Neonatal necrotizing enterocolitis (NEC), which is a disease with a poor prognosis, is considered to be caused by the coincidence of intestinal ischemia-reperfusion injury and systemic inflammation due to the colonization of pathogenic bacteria. Interleukin (IL)-8, a proinflammatory cytokine, plays an important role in the pathophysiology of NEC. It was recently reported that IL-1beta activates the IL-8 gene by regulating the transcriptional nuclear factor kappaB (NF-kappaB) signaling pathways in intestinal cells. The protective role of maternal milk in NEC pathogenesis has been reported in both human and animal studies. In this study, we show that human breast milk dramatically suppressed the IL-1beta-induced activation of the IL-8 gene promoter by inhibiting the activation pathway of NF-kappaB. Moreover, we also show that human breast milk induced the production of IkappaBalpha. These results suggest that human breast milk could be protective and therapeutic in neonates with NEC by inhibiting the activation pathway of NF-kappaB.