Figure 4.

In situ assessment of T-cell density and IC expression profile using immunohistochemistry in the IC^high/low subgroups of CRC identified via flow cytometry. (A) Representative CD3 (left) and CD8 (right) immunostaining of an MSS CRC with an IC^high expression profile. Stained cells appear in brown within tumor glands and in the surrounding stroma; nuclei are counterstained in blue. Bars indicate 100 μm. (B) Box plots in the lower panels indicate the percentages of CD3⁺ or CD8⁺ TILs counted inside tumor glands (intraepithelial TILs) or in the peritumoral stroma using QuPath software, according to IC^low or IC^high subgroups; Mann–Whitney test. MSS tumors are shown in pale green and MSI tumors in dark blue. (C) Representative immunostaining of PD-1, TIGIT and CD94 of the same MSS CRC with an IC^high expression profile, as in (A). Numerous TIGIT⁺ TILs can be seen within the tumor and in the stroma, and only a few PD-1⁺ cells and CD94⁺ TILs. Bars indicate 100 μm. (D) Box plots recapitulate the percentages of cells expressing PD-1, TIGIT or CD94, counted inside tumor glands (intraepithelial TILs) or in the peritumoral stroma using QuPath software, according to IC^low vs. IC^high subgroups; Mann–Whitney test (* p < 0.05; ** p < 0.01). MSS tumors are shown in pale green and MSI tumors in dark blue.