Figure 7. Lactate-induced oxidant production in microvascular endothelial cells is likely to be NADPH oxidase dependent.

. Human micro-vascular endothelial cells (HMVEC-d) purchased from Cambrex Bio Science (Walkersville, MD) and passages 5–9 were used for experiments. Cells were grown in endothelial basal medium 2 (CC-3156) containing EGM-2, MV (SingleQuots, Cambrex, Walkersville, MD, CC-4147), 100 units/ml penicillin, 100 units/ml streptomycin and 0,2 μg/ml amphotericin B. Cultures were maintained at 37° in humidified 95% air and 5% CO₂ atmosphere. Cells were growth-arrested by incubating overnight in endothelial basal medium 2. Nitroblue tetrazolium (NBT) (1 mg/ml) was added followed by lactate (15 mM). pH was adjusted to 7.4. In similar experiments, effect of the NADPH oxidase inhibitor diphenylene iodonium (DPI, 10μM) and the oxidant donor SIN-1 (1 mM; 3-morpholino-syndnonimine) was tested. Note the differences in VEGF in response to lactate-supplementation.