To investigate the formation and effect of nitric oxide (NO) on oxidative injury and anti-virus in human lung epithelial cells (A549) infected with respiratory syncytial virus (RSV) in vitro. A549 cells infected with RSV treated or untreated with Aminoguanidine (AG), the specific inhibitor for inducible nitric oxide synthase, were used to collect cellular supernatants and do the experiments. At indicated time points, the concentrations of NO, hydroxy radical (OH·), superoxide anion (O2.—) and malondialdehyde (MDA) were measured according to the kit reference. The viral titers were evaluated by plaque forming unit (PFU) assay. We found that RSV infection could markedly up-regulate the levels of OH· and O2.— in A549 cellular supernatants since 4 hours after infection. Meanwhile, the levels of MDA increased obviously in a time-dependent manner. The changes of each index varied significantly compared with the control group. But the NO content in the cellular supernatants could significantly decrease when AG inhibitor was used, and the levels of OH·, O2.— and MDA were also decreased accordingly in comparison with the corresponding RSV-infected group. In addition, the titer of virus was increased, which was about 1.5 times as many the RSV group 24 h after RSV infection. The changes of these indices have significantly different compared with the corresponding RSV-infected group. In conclusion, RSV infection could induce high-level NO. The NO may associate with the content of free radical, which could decrease the viral titers in the infection, but lead oxidative injury to normal cells.