Hostname: page-component-76fb5796d-22dnz Total loading time: 0 Render date: 2024-04-26T20:43:53.526Z Has data issue: false hasContentIssue false
  • English
  • Français

Characterization of immune-active peptides obtained from milk fermented by Lactobacillus helveticus

Published online by Cambridge University Press:  18 January 2010

Angela Tellez ,
Milena Corredig ,
Lubov Y Brovko  and
Mansel W Griffiths
Angela Tellez
Affiliation:
Canadian Research Institute for Food Safety, University of Guelph, Guelph, ON N1G 2W1, Canada
Milena Corredig
Affiliation:
Department of Food Science, University of Guelph, Guelph, ON N1G 2W1, Canada
Lubov Y Brovko
Affiliation:
Canadian Research Institute for Food Safety, University of Guelph, Guelph, ON N1G 2W1, Canada
Mansel W Griffiths*
Affiliation:
Canadian Research Institute for Food Safety, University of Guelph, Guelph, ON N1G 2W1, Canada
*
*For correspondence; e-mail: mgriffit@uoguelph.ca
Get access Rights & Permissions [Opens in a new window]

Abstract

The objectives of this research were to confirm the effect of compounds derived from milk fermented by Lactobacillus helveticus (LH-2) on the nonspecific host defence system, and isolate and characterize the active peptides that mediate the immune response. The cell-free supernatant obtained from the fermented milk and its fractions were tested in vitro for immuno-modulating activity using murine macrophages (RAW 264·7 cell line). Cytokine production (Interleukin-6 (IL-6), Tumor Necrosis Factor-α (TNF-α), and Interleukin-1β (IL1-β)), nitric oxide (NO) production and phagocytosis were used as biomarkers. Macrophages stimulated with cell-free supernatant of fermented milk showed higher production of cytokines and NO compared with macrophages stimulated with LPS (Lipopolysaccharide) and a commercial immunomodulator derived from β-casein (f54-59). Phagocytosis was observed by macrophages stimulated with the supernatant. Two of nine fractions collected from the supernatant using size exclusion chromatography produced the highest response when used to stimulate macrophages. The results of the dose-response study of the effect of the fraction with the highest stimulation effect on the production of TNF-α showed a direct correlation between protein concentration and TNF-α release. The fraction contained four novel peptides, three derived from the hydrolysis of β-casein and one from the hydrolysis of α-lactalbumin. These results confirm that fermentation of milk by Lactobacillus helveticus (LH-2) results in the production of specific peptides capable of modulating macrophage activity.

Keywords

Immunomodulationfermented milkMacrophagesbioactive peptides
Type
Research Article
Information
Journal of Dairy Research , Volume 77 , Issue 2 , May 2010 , pp. 129 - 136
Copyright
Copyright © Proprietors of Journal of Dairy Research 2010

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Abbas, AK, Lichtman, AH & Pobe, JS 1994 Cytokines: Cellular and Molecular Immunology. 2nd Edition. W.B. Sanders Co., pp. 240. Philadelphia PA, USAGoogle Scholar
Aderem, A & Underhill, DM 1999 Mechanisms of Phagocytosis in Macrophages. Annual Reviews of Immunology 17 593–623CrossRefGoogle ScholarPubMed
Brovko, L, Vandenende, C, Chu, B, NG, K, Brooks, A & Griffiths, MW 2003 In Vivo Assessment of Effect of Fermented Milk Diet on Course of Infection in Mice with Bioluminescent Salmonella. Journal of Food Protection 66 2160–2163CrossRefGoogle ScholarPubMed
Christensen, JE, Broadbent, JR & Steele, JL 2003 Hydrolysis of Casein-Derived Peptides αS1-Casein(f1-9) and β-Casein(f193-209) by Lactobacillus helveticus Peptidase Deletion Mutants Indicates the Presence of a Previously Undetected Endopeptidase. Environmental Microbiology 69 1283–1286CrossRefGoogle Scholar
Coste, M, Rochet, V, Lonil, J, Molle, D, Bouhallab, S & Tome, D 1992 Identification of C-terminal peptides of bovine//-casein that enhance proliferation of rat lymphocytes. Immunology Letters 33 41–46CrossRefGoogle Scholar
De LeBlanc, A, Matar, C, LeBlanc, N & Perdigon, G 2005 Effects of milk fermented by Lactobacillus helveticus R389 on a murine breast cancer model. Breast Cancer Research 7 477–486CrossRefGoogle Scholar
De LeBlanc, A, Chaves, S, Carmuega, E, Weill, R, Antoine, J & Perdigon, G 2008 Effect of long-term continuous consumption of fermented milk containing probiotic bacteria on mucosal immunity and the activity of peritoneal macrophages. Immunobiology 213 97–108CrossRefGoogle Scholar
de Moreno de LeBlanc, A, Chaves, S, Carmuega, E, Weill, R, Antoine, J & Perdigon, G 2008 Effect of long-term continuous consumption of fermented milk containing probiotic bacteria on mucosal immunity and the activity of peritoneal macrophages. Immunobiology 213 97–108CrossRefGoogle ScholarPubMed
Deutsch, S, Molle, D, Gagnaire, V, Piot, M, Atlan, D & Lortal, S 2000 Hydrolysis of Sequenced β-Casein Peptides Provides New Insight into Peptidase Activity from Thermophilic Lactic Acid Bacteria and Highlights Intrinsic Resistance of Phosphopeptides. Applied and Environmental Microbiology 66 5360–5367CrossRefGoogle ScholarPubMed
FitzGerald, RJ & Meisel, H 2003 Milk protein hydrolysates and bioactive peptides. In Fox, PF and McSweeney, PLH (eds). Advanced dairy chemistry, Vol. 1: Proteins. 3rd Edition. New York, NY, USA: Kluwer AcademicGoogle Scholar
Gasteiger, E, Hoogland, C, Gattiker, A, Duvaud, S, Wilkins, MR, Appel, RD & Bairoch, A 2005 In Walker, JM (eds). Protein Identification and Analysis Tools on the ExPASy Server. Totowa, NJ, USA: Humana Press Inc.CrossRefGoogle Scholar
Herbert, E, Rayas, R & de Giori, G 1997 Characterization of a Cell Membrane-Associated Proteinase from Lactobacillus helveticus CRL 581. Current Microbiology 35 161–164Google Scholar
Jeffrey, B, Van Ostade, X & Lopez, A 1996 Tumour necrosis factor-alpha (TNF-α): The good, the bad and potentially very effective. Immunology and Cell Biology 74 434–443Google Scholar
Jolles, P & Migliore-Samour, D 1988 Casein, a prohormone with an immunomodulasting role for the newborn? Cellular and Molecular Life Science 44 188–193Google Scholar
Kagan, A, Yu, Z, Fishman, G & McDonald, T 2000 The Dominant Negative LQT2 Mutation A561V Reduces Wild-type HERG Expression. The Journal of Biological Chemistry 275 11241–11248CrossRefGoogle ScholarPubMed
Kenny, O, FitzGerald, RJ, O'Cuinn, G, Beresford, T & Jordan, K 2003 Growth phase and growth medium effects on the peptidase activities of Lactobacillus helveticus. International Dairy Journal 13 509–516CrossRefGoogle Scholar
LeBlanc, JG, Matar, C, Valdez, JC, LeBlanc, J & Perdigon, G 2002 Immunomodulating Effects of Peptidic Fractions Issued from Milk Fermented with Lactobacillus helveticus. Journal of Dairy Science 85 2733–2742CrossRefGoogle ScholarPubMed
LeBlanc, J, Fliss, I & Matar, C 2004 Induction of a Humoral Immune Response following an Escherichia coli O157:H7 Infection with an Immunomodulatory Peptidic Fraction Derived from Lactobacillus helveticus-Fermented Milk. Clinical and Diagnostic Laboratory Immunology 11 1171–1181Google ScholarPubMed
Matar, C, Valdez, JC, Medina, M, Rachid, M & Perdigon, G 2001 Immunomodulating effects of milks fermented by Lactobacillus helveticus and its non-proteolytic variant. Journal of Dairy Research 68 601–609CrossRefGoogle ScholarPubMed
Matar, C, LeBlanc, JG, Martin, L & Perdigon, G 2003 Biologically active peptides released from fermented milk: role and functions, In Farnworth, ER (eds), Handbook of fermented functional foods. 1rst Edition. Boca Raton, FL, USA: CRC PressGoogle Scholar
Meisel, H & Bockelmann, W 1999 Bioactive peptides encrypted in milk proteins: proteolytic activation and thropho-functional properties. Antoine van Leeuwenhoek 76 207–315CrossRefGoogle ScholarPubMed
Miettinen, M, Vuopio-varkila, J & Varkila, K 1996 Production of Human Tumor Necrosis Factor Alpha, Interleukin-6 and Interleukin-10 is induced by Lactic Acid Bacteria. Infection and Immunity 64 5403–5405CrossRefGoogle ScholarPubMed
Netea, MG, van der Meer, J, van Deuren, M & Kullberg, B 2003 Proinflammatory cytokines and sepsis syndrome: not enough, or too much of a good thing? Trends in Immunology 24 254–258CrossRefGoogle ScholarPubMed
Ng, KY & Grifffiths, MW 2002 Enhancement of macrophage cytokine release by cell-free fractions o fermented milk. Milchwissenschaft 57 66–70Google Scholar
Parker, D, Migliore-Samour, D, Floch, F, Zerial, A, Werner, G, Jolles, J, Casaretto, M, Zahn, H & Jolles, P 1984 Immunostimulating hexapeptide from human casein: amino acid sequence, synthesis and biological properties. European Journal of Biochemistry 45 677–682CrossRefGoogle Scholar
Perdigon, G, Fuller, R & Rayas, R 2001 Lactic Acid Bacteria and their Effect on the Immune System. Current issues in intestinal microbiology 2 27–42Google ScholarPubMed
Perkins, D, Pappin, D, Creasy, D & Cottrell, J 1999 Probability-based protein identification by searching sequence databases using mass spectrometry data. Electrophoresis 20 3551–35673.0.CO;2-2>CrossRefGoogle ScholarPubMed
Sandre, C, Gleizes, A, Foresties, F, Gorges-Kergot, R, Chilmonczyk, S, Leonil, J, Moreau, MC & Labarre, C 2001 A Peptide Derived from Bovine b-Casein Modulates Functional Properties of Bone Marrow-Derived Macrophages from Germfree and Human Flora-Associated Mice. The Journal of Nutrition 131 2936–2942CrossRefGoogle Scholar
Takahashi, T, Oka, T, Iwana, H, Kuwata, T & Yamamoto, Y 1993 Immune response of mice to orally administered lactic acid bacteria. Bioscience, biotechnology, and biochemistry 57 1557–1560CrossRefGoogle Scholar
Tejada-Simon, MV & Pestka, J 1999 Proinflamatory cytokine and nitric oxide induction in murine macrophages by cell wall and cytoplasmic extracs of Lactic acid bacteria. Journal of Food Protection 62 1435–1444CrossRefGoogle Scholar
Tsao, L, Lin, Ch & Wang, J 2004 Justicidin A Inhibits the Transport of Tumor Necrosis Factor-α to Cell Surface in Lipopolysaccharide-Stimulated RAW 264·7 Macrophages. Molecular pharmacology 65 1063–1069CrossRefGoogle ScholarPubMed
Urban, J, Shepard, HM, Rothstein, JL, Sugarman, BJ & Schreiber, H 1986 Tumor necrosis factor: A potent effector molecule for tumor cell killing by activated macrophages. Immunology 83 5233–5237Google ScholarPubMed
Ustundag, B, Yilmaz, E, Dogan, Y, Akarsu, S, Canatan, H, Halifeoglu, I, Cikim, G & Aygun, D 2005 Levels of Cytokines (IL-1β, IL-2, IL-6, IL-8, TNF-α) and Trace Elements (Zn, Cu) in Breast Milk From Mothers of Preterm and Term Infants. Mediators of Inflammation 6 331–336CrossRefGoogle Scholar
Vinderola, G, Matar, C & Perdigon, G 2007a Milk fermentation products of L. helveticus R389 activate calcineurin as a signal to promote gut mucosal immunity. Immunology 8 1–10Google ScholarPubMed
Vinderola, G, Matar, C, Palacios, J & Perdigon, G 2007b Mucosal immunomodulation by the non-bacterial fraction of milk fermented by L. Helveticus R389. International Journal of Food Microbiology 115 180–186CrossRefGoogle Scholar
Vinderola, G, Matar, C & Perdigon, G 2007c Milk fermented by Lactobacillus helveticus R389 and its non-bacterial fraction confer enhanced protection against Salmonella enteritidis serovar Typhimurium infection in mice. Immunobiology 212 107–118CrossRefGoogle ScholarPubMed
Yasutaka, A & Kiyoshi, O 2002 Endomorphin-2 Modulates Productions of TNF-α, IL-1β, IL-10, and IL-12, and Alters Functions Related to Innate Immune of Macrophages. Inflammation 26 223–232Google Scholar