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Circulating cell-free DNA (cf-DNA) has emerged as a promising biomarker of ageing, tissue damage and cellular stress. However, less is known about health behaviours, ageing phenotypes and metabolic processes that lead to elevated cf-DNA levels. We sought to analyse the relationship of circulating cf-DNA level to age, sex, smoking, physical activity, vegetable consumption, ageing phenotypes (physical functioning, the number of diseases, frailty) and an extensive panel of biomarkers including blood and urine metabolites and inflammatory markers in three human cohorts (N = 5385; 17–82 years). The relationships were assessed using correlation statistics, and linear and penalised regressions (the Lasso), also stratified by sex.cf-DNA levels were significantly higher in men than in women, and especially in middle-aged men and women who smoke, and in older more frail individuals. Correlation statistics of biomarker data showed that cf-DNA level was higher with elevated inflammation (C-reactive protein, interleukin-6), and higher levels of homocysteine, and proportion of red blood cells and lower levels of ascorbic acid. Inflammation (C-reactive protein, glycoprotein acetylation), amino acids (isoleucine, leucine, tyrosine), and ketogenesis (3-hydroxybutyrate) were included in the cf-DNA level-related biomarker profiles in at least two of the cohorts.In conclusion, circulating cf-DNA level is different by sex, and related to health behaviour, health decline and metabolic processes common in health and disease. These results can inform future studies where epidemiological and biological pathways of cf-DNA are to be analysed in details, and for studies evaluating cf-DNA as a potential clinical marker.

Background The cline of human genetic diversity observable across Europe is recapitulated at a micro-geographic scale by variation within the Italian population. Besides resulting from extensive gene flow, this might be ascribable also to local adaptations to diverse ecological contexts evolved by people who anciently spread along the Italian Peninsula. Dissecting the evolutionary history of the ancestors of present-day Italians may thus improve the understanding of demographic and biological processes that contributed to shape the gene pool of European populations. However, previous SNP array-based studies failed to investigate the full spectrum of Italian variation, generally neglecting low-frequency genetic variants and examining a limited set of small effect size alleles, which may represent important determinants of population structure and complex adaptive traits. To overcome these issues, we analyzed 38 high-coverage whole-genome sequences representative of population clusters at the opposite ends of the cline of Italian variation, along with a large panel of modern and ancient Euro-Mediterranean genomes. Results We provided evidence for the early divergence of Italian groups dating back to the Late Glacial and for Neolithic and distinct Bronze Age migrations having further differentiated their gene pools. We inferred adaptive evolution at insulin-related loci in people from Italian regions with a temperate climate, while possible adaptations to pathogens and ultraviolet radiation were observed in Mediterranean Italians. Some of these adaptive events may also have secondarily modulated population disease or longevity predisposition. Conclusions We disentangled the contribution of multiple migratory and adaptive events in shaping the heterogeneous Italian genomic background, which exemplify population dynamics and gene-environment interactions that played significant roles also in the formation of the Continental and Southern European genomic landscapes.

BACKGROUND While insulin resistance (IR) is associated with specific metabolite signatures in adults, there have been few truly longitudinal studies in healthy children, either to confirm which abnormalities are present, or to determine whether they precede or result from IR. Therefore, we investigated the association of serum metabolites with IR in childhood in the Earlybird cohort. METHODS The Earlybird cohort is a well-characterized cohort of healthy children with annual measurements from age 5 to 16 years. For the first time, longitudinal association analyses between individual serum metabolites and homeostatic model assessment (HOMA) of insulin resistance (HOMA-IR) have been performed taking into account the effects of age, growth, puberty, adiposity, and physical activity. RESULTS IR was higher in girls than in boys and was associated with increasing body mass index (BMI). In longitudinal analysis IR was associated with reduced concentrations of branched-chain amino acids (BCAA), 2-ketobutyrate, citrate and 3-hydroxybutyrate, and higher concentrations of lactate and alanine. These findings demonstrate the widespread biochemical consequences of IR for intermediary metabolism, ketogenesis, and pyruvate oxidation during normal child growth and development. CONCLUSIONS Longitudinal analysis can differentiate metabolite signatures that precede or follow the development of greater levels of IR. In healthy normal weight children, higher levels of IR are associated with reduced levels of BCAA, ketogenesis, and fuel oxidation. In contrast, elevated lactate concentrations preceded the rise in IR. These changes reveal the metabolite signature of insulin action during normal growth, and they contrast with previous findings in obese children and adults that represent the consequences of IR and obesity.

Extreme longevity is the paradigm of healthy aging as individuals who reached the extreme decades of human life avoided or largely postponed all major age-related diseases. In this study, we sequenced at high coverage (90X) the whole genome of 81 semi-supercentenarians and supercentenarians [105+/110+] (mean age: 106.6 ± 1.6) and of 36 healthy unrelated geographically matched controls (mean age 68.0 ± 5.9) recruited in Italy. The results showed that 105+/110+ are characterized by a peculiar genetic background associated with efficient DNA repair mechanisms, as evidenced by both germline data (common and rare variants) and somatic mutations patterns (lower mutation load if compared to younger healthy controls). Results were replicated in a second independent cohort of 333 Italian centenarians and 358 geographically matched controls. The genetics of 105+/110+ identified DNA repair and clonal haematopoiesis as crucial players for healthy aging and for the protection from cardiovascular events.

Frailty among elderly people leads to an increased risk for negative health outcomes. To prevent frailty, we need a better understanding of the underlying mechanisms and early detection of individuals at risk. Both may be served by identifying candidate (bio)markers, i.e. biomarkers and markers, for the physical, cognitive, and psychological frailty domains. We used univariate (Rank-ANOVA) and multivariate (elastic net) approaches on the RASIG study population (age range: 35–74 years, n = 2220) of the MARK-AGE study to study up to 331 (bio)markers between individuals with and without frailty for each domain. Biomarkers and markers identified by both approaches were studied further regarding their association with frailty using logistic regression. Univariately, we found lower levels of antioxidants, including β-cryptoxanthin and zeaxanthin, in those who were physically, cognitively or psychologically frail. Additionally, self-reported health was worse in these three frail groups. Multivariately, we observed lower levels of β-cryptoxanthin and zeaxanthin in the cognitively frail. Levels of these carotenoids were inversely associated with the risk of being cognitively frail after adjusting for confounders. Antioxidants and self-reported health are potential (bio)markers to detect persons at risk of becoming frail. The biomarkers identified may indicate the involvement of inflammation in frailty, especially for physical and cognitive frailty.

Aim: There is increasing interest in the profiling and quantitation of methionine pathway metabolites for health management research. Currently, several analytical approaches are required to cover metabolites and co-factors. Results: We report the development and the validation of a method for the simultaneous detection and quantitation of 13 metabolites in red blood cells. The method, validated in a cohort of healthy human volunteers, shows a high level of accuracy and reproducibility. Conclusion: This high-throughput protocol provides a robust coverage of central metabolites and co-factors in one single analysis and in a high-throughput fashion. In large-scale clinical settings, the use of such an approach will significantly advance the field of nutritional research in health and disease.

Metabonomics provides access to the environmental, diet, genetic and metagenetic integrative metabolic phenotype of individuals. Recent advances in spectroscopic and chromatographic techniques can capture metabolites and nutrients within a broad dynamic range. Applications to nutrition sciences have addressed nutrient metabolism and diet-related phenotypes under healthy and pathological conditions. Future molecular stratified nutrition requires development of comprehensive, quantitative profiling of nutrients to match the individual’s specific requirements and metabolic peculiarities (metabolic phenotype) to improve health and dietary management of medical conditions.

Population ageing has emerged as a major demographic trend worldwide due to improved health and longevity. This global ageing phenomenon will have a major impact on health-care systems worldwide due to increased morbidity and greater needs for hospitalization/institutionalization. As the ageing population increases worldwide, there is an increasing awareness not only of increased longevity but also of the importance of "healthy ageing" and "quality of life". Yet, the age related chronic inflammation is believed to be pathogenic with regards to its contribution to frailty and degenerative disorders. In particular, the frailty syndrome is increasingly being considered as a key risk indicator of adverse health outcomes. In addition, elderly may be also prone to be resistant to anabolic stimuli which is likely a key factor in the loss of skeletal muscle mass with ageing. Vital to understand these key biological processes is the development of biological markers, through system biology approaches, aiding at strategies for tailored therapeutic and personalized nutritional program. Overall aim is to prevent or attenuate decline of key physiological functions required to live an active, independent life. This review focus on core indicators of health and functions in older adults, where nutrition and tailored personalized programs could exhibit preventive roles, and where the aid of metabolomics technologies are increasingly displaying potential in revealing key molecular mechanisms/targets linked to specific ageing and/or healthy ageing processes. (C) 2013 Elsevier Ireland Ltd. All rights reserved.

Calorie restriction (CR) has long been used to study lifespan effects and oppose the development of a broad array of age-related biological and pathological changes (increase healthspan). Yet, a comprehensive comparison of the metabolic phenotype across different genetic backgrounds to identify common metabolic markers affected by CR is still lacking. Using a system biology approach comprising metabonomics and liver transcriptomics we revealed the effect of CR across multiple mouse strains (129S1/SvlmJ, C57BL6/J, C3H/HeJ, CBA/J, DBA/2J, JC3F1/J). Oligonucleotide microarrays identified 76 genes as differentially expressed in all six strains confirmed. These genes were subjected to quantitative RT-PCR analysis in the C57BL/6J mouse strain, and a CR-induced change expression was confirmed for 14 genes. To fully depict the metabolic pathways affected by CR and complement the changes observed through differential gene expression, the metabolome of C57BL6/J was further characterized in liver tissues, urine and plasma levels using a combination or targeted mass spectrometry and proton nuclear magnetic resonance spectroscopy. Overall, our integrated approach commonly confirms that energy metabolism, stress response, lipids regulators and the insulin/IGF-1 are key determinants factors involved in CR regulation.

Epidemiological studies consistently find that diets rich in whole-grain (WG) cereals lead to decreased risk of disease compared with refined grain (RG)-based diets. Aside from a greater amount of fiber and micronutrients, possible mechanisms for why WGs may be beneficial for health remain speculative. In an exploratory, randomized, researcher-blinded, crossover trial, we measured metabolic profile differences between healthy participants eating a diet based on WGs compared with a diet based on RGs. Seventeen healthy adult participants (11 female, 6 male) consumed a controlled diet based on either WG-rich or RG-rich foods for 2 wk, followed by the other diet after a 5-wk washout period. Both diets were the same except for the use of WG (150 g/d) or FIG foods. The metabolic profiles of plasma, urine, and fecal water were measured using H-1-nuclear magnetic resonance spectroscopy and gas chromatography-mass spectrometry (plasma only). After 1 wk of intervention, the WG diet led to decreases in urinary excretion of metabolites related to protein catabolism (urea, methylguanadine), lipid (carnitine and acylcarnitines) and gut microbial (4-hydroxyphenylacetate, trimethylacetate, dimethylacetate) metabolism in men compared with the same time point during the FIG intervention. There were no differences between the interventions after 2 wk. Urinary urea, carnitine, and acylcarnitine were lower at wk 1 of the WG intervention relative to the FIG intervention in all participants. Fecal water short-chain fatty acids acetate and butyrate were relatively greater after the WG diet compared to the RG diet. Although based on a small population and for a short time period, these observations suggest that a WG diet may affect protein metabolism.

Metabolism is essential to understand human health. To characterize human metabolism, a high-resolution read-out of the metabolic status under various physiological conditions, either in health or disease, is needed. Metabolomics offers an unprecedented approach for generating system-specific biochemical definitions of a human phenotype through the capture of a variety of metabolites in a single measurement. The emergence of large cohorts in clinical studies increases the demand of technologies able to analyze a large number of measurements, in an automated fashion, in the most robust way. NMR is an established metabolomics tool for obtaining metabolic phenotypes. Here, we describe the analysis of NMR-based urinary profiles for metabolic studies, challenged to a large human study (3007 samples). This method includes the acquisition of nuclear Overhauser effect spectroscopy one-dimensional and J-resolved two-dimensional (J-Res-2D) 1H NMR spectra obtained on a 600 MHz spectrometer, equipped with a 120 μL flow probe, coupled to a flow-injection analysis system, in full automation under the control of a sampler manager. Samples were acquired at a throughput of ∼20 (or 40 when J-Res-2D is included) min/sample. The associated technical analysis error over the full series of analysis is 12%, which demonstrates the robustness of the method. With the aim to describe an overall metabolomics workflow, the quantification of 36 metabolites, mainly related to central carbon metabolism and gut microbial host cometabolism, was obtained, as well as multivariate data analysis of the full spectral profiles. The metabolic read-outs generated using our analytical workflow can therefore be considered for further pathway modeling and/or biological interpretation.

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 (n = 80) were fed a standard chow diet for 7 days prior to randomization into a high-fat (HF) (n = 56) or a low-fat (LF) (n = 24) diet group. The (1)H 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 (n = 10) and SR mice (n = 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.

Metabolomics is recognized as a powerful top-down system biological approach to understand genetic-environment-health paradigms paving new avenues to identify clinically relevant biomarkers. It is nowadays commonly used in clinical applications shedding new light on physiological regulatory processes of complex mammalian systems with regard to disease aetiology, diagnostic stratification and, potentially, mechanism of action of therapeutic solutions. A key feature of metabolomics lies in its ability to underpin the complex metabolic interactions of the host with its commensal microbial partners providing a new way to define individual and population phenotypes. This review aims at describing recent applications of metabolomics in clinical fields with insight into diseases, diagnostics/monitoring and improvement of homeostatic metabolic regulation.

Background Hyperhomocysteinemia is a risk factor for cognitive decline and dementia, including Alzheimer disease (AD). Homocysteine (Hcy) is a sulfur-containing amino acid and metabolite of the methionine pathway. The interrelated methionine, purine, and thymidylate cycles constitute the one-carbon metabolism that plays a critical role in the synthesis of DNA, neurotransmitters, phospholipids, and myelin. In this study, we tested the hypothesis that one-carbon metabolites beyond Hcy are relevant to cognitive function and cerebrospinal fluid (CSF) measures of AD pathology in older adults. Methods Cross-sectional analysis was performed on matched CSF and plasma collected from 120 older community-dwelling adults with (n = 72) or without (n = 48) cognitive impairment. Liquid chromatography-mass spectrometry was performed to quantify one-carbon metabolites and their cofactors. Least absolute shrinkage and selection operator (LASSO) regression was initially applied to clinical and biomarker measures that generate the highest diagnostic accuracy of a priori-defined cognitive impairment (Clinical Dementia Rating-based) and AD pathology (i.e., CSF tau phosphorylated at threonine 181 [p-tau181]/β-Amyloid 1–42 peptide chain [Aβ1–42] >0.0779) to establish a reference benchmark. Two other LASSO-determined models were generated that included the one-carbon metabolites in CSF and then plasma. Correlations of CSF and plasma one-carbon metabolites with CSF amyloid and tau were explored. LASSO-determined models were stratified by apolipoprotein E (APOE) ε4 carrier status. Results The diagnostic accuracy of cognitive impairment for the reference model was 80.8% and included age, years of education, Aβ1–42, tau, and p-tau181. A model including CSF cystathionine, methionine, S-adenosyl-L-homocysteine (SAH), S-adenosylmethionine (SAM), serine, cysteine, and 5-methyltetrahydrofolate (5-MTHF) improved the diagnostic accuracy to 87.4%. A second model derived from plasma included cystathionine, glycine, methionine, SAH, SAM, serine, cysteine, and Hcy and reached a diagnostic accuracy of 87.5%. CSF SAH and 5-MTHF were associated with CSF tau and p-tau181. Plasma one-carbon metabolites were able to diagnose subjects with a positive CSF profile of AD pathology in APOE ε4 carriers. Conclusions We observed significant improvements in the prediction of cognitive impairment by adding one-carbon metabolites. This is partially explained by associations with CSF tau and p-tau181, suggesting a role for one-carbon metabolism in the aggregation of tau and neuronal injury. These metabolites may be particularly critical in APOE ε4 carriers. Electronic supplementary material The online version of this article (doi:10.1186/s13195-017-0270-x) contains supplementary material, which is available to authorized users.

Cooperative effects of magnesium ions and acidic polypeptides originating from a family of proteins known as Asprich (mollusk Atrina rigida) were studied. In our previous studies, these two acidic polypeptides were found to be effective in controlling the morphology of the calcium carbonate mineral, the main inorganic constituent of prismatic layer of the mollusk shell. Since these Asprich sequences are believed to contain a putative magnesium binding domain, the morphology-controlling effects were further investigated with the addition of magnesium ions. The mineral morphology was dramatically changed by the combined influence of each polypeptides and the magnesium ions, substantiating the recognized importance of magnesium in the formation of calcium carbonate-based biominerals.

Recent advances in molecular biology and microbiology have increased awareness on the importance of the gut microbiota to the overall mammalian host's health status. There is therefore increasing interest in nutrition research to characterise the molecular foundations of the gut microbial mammalian crosstalk at both physiological and biochemical pathway levels. Tackling these challenges can be achieved through systems biology strategies based on the measurement of metabolites to assess the highly complex metabolic exchanges between diverse biological compartments, including organs, biofluids and microbial symbionts. By opening a direct biochemical window into the metabolome, metabonomics is uniquely suited for the identification of biomarkers providing better understanding of these complex metabolic processes. Recent applications of top-down system biology based on (1)H NMR spectroscopy coupled to advanced chemometric modelling approaches provided compelling evidence that system-wide and organ-specific changes in biochemical processes may be finely tuned by gut microbial activities. This review aims at describing current advances in NMR-based metabonomics where the main objective is to discern the molecular pathways and biochemical mechanisms under the influence of the gut microbiota. Furthermore, emphasis is given on nutritional approaches, where the quest for homeostatic balance is dependent not only on the host but also on the nutritional modulation of the gut microbiota-host metabolic interactions, using, for instance, probiotics and prebiotics.

The underlying biochemical consequences of inflammatory bowel disease (IBD) on the systemic and gastrointestinal metabolism have not yet been fully elucidated but could help to better understand the disease pathogenesis and to identify tissue-specific markers associated with the different disease stages. Here, we applied a metabonomic approach to monitor metabolic events associated with the gradual development of Crohn's disease (CD)-like ileitis in the TNF(ΔARE/WT) mouse model. Metabolic profiles of different intestinal compartments from the age of 4 up to 24 weeks were generated by combining proton nuclear magnetic resonance ((1)H NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS). From 8 weeks onward, mice developed CD similar to the immune and tissue-related phenotype of human CD with ileal involvement, including ileal histological abnormalities, reduced fat mass and body weight, as well as hallmarks of malabsorption with higher energy wasting. The metabonomic approach highlighted shifts in the intestinal lipid metabolism concomitant to the histological onset of inflammation. Moreover, the advanced disease status was characterized by a significantly altered metabolism of cholesterol, triglycerides, phospholipids, plasmalogens, and sphingomyelins in the inflamed tissue (ileum) and the adjacent intestinal parts (proximal colon). These results describe different biological processes associated with the disease onset, including modifications of the general cell membrane composition, alteration of energy homeostasis, and finally the generation of inflammatory lipid mediators. Taken together, this provides novel insights into IBD-related alterations of specific lipid-dependant processes during inflammatory states.

Insulin/IGF-like signaling (IIS) and nutrient sensing are among the most potent regulators of health status and aging. Here, a global view of the metabolic changes in C. elegans with impaired function of IIS represented by daf-2 and daf-16 and the intestinal di- and tripeptide transport pept-1 was generated using (1)H nuclear magnetic resonance spectroscopic analysis of worm extracts and spent culture media. We showed that specific metabolic profiles were significantly associated with each type of mutant. On the basis of the metabonomics data, selected underlying processes were further investigated using proteomic and transcriptomic approaches. The observed changes suggest a decreased activity of the one carbon metabolism in pept-1(lg601) mutants. Higher concentration of branched-chain amino acids (BCAA) and altered transcript levels of genes involved in BCAA metabolism were observed in long-living strains daf-2(e1370) and daf-2(e1370);pept-1(lg601) when compared to wild types and daf-16(m26);daf-2(e1370);pept-1(lg601) C. elegans, suggesting a DAF-16-dependent mechanism.

In man, living environment and lifestyle influence host physiology and microbial populations, and these interactions determine the health of individuals. In particular, dietary preferences are partially triggered by genetically-mediated taste responses to specific foods and may be reflected in the metabolic phenotype. However, identification of dietary metabolic signatures in free-living populations is made difficult by the relatively low amplitude of nutritional modulations when compared to inherent intra- and inter-individual variability that results from numerous confounding factors, e.g. sampling, lifestyle, gender and ageing. The identification of dietary biomarkers lies in the application of chemometrics to recover key metabolic information from the high density and highly variable metabolic phenotypes. One preferred approach is the use of linear regression models such as Partial Least Squares – Discriminant Analysis, whose reliability depends on appropriate validation procedures. However, the high inter- and intra-individual variability in free-living populations makes difficult the direct application of consolidated validation methodologies such as n-fold cross-validation that often leads to unstable solutions. In the present manuscript, we propose a new validation strategy based on the combination of several methodologies, merging n-fold cross validation and bootstrapping procedures with the confirmation of the reliability of the model through the generation of a set of replicate models. The proposed method was employed to validate the identification of dietary specific metabolic biomarkers in 1 H NMR and UPLC TOFMS urine metabolic profiles obtained from 21 free-living subjects stratified according to their sensitivity to propylthiouracil.

Dietary preferences influence basal human metabolism and gut microbiome activity that in turn may have long-term health consequences. The present study reports the metabolic responses of free living subjects to a daily consumption of 40 g of dark chocolate for up to 14 days. A clinical trial was performed on a population of 30 human subjects, who were classified in low and high anxiety traits using validated psychological questionnaires. Biological fluids (urine and blood plasma) were collected during 3 test days at the beginning, midtime and at the end of a 2 week study. NMR and MS-based metabonomics were employed to study global changes in metabolism due to the chocolate consumption. Human subjects with higher anxiety trait showed a distinct metabolic profile indicative of a different energy homeostasis (lactate, citrate, succinate, trans-aconitate, urea, proline), hormonal metabolism (adrenaline, DOPA, 3-methoxy-tyrosine) and gut microbial activity (methylamines, p-cresol sulfate, hippurate). Dark chocolate reduced the urinary excretion of the stress hormone cortisol and catecholamines and partially normalized stress-related differences in energy metabolism (glycine, citrate, trans-aconitate, proline, beta-alanine) and gut microbial activities (hippurate and p-cresol sulfate). The study provides strong evidence that a daily consumption of 40 g of dark chocolate during a period of 2 weeks is sufficient to modify the metabolism of free living and healthy human subjects, as per variation of both host and gut microbial metabolism.

Metabonomics is a well established analytical approach for the analysis of physiological regulatory processes via the metabolic profiling of biofluids and tissues in living organisms. Its potential is fully exploited in the field of “nutrimetabonomics” that aims at assessing the metabolic effects of active ingredients and foods in individuals. Yet, one of the greatest challenges in nutrition research is to decipher the critical interactions between mammalian organisms and environmental factors, including the gut microbiota. “Nutrimetabonomics” is today foreseen as a powerful approach for future nutritional programs tailored at health maintenance and disease prevention.

Multicompartmental metabolic profiling combined with multivariate data analysis offers a unique opportunity to explore the multidimensional metabolic relationships between various biological matrices. Here, we applied unsupervised chemometric methods for integrating 1H NMR metabolic profiles from mouse plasma, liver, pancreas, adrenal gland and kidney cortex matrices in order to infer intercompartments functional links. Principal Component Analysis (PCA) revealed metabolic differences between matrices but contained limited information on intercompartment metabolic relationships. Multiway PCA enabled the assessment of interindividual metabolic variability across multiple compartments in a single model and, therefore, metabolic correlations between different organs and circulating biofluids. However, this approach does not provide information on the relative contribution of one compartment to another. Integration of metabolic profiles using Multivariate Curve Resolution (MCR) and Parallel Factor Analysis (PARAFAC) methods provided an overview of functional relationships across matrices and enabled the characterization of compartment-specific metabolite signatures, the spectrotypes. In particular, the spectrotypes describe biochemical profiles specific or common to different biological compartments. Consequently, MCR-ALS and PARAFAC appeared to be better adapted for stepwise variable and compartment selection for further correlation analysis. Such a combination of chemometric techniques could provide new research avenues to assess the efficacy of drug or nutritional interventions on targeted organs.

Despite extensive recent reports on combinatorially selected inorganic-binding peptides and their bionanotechnological utility as synthesizers and molecular linkers, there is still only limited knowledge about the molecular mechanisms of peptide binding to solid surfaces. There is, therefore, much work that needs to be carried out in terms of both the fundamentals of solid-binding kinetics of peptides and the effects of peptide primary and secondary structures on their recognition and binding to solid materials. Here we discuss the effects of constraints imposed on FliTrx-selected gold-binding peptide molecular structures upon their quantitative gold-binding affinity. We first selected two novel gold-binding peptide (AuBP) sequences using a FliTrx random peptide display library. These were, then, synthesized in two different forms: cyclic (c), reproducing the original FliTrx gold-binding sequence as displayed on bacterial cells, and linear (l) dodecapeptide gold-binding sequences. All four gold-binding peptides were then analyzed for their adsorption behavior using surface plasmon resonance spectroscopy. The peptides exhibit a range of binding affinities to and adsorption kinetics on gold surfaces, with the equilibrium constant, Keq, varying from 2.5x10(6) to 13.5x10(6) M(-1). Both circular dichroism and molecular mechanics/energy minimization studies reveal that each of the four peptides has various degrees of random coil and polyproline type II molecular conformations in solution. We found that AuBP1 retained its molecular conformation in both the c- and l-forms, and this is reflected in having similar adsorption behavior. On the other hand, the c- and l-forms of AuBP2 have different molecular structures, leading to differences in their gold-binding affinities.

Many of the interesting mechanical and materials properties of the mollusk shell are thought to stem from the prismatic calcite crystal assemblies within this composite structure. It is now evident that proteins play a major role in the formation of these assemblies. Recently, a superfamily of seven conserved prismatic layer-specific mollusk shell proteins, Asprich, were sequenced, and the conserved 42 AA C-terminal sequence region of this protein superfamily was found to introduce surface voids or porosities on calcite crystals in vitro. Using AFM imaging techniques, we further investigate the effect that this 42 AA domain (Fragment-2) and its constituent subdomains, DEAD-17 and Acidic-2, have on the morphology and growth kinetics of calcite dislocation hillocks. We find that Fragment-2 adsorbs on terrace surfaces and pins acute steps, accelerates and then decelerates the growth of obtuse steps, forms clusters and voids on terrace surfaces, and transforms calcite hillock morphology from a rhombohedral fo...

Usually the genetics of human longevity is restricted to the nuclear genome (nDNA). However it is well known that the nDNA interacts with a physically and functionally separated genome, the mitochondrial DNA (mtDNA) that, even if limited in length and number of genes encoded, plays a major role in the ageing process. The complex interplay between nDNA/mtDNA and the environment is most likely involved in phenomena such as ageing and longevity. To this scenario we have to add another level of complexity represented by the microbiota, that is, the whole set of bacteria present in the different part of our body with their whole set of genes. In particular, several studies investigated the role of gut microbiota (GM) modifications in ageing and longevity and an age-related GM signature was found. In this view, human being must be considered as “metaorganism” and a more holistic approach is necessary to grasp the complex dynamics of the interaction between the environment and nDNA-mtDNA-GM of the host during ageing. In this review, the relationship between the three genetics and human longevity is addressed to point out that a comprehensive view will allow the researchers to properly address the complex interactions that occur during human lifespan.

Systems Medicine (SM) can be defined as an extension of Systems Biology (SB) to Clinical-Epidemiological disciplines through a shifting paradigm, starting from a cellular, toward a patient centered framework. According to this vision, the three pillars of SM are Biomedical hypotheses, experimental data, mainly achieved by Omics technologies and tailored computational, statistical and modeling tools. The three SM pillars are highly interconnected, and their balancing is crucial. Despite the great technological progresses producing huge amount of data (Big Data) and impressive computational facilities, the Bio-Medical hypotheses are still of primary importance. A paradigmatic example of unifying Bio-Medical theory is the concept of Inflammaging. This complex phenotype is involved in a large number of pathologies and patho-physiological processes such as aging, age-related diseases and cancer, all sharing a common inflammatory pathogenesis. This Biomedical hypothesis can be mapped into an ecological perspective capable to describe by quantitative and predictive models some experimentally observed features, such as microenvironment, niche partitioning and phenotype propagation. In this article we show how this idea can be supported by computational methods useful to successfully integrate, analyze and model large data sets, combining cross-sectional and longitudinal information on clinical, environmental and omics data of healthy subjects and patients to provide new multidimensional biomarkers capable of distinguishing between different pathological conditions, e.g. healthy versus unhealthy state, physiological versus pathological aging.

Nature's use of biomineralization polypeptides to control and modulate the growth of biogenic minerals is an important process that, if properly understood, could have significant implications for designing and creating new inorganic-based materials. Although the sequences for a number of biomineralization proteins exist, very little is known about the participation of specific amino acids in the mineral modulation process. In this letter, we investigate the impact of global Asp --Asn and Glu --Gln substitutions on two mollusk shell nacre polypeptides, AP7N and n16N. We find that these global substitutions, which remove all anionic Ca(II) binding sites, abolish the expected in vitro mineralization activities associated with each native polypeptide. In addition, the ability of substituted peptides to form complexes with both Ca(II) and Ca(II) metal ion analogs is also abolished. However, some unexpected effects were noted. First, the Asp --Asn, Glu --Gln substituted n16N polypeptide is observed to self-assemble and form biofilms or coatings that appear to mineralize in vitro. Second, both polypeptides are structurally affected by these substitutions, with Asp --Asn substituted AP7N transforming to an alpha helix and Asp --Asn, Glu --Gln substituted n16N transforming to a more unfolded random-coil-like structure. We find that the participation of Asp and Glu residues is crucial to the inherent mineralization activities and conformations of AP7N and n16N polypeptides. Surprisingly, we find that the replacement of anionic residues within biomineralization polypeptides such as n16N still permits mineral modulation, but in a different form that now involves peptide self-association and biofilm formation.

AP7 and AP24 are mollusk shell proteins which are responsible for aragonite polymorph formation and stabilization within the nacre layer of the Pacific red abalone, Haliotis rufescens. It is known that the 30-AA N-terminal mineral modification domains of both proteins (AP7N, AP24N) possess identical multifunctional mineralization capabilities within in vitro assays but differ in terms of rate kinetics, with AP24N > AP7N. In this report, we identify previously unreported molecular features of AP24N and contrast the lowest energy polypeptide backbone structures of AP24N (planar configuration) with that of AP7N ("bent paper clip" configuration) using NMR data and simulated annealing molecular dynamics structure refinement. Like AP7N, we find that AP24N possesses an unfolded conformation, can sequester Ca(II) and other multivalent metal ions, can adsorb onto or within calcite crystals, and possesses anionic and cationic electrostatic "pocket" regions on its molecular surfaces. However, AP24N has some unique features: greater conformational responsiveness to Ca(II), the tendency to form a more planar backbone configuration, and longer anionic and hydrogen-bonding donor/acceptor sequence blocks. We conclude that the presence of unfolded polypeptide conformation, electrostatic surface pockets, and interactive sequence clustering endow both AP7N and AP24N with similar features that lead to comparable effects on crystal morphology and nucleation. However, AP24N possesses longer anionic and hydrogen-bonding sequence clusters and exhibits a tendency to adopt a more planar backbone configuration than AP7N does. We believe that these features facilitate peptide-mineral, peptide-ion, or water cluster interactions, thereby enhancing the mineralization kinetics of AP24N over AP7N.

Many candidate biomarkers of human ageing have been proposed in the scientific literature but in all cases their variability in cross-sectional studies is considerable, and therefore no single measurement has proven to serve a useful marker to determine, on its own, biological age. A plausible reason for this is the intrinsic multi-causal and multi-system nature of the ageing process. The recently completed MARK-AGE study was a large-scale integrated project supported by the European Commission. The major aim of this project was to conduct a population study comprising about 3200 subjects in order to identify a set of biomarkers of ageing which, as a combination of parameters with appropriate weighting, would measure biological age better than any marker in isolation. (C) 2015 The Authors. Published by Elsevier Ireland Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Protein-mediated biomineral formation and polymorph selection in the mollusk shell involves the participation of a number of different proteins. One of these proteins, AP7, has been identified as a participant in nacre (aragonite) formation in the mollusk Haliotis rufescens. However, the role(s) of this protein in aragonite formation is (are) poorly understood, due to the fact that low quantities of this protein are recoverable from the nacre matrix. To overcome this problem, we employed stepwise solid-phase tBOC synthesis to recreate the 66-AA single-chain protein AP7 and utilized this synthetic form for in vitro mineralization studies. We find that the AP7 protein promotes incomplete or interrupted crystal growth and step edge roundening in a concentration-dependent fashion. Using synthetic peptides which represent the 30-AA N-terminal (AP7N) and 36-AA C-terminal (AP7C) subdomains of AP7, we have identified that the mineral modification activity of AP7 is localized to the unstructured, conformationally labile N-terminal subdomain. Interestingly, the 36-AA C-terminal subdomain has no observable direct effect on in vitro calcium carbonate crystal growth; however, we cannot rule out the possibility that AP7C plays an indirect role in AP7 mineralization activity. Qualitative structural studies reveal that AP7, although possessing Zn(II) fingerlike -His-(X) 6 -His, -Cys-(X) 2 -Cys-, and -Cys-(X) 4 -Cys- motifs within its C-terminal region, does not possess the structural characteristics of known Zn(II) finger polypeptides, as evidenced by the presence of an ordered, α-helical conformation within the C-terminal subdomain of apo-AP7. Given its associative nature with AP24 and the multifunctional capabilities of the 30-AA N-terminal domain, it is likely that the AP7 protein possesses multifunctional capabilities with regard to nacre formation within the mollusk shell.

The formation of the mineralized mollusk shell involves the participation specific proteins to control mineral formation and morphology. Recent sequencing studies have identified a subfamily of seven proteins that are specific for the prismatic or calcitic layer of the mollusk Atrina rigida. This subfamily, named Asprich a through g, are polyanionic in sequence and contain conserved sequence regions at the N- and C-termini that are believed to play a role in mineral formation. Using solid-phase peptide synthesis, we have recreated the 42-AA C-terminal domain of Asprich which contains two conserved subdomains, the 17 AA Asp, Glu, Ala repeat (DEAD17) and the Asp, Glu-rich 25 AA acidic-2 region. Using in vitro calcium carbonate mineralization assays, we demonstrate that the 42 AA C-terminal domain exerts effects on the morphology of forming calcite crystals, leading to the formation of porosities and irregular crystal growth. The two subdomains, DEAD17 and acidic-2, exert similar, albeit weaker, effects on c...