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Optimized Intracellular Staining Reveals Heterogeneous Cytokine Production Ability of Murine and Human Hematopoietic Stem and Progenitor Cells
- Source :
- Frontiers in Immunology, Frontiers in Immunology, Vol 12 (2021)
- Publication Year :
- 2021
- Publisher :
- Frontiers Media S.A., 2021.
-
Abstract
- Under stress conditions, hematopoietic stem and progenitor cells (HSPCs) can translate danger signals into a plethora of cytokine signals. These cytokines, or more precisely their combination, instruct HSPCs to modify the magnitude and composition of hematopoietic output in response to the threat, but investigations into the heterogeneous cytokine expression and regulatory mechanisms are hampered by the technical difficulty of measuring cytokine levels in HSPCs at the single-cell level. Here, we optimized a flow cytometry-based method for the simultaneous assessment of multiple intracellular cytokines in HSPCs. By selecting an optimal combination of cytokine restimulation reagents, protein transport inhibitors, and culture supplements, an optimized restimulation protocol for intracellular staining was developed. Using this method, we successfully examined expression levels of granulocyte/macrophage colony-stimulating factor (GM-CSF), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) in murine and human HSPC subsets under steady-state or different stress conditions. Different cytokine expression patterns were observed, suggesting distinct regulatory modes of cytokine production dependent on the HSPC subset, cytokine, disease, organ, and species. Collectively, this technical advance may help to obtain a better understanding of the nature of HSPC heterogeneity on the basis of differential cytokine production.
- Subjects :
- lcsh:Immunologic diseases. Allergy
tumor
medicine.medical_treatment
Immunology
hematopoietic stem and progenitor cells
Cell Culture Techniques
Gene Expression
Biology
Granulocyte
Immunophenotyping
Flow cytometry
Mice
medicine
Methods
cytokine
Animals
Humans
Immunology and Allergy
Macrophage
Progenitor cell
Cells, Cultured
Myelopoiesis
medicine.diagnostic_test
flow cytometry
Cell Differentiation
Hematopoietic Stem Cells
Immunohistochemistry
infection
Cell biology
Haematopoiesis
Cytokine
medicine.anatomical_structure
Cytokines
Tumor necrosis factor alpha
lcsh:RC581-607
Intracellular
Signal Transduction
Subjects
Details
- Language :
- English
- ISSN :
- 16643224
- Volume :
- 12
- Database :
- OpenAIRE
- Journal :
- Frontiers in Immunology
- Accession number :
- edsair.doi.dedup.....7d5c087a3b7df5db54ab0ebfc254947d