Nitric oxide is a strong vasodilatory and anti-inflammatory signaling molecule that plays diverse roles in maintaining vascular homeostasis. Nitric oxide produced by endothelial cells is a critical regulator of this balance, such that endothelial dysfunction is defined as a reduced capacity for nitric oxide production and decreased nitric oxide sensitivity. This ultimately results in an imbalance in vascular homeostasis leading to a prothrombotic, proinflammatory, and less compliant blood vessel wall. Endothelial dysfunction is central in numerous pathophysiologic processes. This article reviews mechanisms governing nitric oxide production and downstream effects, highlighting the role of nitric oxide signaling in organ system pathologies.