Synergistic differentiation-promoting activity of interferon gamma and tumor necrosis factor-alpha: role of receptor regulation on human neuroblasts.

BACKGROUND

Interferon gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF) synergize in inducing human neuroblastoma cells to differentiate terminally in vitro into mature nonproliferating neurons. The mechanisms by which this synergistic activity takes place are still obscure.

PURPOSE

To understand the basis of IFN-gamma-TNF synergism, we investigated the constitutive equipment of receptors to IFN-gamma and TNF in two human neuroblastoma cell lines (i.e., LAN-5 and GI-LI-N) and their quantitative and functional variations following treatment with IFN-gamma or TNF.

METHODS

IFN-gamma receptors and TNF receptors were assessed and functionally characterized by radioreceptor-binding assay before and after treatment of the cells with IFN-gamma or TNF. The TNF receptor subtypes were identified by the reverse transcriptase-polymerase chain reaction, chemical cross-linking of receptors to iodinated TNF, and inhibition of TNF binding by type-specific anti-TNF receptor monoclonal antibodies. The effects of cytokines on cell differentiation were assessed by thymidine incorporation inhibition and morphologic maturation.

RESULTS

No quantitative or functional modification of IFN-gamma receptors was observed in TNF-treated cells. However, after treatment with IFN-gamma, TNF receptor numbers were enhanced to a different extent in both cell lines. The two neuroblastoma cell lines expressed, both constitutively and after IFN-gamma induction, only one species of TNF receptor, i.e., the p80 form in LAN-5 and the p60 form in GI-LI-N. Sequential treatment with IFN-gamma followed by TNF, but not in the opposite order, could reproduce the early effects of differentiation in neuroblastoma cells, supporting a role for TNF receptor up-regulation as a basis for the cooperation between the two cytokines.

CONCLUSION

The results strongly suggest that receptor regulation can be at least one mechanism by which IFN-gamma and TNF exert their synergistic effects. Moreover, it appears that the two TNF receptor types are redundant in signaling neuroblastoma cell differentiation.

IMPLICATIONS

Our findings can provide a guideline for a rational design of experimental differentiation-based therapeutic protocols in patients with neuroblastoma.