Neuroimmune cardiovascular interfaces control atherosclerosis.

Atherosclerotic plaques develop in the inner intimal layer of arteries and can cause heart attacks and strokes1. As plaques lack innervation, the effects of neuronal control on atherosclerosis remain unclear. However, the immune system responds to plaques by forming leukocyte infiltrates in the outer connective tissue coat of arteries (the adventitia)2-6. Here, because the peripheral nervous system uses the adventitia as its principal conduit to reach distant targets7-9, we postulated that the peripheral nervous system may directly interact with diseased arteries. Unexpectedly, widespread neuroimmune cardiovascular interfaces (NICIs) arose in mouse and human atherosclerosis-diseased adventitia segments showed expanded axon networks, including growth cones at axon endings near immune cells and media smooth muscle cells. Mouse NICIs established a structural artery-brain circuit (ABC): abdominal adventitia nociceptive afferents10-14 entered the central nervous system through spinal cord T6-T13 dorsal root ganglia and were traced to higher brain regions, including the parabrachial and central amygdala neurons; and sympathetic efferent neurons projected from medullary and hypothalamic neurons to the adventitia through spinal intermediolateral neurons and both coeliac and sympathetic chain ganglia. Moreover, ABC peripheral nervous system components were activated: splenic sympathetic and coeliac vagus nerve activities increased in parallel to disease progression, whereas coeliac ganglionectomy led to the disintegration of adventitial NICIs, reduced disease progression and enhanced plaque stability. Thus, the peripheral nervous system uses NICIs to assemble a structural ABC, and therapeutic intervention in the ABC attenuates atherosclerosis.

Differential effects of plant and animal fats on obesity-induced dyslipidemia and atherosclerosis in Ldlr-/-.Leiden mice.

Cardiovascular disease (CVD) is closely associated with obesity through risk factors such as dyslipidemia and chronic low-grade inflammation, which may be affected by diet. Dietary fats have been extensively studied in relation to CVD risk, however these studies have not always yielded consistent results, most likely due to lack in control of experimental conditions and confounding factors. Here we studied the effects of different plant and animal fats on dyslipidemia, inflammation, and atherosclerosis. Ldlr-/-.Leiden mice were fed isocaloric energy-dense diets with translational macronutrient composition for 28 weeks. The diets were identical apart from the type of fat they contained: either (1) a mixture of olive and rapeseed oil, (2) sunflower oil, (3) pork fat, (4) beef fat, or (5) milk fat. The fatty acid composition of the diets was determined and effects on circulating lipid and inflammatory risk factors and atherosclerosis were examined, complemented by adipose tissue histology and liver transcriptomics. While visceral fat mass, adipocyte size, and adipose tissue inflammation were not differentially affected by the diets, atherosclerotic lesion load and severity was more pronounced with increasing dietary saturated fatty acid content and decreasing monounsaturated and polyunsaturated fatty acid content, and hence most pronounced with beef and milk fat. These differential effects were accompanied by increases in pro-atherogenic plasma lipids/lipoproteins (e.g., triglycerides, apolipoprotein B), activation of pro-atherogenic cytokine/chemokine signaling pathways in liver, and with circulating pro-atherogenic mediators of inflammation altogether providing a rationale for the differential effects of plant and animal fats.

Metabolic subtypes of patients with NAFLD exhibit distinctive cardiovascular risk profiles.

BACKGROUND AND AIMS: We previously identified subsets of patients with NAFLD with different metabolic phenotypes. Here we align metabolomic signatures with cardiovascular disease (CVD) and genetic risk factors. APPROACH AND RESULTS: We analyzed serum metabolome from 1154 individuals with biopsy-proven NAFLD, and from four mouse models of NAFLD with impaired VLDL-triglyceride (TG) secretion, and one with normal VLDL-TG secretion. We identified three metabolic subtypes: A (47%), B (27%), and C (26%). Subtype A phenocopied the metabolome of mice with impaired VLDL-TG secretion; subtype C phenocopied the metabolome of mice with normal VLDL-TG; and subtype B showed an intermediate signature. The percent of patients with NASH and fibrosis was comparable among subtypes, although subtypes B and C exhibited higher liver enzymes. Serum VLDL-TG levels and secretion rate were lower among subtype A compared with subtypes B and C. Subtype A VLDL-TG and VLDL-apolipoprotein B concentrations were independent of steatosis, whereas subtypes B and C showed an association with these parameters. Serum TG, cholesterol, VLDL, small dense LDL5,6 , and remnant lipoprotein cholesterol were lower among subtype A compared with subtypes B and C. The 10-year high risk of CVD, measured with the Framingham risk score, and the frequency of patatin-like phospholipase domain-containing protein 3 NAFLD risk allele were lower in subtype A. CONCLUSIONS: Metabolomic signatures identify three NAFLD subgroups, independent of histological disease severity. These signatures align with known CVD and genetic risk factors, with subtype A exhibiting a lower CVD risk profile. This may account for the variation in hepatic versus cardiovascular outcomes, offering clinically relevant risk stratification.

Intervention with isoleucine or valine corrects hyperinsulinemia and reduces intrahepatic diacylglycerols, liver steatosis, and inflammation in Ldlr-/-.Leiden mice with manifest obesity-associated NASH.

Non-alcoholic steatohepatitis (NASH) is associated with a disturbed metabolism in liver, insulin resistance, and excessive accumulation of ectopic fat. Branched-chain amino acids (BCAAs) may beneficially modulate hepatic lipids, however, it remains unclear whether individual BCAAs can attenuate already established NASH and associated oxidative-inflammatory stress. After a 26 weeks run-in on fast food diet (FFD), obese Ldlr-/-.Leiden mice were treated for another 12 weeks with either valine or isoleucine (3% of FFD) and then compared to FFD controls. Valine and isoleucine did not affect obesity, dyslipidemia, gut permeability, or fecal fatty acid excretion, but significantly reduced hyperinsulinemia. Valine and isoleucine reduced ALT, CK18-M30, and liver steatosis with a particularly pronounced suppression of the microvesicular component (-61% by valine and -71% by isoleucine). Both BCAAs decreased intrahepatic diacylglycerols and 4-hydroxynonenal immunoreactivity, a marker for oxidative stress-induced lipid peroxidation. Functional genomics analysis demonstrated that valine and isoleucine affected BCAA metabolism genes, deactivated master regulators of anabolic pathways related to steatosis (e.g., SREBPF1), and activated master regulators of mitochondrial biogenesis (e.g., PPARGC1A) and lipid catabolism (e.g., ACOX1, AMPK). This correction of critical metabolic pathways on gene expression level was accompanied by a significant decrease in histological liver inflammation, and suppression of FFD-stimulated cytokine and chemokine proteins KC/CXCL1, MCP-1/CCL2, and MIP-2/CXCL2 and their pathways. In conclusion, dietary intervention with either valine or isoleucine corrected liver diacylglycerols, gene expression of multiple metabolic processes, and reduced NASH histology with profound hepatoprotective effects on oxidative stress and inflammatory proteins.

Milk fat globule membrane attenuates high fat diet-induced neuropathological changes in obese Ldlr-/-.Leiden mice.

BACKGROUND: Milk-fat globule membrane (MFGM) is a complex structure secreted by the mammary gland and present in mammalian milk. MFGM contains lipids and glycoproteins as well as gangliosides, which may be involved in myelination processes. Notably, myelination and thereby white matter integrity are often altered in obesity. Furthermore, MFGM interventions showed beneficial effects in obesity by affecting inflammatory processes and the microbiome. In this study, we investigated the impact of a dietary MFGM intervention on fat storage, neuroinflammatory processes and myelination in a rodent model of high fat diet (HFD)-induced obesity. METHODS: 12-week-old male low density lipoprotein receptor-deficient Leiden mice were exposed to a HFD, a HFD enriched with 3% whey protein lipid concentrate (WPC) high in MFGM components, or a low fat diet. The impact of MFGM supplementation during 24-weeks of HFD-feeding was examined over time by analyzing body weight and fat storage, assessing cognitive tasks and MRI scanning, analyzing myelinization with polarized light imaging and examining neuroinflammation using immunohistochemistry. RESULTS: We found in this study that 24 weeks of HFD-feeding induced excessive fat storage, increased systolic blood pressure, altered white matter integrity, decreased functional connectivity, induced neuroinflammation and impaired spatial memory. Notably, supplementation with 3% WPC high in MFGM components restored HFD-induced neuroinflammation and attenuated the reduction in hippocampal-dependent spatial memory and hippocampal functional connectivity. CONCLUSIONS: We showed that supplementation with WPC high in MFGM components beneficially contributed to hippocampal-dependent spatial memory, functional connectivity in the hippocampus and anti-inflammatory processes in HFD-induced obesity in rodents. Current knowledge regarding exact biological mechanisms underlying these effects should be addressed in future studies.

Heat-Inactivated Akkermansia muciniphila Improves Gut Permeability but Does Not Prevent Development of Non-Alcoholic Steatohepatitis in Diet-Induced Obese Ldlr-/-.Leiden Mice.

The development of non-alcoholic steatohepatitis (NASH) has been associated with alterations in gut microbiota composition and reduced gut barrier function. Akkermansia muciniphila is a gut microbe that is thought to have health-promoting properties, including the ability to improve gut barrier function and host metabolism, both when administered live and after heat-inactivation. We questioned whether heat-inactivated A. muciniphila may reduce NASH development. Ldlr-/-.Leiden mice, a translational, diet-induced model for NASH, were fed a NASH-inducing high-fat diet (HFD) supplemented with heat-inactivated A. muciniphila. After 28 weeks, effects of the treatment on obesity and associated metabolic dysfunction in the gut (microbiota composition and permeability), adipose tissue, and liver were studied relative to an untreated HFD control. Treatment with heat-inactivated A. muciniphila did not affect body weight or adiposity and had no effect on plasma lipids, blood glucose, or plasma insulin. Heat-inactivated A. muciniphila had some minor effects on mucosal microbiota composition in ileum and colon and improved gut barrier function, as assessed by an in vivo functional gut permeability test. Epidydimal white adipose tissue (WAT) hypertrophy and inflammation were not affected, but heat-inactivated A. muciniphila did reduce hypertrophy in the mesenteric WAT which is in close proximity to the intestine. Heat-inactivated A. muciniphila did not affect the development of NASH or associated fibrosis in the liver and did not affect circulating bile acids or markers of liver fibrosis, but did reduce PRO-C4, a type IV collagen synthesis marker, which may be associated with gut integrity. In conclusion, despite beneficial effects in the gut and mesenteric adipose tissue, heat-inactivated A. muciniphila did not affect the development of NASH and fibrosis in a chronic disease setting that mimics clinically relevant disease stages.

Therapeutic Intervention with Anti-Complement Component 5 Antibody Does Not Reduce NASH but Does Attenuate Atherosclerosis and MIF Concentrations in Ldlr-/-.Leiden Mice.

BACKGROUND: Chronic inflammation is an important driver in the progression of non-alcoholic steatohepatitis (NASH) and atherosclerosis. The complement system, one of the first lines of defense in innate immunity, has been implicated in both diseases. However, the potential therapeutic value of complement inhibition in the ongoing disease remains unclear. METHODS: After 20 weeks of high-fat diet (HFD) feeding, obese Ldlr-/-.Leiden mice were treated twice a week with an established anti-C5 antibody (BB5.1) or vehicle control. A separate group of mice was kept on a chow diet as a healthy reference. After 12 weeks of treatment, NASH was analyzed histopathologically, and genome-wide hepatic gene expression was analyzed by next-generation sequencing and pathway analysis. Atherosclerotic lesion area and severity were quantified histopathologically in the aortic roots. RESULTS: Anti-C5 treatment considerably reduced complement system activity in plasma and MAC deposition in the liver but did not affect NASH. Anti-C5 did, however, reduce the development of atherosclerosis, limiting the total lesion size and severity independently of an effect on plasma cholesterol but with reductions in oxidized LDL (oxLDL) and macrophage migration inhibitory factor (MIF). CONCLUSION: We show, for the first time, that treatment with an anti-C5 antibody in advanced stages of NASH is not sufficient to reduce the disease, while therapeutic intervention against established atherosclerosis is beneficial to limit further progression.

Intramuscular short-chain acylcarnitines in elderly people are decreased in (pre-)frail females, but not in males.

This study tested the hypothesis that in human aging, a decreased intramuscular acylcarnitine status is associated with (pre-)frailty, reduced physical performance, and altered mitochondrial function. We used a cross-sectional study design with well-matched fit and (pre-)frail old males and females, using young males and females as healthy controls. Frailty was assessed according to the Fried criteria and physical performance was determined by 400 m walk test, short physical performance battery and handgrip strength. Muscle and plasma acylcarnitine status, and muscle mitochondrial gene expression was analyzed. Results showed that intramuscular total carnitine levels and short-chain acylcarnitine levels were lower in (pre-)frail old females compared to fit old females and young females, whereas no differences were observed in males. The low intramuscular short-chain acylcarnitine levels in females correlated with low physical performance, even after correction for muscle mass (%), and were accompanied with lowered expression of genes involved in mitochondrial energy production and functionality. It is, therefore, concluded that in (pre-)frail old females, intramuscular total carnitine levels and short-chain acylcarnitine levels are decreased, and this decrease is associated with reduced physical performance and low expression of a wide range of genes critical for mitochondrial function. The results stress the importance of taking sex differences into account in aging research.

Propionic acid and not caproic acid, attenuates nonalcoholic steatohepatitis and improves (cerebro) vascular functions in obese Ldlr-/- .Leiden mice.

The obesity epidemic increases the interest to elucidate impact of short-chain fatty acids on metabolism, obesity, and the brain. We investigated the effects of propionic acid (PA) and caproic acid (CA) on metabolic risk factors, liver and adipose tissue pathology, brain function, structure (by MRI), and gene expression, during obesity development in Ldlr-/- .Leiden mice. Ldlr-/- .Leiden mice received 16 weeks either a high-fat diet (HFD) to induce obesity, or chow as reference group. Next, obese HFD-fed mice were treated 12 weeks with (a) HFD + CA (CA), (b) HFD + PA (PA), or (c) a HFD-control group. PA reduced the body weight and systolic blood pressure, lowered fasting insulin levels, and reduced HFD-induced liver macrovesicular steatosis, hypertrophy, inflammation, and collagen content. PA increased the amount of glucose transporter type 1-positive cerebral blood vessels, reverted cerebral vasoreactivity, and HFD-induced effects in microstructural gray and white matter integrity of optic tract, and somatosensory and visual cortex. PA and CA also reverted HFD-induced effects in functional connectivity between visual and auditory cortex. However, PA mice were more anxious in open field, and showed reduced activity of synaptogenesis and glutamate regulators in hippocampus. Therefore, PA treatment should be used with caution even though positive metabolic, (cerebro) vascular, and brain structural and functional effects were observed.

FURIN Inhibition Reduces Vascular Remodeling and Atherosclerotic Lesion Progression in Mice.

Objective- Atherosclerotic coronary artery disease is the leading cause of death worldwide, and current treatment options are insufficient. Using systems-level network cluster analyses on a large coronary artery disease case-control cohort, we previously identified PCSK3 (proprotein convertase subtilisin/kexin family member 3; FURIN) as a member of several coronary artery disease-associated pathways. Thus, our objective is to determine the role of FURIN in atherosclerosis. Approach and Results- In vitro, FURIN inhibitor treatment resulted in reduced monocyte migration and reduced macrophage and vascular endothelial cell inflammatory and cytokine gene expression. In vivo, administration of an irreversible inhibitor of FURIN, α-1-PDX (α1-antitrypsin Portland), to hyperlipidemic Ldlr-/- mice resulted in lower atherosclerotic lesion area and a specific reduction in severe lesions. Significantly lower lesional macrophage and collagen area, as well as systemic inflammatory markers, were observed. MMP2 (matrix metallopeptidase 2), an effector of endothelial function and atherosclerotic lesion progression, and a FURIN substrate was significantly reduced in the aorta of inhibitor-treated mice. To determine FURIN's role in vascular endothelial function, we administered α-1-PDX to Apoe-/- mice harboring a wire injury in the common carotid artery. We observed significantly decreased carotid intimal thickness and lower plaque cellularity, smooth muscle cell, macrophage, and inflammatory marker content, suggesting protection against vascular remodeling. Overexpression of FURIN in this model resulted in a significant 67% increase in intimal plaque thickness, confirming that FURIN levels directly correlate with atherosclerosis. Conclusions- We show that systemic inhibition of FURIN in mice decreases vascular remodeling and atherosclerosis. FURIN-mediated modulation of MMP2 activity may contribute to the atheroprotection observed in these mice.

Combined Treatment with L-Carnitine and Nicotinamide Riboside Improves Hepatic Metabolism and Attenuates Obesity and Liver Steatosis.

Obesity characterized by adiposity and ectopic fat accumulation is associated with the development of non-alcoholic fatty liver disease (NAFLD). Treatments that stimulate lipid utilization may prevent the development of obesity and comorbidities. This study evaluated the potential anti-obesogenic hepatoprotective effects of combined treatment with L-carnitine and nicotinamide riboside, i.e., components that can enhance fatty acid transfer across the inner mitochondrial membrane and increase nicotinamide adenine nucleotide (NAD+) levels, which are necessary for ß-oxidation and the TCA cycle, respectively. Ldlr -/-.Leiden mice were treated with high-fat diet (HFD) supplemented with L-carnitine (LC; 0.4% w/w), nicotinamide riboside (NR; 0.3% w/w) or both (COMBI) for 21 weeks. L-carnitine plasma levels were reduced by HFD and normalized by LC. NR supplementation raised its plasma metabolite levels demonstrating effective delivery. Although food intake and ambulatory activity were comparable in all groups, COMBI treatment significantly attenuated HFD-induced body weight gain, fat mass gain (-17%) and hepatic steatosis (-22%). Also, NR and COMBI reduced hepatic 4-hydroxynonenal adducts. Upstream-regulator gene analysis demonstrated that COMBI reversed detrimental effects of HFD on liver metabolism pathways and associated regulators, e.g., ACOX, SCAP, SREBF, PPARGC1B, and INSR. Combination treatment with LC and NR exerts protective effects on metabolic pathways and constitutes a new approach to attenuate HFD-induced obesity and NAFLD.

Cardiovascular effects among workers exposed to multiwalled carbon nanotubes.

OBJECTIVES: The increase in production of multiwalled carbon nanotubes (MWCNTs) has led to growing concerns about health risks. In this study, we assessed the association between occupational exposure to MWCNTs and cardiovascular biomarkers. METHODS: A cross-sectional study was performed among 22 workers of a company commercially producing MWCNTs (subdivided into lab personnel with low or high exposure and operators), and a gender and age-matched unexposed population (n=42). Exposure to MWCNTs and 12 cardiovascular markers were measured in participants' blood (phase I). In a subpopulation of 13 exposed workers and six unexposed workers, these measures were repeated after 5 months (phase II). We analysed associations between MWCNT exposure and biomarkers of cardiovascular risk, adjusted for age, body mass index, sex and smoking. RESULTS: We observed an upward trend in the concentration of endothelial damage marker intercellular adhesion molecule-1 (ICAM-1), with increasing exposure to MWCNTs in both phases. The operator category showed significantly elevated ICAM-1 geometric mean ratios (GMRs) compared with the controls (phase I: GMR=1.40, P=1.30E-3; phase II: GMR=1.37, P=0.03). The trends were significant both across worker categories (phase I: P=1.50E-3; phase II: P=0.01) and across measured GM MWCNT concentrations (phase I: P=3.00E-3; phase II: P=0.01). No consistent significant associations were found for the other cardiovascular markers. CONCLUSION: The associations between MWCNT exposure and ICAM-1 indicate endothelial activation and an increased inflammatory state in workers with MWCNT exposure.

Diet-Independent Correlations between Bacteria and Dysfunction of Gut, Adipose Tissue, and Liver: A Comprehensive Microbiota Analysis in Feces and Mucosa of the Ileum and Colon in Obese Mice with NAFLD.

Development of non-alcoholic fatty liver disease (NAFLD) is linked to obesity, adipose tissue inflammation, and gut dysfunction, all of which depend on diet. So far, studies have mainly focused on diet-related fecal microbiota changes, but other compartments may be more informative on host health. We present a first systematic analysis of microbiota changes in the ileum and colon using multiple diets and investigating both fecal and mucosal samples. Ldlr-/-.Leiden mice received one of three different energy-dense (ED)-diets (n = 15/group) for 15 weeks. All of the ED diets induced obesity and metabolic risk factors, altered short-chain fatty acids (SCFA), and increased gut permeability and NAFLD to various extents. ED diets reduced the diversity of high-abundant bacteria and increased the diversity of low-abundant bacteria in all of the gut compartments. The ED groups showed highly variable, partially overlapping microbiota compositions that differed significantly from chow. Correlation analyses demonstrated that (1) specific groups of bacteria correlate with metabolic risk factors, organ dysfunction, and NAFLD endpoints, (2) colon mucosa had greater predictive value than other compartments, (3) correlating bacteria differed per compartment, and (4) some bacteria correlated with plasma SCFA levels. In conclusion, this comprehensive microbiota analysis demonstrates correlations between the microbiota and dysfunctions of gut, adipose tissue, and liver, independent of a specific disease-inducing diet.

Effects of Anthocyanin and Flavanol Compounds on Lipid Metabolism and Adipose Tissue Associated Systemic Inflammation in Diet-Induced Obesity.

Background. Naturally occurring substances from the flavanol and anthocyanin family of polyphenols have been proposed to exert beneficial effects in the course of obesity. We hypothesized that their effects on attenuating obesity-induced dyslipidemia as well as the associated inflammatory sequelae especially have health-promoting potential. Methods. Male C57BL/6J mice (n = 52) received a control low-fat diet (LFD; 10 kcal% fat) for 6 weeks followed by 24 weeks of either LFD (n = 13) or high-fat diet (HFD; 45 kcal% fat; n = 13) or HFD supplemented with 0.1% w/w of the flavanol compound epicatechin (HFD+E; n = 13) or an anthocyanin-rich bilberry extract (HFD+B; n = 13). Energy substrate utilization was determined by indirect calorimetry in a subset of mice following the dietary switch and at the end of the experiment. Blood samples were collected at baseline and at 3 days and 4, 12, and 20 weeks after dietary switch and analyzed for systemic lipids and proinflammatory cytokines. Adipose tissue (AT) histopathology and inflammatory gene expression as well as hepatic lipid content were analyzed after sacrifice. Results. The switch from a LFD to a HFD lowered the respiratory exchange ratio and increased plasma cholesterol and hepatic lipid content. These changes were not attenuated by HFD+E or HFD+B. Furthermore, the polyphenol compounds could not prevent HFD-induced systemic rise of TNF-α levels. Interestingly, a significant reduction in Tnf gene expression in HFD+B mice was observed in the AT. Furthermore, HFD+B, but not HFD+E, significantly prevented the early upregulation of circulating neutrophil chemoattractant mKC. However, no differences in AT histopathology were observed between the HFD types. Conclusion. Supplementation of HFD with an anthocyanin-rich bilberry extract but not with the flavanol epicatechin may exert beneficial effects on the systemic early inflammatory response associated with diet-induced obesity. These systemic effects were transient and not observed after prolongation of HFD-feeding (24 weeks). On the tissue level, long-term treatment with bilberry attenuated TNF-α expression in adipose tissue.

Mirtoselect, an anthocyanin-rich bilberry extract, attenuates non-alcoholic steatohepatitis and associated fibrosis in ApoE(∗)3Leiden mice.

BACKGROUND & AIMS: Anthocyanins may have beneficial effects on lipid metabolism and inflammation and are demonstrated to have hepatoprotective properties in models of restraint-stress- and chemically-induced liver damage. However, their potential to protect against non-alcoholic steatohepatitis (NASH) under conditions relevant for human pathogenesis remains unclear. Therefore, we studied the effects of the standardised anthocyanin-rich extract Mirtoselect on diet-induced NASH in a translational model of disease. METHODS: ApoE(∗)3Leiden mice were fed a Western-type cholesterol-containing diet without (HC) or with 0.1% (w/w) Mirtoselect (HCM) for 20weeks to study the effects on diet-induced NASH. RESULTS: Mirtoselect attenuated HC-induced hepatic steatosis, as observed by decreased macro- and microvesicular hepatocellular lipid accumulation and reduced hepatic cholesteryl ester content. This anti-steatotic effect was accompanied by local anti-inflammatory effects in liver, as demonstrated by reduced inflammatory cell clusters and reduced neutrophil infiltration in HCM. On a molecular level, HC diet significantly induced hepatic expression of pro-inflammatory genes Tnf, Emr1, Ccl2, Mpo, Cxcl1, and Cxcl2 while this induction was less pronounced or significantly decreased in HCM. A similar quenching effect was observed for HC-induced pro-fibrotic genes, Acta2 and Col1a1 and this anti-fibrotic effect of Mirtoselect was confirmed histologically. Many of the pro-inflammatory and pro-fibrotic parameters positively correlated with intrahepatic free cholesterol levels. Mirtoselect significantly reduced accumulation and crystallisation of intrahepatic free cholesterol, providing a possible mechanism for the observed hepatoprotective effects. CONCLUSIONS: Mirtoselect attenuates development of NASH, reducing hepatic lipid accumulation, inflammation and fibrosis, possibly mediated by local anti-inflammatory effects associated with reduced accumulation and crystallisation of intrahepatic free cholesterol.

Metabolically induced liver inflammation leads to NASH and differs from LPS- or IL-1ß-induced chronic inflammation.

The nature of the chronic inflammatory component that drives the development of non-alcoholic steatohepatitis (NASH) is unclear and possible inflammatory triggers have not been investigated systematically. We examined the effect of non-metabolic triggers (lipopolysaccharide (LPS), interleukin-1ß (IL-1ß), administered by slow-release minipumps) and metabolic dietary triggers (carbohydrate, cholesterol) of inflammation on the progression of bland liver steatosis (BS) to NASH. Transgenic APOE3*Leiden.huCETP (APOE3L.CETP) mice fed a high-fat diet (HFD) developed BS after 10 weeks. Then, inflammatory triggers were superimposed or not (control) for six more weeks. Mouse livers were analyzed with particular emphasis on hallmarks of inflammation which were defined in human liver biopsies with and without NASH. Livers of HFD-treated control mice remained steatotic and did not progress to NASH. All four inflammatory triggers activated hepatic nuclear factor-κB (NF-κB) significantly and comparably (≥5-fold). However, HFD+LPS or HFD+IL-1ß did not induce a NASH-like phenotype and caused intrahepatic accumulation of almost exclusively mononuclear cells. By contrast, mice treated with metabolic triggers developed NASH, characterized by enhanced steatosis, hepatocellular hypertrophy, and formation of mixed-type inflammatory foci containing myeloperoxidase-positive granulocytes (neutrophils) as well as mononuclear cells, essentially as observed in human NASH. Specific for the metabolic inducers was an activation of the proinflammatory transcription factor activator protein-1 (AP-1), neutrophil infiltration, and induction of risk factors associated with human NASH, that is, dyslipidemia (by cholesterol) and insulin resistance (by carbohydrate). In conclusion, HFD feeding followed by NF-κB activation per se (LPS, IL-1ß) does not promote the transition from BS to NASH. HFD feeding followed by metabolically evoked inflammation induces additional inflammatory components (neutrophils, AP-1 pathway) and causes NASH.

MIF is a noncognate ligand of CXC chemokine receptors in inflammatory and atherogenic cell recruitment.

The cytokine macrophage migration inhibitory factor (MIF) plays a critical role in inflammatory diseases and atherogenesis. We identify the chemokine receptors CXCR2 and CXCR4 as functional receptors for MIF. MIF triggered G(alphai)- and integrin-dependent arrest and chemotaxis of monocytes and T cells, rapid integrin activation and calcium influx through CXCR2 or CXCR4. MIF competed with cognate ligands for CXCR4 and CXCR2 binding, and directly bound to CXCR2. CXCR2 and CD74 formed a receptor complex, and monocyte arrest elicited by MIF in inflamed or atherosclerotic arteries involved both CXCR2 and CD74. In vivo, Mif deficiency impaired monocyte adhesion to the arterial wall in atherosclerosis-prone mice, and MIF-induced leukocyte recruitment required Il8rb (which encodes Cxcr2). Blockade of Mif but not of canonical ligands of Cxcr2 or Cxcr4 in mice with advanced atherosclerosis led to plaque regression and reduced monocyte and T-cell content in plaques. By activating both CXCR2 and CXCR4, MIF displays chemokine-like functions and acts as a major regulator of inflammatory cell recruitment and atherogenesis. Targeting MIF in individuals with manifest atherosclerosis can potentially be used to treat this condition.

Seven robust and easy to obtain biomarkers to measure acute stress.

With the purpose of identifying a sensitive, robust, and easy-to-measure set of biomarkers to assess stress reactivity, we here study a large set of relatively easy to obtain markers reflecting subjective, autonomic nervous system (ANS), endocrine, and inflammatory responses to acute social stress (n = 101). A subset of the participants was exposed to another social stressor the next day (n = 48) while being measured in the same way. Acute social stress was induced following standardized procedures. The markers investigated were self-reported positive and negative affect, heart rate, electrodermal activity, salivary cortisol, and ten inflammatory markers both in capillary plasma and salivary samples, including IL-22 which has not been studied in response to acute stress in humans before. Robust effects (significant effect in the same direction for both days) were found for self-reported negative affect, heart rate, electrodermal activity, plasma IL-5, plasma IL-22, salivary IL-8 and salivary IL-10. Of these seven markers, the participants' IL-22 responses on the first day were positively correlated to those on the second day. We found no correlations between salivary and capillary plasma stress responses for any of the ten cytokines and somewhat unexpectedly, cytokine responses in saliva seemed more pronounced and more in line with previous literature than cytokines in capillary plasma. In sum, seven robust and easy to obtain biomarkers to measure acute stress response were identified and should be used in future stress research to detect and examine stress reactivity. This includes IL-22 in plasma as a promising novel marker.

Translatability of mouse muscle-aging for humans: the role of sex.

Muscle-aging drives sarcopenia and is a major public health issue. Mice are frequently used as a model for human muscle-aging, however, research investigating their translational value is limited. In addition, mechanisms underlying muscle-aging may have sex-specific features in humans, but it is not yet assessed whether these are recapitulated in mice. Here, we studied the effects of aging on a functional, histological and transcriptional level at multiple timepoints in male and female mice (4, 17, 21 and 25 months), with particular emphasis on sex-differences. The effects of natural aging on the transcriptome of quadriceps muscle were compared to humans on pathway level. Significant loss of muscle mass occurred late, at 25 months, in both male (-17%, quadriceps) and female mice (-10%, quadriceps) compared to young control mice. Concomitantly, we found in female, but not male mice, a slower movement speed in the aged groups compared to the young mice (P < 0.001). Consistently, weighted gene co-expression network analysis revealed a stronger association between the aging-related reduction of movement and aging-related changes in muscle transcriptome of female compared to male mice (P < 0.001). In male, but not female mice, major distinctive aging-related changes occurred in the last age group (25 months), which highlights the necessity for careful selection of age using mice as a muscle-aging model. Furthermore, contrasting to humans, more aging-related changes were found in the muscle transcriptome of male mice compared to female mice (4090 vs. 2285 differentially expressed genes at 25 months, respectively). Subsequently, male mice recapitulated more muscle-aging related pathways characteristic for both male and female humans. In conclusion, our data show that sex has a critical effect on the mouse muscle-aging trajectory, although these do not necessarily reflect sex differences observed in the human muscle-aging trajectory.

Long-Term Brain Structure and Cognition Following Bariatric Surgery.

Importance: Weight loss induced by bariatric surgery (BS) is associated with improved cognition and changed brain structure; however, previous studies on the association have used small cohorts and short follow-up periods, making it difficult to determine long-term neurological outcomes associated with BS. Objective: To investigate long-term associations of weight loss after BS with cognition and brain structure and perfusion. Design, Setting, and Participants: This cohort study included participants from the Bariatric Surgery Rijnstate and Radboudumc Neuroimaging and Cognition in Obesity study. Data from participants with severe obesity (body mass index [BMI; calculated as weight in kilograms divided by height in meters squared] >40, or BMI >35 with comorbidities) eligible for Roux-en-Y gastric bypass and aged 35 to 55 years were enrolled from a hospital specialized in BS (Rijnstate Hospital, Arnhem, the Netherlands). Participants were recruited between September 2018 and December 2020 with follow-up till March 2023. Data were collected before BS and at 6 and 24 months after BS. Data were analyzed from March to November 2023. Exposure: Roux-en-Y gastric bypass. Main Outcomes and Measures: Primary outcomes included body weight, BMI, waist circumference, blood pressure, medication use, cognitive performance (20% change index of compound z-score), brain volumes, cortical thickness, cerebral blood flow (CBF), and spatial coefficient of variation (sCOV). Secondary outcomes include cytokines, adipokines, depressive symptoms (assessed using the Beck Depression Inventory), and physical activity (assessed using the Baecke Questionnaire). Results: A total of 133 participants (mean [SD] age, 46.8 [5.7] years; 112 [84.2%] female) were included. Global cognition was at least 20% higher in 52 participants (42.9%) at 24 months after BS. Compared with baseline, at 24 months, inflammatory markers were lower (mean [SD] high-sensitivity C-reactive protein: 4.77 [5.80] µg/mL vs 0.80 [1.09] µg/mL; P < .001), fewer patients used antihypertensives (48 patients [36.1%] vs 22 patients [16.7%]), and patients had lower depressive symptoms (median [IQR] BDI score: 9.0 [5.0-13.0] vs 3.0 [1.0-6.0]; P < .001) and greater physical activity (mean [SD] Baecke score: 7.64 [1.29] vs 8.19 [1.35]; P < .001). After BS, brain structure and perfusion were lower in most brain regions, while hippocampal and white matter volume remained stable. CBF and sCOV did not change in nucleus accumbens and parietal cortex. The temporal cortex showed a greater thickness (mean [SD] thickness: 2.724 [0.101] mm vs 2.761 [0.007] mm; P = .007) and lower sCOV (median [IQR] sCOV: 4.41% [3.83%-5.18%] vs 3.97% [3.71%-4.59%]; P = .02) after BS. Conclusions and Relevance: These findings suggest that BS was associated with health benefits 2 years after surgery. BS was associated with improved cognition and general health and changed blood vessel efficiency and cortical thickness of the temporal cortex. These results may improve treatment options for patients with obesity and dementia.