Endothelial Cell Inhibition of Hypoxia-induced Stimulation of Serotonin Uptake by Vascular Smooth Muscle Cells

Exposure of bovine pulmonary artery smooth muscle cells (SMC) to anoxia for 24 h resulted in an approximate twofold stimulation of serotonin (5-HT) uptake compared with SMC exposed to 20% O2. The stimulation of 5-HT uptake by exposure to anoxia was eliminated when bovine pulmonary artery endothelial cells (EC) were cocultured with SMC. Incubation with EC-conditioned medium produced similar inhibitory effects on 5-HT uptake of SMC exposed to anoxia, a result not seen with SMC-conditioned medium. The inhibitory effect of EC-conditioned medium on the anoxia-stimulated 5-HT uptake was concentration dependent and absent at a dilution of 1:16. The production of the inhibitor was time-dependent. The EC-derived inhibitory factor was heat-stable at 100 degrees C for as long as 10 min and was stable in a pH range from 5.0 to 10.0. Significant losses of inhibitory activity of EC-conditioned medium were observed after treatment with trypsin, pronase E, and proteinase K. The molecular weight on the inhibitory factor from EC-conditioned medium was estimated to be approximately 66,000 by size-exclusion chromatography. The data show that stimulated uptake of 5-HT by SMC under anoxic conditions is under regulation by a protein (or polypeptide) produced by EC.