Fig. 4.

MET and soy factors inhibit colonosphere formation in part through regulation of FASN and PTEN gene expression. a Lun and Gen decreased FASN mRNA abundance and induced PTEN transcript levels in P1 colonospheres formed from unsorted HCT116 cells. Means with distinct letters differed at P < 0.05, relative to untreated cells (n = 4 independent samples). b Colonosphere formation in HCT116 cells treated with Lun, Gen, or MET in the presence of the PTEN inhibitor SF1670. HCT116 cells were pre-treated with SF1670 (2 µM) for 24 h prior to plating under low attachment conditions with added soy factors (Gen, Lun) and MET. P1 colonospheres were quantified after 5 days. *P < 0.05 between control and SF1670-treated cells. Means with distinct letters differed at P < 0.05 (two independent experiments, each in quadruplicates). c Insulin added to 2 µM final concentration increased P1 colonosphere formation (*P < 0.05 between control and insulin-treated group). Treatment with Lun, Gen, or MET reduced colonosphere formation (Gen > Lun = MET) under insulin supplementation. Mean ± SEM are from two independent experiments, each performed in quadruplicate. Means with different letters differed at P < 0.05. d FACS analysis, using stem cell surface antigen markers CD133 and CD44, of attached HCT116 cells treated with Lun, Gen, or MET for 72 h in the presence of insulin (2 μM final concentration). Addition of each factor decreased the percentage of CD133⁺CD44⁺ cells, relative to insulin-treated cells. Data (mean ± SEM) are from three independent experiments, each performed in quadruplicate. *P < 0.05 between control and insulin-treated groups. Means with different letters differed at P < 0.05