Stimulation strategy matters: A B-cell focused single-cell transcriptomic study on in vitro stimulated immune cells
- 1VIB - Ghent University
Abstract
Distinct in vitro B-cell stimulation strategies, defined by stimulation condition and incubation duration, have been described in diverse studies. Often, these approaches are considered interchangeable due to comparable flow-cytometric outputs concerning fluorescence intensities and B-cell subset distributions. However, whether these phenotypic similarities also reflect comparable underlying transcriptional states have not been systematically evaluated yet. In this study, we provide an unbiased comparison of B-cell activation strategies on healthy individuals, resulting in a single-cell transcriptomic atlas (4 donors; ~2,029 cells per sample; 81,160 cells in total with 56.3% annotated as B cells). Two T-cell independent stimulators targeting toll-like receptors (ODN2006 and IFN-{gamma}/R-848), were assessed next to two T-cell dependent stimulators (anti-CD40 and anti-CD3/CD28 antibodies), all incubated for eight and 30 hours. Observed overlap for differential abundance, differential gene expression and cell-cell interactions could directly be linked to T-cell (in)dependent working mechanism. Strategy-specific results uncovered time-point specific transcriptional changes, revealed the largest set of differentially expressed genes for IFN-{gamma}/R-848 and ODN2006, and showed that IFN-{gamma}/R-848 and anti-CD3/CD28 induced the majority of the cell-cell interactions. Ultimately, our reference atlas emphasizes the importance and facilitates the rational selection of optimal B-cell stimulation strategies, particularly for studies aiming to interrogate B-cell dysregulation in immune-mediated diseases.
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BibTeX
@misc{dhamers2026stimulation,
title = {{Stimulation strategy matters: A B-cell focused single-cell transcriptomic study on in vitro stimulated immune cells}},
author = {T. D'hamers and J. Neirinck and M. Buysse and C. De Vriendt and T. Kerre and F. Haerynck and M. Hofmans and C. Bonroy and C. Everaert},
howpublished = {bioRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.22.760934},
url = {https://www.biorxiv.org/content/10.64898/2026.10.22.760934v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Stimulation strategy matters: A B-cell focused single-cell transcriptomic study on in vitro stimulated immune cells
AU - T. D'hamers
AU - J. Neirinck
AU - M. Buysse
AU - C. De Vriendt
AU - T. Kerre
AU - F. Haerynck
AU - M. Hofmans
AU - C. Bonroy
AU - C. Everaert
PY - 2026
DA - 2026/10/22
PB - bioRxiv
DO - 10.64898/2026.10.22.760934
UR - https://www.biorxiv.org/content/10.64898/2026.10.22.760934v1
AB - Distinct in vitro B-cell stimulation strategies, defined by stimulation condition and incubation duration, have been described in diverse studies. Often, these approaches are considered interchangeable due to comparable flow-cytometric outputs concerning fluorescence intensities and B-cell subset distributions. However, whether these phenotypic similarities also reflect comparable underlying transcriptional states have not been systematically evaluated yet. In this study, we provide an unbiased comparison of B-cell activation strategies on healthy individuals, resulting in a single-cell transcriptomic atlas (4 donors; ~2,029 cells per sample; 81,160 cells in total with 56.3% annotated as B cells). Two T-cell independent stimulators targeting toll-like receptors (ODN2006 and IFN-{gamma}/R-848), were assessed next to two T-cell dependent stimulators (anti-CD40 and anti-CD3/CD28 antibodies), all incubated for eight and 30 hours. Observed overlap for differential abundance, differential gene expression and cell-cell interactions could directly be linked to T-cell (in)dependent working mechanism. Strategy-specific results uncovered time-point specific transcriptional changes, revealed the largest set of differentially expressed genes for IFN-{gamma}/R-848 and ODN2006, and showed that IFN-{gamma}/R-848 and anti-CD3/CD28 induced the majority of the cell-cell interactions. Ultimately, our reference atlas emphasizes the importance and facilitates the rational selection of optimal B-cell stimulation strategies, particularly for studies aiming to interrogate B-cell dysregulation in immune-mediated diseases.
ER -