Background Hypoxia is considered a key factor in the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP). However, the specific mechanism driving polypogenesis under hypoxic conditions is unclear. This study aimed to explore hypoxia-induced alterations in the transcriptome of human nasal epithelial cells (HNECs) in vitro. Methods HNECs derived from the tissue of patients with CRSwNP were established as air-liquid interface (ALI) cultures. Confluent cultures were kept submerged or treated with cobalt chloride (CoCl2) to induce hypoxia. Transcriptome analysis was used to identify key mRNAs involved in this process. Real-time PCR (RT-PCR), Western blotting, and immunofluorescence were used to observe the effects of hypoxia on ciliogenesis. Results Numerous genes, biological processes and pathways were altered under submerged culture conditions or after CoCl2 treatment. Analysis of the results under both hypoxic conditions revealed that the transcriptional program responsible for ciliogenesis was significantly impaired. Downregulation of cilia-related genes and inhibition of ciliated cell differentiation under hypoxia were confirmed by RT-PCR, Western blot and immunofluorescence analyses. Conclusion Hypoxia impairs ciliogenesis and ciliary function in HNECs, which might play a role in the pathogenesis of CRSwNP.