Proteomic insights on the metabolism in inflammatory bowel disease

Inflammatory bowel diseases (IBD) are chronic and relapsing inflammatory conditions of the gut that include Crohn's disease and ulcerative colitis. The pathogenesis of IBD is not completely unraveled, IBD are multi-factorial diseases with reported alterations in the gut microbiota, activation of different immune cell types, changes in the vascular endothelium, and alterations in the tight junctions\u2019 structure of the colonic epithelial cells. Proteomics represents a useful tool to enhance our biological understanding and to discover biomarkers in blood and intestinal specimens. It is expected to provide reproducible and quantitative data that can support clinical assessments and help clinicians in the diagnosis and treatment of IBD. Sometimes a differential diagnosis of Crohn's disease and ulcerative colitis and the prediction of treatment response can be deducted by finding meaningful biomarkers. Although some non-invasive biomarkers have been described, none can be considered as the \u201cgold standard\u201d for IBD diagnosis, disease activity and therapy outcome. For these reason new studies have proposed an \u201cIBD signature\u201d, which consists in a panel of biomarkers used to assess IBD. The above described approach characterizes \u201comics\u201d and in this review we will focus on proteomics

Diversity of human skeletal muscle in health and disease: contribution of proteomics

Muscle represents a large fraction of the human body mass. It is an extremely heterogeneous tissue featuring in its contractile structure various proportions of heavy- and light-chain slow type 1 and fast types 2A and 2X myosins, actins, tropomyosins, and troponin complexes as well as metabolic proteins (enzymes and most of the players of the so-called excitation-transcription coupling). Muscle is characterized by wide plasticity, i.e. capacity to adjust size and functional properties in response to endogenous and exogenous influences. Over the last decade, proteomics has become a crucial technique for the assessment of muscle at the molecular level and the investigation of its functional changes. Advantages and shortcomings of recent techniques for muscle proteome analysis are discussed. Data from differential proteomics applied to healthy individuals in normal and unusual environments (hypoxia and cold), in exercise, immobilization, aging and to patients with neuromuscular hereditary disorders (NMDs), inclusion body myositis and insulin resistance are summarized, critically discussed and, when required, compared with homologous data from pertinent animal models. The advantages as well as the limits of proteomics in view of the identification of new biomarkers are evaluate

Endoplasmic Reticulum Associated Aminopeptidase 2 (ERAP2) is released in the secretome of activated MDMs and reduces in vitro HIV-1 infection

Background: Haplotype-specific alternative splicing of the endoplasmic reticulum (ER) aminopeptidase type 2 (ERAP2) gene results in either full-length (FL, haplotype A) or alternatively spliced (AS, haplotype B) mRNA. HapA/HapA homozygous (HomoA) subjects show a reduced susceptibility to HIV-1 infection, probably secondary to the modulation of the antigen processing/presenting machinery. ERAP1 was recently shown to be secreted from the plasma membrane in response to activation; we investigated whether ERAP2 can be released as well and if the secreted form of this enzyme retains its antiviral function. Methods: Human monocyte derived macrophages (MDMs) were differentiated from peripheral blood mononuclear cells (PBMCs) isolated from 6 HomoA healthy controls and stimulated with IFN\u3b3 and LPS. ERAP2-FL secretion was evaluated by mass spectrometry. PBMCs (14 HomoA and 16 HomoB) and CD8-depleted PBMCs (CD8-PBMCs) (4 HomoA and 4 HomoB) were in vitro HIV-infected in the absence/presence of recombinant human ERAP2-FL (rhERAP2) protein; p24 viral antigen quantification was used to assess viral replication. IFN\u3b3 and CD69 mRNA expression, as well as the percentage of perforin-producing CD8+ T Lymphocytes, were analyzed 3 and 7-days post in vitro HIV-1-infection, respectively. The effect of rhERAP2 addition in cell cultures on T cell apoptosis, proliferation, activation, and maturation was evaluated as well on 24 h-stimulated PBMCs. Results: ERAP2 can be secreted from human MDMs in response to IFN\u3b3/LPS stimulation. Notably, the addition of rhERAP2 to PBMC and CD8-PBMC cultures resulted in the reduction of viral replication, though these differences were statistically significant only in PBMCs (p < 0.05 in both HomoA and HomoB). This protective effect was associated with an increase in IFN\u3b3 and CD69 mRNA expression and in the percentage of perforin-expressing CD107+CD8+ cells. RhERAP2 addition also resulted in an increase in CD8+ activated lymphocyte (CD25+HLA-DRII+) and Effector Memory/Terminally differentiated CD8+ T cells ratio. Conclusions: This is the first report providing evidence for the release of ERAP2 in the secretome of immunocompetent cells. Data herein also indicate that exogenous ERAP2-FL exerts its protective function against HIV-1 infection, even in HomoB subjects who do not genetically produce it. Presumably, this defensive extracellular feature is only partially dependent on immune system modulation

Sphingolipid serum profiling in vitamin D deficient and dyslipidemic obese dimorphic adults

Recent studies on Saudi Arabians indicate a prevalence of dyslipidemia and vitamin D deficiency (25(OH)D) in both normal weight and obese subjects. In the present study the sphingolipid pattern was investigated in 23 normolipidemic normal weight (NW), 46 vitamin D deficient dyslipidemic normal weight (-vitDNW) and 60 vitamin D deficient dyslipidemic obese (-vitDO) men and women by HPTLC-primuline profiling and LC-MS analyses. Results indicate higher levels of total ceramide (Cer) and dihydroceramide (dhCers C18\u201322) and lower levels of total sphingomyelins (SMs) and dihydrosphingomyelin (dhSM) not only in -vitDO subjects compared to NW, but also in \u2013vitDNW individuals. A dependency on body mass index (BMI) was observed analyzing specific Cer acyl chains levels. Lower levels of C20 and 24 were observed in men and C24.2 in women, respectively. Furthermore, LC-MS analyses display dimorphic changes in NW, -vitDNW and \u2013vitDO subjects. In conclusion, LC-MS data identify the independency of the axis high Cers, dhCers and SMs from obesity per se. Furthermore, it indicates that long chains Cers levels are specific target of weight gain and that circulating Cer and SM levels are linked to sexual dimorphism status and can contribute to predict obese related co-morbidities in men and women

J Musculoskelet Neuronal Interact

Long-term bed-rest is used to simulate the effect of spaceflight on the human body and test different kinds of countermeasures. The 2nd Berlin BedRest Study (BBR2-2) tested the efficacy of whole-body vibration in addition to high-load resisitance exercise in preventing bone loss during bed-rest. Here we present the protocol of the study and discuss its implementation. Twenty-four male subjects underwent 60-days of six-degree head down tilt bed-rest and were randomised to an inactive control group (CTR), a high-load resistive exercise group (RE) or a high-load resistive exercise with whole-body vibration group (RVE). Subsequent to events in the course of the study (e.g. subject withdrawal), 9 subjects participated in the CTR-group, 7 in the RVE-group and 8 (7 beyond bed-rest day-30) in the RE-group. Fluid intake, urine output and axiallary temperature increased during bed-rest (p or = .17). Body weight changes differed between groups (p < .0001) with decreases in the CTR-group, marginal decreases in the RE-group and the RVE-group displaying significant decreases in body-weight beyond bed-rest day-51 only. In light of events and experiences of the current study, recommendations on various aspects of bed-rest methodology are also discussed

Contribution of Extracellular Matrix and Signal Mechanotransduction to Epithelial Cell Damage in Inflammatory Bowel Disease Patients: A Proteomic Study

This study utilizes 2D-DIGE (difference gel etrophoresis), isotope-coded protein labeling and biochemical assays to characterize protein alteration in ulcerative colitis (UC) and Crohn's disease (CD) in human epithelial cell and mucosal biopsies in inflammatory bowel disease (IBD)-affected patients. The aim of this study is to identify the key molecular signatures involved in epithelial cell structure of IBDs. In non-inflamed UC (QUC) keratins, vimentin, and focal adhesion kinase (7) increased, whereas vinculin and de-tyrosinated \uce\ub1-tubulin decreased; inflammation (IUC) exacerbated molecular changes, being collagen type VI alpha 1 chain (COL6A1), tenascin-C and vimentin increased. In non-inflamed CD (QCD), tenascin C, de-tyrosinated \uce\ub1-tubulin, vinculin, FAK, and Rho-associated protein kinase 1 (ROCK1) decreased while vimentin increased. In inflamed CD (ICD), COL6A1, vimentin and integrin alpha 4 increased. In QUC, cell metabolism is characterized by a decrease of the tricarboxylic acid cycle enzymes and a decrease of short/branched chain specific acyl-CoA dehydrogenase, fatty acid synthase, proliferator-activated receptors alpha, and proliferator-activated receptors gamma. In QCD a metabolic rewiring occurs, as suggested by glycerol-3-phosphate dehydrogenase (GPD2), pyruvate dehydrogenase E1 component subunit beta, NADH dehydrogenase [ubiquinone] iron-sulfur protein 3, and 4-trimethylaminobutyraldehyde dehydrogenase increment, while dihydrolipoyl dehydrogenase decreased. Macroautophagy is activated in QUC and IUC, with increased levels of p62, HSC70, major vault protein, myosin heavy chain 9, whereas it is blunted in QCD and ICD. The differing pattern of extracellular matrix, cytoskeletal derangements, cellular metabolism, and autophagy in UC and CD may contribute to the pathophysiological understanding of these disorders and serve as diagnostic markers in IBD patients

Cancer cell growth and survival as a system-level property sustained by enhanced glycolysis and mitochondrial metabolic remodeling

Systems Biology holds that complex cellular functions are generated as system level properties endowed with robustness, each involving large networks of molecular determinants, generally identified by "omics" analyses. In this paper we describe four basic cancer cell properties that can easily be investigated in vitro: enhanced proliferation, evasion from apoptosis, genomic instability, and inability to undergo oncogene-induced senescence. Focusing our analysis on a K-ras dependent transformation system, we show that enhanced proliferation and evasion from apoptosis are closely linked, and present findings that indicate how a large metabolic remodeling sustains the enhanced growth ability. Network analysis of transcriptional profiling gives the first indication on this remodeling, further supported by biochemical investigations and metabolic flux analysis (MFA). Enhanced glycolysis, down-regulation of TCA cycle, decoupling of glucose and glutamine utilization, with increased reductive carboxylation of glutamine, so to yield a sustained production of growth building blocks and glutathione, are the hallmarks of enhanced proliferation. Low glucose availability specifically induces cell death in K-ras transformed cells, while PKA activation reverts this effect, possibly through at least two mitochondrial targets. The central role of mitochondria in determining the two investigated cancer cell properties is finally discussed. Taken together the findings reported herein indicate that a system level property is sustained by a cascade of interconnected biochemical pathways that behave differently in normal and in transformed cells