Lipopolysaccharide and Tumor Necrosis Factor Stimulate Lung Microvascular Arginine Uptake,a Response Attenuated by Dexamethasone

Abstract

Background: Lipopolysaccharide (LPS), tumor necrosis factor-α, (TNF), and glucocorticoids can modulate endothelial nitric oxide (NO) production. L-Arginine is the exclusive precursor for NO biosynthesis, suggesting that NO generation and arginine transport are intimately linked. Methods: To further study this relationship, we examined the effects of LPS, TNF, and dexamethasone (DEX) on arginine uptake by rat lung microvascular endothelial cells. The transport of radiolabeled arginine was assayed in confluent cells grown in 24-well plates. Results: The bulk (>90%) of arginine transport was mediated by the Na⁺-independent carriers System y⁺ and System b^0,+. Arginine transport was stimulated independently by LPS and TNF, a response first observed at 10 hours. Together, both agents exerted an additive effect on carrier-mediated uptake. The LPS- and TNFinduced increase in arginine transport activity was blocked by cycloheximide and actinomycin D, indicating the requirement for RNA and protein synthesis. The enhancement in transport activity was primarily due to an increase in Systems y⁺ maximal transport capacity (V _max) with no change in transporter affinity and little change in System b^0,+ activity. Treatment of cells with dexamethasone inhibited arginine transport activity in a time-and dose-dependent manner, an event that was abrogated by both actinomycin D and cycloheximide. The combination of DEX and LPS and TNF abrogated each other's antagonistic effects. Conclusions: These data indicate that LPS and TNF additively stimulate arginine transport in lung microvascular endothelial cells via a pathway that requires de novo protein synthesis (possibly of the transporter protein itself) and that this response is attenuated by DEX. (Journal of Parenteral and Enteral Nutrition 20:50-55, 1996)

Get full access to this article

View all access options for this article.

References

Barbul A. : Arginine: Biochemistry, physiology, and therapeutic implications. JPEN 10:227-238, 1986

Moncada S. , Palmer RMJ , Higgs EA : Nitric oxide: Physiology, pathophysiology, and pharmacology. Pharmacol Rev 43:109-142, 1991

Nathan C. : Nitric oxide as a secretory product of mammalian cells. FASEB J 6:3051-3064, 1992

Forstermann U. , Schmidt Hhhw , Pollock JS , et al: Isoforms of nitric oxide synthetase. Characterization and purification from different cell types. Biochem Pharmacol 42:1849-1857, 1991

Henry Y. , Lepoivre M. , Drapier JC , et al: EPR characterization of molecular targets for NO in mammalian cells and organelles. FASEB J 7:1124-1134, 1993

Chen PY , Sanders PW : L-Arginine abrogates salt-sensitive hypertension in Dahl/Rapp rats . J Clin Invest 88:1559-1567, 1991

Griffith OW , Park KH , Levi R. , et al: The role of plasma arginine in nitric oxide synthesis: Studies with arginase-treated rats. IN Biology of Nitric Oxide, Moncada S , Marletta MA , Hibbs JB , (eds). Portland Press, Brookfield, VT, 6-9, 1991

Hinshaw LB , Archer LT , Beller-Todd BK , et al: Survival of primates in LD100 septic shock following steroid/antibiotic therapy. J Surg Res 28:151-170, 1980

Hoffman SL , Punjabi NH , Kamula S. , et al: Reduction of mortality in chloramphenicol-treated severe typhoid fever by high-dose dexamethasone. N Engl J Med 310:82-84, 1984

10.

Nicholson DP : Glucocorticoids in the treatment of shock and the adult respiratory distress syndrome. Clin Chest Med 3:121-132, 1982

11.

White MF : The transport of cationic amino acids across the plasma membrane of mammalian cells. Biochem Biophy Acta 822:355-374, 1985

12.

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

13.

Ochoa JB , Udekwu AO , Billiar TR , et al: Nitrogen oxide levels in patients following trauma and during sepsis. Ann Surg 214:621-626, 1991

14.

Petros A. , Bennet D. , Vallance P. : Effects of nitric oxide synthesis inhibitor on hypotension in patients with septic shock. Lancet 338:1557-1558, 1991

15.

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

16.

Van Winkle LJ , Campione AL , Gorman JM : Na+-independent transport of basic and zwitterionic amino acids in mouse blastocysts by a shared system and by processes which distinguish between these substrates. J Biol Chem 263:3150-3163, 1988

17.

Christensen HN , Liang M. : Transport of diamino acids into the Ehrlich cell. J Biol Chem 241:5542-5551, 1966

18.

Lind DS , Copeland EM , Souba WW : Endotoxin stimulates arginine transport in pulmonary endothelial cells . Surgery 114:199-205, 1993

19.

Cendan JC , Souba WW , Copeland EM , et al: Cytokines regulate endotoxin stimulation of endothelial cell arginine transport. Surgery 117:213-219, 1995

20.

Cendan JC , Souba WW , Copeland EM , et al: Endotoxin-induced nitric oxide production in pulmonary artery endothelial cells is regulated by cytokines. Arch Surg 129:1296-1300, 1994

21.

Knowles RG , Merrett M. , Salter M. , et al: Differential induction of brain, lung, and liver nitric oxide synthetase by endotoxin in the rat. Biochem J 270:833-836, 1990

22.

Radomski MW , Palmer RMJ , Moncada S. : Glucocorticoids inhibit the expression of an inducible, but not the constitutive nitric oxide synthetase in vascular endothelial cells. Proc Natl Acad Sci USA 87:10043-10047, 1990

23.

Stuehr DJ , Griffith OW : Mammalian nitric oxide synthetase. Adv Enzymol 65:287-346, 1992

24.

Ignarro LJ : Biosynthesis and metabolism of endothelium-derived nitric oxide. Annu Rey Pharmacol Toxicol 30:535-560. 1990

25.

Nussler AK di Silvio M. Billiar TR et al: Stimulation of the nitric oxide synthase pathway in human hepatocytes by cytokines and endotoxin. J Exp Med 176:261-266, 1992

26.

Hibbs JB , Westenfelder C. , Taintor R. , et al: Evidence of cytokine-inducible nitric oxide synthesis from L-arginine in patients receiving interleukin-2 therapy. J Clin Invest 89:867-877, 1992

27.

Westenberger U. , Thanner S. Ruf HH , et al: Formation of free radicals and nitric oxide derivative of hemoglobin in rats during shock syndrome. Free Rad Res Commun 11:167-178, 1990

28.

Marletta MA , Yoon PS Iyengar R. : Macrophage oxidation of L-arginine to nitrite and nitrate: nitric oxide is an intermediate. Biochemistry 27:8706-8711, 1988

29.

Rees DD , Cellek S. , Palmer RMJ , et al: Dexamethasone prevents the induction by endotoxin of a nitric oxide synthetase and the association effects on vascular tone. An insight into endotoxin shock. Biochem Biophys Res Commun 173:541-547, 1990

30.

Palmer RMJ , Ashton DS , Moncada S. : Vascular endothelial cells synthesize nitric oxide from L-arginine. Nature 333:664-666, 1988

31.

Schmidt Hhhw , Nau H. , Wittfoht W. , et al: Arginine is a physiological precursor of endothelium-derived nitric oxide. Eur J Pharmacol 154:213-216, 1988

32.

Hibbs JB , Vavrin Z. , Taintor RR : L-Arginine in required for expression of the activated macrophage effector mechanism causing selective metabolic inhibition in target cells . J Immunol 138:550-565, 1987

33.

Iyenger R. , Stuehr DJ , Marletta MA : Macrophage synthesis of nitrite, nitrate and N-nitrosamine: Precursors and role of the respiratory burst. Proc Natl Acad Sci USA 84:6369-6373, 1987

34.

Pacitti AJ , Copeland EM , Souba WW : Stimulation of hepatocyte System y+-mediated L-arginine transport by an inflammatory agent. Surgery 112:403-411, 1992