Breastfeeding protects against mucosal infections in infants. The underlying mechanisms through which immunity develops in human milk following maternal infection with mucosal pathogens are not well understood. We simulated nasal mucosal influenza infection through live attenuated influenza vaccination (LAIV) and compared immune responses in milk to inactivated influenza vaccination (IIV). Transcriptomic analysis was performed on RNA extracted from human milk cells to evaluate differentially expressed genes and pathways on days 1 and 7 post-vaccination. Both LAIV and IIV vaccines induced influenza-specific IgA that persisted for at least 6 months. Regulation of type I interferon production, toll-like receptor, and pattern recognition receptor signaling pathways were highly upregulated in milk on day 1 following LAIV but not IIV at any time point. Upregulation of innate immunity in human milk may provide timely protection against mucosal infections until antigen-specific immunity develops in the human milk-fed infant.