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

Elliott TR, Rayment NB, Hudspith BN, Hands RE, Taylor K, Parkes GC, Prescott NJ, Petrovska L, Hermon-Taylor J, Brostoff J, Boussioutas A, Mathew CG, Bustin SA, Sanderson JD - BMC Gastroenterol (2015)

Bottom Line: In inflamed tissue, E. coli-unladen macrophages expressed high TNFα, IL-23 & iNOS and lower IL-10 & CD163.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.

View Article: PubMed Central - PubMed

Affiliation: Diabetes and Nutritional Sciences Division, King's College London, Franklin Wilkins Building, 150 Stamford Street, London, SE1 9NH, UK. elliott_timothy@hotmail.com.

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.

No MeSH data available.


Related in: MedlinePlus

CD68 staining macrophages (red) can be seen to contain E. coli ( blue ) in the biopsy from a patient with CD ((a) low power (x40), (b) high power (x63), but not in biopsies from a healthy control (x20) (c). Labelled lamina propria macrophages (d) before and (e) after laser capture microdissection
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Fig1: CD68 staining macrophages (red) can be seen to contain E. coli ( blue ) in the biopsy from a patient with CD ((a) low power (x40), (b) high power (x63), but not in biopsies from a healthy control (x20) (c). Labelled lamina propria macrophages (d) before and (e) after laser capture microdissection

Mentions: LP macrophages with or without E. coli co-localisation were identified in mucosal biopsies using previously validated immunolabelled CD68+ [19] and E. coli antibodies [20]. A rapid indirect immunostaining protocol was employed for macrophage-specific CD68 (PG M1) to minimize risk of mRNA degradation [21]. CD68 positive cells were detected using a Vectastain ABC-AP kit and a Vector Red chromogenic substrate. Detection of intracellular E. coli was achieved by co-staining CD68+ macrophages with an anti-E. coli polyclonal antibody and labeled with Vector Blue chromogenic substrate. Staining was visualized using a Zeiss axioplan MOT 400 M microscope. CD68+ cells co-localising with anti-E. coli antibody were termed E. coli-laden macrophages (Fig. 1a, b). CD68+ cells without anti-E. coli antibody co-localisation were termed E. coli-unladen macrophages (Fig. 1c).Fig. 1


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

Elliott TR, Rayment NB, Hudspith BN, Hands RE, Taylor K, Parkes GC, Prescott NJ, Petrovska L, Hermon-Taylor J, Brostoff J, Boussioutas A, Mathew CG, Bustin SA, Sanderson JD - BMC Gastroenterol (2015)

CD68 staining macrophages (red) can be seen to contain E. coli ( blue ) in the biopsy from a patient with CD ((a) low power (x40), (b) high power (x63), but not in biopsies from a healthy control (x20) (c). Labelled lamina propria macrophages (d) before and (e) after laser capture microdissection
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4490755&req=5

Fig1: CD68 staining macrophages (red) can be seen to contain E. coli ( blue ) in the biopsy from a patient with CD ((a) low power (x40), (b) high power (x63), but not in biopsies from a healthy control (x20) (c). Labelled lamina propria macrophages (d) before and (e) after laser capture microdissection
Mentions: LP macrophages with or without E. coli co-localisation were identified in mucosal biopsies using previously validated immunolabelled CD68+ [19] and E. coli antibodies [20]. A rapid indirect immunostaining protocol was employed for macrophage-specific CD68 (PG M1) to minimize risk of mRNA degradation [21]. CD68 positive cells were detected using a Vectastain ABC-AP kit and a Vector Red chromogenic substrate. Detection of intracellular E. coli was achieved by co-staining CD68+ macrophages with an anti-E. coli polyclonal antibody and labeled with Vector Blue chromogenic substrate. Staining was visualized using a Zeiss axioplan MOT 400 M microscope. CD68+ cells co-localising with anti-E. coli antibody were termed E. coli-laden macrophages (Fig. 1a, b). CD68+ cells without anti-E. coli antibody co-localisation were termed E. coli-unladen macrophages (Fig. 1c).Fig. 1

Bottom Line: In inflamed tissue, E. coli-unladen macrophages expressed high TNFα, IL-23 & iNOS and lower IL-10 & CD163.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.

View Article: PubMed Central - PubMed

Affiliation: Diabetes and Nutritional Sciences Division, King's College London, Franklin Wilkins Building, 150 Stamford Street, London, SE1 9NH, UK. elliott_timothy@hotmail.com.

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.

No MeSH data available.


Related in: MedlinePlus