Specific Dietary Preferences Are Linked to Differing Gut Microbial Metabolic Activity in Response to Dark Chocolate Intake

Systems biology approaches are providing novel insights into the role of nutrition for the management of health and disease. In the present study, we investigated if dietary preference for dark chocolate in healthy subjects may lead to different metabolic response to daily chocolate consumption. Using NMR- and MS-based metabolic profiling of blood plasma and urine, we monitored the metabolic response of 10 participants stratified as chocolate desiring and eating regularly dark chocolate (CD) and 10 participants stratified as chocolate indifferent and eating rarely dark chocolate (CI) to a daily consumption of 50 g of dark chocolate as part of a standardized diet over a one week period. We demonstrated that preference for chocolate leads to different metabolic response to chocolate consumption. Daily intake of dark chocolate significantly increased HDL cholesterol by 6% and decreased polyunsaturated acyl ether phospholipids. Dark chocolate intake could also induce an improvement in the metabolism of long chain fatty acid, as noted by a compositional change in plasma fatty acyl carnitines. Moreover, a relationship between regular long-term dietary exposure to a small amount of dark chocolate, gut microbiota, and phenolics was highlighted, providing novel insights into biological processes associated with cocoa bioactives

Metabolic Signatures of Extreme Longevity in Northern Italian Centenarians Reveal a Complex Remodeling of Lipids, Amino Acids, and Gut Microbiota Metabolism

<div><p>The aging phenotype in humans has been thoroughly studied but a detailed metabolic profiling capable of shading light on the underpinning biological processes of longevity is still missing. Here using a combined metabonomics approach compromising holistic ¹H-NMR profiling and targeted MS approaches, we report for the first time the metabolic phenotype of longevity in a well characterized human aging cohort compromising mostly female centenarians, elderly, and young individuals. With increasing age, targeted MS profiling of blood serum displayed a marked decrease in tryptophan concentration, while an unique alteration of specific glycerophospholipids and sphingolipids are seen in the longevity phenotype. We hypothesized that the overall lipidome changes specific to longevity putatively reflect centenarians' unique capacity to adapt/respond to the accumulating oxidative and chronic inflammatory conditions characteristic of their extreme aging phenotype. Our data in centenarians support promotion of cellular detoxification mechanisms through specific modulation of the arachidonic acid metabolic cascade as we underpinned increased concentration of 8,9-EpETrE, suggesting enhanced cytochrome P450 (CYP) enzyme activity. Such effective mechanism might result in the activation of an anti-oxidative response, as displayed by decreased circulating levels of 9-HODE and 9-oxoODE, markers of lipid peroxidation and oxidative products of linoleic acid. Lastly, we also revealed that the longevity process deeply affects the structure and composition of the human gut microbiota as shown by the increased extrection of phenylacetylglutamine (PAG) and p-cresol sulfate (PCS) in urine of centenarians. Together, our novel approach in this representative Italian longevity cohort support the hypothesis that a complex remodeling of lipid, amino acid metabolism, and of gut microbiota functionality are key regulatory processes marking exceptional longevity in humans.</p> </div

Metabolic signature of aging and longevity in serum as per LC/MS eicosanoids profiling.

<p>Reported is median value in ng/100 µl serum among the three age groups. Blue denotes negative/decreased concentration, orange denotes positive/increased correlation, black denotes no changes. All significantly regulated metabolites are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056564#pone.0056564.s008" target="_blank">Table S6</a>.</p

Differences in metabolic profiles as displayed by LC/MS-MS targeted approach between centenarian's offspring (46 subjects average age 68.4 yrs) and offspring of non long-lived parents (42 subjects average age 70.7 yrs).

<p>Bar plots indicating mean (µM) ±standard error. All significantly regulated metabolites and statistical changes are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056564#pone.0056564.s011" target="_blank">Table S9</a>. Significant differences were assessed by Mann-Whitney U test where *p<0.05., **p<0.01, ***p<0.001.</p

Demographic characteristics of the recruited age cohorts.

<p>Values are presented as mean ±SD with the range in parentheses.</p

Early Metabolic Adaptation in C57BL/6 Mice Resistant to High Fat Diet Induced Weight Gain Involves an Activation of Mitochondrial Oxidative Pathways

We investigated the short-term (7 days) and long-term (60 days) metabolic effect of high fat diet induced obesity (DIO) and weight gain in isogenic C57BL/6 mice and examined the specific metabolic differentiation between mice that were either strong-responders (SR), or non-responders (NR) to weight gain. Mice (<i>n</i> = 80) were fed a standard chow diet for 7 days prior to randomization into a high-fat (HF) (<i>n</i> = 56) or a low-fat (LF) (<i>n</i> = 24) diet group. The ¹H NMR urinary metabolic profiles of LF and HF mice were recorded 7 and 60 days after the diet switch. On the basis of the body weight gain (BWG) distribution of HF group, we identified NR mice (<i>n</i> = 10) and SR mice (<i>n</i> = 14) to DIO. Compared with LF, HF feeding increased urinary excretion of glycine conjugates of β-oxidation intermediate (hexanoylglycine), branched chain amino acid (BCAA) catabolism intermediates (isovalerylglycine, α-keto-β-methylvalerate and α-ketoisovalerate) and end-products of nicotinamide adenine dinucleotide (NAD) metabolism (N1-methyl-2-pyridone-5-carboxamide, N1-methyl-4-pyridone-3-carboxamide) suggesting up-regulation of mitochondrial oxidative pathways. In the HF group, NR mice excreted relatively more hexanoylglycine, isovalerylglycine, and fewer tricarboxylic acid (TCA) cycle intermediate (succinate) in comparison to SR mice. Thus, subtle regulation of ketogenic pathways in DIO may alleviate the saturation of the TCA cycle and mitochondrial oxidative metabolism

Markers of longevity as per ¹H-NMR urine profiling.

<p>Bar plots indicating mean (relative concentration) ±standard error. PAG = Phenylacetylglutamine, PCS = p-cresol-sulfate, 2HB = 2-hydroxybenzoate. All significantly regulated metabolites and statistical changes are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056564#pone.0056564.s014" target="_blank">Table S12</a>. Significant differences were assessed by Mann-Whitney U test where ***p<0.001.</p

Spearman correlation map between urine markers of longevity (PAG = phenylacetylglutamine, PCS = p-cresol sulfate, 3-HB = 3-hydroxybenzoate) and order/genus-like bacterial phylogroups.

<p>Blue denotes negative correlation, orange denotes positive correlation, and black denotes no correlation.</p

Comparison of the content and size of different standard file formats for the storage of genomic data.

<p>Comparison of the content and size of different standard file formats for the storage of genomic data.</p

The Use of Non-Variant Sites to Improve the Clinical Assessment of Whole-Genome Sequence Data

<div><p>Genetic testing, which is now a routine part of clinical practice and disease management protocols, is often based on the assessment of small panels of variants or genes. On the other hand, continuous improvements in the speed and per-base costs of sequencing have now made whole exome sequencing (WES) and whole genome sequencing (WGS) viable strategies for targeted or complete genetic analysis, respectively. Standard WGS/WES data analytical workflows generally rely on calling of sequence variants respect to the reference genome sequence. However, the reference genome sequence contains a large number of sites represented by rare alleles, by known pathogenic alleles and by alleles strongly associated to disease by GWAS. It’s thus critical, for clinical applications of WGS and WES, to interpret whether non-variant sites are homozygous for the reference allele or if the corresponding genotype cannot be reliably called. Here we show that an alternative analytical approach based on the analysis of both variant and non-variant sites from WGS data allows to genotype more than 92% of sites corresponding to known SNPs compared to 6% genotyped by standard variant analysis. These include homozygous reference sites of clinical interest, thus leading to a broad and comprehensive characterization of variation necessary to an accurate evaluation of disease risk. Altogether, our findings indicate that characterization of both variant and non-variant clinically informative sites in the genome is necessary to allow an accurate clinical assessment of a personal genome. Finally, we propose a highly efficient extended VCF (eVCF) file format which allows to store genotype calls for sites of clinical interest while remaining compatible with current variant interpretation software.</p></div