Ureteropelvic junction obstruction with primary lymphoedema associated with CELSR1 variants.

BACKGROUND: Primary lymphoedema (PL) is a chronic, debilitating disease caused by developmental and functional defects of the lymphatic system. It is marked by an accumulation of interstitial fluid, fat and tissue fibrosis. There is no cure. More than 50 genes and genetic loci have been linked to PL. We sought to study systematically cell polarity signalling protein Cadherin Epidermal Growth Factor Laminin G Seven-pass G-type Receptor 1 (CELSR1) variants linked to PL. METHODS: We investigated 742 index patients from our PL cohort using exome sequencing. RESULTS: We identified nine variants predicted to cause CELSR1 loss of function. Four of them were tested for nonsense-mediated mRNA decay, but none was observed. Most of the truncated CELSR1 proteins would lack the transmembrane domain, if produced. The affected individuals had puberty/late-onset PL on lower extremities. The variants had a statistically significant difference in penetrance between female patients (87%) and male patients (20%). Eight variant carriers had a kidney anomaly, mostly in the form of ureteropelvic junction obstruction, which has not been associated with CELSR1 before. CELSR1 is located in the 22q13.3 deletion locus of the Phelan-McDermid syndrome. As variable renal defects are often seen in patients with the Phelan-McDermid syndrome, CELSR1 may be the long-sought gene for the renal defects. CONCLUSION: PL associated with a renal anomaly suggests a CELSR1-related cause.

Regenerating islet-derived protein 3α: A promising therapy for diabetes. Preliminary data in rodents and in humans.

The aim of our study was to test the hypothesis that administration of Regenerating islet-derived protein 3α (Reg3α), a protein described as having protective effects against oxidative stress and anti-inflammatory activity, could participate in the control of glucose homeostasis and potentially be a new target of interest in the treatment of type 2 diabetes. To that end the recombinant human Reg3α protein was administered for one month in insulin-resistant mice fed high fat diet. We performed glucose and insulin tolerance tests, assayed circulating chemokines in plasma and measured glucose uptake in insulin sensitive tissues. We evidenced an increase in insulin sensitivity during an oral glucose tolerance test in ALF-5755 treated mice vs controls and decreased the pro-inflammatory cytokine C-X-C Motif Chemokine Ligand 5 (CXCL5). We also demonstrated an increase in glucose uptake in skeletal muscle. Finally, correlation studies using human and mouse muscle biopsies showed negative correlation between intramuscular Reg3α mRNA expression (or its murine isoform Reg3γ) and insulin resistance. Thus, we have established the proof of concept that Reg3α could be a novel molecule of interest in the treatment of T2D by increasing insulin sensitivity via a skeletal muscle effect.

Human and preclinical studies of the host-gut microbiome co-metabolite hippurate as a marker and mediator of metabolic health.

OBJECTIVE: Gut microbial products are involved in regulation of host metabolism. In human and experimental studies, we explored the potential role of hippurate, a hepatic phase 2 conjugation product of microbial benzoate, as a marker and mediator of metabolic health. DESIGN: In 271 middle-aged non-diabetic Danish individuals, who were stratified on habitual dietary intake, we applied 1H-nuclear magnetic resonance (NMR) spectroscopy of urine samples and shotgun-sequencing-based metagenomics of the gut microbiome to explore links between the urine level of hippurate, measures of the gut microbiome, dietary fat and markers of metabolic health. In mechanistic experiments with chronic subcutaneous infusion of hippurate to high-fat-diet-fed obese mice, we tested for causality between hippurate and metabolic phenotypes. RESULTS: In the human study, we showed that urine hippurate positively associates with microbial gene richness and functional modules for microbial benzoate biosynthetic pathways, one of which is less prevalent in the Bacteroides 2 enterotype compared with Ruminococcaceae or Prevotella enterotypes. Through dietary stratification, we identify a subset of study participants consuming a diet rich in saturated fat in which urine hippurate concentration, independently of gene richness, accounts for links with metabolic health. In the high-fat-fed mice experiments, we demonstrate causality through chronic infusion of hippurate (20 nmol/day) resulting in improved glucose tolerance and enhanced insulin secretion. CONCLUSION: Our human and experimental studies show that a high urine hippurate concentration is a general marker of metabolic health, and in the context of obesity induced by high-fat diets, hippurate contributes to metabolic improvements, highlighting its potential as a mediator of metabolic health.

Dominant gut Prevotella copri in gastrectomised non-obese diabetic Goto-Kakizaki rats improves glucose homeostasis through enhanced FXR signalling.

AIMS/HYPOTHESIS: Drug and surgical-based therapies in type 2 diabetes are associated with altered gut microbiota architecture. Here we investigated the role of the gut microbiome in improved glucose homeostasis following bariatric surgery. METHODS: We carried out gut microbiome analyses in gastrectomised (by vertical sleeve gastrectomy [VSG]) rats of the Goto-Kakizaki (GK) non-obese model of spontaneously occurring type 2 diabetes, followed by physiological studies in the GK rat. RESULTS: VSG in the GK rat led to permanent improvement of glucose tolerance associated with minor changes in the gut microbiome, mostly characterised by significant enrichment of caecal Prevotella copri. Gut microbiota enrichment with P. copri in GK rats through permissive antibiotic treatment, inoculation of gut microbiota isolated from gastrectomised GK rats, and direct inoculation of P. copri, resulted in significant improvement of glucose tolerance, independent of changes in body weight. Plasma bile acids were increased in GK rats following inoculation with P. copri and P. copri-enriched microbiota from VSG-treated rats; the inoculated GK rats then showed increased liver glycogen and upregulated expression of Fxr (also known as Nr1h4), Srebf1c, Chrebp (also known as Mlxipl) and Il10 and downregulated expression of Cyp7a1. CONCLUSIONS: Our data underline the impact of intestinal P. copri on improved glucose homeostasis through enhanced bile acid metabolism and farnesoid X receptor (FXR) signalling, which may represent a promising opportunity for novel type 2 diabetes therapeutics.

The Natural Metabolite 4-Cresol Improves Glucose Homeostasis and Enhances ß-Cell Function.

Exposure to natural metabolites contributes to the risk of cardiometabolic diseases (CMDs). Through metabolome profiling, we identify the inverse correlation between serum concentrations of 4-cresol and type 2 diabetes. The chronic administration of non-toxic doses of 4-cresol in complementary preclinical models of CMD reduces adiposity, glucose intolerance, and liver triglycerides, enhances insulin secretion in vivo, stimulates islet density and size, and pancreatic ß-cell proliferation, and increases vascularization, suggesting activated islet enlargement. In vivo insulin sensitivity is not affected by 4-cresol. The incubation of mouse isolated islets with 4-cresol results in enhanced insulin secretion, insulin content, and ß-cell proliferation of a magnitude similar to that induced by GLP-1. In both CMD models and isolated islets, 4-cresol is associated with the downregulated expression of the kinase DYRK1A, which may mediate its biological effects. Our findings identify 4-cresol as an effective regulator of ß-cell function, which opens up perspectives for therapeutic applications in syndromes of insulin deficiency.

Conserved properties of genetic architecture of renal and fat transcriptomes in rat models of insulin resistance.

To define renal molecular mechanisms that are affected by permanent hyperglycaemia and might promote phenotypes relevant to diabetic nephropathy, we carried out linkage analysis of genome-wide gene transcription in the kidneys of F2 offspring from the Goto-Kakizaki (GK) rat model of type 2 diabetes and normoglycaemic Brown Norway (BN) rats. We mapped 2526 statistically significant expression quantitative trait loci (eQTLs) in the cross. More than 40% of eQTLs mapped in the close vicinity of the linked transcripts, underlying possible cis-regulatory mechanisms of gene expression. We identified eQTL hotspots on chromosomes 5 and 9 regulating the expression of 80-165 genes, sex or cross direction effects, and enriched metabolic and immunological processes by segregating GK alleles. Comparative analysis with adipose tissue eQTLs in the same cross showed that 496 eQTLs, in addition to the top enriched biological pathways, are conserved in the two tissues. Extensive similarities in eQTLs mapped in the GK rat and in the spontaneously hypertensive rat (SHR) suggest a common aetiology of disease phenotypes common to the two strains, including insulin resistance, which is a prominent pathophysiological feature in both GK rats and SHRs. Our data shed light on shared and tissue-specific molecular mechanisms that might underlie aetiological aspects of insulin resistance in the context of spontaneously occurring hyperglycaemia and hypertension.

Systems Genetics of Hepatic Metabolome Reveals Octopamine as a Target for Non-Alcoholic Fatty Liver Disease Treatment.

Non-alcoholic fatty liver disease (NAFLD) is often associated with obesity and type 2 diabetes. To disentangle etiological relationships between these conditions and identify genetically-determined metabolites involved in NAFLD processes, we mapped 1H nuclear magnetic resonance (NMR) metabolomic and disease-related phenotypes in a mouse F2 cross derived from strains showing resistance (BALB/c) and increased susceptibility (129S6) to these diseases. Quantitative trait locus (QTL) analysis based on single nucleotide polymorphism (SNP) genotypes identified diet responsive QTLs in F2 mice fed control or high fat diet (HFD). In HFD fed F2 mice we mapped on chromosome 18 a QTL regulating liver micro- and macrovesicular steatosis and inflammation, independently from glucose intolerance and adiposity, which was linked to chromosome 4. Linkage analysis of liver metabolomic profiling data identified a QTL for octopamine, which co-localised with the QTL for liver histopathology in the cross. Functional relationship between these two QTLs was validated in vivo in mice chronically treated with octopamine, which exhibited reduction in liver histopathology and metabolic benefits, underlining its role as a mechanistic biomarker of fatty liver with potential therapeutic applications.

Untargeted Mass Spectrometry Lipidomics identifies correlation between serum sphingomyelins and plasma cholesterol.

BACKGROUND: Lipoproteins are major players in the development and progression of atherosclerotic plaques leading to coronary stenosis and myocardial infarction. Epidemiological, genetic and experimental observations have implicated the association of sphingolipids and intermediates of sphingolipid synthesis in atherosclerosis. We aimed to investigate relationships between quantitative changes in serum sphingolipids, the regulation of the metabolism of lipoproteins (LDL, HDL), and endophenotypes of coronary artery disease (CAD). METHODS: We carried out untargeted liquid chromatography - mass spectrometry (UPLC-MS) lipidomics of serum samples of subjects belonging to a cross-sectional study and recruited on the basis of absence or presence of angiographically-defined CAD, and extensively characterized for clinical and biochemical phenotypes. RESULTS: Among the 2998 spectral features detected in the serum samples, 1328 metabolic features were significantly correlated with at least one of the clinical or biochemical phenotypes measured in the cohort. We found evidence of significant associations between 34 metabolite signals, corresponding to a set of sphingomyelins, and serum HDL cholesterol. Many of these metabolite associations were also observed with serum LDL and total cholesterol levels but not as much with serum triglycerides. CONCLUSION: Among patients with CAD, sphingolipids in the form of sphingomyelins are directly correlated with serum levels of lipoproteins and total cholesterol. Results from this study support the fundamental role of sphingolipids in modulating lipid serum levels, highlighting the importance to identify novel targets in the sphingolipid metabolic pathway for anti-atherogenic therapies.

Implication of gut microbiota metabolites in cardiovascular and metabolic diseases.

Evidence from the literature keeps highlighting the impact of mutualistic bacterial communities of the gut microbiota on human health. The gut microbita is a complex ecosystem of symbiotic bacteria which contributes to mammalian host biology by processing, otherwise, indigestible nutrients, supplying essential metabolites, and contributing to modulate its immune system. Advances in sequencing technologies have enabled structural analysis of the human gut microbiota and allowed detection of changes in gut bacterial composition in several common diseases, including cardiometabolic disorders. Biological signals sent by the gut microbiota to the host, including microbial metabolites and pro-inflammatory molecules, mediate microbiome-host genome cross-talk. This rapidly expanding line of research can identify disease-causing and disease-predictive microbial metabolite biomarkers, which can be translated into novel biodiagnostic tests, dietary supplements, and nutritional interventions for personalized therapeutic developments in common diseases. Here, we review results from the most significant studies dealing with the association of products from the gut microbial metabolism with cardiometabolic disorders. We underline the importance of these postbiotic biomarkers in the diagnosis and treatment of human disorders.

A clinical-scale BioArtificial Liver, developed for GMP, improved clinical parameters of liver function in porcine liver failure.

Liver failure, whether arising directly from acute liver failure or from decompensated chronic liver disease is an increasing problem worldwide and results in many deaths. In the UK only 10% of individuals requiring a liver transplant receive one. Thus the need for alternative treatments is paramount. A BioArtificial Liver machine could temporarily replace the functions of the liver, buying time for the patient's liver to repair and regenerate. We have designed, implemented and tested a clinical-scale BioArtificial Liver machine containing a biomass derived from a hepatoblastoma cell-line cultured as three dimensional organoids, using a fluidised bed bioreactor, together with single-use bioprocessing equipment, with complete control of nutrient provision with feedback BioXpert recipe processes, and yielding good phenotypic liver functions. The methodology has been designed to meet specifications for GMP production, required for manufacture of advanced therapy medicinal products (ATMPs). In a porcine model of severe liver failure, damage was assured in all animals by surgical ischaemia in pigs with human sized livers (1.2-1.6 kg liver weights). The BioArtificial liver (UCLBAL) improved important prognostic clinical liver-related parameters, eg, a significant improvement in coagulation, reduction in vasopressor requirements, improvement in blood pH and in parameters of intracranial pressure (ICP) and oxygenation.

Preculturing Islets With Adipose-Derived Mesenchymal Stromal Cells Is an Effective Strategy for Improving Transplantation Efficiency at the Clinically Preferred Intraportal Site.

We have recently shown that preculturing islets with kidney-derived mesenchymal stromal cells (MSCs) improves transplantation outcome in streptozotocin-diabetic mice implanted with a minimal mass of islets beneath the kidney capsule. In the present study, we have extended our previous observations to investigate whether preculturing islets with MSCs can also be used to enhance islet function at the clinically used intraportal site. We have used MSCs derived from adipose tissue, which are more readily accessible than alternative sources in human subjects and can be expanded to clinically efficacious numbers, to preculture islets throughout this study. The in vivo efficacy of grafts consisting of islets precultured alone or with MSCs was tested using a syngeneic streptozotocin-diabetic minimal islet mass model at the clinically relevant intraportal site. Blood glucose concentrations were monitored for 1 month. The vascularization of islets precultured alone or with MSCs was investigated both in vitro and in vivo, using immunohistochemistry. Islet insulin content was measured by radioimmunoassay. The effect of preculturing islets with MSCs on islet function in vitro was investigated using static incubation assays. There was no beneficial angiogenic influence of MSC preculture, as demonstrated by the comparable vascularization of islets precultured alone or with MSCs, both in vitro after 3 days and in vivo 1 month after islet transplantation. However, the in vitro insulin secretory capacity of MSC precultured islets was superior to that of islets precultured alone. In vivo, this was associated with improved glycemia at 7, 14, 21, and 28 days posttransplantation, in recipients of MSC precultured islets compared to islets precultured alone. The area of individual islets within the graft-bearing liver was significantly higher in recipients of MSC precultured islets compared to islets precultured alone. Our experimental studies suggest that preculturing islets with MSCs represents a favorable strategy for improving the efficiency of clinical islet transplantation.