Extended Data Fig. 2: TCB-triggered T-cell infiltration is not mediated by CD103-E-cadherin interactions.
a and b, Chromogenic staining (brown) with according quantification of tight junctions (ZO-1; nOrgZO-1 = 59, nTumZO-1 = 58) and adherent junctions (e-Cadherin; nOrgZO-1 = 31, nTumZO-1 = 84) in organoids and tumouroids. 20x magnification, scale: 100 µm. Statistical analysis was conducted by an unpaired t-test (two-tailed) and was defined as ****p < 0.0001. Red line displays mean. c, Representative images at 20x magnification of organoids treated with EpCAM and CEA(hi) TCB (1 µg/mL) at 48 h. Top row displays TCBs only, bottom row is co-treated with an αE(CD103)β7-inhibitor cocktail (Etrolizumab (10 µg/mL) and an anti-Integrin β7 monoclonal antibody (10 µg/mL)). E-cadherin+ organoids (magenta) surrounded by CD4+ (orange), CD8+ (turquoise) and CEA+ (yellow) immune cells. Caspase-3 (green) captures TCB-triggered immune-induced apoptosis, nuclei are stained with DAPI (blue). Scale: 100 µm. d, Quantification of caspase+ area per organoid across different TCB treatments (48 h) with (+) and without (-) an αE(CD103)β7-inhibitor cocktail, plotted as boxplot, whiskers showing all points (min to max; nNT TCB = 5; nEpCAM TCB = 3; nCEA TCB = 3). Statistical analysis was conducted by an unpaired t-test (two-tailed) and was found to be non-significant (ns). e, Quantification of T cell infiltration per organoid area across different TCB treatments (48 h) with (+) and without (-) an αE(CD103)β7-inhibitor cocktail, plotted as boxplot, whiskers showing all points (min to max; nNT TCB = 4; nEpCAM TCB = 3; nCEA TCB = 3). Statistical analysis was performed by unpaired t-tests (two-tailed) and was defined as **p < 0.01 and non-significant (ns) (ns). f, FACS staining of CD103 on organoid and tumouroid co-cultured with PBMCs, treated with EpCAM TCB for 72 h. Data are presented as mean values ∓ SD (n = 3). Statistical analysis was conducted by an unpaired t-test (two-tailed) and was found to be non-significant (ns). g, Chromogenic DAB staining of CD103 (brown) in healthy small intestine tissue and different organoid-immune co-cultures, scanned at 20x. Scale: 100 µm. CD103+ immune cells only found in native SI tissue and expressed by tissue-resident memory T cells (TRMs). The displayed experiment in this figure was performed once.
