Fig. 4: GrK⁺CD103^–CD8⁺ Trm cells induce neuronal alterations by engaging PAR-1.

a Graphical overview of the experimental design. Created in BioRender. Terrabuio, E. (2025) https://BioRender.com/ir5h5rf. b Immunofluorescence staining showing GrK granules inside CD103⁺ and CD103^– CD8⁺ Trm cells. Scale bar = 3 µm. c Intracellular Ca²⁺ release in neurons co-cultured with CD103⁺ or CD103^– CD8⁺ Trm cells. The last time point is shown in the right panel. Data are means ± SD from four independent experiments. P-values based on two-way ANOVA-multiple comparisons. Ctrl^– = neurons alone. Ctrl⁺ = ionomycin-stimulated neurons (10 µM). Source data are provided as a Source Data file. d Intracellular Ca²⁺ release in neurons treated with purified active GrK or vehicle. The last time point is shown in the right panel. Data are means ± SD from three independent experiments. P-values based on two-way ANOVA-multiple comparisons. Source data are provided as a Source Data file. e Intracellular Ca²⁺ release in neurons co-cultured for 5 h with CD103^–CD8⁺ Trm cells in the presence/absence of the PAR-1 inhibitor SCH79797 (100 nM). The last time point (300 min) is shown in the right panel. Data are means ± SD from four independent experiments. P-values based on two-way ANOVA-multiple comparisons. Ctrl^– = neurons alone. Ctrl⁺ = ionomycin-stimulated neurons (10 µM). Source data are provided as a Source Data file. f, g Intracellular Ca²⁺ release in neurons cultured for 5 h with purified active GrK alone (150 nM) or with SCH79797 (50 nM, or 100 nM). Ctrl^– = neurons alone. The last time point is shown in the right panel. Data are means ± SD from three independent experiments. P-values based on two-way ANOVA-multiple comparisons. Representative images are shown in (g). Red = intracellular Ca²⁺ release. Scale bar = 20 µm. Source data are provided as a Source Data file. h Immunofluorescence microscopy showing GrK⁺CD103^–CD8⁺ Trm cells near the soma (cell I) and dendrites (cell II) of a PAR-1⁺ neuron. Cell morphology was visualized by wide-field imaging using a DIC filter. Scale bar = 5 µm (or 2 µm for zoomed images I, and II).