Background: IL-8 is an important chemokine implicated in the pathogenesis of chronic rhinosinusitis (CRS), but little is known about epigenetic regulation of IL8 in the pathogenesis of CRS. Objective: We sought to investigate the relationship between the DNA methylation level in the IL8 proximal promoter and CRS in Han Chinese subjects. Methods: Patients with chronic rhinosinusitis with nasal polyps (CRSwNP; n = 187), patients with chronic rhinosinusitis without nasal polyps (CRSsNP; n = 89), and control subjects (n = 57) were enrolled in 2 independent cohorts. Purified human nasal epithelial cells from each participant were assessed for percentage DNA methylation of CpG sites in the IL8 proximal promoter by using bisulfite pyrosequencing and for functional aspects of methylation status by using in vitro assays. Results: DNA methylation of CpG sites 1, 2, and 3, respectively, in the IL8 proximal promoter was significantly decreased in human nasal epithelial cells of patients with CRSwNP compared with that in patients with CRSsNP (P < .001) and control subjects (P < .001). Percentage of DNA methylation of the CpG3 site was correlated negatively with both tissue eosinophilic cationic protein (P < .01) and myeloperoxidase (P < .05) levels. IL-1β (P < .001) and TNF-α (P < .01) significantly increased IL8 expression accompanied by a reduction in methylation at the CpG3 site (P < .001). Electrophoretic mobility shift assays demonstrated that methylation status of CpG3 changed the binding of octamer-binding transcription factor 1 and nuclear factor κB. Conclusion: Decreased DNA methylation of particularly CpG sites in the IL8 proximal promoter might play a role in the pathogenesis of CRSwNP. Keywords: Chronic rhinosinusitis; DNA methylation; Han Chinese; IL-8 proximal promoter; Octamer-binding transcription factor 1; human nasal epithelial cells; nuclear factor κB.