Summary of Study ST002983
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 PR001858. The data can be accessed directly via it's Project DOI: 10.21228/M8JM83 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 | ST002983 |
| Study Title | Deciphering the metabolic heterogeneity of hematopoietic stem cells with single-cell resolution |
| Study Summary | Metabolic status is crucial for stem cell functions; however, the metabolic heterogeneity of endogenous stem cells has never been directly assessed. Here, we develop a platform for high-throughput single-cell metabolomics (hi-scMet) of hematopoietic stem cells (HSCs). By combining flow cytometric isolation and nanoparticle-enhanced laser desorption/ionization mass spectrometry, we routinely detected >100 features from single cells. We mapped the single-cell metabolomes of all hematopoietic cell populations, and HSC subpopulations with different division times, detecting 33 features whose levels exhibited trending changes during HSC proliferation. We found progressive activation of oxidative pentose phosphate pathway (OxiPPP) from dormant to active HSCs. Genetic or pharmacological interference with OxiPPP increased reactive oxygen species level in HSCs, reducing HSC self-renewal upon oxidative stress. Together, our work uncovers the metabolic dynamics during HSC proliferation, reveals a role of OxiPPP for HSC activation, and illustrates the utility of hi-scMet in dissecting metabolic heterogeneity of immunophenotypically defined cell populations. |
| Institute | Shanghai Jiao Tong University |
| Last Name | CAO |
| First Name | JING |
| Address | 1954 Huashan Road, Shanghai, Shanghai, 200030, China |
| caojing1@sjtu.edu.cn | |
| Phone | +8615201957271 |
| Submit Date | 2023-11-21 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | .txt |
| Analysis Type Detail | MALDI |
| Release Date | 2023-11-28 |
| Release Version | 1 |
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Collection:
| Collection ID: | CO003089 |
| Collection Summary: | Bones were rapidly dissected and stored on ice in Ca2+- and Mg2+-free HBSS (ThermoFisher) plus 2% heat-inactivated fetal bovine serum (Gibco). Bone marrow cells were flushed out from the bone and then dissociated to single-cell suspension by gently passing through the needle then filtering through a 70-μm nylon mesh. The following antibodies were used to isolate hematopoietic cells: TER-119-FITC, CD3-FITC, CD5-FITC, CD8-FITC, B220-FITC, Gr-1-FITC, TER-119-APC780, CD3e-APC780, CD5-APC780, CD8-APC780, B220-APC780, Gr-1-APC780, TER-119-biotin, CD3-biotin, B220-biotin, Gr-1-biotin, c-kit-biotin, TER-119-PerCP/Cy5.5, Sca-1-PerCP/Cy5.5, MAC-1-APC, CD48-APC, CD16/32-APC, CD135-APC, CD127-PE, CD34-PE and CD3-PE, CD150-PE. FITC streptavidin, APC/Cy7 streptavidin and/or APC-R700 streptavidin were used for biotin-labeled antibodies. All reagents were acquired from BD Biosciences, eBiosciences or BioLegend. For isolation of HSPCs, cells were incubated with c-kit-biotin and paramagnetic microbeads, and then passed through an autoMACS magnetic separator. For isolation of CLPs, lineage was stained with Ter119-biotin, CD3-biotin, B220-biotin and Gr-1-biotin and paramagnetic microbeads. Then an autoMACS magnetic separator was used to enrich lineage- populations. To minimize metabolic changes, antibody-stained cells were fixed with 2% PFA (Sigma) for 15 minutes on ice. Flow cytometric analysis was performed with a BD LSRFortessa cytometer. To measure ROS levels of HSCs, antibody-stained cells were incubated with 5μm DCFDA for 15 minutes at 37°C before flow cytometric analysis. For metabolic detection, cells were sorted with a FACSAria SORP cytometer into 384-well plates containing 2.5-μl 80% methanol (Sigma) per well, and then centrifuged at 1500g for 5 mins at 4°C. Plates were sunk in liquid nitrogen for 10 mins to lyse cells and kept at -80°C before MS analysis. |
| Sample Type: | Stem cells |
