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1.
Nat Immunol ; 24(9): 1473-1486, 2023 09.
Article in English | MEDLINE | ID: mdl-37580603

ABSTRACT

Omnivorous animals, including mice and humans, tend to prefer energy-dense nutrients rich in fat over plant-based diets, especially for short periods of time, but the health consequences of this short-term consumption of energy-dense nutrients are unclear. Here, we show that short-term reiterative switching to 'feast diets', mimicking our social eating behavior, breaches the potential buffering effect of the intestinal microbiota and reorganizes the immunological architecture of mucosa-associated lymphoid tissues. The first dietary switch was sufficient to induce transient mucosal immune depression and suppress systemic immunity, leading to higher susceptibility to Salmonella enterica serovar Typhimurium and Listeria monocytogenes infections. The ability to respond to antigenic challenges with a model antigen was also impaired. These observations could be explained by a reduction of CD4+ T cell metabolic fitness and cytokine production due to impaired mTOR activity in response to reduced microbial provision of fiber metabolites. Reintroducing dietary fiber rewired T cell metabolism and restored mucosal and systemic CD4+ T cell functions and immunity. Finally, dietary intervention with human volunteers confirmed the effect of short-term dietary switches on human CD4+ T cell functionality. Therefore, short-term nutritional changes cause a transient depression of mucosal and systemic immunity, creating a window of opportunity for pathogenic infection.


Subject(s)
Mucous Membrane , Salmonella typhimurium , Humans , Mice , Animals , T-Lymphocytes , Immunity, Mucosal
2.
Nature ; 625(7994): 385-392, 2024 Jan.
Article in English | MEDLINE | ID: mdl-38123683

ABSTRACT

Digested dietary fats are taken up by enterocytes where they are assembled into pre-chylomicrons in the endoplasmic reticulum followed by transport to the Golgi for maturation and subsequent secretion to the circulation1. The role of mitochondria in dietary lipid processing is unclear. Here we show that mitochondrial dysfunction in enterocytes inhibits chylomicron production and the transport of dietary lipids to peripheral organs. Mice with specific ablation of the mitochondrial aspartyl-tRNA synthetase DARS2 (ref. 2), the respiratory chain subunit SDHA3 or the assembly factor COX10 (ref. 4) in intestinal epithelial cells showed accumulation of large lipid droplets (LDs) in enterocytes of the proximal small intestine and failed to thrive. Feeding a fat-free diet suppressed the build-up of LDs in DARS2-deficient enterocytes, which shows that the accumulating lipids derive mostly from digested fat. Furthermore, metabolic tracing studies revealed an impaired transport of dietary lipids to peripheral organs in mice lacking DARS2 in intestinal epithelial cells. DARS2 deficiency caused a distinct lack of mature chylomicrons concomitant with a progressive dispersal of the Golgi apparatus in proximal enterocytes. This finding suggests that mitochondrial dysfunction results in impaired trafficking of chylomicrons from the endoplasmic reticulum to the Golgi, which in turn leads to storage of dietary lipids in large cytoplasmic LDs. Taken together, these results reveal a role for mitochondria in dietary lipid transport in enterocytes, which might be relevant for understanding the intestinal defects observed in patients with mitochondrial disorders5.


Subject(s)
Dietary Fats , Enterocytes , Lipid Metabolism , Mitochondria , Animals , Mice , Aspartate-tRNA Ligase/metabolism , Chylomicrons/metabolism , Dietary Fats/metabolism , Electron Transport Complex II/metabolism , Endoplasmic Reticulum/metabolism , Enterocytes/metabolism , Enterocytes/pathology , Epithelial Cells/metabolism , Golgi Apparatus/metabolism , Intestines , Lipid Droplets/metabolism , Mitochondria/metabolism , Mitochondria/pathology
3.
Nature ; 609(7926): 361-368, 2022 09.
Article in English | MEDLINE | ID: mdl-35790189

ABSTRACT

Brown adipose tissue (BAT) dissipates energy1,2 and promotes cardiometabolic health3. Loss of BAT during obesity and ageing is a principal hurdle for BAT-centred obesity therapies, but not much is known about BAT apoptosis. Here, untargeted metabolomics demonstrated that apoptotic brown adipocytes release a specific pattern of metabolites with purine metabolites being highly enriched. This apoptotic secretome enhances expression of the thermogenic programme in healthy adipocytes. This effect is mediated by the purine inosine that stimulates energy expenditure in brown adipocytes by the cyclic adenosine monophosphate-protein kinase A signalling pathway. Treatment of mice with inosine increased BAT-dependent energy expenditure and induced 'browning' of white adipose tissue. Mechanistically, the equilibrative nucleoside transporter 1 (ENT1, SLC29A1) regulates inosine levels in BAT: ENT1-deficiency increases extracellular inosine levels and consequently enhances thermogenic adipocyte differentiation. In mice, pharmacological inhibition of ENT1 as well as global and adipose-specific ablation enhanced BAT activity and counteracted diet-induced obesity, respectively. In human brown adipocytes, knockdown or blockade of ENT1 increased extracellular inosine, which enhanced thermogenic capacity. Conversely, high ENT1 levels correlated with lower expression of the thermogenic marker UCP1 in human adipose tissues. Finally, the Ile216Thr loss of function mutation in human ENT1 was associated with significantly lower body mass index and 59% lower odds of obesity for individuals carrying the Thr variant. Our data identify inosine as a metabolite released during apoptosis with a 'replace me' signalling function that regulates thermogenic fat and counteracts obesity.


Subject(s)
Adipocytes, Brown , Adipose Tissue, Brown , Energy Metabolism , Inosine , Adipocytes, Brown/drug effects , Adipocytes, Brown/metabolism , Adipose Tissue, Brown/drug effects , Adipose Tissue, Brown/metabolism , Adipose Tissue, White/drug effects , Adipose Tissue, White/metabolism , Animals , Energy Metabolism/drug effects , Equilibrative Nucleoside Transporter 1/antagonists & inhibitors , Equilibrative Nucleoside Transporter 1/metabolism , Humans , Inosine/metabolism , Inosine/pharmacology , Mice , Obesity/genetics , Obesity/metabolism , Thermogenesis/genetics , Uncoupling Protein 1/metabolism
4.
PLoS Pathog ; 20(9): e1012509, 2024 Sep.
Article in English | MEDLINE | ID: mdl-39241103

ABSTRACT

The replication organelle of hepatitis C virus (HCV), called membranous web, is derived from the endoplasmic reticulum (ER) and mainly comprises double membrane vesicles (DMVs) that concentrate the viral replication complexes. It also tightly associates with lipid droplets (LDs), which are essential for virion morphogenesis. In particular acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1), a rate-limiting enzyme in triglyceride synthesis, promotes early steps of virus assembly. The close proximity between ER membranes, DMVs and LDs therefore permits the efficient coordination of the HCV replication cycle. Here, we demonstrate that exaggerated LD accumulation due to the excessive expression of the DGAT1 isozyme, DGAT2, dramatically impairs the formation of the HCV membranous web. This effect depended on the enzymatic activity and ER association of DGAT2, whereas the mere LD accumulation was not sufficient to hamper HCV RNA replication. Our lipidomics data indicate that both HCV infection and DGAT2 overexpression induced membrane lipid biogenesis and markedly increased phospholipids with long chain polyunsaturated fatty acids, suggesting a dual use of these lipids and their possible competition for LD and DMV biogenesis. On the other hand, overexpression of DGAT2 depleted specific phospholipids, particularly oleyl fatty acyl chain-containing phosphatidylcholines, which, in contrast, are increased in HCV-infected cells and likely essential for viral infection. In conclusion, our results indicate that lipid exchanges occurring during LD biogenesis regulate the composition of intracellular membranes and thereby affect the formation of the HCV replication organelle. The potent antiviral effect observed in our DGAT2 overexpression system unveils lipid flux that may be relevant in the context of steatohepatitis, a hallmark of HCV infection, but also in physiological conditions, locally in specific subdomains of the ER membrane. Thus, LD formation mediated by DGAT1 and DGAT2 might participate in the spatial compartmentalization of HCV replication and assembly factories within the membranous web.


Subject(s)
Diacylglycerol O-Acyltransferase , Endoplasmic Reticulum , Hepacivirus , Triglycerides , Virus Replication , Diacylglycerol O-Acyltransferase/metabolism , Diacylglycerol O-Acyltransferase/genetics , Humans , Hepacivirus/physiology , Virus Replication/physiology , Triglycerides/metabolism , Endoplasmic Reticulum/metabolism , Endoplasmic Reticulum/virology , Hepatitis C/metabolism , Hepatitis C/virology , Lipid Droplets/metabolism , Lipid Droplets/virology
5.
Immunity ; 47(3): 566-581.e9, 2017 09 19.
Article in English | MEDLINE | ID: mdl-28930663

ABSTRACT

Microglia play a pivotal role in the maintenance of brain homeostasis but lose homeostatic function during neurodegenerative disorders. We identified a specific apolipoprotein E (APOE)-dependent molecular signature in microglia from models of amyotrophic lateral sclerosis (ALS), multiple sclerosis (MS), and Alzheimer's disease (AD) and in microglia surrounding neuritic ß-amyloid (Aß)-plaques in the brains of people with AD. The APOE pathway mediated a switch from a homeostatic to a neurodegenerative microglia phenotype after phagocytosis of apoptotic neurons. TREM2 (triggering receptor expressed on myeloid cells 2) induced APOE signaling, and targeting the TREM2-APOE pathway restored the homeostatic signature of microglia in ALS and AD mouse models and prevented neuronal loss in an acute model of neurodegeneration. APOE-mediated neurodegenerative microglia had lost their tolerogenic function. Our work identifies the TREM2-APOE pathway as a major regulator of microglial functional phenotype in neurodegenerative diseases and serves as a novel target that could aid in the restoration of homeostatic microglia.


Subject(s)
Apolipoproteins E/metabolism , Membrane Glycoproteins/metabolism , Microglia/metabolism , Neurodegenerative Diseases/genetics , Neurodegenerative Diseases/metabolism , Receptors, Immunologic/metabolism , Signal Transduction , Transcriptome , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Alzheimer Disease/pathology , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/metabolism , Animals , Apolipoproteins E/deficiency , Apolipoproteins E/genetics , Apoptosis/genetics , Apoptosis/immunology , Cerebral Cortex/metabolism , Cerebral Cortex/pathology , Cluster Analysis , Disease Models, Animal , Encephalomyelitis, Autoimmune, Experimental , Female , Gene Expression Profiling , Gene Expression Regulation , Gene Targeting , Humans , Immune Tolerance , Mice , Mice, Knockout , Mice, Transgenic , Microglia/immunology , Monocytes/immunology , Monocytes/metabolism , Neurodegenerative Diseases/immunology , Neurons/metabolism , Phagocytosis/genetics , Phagocytosis/immunology , Phenotype , Plaque, Amyloid/metabolism , Plaque, Amyloid/pathology , Superoxide Dismutase-1/genetics , Superoxide Dismutase-1/metabolism , Transforming Growth Factor beta/metabolism
6.
Hepatology ; 78(5): 1418-1432, 2023 11 01.
Article in English | MEDLINE | ID: mdl-36053190

ABSTRACT

BACKGROUND AND AIMS: The assembly and secretion of VLDL from the liver, a pathway that affects hepatic and plasma lipids, remains incompletely understood. We set out to identify players in the VLDL biogenesis pathway by identifying genes that are co-expressed with the MTTP gene that encodes for microsomal triglyceride transfer protein, key to the lipidation of apolipoprotein B, the core protein of VLDL. Using human and murine transcriptomic data sets, we identified small leucine-rich protein 1 ( SMLR1 ), encoding for small leucine-rich protein 1, a protein of unknown function that is exclusively expressed in liver and small intestine. APPROACH AND RESULTS: To assess the role of SMLR1 in the liver, we used somatic CRISPR/CRISPR-associated protein 9 gene editing to silence murine Smlr1 in hepatocytes ( Smlr1 -LKO). When fed a chow diet, male and female mice show hepatic steatosis, reduced plasma apolipoprotein B and triglycerides, and reduced VLDL secretion without affecting microsomal triglyceride transfer protein activity. Immunofluorescence studies show that SMLR1 is in the endoplasmic reticulum and Cis-Golgi complex. The loss of hepatic SMLR1 in female mice protects against diet-induced hyperlipidemia and atherosclerosis but causes NASH. On a high-fat, high-cholesterol diet, insulin and glucose tolerance tests did not reveal differences in male Smlr1 -LKO mice versus controls. CONCLUSIONS: We propose a role for SMLR1 in the trafficking of VLDL from the endoplasmic reticulum to the Cis-Golgi complex. While this study uncovers SMLR1 as a player in the VLDL assembly, trafficking, and secretion pathway, it also shows that NASH can occur with undisturbed glucose homeostasis and atheroprotection.


Subject(s)
Atherosclerosis , Lipoproteins, VLDL , Non-alcoholic Fatty Liver Disease , Small Leucine-Rich Proteoglycans , Animals , Female , Humans , Male , Mice , Apolipoproteins B/blood , Atherosclerosis/blood , Atherosclerosis/genetics , Atherosclerosis/metabolism , Atherosclerosis/prevention & control , Leucine , Lipoproteins, VLDL/biosynthesis , Lipoproteins, VLDL/blood , Lipoproteins, VLDL/metabolism , Liver/metabolism , Non-alcoholic Fatty Liver Disease/blood , Non-alcoholic Fatty Liver Disease/genetics , Non-alcoholic Fatty Liver Disease/metabolism , Small Leucine-Rich Proteoglycans/genetics , Small Leucine-Rich Proteoglycans/metabolism , Triglycerides/blood
7.
Lipids Health Dis ; 23(1): 197, 2024 Jun 26.
Article in English | MEDLINE | ID: mdl-38926753

ABSTRACT

BACKGROUND: Lipids, including phospholipids and bile acids, exert various signaling effects and are thought to contribute to the development of coronary artery disease (CAD). Here, we aimed to compare lipidomic and bile acid profiles in the blood of patients with and without CAD stratified by sex. METHODS: From 2015 to 2022, 3,012 patients who underwent coronary angiography were recruited in the INTERCATH cohort. From the overall cohort, subgroups were defined using patient characteristics such as CAD vs. no CAD, 1st vs. 3rd tertile of LDL-c, and female vs. male sex. Hereafter, a matching algorithm based on age, BMI, hypertension status, diabetes mellitus status, smoking status, the Mediterranean diet score, and the intake of statins, triglycerides, HDL-c and hs-CRP in a 1:1 ratio was implemented. Lipidomic analyses of stored blood samples using the Lipidyzer platform (SCIEX) and bile acid analysis using liquid chromatography with tandem mass spectrometry (LC‒MS/MS) were carried out. RESULTS: A total of 177 matched individuals were analyzed; the median ages were 73.5 years (25th and 75th percentile: 64.1, 78.2) and 71.9 years (65.7, 77.2) for females and males with CAD, respectively, and 67.6 years (58.3, 75.3) and 69.2 years (59.8, 76.8) for females and males without CAD, respectively. Further baseline characteristics, including cardiovascular risk factors, were balanced between the groups. Women with CAD had decreased levels of phosphatidylcholine and diacylglycerol, while no differences in bile acid profiles were detected in comparison to those of female patients without CAD. In contrast, in male patients with CAD, decreased concentrations of the secondary bile acid species glycolithocholic and lithocholic acid, as well as altered levels of specific lipids, were detected compared to those in males without CAD. Notably, male patients with low LDL-c and CAD had significantly greater concentrations of various phospholipid species, particularly plasmalogens, compared to those in high LDL-c subgroup. CONCLUSIONS: We present hypothesis-generating data on sex-specific lipidomic patterns and bile acid profiles in CAD patients. The data suggest that altered lipid and bile acid composition might contribute to CAD development and/or progression, helping to understand the different disease trajectories of CAD in women and men. REGISTRATION: https://clinicaltrials.gov/ct2/show/NCT04936438 , Unique identifier: NCT04936438.


Subject(s)
Bile Acids and Salts , Coronary Artery Disease , Lipidomics , Aged , Female , Humans , Male , Middle Aged , Bile Acids and Salts/blood , Cholesterol, HDL/blood , Cholesterol, LDL/blood , Coronary Artery Disease/blood , Sex Characteristics , Sex Factors , Tandem Mass Spectrometry , Triglycerides/blood , Cohort Studies
8.
Neoplasma ; 71(2): 164-179, 2024 Apr.
Article in English | MEDLINE | ID: mdl-38766857

ABSTRACT

Obesity is a major public health concern because it increases the risk of several diseases, including cancer. Crosstalk between obesity and cancer seems to be very complex, and the interaction between adipocytes and cancer cells leads to changes in adipocytes' function and their paracrine signaling, promoting a microenvironment that supports tumor growth. Carbonic anhydrase IX (CA IX) is a tumor-associated enzyme that not only participates in pH regulation but also facilitates metabolic reprogramming and supports the migration, invasion, and metastasis of cancer cells. In addition, CA IX expression, predominantly regulated via hypoxia-inducible factor (HIF-1), serves as a surrogate marker of hypoxia. In this study, we investigated the impact of adipocytes and adipocyte-derived factors on the expression of CA IX in colon and breast cancer cells. We observed increased expression of CA9 mRNA as well as CA IX protein in the presence of adipocytes and adipocyte-derived conditioned medium. Moreover, we confirmed that adipocytes affect the hypoxia signaling pathway and that the increased CA IX expression results from adipocyte-mediated induction of HIF-1α. Furthermore, we demonstrated that adipocyte-mediated upregulation of CA IX leads to increased migration and decreased adhesion of colon cancer cells. Finally, we brought experimental evidence that adipocytes, and more specifically leptin, upregulate CA IX expression in cancer cells and consequently promote tumor progression.


Subject(s)
Adipocytes , Antigens, Neoplasm , Breast Neoplasms , Carbonic Anhydrase IX , Colonic Neoplasms , Paracrine Communication , Animals , Female , Humans , Mice , Adipocytes/metabolism , Antigens, Neoplasm/metabolism , Breast Neoplasms/pathology , Breast Neoplasms/metabolism , Carbonic Anhydrase IX/metabolism , Cell Line, Tumor , Cell Movement , Colonic Neoplasms/pathology , Colonic Neoplasms/metabolism , Culture Media, Conditioned/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Leptin/metabolism , Obesity/metabolism , Tumor Microenvironment
9.
Am J Physiol Endocrinol Metab ; 322(2): E85-E100, 2022 02 01.
Article in English | MEDLINE | ID: mdl-34927460

ABSTRACT

Activation of uncoupling protein 1 (UCP1) in brown adipose tissue (BAT) upon cold stimulation leads to substantial increase in energy expenditure to defend body temperature. Increases in energy expenditure after a high-caloric food intake, termed diet-induced thermogenesis, are also attributed to BAT. These properties render BAT a potential target to combat diet-induced obesity. However, studies investigating the role of UCP1 to protect against diet-induced obesity are controversial and rely on the phenotyping of a single constitutive UCP1-knockout model. To address this issue, we generated a novel UCP1-knockout model by Cre-mediated deletion of exon 2 in the UCP1 gene. We studied the effect of constitutive UCP1 knockout on metabolism and the development of diet-induced obesity. UCP1 knockout and wild-type mice were housed at 30°C and fed a control diet for 4 wk followed by 8 wk of high-fat diet. Body weight and food intake were monitored continuously over the course of the study, and indirect calorimetry was used to determine energy expenditure during both feeding periods. Based on Western blot analysis, thermal imaging and noradrenaline test, we confirmed the lack of functional UCP1 in knockout mice. However, body weight gain, food intake, and energy expenditure were not affected by loss of UCP1 function during both feeding periods. We introduce a novel UCP1-KO mouse enabling the generation of conditional UCP1-knockout mice to scrutinize the contribution of UCP1 to energy metabolism in different cell types or life stages. Our results demonstrate that UCP1 does not protect against diet-induced obesity at thermoneutrality.NEW & NOTEWORTHY We provide evidence that the abundance of UCP1 does not influence energy metabolism at thermoneutrality studying a novel Cre-mediated UCP1-KO mouse model. This model will be a foundation for a better understanding of the contribution of UCP1 in different cell types or life stages to energy metabolism.


Subject(s)
Diet, High-Fat/adverse effects , Obesity/etiology , Obesity/metabolism , Temperature , Uncoupling Protein 1/metabolism , Adipose Tissue, Brown/metabolism , Animals , Calorimetry, Indirect/methods , Disease Susceptibility/metabolism , Eating/genetics , Energy Metabolism/genetics , Female , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Obesity/genetics , Thermogenesis/genetics , Uncoupling Protein 1/genetics , Weight Gain/genetics
10.
Handb Exp Pharmacol ; 270: 289-310, 2022.
Article in English | MEDLINE | ID: mdl-33373032

ABSTRACT

Adipose tissue as a major organ of lipid and lipoprotein metabolism has a major impact on metabolic homeostasis and thus influences the development of atherosclerosis and related cardiometabolic diseases. Unhealthy adipose tissue, which is often associated with obesity and systemic insulin resistance, promotes the development of diabetic dyslipidemia and can negatively affect vascular tissue homeostasis by secreting pro-inflammatory peptides and lipids. Conversely, paracrine and endocrine factors that are released from healthy adipose tissue can preserve metabolic balance and a functional vasculature. In this chapter, we describe adipose tissue types relevant for atherosclerosis and address the question how lipid metabolism as well as regulatory molecules produced in these fat depots can be targeted to counteract atherogenic processes in the vessel wall and improve plasma lipids. We discuss the role of adipose tissues in the action of approved drugs with anti-atherogenic activity. In addition, we present potential novel targets and therapeutic approaches aimed at increasing lipoprotein disposal in adipose tissue, boosting the activity of heat-producing (thermogenic) adipocytes, reducing adipose tissue inflammation, and improving or replacing beneficial hormones released from adipose tissues. Furthermore, we describe the future potential of innovative drug delivery technologies.


Subject(s)
Atherosclerosis , Insulin Resistance , Adipokines , Adipose Tissue , Atherosclerosis/drug therapy , Atherosclerosis/prevention & control , Humans , Thermogenesis
11.
Immunology ; 162(4): 452-463, 2021 04.
Article in English | MEDLINE | ID: mdl-33346377

ABSTRACT

Autoimmune diseases are caused by adaptive immune responses to self-antigens. The development of antigen-specific therapies that suppress disease-related, but not unrelated immune responses in general, is an important goal of biomedical research. We have previously shown that delivery of myelin peptides to liver sinusoidal endothelial cells (LSECs) using LSEC-targeting nanoparticles provides effective protection from CD4 T-cell-driven autoimmune encephalomyelitis. Here, we investigated whether this methodology might also serve antigen-specific treatment of a CD8 T-cell-driven autoimmune disease. As a model for CD8 T-cell-mediated autoimmunity, we used OT-1 T-cell-driven cholangitis in K14-OVAp mice expressing the cognate MHC I-restricted SIINFEKL peptide in cholangiocytes. To study whether peptide delivery to LSECs could modulate cholangitis, SIINFEKL peptide-conjugated nanoparticles were administered intravenously one day before transfer of OT-1 T cells; five days after cell transfer, liver pathology and hepatic infiltrates were analysed. SIINFEKL peptide-conjugated nanoparticles were rapidly taken up by LSECs in vivo, which effectively cross-presented the delivered peptide on MHC I molecules. Intriguingly, K14-OVAp mice receiving SIINFEKL-loaded nanoparticles manifested significantly reduced liver damage compared with vehicle-treated K14-OVAp mice. Mechanistically, treatment with LSEC-targeting SIINFEKL-loaded nanoparticles significantly reduced the number of liver-infiltrating OT-1 T cells, which up-regulated expression of the co-inhibitory receptor PD-1 and down-regulated cytotoxic effector function and inflammatory cytokine production. These findings show that tolerogenic LSECs can effectively internalize circulating nanoparticles and cross-present nanoparticle-bound peptides on MHC I molecules. Therefore, nanoparticle-mediated autoantigen peptide delivery to LSECs might serve the antigen-specific treatment of CD8 T-cell-driven autoimmune disease.


Subject(s)
Autoantigens/administration & dosage , Autoimmune Diseases/immunology , CD8-Positive T-Lymphocytes/immunology , Cholangitis/immunology , Endothelial Cells/immunology , Immunotherapy/methods , Liver/pathology , Magnetic Iron Oxide Nanoparticles/administration & dosage , Ovalbumin/administration & dosage , T-Lymphocytes, Regulatory/immunology , Animals , Autoantigens/chemistry , Autoimmune Diseases/therapy , Cells, Cultured , Cholangitis/therapy , Cross-Priming , Cytotoxicity, Immunologic , Disease Models, Animal , Humans , Immunosuppression Therapy , Liver/blood supply , Magnetic Iron Oxide Nanoparticles/chemistry , Mice , Mice, Transgenic , Ovalbumin/chemistry , Peptide Fragments/administration & dosage , Peptide Fragments/chemistry , Programmed Cell Death 1 Receptor/metabolism
12.
Cell Mol Life Sci ; 77(14): 2839-2857, 2020 Jul.
Article in English | MEDLINE | ID: mdl-31664461

ABSTRACT

Cholesterol accumulation in late endosomes is a prevailing phenotype of Niemann-Pick type C1 (NPC1) mutant cells. Likewise, annexin A6 (AnxA6) overexpression induces a phenotype reminiscent of NPC1 mutant cells. Here, we demonstrate that this cellular cholesterol imbalance is due to AnxA6 promoting Rab7 inactivation via TBC1D15, a Rab7-GAP. In NPC1 mutant cells, AnxA6 depletion and eventual Rab7 activation was associated with peripheral distribution and increased mobility of late endosomes. This was accompanied by an enhanced lipid accumulation in lipid droplets in an acyl-CoA:cholesterol acyltransferase (ACAT)-dependent manner. Moreover, in AnxA6-deficient NPC1 mutant cells, Rab7-mediated rescue of late endosome-cholesterol export required the StAR-related lipid transfer domain-3 (StARD3) protein. Electron microscopy revealed a significant increase of membrane contact sites (MCS) between late endosomes and ER in NPC1 mutant cells lacking AnxA6, suggesting late endosome-cholesterol transfer to the ER via Rab7 and StARD3-dependent MCS formation. This study identifies AnxA6 as a novel gatekeeper that controls cellular distribution of late endosome-cholesterol via regulation of a Rab7-GAP and MCS formation.


Subject(s)
Annexin A6/genetics , Cholesterol/genetics , GTPase-Activating Proteins/genetics , Niemann-Pick Disease, Type C/genetics , rab GTP-Binding Proteins/genetics , Animals , CHO Cells , Carrier Proteins/genetics , Cell Membrane/genetics , Cell Membrane/metabolism , Cholesterol/metabolism , Cricetulus , Endoplasmic Reticulum/genetics , Endoplasmic Reticulum/metabolism , Endosomes/genetics , Endosomes/metabolism , Humans , Membrane Proteins/genetics , Niemann-Pick Disease, Type C/metabolism , Niemann-Pick Disease, Type C/pathology , Protein Domains/genetics , Protein Transport/genetics , RNA, Small Interfering/genetics , rab7 GTP-Binding Proteins
13.
Gut ; 69(4): 665-672, 2020 04.
Article in English | MEDLINE | ID: mdl-31243055

ABSTRACT

BACKGROUND: Patients with primary sclerosing cholangitis (PSC) display an altered colonic microbiome compared with healthy controls. However, little is known on the bile duct microbiome and its interplay with bile acid metabolism in PSC. METHODS: Patients with PSC (n=43) and controls without sclerosing cholangitis (n=22) requiring endoscopic retrograde cholangiography were included prospectively. Leading indications in controls were sporadic choledocholithiasis and papillary adenoma. A total of 260 biospecimens were collected from the oral cavity, duodenal fluid and mucosa and ductal bile. Microbiomes of the upper alimentary tract and ductal bile were profiled by sequencing the 16S-rRNA-encoding gene (V1-V2). Bile fluid bile acid composition was measured by high-performance liquid chromatography mass spectrometry and validated in an external cohort (n=20). RESULTS: The bile fluid harboured a diverse microbiome that was distinct from the oral cavity, the duodenal fluid and duodenal mucosa communities. The upper alimentary tract microbiome differed between PSC patients and controls. However, the strongest differences between PSC patients and controls were observed in the ductal bile fluid, including reduced biodiversity (Shannon entropy, p=0.0127) and increase of pathogen Enterococcus faecalis (FDR=4.18×10-5) in PSC. Enterococcus abundance in ductal bile was strongly correlated with concentration of the noxious secondary bile acid taurolithocholic acid (r=0.60, p=0.0021). CONCLUSION: PSC is characterised by an altered microbiome of the upper alimentary tract and bile ducts. Biliary dysbiosis is linked with increased concentrations of the proinflammatory and potentially cancerogenic agent taurolithocholic acid.


Subject(s)
Bile/microbiology , Cholangitis, Sclerosing/microbiology , Dysbiosis/complications , Microbiota , Adult , Aged , Aged, 80 and over , Bile Ducts/microbiology , Case-Control Studies , Cohort Studies , Duodenum/microbiology , Dysbiosis/pathology , Female , Humans , Male , Middle Aged , Mouth Mucosa/microbiology , Young Adult
14.
J Lipid Res ; 61(11): 1377-1389, 2020 11.
Article in English | MEDLINE | ID: mdl-32769145

ABSTRACT

Adaptive thermogenesis is highly dependent on uncoupling protein 1 (UCP1), a protein expressed by thermogenic adipocytes present in brown adipose tissue (BAT) and white adipose tissue (WAT). Thermogenic capacity of human and mouse BAT can be measured by positron emission tomography-computed tomography quantifying the uptake of 18F-fluodeoxyglucose or lipid tracers. BAT activation is typically studied in response to cold exposure or treatment with ß-3-adrenergic receptor agonists such as CL316,243 (CL). Currently, it is unknown whether cold-stimulated uptake of glucose or lipid tracers is a good surrogate marker of UCP1-mediated thermogenesis. In metabolic studies using radiolabeled tracers, we found that glucose uptake is increased in mildly cold-activated BAT of Ucp1-/- versus WT mice kept at subthermoneutral temperature. Conversely, lower glucose disposal was detected after full thermogenic activation achieved by sustained cold exposure or CL treatment. In contrast, uptake of lipoprotein-derived fatty acids into chronically activated thermogenic adipose tissues was substantially increased in UCP1-deficient mice. This effect is linked to higher sympathetic tone in adipose tissues of Ucp1-/- mice, as indicated by elevated levels of thermogenic genes in BAT and WAT. Thus, glucose and lipoprotein handling does not necessarily reflect UCP1-dependent thermogenic activity, but especially lipid uptake rather mirrors sympathetic activation of adipose tissues.


Subject(s)
Adipose Tissue, Brown/chemistry , Glucose/metabolism , Lipoproteins/metabolism , Thermogenesis , Uncoupling Protein 1/metabolism , Adipose Tissue, Brown/metabolism , Animals , Lipoproteins/chemistry , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Uncoupling Protein 1/deficiency
15.
Purinergic Signal ; 16(4): 529-542, 2020 12.
Article in English | MEDLINE | ID: mdl-33025427

ABSTRACT

Several studies suggest a role of extracellular adenine nucleotides in regulating adipose tissue functions via the purinergic signaling network. Metabolic studies in mice with global deletion of the purinergic receptor P2X7 on the C57BL/6 background indicate that this receptor has only a minor role in adipose tissue for diet-induced inflammation or cold-triggered thermogenesis. However, recent data show that a polymorphism (P451L) present in C57BL/6 mice attenuates P2X7 receptor function, whereas BALB/c mice express the fully functional P451 allele. To determine the potential role of P2rx7 under metabolic and thermogenic stress conditions, we performed comparative studies using male P2rx7 knockout (KO) and respective wild-type controls on both BALB/c and C57BL/6 backgrounds. Our data show that adipose P2rx7 mRNA levels are increased in obese mice. Moreover, P2rx7 deficiency results in reduced levels of circulating CCL2 and IL6 with a moderate effect on gene expression of pro-inflammatory markers in white adipose tissue and liver of BALB/c and C57BL/6 mice. However, P2X7 expression does not alter body weight, insulin resistance, and hyperglycemia associated with high-fat diet feeding on both genetic backgrounds. Furthermore, deficiency of P2rx7 is dispensable for energy expenditure at thermoneutral and acute cold exposure conditions. In summary, these data show that-apart from a moderate effect on inflammatory cytokines-P2X7 plays only a minor role in inflammatory and thermogenic effects of white and brown adipose tissue even on the BALB/c background.


Subject(s)
Adipose Tissue, Brown/metabolism , Adipose Tissue, White/metabolism , Energy Metabolism/physiology , Homeostasis/physiology , Receptors, Purinergic P2X7/metabolism , Animals , Male , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout , Obesity/metabolism
16.
Nature ; 509(7502): 641-4, 2014 May 29.
Article in English | MEDLINE | ID: mdl-24747400

ABSTRACT

Despite the introduction of antiproliferative drug-eluting stents, coronary heart disease remains the leading cause of death in the United States. In-stent restenosis and bypass graft failure are characterized by excessive smooth muscle cell (SMC) proliferation and concomitant myointima formation with luminal obliteration. Here we show that during the development of myointimal hyperplasia in human arteries, SMCs show hyperpolarization of their mitochondrial membrane potential (ΔΨm) and acquire a temporary state with a high proliferative rate and resistance to apoptosis. Pyruvate dehydrogenase kinase isoform 2 (PDK2) was identified as a key regulatory protein, and its activation proved necessary for relevant myointima formation. Pharmacologic PDK2 blockade with dichloroacetate or lentiviral PDK2 knockdown prevented ΔΨm hyperpolarization, facilitated apoptosis and reduced myointima formation in injured human mammary and coronary arteries, rat aortas, rabbit iliac arteries and swine (pig) coronary arteries. In contrast to several commonly used antiproliferative drugs, dichloroacetate did not prevent vessel re-endothelialization. Targeting myointimal ΔΨm and alleviating apoptosis resistance is a novel strategy for the prevention of proliferative vascular diseases.


Subject(s)
Aorta/injuries , Arteries/injuries , Constriction, Pathologic/prevention & control , Dichloroacetic Acid/pharmacology , Dichloroacetic Acid/therapeutic use , Tunica Intima/drug effects , Tunica Intima/pathology , Angioplasty, Balloon/adverse effects , Animals , Aorta/drug effects , Aorta/pathology , Apoptosis/drug effects , Arteries/drug effects , Arteries/pathology , Cell Proliferation/drug effects , Constriction, Pathologic/pathology , Coronary Vessels/drug effects , Coronary Vessels/injuries , Coronary Vessels/pathology , Disease Models, Animal , Enzyme Activation/drug effects , Gene Knockdown Techniques , Humans , Hyperplasia/drug therapy , Hyperplasia/pathology , Iliac Artery/drug effects , Iliac Artery/injuries , Iliac Artery/pathology , Mammary Arteries/drug effects , Mammary Arteries/injuries , Mammary Arteries/pathology , Membrane Potential, Mitochondrial/drug effects , Mitochondria, Heart/drug effects , Mitochondria, Heart/metabolism , Myocytes, Smooth Muscle/drug effects , Myocytes, Smooth Muscle/pathology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein Serine-Threonine Kinases/deficiency , Protein Serine-Threonine Kinases/genetics , Pyruvate Dehydrogenase Acetyl-Transferring Kinase , Rabbits , Rats , Secondary Prevention , Stents/adverse effects , Swine , Tunica Intima/injuries
17.
Alzheimers Dement ; 16(9): 1248-1258, 2020 09.
Article in English | MEDLINE | ID: mdl-32588544

ABSTRACT

INTRODUCTION: Apolipoprotein E (apoE) is a carrier for brain lipids and the most important genetic risk factor for Alzheimer's disease (AD). ApoE binds the receptor sortilin, which mediates uptake of apoE-bound cargo into neurons. The significance of this uptake route for brain lipid homeostasis and AD risk seen with apoE4, but not apoE3, remains unresolved. METHODS: Combining neurolipidomics in patient specimens with functional studies in mouse models, we interrogated apoE isoform-specific functions for sortilin in brain lipid metabolism and AD. RESULTS: Sortilin directs the uptake and conversion of polyunsaturated fatty acids into endocannabinoids, lipid-based neurotransmitters that act through nuclear receptors to sustain neuroprotective gene expression in the brain. This sortilin function requires apoE3, but is disrupted by binding of apoE4, compromising neuronal endocannabinoid metabolism and action. DISCUSSION: We uncovered the significance of neuronal apoE receptor sortilin in facilitating neuroprotective actions of brain lipids, and its relevance for AD risk seen with apoE4.


Subject(s)
Adaptor Proteins, Vesicular Transport/metabolism , Apolipoprotein E4 , Endocannabinoids/metabolism , Lipid Metabolism , Neurons/metabolism , Neuroprotection , Adaptor Proteins, Vesicular Transport/genetics , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Animals , Apolipoprotein E3/genetics , Apolipoprotein E3/metabolism , Apolipoprotein E4/genetics , Apolipoprotein E4/metabolism , Biological Transport , Brain/metabolism , Humans , Mice , Signal Transduction
18.
Am J Physiol Endocrinol Metab ; 316(3): E487-E503, 2019 03 01.
Article in English | MEDLINE | ID: mdl-30576247

ABSTRACT

The possibility that recruitment and activation of brown adipose tissue (BAT) thermogenesis could be beneficial for curtailing obesity development in humans prompts a need for a better understanding of the control of these processes [that are often referred to collectively as diet-induced thermogenesis (DIT)]. Dietary conditions are associated with large changes in blood-borne factors that could be responsible for BAT recruitment, but BAT is also innervated by the sympathetic nervous system. To examine the significance of the innervation for DIT recruitment, we surgically denervated the largest BAT depot, i.e., the interscapular BAT depot in mice and exposed the mice at thermoneutrality to a high-fat diet versus a chow diet. Denervation led to an alteration in feeding pattern but did not lead to enhanced obesity, but obesity was achieved with a lower food intake, as denervation increased metabolic efficiency. Conclusively, denervation totally abolished the diet-induced increase in total UCP1 protein levels observed in the intact mice, whereas basal UCP1 expression was not dependent on innervation. The denervation of interscapular BAT did not discernably hyper-recruit other BAT depots, and no UCP1 protein could be detected in the principally browning-competent inguinal white adipose tissue depot under any of the examined conditions. We conclude that intact innervation is essential for diet-induced thermogenesis and that circulating factors cannot by themselves initiate recruitment of brown adipose tissue under obesogenic conditions. Therefore, the processes that link food intake and energy storage to activation of the nervous system are those of significance for the further understanding of diet-induced thermogenesis.


Subject(s)
Adipose Tissue, Brown/innervation , Obesity/metabolism , Sympathectomy , Thermogenesis/physiology , Uncoupling Protein 1/metabolism , Adipose Tissue, Brown/metabolism , Adipose Tissue, White/metabolism , Animals , Calorimetry, Indirect , Diet , Diet, High-Fat , Energy Intake , Male , Mice
19.
J Hepatol ; 71(4): 783-792, 2019 10.
Article in English | MEDLINE | ID: mdl-31207266

ABSTRACT

BACKGROUND & AIMS: T cells are central mediators of liver inflammation and represent potential treatment targets in cholestatic liver disease. Whereas emerging evidence shows that bile acids (BAs) affect T cell function, the role of T cells for the regulation of BA metabolism is unknown. In order to understand this interplay, we investigated the influence of T cells on BA metabolism in a novel mouse model of cholangitis. METHODS: Mdr2-/- mice were crossed with transgenic K14-OVAp mice, which express an MHC class I restricted ovalbumin peptide on biliary epithelial cells (Mdr2-/-xK14-OVAp). T cell-mediated cholangitis was induced by the adoptive transfer of antigen-specific CD8+ T cells. BA levels were quantified using a targeted liquid chromatography-mass spectrometry-based approach. RESULTS: T cell-induced cholangitis resulted in reduced levels of unconjugated BAs in the liver and significantly increased serum and hepatic levels of conjugated BAs. Genes responsible for BA synthesis and uptake were downregulated and expression of the bile salt export pump was increased. The transferred antigen-specific CD8+ T cells alone were able to induce these changes, as demonstrated using Mdr2-/-xK14-OVAp recipient mice on the Rag1-/- background. Mechanistically, we showed by depletion experiments that alterations in BA metabolism were partly mediated by the proinflammatory cytokines TNF and IFN-γ in an FXR-dependent manner, a process that in vitro required cell contact between T cells and hepatocytes. CONCLUSION: Whereas it is known that BA metabolism is dysregulated in sepsis and related conditions, we have shown that T cells are able to control the synthesis and metabolism of BAs, a process which depends on TNF and IFN-γ. Understanding the effect of lymphocytes on BA metabolism will help in the design of combined treatment strategies for cholestatic liver diseases. LAY SUMMARY: Dysregulation of bile acid metabolism and T cells can contribute to the development of cholangiopathies. Before targeting T cells for the treatment of cholangiopathies, it should be determined whether they exert protective effects on bile acid metabolism. Herein, we demonstrate that T cell-induced cholangitis resulted in decreased levels of harmful unconjugated bile acids. T cells were able to directly control synthesis and metabolism of bile acids, a process which was dependent on the proinflammatory cytokines TNF and IFN-γ. Understanding the effect of lymphocytes on bile acid metabolism will help in the design of combined treatment strategies for cholestatic liver diseases.


Subject(s)
Bile Acids and Salts , Cholangitis , Interferon-gamma/immunology , T-Lymphocytes , Tumor Necrosis Factor-alpha/immunology , ATP Binding Cassette Transporter, Subfamily B/genetics , Animals , Bile Acids and Salts/biosynthesis , Bile Acids and Salts/metabolism , Biosynthetic Pathways/immunology , Cholangitis/immunology , Cholangitis/metabolism , Cholangitis/pathology , Mice , Mice, Knockout , Mice, Transgenic , Models, Animal , Serpins/genetics , T-Lymphocytes/metabolism , T-Lymphocytes/pathology , ATP-Binding Cassette Sub-Family B Member 4
20.
Arterioscler Thromb Vasc Biol ; 38(7): 1440-1453, 2018 07.
Article in English | MEDLINE | ID: mdl-29853565

ABSTRACT

OBJECTIVE: Studies into the role of LRP1 (low-density lipoprotein receptor-related protein 1) in human lipid metabolism are scarce. Although it is known that a common variant in LRP1 (rs116133520) is significantly associated with HDL-C (high-density lipoprotein cholesterol), the mechanism underlying this observation is unclear. In this study, we set out to study the functional effects of 2 rare LRP1 variants identified in subjects with extremely low HDL-C levels. APPROACH AND RESULTS: In 2 subjects with HDL-C below the first percentile for age and sex and moderately elevated triglycerides, we identified 2 rare variants in LRP1: p.Val3244Ile and p.Glu3983Asp. Both variants decrease LRP1 expression and stability. We show in a series of translational experiments that these variants culminate in reduced trafficking of ABCA1 (ATP-binding cassette A1) to the cell membrane. This is accompanied by an increase in cell surface expression of SR-B1 (scavenger receptor class B type 1). Combined these effects may contribute to low HDL-C levels in our study subjects. Supporting these findings, we provide epidemiological evidence that rs116133520 is associated with apo (apolipoprotein) A1 but not with apoB levels. CONCLUSIONS: This study provides the first evidence that rare variants in LRP1 are associated with changes in human lipid metabolism. Specifically, this study shows that LRP1 may affect HDL metabolism by virtue of its effect on both ABCA1 and SR-B1.


Subject(s)
ATP Binding Cassette Transporter 1/metabolism , Cholesterol, HDL/metabolism , Fibroblasts/metabolism , Genetic Variation , Hypoalphalipoproteinemias/blood , Hypoalphalipoproteinemias/genetics , Low Density Lipoprotein Receptor-Related Protein-1/genetics , Scavenger Receptors, Class B/metabolism , Apolipoprotein A-I/blood , Cell Line, Tumor , Cell Membrane/metabolism , Genetic Predisposition to Disease , HEK293 Cells , Humans , Hypoalphalipoproteinemias/diagnosis , Liver/metabolism , Low Density Lipoprotein Receptor-Related Protein-1/metabolism , Phenotype , Prospective Studies , Protein Stability , Protein Transport , Triglycerides/blood
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