Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1995 Feb 14;92(4):1147–1151. doi: 10.1073/pnas.92.4.1147

Development and characterization of essential fatty acid deficiency in human endothelial cells in culture.

R Lerner 1, P Lindström 1, A Berg 1, E Johansson 1, K Rosendahl 1, J Palmblad 1
PMCID: PMC42655  PMID: 7862650

Abstract

We induced an essential fatty acid deficiency (EFAD) in human umbilical vein endothelial cells by culture in medium with 20% (vol/vol) delipidated fetal calf serum. EFAD, reflected by decreased cellular linoleic acid (18:2 omega 6) and arachidonic acid (20:4 omega 6) and emergence of the oleic acid derivative 5,8,11-eicosatrienoic acid (20:3 omega 9; Mead's acid), was evident after 1 week of culture and became pronounced after 2 weeks. Beyond that time point, control cells (cultured in 20% normal fetal calf serum) grew deficient of 18:2 omega 6, and EFAD cells died. 18:2 omega 6 addition to EFAD cells resulted in dose-dependent increases of 18:2 omega 6 and 20:4 omega 6. 20:4 omega 6 or 5,8,11,14,17-eicosapentaenoic acid (20:5 omega 3) additions resulted in normalization of these acids, and conversion of 20:5 omega 3 to 4,7,10,13,16,19-docosahexaenoic acid (22:6 omega 3) was noted. Agonist-induced increases in concentrations of prostacycline (prostaglandin I2; PGI2) and cytosolic Ca2+, [Ca2+]i, were reduced in EFAD cells and not restored by 18:2 omega 6 or 20:4 omega 6 additions. Change of the medium in EFAD cultures 1 day before the experiments decreased 20:3 omega 9 and normalized the PGI2 production and [Ca2+]i changes, whereas addition of 20:3 omega 9 to control cells impaired the [Ca2+]i response, indicating a suppressive effect of 20:3 omega 9. Thus, EFAD in endothelial cells is associated with abnormalities of eicosanoid and second-messenger production partly attributable to 20:3 omega 9 accumulation. Moreover, the gradual emergence of 18:2 omega 6 deficiency in regularly grown control cells underlines the need for careful analysis of fatty acids in long-term cell cultures.

Full text

PDF
1147

Images in this article

1150Fig. 5
on p.1150
1150Fig. 6
on p.1150

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bjerve K. S., Mostad I. L., Thoresen L. Alpha-linolenic acid deficiency in patients on long-term gastric-tube feeding: estimation of linolenic acid and long-chain unsaturated n-3 fatty acid requirement in man. Am J Clin Nutr. 1987 Jan;45(1):66–77. doi: 10.1093/ajcn/45.1.66. [DOI] [PubMed] [Google Scholar]
  2. Bonta I. L., Parnham M. J., Adolfs M. J. Reduced exudation and increased tissue proliferation during chronic inflammation in rats deprived of endogenous prostaglandin precursors. Prostaglandins. 1977 Aug;14(2):295–307. doi: 10.1016/0090-6980(77)90175-7. [DOI] [PubMed] [Google Scholar]
  3. Cham B. E., Knowles B. R. A solvent system for delipidation of plasma or serum without protein precipitation. J Lipid Res. 1976 Mar;17(2):176–181. [PubMed] [Google Scholar]
  4. Denko C. W. Effect of prostaglandins in urate crystal inflammation. Pharmacology. 1974;12(6):331–339. doi: 10.1159/000136556. [DOI] [PubMed] [Google Scholar]
  5. Evans J. F., Nathaniel D. J., Zamboni R. J., Ford-Hutchinson A. W. Leukotriene A3. A poor substrate but a potent inhibitor of rat and human neutrophil leukotriene A4 hydrolase. J Biol Chem. 1985 Sep 15;260(20):10966–10970. [PubMed] [Google Scholar]
  6. Freed M. S., Spaethe S. M., Lefkowith J. B., Saffitz J. E., Needleman P. Essential fatty acid deficiency inhibits early but not late leukocyte infiltration in rabbit myocardial infarcts. Prostaglandins. 1989 Jul;38(1):33–44. doi: 10.1016/0090-6980(89)90014-2. [DOI] [PubMed] [Google Scholar]
  7. Grynkiewicz G., Poenie M., Tsien R. Y. A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem. 1985 Mar 25;260(6):3440–3450. [PubMed] [Google Scholar]
  8. Gyllenhammar H., Palmblad J. Linoleic acid-deficient rat neutrophils show decreased bactericidal capacity, superoxide formation and membrane depolarization. Immunology. 1989 Apr;66(4):616–620. [PMC free article] [PubMed] [Google Scholar]
  9. Gyllenhammar H., Palmblad J., Ringertz B., Hafström I., Borgeat P. Rat neutrophil function, and leukotriene generation in essential fatty acid deficiency. Lipids. 1988 Feb;23(2):89–95. doi: 10.1007/BF02535286. [DOI] [PubMed] [Google Scholar]
  10. Gyllenhammar H., Ringertz B., Becker W., Svensson J., Palmblad J. Essential fatty acid deficiency in rats: effects on arachidonate metabolism, generation of cyclooxygenase products and functional responses in neutrophils. Immunol Lett. 1986 Oct;13(4):185–189. doi: 10.1016/0165-2478(86)90053-2. [DOI] [PubMed] [Google Scholar]
  11. HANSEN A. E., WIESE H. F. Fat in the diet in relation to nutrition of the dog. I. Characteristic appearance and gross changes of animals fed diets with and without fat. Tex Rep Biol Med. 1951;9(3):491–515. [PubMed] [Google Scholar]
  12. Hurd E. R., Johnston J. M., Okita J. R., MacDonald P. C., Ziff M., Gilliam J. W. Prevention of glomerulonephritis and prolonged survival in New Zealand Black/New Zealand White F1 hybrid mice fed an essential fatty acid-deficient diet. J Clin Invest. 1981 Feb;67(2):476–485. doi: 10.1172/JCI110056. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jaffe E. A., Nachman R. L., Becker C. G., Minick C. R. Culture of human endothelial cells derived from umbilical veins. Identification by morphologic and immunologic criteria. J Clin Invest. 1973 Nov;52(11):2745–2756. doi: 10.1172/JCI107470. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. James M. J., Gibson R. A., Neumann M. A., Cleland L. G. Effect of dietary supplementation with n-9 eicosatrienoic acid on leukotriene B4 synthesis in rats: a novel approach to inhibition of eicosanoid synthesis. J Exp Med. 1993 Dec 1;178(6):2261–2265. doi: 10.1084/jem.178.6.2261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Laposata M., Prescott S. M., Bross T. E., Majerus P. W. Development and characterization of a tissue culture cell line with essential fatty acid deficiency. Proc Natl Acad Sci U S A. 1982 Dec;79(24):7654–7658. doi: 10.1073/pnas.79.24.7654. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lefkowith J. B. Essential fatty acid deficiency inhibits the in vivo generation of leukotriene B4 and suppresses levels of resident and elicited leukocytes in acute inflammation. J Immunol. 1988 Jan 1;140(1):228–233. [PubMed] [Google Scholar]
  17. Lefkowith J. B., Jakschik B. A., Stahl P., Needleman P. Metabolic and functional alterations in macrophages induced by essential fatty acid deficiency. J Biol Chem. 1987 May 15;262(14):6668–6675. [PubMed] [Google Scholar]
  18. Lefkowith J. B., Rogers M., Lennartz M. R., Brown E. J. Essential fatty acid deficiency impairs macrophage spreading and adherence. Role of arachidonate in cell adhesion. J Biol Chem. 1991 Jan 15;266(2):1071–1076. [PubMed] [Google Scholar]
  19. Lepage G., Roy C. C. Direct transesterification of all classes of lipids in a one-step reaction. J Lipid Res. 1986 Jan;27(1):114–120. [PubMed] [Google Scholar]
  20. Lindström P., Lerner R., Palmblad J., Patarroyo M. Rapid adhesive responses of endothelial cells and of neutrophils induced by leukotriene B4 are mediated by leucocytic adhesion protein CD18. Scand J Immunol. 1990 Jun;31(6):737–744. doi: 10.1111/j.1365-3083.1990.tb02825.x. [DOI] [PubMed] [Google Scholar]
  21. Marshall L. A., Johnston P. V. The influence of dietary essential fatty acids on rat immunocompetent cell prostaglandin synthesis and mitogen-induced blastogenesis. J Nutr. 1985 Dec;115(12):1572–1580. doi: 10.1093/jn/115.12.1572. [DOI] [PubMed] [Google Scholar]
  22. Palmblad J., Gyllenhammar H., Ringertz B., Nilsson E., Cottell B. Leukotriene B4 triggers highly characteristic and specific functional responses in neutrophils: studies of stimulus specific mechanisms. Biochim Biophys Acta. 1988 Aug 19;971(1):92–102. doi: 10.1016/0167-4889(88)90165-6. [DOI] [PubMed] [Google Scholar]
  23. Palmblad J., Wannemacher R. W., Salem N., Jr, Kuhns D. B., Wright D. G. Essential fatty acid deficiency and neutrophil function: studies of lipid-free total parenteral nutrition in monkeys. J Lab Clin Med. 1988 Jun;111(6):634–644. [PubMed] [Google Scholar]
  24. Paulsrud J. R., Pensler L., Whitten C. F., Stewart S., Holman R. T. Essential fatty acid deficiency in infants induced by fat-free intravenous feeding. Am J Clin Nutr. 1972 Sep;25(9):897–904. doi: 10.1093/ajcn/25.9.897. [DOI] [PubMed] [Google Scholar]
  25. Ramesha C. S., Pickett W. C. Platelet-activating factor and leukotriene biosynthesis is inhibited in polymorphonuclear leukocytes depleted of arachidonic acid. J Biol Chem. 1986 Jun 15;261(17):7592–7595. [PubMed] [Google Scholar]
  26. Schreiner G. F., Flye W., Brunt E., Korber K., Lefkowith J. B. Essential fatty acid depletion of renal allografts and prevention of rejection. Science. 1988 May 20;240(4855):1032–1033. doi: 10.1126/science.3285468. [DOI] [PubMed] [Google Scholar]
  27. Schreiner G. F., Rovin B., Lefkowith J. B. The antiinflammatory effects of essential fatty acid deficiency in experimental glomerulonephritis. The modulation of macrophage migration and eicosanoid metabolism. J Immunol. 1989 Nov 15;143(10):3192–3199. [PubMed] [Google Scholar]
  28. Spector A. A., Kaduce T. L., Hoak J. C., Fry G. L. Utilization of arachidonic and linoleic acids by cultured human endothelial cells. J Clin Invest. 1981 Oct;68(4):1003–1011. doi: 10.1172/JCI110322. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Stenson W. F., Prescott S. M., Sprecher H. Leukotriene B formation by neutrophils from essential fatty acid-deficient rats. J Biol Chem. 1984 Oct 10;259(19):11784–11789. [PubMed] [Google Scholar]
  30. Wene J. D., Connor W. E., DenBesten L. The development of essential fatty acid deficiency in healthy men fed fat-free diets intravenously and orally. J Clin Invest. 1975 Jul;56(1):127–134. doi: 10.1172/JCI108061. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Wickham N. W., Vercellotti G. M., Moldow C. F., Visser M. R., Jacob H. S. Measurement of intracellular calcium concentration in intact monolayers of human endothelial cells. J Lab Clin Med. 1988 Aug;112(2):157–167. [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES