Ewing’s sarcoma and peripheral primitive neuroectodermal tumor cells produce large quantities of bioactive tumor necrosis factor-α (tnf-α) after radiation exposure

Abstract

Purpose In the present study, we examined human Ewing’s sarcoma (ES) and peripheral primitive neuroectodermal tumor (pPNET) cell lines that are able to produce TNF-α as part of the response to irradiation. Radiation-induced tumor cell production of TNF-α may enhance irradiation efficacy and improve the effect of local tumor irradiation. On the other hand, radiation-induced tumor cell production of TNF-α may adversely affect the normal tissue. Methods and materials Twelve different ES/pPNET cell lines were investigated in vitro and (after establishment as tumor xenografts in athymic nude mice) in vivo for their TNF-α mRNA expression (real-time quantitative reverse transcriptase polymerase chain reaction) and TNF-α protein production ( in vitro: enhanced amplified sensitivity immunoassay; in vivo: immunohistochemistry) after exposure to different irradiation doses (2, 5, 10, 20, 30, or 40 Gy) and after different time intervals (1, 3, 6, 12, 24, 48, or 72 h after irradiation). The bioactivity of the TNF-α protein was evaluated in chromogenic cytotoxicity and neutralization assays. Results Nine out of 12 ES/pPNET cell lines express constitutively significant quantities of bioactive TNF-α in vitro. ES/pPNET cells originating from primary tumors secreted higher TNF-α levels than cells derived from metastatic lesions. In 5 of the 9 TNF-α–producing cell lines, TNF-α mRNA and protein levels were upregulated after irradiation exposure in a time- and dose-dependent manner. After establishment of the ES/pPNET cell lines in athymic nude mice, the radiation-induced TNF-α release could be demonstrated also in the xenograft tumors in vivo (analogous to the in vitro experiments). Using the same methods for quantitative analysis, it was determined that the TNF-α expression of the radiation-responsive tumor cells was up to 2000-fold higher compared to the maximal radiation-induced TNF-α release in normal lung tissue measured during the pneumonic phase. Conclusion Certain ES/pPNET cell lines produce extremely large quantities of bioactive TNF-α after radiation exposure in a time- and dose-dependent manner. Radiation-induced TNF-α production of tumor cells may be of paramount importance in respect to not only tumor behavior, but also to potential damage to normal tissue and the clinical status of the host.