Fig. 2: TRF reduces Py230 mammary tumor growth in two obese postmenopausal mouse models.

a, d Py230 tumor volume over time in OVX or VCD mice on NC, AL, and TRF (number of mice n = 8 for NC, n = 10 for AL, n = 10 for TRF in a; n = 7 for NC, n = 7 for AL, n = 7 for TRF in d; **p < 0.01, ***p < 0.001, ****p < 0.0001 by two-way ANOVA with Tukey’s multiple comparisons test). b, e Individual tumor weights at the end of the study in OVX or VCD mice (number of tumors n = 32 for NC, n = 40 for AL, n = 40 for TRF in a; n = 27 for NC, n = 28 for AL, n = 32 for TRF in d; ****p < 0.0001 by one-way ANOVA with Tukey’s multiple comparisons test). c, f Correlation plot of mean tumor weight per mouse vs. fasting insulin in OVX or VCD mice (for c, number of mice n = 6 for NC, n = 7 for AL, n = 8 for TRF; for f, n = 7 for NC, n = 7 for AL, n = 8 for TRF). Data were analyzed by linear regression using Pearson correlation. g Representative H&E, Ki67 and CD31 staining of Py230 tumor tissue in OVX mice on NC, AL, and TRF. Scale bar, 50 µm. Insets in H&E images show higher magnification of a mitotic cell. Data shown are representative of two independent experiments. h–j Quantification of mitoses (n = 27 for NC, n = 38 for AL, n = 36 for TRF), Ki67-positive cells (n = 31 for NC, n = 36 for AL, n = 32 for TRF), and CD31 staining (n = 27 for NC, n = 27 for AL, n = 21 for TRF). Data are presented as mean ± SEM per high-powered field (hpf) or 5hpf for mitoses, with 4 hpf per section (n = number of tumors). Asterisks indicate significance; *p < 0.05, ****p < 0.0001 by one-way ANOVA with Tukey’s multiple comparisons test. k Akt(Ser473), GSK3β(Ser9), and ERK(Thr202/Tyr204) phosphorylation by western blot in Py230 tumor extracts. l–n Quantification of Akt, GSK3β, and ERK phosphorylation normalized to total protein (mean ± SEM, n = 3, two-tailed t test). Data are presented as mean ± SEM. Asterisks show statistical significance TRF vs AL or as shown.