Macrophages distributed in tissues throughout the body contribute to homeostasis. In the inflammatory state, macrophages undergo mechanical stress that regulates the signal transduction of immune responses and various cellular functions. However, the effects of the inflammatory response on macrophages under physiological cyclic stretch are unclear. We found that physiological cyclic stretch suppresses inflammatory cytokine expression in macrophages by regulating NF-κB activity. NF-κB phosphorylation at Ser536 in macrophages was inhibited, suggesting that tank-binding kinase (TBK1) regulates NF-κB activity during physiological stress. Moreover, TBK1 expression was suppressed by physiological stretch, and TBK1 knockdown by siRNA induced the suppression of NF-κB phosphorylation at Ser536. In conclusion, physiological stretch triggers suppression of a TBK1-dependent excessive inflammatory response, which may be necessary to maintain tissue homeostasis.