INHIBITION OF PULMONARY MICROVASCULAR ENDOTHELIAL GLUTAMINE TRANSPORTBY GLUCOCORTICOIDS AND ENDOTOXIN

Background: During septic states, the lungs produce increased amounts of glutamine, an event that is mediated by both endotoxin and the gluc ocorticoid hormones and is presumed to be due to accelerated intracell ular glutamine biosynthesis. Because enhanced net glutamine release in vivo could also be due to a decrease in cellular uptake, we assayed g lutamine transport in cultured rat microvascular pulmonary endothelial cells. Methods: The effect of Escherichia coli endotoxin (LPS, 1 mu g /mL), various cytokines, and dexamethasone (DEX, 0.1 mu mol/L) on glut amine transport activity was studied in rat lung microvascular endothe lial cells grown in varying glutamine concentrations (0, 0.1, 0.5, and 2 mmol/L). Experiments were also performed in cells treated with cycl oheximide, actinomycin D, or chelerythrine chloride. Results: More tha n 90% of glutamine transport was mediated by the Na+-dependent transpo rt system ASC. DEX and LPS inhibited endothelial glutamine uptake in a time- and dose-dependent manner, a response that was only observed wh en incubation medium contained the lower concentrations of glutamine. Neither DEX nor LPS altered transport activity in cells cultured in me dium containing 2 mmol glutamine/L. There was no synergistic or additi ve effect when both compounds were added together. The cytokines tumor necrosis factor alpha, interleukin (IL) 1, IL-2, and IL-6 did not alt er glutamine transport. Both DEX and LPS inhibited glutamine transport by decreasing transporter maximal transport velocity (V-max) without affecting transporter affinity (K-m) Cycloheximide and actinomycin D a brogated the inhibition of transport activity that was observed in DEX - or LPS-treated cells, whereas the protein kinase C inhibitor chelery thrine chloride had no effect on either control or stimulated glutamin e transport. Conclusions: These data suggest that DEX and LPS ''downre gulate'' glutamine uptake by lung microvascular endothelial cells by i nducing the synthesis of an inhibitory protein that modulates the acti vity of the System ASC protein. This response in vitro appears to be i nfluenced by the extracellular glutamine concentration. This decrease in microvascular endothelial glutamine transport may be one mechanism by which net lung glutamine release is enhanced during critical illnes s.