TY - JOUR
T1 - Minimalistic In Vitro Culture to Drive Human Naive B Cell Differentiation into Antibody-Secreting Cells
AU - Unger, Peter-Paul A.
AU - Verstegen, Niels J. M.
AU - Marsman, Casper
AU - Jorritsma, Tineke
AU - Rispens, Theo
AU - ten Brinke, Anja
AU - van Ham, S. Marieke
N1 - Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. Copyright: Copyright 2021 Elsevier B.V., All rights reserved.
PY - 2021/5/12
Y1 - 2021/5/12
N2 - High-affinity antibody-secreting cells (ASC) arise from terminal differentiation of B-cells after coordinated interactions with T follicular helper (Tfh) cells in germinal centers (GC). Elucidation of cues promoting human naive B-cells to progress into ASCs is challenging, as this process is notoriously difficult to induce in vitro while maintaining enough cell numbers to investigate the differentiation route(s). Here, we describe a minimalistic in vitro culture system that supports efficient differentiation of human naive B-cells into antibody-secreting cells. Upon initial stimulations, the interplay between level of CD40 costimulation and the Tfh cell-associated cytokines IL-21 and IL-4 determined the magnitude of B-cell expansion, immunoglobulin class-switching and expression of ASC regulator PRDM1. In contrast, the B-cell-specific transcriptional program was maintained, and efficient ASC formation was hampered. Renewed CD40 costimulation and Tfh cytokines exposure induced rapid secondary STAT3 signaling and extensive ASC differentiation, accompanied by repression of B-cell identity factors PAX5, BACH2 and IRF8 and further induction of PRDM1. Our work shows that, like in vivo, renewed CD40L costimulation also induces efficient terminal ASC differentiation after initial B-cell expansion in vitro. This culture system for efficient differentiation of human naive B-cells into ASCs, while also maintaining high cell numbers, may form an important tool in dissecting human naive B-cell differentiation, thereby enabling identification of novel transcriptional regulators and biomarkers for desired and detrimental antibody formation in humans.
AB - High-affinity antibody-secreting cells (ASC) arise from terminal differentiation of B-cells after coordinated interactions with T follicular helper (Tfh) cells in germinal centers (GC). Elucidation of cues promoting human naive B-cells to progress into ASCs is challenging, as this process is notoriously difficult to induce in vitro while maintaining enough cell numbers to investigate the differentiation route(s). Here, we describe a minimalistic in vitro culture system that supports efficient differentiation of human naive B-cells into antibody-secreting cells. Upon initial stimulations, the interplay between level of CD40 costimulation and the Tfh cell-associated cytokines IL-21 and IL-4 determined the magnitude of B-cell expansion, immunoglobulin class-switching and expression of ASC regulator PRDM1. In contrast, the B-cell-specific transcriptional program was maintained, and efficient ASC formation was hampered. Renewed CD40 costimulation and Tfh cytokines exposure induced rapid secondary STAT3 signaling and extensive ASC differentiation, accompanied by repression of B-cell identity factors PAX5, BACH2 and IRF8 and further induction of PRDM1. Our work shows that, like in vivo, renewed CD40L costimulation also induces efficient terminal ASC differentiation after initial B-cell expansion in vitro. This culture system for efficient differentiation of human naive B-cells into ASCs, while also maintaining high cell numbers, may form an important tool in dissecting human naive B-cell differentiation, thereby enabling identification of novel transcriptional regulators and biomarkers for desired and detrimental antibody formation in humans.
KW - 3T3 Cells
KW - Animals
KW - Antibodies/chemistry
KW - Antibody Formation
KW - B-Lymphocytes/cytology
KW - CD40 Antigens/metabolism
KW - Cell differentiation
KW - Costimulatory molecules
KW - Cytokines
KW - Cytokines/metabolism
KW - Enzyme-Linked Immunosorbent Assay
KW - Gene Expression Regulation
KW - Germinal Center/immunology
KW - Humans
KW - Immunoglobulin G/immunology
KW - Immunoglobulin M/immunology
KW - In Vitro Techniques
KW - Lymphocyte Activation/immunology
KW - Mice
KW - NIH 3T3 Cells
KW - Phosphorylation
KW - Positive Regulatory Domain I-Binding Factor 1/metabolism
KW - Signal Transduction
KW - T-Lymphocytes, Helper-Inducer/cytology
KW - Transcription factors
UR - http://www.scopus.com/inward/record.url?scp=85107423833&partnerID=8YFLogxK
UR - https://pure.uva.nl/ws/files/127368908/Supplementary_Materials_Minimalistic_In_Vitro_Culture.pdf
U2 - https://doi.org/10.3390/cells10051183
DO - https://doi.org/10.3390/cells10051183
M3 - Article
C2 - 34066151
SN - 2073-4409
VL - 10
JO - Cells
JF - Cells
IS - 5
M1 - 1183
ER -