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Osteoarthritis (OA) is a chronic and degenerative joint disease affecting more than 500 million patients worldwide with no disease-modifying treatment approved to date. Several publications report on the transforming growth factor β-activated kinase 1 (TAK1) as a potential molecular target for OA, with complementary anti-catabolic and anti-inflammatory effects. We report herein on the development of TAK1 inhibitors with physicochemical properties suitable for intra-articular injection, with the aim to achieve high drug concentration at the affected joint, while avoiding severe toxicity associated with systemic inhibition. More specifically, reducing solubility by increasing crystallinity, while maintaining moderate lipophilicity proved to be a good compromise to ensure high and sustained free drug exposures in the joint. Furthermore, structure-based design allowed for an improvement of selectivity versus interleukin-1 receptor-associated kinases 1 and 4 (IRAK1/4). Finally, TAK-756 was discovered as a potent TAK1 inhibitor with good selectivity versus IRAK1/4 as well as excellent intra-articular pharmacokinetic properties.

Mental health conditions constitute a major burden of disease for adolescents globally and can lead to significant adverse consequences. This systematic review aimed to identify if psychosocial interventions are effective in preventing mental health conditions in adolescents already experiencing emotional problems. We searched for randomized controlled trials comparing psychosocial interventions for preventing mental health conditions with care as usual in adolescents aged 10-19 who are experiencing symptoms of emotional problems. We searched PubMed/Medline, PsycINFO, ERIC, EMBASE, and ASSIA databases to identify studies. We found 82 eligible studies (n = 13,562 participants). Findings show that interventions can reduce mental health conditions and increase positive mental health. Across all reported time points, psychosocial interventions showed significant, small-to moderate-sized beneficial effects on preventing mental health conditions (SMD: -0.26, 95% CI [-0.42, -0.19] and small positive effects on positive mental health (SMD: 0.17, 95% CI [0.097, 0.29]. There were no statistically significant pooled findings suggesting that psychosocial interventions had either a positive or negative effect on self-harm or suicide; aggressive, disruptive and oppositional behavior; substance use; or school attendance. Despite the positive findings, a critical gap exists in the design of effective psychosocial interventions to reduce self-harm and suicide, and other risk behaviors in adolescents with symptoms of emotional problems., (Copyright © 2024 Society for Adolescent Health and Medicine. Published by Elsevier Inc. All rights reserved.)

TGFβ-signaling regulates cancer progression by controlling cell division, migration, and death. These outcomes are mediated by gene expression changes, but the mechanisms of decision-making toward specific fates remain unclear. Here, we combine SMAD transcription factor imaging, genome-wide RNA sequencing, and morphological assays to quantitatively link signaling, gene expression, and fate decisions in mammary epithelial cells. Fitting genome-wide kinetic models to our time-resolved data, we find that most of the TGFβ target genes can be explained as direct targets of SMAD transcription factors, whereas the remainder show signs of complex regulation, involving delayed regulation and strong amplification at high TGFβ doses. Knockdown experiments followed by global RNA sequencing revealed transcription factors interacting with SMADs in feedforward loops to control delayed and dose-discriminating target genes, thereby reinforcing the specific epithelial-to-mesenchymal transition at high TGFβ doses. We identified early repressors, preventing premature activation, and a late activator, boosting gene expression responses for a sufficiently strong TGFβ stimulus. Taken together, we present a global view of TGFβ-dependent gene regulation and describe specificity mechanisms reinforcing cellular decision-making., (© 2024 Hartmann et al.)

Background and Aims: Severe acute respiratory syndrome coronavirus (SARS-CoV-2) preferentially infects the respiratory tract; however, several studies have implicated a multi-organ involvement. Hepatic dysfunctions caused by SARS-CoV-2 infection have been increasingly recognized and described to correlate with disease severity. To elucidate molecular factors that could contribute towards hepatic infection, we concentrated on microRNAs (miRNAs), a class of small non-coding RNAs that modulate various cellular processes and which are reported to be differentially regulated during liver injury. We aimed to study the infection of primary human hepatocytes (PHH) with SARS-CoV-2 and to evaluate the potential of miRNAs for modulating viral infection., Methods: We analysed liver autopsies from a coronavirus disease 19 (COVID-19)-positive cohort for the presence of viral RNA using Nanopore sequencing. PHH were used for the infection with SARS-CoV-2. The candidate miRNAs targeting angiotensin converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) were identified using in silico approaches. To discover the potential regulatory mechanism, transfection experiments, qRT-PCRs, western blots and luciferase reporter assays were performed., Results: We could detect SARS-CoV-2 RNA in COVID-19-positive liver autopsies. We show that PHH express ACE2 and TMPRSS2 and can be readily infected with SARS-CoV-2, resulting in robust replication. Transfection of selected miRNA mimics reduced SARS-CoV-2 receptor expression and SARS-CoV-2 burden in PHH. In silico and biochemical analyses supported a potential direct binding of miR-141-3p to the SARS-CoV-2 genome., Conclusion: We confirm that PHH are susceptible to SARS-CoV-2 infection and demonstrate selected miRNAs targeting SARS-CoV-2 entry factors and/or the viral genome reduce viral loads. These data provide novel insights into hepatic susceptibility to SARS-CoV-2 and associated dysfunctions in COVID-19., (© 2024 The Author(s). Liver International published by John Wiley & Sons Ltd.)

Glycan-protein interactions play a key role in various biological processes from fertilization to infections. Many of these interactions take place at the glycocalyx-a heavily glycosylated layer at the cell surface. Despite its significance, studying the glycocalyx remains challenging due to its complex, dynamic, and heterogeneous nature. This study introduces a glycocalyx model allowing for the first time to control spatial organization and heterogeneity of the glycan moieties. Glycan-mimetics with lipid-moieties that partition into either liquid-ordered (Lo, lipid rafts) or liquid-disordered (Ld) phases of giant unilamellar vesicles (GUVs), which serve as simplified cell membrane models mimicking lipid rafts, are developed. This phase-specific allocation allows controlled placement of glycan motifs in distinct membrane environments, creating heteromultivalent systems that replicate the natural glycocalyx's complexity. We show that phase localization of glycan mimetics significantly influences recruitment of protein receptors to the membrane. Glycan-conjugates in the ordered phase demonstrate enhanced lectin binding, supporting the idea that raft-like domains facilitate stronger receptor interactions. This study provides a platform for systematically investigating spatial and dynamic presentation of glycans in biological systems and presents the first experimental evidence that glycan accumulation in lipid rafts enhances receptor binding affinity, offering deeper insights into the glycocalyx's functional mechanisms., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

This work presents a versatile strategy for the synthesis of dual stimuli-responsive amphiphilic glycomacromolecules with tailored release properties. Amphiphilic precision glycomacromolecules (APGs) derived from tailor-made building blocks using solid phase polymer synthesis form glycofunctionalized micelles, a versatile class of materials with applications in drug delivery, as antiinfection agents as well as simple cell mimetics. In this work, this concept is extended by integrating cleavable building blocks into APGs now allowing stimuli-responsive release of glycan ligands or destruction of the micelles. This study incorporates a newly designed acid-labile building block, 4-(4-(((((9H-fluoren-9-yl)methoxy)carbonyl)amino)methyl)-1,3-dioxolan-2-yl)benzoic acid (DBA), suitable also for other types of solid phase or amide chemistry, and an established UV-cleavable 2-nitrobenzyl linker (PL). The results demonstrate that both linkers can be cleaved independently and thus allow dual stimuli-responsive release from the APG micelles. By choosing the APG design e.g., placing the cleavable linkers between glycomacromolecular blocks presenting different types of carbohydrates, they can tune APG and micellar stability as well as the interaction and cluster formation with a carbohydrate-recognizing lectin. Such dual-responsive glycofunctionalized micelles have wide potential for use in drug delivery applications or for the development as anti-adhesion agents in antiviral and antibacterial treatments., (© 2024 The Author(s). Macromolecular Rapid Communications published by Wiley‐VCH GmbH.)

Protein-polymer conjugates combine properties of biopolymers and synthetic polymers, such as specific bioactivity and increased stability, with great benefits for various applications from catalysis to biomedicine. Furthermore, polymer conjugation can mimic important posttranslational modifications of proteins such as glycosylation. There are typically two approaches to create protein-polymer conjugates: the protein is functionalized in advance with an initiator for a grafting-from method or a previously produced polymer is conjugated to the protein via a grafting-to method. In this study, we present a new approach that uses native proteins and allows for direct grafting-from using a thiol-induced, light-activated controlled radical polymerization (TIRP) that is initiated at thiols from specific cysteine residues of the protein. This straightforward method is employed to introduce polymers onto proteins and enzymes without any prior protein modifications, it works in aqueous buffer and maintains the protein's native structure and activity. The resulting protein-polymer conjugates exhibit high molar masses and low dispersities. We demonstrate the versatility of this approach by introducing different types of polymers such as hydrophilic poly(2-hydroxyethyl acrylate) (pHEAA), temperature-responsive poly( N -isopropylacrylamide) (pNIPAM) as well as glycopolymers mimicking the natural protein glycosylation and enabling selective interactions. We present successful combinations of the protein and polymer functions e.g. , temperature-induced aggregation leading to an increase in enzyme activity and the introduction of artificial glycosylation inducing specific protein-protein cluster formation and giving straightforward access to glycosurfaces. Based on this straightforward, potentially scalable yet highly controlled synthesis of protein-polymer conjugates, various areas of applications are envisioned ranging from biomedicine to material sciences., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Plant peptides communicate by binding to a large family of receptor-like kinases (RLKs), and they share a conserved binding mechanism, which may account for their promiscuous interaction with several RLKs. In order to understand the in vivo binding specificity of the CLAVATA3/EMBRYO SURROUNDING REGION-RELATED peptide family in Arabidopsis, we have developed a novel set of CLAVATA3 (CLV3)-based peptide tools. After carefully evaluating the CLE peptide binding characteristics, using solid phase synthesis process, we modified the CLV3 peptide and attached a fluorophore and a photoactivable side group. We observed that the labeled CLV3 shows binding specificity within the CLAVATA1 clade of RLKs while avoiding the distantly related PEP RECEPTOR clade, thus resolving the contradictory results obtained previously by many in vitro methods. Furthermore, we observed that the RLK-bound CLV3 undergoes clathrin-mediated endocytosis and is trafficked to the vacuole via ARA7 (a Rab GTPase)-labeled endosomes. Additionally, modifying CLV3 for light-controlled activation enabled spatial and temporal control over CLE signaling. Hence, our CLV3 macromolecular toolbox can be used to study rapid cell specific down-stream effects. Given the conserved binding properties, in the future our toolbox can also be used as a template to modify other CLE peptides., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

The glycocalyx, a complex carbohydrate layer on cell surfaces, plays a crucial role in various biological processes. Understanding native glycocalyces' complexity is challenging due to their intricate and dynamic nature. Simplified mimics of native glycocalyces offer insights into glycocalyx functions but often lack molecular precision and fail to replicate key features of the natural analogues like molecular crowding and heteromultivalency. We introduce membrane-anchoring precision glycomacromolecules synthesized via solid-phase polymer synthesis (SPPoS) and thiol-induced, light-activated controlled radical polymerization (TIRP), enabling the construction of crowded and heteromultivalent glycocalyx mimetics with varying molecular weights and densities in giant unilamellar vesicles (GUVs). The incorporation and dynamics of glycomacromolecules in the GUVs are examined via microscopy and fluorescence correlation spectroscopy (FCS) and studies on lectin-carbohydrate-mediated adhesion of GUVs reveal inhibitory and promotional adhesion effects corresponding to different glycocalyx mimetic compositions, bridging the gap between synthetic models and native analogues.

This study employed a cross-sectional, qualitative individual interview methodology to explore South African women with physical disabilities' experiences of intimate partner and sexual violence, inclusive of non-consensual and coerced sexual intercourse. For the participants, disability was a factor that intersected with gender norms to create vulnerability to abuse, and that patriarchal ideologies constructing how women should perform their gendered roles in marriage or sexual partnerships, as well as disability stigma, exacerbated this vulnerability. It is important to develop understandings of the different risk factors for violence - at the individual level and in the context of dyadic relationships - to develop programming to better support women., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Monitoring health conditions in neonates for early therapeutic intervention in case deviations from physiological conditions is crucial for their long-term development. Due to their immaturity preterm born neonates are dependent on particularly careful physical and neurological diagnostic methods. Ideally, these should be noninvasive, noncontact, and radiation free. Infrared spectroscopy was used to analyze exhaled breath from 71 neonates with a special emphasis on preterm infants, as a noninvasive, noncontact, and radiation-free diagnostic tool. Passive sample collection was performed by skilled clinicians. Depending on the mode of respiratory support of infants, four different sampling procedures were adapted to collect exhaled breath. With the aid of appropriate reference samples, infrared spectroscopy has successfully demonstrated its effectiveness in the analysis of breath samples of neonates. The discernible increase in concentrations of carbon dioxide, carbon monoxide, and methane in collected samples compared to reference samples served as compelling evidence of the presence of exhaled breath. With regard to technical hurdles and sample analysis, samples collected from neonates without respiratory support proved to be more advantageous compared to those obtained from intubated infants and those with CPAP (continuous positive airway pressure). The main obstacle lies in the significant dilution of exhaled breath in the case of neonates receiving respiratory support. Metabolic analysis of breath samples holds promise for the development of noninvasive biomarker-based diagnostics for both preterm and sick neonates provided an adequate amount of breath is collected., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

Ex vivo expansion of human CD34+ hematopoietic stem and progenitor cells remains a challenge due to rapid differentiation after detachment from the bone marrow niche. In this study, we assessed the capacity of an inducible fusion protein to enable sustained ex vivo proliferation of hematopoietic precursors and their capacity to differentiate into functional phagocytes. We fused the coding sequences of an FK506-Binding Protein 12 (FKBP12)-derived destabilization domain (DD) to the myeloid/lymphoid lineage leukemia/eleven nineteen leukemia (MLL-ENL) fusion gene to generate the fusion protein DD-MLL-ENL and retrovirally expressed the protein switch in human CD34+ progenitors. Using Shield1, a chemical inhibitor of DD fusion protein degradation, we established large-scale and long-term expansion of late monocytic precursors. Upon Shield1 removal, the cells lost self-renewal capacity and spontaneously differentiated, even after 2.5 y of continuous ex vivo expansion. In the absence of Shield1, stimulation with IFN-γ, LPS, and GM-CSF triggered terminal differentiation. Gene expression analysis of the obtained phagocytes revealed marked similarity with naïve monocytes. In functional assays, the novel phagocytes migrated toward CCL2, attached to VCAM-1 under shear stress, produced reactive oxygen species, and engulfed bacterial particles, cellular particles, and apoptotic cells. Finally, we demonstrated Fcγ receptor recognition and phagocytosis of opsonized lymphoma cells in an antibody-dependent manner. Overall, we have established an engineered protein that, as a single factor, is useful for large-scale ex vivo production of human phagocytes. Such adjustable proteins have the potential to be applied as molecular tools to produce functional immune cells for experimental cell-based approaches., Competing Interests: Competing interests statement:The authors declare no competing interest.

Escherichia coli and other bacteria use adhesion receptors, such as FimH, to attach to carbohydrates on the cell surface as the first step of colonization and infection. Efficient inhibitors that block these interactions for infection treatment are multivalent carbohydrate-functionalized scaffolds. However, these multivalent systems often lead to the formation of large clusters of bacteria, which may pose problems for clearing bacteria from the infected site. Here, we present Man-containing Janus particles (JPs) decorated on one side with glycomacromolecules to target Man-specific adhesion receptors of E. coli . On the other side, poly( N -isopropylacrylamide) is attached to the particle hemisphere, providing temperature-dependent sterical shielding against binding and cluster formation. While homogeneously functionalized particles cluster with multiple bacteria to form large aggregates, glycofunctionalized JPs are able to form aggregates only with individual bacteria. The formation of large aggregates from the JP-decorated single bacteria can still be induced in a second step by increasing the temperature and making use of the collapse of the PNIPAM hemisphere. This is the first time that carbohydrate-functionalized JPs have been derived and used as inhibitors of bacterial adhesion. Furthermore, the developed JPs offer well-controlled single bacterial inhibition in combination with cluster formation upon an external stimulus, which is not achievable with conventional carbohydrate-functionalized particles.

A cationic, dendrimer-like oligo(aminoamide) carrier with four-arm topology based on succinoyl tetraethylene pentamine and histidines, cysteines, and N -terminal azido-lysines was screened for plasmid DNA delivery on various cell lines. The incorporated azides allow modification with various shielding agents of different polyethylene glycol (PEG) lengths and/or different ligands by copper-free click reaction, either before or after polyplex formation. Prefunctionalization was found to be advantageous over postfunctionalization in terms of nanoparticle formation, stability, and efficacy. A length of 24 ethylene oxide repetition units and prefunctionalization of ≥50% of azides per carrier promoted optimal polyplex shielding. PEG shielding resulted in drastically reduced DNA transfer, which could be successfully restored by active lectin targeting via novel GalNAc or mannose ligands, enabling enhanced receptor-mediated endocytosis of the carrier system. The involvement of the asialoglycoprotein receptor (ASGPR) in the uptake of GalNAc-functionalized polyplexes was confirmed in the ASGPR-positive hepatocarcinoma cell lines HepG2 and Huh7. Mannose-modified polyplexes showed superior cellular uptake and transfection efficacy compared to unmodified and shielded polyplexes in mannose-receptor-expressing dendritic cell-like DC2.4 cells.

An advanced design of the analytical ultracentrifuge with multiwavelength emission detection (MWE-AUC) is presented which offers outstanding performance concerning the spectral resolution and range flexibility as well as the quality of the data acquired. The excitation by a 520 nm laser is complemented with a 405 nm laser. An external spectrograph with three switchable tunable gratings permits optimisation of the spectral resolution in an order of magnitude range while keeping the spectral region broad. The new system design leads also to a significant reduction of systematic signal noise and allows the assessment and control of inner filter effects. Details regarding the very large signal dynamic range are presented, an important aspect when studying samples in a broad concentration range of up to five orders of magnitude. Our system is validated by complementary studies on two biological systems, fluorescent BSA and GFP, using the commercial Optima AUC with absorbance detection for comparison. Finally, we demonstrate the capabilities of our second generation MWE-AUC with respect to multiwavelength characterisation of gold nanoclusters, which exhibit specific fluorescence depending on their structure. Overall, this work depicts an important stepping stone for the concept of multiwavelength emission detection in AUC. The MWE-AUC developed, being to our knowledge the first and sole one of its kind, has reached the development level suitable for the future in-depth studies of size-, shape- and composition-dependent emission properties of colloids., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Despite promising preclinical and earlier clinical data, a recent phase III trial on the anti-β7 integrin antibody etrolizumab in Crohn's disease (CD) did not reach its primary endpoint. The mechanisms leading to this outcome are not well understood. Here we characterize the β7 + T cell compartment from patients with CD in comparison to cells from individuals without inflammatory bowel disease. By flow cytometric, transcriptomic and functional profiling of circulating T cells, we find that triple-integrin-expressing (α4 + β7 + β1 hi ) T cells have the potential to home to the gut despite α4β7 blockade and have a specific cytotoxic signature. A subset of triple-integrin-expressing cells readily acquires αE expression and could be co-stimulated via E-Cadherin-αEβ7 interactions in vitro. Etrolizumab-s fails to block such αEβ7 signalling at high levels of T cell stimulation. Consistently, in CD patients treated with etrolizumab, T cell activation correlates with cytotoxic signatures. Collectively, our findings might add one important piece to the puzzle to explain phase III trial results with etrolizumab, while they also highlight that αEβ7 remains an interesting target for future therapeutic approaches in inflammatory bowel disease., (© 2024. The Author(s).)

Objective: Digital interventions hold important potential for supporting parents when face-to-face interventions are unavailable. We assessed the feasibility and effectiveness of a digital parenting intervention in Zambia and Tanzania., Methods: Using a randomised controlled trial, we evaluated the Sharing Stories digital parenting intervention for caregivers of children aged 9-32 months with access to a smartphone in their household. Caregivers were stratified based on child age and randomly assigned to the intervention or waitlist control arm. The intervention was delivered via facilitated WhatsApp groups over 6 weeks to promote caregiver wellbeing and responsive caregiving through shared reading activities. Primary outcomes were caregiver-reported responsive caregiving, child language and socio-emotional development. Secondary outcomes were caregiver mental health and parental stress. Masked assessors conducted assessments at baseline and immediate follow-up., Results: Between October 2020 and March 2021, we randomly assigned 494 caregiver-child dyads to the intervention ( n  = 248) or waitlist control ( n  = 246) arm. Caregivers in the intervention group reported more responsive caregiving (OR = 2.55, 95% CI: 1.15-5.66, p  = 0.02), time reading or looking at books (β = 0.45, p  = 0.04) and telling stories (β = 0.72, p  = 0.002). Intervention caregivers reported significantly lower symptoms of depression (β = -0.64, p  = 0.05) and anxiety (β = -0.65, p  = 0.02). Child development and parental stress did not differ significantly between groups., Conclusions: Digital parenting interventions using WhatsApp can effectively promote responsive caregiving and caregiver mental health in low-resource settings, with great potential for scalability., Trial Registration: ISRCTN database, ISRCTN77689525., Competing Interests: The authors declared no potential conflicts of interest with respect to the research, authorship and/or publication of this article., (© The Author(s) 2023.)

The straightforward synthesis of polyamide-derived statistical copolymers with catechol, amine, amide and hydroxy residues via free radical polymerization is presented. In particular, catechol, amine and amide residues are present in natural mussel foot proteins, enabling strong underwater adhesion due to synergistic effects where cationic residues displace hydration and ion layers, followed by strong short-rang hydrogen bonding between the catechol or primary amides and SiO 2 surfaces. The present study is aimed at investigating whether such synergistic effects also exist for statistical copolymer systems that lack the sequence-defined positioning of functional groups in mussel foot proteins. A series of copolymers is established and the adsorption in saline solutions on SiO 2 is determined by quartz crystal microbalance measurements and ellipsometry. These studies confirm a synergy between cationic amine groups with catechol units and primary amide groups via an increased adsorptivity and increased polymer layer thicknesses. Therefore, the free radical polymerization of catechol, amine and amide monomers as shown here may lead to simplified mussel-inspired adhesives that can be prepared with the readily scalable methods required for large-scale applications.

Survivin, a well-known member of the inhibitor of apoptosis protein family, is upregulated in many cancer cells, which is associated with resistance to chemotherapy. To circumvent this, inhibitors are currently being developed to interfere with the nuclear export of survivin by targeting its protein-protein interaction (PPI) with the export receptor CRM1. Here, we combine for the first time a supramolecular tweezer motif, sequence-defined macromolecular scaffolds, and ultrasmall Au nanoparticles (us-AuNPs) to tailor a high avidity inhibitor targeting the survivin-CRM1 interaction. A series of biophysical and biochemical experiments, including surface plasmon resonance measurements and their multivalent evaluation by EVILFIT, reveal that for divalent macromolecular constructs with increasing linker distance, the longest linkers show superior affinity, slower dissociation, as well as more efficient PPI inhibition. As a drawback, these macromolecular tweezer conjugates do not enter cells, a critical feature for potential applications. The problem is solved by immobilizing the tweezer conjugates onto us-AuNPs, which enables efficient transport into HeLa cells. On the nanoparticles, the tweezer valency rises from 2 to 16 and produces a 100-fold avidity increase. The hierarchical combination of different scaffolds and controlled multivalent presentation of supramolecular binders was the key to the development of highly efficient survivin-CRM1 competitors. This concept may also be useful for other PPIs.

Graft-versus-host disease (GVHD) is a major risk of the administration of allogeneic chimeric antigen receptor (CAR)-redirected T cells to patients who are HLA unmatched. Gene editing can be used to disrupt potentially alloreactive T-cell receptors (TCRs) in CAR T cells and reduce the risk of GVHD. Despite the high knockout rates achieved with the optimized methods, a subsequent purification step is necessary to obtain a safe allogeneic product. To date, magnetic cell separation (MACS) has been the gold standard for purifying TCRα/β- CAR T cells, but product purity can still be insufficient to prevent GVHD. We developed a novel and highly efficient approach to eliminate residual TCR/CD3+ T cells after TCRα constant (TRAC) gene editing by adding a genetically modified CD3-specific CAR NK-92 cell line during ex vivo expansion. Two consecutive cocultures with irradiated, short-lived, CAR NK-92 cells allowed for the production of TCR- CAR T cells with <0.01% TCR+ T cells, marking a 45-fold reduction of TCR+ cells compared with MACS purification. Through an NK-92 cell-mediated feeder effect and circumventing MACS-associated cell loss, our approach increased the total TCR- CAR T-cell yield approximately threefold while retaining cytotoxic activity and a favorable T-cell phenotype. Scaling in a semiclosed G-Rex bioreactor device provides a proof-of-principle for large-batch manufacturing, allowing for an improved cost-per-dose ratio. Overall, this cell-mediated purification method has the potential to advance the production process of safe off-the-shelf CAR T cells for clinical applications., (© 2023 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Background: Latinx youth are a population of concern, at elevated risk for chronic diseases and with poor adherence to dietary recommendations., Objectives: To examine Latinx seventh-grade students' perceptions of the factors that influence their diet and eating behaviors., Design: This qualitative research used focus groups and an inductive content analysis approach., Participants/setting: Five sex-stratified focus groups (three groups with females) with 35 primarily Latinx seventh-grade students were conducted at two local Title 1 public middle schools in a large metropolitan area of the Southwestern United States., Main Outcome Measures: The discussion protocol included questions about participants' food choices, the role of their parents in their diet, and healthy body-related concerns among their peers., Analyses: Verbatim transcripts were coded in NVivo 12 on the basis of specificity, extensiveness, and frequency. Themes emerged from group dialogue, detailed conversations, and predominant topics of discussion, and aligned with ecological systems theory., Results: Participants referred to factors influencing Latinx seventh-grade students' eating behaviors at the individual, family, household, and school levels. At the individual level, participants described their eating as unhealthy and perceived it as determined by taste, convenience, ease of preparation, and home availability. Participants expressed concerns about diabetes because of their body weight and family history, and identified those concerns as reasons for acceptance of healthy foods and the desire for parents to model healthy eating behaviors. Family-level factors perceived as influencing dietary behaviors included the role of parents as providers of food and models of unhealthy eating, budget constraints, and availability (or lack thereof) of healthy foods at home. Similarly, the identified school-level factors aligned with availability and quality of foods in that environment., Conclusions: Family- and household-related factors emerged as important influences on seventh-grade students' dietary behaviors. Future diet interventions should incorporate strategies targeting these multiple-level factors that influence dietary intake for Latinx youth and that address the concerns related to disease risk., (Copyright © 2023 Academy of Nutrition and Dietetics. Published by Elsevier Inc. All rights reserved.)

This study presents the preparation and phase behavior of glycan-functionalized polyelectrolytes for capturing carbohydrate-binding proteins and bacteria in liquid condensate droplets. The droplets are formed by complex coacervation of poly(active ester)-derived polyanions and polycations. This approach allows for a straightforward modular introduction of charged motifs and specifically interacting units; mannose and galactose oligomers are used here as first examples. The introduction of carbohydrates has a notable effect on the phase separation and the critical salt concentration, potentially by reducing the charge density. Two mannose binding species, concanavalin A (ConA) and Escherichia coli , are shown to not only specifically bind to mannose-functionalized coacervates but also to some degree to unfunctionalized, carbohydrate-free coacervates. This suggests non-carbohydrate-specific charge-charge interactions between the protein/bacteria and the droplets. However, when mannose interactions are inhibited or when non-binding galactose-functionalized polymers are used, interactions are significantly weakened. This confirms specific mannose-mediated binding functionalization and suggests that introducing carbohydrates reduces non-specific charge-charge interactions by a so far unidentified mechanism. Overall, the presented route toward glycan-presenting polyelectrolytes enables new functional liquid condensate droplets with specific biomolecular interactions.

Iron (Fe) is a required micronutrient in plants for the production of chlorophyll and transport of oxygen. A commonly used surrogate for measuring nutrient levels is the measurement of electrical conductivity or total dissolved solids, but this technique is not selective towards any particular dissolved ion. In this study, using a conventional microwave, fluorescent carbon dots (CDs) are produced from glucose and a household cleaning product and applied towards monitoring dissolved ferric iron levels in hydroponic systems through fluorescent quenching. The produced particles have an average size of 3.19 ± 0.76 nm with a relatively high degree of oxygen surface groups. When using an excitation of 405 nm, a broad emission peak is centered at approximately 500 nm. A limit-of-detection of 0.196 ± 0.067 ppm (3.51 ± 1.21 μM) with minimal interference from common heavy metal quenchers and ions found in hydroponic systems was determined. Butterhead lettuce was grown while discretely monitoring iron levels via the CDs for three separate weeks of growth. The CDs displayed a non-significant difference ( p > 0.05) in performance when compared to a standard method. These results along with a simple and relatively low-cost production method make the CDs in this study a promising tool for monitoring iron levels in hydroponic systems., Competing Interests: The authors declare no competing financial interest., (This journal is © The Royal Society of Chemistry.)

Lectin-glycan interactions are at the heart of a multitude of biological events. Glycans are usually presented in a multivalent manner on the cell surface as part of the so-called glycocalyx, where they interact with other entities. This multivalent presentation allows us to overcome the typical low affinities found for individual glycan-lectin interactions. Indeed, the presentation of glycans may drastically impact their binding by lectins, highly affecting the corresponding binding affinity and even selectivity. In this context, we herein present the study of the interaction of a variety of homo- and heteromultivalent lactose-functionalized glycomacromolecules and their lipid conjugates with two human galectins. We have employed as ligands the glycomacromolecules, as well as liposomes decorated with those structures, to evaluate their interactions in a cell-mimicking environment. Key details of the interaction have been unravelled by NMR experiments, both from the ligand and receptor perspectives, complemented by cryo-electron microscopy methods and molecular dynamics simulations., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

Organisms require micronutrients, and Arabidopsis (Arabidopsis thaliana) IRON-REGULATED TRANSPORTER1 (IRT1) is essential for iron (Fe2+) acquisition into root cells. Uptake of reactive Fe2+ exposes cells to the risk of membrane lipid peroxidation. Surprisingly little is known about how this is avoided. IRT1 activity is controlled by an intracellular variable region (IRT1vr) that acts as a regulatory protein interaction platform. Here, we describe that IRT1vr interacted with peripheral plasma membrane SEC14-Golgi dynamics (SEC14-GOLD) protein PATELLIN2 (PATL2). SEC14 proteins bind lipophilic substrates and transport or present them at the membrane. To date, no direct roles have been attributed to SEC14 proteins in Fe import. PATL2 affected root Fe acquisition responses, interacted with ROS response proteins in roots, and alleviated root lipid peroxidation. PATL2 had high affinity in vitro for the major lipophilic antioxidant vitamin E compound α-tocopherol. Molecular dynamics simulations provided insight into energetic constraints and the orientation and stability of the PATL2-ligand interaction in atomic detail. Hence, this work highlights a compelling mechanism connecting vitamin E with root metal ion transport at the plasma membrane with the participation of an IRT1-interacting and α-tocopherol-binding SEC14 protein., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2022. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Pathogens including viruses, bacteria, fungi, and parasites continue to shape our lives in profound ways every day. As we have learned to live in parallel with pathogens, we have gained a better understanding of the rules of engagement for how they bind, adhere, and invade host cells. One such mechanism involves the exploitation of host cell surface glycans for attachment/adhesion, one of the first steps of infection. This knowledge has led to the development of glycan-based diagnostics and therapeutics for the treatment and prevention of infection. One class of compounds that has become increasingly important are the glycopolymers. Glycopolymers are macromolecules composed of a synthetic scaffold presenting carbohydrates as side chain motifs. Glycopolymers are particularly attractive because their properties can be tuned by careful choice of the scaffold, carbohydrate/glycan, and overall presentation. In this review, we highlight studies over the past ten years that have examined the role of glycopolymers in pathogen adhesion and host cell infection, biofilm formation and removal, and drug delivery with the aim of examining the direct effects of these macromolecules on pathogen engagement. In addition, we also examine the role of glycopolymers as diagnostics for the detection and monitoring of pathogens.

Background: Myelodysplastic syndromes (MDS) are a heterogenous collection of clonal bone marrow diseases characterized by cytopenias, abnormal karyotypes, molecular abnormalities, and dysplasia by flow cytometry and/or morphology. The progression of MDS to severe cytopenias and/or overt leukemia is associated with the accumulation of additional cytogenetic abnormalities, suggesting clonal evolution. The impact of these accumulated abnormalities on myeloid maturation and the severity of the disease is poorly understood., Methods: Bone marrow specimens from 16 patients with cytogenetic abnormalities were flow cytometrically sorted into three myeloid populations: progenitors, immature myeloid cells, and mature myeloid cells. Fluorescence in situ hybridization analysis was performed on each to determine the distribution of chromosomal abnormalities during myeloid maturation., Results: Our findings revealed three distinct distributions of cytogenetic abnormalities across myeloid maturation, each of which corresponded to specific cytogenetic abnormalities. Group 1 had continuous distribution across all maturational stages and contained patients with a single cytogenetic aberration associated with good-to-intermediate prognosis; Group 2 had accumulation of abnormalities in immature cells and contained patients with high-risk monosomy 7; and Group 3 had abnormalities defining the founding clone equally distributed across maturational stages while subclonal abnormalities were enriched in progenitor cells and contained patients with multiple, non-monosomy 7, abnormalities with evidence of clonal evolution., Conclusions: Our findings demonstrate that low-risk abnormalities (e.g., del(20q) and trisomy 8) occurring in the founding clone display a markedly different disease etiology, with respect to myeloid maturation, than monosomy 7 or abnormalities acquired in subclones, which result in a disruption of myeloid cell maturation in MDS., (© 2021 International Clinical Cytometry Society.)

Cases of myocarditis, diagnosed clinically by laboratory tests and imaging have been described in the context of mRNA-based anti-SARS-CoV-2 vaccination. Autopsy-based description of detailed histological features of vaccine-induced myocarditis is lacking. We describe the autopsy findings and common characteristics of myocarditis in untreated persons who received anti-SARS-CoV-2 vaccination. Standardized autopsies were performed on 25 persons who had died unexpectedly and within 20 days after anti-SARS-CoV-2 vaccination. In four patients who received a mRNA vaccination, we identified acute (epi-)myocarditis without detection of another significant disease or health constellation that may have caused an unexpected death. Histology showed patchy interstitial myocardial T-lymphocytic infiltration, predominantly of the CD4 positive subset, associated with mild myocyte damage. Overall, autopsy findings indicated death due to acute arrhythmogenic cardiac failure. Thus, myocarditis can be a potentially lethal complication following mRNA-based anti-SARS-CoV-2 vaccination. Our findings may aid in adequately diagnosing unclear cases after vaccination and in establishing a timely diagnosis in vivo, thus, providing the framework for adequate monitoring and early treatment of severe clinical cases., (© 2022. The Author(s).)

Recently we have uncovered a non-enzymatic multi-step cycle for the regeneration of ergothioneine (ET), after reaction with noxious singlet oxygen ( 1 O 2 ), by glutathione (GSH). When living cells were loaded with ET labeled with deuterium and N-15 atoms (D5-ET) and exposed to light in the presence of a photosensitizer, no loss of deuterium at position 5 of the imidazole ring was observed, in contradiction to our previous mechanistic proposal. Therefore, it was necessary to reexamine the in vitro products of ET and 1 O 2 by liquid chromatography coupled to high resolution mass spectrometry. Pure 1 O 2 was generated by thermolysis at 37 °C of the endoperoxide DHPNO 2 . The use of D5-ET enabled us to revise and extend the reaction scheme. On the main pathway, 1 O 2 attacks the imidazole ring, and the hydroperoxide intermediates are reduced rapidly by ET or GSH via different mechanisms. The intramolecular water elimination from the 5-hydroperoxide described previously is slower and not a part of the cycle. On another side path, 1 O 2 attacks the sulfur of ET to form a sulfine (S-oxide). The reduction of the sulfine also allows for the complete regeneration of ET. Experiments with methanol instead of water as solvent revealed that, in the absence of GSH, ET was attacked 6 times more frequently at the ring than at the sulfur. In the presence of 1 mM GSH or higher, both side paths were abandoned. ET efficiently captures 1 O 2 with its ring and can then be regenerated to a large extent by GSH, without enzyme involvement., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The new oxygen-bridged geminal Si/P Frustrated Lewis Pair (FLP) tBu 2 P-O-Si(C 2 F 5 ) 3 (2) is able to reversibly bind carbon dioxide at ambient temperature. We compared its reactivity towards benzil, but-3-en-2-one, nitriles and phenylacetylene to that of the Al/P FLP tBu 2 P-O-AlBis 2 (Bis=-CH(SiMe 3 ) 2 ) (1). When reacted with benzil, both, 1 and 2, form the 1,2-addition product, but in the Si/P FLP 2, the second carbonyl function additionally binds to the silicon atom. With but-3-en-2-one 2 forms the 1,2-addition product, while 1 binds in 1,4-position. The reaction with acetonitrile yielded an unexpected etheneimine adduct for both systems, while only 1 reacted with tert-butylnitrile. With benzonitrile and acrylonitrile, 2 showed reversible addition to the C≡N bond and 1 forms a stable adduct with benzonitrile. Solely 1 shows reactivity towards phenylacetylene affording a mixture of the CH deprotonation adduct tBu 2 P(H)-O-AlBis 2 (CCPh) and the FLP -C≡C 1,2-addition adduct under ring formation. All compounds were characterized by multinuclear NMR spectroscopy, XRD and elemental analysis., (© 2022 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

Artisanal and small-scale gold mining (ASGM) is the largest anthropogenic source of mercury emissions globally. Concern over mercury pollution increases due to its long-term impacts on human health and aquatic and terrestrial ecosystems. Using a participatory research methodology, we gathered social and behavioral information regarding daily practices and water usage by an ASGM community in Suárez, Colombia. Based on this information, we identified 18 sampling sites of water sources commonly used by the community. The samples were analyzed for total mercury, total coliforms, pH, electrical conductivity, and total dissolved oxygen. Physicochemical and microbiological parameters from the water assessment were compared with the drinking water thresholds set by the Colombian regulatory agencies, the EPA, and the WHO. Our results showed that the majority of the samples do not meet one or more quality and safety standards. On average, the sampling sites showed total mercury levels below the regulatory limits; however, the data had considerable variability, and in many cases, individual observations fell above the maximum concentration limit for drinking water. We discuss these results within the larger framework of the regulatory gaps for human and environmental protection in ASGM contexts. The total lack of water, sanitation, and hygiene infrastructure, combined with the long-term consumption of sublethal doses of mercury and other water contaminants, constitutes a significant threat to the well-being of communities and territories that necessitates further research and intervention by institutional authorities., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Glycoconjugates are a versatile class of bioactive molecules that have found application as vaccines and antivirals and in cancer therapy. Their synthesis typically involves elaborate functionalization and use of protecting groups on the carbohydrate component in order to ensure efficient and selective conjugation. Alternatively, non-functionalized, non-protected carbohydrates isolated from biological sources or derived through biotechnological methods can be directly conjugated via N -methyloxyamine groups. In this study, we introduce such N -methyloxyamine groups into a variety of multivalent scaffolds─from small to oligomeric to polymeric scaffolds─making use of solid-phase polymer synthesis to assemble monodisperse sequence-defined macromolecules. These scaffolds are then successfully functionalized with different types of human milk oligosaccharides deriving a library of homo- and heteromultivalent glycoconjugates. Glycomacromolecules presenting oligosaccharide side chains with either α2,3- or α2,6-linked terminal sialic acid are used in a binding study with two types of polyomavirus capsid proteins showing that the multivalent presentation through the N -methyloxyamine-derived scaffolds increases the number of contacts with the protein. Overall, a straightforward route to derive glycoconjugates from complex oligosaccharides with high variability yet control in the multivalent scaffold is presented, and applicability of the derived structures is demonstrated.

Radiotherapy can act as an in situ vaccine, activating preventive tumor-specific immune responses in patients. Although carbon ion radiotherapy has superior biophysical properties over conventional photon irradiation, the immunological effects induced by this radiation type are poorly understood. Multiple strategies combining radiotherapy with immune checkpoint inhibition (radioimmunotherapy) to enhance antitumor immunity have been described; however, immune cell composition in tumors following radioimmunotherapy with carbon ions remains poorly explored. We developed a bilateral tumor model based on time-shifted subcutaneous injection of murine Her2+ EO771 tumor cells into immune-competent mice followed by selective irradiation of the primary tumor. αCTLA4-, but not αPD-L1-based radioimmunotherapy, induced complete tumor rejection and mediated the eradication of even non-irradiated, distant tumors. Cured mice were protected against the EO771 rechallenge, indicating long-lasting, tumor-specific immunological memory. Single-cell RNA sequencing and flow cytometric analyses of irradiated tumors revealed activation of NK cells and distinct tumor-associated macrophage clusters with upregulated expression of TNF and IL1 responsive genes. Distant tumors in the irradiated mice showed higher frequencies of naïve T cells activated upon the combination with CTLA4 blockade. Thus, radioimmunotherapy with carbon ions plus CTLA4 inhibition reshapes the tumor-infiltrating immune cell composition and can induce complete rejection even of non-irradiated tumors. Our data suggest combining radiotherapy approaches with CTLA4 blockade to achieve durable antitumor immunity. Evaluation of future radioimmunotherapy approaches should not be restricted to immunological impact at the irradiation site but should also consider systemic immunological effects on non-irradiated tumors., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: J.D. has received research grants from Siemens Healthcare GmbH, Solution Akademia GmbH, ViewRay, The Clinical Research Institute GmbH, Accuray International Sàrl, RaySearch Laboratories AB, Vision RT, Merck Serono GmbH, Astellas Pharma GmbH, AstraZeneca GmbH, Ergomed, Quintiles GmbH, and Pharmaceutical Research Associates GmbH and declares grant support from EMD/Merck KGaA to his institution to conduct experiments not directly related to the published study. A.A. has received research grants from Merck KGaA, FibroGen, and Bayer; has a consulting or advisory role with Roche, Merck KGaA, Merck Serono, FibroGen, BMS Brazil, Bayer Health, and BioMedX; and declares grant support from EMD/Merck KGaA to his institution to conduct experiments not directly related to the published study. All other authors declare that they have no competing interests., (Copyright © 2022. Published by Elsevier B.V.)

Sialoglycans play a key role in many biological recognition processes and sialylated conjugates of various types have successfully been applied, e.g., as antivirals or in antitumor therapy. A key feature for high affinity binding of such conjugates is the multivalent presentation of sialoglycans which often possess synthetic challenges. Here, the combination is described of solid phase polymer synthesis and enzymatic sialylation yielding 3'-sialyllactose-presenting precision glycomacromolecules. CMP-Neu5Ac synthetase from Neisseria meningitidis (NmCSS) and sialyltransferase from Pasteurella multocida (PmST1) are combined in a one-pot reaction giving access to sequence-defined sialylated macromolecules. Surprisingly, when employing Tris(hydroxymethyl)aminomethane (Tris) as a buffer, formation of significant amounts of α-linked Tris-sialoside is observed as a side reaction. Further exploring and exploiting this unusual sialylation reaction, different neoglycosidic structures are synthesized showing that PmST1 can be used to derive both, sialylation on natural carbohydrates as well as on synthetic hydroxylated scaffolds., (© 2022 The Authors. Macromolecular Bioscience published by Wiley-VCH GmbH.)

Background: Historically, complex media are used for the cultivation of Gluconobacter oxydans in industry and research. Using complex media has different drawbacks like higher costs for downstream processing and significant variations in fermentation performances. Synthetic media can overcome those drawbacks, lead to reproducible fermentation performances. However, the development of a synthetic medium is time and labour consuming. Detailed knowledge about auxotrophies and metabolic requirements of G. oxydans is necessary. In this work, we use a systematic approach applying the in-house developed μRAMOS technology to identify auxotrophies and develop a defined minimal medium for cultivation of G. oxydans fdh, improving the production process of the natural sweetener 5-ketofructose., Results: A rich, defined synthetic medium, consisting of 48 components, including vitamins, amino acids and trace elements, was used as a basis for medium development. In a comprehensive series of experiments, component groups and single media components were individually omitted from or supplemented to the medium and analysed regarding their performance. Main components like salts and trace elements were necessary for the growth of G. oxydans fdh, whereas nucleotides were shown to be non-essential. Moreover, results indicated that the amino acids isoleucine, glutamate and glycine and the vitamins nicotinic acid, pantothenic acid and p-aminobenzoic acid are necessary for the growth of G. oxydans fdh. The glutamate concentration was increased three-fold, functioning as a precursor for amino acid synthesis. Finally, a defined minimal medium called 'Gluconobacter minimal medium' was developed. The performance of this medium was tested in comparison with commonly used media for Gluconobacter. Similar/competitive results regarding cultivation time, yield and productivity were obtained. Moreover, the application of the medium in a fed-batch fermentation process was successfully demonstrated., Conclusion: The systematic investigation of a wide range of media components allowed the successful development of the Gluconobacter minimal medium. This chemically defined medium contains only 14 ingredients, customised for the cultivation of G. oxydans fdh and 5-ketofructose production. This enables a more straightforward process development regarding upstream and downstream processing. Moreover, metabolic demands of G. oxydans were identified, which further can be used in media or strain development for different processes., (© 2022. The Author(s).)

Many natural proteins contain flexible loops utilizing well-defined complementary surface regions of their interacting partners and usually undergo major structural rearrangements to allow perfect binding. The molecular recognition of such flexible structures is still highly challenging due to the inherent conformational dynamics. Notably, protein-protein interactions are on the other hand characterized by a multivalent display of complementary binding partners to enhance molecular affinity and specificity. Imitating this natural concept, we here report the rational design of advanced multivalent supramolecular tweezers that allow addressing two lysine and arginine clusters on a flexible protein surface loop. The protease Taspase 1, which is involved in cancer development, carries a basic bipartite nuclear localization signal (NLS) and thus interacts with Importin α, a prerequisite for proteolytic activation. Newly established synthesis routes enabled us to covalently fuse several tweezer molecules into multivalent NLS ligands. The resulting bi- up to pentavalent constructs were then systematically compared in comprehensive biochemical assays. In this series, the stepwise increase in valency was robustly reflected by the ligands' gradually enhanced potency to disrupt the interaction of Taspase 1 with Importin α, correlated with both higher binding affinity and inhibition of proteolytic activity.

Clonal hematopoiesis (CH) is characterized by somatic mutations in blood cells of individuals without hematologic disease. While the mutational landscape of CH in peripheral blood (PB) has been well characterized, detailed analyses addressing its spatial and cellular distribution in the bone marrow (BM) compartment are sparse. We studied CH driver mutations in healthy individuals (n = 261) across different anatomical and cellular compartments. Variant allele frequencies were higher in BM than PB and positively correlated with the number of driver variants, yet remained stable during a median of 12 months of follow-up. In CH carriers undergoing simultaneous bilateral hip replacement, we detected ASXL1-mutant clones in one anatomical location but not the contralateral side, indicating intra-patient spatial heterogeneity. Analyses of lineage involvement in ASXL1-mutated CH showed enriched clonality in BM stem and myeloid progenitor cells, while lymphocytes were particularly involved in individuals carrying the c.1934dupG variant, indicating different ASXL1 mutations may have distinct lineage distribution patterns. Patients with overt myeloid malignancies showed higher mutation numbers and allele frequencies and a shifting mutation landscape, notably characterized by increasing prevalence of DNMT3A codon R882 variants. Collectively, our data provide novel insights into the genetics, evolution, and spatial and lineage-specific BM involvement of CH., (© 2022. The Author(s).)

Heparin (HP) and heparan sulfate (HS) are linear, anionically charged polysaccharides well-known for their diverse biological activities. While HP is generally localized in mast cells and in connective tissues, HS is part of the glycocalyx and involved in the attachment of viruses to host cells, constituting the first step of an infection. HP and HS also exhibit antiviral activity by blocking viral receptors, thereby inhibiting viruses from engaging with host cells. Inspired by their structural features, such as their high molecular weight and polyanionic character, various synthetic polymers mimicking HP/HS have been developed and used as model systems to study bioactivity, as well as for therapeutic applications. This Perspective provides an overview of the roles of HP/HS in viral engagement, and examines historical and recent approaches toward oligo-/polysaccharide, glycopolymer, and anionic polymer HP/HS mimetics. An overview of current applications and future prospects of these molecules is provided, demonstrating their potential in addressing current and future epidemics and pandemics., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Sulfated glycosaminoglycans (sGAGs) such as heparan sulfate (HS) are structurally diverse linear polysaccharides that are involved in many biological processes and have gained interest as antiviral compounds. Their recognition is driven by a complex orchestra of structural parameters that are still under intense investigation. One distinct characteristic is the incorporation of sulfation patterns including highly sulfated and non-sulfated sequences that provide variations in flexibility and conformation, which in turn impact the biological function of sGAGs. However, these distinct features have not yet been fully realized in the synthetic preparation of sGAG mimetics. Here, we present the synthesis of three groups of sulfated glycomacromolecules as sGAG mimetics: (i) globally sulfated glycooligomers, (ii) glycooligomers with sequence-defined sulfation patterns, and (iii) a globally sulfated glycooligomer-oligo- L -proline hybrid structure. The complete synthesis, including chemical sulfation, was conducted on solid support, enabled by the introduction of a commercially available photocleavable linker allowing for the preservation of sensitive sulfates during cleavage of the products. Structures were obtained in good purity and with high degrees of sulfation demonstrating the wide applicability of this methodology to prepare tailor-made sulfated glycomacromolecules and similar sGAG mimetics. Structures were tested for their anticoagulant properties showing activity similar to their natural HS counterpart and significantly lower than HP.

Biobanking plays a critical role in diagnostics, biomarker research and development of novel treatment approaches for various diseases. In urgent need of understanding, preventing and treating coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the importance of biobanking including data sharing and management further increased. To provide high quality tissue biomaterials and data for research and public health, the COVID-19 Autopsy and Biosample Registry was established in the state of Baden-Wuerttemberg (BW) in Germany, combining expertise and technologies of the Institutes of Pathology of the five university hospitals in BW (Heidelberg, Tübingen, Ulm, Freiburg, Mannheim). The COVID-19 Autopsy and Biosample Registry BW comprises tissue samples from autopsies and associated data of deceased patients in the context of SARS-CoV-2 infection and/or vaccination against SARS-CoV-2. The aim is to collect autopsy biospecimens, associated clinical and diagnostic data in a timely manner, register them, make them accessible for research projects and thus to support especially tissue-related research addressing COVID-19. By now, the BW network holds multiple collaborations and supported numerous publications to increase the understanding of COVID-19 disease. The achievements of the BW network as a landmark biobanking model project represent a potential blueprint for future disease-related biobanking and registry effort., Competing Interests: Declaration of interest None., (Copyright © 2022 The Authors. Published by Elsevier GmbH.. All rights reserved.)

We analyze the influence of music in a network of FitzHugh-Nagumo oscillators with empirical structural connectivity measured in healthy human subjects. We report an increase of coherence between the global dynamics in our network and the input signal induced by a specific music song. We show that the level of coherence depends crucially on the frequency band. We compare our results with experimental data, which also describe global neural synchronization between different brain regions in the gamma-band range in a time-dependent manner correlated with musical large-scale form, showing increased synchronization just before transitions between different parts in a musical piece (musical high-level events). The results also suggest a separation in musical form-related brain synchronization between high brain frequencies, associated with neocortical activity, and low frequencies in the range of dance movements, associated with interactivity between cortical and subcortical regions., 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. The handling editor KL declared a past collaboration with the authors JS and ES., (Copyright © 2022 Sawicki, Hartmann, Bader and Schöll .)

Adjustment of key parameters of electrical stunning of pigs is necessary to better respect animal welfare and avoid impairments of meat quality. In the present study, we analysed the effect of genetic line, stunning program and genetic line × stunning program interactions on stunning effectiveness, carcass and meat quality traits in 10,338 pigs from five different genetic lines in a German slaughterhouse using electrical stunning (two-cycle method head/heart current). Genetic line, stunning program and its interaction significantly influenced stunning effectiveness and pH in the semimembranosus muscle. Backup stunning rates ranged from 1.59 to 5.01% in the five genetic lines, and were significantly decreased (2.85%) in program 1 using a current of 1.3 to 1.35 A, 800 Hz and 1 s for the head-cycle compared to program 2 (4.73%) with 1.8 A, 100 Hz and 4.5 s. We suggest to consider genetic line × stunning program interactions when adjusting officially defined stunning parameters for head-heart electrical stunning., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

In this contribution, we report on the "free" [P(C 2 F 5 ) 2 F 2 ] - ion and its ligand properties in transition metal complex chemistry. For this purpose, Ag[P(C 2 F 5 ) 2 F 2 ] was treated with [{(Et 2 N) 3 P═N} 3 P═NHC(CH 3 ) 3 ]Cl ([EtP 4 H]Cl) to yield [EtP 4 H][P(C 2 F 5 ) 2 F 2 ], featuring a weakly coordinating phosphazenium cation. Due to the weak interaction between the cation and anion, the [P(C 2 F 5 ) 2 F 2 ] - ion meets the so-called pseudo-gas-phase conditions. To determine the Tolman electronic parameter, [EtP 4 H][Ni(CO) 3 {P(C 2 F 5 ) 2 F 2 }] was prepared from [EtP 4 H][P(C 2 F 5 ) 2 F 2 ] and [Ni(CO) 4 ], facilitating the classification of the P(C 2 F 5 ) 2 F 2 moiety as a moderately π-acidic ligand. By treatment of [EtP 4 H][P(C 2 F 5 ) 2 F 2 ] with [AuCl(tht)], the neutral tetrahydrothiophene ligand was substituted by the phosphoranide ion, yielding [EtP 4 H][AuCl{P(C 2 F 5 ) 2 F 2 }]. When Ag[P(C 2 F 5 ) 2 F 2 ] was treated with [AuCl(tht)], on the other hand, the chloride was substituted. Transmetalation reactions of this type proved to be an efficient transfer method of the P(C 2 F 5 ) 2 F 2 moiety, as further demonstrated by the reactions of Ag[P(C 2 F 5 ) 2 F 2 ] with [FeCl(CO) 2 Cp], [FeCl(CO) 2 Cp*], and [PdCl 2 (NCMe) 2 ]. Surprisingly, P(C 2 F 5 ) 2 F demonstrated fluorinating abilities toward [FeCl(CO) 2 Cp], [FeCl(CO) 2 Cp*], [AuCl(tht)], and [PdCl 2 (NCMe) 2 ]. Apparently, fluorido transition metal complexes were generated in situ under the formation of P(C 2 F 5 ) 2 Cl. The fluorido iron and palladium complexes transfer their fluoride ions onto P(C 2 F 5 ) 2 F, yielding the respective phosphoranido complexes, featuring the P(C 2 F 5 ) 2 F 2 moiety.

The tetrazine/ trans -cyclooctene ligation stands out from the bioorthogonal toolbox due to its exceptional reaction kinetics, enabling multiple molecular technologies in vitro and in living systems. Highly reactive 2-pyridyl-substituted tetrazines have become state of the art for time-critical processes and selective reactions at very low concentrations. It is widely accepted that the enhanced reactivity of these chemical tools is attributed to the electron-withdrawing effect of the heteroaryl substituent. In contrast, we show that the observed reaction rates are way too high to be explained on this basis. Computational investigation of this phenomenon revealed that distortion of the tetrazine caused by intramolecular repulsive N-N interaction plays a key role in accelerating the cycloaddition step. We show that the limited stability of tetrazines in biological media strongly correlates with the electron-withdrawing effect of the substituent, while intramolecular repulsion increases the reactivity without reducing the stability. These fundamental insights reveal thus far overlooked mechanistic aspects that govern the reactivity/stability trade-off for tetrazines in physiologically relevant environments, thereby providing a new strategy that may facilitate the rational design of these bioorthogonal tools.

In all vertebrates including mammals, the ergothioneine transporter ETT (obsolete name OCTN1; human gene symbol SLC22A4) is a powerful and highly specific transporter for the uptake of ergothioneine (ET). ETT is not expressed ubiquitously and only cells with high ETT cell-surface levels can accumulate ET to high concentration. Without ETT, there is no uptake because the plasma membrane is essentially impermeable to this hydrophilic zwitterion. Here, we review the substrate specificity and localization of ETT, which is prominently expressed in neutrophils, monocytes/macrophages, and developing erythrocytes. Most sites of strong expression are conserved across species, but there are also major differences. In particular, we critically analyze the evidence for the expression of ETT in the brain as well as recent data suggesting that the transporter SLC22A15 may also transport ET. We conclude that, to date, ETT remains the only well-defined biomarker for intracellular ET activity. In humans, the ability to take up, distribute, and retain ET depends principally on this transporter., (© 2022 Federation of European Biochemical Societies.)

Streptomyces species are intensively studied for their ability to produce a variety of natural products. However, conditions influencing and leading to product formation are often not completely recognized. Therefore, in this study, high-throughput online monitoring is presented as a powerful tool to gain in-depth understanding of the cultivation of the model organism Streptomyces coelicolor A3(2). Through online measurements of oxygen transfer rate and autofluorescence, valuable information about availability of nutrients and product formation patterns of the pigments actinorhodin and undecylprodigiosin can be obtained and explained. Therefore, it is possible to determine the onset of pigmentation and to study in detail the influencing factors thereof. One factor identified in this study is the filling volume of the cultivation vessel. Slight variations led to varying pigmentation levels. By combining optical and metabolic online monitoring techniques, the correlation of the filling volume with pigmentation could be explained as a result of different growth trajectories caused by varying specific power inputs and their influence on the pellet formation of the filamentous system. Finally, experiments with the addition of supernatant from unpigmented and pigmented cultures could highlight the applicability of the presented approach to study quorum sensing and cell-cell interaction., Competing Interests: The authors have declared no conflicts of interest., (© 2022 The Authors. Engineering in Life Sciences published by Wiley‐VCH GmbH.)

Background: Acquiring a physical disability in adulthood necessitates a range of adjustments, with past research suggesting that some challenges encountered are unique to women. Moreover, several factors may complicate adjustment to an altered embodiment and difficulties in functioning after an accident, including insufficient rehabilitation and support services and problematic societal attitudes towards disability. In addition, women with disabilities are often excluded from health and social policy and programme development, an oversight that can result in support gaps., Objectives: This article presents the self-identified priority interventions of women with road accident-acquired physical disabilities in South Africa., Methods: We conducted interviews with 18 women with road accident-acquired physical disabilities. The participants were recruited via snowball sampling. Interviews were conducted by experienced interviewers, who were home language speakers of the participants' preferred language of communication. The interview recordings were transcribed, translated, and coded by trained, independent researchers., Results: Study participants identified three key areas of intervention requiring consideration in supportive intervention planning: the acute post-injury environment and healthcare infrastructure, transitional services and social inclusion interventions. These were identified as overlooked areas in which they required support to successfully adapt to limitations in functioning., Conclusion: To develop inclusive, accessible, and practical policy and programming for people with disabilities, exercises like those outlined in this research - eliciting intervention ideas from lived experience - should be conducted as they highlight actionable priorities for programming., Competing Interests: The authors declare that they have no financial or personal relationships that may have inappropriately influenced them in writing this article., (© 2022. The Authors.)

African swine fever virus (ASFV) remains a threat to global pig populations. Infections with ASFV lead to a hemorrhagic disease with up to 100% lethality in Eurasian domestic and wild pigs. Although myeloid cells are the main target cells for ASFV, T cell responses are impacted by the infection as well. The complex responses remain not well understood, and, consequently, there is no commercially available vaccine. Here, we review the current knowledge about the induction of antiviral T cell responses by cells of the myeloid lineage, as well as T cell responses in infected animals, recent efforts in vaccine research, and T cell epitopes present in ASFV.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the novel coronavirus responsible for COVID-19. Infection in humans requires angiotensin-converting enzyme II (hACE2) as the point of entry for SARS-CoV-2. PCR testing is generally definitive but expensive, although it is highly sensitive and accurate. Biosensor-based monitoring could be a low-cost, accurate, and non-invasive approach to improve testing capacity. We develop a capacitive hACE2 biosensor for intact SARS-CoV-2 detection in saliva. Laser-induced graphene (LIG) electrodes were modified with platinum nanoparticles. The quality control of LIG electrodes was performed using cyclic voltammetry. Truncated hACE2 was used as a biorecognition element and attached to the electrode surface by streptavidin-biotin coupling. Biolayer interferometry was used for qualitative interaction screening of hACE2 with UV-attenuated virions. Electrochemical impedance spectroscopy (EIS) was used for signal transduction. Truncated hACE2 binds wild-type SARS-CoV-2 and its variants with greater avidity than human coronavirus (common cold virus). The limit of detection (LoD) is estimated to be 2,960 copies/ml. The detection process usually takes less than 30 min. The strength of these features makes the hACE2 biosensor a potentially low-cost approach for screening SARS-CoV-2 in non-clinical settings with high demand for rapid testing (for example, schools and airports)., Competing Interests: Conflict of Interest: 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.