Peripheral blood cultured mast cells: Phenotypic and functional outcomes of different culture protocols.
Cells, Cultured
/ immunology
Culture Media
/ metabolism
Flow Cytometry
Healthy Volunteers
Humans
Immunophenotyping
Interleukin-6
/ metabolism
Mast Cells
/ immunology
Nerve Tissue Proteins
/ metabolism
Primary Cell Culture
/ methods
Receptors, G-Protein-Coupled
/ metabolism
Receptors, Neuropeptide
/ metabolism
Time Factors
Flow cytometry
Human mast cell
IgE
IgE-dependent activation
MRGPRX2
Mast cell activation
Journal
Journal of immunological methods
ISSN: 1872-7905
Titre abrégé: J Immunol Methods
Pays: Netherlands
ID NLM: 1305440
Informations de publication
Date de publication:
05 2021
05 2021
Historique:
received:
30
12
2020
revised:
08
02
2021
accepted:
18
02
2021
pubmed:
2
3
2021
medline:
25
9
2021
entrez:
1
3
2021
Statut:
ppublish
Résumé
Mast cells (MCs) play a pivotal role in innate and adaptive immune responses. However, MCs are also involved in different pathologic conditions. Studies on the mechanisms that govern human MC functions are impeded by their limited and difficult recovery. Therefore, several research groups have developed protocols to culture human MCs from progenitor cells. These protocols vary with respect to culture duration and used maturation cytokines. How MCs obtained by different protocols differ in phenotype and functionality is currently unknown. To compare different protocols for the generation of human MCs from peripheral blood progenitors. Thirteen paired human MC cultures were investigated. MCs were cultured form CD34 Cell cultures for 4 weeks in the presence of IL-6 consistently yielded the highest numbers of MCs. MCs cultured for 8 weeks with IL-6 showed more autofluorescence significantly impeding correct analyses of FcεRI and CD32. The density of FcεRI and CD32 was comparable between the different culture conditions. MRGPRX2 expression was significantly higher in the 8 week cultures. The density of CD300a was increased in the cultures with IL-6. Cells cultured for 8 weeks were more responsive to MRGPRX2 activation. In contrast, the 4-weeks cultures with IL-6 showed significantly higher allergen-specific activation. Four weeks of culture with IL-6 are sufficient to generate sizeable numbers of human mast cells from blood progenitors, thereby enabling simultaneous exploration of allergen-specific sIgE/FcεRI cross-linking and non-specific activation via MRGPRX2.
Sections du résumé
BACKGROUND
Mast cells (MCs) play a pivotal role in innate and adaptive immune responses. However, MCs are also involved in different pathologic conditions. Studies on the mechanisms that govern human MC functions are impeded by their limited and difficult recovery. Therefore, several research groups have developed protocols to culture human MCs from progenitor cells. These protocols vary with respect to culture duration and used maturation cytokines. How MCs obtained by different protocols differ in phenotype and functionality is currently unknown.
OBJECTIVE
To compare different protocols for the generation of human MCs from peripheral blood progenitors.
METHODS
Thirteen paired human MC cultures were investigated. MCs were cultured form CD34
RESULTS
Cell cultures for 4 weeks in the presence of IL-6 consistently yielded the highest numbers of MCs. MCs cultured for 8 weeks with IL-6 showed more autofluorescence significantly impeding correct analyses of FcεRI and CD32. The density of FcεRI and CD32 was comparable between the different culture conditions. MRGPRX2 expression was significantly higher in the 8 week cultures. The density of CD300a was increased in the cultures with IL-6. Cells cultured for 8 weeks were more responsive to MRGPRX2 activation. In contrast, the 4-weeks cultures with IL-6 showed significantly higher allergen-specific activation.
CONCLUSION
Four weeks of culture with IL-6 are sufficient to generate sizeable numbers of human mast cells from blood progenitors, thereby enabling simultaneous exploration of allergen-specific sIgE/FcεRI cross-linking and non-specific activation via MRGPRX2.
Identifiants
pubmed: 33647250
pii: S0022-1759(21)00048-X
doi: 10.1016/j.jim.2021.113003
pii:
doi:
Substances chimiques
Culture Media
0
IL6 protein, human
0
Interleukin-6
0
MRGPRX2 protein, human
0
Nerve Tissue Proteins
0
Receptors, G-Protein-Coupled
0
Receptors, Neuropeptide
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
113003Informations de copyright
Copyright © 2021 Elsevier B.V. All rights reserved.