Hyperoxic acute lung injury (HALI) is characterized by inflammation and epithelial cell death. CLOCK genes are master regulators of circadian rhythm also implicated in inflammation and lung diseases. However, the relationship of CLOCK genes in hyperoxia-induced lung injury has not been studied. This study will determine if HALI alters CLOCK gene expression. To test this, wild-type and NALP3^–/– mice were exposed to room air or hyperoxia for 24, 48, or 72 hr. In addition, mice were exposed to different concentrations of hyperoxia (50, 75, 100% O₂) or room air for 72 hr. The mRNA and protein levels of lung CLOCK genes, based on qPCR and Western blot analysis respectively, and their target genes are significantly elevated in mice exposed to hyperoxia compared to controls. Alterations in CLOCK genes are associated with increased inflammatory markers in bronchoalveolar lavage fluid of hyperoxic mice compared to controls. Histological examination of mice lungs exposed to hyperoxia show increased inflammation and alveolar congestion compared to controls. Our results indicate sequential increase in CLOCK gene expression in lungs of mice exposed to hyperoxia compared to controls. Additionally, data suggests dose-dependent increase in CLOCK gene expression with increased oxygen concentrations. To validate if the expression changes related to CLOCK genes are indeed associated with inflammation, NALP3^-/- was introduced to analyze loss of function in inflammation. Western blot analysis showed significant CLOCK gene downregulation in NALP3^-/- mice compared to wild-type controls. Together, our results demonstrate that hyperoxia-mediated lung inflammation is associated with alterations in CLOCK gene expression.