Abstract
Mucosa-associated Escherichia coli are increased in Crohn’s disease (CD) and colorectal cancer (CRC). CD isolates replicate within macrophages but the specificity of this effect for CD and its mechanism are unclear. Gentamicin exclusion assay was used to assess E. coli replication within J774.A1 murine macrophages. E. coli growth was assessed following acid, low-nutrient, nitrosative, oxidative and superoxide stress, mimicking the phagolysosome. Twelve of 16 CD E. coli isolates replicated >2-fold within J774.A1 macrophages; likewise for isolates from 6/7 urinary tract infection (UTI), 8/9 from healthy subjects, compared with 2/6 ulcerative colitis, 2/7 colorectal cancer and 0/3 laboratory strains. CD mucosal E. coli were tolerant of acidic, low-nutrient, nitrosative and oxidative stress. Replication within macrophages correlated strongly with tolerance to superoxide stress (rho = 0.44, p = 0.0009). Exemplar CD E. coli HM605 and LF82 were unable to survive within Nfκb1-/- murine bone marrow-derived macrophages. In keeping with this, pre-incubation of macrophages with hydrocortisone (0.6 µM for 24 h) caused 70.49 ± 12.11% inhibition of intra-macrophage replication. Thus, CD mucosal E. coli commonly replicate inside macrophages, but so do some UTI and healthy subject strains. Replication correlates with resistance to superoxide and is highly dependent on macrophage NF-κB signalling. This may therefore be a good therapeutic target.
Highlights
Mucosa-associated E. coli have been found in increased numbers on the ileal and colonic mucosae, including the inner adherent mucus layer, of patients with Crohn’s disease (CD) [1,2,3,4,5,6] or colorectal cancer (CRC) [3,4,5,6,7], and to a lesser extent, patients with ulcerative colitis (UC) [8,9]
A high proportion of CD mucosal E. coli strains adhere to, and invade, intestinal epithelial cell-lines Caco-2 and Int-407, and induce release of pro-inflammatory cytokines [2,3,4,5,6,10]; it has been noted that the level of invasion into epithelial cell lines is strongly dependent on the cell line chosen for experimental study [3]
In vitro studies using the paradigm CD E. coli strains from the ileum (LF82) [1] and from the colon (HM605) [3] showed that they possess the ability to replicate within murine and human
Summary
Mucosa-associated E. coli have been found in increased numbers on the ileal and colonic mucosae, including the inner adherent mucus layer, of patients with Crohn’s disease (CD) [1,2,3,4,5,6] or colorectal cancer (CRC) [3,4,5,6,7], and to a lesser extent, patients with ulcerative colitis (UC) [8,9]. CD isolates (and CRC isolates) more commonly possess key virulence genes that drive common phenotypic/pathogenic actions, including epithelial cell adherence and invasion, entry via microfold (M) cells of the follicle-associated epithelium, enhanced angiogenic potential and genotoxicity [34,35,36,37,38]. They are possibly best defined as pathobionts—organisms with the potential for causing disease that likely live as symbionts under circumstances of normal gut health. We have assessed their tolerance of stress conditions characteristic of the macrophage phagolysosomes and the role of the classical NF-κB pathway activation in this process
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