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Central European Journal of Immunology
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vol. 34

Review paper
Immunonutrition and lipopolysaccharide – induced Toll-like receptor signaling

Robert Słotwiński
Sylwia M. Słotwińska
Barbara J. Bałan
Sylwia Kędziora

Centr Eur J Immunol 2009; 34 (2): 137-142
Online publish date: 2009/05/20
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Malnutrition is a major global public health problem and can be defined as a state of nutrition in which a deficiency of energy, protein and other nutrients like arginine, glutamine, fatty acids, vitamins and trace elements causes measurable effects on body and tissue function and clinical outcome. Surgical trauma increases immune system suppression and deepens disease related malnutrition. The immune disorders and malnutrition worsen in the early postoperative period, considerably affecting the process of wound healing, intestinal barrier function and the number of post-operative infections. Infections in malnourished surgical patients increase morbidity and mortality rate. Despite advances in treatment, there is still no therapy available to efficiently reduce the excessive inflammatory response, which can increase the risk of multiple organ failure (MOF) [1]. The promising results of experimental studies on treating severe infections with LPS inhibitors, TNF-α, IL-1, PAF, NO, arachidonic acid metabolites, complement component inhibitors or free radicals did not considerably reduce the mortality rate in septic patients [2]. Other strategies for the treatment of sepsis in surgical patients based on the attempts to block LPS-binding receptors and on blocking signaling pathway proteins for antibacterial response (e.g. blockade of TLR4, caspases, Fasl-Fas or NF-kB activity and blocking of HMGB1- high mobility group box1 pathway) and on attempting to regulate the neutrophil and lymphocyte apoptosis (e.g. by over expression of anti-apoptotic proteins such as Bcl-2) are still subject of experimental research [2-6]. The aim of this study is to efficiently reduce the excessive inflammatory response, above all reducing the activation of nuclear factor kB (NF-kB), the production of post-inflammatory cytokines (TNF, IL-1, IL-6), chemokines and adhesive molecules.
More efficient therapy consists influences the mechanisms of inflammatory response to a massive infection. After a major surgery complicated by severe infection special attention should be paid to modulation of the expression of signaling pathway proteins in cells that take part in early (innate) immune response to infection by applying immunonutrition. It is well known that neutrophils and monocytes/macrophages that take part in innate immune response to trauma and infection play a significant role in the elimination of microorganisms and in local and systemic inflammatory response regulation (SIRS – systemic inflammatory response syndrome) that increases the risk of MOF [7]. The disorders of phagocytosis and microorganism elimination in the site of bacterial penetration (extensive surgical wound, catheter in a large vein) intensify the pro- and anti-inflammatory response (CARS- compensatory anti-inflammatory response), which intensifies post-operative immunosuppression and may result in immunity breakdown [8]. These issues opens the discussion if the re-programming of signal transduction pathways in intestinal mucosa and innate immunity cells of septic patients after immunonutrition contribute to the attenuation of local and systemic hyperinflammatory response in massive bacterial load?
The immunomodulatory action of unsaturated fatty acids affects the decrease in activity of neutrophils, monocytes, lymphocytes, and the production of cytokines [9-12]. Immunostimulatory action of amino acids increases the phagocytal activity of leukocytes, enhances immunity to infections and accelerates wound healing [13-19]. In randomized studies it has been found that enteral immunonutrition improves the clinical course, decreases the frequency of severe infections, shortens hospital stays, reduces treatment costs and significantly decreases mortality in severe ill patients with MOF [20-25]. These benefits were found to be most impressive in surgical patients. In patients with severe trauma and infection receiving immunonutrition significant decrease in the duration of SIRS and in the frequency of MOF have been found [20, 23, 24, 26]. In the clinical setting immunonutrition (with arginine, nucleic acids and n-3 fatty acids) reduced infections complications in critically ill patients after trauma and cancer surgery [22, 23, 25, 27, 28]. The studies have been performed in various populations patients, which makes it difficult to compare their results. The most frequently included patients treated in intensive care units. In the majority of those studies, the changes in nutritional and immunity status in the course of immunonutrition and infection have not been monitored. Despite the advantage of the positive effects of immunonutrition on the treatment of surgical patients, the impact of this nutrition on the immune system still remains unclear. A better knowledge of advantages of immunomodulating nutrition in treating surgical infections requires studying the changes in the expression of signaling cascade proteins associated with their stimulation account not only for pathological inflammatory response to trauma or infection, but they can also have a protective action (e.g. increasing the apoptosis of selected cells, stimulation of signaling pathway inhibitors).
In regulating the mechanisms of local and systemic inflammatory response to a massive infection in surgical patients a significant role is played by Toll-like receptors (TLRs) expressioned in gut mucosa cells and the cells that take part in innate response to infection. Some studies performed show that trauma reduces, whereas severe infection increases the expression of TLRs recognizing bacterial antigens (e.g. LPS, peptidoglycan) [29-32]. As compared with healthy people, the expression of TLR4 in the monocytes of trauma patients was reduced [29]. In experimental studies the lack of TLRs increased the susceptibility to infections in mice [33] and caused disorders in inflammatory mediator secretion, disorders in phagocytosis and antigen presentation [34-36]. The experimental findings suggest that TLR4 plays a key role in regulating the expression of inflammatory cytokines in the lung during endotoxic shock [37]. Six hours of LPS administration induced a significant increase in pulmonary TNF-α, IL-1b and IL-6 mRNA in control (TLR4+) mice compared to TLR4 –deficient mice.
To date, several randomized clinical trials have evaluated the efficacy of arginine, glutamine, omega-3 fatty acids, nucleotides and trace elements with antioxidant properties in critically ill patients with trauma and/or infections, but the basic molecular mechanisms that can attenuate the overwhelming inflammatory response in sepsis are still unclear. In malnourished surgical patients with infections, the direct factor that intensifies the failure of local “first line” antibacterial defense can be the disorders of pathogen-associated molecular pattern (PAMPs) (e.g. LPS, peptidoglycan, teichoic acids, bacterial DNA) recognition by innate immunity cells. The hypothesis that one of the main reasons for false recognition of bacterial antigens by immune system cells (mainly by phagocytic cells) is malnutrition is highly probable. The deficiency of immunoactive nourishing substances (e.g. glutamine, fatty acids) can intensify the disorders of expression of bacterial antigen binding extracellular receptors and intracellular proteins/receptors. The excessive accumulation of bacterial wall fragments and the microorganisms being proliferated in tissues intensify the local inflammatory response and increase the release of cytokines into the blood.
Glutamine is an important energy source for lymphatic tissue and glutamine-enriched enteral nutrition has been found to reduce the incidence of sepsis in trauma patients, due to maintaining the integrity of intestinal mucosa [38-40]. Low plasma glutamine concentrations (<0.42 mM) at admission to intensive care units were associated with higher severity of illness and higher mortality rates [41]. The results of recent studies show the regulative glutamine impact on inflammatory response in severe infections and indicate that it is necessary to administer high doses (e.g. in parenteral administration 0.35g/kg–1/day–1) to obtain a better therapeutic effect [39, 42, 43]. Some most recent experimental studies show that the enteral administration of glutamine reduces the increased TLR4 expression, signal adaptor protein MyD88 (myeloid differentiation factor 88) and TRAF6mRNA (TNF-α receptor-associated factor 6) in intestinal mucosa as a response to LPS induced endotoxemia in rats (Fig. 1) [5]. In addition, the above-mentioned studies found a decreased injury to the mucous membrane of the small intestine. The effect of glutamine on intestinal TRL4 expression may be considered as a mechanism via which immunonutrition helps in the recovery of critically ill and septic patients. The mechanisms by which glutamine prevents the occurrence of infection are still unclear, but it is well known that in surgical or burn patients glutamine decreases the production of pro-inflammatory cytokines [44] and improves the bactericidal function of neutrophils [46].
The anti-inflammatory action of unsaturated fatty acids (mainly n-3 PUFAs) and their application in treating surgical infections and early sepsis (in the first phase of sepsis syndrome) still seem to be very interesting. In traumatized and surgical patients an enteral diet containing n-3 fatty acids significantly reduced infectious complications and septic events [23, 47, 48]. Enhanced survival and reduced lung failure after enteral or parenteral usage of n-3 lipids was observed in experimental models of sepsis [49-51]. Interestingly, by incorporation into various membrane (phospho)-lipid pools, n-3 fatty acids may affect lipid-signaling events in different cell types [52, 53]. The omega-3 fatty acids have also an ability to selectively suppress the signaling cascade associated with innate antibacterial response (mainly leukocytes and macrophages), independently at subsequent stages: a) endotoxin interaction with TRL4, b) activation of inhibitor phosphorylation kinases of the NF-kB (IkB) transcription factor and c) translocation to nucleus and connecting NFkB to an appropriate DNA sequence (suppressing the transcription of inflammatory response mediator genes) (Fig. 1) [54-61].
It was indicated that the enteral administration of diet enriched in unsaturated fatty acids (EPA) and glutamine in septic patients treated in intensive care units reduced the inflammatory response and mortality rate caused by acute lung injury (acute respiratory distress syndrome – ARDS) [62, 63]. The enteral administration of n-3 acids in septic patients modulated the functions of neutrophils, changed the disadvantageous proportion of n-6 acids to n-3 in the direction of higher concentration of n-3 acids, which was associated with lower concentrations of pro-inflammatory cytokines [64, 65]. These findings indicate that immunomodulating nutrition may be an effective means of influencing the inflammatory response, particularly for those pathways affected by TLR4 signaling.
Our previous study has clearly indicated that the anti-inflammatory mechanisms are activated early in malnourished patients after pancreaticoduodenectomy receiving enteral immunonutrition [66]. Early enteral immunonutrition (with glutamine, arginine and n-3 fatty acids) in comparison to standard nutrition has an immunomodulative effect on the changes in the immune response after extensive surgical trauma. These consist in selective stimulation of IL-6, IL-8, IL-10 and IL-1ra production and down-regulation of IL-1 beta and TNF-α production. The temporary increase in IL-1ra concentration between post-operative days 7-14 obtained as a result of enteral immunonutrition decreases the inflammatory response to extensive surgical trauma and shortens its duration; this accelerates the wound healing process/tissue regeneration and may help avoid late complications (fistulas, abscesses).

The above-presented results show that to improve outcomes in the group of malnourished surgical patients suffering from severe infections more attention should be devoted to explaining the molecular mechanisms regulating the innate antibacterial response. One of the preconditions to provide progress in treating the most severely ill patients is to find out more about the impact of the state of nutrition, severe infections and immunonutrition on the expression of selected signaling pathway proteins of innate antibacterial response cells. Attempts to modulate the innate antibacterial immune response by applying immunonutrition are promising and indicate that in the future it can be a valuable supplement of the therapy using a blockade of selected signaling pathways to reduce the life-threatening effects of massive infection, including mainly the increased inflammatory response.

1. Eichacker PQ, Parent C, Kalil A et al. (2002): Risk and the efficacy of antiinflammatory agents: retrospective and confirmatory studies of sepsis. Am J Respir Crit Care Med 166: 1197-1205.
2. Riedemann NC, Guo RF, Ward PA (2003): Novel strategies for the treatment of sepsis. Nat Med 9: 517-524.
3. Lee JY, Sohn KH, Rhee SH et al. (2001): Saturated fatty acids, but not unsaturated fatty acids, induce the expression of cyclooxygenase-2 mediated through Toll-like receptor 4. J Biol Chem 276: 16683-16689.
4. Singer P, Shapiro H, Theilla M et al. (2008) Anti-inflammatory properties of omega-3 fatty acids in critical illness: novel mechanisms and integrative perspective Intensive Care Med 34: 1580-1592.
5. Kessel A, Toubi E, Pavlotzky E et al. (2008): Treatment with glutamine is associated with down-regulation of Toll-like receptor-4 and myeloid differentiation factor 88 expression and decrease in intestinal mucosal injury caused by lipopolysaccharide endotoxaemia in a rat. Clin Exp Immunol 151: 341-347.
6. Chang WK, Yang KD, Chuang H et al. (2002): Glutamine Protects Activated Human T Cells from Apoptosis by Up-Regulating Glutathione and Bcl-2 Levels. Clin Immunol 104: 151-160.
7. Cohen J (2002): The immunopathogenesis of sepsis. Nature 420: 885-891.
8. Hotchkiss RS, Karl IE (2003): The pathophysiology and treatment of sepsis. N Engl J Med 348: 138-150.
9. Wachtler P, Konig W, Senkal M et al. (1997): Influence of total parenteral nutrition enriched with n-3 fatty acids on leucotriene synthesis of peripheral leucocytes and systemic cytokine levels in patients with major surgery. J Trauma 42: 191-198.
10. Caughey GC, Mantzioris E, Gibson RA et al. (1996): The effect on human tumor necrosis factor a and interleukin 1b production of diets enriched in n-3 fatyy acids vegetable oil or fish oil. Am J Clin Nutrition 63: 116-122.
11. Sperling RI, Benincaso AI, Knoell CT et al. (1993): Dietary n-3 polyunsaturated fatty acids inhibit phosphoinositide formation and chemotaxis in neutrophils. J Clin Invest 91: 651-660.
12. Fisher M, Levine PH, Weiner BH et al. (1990): Dietary n-3 fatty acid supplementation reduces superxide production and chemiluminescence in monocytes-enriched preparation of leucocytes. Am J Clin Nutrition 51: 804-808.
13. Gianotti L, Alexander JW, Pyles T et al. (1993): Arginine-supplemented diets improve survival in gut-derived sepsis and peritonitis by modulating bacterial clearance: the role of nitric oxide. Annals of Surgery 217: 644-653.
14. Reynolds JV, Daly JM, Zhang S et al. (1988): Immunomodulatory effects of arginine. Surgery 104: 142-151.
15. Kirk SJ, Hurson M, Regan MC et al. (1993): Arginine stimulates wound healing and immune function in elderly human beings. Surgery 114: 155-160.
16. Azzara A, Carulli G, Sbrana S et al. (1995): Effects of lysine-arginine association on immune functions in patients with recurrent infections. Drugs Under Exper Clin Res 21: 71-78.
17. Moffat FL, Han T, Li ZM et al. (1996): Supplemental L-arginine HCL augments bacterial phagocytosis in human polymorphonuclear leukocytes. J Cell Physiol 168: 26-33.
18. Spittler A, Winkler S, Gotzinger P et al. (1995): Influence of glutamine on the phenotype and function of human monocytes. Blood 86: 1564-1569.
19. Wiebke EA, Grieshop NA, Sinder RA et al. (1997): Effects of L-arginine supplementation on human lymphocyte proliferation in response to nonspecific and alloantigenic stimulation. J Surg Res 70: 89-94.
20. Atkinson S, Sieffert E, Bihari D (1998): Guy’s Hospital Intensive Care Group. A prospective, randomized, double-blind, controlled clinical trial of enteral immunonutrition in the critically ill. Crit Care Med 26: 1164-1172.
21. Griffiths RD, Jones C, Allan Palmer TE (1997): Six month outcome of critically ill patients given glutamine-supplemented parenteral nutrition. Nutrition 13: 295-302.
22. Marik PE, Zaloga GP (2008): Immunonutrition in critically ill patients: a systemic review and analysis of the litetarure. Intensive Care Med 34: 1980-1990.
23. Kudsk KA, Minard G, Croce MA et al. (1996): A randomized trial of isonitrogenous enteral diets after severe trauma. An immune-enhancing diet reduces septic complications. Ann Surg 224: 531-543.
24. Weimann A, Bastian L, Bischoff WE et al. (1998): Influence of arginine, omega-3 fatty acids and nucleotide-supplemented enteral support on systemic inflammatory response syndrome and multiple organ failure in patients after severe trauma. Nutrition 14: 165-172.
25. Galban C, Montejo JC, Mesejo A et al. (2000): An immune-enhancing enteral diet reduces mortality and episodes of bacteremia in septic intensive care unit patients. Crit Care Med 28: 643-648.
26. Jolliet P (1999): Immunonutrition in critically ill. Intensive Care Med 25: 631-638.
27. Senkal M, Zumtobel V, Bauer KH et al. (1999): Outcome and cost-effectiveness of perioperative enteral immunonutrition in patients undergoing upper gastrointestinal surgery. Arch Surg 134: 1309-1316.
28. Bower RH, Cerra FB, Bershadsky B et al. (1995): Early enteral administration of a formula (Impact) supplemented with arginine, nucleotides, and fish oil in intensive care unit patients: results of a multicenter, prospective, randomized, clinical trial. Crit Care Med 23: 436-439.
29. Adib-Conquy M, Moine P, Asehnoune K et al. (2003): Toll Like receptor mediated tumor necrosis factorand interleukin 10 production differ during systemic inflammation. Am
J Respir Crit Care Med 168: 158-164.
30. Rodríguez Salgado A, Regueiro V, Pérez Bárcena J et al. (2006): La glutamina como compuesto modulador de la respuesta inflamatoria en pacientes críticos con nutrición parenteral: efecto sobre la expresión y funcionalidad de los receptores toll-like. Datos preliminares. Nutr Hosp 21 (Suppl 1): 65.
31. Brandl K, Gluck T, Huber C et al. (2005): TLR4 surface display is increased in septic patients. Eur J Med Res 10: 319-324.
32. Adib-Conquy M, Adrie C, Fitting C et al. (2006): Up regulation of MyD88 and SIGIRR, molecules inhibiting Toll Like receptor signaling in monocytes from septic patients. Crit Care Med 34: 2377-2385.
33. Qureshi ST, Medzhitov R (2003): Toll-like receptors and their role in experimental models of microbial infection. Genes Immun 4: 87-94.
34. Medzhitov R, Janeway C (2000): Innate immune recognition: Mechanism and pathways. Immunol Rev 173: 89-97.
35. Akira S. Toll-like receptor signaling (2003): J Biol Chem 278: 105-108.
36. Blander FM, Medzhitov R (2004): Regulation of phagosome maturation by signals from toll-like receptors. Science 304: 1014-1018.
37. Baumgarten G, Knuefermann P, Wrigge H et al. (2006): Role of Toll-like receptor 4 for the pathogenesis of acute lung injury in Gram-negative sepsis. Eur J Anaesthesiol 23: 1041-1048.
38. Foitzik T (2001): Pancreatitis and nutrition. Significance of the gastrointestinal tract and nutrition for septic complications. Zentralbl Chir 4: 126-129.
39. Houdijk APJ, Rijnsburger ER, Jansen J et al. (1998): Randomised trial of glutamine–enriched enteral nutrition on infectious morbidity in patients with multiple trauma. Lancet 352: 772-776.
40. Novak F, Heyland DK, Avenell A et al. (2002): Glutamine supplementation in serious illness: a systematic review of the evidence. Crit Care Med 30: 2022-2029.
41. Oudemans-van Straaten HM, Bosman RJ, Treskes M et al. (2001): Plasma glutamine depletion and patient outcome in acute ICU admissions. Intensive Care Med 27: 84-90.
42. Wischmeyer PE, Kahana MD, Wolfson R et al. (2001): Glutamine induces heat shock protein and protects against endotoxin shock in the rat. J Appl Physiol 90: 2403-2410.
43. Wischmeyer PE, Riehm J, Singleton KD et al. (2003): Glutamine attenuates tumour necrosis factor-a release and enhances heat shock protein 70 in human peripheral blood mononuclear cells. Nutrition 19: 1-6.
44. O’Riordain MG, De Beaux A, Fearon KC (1996): Effect of glutamine on immune function in the surgical patient. Nutrition 12: S82-84.
45. Aosasa S, Mochizuki H, Yamamoto T et al. (1999): A clinical study of the effectiveness of oral glutamine supplementation during total parenteral nutrition: influence on mesenteric mononuclear cells. JPEN 23: S41-44.
46. Ogle CK, Ogle JD, Mao JX et al. (1994): Effect of glutamine on phagocytosis and bacterial killing by normal and pediatric burn patient neutrophils. JPEN 18: 128-133.
47. Braga M, Vignali A, Gianotti L et al. (1996): Immune and nutritional effects of early enteral nutrition after major abdominal operations. Eur J Surg 162: 105-112.
48. Senkal M, Mumme A, Eickhoff U et al. (1997): Early postoperative enteral immunonutrition: clinical outcome and cost-comparison analysis in surgical patients. Crit Care Med 25: 1489-1496.
49. Barton RG, Wells CL, Carlson A et al. (1991): Dietary omega-3 fatty acids decrease mortality and kupffer cell prostaglandin E2 production in a rat model of chronic sepsis. J Trauma 31: 768-774.
50. Johnson JA, Griswold JA, Muakkassa FF (1993): Essential fatty acids influence survival in sepsis. J Trauma 35: 128-131.
51. Grimminger F, Wahn H, Mayer K et al. (1997): Impact of arachidonic versus eicosapentaenoic acid on exotoxin-induces lung vascular leakage. Am J Respir Crit Care Med 155: 513-519.
52. Chakrabati R, Hubbard NE, Lin D et al. (1997): Alteration of platelet-activating factor-induced signal transduction in macrophages by n-3fatty acids. Cell Immunol 175: 76-84.
53. Diep QN, Intengan HD, Schiffrin EL (2000): Endothelin-1 attenuates n-3 fatty acid-induced apoptosis by inhibition of caspase 3. Hypertension 35: 278-291.
54. Calder PC (2006): N-3 polyunsaturated fatty acids, inflammation, and inflammatory diseases. Am J Clin Nutr 83 (6 Suppl): 1505S-1519S.
55. Zhao Y, Joshi-Barve S, Barve S et al. (2004): Eicosapentaenoic acid prevents LPS-induced TNF-αlpha expression by preventing NF-kappaB activation. J Am Coll Nutr 23: 71-78.
56. Weldon SM, Mullen AC, Loscher CE et al. (2007): Docosahexaenoic acid induces an anti-inflammatory profile in lipopolysaccharide-stimulated human THP-1 macrophages more effectively than eicosapentaenoic acid. J Nutr Biochem 18: 250-258.
57. Novak TE, Babcock TA, Jho DH et al. (2003): NF-kappa B inhibition by omega -3 fatty acids modulates LPS-stimulated macrophage TNF-αlpha transcription. Am J Physiol Lung Cell Mol Physiol 284: L84-L89.
58. Moon DO, Kim KC, Jin CY et al. (2007): Inhibitory effects of eicosapentaenoic acid on lipopolysaccharide-induced activation in BV2 microglia. Int Immunopharmacol 7: 222-229.
59. Wendel M, Paul R, Heller AR (2007): Lipoproteins in inflammation and sepsis. II. Clinical aspects. Intensive Care Med 33: 25-35.
60. Mishra A, Chaudhary A, Sethi S (2004): Oxidized omega-3 fatty acids inhibit NFkappaB activation via a PPARalpha-dependent pathway. Arterioscler Thromb Vasc Biol 24: 1621-1627.
61. Li H, Ruan XZ, Powis SH et al. (2005): EPA and DHA reduce LPS-induced inflammation responses in HK-2 cells: evidence for a PPAR-gamma-dependent mechanism. Kidney Int 67: 867-874.
62. Ponte-Arruda A, Aragao AM, Albuquerque JD (2006): Effects of enteral feeding with Eicosapentaenoic acid, gamma-linolenic acid, and antioxidants in mechanically ventiled patients with severe sepsis and septic shock. Crit Care Med 34: 2325-2333.
63. Singer P, Theilla M, Fisher H et al. (2006): Benefit of an enteral diet enriched with eicosapentaenoic acid and gamma-linolenic acid in ventilated patients with acute lung injury. Crit Care Med 34: 1033-1038.
64. Mayer K, Fegbeutel C, Hattar K et al. (2003): Omega-3 vs. Omega-6 lipid emulsions exert differential influence on neutrophils in septic shock patients: impact on plasma fatty acids and lipid mediator generation. Intensive Care Med 29: 1472-1481.
65. Mayer K, Gokorsch S, Fegbeutel C et al. (2003): Parenteral nutrition with fish oil modulates cytokine response in patients with sepsis. Am J Respir Crit Care Med 167: 1321-1328.
66. Słotwiński R, Olszewski WL, Słodkowski M et al. (2007): Can the Interleukin-1 Receptor Antagonist (IL-1ra) Be a Marker of Anti-Inflammatory Response to Enteral Immunonutrition in Malnourished Patients after Pancreaticoduodenectomy?
J Pancreas 8: 759-769.
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