The presence of multiple areas of hypoxia (low oxygen tension) is a hallmark feature of human and experimental tumours. Monocytes are continually recruited into tumours where they differentiate into tumour-associated macrophages (TAM) and often accumulate in hypoxic and/or necrotic areas. A number of recent studies have shown that macrophages respond to hypoxia by up-regulating transcription factors such as HIF-1alpha and HIF-2alpha, which in turn up-regulate the expression of a broad array of mitogenic, pro-invasive, pro-angiogenic and pro-metastatic genes. Here we show that primary human macrophages but not monocytes rapidly up-regulate HIF-1alpha and HIF-2alpha proteins upon exposure to hypoxia, and that these proteins then translocate to the nucleus. We also demonstrate differences in the temporal expression and responses to re-oxygenation for HIF-1alpha and HIF-2alpha in macrophages. Here we found that, compared to HIF-1alpha, HIF-2alpha expression was prolonged and persisted with re-oxygenation. ATF-4 and Egr-1 were also found to be hypoxia-responsive transcription factors in macrophages but not monocytes, but only early after exposure to hypoxia. Taken together, these findings indicate that a number of transcription factors work together in a tightly regulated fashion to control macrophage activities in ischaemic areas of diseased tissues.