Atypical memory B cell clonal expansion and inflammatory programs associate with platelet-activating antibody development in COVID-19
Nathan Witman, Mei Yu, Yuqi Zhang, Kexin Gai, Yuhong Chen, Lu Zhou, Christine Nguyen, Wen Zhu, Yongwei Zheng, Shawn Jobe, Mary Beth Graham, Weiguo Cui, Demin Wang, Renren Wen
Nathan Witman, Mei Yu, Yuqi Zhang, Kexin Gai, Yuhong Chen, Lu Zhou, Christine Nguyen, Wen Zhu, Yongwei Zheng, Shawn Jobe, Mary Beth Graham, Weiguo Cui, Demin Wang, Renren Wen
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Research Article Immunology Vascular biology

Atypical memory B cell clonal expansion and inflammatory programs associate with platelet-activating antibody development in COVID-19

Abstract

Patients with COVID-19 who develop platelet-activating antibodies represent a subset at heightened thrombotic risk, yet the immune features associated with this response remains to be defined. We applied single-cell RNA-seq of B and T cells, single B cell V(D)J-seq, and plasma cytokine and chemokine analysis to define immune signatures distinguishing patients who did (PEA+) or did not (PEA–) develop these antibodies. Patients positive for PEA showed prominent transcriptional enrichment of inflammatory, antigen presentation, and B cell receptor signaling pathways within antigen-experienced B cell subsets. Expanded B cell clones in patients positive for PEA were disproportionately enriched within atypical memory B cells and exhibited upregulated IFN-γ–response signatures, increased proliferative mutational patterns, limited class switching, and a significant overrepresentation of RKH/Y5 heavy-chain motifs associated with platelet-activating antibodies, consistent with an extrafollicular-biased response. Parallel T cell profiling revealed IL-12 pathway enrichment across most T cell subsets, increased IFN-γ transcription, and elevated plasma levels of Th1-associated cytokines in patients positive for PEA. Collectively, these data highlight a coordinated inflammatory environment marked by Th1-skewed T cell activation and selective expansion of atypical memory B cell clones carrying RKH/Y5 motifs, defining immunologic features associated with platelet-activating antibody development in COVID-19.

Authors

Nathan Witman, Mei Yu, Yuqi Zhang, Kexin Gai, Yuhong Chen, Lu Zhou, Christine Nguyen, Wen Zhu, Yongwei Zheng, Shawn Jobe, Mary Beth Graham, Weiguo Cui, Demin Wang, Renren Wen

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Figure 1

Comparable B cell clusters and subpopulations in PEA+ and PEA– patients.

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Comparable B cell clusters and subpopulations in PEA+ and PEA– patients....
CD19+ B cells were sorted by FACS from PBMCs of 6 PEA+ patients and 5 PEA– patients, followed by scRNA-seq and scV(D)J-seq. (A) Schematic overview of patient selection and the scRNA-seq workflow. (B) UMAP visualization of CD19+ B cell clusters by gene expression. UMAP visualization of all CD19+ B cells (n = 29,505), colored by 6 identified clusters based on 5′ gene expression profiles and annotated according to B cell subset identities. (C) Dot plot showing the expression of key marker genes used to validate cluster annotations. (D) PEA status–specific UMAP of CD19+ B cells. UMAP plots from B split by PEA status, showing CD19+ B cells from PEA+ (n = 14,414) and PEA– (n = 15,019) patients. (E) Comparison of B cell clusters between PEA+ and PEA– patients represented as percentage of total CD19+ cells. Statistical analysis was performed using global multinomial logistic regression comparisons. (F) Representative flow cytometry gating strategy for B cell subsets between PEA+, PEA–, and healthy donor (HD) groups. Data represent a single experiment. (G) Flow cytometry analysis of B cell subsets between PEA+ (n = 6), PEA– (n = 5), and HD (n = 5). Percents are presented of their parent populations gated from F. Statistical comparisons were generated with Wilcoxon rank-sum test with Bonferroni correction; *P < 0.05.