Emerging technologies allow the high-throughput profiling of metabolic status from a blood specimen (metabolomics). We investigated whether metabolite profiles could predict the development of diabetes. Among 2,422 normoglycemic individuals followed for 12 years, 201 developed diabetes. Amino acids, amines, and other polar metabolites were profiled in baseline specimens using liquid chromatography-tandem mass spectrometry. Cases and controls were matched for age, body mass index and fasting glucose. Five branched-chain and aromatic amino acids had highly-significant associations with future diabetes: isoleucine, leucine, valine, tyrosine, and phenylalanine. A combination of three amino acids predicted future diabetes (>5-fold higher risk for individuals in top quartile). The results were replicated in an independent, prospective cohort. These findings underscore the potential importance of amino acid metabolism early in the pathogenesis of diabetes, and suggest that amino acid profiles could aid in diabetes risk assessment.

Circulating proteins are vital in human health and disease and are frequently used as biomarkers for clinical decision-making or as targets for pharmacological intervention. Here we map and replicate protein quantitative trait loci (pQTL) for 90 cardiovascular proteins in over 30,000 individuals, resulting in 451 pQTLs for 85 proteins. For each protein we further perform pathway mapping to obtain trans-pQTL gene and regulatory designations. We substantiate these regulatory findings with orthogonal evidence for trans-pQTLs using mouse knock-down experiments (ABCA1, TRIB1) and clinical trial results (CCR2, CCR5), with consistent regulation. Finally we evaluate known drug targets, and suggest new target candidates or repositioning opportunities using Mendelian randomization. This identifies 11 proteins with causal evidence of involvement in human disease that have not previously been targeted, including (gene symbols) EGF, IL16, PAPPA, SPON1, F3, ADM, CASP8, CHI3L1, CXCL16, GDF15, and MMP12. Taken together these findings demonstrate the utility of largescale mapping the genetics of the proteome, and provide a resource for future precision studies of circulating proteins in human health.

BackgroundType 2 diabetes is associated with obesity, ectopic lipid accumulation and low-grade inflammation. A dysfunctional gut microbiota has been suggested to participate in the pathogenesis of the disease. Green tea is rich in polyphenols and has previously been shown to exert beneficial metabolic effects. Lactobacillus plantarum has the ability to metabolize phenolic acids. The health promoting effect of whole green tea powder as a prebiotic compound has not been thoroughly investigated previously.MethodsC57BL/6J mice were fed a high-fat diet with or without a supplement of 4% green tea powder (GT), and offered drinking water supplemented with Lactobacillus plantarum DSM 15313 (Lp) or the combination of both (Lp + GT) for 22 weeks. Parameters related to obesity, glucose tolerance, lipid metabolism, hepatic steatosis and inflammation were examined. Small intestinal tissue and caecal content were collected for bacterial analysis.ResultsMice in the Lp + GT group had significantly more Lactobacillus and higher diversity of bacteria in the intestine compared to both mice in the control and the GT group. Green tea strongly reduced the body fat content and hepatic triacylglycerol and cholesterol accumulation. The reduction was negatively correlated to the amount of Akkermansia and/or the total amount of bacteria in the small intestine. Markers of inflammation were reduced in the Lp + GT group compared to control. PLS analysis of correlations between the microbiota and the metabolic variables of the individual mice showed that relatively few components of the microbiota had high impact on the correlation model.ConclusionsGreen tea powder in combination with a single strain of Lactobacillus plantarum was able to promote growth of Lactobacillus in the intestine and to attenuate high fat diet-induced inflammation. In addition, a component of the microbiota, Akkermansia, correlated negatively with several metabolic parameters known to be risk factors for the development of type 2 diabetes.

BackgroundDiabetes mellitus is linked to premature mortality of virtually all causes. Furin is a proprotein convertase broadly involved in the maintenance of cellular homeostasis; however, little is known about its role in the development of diabetes mellitus and risk of premature mortality.ObjectivesTo test if fasting plasma concentration of furin is associated with the development of diabetes mellitus and mortality.MethodsOvernight fasted plasma furin levels were measured at baseline examination in 4678 individuals from the population‐based prospective Malmö Diet and Cancer Study. We studied the relation of plasma furin levels with metabolic and hemodynamic traits. We used multivariable Cox proportional hazards models to investigate the association between baseline plasma furin levels and incidence of diabetes mellitus and mortality during 21.3–21.7 years follow‐up.ResultsAn association was observed between quartiles of furin concentration at baseline and body mass index, blood pressure and plasma concentration of glucose, insulin, LDL and HDL cholesterol (|0.11| ≤ β ≤ |0.31|, P < 0.001). Plasma furin (hazard ratio [HR] per one standard deviation increment of furin) was predictive of future diabetes mellitus (727 events; HR = 1.24, CI = 1.14–1.36, P < 0.001) after adjustment for age, sex, body mass index, systolic and diastolic blood pressure, use of antihypertensive treatment, alcohol intake and fasting plasma level of glucose, insulin and lipoproteins cholesterol. Furin was also independently related to the risk of all‐cause mortality (1229 events; HR = 1.12, CI = 1.05–1.19, P = 0.001) after full multivariable adjustment.ConclusionIndividuals with high plasma furin concentration have a pronounced dysmetabolic phenotype and elevated risk of diabetes mellitus and premature mortality.

We discovered associations between four gut microbiota genera (Blautia, Dorea, Ruminococcus, and SHA-98) and BMI-predictive plasma metabolites, including glutamate and BCAAs. Thus, these metabolites could be mediators between gut microbiota and obesity, pointing to potential future opportunities for targeting the gut microbiota in prevention of obesity.

AimsWe tested whether characteristic changes of the plasma lipidome in individuals with comparable total lipids level associate with future cardiovascular disease (CVD) outcome and whether 23 validated gene variants associated with coronary artery disease (CAD) affect CVD associated lipid species.Methods and ResultsScreening of the fasted plasma lipidome was performed by top-down shotgun analysis and lipidome compositions compared between incident CVD cases (n = 211) and controls (n = 216) from the prospective population-based MDC study using logistic regression adjusting for Framingham risk factors. Associations with incident CVD were seen for eight lipid species (0.21≤q≤0.23). Each standard deviation unit higher baseline levels of two lysophosphatidylcholine species (LPC), LPC16∶0 and LPC20∶4, was associated with a decreased risk for CVD (P = 0.024–0.028). Sphingomyelin (SM) 38∶2 was associated with increased odds of CVD (P = 0.057). Five triglyceride (TAG) species were associated with protection (P = 0.031–0.049). LPC16∶0 was negatively correlated with the carotid intima-media thickness (P = 0.010) and with HbA1c (P = 0.012) whereas SM38∶2 was positively correlated with LDL-cholesterol (P = 0.0*10−6) and the q-values were good (q≤0.03). The risk allele of 8 CAD-associated gene variants showed significant association with the plasma level of several lipid species. However, the q-values were high for many of the associations (0.015≤q≤0.75). Risk allele carriers of 3 CAD-loci had reduced level of LPC16∶0 and/or LPC 20∶4 (P≤0.056).ConclusionOur study suggests that CVD development is preceded by reduced levels of LPC16∶0, LPC20∶4 and some specific TAG species and by increased levels of SM38∶2. It also indicates that certain lipid species are intermediate phenotypes between genetic susceptibility and overt CVD. But it is a preliminary study that awaits replication in a larger population because statistical significance was lost for the associations between lipid species and future cardiovascular events when correcting for multiple testing.

Unbiased, "nontargeted" metabolite profiling techniques hold considerable promise for biomarker and pathway discovery, in spite of the lack of successful applications to human disease. By integrating nontargeted metabolomics, genetics, and detailed human phenotyping, we identified dimethylguanidino valeric acid (DMGV) as an independent biomarker of CTdefined nonalcoholic fatty liver disease (NAFLD) in the offspring cohort of the Framingham Heart Study (FHS) participants. We verified the relationship between DMGV and early hepatic pathology. Specifically, plasma DMGV levels were correlated with biopsy-proven nonalcoholic steatohepatitis (NASH) in a hospital cohort of individuals undergoing gastric bypass surgery, and DMGV levels fell in parallel with improvements in post-procedure cardiometabolic parameters. Further, baseline DMGV levels independently predicted future diabetes up to 12 years before disease onset in 3 distinct human cohorts. Finally, we provide all metabolite peak data consisting of known and unidentified peaks, genetics, and key metabolic parameters as a publicly available resource for investigations in cardiometabolic diseases.

The small GTPases Ras or Rap1 were suggested to mediate the stimulatory effect of some G protein-coupled receptors on ERK activity in neuronal cells. Accordingly, we reported here that pituitary adenylate cyclase-activating polypeptide (PACAP), whose G protein-coupled receptor triggers neuronal differentiation of the PC12 cell line via ERK1/2 activation, transiently activated Ras and induced the sustained GTP loading of Rap1. Ras mediated peak stimulation of ERK by PACAP, whereas Rap1 was necessary for the sustained activation phase. However, PACAP-induced GTP-loading of Rap1 was not sufficient to account for ERK activation by PACAP because 1) PACAP-elicited Rap1 GTP-loading depended only on phospholipase C, whereas maximal stimulation of ERK by PACAP also required the activity of protein kinase A (PKA), protein kinase C (PKC), and calcium-dependent signaling; and 2) constitutively active mutants of Rap1, Rap1A-V12, and Rap1B-V12 only minimally stimulated the ERK pathway compared with Ras-V12. The effect of Rap1A-V12 was dramatically potentiated by the concurrent activation of PKC, the cAMP pathway, and Ras, and this potentiation was blocked by dominant-negative mutants of Ras and Raf. Thus, this set of data indicated that GPCR-elicited GTP loading of Rap1 was not sufficient to stimulate efficiently ERK in PC12 cells and required the permissive co-stimulation of PKA, PKC, or Ras.Since its original establishment by Greene and Tischler (1), the PC12 pheochromocytoma cell line is a widely used model of neuronal differentiation. The first isolated neurotrophin, nerve growth factor (NGF) 1 (2), was shown to induce a neuronal like phenotype characterized by neurite outgrowth (1). The effect of NGF is dependent on a long lasting activation of the Ras-Raf-MEK-ERK pathway (3). Activation of the cAMP pathway was also reported to induce PC12 differentiation (4,5). In line with the demonstrated role of ERK activation in NGF-induced PC12 differentiation, cAMP analogues, and forskolin, a direct activator of adenylate cyclase (AC) was shown to stimulate ERK activity in this cell line (5, 6). Similarly, mobilization of calcium and/or stimulation of the diacylglycerol (DAG) production results in ERK activation and eventually neurite outgrowth (7, 8).The mechanisms responsible for the control of the ERK pathway by receptor tyrosine kinases, cAMP analogues, and calcium/DAG were intensively investigated, and Ras-like small GTP-binding proteins emerged as key elements in this pathway. The products of the H-Ras gene was shown to stimulate the activity of the MEK kinase Raf-1 following activation of receptor tyrosine kinases (9). Cyclic AMP and calcium were also suggested to control the activity of Ras in some cell types (7, 10), resulting in ERK activation. More recently, the Ras superfamily member Rap1 and the protein kinase B-Raf were suggested to link PKA activation by cAMP analogues to MEK1 stimulation in neuronal cells (11). On the other hand, several mechanisms have been proposed for calcium-induced ERK activation, including ac...