Synergistic Antiproliferative Effects of Combined γ-Tocotrienol and PPARγ Antagonist Treatment Are Mediated through PPARγ-Independent Mechanisms in Breast Cancer Cells
Figure 10
(a) Effect of γ-tocotrienol, rosiglitazone, and GW9662 given alone or in combination on PGD2 synthesis and (b) treatment effect of 15d-PGJ2 in PPARγ positive MCF-7 and MDA-MB-231 cells. MCF-7 and MDA-MB-231 cells were plated at a density of 05 cells/well (3 replicates per group) in 6-well plates in 2 mL antibiotic-free media. Transfections were performed using 5 μL lipofectamine 2000 according to the manufacturer’s protocol. Briefly, for each well to be transfected, 100 pmol of the scrambled or PPARγ siRNAs was diluted with 2 mL of media. After 6 h transfection, the medium was replaced with fresh growth media containing 10% FBS and cells were cultured for 18 h. Cells were then exposed to control or treatment media for a 4-day culture period. Afterwards whole cell lysates were assayed for PGD2 according to the manufacturer’s protocol. Vertical bars indicate the amount of PGD2 synthesized (pg/mL) ± SEM in each treatment group. For 15d-PGJ2 effect, MCF-7 and MDA-MB-231 cells were plated at a density of 04 cells/well (3 replicates per group) in 96-well plates and allowed to adhere overnight. Transfections were performed using 0.25 μL lipofectamine 2000. For each well of cells to be transfected, 5 pmol of the scrambled or PPARγ siRNAs was diluted with 100 μL of media. After 6 h transfection, the medium was replaced with fresh growth media containing 10% fetal bovine serum and cells were cultured for 18 h. Cells were then exposed to 100 μL of control or treatment media for a 4-day culture period. Afterwards viable cell number was determined using MTT colorimetric assay. Vertical bars indicate mean cell count ± SEM in each treatment group. * as compared with vehicle-treated controls.
