Located in the innermost layer of the vasculature and directly interacting with blood flow, endothelium integrates various biochemical and biomechanical signals to maintain barrier function with selective permeability, vascular tone, blood fluidity, and vascular formation. Endothelial cells respond to laminar and disturbed flow by structural and functional adaption, which involves reprogramming gene expression, cell proliferation and migration, senescence, autophagy and cell death, as well as synthesizing signal molecules (nitric oxide and prostanoids, etc) that act in manners of autocrine, paracrine, or juxtacrine. Inflammation occurs after infection or tissue injury. Dysregulated inflammatory response participates in pathogenesis of many diseases. Endothelial cells exposed to inflammatory stimuli from the circulation or the microenvironment exhibit impaired vascular tone, increased permeability, elevated procoagulant activity, and dysregulated vascular formation, collectively contributing to the development of vascular diseases. Understanding the endothelial response to pathophysiological stress of hemodynamics and inflammation provides mechanistic insights into cardiovascular diseases, as well as therapeutic opportunities.