T cell exhaustion markers and populations
As there is no clear consensus, and just little information about the
specific marker combination that describes specific subpopulations of
exhausted T cells in patients with SLE, we initially used an
unsupervised approach to explore the behavior and clustering of surface
markers and transcription factors that have been reported as relevant in
exhausted T cells (see methods for the complete list of markers
included). As a first step, we generated tSNE maps using concatenated
files with an equal number of CD3+ cells from all the patients included
in each group. Figures 1A and 1B show the tSNE plots for each group, and
the distribution of the included markers across the plots. In order to
identify the clustering of these markers in subpopulations, we used a
self-organizing maps algorithm [11]. We could identify five
subpopulations of CD8+, seven of CD4+ cells and three CD3+CD4-CD8-. The
heatmaps and tables showing the relative expression of markers in each
cluster across all the identified subpopulations are shown in figure 1C.
Overlapping of these populations on the tSNE plots are shown in 1D.
Histograms represent the relative expression of a CD3+ CD8+ cell
population that seems to be exclusive to the PR-SLE group, in both in
panel II and III (2.86% and 3.75% respectively) and which expresses
significantly transcription factors and surface markers associated with
senescence and depletion.
Populations differentially represented in the PR when compared
with the Act-SLE group .
Within the CD4+ compartment, a population of Tbet+CD45RO+ cells
identified in the unsupervised analysis, which represents 10.20%
(1.74–30.50) of the CD4+ cells in the PR group, was significantly
increased compared with the Act-SLE (1.68, IQR: 0.42–2.83) and HS
(4.08, IQR: 1.64–5.48). The CD4+EOMES+ cells were also increased in the
PR group compared with Act-SLE and HS. The other population that was
significantly different between PR and Act-SLE was the CD4+Tbet+ cells,
which were increased in the PR group, and similar to the HS.
Within the CD8+ T cells, the proportion of PD1+ cells was apparently
increased only in the PR group, even though the difference was only
significant when compared with the HS. The CD8+2B4+ cells, identified
through the unsupervised clustering, were increased in the PR vs Act-SLE
group. The most striking difference was observed in the EOMES+ cells,
with a median of 37.6 (24.9–53.2) in the PR group, vs 12.9 (6.78–24.8)
in the HS and 8.13 (2.33–20.5) in the Act-SLE groups.