Cardiac biopsies reveal differences in transcriptomics between left and right ventricle in patients with or without diagnostic signs of heart failure.

New or mild heart failure (HF) is mainly caused by left ventricular dysfunction. We hypothesised that gene expression differ between the left (LV) and right ventricle (RV) and secondly by type of LV dysfunction. We compared gene expression through myocardial biopsies from LV and RV of patients undergoing elective coronary bypass surgery (CABG). Patients were categorised based on LV ejection fraction (EF), diastolic function and NT-proBNP into pEF (preserved; LVEF ≥ 45%), rEF (reduced; LVEF < 45%) or normal LV function. Principal component analysis of gene expression displayed two clusters corresponding to LV and RV. Up-regulated genes in LV included natriuretic peptides NPPA and NPPB, transcription factors/coactivators STAT4 and VGLL2, ion channel related HCN2 and LRRC38 associated with cardiac muscle contraction, cytoskeleton, and cellular component movement. Patients with pEF phenotype versus normal differed in gene expression predominantly in LV, supporting that diastolic dysfunction and structural changes reflect early LV disease in pEF. DKK2 was overexpressed in LV of HFpEF phenotype, potentially leading to lower expression levels of ß-catenin, α-SMA (smooth muscle actin), and enhanced apoptosis, and could be a possible factor in the development of HFpEF. CXCL14 was down-regulated in both pEF and rEF, and may play a role to promote development of HF.

Expression of mammalian cell entry genes in clinical isolates of M. tuberculosis and the cell entry potential and immunological reactivity of the Rv0590A protein.

Mammalian cell entry (mce) operons play a vital role in cell invasion and survival of M. tuberculosis. Of the mce genes, the function of Rv0590A is still unknown. The present study was performed to investigate the function and immunogenic properties of the protein Rv0590A. Human leukemia monocytic cell line (THP-1) derived macrophages were infected with M. tuberculosis H37Rv at 3, 6, and 24 h of infection. The maximum colony forming units (CFU) were observed at 6 h (p < 0.005), followed by 3 h after infection. M. tuberculosis H37Rv and clinical isolates representative of Delhi/CAS, EAI, Beijing, Haarlem and Euro-American-superlineage were included in the study for expression analysis of mce1A, mce2A, mce3A, mce4A, and Rv0590A genes. Maximum upregulation of all mce genes was observed at 3 h of infection. All the five clinical isolates and H37Rv upregulated Rv0590A at various time points. Macrophage infection with M. tuberculosis H37Rv-overexpressing Rv0590A gene showed higher intracellular CFU as compared to that of wild-type H37Rv. Further, purified Rv0590A protein stimulated the production of TNFα, IFNγ, and IL-10 in macrophages. Thus, Rv0590A was found to be involved in cell invasion and showed good immunological response.

Role of cytokine in malignant T-cell metabolism and subsequent alternation in T-cell tumor microenvironment.

T cells are an important component of adaptive immunity and T-cell-derived lymphomas are very complex due to many functional sub-types and functional elasticity of T-cells. As with other tumors, tissues specific factors are crucial in the development of T-cell lymphomas. In addition to neoplastic cells, T- cell lymphomas consist of a tumor micro-environment composed of normal cells and stroma. Numerous studies established the qualitative and quantitative differences between the tumor microenvironment and normal cell surroundings. Interaction between the various component of the tumor microenvironment is crucial since tumor cells can change the microenvironment and vice versa. In normal T-cell development, T-cells must respond to various stimulants deferentially and during these courses of adaptation. T-cells undergo various metabolic alterations. From the stage of quiescence to attention of fully active form T-cells undergoes various stage in terms of metabolic activity. Predominantly quiescent T-cells have ATP-generating metabolism while during the proliferative stage, their metabolism tilted towards the growth-promoting pathways. In addition to this, a functionally different subset of T-cells requires to activate the different metabolic pathways, and consequently, this regulation of the metabolic pathway control activation and function of T-cells. So, it is obvious that dynamic, and well-regulated metabolic pathways are important for the normal functioning of T-cells and their interaction with the microenvironment. There are various cell signaling mechanisms of metabolism are involved in this regulation and more and more studies have suggested the involvement of additional signaling in the development of the overall metabolic phenotype of T cells. These important signaling mediators include cytokines and hormones. The impact and role of these mediators especially the cytokines on the interplay between T-cell metabolism and the interaction of T-cells with their micro-environments in the context of T-cells lymphomas are discussed in this review article.

Evaluating the efficacy of stool sample on Xpert MTB/RIF Ultra and its comparison with other sample types by meta-analysis for TB diagnostics.

Precise and timely detection of tuberculosis (TB) is crucial to reduce transmission. This study aims to assess the accuracy of Xpert MTB/RIF Ultra on stool samples and systematically review the performance of Xpert MTB/RIF Ultra with different sample types by meta-analysis. Stool samples of smear-negative pulmonary TB (PTB), cervical lymph node TB, and abdominal TB patients were tested on the Xpert MTB/RIF Ultra system. Meta-analysis was performed on a set of 44 studies. Data were grouped by sample type, and the pooled sensitivity and specificity of Xpert MTB/RIF Ultra were calculated. The sensitivity of Xpert MTB/RIF Ultra with stool samples was 100% for smear-negative PTB, 27.27% for cervical lymph node TB, and 50% for abdominal TB patients, with 100% specificity for all included TB groups. The summary estimate for all PTB samples showed 84.2% sensitivity and 94.5% specificity, and EPTB samples showed 88.6% sensitivity and 96.4% specificity. Among all sample types included in our meta-analysis, urine showed the best performance for EPTB diagnosis. This pilot study supports the use of stool as an alternative non-invasive sample on Xpert MTB/RIF Ultra for rapid testing, suitable for both PTB and EPTB diagnosis.

Metabolomic Profile in HFpEF vs HFrEF Patients.

BACKGROUND: Heart failure with preserved ejection fraction (HFpEF) and HF with reduced ejection fraction (HFrEF) are associated with metabolic derangements, which may have different pathophysiological implications. METHODS AND RESULTS: In new-onset HFpEF (EF of ≥50%, n = 46) and HFrEF (EF of <40%, n = 75) patients, 109 endogenous plasma metabolites including amino acids, phospholipids and acylcarnitines were assessed using targeted metabolomics. Differentially altered metabolites and associations with clinical characteristics were explored. Patients with HFpEF were older, more often female with hypertension, atrial fibrillation, and diabetes compared with patients with HFrEF. Patients with HFpEF displayed higher levels of hydroxyproline and symmetric dimethyl arginine, alanine, cystine, and kynurenine reflecting fibrosis, inflammation and oxidative stress. Serine, cGMP, cAMP, l-carnitine, lysophophatidylcholine (18:2), lactate, and arginine were lower compared with patients with HFrEF. In patients with HFpEF with diabetes, kynurenine was higher (P = .014) and arginine lower (P = .014) vs patients with no diabetes, but did not differ with diabetes status in HFrEF. Decreasing kynurenine was associated with higher eGFR only in HFpEF (Pinteraction = .020). CONCLUSIONS: Patients with new-onset HFpEF compared with patients with new-onset HFrEF display a different metabolic profile associated with comorbidities, such as diabetes and kidney dysfunction. HFpEF is associated with indices of increased inflammation and oxidative stress, impaired lipid metabolism, increased collagen synthesis, and downregulated nitric oxide signaling. Together, these findings suggest a more predominant systemic microvascular endothelial dysfunction and inflammation linked to increased fibrosis in HFpEF compared with HFrEF. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov NCT03671122 https://clinicaltrials.gov.

Intercalating the Role of MicroRNAs in Cancer: As Enemy or Protector.

OBJECTIVE: The transformation in cells at genetic levels stimulatesthe proliferation of cancer. The current review highlights the role of miRNA in management of cancer by altering processes of body at cellular levels. METHODS: A deep research on the literature available till date for miRNA in cancer was conducted using various medical sites like PubMed, MEDLINE from internet and data was collected. The articles were majorly preferred in English language. RESULTS: The development of normal cells into cancerous cells is a multivalent procedure highlighting numerous responsible factors. During the progression of cancer, the role of oncogene and tumor suppressor genes outshines at different levels of tumorogenesis. Metastasis poses highest threat in cancer progression and fabricates obstacles to clinicians and researchers in preventing formation of tumor on secondary sites. The mesenchymal-epithelial transition (MET) and epithelial mesenchymal transition (EMT) induce dissemination and ultimately progression of cancer. CONCLUSION: A comprehensive knowledge of the altered genes and the mechanism by which they induce formation of tumor is essential as they contribute in proliferating cancer at various stages, aggravating clinical symptoms. Hence miRNAs can be efficiently employed as an emerging treatment therapy for cancer.

A Multi-Omics Approach to Liver Diseases: Integration of Single Nuclei Transcriptomics with Proteomics and HiCap Bulk Data in Human Liver.

The liver is the largest solid organ and a primary metabolic hub. In recent years, intact cell nuclei were used to perform single-nuclei RNA-seq (snRNA-seq) for tissues difficult to dissociate and for flash-frozen archived tissue samples to discover unknown and rare cell subpopulations. In this study, we performed snRNA-seq of a liver sample to identify subpopulations of cells based on nuclear transcriptomics. In 4282 single nuclei, we detected, on average, 1377 active genes and we identified seven major cell types. We integrated data from 94,286 distal interactions (p < 0.05) for 7682 promoters from a targeted chromosome conformation capture technique (HiCap) and mass spectrometry proteomics for the same liver sample. We observed a reasonable correlation between proteomics and in silico bulk snRNA-seq (r = 0.47) using tissue-independent gene-specific protein abundancy estimation factors. We specifically looked at genes of medical importance. The DPYD gene is involved in the pharmacogenetics of fluoropyrimidine toxicity and some of its variants are analyzed for clinical purposes. We identified a new putative polymorphic regulatory element, which may contribute to variation in toxicity. Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and we investigated all known risk genes. We identified a complex regulatory landscape for the SLC2A2 gene with 16 candidate enhancers. Three of them harbor somatic motif breaking and other mutations in HCC in the Pan Cancer Analysis of Whole Genomes dataset and are candidates to contribute to malignancy. Our results highlight the potential of a multi-omics approach in the study of human diseases.

rs953413 Regulates Polyunsaturated Fatty Acid Metabolism by Modulating ELOVL2 Expression.

Long-chain polyunsaturated fatty acids (LC-PUFAs) influence human health in several areas, including cardiovascular disease, diabetes, fatty liver disease, and cancer. ELOVL2 encodes one of the key enzymes in the in vivo synthesis of LC-PUFAs from their precursors. Variants near ELOVL2 have repeatedly been associated with levels of LC-PUFA-derived metabolites in genome-wide association studies (GWAS), but the mechanisms behind these observations remain poorly defined. In this study, we found that rs953413, located in the first intron of ELOVL2, lies within a functional FOXA and HNF4α cooperative binding site. The G allele of rs953413 increases binding of FOXA1/FOXA2 and HNF4α to an evolutionarily conserved enhancer element, conferring allele-specific upregulation of the rs953413-associated gene ELOVL2. The expression of ELOVL2 was significantly downregulated by both FOXA1 and HNF4α knockdown and CRISPR/Cas9-mediated direct mutation to the enhancer element. Our results suggest that rs953413 regulates LC-PUFAs metabolism by altering ELOVL2 expression through FOXA1/FOXA2 and HNF4α cooperation.

Transcriptomics of cardiac biopsies reveals differences in patients with or without diagnostic parameters for heart failure with preserved ejection fraction.

Heart failure affects 2-3% of adult Western population. Prevalence of heart failure with preserved left ventricular (LV) ejection fraction (HFpEF) increases. Studies suggest HFpEF patients to have altered myocardial structure and functional changes such as incomplete relaxation and increased cardiac stiffness. We hypothesised that patients undergoing elective coronary bypass surgery (CABG) with HFpEF characteristics would show distinctive gene expression compared to patients with normal LV physiology. Myocardial biopsies for mRNA expression analysis were obtained from sixteen patients with LV ejection fraction ≥45%. Five out of 16 patients (31%) had echocardiographic characteristics and increased NTproBNP levels indicative of HFpEF and this group was used as HFpEF proxy, while 11 patients had Normal LV physiology. Utilising principal component analysis, the gene expression data clustered into two groups, corresponding to HFpEF proxy and Normal physiology, and 743 differentially expressed genes were identified. The associated top biological functions were cardiac muscle contraction, oxidative phosphorylation, cellular remodelling and matrix organisation. Our results also indicate that upstream regulatory events, including inhibition of transcription factors STAT4, SRF and TP53, and activation of transcription repressors HEY2 and KDM5A, could provide explanatory mechanisms to observed gene expression differences and ultimately cardiac dysfunction in the HFpEF proxy group.

MicroRNAs as Key Regulators of Ovarian Cancers.

The tumor microenvironment can be realistically viewed as an active battle ground between the host immune system and the growing tumor cells. This reactive space surrounding the tumor possesses several possibilities and facilitates the progression of a tumor from a neoplastic stage to that of metastasis. The contemporary approach of understanding the cancer biology from a "within the cell" perspective has been largely challenged with complex and intricate "outside the cell" events. Thus understanding the biology of the tumor microenvironment has been of scientific and clinical interest. Small non-coding microRNAs with a pleotropic and wide range of cellular gene targets can be reasonably hypothesized to regulate the events of carcinogenesis and progression. MicroRNAs have been investigated in different cancer models, and evidence of their involvement in the regulation of the tumor microenvironment has been of much interest. In particular, a major interest has been exploring the role of the tumor microenvironment in regulating the interaction of cancer cells with surrounding stromal components and the effect of such interactions on the cancer cells. Fine-tuned regulation by these microRNAs extends our contemporary understanding of these small biomolecules in epigenetic regulations. This review focuses on microRNAs that are dysregulated in ovarian carcinomas, their effect on the components of the tumor microenvironment, and the correlation of their heterogeneous expression profiles with disease severity and prognosis in patients. In addition, this paper also discusses the differential expression of exosomal microRNAs that are known to link the cancer cell with its microenvironment, facilitating the development of an improved prognostic/diagnostic marker and effective therapeutic regime.

Liver macrophages regulate systemic metabolism through non-inflammatory factors.

Liver macrophages (LMs) have been proposed to contribute to metabolic disease through secretion of inflammatory cytokines. However, anti-inflammatory drugs lead to only modest improvements in systemic metabolism. Here we show that LMs do not undergo a proinflammatory phenotypic switch in obesity-induced insulin resistance in flies, mice and humans. Instead, we find that LMs produce non-inflammatory factors, such as insulin-like growth factor-binding protein 7 (IGFBP7), that directly regulate liver metabolism. IGFBP7 binds to the insulin receptor and induces lipogenesis and gluconeogenesis via activation of extracellular-signal-regulated kinase (ERK) signalling. We further show that IGFBP7 is subject to RNA editing at a higher frequency in insulin-resistant than in insulin-sensitive obese patients (90% versus 30%, respectively), resulting in an IGFBP7 isoform with potentially higher capacity to bind to the insulin receptor. Our study demonstrates that LMs can contribute to insulin resistance independently of their inflammatory status and indicates that non-inflammatory factors produced by macrophages might represent new drug targets for the treatment of metabolic diseases.

Cognate sensor kinase-independent activation of Mycobacterium tuberculosis response regulator DevR (DosR) by acetyl phosphate: implications in anti-mycobacterial drug design.

The DevRS/DosT two-component system is essential for mycobacterial survival under hypoxia, a prevailing stress within granulomas. DevR (also known as DosR) is activated by an inducing stimulus, such as hypoxia, through conventional phosphorylation by its cognate sensor kinases, DevS (also known as DosS) and DosT. Here, we show that the DevR regulon is activated by acetyl phosphate under 'non-inducing' aerobic conditions when Mycobacterium tuberculosis devS and dosT double deletion strain is cultured on acetate. Overexpression of phosphotransacetylase caused a perturbation of the acetate kinase-phosphotransacetylase pathway, a decrease in the concentration of acetyl phosphate and dampened the aerobic induction response in acetate-grown bacteria. The operation of two pathways of DevR activation, one through sensor kinases and the other by acetyl phosphate, was established by an analysis of wild-type DevS and phosphorylation-defective DevSH395Q mutant strains under conditions partially mimicking a granulomatous-like environment of acetate and hypoxia. Our findings reveal that DevR can be phosphorylated in vivo by acetyl phosphate. Importantly, we demonstrate that acetyl phosphate-dependent phosphorylation can occur in the absence of DevR's cognate kinases. Based on our findings, we conclude that anti-mycobacterial therapy should be targeted to DevR itself and not to DevS/DosT kinases.

Alterations in the hepatic transcriptional landscape after RNAi mediated ApoB silencing in cynomolgus monkeys.

The greater genomic conservation between humans and non-human primates (NHP) enables target validation studies for developing of therapeutic strategies for human diseases. Together with predicting activity and potential adverse clinical signs, the inclusion of NHP testing bequeaths to efficacy models for dose titration and pharmacodynamic effects. We have used lipid nanoparticle encapsulated siRNA to silence ApoB in the liver and assessed the phenotypic effects on serum lipids with various levels of hepatic ApoB mRNA knockdown in healthy lean cynomolgus monkeys. ApoB siRNA dosed animals demonstrated significant reductions of hepatic ApoB mRNA and serum APOB protein, with a substantial lowering of plasma lipid levels without obvious signs of toxicity. Microarray based assessment of ApoB siRNA mediated effects revealed a number of differentially expressed genes which mapped onto biological pathways and processes related to lipid and cholesterol metabolism. Furthermore, we identified potential targets and cellular effects that could be studied for therapeutic benchmarking of APOB mediated effects. The network of ApoB regulated genes should be of significance for the understanding and development of novel hypercholesterolemia therapies.

Combination of chemical genetics and phosphoproteomics for kinase signaling analysis enables confident identification of cellular downstream targets.

Delineation of phosphorylation-based signaling networks requires reliable data about the underlying cellular kinase-substrate interactions. We report a chemical genetics and quantitative phosphoproteomics approach that encompasses cellular kinase activation in combination with comparative replicate mass spectrometry analyses of cells expressing either inhibitor-sensitive or resistant kinase variant. We applied this workflow to Plk1 (Polo-like kinase 1) in mitotic cells and induced cellular Plk1 activity by wash-out of the bulky kinase inhibitor 3-MB-PP1, which targets a mutant kinase version with an enlarged catalytic pocket while not interfering with wild-type Plk1. We quantified more than 20,000 distinct phosphorylation sites by SILAC, approximately half of which were measured in at least two independent experiments in cells expressing mutant and wild-type Plk1. Based on replicate phosphorylation site quantifications in both mutant and wild-type Plk1 cells, our chemical genetic proteomics concept enabled stringent comparative statistics by significance analysis of microarrays, which unveiled more than 350 cellular downstream targets of Plk1 validated by full concordance of both statistical and experimental data. Our data point to hitherto poorly characterized aspects in Plk1-controlled mitotic progression and provide a largely extended resource for functional studies. We anticipate the described strategies to be of general utility for systematic and confident identification of cellular protein kinase substrates.

Quantitative analysis of kinase-proximal signaling in lipopolysaccharide-induced innate immune response.

The innate immune system senses invariant microbial components via toll-like receptors (TLRs) to elicit a host defense program against invading pathogens. Lipopolysaccharide (LPS), a constituent of Gram-negative bacteria, is recognized by TLR4 and triggers protein kinase signaling to orchestrate immune responses such as inflammatory cytokine production. To analyze kinase-proximal signaling in murine macrophages, we performed prefractionation experiments with immobilized kinase inhibitors to enrich for protein kinases and their interaction partners. In conjunction with SILAC-based quantitative mass spectrometry and phosphopeptide enrichment, we recorded five time point profiles for more than 850 distinct phosphorylation events on protein kinases and copurifying factors. More than 15% exhibited significant changes and many of those mapped to LPS-regulated kinase networks. We identified many unreported TLR signaling events including LPS-triggered phosphorylations of Akt substrates, which point to previously unknown molecular mechanisms in innate immune response. We further detected extensive phosphoregulation of TANK-binding kinase 1, inhibitor of nuclear factor-kappaB kinase epsilon and their associating scaffolding factors, and none of these events were known despite the key roles of these proteins in LPS signaling. Thus, our data expands previous knowledge for functional analyses of innate immune response.

Quantitative phosphoproteomics reveals widespread full phosphorylation site occupancy during mitosis.

Eukaryotic cells replicate by a complex series of evolutionarily conserved events that are tightly regulated at defined stages of the cell division cycle. Progression through this cycle involves a large number of dedicated protein complexes and signaling pathways, and deregulation of this process is implicated in tumorigenesis. We applied high-resolution mass spectrometry-based proteomics to investigate the proteome and phosphoproteome of the human cell cycle on a global scale and quantified 6027 proteins and 20,443 unique phosphorylation sites and their dynamics. Co-regulated proteins and phosphorylation sites were grouped according to their cell cycle kinetics and compared to publicly available messenger RNA microarray data. Most detected phosphorylation sites and more than 20% of all quantified proteins showed substantial regulation, mainly in mitotic cells. Kinase-motif analysis revealed global activation during S phase of the DNA damage response network, which was mediated by phosphorylation by ATM or ATR or DNA-dependent protein kinases. We determined site-specific stoichiometry of more than 5000 sites and found that most of the up-regulated sites phosphorylated by cyclin-dependent kinase 1 (CDK1) or CDK2 were almost fully phosphorylated in mitotic cells. In particular, nuclear proteins and proteins involved in regulating metabolic processes have high phosphorylation site occupancy in mitosis. This suggests that these proteins may be inactivated by phosphorylation in mitotic cells.

Lysine acetylation targets protein complexes and co-regulates major cellular functions.

Lysine acetylation is a reversible posttranslational modification of proteins and plays a key role in regulating gene expression. Technological limitations have so far prevented a global analysis of lysine acetylation's cellular roles. We used high-resolution mass spectrometry to identify 3600 lysine acetylation sites on 1750 proteins and quantified acetylation changes in response to the deacetylase inhibitors suberoylanilide hydroxamic acid and MS-275. Lysine acetylation preferentially targets large macromolecular complexes involved in diverse cellular processes, such as chromatin remodeling, cell cycle, splicing, nuclear transport, and actin nucleation. Acetylation impaired phosphorylation-dependent interactions of 14-3-3 and regulated the yeast cyclin-dependent kinase Cdc28. Our data demonstrate that the regulatory scope of lysine acetylation is broad and comparable with that of other major posttranslational modifications.

Comparative proteomic phenotyping of cell lines and primary cells to assess preservation of cell type-specific functions.

Biological experiments are most often performed with immortalized cell lines because they are readily available and can be expanded without limitation. However, cell lines may differ from the in vivo situation in important aspects. Here we introduce a straightforward methodology to compare cell lines to their cognate primary cells and to derive a comparative functional phenotype. We used SILAC (stable isotope labeling by amino acids in cell culture) for quantitative, mass spectrometry-based comparison of the hepatoma cell line Hepa1-6 with primary hepatocytes. The resulting quantitative proteome of 4,063 proteins had an asymmetric distribution, with many proteins down-regulated in the cell line. Bioinformatic analysis of the quantitative proteomics phenotypes revealed that Hepa1-6 cells were deficient in mitochondria, reflecting re-arrangement of metabolic pathways, drastically up-regulate cell cycle-associated functions and largely shut down drug metabolizing enzymes characteristic for the liver. This quantitative knowledge of changes provides an important basis to adapt cell lines to more closely resemble physiological conditions.

Combined use of RNAi and quantitative proteomics to study gene function in Drosophila.

RNA interference is a powerful way to study gene function and is frequently combined with microarray analysis. Here we introduce a similar technology at the protein level by simultaneously applying Stable Isotope Labeling by Amino acids in Cell culture (SILAC) and RNA interference (RNAi) to Drosophila SL2 cells. After knockdown of ISWI, an ATP-hydrolyzing motor of different chromatin remodeling complexes, we obtained a quantitative proteome of more than 4,000 proteins. ISWI itself was reduced 10-fold as quantified by SILAC. Several hundred proteins were significantly regulated and clustered into distinct functional categories. Acf-1, a direct interaction partner of ISWI, is severely depleted at the protein, but not the transcript, level; this most likely results from reduced protein stability. We found little overall correlation between changes in the transcriptome and proteome with many protein changes unaccompanied by message changes. However, correlation was high for those mRNAs that changed significantly by microarray.

Integrated analysis of the cerebrospinal fluid peptidome and proteome.

Cerebrospinal fluid (CSF) is the only body fluid in direct contact with the brain and thus is a potential source of biomarkers. Furthermore, CSF serves as a medium of endocrine signaling and contains a multitude of regulatory peptides. A combined study of the peptidome and proteome of CSF or any other body fluid has not been reported previously. We report confident identification in CSF of 563 peptide products derived from 91 precursor proteins as well as a high confidence CSF proteome of 798 proteins. For the CSF peptidome, we use high accuracy mass spectrometry (MS) for MS and MS/MS modes, allowing unambiguous identification of neuropeptides. Combination of the peptidome and proteome data suggests that enzymatic processing of membrane proteins causes release of their extracellular parts into CSF. The CSF proteome has only partial overlap with the plasma proteome, thus it is produced locally rather than deriving from plasma. Our work offers insights into CSF composition and origin.