Skip to main content
Top
Published in: BMC Gastroenterology 1/2015

Open Access 01-12-2015 | Research article

Lamina propria macrophage phenotypes in relation to Escherichia coli in Crohn’s disease

Authors: Timothy R. Elliott, Neil B. Rayment, Barry N. Hudspith, Rebecca E. Hands, Kirstin Taylor, Gareth C. Parkes, Natalie J. Prescott, Liljana Petrovska, John Hermon-Taylor, Jonathan Brostoff, Alex Boussioutas, Christopher G. Mathew, Stephen A. Bustin, Jeremy D. Sanderson

Published in: BMC Gastroenterology | Issue 1/2015

Login to get access

Abstract

Background

Abnormal handling of E. coli by lamina propria (LP) macrophages may contribute to Crohn’s disease (CD) pathogenesis. We aimed to determine LP macrophage phenotypes in CD, ulcerative colitis (UC) and healthy controls (HC), and in CD, to compare macrophage phenotypes according to E. coli carriage.

Methods

Mucosal biopsies were taken from 35 patients with CD, 9 with UC and 18 HCs. Laser capture microdissection was used to isolate E. coli-laden and unladen LP macrophages from ileal or colonic biopsies. From these macrophages, mRNA was extracted and cytokine and activation marker expression measured using RT-qPCR.

Results

E. coli-laden LP macrophages were identified commonly in mucosal biopsies from CD patients (25/35, 71 %), rarely in UC (1/9, 11 %) and not at all in healthy controls (0/18). LP macrophage cytokine mRNA expression was greater in CD and UC than healthy controls. In CD, E. coli-laden macrophages expressed high IL-10 & CD163 and lower TNFα, IL-23 & iNOS irrespective of macroscopic inflammation. In inflamed tissue, E. coli-unladen macrophages expressed high TNFα, IL-23 & iNOS and lower IL-10 & CD163. In uninflamed tissue, unladen macrophages had low cytokine mRNA expression, closer to that of healthy controls.

Conclusion

In CD, intra-macrophage E. coli are commonly found and LP macrophages express characteristic cytokine mRNA profiles according to E. coli carriage. Persistence of E. coli within LP macrophages may provide a stimulus for chronic inflammation.
Literature
1.
go back to reference Smith AM, Rahman FZ, Hayee B, et al. Disordered macrophage cytokine secretion underlies impaired acute inflammation and bacterial clearance in Crohn’s disease. J Exp Med. 2009;206:1883–97.CrossRefPubMedPubMedCentral Smith AM, Rahman FZ, Hayee B, et al. Disordered macrophage cytokine secretion underlies impaired acute inflammation and bacterial clearance in Crohn’s disease. J Exp Med. 2009;206:1883–97.CrossRefPubMedPubMedCentral
2.
go back to reference Homer CR, Richmond AL, Rebert NA, et al. ATG16L1 and NOD2 interact in an autophagy-dependent antibacterial pathway implicated in Crohn’s disease pathogenesis. Gastroenterology. 2010;139:1630–41. 1641 e1631-1632.CrossRefPubMedPubMedCentral Homer CR, Richmond AL, Rebert NA, et al. ATG16L1 and NOD2 interact in an autophagy-dependent antibacterial pathway implicated in Crohn’s disease pathogenesis. Gastroenterology. 2010;139:1630–41. 1641 e1631-1632.CrossRefPubMedPubMedCentral
4.
go back to reference Lamps LW, Madhusudhan KT, Havens JM, et al. Pathogenic Yersinia DNA is detected in bowel and mesenteric lymph nodes from patients with Crohn’s disease. Am J Surg Pathol. 2003;27:220–7.CrossRefPubMed Lamps LW, Madhusudhan KT, Havens JM, et al. Pathogenic Yersinia DNA is detected in bowel and mesenteric lymph nodes from patients with Crohn’s disease. Am J Surg Pathol. 2003;27:220–7.CrossRefPubMed
5.
go back to reference Liu Y, van Kruiningen HJ, West AB, et al. Immunocytochemical evidence of Listeria, Escherichia coli, and Streptococcus antigens in Crohn’s disease. Gastroenterology. 1995;108:1396–404.CrossRefPubMed Liu Y, van Kruiningen HJ, West AB, et al. Immunocytochemical evidence of Listeria, Escherichia coli, and Streptococcus antigens in Crohn’s disease. Gastroenterology. 1995;108:1396–404.CrossRefPubMed
6.
go back to reference Martin HM, Campbell BJ, Hart CA, et al. Enhanced Escherichia coli adherence and invasion in Crohn’s disease and colon cancer. Gastroenterology. 2004;127:80–93.CrossRefPubMed Martin HM, Campbell BJ, Hart CA, et al. Enhanced Escherichia coli adherence and invasion in Crohn’s disease and colon cancer. Gastroenterology. 2004;127:80–93.CrossRefPubMed
7.
go back to reference Darfeuille-Michaud A, Boudeau J, Bulois P, et al. High prevalence of adherent-invasive Escherichia coli associated with ileal mucosa in Crohn’s disease. Gastroenterology. 2004;127:412–21.CrossRefPubMed Darfeuille-Michaud A, Boudeau J, Bulois P, et al. High prevalence of adherent-invasive Escherichia coli associated with ileal mucosa in Crohn’s disease. Gastroenterology. 2004;127:412–21.CrossRefPubMed
8.
go back to reference Glasser AL, Boudeau J, Barnich N, et al. Adherent invasive Escherichia coli strains from patients with Crohn’s disease survive and replicate within macrophages without inducing host cell death. Infect Immun. 2001;69:5529–37.CrossRefPubMedPubMedCentral Glasser AL, Boudeau J, Barnich N, et al. Adherent invasive Escherichia coli strains from patients with Crohn’s disease survive and replicate within macrophages without inducing host cell death. Infect Immun. 2001;69:5529–37.CrossRefPubMedPubMedCentral
9.
go back to reference Baumgart M, Dogan B, Rishniw M, et al. Culture independent analysis of ileal mucosa reveals a selective increase in invasive Escherichia coli of novel phylogeny relative to depletion of Clostridiales in Crohn’s disease involving the ileum. ISME J. 2007;1:403–18.CrossRefPubMed Baumgart M, Dogan B, Rishniw M, et al. Culture independent analysis of ileal mucosa reveals a selective increase in invasive Escherichia coli of novel phylogeny relative to depletion of Clostridiales in Crohn’s disease involving the ileum. ISME J. 2007;1:403–18.CrossRefPubMed
10.
go back to reference Ryan P, Kelly RG, Lee G, et al. Bacterial DNA within granulomas of patients with Crohn’s disease--detection by laser capture microdissection and PCR. Am J Gastroenterol. 2004;99:1539–43.CrossRefPubMed Ryan P, Kelly RG, Lee G, et al. Bacterial DNA within granulomas of patients with Crohn’s disease--detection by laser capture microdissection and PCR. Am J Gastroenterol. 2004;99:1539–43.CrossRefPubMed
11.
go back to reference Mylonaki M, Rayment NB, Rampton DS, et al. Molecular characterization of rectal mucosa-associated bacterial flora in inflammatory bowel disease. Inflamm Bowel Dis. 2005;11:481–7.CrossRefPubMed Mylonaki M, Rayment NB, Rampton DS, et al. Molecular characterization of rectal mucosa-associated bacterial flora in inflammatory bowel disease. Inflamm Bowel Dis. 2005;11:481–7.CrossRefPubMed
12.
go back to reference Elliott TR, Hudspith BN, Wu G, Cooley M, Parkes GC, Quiñones B, et al. Quantification and characterization of mucosa-associated and intracellular Escherichia coli in inflammatory bowel disease. Inflamm Bowel Dis. 2013;19:2326–38.CrossRefPubMed Elliott TR, Hudspith BN, Wu G, Cooley M, Parkes GC, Quiñones B, et al. Quantification and characterization of mucosa-associated and intracellular Escherichia coli in inflammatory bowel disease. Inflamm Bowel Dis. 2013;19:2326–38.CrossRefPubMed
13.
go back to reference Gratchev A, Kzhyshkowska J, Kothe K, et al. Mphi1 and Mphi2 can be re-polarized by Th2 or Th1 cytokines, respectively, and respond to exogenous danger signals. Immunobiology. 2006;211:473–86.CrossRefPubMed Gratchev A, Kzhyshkowska J, Kothe K, et al. Mphi1 and Mphi2 can be re-polarized by Th2 or Th1 cytokines, respectively, and respond to exogenous danger signals. Immunobiology. 2006;211:473–86.CrossRefPubMed
15.
go back to reference Smith PD, Smythies LE, Shen R, et al. Intestinal macrophages and response to microbial encroachment. Mucosal Immunol. 2011;4:31–42.CrossRefPubMed Smith PD, Smythies LE, Shen R, et al. Intestinal macrophages and response to microbial encroachment. Mucosal Immunol. 2011;4:31–42.CrossRefPubMed
16.
go back to reference Kamada N, Hisamatsu T, Okamoto S, et al. Unique CD14 intestinal macrophages contribute to the pathogenesis of Crohn disease via IL-23/IFN-gamma axis. J Clin Invest. 2008;118:2269–80.PubMedPubMedCentral Kamada N, Hisamatsu T, Okamoto S, et al. Unique CD14 intestinal macrophages contribute to the pathogenesis of Crohn disease via IL-23/IFN-gamma axis. J Clin Invest. 2008;118:2269–80.PubMedPubMedCentral
17.
go back to reference Daperno M, D’Haens G, Van Assche G, et al. Development and validation of a new, simplified endoscopic activity score for Crohn’s disease: the SES-CD. Gastrointest Endosc. 2004;60:505–12.CrossRefPubMed Daperno M, D’Haens G, Van Assche G, et al. Development and validation of a new, simplified endoscopic activity score for Crohn’s disease: the SES-CD. Gastrointest Endosc. 2004;60:505–12.CrossRefPubMed
18.
19.
go back to reference Falini B, Flenghi L, Pileri S, et al. PG-M1: A new monoclonal antibody directed against a fixative-resistant epitope on the macrophage-restricted form of the CD68 molecule. Am J Pathol. 1993;142:1359–72.PubMedPubMedCentral Falini B, Flenghi L, Pileri S, et al. PG-M1: A new monoclonal antibody directed against a fixative-resistant epitope on the macrophage-restricted form of the CD68 molecule. Am J Pathol. 1993;142:1359–72.PubMedPubMedCentral
20.
go back to reference Berry RE et al. Urothelial cultures support intracellular bacterial community formation by uropathogenic Escherichia coli. Infect Immun. 2009;77:2762–72.CrossRefPubMedPubMedCentral Berry RE et al. Urothelial cultures support intracellular bacterial community formation by uropathogenic Escherichia coli. Infect Immun. 2009;77:2762–72.CrossRefPubMedPubMedCentral
21.
go back to reference Trogan E, Fisher EA. Laser capture microdissection for analysis of macrophage gene expression from atherosclerotic lesions. Methods Mol Biol. 2005;293:221–31. Review.PubMed Trogan E, Fisher EA. Laser capture microdissection for analysis of macrophage gene expression from atherosclerotic lesions. Methods Mol Biol. 2005;293:221–31. Review.PubMed
22.
go back to reference Fleige S, Pfaffl MW. RNA integrity and the effect on the real-time qRT-PCR performance. Mol Aspects Med. 2006;27(2-3):126–39.CrossRefPubMed Fleige S, Pfaffl MW. RNA integrity and the effect on the real-time qRT-PCR performance. Mol Aspects Med. 2006;27(2-3):126–39.CrossRefPubMed
23.
go back to reference Bustin SA, Benes V, Garson JA, et al. The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem. 2009;55(4):611–22.CrossRefPubMed Bustin SA, Benes V, Garson JA, et al. The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem. 2009;55(4):611–22.CrossRefPubMed
24.
go back to reference Bustin SA. Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems. J Mol Endocrinol. 2002;29(1):23–39.CrossRefPubMed Bustin SA. Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems. J Mol Endocrinol. 2002;29(1):23–39.CrossRefPubMed
25.
go back to reference Rayment N, Mylonaki M, Hudspith B, et al. Co-localisation of E. coli with macrophages in lamina propria in patients with active inflammatory bowel disease (IBD). Gut. 2003;52:A13. Rayment N, Mylonaki M, Hudspith B, et al. Co-localisation of E. coli with macrophages in lamina propria in patients with active inflammatory bowel disease (IBD). Gut. 2003;52:A13.
26.
go back to reference Woo PC, Lau SK, Teng JL, et al. Then and now: use of 16S rDNA gene sequencing for bacterial identification and discovery of novel bacteria in clinical microbiology laboratories. Clin Microbiol Infect. 2008;14(10):908–34.CrossRefPubMed Woo PC, Lau SK, Teng JL, et al. Then and now: use of 16S rDNA gene sequencing for bacterial identification and discovery of novel bacteria in clinical microbiology laboratories. Clin Microbiol Infect. 2008;14(10):908–34.CrossRefPubMed
27.
go back to reference Elliott TR, Hudspith BN, Rayment NB et al. Defective macrophage handling of Escherichia coli in Crohn’s disease. J Gastroenterol Hepatol. 2015 Mar 23. doi: 10.1111/jgh.12955. [Epub ahead of print] Elliott TR, Hudspith BN, Rayment NB et al. Defective macrophage handling of Escherichia coli in Crohn’s disease. J Gastroenterol Hepatol. 2015 Mar 23. doi: 10.1111/jgh.12955. [Epub ahead of print]
28.
go back to reference Prescott NJ, Fisher SA, Franke A, et al. A nonsynonymous SNP in ATG16L1 predisposes to ileal Crohn’s disease and is independent of CARD15 and IBD5. Gastroenterology. 2007;132:1665–71.CrossRefPubMed Prescott NJ, Fisher SA, Franke A, et al. A nonsynonymous SNP in ATG16L1 predisposes to ileal Crohn’s disease and is independent of CARD15 and IBD5. Gastroenterology. 2007;132:1665–71.CrossRefPubMed
29.
go back to reference Sasaki M, Sitaraman SV, Babbin BA, et al. Invasive Escherichia coli are a feature of Crohn’s disease. Lab Invest. 2007;87:1042–54.CrossRefPubMed Sasaki M, Sitaraman SV, Babbin BA, et al. Invasive Escherichia coli are a feature of Crohn’s disease. Lab Invest. 2007;87:1042–54.CrossRefPubMed
30.
go back to reference Tricarico C et al. Quantitative real-time reverse transcription polymerase chain reaction: normalization to rRNA or single housekeeping genes is inappropriate for human tissue biopsies. Anal Biochem. 2002;309(2):293–300.CrossRefPubMed Tricarico C et al. Quantitative real-time reverse transcription polymerase chain reaction: normalization to rRNA or single housekeeping genes is inappropriate for human tissue biopsies. Anal Biochem. 2002;309(2):293–300.CrossRefPubMed
31.
go back to reference Dheda K et al. Validation of housekeeping genes for normalizing RNA expression in real-time PCR. Biotechniques. 2004;37(1):112. -4, 116, 118-9.PubMed Dheda K et al. Validation of housekeeping genes for normalizing RNA expression in real-time PCR. Biotechniques. 2004;37(1):112. -4, 116, 118-9.PubMed
32.
go back to reference Bustin SA, Benes V, Nolan T, Pfaffl MW. Quantitative real-time RT-PCR – a perspective. J Mol Endocrinol. 2005;34:597–601.CrossRefPubMed Bustin SA, Benes V, Nolan T, Pfaffl MW. Quantitative real-time RT-PCR – a perspective. J Mol Endocrinol. 2005;34:597–601.CrossRefPubMed
33.
go back to reference Nolan T, Hands RE, Bustin SA. Quantification of mRNA using real-time RT-PCR. Nat Protoc. 2006;1:1559–82.CrossRefPubMed Nolan T, Hands RE, Bustin SA. Quantification of mRNA using real-time RT-PCR. Nat Protoc. 2006;1:1559–82.CrossRefPubMed
34.
go back to reference Smythies LE, Sellers M, Clements RH, et al. Human intestinal macrophages display profound inflammatory anergy despite avid phagocytic and bacteriocidal activity. J Clin Invest. 2005;115:66–75.CrossRefPubMedPubMedCentral Smythies LE, Sellers M, Clements RH, et al. Human intestinal macrophages display profound inflammatory anergy despite avid phagocytic and bacteriocidal activity. J Clin Invest. 2005;115:66–75.CrossRefPubMedPubMedCentral
35.
go back to reference Mege JL, Meghari S, Honstettre A, et al. The two faces of interleukin 10 in human infectious diseases. Lancet Infect Dis. 2006;6:557–69.CrossRefPubMed Mege JL, Meghari S, Honstettre A, et al. The two faces of interleukin 10 in human infectious diseases. Lancet Infect Dis. 2006;6:557–69.CrossRefPubMed
36.
go back to reference Park HJ, Lee SJ, Kim SH, et al. IL-10 inhibits the starvation induced autophagy in macrophages via class I phosphatidylinositol 3-kinase (PI3K) pathway. Mol Immunol. 2011;48:720–7.CrossRefPubMed Park HJ, Lee SJ, Kim SH, et al. IL-10 inhibits the starvation induced autophagy in macrophages via class I phosphatidylinositol 3-kinase (PI3K) pathway. Mol Immunol. 2011;48:720–7.CrossRefPubMed
37.
go back to reference Moos V, Schmidt C, Geelhaar A, et al. Impaired immune functions of monocytes and macrophages in Whipple’s disease. Gastroenterology. 2010;138:210–20.CrossRefPubMed Moos V, Schmidt C, Geelhaar A, et al. Impaired immune functions of monocytes and macrophages in Whipple’s disease. Gastroenterology. 2010;138:210–20.CrossRefPubMed
38.
go back to reference Andersen V, Nimmo E, Krarup HB, et al. Cyclooxygenase-2 (COX-2) polymorphisms and risk of inflammatory bowel disease in a Scottish and Danish case-control study. Inflamm Bowel Dis. 2011;17:937–46.CrossRefPubMed Andersen V, Nimmo E, Krarup HB, et al. Cyclooxygenase-2 (COX-2) polymorphisms and risk of inflammatory bowel disease in a Scottish and Danish case-control study. Inflamm Bowel Dis. 2011;17:937–46.CrossRefPubMed
39.
go back to reference Subramanian S, Roberts CL, Hart CA, et al. Replication of colonic Crohn’s disease mucosal escherichia coli isolates within macrophages and their susceptibility to antibiotics. Antimicrob Agents Chemother. 2008;52:427–34.CrossRefPubMed Subramanian S, Roberts CL, Hart CA, et al. Replication of colonic Crohn’s disease mucosal escherichia coli isolates within macrophages and their susceptibility to antibiotics. Antimicrob Agents Chemother. 2008;52:427–34.CrossRefPubMed
Metadata
Title
Lamina propria macrophage phenotypes in relation to Escherichia coli in Crohn’s disease
Authors
Timothy R. Elliott
Neil B. Rayment
Barry N. Hudspith
Rebecca E. Hands
Kirstin Taylor
Gareth C. Parkes
Natalie J. Prescott
Liljana Petrovska
John Hermon-Taylor
Jonathan Brostoff
Alex Boussioutas
Christopher G. Mathew
Stephen A. Bustin
Jeremy D. Sanderson
Publication date
01-12-2015
Publisher
BioMed Central
Published in
BMC Gastroenterology / Issue 1/2015
Electronic ISSN: 1471-230X
DOI
https://doi.org/10.1186/s12876-015-0305-3

Other articles of this Issue 1/2015

BMC Gastroenterology 1/2015 Go to the issue