Figure 1From: Phosphoinositide 3-kinase targeting by the β galactoside binding protein cytokine negates akt gene expression and leads aggressive breast cancer cells to apoptotic deathEvents leading to apoptotic death in BT474 and SKBR3 breast cancer cells treated with human recombinant β galactoside binding protein (Hu-r-βGBP). (a-b)Rate of cell proliferation and dose response to Hu-r-βGBP. Dotted lines in growth curves define the time of occurrence of apoptotic events. Values are means of triplicate cultures ± standard error of the mean (SEM). (c-d)Sequential expression of apoptotic events after treatment with 30 nM Hu-r-βGBP. Top to bottom: loss of mitochondrial membrane potential assessed by tetramethylrhodamine ester (TMRE); phosphatidylserine orientation at the plasma membrane assessed by annexin V staining; caspase 3 activity and DNA fragmentation (terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labelling (TUNEL)). Apoptotic cells are in circled areas. Values are percentages of cells in apoptosis. Left panels: controls, right panels: cells treated with 30 nM Hu-r-βGBP. (e, h)Inhibition of class IA phosphoinositide 3-kinase (PI3K) activity by 30 nM Hu-r-βGBP measured as phosphatidylinositol (3,4,5)-trisphosphate (PIP3) generated from immunoprecipitated PI3K in a kinase assay at 30°C, and assessed in a competitive ELISA. Values are means from triplicate readings ± SEM. (f, i) akt mRNA and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA levels. Left panels: controls, right panels: cells treated with 30 nM Hu-r-βGBP. (g, j)Phosphorylated (Ser473) Akt and total Akt protein. Left panels: controls, right panels: cells treated with 30 nM Hu-r-βGBP. (a-j) Hu-r-βGBP added at 3 h after seeding. The data are representative of results obtained in three separate experiments.Back to article page