Fig 1: Calcipotriol-mediated mammary tumor suppression and IL-24 induction rely on TSLP and CD4+ T cells.(A) Schematic diagram outlining the experimental approach to assess the mechanism of topical calcipotriol treatment in inhibiting mammary tumor growth. WT and TslprKO mice served as tumor recipients, which were treated with anti-CD4 Ab (αCD4 Ab) or IgG control every 5 days starting 1 day before tumor cell implantation. Two days after orthotopic PyMt mammary tumor cell implantation, mice were topically treated with 20 nmol calcipotriol or EtOH every 2 days. (B and C) PyMt mammary tumor growth in WT and TslprKO mice subjected to calcipotriol versus EtOH treatments, with or without CD4+ T cell depletion, shown as (B) mean tumor volumes + SD and (C) spider plot. WT + EtOH + IgG (n = 6), WT + calcipotriol + IgG (n = 7), WT + calcipotriol + αCD4 Ab (n = 7), and TslprKO + calcipotriol + IgG (n = 7). (D) Representative IF images of mammary tumors stained for CD4 and IL-24. (E and F) Quantification of (E) CD4+ T cells and (F) IL-24+ cells in WT mammary tumors treated with EtOH + IgG, calcipotriol + IgG, or calcipotriol + αCD4 Ab. Each dot represents a high-power field (HPF) image. Three HPF images from 5 tumors are included in each group. (G) Correlation between the presence of CD4+ T cells and IL-24+ cells in WT mammary tumors. Bar graphs show mean + SD, 1-way ANOVA (E and F), 2-way ANOVA (B and C), and Spearman’s rank correlation analysis (G). Scale bar: 100 μm (D).
Fig 2: Topical calcipotriol treatment inhibits PyMt mammary tumor growth in WT mice.(A) Schematic diagram outlining the experimental design used to test the efficacy of topical calcipotriol treatment in blocking mammary tumor growth. Six- to eight-week-old WT mice were used as tumor recipients. The animals were treated with 20 nmol calcipotriol or EtOH topically every 2 days, starting at 2 days after orthotopic PyMt mammary tumor cell implantation. (B) TSLP levels in the circulation after topical calcipotriol versus EtOH treatment. (C–E) Topical calcipotriol treatment effect on PyMt mammary tumor growth shown as (C) representative macroscopic images of the mammary tumors at the endpoint (circles highlight the tumors), (D) mean tumor volumes + SD, and (E) spider plot of tumor volume over time (n = 10 in each group). (F–J) Tumor-infiltrating CD3+ T cells, CD4+ T cells, and CD8+ T cells shown as (F) representative immunofluorescence (IF) images of CD3-, CD4-, and CD8-stained tumor tissues; (G) percentage of CD3+ T cells of total cells; (H) percentage of CD4+ T cells of total cells; (I) percentage of CD8+ T cells of total cells; and (J) The ratio of CD4+ T to CD8+ T cells in calcipotriol- and EtOH-treated mammary tumors (n = 8 in each group). Bar graphs show mean + SD. Mann-Whitney U test (B and G–J) and 2-way ANOVA (D and E). Scale bars: 1 cm (C); 100 μm (F).
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