bluuebunny/crossref_metadata_2025_split huggingface dataset:
Human iPSC-derived Microglial Cells Integrated into Mouse Retina and Recapitulated Features of Endogenous Microglia
Authors: Ma Wenxin , Zhao Lian , Xu Biying , Fariss Robert N. , Redmond T. Michael , Zou Jizhong , Wong Wai T. , Li Wei 
Published: August 2023

AbstractMicroglia exhibit both maladaptive and adaptive roles in the pathogenesis of neurodegenerative diseases and have emerged as a cellular target for central nervous system (CNS) disorders, including those affecting the retina. Replacing maladaptive microglia, such as those impacted by aging or over-activation, with exogenous microglia that can enable adaptive functions has been proposed as a potential therapeutic strategy for neurodegenerative diseases. To investigate microglia replacement as an approach for retinal diseases, we first employed a protocol to efficiently generate human-induced pluripotent stem cells (hiPSC)-derived microglia in quantities sufficient for in vivo transplantation. These cells demonstrated expression of microglia-enriched genes and showed typical microglial functions such as LPS-induced responses and phagocytosis. We then performed xenotransplantation of these hiPSC-derived microglia into the subretinal space of adult mice whose endogenous retinal microglia have been pharmacologically depleted. Long-term analysis post-transplantation demonstrated that transplanted hiPSC-derived microglia successfully integrated into the neuroretina as ramified cells, occupying positions previously filled by the endogenous microglia and expressed microglia homeostatic markers such as P2ry12 and Tmem119. Further, these cells were found juxtaposed alongside residual endogenous murine microglia for up to eight months in the retina, indicating their ability to establish a stable homeostatic statein vivo. Following retinal pigment epithelial (RPE) cell injury, transplanted microglia demonstrated responses typical of endogenous microglia, including migration, proliferation, and phagocytosis. Our findings indicate the feasibility of microglial transplantation and integration in the retina and suggest that modulating microglia through replacement may be a therapeutic strategy for treating neurodegenerative retinal diseases.
Identifying repurposed drugs with moderate anti-influenza virus activity through computational prioritization of drug-target pairs
Authors: Taye Biruhalem , Thünauer Roland , Sugrue Richard J , Maurer-Stroh Sebastian , Kosinski Jan 
Published: August 2023

AbstractInfluenza A virus (IAV) causes up to five million cases of severe illness and half a million deaths worldwide each year. While there are a few clinically approved drugs for treating IAV, they are challenged by the rapid evolution of the virus leading to emergence of drug resistance and the adverse effects of the drugs. Targeting host cellular factors that support virus replication could limit resistance, increase the broad-spectrum antiviral properties of drugs, and benefit from repurposing drugs already existing against those factors. However, selecting the right drug-target pairs with low toxicity and minimal adverse effects has been challenging, even though hundreds of cellular host factors have been identified. In this study, we applied a computational and knowledge-based drug-target prioritization approach to identify promising drug-target pairs. We selected five pairs for experimental validation: telmisartan-Angiotensin II receptor, type 1 (AGTR1), metoclopramide hydrochloride-Cholinergic receptor muscarinic 1 (CHRM1), cefepime hydrochloride-phosphogluconate dehydrogenase (PGD), ranolazine dihydrochloride-sodium channel voltage-gated type v alpha subunit (SCN5A), and ofloxacin-topoisomerase II alpha 170kDa (TOP2A). Except for cefepime hydrochloride, all four drugs showed significant plaque reduction in Madin Darby canine kidney (MDCK) cells. In the immunofluorescence assay, metoclopramide hydrochloride, ranolazine dihydrochloride, and telmisartan showed antiviral activity in MDCK and/or adenocarcinoma human alveolar basal epithelial (A549) cell lines. In conclusion, our approach can prioritize and identify drugs with antiviral activity against influenza virus. Refining and strengthening such approaches could be valuable for rapid antiviral discovery and pandemic preparedness.HighlightsComputational drug-target prioritization indicated drugs for validationTelmisartan showed anti-influenza virus activity in MDCK cellsMetoclopramide and ranolazine showed anti-influenza activity in A549 and MDCK cells
Humoral correlates of protection againstMycobacterium tuberculosisfollowing intravenous Bacille Calmette-Guérin vaccination in rhesus macaques
Authors: Irvine Edward B. , Darrah Patricia A. , Wang Shu , Wang Chuangqi , McNamara Ryan P. , Roederer Mario , Seder Robert A. , Lauffenburger Douglas A. , Flynn JoAnne L. , Fortune Sarah M. , Alter Galit 
Published: August 2023

ABSTRACTAltering the route of Bacille Calmette-Guérin (BCG) immunization from low-dose intradermal vaccination to high-dose intravenous (IV) vaccination resulted in a high level of protection againstMycobacterium tuberculosis(Mtb) infection, providing an opportunity to uncover immune correlates and mechanisms of protection. In addition to strong T cell immunity, IV BCG vaccination was associated with a robust expansion of humoral immune responses that tracked with bacterial control. However, given the near complete protection afforded by high-dose IV BCG immunization, a precise correlate of immune protection was difficult to define. Here we leveraged plasma and bronchoalveolar lavage fluid (BAL) from a cohort of rhesus macaques that received decreasing doses of IV BCG and aimed to define the correlates of immunity across macaques that experienced immune protection or breakthrough infection followingMtbchallenge. We show an IV BCG dose-dependent induction of mycobacterial-specific humoral immune responses, both in the plasma and in the airways. Moreover, antibody responses at peak immunogenicity significantly predicted bacterial control following challenge. Multivariate analyses revealed antibody-mediated complement and NK cell activating humoral networks as key functional signatures associated with protective immunity. Collectively, this work extends our understanding of humoral biomarkers and potential mechanisms of IV BCG mediated protection againstMtb.
Epi-microRNA mediated metabolic reprogramming ensures affinity maturation
Authors: Nakagawa Rinako , Llorian Miriam , Varsani-Brown Sunita , Chakravarty Probir , Camarillo Jeannie M. , Barry David , George Roger , Blackledge Neil P. , Duddy Graham , Kelleher Neil L. , Klose Robert J. , Turner Martin , Calado Dinis P. 
Published: August 2023

AbstractTo increase antibody affinity against pathogens, positively selected GC-B cells initiate cell division in the light zone (LZ) of germinal centres (GCs). Among those, higher-affinity clones migrate to the dark zone (DZ) and vigorously proliferate by relying on oxidative phosphorylation (OXPHOS). However, it remains unknown how positively selected GC-B cells adapt their metabolism for cell division in the glycolysis-dominant, cell cycle arrest-inducing, hypoxic LZ microenvironment. Here, we show that microRNA (miR)-155 mediates metabolic reprogramming during positive selection to protect high-affinity clones. Transcriptome examination and mass spectrometry analysis revealed that miR-155 regulates H3K36me2 levels by directly repressing hypoxia-induced histone lysine demethylase,Kdm2a. This is indispensable for enhancing OXPHOS through optimizing the expression of vital nuclear mitochondrial genes under hypoxia. The miR-155-Kdm2ainteraction is crucial to prevent excessive production of reactive oxygen species and apoptosis. Thus, miR-155-mediated epigenetic regulation promotes mitochondrial fitness in high-affinity clones, ensuring their expansion and consequently affinity maturation.
Biophysical basis for the induction of glioblastoma-like phenotype in astrocytes
Authors: Starck Laurent , Sarem Melika , Heimrich Bernd , Sawarkar Ritwick , Ritz Marie-Françoise , Hutter Gregor , Shastri V. Prasad 
Published: August 2023

AbstractWhile the direct biological factors underlying the progression of GBM, an aggressive form of brain cancer, have been extensively studied, emerging evidence suggests that indirect biological triggers, such as traumatic brain injury (TBI), may also have a role. Since reactive astrocytes are associated with TBI, and astroglial cells are the source of proteoglycans which contribute to changes in biophysical characteristics (stochastic topography, stiffness) of the brain, we postulated a role for stochastic nanoroughness in the induction of glioma. Using a model system to emulate such physical cues, we demonstrate that human cortical astrocytes undergo spontaneous organization into spheroids in response to nanoroughness and retain the spheroid phenotype even upon withdrawal of the physical cues. Furthermore, spheroids serve as aggregation foci for naïve astrocytes; express activated MMP2, and disseminate upon implantation in mouse brain. RNA-seq revealed a tumoral phenotype with a gene expression pattern involving p53, ADAMTS proteases and fibronectin. Moreover, nanoroughness mediates a cross-talk between cancer cells and astrocytes through induced senescence. These findings implicate a role for stochastic biophysical cues in driving a potential malignant transformation of astrocytes.
Psychopathic traits predict neural responses to emotional movies in the general population
Authors: Aksiuto Anna , Lahnakoski Juha M. , Kauttonen Janne , Saarimäki Heini , Volynets Sofia , Nummenmaa Lauri , Sams Mikko , Jääskeläinen Iiro P. 
Published: August 2023

ABSTRACTPsychopathic personality traits such as callousness, manipulativeness, and lack of empathy are continuously distributed in non-institutionalized populations. Here, we show that brain responses to emotional movies in healthy females vary as a function of primary (PP) and secondary psychopathy (SP) traits. Healthy female volunteers (n=50) with variable scorings in Levenson Self-Report Psychopathy Scale watched a set of emotional movie clips (n=39) during functional magnetic resonance imaging of brain activity. We found that participants low on primary and secondary psychopathy traits showed higher cortical and subcortical activity when watching emotional movies, what possibly indicates the somatosensory simulation and empathizing with the emotions of movie characters. On the other hand, the individuals with high primary and secondary psychopathy traits presented a lack of vicarious engagement during watching movies and they manifested brainstem-based heightened response towards thread-related stimuli, which could indicate potent autonomic reactivity. Overall, our results indicate that variability in psychopathic traits in the general population is manifested as differential neural responses to emotional signals, and confirm previous findings of there being separate PP and SP personality characteristics due to the dissociable brain activity patterns while viewing emotional movies.
Perceptual axioms are irreconcilable with Euclidean geometry
Authors: Zeki Semir , Hale Zachary F , Beyh Ahmad , Rasche Samuel E 
Published: August 2023

AbstractThere are different definitions of axioms, but the one that seems to have general approval is that axioms are statements whose truths are universally accepted but cannot be proven; they are the foundation from which further propositional truths are derived. Previous attempts, led by David Hilbert, to show that all of mathematics can be built into an axiomatic system that is complete and consistent failed when Kurt Gödel proved that there will always be statements which are known to be true but can never be proven within the same axiomatic system. But Gödel and his followers took no account of brain mechanisms that generate and mediate logic. In this largely theoretical paper, but backed by previous experiments and our new ones reported below, we show that in the case of so-called “optical illusions” there exists a significant and irreconcilable difference between their visual perception and their description according to Euclidean geometry; when participants are asked to adjust, from an initial randomised state, the perceptual geometric axioms to conform to the Euclidean description, the two never match, although the degree of mismatch varies between individuals. These results provide evidence that perceptual axioms, or statements known to be perceptually true, cannot be described mathematically. Thus the logic of the visual perceptual system is irreconcilable with the cognitive (mathematical) system and cannot be updated even when knowledge of the difference between the two is available. Hence no one brain reality is more “objective” than any other.
Optimising CNT-FET Biosensor Design: Predictive Modelling of Biomolecular Electrostatic Gating and its Application to Beta-Lactamase Detection
Authors: Gwyther Rebecca E.A. , Côté Sébastien , Lee Chang-Seuk , Ramakrishnan Krithika , Palma Matteo , Jones D. Dafydd 
Published: August 2023

AbstractCarbon nanotube field effect transistor (CNT-FET) setups hold great promise for constructing next generation miniaturised biosensors whereby a biomolecular event gates conductance. The main issue is predicting how proteins, with their innate mosaic and distinctive electrostatic surfaces, interact with and thus modulate conductance of the CNT-FET. To overcome this barrier, we used advanced sampling molecular dynamics combined with non-canonical amino acid chemistry, to model the protein electrostatic potential imparted on SWCNTs. Here, we focused our efforts using β-lactamase binding protein (BLIP2) as the receptor due to its potential as a biosensor for the most common antibiotic degrading enzymes, the β-lactamases (BLs). Modelling was confirmed experimentally by attaching BLIP2 at single designed residues positions directly to SWCNTs using genetically encoded phenyl azide photochemistry. Our devices were able to successfully detect the two different BLs, TEM-1 and KPC-2, with each BL generating distinct conductance profiles due to differences in their unique surface electrostatic profiles presented close to the SWCNT surface. The changes in conductance closely matched the predicted electrostatic profile sampled by the SWCNTs on BL binding. Thus, our modelling approach combined with new and straight-forward residue-specific receptor attachment techniques, provides a general approach for more effective and optimal CNT-FET biosensor construction.
RNA binding protein IMP1/ZBP1 drivesActbmRNA localization and local translation in microglial peripheral processes and mediates morphological changes, motility and phagocytosis in response to inflammation
Authors: Imaz-Iruretagoyena Josune , Blanco-Urrejola Maite , Núñez-García Irene , García-Toledo Irene , Fernández-Beltrán Luis C , Márquez Mar , Corrochano Silvia , Sierra Amanda , Baleriola Jimena 
Published: August 2023

ABSTRACTPolarized cells in the brain, such as neurons and glia, rely on the asymmetric distribution of their proteins compartmentalizing the function of dendrites, axons, glial projections and endfeet. Subcellular proteomes can be assembled either by the transport of proteins synthesized in the cell soma or by the delivery of mRNAs to target compartments where they are locally translated into protein. This latter mechanism is known as local protein synthesis or local translation, and it has been best studied in neurons. Increasing evidence suggest it is also required to maintain local protein homeostasis in glial cells, however, in microglia, local translation remains largely unexplored. Given the scant evidence, we aimed at exploring the existence of local translation in peripheral microglial processes (PeMPs) and unravel its functional significance in response to inflammation, a major hallmark of neurodegenerative diseases. We report that local translation in PeMPs is enhanced by triggering a microglial inflammatory response with bacterial lipopolysaccharides (LPS). We found thatActbmRNA polarizes to PeMPs and is locally translated upon LPS exposure. mRNA localization in eukaryotic cells is driven by RNA binding proteins. Interestingly, downregulation of theActbbinding protein IMP1/ZBP1 impairedActbmRNA polarization and its localized translation, and led to defects in filopodia distribution, PeMP motility, lamellar directed migration and phagocytosis in microglia. Thus, our work contributes to recent findings that mRNA localization and localized translation occur in microglia and gives a mechanistic insight into the relevance of this molecular mechanism in fundamental microglial functions in response to inflammation.
Selective inhibition of hsp90 paralogs: Uncovering the role of helix 1 in Grp94-selective ligand binding
Authors: Que Nanette L. S. , Seidler Paul M. , Aw Wen J. , Chiosis Gabriela , Gewirth Daniel T. 
Published: August 2023

AbstractGrp94 is the endoplasmic reticulum paralog of the hsp90 family of chaperones, which have been targeted for therapeutic intervention via their highly conserved ATP binding sites. The design of paralog-selective inhibitors relies on understanding the protein structural elements that drive higher affinity in selective inhibitors. Here, we determined the structures of Grp94 and Hsp90 in complex with the Grp94-selective inhibitor PU-H36, and of Grp94 with the non-selective inhibitor PU-H71. In Grp94, PU-H36 derives its higher affinity by utilizing Site 2, a Grp94-specific side pocket adjoining the ATP binding cavity, but in Hsp90 PU-H36 occupies Site 1, a side pocket that is accessible in all paralogs with which it makes lower affinity interactions. The structure of Grp94 in complex with PU-H71 shows only Site 1 binding. While changes in the conformation of helices 4 and 5 in the N-terminal domain occur when ligands bind to Site 1 of both Hsp90 and Grp94, large conformational shifts that also involve helix 1 are associated with the engagement of the Site 2 pocket in Grp94 only. Site 2 in Hsp90 is blocked and its helix 1 conformation is insensitive to ligand binding. To understand the role of helix 1 in ligand selectivity, we tested the binding of PU-H36 and other Grp94-selective ligands to chimeric Grp94/Hsp90 constructs. These studies show that helix 1 is the major determinant of selectivity for Site 2 targeted ligands, and also influences the rate of ATPase activity in Hsp90 paralogs.
Intra-host variation in the spike S1/S2 region of a feline coronavirus type-1 in a cat with persistent infection
Authors: Olarte-Castillo Ximena A. , Licitra Beth. N. , André Nicole M. , Sierra Maria A. , Mason Christopher E. , Goodman Laura B. , Whittaker Gary R. 
Published: August 2023

AbstractFeline coronavirus type 1 (FCoV-1) is widely known for causing feline infectious peritonitis (FIP), a systemic infection that is often fatal, with the virus known as the FIPV biotype. However, subclinical disease also occurs, in which cats may not show signs and intermittently shed the virus, including in feces, possibly for long periods of time. This virus is known as the FECV biotype. Progression of FECV to FIPV has been linked to several genomic changes, however a specific region of the viral spike protein at the interface of the spike S1 and S2 domains has been especially implicated. In this study, we followed a cat (#576) for six years from 2017, at which time FCoV-1 was detected in feces and conjunctival swabs, until 2022, when the animal was euthanized based on a diagnosis of alimentary small cell lymphoma. Over this time period, the cat was clinically diagnosed with inflammatory bowel disease and chronic rhinitis, and cardiac problems were also suspected. Using hybridization capture targeting the spike (S) gene of FCoV followed by next-generation sequencing, we screened 27 clinical samples. We detected FCoV-1 in 4 samples taken in 2017 (intestine and nasal tissue, feces, and conjunctiva), and 3 samples taken in 2022 (feces, and intestinal and heart tissue), but not in fecal samples taken in 2019 and 2020. Next, we focused on the S1/S2 region within S, which contains the furin cleavage site (FCS), a key regulator of viral transmission and pathogenesis. We show that the FCoV-1 variants obtained from feces in 2017 and 2022 were identical, while the ones from conjunctiva (2017), heart (2022), and intestine (2017 and 2022) were distinct. Sequence comparison of all the variants obtained showed that most of the non-synonymous changes in the S1/S2 region occur within the FCS. In the heart, we found two variants that differed by a single nucleotide, resulting in distinct FCS motifs that differ in one amino acid. It is predicted that one of these FCS motifs will down-regulate spike cleavability. The variant from the conjunctiva (2017) had a 6-nucleotide in-frame insertion that resulted in a longer and more exposed S1/S2 loop, which is predicted to be more accessible to the furin protease. Our studies indicate that FCoV-1 can independently persist in the gastrointestinal tract and heart of a cat over a long period of time without evidence of typical FIP signs, with intermittent viral shedding from the gastrointestinal and respiratory tracts.
IST1 regulates select recycling pathways
Authors: Clippinger Amy K. , Naismith Teresa V. , Yoo Wonjin , Jansen Silvia , Kast David J. , Hanson Phyllis I. 
Published: August 2023

AbstractESCRTs (Endosomal Sorting Complex Required for Transport) are a modular set of protein complexes with membrane remodeling activities that include the formation and release of intralumenal vesicles (ILVs) to generate multivesicular endosomes. While most of the 12 ESCRT-III proteins are known to play roles in ILV formation, IST1 has been associated with a wider range of endosomal remodeling events. Here, we extend previous studies of IST1 function in endosomal trafficking and confirm that IST1, along with its binding partner CHMP1B, contributes to scission of early endosomal carriers. Depleting IST1 impaired delivery of transferrin receptor from early/sorting endosomes to the endocytic recycling compartment and instead increased its rapid recycling to the plasma membrane via peripheral endosomes enriched in the clathrin adaptor AP-1. IST1 is also important for export of mannose 6-phosphate receptor from early/sorting endosomes. Examination of IST1 binding partners on endosomes revealed that IST1 interacts with the MIT domain-containing sorting nexin SNX15, a protein previously reported to regulate endosomal recycling. Our kinetic and spatial analyses establish that SNX15 and IST1 occupy a clathrin-containing subdomain on the endosomal perimeter distinct from those previously implicated in cargo retrieval or degradation. Using live-cell microscopy we see that SNX15 and CHMP1B alternately recruit IST1 to this subdomain or the base of endosomal tubules. These findings indicate that IST1 contributes to a subset of recycling pathways from the early/sorting endosome.Abstract Figure
MicroED as a powerful tool for structure determination of macrocyclic drug compounds directly from their powder formulations
Authors: Danelius E , Bu G , Wieske H , Gonen T 
Published: August 2023

AbstractMacrocycles are important drug leads with many advantages including the ability to target flat and featureless binding sites as well as act as molecular chameleons and thereby reach intracellular targets. However, due to their complex structures and inherent flexibility, macrocycles are difficult to study structurally and there are limited structural data available. Herein, we use the cryo-EM method MicroED to determine the novel atomic structures of several macrocycles which have previously resisted structural determination. We show that structures of similar complexity can now be obtained rapidly from nanograms of material, and that different conformations of flexible compounds can be derived from the same experiment. These results will have impact on contemporary drug discovery as well as natural product exploration.