In vitro cell culture methods for investigating Campylobacter invasion mechanisms

Friis, Lorna M.; Pin, Carmen; Pearson, Bruce M.; Wells, Jerry M.

doi:10.1016/j.mimet.2004.12.003

Cited by 73 publications

(73 citation statements)

References 90 publications

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“…The more adhesive and invasive strains, appeared to be the ones which also induced the highest levels of IL-8 in INT-407 cells, as previously shown by Hickey et al [28,29]. Next to strain-dependency, the level of adhesion and invasion was also found to be dose dependent as previously shown [13][14][15]. Our data illustrate the importance of measuring the invasion and adhesion at different doses when comparing strains.…”

Section: Discussionsupporting

confidence: 87%

“…To avoid the misleading effect of a few culturable bacteria, in our study the infectivity of culturable C. jejuni suspensions and the infectivity of the same culturable C. jejuni suspensions supplemented with a high number of non-culturable C. jejuni, formed at 4 xC, were measured. The comparison was made for different doses of culturable C. jejuni in the absence and presence of non-culturable C. jejuni, as the infectivity might be dose-dependent [13][14][15].…”

Section: Introductionmentioning

confidence: 99%

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Lack of response of INT-407 cells to the presence of non-culturableCampylobacter jejuni

2007

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SUMMARYMany contradictory articles on the infectivity of non-culturable Campylobacter jejuni can be found. We studied the effect of non-culturable C. jejuni in an in vitro assay. To prevent the potential effect of a few culturable bacteria in the non-culturable suspension, INT-407 cells, which mimic the outer cell layer in the small intestines, were exposed to culturable C. jejuni suspensions with or without non-culturable C. jejuni. The number of bacteria adhering to and/or invading INT-407 cells and the IL-8 secretion were measured. No differences were found between bacterial suspensions with or without non-culturable C. jejuni added. These findings show that non-culturable C. jejuni do not adhere to or invade INT-407 cells and do not induce an immune response. As previous studies showed a correlation between the used in vitro assays and the effect in vivo, our study strongly suggests that culturability is a good indicator of the risk for C. jejuni infection.

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Section: Discussionsupporting

confidence: 87%

Section: Introductionmentioning

confidence: 99%

Lack of response of INT-407 cells to the presence of non-culturableCampylobacter jejuni

2007

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“…It is likely that C. jejuni's invasive nature and disruption of epithelial barrier integrity combined with the stimulation of host proinflammatory cytokines and defensins play a composite role in gastroenteritis (11,15,37,64). However, our understanding of the sequence of events leading to postinfectious sequelae (for example, reactive arthritis and irritable bowel syndrome), now more frequently being associated with the bacterium (38,51), is minimal.…”

Section: Discussionmentioning

confidence: 99%

“…Cells were infected with 1 ϫ 10 9 exponentially growing C. jejuni cells resuspended in 500 l infection medium to establish a multiplicity of infection (MOI) of 1,000. This MOI was determined to be optimal for maximum Campylobacter invasion and for making strain-to-strain comparisons (15). Phorbol 12-myristate 13-acetate (PMA) (500 ng ml Ϫ1 ), Escherichia coli 0111:B4 and 055:B5 LPS (100 ng ml Ϫ1 ), and IL-1␤ (10 ng ml Ϫ1 ) were used as controls.…”

Section: Methodsmentioning

confidence: 99%

Campylobacter jejuni Drives MyD88-Independent Interleukin-6 Secretion via Toll-Like Receptor 2

Friis¹,

Keelan

Taylor³

2009

Infect Immun

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Gastrointestinal disease caused by Campylobacter jejuni is characterized by localized inflammation and the destruction of the epithelial cell barrier that forms host innate protection against pathogens. This can lead to an imbalance in fluid transport across the gastrointestinal tract, resulting in severe diarrhea. The mechanisms of host cell receptor recognition of C. jejuni and downstream immune signaling pathways leading to this inflammatory disease, however, remain unclear. The aim of this study was to analyze the mechanisms involved in C. jejuni induction of the acute-phase inflammatory response regulator interleukin-6 (IL-6). Polarized intestinal epithelial Caco-2 monolayers responded to infections with Salmonella enterica serovar Typhimurium and eight isolates of C. jejuni by an increase in levels of expression and secretion of IL-6. No such IL-6 response, however, was produced upon infection with the human commensal organism Lactobacillus rhamnosus GG. The IL-6 signaling pathway was further characterized using short interfering RNA complexes to block gene expression. The inhibition of myeloid differentiation primary response protein 88 (MyD88) expression in this manner did not affect C. jejuni-induced IL-6 secretion, suggesting a MyD88-independent route to IL-6 signal transduction in C. jejuni-infected human epithelial cells. However, a significant reduction in levels of IL-6 was evident in the absence of Toll-like receptor 2 (TLR-2) expression, implying a requirement for TLR-2 in C. jejuni recognition. Caco-2 cells were also treated with heat-inactivated and purified membrane components of C. jejuni to isolate the factor responsible for triggering IL-6 signaling. The results demonstrate that C. jejuni surface polysaccharides induce IL-6 secretion from intestinal epithelial cells via TLR-2 in a MyD88-independent manner.

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“…In the absence of a suitable animal model that mimics human disease (Newell, 2001), invasion has been studied using in vitro cell culture. There have been many reports of in vitro cell culture models of invasion for campylobacters and these have recently been reviewed (Friis et al, 2005). As in other enteropathogens, virulence varies between strains of C. jejuni.…”

Section: Introductionmentioning

confidence: 99%

Transposon mutagenesis in a hyper-invasive clinical isolate of Campylobacter jejuni reveals a number of genes with potential roles in invasion

et al. 2010

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Transposon mutagenesis has been applied to a hyper-invasive clinical isolate of Campylobacter jejuni, 01/51. A random transposon mutant library was screened in an in vitro assay of invasion and 26 mutants with a significant reduction in invasion were identified. Given that the invasion potential of C. jejuni is relatively poor compared to other enteric pathogens, the use of a hyper-invasive strain was advantageous as it greatly facilitated the identification of mutants with reduced invasion. The location of the transposon insertion in 23 of these mutants has been determined; all but three of the insertions are in genes also present in the genome-sequenced strain NCTC 11168. Eight of the mutants contain transposon insertions in one region of the genome (∼14 kb), which when compared with the genome of NCTC 11168 overlaps with one of the previously reported plasticity regions and is likely to be involved in genomic variation between strains. Further characterization of one of the mutants within this region has identified a gene that might be involved in adhesion to host cells.

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In vitro cell culture methods for investigating Campylobacter invasion mechanisms

Cited by 73 publications

References 90 publications

Lack of response of INT-407 cells to the presence of non-culturableCampylobacter jejuni

Lack of response of INT-407 cells to the presence of non-culturableCampylobacter jejuni

Campylobacter jejuni Drives MyD88-Independent Interleukin-6 Secretion via Toll-Like Receptor 2

Transposon mutagenesis in a hyper-invasive clinical isolate of Campylobacter jejuni reveals a number of genes with potential roles in invasion

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