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In vitro modeling of host-parasite interactions: the 'subgingival' biofilm challenge of primary human epithelial cells.

Guggenheim B, Gmür R, Galicia JC, Stathopoulou PG, Benakanakere MR, Meier A, Thurnheer T, Kinane DF - BMC Microbiol. (2009)

Bottom Line: The new model takes into account that the microbial challenge derives from a biofilm community and not from planktonically cultured bacterial strains.It will facilitate easily the introduction of additional host cells such as neutrophils for future biofilm:host cell challenge studies.Our methodology may generate particular interest, as it should be widely applicable to other biofilm-related chronic inflammatory diseases.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute for Oral Biology, Section for Oral Microbiology and General Immunology, University of Zürich, Plattenstrasse 11, CH-8032 Zürich, Switzerland. bernie@zzmk.uzh.ch

ABSTRACT

Background: Microbial biofilms are known to cause an increasing number of chronic inflammatory and infectious conditions. A classical example is chronic periodontal disease, a condition initiated by the subgingival dental plaque biofilm on gingival epithelial tissues. We describe here a new model that permits the examination of interactions between the bacterial biofilm and host cells in general. We use primary human gingival epithelial cells (HGEC) and an in vitro grown biofilm, comprising nine frequently studied and representative subgingival plaque bacteria.

Results: We describe the growth of a mature 'subgingival' in vitro biofilm, its composition during development, its ability to adapt to aerobic conditions and how we expose in vitro a HGEC monolayer to this biofilm. Challenging the host derived HGEC with the biofilm invoked apoptosis in the epithelial cells, triggered release of pro-inflammatory cytokines and in parallel induced rapid degradation of the cytokines by biofilm-generated enzymes.

Conclusion: We developed an experimental in vitro model to study processes taking place in the gingival crevice during the initiation of inflammation. The new model takes into account that the microbial challenge derives from a biofilm community and not from planktonically cultured bacterial strains. It will facilitate easily the introduction of additional host cells such as neutrophils for future biofilm:host cell challenge studies. Our methodology may generate particular interest, as it should be widely applicable to other biofilm-related chronic inflammatory diseases.

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Cytokine production in biofilm-challenged HGEC cultures. (A) Supernatant IL-1 and IL-6 levels, and (B) IL-8 levels in challenged HGEC cultures and controls. Unchallenged cells were used as a negative control. Values represent the mean ± SD of triplicate ELISA assays performed after 4 and 24 h of co-culture.
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Figure 4: Cytokine production in biofilm-challenged HGEC cultures. (A) Supernatant IL-1 and IL-6 levels, and (B) IL-8 levels in challenged HGEC cultures and controls. Unchallenged cells were used as a negative control. Values represent the mean ± SD of triplicate ELISA assays performed after 4 and 24 h of co-culture.

Mentions: HGEC were challenged with 'subgingival' biofilms for 4 and 24 h. At 4 h, the primary (IL-1β) and secondary cytokine (IL-6) and chemokine (IL-8) responses were significantly elevated (Fig. 4A &4B). At 24 h, however, the level of all cytokines had subsided significantly. To test our hypothesis that this decline of cytokine levels is due to biofilm-mediated degradation, supernatant from HGEC, pre-challenged with heat-killed planktonic P. gingivalis ATCC 33277 for establishing cytokine production, was incubated either with biofilms on HA discs, filtered supernatant from 64.5 h biofilm cultures, or with KSFM medium in which biofilms had been incubated for 24 h (filtered or unfiltered). Assays performed after 1 min, 30 min, 1, 2, and 4 h of exposure to the biofilm showed that IL-1β, IL-6 and IL-8 degradation began immediately and progressively increased to reach approximately 25% after 4 h (Table 2). When filtered biofilm culture supernatant or KSFM were tested, IL-6 and IL-8 degradation was not observed, while the rate of IL-1β degradation was significantly reduced up to 2 h but eventually reached levels similar to that induced by the biofilm at 4 h. This suggests that the presence of biofilm bacteria is necessary for IL-6 and IL-8 degradation, while IL-1β is more susceptible to degradation by a soluble component of the biofilm culture.


In vitro modeling of host-parasite interactions: the 'subgingival' biofilm challenge of primary human epithelial cells.

Guggenheim B, Gmür R, Galicia JC, Stathopoulou PG, Benakanakere MR, Meier A, Thurnheer T, Kinane DF - BMC Microbiol. (2009)

Cytokine production in biofilm-challenged HGEC cultures. (A) Supernatant IL-1 and IL-6 levels, and (B) IL-8 levels in challenged HGEC cultures and controls. Unchallenged cells were used as a negative control. Values represent the mean ± SD of triplicate ELISA assays performed after 4 and 24 h of co-culture.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2818713&req=5

Figure 4: Cytokine production in biofilm-challenged HGEC cultures. (A) Supernatant IL-1 and IL-6 levels, and (B) IL-8 levels in challenged HGEC cultures and controls. Unchallenged cells were used as a negative control. Values represent the mean ± SD of triplicate ELISA assays performed after 4 and 24 h of co-culture.
Mentions: HGEC were challenged with 'subgingival' biofilms for 4 and 24 h. At 4 h, the primary (IL-1β) and secondary cytokine (IL-6) and chemokine (IL-8) responses were significantly elevated (Fig. 4A &4B). At 24 h, however, the level of all cytokines had subsided significantly. To test our hypothesis that this decline of cytokine levels is due to biofilm-mediated degradation, supernatant from HGEC, pre-challenged with heat-killed planktonic P. gingivalis ATCC 33277 for establishing cytokine production, was incubated either with biofilms on HA discs, filtered supernatant from 64.5 h biofilm cultures, or with KSFM medium in which biofilms had been incubated for 24 h (filtered or unfiltered). Assays performed after 1 min, 30 min, 1, 2, and 4 h of exposure to the biofilm showed that IL-1β, IL-6 and IL-8 degradation began immediately and progressively increased to reach approximately 25% after 4 h (Table 2). When filtered biofilm culture supernatant or KSFM were tested, IL-6 and IL-8 degradation was not observed, while the rate of IL-1β degradation was significantly reduced up to 2 h but eventually reached levels similar to that induced by the biofilm at 4 h. This suggests that the presence of biofilm bacteria is necessary for IL-6 and IL-8 degradation, while IL-1β is more susceptible to degradation by a soluble component of the biofilm culture.

Bottom Line: The new model takes into account that the microbial challenge derives from a biofilm community and not from planktonically cultured bacterial strains.It will facilitate easily the introduction of additional host cells such as neutrophils for future biofilm:host cell challenge studies.Our methodology may generate particular interest, as it should be widely applicable to other biofilm-related chronic inflammatory diseases.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute for Oral Biology, Section for Oral Microbiology and General Immunology, University of Zürich, Plattenstrasse 11, CH-8032 Zürich, Switzerland. bernie@zzmk.uzh.ch

ABSTRACT

Background: Microbial biofilms are known to cause an increasing number of chronic inflammatory and infectious conditions. A classical example is chronic periodontal disease, a condition initiated by the subgingival dental plaque biofilm on gingival epithelial tissues. We describe here a new model that permits the examination of interactions between the bacterial biofilm and host cells in general. We use primary human gingival epithelial cells (HGEC) and an in vitro grown biofilm, comprising nine frequently studied and representative subgingival plaque bacteria.

Results: We describe the growth of a mature 'subgingival' in vitro biofilm, its composition during development, its ability to adapt to aerobic conditions and how we expose in vitro a HGEC monolayer to this biofilm. Challenging the host derived HGEC with the biofilm invoked apoptosis in the epithelial cells, triggered release of pro-inflammatory cytokines and in parallel induced rapid degradation of the cytokines by biofilm-generated enzymes.

Conclusion: We developed an experimental in vitro model to study processes taking place in the gingival crevice during the initiation of inflammation. The new model takes into account that the microbial challenge derives from a biofilm community and not from planktonically cultured bacterial strains. It will facilitate easily the introduction of additional host cells such as neutrophils for future biofilm:host cell challenge studies. Our methodology may generate particular interest, as it should be widely applicable to other biofilm-related chronic inflammatory diseases.

Show MeSH
Related in: MedlinePlus