Search

Abstract Ligand-induced activation of G protein-coupled receptors (GPCRs) can initiate signaling through multiple distinct pathways with differing biological and physiological outcomes. There is intense interest in understanding how variation in GPCR ligand structure can be used to promote pathway selective signaling (“biased agonism”) with the goal of promoting desirable responses and avoiding deleterious side effects. Here we present an approach in which a conventional peptide ligand for the type 1 parathyroid hormone receptor (PTHR1) is converted from an agonist which induces signaling through all relevant pathways to a compound that is highly selective for a single pathway. This is achieved not through variation in the core structure of the agonist, but rather by linking it to a nanobody tethering agent that binds with high affinity to a separate site on the receptor not involved in signal transduction. The resulting conjugate represents the most biased agonist of PTHR1 reported to date. This approach holds promise for facile generation of pathway selective ligands for other GPCRs.

Summary: Here, we present a protocol for the development of mRNA-loaded lipid nanoparticle (LNP) vaccines for target antigen sequences of interest. We describe key steps required to design and synthesize mRNA constructs, their LNP encapsulation, and mouse immunization. We then detail quality control assays to determine RNA purity, guidelines to measure RNA immunogenicity using in vitro reporter systems, and a technique to evaluate antigen-specific T cell responses following immunization. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

IntroductionWhite matter hyperintensities (WMH) are associated with key dementia etiologies, in particular arteriolosclerosis and amyloid pathology. We aimed to identify WMH locations associated with vascular risk or cerebral amyloid-β1-42 (Aβ42)-positive status.MethodsIndividual patient data (n = 3,132; mean age 71.5 ± 9 years; 49.3% female) from 11 memory clinic cohorts were harmonized. WMH volumes in 28 regions were related to a vascular risk compound score (VRCS) and Aß42 status (based on cerebrospinal fluid or amyloid positron emission tomography), correcting for age, sex, study site, and total WMH volume.ResultsVRCS was associated with WMH in anterior/superior corona radiata (B = 0.034/0.038, p

A viscoplastic model for representing plastic silts and clays in geotechnical static slope stability applications is presented. The PM4SiltR model builds on the stress ratio-controlled, critical state-based, bounding surface plasticity model PM4Silt and is coded as a dynamic link library for use in the finite difference program FLAC 8.1. PM4SiltR incorporates strain rate-dependent shear strength, stress relaxation, and creep using a consistency approach combined with an internal strain rate and auto-decay process. The model does not include a cap, and as such cannot simulate strain rate-dependent consolidation under increasing overburden stress. Six parameters control the viscous response for PM4SiltR, while the parameters controlling the nonviscous components of the response are the same as for PM4Silt. Single element simulations are presented to illustrate the influence of viscoplasticity on the constitutive response in direct simple shear loading and undrained creep. Single element responses are shown to be consistent with observed experimental results. Simulations of a hypothetical tailings dam constructed using the upstream method are performed to illustrate use of PM4SiltR at field scale. Results of field-scale simulations show PM4SiltR can model undrained creep and progressive failure leading to delayed slope instability after relatively minor changes in loading conditions at field scale.

Potassium (K) nutrition and drought stress affect soybean (Glycine max (L.) Merr.) vigor and productivity through the combined impacts on water regulation. A study was conducted with soybean grown in 18.9‐L buckets under a rain out shelter to determine how the interaction between these crop stresses at various growth stages influences the crop leaf K concentration, biomass production, total K uptake (TKU), grain yield, and temperature of the uppermost fully expanded trifoliate. Treatments included soybean grown with and without preplant fertilizer K, soil moisture at 50% (drought) or 80% (well‐watered) field capacity, imposed drought during vegetative growth (V3–V7), flowering (R1–R3), pod development (R4–early R6), and seed development (R5–mid‐R6) on two different silt loam soils. Widespread K deficiencies were observed during the study across all treatments. Drought stress significantly (p < 0.05) reduced the TKU, aboveground biomass production, and grain yield. The crop growth stage when drought stress was imposed was a significant factor, with greater reductions in plant response parameters when stress was imposed during reproductive growth. Preplant fertilizer K increased trifoliolate K concentrations and TKU in drought conditions, but did not increase the grain yield of well‐watered soybean. Leaf temperature increased when under drought stress compared to well‐watered soybean (p < 0.0001) but the impact of crop K nutrition status on leaf temperature was inconclusive because of widespread K deficiencies. Results emphasize the complexity of the interactions between K nutrition and drought stress in soybean, as drought stress impeded K uptake, exacerbated K deficiencies, and limited yield. Core Ideas: Drought stress exacerbated the impacts of K deficiency in soybean.Leaf temperature increased when under drought stress across growth stages but was inconclusive when K deficient.Total K uptake and biomass production are affected by both K availability and drought stress across growth stages.Drought stress during reproductive stages reduces total K uptake, biomass, and grain yield to a greater degree. [ABSTRACT FROM AUTHOR]

A viscoplastic model for representing plastic silts and clays in geotechnical static slope stability applications is presented. The PM4SiltR model builds on the stress ratio-controlled, critical state-based, bounding surface plasticity model PM4Silt and is coded as a dynamic link library for use in the finite difference program FLAC 8.1. PM4SiltR incorporates strain rate-dependent shear strength, stress relaxation, and creep using a consistency approach combined with an internal strain rate and auto-decay process. The model does not include a cap, and as such cannot simulate strain rate-dependent consolidation under increasing overburden stress. Six parameters control the viscous response for PM4SiltR, while the parameters controlling the nonviscous components of the response are the same as for PM4Silt. Single element simulations are presented to illustrate the influence of viscoplasticity on the constitutive response in direct simple shear loading and undrained creep. Single element responses are shown to be consistent with observed experimental results. Simulations of a hypothetical tailings dam constructed using the upstream method are performed to illustrate use of PM4SiltR at field scale. Results of field-scale simulations show PM4SiltR can model undrained creep and progressive failure leading to delayed slope instability after relatively minor changes in loading conditions at field scale. [ABSTRACT FROM AUTHOR]

Prior work has shown Americans have higher levels of memory specificity than East Asians. Neuroimaging studies have not investigated mechanisms that account for cultural differences at retrieval. In this study, we use fMRI to assess whether mnemonic discrimination, distinguishing novel from previously encountered stimuli, accounts for cultural differences in memory. Fifty-five American and 55 Taiwanese young adults completed an object recognition paradigm testing discrimination of old targets, similar lures and novel foils. Mnemonic discrimination was tested by comparing discrimination of similar lures from studied targets, and results showed the relationship between activity in right fusiform gyrus and behavioural discrimination between target and lure objects differed across cultural groups. Parametric modulation analyses of activity during lure correct rejections also indicated that groups differed in left superior parietal cortex response to variations in lure similarity. Additional analyses of old vs. new activity indicated that Americans and Taiwanese differ in the neural activity supporting general object recognition in the hippocampus, left inferior frontal gyrus and middle frontal gyrus. Results are juxtaposed against comparisons of the regions activated in common across the two cultures. Overall, Americans and Taiwanese differ in the extent to which they recruit visual processing and attention modulating brain regions. [ABSTRACT FROM AUTHOR]

This interview with art therapist Gary Nash explores his thoughts on the field, both successes and challenges, including his work with response art and as a portfolio art therapist. Additionally, Nash offers his observations about art-based research based on his many years of research experience. Nash provides some background on his interest in and approach to art therapy, response art and art-based research. Finally, Nash shares how Nature influences and inspires his more recent work in environmental arts therapy. [ABSTRACT FROM AUTHOR]

This editorial celebrates the Journal of Applied Arts & Health's (JAAH) fifteenth anniversary, reflecting on its evolution and impact at the intersection of art, health and well-being. It reflects on the previous JAAH Special Issue (14:3) paying homage to Emeritus University Professor Shaun McNiff, a trailblazer in the field, whose fifty-plus-year career is highlighted. The journal's role in promoting understanding of art's diverse benefits, particularly in healthcare, is emphasized. Amid global challenges, the editorial acknowledges the world's ongoing shifts and challenges to health and well-being, including the impact of the COVID-19 pandemic, economic disruptions, geopolitical tensions and climate change concerns. It recognizes the role of JAAH in fostering exploration, understanding and advocacy in applied arts and health, while calling for continued innovation and inquiry in the next fifteen years. The editorial introduces the current journal issue's content, which covers a range of research projects, an interview and reviews. [ABSTRACT FROM AUTHOR]

Antibodies have proven highly valuable for therapeutic development; however, they are typically poor candidates for applications that require activation of G protein‐coupled receptors (GPCRs), the largest collection of targets for clinically approved drugs. Nanobodies (Nbs), the smallest antibody fragments retaining full antigen‐binding capacity, have emerged as promising tools for pharmacologic applications, including GPCR modulation. Past work has shown that conjugation of Nbs with ligands can provide GPCR agonists that exhibit improved activity and selectivity compared to their parent ligands. The neurokinin‐1 receptor (NK1R), a GPCR targeted for the treatment of pain, is activated by peptide agonists such as Substance P (SP) and neurokinin A (NKA), which induce signaling through multiple pathways (Gs, Gq and β‐arrestin). In this study, we investigated whether conjugating NK1R ligands with Nbs that bind to a separate location on the receptor would provide chimeric compounds with distinctive signaling properties. We employed sortase A‐mediated ligation to generate several conjugates consisting of Nbs linked to NK1R ligands. Many of these conjugates exhibited divergent and unexpected signaling properties and transcriptional outputs. For example, some Nb‐NKA conjugates showed enhanced receptor binding capacity, high potency partial agonism, prolonged cAMP production, and an increase in transcriptional output associated with Gs signaling; whereas other conjugates were virtually inactive. Nanobody conjugation caused only minor alterations in ligand‐induced upstream Gq signaling with unexpected enhancements in transcriptional (downstream) responses. Our findings underscore the potential of nanobody conjugation for providing compounds with advantageous properties such as biased agonism, prolonged duration of action, and enhanced transcriptional responses. These compounds hold promise not only for facilitating fundamental research on GPCR signal transduction mechanisms but also for the development of more potent and enduring therapeutics. [ABSTRACT FROM AUTHOR]

Vaccination is considered one of the major milestones in modern medicine, facilitating the control and eradication of life‐threatening infectious diseases. Vaccine adjuvants are a key component of many vaccines, serving to steer antigen‐specific immune responses and increase their magnitude. Despite major advances in the field of adjuvant research over recent decades, our understanding of their mechanism of action remains incomplete. This hinders our capacity to further improve these adjuvant technologies, so addressing how adjuvants induce and control the induction of innate and adaptive immunity is a priority. Investigating how adjuvant physicochemical properties, such as size and charge, exert immunomodulatory effects can provide valuable insights and serve as the foundation for the rational design of vaccine adjuvants. Most clinically applied adjuvants are particulate in nature and polymeric particulate adjuvants present advantages due to stability, biocompatibility profiles, and flexibility in terms of formulation. These properties can impact on antigen release kinetics and biodistribution, cellular uptake and targeting, and drainage to the lymphatics, consequently dictating the induction of innate, cellular, and humoral adaptive immunity. A current focus is to apply rational design principles to the development of adjuvants capable of eliciting robust cellular immune responses including CD8+ cytotoxic T‐cell and Th1‐biased CD4+ T‐cell responses, which are required for vaccines against intracellular pathogens and cancer. This review highlights recent advances in our understanding of how particulate adjuvants, especially polymer‐based particulates, modulate immune responses and how this can be used as a guide for improved adjuvant design. [ABSTRACT FROM AUTHOR]

Chondrogenic induction of bone-marrow-derived stromal cells (BMSCs) is typically accomplished with medium supplemented with growth factors (GF) from the transforming growth factor-beta (TGF-β)/bone morphogenetic factor (BMP) superfamily. In a previous study, we demonstrated that brief (1–3 days) stimulation with TGF-β1 was sufficient to drive chondrogenesis and hypertrophy using small-diameter microtissues generated from 5000 BMSC each. This biology is obfuscated in typical large-diameter pellet cultures, which suffer radial heterogeneity. Here, we investigated if brief stimulation (2 days) of BMSC microtissues with BMP-2 (100 ng/mL) or growth/differentiation factor (GDF-5, 100 ng/mL) was also sufficient to induce chondrogenic differentiation, in a manner comparable to TGF-β1 (10 ng/mL). Like TGF-β1, BMP-2 and GDF-5 are reported to stimulate chondrogenic differentiation of BMSCs, but the effects of transient or brief use in culture have not been explored. Hypertrophy is an unwanted outcome in BMSC chondrogenic differentiation that renders engineered tissues unsuitable for use in clinical cartilage repair. Using three BMSC donors, we observed that all GFs facilitated chondrogenesis, although the efficiency and the necessary duration of stimulation differed. Microtissues treated with 2 days or 14 days of TGF-β1 were both superior at producing extracellular matrix and expression of chondrogenic gene markers compared to BMP-2 and GDF-5 with the same exposure times. Hypertrophic markers increased proportionally with chondrogenic differentiation, suggesting that these processes are intertwined for all three GFs. The rapid action, or "temporal potency", of these GFs to induce BMSC chondrogenesis was found to be as follows: TGF-β1 > BMP-2 > GDF-5. Whether briefly or continuously supplied in culture, TGF-β1 was the most potent GF for inducing chondrogenesis in BMSCs. [ABSTRACT FROM AUTHOR]

Purpose To assess sickness absence and transitions from employment for employees with arthritis compared with employees without arthritis over time.Methods We use 10 waves of the UK Household Longitudinal Survey (2009–2019). The sample (n=38 928) comprises employees aged 50 years to state retirement age. Arthritis was self-reported and could refer to people with conditions under the umbrella term ‘inflammatory arthritis’ or osteoarthritis (hereafter ‘arthritis’). Weighted random-effects multivariable linear probability models were estimated for two employment-related measures (1) sickness absence and (2) transitions from employment to: (a) unemployment; (b) long-term sick; (c) early retirement. These were regressed against a variable for arthritis and confounding factors (age, socioeconomic job classification, employing sector, year and additional health conditions). Additional analyses examined an interaction between the variable arthritis and these factors to test whether the effect of arthritis differs between these groups.Results Employees reporting having arthritis were more likely to have sickness absence (1.35 percentage points greater rate (95% CI (0.92, 1.78)) and to transition to long-term sick (0.79 percentage points (0.46, 1.13)) and early retirement (0.58 percentage points (0.05, 1.11)). No effect was found for transitions to unemployment. There was limited evidence that the effects of arthritis vary for employees in different socioeconomic classifications.Conclusions Employees living with arthritis have higher rates of sickness absence and greater rates of transitions from employment to long-term sick and early retirement. Further work could look at ways to quantify the implications for individuals, employers and the state and ways to alleviate the effects of living with arthritis on work participation.

Background Physical activity referral schemes have been used extensively as one pathway to support behaviour change in people with long term conditions. Best practice guidance, across countries, recommend that schemes use behaviour change practices. The effectiveness of these schemes is inconsistent, yet, little is known about the implementation of specific approaches, or what influences practitioner’s delivery. This article provides a narrative review of evidence exploring the implementation of behaviour change practices in physical activity referral schemes.Methods An electronic search of three databases (PubMed, Scopus, Google Scholar) was undertaken. A menu of iterative techniques was also applied from the CLUSTER approach to increase coverage.Results A total of 45 eligible articles was included covering diverse research designs. Enduring issues with the literature pertain to the insufficient emphasis on implementation, a conflation of behaviour change practice, and an inconsistency of scheme components. Against this backdrop, diverse factors within practitioner, attendee, partnership, work environment, and organisational domains influence the implementation of behaviour change practices.Conclusion The translation of behaviour change practices to applied physical activity settings must tend to the multilevel factors which have the potential to influence the quality of behaviour change implementation.

IntroductionConventional 2D intestinal epithelial cell lines have been widely used in investigating intestinal functions, yet with limitations in recapitulating the in vivo gut physiology of chickens. A recently established chicken enteroid model with apical-out nature and the presence of leukocyte components represents intestinal mucosal functions. The objectives of this study were to 1) evaluate basic gut nutrient transport and barrier functions in this model and 2) identify the model’s effectiveness in studying inflammation and oxidative stress responses.MethodsEnteroids were generated from individual villus units isolated from the small intestine of Cobb500 broiler embryos. Enteroid viability, morphology, and epithelial cell markers were monitored; barrier function was evaluated based on the permeability to fluorescein isothiocyanate–dextran (FD4) with or without EDTA and lipopolysaccharide (LPS) challenges; nutrient transport was evaluated by fluorescence-labeled glucose (2NBD-G) with or without transporter blockade; the oxidative status was indicated by reactive oxygen species (ROS). Inflammatory and oxidative challenges were induced by LPS and menadione treatment, respectively. Selected marker gene expressions, including tight junction proteins (CLDN-1, CLDN-2, ZO-1, and OCCL), epithelial cell markers (Lgr-5, LYZ, and MUC-2), cytokines (IL-1β, IL-6, IL-8, IL-10, TNF-α, and INF-γ), and antioxidant enzymes (Nrf-2, catalase, and SOD), were determined by using RT-qPCR. Data were analyzed by one-way ANOVA among treatment groups.ResultsEnteroid cell activity was stable from day (d) 2 to d 6 and declined at d 7. Epithelial cell marker and cytokine expressions were stable from d 4 to d 6. FD4 permeability was increased after the EDTA treatment (P ≤ 0.05). Transporter-mediated 2NBD-G absorption was observed, which was reduced with glucose transporter blockade (P ≤ 0.05). Enteroids showed classic responses to LPS challenges, including upregulated gene expressions of IL-1β and IL-6, downregulated gene expressions of ZO-1 and OCCL, and increased FD4 permeability (P ≤ 0.05). Enteroids showed increased ROS generation (P ≤ 0.05) in response to oxidative stress.DiscussionIn conclusion, this apical-out enteroid model is a stable alternative in vitro model that exhibits intestinal barrier, nutrient transport, oxidation, and inflammation functions. With this enteroid model, we developed two challenge protocols for evaluating intestinal functions under oxidative stress and inflammation conditions.

The emerging fungal pathogen Batrachochytrium salamandrivorans (Bsal) threatens the diversity of amphibians, particularly in North America where it is projected to invade. Amphibian skin defenses include a mucosal layer containing microorganisms that can potentially modulate host response to pathogens such as Bsal. In this study, we focused on the composition of the skin microbiome across life stages of spotted salamanders (Ambystoma maculatum). We also evaluated the stress hormone corticosterone and skin microbiome response to inoculations with Bsal and probiotics at both the larval and juvenile developmental stages, and the response to different environmental conditions. Results indicated that both bacterial and fungal communities found on the skin significantly differed in structure and diversity between life stages of A. maculatum. Exposure to three different probiotics (Bacillus thuringiensis, Chryseobacterium rhizoplanae, and Penicillium sp.) and Bsal evoked shifts in the microbiome of larvae and juveniles, and the metabolite profile of the larval mucosal layer of A. maculatum. Despite changes in the microbiome, all tested probiotics and Bsal were unable to persist on the skin. Larval bacterial microbiomes shifted in response to Bsal and B. thuringiensis with no significant impacts on antifungal function or bacteria richness, however fungi strongly responded to Bsal and B. thuringiensis application. This indicates that developmental shifts in the microbiome can be initiated by microbial applications such as B. thuringiensis, a widely used mosquito larvicide. Overall, experimental results indicate that life stage, growth and development, and environmental conditions appeared to be the main factors driving changes in the amphibian skin microbiome and potential anti-Batrachochytrium function.