Summary of Study ST002726
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.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
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.
| Study ID | ST002726 |
| Study Title | Metabolic and Proteomic Divergence is Present in Circulating Monocytes and Tissue Resident Macrophages from Berkeley Sickle Cell Anemia and B-thalassemia mice (Spleen) |
| Study 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 |
| francesca.cendali@cuanschutz.edu | |
| Phone | 3037246131 |
| Submit Date | 2023-06-05 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-06-20 |
| Release Version | 1 |
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: | SU002832 |
| 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 |
|---|---|---|
| SA273964 | 44 | Berk |
| SA273965 | 43 | Berk |
| SA273966 | 42 | Berk |
| SA273967 | 46 | Berk |
| SA273968 | 48 | Berk |
| SA273969 | 50 | Berk |
| SA273970 | 49 | Berk |
| SA273971 | 41 | Berk |
| SA273972 | 47 | Berk |
| SA273973 | 45 | Berk |
| SA273974 | 55 | Beta Thal |
| SA273975 | 54 | Beta Thal |
| SA273976 | 53 | Beta Thal |
| SA273977 | 56 | Beta Thal |
| SA273978 | 57 | Beta Thal |
| SA273979 | 60 | Beta Thal |
| SA273980 | 59 | Beta Thal |
| SA273981 | 58 | Beta Thal |
| SA273982 | 52 | Beta Thal |
| SA273983 | 51 | Beta Thal |
| SA273984 | 37 | WT |
| SA273985 | 38 | WT |
| SA273986 | 39 | WT |
| SA273987 | 31 | WT |
| SA273988 | 30 | WT |
| SA273989 | 36 | WT |
| SA273990 | 32 | WT |
| SA273991 | 33 | WT |
| SA273992 | 34 | WT |
| SA273993 | 35 | WT |
| SA273994 | 40 | WT |
| Showing results 1 to 31 of 31 |
Collection:
| Collection ID: | CO002825 |
| Collection Summary: | The spleen organ tissue were harvested, finely minced, and each organ placed individually in 2mL Eppendorf tubes containing 1mL of DMEM with Collagenase D (Sigma Aldrich, product #11088866001). The tissues were incubated at 37C with agitation for 30 minutes. After incubation, 100 uL of 0.1M EDTA was added to the tissue-containing tubes and placed on ice. A single cell suspension was created by addition of Hanks buffered salt solution (HBSS, Corning, product #MT21022CV), passing through a 100 um filter, and collected in a 15 mL conical tube. The cell suspension was spun at 500g for 5 minutes and the supernatant discarded. The remaining tissue/cell solution was resuspended in 5ml of RBC lysis buffer (Invitrogen eBiosciences, product #00-4333-57), incubated at room temperature for 15 minutes, and centrifuged at 500g for 5 minutes, this step was repeated if RBC presence was sustained. Next, the cells were washed with Miltenyi buffer (HBSS, 0.5M EDTA, Fetal bovine serum), resuspended in 180uL Miltenyi buffer, and incubated on ice with CD11b Microbeads (Miltenyi Biotech, product #130-093-636) for 15minutes. Positive cells were isolated using magnetic LS columns (Miltenyi Biotech, product #130-042-401), collected in 2 mL Eppendorf tubes, counted, and then final pellet aliquots were frozen in liquid nitrogen and stored at -80C. continuously using LabScribe2 and analyzed offline. |
| Sample Type: | Macrophages |
Treatment:
| Treatment ID: | TR002841 |
| 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: | SP002838 |
| Sampleprep Summary: | Spleen macrophages 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 | AN004418 | AN004419 |
|---|---|---|
| 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: | CH003317 |
| 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: | CH003318 |
| 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: | 10% water; 0.1% formic acid |
| Solvent B: | 100% acetonitrile; 0.1% formic acid |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS004165 |
| Analysis ID: | AN004418 |
| 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: | MS004166 |
| Analysis ID: | AN004419 |
| 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 |
