Isolation and characterisation of PR3-specific B cells and their immunoglobulin sequences

Area of Science:

• Immunology
• Autoimmunity

Background:

• Anti-neutrophil autoantibodies, particularly anti-proteinase 3 (PR3) autoantibodies, are critical biomarkers for diagnosing and monitoring granulomatosis with polyangiitis (GPA).
• Despite their diagnostic importance, the specific B cell receptor sequences encoding PR3 autoantibodies have remained largely uncharacterized.

Purpose of the Study:

• To isolate and functionally characterize B cells that specifically recognize PR3 from the peripheral blood of patients with established PR3 autoantibodies.
• To gain insights into the B cell repertoire involved in PR3 autoimmunity.

Main Methods:

• Peripheral blood mononuclear cells (PBMCs) from seven GPA patients with PR3 autoantibodies were isolated.
• PR3-binding B cells were identified using PR3 staining, followed by single-cell sorting.
• Transcriptome sequencing and subsequent expression of immunoglobulin genes allowed for antibody generation and specificity testing via ELISA.

Main Results:

• Nineteen PR3-specific B cells were identified at a low frequency (0.0075%) in the peripheral blood of a single patient.
• These B cells were polyclonal, predominantly IgG4+, and exhibited specific features including lambda light chain pairing, IGHJ6 gene usage, and distinct CDRH3 lengths.
• The identified B cells showed limited somatic hypermutation and variable reduction in PR3 binding upon reversion to their germline configuration.

Conclusions:

• The study demonstrates the feasibility, albeit challenging, of identifying PR3-specific B cells in the peripheral circulation of GPA patients.
• The characteristics of these B cells suggest that PR3-self reactivity might be initiated early in the B cell development pathway.