Morphological and biochemical changes induced by arsenic trioxide in neuroblastoma cell lines

Arsenic trioxide has recently been shown to inhibit growth and induce apoptosis in a variety of hematologic malignancies, but very little is known about its effects on solid tumors and especially on neuroblastoma cells that have self-differentiating characteristics. To demonstrate the growth inhibition induced in neuroblastoma cells (the SH-SY5Y and SK-N-AS cell line) and acute promyelocytic leukemia cells (HL-60) by arsenic trioxide (As₂O ₃), the viable cell numbers were counted after trypan blue staining. Apoptosis was assessed by the cell morphology, by flow cytometry with annexin-V staining, and by Western blot analysis for the apoptosis-relafed proteins (bcl-2 and PARP). To decide the dose for the clinical application of AS ₂O₃, normal peripheral blood lymphocytes were also examined. The growth and survival of the SH-SY5Y and SK-N-AS cells were markedly inhibited by As₂O₃ treatment at a 3 μM concentration before the changes of the normal lymphocytes were observed. The apoptotic cells showed a shrunken cell nucleus, and an increase in the number and balloon-like swelling of the mitochondria at 72 h after the As₂O₃ was added. Apoptosis of the annexin-V-positive cell proportion in the neuroblastoma cell lines was increased with increasing the exposure time and the concentration of As₂O₃, just like the HL-60 cells. Bcl-2 downregulation and PARP degradation were also noted all the cell lines, but these changes were not statistically significant among the 3 cell lines. Taken together, these results indicate that As₂O₃ is an excellent candidate as a therapeutic agent for the treatment of neuroblastoma.