Kefir peptides, generated by kefir grain fermentation of milk proteins, showed positive antioxidant effects, lowered blood pressure and modulated the immune response. In this study, kefir peptide was evaluated regarding their anti-inflammatory effects on particulate matter <4 μm (PM4.0)-induced lung inflammation in NF-κB-luciferase+/+ transgenic mice. The lungs of mice under 20 mg/kg or 10 mg/kg PM4.0 treatments, both increased significantly the generation of reactive oxygen species (ROS) and inflammatory cytokines; increased the protein expression levels of p-NF-κB, NLRP3, caspase-1, IL-1β, TNF-α, IL-6, IL-4 and α-SMA. Thus, we choose the 10 mg/kg of PM4.0 for animal trials; the mice were assigned to four treatment groups, including control group (saline treatment), PM4.0 + Mock group (only PM4.0 administration), PM4.0 + KL group (PM4.0 + 150 mg/kg low-dose kefir peptide) and PM4.0 + KH group (PM4.0 + 500 mg/kg high-dose kefir peptide). Data showed that treatment with both doses of kefir peptides decreased the PM4.0-induced inflammatory cell infiltration and the expression of the inflammatory mediators IL-lβ, IL-4 and TNF-α in lung tissue by inactivating NF-κB signaling. The oral administrations of kefir peptides decrease the PM4.0-induced lung inflammation process through the inhibition of NF-κB pathway in transgenic luciferase mice, proposing a new clinical application to particulate matter air pollution-induced pulmonary inflammation.