Production of cytokines by in vitro stimulated naive B lymphocytes and memory B cells
Abstract
Introduction. Much attention is paid to the description of the role of individual cytokines in the induction of the differentiation of various subpopulations of B cells into plasmablasts and plasma cells. In contrast, little is known about which cytokines are produced by B-lymphocytes themselves and their subpopulations.
The aim of the study was to determine the cytokine profile of human B-lymphocytes during in vitro stimulation. The objectives of the study also included a comparative study of the spectrum of cytokines secreted by naive B-lymphocytes, as well as memory B-cells with switched and unswitched Ig synthesis.
Material and methods. Subpopulations of naive B-lymphocytes, as well as memory B-cells with switched (IgG+CD27+) and non-switched (IgM+CD27+) Ig synthesis were isolated using a flow cytometer. The isolated B lymphocytes were stimulated in vitro in the presence of feeder cells carrying the CD40L molecule and exogenous IL-21. Supernatants collected from stimulated B cells were analyzed for the presence of cytokines. The study was screening in nature. In order to cover the widest possible panel of lymphokines, a high-performance method of multiplex analysis was used in the work. The tested panel included the following lymphokines: EGF, Eotaxin, G-CSF, GM-CSF, IFN-α2, IFN-γ, IL-10, IL-12P40, IL-12P70, IL-13, IL-15, IL-17A, IL-1RA, IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IP-10, MCP-1, MIP-1α, MIP-1β, RANTES, TNFα, TNFβ, VEGF, FGF-2, TGF-α, Flt-3L, Fractalkine, GRO, MCP-3, MDC, PDGF-AA, PDGF-AB/BB and IL-9.
Results. Three subpopulations of stimulated B lymphocytes were obtained and functionally characterized: naive B-lymphocytes, memory B cells with switched (IgG+CD27+) and unswitched (IgM+CD27+) Ig synthesis. Stimulated B lymphocytes were characterized by active proliferation, acquired the plasmablast phenotype and secreted Ig. B lymphocytes stimulated in vitro in the IL-21/CD40L system produced a wide range of cytokines. IP-10, MDC and MCP-1 were secreted to the greatest extent. The secretion of IL-17A, IL-12P70, TGFα, IFN-γ, IL-3, IL-5 and IL-1β was below the level of detection. Naive B cells secreted the cytokines IL-10, MCP-3, MDC, IL-1α, MCP-1, TNFα and TNFβ more actively than memory B cells. For the cytokines IL-10, MDC, MCP-1, TNFα, and TNFβ, a significant difference was observed for both subpopulations of memory B cells. For MCP-3, a significant difference was observed only for memory B cells with switched Ig synthesis, and for IL-1α only when compared with memory B cells with unswitched Ig synthesis.
Conclusion. Cytokine profiles of activated B lymphocytes and their subpopulations were determined. The obtained results show that the production of cytokines by B cells is largely dependent on the activation and differentiation of B lymphocytes.
Keywords:B-lymphocytes; IL-21/CD40L stimulation; cytokines; Luminex assay; B memory cells; plasmablasts; ELISPOT
For citation: Byazrova M.G., Poroshina A.S., Sukhova M.M., Frolov E.A., Shilkina A.B., Latysheva E.A., Latysheva T.V., Filatov A.V. Production of cytokines by in vitro stimulated naive B lymphocytes and memory B cells. Immunologiya. 2023; 44 (5): 575–85. DOI: https://doi.org/10.33029/0206-4952-2023-44-5-575-585 (in Russian)
Funding. The study was supported by the grant of Russian Science Foundation (RSF) No. 23-15-00289, https://rscf.ru/project/23-15-00289/.
Conflict of interests. The authors declare no conflict of interests.
Authors’ contribution. Collection and processing of material – Byazrova M.G., Poroshina A.S., Sukhova M.M., Shilkina A.B.; text writing, editing – Frolov E.A., Filatov A.V.; the final version of the text – Latysheva E.A., Latysheva T.V., Filatov A.V.
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