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Vol. 99, No. 3, 2011
Issue release date: March 2011
Section title: Original Paper
Free Access
Neonatology 2011;99:192–201
(DOI:10.1159/000314109)

Breast Milk-Transforming Growth Factor-β2 Specifically Attenuates IL-1β-Induced Inflammatory Responses in the Immature Human Intestine via an SMAD6- and ERK-Dependent Mechanism

Rautava S.a, b · Nanthakumar N.N.a · Dubert-Ferrandon A.a · Lu L.a · Rautava J.a · Walker W.A.a
aDevelopmental Gastroenterology Laboratory, Division of Pediatric Gastroenterology, Massachusetts General Hospital for Children, Charlestown, Mass., USA; bDepartment of Paediatrics, Turku University Central Hospital, Turku, Finland
email Corresponding Author

Abstract

Background: Breast milk is known to protect the infant against infectious and immuno-inflammatory diseases, but the mechanisms of this protection are poorly understood. Objectives: We hypothesized that transforming growth factor-β2 (TGF-β2), an immunoregulatory cytokine abundant in breast milk, may have a direct anti-inflammatory effect on immature human intestinal epithelial cells (IECs). Methods: Human fetal ileal organ culture, primary human fetal IECs, and the human fetal small intestinal epithelial cell line H4 were stimulated with interleukin 1β (IL-1β) with or without TGF-β2. Pro-inflammatory cytokine secretion and mRNA expression were measured by ELISA and quantitative real-time polymerase chain reaction, respectively. Alterations in ERK signalling were detected from IECs by immunoblotting and in fetal ileal tissue culture by immunohistochemistry. SMAD6 knockdown was performed by transfecting the cells with SMAD6 siRNA. Results: TGF-β2 significantly attenuated IL-1β-induced pro-inflammatory cytokine production in fetal intestinal organ culture and the cell culture models. In addition, TGF-β2 reduced the IL-1β-induced IL-8 and IL-6 mRNA response in H4 cells. TGF-β2 markedly inhibited IL-1β-induced phosphorylation of ERK, which was necessary for the cytokine response. The inhibitory effect of TGF-β2 on IL-1β-induced cytokine production was completely abrogated by SMAD6 siRNA knockdown. Conclusions: TGF-β2 attenuates IL-1β-induced pro-inflammatory cytokine production in immature human IECs by inhibiting ERK signalling. The anti-inflammatory effect of TGF-β2 is dependent on SMAD6. Breast milk TGF-β2 may provide the neonate with important immunoregulatory support. TGF-β2 might provide a novel means to improve intestinal immunophysiology in premature neonates.

© 2010 S. Karger AG, Basel


  

Key Words

  • Transforming growth factor-β
  • Immature intestinal epithelium
  • Modulation of immune response

References

  1. Holt PG, Jones CA: The development of the immune system during pregnancy and early life. Allergy 2000;55:688–697.
  2. Quigley MA, Henderson G, Anthony MY, McGuire W: Formula milk versus donor breast milk for feeding preterm or low birth weight infants. Cochrane Database Syst Rev 2007:CD002971.
  3. Rautava S, Walker WA: Academy of Breastfeeding Medicine founder’s lecture 2008: breastfeeding – an extrauterine link between mother and child. Breastfeed Med 2009;4:3–10.
  4. Newburg DS, Walker WA: Protection of the neonate by the innate immune system of developing gut and of human milk. Pediatr Res 2007;61:2–8.
  5. Li MO, Wan YY, Sanjabi S, Robertson AK, Flavell RA: Transforming growth factor-β regulation of immune responses. Annu Rev Immunol 2006;24:99–146.
  6. Planchon SM, Martins CA, Guerrant RL, Roche JK: Regulation of intestinal epithelial barrier function by TGF-β1. Evidence for its role in abrogating the effect of a T cell cytokine. J Immunol 1994;153:5730–5739.
  7. Penttila IA, van Spriel AB, Zhang MF, Xian CJ, Steeb CB, Cummins AG, Zola H, Read LC: Transforming growth factor-β levels in maternal milk and expression in postnatal rat duodenum and ileum. Pediatr Res 1998;44:524–531.
  8. Zhang MF, Zola H, Read LC, Penttila IA: Localization of transforming growth factor-β receptor types I, II, and III in the postnatal rat small intestine. Pediatr Res 1999;46:657–665.
  9. Nanthakumar NN, Fusunyan RD, Sanderson I, Walker WA: Inflammation in the developing human intestine: a possible pathophysiologic contribution to necrotizing enterocolitis. Proc Natl Acad Sci USA 2000;97:6043–6048.
  10. Quaroni A, Beaulieu JF: Cell dynamics and differentiation of conditionally immortalized human intestinal epithelial cells. Gastroenterology 1997;113:1198–1213.
  11. Sanderson IR, Ezzell RM, Kedinger M, Erlanger M, Xu ZX, Pringault E, Leon-Robine S, Louvard D, Walker WA: Human fetal enterocytes in vitro: modulation of the phenotype by extracellular matrix. Proc Natl Acad Sci USA 1996;93:7717–7722.
  12. Claud EC, Savidge T, Walker WA: Modulation of human intestinal epithelial cell IL-8 secretion by human milk factors. Pediatr Res 2003;53:419–425.
  13. Rautava J, Jee KJ, Miettinen PJ, Nagy B, Myllykangas S, Odell EW, Soukka T, Morgan PR, Heikinheimo K: ERBB receptors in developing, dysplastic and malignant oral epithelia. Oral Oncol 2008;44:227–235.
  14. Luo X, Zhang Q, Liu V, Xia Z, Pothoven KL, Lee C: Cutting Edge: TGF-β-induced expression of Foxp3 in T cells is mediated through inactivation of ERK. J Immunol 2008;180:2757–2761.
  15. Schmierer B, Hill CS: TGFβ-SMAD signal transduction: molecular specificity and functional flexibility. Nat Rev Mol Cell Biol 2007;8:970–982.
  16. Choi KC, Lee YS, Lim S, Choi HK, Lee CH, Lee EK, Hong S, Kim IH, Kim SJ, Park SH: Smad6 negatively regulates interleukin 1-receptor-Toll-like receptor signaling through direct interaction with the adaptor Pellino-1. Nat Immunol 2006;7:1057–1065.
  17. Ahmad R, Sylvester J, Zafarullah M: MyD88, IRAK1 and TRAF6 knockdown in human chondrocytes inhibits interleukin-1-induced matrix metalloproteinase-13 gene expression and promoter activity by impairing MAP kinase activation. Cell Signal 2007;19:2549–2557.
  18. Verhasselt V, Milcent V, Cazareth J, Kanda A, Fleury S, Dombrowicz D, Glaichenhaus N, Julia V: Breast milk-mediated transfer of an antigen induces tolerance and protection from allergic asthma. Nat Med 2008;14:170–175.
  19. Kalliomaki M, Ouwehand A, Arvilommi H, Kero P, Isolauri E: Transforming growth factor-β in breast milk: a potential regulator of atopic disease at an early age. J Allergy Clin Immunol 1999;104:1251–1257.
  20. Oddy WH, Halonen M, Martinez FD, Lohman IC, Stern DA, Kurzius-Spencer M, Guerra S, Wright AL: TGF-β in human milk is associated with wheeze in infancy. J Allergy Clin Immunol 2003;112:723–728.
  21. Penttila IA, Flesch IE, McCue AL, Powell BC, Zhou FH, Read LC, Zola H: Maternal milk regulation of cell infiltration and interleukin 18 in the intestine of suckling rat pups. Gut 2003;52:1579–1586.
  22. Ozawa T, Miyata M, Nishimura M, Ando T, Ouyang Y, Ohba T, Shimokawa N, Ohnuma Y, Katoh R, Ogawa H, Nakao A: Transforming growth factor-β activity in commercially available pasteurized cow milk provides protection against inflammation in mice. J Nutr 2009;139:69–75.
  23. Tooley KL, El-Merhibi A, Cummins AG, Grose RH, Lymn KA, DeNichilo M, Penttila IA: Maternal milk, but not formula, regulates the immune response to β-lactoglobulin in allergy-prone rat pups. J Nutr 2009;139:2145–2151.
  24. Penttila I: Effects of transforming growth factor-β and formula feeding on systemic immune responses to dietary β-lactoglobulin in allergy-prone rats. Pediatr Res 2006;59:650–655.
  25. Rautava S, Kalliomaki M, Isolauri E: Probiotics during pregnancy and breast-feeding might confer immunomodulatory protection against atopic disease in the infant. J Allergy Clin Immunol 2002;109:119–121.

  

Author Contacts

Samuli Rautava, MD, PhD
Department of Paediatrics
Turku University Central Hospital
Kiinamyllynkatu 4–8, FI–20520 Turku (Finland)
Tel. +358 2 313 0000, Fax +358 2 313 1463, E-Mail samrau@utu.fi

  

Article Information

Received: February 2, 2010
Accepted after revision: April 20, 2010
Published online: September 25, 2010
Number of Print Pages : 10
Number of Figures : 7, Number of Tables : 0, Number of References : 25

  

Publication Details

Neonatology (Fetal and Neonatal Research)

Vol. 99, No. 3, Year 2011 (Cover Date: March 2011)

Journal Editor: Halliday H.L. (Belfast), Speer C.P. (Würzburg)
ISSN: 1661-7800 (Print), eISSN: 1661-7819 (Online)

For additional information: http://www.karger.com/NEO


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References

  1. Holt PG, Jones CA: The development of the immune system during pregnancy and early life. Allergy 2000;55:688–697.
  2. Quigley MA, Henderson G, Anthony MY, McGuire W: Formula milk versus donor breast milk for feeding preterm or low birth weight infants. Cochrane Database Syst Rev 2007:CD002971.
  3. Rautava S, Walker WA: Academy of Breastfeeding Medicine founder’s lecture 2008: breastfeeding – an extrauterine link between mother and child. Breastfeed Med 2009;4:3–10.
  4. Newburg DS, Walker WA: Protection of the neonate by the innate immune system of developing gut and of human milk. Pediatr Res 2007;61:2–8.
  5. Li MO, Wan YY, Sanjabi S, Robertson AK, Flavell RA: Transforming growth factor-β regulation of immune responses. Annu Rev Immunol 2006;24:99–146.
  6. Planchon SM, Martins CA, Guerrant RL, Roche JK: Regulation of intestinal epithelial barrier function by TGF-β1. Evidence for its role in abrogating the effect of a T cell cytokine. J Immunol 1994;153:5730–5739.
  7. Penttila IA, van Spriel AB, Zhang MF, Xian CJ, Steeb CB, Cummins AG, Zola H, Read LC: Transforming growth factor-β levels in maternal milk and expression in postnatal rat duodenum and ileum. Pediatr Res 1998;44:524–531.
  8. Zhang MF, Zola H, Read LC, Penttila IA: Localization of transforming growth factor-β receptor types I, II, and III in the postnatal rat small intestine. Pediatr Res 1999;46:657–665.
  9. Nanthakumar NN, Fusunyan RD, Sanderson I, Walker WA: Inflammation in the developing human intestine: a possible pathophysiologic contribution to necrotizing enterocolitis. Proc Natl Acad Sci USA 2000;97:6043–6048.
  10. Quaroni A, Beaulieu JF: Cell dynamics and differentiation of conditionally immortalized human intestinal epithelial cells. Gastroenterology 1997;113:1198–1213.
  11. Sanderson IR, Ezzell RM, Kedinger M, Erlanger M, Xu ZX, Pringault E, Leon-Robine S, Louvard D, Walker WA: Human fetal enterocytes in vitro: modulation of the phenotype by extracellular matrix. Proc Natl Acad Sci USA 1996;93:7717–7722.
  12. Claud EC, Savidge T, Walker WA: Modulation of human intestinal epithelial cell IL-8 secretion by human milk factors. Pediatr Res 2003;53:419–425.
  13. Rautava J, Jee KJ, Miettinen PJ, Nagy B, Myllykangas S, Odell EW, Soukka T, Morgan PR, Heikinheimo K: ERBB receptors in developing, dysplastic and malignant oral epithelia. Oral Oncol 2008;44:227–235.
  14. Luo X, Zhang Q, Liu V, Xia Z, Pothoven KL, Lee C: Cutting Edge: TGF-β-induced expression of Foxp3 in T cells is mediated through inactivation of ERK. J Immunol 2008;180:2757–2761.
  15. Schmierer B, Hill CS: TGFβ-SMAD signal transduction: molecular specificity and functional flexibility. Nat Rev Mol Cell Biol 2007;8:970–982.
  16. Choi KC, Lee YS, Lim S, Choi HK, Lee CH, Lee EK, Hong S, Kim IH, Kim SJ, Park SH: Smad6 negatively regulates interleukin 1-receptor-Toll-like receptor signaling through direct interaction with the adaptor Pellino-1. Nat Immunol 2006;7:1057–1065.
  17. Ahmad R, Sylvester J, Zafarullah M: MyD88, IRAK1 and TRAF6 knockdown in human chondrocytes inhibits interleukin-1-induced matrix metalloproteinase-13 gene expression and promoter activity by impairing MAP kinase activation. Cell Signal 2007;19:2549–2557.
  18. Verhasselt V, Milcent V, Cazareth J, Kanda A, Fleury S, Dombrowicz D, Glaichenhaus N, Julia V: Breast milk-mediated transfer of an antigen induces tolerance and protection from allergic asthma. Nat Med 2008;14:170–175.
  19. Kalliomaki M, Ouwehand A, Arvilommi H, Kero P, Isolauri E: Transforming growth factor-β in breast milk: a potential regulator of atopic disease at an early age. J Allergy Clin Immunol 1999;104:1251–1257.
  20. Oddy WH, Halonen M, Martinez FD, Lohman IC, Stern DA, Kurzius-Spencer M, Guerra S, Wright AL: TGF-β in human milk is associated with wheeze in infancy. J Allergy Clin Immunol 2003;112:723–728.
  21. Penttila IA, Flesch IE, McCue AL, Powell BC, Zhou FH, Read LC, Zola H: Maternal milk regulation of cell infiltration and interleukin 18 in the intestine of suckling rat pups. Gut 2003;52:1579–1586.
  22. Ozawa T, Miyata M, Nishimura M, Ando T, Ouyang Y, Ohba T, Shimokawa N, Ohnuma Y, Katoh R, Ogawa H, Nakao A: Transforming growth factor-β activity in commercially available pasteurized cow milk provides protection against inflammation in mice. J Nutr 2009;139:69–75.
  23. Tooley KL, El-Merhibi A, Cummins AG, Grose RH, Lymn KA, DeNichilo M, Penttila IA: Maternal milk, but not formula, regulates the immune response to β-lactoglobulin in allergy-prone rat pups. J Nutr 2009;139:2145–2151.
  24. Penttila I: Effects of transforming growth factor-β and formula feeding on systemic immune responses to dietary β-lactoglobulin in allergy-prone rats. Pediatr Res 2006;59:650–655.
  25. Rautava S, Kalliomaki M, Isolauri E: Probiotics during pregnancy and breast-feeding might confer immunomodulatory protection against atopic disease in the infant. J Allergy Clin Immunol 2002;109:119–121.