Ectopic Spns2 expression induces apoptosis in NSCLC cells.
(A), Quantification of Casp3 positive cells shown in D. Vehicle (pEGFP) transfected cells were used as control (Con). N>100 cells are counted from three separate experiments. (B), Western blot analyses for cell death markers. A549 cells were transiently transfected as in A. Cell lysates were collected and blotted with the antibodies indicated. Vehicle (pEGFP) transfected cells were used as a negative control and Actin was used as a loading control. (C), Representative images of FLICA staining of Spns2 positive cells. Live A549 cells were incubated with FLICA for 1 hour 48 hours after transfection. Cells were fixed and micrographic images taken. Scale bar is 10 µm. (D), Quantification of FLICA positive cells shown in C. N>100 cells from three separate experiments were counted double blindly. (E), Spns2 expression reduced viable cell number in A549 cells. Cells were transfected as in A. 48 hours later, the relative live cell number was measured by a CCK-8 kit using a microplate reader. N = 4. (F), Spns2 expression reduced cell number in H1299 cells. Live H1299 cell number were measured as in I. N = 4. In B, E, H, I, J, error bars represent SD, *, p<0.05, **, p<0.01.