Single-cell landscape analysis unravels molecular programming of the human B cell compartment in chronic GVHD
Jonathan C. Poe, Jiyuan Fang, Dadong Zhang, Marissa R. Lee, Rachel A. DiCioccio, Hsuan Su, Xiaodi Qin, Jennifer Y. Zhang, Jonathan Visentin, Sonali J. Bracken, Vincent T. Ho, Kathy S. Wang, Jeremy J. Rose, Steven Z. Pavletic, Frances T. Hakim, Wei Jia, Amy N. Suthers, Itaevia M. Curry-Chisolm, Mitchell E. Horwitz, David A. Rizzieri, William C. McManigle, Nelson J. Chao, Adela R. Cardones, Jichun Xie, Kouros Owzar, Stefanie Sarantopoulos
Jonathan C. Poe, Jiyuan Fang, Dadong Zhang, Marissa R. Lee, Rachel A. DiCioccio, Hsuan Su, Xiaodi Qin, Jennifer Y. Zhang, Jonathan Visentin, Sonali J. Bracken, Vincent T. Ho, Kathy S. Wang, Jeremy J. Rose, Steven Z. Pavletic, Frances T. Hakim, Wei Jia, Amy N. Suthers, Itaevia M. Curry-Chisolm, Mitchell E. Horwitz, David A. Rizzieri, William C. McManigle, Nelson J. Chao, Adela R. Cardones, Jichun Xie, Kouros Owzar, Stefanie Sarantopoulos
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Resource and Technical Advance Immunology

Single-cell landscape analysis unravels molecular programming of the human B cell compartment in chronic GVHD

Abstract

Alloreactivity can drive autoimmune syndromes. After allogeneic hematopoietic stem cell transplantation (allo-HCT), chronic graft-versus-host disease (cGVHD), a B cell–associated autoimmune-like syndrome, commonly occurs. Because donor-derived B cells continually develop under selective pressure from host alloantigens, aberrant B cell receptor (BCR) activation and IgG production can emerge and contribute to cGVHD pathobiology. To better understand molecular programing of B cells in allo-HCT, we performed scRNA-Seq analysis on high numbers of purified B cells from patients. An unsupervised analysis revealed 10 clusters, distinguishable by signature genes for maturation, activation, and memory. Within the memory B cell compartment, we found striking transcriptional differences in allo-HCT patients compared with healthy or infected individuals, including potentially pathogenic atypical B cells (ABCs) that were expanded in active cGVHD. To identify intrinsic alterations in potentially pathological B cells, we interrogated all clusters for differentially expressed genes (DEGs) in active cGVHD versus patients who never had signs of immune tolerance loss (no cGVHD). Active cGVHD DEGs occurred in both naive and BCR-activated B cell clusters. Remarkably, some DEGs occurred across most clusters, suggesting common molecular programs that may promote B cell plasticity. Our study of human allo-HCT and cGVHD provides understanding of altered B cell memory during chronic alloantigen stimulation.

Authors

Jonathan C. Poe, Jiyuan Fang, Dadong Zhang, Marissa R. Lee, Rachel A. DiCioccio, Hsuan Su, Xiaodi Qin, Jennifer Y. Zhang, Jonathan Visentin, Sonali J. Bracken, Vincent T. Ho, Kathy S. Wang, Jeremy J. Rose, Steven Z. Pavletic, Frances T. Hakim, Wei Jia, Amy N. Suthers, Itaevia M. Curry-Chisolm, Mitchell E. Horwitz, David A. Rizzieri, William C. McManigle, Nelson J. Chao, Adela R. Cardones, Jichun Xie, Kouros Owzar, Stefanie Sarantopoulos

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Figure 5

Cluster 5 contains both typical CD27+ memory B cells and ABC subsets, with the latter expanded in active cGVHD.

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Cluster 5 contains both typical CD27+ memory B cells and ABC subsets, wi...
(A) B cells expressing CD27 or ITGAX transcripts are enriched in cluster 5 in 2 separate regions. Shown are UMAP density plots from the scRNA-Seq data set displaying relative transcript density for the gene indicated, at the single-cell level. The dashed line approximates the perimeter of cluster 5 (enlarged for clarity). Gray arrows indicate a region enriched for CD27-expressing B cells, and black arrows indicate a region enriched for ITGAX-expressing B cells. (B and C) Flow cytometric analysis to identify ABCs, CD27+ memory B cells, and CD11c+CD27+ memory B cells in PBMC samples from allo-HCT patients with active cGVHD (n = 6) or no cGVHD (n = 6) or from HDs (n = 4). PBMCs were pregated on live (7-AAD–) CD19+ B cells. Dot plots in B show representative individuals from each group. In C, results from all groups for the CD11c+CD27– population, CD11c+CD27+ (DP) population, and CD11c–CD27+ population (as gated in B) are represented. Statistical comparison: 1-way ANOVA with Tukey’s multiple comparisons test (GraphPad Prism 9 software; *, P < 0.05; **, P < 0.01; ***, P < 0.001; error bars represent mean ± SEM). (D and E) DN2 B cells are expanded in allo-HCT patients with active cGVHD. (D) Representative FACS plots from PBMCs of allo-HCT patients with active or no cGVHD, and from an HD, generated after first gating on viable (7-AAD–), CD19+ B cells, then CD11c+CD21– cells (upper panel), followed by DN2 B cell identification in all groups (CD11c+CD21–CD27–IgD–), as depicted in magenta for active cGVHD. (E) Statistical comparison between groups (active cGVHD, n = 6; no cGVHD n = 6, HDs, n = 4) was performed using a 1-way ANOVA with Tukey’s multiple comparisons test (GraphPad Prism 9 software; *, P < 0.05; **, P < 0.01; ***, P < 0.001; error bars represent mean ± SEM).