Dataset Information

Serial-Omics of Equine Urine - Serial-omics characterization of equine urine

ABSTRACT: Horse urine is easily collected and contains molecules readily measurable using mass spectrometry that can be used as biomarkers representative of health, disease or drug tampering. This study aimed at analyzing microliter levels of horse urine to purify, identify and quantify proteins, polar metabolites and non-polar lipids. Urine from a healthy 12 year old quarter horse mare on a diet of grass hay and vitamin/mineral supplements with limited pasture access was collected for serial-omics characterization. The urine was treated with methyl tert-butyl ether (MTBE) and methanol to partition into three distinct layers for protein, non-polar lipid and polar metabolite content from a single liquid-liquid extraction and was repeated two times. Each layer was analyzed by high performance liquid chromatography – high resolution tandem mass spectrometry (LC-MS/MS) to obtain protein sequence and relative protein levels as well as identify and quantify small polar metabolites and lipids. The results show 46 urine proteins, many related to normal kidney function, structural and circulatory proteins as well as 474 small polar metabolites but only 10 lipid molecules. Metabolites were mostly related to urea cycle and ammonia recycling as well as amino acid related pathways, plant diet specific molecules, etc. The few lipids represented triglycerides and phospholipids. These data show a complete mass spectrometry based –omics characterization of equine urine from a single 333 uL mid-stream urine aliquot. The data can be used as a baseline for healthy urine composition and analyses can be used to monitor disease progression, health status, monitor drug use, etc.

INSTRUMENT(S): LTQ Orbitrap Elite, Q TRAP, Q Exactive

ORGANISM(S): Equus Caballus (horse)

TISSUE(S): Urine

SUBMITTER: John Asara

LAB HEAD: John M Asara

PROVIDER: PXD007646 | Pride | 2018-07-23

REPOSITORIES: Pride

ACCESS DATA

Publications

Serial-omics characterization of equine urine.

Yuan Min M Breitkopf Susanne B SB Asara John M JM

PloS one 20171013 10

Horse urine is easily collected and contains molecules readily measurable using mass spectrometry that can be used as biomarkers representative of health, disease or drug tampering. This study aimed at analyzing microliter levels of horse urine to purify, identify and quantify proteins, polar metabolites and non-polar lipids. Urine from a healthy 12 year old quarter horse mare on a diet of grass hay and vitamin/mineral supplements with limited pasture access was collected for serial-omics charac ...[more]

PMID: 29028822

Similar Datasets

Project description:To cause disease, Salmonella enterica serovar Typhimurium requires two type-III secretion systems, encoded on Salmonella Pathogenicity Islands 1 and 2 (SPI-1 and -2). These secretion systems serve to deliver virulence proteins, termed effectors, into the host cell cytosol. While the importance of these effector proteins to promote colonization and replication within the host has been established, the specific roles of individual secreted effectors in the disease process are not well understood. In this study, we used an in vivo gallbladder epithelial cell infection model to study the function of the SPI-2-encoded effector, SseL. Deletion of the sseL gene resulted in bacterial filamentation and elongation and unusual localization of Salmonella within infected epithelial cells. Infection with the ?sseL strain also caused dramatic changes in lipid metabolism and led to massive accumulation of lipid droplets in infected cells. Some of these changes were investigated through metabolomics of gallbladder tissue. This phenotype was directly attributed to the deubiquitinase activity of SseL, as a Salmonella strain carrying a single point mutation in the catalytic cysteine resulted in the same phenotype as the deletion mutant. Excessive buildup of lipids due to the absence of a functional sseL gene was also observed in S. Typhimurium-infected livers. These results demonstrate that SseL alters host lipid metabolism in infected epithelial cells by modifying ubiquitination patterns of cellular targets. Female C57BL/6 mice were infected with the indicated strain of Salmonella enterica serovar Typhimurium by oral gavage. Four gallbladders were collected and pooled per sample group and metabolites extracted using a mixture of methanol and chloroform. Extracts were infused into a 12-T Apex-Qe hybrid quadrupole-FT-ICR mass spectrometer equipped with an Apollo II electrospray ionization source, a quadrupole mass filter and a hexapole collision cell. Raw mass spectrometry data were processed as described elsewhere (Han et al. 2008. Metabolomics. 4:128-140). To identify differences in metabolite composition between different groups of samples, we filtered the list of masses for metabolites which were present on one set of samples but not the other. Additionally, we calculated the ratios between averaged intensities of metabolites from each group of mice. To assign possible metabolite identities, monoisotopic neutral masses of interest were queried against MassTrix (http://masstrix.org). Masses were searched against the Mus musculus database within a mass error of 3 ppm.

Project description:Lower urinary tract symptoms (LUTS) relate to storage and/or voiding disturbances which are common among aging men. Disease etiology is largely unknown, and diagnosis is currently based on subjective symptom scores. While mainstay treatments can be ineffective and bothersome, biomarker discovery efforts may facilitate objective diagnostic criteria for personalized medicine and new potential druggable pathways. Since inflammation is one of its plausible etiologies, the goal of this project is to explore urinary metabolite and protein biomarkers as a non-invasive method to diagnose of LUTS which is induced by inflammation. Herein, a bacterial-induced prostatic inflammation model was established with C57BL/6J mice infected with uropathogenic E. coli 1677. Mouse urine samples were collected from control and treatment mice. Mass Spectrometry-based multi-omics analysis was employed here to discover urinary protein and metabolite-based biomarkers specific to prostatic inflammation-induced LUTS. We investigated the use of isobaric dimethylated leucine (DiLeu) labeling for metabolite identification and quantification for the mouse urine samples, and because of the use of DiLeu labeling, we combined metabolomics and proteomics on the same LC-MS platform. Overall, a total of 143 amine-containing metabolites and 1058 urinary proteins were identified and quantified, among them, 14 metabolites and 168 proteins were significantly changed by the inflammation treatment. Five metabolic pathways and four inflammatory-related biological processes were potentially disrupted. After compare with LUTS patient and other previously report, metabolite creatine and protein Polymeric Ig receptor presents especially attractive combined biomarker for prostatic inflammation induced LUTS and could be further used as the LUTS patient stratification. Overall, this study found urine metabolomics and proteomics to be an informative and noninvasive method for candidate biomarkers determination and etiologies classification of LUTS induced by inflammation.

Project description:Low molecular weight (LMW) peptides and proteins (LMWPs, < 30 kDa) in human plasma, serving as potential biomarkers or drug targets, are endowed with a wealth of biological and clinical research values. However, the extraction of LMW species is retarded by high-abundance proteins (HAPs) and therein large dynamic concentration ranges of proteins in plasma. Here we present an optimized MeOH/MTBE-based organic solvent precipitation approach that combining the efficient protein precipitation and effective delipidation (referred here as MeOH/MTBE) to enrich LMWPs from human plasma. The Tricine-SDS-PAGE analysis showed that MeOH/MTBE can not only maximally deplete HAPs but also effectively extract LMWPs from human plasma. By coupling with Top-Down Proteomics (TDP) strategy, MeOH/MTBE enabled the identification of 725 peptides derived from 158 proteins on average from human plasma, which should represent the largest number to our best knowledge with regard to TDP-based single LC-MS/MS acquisition. When combined with Bottom-Up Proteomics (BUP) strategy, MeOH/MTBE allowed for the identification of 376 proteins from human plasma. Notably, 155 out of 376 identified proteins (41.2%) were found to be with a molecular weight under 30 kDa, which further proved the efficient enrichment of LMWPs from human plasma by MeOH/MTBE approach. Moreover, concentrations of the identified proteins were found to span 5-6 orders of magnitude according to the PaxDb dataset. Additionally, many low abundant peptides including 25 neuropeptides and 19 smORF-encoded polypeptides (SEPs) derived from long noncoding RNAs (lncRNAs) were detected. Taken together, the MeOH/MTBE approach described here represents a simple, rapid and reproducible methodology for efficient and sensitive analysis of LMWPs and low abundant proteins from human plasma, which will greatly facilitate the selective identification and quantification of LMWPs from human plasma and accelerate the discovery rate of potential biomarkers and drug targets.

Project description:The Mongolian horse has the characteristics of cold resistance, rough feeding resistance, good endurance, strong disease resistance and strong gregariousness. The normal production of sperm in male mammals is the basis of reproduction, and spermatogenesis includes a mitosis, meiosis twice and metamorphosis of sperm cells. The MEI1 gene is involved in the meiosis cycle and is required for normal meiosis chromosome association. Previous studies have shown that alternative splicing of MEI1 gene can promote spermatogenesis in Mongolian horses. In this study, the expression of MEI1 gene was determined by immunofluorescence in the third generation Mongolian horse testicular sertoli cells. Two overexpressed lentiviral vectors with MXE and SE events of MEI1 gene were constructed and successfully infected into sertoli cells. CCK-8 was used to detect the proliferation and activity of infected cells, transcripomics and metabolomics were used to screen and annotate differential genes and differential metabolites in the two sertoli cells with MXE and SE events. After 72 h of virus infection, 193 differentially expressed genes and 11360 differentially expressed metabolites were screened, 109 up-regulated genes including MEI1 and 84 down-regulated genes, 7494 up-regulated metabolites and 3866 down-regulated metabolites. Differential genes and differential metabolites were mainly enriched in several pathways related to spermatogenesis. Differential genes such as IL31RA, ATP2B3, CASQ2 were highly expressed in SE events, IL11, PRLR, CCR7 were highly expressed in MXE events. Metabolites such as folic acid and spermine were highly expressed in SE events, citric acid and glutathione were highly expressed in MXE events. This suggested that both MXE and SE events of MEI1 gene can promote the activity of spermatogenesis signaling pathway, and the gene influences spermatogenesis by regulating the expression of metabolites. In summary, MXE and SE events of MEI1 gene can promote the expression of spermatogen-related genes and metabolites, and genes affect spermatogenesis by regulating the expression of metabolites. The results of this study laid a theoretical foundation for exploring the regulatory mechanism of different alternative splicing events on spermatogenesis in Mongolian horse.

Dataset Information

Serial-Omics of Equine Urine - Serial-omics characterization of equine urine

Publications

Serial-omics characterization of equine urine.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure