Muscle cells, including human airway smooth muscle cells (HASMCs) express ankyrin repeat protein 1 (Ankrd1), a member of ankyrin repeat protein family. Ankrd1 efficiently interacts with the type III intermediate filament desmin. Our earlier study showed that desmin is an intracellular load-bearing protein that influences airway compliance, lung recoil, and airway contractile responsiveness. These results suggest that Ankrd1 and desmin may play important roles on ASMC homeostasis. Here we show that small interfering (si)RNA-mediated knockdown of the desmin gene in HASMCs, recombinant HASMCs (reHASMCs), up-regulates Ankrd1 expression. Moreover, loss of desmin in HASMCs increases the phosphorylation of Akt, inhibitor of κB kinase (IKK)-α, and inhibitor of κB (IκB)-α proteins, leading to NF-κB activation. Treatment of reHASMCs with Akt, IKKα, IκBα, or NF-κB inhibitor inhibits the loss of desmin-induced Ankrd1 up-regulation, suggesting Akt/NF-κB-mediated Ankrd1 regulation. Transfection of reHASMCs with siRNA specific for p50 or p65 corroborates the NF-κB-mediated Ankrd1 regulation. Luciferase reporter assays show that NF-κB directly binds on Ankrd1 promoter and up-regulates Ankrd1 levels. Overall, our data provide a new link between desmin and Ankrd1 regulation, which may be important for ASMC homeostasis.