The etiology of acute liver failure (ALF) remains elusive in almost half of affected children. We hypothesized that inherited mitochondrial and fatty acid oxidation disorders were occult etiological factors in patients with idiopathic ALF and impaired energy metabolism.

Alterations of the innate immunity contribute to the development of spontaneous bacterial peritonitis (SBP) in liver cirrhosis. Given its role in immune signaling, antimicrobial function, and macrophage differentiation, we hypothesized that genetic polymorphisms of TRAF6 modulate the risk of SBP. Thus, we determined theTRAF6 haplotype in 432 patients with cirrhosis and ascites using the haplotype-tagging single nucleotide polymorphisms rs331457 and rs5030419. In addition, peritoneal macrophages were immunomagnetically isolated and characterized. Overall, 122 (28%) patients had an episode of SBP. In the combined prospective-retrospective analysis the frequency of SBP differed between the four haplotypes (P = 0.014) and was the highest in 102 patients carrying the rs331457 but not the rs5030419 variant, when compared to other haplotypes (odds ratio 1.95 [1.22-3.12]) or to the wild-type (odds ratio 1.71 [1.04-2.82]). This association was confirmed in multivariate logistic regression (adjusted odds ratio 2.00 [1.24-3.22]) and in prospective sensitivity analysis (hazard ratio 2.09 [1.08-4.07]; P = 0.03). The risk haplotype was associated with lower concentrations of the immune activation marker soluble CD87 in ascitic fluid and with a decreased expression of IL-6 and CXCL8 in isolated peritoneal macrophages. In conclusion, genetic polymorphisms of TRAF6 are associated with decreased peritoneal immune activation and an increased risk of SBP.

Comprehensive proteomics profiling may offer new insights into the dysregulated metabolic milieu of type 2 diabetes, and in the future, serve as a useful tool for personalized medicine. This calls for a better understanding of circulating protein patterns at the early stage of type 2 diabetes as well as the dynamics of protein patterns during changes in metabolic status.

G protein-coupled receptors (GPCRs) are key regulatory proteins of immune cell function inducing signaling in response to extracellular (pathogenic) stimuli. Although unrelated, hydroxycarboxylic acid receptor 3 (HCA3) and GPR84 share signaling via Gαi/o proteins and the agonist 3-hydroxydecanoic acid (3HDec). Both receptors are abundantly expressed in monocytes, macrophages and neutrophils but have opposing functions in these innate immune cells. Detailed insights into the molecular mechanisms and signaling components involved in immune cell regulation by GPR84 and HCA3 are still lacking. Here, we report that GPR84-mediated pro-inflammatory signaling depends on coupling to the hematopoietic cell-specific Gα15 protein in human macrophages, while HCA3 exclusively couples to Gαi protein. We show that activated GPR84 induces Gα15-dependent ERK activation, increases intracellular Ca2+ and IP3 levels as well as ROS production. In contrast, HCA3 activation shifts macrophage metabolism to a less glycolytic phenotype, which is associated with anti-inflammatory responses. This is supported by an increased release of anti-inflammatory IL-10 and a decreased secretion of pro-inflammatory IL-1β. In primary human neutrophils, stimulation with HCA3 agonists counteracts the GPR84-induced neutrophil activation. Our analyses reveal that 3HDec acts solely through GPR84 but not HCA3 activation in macrophages. In summary, this study shows that HCA3 mediates hyporesponsiveness in response to metabolites derived from dietary lactic acid bacteria and uncovers that GPR84, which is already targeted in clinical trials, promotes pro-inflammatory signaling via Gα15 protein in macrophages.

Background The association between common carotid artery intima-media thickness (CCA-IMT) and incident carotid plaque has not been characterized fully. We therefore aimed to precisely quantify the relationship between CCA-IMT and carotid plaque development. Methods and Results We undertook an individual participant data meta-analysis of 20 prospective studies from the Proof-ATHERO (Prospective Studies of Atherosclerosis) consortium that recorded baseline CCA-IMT and incident carotid plaque involving 21 494 individuals without a history of cardiovascular disease and without preexisting carotid plaque at baseline. Mean baseline age was 56 years (SD, 9 years), 55% were women, and mean baseline CCA-IMT was 0.71 mm (SD, 0.17 mm). Over a median follow-up of 5.9 years (5th-95th percentile, 1.9-19.0 years), 8278 individuals developed first-ever carotid plaque. We combined study-specific odds ratios (ORs) for incident carotid plaque using random-effects meta-analysis. Baseline CCA-IMT was approximately log-linearly associated with the odds of developing carotid plaque. The age-, sex-, and trial arm-adjusted OR for carotid plaque per SD higher baseline CCA-IMT was 1.40 (95% CI, 1.31-1.50; I2=63.9%). The corresponding OR that was further adjusted for ethnicity, smoking, diabetes, body mass index, systolic blood pressure, low- and high-density lipoprotein cholesterol, and lipid-lowering and antihypertensive medication was 1.34 (95% CI, 1.24-1.45; I2=59.4%; 14 studies; 16 297 participants; 6381 incident plaques). We observed no significant effect modification across clinically relevant subgroups. Sensitivity analysis restricted to studies defining plaque as focal thickening yielded a comparable OR (1.38 [95% CI, 1.29-1.47]; I2=57.1%; 14 studies; 17 352 participants; 6991 incident plaques). Conclusions Our large-scale individual participant data meta-analysis demonstrated that CCA-IMT is associated with the long-term risk of developing first-ever carotid plaque, independent of traditional cardiovascular risk factors.

Reduced lung function is associated with cardiovascular mortality, but the relationships with atherosclerosis are unclear. The population-based Swedish CArdioPulmonary BioImage study measured lung function, emphysema, coronary CT angiography, coronary calcium, carotid plaques and ankle-brachial index in 29,593 men and women aged 50-64 years. The results were confirmed using 2-sample Mendelian randomization. Lower lung function and emphysema were associated with more atherosclerosis, but these relationships were attenuated after adjustment for cardiovascular risk factors. Lung function was not associated with coronary atherosclerosis in 14,524 never-smokers. No potentially causal effect of lung function on atherosclerosis, or vice versa, was found in the 2-sample Mendelian randomization analysis. Here we show that reduced lung function and atherosclerosis are correlated in the population, but probably not causally related. Assessing lung function in addition to conventional cardiovascular risk factors to gauge risk of subclinical atherosclerosis is probably not meaningful, but low lung function found by chance should alert for atherosclerosis.

Like in other vertebrates, the anterior part of the telencephalon of amphibians mainly consists of the olfactory bulb (OB), but different from higher vertebrates, the lateral telencephalic ventricles of larval Xenopus laevis expand deep into the anterior telencephalon. The neurogenic periventricular zone (PVZ) of the lateral ventricles generates new OB neurons throughout the animal's lifetime. We investigated the ultrastructural organization of the PVZ and found that within a time period of 24 h, 42.54 ± 6.65% of all PVZ cells were actively proliferating. Functional purinergic receptors are widespread in the central nervous system and their activation has been associated with many critical physiological processes, including the regulation of cell proliferation. In the present study we identified and characterized the purinergic system of the OB and the PVZ. ATP and 2MeSATP induced strong [Ca(2+)](i) increases in cells of both regions, which could be attenuated by purinergic antagonists. However, a more thorough pharmacological investigation revealed clear differences between the two brain regions. Cells of the OB almost exclusively express ionotropic P2X purinergic receptor subtypes, whereas PVZ cells express both ionotropic P2X and metabotropic P1 and P2Y receptor subtypes. The P2X receptors expressed in the OB are evidently not involved in the immediate processing of olfactory information.

The present study aims to describe accelerometer-assessed physical activity (PA) patterns and fulfillment of PA recommendations in a large sample of middle-aged men and women, and to study differences between subgroups of socio-demographic, socio-economic, and lifestyle-related variables. A total of 27 890 (92.5% of total participants, 52% women, aged 50-64 years) middle-aged men and women with at least four days of valid hip-worn accelerometer data (Actigraph GT3X+, wGT3X+ and wGT3X-BT) from the Swedish CArdioPulmonary bioImage Study, SCAPIS, were included. In total, 54.5% of daily wear time was spent sedentary, 39.1% in low, 5.4% in moderate, and only 0.1% in vigorous PA. Male sex, higher education, low financial strain, born in Sweden, and sedentary/light working situation were related to higher sedentary time, but also higher levels of vigorous PA. High BMI and having multiple chronic diseases associated strongly with higher sedentary time and less time in all three PA intensities. All-year physically active commuters had an overall more active PA pattern. The proportion fulfilling current PA recommendations varied substantially (1.4% to 92.2%) depending on data handling procedures and definition used. Twenty-eight percent was defined as having an "at-risk" behavior, which included both high sedentary time and low vigorous PA. In this large population-based sample, a majority of time was spent sedentary and only a fraction in vigorous PA, with clinically important variations between subgroups. This study provides important reference material and emphasizes the importance of a comprehensive assessment of all aspects of the individual PA pattern in future research and clinical practice.

The adaptation of cellular metabolism is considered a hallmark of cancer. Oncogenic signaling pathways support tumorigenesis and cancer progression through the induction of certain metabolic phenotypes associated with altered regulation of key metabolic enzymes. Hydroxycarboxylic acid receptor 2 (HCA2) is a G protein-coupled receptor previously shown to act as a tumor suppressor. Here, we aimed to unveil the connection between cellular metabolism and HCA2 in BT-474 cells. Moreover, we intend to clarify how well this metabolic phenotype is reflected in transcriptional changes and metabolite levels as determined by global metabolomics analyses.

The interplay of microbiota and the human host is physiologically crucial in health and diseases. The beneficial effects of lactic acid bacteria (LAB), permanently colonizing the human intestine or transiently obtained from food, have been extensively reported. However, the molecular understanding of how LAB modulate human physiology is still limited. G protein-coupled receptors for hydroxycarboxylic acids (HCAR) are regulators of immune functions and energy homeostasis under changing metabolic and dietary conditions. Most mammals have two HCAR (HCA1, HCA2) but humans and other hominids contain a third member (HCA3) in their genomes. A plausible hypothesis why HCA3 function was advantageous in hominid evolution was lacking. Here, we used a combination of evolutionary, analytical and functional methods to unravel the role of HCA3 in vitro and in vivo. The functional studies included different pharmacological assays, analyses of human monocytes and pharmacokinetic measurements in human. We report the discovery of the interaction of D-phenyllactic acid (D-PLA) and the human host through highly potent activation of HCA3. D-PLA is an anti-bacterial metabolite found in high concentrations in LAB-fermented food such as Sauerkraut. We demonstrate that D-PLA from such alimentary sources is well absorbed from the human gut leading to high plasma and urine levels and triggers pertussis toxin-sensitive migration of primary human monocytes in an HCA3-dependent manner. We provide evolutionary, analytical and functional evidence supporting the hypothesis that HCA3 was consolidated in hominids as a new signaling system for LAB-derived metabolites.

The link between the gut microbiota and type 2 diabetes (T2D) warrants further investigation because of known confounding effects from antidiabetic treatment. Here, we profiled the gut microbiota in a discovery (n = 1,011) and validation (n = 484) cohort comprising Swedish subjects naive for diabetes treatment and grouped by glycemic status. We observed that overall gut microbiota composition was altered in groups with impaired glucose tolerance, combined glucose intolerance and T2D, but not in those with impaired fasting glucose. In addition, the abundance of several butyrate producers and functional potential for butyrate production were decreased both in prediabetes and T2D groups. Multivariate analyses and machine learning microbiome models indicated that insulin resistance was strongly associated with microbial variations. Therefore, our study indicates that the gut microbiota represents an important modifiable factor to consider when developing precision medicine approaches for the prevention and/or delay of T2D.

Medium-chain fatty acids and their 3-hydroxy derivatives are metabolites endogenously produced in humans, food-derived or originating from bacteria. They activate G protein-coupled receptors, including GPR84 and HCA3, which regulate metabolism and immune functions. Although both receptors are coupled to Gi proteins, share at least one agonist and show overlapping tissue expression, GPR84 exerts pro-inflammatory effects whereas HCA3 is involved in anti-inflammatory responses. Here, we analyzed signaling kinetics of both HCA3 and GPR84, to unravel signal transduction components that may explain their physiological differences.

Interactions between the gut microbiota, diet, and the host potentially contribute to the development of metabolic diseases. Here, we identify imidazole propionate as a microbially produced histidine-derived metabolite that is present at higher concentrations in subjects with versus without type 2 diabetes. We show that imidazole propionate is produced from histidine in a gut simulator at higher concentrations when using fecal microbiota from subjects with versus without type 2 diabetes and that it impairs glucose tolerance when administered to mice. We further show that imidazole propionate impairs insulin signaling at the level of insulin receptor substrate through the activation of p38γ MAPK, which promotes p62 phosphorylation and, subsequently, activation of mechanistic target of rapamycin complex 1 (mTORC1). We also demonstrate increased activation of p62 and mTORC1 in liver from subjects with type 2 diabetes. Our findings indicate that the microbial metabolite imidazole propionate may contribute to the pathogenesis of type 2 diabetes.

Carotid intima-media thickness (CIMT) is a marker of subclinical organ damage and predicts cardiovascular disease (CVD) events in the general population. It has also been associated with vascular risk in people with diabetes. However, the association of CIMT change in repeated examinations with subsequent CVD events is uncertain, and its use as a surrogate end point in clinical trials is controversial. We aimed at determining the relation of CIMT change to CVD events in people with diabetes.

Cyanobacteria have evolved over billions of years to adapt and survive in diverse climates. Environmentally, this presents a huge challenge because cyanobacteria can now rapidly form algae blooms that are detrimental to aquatic life. In addition, many cyanobacteria produce toxins, making them hazardous to animals and humans that they encounter. Rapid identification of cyanobacteria is essential to monitor and prevent toxic algae blooms. Here, we show for the first time how native mass spectrometry can quickly and precisely identify cyanobacteria from diverse aquatic environments. By monitoring phycobiliproteins, abundant protein complexes within cyanobacteria, simple, easy-to-understand mass spectral "fingerprints" were created that were unique to each species. Moreover, our method is 10-fold more sensitive than the current MALDI-TOF mass spectrometric methods, meaning that cyanobacteria can be monitored using this technology prior to bloom formation. Together, the data show great promise for the simultaneous detection and identification of co-existing cyanobacteria in situ.

Increased extracellular Ca2+ concentrations ([Ca2+]ex) trigger activation of the NLRP3 inflammasome in monocytes through calcium-sensing receptor (CaSR). To prevent extraosseous calcification in vivo, the serum protein fetuin-A stabilizes calcium and phosphate into 70-100 nm-sized colloidal calciprotein particles (CPPs). Here we show that monocytes engulf CPPs via macropinocytosis, and this process is strictly dependent on CaSR signaling triggered by increases in [Ca2+]ex. Enhanced macropinocytosis of CPPs results in increased lysosomal activity, NLRP3 inflammasome activation, and IL-1β release. Monocytes in the context of rheumatoid arthritis (RA) exhibit increased CPP uptake and IL-1β release in response to CaSR signaling. CaSR expression in these monocytes and local [Ca2+] in afflicted joints are increased, probably contributing to this enhanced response. We propose that CaSR-mediated NLRP3 inflammasome activation contributes to inflammatory arthritis and systemic inflammation not only in RA, but possibly also in other inflammatory conditions. Inhibition of CaSR-mediated CPP uptake might be a therapeutic approach to treating RA.

(1) Background: Rapid microglial proliferation contributes to the complex responses of the innate immune system in the brain to various neuroinflammatory stimuli. Here, we investigated the regulatory function of phosphoinositide 3-kinase γ (PI3Kγ) and reactive oxygen species (ROS) for rapid proliferation of murine microglia induced by LPS and ATP. (2) Methods: PI3Kγ knockout mice (PI3Kγ KO), mice expressing catalytically inactive PI3Kγ (PI3Kγ KD) and wild-type mice were assessed for microglial proliferation using an in vivo wound healing assay. Additionally, primary microglia derived from newborn wild-type, PI3Kγ KO and PI3Kγ KD mice were used to analyze PI3Kγ effects on proliferation and cell viability, senescence and cellular and mitochondrial ROS production; the consequences of ROS production for proliferation and cell viability after LPS or ATP stimulation were studied using genetic and pharmacologic approaches. (3) Results: Mice with a loss of lipid kinase activity showed impaired proliferation of microglia. The prerequisite of induced microglial proliferation and cell viability appeared to be PI3Kγ-mediated induction of ROS production. (4) Conclusions: The lipid kinase activity of PI3Kγ plays a crucial role for microglial proliferation and cell viability after acute inflammatory activation.

Leptin and insulin resistance are being discussed to be involved in the pathogenesis of nonalcoholic fatty liver disease, which is frequently characterized by moderately elevated transaminases. However, longitudinal studies proving an association between leptin, insulin resistance, and transaminases are scarce. We examined weight status, aspartate aminotransferase (AST), alanine aminotransferase (ALT), leptin, glucose, and insulin in 180 overweight children at baseline and 1 year later. Relationships between these parameters at baseline and their changes in the course of 1 year were determined by multiple regression analysis adjusted for age, sex, pubertal stage, and body mass index (BMI). Leptin but not homeostasis model assessment of insulin resistance index correlated significantly to transaminases in both cross-sectional and longitudinal analyses. The same findings were observed in 30 children with suspected nonalcoholic fatty liver disease by ultrasound. The 130 children who participated in a 1-year lifestyle intervention reduced their overweight (standard deviation score [SDS]-BMI, -0.37 +/- 0.11). In the course of 1 year, their changes of transaminases depended on change of weight status (SDS-BMI decrease >0.5: ALT 12 [10-15] --> 9 [8-13] U/L, AST 11 [9-12] --> 9 [8-12] U/L; SDS-BMI decrease >0 but 16 [12-26] U/L, AST 10 [8-14] --> 10 [8-24] U/L; no SDS-BMI decrease: ALT 13 [11-20] --> 20[13-33] U/L, AST 11 [9-21] --> 15 [9-24] U/L; data as median and interquartile range). The 50 children without intervention increased their SDS-BMI (+0.02 +/- 0.18) and transaminases (ALT 14 [11-18] --> 19 [15-25] U/L, AST 10 [8-15] --> 16 [10-25] U/L). These findings suggest that leptin may be involved in the pathogenesis of liver diseases. However, to test this hypothesis, careful histologic assessments in correlation to leptin levels are needed.

MIR143 is pathologically downregulated and may function as a tumor suppressor in prostate cancer. Likewise, the urokinase plasminogen activator receptor (UPAR) is overexpressed in prostate carcinoma, representing a negative prognostic marker and putative therapeutic target gene. In this paper, we establish UPAR as a new direct target of MIR143. Luciferase reporter gene constructs identify one of the two in silico-predicted binding sites as functionally relevant for direct MIR143 binding to the 3' UTR, and, concomitantly, transfection of MIR143 reduces UPAR protein levels in prostate carcinoma cells in vitro. Inhibitory effects on cell proliferation and colony formation, spheroid growth and integrity, and cell viability are extensively analyzed, and they are compared to direct small interfering RNA (siRNA)-mediated uPAR knockdown or combined microRNA (miRNA)-siRNA treatment. Switching to a therapeutically more relevant in vivo model, we demonstrate tumor-inhibitory effects of MIR143 replacement therapy by systemic treatment of mice bearing subcutaneous PC-3 tumor xenografts with MIR143 formulated in polymeric nanoparticles. This efficient, nanoparticle-mediated delivery of intact MIR143 mediates the marked downregulation of uPAR protein, but not mRNA levels, thus indicating translational inhibition rather than mRNA degradation. In summary, we identify UPAR as a direct target gene of MIR143, and we establish the therapeutic anti-tumor potential of nanoparticle-based MIR143 replacement in prostate cancer.

The danger signal extracellular calcium is pathophysiologically increased in the synovial fluid of patients with rheumatoid arthritis (RA). Calcium activates the NLRP3-inflammasome via the calcium-sensing receptor in monocytes/macrophages primed by lipopolysaccharide, and this effect is mediated by the uptake of calciprotein particles (CPPs) formed out of calcium, phosphate, and fetuin-A. Aim of the study was to unravel the influence of calcium on monocytes when the priming signal is not present. Monocytes were isolated from the blood of healthy controls and RA patients. Macrophages were characterized using scRNA-seq, DNA microarray, and proteomics. Imaging flow cytometry was utilized to study intracellular events. Here we show that extracellular calcium and CPPs lead to the differentiation of monocytes into calcium-macrophages when the priming signal is absent. Additional growth factors are not needed, and differentiation is triggered by calcium-dependent CPP-uptake, lysosomal alkalization due to CPP overload, and TFEB- and STAT3-dependent increased transcription of the lysosomal gene network. Calcium-macrophages have a needle-like shape, are characterized by excessive, constitutive SPP1/osteopontin production and a strong pro-inflammatory cytokine response. Calcium-macrophages differentiated out of RA monocytes show a stronger manifestation of this phenotype, suggesting the differentiation process might lead to the pro-inflammatory macrophage response seen in the RA synovial membrane.