Several lines of evidence indicate that hypoxia is a stimulus to vascular smooth muscle cell (VSMC) proliferation that occurs in pulmonary hypertension. The present study tested the hypothesis that low O₂ tension directly stimulates human VSMC proliferation by inducing them to produce interleukin (IL)-1, a potent autocrine growth factor for human VSMC. Human VSMC derived from pulmonary artery, aorta, or saphenous vein were incubated in either a normal in vitro O₂environment (20% O₂) or in chambers containing low (∼1%) or moderate (5%) O₂. Levels of IL-1α and IL-1β mRNA increased in human VSMC after 24–48 h of incubation in low O₂ compared with levels in normoxic cells and then decreased upon subsequent reoxygenation. Levels of cell-associated IL-1α also increased progressively after 24–48 h in low O₂; however, detectable IL-1α was not released from the cells in the media. IL-1β was detectable in cell lysates and supernatants; however, the levels were not affected by exposure to low O₂. mRNA encoding for tumor necrosis factor-α (TNF-α), a related cytokine and VSMC mitogen, was not detectable in human VSMC exposed to either low or 20% O₂. Proliferation of human VSMC was not stimulated during exposure to low O₂, despite the fact that cells remained responsive to the mitogenic effect of exogenous IL-1. Interestingly, however, exposure to 5% O₂ enhanced proliferation of human VSMC but did not induce IL-1α production. Inhibition of IL-1 binding to the type I IL-1 receptor by exogenous addition of IL-1-receptor antagonist (10 μg/ml) did not attenuate the proliferation rates of human VSMC incubated in 20%, 5%, or low O₂ or in human VSMC that were reoxygenated after exposure to low O₂. These results demonstrate two direct and distinct effects of hypoxia on VSMC. Exposure to moderately low O₂ tension induces VSMC proliferation, independent of IL-1, whereas exposure to very low O₂ tension induces production of IL-1α.