To Örebro University

Shifting to a plant-based diet naturally alters protein source choices. In many countries, protein from yellow pea is widely used as a main ingredient in meat alternatives. Still, its biological effects, especially regarding gastrointestinal health, remain incompletely understood. The aim of our study was to investigate how a weekly increase in the intake of a well-characterized pea protein isolate affects surrogate markers of health, fecal short-chain fatty acids and gut microbiota composition in healthy individuals. Male and female adults (N = 29) participated in this exploratory intervention study. A 4-week pre-intervention period for questionnaires and fecal samples collection was followed by a 4-week supplementation. Participants consumed isolated pea protein in weekly increasing amounts, starting from 0.25 g per kg body mass per day in week 5 to 1.00 g per kg body mass per day in week 8. Questionnaire data, fecal samples as well as fasting blood and 24 h urine samples were collected weekly. Data from biological samples and questionnaires confirmed a healthy study population and compliance. Fecal calprotectin levels significantly increased only in a subset of participants, which was accompanied by higher fecal water cytotoxicity in vitro. Short-chain fatty acids mainly rose in those subjects with stable calprotectin levels. Relative abundances of Limosilactobacillus frumenti, Odoribacter splanchnicus and Lactobacillus crispatus increased significantly in the total population during the intervention while the relative abundance of Bifidobacterium longum and Bifidobacterium catenulatum decreased. Our results indicate that an increased intake of pea protein isolate affects the growth of certain beneficial bacterial strains and differentially influences markers related to gut inflammation in healthy individuals.

Per- and polyfluoroalkyl substances (PFAS) are a family of persistent chemicals that continue to be released pervasively into the environment, leading to widespread human exposure. Emerging epidemiological evidence shows adverse effects on liver lipids; however, past toxicological studies have been limited by a focus on peroxisome proliferator activated receptor α (PPARα) driven effects on triglycerides in rodent systems. Here, we use a more agonostic approach incorporating lipidomics and transcriptomics to test the hypothesis that activation of human PPARα by PFOA, disrupts liver lipid homeostasis, broadly, similar to that seen in human liver diseases. Female and male mice expressing human PPAR α or that were PPARα null were fed a What We Eat In America diet and exposed to PFOA via drinking water for 6 weeks. Serum PFOA concentrations averaged 48 ± 9 μg/mL. PFOA changed the expression of ~2000 hepatic genes with expression of a larger number of genes in hPPARα versus PPARα null mice. In this occupational level PFOA exposure scenario, less than 60 % of transcriptional changes induced by PFOA depended on PPARα expression. CAR was another major molecular initiating event, with other transcription factors pathways more likely to be modulated downstream of hPPARα activation. In hPPARα mice of both sexes, PFOA increased total liver lipids. In addition to triacylglycerides, lipid classes strongly altered by PFOA exposure predominantly belong to phosphatidylcholine and sphingolipid classes. PFOA significantly decreased sphingomyelin abundance and increased ceramide abundance regardless of genotype, which coincided with an increase in expression of SMase, the enzyme that converts sphingomyelin to ceramide. These results highlight the ability of PFOA to modulate liver lipids beyond triacylglycerides in both an hPPARα-dependent and -independent manner.

The deep terrestrial subsurface (DTS) biosphere consists of a variety of distinct microbial taxa, mostly bacterial. The mechanisms by which microbes dynamically manage the uptake and concurrent utilization of nutrients within the DTS environments remain largely unexplored. Here, we examined the utilization patterns of amino acids and other polar metabolites in cultured DTS bacterial communities to investigate the adaptive responses and metabolic pathways employed under varying nutrient conditions to gain insight into how environmental shifts impact the metabolism of these communities. Previously, we found that changes in growth conditions affected the composition and size of the bacterial communities enriched from these oligotrophic, anoxic environments and induced changes in the production of primary and secondary metabolites. In the present study, metabolic fingerprinting was used to investigate the primary and secondary metabolite utilization and main metabolic pathways present in the enriched DTS bacterial consortium originating from the deep bedrock of the Fennoscandian Shield. We found that especially amino acids were predominantly degraded under different nutrient conditions. Notably, the degradation of phenylalanine and valine constituted a 'core' metabolic process that remained unaffected by variations in available nutrients within this community. Further, the most significant metabolic pathways employed were those connected to phenylalanine, cysteine and methionine.

Emerging studies reveal that gut microbes can conjugate diverse amino acids to bile acids, known as microbially conjugated bile acids. However, their regulation and health effects remain unclear. Here, we analyzed early-life microbially conjugated bile acid patterns and their link to islet autoimmunity. We quantified 110 microbial bile acids in 303 stool samples collected longitudinally (3-36 months) from children who developed one or more islet autoantibodies and controls who remained autoantibody-negative. We identified distinct age-dependent trajectories of these bile acid amidates and correlated them with gut microbiome composition. We found that altered levels of ursodeoxycholic and deoxycholic acid conjugates were linked to islet autoimmunity as well as modulated monocyte activation in response to immunostimulatory lipopolysaccharide and Th17/Treg cell balance. These findings suggest that microbially conjugated bile acids influence immune development and type 1 diabetes risk.

Background & Aims: Granular detail about the location and nature of liver cell interactions and the metabolic, inflammatory and fibrogenic pathways driving progressive fibrosis in metabolic dysfunction-associated steatotic liver disease (MASLD) is needed to deliver novel therapeutic targets.

Methods: We generated Visium spatial transcriptomic data from 33 human liver biopsies across the spectrum of MASLD. Gene expression data were overlaid with histological annotations to integrate spatial molecular and histopathological insights to interrogate disease progression. Differential gene expression and pathway, cellular deconvolution and ligand-receptor interaction analyses were conducted for each annotated anatomical category, with specific protein expression validated using immunohistochemistry staining.

Results: Unsupervised clustering based on gene expression data classified the annotated spots into 2 main clusters enriched for fibro-inflammatory vs parenchymal regions. Transcriptomic cellular deconvolution aligned well with manually annotated histopathological features. Fibrotic regions were enriched for genes involved in extracellular matrix/receptor interactions and inflammatory pathways (Benjamini-Hochberg adjusted p-values < 0.05), underscoring known pathological mechanisms. We also identified immunoglobulin gene induction in late-stage fibrosis, which was spatially associated with a senescence signature, as has previously been reported in aging tissues. Dynamic changes in metabolic gene expression from early to late fibrosis were observed, suggesting MASLD progression is accompanied by a decline in normal liver metabolic function and reprogramming of metabolic fuel utilisation from oxidative to glycolytic metabolism, which may be both a cause and a consequence of senescence.

Conclusions: Taken together, our valuable discovery dataset highlights the complex crosstalk between metabolic perturbations and inflammation underpinning fibrosis progression in MASLD.

Impact and implications: Metabolic associated steatotic liver disease has a complex pathogenesis driven by cell and matrix interactions in inflammatory niches. In this study we identify a senescence signature in fibroinflammatory regions, characterised by high immunoglobulin expression and associated with a shift from oxidative to glycolytic metabolism. We identify spatially co-expressed ligand-receptor pairs, including senescence-associated factors, correlated with progressive fibrosis. This discovery dataset highlights the complex crosstalk between metabolic perturbations and inflammation underpinning fibrosis progression in MASLD and lays the groundwork for future research into the role of senescence in MASLD.

Type 1 diabetes (T1D) is increasing globally, yet the earliest biological determinants remain poorly defined, particularly in general population studies. We studied the Swedish population-based ABIS birth cohort (n = 16,683) to identify early-life risk factors. Olink proteomic analysis (n = 286 controls, n = 146 cases) of inflammatory signals at birth shows differential abundance years before diagnosis (mean age 12.6 years), with proteins enriched for neutrophil migration, cytotoxicity, extracellular matrix remodeling, and immune regulation. Several markers remain significant in spite of prenatal and perinatal factors including family history of diabetes, and are associated with differences in compounds like stearic acid, lysine, glutamine, and persistent, environmental toxicants perfluorodecylethanoic acid and perfluorooctane sulfonate (PFOS). Using machine learning, we identify a protein subset that predicts T1D with high accuracy (AUC = 0.89 ± 0.02), independently of HLA genetic risk. These findings suggest that innate and tissue-remodeling pathways are perturbed at birth, possibly reflecting early β-cell vulnerability. Identifying these disruptions at birth with a non-invasive method opens a window for prevention, protecting β-cells before the inflammatory attack on islets begins.

Per- and polyfluoroalkyl substances (PFAS) are persistent environmental pollutants with known bioaccumulative properties and increasing recognition for their potential to disrupt metabolic pathways, including bile acid transport and lipid metabolism. Despite growing global attention, data on PFAS exposure and distribution in the Balkan region, including Serbia, remain limited. This work presents preliminary findings on PFAS presence and explores potential associations with bile acid metabolism in patients with metabolic dysfunction-associated steatotic liver disease (MASLD) and age-, BMI-, and sex-matched controls, as part of a broader multi-omics investigation. Plasma samples were collected from 122 participants (MASLD patients and matched controls) and metabolites were extracted using a monophasic solution of methanol, methyl tert-butyl ether, and isopropanol (20:15:15) containing internal standards. Extracts were analyzed using ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry on a C18 column. Data processing was performed using MZmine 3 software, and compound annotation was based on an in-house spectral library. Among the detected environmental contaminants, three PFAS compounds—perfluorohexane sulfonic acid (PFHxS) and both linear and branched isomers of perfluorooctane sulfonic acid (PFOS)—were consistently identified across all samples, suggesting widespread low-level exposure in the cohort, with no significant differences in PFAS abundance between MASLD patients and healthy controls. These preliminary results highlight the presence of PFAS compounds in all participants, reflecting a baseline environmental exposure within this population. Although no clear associations with MASLD status were observed, the integration of environmental contaminant data into multi-omics frameworks is critical for uncovering subtle metabolic perturbations. Ongoing analyses will focus on profiling secondary bile acids and microbiota-derived metabolites to explore potential downstream effects of PFAS exposure and their relevance to MASLD pathophysiology.

Biocides are crucial in industrial applications to minimize microbial growth and prevent product spoilage. Water-based construction coatings are susceptible to microbial contamination during manufacturing and storage and this adversely impacts product properties, reduces shelf-life and leads to substantial commercial losses. The future trend to lower the biocide concentrations in water-based coatings raises concerns about the emergence of biocide-resistant microbes. This study aims to identify and characterize the biocide-resistant microbe isolated from construction water-based coating materials to better understand its mechanisms of resistance. A total of 63 samples were collected from spoiled products, raw materials, and water from a manufacturing facility, and Pseudomonas oleovorans P4A were identified in all biocides-treated samples. A comparison between a P. oleovorans reference strain, 1045, and the P4A isolate revealed distinct colony morphology, growth rate and sensitivity to biocides and antibiotics. The P4A isolate was 3-fold more resistant to 5-chloro-2-methyl-isothiazolin-3-one (CMIT) and 1.5-fold more resistant to benzothiazolinone (BIT) compared to the reference strain. Conversely, it was 1.4-fold more sensitive to methylisothiazolinone (MIT) compared to the reference strain. No cross-resistance to antibiotics was observed. Metabolomic analysis using liquid chromatography combined with high-resolution mass spectrometry of lipids and polar metabolites showed that P4A had a relatively higher amount of lipids, while 1045 had a relatively higher amount of polar metabolites identified. A significant difference in lipid composition, specifically in diacylglycerol, phosphatidic acid, phosphatidylcholine, and phosphatidylserine was observed between P. oleovorans strains 1045 and P4A. These distinctions highlight increased lipid metabolism in P. oleovorans P4A and this may contribute to its adaptation to biocides. Microbial resistance can directly affect the effectiveness of these products, leading to an increased need for frequent maintenance and replacement, safety concerns, and environmental implications.

Background: Poly- and perfluoroalkyl substances (PFAS) are persistent pollutants affecting wildlife and biodiversity. Perfluorooctane sulfonic acid (PFOS) and one of its short-chain substitutes, perfluorobutane sulfonic acid (PFBS), are widely found in environmental components, especially in water. PFOS has been highlighted as causing deleterious effects on various organisms while PFBS adversity is suspected but requires further investigation. In this study, zebrafish embryos were exposed from 2 h post-fertilization to 28 days post-fertilization to two different concentrations (0.2 mu g/L and 2 mu g/L) of PFOS or PFBS. We then investigated the impacts of these early exposures later in life on adult fish fitness, growth, morphology, behaviour, and liver lipidomic profiles.

Results: PFOS exposure significantly reduced egg production, and both PFOS and PFBS altered growth patterns, organ development, and anxiety-like behaviour. Lipidomic analyses revealed persistent shifts in liver lipid composition that correspond to these phenotypic changes.

Conclusions: Taken together, our findings indicate that early-life exposure to low levels of PFOS and PFBS leads to long-term, sex-specific impairments in zebrafish physiology and behaviour, with disruptions in lipid metabolism emerging as a potential underlying mechanism.

Background and aims: Phosphatidylcholines (PCs) are a major hepatic reservoir of polyunsaturated fatty acids (PUFAs). In murine models of metabolic dysfunction-associated steatotic liver disease (MASLD), liver injury is induced by PUFA-PC depletion. The lipid droplet enzyme HSD17B13 metabolizes PUFA derivatives, and its loss-of-function variant rs72613567:TA protects against steatohepatitis (MASH) and fibrosis, although the mechanism underlying this protection remains unknown. We investigated whether hepatic PUFA-PC metabolism is influenced by MASLD or the protective HSD17B13 variant.

Method: Obese patients with a liver biopsy (n = 135) underwent genotyping for HSD17B13 rs72613567:TA and analysis of the hepatic lipidome by UPLC-MS. A state-of-the-art deep learning image analysis method (Aiforia Technologies) quantified steatosis via the hepatic parenchymal fat fraction. Using a recruit-by-genotype approach, we studied homozygous rs72613567:TA carriers (n = 13) and non-carriers (n = 13) to determine whether the variant affects incorporation of 13C-labeled PUFAs linoleic acid (LA) and alphalinolenic acid (ALA) into triglycerides (TGs) and phospholipids (PLs) secreted by the liver in very-low-density lipoprotein (VLDL). We tested whether targeting HSD17B13 with a small molecule inhibitor, INI-822, affects PC metabolism using two models: in vitro, a primary human liver-on-a-chip system challenged with high-fat media for 20 days; and in vivo, Zucker obese rats fed either an atherogenic diet or a choline-deficient, L-amino acid-defined high-fat diet for 21 days.

Results: Human MASLD livers were characterized by a marked depletion of hepatic PUFA-PCs containing 5–8 double bonds. Steatosis had a particularly accentuated effect to lower PUFA-PCs in individuals without the HSD17B13 rs72613567:TA variant. However, this effect was abolished in variant carriers due to markedly increased concentrations of hepatic PUFA-PCs compared to non-carriers. Homozygous carriers had significantly decreased incorporation of [U-13 C]LA into VLDL-TG (P < 0.001) and of [U-13 C] LA and[U-13 C]ALA into VLDL-PL (P = 0.01 and 0.05), consistent with retention of these PUFAs in the liver. In vitro, inhibition of HSD17B13 by INI-822 in the human liver-on-a-chip system lowered fibrotic markers while stabilizing choline utilization and increasing PC concentrations. In vivo, inhibition of HSD17B13 in Zucker obese rats fed MASH-inducing diets reduced liver enzymes and dose-dependently increased hepatic PC concentrations.

Conclusion: In humans, HSD17B13 rs72613567:TA prevents MASLD-induced hepatic PUFA-PC depletion by retaining PUFAs within the liver. Pharmacological inhibition of HSD17B13 recapitulates the human phenotype in vitro and in vivo. These findings suggest tha thepatic PC enrichment is central to the protective effects associated with HSD17B13 loss-of-function.