Figure 5: MAIT cell neighborhood analysis identifies the cellular interaction network underlying the immunosuppressive MAIT cell niche.

(A) Schematic of S³-CIMA (Supervised Spatial Single Cell Imaging Analysis) algorithm used on CODEX single-cell data to investigate the MAIT cell neighborhoods in different tissue regions.

(B) Boxplots showing frequencies of selected cells from different tissue regions compared with randomly selected cells in the background.

(C) Bubble plot displaying S³-CIMA classification of cell types in whole tissue sections with neighborhood size k=40 nearest neighboring cells. Y-Axis shows the ratio of the number of all selected cell type CT in nearest neighbor of the anchor cell $\left(𝒦n{n}_{CT}^S:\right)$ to the number of all cell type CT in nearest neighbor of the anchor cell$\left(𝒦n{n}_{CT}\right)$. X-axis shows the ratio of the Number of all selected cell type CT $\left({𝒦}_{CT}^S:\right)$ to the number of all cells of cell type CT ${𝒦}_{CT}$. Color-coding corresponds to the enrichment score (ES) as displayed in (C). Size of the bubble displays the ratio: number of all selected cell type CT in nearest neighbor of the anchor cell $\left(𝒦n{n}_{CT}^S\right)$ to the number of all selected cells in nearest neighbor of the anchor cell $𝒦n{n}^S$ or $\frac{𝒦n{n}_{CT}^S}{𝒦n{n}^S}$

(D) Waterfall plot displaying enrichment score of indicated cell populations in the MAIT cell neighborhood (data for adjacent liver is shown) as selected by S³-CIMA. Values >1 indicate specific enrichment in the MAIT cell niche.

(E) Five-color overlay images of a CODEX datasets displaying interactions between PD-L1⁺ (turquoise) CD163⁺ (pink) macrophages and PD-1 (yellow) on MAIT cells (TCRVa7.2^+, red) in the adjacent liver. Examples for 4 different patient samples (are shown.

(F) Mean expression of PD-L1 on selected cell populations in the normal liver tissue region as determined by CODEX imaging.

(G) Representative histograms showing M2 polarization of TAMs (defined as CD45⁺CD68⁺) as determined by expression of CD163 (left) and PD-L1 (right) on macrophages from healthy PBMCs (grey), HCC patient PBMCs (black), and adjacent, non-tumor tissue (red).