The vascular endothelium plays a crucial role in the regulation of vascular homeostasis and in preventing the initiation and progress of cardiovascular disease by controlling mechanical functions of the underlying vascular smooth muscle. Three vasodilators: nitric oxide (NO), prostacyclin, and endothelium-derived hyperpolarizing factor, produced by the endothelium, underlie this activity. These substances act in a co-ordinated interactive manner to maintain normal endothelial function and operate as support mechanisms when one pathway malfunctions. In this review, we discuss recent advances in our understanding of how gender influences the interaction of these factors resulting in the vascular protective effects seen in pre-menopausal women. We also discuss how endothelial NO synthase (NOS) can act in both a pro- and anti-inflammatory action and therefore is likely to be pivotal in the initiation and time course of an inflammatory response, particularly with respect to inflammatory cardiovascular disorders. Finally, we review recent evidence demonstrating that it is not solely NOS-derived NO that mediates many of the beneficial effects of the endothelium, in particular, nitrite acts as a store of NO released during pathological episodes associated with NOS inactivity (ischemia/hypoxia). Each of these more recent findings has emphasized new pathways involved in endothelial biology, and following further research and understanding of the significance and mechanisms of these systems, it is likely that new and improved treatments for cardiovascular disease will result.