Delineation of the Human Germinal Centre Immune Landscape Using Multiplex Imaging Analysis

Area of Science:

• Immunology
• Cell Biology
• Pathology

Background:

• Follicular immune dynamics, particularly germinal centers, are vital for antibody development.
• In situ characterization of these cells is crucial for understanding immune responses and diseases.

Purpose of the Study:

• To develop and validate a multiplex immunofluorescence imaging pipeline for analyzing human follicular immune cell subsets.
• To investigate the in situ phenotypic heterogeneity and localization of follicular helper CD4 T (T_FH) cells.
• To explore the functional heterogeneity of T_FH cells and compare immune cell densities between tonsils and lymph nodes.

Main Methods:

• Development of a multiplex immunofluorescence imaging pipeline.
• Analysis of human follicular adaptive and innate immune cell subsets in tonsils and reactive lymph nodes.
• Application of cell clustering analysis and multiplex RNAscope/protein imaging assays.

Main Results:

• Revealed in situ phenotypic heterogeneity and differential localization of T_FH cell subsets in tonsils and lymph nodes.
• Identified specific T_FH subsets with varying prevalence between tonsils and lymph nodes via clustering analysis.
• Demonstrated functional heterogeneity of T_FH cells using multiplex RNAscope/protein imaging.
• Found no significant differences in follicular innate immune cell densities between tonsils and lymph nodes.

Conclusions:

• A combinatory experimental approach provides comprehensive analysis of human follicular and germinal center immune dynamics.
• This pipeline can be utilized to further understand the pathogenesis of diseases like HIV and lymphomas.
• The study highlights the importance of characterizing T_FH cell heterogeneity for immunological research.