Figure 3.

Immune preconditioning modulates the composition of the thyroidal immune cell infiltrate, treatment with ICIs does not. UMAP plot representing the gene expression profile of CD45⁺ cells isolated from the thyroids of animals with concomitant IET (A–D) or pre-existing IET (E–H). The thyroids from two animals for each experimental condition were digested and CD45⁺ cells were sorted by flow cytometry. Cells from each experimental condition were labeled with hashtag antibodies and pooled into a single reaction/sequencing library per experimental arm. Immune cell types were identified by unsupervised graph-based clustering integrated with biological knowledge. Six different immune cell types were identified: B cells, dendritic cells, macrophages, neutrophils, stromal cells, and T cells. A small fraction of CD45⁺ cells did not fall into any of these six categories and was labeled as unassigned or N/A. The relative representation and the global gene expression profile of the 6 different immune cell types identified appeared to be markedly different in “Control concomitant IET” and “Control pre-existing IET” (A vs E). However, within each experimental arm, treatment with the different tested ICIs did not appear to induce marked changes in the relative representation or in the global gene expression profile of any of the identified immune cell types (i.e., plot A is similar to plots B–D while plot E is similar to plots F–H). ICI, iodine exacerbated thyroiditis; IET, iodine exacerbated thyroiditis; N/A, not available; UMAP, uniform manifold approximation and projection.PD-1, programmed death-1; LAG-3, lymphocyte activation gene 3; TIM-3, T-cell immunoglobulin and mucin domain 3.