Fig. 7. CPT1A is required to mediate the tumor promoting effect of adipocytes in vivo.

a Control and CPT1A knockdown SW480 cells were mixed with Matrigel alone or in combination with adipocytes and injected subcutaneously into NSG mice. The size of the tumors was measured every 5 days for 51 days. Data represent the mean ± SEM (n = 6, for sh-Control and sh-CPT1A group; and n = 7 for sh-Control + adipocytes and sh-CPT1A + adipocytes group, ^#p < 0.05 and *p < 0.01; NS = not significant). b Tumor tissues from three mice of each group were analyzed for the levels of CPT1A, active-β-catenin, and β-catenin using Western blotting. c Quantitative analysis of relative active-β-catenin levels in xenograft tumors from four different groups of mice. The levels of active-β-catenin were normalized to total β-catenin in each sample. Data represent the mean ± SD (n = 3, *p < 0.01). d Tumor tissues from three mice of each group were analyzed for the expression of CPT1A, LGR5, and MYC using RT-PCR. Data represent the mean ± SD (n = 3, ^#p < 0.05, *p < 0.01, §p < 0.001, and ^¶p < 0.0001). e Tumor initiation experiments were performed using control and CPT1A knockdown SW480 cells. Cells were mixed with Matrigel alone or in combination with adipocytes and injected into NSG mice at 100 or 1000 cells per site and total eight injections were used for each cell group. The number of tumors formed was determined 3 months post inoculation. The stem cell frequency was calculated using extreme limiting dilution analysis (ELDA) (&p = 0.01, comparing sh-control group with sh-control + adipocytes group; and **p = 0.0003, comparing sh-control + adipocytes group with sh-CPT1A + adipocytes group).