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Functional & Integrative Genomics

, Volume 14, Issue 4, pp 657–671 | Cite as

Gut response induced by weaning in piglet features marked changes in immune and inflammatory response

  • Lorenzo Bomba
  • Andrea Minuti
  • Sonia J. Moisá
  • Erminio Trevisi
  • Elisa Eufemi
  • Michela Lizier
  • Fatima Chegdani
  • Franco Lucchini
  • Marcin Rzepus
  • Aldo Prandini
  • Filippo Rossi
  • Raffaele Mazza
  • Giuseppe Bertoni
  • Juan J. Loor
  • Paolo Ajmone-Marsan
Original Paper

Abstract

At weaning, piglets are exposed to many stressors, such as separation from the sow, mixing with other litters, end of lactational immunity, and a change in their environment and gut microbiota. The sudden change of feeding regime after weaning causes morphological and histological changes in the small intestine which are critical for the immature digestive system. Sixteen female piglets were studied to assess the effect of sorbic acid supplementation on the small intestine tissue transcriptome. At weaning day (T0, piglet age 28 days), four piglets were sacrificed and ileal tissue samples collected. The remaining 12 piglets were weighed and randomly assigned to different postweaning (T5, piglet age 33 days) diets. Diet A (n = 6) contained 5 g/kg of sorbic acid. In diet B (n = 6), the organic acids were replaced by barley flour. Total RNA was isolated and then hybridized to CombiMatrix CustomArray™ 90-K platform microarrays, screening about 30 K genes. Even though diet had no detectable effect on the transcriptome during the first 5 days after weaning, results highlighted some of the response mechanisms to the stress of weaning occurring in the piglet gut. A total of 205 differentially expressed genes were used for functional analysis using the bioinformatics tools BLAST2GO, Ingenuity Pathway Analysis 8.0, and Dynamic Impact Approach (DIA). Bioinformatic analysis revealed that apoptosis, RIG-I-like, and NOD-like receptor signaling were altered as a result of weaning. Interferons and caspases gene families were the most activated after weaning in response to piglets to multiple stressors. Results suggest that immune and inflammatory responses were activated and likely are a cause of small intestine atrophy as revealed by a decrease in villus height and villus/crypt ratio.

Keywords

Transcriptomics Weaning Pig Inflammation Gut Differential gene expression 

Notes

Acknowledgments

We thank Alberto Ferrarini (Functional Genomics Center, University of Verona, Italy) for his technical assistance in the microarray experiment. This work was conducted in the framework of the project “Nutrigenomics” supported by the “Fondazione Romeo ed Enrica Invernizzi,” Milan, Italy.

Supplementary material

10142_2014_396_MOESM1_ESM.pdf (275 kb)
Online Resource 1 Principal Component Analysis (PCA) for clusterig gene expression data: a) Principal Component Analysis (PCA) for clusterig gene expression data of 12 selected piglet. In the analysis were included the 4 piglets of T0 group (colored in yellow) , the 4 healthies animal of T5 group (colored in red) and the 4 most diarrheal subjects of T5 group (colored in blue). The animals were selected based on the clinical and hematological measurament. b) percentage of the total variance explained by each principal component. (PDF 275 kb)
10142_2014_396_MOESM2_ESM.xls (108 kb)
Online Resource 2 Differentially expressed genes (DGE) comparing T5 with T0: List of the 205 DGE comparing T5 with T0 together with log fold-change and qvalue (XLS 107 kb)
10142_2014_396_MOESM3_ESM.pdf (187 kb)
Online Resource 3 Differentially expressed genes (DGE) mapped and annotated using the IPA software: Differentially expressed genes (DGE) found in the ileum of piglets, in response to weaning that were mapped and annotated using the IPA software. The accession number derived from a blast to the homology sequence in human NCBI nr database. (PDF 186 kb)
10142_2014_396_MOESM4_ESM.pdf (92 kb)
Online Resource 4 Networks ranking: Networks ranking with relative molecules symbols list, Score value, number of molecules belonging to the dataset (Focus molecules) and top function for each network. (PDF 91 kb)
10142_2014_396_MOESM5_ESM.xls (1.3 mb)
Online Resource 5 KEGG pathways graphs: KEGG pathways graphs with the fold change value of the DEG denoted with colour scales. (XLS 1343 kb)
10142_2014_396_MOESM6_ESM.xls (132 kb)
Online Resource 6 Dynamic Impact Approach (DIA) based on DAVID and Interpro (IP): Results of flux and impact of DAVID Biological Process (BP), Cellular Component (CC), Molecular Function (MF), and IP uncovered by the DIA with impact value above the 30 % of the maximum total impact in swine transcriptome. (XLS 131 kb)
10142_2014_396_MOESM7_ESM.xls (68 kb)
Online Resource 7 Connections between the most relevant GO Terms from DIA results: Connections between the most relevant GO Terms from DIA results for each DEG present in the most impacted KEGG pathways. each gene is associated with the KEGG pathway where it has a role, and each GO Term (biological processes, molecular function, cellular component, and Interpro) sorted from higher to lower impact. (XLS 68 kb)

References

  1. Benjamini Y, Hochberg Y (1995) Controlling the false discovery rate: a practical and powerful approach to multiple testing. J R Stat Soc Ser B 57(1):289–300Google Scholar
  2. Bionaz M, Periasamy K, Rodriguez-Zas SL, Everts RE, Lewin HA, Hurley WL, Loor JJ (2012a) Old and new stories: revelations from functional analysis of the bovine mammary transcriptome during the lactation cycle. PLoS One 7:e33268. doi: 10.1371/journal.pone.0033268 PONE-D-12-02397 PubMedCentralPubMedCrossRefGoogle Scholar
  3. Bionaz M, Periasamy K, Rodriguez-Zas SL, Hurley WL, Loor JJ (2012b) A novel dynamic impact approach (DIA) for functional analysis of time-course omics studies: validation using the bovine mammary transcriptome. PLoS One 7:e32455. doi: 10.1371/journal.pone.0032455 PONE-D-11-19484 PubMedCentralPubMedCrossRefGoogle Scholar
  4. Bolstad BM, Irizarry RA, Astrand M, Speed TP (2003) A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics 19:185–193PubMedCrossRefGoogle Scholar
  5. Boudry G, Peron V, Le Huerou-Luron I, Lalles JP, Seve B (2004) Weaning induces both transient and long-lasting modifications of absorptive, secretory, and barrier properties of piglet intestine. J Nutr 134:2256–2262PubMedGoogle Scholar
  6. Burnell TW, Cromwell GL, Stahly TS (1988) Effects of dried whey and copper sulfate on the growth responses to organic acid in diets for weanling pigs. J Anim Sci 66:1100–1108PubMedGoogle Scholar
  7. Caswell JL, Middleton DM, Gordon JR (2001) The importance of interleukin-8 as a neutrophil chemoattractant in the lungs of cattle with pneumonic pasteurellosis. Can J Vet Res 65:229–232PubMedCentralPubMedGoogle Scholar
  8. Dieckgraefe BK, Stenson WF, Korzenik JR, Swanson PE, Harrington CA (2000) Analysis of mucosal gene expression in inflammatory bowel disease by parallel oligonucleotide arrays. Physiol Genomics 4:1–11PubMedGoogle Scholar
  9. Donovan J, Dufner M, Korennykh A (2013) Structural basis for cytosolic double-stranded RNA surveillance by human oligoadenylate synthetase 1. Proc Natl Acad Sci U S A 110:1652–1657. doi: 10.1073/pnas.1218528110 PubMedCentralPubMedCrossRefGoogle Scholar
  10. Evans ME, Jones DP, Ziegler TR (2003) Glutamine prevents cytokine-induced apoptosis in human colonic epithelial cells. J Nutr 133:3065–3071PubMedGoogle Scholar
  11. Gu YR, Li MZ, Zhang K, Chen L, Jiang AA, Wang JY, Li XW (2011) Evaluation of endogenous control genes for gene expression studies across multiple tissues and in the specific sets of fat- and muscle-type samples of the pig. J Anim Breed Genet 128:319–325PubMedCrossRefGoogle Scholar
  12. Haas MJ, Mazza AD, Wong NC, Mooradian AD (2011) Inhibition of apolipoprotein A-I gene expression by obesity-associated endocannabinoids. Obesity (Silver Spring) 20:323Google Scholar
  13. Haller O, Kochs G, Weber F (2007) Interferon, Mx, and viral countermeasures. Cytokine Growth Factor Rev 18:425–433PubMedCrossRefGoogle Scholar
  14. Hampson DJ, Kidder DE (1986) Influence of creep feeding and weaning on brush border enzyme activities in the piglet small intestine. Res Vet Sci 40:24–31PubMedGoogle Scholar
  15. Hampson DJ, Smith WC (1986) Influence of creep feeding and dietary intake after weaning on malabsorption and occurrence of diarrhoea in the newly weaned pig. Res Vet Sci 41:63–69PubMedGoogle Scholar
  16. Hou F, Sun L, Zheng H, Skaug B, Jiang QX, Chen ZJ (2011) MAVS forms functional prion-like aggregates to activate and propagate antiviral innate immune response. Cell 146:448–461PubMedCentralPubMedCrossRefGoogle Scholar
  17. Kato H et al (2008) Length-dependent recognition of double-stranded ribonucleic acids by retinoic acid-inducible gene-I and melanoma differentiation-associated gene 5. J Exp Med 205:1601–1610. doi: 10.1084/jem.20080091 PubMedCentralPubMedCrossRefGoogle Scholar
  18. Krejs GJ (1986) Physiological role of somatostatin in the digestive tract: gastric acid secretion, intestinal absorption, and motility. Scand J Gastroenterol Suppl 119:47–53PubMedCrossRefGoogle Scholar
  19. Lalles JP, Bosi P, Smidt H, Stokes CR (2007) Nutritional management of gut health in pigs around weaning. Proc Nutr Soc 66:260–268PubMedCrossRefGoogle Scholar
  20. Luo ZF et al (2011) Sorbic acid improves growth performance and regulates insulin-like growth factor system gene expression in swine. J Anim Sci 89:2356–2364PubMedCrossRefGoogle Scholar
  21. McCracken BA, Gaskins HR, Ruwe-Kaiser PJ, Klasing KC, Jewell DE (1995) Diet-dependent and diet-independent metabolic responses underlie growth stasis of pigs at weaning. J Nutr 125:2838–2845PubMedGoogle Scholar
  22. McCracken BA, Spurlock ME, Roos MA, Zuckermann FA, Gaskins HR (1999) Weaning anorexia may contribute to local inflammation in the piglet small intestine. J Nutr 129:613–619PubMedGoogle Scholar
  23. McLamb BL, Gibson AJ, Overman EL, Stahl C, Moeser AJ (2013) Early weaning stress in pigs impairs innate mucosal immune responses to enterotoxigenic E. coli challenge and exacerbates intestinal injury and clinical disease. PLoS One 8:e59838PubMedCentralPubMedCrossRefGoogle Scholar
  24. Mettlen M, Pucadyil T, Ramachandran R, Schmid SL (2009) Dissecting dynamin’s role in clathrin-mediated endocytosis. Biochem Soc Trans 37:1022–1026. doi: 10.1042/BST0371022 BST0371022 PubMedCentralPubMedCrossRefGoogle Scholar
  25. Palm M, Garigliany MM, Cornet F, Desmecht D (2010) Interferon-induced Sus scrofa Mx1 blocks endocytic traffic of incoming influenza A virus particles. Vet Res 41:29PubMedCentralPubMedCrossRefGoogle Scholar
  26. Partanen KH, Mroz Z (1999) Organic acids for performance enhancement in pig diets. Nutr Res Rev 12:117–145PubMedCrossRefGoogle Scholar
  27. Pfaffl MW, Horgan GW, Dempfle L (2002) Relative expression software tool (REST) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res 30:e36PubMedCentralPubMedCrossRefGoogle Scholar
  28. Pie S, Lalles JP, Blazy F, Laffitte J, Seve B, Oswald IP (2004) Weaning is associated with an upregulation of expression of inflammatory cytokines in the intestine of piglets. J Nutr 134:641–647PubMedGoogle Scholar
  29. Piva A, Grilli E, Fabbri L, Pizzamiglio V, Campani I (2007) Free versus microencapsulated organic acids in medicated or not medicated diet for piglets livestock. Science 108:214–217Google Scholar
  30. Pluske JR, Thompson MJ, Atwood CS, Bird PH, Williams IH, Hartmann PE (1996) Maintenance of villus height and crypt depth, and enhancement of disaccharide digestion and monosaccharide absorption, in piglets fed on cows' whole milk after weaning. Br J Nutr 76:409–422PubMedCrossRefGoogle Scholar
  31. Pluske J, Hampson D, Williams I (1997) Factors influencing the structure and function of the small intestine in the weaned pig: a review. Livest Prod Sci 51:215–236CrossRefGoogle Scholar
  32. Risley CR, Kornegay ET, Lindemann MD, Wood CM, Eigel WN (1992) Effect of feeding organic acids on selected intestinal content measurements at varying times postweaning in pigs. J Anim Sci 70:196–206PubMedGoogle Scholar
  33. Rouillard JM, Zuker M, Gulari E (2003) OligoArray 2.0: design of oligonucleotide probes for DNA microarrays using a thermodynamic approach. Nucleic Acids Res 31:3057–3062PubMedCentralPubMedCrossRefGoogle Scholar
  34. Schroyen M, Stinckens A, Verhelst R, Geens M, Cox E, Niewold T, Buys N (2012) Susceptibility of piglets to enterotoxigenic Escherichia coli is not related to the expression of MUC13 and MUC20. Anim Genet 43:324–327. doi: 10.1111/j.1365-2052.2011.02241.x PubMedCrossRefGoogle Scholar
  35. Sever S, Damke H, Schmid SL (2000) Dynamin: GTP controls the formation of constricted coated pits, the rate limiting step in clathrin-mediated endocytosis. J Cell Biol 150:1137–1148PubMedCentralPubMedCrossRefGoogle Scholar
  36. Skinner JG, Brown RA, Roberts L (1991) Bovine haptoglobin response in clinically defined field conditions. Vet Rec 128:147–149PubMedCrossRefGoogle Scholar
  37. Sodek J, Batista Da Silva AP, Zohar R (2006) Osteopontin and mucosal protection. J Dent Res 85:404–415PubMedCrossRefGoogle Scholar
  38. Spreeuwenberg MA, Verdonk JM, Gaskins HR, Verstegen MW (2001) Small intestine epithelial barrier function is compromised in pigs with low feed intake at weaning. J Nutr 131:1520–1527PubMedGoogle Scholar
  39. Sunderman FW Jr, Nomoto S (1970) Measurement of human serum ceruloplasmin by its p-phenylenediamine oxidase activity. Clin Chem 16:903–910PubMedGoogle Scholar
  40. Tamassia N et al (2008) Activation of an immunoregulatory and antiviral gene expression program in poly(I:C)-transfected human neutrophils. J Immunol 181:6563–6573PubMedCrossRefGoogle Scholar
  41. Thornton JR, Willoughby RA, McSherry BJ (1972) Studies on diarrhea in neonatal calves: the plasma proteins of normal and diarrheic calves during the first ten days of age. Can J Comp Med 36:17–25PubMedCentralPubMedGoogle Scholar
  42. Tsiloyiannis VK, Kyriakis SC, Vlemmas J, Sarris K (2001) The effect of organic acids on the control of porcine post-weaning diarrhoea. Res Vet Sci 70:287–293PubMedCrossRefGoogle Scholar
  43. van Beers-Schreurs HM, Nabuurs MJ, Vellenga L, der Valk HJ K-v, Wensing T, Breukink HJ (1998) Weaning and the weanling diet influence the villous height and crypt depth in the small intestine of pigs and alter the concentrations of short-chain fatty acids in the large intestine and blood. J Nutr 128:947–953PubMedGoogle Scholar
  44. Vega-Lopez MA, Bailey M, Telemo E, Stokes CR (1995) Effect of early weaning on the development of immune cells in the pig small intestine. Vet Immunol Immunopathol 44:319–327PubMedCrossRefGoogle Scholar
  45. Wang J et al (2008) Gene expression is altered in piglet small intestine by weaning and dietary glutamine supplementation. J Nutr 138:1025–1032PubMedCrossRefGoogle Scholar
  46. Wies E, Wang MK, Maharaj NP, Chen K, Zhou S, Finberg RW, Gack MU (2013) Dephosphorylation of the RNA sensors RIG-I and MDA5 by the phosphatase PP1 is essential for innate immune signaling. Immunity 38:437–449. doi: 10.1016/j.immuni.2012.11.018 S1074-7613(13)00095-2 PubMedCentralPubMedCrossRefGoogle Scholar
  47. Zentek J, Ferrara F, Pieper R, Tedin L, Meyer W, Vahjen W (2013) Effects of dietary combinations of organic acids and medium chain fatty acids on the gastrointestinal microbial ecology and bacterial metabolites in the digestive tract of weaning piglets. J Anim Sci 91:3200–3210PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Lorenzo Bomba
    • 1
    • 2
  • Andrea Minuti
    • 1
    • 2
  • Sonia J. Moisá
    • 3
  • Erminio Trevisi
    • 1
    • 2
  • Elisa Eufemi
    • 1
  • Michela Lizier
    • 4
    • 2
  • Fatima Chegdani
    • 5
    • 2
  • Franco Lucchini
    • 6
    • 2
  • Marcin Rzepus
    • 7
  • Aldo Prandini
    • 7
    • 2
  • Filippo Rossi
    • 7
    • 2
  • Raffaele Mazza
    • 8
  • Giuseppe Bertoni
    • 1
    • 2
  • Juan J. Loor
    • 3
  • Paolo Ajmone-Marsan
    • 1
    • 2
  1. 1.Istituto di ZootecnicaUniversità Cattolica del Sacro CuorePiacenzaItaly
  2. 2.Centro di Ricerca Nutrigenomica e Proteomica—PRONUTRIGENUniversità Cattolica del Sacro CuorePiacenzaItaly
  3. 3.Mammalian NutriPhysioGenomicsUniversity of IllinoisUrbanaUSA
  4. 4.Istituto Clinico Humanitas IRCSSRozzanoItaly
  5. 5.Laboratoire de Physiologie et Génétique MoléculaireUniversité Hassan II Aïn ChockCasablancaMorocco
  6. 6.Centro Ricerche Biotecnologiche, Istituto di MicrobiologiaUniversità Cattolica del Sacro CuoreCremonaItaly
  7. 7.Istituto di Scienze degli Alimenti e della NutrizioneUniversità Cattolica del Sacro CuorePiacenzaItaly
  8. 8.Associazione Italiana AllevatoriRomeItaly

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