Extended Data Fig. 8: Sucralose treatment in mice reduces antigen-specifc CD8⁺ T cell responses.

a) Representative density plot of intratumoral CD8⁺ T cells recognizing the MHC-peptide tetramer complex (K^b-OVA) from EL4^OVA tumours from mice fed water or 0.72 μg ml⁻¹ Scrl 10 days post challenge. (b, c) Mice fed water or 0.17 μg ml⁻¹ Scrl . N = 8 biological replicates per group and each dot represents an individual mouse b) Intratumoral frequency of CD8⁺ T cells recognizing the MHC-peptide tetramer complex (K^b-OVA) from EL4^OVA tumours. c) Intratumoral frequency of CD8⁺IFNγ⁺ T cells from tumour bearing mice post re-stimulation with the OVA peptide. d) Cytotoxic T cell assay of OT-I T cells activated with OVA-peptide either in TCM with (Scrl) or without (Ctrl) 0.5 mM Scrl followed by co-culture with EL4^OVA cancer cells. N = 5 for technical replicates. (e–h) Splenocytes from C57BL/6J mice fed water or 0.72 mg ml⁻¹ Scrl followed by bacterial challenge with LmOVA (10⁵ CFU permouse). N = 7 biological replicates per group. e) Total splenocyte numbers 7 days post infection. f) Representative density plot of OVA-specific CD8⁺ T cells identified by K^b-OVA and CD8⁺ surface staining. g) Frequency of CD8⁺ T cells re-stimulated with OVA-peptide and analyzed for IFNγ expression. h) Absolute numbers of splenic CD8⁺ T cells in Extended Data Fig. 8g. i) The bacterial load (colony forming units per mg of tissue) of the liver (left) and spleen (right) at day 3 post LmOVA (10⁵ CFU per mouse) infection of mice fed water or 0.72 mg ml⁻¹ Scrl; n = 6 (spleen) or 7 (liver) of biological replicates per group. j) Absolute cell number of Jurkat T cells grown in media with or without Scrl as indicated at day 5. N = 3 technical replicates. k) Absolute splenic numbers of CD8⁺ T cells producing IFNγ and Granzyme B (GZMB) in response to OVA-peptide re-stimulation of mice infected with LmOVA (10⁵ CFU per mouse) 7 days post infection. Mice were fed water (n = 7), 0.72 mg ml⁻¹ Scrl for 2 weeks (n = 6) or 0.72 mg ml⁻¹ Scrl treatment for 2 weeks followed by one-week water washout (n = 7, Scrl off). Data presented as mean value ± s.e.m. (b-e, g-k). Each dot represents a sample derived from an individual mouse (b, c, e, g, h, i, k). Statistical significance was tested using single or multiple unpaired 2-tailed Student’s t test (b-e, g-i) or a One-way ANOVA with a Tukey’s Multiple Comparison ( j, k). Data representative of 2 (d, e, i, k) or 3 ( j) independent experiments.

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