Figure 4

Over-expression of dnTCF1 lead to reduced viability and increased cell death rate in LS174T cells. Over-expression of dnTCF1 significantly increases the rate of cell death in LS174T cells 48 hr after induction (p < 0.005, student's t-test). This was determined using two cell death assays based on either trypan blue dye or DNA-intercalating dyes, propidium iodide (PI) and hoechst 342 (HST). LS174T control and dnTCF1 cell lines were cultured in the presence of induction. After 24 hr and 48 hr, floating and attached cells were stained with the respective dyes. Data from both assays are presented as the mean ± standard deviation from 3 separate experiments. (A) PI/HST assay: Cells were stained with PI (stains apoptotic/necrotic cells) and HST (stains viable cells). Percentage of PI stained cells relative to total cell number reflect the extent of cell death. (B) Trypan blue dye exclusion assay: Cells were stained with the trypan blue dye. Percentage of trypan blue stained cells (stains dead cells) relative to total cell number reflect extent of cell death. (C) MTT viability assay: The cells' capability to metabollically convert the MTT substrate was quantified, 48 hr after either induction (+dox) or no induction (-dox), by solubilizing MTT formazan crystals and performing spectrophotometry at 540 nm. Cell viability, which is proportional to the absorbance at 540 nm, decreases in dnTCF1 over-expressing LS174T cells relative to control cells.