Endothelial cell function and metabolism are closely linked to differential use of energy substrate sources and combustion. While endothelial cell migration is promoted by 2-phosphofructokinase-6/fructose-2,6-bisphosphatase (PFKFB3)-driven glycolysis, proliferation also depends on fatty acid oxidation for dNTP synthesis. We show that inflammatory activation of human umbilical vein endothelial cells (HUVECs) by interleukin-1β (IL-1β), despite inhibiting proliferation, promotes a shift toward more metabolically active phenotype. This was reflected in increased cellular glucose uptake and consumption, which was preceded by an increase in PFKFB3 mRNA and protein expression. However, despite a modest increase in extracellular acidification rates, the increase in glycolysis did not correlate with extracellular lactate accumulation. Accordingly, IL-1β stimulation also increased oxygen consumption rate, but without a concomitant rise in fatty acid oxidation. Together, this suggests that the IL-1β-stimulated energy shift is driven by shunting of glucose-derived pyruvate into mitochondria to maintain elevated oxygen consumption in HUVECs. We also revealed a marked donor-dependent variation in the amplitude of the metabolic response to IL-1β and postulate that the donor-specific response should be taken into account when considering targeting dysregulated endothelial cell metabolism.