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Interactions of anaerobic bacteria with dental stem cells: an in vitro study.

Biedermann A, Kriebel K, Kreikemeyer B, Lang H - PLoS ONE (2014)

Bottom Line: Similar results were observed for host cell internalization (F. nucleatum: 0.002% internalization into hDFSC; 0.09% internalization into Ca9-22).The IL-10 response of the differentiated cells was found to be low in relation to their pro-inflammatory IL-8 response.The results indicate that dental stem cells are less prone to interactions with pathogenic bacteria than differentiated cells in an anaerobic environment.

View Article: PubMed Central - PubMed

Affiliation: Department of Operative Dentistry and Periodontology, University of Rostock, Rostock, Germany.

ABSTRACT

Background: In patients with periodontitis, it is highly likely that local (progenitor) cells encounter pathogenic bacteria. The purpose of this in vitro study was to elucidate how human dental follicle stem cells (hDFSC) react towards a direct challenge with anaerobic periodontal pathogens under their natural oxygen-free atmosphere. HDFSC were compared to human bone marrow mesenchymal stem cells (hBMSC) and differentiated primary human gingival fibroblasts (hGiF), as well as permanent gingival carcinoma cells (Ca9-22).

Methodology/principal findings: The different cell types were investigated in a co-culture system with Porphyromonas gingivalis (P. gingivalis) and Fusobacterium nucleatum (F. nucleatum). The viability of the cells and pathogens under anaerobic conditions, as well as interactions in terms of adherence and internalization, were examined. Additionally, the release of pro-inflammatory interleukin-8 (IL-8) and anti-inflammatory interleukin-10 (IL-10) was quantified via enzyme-linked immunosorbent assay. The bacteria adhered less efficiently to hDFSC compared to Ca9-22 (P. gingivalis: 0.18% adherence to hDFSC; 3.1% adherence to Ca9-22). Similar results were observed for host cell internalization (F. nucleatum: 0.002% internalization into hDFSC; 0.09% internalization into Ca9-22). Statistically significantly less IL-8 was secreted from hDFSC after stimulation with F. nucleatum and P. gingivalis in comparison with hGiF (F. nucleatum: 2080.0 pg/ml--hGiF; 19.7 pg/ml--hDFSC). The IL-10 response of the differentiated cells was found to be low in relation to their pro-inflammatory IL-8 response.

Conclusions/significance: The results indicate that dental stem cells are less prone to interactions with pathogenic bacteria than differentiated cells in an anaerobic environment. Moreover, during bacterial challenge, the stem cell immune response seems to be more towards an anti-inflammatory reaction. For a potential future therapeutic use of hDFSC, these findings support the idea of a save application.

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Bacterial growth in cell culture medium.Growth rates of bacteria in DMEM cell culture medium under anaerobic conditions (10% CO2, 10% H2, 80% N2), 37°C. Bacteria were grown in PYG medium supplemented with 5 µg/ml hemin and 1% vitamin K to the stationary phase. Subsequently, they were centrifuged, washed in PBS, and each bacterial suspension was diluted in DMEM 1∶10. The OD600 was measured at 600 nm from timepoint 0 every hour to timepoint 12 h with a final measurement after 24 h.
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pone-0110616-g002: Bacterial growth in cell culture medium.Growth rates of bacteria in DMEM cell culture medium under anaerobic conditions (10% CO2, 10% H2, 80% N2), 37°C. Bacteria were grown in PYG medium supplemented with 5 µg/ml hemin and 1% vitamin K to the stationary phase. Subsequently, they were centrifuged, washed in PBS, and each bacterial suspension was diluted in DMEM 1∶10. The OD600 was measured at 600 nm from timepoint 0 every hour to timepoint 12 h with a final measurement after 24 h.

Mentions: The bacterial growth was determined in cell culture medium under anaerobic conditions. All four tested species showed normal bacterial growth (Fig. 2). From these results, we concluded that co-culture with anaerobic bacteria and hDFSC (and hBMSC, hGiF, or Ca9-22) was possible. Live/dead staining and fluorescence microscopy confirmed the survival of bacteria and hDFSC in the co-culture system (data not shown).


Interactions of anaerobic bacteria with dental stem cells: an in vitro study.

Biedermann A, Kriebel K, Kreikemeyer B, Lang H - PLoS ONE (2014)

Bacterial growth in cell culture medium.Growth rates of bacteria in DMEM cell culture medium under anaerobic conditions (10% CO2, 10% H2, 80% N2), 37°C. Bacteria were grown in PYG medium supplemented with 5 µg/ml hemin and 1% vitamin K to the stationary phase. Subsequently, they were centrifuged, washed in PBS, and each bacterial suspension was diluted in DMEM 1∶10. The OD600 was measured at 600 nm from timepoint 0 every hour to timepoint 12 h with a final measurement after 24 h.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4219685&req=5

pone-0110616-g002: Bacterial growth in cell culture medium.Growth rates of bacteria in DMEM cell culture medium under anaerobic conditions (10% CO2, 10% H2, 80% N2), 37°C. Bacteria were grown in PYG medium supplemented with 5 µg/ml hemin and 1% vitamin K to the stationary phase. Subsequently, they were centrifuged, washed in PBS, and each bacterial suspension was diluted in DMEM 1∶10. The OD600 was measured at 600 nm from timepoint 0 every hour to timepoint 12 h with a final measurement after 24 h.
Mentions: The bacterial growth was determined in cell culture medium under anaerobic conditions. All four tested species showed normal bacterial growth (Fig. 2). From these results, we concluded that co-culture with anaerobic bacteria and hDFSC (and hBMSC, hGiF, or Ca9-22) was possible. Live/dead staining and fluorescence microscopy confirmed the survival of bacteria and hDFSC in the co-culture system (data not shown).

Bottom Line: Similar results were observed for host cell internalization (F. nucleatum: 0.002% internalization into hDFSC; 0.09% internalization into Ca9-22).The IL-10 response of the differentiated cells was found to be low in relation to their pro-inflammatory IL-8 response.The results indicate that dental stem cells are less prone to interactions with pathogenic bacteria than differentiated cells in an anaerobic environment.

View Article: PubMed Central - PubMed

Affiliation: Department of Operative Dentistry and Periodontology, University of Rostock, Rostock, Germany.

ABSTRACT

Background: In patients with periodontitis, it is highly likely that local (progenitor) cells encounter pathogenic bacteria. The purpose of this in vitro study was to elucidate how human dental follicle stem cells (hDFSC) react towards a direct challenge with anaerobic periodontal pathogens under their natural oxygen-free atmosphere. HDFSC were compared to human bone marrow mesenchymal stem cells (hBMSC) and differentiated primary human gingival fibroblasts (hGiF), as well as permanent gingival carcinoma cells (Ca9-22).

Methodology/principal findings: The different cell types were investigated in a co-culture system with Porphyromonas gingivalis (P. gingivalis) and Fusobacterium nucleatum (F. nucleatum). The viability of the cells and pathogens under anaerobic conditions, as well as interactions in terms of adherence and internalization, were examined. Additionally, the release of pro-inflammatory interleukin-8 (IL-8) and anti-inflammatory interleukin-10 (IL-10) was quantified via enzyme-linked immunosorbent assay. The bacteria adhered less efficiently to hDFSC compared to Ca9-22 (P. gingivalis: 0.18% adherence to hDFSC; 3.1% adherence to Ca9-22). Similar results were observed for host cell internalization (F. nucleatum: 0.002% internalization into hDFSC; 0.09% internalization into Ca9-22). Statistically significantly less IL-8 was secreted from hDFSC after stimulation with F. nucleatum and P. gingivalis in comparison with hGiF (F. nucleatum: 2080.0 pg/ml--hGiF; 19.7 pg/ml--hDFSC). The IL-10 response of the differentiated cells was found to be low in relation to their pro-inflammatory IL-8 response.

Conclusions/significance: The results indicate that dental stem cells are less prone to interactions with pathogenic bacteria than differentiated cells in an anaerobic environment. Moreover, during bacterial challenge, the stem cell immune response seems to be more towards an anti-inflammatory reaction. For a potential future therapeutic use of hDFSC, these findings support the idea of a save application.

Show MeSH
Related in: MedlinePlus