Fig. 7
From: C1Q+ TPP1+ macrophages promote colon cancer progression through SETD8-driven p53 methylation
SETD8 genetic inhibition in CSCs and macrophages impairs tumor growth, metastasis formation and prolongs murine survival in in vivo models of CRC. A, B In vivo whole-body imaging analysis (A) and kinetics of tumor formation (B) detected by in vivo imaging analysis at the indicated time following orthotopic injection of CR-CSphC#8 stably transduced with doxycycline (DOXY)-inducible vectors pTRIPZ and shSETD8. Time −2 weeks indicates the day of cell implantation. After 2 weeks, DOXY was added to both groups of mice (pTRIPZ and shSETD8). Bars show the tumor size average of 6 mice/group ± SEM. Slopes of the growth rate were compared by Mann–Whitney test. C Immunohistochemical analysis of SETD8, p53K382me1 and CK20 on 2 tumors randomly chosen from each group divided as in (B) collected 4 weeks after the injection and 2 weeks after doxy treatment. Scale bars, 40 µm. D Xenograft tumor size in mice subcutaneously co-injected with CR-CSphC#8 and THP1 cells. CR-CSphC#8 and THP1 cells were both stably transduced with doxycycline (DOXY)-inducible vectors pTRIPZ and shSETD8 alone or in combination with shTP53. Day 0 indicates the day of cell implantation. Doxycycline (DOXY) was added after tumors reached 75-100 mm3, 12 days after the injection. Bars show the tumor size average of 6 mice/group ± SEM. Slopes of the growth rate were compared by t-test. E Immunohistochemical analysis of CK20 and CD68 on a tumor generated from the subcutaneous co-injection of CR-CSphC#8 and THP1 cells stably transduced with doxycycline (DOXY)-inducible vector pTRIPZ. Scale bars, 40 µm. F Kaplan-Meier graphs showing the murine survival upon SETD8 silencing alone or in combination with shTP53 in mice treated as in (D). The statistical significance between two treatment groups was evaluated using a log rank test. G Kinetics of metastasis formation detected by in vivo imaging analysis at the indicated time following spleen injection of CR-CSphC#8 stably transduced with doxycycline (DOXY)-inducible vectors pTRIPZ and shSETD8. Data are expressed as mean ± SD of 6 mice analyzed. Slopes of the growth rate were compared by t-test. H In vivo whole-body imaging analysis of mice treated as in (G) and analyzed at the indicated time points after splenectomy. I Photons count of all metastatic sites (intestine, lung and liver) in mice following spleen injection of CR-CSphC#8 and THP1 cells both stably transduced with doxycycline (DOXY)-inducible vectors pTRIPZ and shSETD8. Data are expressed as mean ± SD of 6 mice analyzed. The statistical significance between the treatment groups was evaluated using a t-test (upper panel). Representative in vivo imaging analysis of metastatic foci in the intestine, lung and liver of mice treated as indicated (lower panels). See also Figures S6 and S7