Deep sequencing of Phox2a nuclei reveals five classes of anterolateral system neurons.

The anterolateral system (ALS) is a major ascending pathway from the spinal cord that projects to multiple brain areas and underlies the perception of pain, itch, and skin temperature. Despite its importance, our understanding of this system has been hampered by the considerable functional and molecular diversity of its constituent cells. Here, we use fluorescence-activated cell sorting to isolate ALS neurons belonging to the Phox2a-lineage for single-nucleus RNA sequencing. We reveal five distinct clusters of ALS neurons (ALS1-5) and document their laminar distribution in the spinal cord using in situ hybridization. We identify three clusters of neurons located predominantly in laminae I-III of the dorsal horn (ALS1-3) and two clusters with cell bodies located in deeper laminae (ALS4 and ALS5). Our findings reveal the transcriptional logic that underlies ALS neuronal diversity in the adult mouse and uncover the molecular identity of two previously identified classes of projection neurons. We also show that these molecular signatures can be used to target groups of ALS neurons using retrograde viral tracing. Overall, our findings provide a valuable resource for studying somatosensory biology and targeting subclasses of ALS neurons.

Characterisation of anhydro-sialic acid transporters from mucosa-associated bacteria.

Sialic acid (Sia) transporters are critical to the capacity of host-associated bacteria to utilise Sia for growth and/or cell surface modification. While N-acetyl-neuraminic acid (Neu5Ac)-specific transporters have been studied extensively, little is known on transporters dedicated to anhydro-Sia forms such as 2,7-anhydro-Neu5Ac (2,7-AN) or 2,3-dehydro-2-deoxy-Neu5Ac (Neu5Ac2en). Here, we used a Sia-transport-null strain of Escherichia coli to investigate the function of members of anhydro-Sia transporter families previously identified by computational studies. First, we showed that the transporter NanG, from the Glycoside-Pentoside-Hexuronide:cation symporter family, is a specific 2,7-AN transporter, and identified by mutagenesis a crucial functional residue within the putative substrate-binding site. We then demonstrated that NanX transporters, of the Major Facilitator Superfamily, also only transport 2,7-AN and not Neu5Ac2en nor Neu5Ac. Finally, we provided evidence that SiaX transporters, of the Sodium-Solute Symporter superfamily, are promiscuous Neu5Ac/Neu5Ac2en transporters able to acquire either substrate equally well. The characterisation of anhydro-Sia transporters expands our current understanding of prokaryotic Sia metabolism within host-associated microbial communities.

PCSK9 inhibition attenuates alcohol-associated neuronal oxidative stress and cellular injury.

Alcohol Use Disorder (AUD) is a persistent condition linked to neuroinflammation, neuronal oxidative stress, and neurodegenerative processes. While the inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) has demonstrated effectiveness in reducing liver inflammation associated with alcohol, its impact on the brain remains largely unexplored. This study aimed to assess the effects of alirocumab, a monoclonal antibody targeting PCSK9 to lower systemic low-density lipoprotein cholesterol (LDL-C), on central nervous system (CNS) pathology in a rat model of chronic alcohol exposure. Alirocumab (50 mg/kg) or vehicle was administered weekly for six weeks in 32 male rats subjected to a 35 % ethanol liquid diet or a control liquid diet (n = 8 per group). The study evaluated PCSK9 expression, LDL receptor (LDLR) expression, oxidative stress, and neuroinflammatory markers in brain tissues. Chronic ethanol exposure increased PCSK9 expression in the brain, while alirocumab treatment significantly upregulated neuronal LDLR and reduced oxidative stress in neurons and brain vasculature (3-NT, p22phox). Alirocumab also mitigated ethanol-induced microglia recruitment in the cortex and hippocampus (Iba1). Additionally, alirocumab decreased the expression of pro-inflammatory cytokines and chemokines (TNF, CCL2, CXCL3) in whole brain tissue and attenuated the upregulation of adhesion molecules in brain vasculature (ICAM1, VCAM1, eSelectin). This study presents novel evidence that alirocumab diminishes oxidative stress and modifies neuroimmune interactions in the brain elicited by chronic ethanol exposure. Further investigation is needed to elucidate the mechanisms by which PCSK9 signaling influences the brain in the context of chronic ethanol exposure.

Something in Our Ears Is Oscillating, but What? A Modeller's View of Efforts to Model Spontaneous Emissions.

When David Kemp discovered "spontaneous ear noise" in 1978, it opened up a whole new perspective on how the cochlea works. The continuous tonal sound emerging from most healthy human ears, now called spontaneous otoacoustic emissions or SOAEs, was an unmistakable sign that our hearing organ must be considered an active detector, not just a passive microphone, just as Thomas Gold had speculated some 30 years earlier. Clearly, something is oscillating as a byproduct of that sensitive inbuilt detector, but what exactly is it? Here, we give a chronological account of efforts to model SOAEs as some form of oscillator, and at intervals, we illustrate key concepts with numerical simulations. We find that after many decades there is still no consensus, and the debate extends to whether the oscillator is local, confined to discrete local sources on the basilar membrane, or global, in which an assembly of micro-mechanical elements and basilar membrane sections, coupled by inner ear fluid, interact over a wide region. It is also undecided whether the cochlear oscillator is best described in terms of the well-known Van der Pol oscillator or the less familiar Duffing or Hopf oscillators. We find that irregularities play a key role in generating the emissions. This paper is not a systematic review of SOAEs and their properties but more a historical survey of the way in which various oscillator configurations have been applied to modelling human ears. The conclusion is that the difference between the local and global approaches is not clear-cut, and they are probably not mutually exclusive concepts. Nevertheless, when one sees how closely human SOAEs can be matched to certain arrangements of oscillators, Gold would no doubt say we are on the right track.

Pyoderma Gangrenosum in a 75-Year-Old Male With Follicular Diffuse Large B-cell Lymphoma.

Pyoderma gangrenosum (PG) is a rare neutrophilic dermatosis that characteristically presents with progressive ulcerative lesions. The association of PG with hematological malignancies remains unclear due to its varied clinical presentation. Herein, we report the unusual case of PG in a 75-year-old male with stage III follicular diffuse large B-cell lymphoma. Seven days subsequent to his first dose of rituximab, cyclophosphamide, doxorubicin, vincristine, prednisolone (R-CHOP) therapy, he presented to the emergency department with generalized malaise, bilateral lower extremity edema, and ecchymoses with ulcerative wounds on the dorsal of his feet. Due to the rapid progression of the patient's dermatological manifestations and declining clinical status, he required serial surgical wound debridement and a biopsy, which revealed an occlusive vasculopathy with dermal and epidermal necrosis. These pathological findings, along with the patient's clinical presentation, led to the diagnosis of PG. The patient was treated with negative pressure wound therapy, steroids, and tacrolimus ointment, which led to a marked improvement in the appearance of the patient's dermatological features and clinical status.

Extending intersectional multilevel analysis of individual heterogeneity and discriminatory accuracy (MAIHDA) to study individual longitudinal trajectories, with application to mental health in the UK.

The intersectional Multilevel Analysis of Individual Heterogeneity and Discriminatory Accuracy (MAIHDA) approach is gaining prominence in health sciences and beyond, as a robust quantitative method for identifying intersectional inequalities in a range of individual outcomes. However, it has so far not been applied to longitudinal data, despite the availability of such data, and growing recognition that intersectional social processes and determinants are not static, unchanging phenomena. Drawing on intersectionality and life course theories, we develop a longitudinal version of the intersectional MAIHDA approach, allowing the analysis not just of intersectional inequalities in static individual differences, but also of life course trajectories. We discuss the conceptualization of intersectional groups in this context: how they are changeable over the life course, appropriate treatment of generational differences, and relevance of the age-period-cohort identification problem. We illustrate the approach with a study of mental health using United Kingdom Household Longitudinal Study data (2009-2021). The results reveal important differences in trajectories between generations and intersectional strata, and show that trajectories are partly multiplicative but mostly additive in their intersectional inequalities. This article provides an important and much needed methodological contribution, enabling rigorous quantitative, longitudinal, intersectional analyses in social epidemiology and beyond.

A tutorial for conducting intersectional multilevel analysis of individual heterogeneity and discriminatory accuracy (MAIHDA).

Intersectional multilevel analysis of individual heterogeneity and discriminatory accuracy (I-MAIHDA) is an innovative approach for investigating inequalities, including intersectional inequalities in health, disease, psychosocial, socioeconomic, and other outcomes. I-MAIHDA and related MAIHDA approaches have conceptual and methodological advantages over conventional single-level regression analysis. By enabling the study of inequalities produced by numerous interlocking systems of marginalization and oppression, and by addressing many of the limitations of studying interactions in conventional analyses, intersectional MAIHDA provides a valuable analytical tool in social epidemiology, health psychology, precision medicine and public health, environmental justice, and beyond. The approach allows for estimation of average differences between intersectional strata (stratum inequalities), in-depth exploration of interaction effects, as well as decomposition of the total individual variation (heterogeneity) in individual outcomes within and between strata. Specific advice for conducting and interpreting MAIHDA models has been scattered across a burgeoning literature. We consolidate this knowledge into an accessible conceptual and applied tutorial for studying both continuous and binary individual outcomes. We emphasize I-MAIHDA in our illustration, however this tutorial is also informative for understanding related approaches, such as multicategorical MAIHDA, which has been proposed for use in clinical research and beyond. The tutorial will support readers who wish to perform their own analyses and those interested in expanding their understanding of the approach. To demonstrate the methodology, we provide step-by-step analytical advice and present an illustrative health application using simulated data. We provide the data and syntax to replicate all our analyses.

Clarifications on the intersectional MAIHDA approach: A conceptual guide and response to Wilkes and Karimi (2024).

Intersectional Multilevel Analysis of Individual Heterogeneity and Discriminatory Accuracy (MAIHDA) has been welcomed as a new gold standard for quantitative evaluation of intersectional inequalities, and it is being rapidly adopted across the health and social sciences. In their commentary "What does the MAIHDA method explain?", Wilkes and Karimi (2024) raise methodological concerns with this approach, leading them to advocate for the continued use of conventional single-level linear regression models with fixed-effects interaction parameters for quantitative intersectional analysis. In this response, we systematically address these concerns, and ultimately find them to be unfounded, arising from a series of subtle but important misunderstandings of the MAIHDA approach and literature. Since readers new to MAIHDA may share confusion on these points, we take this opportunity to provide clarifications. Our response is organized around four important clarifications: (1) At what level are the additive main effect variables defined in intersectional MAIHDA models? (2) Do MAIHDA models have problems with collinearity? (3) Why does the Variance Partitioning Coefficient (VPC) tend to be small, and the Proportional Change in Variance (PCV) tend to be large in MAIHDA? and (4) What are the goals of MAIHDA analysis?

Lights, Sirens, and Load: Anticipatory emergency medical treatment planning causes cognitive load during emergency response driving among paramedicine students.

Paramedics face various unconventional and secondary task demands while driving ambulances, leading to significant cognitive load, especially during lights-and-sirens responses. Previous research suggests that high cognitive load negatively affects driving performance, increasing the risk of accidents, particularly for inexperienced drivers. The current study investigated the impact of anticipatory treatment planning on cognitive load during emergency driving, as assessed through the use of a driving simulator. We recruited 28 non-paramedic participants to complete a simulated baseline drive with no task and a cognitive load manipulation using the 1-back task. We also recruited 18 paramedicine students who completed a drive while considering two cases they were travelling to: cardiac arrest and infant seizure, representing varying difficulty in required treatment. The results indicated that both cases imposed considerable cognitive load, as indicated by NASA Task Load Index responses, comparable to the 1-back task and significantly higher than driving with no load. These findings suggest that contemplating cases and treatment plans may impact the safety of novice paramedics driving ambulances for emergency response. Further research should explore the influence of experience and the presence of a second individual in the vehicle to generalise to broader emergency response driving contexts.

Major Psychiatric Disorders, Substance Use Behaviors, and Longevity.

Importance: Observational studies suggest that major psychiatric disorders and substance use behaviors reduce longevity, making it difficult to disentangle their relationships with aging-related outcomes. Objective: To evaluate the associations between the genetic liabilities for major psychiatric disorders, substance use behaviors (smoking and alcohol consumption), and longevity. Design, Settings, and Participants: This 2-sample mendelian randomization (MR) study assessed associations between psychiatric disorders, substance use behaviors, and longevity using single-variable and multivariable models. Multiomics analyses were performed elucidating transcriptomic underpinnings of the MR associations and identifying potential proteomic therapeutic targets. This study sourced summary-level genome-wide association study (GWAS) data, gene expression, and proteomic data from cohorts of European ancestry. Analyses were performed from May 2022 to November 2023. Exposures: Genetic susceptibility for major depression (n = 500 199), bipolar disorder (n = 413 466), schizophrenia (n = 127 906), problematic alcohol use (n = 435 563), weekly alcohol consumption (n = 666 978), and lifetime smoking index (n = 462 690). Main Outcomes and Measures: The main outcome encompassed aspects of health span, lifespan, and exceptional longevity. Additional outcomes were epigenetic age acceleration (EAA) clocks. Results: Findings from multivariable MR models simultaneously assessing psychiatric disorders and substance use behaviorsm suggest a negative association between smoking and longevity in cohorts of European ancestry (n = 709 709; 431 503 [60.8%] female; ß, -0.33; 95% CI, -0.38 to -0.28; P = 4.59 × 10-34) and with increased EAA (n = 34 449; 18 017 [52.3%] female; eg, PhenoAge: ß, 1.76; 95% CI, 0.72 to 2.79; P = 8.83 × 10-4). Transcriptomic imputation and colocalization identified 249 genes associated with smoking, including 36 novel genes not captured by the original smoking GWAS. Enriched pathways included chromatin remodeling and telomere assembly and maintenance. The transcriptome-wide signature of smoking was inversely associated with longevity, and estimates of individual smoking-associated genes, eg, XRCC3 and PRMT6, aligned with the smoking-longevity MR analyses, suggesting underlying transcriptomic mediators. Cis-instrument MR prioritized brain proteins associated with smoking behavior, including LY6H (ß, 0.02; 95% CI, 0.01 to 0.03; P = 2.37 × 10-6) and RIT2 (ß, 0.02; 95% CI, 0.01 to 0.03; P = 1.05 × 10-5), which had favorable adverse-effect profiles across 367 traits evaluated in phenome-wide MR. Conclusions: The findings suggest that the genetic liability of smoking, but not of psychiatric disorders, is associated with longevity. Transcriptomic associations offer insights into smoking-related pathways, and identified proteomic targets may inform therapeutic development for smoking cessation strategies.

Host and pathogen factors that influence variability of Mycobacterium tuberculosis lipid body content in sputum from patients with tuberculosis: an observational study.

BACKGROUND: High proportions of Mycobacterium tuberculosis cells in sputum containing triacylglycerol-rich lipid bodies have been shown to be associated with treatment failure or relapse following antituberculous chemotherapy. Although lipid body determination is a potential biomarker for supporting clinical trial and treatment decisions, factors influencing variability in sputum frequencies of lipid body-positive (%LB+) M tuberculosis in patients are unknown. We aimed to test our hypothesis that exposure to host-generated NO and M tuberculosis strains are factors associated with differences in sputum %LB+. METHODS: In this observational study, we determined %LB+ frequencies before treatment by microscopy in patients with smear-positive tuberculosis from two separate prospective observational study settings (Gondar, Ethiopia, recruited between May 1, 2010, and April 30, 2011, and Fajara, The Gambia, who provided sputum samples before treatment between May 5, 2010, and Dec 22, 2011). In Ethiopia, fractional exhaled nitric oxide (FeNO) was measured as a biomarker of host NO, and M tuberculosis strain differences were determined by spoligotyping. Treatment response was assessed by percentage weight change after 7 months. In The Gambia, treatment responses were assessed as change in BMI and radiographic burden of disease after 6 months. Sputum M tuberculosis isolates were studied in vitro for their %LB+ and triacylglycerol synthase 1 (tgs1) mRNA responses to NO exposure. Propidium iodide staining was used as a measure of NO strain toxicity. Correlation between in vitro %LB+ frequencies following NO exposure and those of the same strain in sputum was examined with linear regression and Dunnett's multiple comparison test. FINDINGS: In Ethiopia, 73 patients who were smear positive for pulmonary tuberculosis were recruited (43 [59%] were male and 30 [41%] were female). Of these, the %LB+ in the sputum of 59 patients showed linear correlation with log10 FeNO (r2=0·28; p<0·0001) and an association with strain spoligotype was suggested. Seven M tuberculosis strains from The Gambia showed different dose-responses to NO in vitro, demonstrated by changing lipid body content, tgs1 transcription, and bacterial toxicity. In sputum %LB+ frequencies correlated with in vitro %LB+ responses to NO of the corresponding isolate. In a subset of 34 patients across both cohorts, higher sputum %LB+ frequencies before treatment were associated with weaker responses to treatment than lower sputum %LB+ frequencies. INTERPRETATION: M tuberculosis strain and exposure to host-generated NO are associated with sputum %LB+. Our results support the use of M tuberculosis strain-dependent sputum %LB+ as a predictive biomarker of treatment response. FUNDING: The Medical Research Council, the University of Leicester, and the University of Gondar.

Comparative sanitation data from high-frequency phone surveys across 3 countries.

With less than half of the world's urban population having safely managed sanitation due to the high cost and difficulty of building sewers and treatment plants, many rely on off-grid options like pit latrines and septic tanks, which are hard to empty and often lead to illegal waste dumping; this research focuses on container-based sanitation (CBS) as an emerging off-grid solution. Off-grid sanitation refers to waste management systems that operate independently of centralized infrastructure and CBS is a service providing toilets that collect human waste in sealable containers, which are regularly emptied and safely disposed of. These data relate to a project investigating CBS in Kenya, Peru, and South Africa, focusing on how different user groups access and utilize sanitation - contrasting CBS with other types. Participants, acting as citizen scientists, collected confidential data through a dedicated smartphone app designed by the authors and external contractors. This project aimed to explore the effective scaling, management, and regulation of off-grid sanitation systems, relevant to academics in urban planning, water and sanitation services, institutional capability, policy and governance, and those addressing inequality and poverty reduction. The 12-month data collection period offered participants small incentives for weekly engagement, in a micro payment for micro tasks approach. Participants were randomly selected, attended a training workshop, and (where needed) were given a smartphone which they could keep at the end of the project. We conducted weekly smartphone surveys in over 300 households across informal settlements. These surveys aimed to understand human-environment interactions by capturing daily life, wellbeing, income, infrastructural service use, and socioeconomic variables at a weekly resolution, contributing to more informed analyses and decision-making. The smartphone-based approach offers efficient, cost-effective, and flexible data collection, enabling extensive geographical coverage, broad subject areas, and frequent engagement. The Open Data Kit (ODK) tools were used to support data collection in the resource-constrained environment with limited or intermittent connectivity.

Increased Synthetic Cytotoxicity of Combinatorial Chemoradiation Therapy in Homologous Recombination Deficient Tumors.

PURPOSE: Homologous recombination deficient (HRD) tumors are exquisitely sensitive to platinum-based chemotherapy and when combined with radiation therapy (RT), leads to improved overall survival in multiple cancer types. Whether a subset of tumors with distinct molecular characteristics demonstrate increased benefit from cisplatin and RT (c-RT) is unclear. We hypothesized that HRD tumors, whether associated with BRCA mutations or genomic scars of HRD, exhibit exquisite sensitivity to c-RT, and that HRD may be a significant driver of c-RT benefit. METHODS AND MATERIALS: Sensitivity to c-RT was examined using isogenic and sporadic breast cancer cell lines. HRD was assessed using 4 assays: RT-induced Rad51 foci, a DR-GFP reporter assay, a genomic scar score (large-scale state transitions [LST]), and clonogenic survival assays. Whole-genome sequencing of 4 breast tumors from a phase 2 clinical trial of neoadjuvant c-RT in triple-negative breast cancer was performed and HRD was defined using HRDetect. RESULTS: BRCA1/2 deficient cell lines displayed functional HRD based on the Rad51 functional assay, with c-RT to RT or cisplatin interaction ratios (IR) of 1.11 and 26.84 for the BRCA1 isogenic pair at 2 µM cisplatin and 6 Gy, respectively. The highest LST lines demonstrated HRD and synthetic cytotoxicity to c-RT with IR at 2 Gy and cisplatin 20 µM of 7.50, and the lowest LST line with IR of 0.65. Of 4 evaluable patients in the phase 2 trial, one achieved a pathologic complete response with corresponding HRD based on multiple genomic scar scores including HRDetect and LST scores, compared with patients without a pathologic complete response. CONCLUSIONS: HRD breast cancers, whether identified by BRCA1/2 mutation status, functional tests, or mutational signatures, appear to be significantly more sensitive to c-RT compared with isogenic controls or tumors without HRD mutational signatures. HRD tumors may be exquisitely sensitive to c-RT which warrants further clinical investigation to guide a precision oncology approach.