Background: Hypoxia‐inducible factor (HIF)‐1 is a key transcription factor responding to hypoxia. It is composed of an oxygen‐sensitive α subunit (HIF‐1α) and a constitutively expressed β subunit. Increasing evidence indicates an essential role for HIF‐1α in infection and immunity. Because inflamed periodontium is thought to be hypoxic, we hypothesize that HIF‐1α is expressed and related to its upstream regulator tumor necrosis factor (TNF)‐α and downstream effecter vascular endothelial growth factor (VEGF). Methods: Human gingival biopsies were collected from advanced periodontitis sites and clinically healthy sites, and immunohistochemically examined for HIF‐1α and VEGF peptides. The messenger ribonucleic acid (mRNA) and protein levels of HIF‐1α, VEGF, and TNF‐α in the biopsies were then assessed by reverse transcription polymerase chain reaction and Western blotting. Results: HIF‐1α–positive immunoreactivity was detected in the nuclei of epithelial and endothelial cells. In periodontal pockets, there was a marked increase in the proportion of fibroblast‐like cells and leukocyte‐like cells expressing HIF‐1α. Protein levels of HIF‐1α, VEGF, and TNF‐α were significantly higher in periodontal pockets than in control gingival samples. The mRNA expression of VEGF and TNF‐α was also increased in periodontal pockets. Conclusion: HIF‐1α is expressed in healthy and diseased periodontium and may be related to TNF‐α and VEGF function during periodontitis.