Background: many previous studies have demonstrated the therapeutic potential ofN. sativatotal oil fractions, neutral lipids (NLs), glycolipids (GLs), phospholipids (PLs), and unsaponifiable (IS) in asthma patients. We therefore tested its effect on airway smooth muscle (ASM) cells by observing its ability to regulate the production of glucocorticoid (GC)-insensitive chemokines in cells treated with TNF-α/IFN-γ, and its antioxidative and reactive oxygen species (ROS) scavenging properties. Materials and methods: the cytotoxicity ofN. sativaoil fractions was assessed using an MTT assay. ASM cells were treated with TNF-α/IFN-γ for 24 h in the presence of different concentrations ofN. sativaoil fractions. An ELISA assay was used to determine the effect ofN. sativaoil fractions on chemokine production (CCL5, CXCL-10, and CXCL-8). The scavenging effect ofN. sativaoil fractions was evaluated on three reactive oxygen species (ROS), O2•−, OH•, and H2O2. Results: our results show that differentN. sativaoil fractions used at 25 and 50 µg/mL did not affect cell viability. All fractions ofN. sativaoil inhibited chemokines in a concentration-dependent manner. Interestingly, the total oil fraction showed the most significant effect of chemokine inhibition, and had the highest percentage of ROS scavenging effect. Conclusion: these results suggest thatN. sativaoil modulates the proinflammatory actions of human ASM cells by inhibiting the production of GC-insensitive chemokines.