Revisiting biomarker discovery by plasma proteomics

Clinical analysis of blood is the most widespread diagnostic procedure in medicine, and blood biomarkers are used to categorize patients and to support treatment decisions. However, existing biomarkers are far from comprehensive and often lack specificity and new ones are being developed at a very slow rate. As described in this review, mass spectrometry (MS)-based proteomics has become a powerful technology in biological research and it is now poised to allow the characterization of the plasma proteome in great depth. Previous "triangular strategies" aimed at discovering single biomarker candidates in small cohorts, followed by classical immunoassays in much larger validation cohorts. We propose a "rectangular" plasma proteome profiling strategy, in which the proteome patterns of large cohorts are correlated with their phenotypes in health and disease. Translating such concepts into clinical practice will require restructuring several aspects of diagnostic decision-making, and we discuss some first steps in this direction

Effects of a Follow-On Formula Containing Isomaltulose (Palatinose) on Metabolic Response, Acceptance, Tolerance and Safety in Infants: A Randomized-Controlled Trial

UNLABELLED:Effects of the dietary glycaemic load on postprandial blood glucose and insulin response might be of importance for fat deposition and risk of obesity. We aimed to investigate the metabolic effects, acceptance and tolerance of a follow-on formula containing the low glycaemic and low insulinaemic carbohydrate isomaltulose replacing high glycaemic maltodextrin. Healthy term infants aged 4 to 8 completed months (n = 50) were randomized to receive the intervention follow-on formula (IF, 2.1g isomaltulose (Palatinose™)/100mL) or an isocaloric conventional formula (CF) providing 2.1g maltodextrin/100mL for four weeks. Plasma insulinaemia 60 min after start of feeding (primary outcome) was not statistically different, while glycaemia adjusted for age and time for drinking/volume of meal 60 min after start of feeding was 122(105,140) mg/dL in IF (median, interquartile range) and 111(100,123) in CF (p = 0.01). Urinary c-peptide:creatinine ratio did not differ (IF:81.5(44.7, 96.0) vs. CF:56.8(37.5, 129),p = 0.43). Urinary c-peptide:creatinine ratio was correlated total intake of energy (R = 0.31,p = 0.045), protein (R = 0.42,p = 0.006) and fat (R = 0.40,p = 0.01) but not with carbohydrate intake (R = 0.22,p = 0.16). Both formulae were well accepted without differences in time of crying, flatulence, stool characteristics and the occurrence of adverse events. The expected lower postprandial plasma insulin and blood glucose level due to replacement of high glycaemic maltodextrin by low glycaemic isomaltulose were not observed in the single time-point blood analysis. In infants aged 4 to 8 completed months fed a liquid formula, peak blood glucose might be reached earlier than 60 min after start of feeding. Non-invasive urinary c-peptide measurements may be a suitable marker of nutritional intake during the previous four days in infants. TRIAL REGISTRATION:ClinicalTrials.gov NCT01627015

Association of the habitual dietary intake with the fatty liver index and effect modification by metabotypes in the population-based KORA-Fit study

Background Non-alcoholic fatty liver disease (NAFLD) is an emerging threat for public health with diet being a major risk factor in disease development and progression. However, the effects of habitual food consumption on fatty liver are still inconclusive as well as the proposed role of the individuals’ metabolic profiles. Therefore, the aim of our study is to examine the associations between diet and NAFLD with an emphasis on the influence of specific metabotypes in the general population. Methods A total of 689 participants (304 men and 385 women) of the KORA-Fit (S4) survey, a follow-up study of the population-based KORA cohort study running in the Region of Augsburg, Germany, were included in this analysis. Dietary information was derived from repeated 24-h food lists and a food frequency questionnaire. The intake of energy and energy-providing nutrients were calculated using the national food composition database. The presence of fatty liver was quantified by the fatty liver index (FLI), and metabotypes were calculated using K-means clustering. Multivariable linear regression models were used for the analysis of habitual food groups and FLI; for the evaluation of macronutrients, energy substitution models were applied. Results A higher consumption of nuts and whole grains, and a better diet quality (according to Alternate Healthy Eating Index and Mediterranean Diet Score) were associated with lower FLI values, while the intake of soft drinks, meat, fish and eggs were associated with a higher FLI. The isocaloric substitution of carbohydrates with polyunsaturated fatty acids was associated with a decreased FLI, while substitution with monounsaturated fatty acids and protein showed increased FLI. Statistically significant interactions with the metabotype were observed for most food groups. Conclusion The consumption of plant-based food groups, including nuts and whole grains, and diet quality, were associated with lower FLI values, whereas the intake of soft drinks and products of animal origin (meat, fish, eggs) were associated with a higher FLI. The observed statistically significant interactions with the metabotype for most food groups could help to develop targeted prevention strategies on a population-based level if confirmed in

Are C-Reactive Protein Associated Genetic Variants Associated with Serum Levels and Retinal Markers of Microvascular Pathology in Asian Populations from Singapore?

Introduction:C-reactive protein (CRP) levels are associated with cardiovascular disease and systemic inflammation. We assessed whether CRP-associated loci were associated with serum CRP and retinal markers of microvascular disease, in Asian populations.Methods:Genome-wide association analysis (GWAS) for serum CRP was performed in East-Asian Chinese (N = 2,434) and Malays (N = 2,542) and South-Asian Indians (N = 2,538) from Singapore. Leveraging on GWAS data, we assessed, in silico, association levels among the Singaporean datasets for 22 recently identified CRP-associated loci. At loci where directional inconsistencies were observed, quantification of inter-ethnic linkage disequilibrium (LD) difference was determined. Next, we assessed association for a variant at CRP and retinal vessel traits [central retinal artery equivalent (CRAE) and central retinal vein equivalent (CRVE)] in a total of 24,132 subjects of East-Asian, South-Asian and European ancestry.Results:Serum CRP was associated with SNPs in/near APOE, CRP, HNF1A and LEPR (p-values ≤4.7×10-8) after meta-analysis of Singaporean populations. Using a candidate-SNP approach, we further replicated SNPs at 4 additional loci that had been recently identified to be associated with serum CRP (IL6R, GCKR, IL6 and IL1F10) (p-values ≤0.009), in the Singaporean datasets. SNPs from these 8 loci explained 4.05% of variance in serum CRP. Two SNPs (rs2847281 and rs6901250) were detected to be significant (p-value ≤0.036) but with opposite effect directions in the Singaporean populations as compared to original European studies. At these loci we did not detect significant inter-population LD differences. We further did not observe a significant association between CRP variant and CRVE or CRAE levels after meta-analysis of all Singaporean and European datasets (p-value >0.058).Conclusions:Common variants associated with serum CRP, first detected in primarily European studies, are also associated with CRP levels in East-Asian and South-Asian populations. We did not find a causal link between CRP and retinal measures of microvascular disease

Alteration of Multiple Leukocyte Gene Expression Networks is Linked with Magnetic Resonance Markers of Prognosis After Acute ST-Elevation Myocardial Infarction

Prognostic relevant pathways of leukocyte involvement in human myocardial ischemic-reperfusion injury are largely unknown. We enrolled 136 patients with ST-elevation myocardial infarction (STEMI) after primary angioplasty within 12 h after onset of symptoms. Following reperfusion, whole blood was collected within a median time interval of 20 h (interquartile range: 15-25 h) for genome-wide gene expression analysis. Subsequent CMR scans were performed using a standard protocol to determine infarct size (IS), area at risk (AAR), myocardial salvage index (MSI) and the extent of late microvascular obstruction (lateMO). We found 398 genes associated with lateMO and two genes with IS. Neither AAR, nor MSI showed significant correlations with gene expression. Genes correlating with lateMO were strongly related to several canonical pathways, including positive regulation of T-cell activation (p = 3.44 x 10(-5)), and regulation of inflammatory response (p = 1.86 x 10(-3)). Network analysis of multiple gene expression alterations associated with larger lateMO identified the following functional consequences: facilitated utilisation and decreased concentration of free fatty acid, repressed cell differentiation, enhanced phagocyte movement, increased cell death, vascular disease and compensatory vasculogenesis. In conclusion, the extent of lateMO after acute, reperfused STEMI correlated with altered activation of multiple genes related to fatty acid utilisation, lymphocyte differentiation, phagocyte mobilisation, cell survival, and vascular dysfunction

Serum concentrations of TNF-α, sTNF-R p55 and p75 and post-traumatic stress in German soldiers

Growing evidence suggests involvement of the tumor necrosis factor (TNF)-α system in the pathophysiology of psychiatric disorders. Research into post-traumatic stress disorder (PTSD) has investigated serum levels of TNF-α, but not to date its soluble receptors sTNF-R p55 and sTNF-R p75. We examined serum levels of TNF-α, sTNF-R p55 and sTNF-R p75 in 135 male German soldiers 70 of whom had been deployed abroad and 65 in Germany only. Post-traumatic stress symptoms were measured using the Post-traumatic Stress Diagnostic Scale (PDS) and the Trier Inventory for the Assessment of Chronic Stress (TICS). Correlational analysis controlling for multiple testing, showed no significant Spearman rank correlations between PDS or TICS scores and serum levels of TNF-α, sTNF-R p55 or sTNF-R p75, either in the full sample or in the group of soldiers who had been deployed abroad. ANCOVAs showed no significant differences between soldiers with or without a PDS-derived diagnosis of PTSD, or between soldiers with or without deployment abroad, after controlling for age, smoking and body mass index (BMI). These results suggest that the TNF-α system, as reflected by TNF-α, sTNF-R p55 and sTNF-R p75 serum levels, does not play a major role in the pathophysiology and development of PTSD symptoms as measured by the PDS and the TICS. However, several methodological and contextual issues have to be considered.</p

Epigenetic signatures at AQP3 and SOCS3 engage in low-grade inflammation across different tissues

Background Elevated levels of C-reactive protein (CRP, determined by a high-sensitivity assay) indicate low-grade inflammation which is implicated in many age-related disorders. Epigenetic studies on CRP might discover molecular mechanisms underlying CRP regulation. We aimed to identify DNA methylation sites related to CRP concentrations in cells and tissues regulating low-grade inflammation. Results Genome-wide DNA methylation was measured in peripheral blood in 1,741 participants of the KORA F4 study using Illumina HumanMethylation450 BeadChip arrays. Four CpG sites (located at BCL3, AQP3, SOCS3, and cg19821297 intergenic at chromosome 19p13.2, P ≤ 1.01E-07) were significantly hypomethylated at high CRP concentrations independent of various confounders including age, sex, BMI, smoking, and white blood cell composition. Findings were not sex-specific. CRP-related top genes were enriched in JAK/ STAT pathways (Benjamini-Hochberg corrected P &lt;0.05). Results were followed-up in three studies using DNA from peripheral blood (EPICOR, n = 503) and adipose tissue (TwinsUK, n = 368) measured as described above and from liver tissue (LMU liver cohort, n = 286) measured by MALDI-TOF mass spectrometry using EpiTYPER. CpG sites at the AQP3 locus (significant p-values in peripheral blood = 1.72E-03 and liver tissue = 1.51E-03) and the SOCS3 locus (p-values in liver &lt;2.82E-05) were associated with CRP in the validation panels. Conclusions Epigenetic modifications seem to engage in low-grade inflammation, possibly via JAK/ STAT mediated pathways. Results suggest a shared relevance across different tissues at the AQP3 locus and highlight a role of DNA methylation for CRP regulation at the SOCS3 locus.</p

Polymorphism of the CD36 Gene and Cardiovascular Risk Factors in Patients with Coronary Artery Disease Manifested at a Young Age

This study investigates potential associations between CD36 gene variants and the presence of risk factors in Caucasians with coronary artery disease (CAD) manifested at a young age. The study group consisted of 90 patients; the men were ≤ 50 years old and the women were ≤ 55 years old. Amplicons of exons 4 and 5 including fragments of introns were analyzed by DHPLC. Two polymorphisms were found: IVS3-6 T/C (rs3173798) and IVS4-10 G/A (rs3211892). The C allele of the IVS3-6 T/C polymorphism was associated with higher prevalence of obesity and diabetes, higher hsCRP, lower Lp(a) serum concentrations, and younger age at myocardial infarction. The A allele of the IVS4-10 G/A polymorphism was associated with older age of myocardial infarction and higher white blood cell count. The functional role of CD36 polymorphisms in CAD development needs further research