Fig. 2

Caspase-1 inhibition prevents PPARγ cleavage and then blocks TAM differentiation. a THP-1 cells were cultured alone or cocultured with MCF-7 or cocultured in the presence of YVAD, VAD or NCX-4016 for 48 h, and the expression of pro-tumoral genes and CD36 were assessed by RT-qPCR (n = 3). b, c The levels of VEGF and IL-6 in concentrated cell culture supernatant were measured by ELISA (n = 3). d–f The pro-angiogenesis, pro-invasion and pro-migration ability of CCM_Ctrl, CCM_Coculture, CCM_YVAD, CCM_VAD and CCM_NCX-4016 was tested (n = 3). g Peritoneal macrophages were isolated from wild type (Co-Casp1-WT) or caspase-1 ^−/− mice (Co-Casp1-KO) and cocultured with 4T1. After 2 days, pro-tumoral genes and Cd36 were determined by RT-qPCR, using wild type peritoneal macrophages cultured alone as control (Ctrl). Data were generated and analyzed by Step one plus Real-Time PCR-system (AB Applied biosystem) (n = 3). h Caspase-1 expression in BMDMs from wild type or caspase-1 ^−/− mice was determined by immunoblot. i, j EO771 cancer cells were implanted in the flank of nude mice alone or in combination with WT or caspase-1-deficient (KO) BMDMs. Tumors were resected and measured 21 days later (in j, n = 8). Scale bar, 2 cm. k TAMs were sorted from the total tumor tissue cells by magnetic cell sorting using MACS, and TAM percentage in mixed cells was analyzed (n = 8). l Caspase-1 expression in TAMs was analyzed by immunoblot. m PPARγ and its fragment in TAMs were analyzed by immunoblot. n Nude mice body weight was analyzed (n = 8). o Tumor growth was monitored by measuring volumes every 2 days with a caliber rule. Tumor volumes were calculated by the formula, 1/2(a × b²), where a is the long diameter and b is the short diameter (in mm; n = 8). All statistic data in this figure represent means ± s.e.m., (*P < 0.05, **P < 0.01, ***P < 0.001, ns, not significant, one way ANOVA for multiple comparisons)