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PP-008 Survival of Escherichia coli in monocytic cells in Crohn's disease: evidence of reduced killing capacity
  1. B N Hudspith,
  2. K Blackie,
  3. N B Rayment,
  4. T Elliott,
  5. J Brostoff,
  6. J Hermon-Taylor,
  7. J D Sanderson
  1. Nutrional Sciences Division, King's College London, London, UK

Abstract

Introduction Whist the exact cause of Crohn's disease (CD) remains unclear, an innate defect in the handling of intracellular bacteria appears to be involved. Evidence for this comes from 1. genome wide association studies implicating a role for genes involved in bacteria-immune cell interactions including NOD-2 and the autophagy related genes, ATG16L1 and IRGM, 2. an observed defect in phagocyte recruitment and 3. the consistent demonstration of viable E coli within lamina propria macrophages in CD. It is not known if this latter observation is due to increased uptake of E coli or a failure to achieve effective phagolysosomal killing of bacteria once within the cell.

Methods To examine phagocytosis and oxidative burst activity of phagocytic cells in peripheral blood of 16 healthy individuals and 16 patients with CD. Phagocytic activity was assessed by the number of cells taking up fluorescently tagged E coli (Phagotest) and the potential role of defective reactive oxygen radical (ROR) production involved in bacterial killing (Bursttest). This is measured by the conversion of non-fluorescent dihydrorhodamine 123 to the fluorescent rhodamine 123 by ROR's. The leucocytes were challenged with three strains of E coli, a heat killed sample, a laboratory strain (K12) and a strain isolated from a patient with CD. Results were obtained using florescent markers measured by flow cytometery.

Results Using the phagotest, to measure the uptake of bacteria no difference was seen between CD and controls and did not vary according to strain of E coli used. However, although the number of monocytes expressing ROR's was similar in both groups, their was a significantly level of ROR's produced in cells from CD compared to controls when viable bacteria were used (E coli K12: control 98.5±7.4 vs CD 68.1±13.2, p=0.04, and the CD isolate: control cells 101.1±7.4 vs CD 59.2±12.2, p=0.010). No difference was seen in the granulocytes between CD and controls for either phagocytosis or ROR's.

Conclusion The ability of monocytes from CD patients to engulf E coli appears to be normal. However, there is good evidence of an impaired capacity to kill these bacteria as measured by reduced levels of ROR's. These results points to either an inability to complete the process of phagolysosome formation or defects in the enzyme pathways involved in formation of reactive oxygen products that are induced by phagocytosis and support the concept of impaired innate immunity in CD.

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