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Immunochemical detection of CD14 on human gingival fibroblasts in vitro
1Department of Periodontology, Kagoshima University Dental School, Japan.
Abstract:
The activation of monocytes and macrophages induced by lipopolysaccharide has been shown to contribute to the binding of lipopolysaccharide and lipopolysaccharide-binding protein complex to the cell surface CD14 molecule. To clarify the mechanism of the lipopolysaccharide-induced modulation of the function of gingival fibroblasts, we investigated the effect of anti-CD14 on interleukin 6 (IL-6) production on human gingival fibroblasts in vitro. Immunochemical staining revealed weak positivity for CD14 on fibroblasts from healthy gingiva, while strong positivity for CD14 was found on fibroblasts from inflamed gingiva. Western blot profiles of the fibroblasts and monocytes showed a CD14-positive reaction at 55 kDa. Fluorescein isothiocyanate-conjugated Escherichia coli lipopolysaccharide bound to fibroblasts more strongly in the presence of 10% fetal bovine serum than without serum. This binding, as well as IL-6 production, was blocked by anti-CD14 monoclonal antibody. The results showed that CD14 was present on human gingival fibroblasts, which suggests that lipopolysaccharide modulation of gingival fibroblast function depends on CD14.
Insights
Lipopolysaccharide (LPS) activates immune cells via CD14. This study shows CD14 on gingival fibroblasts mediates LPS binding and interleukin-6 (IL-6) production, crucial for gum inflammation.
Area of Science:
- Oral biology
- Immunology
- Cell biology
Background:
- Lipopolysaccharide (LPS) activates monocytes and macrophages through CD14.
- The role of CD14 in modulating gingival fibroblast function by LPS is not fully understood.
Purpose of the Study:
- To investigate the presence and function of CD14 on human gingival fibroblasts.
- To clarify the mechanism of LPS-induced modulation of gingival fibroblast function.
Main Methods:
- Immunochemical staining and Western blot analysis to detect CD14 expression.
- In vitro experiments using fluorescein isothiocyanate-conjugated E. coli LPS to assess binding to fibroblasts.
- Assessing the effect of anti-CD14 monoclonal antibody on LPS binding and IL-6 production.
Main Results:
- CD14 was detected on human gingival fibroblasts, with higher expression in inflamed gingiva.
- LPS binding to fibroblasts was enhanced by fetal bovine serum and blocked by anti-CD14 antibody.
- Anti-CD14 antibody inhibited LPS-induced IL-6 production by gingival fibroblasts.
Conclusions:
- CD14 is expressed on human gingival fibroblasts.
- CD14 plays a critical role in mediating LPS binding and subsequent IL-6 production in gingival fibroblasts.
- These findings suggest CD14 is involved in the inflammatory response of gingival fibroblasts to LPS.