To Örebro University

INTRODUCTION: We have recently described relationships between environmental contaminants (ECs) and sex-hormones in the elderly. The present study investigates whether blood concentrations of four classes of ECs also associated with concentrations of five adrenal steroids (AS), cortisol and its four precursors.

METHODS: Plasma samples from participants of the cross-sectional population-based Prospective Investigation of the Vasculature in Uppsala Seniors study (PIVUS, 70-year-old men and women, n = 950) were analyzed using validated mass spectrometry-based methods for four AS: pregnenolone (Pregn), 17 hydroxy Pregn (17OH-Pregn), 17 hydroxy progesterone (17OH-Prog), 11 deoxycortisol; 21 persistent organic pollutants (POP); 8 perfluoroalkyl substances (PFAS); 4 phthalates and 10 metals. Cortisol was analyzed using competitive ELISA.

RESULTS: Results of this study provide evidence that ECs of all evaluated classes were related to AS concentrations in a complex manner, suggesting structure-activity relationships influencing AS biosynthesis. Among the steroids analyzed, Pregn exhibited the strongest associations with ECs concentrations, while cortisol was the least affected. The most pronounced relationships with Pregn, 17OH-Prog, and 17OH-Pregn were observed with highly chlorinated PCBs, all analyzed phthalates, and metals, aluminum, cadmium, copper, mercury and nickel. These associations were generally consistent between men and women, although stronger associations between the ECs and Pregn, 17OH-Pregn, and 17OH-Prog were observed in women. Multivariable regression models further revealed several non-linear relationships between ECs and AS concentrations.

CONCLUSIONS: Our data suggest that in elderly, concentrations of many ECs are associated with concentrations of AS of cortisol pathway. The impact of such relationships on diseases associated with AS biosynthesis remains to be established.

Per- and polyfluoroalkyl substances (PFASs) are persistent environmental pollutants linked to adverse health outcomes, notably kidney disease. This study examines the effects of hemodialysis and dialysis membrane characteristics on the clearance of eight specific PFASs in patients undergoing hemodialysis. A cohort of 301 chronic hemodialysis patients, each treated for a minimum of 90 days, was analyzed. Automated column-switching ultra-performance liquid chromatography-tandem mass spectrometry was utilized to detect the PFASs, and multivariable linear regression models assessed the association between PFAS levels and clinical nutritional biochemistry markers. Results revealed significantly lower PFAS concentrations in the FILTRYZER dialyzer membrane compared to Polyamix, with no notable differences across varying membrane surface areas. Moreover, significant correlations were observed between nutritional markers-such as albumin, uric acid, and the normalized protein catabolic rate-and the levels of perfluoroheptanoic acid (PFHpA), perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), and perfluoroundecanoic acid (PFUnDA). The findings indicate that PFAS levels in hemodialysis patients are influenced by the membrane composition and properties but are unaffected by the surface area of the membranes.

BACKGROUND AND AIMS: The diagnostic and prognostic properties of anti-integrin αvβ6 IgG autoantibodies in ulcerative colitis (UC) are poorly understood. We aimed to assess the diagnostic performance of anti-integrin αvβ6 autoantibodies and examine their association with disease outcomes.

METHODS: Serum samples from a Swedish inception cohort of patients with suspected inflammatory bowel disease (IBD, n=473) were analysed using an in-house fluorescence enzyme immunoassay based on EliA™technology. Findings were validated in a Norwegian population-based inception cohort (n=570). Diagnostic performance was assessed by calculating the area under the curve (AUC) with 95% confidence intervals (CIs) and determining sensitivity and specificity. Reclassification was evaluated using the net reclassification index.

RESULTS: In the discovery cohort, patients with UC, IBD-unclassified, or colonic Crohn's disease exhibited higher median autoantibody levels compared to symptomatic and healthy controls. In the validation cohort, the autoantibody demonstrated 79% sensitivity and 94% specificity for UC vs symptomatic controls at a cut-off of 400 UA/l. Its diagnostic performance (AUC=0.92, 95%CI 0.89-0.95) was superior to hs-CRP (AUC=0.65, 95%CI 0.60-0.70, P<0.001) and faecal calprotectin (fcalpro) (AUC=0.88, 95%CI 0.84-0.92, P=0.09). Combining the autoantibody with fcalpro further improved diagnostic accuracy (AUC=0.97, 95%CI 0.95-0.98) and patient reclassification (P<0.001). Autoantibody positivity was associated with a severe phenotype of UC, characterised by increased inflammatory activity and higher IL-17A and granzyme B levels. Higher autoantibody levels were linked to an aggressive disease course, remaining stable in aggressive UC but decreasing in indolent disease (P=0.003).

CONCLUSIONS: Anti-integrin αvβ6 is a reliable diagnostic and prognostic marker for UC, with potential clinical implementation.

Trimethylamine-N-oxide (TMAO), a gut microbiota-derived dietary metabolite, is linked to progression of chronic kidney disease (CKD). Megalin, a renal proximal tubule receptor crucial for albumin reabsorption, also plays a role in CKD. However, the relationship between them is not well explored. The aim of this study was to investigate if there are any correlations between the levels of TMAO, megalin, lysine and markers of tubular damage in CKD. Urinary metabolites (TMAO, choline, L-carnitine, betaine, lysine) and tubular markers (megalin, albumin, EGF, MCP-1) were quantified by LC-MS/MS and ELISA. Associations were evaluated using analysis of covariance (ANCOVA) adjusted for age and diabetes, with false discovery rate correction. Compared with controls, CKD patients showed higher urinary choline (FDR < 0.001), betaine (FDR = 0.007), lysine (FDR = 0.005), and soluble megalin (FDR < 0.001) but lower EGF and EGF/MCP-1 ratio (both FDR < 0.001). Correlation analyses revealed that serum TMAO was positively associated with soluble megalin and negatively with EGF/MCP-1 ratio. Choline, L-carnitine, and betaine were positively correlated with megalin. This cross-sectional study identifies associations between urinary metabolites, megalin, and tubular injury markers in advanced CKD. Although causality cannot be inferred, the results point to a potential metabolic-tubular link that should be explored in future longitudinal and mechanistic studies.

BACKGROUND: Perfluoroalkyl substances (PFAS) constitute a diverse group of chemical compounds used in various consumer products. While the associations between PFAS and certain adverse human health effects are well-documented, their impact on kidney function remains less known.

OBJECTIVE: The main aim of this study is to investigate the relationship between PFAS levels and kidney function (estimated glomerular filtration rate [eGFR]) utilizing a longitudinal design.

METHODS: The population-based Prospective Investigation of the Vasculature in Uppsala Seniors (PIVUS) study included 997 individuals at baseline (all aged 70 years, 50% females). Follow-up investigations were performed at 75 and 80 years of age. Seven major PFAS were determined in plasma using ultra-high performance liquid chromatography-mass spectrometry. Longitudinal and cross-sectional associations between PFAS and eGFR were analyzed using linear regression and mixed effects models following adjustment for sex, HDL and LDL-cholesterol, triglycerides, glucose, BMI, statin use and smoking.

RESULTS: Longitudinal models demonstrated statistically significant positive associations between perfluoroundecanoic acid (PFUnDA), perfluorononanoic acid (PFNA), and perfluorodecanoic acid (PFDA)and eGFR (all P < 0.001). The associations between linear perfluorooctane sulfonic acid (PFOS) and perfluorohexanesulfonic acid (PFHxS) followed a similar trend. In contrast, an inverse relationship between perfluoroheptanoic acid (PFHpA) and perfluorooctanesulfonamide (PFOSA) with eGFR was observed. The findings were largely corroborated by cross-sectional analyses.

IMPACT STATEMENT: This longitudinal study found that changes in certain PFAS concentrations were positively associated with the change in kidney function, though the direction of association varied across PFAS. These findings were further supported by cross-sectional analysis. The complexity of associations remains incompletely understood as some PFAS showed positive associations while others were inverse. Further longitudinal studies with repeated measures are needed to better elucidate the relationship between PFAS exposure and kidney function.

Steroid hormone concentrations reflect diverse physiological and pathological processes and have been recognized as valuable biomarkers for disease, with growing interest in their potential for patient stratification in precision medicine. Salivary steroid concentrations should reflect the free (biologically active) steroids in circulation, as steroids in the bloodstream passively diffuse to saliva. This allows for the direct measurement of free steroids without transporter protein-bound hormones (inactive form). However, implementation of salivary steroid quantification in larger studies remains limited by challenges associated with sample preparation. We began by reviewing the literature on relevant sample preparation approaches and identified commonly used 96-well solid phase extraction (SPE) methods for further evaluation. Three sample preparation methods were selected and assessed in terms of internal standard recovery and matrix effects, and their response was also compared using electrospray (ESI) versus UniSpray ionization (USI) liquid chromatography-tandem mass spectrometry (LC-MS/MS). We demonstrate a sensitive and rapid high-throughput method with Oasis HLB mu Elution SPE of 200 mu L saliva in 96-well format and USI-LC-MS/MS for major steroids (testosterone, androstenedione, cortisone, cortisol, and progesterone) in saliva. The method achieved optimal recovery (77%), matrix effects (33%), and sensitivity with detection limits (MDL) ranging between 1.1 and 3.0 pg/mL and linearity (r2 = 0.99). Intra-plate and inter-plate coefficient of variation (CV) was below 7% and 20% using USI. USI provided a higher (2.0-2.8-fold) response than ESI and higher signal-to-noise ratio (S/N). The method was then applied to 97 authentic saliva samples (41 male and 56 female) and significant correlations between age and BMI, and androgen levels were observed in both sexes. The proposed 96-well SPE USI-LC-MS/MS method is well-suited for determining steroid hormones in saliva with potential usefulness in large-scale studies and clinical settings.

Metal additive manufacturing (AM), also known as industrial 3D printing, has revolutionized modern industry, enabling the creation of complex, high-performance components across sectors such as aerospace, automotive, and biomedicine. While the printing process itself is often well-contained, a critical and understudied phase – post-processing – has emerged as a source of potentially hazardous airborne particulate matter. These emissions may pose health risks to workers, particularly through interaction with the immune system, which serves as the body's first line of defense and a sentinel of environmental stressors. Yet, limited data exist on the physicochemical properties and immunotoxicological impact of these particles. This study aimed to assess the immunological consequences of particle emissions released during the post-processing of metallic AM alloys, using a human macrophage model and a multi-omics framework.

Airborne particles were collected directly from an operational AM facility using a cascade impactor, separating them into five size fractions, ranging from coarse (>2.5 μm) to nanoscale (<250 nm). A comprehensive physicochemical characterization was performed using scanning electron microscopy with energy-dispersive spectroscopy and X-ray photoelectron spectroscopy. The emitted particles were highly heterogeneous, with irregular, sharp morphologies, and exhibited increased surface oxidation compared to virgin feedstock powders. Functional toxicological assessments were performed in human macrophages, including transmission electron microscopy to evaluate particle uptake. Macrophages, both resting and lipopolysaccharide-primed, displayed potent and dose-dependent inflammatory responses, as seen by elevated secretion of several cytokines (e.g., IL-1β, IL-6). RNA sequencing revealed profound alterations in macrophage gene expression, including dysregulation of NF-κB signaling, cellular senescence, and lipid metabolism pathways. Gene set enrichment analysis indicated broader perturbations in immune regulation and macrophage homeostasis. Non-targeted metabolomics demonstrated significant changes in intracellular metabolic profiles. Specifically, there was an upregulation of numerous lipids and a suppression of several metabolites involved in immunomodulation and cellular energy homeostasis, including tryptophan, NAD, and phenylalanine. Integrated multi-omics analysis revealed a coordinated crosstalk between transcriptional and metabolic responses, pointing to an acute and multifaceted inflammatory reprogramming of macrophages in response to post-processing AM particles.

In conclusion, this study provides the first integrative multi-omics characterization of human immune cell responses to airborne particulate emissions from metal AM post-processing. These results not only advance the field of nanosafety in industrial AM environments but also underscore the urgent need for targeted risk mitigation strategies during post-processing.

BACKGROUND: Per- and polyfluoroalkyl substances (PFAS) are persistent and ubiquitous chemicals posing environmental and health risks. Impact on the human immune system is of particular concern, especially during fetal immune development. Alterations to fetal immune development can impact immunity later in life, e.g., the response to vaccines and pathogens. OBJECTIVES: This study investigated the association between PFAS concentrations in healthy pregnant women from Hamburg, Germany, and antibody levels to routine vaccines in childhood and occurrence of childhood infections.

METHODS: Mid-pregnancy serum samples from 152 mothers-child pairs were analyzed for 18 PFAS compounds, and antibody levels to measles, mumps, rubella, diphtheria, and tetanus were assessed at age 5. Maternal questionnaires provided data on childhood infections each year at age 1-5. Linear and Poisson regression models were adjusted for maternal age, education, parity, and breastfeeding duration. Weighted quantile sum (WQS) regression was used to assess the PFAS mixture.

RESULTS: Higher PFAS concentrations were associated with lower antibody titers at age 5 years, particularly for mumps, tetanus, diphtheria, and rubella. Several PFAS were also linked to increased childhood infections, especially respiratory infections, during ages 3 and 4 years. WQS regression revealed a negative association between combined PFAS and tetanus titers.

CONCLUSIONS: Maternal PFAS concentrations during pregnancy are inversely associated with antibody levels in children and positively associated with increased childhood infections, notably respiratory infections. These findings underscore the importance of understanding environmental exposures' impact on immune responses and call for continued monitoring of PFAS in both the environment and human populations to mitigate health risks.

INTRODUCTION: Dalbavancin is a lipoglycopeptide with an exceptionally long half-life that allows simplified administration, which may be of value in long-term treatment of bone and joint infections, such as prosthetic joint infections (PJIs). The objective was to determine trough (Cmin) values of dalbavancin during long-term PJI treatment according to the recommendation of the Swedish National Guidelines for Bone and Joint Infections: a loading dose of 1,500 mg on day 1 and another 1,500 mg on days 8-14, followed by day 28 administration of 1,000 mg every two weeks or 500 mg per week. PATIENTS/

METHODS: Twelve patients with PJI treated with at least six doses of dalbavancin were prospectively followed up, serum samples were collected, and renal function was investigated. Dalbavancin concentrations were measured using ultra-high pressure liquid chromatography coupled with unispray tandem mass spectrometry (UHPLC-MS/MS).

RESULTS: The median serum concentration (Cmin) 14 days after the first 1,500 mg dose was 36.3 mg/L (range: 6.6-62.4 mg/L). The median trough value at the date of the last given dose (1,000 mg) after a total of 6-7 doses was 53.6 mg/L (range: 32.0-97.5 mg/L). Three patients showed a tendency towards successive accumulation of dalbavancin during treatment. None of the patients showed any significant impairment in renal function.

CONCLUSIONS: Therapeutic drug monitoring during long-term dalbavancin treatment is recommended to avoid the risk of accumulation and unnecessarily high trough levels. In many cases, such monitoring can allow the dosing interval to be extended.

Isotope dilution ultrahigh-performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) is commonly used for trace analysis of polyfluoroalkyl and perfluoroalkyl substances (PFAS) in difficult matrices. Commercial nontargeted analysis of major PFAS where relative concentrations are obtained cost effectively is rapidly emerging and is claimed to provide comparable results to that of absolute quantification using matrix matched calibration and isotope dilution UHPLC-MS/MS. However, this remains to be demonstrated on a large scale. We aimed to assess the performance of a targeted absolute quantification isotope dilution LC-MS/MS assay versus a commercial nontargeted relative quantification assay for detection of three major PFAS in human blood. We evaluated a population-based cohort of 503 individuals. Correlations were assessed using Spearman's rank correlation coefficients (rho). Precision and bias were assessed using Bland-Altman plots. For perfluorooctane sulfonic acid, the median concentrations were 5.10 ng/mL (interquartile range [IQR] 3.50-7.24 ng/mL), the two assays correlated with rho 0.83. For perfluorooctanoic acid, the median concentrations were 2.14 ng/mL (IQR 1.60-3.0 ng/mL), the two assays correlated with rho 0.92. For perfluorohexanesulfonate, the median concentrations were 5.5 ng/mL (IQR 2.50-11.61 ng/mL), the two assays correlated with rho 0.96. The Bland-Altman statistical test showed agreement of the mean difference for the majority of samples (97-98%) between the two assays. Absolute plasma concentrations of PFAS obtained using matrix matched calibration and isotope dilution UHPLC-MS/MS show agreement with relative plasma concentrations from a nontargeted commercial platform by Metabolon. We observed striking consistency between the two assays when examining the associations of the three PFAS with cholesterol, offering additional confidence in the validity of utilizing the nontargeted approach for correlations with various health phenotypes.