Prideapplication/xmlftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_allfractions__FDR.xlsxftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_allfraction_allspecies__FDR.xlsxftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_all_fractions_all_species__FDR.xlsxftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/Chondroitinase_FDR.xlsxftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/Chondroitinase_100_FDR.xlsxftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_allfraction__FDR.xlsxftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/GndHCl_FDR.xlsxftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/Chondroitinase_100.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_002.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_015.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_018.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_006.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_005.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_003.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/Chondroitinase_100.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_008.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_009.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_018.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_015.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_001.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_012.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_018.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_005.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_013.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_014.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/Chondroitinase.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_007.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_002.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_010.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_003.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_015.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_016.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_018.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_004.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/GndHCl.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_001.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_002.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_015.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_010.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_002.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_001.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_017.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/Chondroitinase.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_017.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_016.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_009.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_008.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_014.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch1_fraction_001.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_003.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_016_3rd.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_006.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_009.wiffftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_011.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD004922/batch2_fraction_009.wiff.scanftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/09/PXD00492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SantaMass SpectrometryShotgun proteomicsNot availableextracellular matrix (ecm)nucleus pulpous (np)Intervertebral disc (ivd)Intervertebral disc (ivd); extracellular matrix (ecm); nucleus pulpous (np); itraqitraqhttp://www.ebi.ac.uk/pride/archive/projects/PXD004922Nucleus PulposusProtein was extracted for 24 h at 4 ºC in a buffer containing guanidine hydrochloride 4M, Ethylenediamine tetraacetic acid (EDTA) 10mM (Sigma), sodium acetate 50 mM (Merck), dithiothreitol (DTT) 10mg/mL (Sigma), phenylmethylsulfonyl fluoride (PMSF) 1mM (Sigma) and cOmplete, Mini, EDTA-free Protease Inhibitor Cocktail inhibitor 20μL/mL (Sigma). After centrifugation, supernatants were precipitated in 9 volumes of ethanol for 2h at -20 ºC. Precipitated proteins were centrifuged at 20,000xg for 20 min and ressuspended in 0.5 M of Triethylammonium bicarbonate (TEAB) buffer (Sigma) pH 8.5, vortexed and sonicated. Protein content was quantified using 2D Quant Kit (GE Healthcare). The volume corresponding to 100 μg of protein from each sample was concentrated (Concentrator Plus, Eppendorf) at 60C and resuspended in TEAB 0.5M to a final volume of 45 μL. A pool of all samples was performed in order to reach a total of 200 μg and two other pools (one of foetus and other of old samples), were constituted with 100 μg of protein in each. Pooled samples were also concentrated and adjusted to 45 μL with TEAB 0.5 M and 8 μL of the reducing agent tris(2-carboxyethyl)phosphine (TCEP) 50 mM (Sigma) were added. Samples were sonicated (Vibra Cell 750 watt, Sonics) in a cuphorn for 1 min at 20% amplitude. 4 μL of 200 mM methyl methanethiosulfonate (MMTS) (Sigma) were added for 10 min at room temperature. Following addition of TEAB 0.5M to a final volume of 95 µL, protein digestion was performed by adding 5 μL of trypsin (Roche) at 0.5 μg/μL. Next, samples were incubated for 16h at 37 C. After digestion, 2 μL of formic acid (FA) 100% were added and they were dried for 1 h at 60C. Samples were solubilized in 75 μL of 70% isopropanol and 30% TEAB 1M and sonicated in a cuphorn for 10 min at 20 % amplitude, with pulses of 1 sec on and 1 second off. The digested peptides were then labelled with the iTRAQ (8-plex) tags according to manufacturer´s instructions (Sciex). Two iTRAQ 8-plex batches were used where the pool of all samples was repeated in both iTRAQ batches to serve as normalization channel. The labelled samples of each batch were combined into one sample mixture. About 650 µg of sample were solubilized in 2% ACN in 72 mM TEAB and analyzed by 2D LC-MS/MS, with a high pH reverse phase chromatography as first dimension and then analysed by LC-MS. The first dimension chromatography was performed in UltimateTM3000 LC (LC Packings, Dionex) with two online Aeris 3.6 µm XB-C18 columns (15 cm x 2.10 mm) (Phenomenex), using 72 mM TEAB pH 8.5 as mobile phase A and 72 mM TEAB in ACN pH 8.5 as mobile phase B (10 minutes with 2% mobile phase B followed by a linear gradient until 45% mobile phase B during 60 minutes, then followed by column wash and reequilibration). Throughout the chromatographic separation, 75 fractions were collected and then joined into 18 samples, evaporated and prepared for LC-MS/MS as described above. For the LC-MS analysis the peptides were resolved by liquid chromatography (nanoLC Ultra 2D, Eksigent) on a ChromXP™ C18AR reverse phase column (300 μm ID × 15 cm length, 3 μm particles, 120 Å pore size, Eksigent) at 5 μL/min. Peptides were eluted into the mass spectrometer with an ACN gradient in 0.1% FA (2% to 30% ACN, in a linear gradient for 40 min, followed by a column wash and equilibration step), using an electrospray ionization source (DuoSpray™ Source, AB Sciex). The mass spectrometer (Triple TOF™ 5600 System; AB Sciex) was programmed for scanning full spectra (350-1250 m/z) for 250 ms, followed by up to 30 MS/MS scans (100–1500 m/z for 100 ms each). Candidate ions with a charge state between +2 and +5 and a minimum threshold of 70 counts per second were isolated for fragmentation and two MS/MS spectra were collected before adding those ions to the exclusion list for 15 seconds (mass spectrometer operated by Analyst TF 1.6, AB Sciex). Specific iTRAQ rolling collision energy was used. For the preliminary test aiming to understand if the use of chondroitinase in sample processing was advantageous, the procedure adopted was very similar to the one described above, varying only in the usage of chondroitinase and also the analysis of a fraction (“<100”) filtered with a 100kDa cut-off. The LC-MS/MS startegy for protein identification was as described above with sight modifications (for iTRAQ was not used). The chromatographic run consisted on a linear gradient from 2% to 30% ACN in 0.1% FA for 80 minutes followed by a column wash and equilibration step. The mass spectrometer was programmed for scanning full spectra (350-1250 m/z) for 250 ms, followed by up to 20 MS/MS scans (100–1500 m/z for 75 ms each). Candidate ions with a charge state between +2 and +5 and a minimum threshold of 70 counts per second were isolated for fragmentation and one MS/MS spectra were collected before adding those ions to the exclusion list for 15 seconds.PrideiTRAQNo PTMs are included in the datasetFollowing LC-MS analysis, the peptide and protein identification and protein relative quantification was performed with ProteinPilot™ software (v4.5, AB Sciex). The search parameters used were the following: SwissProt database (release 2012_06), against Bos Taurus or all species, using MMTS alkylated cysteines and iTRAQ labelled peptides as fixed modifications. An independent False Discovery Rate (FDR) analysis using the target-decoy approach provided with ProteinPilot software was used to assess the quality of the identifications and positive identifications were considered when identified proteins reached a 5% local FDR with at least one peptide that reached a 5% local FDR. Preliminary tests - Following LC-MS analysis, the peptide and protein identification and protein relative quantification was performed with ProteinPilot™ software (v4.5, AB Sciex). The search parameters used were the following: SwissProt database (release 2012_06), against Bos Taurus, using MMTS alkylated cysteines as fixed modifications. An independent False Discovery Rate (FDR) analysis using the target-decoy approach provided with ProteinPilot software was used to assess the quality of the identifications and positive identifications were considered when identified proteins reached a 1% global FDR with at least one peptide that reached a 5% local FDR.ProteomicsBruno ManadasTripleTOF 5600Center for Neuroscience and Cell BiologyPARTIALBos Taurus (bovine)28912585 Caldeira J, Santa C, Osório H, Molinos M, Manadas B, Gonçalves R, Barbosa M. Matrisome Profiling During Intervertebral Disc Development And Ageing. Sci Rep. 2017 7(1):11629 10.1038/s41598-017-11960-0catiajmsanta@gmail.comBiologicalBiomedicalCenter for Neuroscience and Cell BiologyPortugalIntervertebral disc (IVD) degeneration is often the cause of low back pain. Degeneration occurs with age and is accompanied by extracellular matrix (ECM) depletion, culminating in nucleus pulpous (NP) extrusion and IVD destruction. The changes that occur in the disc with age have been under investigation. However, a thorough study of ECM profiling is needed, to better understand IVD development and age-associated degeneration. As so, iTRAQ LC-MS/MS analysis of foetus, young and old bovine NPs, was performed to define the NP matrisome. The enrichment of Collagen XII and XIV in foetus, Fibronectin and Prolargin in elder NPs and Collagen XI in young ones was independently validated. This study provides the first matrisome database of healthy discs during development and ageing, which is key to determine the pathways and processes that maintain disc homeostasis. The factors identified may help to explain age-associated IVD degeneration or constitute putative effectors for disc regeneration.Matrisome Profiling During Intervertebral Disc Development And Ageing.Caldeira Joana J, Santa Cátia C, Osório Hugo H, Molinos Maria M, Manadas Bruno B, Gonçalves Raquel R, Barbosa Mário Msodium salt, ammonium formate, 13C-labeled, cadmium salt, ion, Ethylenediaminetetraacetic acid, magnesium formate, zinc salt, (ethylenedinitrilo)tetraacetic acid, dmTAF[[II]]230, Polypeptides, protein polypeptide chains, KL receptor activity, hydrogen chloride, cobalt(II) formate dihydrate, sense of sight, GRP1, Grp1, B1, SCO5, SEC, 1, 2, SCO1, Gsfsow3, MMTS, 2-diaminoethane, (R*, ethylenediamine, Grain, pore, C79325, TFIID TAF250, cel, DmelCG13176, N, PTPSTEP, sec, Fluoride, ORF19, composition, proteins, W, SURGICAL AND MEDICAL PROCEDURES, AI047805, ethyl diamine, Benzenemethanesulfonyl Fluoride, Dehydrated ethanol, Wasserstoffchlorid, column, ACN, sample, Acn, D1, methyl methanesulfonothioate, Bs, CT, s, z, acidum edeticum, nickel salt, GPH, spiritus vini, LC-MS2, dTAF[[II]]230, Degradations, ethylenediamine monohydrochloride, cobalt (+2) salt, acid, Liquid Chromatography, Ct, 2'', TAF200, Guanidine Monohydroiodine, 3H-labeled cpd, Procedure, Absolute Alcohol, PBT, sodium (4:1:1) salt, ethane-1, Dm1, magnesium salt, ethylenediamine dihydrobromide, AI662483, Chlorwasserstoff, vision, 1110049F14Rik, [OEtH], Isopropanol, Acid, pbt, CG11628, galactose-6-sulfate sulfatase activity, content, Protein Digestions, formate, pooled, Guanidine Phosphate, methyl methanethiolsulfonate, cromium (+3), Striatum-enriched protein-tyrosine phosphatase, N'-1, beta Trypsin, alcohol etilico, Methylcarbinol, Taf250, liquid chromatography-tandem mass spectroscopy, 100kDa, ammonium (4:1) salt, chlorane, lead (+2) salt, TAF230, krk1, Cleland Reagent, l(1)VE614, nickel formate dihydrate, GRP1/cytohesin 1, lead formate, chlorure d'hydrogene, etanol, aluminum salt, acetylgalactosamine 6-sulfatase activity, MCPSF, 4-dimercapto-, Sodium Acetate Trihydrate, chondroitinsulfatase, R*)-, protein-containing complex, ethylenediamine dinitrate, thomson, body system, EDTA tetraanion, CG7826, mass-to-charge ratio, CG11633, Guanidium, LC-MSMS, ethylenediamine (1:1) sulfate, Gene Products, CG7835, CG42273, system, T6K12_14, iones., Monohydroiodine, anatomical systems, 2'''-(ethane-1, dTAF[[II]]250, FAM39E, 2-Ethane diylbis-(N-(carboxymethyl)glycine), Acinus, Hydrogen chloride, thallium (+1) salt, interventionDescription, ethylenediamine tetraacetic acid tetraanion, cell, Isopropyl, ethanol, beta-Trypsin, rubidium salt, Sodium Acetate, methanoic acid, ESI, ions, kf, acinusL, dTAF250, C2H5OH, Monohydrobromide, Guanidinium Chloride, acinusS, N-acetyl-D-galactosamine-6-sulfate 6-sulfohydrolase activity, ethylenediamine phosphate, peptidos, Saeure, Chromatographies, Anhydrous, PTHB1, Drl, inhibitors, proto-oncogene c-Kit, Guanidine Monohydrate, Guanidine Nitrate, Proteins, Clelands Reagent, stepk, chloridohydrogen, BG:DS00004.13, potassium formate, copper (+2) salt, Mischung, compositionality, buffer, Digestions, Cell, mKIAA0670, Mdu, dTAF230, N-acetylgalactosamine-6-sulfate sulfatase activity, polypeptide, chondroitin sulfatase, Wash2, Wash1, native protein, Temperatures, Proteolyses, KIT ligand receptor activity, mda, C18, cupric formate, chemical analysis, alcool ethylique, TAF[[II]]250/230, Nitrate, ethylenediamine dihydrochloride, aluminum formate, ami, XKrk1, T6K12.14, Cleland's, Intervention, Guanidine Monohydrobromide, AU020952, Taf[[II]]250, Guanidine, lithium formate, Guanidine Sulfate, Saeuren, med, 2610024B04Rik, Chloride, pore-forming toxin activity, CD117, DmelCG11628, ME-IV, Cleland's Reagent, Gene Proteins, Rubbing, C-Kit, Ssm, structure, xkl-1, edamine, liquid chromatography tandem mass spectrometry, Sulfate, hydrochloric acid, Benzenemethanesulfonyl, channel-forming toxin activity, ethylenediaminetetraacetate tetraanion, liquid chromatography tandem mass spectroscopy, ethylenediamine sulfate, Grain Alcohol, determination, zinc formate, Monohydrate, Guanidine Sulfate (2:1), lead salt, protein, Hydrogenchlorid, Xkl-1, 1-hydroxyethane, composed of, CG11121, alcohol, peptide, Cut, Ethanol, Gsfsco1, old, ammonium tetraformate, peptido, BANF, l(1)7Ba, l(1)7Bb, Alkohol, Phenylmethylsulfonyl, Min, 2', protein aggregate, Gsfsco5, DmelCG42273, Edetic acid, SOW3, 2-diyldinitrilo)tetraacetate, LCMSMS, ion(4-), peptides, Ethyl, 2610036I19Rik, 2610510L13Rik, min, chondroitinase, mAPC, Phosphate, Aethanol, copper, [CH2Me(OH)], free, SO, Protein Degradation, 2-ethanediamine, CG11387, fSAP152, lithium salt, Sputolysin, l(2)k08110, Guanidine Sulfate (1:1), TEAB, (ethane-1, ethylenediamine conjugate acid, Sl, ammonium (2:1) salt, So, ethylenediaminetetraacetate, Monohydrochloride, Alcohol, LC-MS/MS, Dmel_CG7826, Tr-kit, Th, {[-(BIS-CARBOXYMETHYL-AMINO)-ETHYL]-CARBOXYMETHYL-AMINO}-ACETIC ACID, mKIAA1367, Ethyl alcohol, TAFII-250, TAF250/230, Isopropyl Alcohol, TAFII250, batch, Cleland, Ionen, Digestion, 2-diamine, Rubbing Alcohol, 3.1.3.48, kl1-A, N-acetylgalactosamine 6-sulfatase activity, KIT, WASH, BcDNA:GH10590, ETHANOL, nickel (+2) salt, Dmel_CG7835, Mnb, MNB, tyrosine-protein kinase Kit, ammonium salt, EtOH, Step, acide, HCl, cobaltous formate, acids, Guanidine Monohydrochloride, 3-Butanediol, acido, kit, AW124434, CG17603, TAF[[II]], organ system, sensory visual perception, ethylenediamine dihydroiodide, STEP, SR3-5, copper salt, Polypeptide, d230, Reagent, Degradation, cesium salt, SCF receptor activity, Gene, dTAFII250, ethylenediaminetetraacetic acid, ACINUS, sight, EfW1, LC-MS-MS, Buffer, formic acid, Intervention or Procedure, cytohesin/GRP1, Aethylalkohol, potassium salt, polypeptide chain, scfr, DmelCG11121, dmTAF1, Phenylmethanesulfonyl Fluoride, Taf230, 14C-labeled, p63, Acide ethylenediaminetetracetique, p65, SCFR, cloruro de hidrogeno, acide edetique, Pulses, TAF250, Guanidium Chloride, Fdc, Taf200, somda, ethylenediamine (1:1) sulfite, Absolute, iones, Selb, inhibiteur, l(2)SH2 0323, Interventional, Taf1p, calcium formate, Protein Degradations, edta, 2-diyldinitrilo)tetraacetic acid, Benzenemethanesulfonyl fluoride, inhibidor, Neural-specific protein-tyrosine phosphatase, DmelCG11387, EDTA, cromium (+3) salt, 2 Propanol, TAF, Guanidine Hydrochloride, sodium formate, TCEP, Cpsf, ethylenediamine dihydrogen iodide, Intervention Strategies, DYRK1, TAF[[II]]250, ethylenediamine hydrochloride, nickel formate, protein complex, strontium formate, inhibitor, l(3)84Ab, Protein Digestion, strontium salt, Guanidinium, Peptide, acido edetico, [HCl], LC/MS/MS, secret agent, natural protein, p230, Protein, Hydrochloride, Guanidine Sulfite (1:1), TFIID, Dyrk1, Phenylmethanesulfonyl, connected anatomical system, l(2)SH0323, CG13176, Ethyl Alcohol, CYH1, c-KIT, TAF[[II]]230, CC1, H4edta, Tripcellim, TAF[II]250, sample population, chromic formate, Protein Gene Products, incised, Trypure, hydroxyethane, c-kit, DmelCG17603, Ion, calcium salt, Peptid, edetic acid, assay, SCH, TAF1spinal disc, intervertebral disc, Intervertebral, development, matrisome, single-organism developmental process, intervertebral disk, Intervertebral Discs, ageing., Intervertebral Disk, Extracellular, intervertebral fibrocartilage, Discs, discus intervertebralis, Matrix, Intervertebral Disks, Disc, Extracellular Matrices, Disks, Matrices, spinal disk, proteinaceous extracellular matrix, DiskApplications Software, Open Source, dairy cow, Computer Software, determination, cow, False, protein complex, T5E21.12, Proteins, Gene, protein, Computer, protein-containing complex, Peptide, Source Softwares, Software Tools, Tool, Programs, polypeptide, peptide, Polypeptides, Program, Computer Applications, protein polypeptide chains, Software Tool, ClvPrd, peptido, native protein, natural protein, polypeptide chain, cattle, Bos primigenius taurus, chemical analysis, Protein, Computer Applications Software, Computer Applications Softwares, Gene Products, domestic cattle, Software Engineering, Softwares, protein aggregate, Software, fixed, Computer Program, Application, Data Base, T5E21_12, Open Source Softwares, proportion, Software Applications, F, peptides, Source Software, Software Application, Open, Engineering, Bos bovis, proportionality, Open Source Software, Computer Programs and Programming, rate, proteins, Computer Software Application, Protein Gene Products, Computer Programs, Gene Proteins, Bos Tauurus, Applications, oxen, Applications Softwares, Tools, ox, quotient, Peptid, peptidos, domestic cow, assay, Polypeptide, species, bovine, Peptide., Computer Software Applications, ratioprojections, liquid chromatography tandem mass spectroscopy, dairy cow, single-organism developmental process, Posterior Compartment, metazoans, Backaches, protein, composed of, Atomkern, Collagen Type-XI, CG11121, protein polypeptide chains, old, alpha2 Subunit, Intervertebral Discs, Intervertebral Disk, horsetail nucleus, alpha3 Chain, Extracellular Matrices, protein aggregate, Fs(3)Hor, Collagen alpha1(XI), cold-insoluble globulin, Animalia, average, Group, treatment, neuronal nucleus, nucleus atomi, DmelCG2684, LCMSMS, Bos bovis, composition, Aches, proteins, NTef2, SO, Social, Bos Tauurus, oxen, Procollagen Type XI, GPIa*, FN1, alpha2 Chain, papilla, Backache, disease management, domestic cow, Collagen, Selective Attentions, associated, So, LC-MS2, Lower, Lower Back, anatomical protrusion, lamina, LC-MS/MS, flanges, Attention Focus, Normalcy, Lower back pain, Fs(3)Sz11, Ache, CIG, Low Backaches, Attention, Pains, shelf, precocious, Discs, 1.3.8.4, Back Aches, cell nucleus, IVD, Individual, Mechanical, spinal disc, FN, content, shelves, Pain, common, spinal disk, projection, ridge, Age, early, Health, Endogenous Regeneration, spine, liquid chromatography-tandem mass spectroscopy, Horka, CG2684, Fs(3)Horka, Regeneration, bovine, alpha1 Subunit, Intervertebral, 1300016K07Rik, Collagen alpha3(XI), advanced, noyau atomique, lamellae, Back Pain, number, Matrix, Gene, Kern, Selective, Matrices, protein-containing complex, Normalcies, animals, process of organ, presence, LC-MS-MS, Lower Back Pain, protrusion, lamella, AI463340, polypeptide chain, Type XI, cattle, DmelCG11121, Recurrent, LC-MSMS, Type XI Collagen, intervertebral fibrocartilage, Gene Products, Intervertebral Disks, Low, DmF2, Animal, nucleo atomico, Back Ache, Postural, nuclei, Normalities, lod, intervertebral disc, study, matrisome, somda, intervertebral disk, alpha1 Chain, nucleus of CNS, noyau, nucleo, ridges, Age Group, Lumbago, Postural Low Back Pain, Social Attention, nucleus, Low Back Pain, Extracellular, alpha3 Subunit, ECM, Low Backache, laminae, Low Back Pains, Drl, Individual Health, Collagen alpha2(XI), cow, protein complex, degeneration, anatomical process, Proteins, discus intervertebralis, Disc, Mechanical Low Back Pain, compositionality, LBP, Disk, EMILIN4, Mdu, development, count in organism, LC/MS/MS, Selective Attention, native protein, natural protein, nervous system nucleus, Bos primigenius taurus, mda, Low Back Ache, Metazoa, Protein, domestic cattle, Focus of Attention, Back Pains, Disks, flange, ami, organ process, Type XI Procollagen, Regenerations, 6720455E18Rik, Lower Back Pains, Normality, med, Collagen Type XI, Low Back, nucleus of neuraxis, Lds, neuraxis nucleus, Protein Gene Products, multicellular animals, ageing, processes, process, Gene Proteins, Procollagen, ox, structure, Recurrent Low Back Pain, processus, liquid chromatography tandem mass spectrometry, Endogenous., quantitative, MMRN, Low Back Aches, proteinaceous extracellular matrix, presence or absence in organismliquid chromatography tandem mass spectroscopy, projections, dairy cow, Collagens, single-organism developmental process, determination, Posterior Compartment, Backaches, Atomkern, CG11121, old, Intervertebral Discs, Intervertebral Disk, horsetail nucleus, death-inducing signalling complex, Extracellular Matrices, Zyderm, Turner-Kiser syndrome, cold-insoluble globulin, neuronal nucleus, nucleus atomi, LCMSMS, Microfibril Collagen Hemostat, discus, Bos bovis, Aches, inhibition of homeostatic process, SO, Bos Tauurus, oxen, GPIa*, FN1, Collagen Fleece, papilla, Backache, homeostasis, domestic cow, Collagen, associated, hereditary onychoostedysplasia, So, LC-MS2, Lower, Lower Back, anatomical protrusion, LC-MS/MS, lamina, flanges, Lower back pain, Ache, CIG, Low Backaches, Pains, positive regulation of homeostatic process, shelf, Discs, 1.3.8.4, Back Aches, cell nucleus, Disco, IVD, Mechanical, spinal disc, FN, shelves, NPS, NPS 1, Pain, spinal disk, iliac horn syndrome, projection, ridge, Endogenous Regeneration, spine, liquid chromatography-tandem mass spectroscopy, Regeneration, Collagen Felt, death receptor-induced signalling complex, death receptor-induced signaling complex, bovine, Intervertebral, 1300016K07Rik, noyau atomique, lamellae, Back Pain, Matrix, alpha-Collagen, Kern, Matrices, process of organ, LC-MS-MS, Lower Back Pain, protrusion, lamella, AI463340, Fong disease, nail-patella syndrome, Microfibril, DmelCG11121, cattle, Recurrent, Dermodress, Collastat, LC-MSMS, intervertebral fibrocartilage, Intervertebral Disks, Low, nucleo atomico, Back Ache, Postural, nuclei, intervertebral disc, study, matrisome, somda, intervertebral disk, nucleus of CNS, noyau, nucleo, Avitene, arthro-onychodysplasia, ridges, alpha Collagen, Lumbago, Postural Low Back Pain, DmelCG9908, nucleus, Low Back Pain, Extracellular, ECM, negative regulation of homeostatic process, Low Backache, nail patella syndrome, osteo-onychodysplasia, laminae, Collagenfleece, Low Back Pains, Drl, Autoregulation, cow, degeneration, anatomical process, discus intervertebralis, Disc, Mechanical Low Back Pain, LBP, Disk, EMILIN4, Mdu, development, NPS1, LC/MS/MS, DISC protein complex, nervous system nucleus, mda, Bos primigenius taurus, Low Back Ache, chemical analysis, Collagen Hemostat, disk, domestic cattle, DISC, Back Pains, Disks, regulation of homeostatic process, disc, ami, flange, Pangen, Data Base, organ process, Regenerations, 6720455E18Rik, Lower Back Pains, med, other collagen, Low Back, Avicon, nucleus of neuraxis, hereditary Osteo-onychodysplasia, CG9908, neuraxis nucleus, Turner-Kieser syndrome, onychoosteodysplasia, ageing, processes, process, activation of homeostatic process, ox, Recurrent Low Back Pain, liquid chromatography tandem mass spectrometry, processus, Endogenous., assay, MMRN, Low Back Aches, proteinaceous extracellular matrixspinal disc, intervertebral disc, Intervertebral, development, matrisome, single-organism developmental process, intervertebral disk, Intervertebral Discs, ageing., Intervertebral Disk, Extracellular, intervertebral fibrocartilage, Discs, discus intervertebralis, Matrix, Intervertebral Disks, Disc, Extracellular Matrices, Disks, Matrices, spinal disk, proteinaceous extracellular matrix, Disk318trueMatrisome remodelling during Intervertebral Disc development and ageingIntervertebral disc (IVD) is often the cause of low back pain. Degeneration occurs with age and is accompanied by extracellular matrix (ECM) depletion, culminating in nucleus pulpous (NP) extrusion and IVD destruction. Although the changes that occur with ageing in the IVD have been under study, not much attention has been given to ECM remodelling during normal development. Therefore, a thorough study of ECM composition and a comprehensive view on how this microenvironment is affected in normal ageing conditions is needed, to better understand the processes involved in IVD development and in age-associated degeneration. We have compared the NP matrisome of bovine IVDs from foetus, young and old animals by quantitative iTRAQ LC-MS/MS. Protein expression levels of the most interesting candidates were validated by Western Blot analysis. In total, 161 bovine proteins were identified. Of these, 77 molecules were common to the three different age groups, of which 36 defined the NP matrisome. Differential expression levels obtained for Collagen Type XI, XII, XIV, Fibronectin and Prolargin were independently confirmed. Herein, we demonstrate a shift in NP matrisome signatures that occurs in healthy bovine IVDs with development and ageing. Thus, this study provides insight into factors that may explain age-associated IVD degeneration, and which are putative targets for therapy. It also highlights key molecules, characteristic of early developmental stages, which constitute potential effectors in IVD regeneration.2017-09-252016-09-12PXD00492244664499175582580388043330295358511145120678110806124619624619728421880903206375811363350906836351524260275838538791187947398007NCBITaxon:128057230711143015557146835145262364927467073641135461388580358233888691568895399784475187043511232946928123977133103430436548135833925198183659219813143205965581914936329572637371922226938124781866132838828139558434513586722294914081691804541619341390363453058611204162326442104117061274432108595614963674283166694369455972945281132081003923751463308793694478420896431744712704459741194669400667145458793299352932010198126469041066114575512165994048339251216597216595160621213312831282128033952321832172920435315228112454349825747874980197525555362312951053948824261914548140479NCBITaxon:21575515343989903712059687623193516309800588845581817808173630943179698936160488566140985956641624251505907118702118705320496306386323555432046632226186633129945658606884535121612956717136044100160235870026420369456941016763410210761277351279334281461002935786174633132529111870021576418096997457744491566667547604452467675663366411363367102398709NCBITaxon:4055932582359092788396526498217498216347256569156934982116544872056971089448663661410NCBITaxon:49328715568910245980810005894526808732311561263730391141554219325552187268723872487298727123598755529547624223141463055180066556484198822299767982510216998231174673897641155744689444474153NCBITaxon:5055796016776744685960065349118226370824812991145784586326547876569835566369838192011164034624136810893161001428099612NCBITaxon:548681NCBITaxon:5820546414307747946409860995865228319662776689605960615729513722187594467087562270861711013354833090349741387834306154319474703342499859961588655974649987039202111072878272154078211908570752061894804189853502471587796271902528674636096469644375939146247274612874601134506253452659197221963852638588596NCBITaxon:61575052110517237263419610526419407722774694163481123869NCBITaxon:2703612506174767927459615384005257313136862697969903469417134434636034751536329644042101851097606331492862879669968583558595968912801726071018568729411009118696141633316780781186941436183NCBITaxon:6191118698840231719118691443906NCBITaxon:102982100071257118969419080218787870384766455347095145122578656870861822607046183260705260707NCBITaxon:780NCBITaxon:1502451516658457702921158612184520928574707856100761921263854504457383379192875NCBITaxon:10239NCBITaxon:71428291268063130957044830064110555241590841228233446900815025754122810430085210977970671356225833136185509924NCBITaxon:569382688306405277965833424090615101515NCBITaxon:6312406744824281307670708761383708281677285035158199011951953370249240536088817562615835583337602126719285321773572671193501846451772177437371137121534811215323957676901472664709157125627457903113335508596285963479249130821203511082332648373153214693NCBITaxon:9615318586141262200298131449376083334111497083332101841385691006581237561275045426662551519788749200375036009412934971530094114903524725175014012573983649908515151438992NCBITaxon:11552374539883747445446415259311792866432359839062887051580119459932716011497861603293338922853422999810894535936457428135122511775058NCBITaxon:1040750277916797181673871349158910306502780291595412629158515849327159570210022612876891355889309451464792031242164133280382803519284343985808946210803481793921182590287889295027906901749342725632273218903829286757221247190400727NCBITaxon:579653745711431123352114020641198108713554771424507229533355571415429315754622421392139011431933829355083329230741129221148489616010677252649997574472758024055071762993665812140535762294128160783957632066724005402965431843183224659620575549922957575759284593307972113549706671436733115955644421432138184922311240607210969762261111795752981818431923347986626528104660104421454747962678253567097477317513565050NCBITaxon:131360769938942468879552014887519795962141421972890922853164351867631854315206494089414424232NCBITaxon:177321409211577892979324470420755910117749906749907312989913112273170187367451011610358132842626587210360983341392696113079813367953588931640710340153033373301581913064139925306883910337212336819940445861153571147161224911131316634305312017554993512993850573176669642688451513458421572678728623862111510416079129831539222746891198351125522856253232136167633631689704132888188551167610807723264248618140843439376520965731116234273068808637721666898626NCBITaxon:2697049339862410312NCBITaxon:1389975864486433197291763438637256737863912334351052311009051414054493179986865540553486545147124546676056810429863571246791884019373345087711982159739757429719133734275779850762554702563188657623951655865886223864946454634076NCBITaxon:1107128835649184588NCBITaxon:592011087439145943267671111678107740844087977895349679440812269003803943512814595361235178616243230399668695978540964097954186942003089544243277869762791292499545115574840905180426430409286924093200302284812313625426428150475102993886538626290341554051208979662877323966920297936289955793157137117008930621864419548663436575584628238235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