Signal Transduction and Targeted Therapy

There is an unmet need to identify biomarkers of active thyroid eye disease (TED). scRNAseq revealed that orbital fibroblasts from orbital decompressions in people with TED express high levels of thyroid hormone receptors, growth factor receptors, including insulin-like growth factor 1 receptor (IGF1R), and extracellular matrix proteins including SPARC (osteonectin), whereas orbital fat endothelial cells expressed thyroid peroxidase (TPO). SPARC was significantly raised in the serum of people with thyroid disease compared to healthy controls. Furthermore, those with moderate, severe and sight threatening TED had higher SPARC levels than those with thyroid disease but free of TED or mild TED. Free-triiodothyronine (FT3) levels were positively correlated with SPARC in moderate-sight threatening TED. SPARC and IGF1 were positively correlated across people with thyroid disease alone, as well as TED. Thyroid stimulating hormone (TSH) levels were negatively correlated with SPARC in moderate-sight threatening TED. When participants were followed longitudinally, SPARC decreased after the active phase of TED. At the protein level, immunohistochemistry indicated that SPARC was heterogeneously expressed by fibroblasts in both control and TED orbital fat. SPARC is a key mediator of fibrosis and deposition of extracellular matrix and the correlation of SPARC serum levels to TED status and FT3 make it a promising biomarker of active TED.

ObjectivePost-thrombotic syndrome (PTS), a common complication of deep vein thrombosis, lacks objective diagnostic biomarkers and its molecular mechanisms remain poorly understood. This study aimed to identify plasma biomarkers and clarify pathways using integrated multi-omics and machine learning. MethodsProteomic and metabolomic profiling of 75 PTS patients and 75 controls was performed. Differential expression analysis, pathway enrichment, and protein-metabolite network analysis were conducted. A multi-algorithm machine learning with 8 feature selection methods prioritized biomarkers. Validations and 14 models were assessed. Results1,104 proteins and 1,891 metabolites were differentially expressed. Citrate cycle and unsaturated fatty acid biosynthesis were enriched. Three proteins, namely DIP2B, KNG1, and SUCLG2, were consistently selected as core biomarkers. All of these proteins were significantly downregulated in PTS and externally validated. A random forest model utilizing these proteins achieved an accuracy of 97.7% in independent testing, with SUCLG2 being the most influential predictor. ConclusionThis study identifies a novel three - protein biomarker panel for the diagnosis of PTS and reveals an immunometabolic axis in the pathogenesis of PTS, which links inflammatory regulation with mitochondrial energy metabolism. These findings provide valuable insights into the development of diagnostic tools and targeted therapeutic approaches.

PurposeSepsis-associated immunothrombosis significantly contributes to high mortality, yet the role of N-glycosylation in this process remains poorly understood. This study aimed to comprehensively profile the plasma N-glycosylation landscape in sepsis and elucidate how its specific reprogramming in the complement and coagulation cascades influences immunothrombotic balance and patient outcomes. MethodsWe performed in-depth 4D-DIA proteomic and N-glycomic analyses on plasma from 43 sepsis patients and 9 healthy controls. Differential expression, weighted gene co-expression network analysis (WGCNA), and protein-glycosylation correlation analyses were used to characterize molecular features. Clinical relevance was assessed via correlation and survival analyses. ResultsExtensive N-glycosylation reprogramming was observed in sepsis plasma,with marked enrichment in complement and coagulation pathways(KEGG p=7.76x10- {superscript 2}{superscript 1}).Pro-coagulant proteins(eg,vWF,fibrinogen)showed increased abundance together with enhanced site-specific glycosylation,potentially amplifying their activity.In contrast,key anticoagulant proteins(eg,SERPINC1)displayed unchanged glycosylation at critical sites despite abundance changes,which may impair function.Survival analysis revealed distinct prognostic values of glycoproteins and specific glycosylation sites.For instance,high vWF protein levels predicted mortality(HR=2.83),whereas elevated glycosylation at vWF N211 was associated with improved survival(HR=0.135),suggesting a negative regulatory role.These glycosylation markers correlated closely with disease severity and prognosis,representing potential early-warning biomarkers independent of current clinical coagulation indicators. ConclusionOur study demonstrates widespread reprogramming of the plasma proteome and N-glycome in sepsis.We propose that decoupling of protein function from abundance through N-glycosylation in the complement-coagulation network contributes to immunothrombotic imbalance.Specific N-glycosylation sites may serve as novel prognostic biomarkers,offering new perspectives for early risk stratification and glycosylation-targeted therapies in sepsis. Key PointsO_LISepsis plasma exhibits specific N-glycosylation reprogramming overwhelmingly focused on the complement and coagulation cascade. C_LIO_LIA dominant "glycosylation-dominated co-upregulation" mode in procoagulant factors, coupled with a "silent" glycosylation state in key anticoagulants, drives prothrombotic imbalance. C_LIO_LISite-specific N-glycosylation levels provide prognostic information distinct from, and often superior to, their carrier protein abundance, offering novel early-risk biomarkers. C_LI

ObjectiveMyositis-associated interstitial lung disease (myositis-ILD) consists of two predominant radiologic patterns of lung injury--nonspecific interstitial pneumonia (NSIP) and organizing pneumonia (OP)--that oftentimes coexist. However, it remains unclear whether either is associated with clinical outcomes. We aimed to assess the therapeutic response in patients with NSIP-compared to those with OP-predominant myositis-ILD. MethodsThis retrospective, single-center cohort study recruited participants from the Northwestern University ILD Registry with a circulating myositis-associated antibody, ILD, and at least 6 months of follow-up while on immunomodulatory therapy during a 24-month observation period after diagnosis. Two thoracic radiologists determined the predominant radiologic pattern (NSIP or OP). The primary outcome was the absolute change in forced vital capacity (FVC) at 24 months post-diagnosis. Secondary outcomes included changes in the diffusing capacity of the lung for carbon monoxide (DLCO) and radiologic qualitative and quantitative measures of lung injury. ResultsForty-one participants were included in analyses. 71% had an OP-predominant while 29% had an NSIP-predominant radiologic pattern of lung injury. Both exposure cohorts had improvement in mean absolute FVC (OP cohort = +0.18L [p=0.005], NSIP cohort = +0.24L [p=0.07]) over the 24-month observation period. The OP (p<0.05) but not the NSIP cohort (p=0.20) had an increase in DLCO. The OP cohort demonstrated improvement in the qualitative assessment of follow-up imaging (p<0.05), driven by quantitative improvement in groundglass/consolidative opacities (p=0.006). A subset of participants demonstrated features of NSIP/OP overlap and had greater baseline radiologic severity of lung injury. ConclusionPatients with circulating myositis-associated antibodies and an OP-predominant pattern of lung injury may have a more favorable response to therapy than those with NSIP. Further studies are needed to validate our findings and delineate other features cognate with these associations. Significance and InnovationsO_LIRadiologic phenotyping may predict therapeutic response in myositis-ILD. This study demonstrates that an OP-predominant computed tomography (CT) pattern of lung injury is associated with greater improvement in lung function and radiologic signs of inflammation over 24 months on at least 6 months of immunomodulatory therapy compared with an NSIP-predominant pattern, suggesting that CT pattern may provide clinically meaningful prognostic information. C_LIO_LIFirst study to integrate blinded qualitative radiologic adjudication with quantitative CT scoring in myositis-ILD. By combining dual-radiologist review with Kazerooni quantitative scoring and longitudinal pulmonary function testing, this study offers a rigorous and multidimensional assessment of treatment response. C_LIO_LIExpands risk stratification beyond antibody-based toward imaging-based phenotyping strategies. In a heterogeneous population defined by diverse myositis-associated antibodies, this work introduces radiologic pattern as a practical and accessible framework for anticipating treatment responsiveness. C_LIO_LIProvides hypothesis-generating data for precision management in myositis-ILD. The findings support the concept that imaging-defined subgroups may exhibit differential therapeutic trajectories, laying groundwork for future multicenter studies integrating CT phenotype, antibody profile, and treatment strategy. C_LI

Respiratory monitoring in daily-life settings is important for health assessment, yet extracting physiologically interpretable information from breathing signals under natural conditions remains challenging, as breathing is inherently dynamic and strongly modulated by behavior. Here, a portable breathing monitoring device based on a flexible lead zirconate titanate sensor is developed to address this challenge. By exploiting polarity-opposed piezoelectric and pyroelectric responses through sensor orientation, the recorded breathing waveform exhibits a characteristic dual-component structure, consisting of a narrow transient spike followed by a broad quasi-steady peak within each breathing phase. This intrinsic waveform structure enables phase-resolved quantification of how breathing effort is distributed between transient and quasi-steady components during inhalation and exhalation. Pilot measurements in healthy subjects and patients with chronic obstructive pulmonary disease or asthma reveal systematic shifts toward transient-enhanced breathing in patients, providing clearer differentiation than conventional descriptors based on breathing duration or amplitude. By transforming complex breathing dynamics into stable and physiologically meaningful signal components under daily-life conditions, this dual-response sensing approach enables more robust access to function-related changes in natural breathing.

This exploratory analysis of PAX LC, a Phase 2, 1:1 randomized, double-blind, superiority, placebo-controlled trial examined whether treatment with nirmatrelvir/ritonavir (NMV/r) versus placebo/ritonavir (PBO/r) in individuals with Long COVID could reveal immune features associated with symptom improvement. Eighty-two participants (n=45 PBO/r; n=37 NMV/r) provided blood samples at baseline (Day 0) and post-treatment (Day 28). Baseline demographic and immunological phenotypes were similar in the two groups. No significant differences were observed in major immune cell populations or organ function markers between NMV/r vs. PBO/r groups, or before vs. after the treatment. Modest hematologic changes were noted in the NMV/r arm. SARS-CoV-2-specific IgG levels remained constant, with changes in total immunoglobulin subtypes and isotypes in both arms. Both arms showed similar shifts in cytokine levels. Notably, the levels of S1 and Spike proteins in circulation remained unchanged post-treatment. Regardless of the treatment arm, participants with self-reported symptom improvement showed reductions in the level of the inflammatory chemokine RANTES. Taken together, the findings of this study demonstrate limited virological and immunological changes in response to nirmatrelvir, contributing insights into the reason for the lack of benefit of the 15-day NMV/r treatment in Long COVID.

Background and AimsThe prognosis of interstitial lung diseases (ILDs) other than idiopathic pulmonary fibrosis (IPF) has not been studied as extensively as IPF. This study aimed to evaluate baseline factors associated with mortality in non-IPF ILD, including demographic characteristics, respiratory function test (RFT), comorbidities, and ILD subtypes. MethodsThis retrospective cohort study analysed prospectively collected data of patients with non-IPF ILD at a single tertiary centre in Malaysia (2010-2023). Patients without baseline RFT or HRCT were excluded. Survival was assessed using Kaplan-Meier analysis, and mortality predictors were identified using Cox regression. ResultsThe mean age was 60 {+/-} 15 years, with a male-to-female ratio of 1:3. Indian ethnicity constituted the largest group (n = 109, 47.6%). The mean baseline forced vital capacity (FVC) was 53.3 {+/-} 21% predicted. An FVC <50% predicted, age [&ge;]50 years at diagnosis, specific ILD subtypes, and ethnicity were independently associated with mortality. Compared with Malays, both Chinese (hazard ratio [HR] 9.86, 95% confidence interval [CI] 1.27-76.89, p = 0.037) and Indians (HR 8.59, 95% CI 1.14-64.69, p = 0.001) were associated with a higher risk of death. Kaplan-Meier analysis demonstrated significant differences in survival across non-IPF ILD subtypes (log-rank p = 0.048), with hypersensitivity pneumonitis showing the poorest prognosis (mean survival 6.1 years). ConclusionEthnicity emerged as an independent prognostic factor for mortality in non-IPF ILD. The underlying mechanisms remain unclear and may reflect differences in genetic variation, cultural factors, or environmental exposures. Larger prospective studies are required to validate these findings.

Background The Hump-nosed pit viper is a recognized but neglected medically significant species causing morbidity and mortality, with non-availability of a specific antivenom. There are many gaps in our understanding of its envenomation, including burden, clinical syndrome, complications and management. Methodology The study is a retrospective sub analysis of the Prospective VENOMS registry and hospital records of Hump Nosed Pit Viper envenomation from a single tertiary care center in coastal Karnataka from May 2018 to March 2024. Epidemiology, syndrome, complications and treatment strategies have been described. A linear mixed model analysis was conducted to study the effect of different therapeutic interventions in combating venom induced consumptive coagulopathy (VICC) Principal Findings Of 46 cases, 24 patients had VICC. The most common complications were AKI (21.7%), TMA (10.9%) and stroke (4.4%). Anaphylaxis to ASV (23.9%) was the most common therapeutic complication. Therapeutic interventions included ASV, administration of blood products and therapeutic plasma exchange along with supportive care. The linear mixed model revealed that administration of blood products (p=<0.001) had the strongest influence on the INR value, however, often resulting in a transient decline in INR value. ASV (p=0.052) caused only marginally significant change in INR. The role of TPE could not be statistically inferred, however, individual cases with severe VICC improved without complications, therefore it required further study but can be considered in critical cases. Conclusions/Significance This study describes the syndrome of hump-nosed pit viper envenomation, while highlighting the urgent need for a species-specific antivenom, recommends treatment strategies that can be used in the interim. Additionally, geo-spatial mapping draws attention to hotspots and the hypothesis that HNPV in coastal Karnataka have regionally distinct toxicity trends.

Newly emerging influenza virus strains pose a constant threat as they encounter a population lacking neutralizing antibodies against the new strain. However, cross-reactive non-neutralizing antibodies (nnABs) may be present and assist in mitigating disease symptoms via various effector mechanisms, including antibody-dependent cellular cytotoxicity (ADCC). Although nnABs to influenza virus have received more attention lately, little information is available on their age-related prevalence, steady-state levels, functional properties, and changes in these parameters over time. Using longitudinal samples from adolescents, adults, and older adults, collected before and after the 2009 swine flu pandemic, we comprehensively characterized the specificity and functionality of nnAB responses against H1N1 pandemic 2009 (H1N1pdm09) virus. Remarkably, all participants exhibited cross-reactive antibodies to this virus before having encountered it through infection or vaccination, with the highest baseline levels observed in older adults. The levels of these IgG antibodies showed a strong correlation with engagement of fragment crystallizable {gamma} receptor IIIa (Fc{gamma}RIIIa) and ADCC activity, both of which were notably lower in adolescents compared to adults and older adults. Without infection or vaccination, average amounts of H1N1pdm09-reactive antibodies remained relatively stable on population level over the 5-year study period. However, on an individual level, substantial increases and decreases occurred. H1N1pdm09 infection or vaccination significantly enhanced specific antibody levels and the Fc{gamma}RIIIa-engaging capacity of these antibodies in all age groups. ADCC-mediating antibodies increased however only in adolescents, reaching the same level as observed in the adult groups. Taken together, our results demonstrate the presence of cross-reactive, non-neutralizing, functional, and boostable antibodies against a never-encountered influenza virus strain across all age groups. These antibodies can potentially contribute to protection from severe disease. Accordingly, in case of a newly emerging virus, their further enhancement by vaccination could be beneficial as an immediate protective measure before a strain-specific vaccine becomes available. Author summaryNearly everyone has contracted influenza and/or has been vaccinated against influenza several times over the years. While the antibodies raised during these earlier encounters will not prevent infection by a newly emerging influenza virus strain, they can help to protect from severe disease. Therefore, it is important to determine the prevalence and quantity of these antibodies, understand their mechanisms of action, assess their persistence over time, and examine potential age-related differences in these parameters. We studied antibody responses to the H1N1pdm09 virus in blood samples of young, adult, and older adult individuals from a large cohort study. Irrespective of age, all blood samples contained antibodies that reacted with a never-before-encountered influenza virus strain. The amounts of these antibodies were initially lower in adolescents but with time increased, reaching the same levels as observed in adults. Importantly, infection with or vaccination against the new virus strengthened the responses in all age groups. We conclude that boosting such broadly-reactive antibodies through vaccination could serve as an immediate strategy when a new virus emerges, buying critical time to develop a more specific vaccine.

BackgroundSevere Fever with Thrombocytopenia Syndrome (SFTS) is an acute infectious disease with high mortality. This study aimed to develop a quantitative scoring system for grading SFTS severity using dynamic clinical data. MethodsA retrospective study included 547 confirmed SFTS patients from two hospitals. Clinical data were collected over a 14-day course (divided into four phases). Patients were grouped into survivors (n=451) and non-survivors (n=96). Statistical analyses, including Kaplan-Meier curves and log-rank tests, were performed. An external validation cohort of 44 new patients was used to validate the scoring system via C-statistic, calibration curves, and decision curve analysis (DCA). ResultsOf 547 patients, 96 (17.55%) were non-survivors. Multivariate logistic regression identified six independent prognostic factors across phases: age, platelet (PLT), aspartate aminotransferase (AST), and creatinine (Cr) (days 5-7); age, red blood cell distribution width (RDW), Cr, and lactate dehydrogenase (LDH) (days 8-10); Cr and LDH (days 11-14). A scoring system (0-11 points) was developed, stratifying patients into low (0-3), intermediate (4-7), and high (8-11) risk groups, with adverse outcome rates of 1.04%, 22.92%, and 76.04%, respectively. Kaplan-Meier curves showed significant prognostic differences (log-rank P<0.001). External validation (44 cases) confirmed excellent performance: AUC 0.810-0.952, good calibration (Hosmer-Lemeshow P>0.05), and net clinical benefit (DCA Eavg 0.068-0.098, Emax 0.422-0.559). ConclusionA dynamic SFTS severity scoring system was developed and validated. Internal and external validation confirmed its reliability and clinical utility, providing a simple, practical tool for timely assessment and early intervention.

BackgroundMultiple sclerosis (MS) is an immune-mediated demyelinating disease of the central nervous system affecting 2.8 million people worldwide. Both genetic and environmental factors contribute to MS risk, with Epstein-Barr virus (EBV) infection being an important environmental factor. To better clarify the role of EBV in MS, we examined its impact on gene expression, chromatin accessibility, and transcription factor binding in primary B cells and EBV-transformed B cells derived from patients with MS and healthy controls. ResultsRNA-seq and ATAC-seq analyses revealed extensive MS-dependent gene expression and chromatin accessibility differences in EBV-transformed, but not in primary B cells. These changes are largely accounted for by the expression levels of EBNA2, an EBV-encoded transcriptional regulator previously implicated in MS. ChIP-seq analysis revealed that EBNA2 binding with its interacting human partners RBPJ, EBF1, and PU.1 is highly enriched at MS genetic risk loci, with extensive EBNA2 allelic binding and increased enrichment at MS genetic risk loci in MS-derived cells. ConclusionsOur findings demonstrate that enhanced EBNA2 activity in MS alters human gene expression, chromatin accessibility, and transcription factor binding in an MS-dependent manner. Collectively, this study provides new insights into the molecular mechanisms through which EBV, particularly EBNA2, interacts with host genetic risk to contribute to MS pathogenesis.

BackgroundBoth hemorrhagic fever with renal syndrome (HFRS) and scrub typhus (ST) are acute zoonotic infectious diseases. There is an overlap in their epidemiological characteristics and clinical manifestations, posing challenges for early differential diagnosis. This study aims to identify predictive factors for these two diseases to provide a basis for early diagnosis. Method/FindingsA retrospective analysis was conducted on the clinical data of patients diagnosed with HFRS and ST at the First Affiliated Hospital of Dali University. Logistic regression analysis was employed to explore independent risk factors for the early differential diagnosis of these two diseases, and a nomogram model was constructed based on these risk factors. The performance of the model was evaluated using the area under the receiver operating characteristic curve (AUC). The nomogram was utilized to visually present the predictive variables. Decision curve analysis (DCA) was performed to assess the clinical utility of the model. ResultsA total of 235 patients each with HFRS and ST were included in this study. After adjusting for confounding factors, the results of multivariate logistic regression analysis revealed that sex (male) (adjusted odds ratio [ajOR]: 2.093, 95% confidence interval [CI]: 1.107 - 3.957, P = 0.018), positive proteinuria (ajOR: 4.937, 95% CI: 2.427 - 10.042, P < 0.001), creatinine (CREA) (ajOR: 1.009, 95% CI: 1.003 - 1.015, P = 0.005), heart rate (ajOR: 0.981, 95% CI: 0.966 - 0.997, P = 0.018), and conjunctival congestion (ajOR: 16.167, 95% CI: 5.326 - 49.072, P < 0.001) were independent risk factors for differentiating HFRS from ST. The AUC of the model constructed based on these five independent risk factors was 0.856. ConclusionSex (male), positive proteinuria, elevated CREA, decreased heart rate, and conjunctival congestion are effective predictive factors.

BackgroundDengue virus (DENV) appears to manipulate several cellular metabolic pathways to permit its replication and immune evasion in the host. Here, we employed high-resolution mass spectrometry (HR-MS) to investigate the serum metabolomic landscape of clinical DENV infection. MethodsSerum specimens from primary dengue (n=11), secondary dengue (n=9) samples, and healthy controls (n=10) were used for untargeted and targeted metabolomic quantification on a Waters Xevo G2-XS QTof Mass Spectrometer. The binding potential of selected ligands against DENV NS1, NS3, and NS5 was evaluated. Crystal structures were retrieved from Protein Data Bank and prepared using the Schrodingers protein preparation wizard. Based on findings from untargeted metabolomics, we validated certain bioactive lipid metabolites using commercial enzyme immunoassays. ResultsSerum metabolomic profiling revealed multiple distinct patterns for primary and secondary dengue versus controls. A consistent peak was observed at 2.06 mins across all samples. Certain bioactive lipid metabolites, such as, lysophospholipids, phosphatidylcholines, phosphatidylserines, and phosphatidylinositols, were detected alongside carnitine fragments, ceramides, diacylglycerols (DAGs), and bile acid conjugates in dengue. Molecular docking showed that DAG consistently exhibited strong binding to all the DENV proteins. Notably, LPC 22:6 showed a selectively strong affinity for NS5. Enzyme validation showed that in the secondary dengue cohort, LPC was significantly elevated than primary and healthy controls (p<0.05). ConclusionsOur investigations of the metabolomic landscaping, unveiled certain characteristic anabolic shift revealing metabolic vulnerabilities in clinical DENV infection, warranting investigations for use as potential biomarkers of inflammation in disease diagnosis and prognosis. Author summaryDengue is a mosquito-borne tropical viral infection that can range in severity from asymptomatic to life-threatening manifestations. Dengue virus (DENV) hijacks cellular machinery to sustain its survival in the host. Using high-resolution mass spectrometry (HR-MS), we studied the serum metabolomic imprints of dengue infection. The binding ability of selected metabolomic ligands against DENV NS1, NS3, and NS5 was studied. We found several distinct retention patterns for the dengue cases, with a consistent peak at 2.06 min across all samples. Further, several bioactive lipid metabolites were detected in the dengue infected cohort. Our molecular docking studies showed that diacylglycerol, a lipid metabolite exhibited strong binding with all the DENV proteins. We concluded that certain unique lipid metabolomic imprints exist in clinical DENV infection. The identified metabolomic signatures reveal significant potential for metabolomics to elucidate host-virus interactions, contributing to the advancement of antiviral and symptomatic treatments, along with prognostic or diagnostic biomarkers of dengue disease.

BackgroundThere is a need for synthetic peptide-based serologic assays that exploit avidity to replace whole antigens while enabling low-cost diagnostics in resource-limited settings. ObjectiveTo evaluate the diagnostic accuracy of a polymeric peptide-based ELISA leveraging avidity to enhance signal. MethodA 15-member SARS-CoV-2 peptide library corresponding to multiple epitope clusters and proteins was screened by indirect ELISA using pooled sera from RT-PCR-confirmed COVID-19 patients to identify peptides with possible diagnostic utility. The identified lead candidate, S559, possessed terminal cysteine-substitution to allow disulfide polymerization, and the resulting avidity gain was evaluated by comparing the apparent dissociation constant (KDapp) before and after depolymerization with N-acetylcysteine. The performance of an optimized ELISA using S559 was evaluated on 1,222 prospectively collected COVID-19 serum samples and 218 biobanked pre-COVID control serum samples. ResultsPolymeric S559 with a KDapp of 29.26 nM-1was demonstrated to have a 218% avidity gain relative to the completely depolymerized form. At pre-defined thresholds, the optimized S559 ELISA has a sensitivity and specificity of 83.39% (95%CI: 81.18% and 85.43%) and 96.79% (95%CI: 93.50% and 98.70%), respectively. At post hoc thresholds determined by Youden index, sensitivity and specificity reached 95.01 (95% CI: 93.63% - 96.16%) and 100.00% (95% CI: 98.32% - 100.00%), respectively. ConclusionHomomultivalent epitope presentation using polymeric S559 allows a highly specific immunoassay using human sera that may have important value in detecting antibodies, whether for diagnosing infection, confirming vaccination status or conducting surveillance.

A role for substance P in promoting neurogenic inflammation and pain has been described in sickle cell disease (SCD). However its origin and contribution to SCD pathophysiology remain unclear. We measured substance P level in plasma from 225 patients with SCD and observed the highest concentrations during acute chest syndrome (ACS). Therefore, we tested the hypothesis that substance P may induce ACS. In transgenic sickle mice, unlike control mice, intravenous injection of substance P caused lethal crises with dose-dependent acute lung injuries. Activation of Fc{varepsilon}R1 with MAR-1 had similar effects, suggesting a role for mast cell or basophil activation and degranulation. Pretreatment of sickle mice with cromolyn, a stabilizer of mast cells and basophils, prevented lethal crisis and lung injuries induced by substance P injection. In SCD patients, blood cellular histamine levels and increased histidine decarboxylase activity were consistent with an involvement of circulating basophils. Flow cytometry analysis revealed higher basophil counts with increased activation and degranulation markers in patients compared with healthy controls. During vaso-occlusive crisis, absolute basophil counts tended to decrease, suggesting their recruitment outside the vascular compartment. The same results were observed in sickle mice after hypoxia-reoxygenation, intravenous hemin injection or substance P injection. Immunohistochemistry revealed the presence of mast cells and basophils in the lungs of sickle mice, but not in control mice, with further basophil recruitment and degranulation after intravenous substance P injection. In SCD patients, we observed extremely high levels of substance P in the sputum collected during ACS, consistently with mast cell and basophil degranulation in the lungs. In vitro, substance P was shown to be a potent chemoattractant for basophils via NK1R. Gene expression analysis on sorted circulating basophils from SCD patients revealed an increased expression of several chemokine receptors, including CCR3 and FPR1, which was confirmed by spectral flow cytometry and could contribute to the recruitment of basophils in the lungs. The two substance P receptors, NK1R and MRGPRX2, were also overexpressed, promoting the vicious cycle of substance P release and pain in SCD patients. Our results reveal a novel mechanism that contributes to the understanding of ACS pathogenesis and highlights the potential role of mast cells and basophils in SCD pathophysiology.

The host response to Neisseria gonorrhoeae is variable, and understanding its systemic and local components is critical to understanding anti-gonococcal immunity for vaccine development. We used a controlled human infection model of male gonococcal urethritis in naive volunteers in combination with multiplex cytokine analyte analysis of blood and urine specimens taken before infection, at the time of acute symptoms, and after curative treatment of N. gonorrhoeae to study responses to early infection. (This study utilized data and specimens from all 11 participants assigned to control arms of two previous randomized clinical trials). All 11 participants developed urethritis between 2 and 5 days post inoculation with N. gonorrhoeae strain FA1090, with a majority having visible discharge by day 3. In urine, we found increases in IL-1RA, G-CSF, and chemokines CXCL10, CCL4, CCL11, GRO/{beta}/{gamma}, and IL-8/CXCL8, with IL-1RA and CCL4 showing direct correlation with the degree of pyuria at the time of infection. Contrary to a prior study using the human challenge model and N. gonorrhoeae strain MS11mkC, we did not see similar increases in urine IL-6, TNF-, or IL-1{beta}, although differences in IL-6, TNF- were observed in participants with later development of infection. Additionally, plasma cytokine levels were unchanged in this cohort over the course of their infection, suggesting these infections were confined to the urethra. We propose that differences in strain virulence or the threshold to define a clinical case may be responsible for this discrepancy, meriting further study and continued use of non-invasive inflammatory markers to study local effects in addition to systemic effects of gonococcal infection. Author SummaryGonorrhea, caused by the bacterium Neisseria gonorrhoeae, remains a global public health concern, yet repeated infections are common and no vaccine is available. A key challenge for vaccine development is limited understanding of how the human immune system responds during early infection, when bacteria are confined to the urethra, vagina, or other mucosal sites. To address this gap, we studied immune responses in a controlled human infection model in which male volunteers with no prior exposure were experimentally infected with N. gonorrhoeae into their urethra. Immune signaling molecules were measured in urine and blood samples collected before infection, during symptoms, and after antibiotic treatment. All participants developed urethral inflammation within a few days of infection. We observed marked increases in multiple inflammatory cytokines in urine, some which correlated with the degree of neutrophils in their urine. In contrast, immune markers in the bloodstream remained largely unchanged. These findings suggest that early infection with the N. gonorrhoeae strain tested triggers a strong localized immune response without widespread systemic inflammation. Our results highlight the value of urine-based, non-invasive sampling and demonstrate the power of human challenge models for studying early immune responses that have been difficult to characterize in animal systems.

Background: Interstitial lung abnormalities (ILA) are radiologic findings of increased lung density or fibrosis in individuals without clinical interstitial lung disease (ILD) and are associated with increased mortality and progression to ILD. Understanding physiologic trajectories of lung function preceding ILA diagnosis may illuminate early mechanisms of lung injury. Methods: We recruited participants from the Coronary Artery Risk Development in Young Adults (CARDIA) Lung Study, a prospective cohort of adults enrolled at ages 18 to 30 years and followed longitudinally for 25 years. Percent predicted forced vital capacity (ppFVC) was measured at five study visits over 20 years. Individual ppFVC trajectories were estimated using random coefficient models. Person specific slopes were incorporated into logistic regression models to examine associations with visually detected ILA on chest CT at exam year 25. Models were adjusted for age, sex, race, body mass index, pack years of smoking, and study center. Results: Among 3,136 participants with complete data, 57 (1.8%) had ILA at mean age 51 years. In univariable and multivariable models, individuals with ILA had greater cumulative decline in ppFVC over the 20 years preceding diagnosis. Each 10% absolute decline in ppFVC was associated with more than twice the odds of ILA (adjusted OR 2.21; 95% confidence interval 1.47, 3.31; p = 0.0001). Conclusions: Greater longitudinal decline in FVC from early adulthood was strongly associated with the presence of ILA at midlife. These findings suggest that physiologic impairments precede radiologic evidence of subclinical parenchymal lung abnormalities, underscoring the potential of life course lung function trajectories to identify individuals at risk for developing ILD.

ObjectiveThis study explored the recovery experiences of individuals who report having (largely) recovered from long covid and who attributed their improvement to mind-body approaches. Design, setting and participantsWe conducted an explorative qualitative study using purposive recruitment through social media and snowball sampling. Eighteen adult women (aged 37-62 years), who self-identified as having had long covid and having substantially recovered through mind-body approaches participated in semi-structured interviews. Data were analysed using Saunders practical thematic analysis. ResultsDespite variation in personal narratives, a common trajectory emerged: participants moved away from a biomedical explanatory model towards one centred on nervous system dysregulation. This shift, sometimes following initial scepticism, was often described as a turning point, sparking hope and motivation to engage in self-directed strategies. Recovery was not linear but an iterative process, involving cycles of practice, reflection (especially when progress stagnated) and adaptation of mind-body techniques. Over time, participants gained insights into contributing factors and, in many cases, made intentional life changes to support ongoing recovery. These patterns echo findings from previous research on mind-body approaches in chronic pain and chronic fatigue, and align with neuroscientific perspectives on symptom generation. Most participants navigated this process without formal clinical support, relying instead on online communities and actively avoiding sources of (biomedical) information that conflicted with their new understanding. ConclusionsWhile causal inferences cannot be drawn from qualitative data, this study highlights potential mechanisms that may underpin recovery for people with long covid using mind-body approaches. Further research is needed to develop structured interventions, and to evaluate their efficacy and safety. Future research should also explore how prevailing narratives within healthcare and society influence treatment engagement and recovery trajectories. STRENGTHS AND LIMITATIONS OF THIS STUDYO_LIThis is the first study exploring experiences of recovery from long covid using mind-body approaches. C_LIO_LIIn-depth, real-world accounts capture the lived-experiences over time and allow in-depth exploration if the recovery process, while the semi-structured design facilitates the emergence of themes rarely captured in clinical research. C_LIO_LIGeneralisability is limited due to self-identified long covid status, lack of formal diagnostic verification, absence of strict definitions of mind-body approaches and recovery, and a relatively homogenous sample (mostly highly educated women). C_LI

Rapid prediction of the severity of acute coronary syndrome (ACS) is crucial for appropriate intervention in emergency department. Neutrophils (Neu), lymphocytes (Lym) and monocytes (Mon) and their ratios (Neu/Lym, NLR; Mon/Lym, MLR NeuxMon/Lym, SIRI) are acknowledged to be associated with the prediction of the severity and adverse outcome of ACS patients. Here, we analysed retrospectively eosinophils (Eos) and Eos-derived novel ratios (Neu/Eos, NER; Mon/Eos, MER; Neu x Mon/Eos, SIII; Neu/Eos x Lym, NEL; Mon/Eos x Lym, MEL; Neu x Mon/Eos x Lym, SV) of first admitted 1053 ACS patients within 24 hours of symptom onset to predict ST-segment elevation of myocardial infarction (STEMI), high Gensini score (H) and cardiac dysfunction (Killip Classification l to III grades). Results showed that Eos was significantly decreased in ST (n=227), Gensini (H) (n=311) and Killip I group (n=237) (P<0.05). All Eos-derived ratios (NER, MER, SIII, NEL, MEL, SV) were significantly higher with diagnostic severity (ST, Gensini (H), and Killip I group (P<0.05). ROC analysis revealed that SIII and SV predicted ST and Gensini (H) with high specificity and sensitivity, which were similar to that of NLR, MLR and SIRI. Conclusion: Eos and Eos-derived ratios, SIII and SV in particular, are strongly linked to the prediction of the severity of ACS, along with those of well-established leukocyte ratios. The new ratios of Eos hold significant importance in emergency department for quick evaluation of ACS patients.

Amatoxin-induced acute liver failure complicates misidentified foraged mushroom ingestion worldwide; abrupt multisystem collapse punctuates apparent improvement. Our prospective single-arm clinical trial investigated proactive toxicokinetic-based management to preserve elimination capacity: sustained enhanced hydration to maintain renal clearance; fasting plus octreotide to suppress meal-driven enterohepatic circulation; and intravenous silibinin to inhibit OATP1B3-mediated hepatic uptake, enabling safe passage and elimination of gallbladder-confined amatoxin-laden bile. Safety population (N=99) transplant-free recovery (TFR): 88.0% (87 recoveries, 6 transplants, 6 deaths). Protocol-adherent Efficacy population (n=86) TFR: 98.8% (85 recoveries, 1 transplant, 0 deaths). Multivariable analysis identified uninterrupted hydration as strongest TFR predictor (P<0.001), followed by earlier silibinin initiation (P=0.003); octreotide shortened INR recovery by 11 hours (P=0.033). These findings support a toxin elimination model in which preserved renal clearance and biliary sequestration are central recovery determinants. The kinetic balance between renal clearance and hepatic uptake governs both recovery and collapse.