in situ differentiation of B cells cross-reactive to Epstein-Barr virus and autoantigens in hidradenitis suppurativa Public

Abstract

Hidradenitis suppurativa (HS) is a chronic inflammatory skin disease characterized by painful nodules, abscesses, and tunnels involving intertriginous skin. HS negatively impacts the mental wellbeing and economic and educational standing of sufferers. HS is not uncommon with an estimated prevalence of 0.1% in the United States. Prevalence is highest among Black women. While the etiology of HS remains unknown, emerging evidence suggests an autoimmune component. B cells are known to infiltrate affected skin and the presence of autoantibodies correlates with disease severity. To understand the B cell contribution to pathogenesis, we conducted single cell transcriptomics and repertoire analysis on B cells and plasma cells from HS surgical excisions and peripheral blood. We found that infiltrating B cells exhibit a chronic activation phenotype and contribute to the inflammatory milieu of the skin by producing TNFa. A large subset of infiltrating B cells has immunosuppressive potential, producing anti-inflammatory TGBb. Infiltrating B cells undergo antigen-driven in situ clonal expansion and plasma cell differentiation, which is associated with inflammatory cytokine production. A serological screen revealed increased reactivity to Epstein-Barr virus (EBV) epitopes in HS patients compared to healthy controls. Subsequently, we identified clonally expanded and differentiated autoreactive B cells that cross-reacted with EBV protein EBNA1, and autoantigens DNA topoisomerase I (SCL-70) and P21 activated kinase 4 (PAK4). Our findings suggest that chronic activation, differentiation, and clonal expansion of autoreactive B cells contribute to HS pathogenesis and that EBV reactivation may be a factor in HS etiology.

Table of Contents

Table of Contents

 

Preface: A History Lesson........................................................................................................... 3

Chapter 1: Introduction............................................................................................................... 5

Self-tolerance................................................................................................................... 5

Autoimmune Disease Etiology.................................................................................. 10

Pathogenic Mechanisms and Treatment Strategies............................................. 16

Epstein-Barr Virus and Autoimmune Disease...................................................... 22

Hidradenitis Suppurativa........................................................................................... 27

Works Cited.................................................................................................................... 31

Chapter 2: in situ differentiation of B cells autoreactive to Epstein-Barr virus and autoantigens in hidradenitis suppurativa 35

Abstract........................................................................................................................... 35

Graphical Abstract....................................................................................................... 36

Introduction................................................................................................................... 37

Results............................................................................................................................. 39

Transitional B cell populations are present in HS skin................................................. 39

B cells and ASC populations in HS skin are transcriptionally distinct from peripheral counterparts 41

Infiltrating B cells undergo in situ plasma cell differentiation and have immunomodulatory potential            43

B cells and ASC in the skin are class-switched and clonally expanded........................... 46

Class-switched and mutate infiltrating B cells are preferentially selected for differentiation                47

Expanded and differentiated clones from HS skin are cross-reactive to EBV EBNA1, human SCL-70 and PAK4         49

Discussion....................................................................................................................... 51

Methods.......................................................................................................................... 56

Tables.............................................................................................................................. 63

Table 1: Cluster classification..................................................................................... 63

Table 2: Clinical characteristics of study participants.................................................. 64

Figures............................................................................................................................ 65

Figure 1: HS skin and peripheral B cells form clusters based on differential gene expression               65

Figure 2: B cells in HS skin express markers and transcription factors associated with chronic activation     66

Figure 3: RNA velocity demonstrates B cell to plasma cell differentiation and cytokine production in HS skin             67

Figure 4: HS skin B cells are class-switched and clonally expanded.............................. 68

Figure 5: Expanded clones in the skin are highly mutated, intra-clonally homogenous, and are primarily of a single isotype 69

Figure 6: Proteome-wide screening reveals clonally expanded clones with cross-reactivity to EBV and autoantigens                 71

Supplemental Figures................................................................................................. 72

Supplemental figure 1: scRNA-seq quality control process........................................... 72

Supplemental figure 2: Expression of genes of interest................................................ 74

Supplemental figure 3: Differential gene expression between skin compartment clusters 75

Supplemental figure 4: Chemokine receptor and ligand expression.............................. 76

Supplemental figure 5: RNA velocity estimation with UniTVelo................................... 77

Supplemental figure 6: Splice dynamics in RNA velocity analysis................................. 78

Supplemental figure 7: Comparison of TNFa+ and TFBb+ skin B cells......................... 79

Supplemental figure 8: Comparison of VirScan PhIP-seq reactivity of MS patients and HS epitope spread   80

Supplemental figure 9: Control monoclonal antibody reactivity in HuProt antigen screen 81

Works cited.................................................................................................................... 82

Chapter 3: Discussion................................................................................................................ 87

About this Dissertation

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School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Immunology and Molecular Pathogenesis
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Health Sciences, Immunology
Mot-clé	hidradenitis suppurativa autoimmunity epstein-barr virus skin immunology
Committee Chair / Thesis Advisor	Joshy Jacob, Emory University
Committee Members	Frances Eun-Hyung Lee, Emory University Jeremy Boss, Emory University Eliver Ghosn, Emory University Max Cooper, Emory University

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