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Conventional and computational flow cytometry analyses reveal sustained human intrathymic T cell development from birth until puberty

Marieke Lavaert (UGent) , Brecht Valcke, Bart Vandekerckhove (UGent) , Georges Leclercq (UGent) , Kai Ling Liang (UGent) and Tom Taghon (UGent)
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Abstract
The thymus is the organ where subsets of mature T cells are generated which subsequently egress to function as central mediators in the immune system. While continuously generating T cells even into adulthood, the thymus does undergo involution during life. This is characterized by an initial rapid decrease in thymic cellularity during early life and by a second age-dependent decline in adulthood. The thymic cellularity of neonates remains low during the first month after birth and the tissue reaches a maximum in cellularity at 6 months of age. In order to study the effect that this first phase of thymic involution has on thymic immune subset frequencies, we performed multi-color flow cytometry on thymic samples collected from birth to 14 years of age. In consideration of the inherent limitations posed by conventional flow cytometry analysis, we established a novel computational analysis pipeline that is adapted from single-cell transcriptome sequencing data analysis. This allowed us to overcome technical effects by batch correction, analyze multiple samples simultaneously, limit computational cost by subsampling, and to rely on KNN-graphs for graph-based clustering. As a result, we successfully identified rare, distinct and gradually developing immune subsets within the human thymus tissues. Although the thymus undergoes early involution from infanthood onwards, our data suggests that this does not affect human T-cell development as we did not observe significant alterations in the proportions of T-lineage developmental intermediates from birth to puberty. Thus, in addition to providing an interesting novel strategy to analyze conventional flow cytometry data for the thymus, our work shows that the early phase of human thymic involution mainly limits the overall T cell output since no obvious changes in thymocyte subsets could be observed.
Keywords
human thymopoiesis, aging, computational flow cytometry, puberty, thymic involution, HEMATOPOIETIC STEM, BONE-MARROW, IN-VITRO, THYMUS, DIFFERENTIATION, REGENERATION, VISUALIZATION, PROGENITORS, EXPRESSION, DYNAMICS

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MLA
Lavaert, Marieke, et al. “Conventional and Computational Flow Cytometry Analyses Reveal Sustained Human Intrathymic T Cell Development from Birth until Puberty.” FRONTIERS IN IMMUNOLOGY, vol. 11, 2020, doi:10.3389/fimmu.2020.01659.
APA
Lavaert, M., Valcke, B., Vandekerckhove, B., Leclercq, G., Liang, K. L., & Taghon, T. (2020). Conventional and computational flow cytometry analyses reveal sustained human intrathymic T cell development from birth until puberty. FRONTIERS IN IMMUNOLOGY, 11. https://doi.org/10.3389/fimmu.2020.01659
Chicago author-date
Lavaert, Marieke, Brecht Valcke, Bart Vandekerckhove, Georges Leclercq, Kai Ling Liang, and Tom Taghon. 2020. “Conventional and Computational Flow Cytometry Analyses Reveal Sustained Human Intrathymic T Cell Development from Birth until Puberty.” FRONTIERS IN IMMUNOLOGY 11. https://doi.org/10.3389/fimmu.2020.01659.
Chicago author-date (all authors)
Lavaert, Marieke, Brecht Valcke, Bart Vandekerckhove, Georges Leclercq, Kai Ling Liang, and Tom Taghon. 2020. “Conventional and Computational Flow Cytometry Analyses Reveal Sustained Human Intrathymic T Cell Development from Birth until Puberty.” FRONTIERS IN IMMUNOLOGY 11. doi:10.3389/fimmu.2020.01659.
Vancouver
1.
Lavaert M, Valcke B, Vandekerckhove B, Leclercq G, Liang KL, Taghon T. Conventional and computational flow cytometry analyses reveal sustained human intrathymic T cell development from birth until puberty. FRONTIERS IN IMMUNOLOGY. 2020;11.
IEEE
[1]
M. Lavaert, B. Valcke, B. Vandekerckhove, G. Leclercq, K. L. Liang, and T. Taghon, “Conventional and computational flow cytometry analyses reveal sustained human intrathymic T cell development from birth until puberty,” FRONTIERS IN IMMUNOLOGY, vol. 11, 2020.
@article{8672252,
  abstract     = {{The thymus is the organ where subsets of mature T cells are generated which subsequently egress to function as central mediators in the immune system. While continuously generating T cells even into adulthood, the thymus does undergo involution during life. This is characterized by an initial rapid decrease in thymic cellularity during early life and by a second age-dependent decline in adulthood. The thymic cellularity of neonates remains low during the first month after birth and the tissue reaches a maximum in cellularity at 6 months of age. In order to study the effect that this first phase of thymic involution has on thymic immune subset frequencies, we performed multi-color flow cytometry on thymic samples collected from birth to 14 years of age. In consideration of the inherent limitations posed by conventional flow cytometry analysis, we established a novel computational analysis pipeline that is adapted from single-cell transcriptome sequencing data analysis. This allowed us to overcome technical effects by batch correction, analyze multiple samples simultaneously, limit computational cost by subsampling, and to rely on KNN-graphs for graph-based clustering. As a result, we successfully identified rare, distinct and gradually developing immune subsets within the human thymus tissues. Although the thymus undergoes early involution from infanthood onwards, our data suggests that this does not affect human T-cell development as we did not observe significant alterations in the proportions of T-lineage developmental intermediates from birth to puberty. Thus, in addition to providing an interesting novel strategy to analyze conventional flow cytometry data for the thymus, our work shows that the early phase of human thymic involution mainly limits the overall T cell output since no obvious changes in thymocyte subsets could be observed.}},
  articleno    = {{1659}},
  author       = {{Lavaert, Marieke and Valcke, Brecht and Vandekerckhove, Bart and Leclercq, Georges and Liang, Kai Ling and Taghon, Tom}},
  issn         = {{1664-3224}},
  journal      = {{FRONTIERS IN IMMUNOLOGY}},
  keywords     = {{human thymopoiesis,aging,computational flow cytometry,puberty,thymic involution,HEMATOPOIETIC STEM,BONE-MARROW,IN-VITRO,THYMUS,DIFFERENTIATION,REGENERATION,VISUALIZATION,PROGENITORS,EXPRESSION,DYNAMICS}},
  language     = {{eng}},
  pages        = {{13}},
  title        = {{Conventional and computational flow cytometry analyses reveal sustained human intrathymic T cell development from birth until puberty}},
  url          = {{http://dx.doi.org/10.3389/fimmu.2020.01659}},
  volume       = {{11}},
  year         = {{2020}},
}

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