Purpose: To investigate the signal factor and its function in the medium-mediated bystander eVect during heavy-ion irradiation of human salivary gland (HSG) neoplastic cells. Materials and methods: Unirradiated recipient HSG cells were co-cultivated with HSG donor cells irradiated with 290MeV/u carbon beams having diVerent LET values. Cell proliferation and micronucleus (MN) induction in recipient cells with and without treatment of a NO scavenger (PTIO) were measured and the concentration of nitrite in the co-culture medium was detected. As a direct control, the eVects of a nitric oxide (NO) generator (sper/NO) on cell proliferation and MN induction were also examined. Results: Increases in cell proliferation and MN induction were found in the recipient HSG cells as a result of co-culturing and cell proliferation was obviously enhanced during a further subculture. In comparison with 13 keV/mm, 100 keV/mm carbon-ion irradiation was found to be a more eYcient inducer of the medium-mediated bystander eVect. The treatment of cells by PTIO resulted in elimination of such eVects, which supports a rooxliedifzoartioNnOprionduthcteomf NedOiu,mni-tmriteediwaatesddebtyescttaenddienr theVeeccot.-cAulstuarne medium, and the dose–response for its concentration was similar to that of cell proliferation and MN induction in the recipient cells. When the HSG cells were treated by sper/NO with a concentration of less than 20 mm, cell proliferation was enhanced, whereas MN increased along with sper/NO concentration. Conclusion: NO participated in the medium-mediated bystander incideVects on cell proliferation and MN induction, depending on the LET of irradiation.