Figure 4
From: Doxorubicin and etoposide sensitize small cell lung carcinoma cells expressing caspase-8 to TRAIL
The role of caspases and mitochondria in apoptosis induced by combination of TRAIL and doxorubicin in U1690 and U1906 SCLC cell lines. (A) U1690 cells (%) with characteristic apoptotic nuclear morphology (DAPI staining) after pre-treatment (1 h) with z-VAD-fmk (3 μM) or vehicle (DMSO), and subsequent treatment (24 h) with TRAIL (100 ng/ml), doxorubicin (1 μM), or their combination. Results are means ± S.E.M. of three independent experiments. Statistical significance: P < 0.05, (*) versus control, (+) versus TRAIL, (x) versus doxorubicin, (o) versus appropriate sample without z-VAD-fmk. (B) Cleavage of PARP, caspase-8 (12 h) or lamin A, caspase-2, and caspase-3 (24 h) in U1690 cells treated as indicated in (A). An equal loading was verified using G3PDH or anti-β-actin antibody. (C) The cleavage of Bid, level of Bcl-2, Bcl-XL or Mcl-1 protein (whole cell lysates), and Bax translocation (mitochondrial fraction) in U1690 cells treated (24 h) with TRAIL (100 ng/ml), doxorubicin (1 μM) or their combination. An equal loading was verified using anti-β-actin (whole cell lysates) or anti-TOM (mitochondrial fraction) antibody. (D) The release of cytochrome c into the cytoplasm of U1690 cells treated (12, 16, 20, 24 h) with TRAIL (100 ng/ml), doxorubicin (1 μM) or their combination. An equal loading was verified using anti-G3PDH antibody. Results are representative of three independent experiments. (E) Percentage of U1906 cells with subdiploid DNA content and decreased mitochondrial membrane potential after treatment (24 h) with TRAIL (100 ng/ml), doxorubicin (0.5 μM), etoposide (2.5 μM) or their combination with TRAIL.