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Lymphocyte Isolation from Human Skin for Phenotypic Analysis and Ex Vivo Cell Culture
Published on: April 8, 2016
Migratory properties and functional capacities of human skin dendritic cells
C D Richters1, M J Hoekstra, J Van Baare
1Department of Cell Biology, Faculty of Medicine, Vrije Universiteit, Amsterdam, The Netherlands.
Insights
Human skin explants release CD1a+ dendritic cells, T cells, and macrophages. CD1a+ cells, crucial for immune response, retain potent allo-activating capacities regardless of CD1b+ expression.
Area of Science:
- Immunology
- Dermatology
- Cell Biology
Background:
- Dendritic cells (DCs) play a critical role in immune responses.
- CD1 molecules (CD1a, CD1b) are involved in antigen presentation.
- Understanding DC subsets in human skin is vital for immunology.
Purpose of the Study:
- Characterize spontaneously migrating cells from human skin explants.
- Investigate the phenotype and function of CD1a+ and CD1b+ dendritic cells.
- Determine the role of CD1b+ expression on CD1a+ cell function.
Main Methods:
- Human skin explants cultured in vitro.
- Phenotyping of migrating cells using flow cytometry (CD1a, CD1b markers).
- Assessment of allogeneic T cell stimulation capacity of purified CD1a+ cells.
Main Results:
- Migrating cells primarily comprised CD1a+ dendritic cells, T cells, and macrophages.
- A subset of CD1a+ cells co-expressed CD1b+ early in culture.
- CD1a+ cells demonstrated potent allo-activating capacity, irrespective of CD1b+ expression.
Conclusions:
- CD1a+ dendritic cells are key migrating cells from human skin.
- CD1b+ expression on CD1a+ cells may indicate active migration.
- CD1a+ cells possess robust T cell stimulatory functions, independent of CD1b+ status.
Abstract:
The different cell types which migrated 'spontaneously' out of human skin explants during different periods of culture were characterized. Before culture, CD1a+ dendritic cells were observed not only in the epidermis but also in the dermis, whereas CD1b+ dendritic cells were present exclusively in the dermis. The populations of migrating cells were harvested and phenotyped on 3 successive days of culture. They always contained high percentages of CD1a+ cells. The other cells that migrated were T cells and macrophages. A relatively high proportion of the CD1a+ cells that migrated during the first 24 h culture period was also CD1b+. The number of cells which were positive for both CD1a and CD1b decreased in the following 2 days of culture. However, the purified CD1a+ cell populations isolated on the 3 consecutive days did not show any difference in their capacity to stimulate allogeneic T cells. The CD1a+ cells possess potent allo-activating capacities that are independent of whether or not they are positive for CD1b+. Three days after culture about half of the CD1a+ cells were still present in the epidermis and dermis, but no CD1b+ cells could be detected in the dermis. This suggests that the CD1b+ cells represent a population of active migrating cells.
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