Bcl6 expression is associated with a distinct immune landscape and spatial transcriptome in COVID-19
Cloé Brenna, Bernat Bramon Mora, Kalliopi Ioannidou, Julien Bodelet, Mia L. Siebmanns, Simon Burgermeister, Spiros Georgakis, Michail Orfanakis, Yannick D. Muller, Nazanin Sédille, Matthew J. Feinstein, Jon W. Lomasney, Oliver Y. Chén, Giuseppe Pantaleo, Sabina Berezowska, Laurence de Leval, Raphael Gottardo, Constantinos Petrovas
Cloé Brenna, Bernat Bramon Mora, Kalliopi Ioannidou, Julien Bodelet, Mia L. Siebmanns, Simon Burgermeister, Spiros Georgakis, Michail Orfanakis, Yannick D. Muller, Nazanin Sédille, Matthew J. Feinstein, Jon W. Lomasney, Oliver Y. Chén, Giuseppe Pantaleo, Sabina Berezowska, Laurence de Leval, Raphael Gottardo, Constantinos Petrovas
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Research Article Immunology Infectious disease

Bcl6 expression is associated with a distinct immune landscape and spatial transcriptome in COVID-19

Abstract

The regulation of follicular (F) and germinal center (GC) immune reactivity in human lymph nodes (LNs), particularly during the acute stages of viral infection, remains poorly understood. We have analyzed lung-draining lymph nodes (LD-LNs) from COVID-19 autopsies using multiplex imaging and spatial transcriptomics to examine the immune landscape with respect to follicular immune reactivity. We identified 3 groups of donors based on the Bcl6 prevalence of their reactive follicles (RFs): RF-Bcl6no/lo, RF-Bcl6int, and RF-Bcl6hi. A distinct B/Tfh immune landscape, associated with increased prevalence of proliferating B cell and Tfh cell subsets, was found in RF-Bcl6hi LD-LNs. The comparison between LD-LNs and subdiaphragmatic (SD) LNs from the same donor revealed a divergent Bcl6 expression between the 2 anatomical sites. LD-LN Bcl6 expression was also associated with a distinct spatial transcriptomic profile. TH1-associated genes/pathways (e.g., CXCR3, STAT5, TNF signaling) were significantly upregulated in RF-Bcl6no/lo tissues, while the RF-Bcl6hi tissues exhibited significant upregulation of GC-promoting genes/pathways (e.g., CXCL13, B-cell receptor signaling). Our findings reveal a heterogeneous F/GC landscape in COVID-19 LD-LNs, highlighting specific molecular targets and pathways that could regulate human F/GC immune dynamics during acute viral infections.

Authors

Cloé Brenna, Bernat Bramon Mora, Kalliopi Ioannidou, Julien Bodelet, Mia L. Siebmanns, Simon Burgermeister, Spiros Georgakis, Michail Orfanakis, Yannick D. Muller, Nazanin Sédille, Matthew J. Feinstein, Jon W. Lomasney, Oliver Y. Chén, Giuseppe Pantaleo, Sabina Berezowska, Laurence de Leval, Raphael Gottardo, Constantinos Petrovas

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Figure 2

RF-Bcl6hi LD-LNs harbor higher densities of proliferating Bcl6hi and CD57hi Tfh cells.

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RF-Bcl6hi LD-LNs harbor higher densities of proliferating Bcl6hi and CD5...
(A) Representative fluorescence images of follicular areas from RF-Bcl6no/lo (Donor 2), RF-Bcl6int (Donor 16), and RF-Bcl6hi (Donor 17) tissues showing PD1hi (orange), CD57hi (yellow), Bcl6hi (cyan), Ki67hi (white), and merged images, highlighting Tfh cell subset differences. Scale bars: 100 μm. (B) Histocytometry gating scheme for CD4 T cell subset identification within CD20hi/dim follicular areas, with prevalence calculated for all follicles combined (All F). (C) Dot plots showing absolute counts of Tfh cell subsets (PD1hi, PD1hiCD57hi, PD1hiKi67hi, PD1hiKi67hiBcl6hi) per follicle from representative donors (RF-Bcl6no/lo, Donor 4; RF-Bcl6int, Donor 13; RF-Bcl6hi, Donor 22). (D) Dot plots of normalized Tfh cell subset densities (cells/μm²) — PD1hi, PD1hi Ki67hi, PD1hiCD57hiBcl6hi, and PD1hiKi67hiBcl6hi — measured in merged follicular areas from each donor. Groups are color-coded as follows: RF-Bcl6no/lo (purple), RF-Bcl6int (pink), RF-Bcl6hi (green), and HLNs from non-COVID-19–infected controls (blue). Light blue dot within the RF-Bcl6no/lo group corresponds to an axillary LN (Donor 25); yellow dot corresponds to para-aortic and mediastinal LNs (Donor 29); gray dots within the RF-Bcl6int group represent mediastinal LN (Donors 24-a and 24-b). (E) SQL clustering projection of B cell and Tfh cell subsets based on the expression of CD20hi/dim, CD20hi/dimKi67hi, CD20hi/dimKi67hiBcl6hi, CD4hi, PD1hi, PD1hiKi67hi, PD1hiCD57hi, and PD1hiKi67hiBcl6hi. Data points are color-coded by RF group: RF-Bcl6no/lo (purple), RF-Bcl6int (pink), and RF-Bcl6hi (green). HLNs from non-COVID-19–infected control donors are shown as blue diamonds. The projection displays SQL Factor 1 versus SQL Factor 3, which capture the main variance across immune cell phenotypes and best separate the groups. Mann-Whitney U test; *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. Raw P values are shown on the graph, and both raw and FDR-adjusted P values are reported in Supplemental Table 11.