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NanoLC-MSMS TMT comparison of native and in vitro culture-derived neutrophils

ABSTRACT: Triplicate samples of donor-matched native neutrophils (purified through fluorescence-assisted flow cytometry using CD66b as a marker) and 18-day in vitro cultured neutrophils (derived from CD34+ cells) were analysed through TMT-based quantitative proteomics to examine possible protein abundance changes underlying neutrophil cell culture.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture, Neutrophil, Blood

DISEASE(S): Disease Free

SUBMITTER: Pedro Moura

LAB HEAD: Ash Toye

PROVIDER: PXD052008 | Pride | 2024-06-02

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	190620_6Plex_TMT.pdResult	Other
	190620_6Plex_TMT.pdResultView	Other
	190620_6Plex_TMT.xlsx	Xlsx
	190620_6Plex_TMT_MSMSSpectrumInfo.txt	Txt
	190620_6Plex_TMT_PS.msf	Msf

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1 - 5 of 27

Publications

Neutrophils cultured ex vivo from CD34<sup>+</sup> stem cells are immature and genetically tractable.

Naveh Claire A CA Roberts Kiran K Zakrzewski Przemysław P Rice Christopher M CM Ponce-Garcia Fernando M FM Fleming Kathryn K Thompson Megan M Panyapiean Nawamin N Jiang Huan H Diezmann Stephanie S Moura Pedro L PL Toye Ashley M AM Amulic Borko B

Journal of translational medicine 20240531 1

<h4>Background</h4>Neutrophils are granulocytes with essential antimicrobial effector functions and short lifespans. During infection or sterile inflammation, emergency granulopoiesis leads to release of immature neutrophils from the bone marrow, serving to boost circulating neutrophil counts. Steady state and emergency granulopoiesis are incompletely understood, partly due to a lack of genetically amenable models of neutrophil development.<h4>Methods</h4>We optimised a method for ex vivo produc ...[more]

PMID: 38822352

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NanoLC-MSMS TMT comparison of in vitro cultured/native reticulocytes and erythrocytes

Project description:Triplicate samples of donor-matched native reticulocytes (CD71+ cells), erythrocytes (CD71- cells) and in vitro culture-derived reticulocytes (derived from CD34+ cells) were analysed through TMT-based quantitative proteomics to examine possible protein abundance changes underlying reticulocyte maturation.

2018-08-03 | PXD009023 | Pride

NanoLC-MSMS TMT comparison of circulated and unstimulated in vitro cultured reticulocytes

Project description:Triplicate samples of unstimulated in vitro culture-derived reticulocytes (derived from CD34+ cells) and cultured reticulocytes circulated overnight in an ex vivo circulation system were analysed through TMT-based quantitative proteomics to explore the impact of shear stress on reticulocyte protein abundance.

2018-08-03 | PXD009024 | Pride

Characterisation of proteome of a novel Escherichia coli strain exhibiting mucoviscous phenotype.

Project description:Protein expression by E. coli 26561 during the late-exponential phase of cultures under anaerobic conditions was examined. E. coli 26561 is a multidrug resistant (MDR) and shows an unusual hyper-mucoviscous phenotype. Resistance includes ESBL (CTX-M-14) and proteome was determined with and without exposure to sub-MIC concentrations of the 3rd generation cephalosporin ceftazidime. Ceftazidime exposure was at two sub-MIC levels, specifically 0.25x MIC (samples 5-7), 0.5x MIC (samples 8 - 10); samples 1-4 provided the unexposed Control. Both whole and phospho-enriched fractions for each sample were analysed. Quantification of peptides was assessed using 10-plex TMT labelling in conjunction with an Orbitrap Fusion Tribrid. Raw data produced by the Orbitrap were processed using Max Quant 1.5.4.7 using the included Andromeda search engine. Peptides were searched against our own database of E. coli 26561 proteins which was produced from a hybrid assembly of our reads obtained from MiSeq and PacBio sequencing platforms.

2018-08-22 | PXD008339 | Pride

ChAdOx infection of human cell lines A549 MRC-5

Project description:Total proteome and phospho proteome of two human cell lines infected with the ChAdOx1 nCoV-19 virus over 72 hours

2021-01-11 | PXD023286 | Pride

Enhancement of red blood cell transfusion compatibility using CRISPR-mediated erythroblast gene editing

Project description:Regular blood transfusion is the cornerstone of care for patients with red blood cell (RBC) disorders such as thalassemia or sickle cell disease. With repeated transfusion, alloimmunisation often occurs due to incompatibility at the level of minor blood group antigens. We use CRISPR-mediated genome editing of an immortalised human erythroblast cell line (BEL-A) to generate multiple enucleation competent cell lines deficient in individual blood groups. Edits are combined to generate a single cell line deficient in multiple antigens responsible for the most common transfusion incompatibilities: ABO (Bombay phenotype), Rh (Rhnull), Kell (K0), Duffy (Duffynull), GPB (S- s- U-). These cells can be differentiated to generate deformable reticulocytes, illustrating the capacity for coexistence of multiple rare blood group antigen null phenotypes. This study provides the first proof-of-principle demonstration of combinatorial CRISPR-mediated blood group gene editing to generate customisable or multi-compatible RBCs for diagnostic reagents or recipients with complicated matching requirements.

2018-04-26 | PXD009291 | Pride

Glutamate as an extracellular signalling molecule in unicellular protists

Project description:Glutamate supplied to Phaeodactylum tricornutum at concentrations of 25 uM and 250 uM. Samples were taken 4 hours after glutamate addition.

2026-05-28 | PXD074385 | Pride

Placental macrophage (Hofbauer cell) response to viral and bacterial ligands is influenced by fetal sex

Project description:Bacterial and viral infections of the placenta are associated with inflammation and adverse pregnancy outcomes. Hofbauer cells (HBCs) are specialised fetal-origin macrophages in the placental villi and are proposed to protect the fetus from vertical transmission of pathogens; however, they are poorly understood. Here, we have performed quantitative proteomics on term HBCs under resting conditions and following exposure to bacterial and viral pathogen associated molecular patterns (PAMPs), and investigated the contribution of fetal sex to these responses. Resting HBCs expressed a plethora of proteins pertinent to macrophage function, including chemokines, cytokines, Toll-like receptors, and classical and non-classical major histocompatibility complex class I and II molecules. HBCs mounted divergent responses to bacterial versus viral PAMPs but exhibited protein expression changes suggestive of a switch towards a more pro-inflammatory phenotype. A comparison between male and female HBCs, showed that the latter mounted a much stronger and wider response. Sexual dimorphism in HBCs was primarily associated with lipid metabolism in males and cytoskeleton organisation in females. We provide a novel and comprehensive understanding regarding the phenotype of term placental macrophages and their sex-dependent responses to infectious triggers.

2022-10-31 | PXD033006 | Pride

NanoLC-MSMS TMT whole cell comparison of ex vivo derived macrophage subtypes.

Project description:Triplicate samples of donor matched ex vivo generated macrophage subtypes M1, M2a, M2c compared against the M0 control analysed using TMT-based 11plex quantitative proteomics.

2023-09-04 | PXD032801 | Pride

The effect of GLP-1 receptor agonist liraglutide on the rat neurointermediate lobe phosphoproteome

Project description:Obesity and its co-morbidities, such as diabetes and hypertension, can significantly reduce a person’s quality of life and place huge pressure on healthcare resources. When we eat a meal our gut and brain release hormones to control the amount of food and fluid we ingest to prevent overeating. One of these hormones is called glucagon-like peptide 1 (GLP-1) and is released from intestinal cells in response to food intake, but also produced and released in the brain. Drug analogues of GLP-1 are already in use in the clinic to treat both diabetes and obesity. The aim of this work was to obtain fundamental knowledge about a GLP-1 receptor population in nerve terminals of the posterior pituitary gland. We have investigated the pharmacological actions of GLP-1 using a selective receptor agonist called liraglutide, a drug that is approved for diabetes and obesity treatment in humans. Our work has focussed on the phosphoproteome of the neurointermediate lobe (posterior pituitary + intermediate lobe) of the rat pituitary gland 30 minutes after intraperitoneal injection of liraglutide (100 µg/kg) compared to vehicle controls (n = 6 animals per group). New understanding of this GLP-1 receptor population is essential for our knowledge of current treatments of diabetes and obesity that use stable peptide analogues in humans.

2023-03-02 | PXD037495 | Pride

Chemical degradation of BTK/TEC as a novel approach to inhibit platelet function.

Project description:Determination of the effect of the generic tyrosine kinase degrader TL12-186, and the BTK degraders DD-04-15 and DD-03-171 on human platelet protein content and function.

2023-03-02 | PXD034440 | Pride