Neuroblastoma and Hepatocyte Coculture Conditioned Media Alter Apoptosis

Background. Neuroblastoma is a childhood tumor that often displays unusual biological behavior. The tumor may present with widespread metastases that are unresponsive to aggressive treatment. At other times, both the metastases and the primary tumor may spontaneously regress without treatment. Apoptosis, or programmed cell death, is thought to play a role in the dichotomous behavior of neuroblastoma. We hypothesize that neuroblastoma cells will interact with host tissues to release mediators that affect apoptosis. Materials and methods. Human neuroblastoma cells and human Chang hepatocytes are grown in a noncontact, coculture system. After incubation for 4 days, the medium from the coculture system is collected. Neuroblastoma cells and Chang hepatocytes are then plated separately with the conditioned medium and their own standard growth medium as controls. After 4 days, these cells are harvested and cytospins made for immunostaining. Tumor necrosis factor α (TNF-α), Fas ligand, and Bcl-2, are measured with immunohistochemistry. Apoptosis is detected with the TUNEL method. Immunostaining data are interpreted with computer image analysis and reported as stain index. TUNEL data are reported as percentage apoptotic cells. All data are reported as means ± SEM. Statistical analysis is performed and P < 0.05 considered significant. Results. Chang hepatocytes grown in the coculture conditioned media have an increase in TNF-α and Fas ligand. The neuroblastoma cells have a significant decrease in Fas ligand. There is a significant increase in the number of apoptotic hepatocytes when they are cultured in the conditioned media. In contrast, the neuroblastoma cells grown in the coculture conditioned media show no increase in apoptosis. Finally, Bcl-2 is significantly increased in the neuroblastoma cells cultured in the conditioned media. Conclusions. Neuroblastoma cells grown in coculture conditioned media show increased expression of Bcl-2 and decreased Fas ligand levels. These changes should diminish apoptosis activity in the tumor cells. In contrast, the conditioned media induce elevated levels of proapoptotic mediators in the Chang hepatocytes. A tumor's ability to successfully metastasize may be dependent on mediators generated in the tumor–host interaction, and may not be just an independent characteristic of the tumor itself.