Summary of Study ST002725

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001691. The data can be accessed directly via it's Project DOI: 10.21228/M84B0K This work is supported by NIH grant, U2C- DK119886.

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This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST002725
Study TitleMetabolic and Proteomic Divergence is Present in Circulating Monocytes and Tissue Resident Macrophages from Berkeley Sickle Cell Anemia and B-thalassemia mice (PBMC's)
Study SummarySickle cell disease and Beta-thalassemia represent hemoglobinopathies arising from dysfunctional or under produced beta-globin chains, respectively. In both diseases, red blood cell injury and anemia are the impetus for end organ injury. Because persistent erythrophagocytosis is a hallmark of these genetic maladies it is critical to understand how macrophage phenotype polarizations in tissue compartments can inform on disease progression. Murine models of sickle cell disease and Beta-thalassemia allow for a basic understanding of mechanisms and provide for translation to human disease. A multi-omics approach to understanding macrophage metabolism and protein changes in two murine models of beta-globinopathy was performed on peripheral blood mononuclear cells as well as spleen and liver macrophages isolated from Berkley sickle cell disease (Berk-ss) and heterozygous B1/B2 globin gene deletion (Hbbth3/+) mice. Results from these experiments revealed the metabolome and proteome of macrophages are polarized to a distinct phenotype in Berk-ss and Hbbth3/+ compared each other and their common background mice (C57BL6/J). Further, spleen and liver macrophages revealed distinct disease specific phenotypes, suggesting macrophages become differentially polarized and reprogrammed within tissue compartments. We conclude that tissue recruitment, polarization, metabolic and proteomic reprogramming of macrophages in Berk-ss and Hbbth3/+ mice may be relevant to disease to progression in other tissue.
Institute
University of Colorado School of Medicine
LaboratoryLaboratory of Angelo D'Alessandro in collaboratation with David Irwin
Last NameCendali
First NameFrancesca
Address13199 East Montview Boulevard, Aurora, CO, 80045, USA
Emailfrancesca.cendali@cuanschutz.edu
Phone3037246131
Submit Date2023-06-05
Raw Data AvailableYes
Raw Data File Type(s)raw(Thermo)
Analysis Type DetailLC-MS
Release Date2023-06-21
Release Version1
Francesca Cendali Francesca Cendali
https://dx.doi.org/10.21228/M84B0K
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:


Project:

Project ID:PR001691
Project DOI:doi: 10.21228/M84B0K
Project Title:Metabolic and Proteomic Changes in Sickle Cell Disease and B-thalassemia Mouse Splenic and Hepatic Macrophages and Peripheral Blood Mononuclear cells
Project Summary:Sickle cell disease and Beta-thalassemia represent hemoglobinopathies arising from dysfunctional or under produced beta-globin chains, respectively. In both diseases, red blood cell injury and anemia are the impetus for end organ injury. Because persistent erythrophagocytosis is a hallmark of these genetic maladies it is critical to understand how macrophage phenotype polarizations in tissue compartments can inform on disease progression. Murine models of sickle cell disease and Beta-thalassemia allow for a basic understanding of mechanisms and provide for translation to human disease. A multi-omics approach to understanding macrophage metabolism and protein changes in two murine models of beta-globinopathy was performed on peripheral blood mononuclear cells as well as spleen and liver macrophages isolated from Berkley sickle cell disease (Berk-ss) and heterozygous B1/B2 globin gene deletion (Hbbth3/+) mice. Results from these experiments revealed the metabolome and proteome of macrophages are polarized to a distinct phenotype in Berk-ss and Hbbth3/+ compared each other and their common background mice (C57BL6/J). Further, spleen and liver macrophages revealed distinct disease specific phenotypes, suggesting macrophages become differentially polarized and reprogrammed within tissue compartments. We conclude that tissue recruitment, polarization, metabolic and proteomic reprogramming of macrophages in Berk-ss and Hbbth3/+ mice may be relevant to disease to progression in other tissue.
Institute:University of Colorado School of Medicine
Laboratory:Laboratory of Angelo D'Alessandro in collaboratation with David Irwin
Last Name:Cendali
First Name:Francesca
Address:13199 East Montview Boulevard, Aurora, CO, 80045, USA
Email:francesca.cendali@cuanschutz.edu
Phone:3037246131

Subject:

Subject ID:SU002831
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090

Factors:

Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id Factor
SA27393514Berk
SA27393613Berk
SA27393716Berk
SA27393817Berk
SA27393918Berk
SA27394012Berk
SA27394115Berk
SA27394224Beta Thal
SA27394323Beta Thal
SA27394422Beta Thal
SA27394521Beta Thal
SA27394625Beta Thal
SA27394726Beta Thal
SA27394829Beta Thal
SA27394928Beta Thal
SA27395027Beta Thal
SA27395120Beta Thal
SA27395219Beta Thal
SA2739538WT
SA27395411WT
SA27395510WT
SA2739562WT
SA2739577WT
SA2739586WT
SA2739593WT
SA2739604WT
SA2739615WT
SA2739621WT
SA2739639WT
Showing results 1 to 29 of 29

Collection:

Collection ID:CO002824
Collection Summary:Whole blood samples (1 mL) were obtained from animals via cardiac puncture, using a syringe with a 26-gauge needle, and placed in an EDTA treated tube. The blood was transferred to a 15 mL conical tube and diluted 2:1, sterile PBS: blood, and gently mixed. Lympholyte® Mammal Cell Separation media (Cedarlane Labs, product # CL5115) was gently added to the bottom of the blood solution and spun at 1400 rpm for 30 minutes in a refrigerated centrifuge. After centrifugation, layers were visualized, the PBMC layer (midlayer) was extracted and resuspended in a new tube. The isolated PBMC’s were washed using ~14mL sterile PBS, spun at 1800 rpm for 10 minutes, excess PBS was removed, cells were resuspended in 1-2mL for counting, and the final pellet was frozen in liquid nitrogen and stored at -80C.
Sample Type:Mononuclear cells

Treatment:

Treatment ID:TR002840
Treatment Summary:Eight- to ten-week-old female C57Bl/6 WT, Berk-ss, or Hbbth3/+ mice were either obtained from Jackson Laboratories (Bar Harbor, ME, USA) or our in-house Berk SCD mouse colony. Mice were housed and bred in an AAALAC accredited animal facility at the University of Colorado, Denver, Anschutz Medical campus and were maintained on a 12:12 light-dark cycle with food and water available ad libitum. Female heterozygous Berk-ss mice were bred with male homozygous Berkss mice to generate homozygous offspring. Specifically, Berk-ss mice with genotype Tg(HuminiLCR α1 Gγ Aγ δ βs ) Hba0/0 Hbb0/0 and the hemizygous with genotype Tg(Hu-miniLCR α1 Gγ Aγ δ βs ) Hba0/0 Hbb0 Hbb+ were littermates. Genotyping of mice used for breeding and experiments was performed by TransnetYX (Cordova, TN, USA). A total of 21 mice (C57Bl/6: n=10, Berk-ss mice: n=11, Hbbth3/+ =10) were used in the present investigation and levels of discomfort and distress were monitored daily by the in-house animal care staff, with a veterinarian available as needed. All experimental procedures were conducted under the guidelines recommended by The Journal of Physiology (11), the National Institutes of Health and were approved by the Institutional Animal Care and Use Committee at the University of Colorado, Denver, Anschutz Medical Campus.

Sample Preparation:

Sampleprep ID:SP002837
Sampleprep Summary:Spleen and liver macrophages and peripheral blood mononuclear cells (PBMCs) were extracted in methanol:acetonitrile:water (5:3:2 v/v/v – at a 1x10^6 cells/ml) ). The samples were vortexed for 30mins at 4°C. The samples were then spun down at 18,213 g for 10 minutes, 4°C and 50uL of supernatant was transferred to autosampler vial.

Combined analysis:

Analysis ID AN004416 AN004417
Analysis type MS MS
Chromatography type Reversed phase Reversed phase
Chromatography system Thermo Vanquish Thermo Vanquish
Column Phenomenex Kinetex C18 (150 x 2.1mm,1.7um) Phenomenex Kinetex C18 (150 x 2.1mm,1.7um)
MS Type ESI ESI
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap
Ion Mode NEGATIVE POSITIVE
Units :Peak area :Peak area

Chromatography:

Chromatography ID:CH003315
Chromatography Summary:Negative ion Mode
Instrument Name:Thermo Vanquish
Column Name:Phenomenex Kinetex C18 (150 x 2.1mm,1.7um)
Column Temperature:45
Flow Gradient:0-0.5 min 0% B, 0.5-1.1 min 0-100% B, 1.1-2.75 min hold at 100% B, 2.75-3 min 100-0% B, 3-5 min hold at 0% B
Flow Rate:0.450ml/min
Solvent A:95% water/5% acetonitrile; 1 mM ammonium acetate
Solvent B:95% acetonitrile/5% water; 1 mM ammonium acetate
Chromatography Type:Reversed phase
  
Chromatography ID:CH003316
Chromatography Summary:Positive Ion Mode
Instrument Name:Thermo Vanquish
Column Name:Phenomenex Kinetex C18 (150 x 2.1mm,1.7um)
Column Temperature:45
Flow Gradient:0-0.5 min 5% B, 0.5-1.1 min 5-95% B, 1.1-2.75 min hold at 95% B, 2.75-3 min 95-5% B, 3-5 min hold at 5% B.
Flow Rate:0.450ml/min
Solvent A:100% water; 0.1% formic acid
Solvent B:100% acetonitrile; 0.1% formic acid
Chromatography Type:Reversed phase

MS:

MS ID:MS004163
Analysis ID:AN004416
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Resolution 70,000, scan range 65-900 m/z, maximum injection time 200 ms, microscans 2, automatic gain control (AGC) 3 x 10^6 ions, source voltage 4.0 kV, capillary temperature 320 C, and sheath gas 45, auxiliary gas 15, and sweep gas 0 (all nitrogen). Data converted to mzXML using RawConverter. Metabolites were annotated and integrated using Maven in conjunction with the KEGG database.
Ion Mode:NEGATIVE
  
MS ID:MS004164
Analysis ID:AN004417
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Resolution 70,000, scan range 65-900 m/z, maximum injection time 200 ms, microscans 2, automatic gain control (AGC) 3 x 10^6 ions, source voltage 4.0 kV, capillary temperature 320 C, and sheath gas 45, auxiliary gas 15, and sweep gas 0 (all nitrogen). Data converted to mzXML using RawConverter. Metabolites were annotated and integrated using Maven in conjunction with the KEGG database.
Ion Mode:POSITIVE
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