eIF2B localization and its regulation during the integrated stress response is cell-type specific.

Eukaryotic initiation factor 2B (eIF2B) controls translation initiation by recycling inactive eIF2-GDP to active eIF2-GTP. Under cellular stress, the integrated stress response (ISR) is activated inhibiting eIF2B activity resulting in the translation attenuation and reprogramming of gene expression to overcome the stress. The ISR can dictate cell fate wherein chronic activation has pathological outcomes. Vanishing white matter disease (VWMD) is a chronic ISR-related disorder with mutations in eIF2B targeting astrocyte and oligodendrocyte cells. Regulation of eIF2B localization (eIF2B bodies) has been implicated in the ISR. We present evidence that neuronal and glial cell types possess distinct patterns of eIF2B bodies which change in a manner correlating to acute and chronic ISR activation. We also demonstrate that while neural and glial cell types respond similarly to the acute induction of the ISR a chronic ISR exerts cell-type specific differences. These findings provide key insights into neural cell responses and adaptation to cellular stress.

Shear Stress Induces a Time-Dependent Inflammatory Response in Human Monocyte-Derived Macrophages.

Macrophages are innate immune cells that are known for their extreme plasticity, enabling diverse phenotypes that lie on a continuum. In a simplified model, they switch between pro-inflammatory (M1) and anti-inflammatory (M2) phenotypes depending on surrounding microenvironmental cues, which have been implicated in disease outcomes. Although considerable research has been focused on macrophage response to biochemical cues and mechanical signals, there is a scarcity of knowledge surrounding their behavior in response to shear stress. In this study, we applied varying magnitudes of shear stress on human monocyte-derived macrophages (MDMs) using a cone-and-plate viscometer and evaluated changes in morphology, gene expression, protein expression, and cytokine secretion over time. MDMs exposed to shear stress exhibited a rounder morphology compared to statically-cultured controls. RT-qPCR results showed significant upregulation of TNF-α, and analysis of cytokine release revealed increased secretion of IL-8, IL-18, fractalkine, and other chemokines. The upregulation of pro-inflammatory factors was evident with both increasing magnitudes of shear and time. Taken together, these results indicate that prolonged shear exposure induced a pro-inflammatory phenotype in human MDMs. These findings have implications for medical technology development, such as in situ vascular graft design wherein macrophages are exposed to shear and have been shown to affect graft resorption, and in delineating disease pathophysiology, for example to further illuminate the role of macrophages in atherosclerosis where shear is directly related to disease outcome.

Digital video analysis reveals gait parameters that predict performance in the jumping test phase of three-day eventing.

In international equestrian sport, visual inspections assess gait and lameness to protect the welfare of performance horses during competition. Horses competing internationally in three-day eventing must pass two mandatory inspections (pre-competition and post-cross country) before attempting the final phase: the jumping test (JT). We hypothesized that digitally quantifying objective gait parameters captured during the two mandatory inspections will identify locomotor characteristics that predict success during the jumping test. Utilizing the DeepLabCut (DLC) software package for labeling of anatomical landmarks and a custom analysis pipeline we calculated gait parameters for 194 competition horses at the trot. During the pre-competition inspection, relative trot speed was significantly associated (P = 0.0060, GLMM), and the forelimb travel trended towards significance (P =0.0800, GLMM), with achieving a clear round in the later jumping test. Post-cross country, the forelimb travel significantly predicted JT results (P = 0.0188, GLMM). As our parameters are scaled for body size, these parameters may indicate conformational characteristics for superior jumping ability and overall athletic fitness. Within each competitive effort, comparisons of the post-cross country and pre-competition observations revealed that the change in speed and duty factor were significantly different in the group that accrued jumping faults (P = 0.00376 and P = 0.02430, GLMM), perhaps capturing locomotor signs of exercise fatigue. Further work employing these approaches to better understand competition performance will encourage the use of objective measures to protect sport horse welfare, as well as provide an advantageous tool for gait evaluation in the horse.

People with short symptom duration of knee osteoarthritis benefit more from exercise therapy than people with longer symptom duration: An individual participant data meta-analysis from the OA trial bank.

OBJECTIVE: To investigate whether duration of knee symptoms influenced the magnitude of the effect of exercise therapy compared to non-exercise control interventions on pain and physical function in people with knee osteoarthritis (OA). METHOD: We undertook an individual participant data (IPD) meta-analysis utilising IPD stored within the OA Trial Bank from randomised controlled trials (RCTs) comparing exercise to non-exercise control interventions among people with knee OA. IPD from RCTs were analysed to determine the treatment effect by considering both study-level and individual-level covariates in the multilevel regression model. To estimate the interaction effect (i.e., treatment x duration of symptoms (dichotomised)), on self-reported pain or physical function (standardised to 0-100 scale), a one-stage multilevel regression model was applied. RESULTS: We included IPD from 1767 participants with knee OA from 10 RCTs. Significant interaction effects between the study arm and symptom duration (≤1 year vs >1 year, and ≤2 years vs>2 years) were found for short- (∼3 months) (Mean Difference (MD) -3.57, 95%CI -6.76 to -0.38 and -4.12, 95% CI-6.58 to -1.66, respectively) and long-term (∼12 months) pain outcomes (MD -8.33, 95%CI -12.51 to -4.15 and -8.00, 95%CI -11.21 to -4.80, respectively), and long-term function outcomes (MD -5.46, 95%CI -9.22 to -1.70 and -4.56 95%CI -7.33 to-1.80, respectively). CONCLUSIONS: This IPD meta-analysis demonstrated that people with a relatively short symptom duration benefit more from therapeutic exercise than those with a longer symptom duration. Therefore, there seems to be a window of opportunity to target therapeutic exercise in knee OA.

A novel system to culture human intestinal organoids under physiological oxygen content to study microbial-host interaction.

Mechanistic investigation of host-microbe interactions in the human gut are hindered by difficulty of co-culturing microbes with intestinal epithelial cells. On one hand the gut bacteria are a mix of facultative, aerotolerant or obligate anaerobes, while the intestinal epithelium requires oxygen for growth and function. Thus, a coculture system that can recreate these contrasting oxygen requirements is critical step towards our understanding microbial-host interactions in the human gut. Here, we demonstrate Intestinal Organoid Physoxic Coculture (IOPC) system, a simple and cost-effective method for coculturing anaerobic intestinal bacteria with human intestinal organoids (HIOs). Using commensal anaerobes with varying degrees of oxygen tolerance, such as nano-aerobe Bacteroides thetaiotaomicron and strict anaerobe Blautia sp., we demonstrate that IOPC can successfully support 24-48 hours HIO-microbe coculture. The IOPC recapitulates the contrasting oxygen conditions across the intestinal epithelium seen in vivo. The IOPC cultured HIOs showed increased barrier integrity, and induced expression of immunomodulatory genes. A transcriptomic analysis suggests that HIOs from different donors show differences in the magnitude of their response to coculture with anaerobic bacteria. Thus, the IOPC system provides a robust coculture setup for investigating host-microbe interactions in complex, patient-derived intestinal tissues, that can facilitate the study of mechanisms underlying the role of the microbiome in health and disease.

Digital Pathology Identifies Associations between Tissue Inflammatory Biomarkers and Multiple Sclerosis Outcomes.

BACKGROUND: Multiple sclerosis (MS) is a clinically heterogeneous disease underpinned by inflammatory, demyelinating and neurodegenerative processes, the extent of which varies between individuals and over the course of the disease. Recognising the clinicopathological features that most strongly associate with disease outcomes will inform future efforts at patient phenotyping. AIMS: We used a digital pathology workflow, involving high-resolution image acquisition of immunostained slides and opensource software for quantification, to investigate the relationship between clinical and neuropathological features in an autopsy cohort of progressive MS. METHODS: Sequential sections of frontal, cingulate and occipital cortex, thalamus, brain stem (pons) and cerebellum including dentate nucleus (n = 35 progressive MS, females = 28, males = 7; age died = 53.5 years; range 38-98 years) were immunostained for myelin (anti-MOG), neurons (anti-HuC/D) and microglia/macrophages (anti-HLA). The extent of demyelination, neurodegeneration, the presence of active and/or chronic active lesions and quantification of brain and leptomeningeal inflammation was captured by digital pathology. RESULTS: Digital analysis of tissue sections revealed the variable extent of pathology that characterises progressive MS. Microglia/macrophage activation, if found at a higher level in a single block, was typically elevated across all sampled blocks. Compartmentalised (perivascular/leptomeningeal) inflammation was associated with age-related measures of disease severity and an earlier death. CONCLUSION: Digital pathology identified prognostically important clinicopathological correlations in MS. This methodology can be used to prioritise the principal pathological processes that need to be captured by future MS biomarkers.

Technique for Rapidly Forming Networks of Microvessel-Like Structures.

Modeling organ-blood barriers through the inclusion of microvessel networks within in vitro tissue models could lead to more physiologically accurate results, especially since organ-blood barriers are crucial to the normal function, drug transport, and disease states of vascularized organs. Microvessel networks are difficult to form, since they push the practical limits of most fabrication methods, and it is difficult to coax vascular cells to self-assemble into structures larger than capillaries. Here, we present a method for rapidly forming networks of microvessel-like structures using sacrificial alginate structures. Specifically, we encapsulated endothelial cells within short alginate threads, and then embedded them in collagen gel. Following enzymatic degradation of the alginate, the collagen gel contained a network of hollow channels seeded with cells, all surrounding a perfusable central channel. This method uses a 3D-printed coaxial extruder and syringe pumps to generate short threads in a way that is repeatable and easily transferrable to other labs. The cell-laden, sacrificial alginate threads can be frozen after fabrication and thawed before embedding without significant loss of cell viability. The ability to freeze the threads enables future scale-up and ease of use. Within millifluidic devices that restrict access to media, the threads enhance cell survival under static conditions. These results indicate the potential for use of this method in a range of tissue engineering applications.

Still a 'boys' club': a qualitative analysis of how gender affects a career in anaesthesia in Australia and Aotearoa New Zealand.

Gender inequity remains an issue in anaesthesia despite increasing numbers of women training and achieving fellowship in the speciality. Women are under-represented in all areas of anaesthetic research, academia and leadership. The Gender Equity Subcommittee of the Australian and New Zealand College of Anaesthetists recently conducted a survey asking "Does gender still matter in the pursuit of a career in anaesthesia in 2022?". The survey was distributed to a randomly selected sample of 1225 anaesthetic consultants and completed by 470 respondents (38% response rate) with 793 free-text comments provided. Three overarching themes were identified: gender effects on the career and family interface; women do not fit the mould; and gender equity changes the status quo. Women respondents described a need to make a choice between career and family, which was not described by men, as well as stigmatisation of part-time work, a lack of access to challenging work and negative impacts of parental leave. Women respondents also described a sense of marginalisation within anaesthesia due to a 'boys' club' mentality, a lack of professional respect and insufficient structural supports for women in leadership. This was compounded for women from ethnically and culturally diverse backgrounds. A need for specific strategies to support anaesthetic careers for women was described as well as normalisation of flexibility in workplaces, combined with a broadening of our definition of success to allow people of all genders to experience fulfilment both at home and at work. This study is the first published qualitative data on factors affecting gender equity for anaesthetists in Australia and Aotearoa New Zealand. It highlights the need for further exploration, as well providing a foundation for changes in attitude and structural changes towards advancing gender equity.

A family reunion of "clinical cousins": Attachment and emotion in four family-oriented therapy models.

Attachment theory and the science of emotion provide a strong foundation for intervention at the family system level. Four therapeutic models in particular, Attachment-Based Family Therapy, Emotion-Focused Family Therapy, Dyadic Developmental Psychotherapy, and Emotionally Focused Family Therapy, demonstrate how a broad and accurate view of attachment relationships and emotion can be utilized to effectively intervene for a variety of presenting problems in a relational and empathic way for all involved. This paper continues a conversation that began at the Summit for Attachment and Emotion in Family Therapy in 2021 and aims to foster openness, collaboration, and affirmation between four different models of family therapy with shared theoretical roots. The presenters at the Summit and the authors of this paper view similarities across these models as validating and differences as opportunities to serve more families in unique ways, learning from one another's creativity to promote healing within families in the most effective and efficient ways possible. The paper frames the value of attachment theory and emotion science for family therapy, discusses the importance of learning from a variety of models with shared theoretical roots, presents brief summaries of the four models presented at the Summit, compares the models for similarities and complementarities, and shares highlights from each of the presenters from the Summit.

Insecticide Susceptibilities and Enzyme Activities of Four Stink Bug Populations in Mississippi, USA.

In Mississippi, the Pentatomidae complex infesting soybean is primarily composed of Euschistus servus, Nezara viridula, Chinavia hilaris, and Piezodorus guildinii. This study employed spray bioassays to evaluate the susceptibilities of these stink bugs to seven commonly used formulated insecticides: oxamyl, acephate, bifenthrin, λ-cyhalothrin, imidacloprid, thiamethoxam, and sulfoxaflor. Stinks bugs were collected from soybeans in Leland, MS, USA during 2022 and 2023, as well as from wild host plants in Clarksdale, MS. There was no significant difference in the susceptibility of C. hilaris to seven insecticides between two years, whereas P. guildinii showed slightly increased susceptibility to neonicotinoids in 2023. Among all four stink bug species, susceptibility in 2022 was ranked as P. guildinii ≤ C. hilaris ≈ N. viridula, while in 2023, it was ranked as P. guildinii ≤ C. hilaris ≤ E. Servus. Additionally, populations of E. servus and P. guildinii collected from Clarksdale exhibited high tolerance to pyrethroids and neonicotinoids. Moreover, populations of E. servus and P. guildinii from SIMRU-2022 and Clarksdale-2023 showed elevated esterase and cytochrome P450 activity, respectively. These findings from spray bioassays and enzyme activity analyses provide a baseline for monitoring insecticide resistance in Pentatomidae and can guide insecticide resistance management strategies for Mississippi soybean.