with high cell surface α2,6 sialylation and low α1,6 fucosylation. Paired studies further revealed that ex vivo B cell-produced IgG carried significantly less sialylation compared to IgG isolated from the plasma of matched animals, providing evidence that sialylation increases after release in vivo. Finally, confocal analyses demonstrated that IgG poorly localized to subcellular compartments rich in sialylation and ST6Gal1, and strongly to regions rich in fucosylation and Fut8. These findings support a model in which IgG subcellular trafficking diverges from the canonical secretory pathway by promoting Fut8-mediated core fucosylation and limiting exposure to and modification by ST6Gal1, providing a mechanism for why B cell-expressed ST6Gal1 is dispensable for IgG sialylation in vivo.
The RNA helicase DDX39B activates FOXP3 RNA splicing to control T regulatory cell fate
Authors: Hirano Minato , Galarza-Muñoz Gaddiel , Schott Geraldine , Wang Liuyang , Nagasawa Chloe , Antonia Alejandro L. , Jain Vaibhav , Yu Xiaoying , Widen Steven G. , Briggs Farren B.S. , Gregory Simon G. , Ko Dennis C. , Samuel Fagg W. , Bradrick Shelton S. , Garcia-Blanco Mariano A.
Published: January 2022

Genes associated with increased susceptibility to multiple sclerosis (MS) have been identified, but their functions are incompletely understood. One of these genes codes for the RNA helicase DExD/H-Box Polypeptide 39B (DDX39B), which shows genetic and functional epistasis with interleukin-7 receptor-α gene (IL7R) in MS-risk. Based on evolutionary and functional arguments we postulated that DDX39B enhances immune tolerance decreasing MS risk. Consistent with such a role we show that DDX39B controls the expression of many MS susceptibility genes and important immune-related genes. Among these we identified ForkheadBox P3 (FOXP3), which codes for the master transcriptional factor in CD4+/CD25+ T regulatory cells. DDX39B knockdown led to loss of immune-regulatory and gain of immune-effector expression signatures. Splicing of FOXP3 introns, which belong to a previously unrecognized subclass of introns with C-rich polypyrimidine tracts, was exquisitely sensitive to DDX39B levels. Given the importance of FOXP3 in autoimmunity, this work cements DDX39B as an important guardian of immune tolerance.
Deep Learning Approaches to Identify Patients within the Thrombolytic Treatment Window
Authors: Polson Jennifer S , Zhang Haoyue , Nael Kambiz , Salamon Noriko , Yoo Bryan Y , El-Saden Suzie , Starkman Sidney , Kim Namkug , Kang Dong-Wha , Speier William F , Arnold Corey W
Published: January 2022

1.

Background

Treatment of acute ischemic stroke is heavily contingent upon time, as there is a strong relationship between time clock and tissue progression. We sought to a develop a deep learning algorithm for classifying time since stroke (TSS) from MR images by comparison to neuroradiologist assessments of imaging signal mismatch and evaluation on external data.

Methods

This retrospective study involved patients who underwent MRI from 2011-2019. Models were trained to classify TSS within 4.5 hours; performance metrics with confidence intervals were reported on both internal and external evaluation sets.

Results

A total of 772 patients (66 ± 9 years, 319 women) were used for model development and evaluation. Three board-certified neuroradiologists’ assessments, based on majority vote, yielded a sensitivity of 0.62, a specificity of 0.86, and a Fleiss’ kappa of 0.46. The deep learning method performed similarly to radiologists and outperformed previously reported methods, with the best model achieving an average evaluation accuracy, sensitivity, and specificity of 0.726, 0.712, and 0.741, on an internal cohort and 0.724, 0.757, and 0.679, respectively, on an external, unseen evaluation cohort from another institution.

Conclusion

This model achieved higher generalization performance on external evaluation datasets than the current state of the art for TSS classification.
Impact of voluntary risk-mitigation behaviour on transmission of the Omicron SARS-CoV-2 variant in England
Authors: Brooks-Pollock Ellen , Northstone Kate , Pellis Lorenzo , Scarabel Francesca , Thomas Amy , Nixon Emily , Matthews David A. , Bowyer Vicky , Garcia Maria Paz , Steves Claire J. , Timpson Nicholas J. , Danon Leon
Published: January 2022

Background

The Omicron variant of SARS-CoV-2 infection poses substantial challenges to public health. In England, “plan B” mitigation measures were introduced in December 2021 including increased home working and face coverings in shops, but stopped short of restrictions on social contacts. The impact of voluntary risk mitigation behaviours on future SARS-CoV-2 burden is unknown.

Methods

We developed a rapid online survey of risk mitigation behaviours during the winter 2021 festive period and deployed in two longitudinal cohort studies in the UK (Avon Longitudinal Study of Parents and Children (ALSPAC) and TwinsUK/Covid Symptom Study (CSS) Biobank) in December 2021. Using an individual-based, probabilistic model of COVID-19 transmission between social contacts with SARS-CoV-2 Omicron variant parameters and realistic vaccine coverage in England, we describe the potential impact of the SARS-CoV-2 Omicron wave in England in terms of the effective reproduction number and cumulative infections, hospital admissions and deaths. Using survey results, we estimated in real-time the impact of voluntary risk mitigation behaviours on the Omicron wave in England, if implemented for the entire epidemic wave.

Results

Over 95% of survey respondents (NALSPAC=2,686 and NTwins=6,155) reported some risk mitigation behaviours, with vaccination and using home testing kits reported most frequently. Less than half of those respondents reported that their behaviour was due to “plan B”. We estimate that without risk mitigation behaviours, the Omicron variant is consistent with an effective reproduction number between 2.5 and 3.5. Due to the reduced vaccine effectiveness against infection with the Omicron variant, our modelled estimates suggest that between 55% and 60% of the English population could be infected during the current wave, translating into between 15,000 and 46,000 cumulative deaths, depending on assumptions about vaccine effectiveness. We estimate that voluntary risk reduction measures could reduce the effective reproduction number to between 1.8 and 2.2 and reduce the cumulative number of deaths by up to 24%.

Conclusions

We conclude that voluntary measures substantially reduce the projected impact of the SARS-CoV-2 Omicron variant, but that voluntary measures alone would be unlikely to completely control transmission.
Patterns of treatment and outcome in patients with unresectable or inoperable esophageal cancer: A real world data
Authors: Chauhan Richa , Trivedi Vinita , Rani Rita , Singh Usha
Published: January 2022

Background

Esophageal cancer is the eighth most common cancer in the world with a high mortality rate. Surgery, radiation and chemotherapy have been tried in various combinations to improve on the survival rates. Our study provides real world data from a South Asian country on patterns of treatment and outcome in patients with unresectable or inoperable esophageal cancer.

Materials and Methods

This study is a retrospective analysis of all consecutive esophageal cancer patients, with medically inoperable or unresectable disease, and treated with conformal radical radiotherapy at a tertiary cancer center from January, 2016 to December, 2017. Data regarding patients’ age, histology, location, pre-treatment imaging, disease stage, treatment details, compliance and response to treatment and status at last follow-up were retrieved from their file. Continuous and categorical variables were summarized by descriptive statistics.

Results

A total of 100 esophageal cancer patients with a mean age of 60.24 years were included in the study. 60% of the patients were male and upper one-third was the most common site involved. Squamous cell carcinoma was reported in 83% of the patients. About 70% of the patients had a T3/T4 disease and 44% also had nodal metastasis. The radiation dose ranged from 45Gy – 63Gy (median = 59.4Gy). Further, 15% and 54% of the patients received neoadjuvant and concurrent chemotherapy respectively. Radiation compliance was seen in 90% of the patients. With a median follow-up of 7 months (range 3-58 months), 80% of the patients were alive with 32.22% having no evidence of disease. Distant metastases and loco regional failure was seen in 32.22% and 28% of the patients respectively.

Conclusion

Our study showed that esophageal cancer is more common in elderly males. Adherence to a uniform treatment protocol using concurrent chemo radiation is difficult in clinical practice especially in resource constrained set up. Both distant metastases and loco regional failure continues to be a matter of concern. Further improvement in local control must be evaluated by either radiation dose escalation or novel combinations with chemotherapy and immunotherapy in large, multi centric trial settings.
SARS-CoV-2 anti-spike antibody levels following second dose of ChAdOx1 nCov-19 or BNT162b2 in residents of long-term care facilities in England (VIVALDI)
Authors: Stirrup Oliver , Krutikov Maria , Tut Gokhan , Palmer Tom , Bone David , Bruton Rachel , Fuller Chris , Azmi Borscha , Lancaster Tara , Sylla Panagiota , Kaur Nayandeep , Spalkova Eliska , Bentley Christopher , Amin Umayr , Jadir Azar , Hulme Samuel , Giddings Rebecca , Nacer-Laidi Hadjer , Baynton Verity , Irwin-Singer Aidan , Hayward Andrew , Moss Paul , Copas Andrew , Shallcross Laura
Published: January 2022

Background

General population studies have shown strong humoral response following SARS-CoV-2 vaccination with subsequent waning of anti-spike antibody levels. Vaccine-induced immune responses are often attenuated in frail and older populations such as Long-Term Care Facility (LTCF) residents but published data are scarce.

Methods

VIVALDI is a prospective cohort study in England which links serial blood sampling in LTCF staff and residents to routine healthcare records. We measured quantitative titres of SARS-CoV-2 anti-spike antibodies in residents and staff following second vaccination dose with ChAdOx1 nCov-19 (Oxford-AstraZeneca) or BNT162b2 (Pfizer-BioNTech). We investigated differences in peak antibody levels and rates of decline using linear mixed effects models.

Results

We report on 1317 samples from 402 residents (median age 86 years, IQR 78-91) and 632 staff (50 years, 37-58), ≤280 days from second vaccination dose. Peak antibody titres were 7.9-fold higher after Pfizer-BioNTech vaccine compared to Oxford-AstraZeneca (95%CI 3.6-17.0; P<0.01) but rate of decline was increased, and titres were similar at 6 months. Prior infection was associated with higher peak antibody levels in both Pfizer-BioNTech (2.8-fold, 1.9-4.1; P<0.01) and Oxford-AstraZeneca (4.8-fold, 3.2-7.1; P<0.01) recipients and slower rates of antibody decline. Increasing age was associated with a modest reduction in peak antibody levels for Oxford-AstraZeneca recipients.

Conclusions

Double-dose vaccination elicits robust and stable antibody responses in older LTCF residents, suggesting comparable levels of vaccine-induced immunity to that in the general population. Antibody levels are higher after Pfizer-BioNTech vaccination but fall more rapidly compared to Oxford-AstraZeneca recipients and are enhanced by prior infection in both groups.
VEGF-C expression in TAMs rewires the metastatic fate of breast cancer cells
Authors: Banerjee Kaveri , Kerzel Thomas , Bekkhus Tove , de Souza Ferreira Sabrina , Wallmann Tatjana , Wallerius Majken , Landwehr Laura-Sophie , Agardy Dennis Alexander , Schauer Nele , Malmerfeldt Anna , Bergh Jonas , Bartish Margarita , Hartman Johan , Östman Arne , Squadrito Mario Leonardo , Rolny Charlotte
Published: January 2022

Expression of pro-lymphangiogenic vascular endothelial growth factor C (VEGF-C) in primary tumors correlates with the occurrence of proximal lymph node metastasis in most solid cancer types. However, the role of VEGF-C in regulating tumor cell dissemination to distant organs is currently unclear. Perivascular tumor-associated macrophages (TAMs) are key regulators of hematogenous cancer cell spreading, forming tumor microenvironment of metastasis (TMEM) doorways for breast cancer cells to intravasate tumor blood vessels and fuel distant metastases. Using an experimental breast cancer (BC) model, we show here that TAMs expressing VEGF-C decrease cancer cell dissemination to the lung while enhancing lymph node metastasis. These TAMs express podoplanin and associate with normalized tumor blood vessels expressing VEGFR3. Further clinical data reveal that VEGF-C+TAMs correlate inversely with malignant grade and with the occurrence of TMEM complexes in a cohort of BC patients. Thus, our study displays an apparently paradoxical role of VEGF-C expressing TAMs in redirecting cancer cells to preferentially disseminate to the lymph nodes, at least in part, by normalizing tumor blood vessels and promoting lymphangiogenesis.
A pipeline to track unlabeled cells in wide migration chambers using pseudofluorescence
Authors: Paola Antonello , Thelen Marcus , Krause Rolf , Ulisse Pizzagalli Diego
Published: January 2022

Cell migration is a pivotal biological process, whose dysregulation is found in many diseases including inflammation and cancer. Advances in microscopy technologies allow now to study cell migration in vitro, within microenvironments that resemble in vivo conditions. However, when cells are observed within large 3D migration chambers at low magnification and for extended periods of time, data analysis becomes difficult. Indeed, cell detection and tracking are hampered due to the large pixel size, the possible low signal-to-noise ratio and distortions in the cell shape due to changes in the z-axis position. Although fluorescent staining can be used to facilitate cell detection, it may alter cell behavior and suffer from fluorescence loss over time (photobleaching).

Here we describe the application of an image analysis pipeline based on deep learning to convert the transmitted light signal from unlabeled lymphoma cells to pseudofluorescence. Such pipeline confers a significant improvement in tracking accuracy while not suffering from photobleaching. This is reflected in the possibility of tracking cells for three-fold longer periods of time.
Genetic modifiers of Cep290-mediated retinal degeneration
Authors: Meyer K.J. , Whitmore S.S. , Burnight E.R. , Riker M. , Mercer H.E. , Ajose A. , Howell G.R. , Stone E.M. , Mullins R.F. , Tucker B.A. , Scheetz T.E. , Anderson M.G.
Published: January 2022

Mutations in CEP290 cause up to 30% of cases of Leber congenital amaurosis (LCA), a severe childhood blindness resulting from abnormalities in the photoreceptor connecting cilia that lead to rapid retinal degeneration. Like many genetic diseases, CEP290-LCA has considerable variable expressivity, indicating the presence of other factors that influence phenotypic outcome. Here, we have undertaken a phenotype-driven approach in mice to identify genetic modifiers of CEP290-mediated retinal degeneration through backcrosses and intercrosses between BXD24-Cep290rd16 mice and the genetically distinct inbred CAST strain to introduce genetic variation. Optical coherence tomography was used to quantitatively measure retinal thickness as a surrogate indication of photoreceptor degeneration in the resulting rd16-mutant mice. It was readily apparent that CEP290-mediated retinal degeneration in the resulting mice is sensitive to genetic background, with some mice exhibiting relatively thick laminated retinas and others having thin retinas with advanced disease. Quantitative trait locus (QTL) analysis identified multiple genomic loci capable of influencing the retinal degeneration phenotype in Cep290-mutant mice that together account for 71.7% of the phenotypic variation in retinal thickness observed in our population. Following the QTL analysis, two suppressor loci were studied in detail through a combination of physical and molecular approaches to narrow the critical region for each QTL and identify the probable causative genetic variations.
Single-cell RNA sequencing identifies phenotypically, functionally, and anatomically distinct stromal niche populations in human bone marrow
Authors: Li Hongzhe , Bräunig Sandro , Dhapolar Parashar , Karlsson Göran , Lang Stefan , Scheding Stefan
Published: January 2022

Hematopoiesis is regulated by the bone marrow (BM) stroma. However, cellular identities and functions of the different BM stromal elements in humans remain poorly defined. Based on single-cell RNA sequencing, we systematically characterized the BM stromal compartment which led to the identification of six transcriptionally and functionally distinct stromal cell populations. Stromal cell differentiation hierarchy was recapitulated based on RNA velocity analysis, in vitro proliferation capacities and differentiation potentials. Potential key factors that govern the transition from stem and progenitor cells to fate- committed cells were identified. In silico cell-cell communication prediction and in situ localization analyses demonstrated distinct hematopoietic stromal cell niches in specific BM locations, which used either the CXCL12 or SPP1 axis as the major hematopoiesis-regulating mechanism. These findings provide the basis for a comprehensive understanding of the cellular complexity of the human BM microenvironment and the intricate stroma- hematopoiesis crosstalk mechanisms, thus refining our current view on hematopoietic niche organization.
A peptide-mediated, multilateral molecular dialogue for the coordination of pollen wall formation
Authors: Truskina Jekaterina , Brück Stefanie , Stintzi Annick , Boeuf Sophy , Doll Nicolas M. , Fujita Satoshi , Geldner Niko , Schaller Andreas , Ingram Gwyneth C.
Published: January 2022

The surface of pollen grains is reinforced by pollen wall components produced non-cell autonomously by tapetum cells that surround developing pollen within the male floral organ, the anther. Here we show that tapetum activity is regulated by the GASSHO (GSO) receptor-like kinase pathway, controlled by two sulfated peptides, CASPARIAN STRIP INTEGRITY FACTOR 3 (CIF3) and CIF4, the precursors of which are expressed in the tapetum itself. Coordination of tapetum activity with pollen grain development depends on the action of subtilases, including AtSBT5.4, which are produced stage-specifically by developing pollen grains. Tapetum-derived CIF precursors are processed by subtilases, triggering GSO-dependent tapetum activation. We show that the GSO receptors act from the middle layer, a tissue surrounding the tapetum and developing pollen. Three concentrically organized cell types therefore cooperate to coordinate pollen wall deposition through a multilateral molecular dialog.

Significance Statement

Pollen viability depends on a tough external barrier called the pollen wall. Pollen wall components are produced by tapetum cells which surround developing pollen grains within the anther. Precise coordination of tapetum activity with pollen grain development is required to ensure effective pollen wall formation. Here we reveal that this is achieved through a multidirectional dialogue involving three distinct cell types. We show that peptide precursors from the tapetum are activated by proteases produced stage-specifically in developing pollen grains. Unexpectedly, we found that activated peptides are perceived not in the tapetum, but in the middle layer, which encloses the developing tapetum and pollen grains, revealing an unsuspected role for this enigmatic cell layer in the control of tapetum development.
Entorhinal layer 6b subplate neurons govern spatial learning and memory
Authors: Ben-Simon Yoav , Kaefer Karola , Velicky Philipp , Csicsvari Jozsef , Danzl Johann G. , Jonas Peter
Published: January 2022

The mammalian hippocampal formation (HF) plays a key role in several higher brain functions, such as spatial coding, learning and memory. Its simple circuit architecture is often viewed as a “trisynaptic loop”, processing input originating from the superficial layers of the entorhinal cortex (EC) in a linear fashion, and sending it back to its deeper layers. Here, we show that contrary to this canonical view, excitatory neurons in the deepest layer of the mouse EC, exhibiting the unique morphology and molecular profile of layer 6b cortical subplate neurons (SPNs), project to all sub-regions comprising the HF, with a preference towards CA3 pyramidal neurons, and receive input from the CA1, thalamus and claustrum. Furthermore, their output is characterized by unique slow-decaying excitatory post-synaptic currents (EPSCs), capable of driving plateau-like potentials in their target cells. Optogenetic inhibition of the EC-6b pathway markedly affects spatial coding in CA1 pyramidal neurons, while cell ablation impairs not only acquisition of new spatial memories, but also degradation of previously acquired ones. Our results provide the first evidence of a functional role for the cortical layer 6b neurons in the adult brain and elucidate a critical novel element determining cortico-hippocampal activity patterns.
Fgfr2b signaling is essential for the maintenance of the alveolar epithelial type 2 lineage during lung homeostasis in mice
Authors: Ahmadvand Negah , Lingampally Arun , Khosravi Farhad , Vazquez-Armendariz Ivonne , Rivetti Stefano , Wilhelm Jochen , Herold Susanne , Barreto Guillermo , Koepke Janine , Samakovlis Christos , Carraro Gianni , Zhang Jin-San , Al Alam Denise , Bellusci Saverio
Published: January 2022

Fibroblast growth factor receptor 2b (Fgfr2b) signaling is essential throughout lung development to form the alveolar epithelial lineage. However, its role in alveolar epithelial type 2 cells (AT2s) homeostasis was recently considered dispensable. SftpcCreERT2; tdTomatoflox/flox mice were used to delete Fgfr2b expression in cells belonging to the AT2 lineage, which contains mature AT2s and a novel SftpcLow lineage-traced population called “injury activated alveolar progenitors” or IAAPs. Upon continuous tamoxifen exposure for either one or two weeks to delete Fgfr2b, a shrinking of the AT2s is observed. Mature AT2s exit the cell cycle, undergo apoptosis and fail to form alveolospheres in vitro. However, the lung morphometry appears normal, suggesting the involvement of compensatory mechanisms. In mutant lungs, IAAPs which escaped Fgfr2b deletion expand, display enhanced alveolosphere formation in vitro and increase drastically their AT2 signature suggesting differentiation towards mature AT2s. Interestingly, a significant increase in AT2s and decrease in IAPPs occurs after a one-week tamoxifen exposure followed by an eight-week chase period. While mature AT2s partially recover their alveolosphere formation capabilities, the IAAPs no longer display this property. Single-cell RNA seq analysis confirms that AT2s and IAAPs represent stable and distinct cell populations and recapitulate some of their characteristics observed in vivo. Our results underscore the essential role played by Fgfr2b signaling in the maintenance of the AT2 lineage in the adult lung and suggest that the IAAPs could represent a new population of AT2 progenitors.
Interaction of the Proline-Rich Domain of Fission Yeast WASp (Ws1p1) with Actin Filaments
Authors: Rosenbloom Aaron D. , Pollard Thomas D.
Published: January 2022

Background

The Wiskott-Aldrich Syndrome protein (WASp) family of proteins plays a crucial role in the activation of the Arp2/3 (actin-related protein 2/3) complex to promote the branching of actin filaments. The proline-rich domain (PRD) of WASp is known to contribute to branching nucleation but was overlooked, until experiments showed that the PRD of budding yeast Las17 can bind actin filaments (1).

Methods

We purified recombinant proline-rich domains from fission yeast S. pombe Wsp1 and budding yeast S. cerevisiae Las17 to test in biochemical assays of actin binding and polymerization.

Results

The PRD of the S. pombe Wsp1 binds actin filaments with micromolar affinity. The PRDs of both Wsp1 and Las17 slowed the rate of actin filament elongation by Mg-ATP-actin monomers by half and slowed the spontaneous polymerization of Mg-ATP-actin monomers modestly.

Conclusion

The affinity of PRDs of WASp-family proteins for actin filaments is high enough to contribute to the reported stimulation of actin filament branching by Arp2/3 complex.
Anti-SARS-CoV-2 swine glyco-humanized polyclonal antibody XAV-19 retains neutralizing activity against SARS-CoV-2 B.1.1.529 (Omicron)
Authors: Vanhove Bernard , Marot Stéphane , Evanno Gwénaëlle , Malet Isabelle , Rouvray Gaëtane , Shneiker Françoise , Mevel Edwige , Ciron Carine , Rousse Juliette , Royer Pierre-Joseph , Lheriteau Elsa , Raffi François , Duvaux Odile , Marcelin Anne-Geneviève , Calvez Vincent
Published: January 2022

B.1.1.529 is the SARS-CoV-2 variant designated Omicron by the WHO in November 2021. It is a highly divergent variant with a high number of mutations, including 26-32 mutations in the spike protein among which 15 in the Receptor Binding Domain (RBD) including at the human angiotensin converting enzyme 2 (ACE-2) receptor interacting interface. Because of a decreased affinity for the ACE-2 receptor and a geometric reorganization of the S1-S2 cleavage site, the Omicron variant is predicted to not have a significant infectivity advantage over the delta variant and to be less pathogenic than Delta. However, in Omicron, neutralizing epitopes are greatly affected, suggesting that current vaccines and neutralizing monoclonal antibodies might confer reduced protection against this variant. In contrast, we and others previously demonstrated that polyclonal antibodies against SARS-CoV-2 RBD obtained from hyperimmunized animal hosts do maintain their neutralizing properties against Alpha to Delta. Here, we confirmed these findings by showing that XAV-19, a swine glyco-humanized polyclonal antibody retains full neutralizing activity against Omicron.
Single-Cell Analysis Reveals Distinct Immune and Smooth Muscle Cell Populations that Contribute to Chronic Thromboembolic Pulmonary Hypertension
Authors: Viswanathan Gayathri , Kirshner Hélène Fradin , Nazo Nour , Ganapathi Asvin , Cummings Ian , Choi Issac , Warman Anmol , Almeida-Peters Susana , Haney John , Corcoran David , Yu Yen-Rei , Rajagopal Sudarshan
Published: January 2022

Chronic thromboembolic pulmonary hypertension (CTEPH) is a sequelae of acute pulmonary embolism (PE) in which the PE remodels into a chronic scar in the pulmonary arteries. This results in vascular obstruction, small vessel arteriopathy and pulmonary hypertension. Our current understanding of CTEPH pathobiology is primarily derived from cell-based studies limited by the use of specific cell markers or phenotypic modulation in cell culture. Here we used single cell RNA sequencing (scRNAseq) of tissue removed at the time of pulmonary thromboendarterectomy (PTE) surgery to identify the multiple cell types, including macrophages, T cells, and smooth muscle cells, that comprise CTEPH thrombus. Notably, multiple macrophage subclusters were identified but broadly split into two categories, with the larger group characterized by an upregulation of inflammatory signaling predicted to promote pulmonary vascular remodeling. Both CD4+ and CD8+ T cells were identified and likely contribute to chronic inflammation in CTEPH. Smooth muscle cells were a heterogeneous population, with a cluster of myofibroblasts that express markers of fibrosis and are predicted to arise from other smooth muscle cell clusters based on pseudotime analysis. Additionally, cultured endothelial, smooth muscle and myofibroblast cells isolated from CTEPH thrombus have distinct phenotypes from control cells with regards to angiogenic potential and rates of proliferation and apoptosis. Lastly, our analysis identified protease-activated receptor 1 (PAR1) as a potential therapeutic target that links thrombosis to chronic PE in CTEPH, with PAR1 inhibition decreasing smooth muscle cell and myofibroblast proliferation and migration. These findings suggest a model for CTEPH similar to atherosclerosis, with chronic inflammation promoted by macrophages and T cells driving vascular remodeling through smooth muscle cell modulation, and suggest new approaches for pharmacologically targeting this disease.
Temperature modulates PVN pre-sympathetic neurones via transient receptor potential ion channels
Authors: O’Brien Fiona , Feetham Claire , Staunton Caroline , Hext Katharine , Barrett-Jolley Richard
Published: January 2022

Background and purpose

The paraventricular nucleus (PVN) of the hypothalamus plays a vital role in maintaining homeostasis and controls cardiovascular function via autonomic pre-sympathetic neurones. We have previously shown that coupling between transient receptor potential cation channel subfamily V member 4 (Trpv4) and small-conductance calcium-activated potassium channels (SK) in the PVN facilitate osmosensing. TRP channels are also thermosensitive and therefore, in this report we investigated the temperature sensitivity of PVN neurones.

Experimental approach

We identified TRP channel mRNA in mouse PVN using quantitative reverse transcription-PCR (RT-PCR). Using cell-attached patch-clamp electrophysiology, we characterised the thermosensitivity of Trpv4-like ion channels on mouse PVN neurones. Following recovery of temperature sensitive single channel kinetic schema, we constructed a novel and predictive stochastic mathematical model of these neurones. We then validated this model with electrophysiological recordings of action current frequency from mouse PVN neurones.

Results

We identified 7 TRP channel genes in the PVN with known thermosensitive capabilities. Trpv4 was the most abundant of these and was easily identified at the single channel level using cell-attached patch-clamp electrophysiology on PVN neurones. We investigated the thermosensitivity of these Trpv4-like channels; open probability (Po) markedly decreased when temperature was decreased, mediated by a decrease in mean open dwell times. Our neuronal model predicted that PVN spontaneous action current frequency (ACf) would increase as temperature is decreased and in our electrophysiological experiments, we found that ACf from PVN neurones was significantly higher at lower temperatures. The broad-spectrum channel blocker, gadolinium (100 μM), was used to block the warm-activated Ca2+-permeable Trpv4 and Trpv3 channels. In the presence of gadolinium (100 μM), the temperature effect was largely retained. Using econazole (10 μM), a blocker of Trpm2, we found there were significant increases in overall ACf and the temperature effect was inhibited.

Conclusion

Our work identified Trpv4 mRNA as an abundantly expressed thermosensitive TRP channel gene in the PVN and this ion channel contributes to the intrinsic thermosensitive properties of PVN neurones. At physiological temperatures (37°C), we observed relatively low ACf primarily due to the activity of Trpm2 channels, whereas at room temperature, where most of the previous characterisation of PVN neuronal activity has been performed, ACf is much higher, and appears to be predominately due to reduced Trpv4 activity. This work gives insight into the fundamental mechanisms by which the body decodes temperature signals and maintains homeostasis.