Inhibition of Pulmonary Microvascular Endothelial Glutamine Transport by Glucocorticoids and Endotoxin

Abstract

Background: During septic states, the lungs produce increased amounts of glutamine, an event that is mediated by both endotoxin and the glucocorticoid hormones and is presumed to be due to accelerated intracellular glutamine biosynthesis. Because enhanced net glutamine release in vivo could also be due to a decrease in cellular uptake, we assayed glutamine transport in cultured rat microvascular pulmonary endothelial cells. Methods: The effect of Escherichia coli endotoxin (LPS, 1 μg/mL), various cytokines, and dexamethasone (DEX, 0.1 μmol/L) on glutamine transport activity was studied in rat lung microvascular endothelial cells grown in varying glutamine concentrations (0, 0.1, 0.5, and 2 mmol/L). Experiments were also performed in cells treated with cycloheximide, actinomycin D, or chelerythrine chloride. Results: More than 90% of glutamine transport was mediated by the Na⁺-dependent transport system ASC. DEX and LPS inhibited endothelial glutamine uptake in a time- and dose-dependent manner, a response that was only observed when incubation medium contained the lower concentrations of glutamine. Neither DEX nor LPS altered transport activity in cells cultured in medium containing 2 mmol glutamine/L. There was no synergistic or additive effect when both compounds were added together. The cytokines tumor necrosis factor α, interleukin (IL) 1, IL-2, and IL-6 did not alter 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 abrogated the inhibition of transport activity that was observed in DEX- or LPStreated cells, whereas the protein kinase C inhibitor chelerythrine chloride had no effect on either control or stimulated glutamine transport. Conclusions: These data suggest that DEX and LPS "down-regulate" glutamine uptake by lung microvascular endothelial cells by inducing the synthesis of an inhibitory protein that modulates the activity of the System ASC protein. This response in vitro appears to be influenced 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 illness. (Journal of Parenteral and Enteral Nutrition 19:477-481, 1995)

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References

Souba WW : Glutamine: Physiology, Biochemistry and Nutrition in Critical Illness . Landes RG Co, Austin, TX, 1992, pp 3-7

Souba WW , Klimberg VS , Plumley DA , et al: The role of glutamine in maintaining a healthy and supporting the metabolic response to injury and infection. J Surg Res 48:383-391, 1990

Brand K. , Fekl W. , von Hintzenstern J. , et al: Metabolism of glutamine in lymphocytes. Metabolism 38(suppl):29-33, 1989

Caldwell MD : Local glutamine metabolism in wounds and inflammation. Metabolism 38(suppl):34S-39S, 1989

Ardawi MSM , Majzoub MF : Glutamine metabolism in skeletal muscle of septic rat. Metabolism 40:155-164, 1991

Plumley DA , Austgen TR , Salloum RM , et al: The role of the lungs in maintaining amino acid homeostasis . JPEN 14:569-573, 1990

Souba WW , Herskowitz K. , Plumley DA : Lung glutamine metabolism. JPEN 14(suppl):68S-70S, 1990

Crapo JD , Peters-Golden M. , Marsh-Salin J. , et al. Pathologic changes in the lungs of oxygen-adapted rats. Lab Invest 39:640-653, 1978

Abcouwer S. , Lukaszewicz G. , Ryan U. , et al: Molecular regulation of lung glutamine synthetase expression . Surgery 118: 325-335, 1995

10.

Ryan US Isolation and culture of pulmonary endothelial cells. Environ Health Perspec 56:103-114, 1984

11.

Pan M. , Wasa M. , Souba WW : TNF-stimulated arginine transport by human vascular endothelium requires activation of protein kinase C. Am Surg 221:590-601, 1995

12.

Kilberg MS , Handlogten ME , Christensen HS. Characteristic of System ASC for transport of neutral amino acids in the isolated rat hepatocyte. J Biol Chem 256:3304-3312, 1981

13.

Plumley DA , Souba WW , Hautamaki RD , et al: Accelerated lung amino acid release in hyperdynamic septic surgical patients. Arch Surg 125:57-61, 1990

14.

Austgen TA , Chen MK , Salloum RM , et al: Glutamine metabolism by the endotoxin-injured lung. J Trauma 31:1068-1075, 1991

15.

Souba WW , Pacitti AJ : How amino acids get into cells: Mechanism, models, menus, and mediators . JPEN 16:569-578, 1992

16.

Scheltinga MR , Young LS , Benfell K. , et al: Glutamine-enriched intravenous feedings attenuate extracellular fluid expansion after standard stress. Ann Surg 214:.385-395, 1991

17.

Herskowitz K. , Bode BP , Block ER , et al: The effects of endotoxin on glutamine transport by pulmonary artery endothelial cells . J Surg Res 50:356-361, 1991