Oral Biofilm-stimulated Human Gingival Epithelium Differentially Regulates Inflammatory Responses in Co-cultured Immune Cells (original) (raw)

Brown, Jason ORCID: https://orcid.org/0000-0002-8168-5333, Johnston, William, Delaney, Christopher, Butcher, John, Khan, Shaz, Bradshaw, David, Culshaw, Shauna ORCID: https://orcid.org/0000-0002-9653-5629 and Ramage, Gordon ORCID: https://orcid.org/0000-0002-0932-3514(2019) Oral Biofilm-stimulated Human Gingival Epithelium Differentially Regulates Inflammatory Responses in Co-cultured Immune Cells. Montana Biofilm Science & Technology Meeting, Montana, USA, 16-18 July 2019.

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Abstract

Introduction: Oral diseases including gingivitis and periodontitis (PD) arise through a dysregulated immune response to microbial challenge. The gingival epithelium provides the first line of defence against the oral microbiota, acting as a physical and immunological barrier in the oral cavity. Communication between the gingival epithelium and the immune cells patrolling the epithelial barrier is vital in maintaining symbiosis between the host and microbiota. Here, we combined a 3D model of immune cells, gingival epithelium and complex biofilms in order to study the bacterial-host cell interactions (Figure 1). Methods: In vitro grown ‘health-associated’, gingivitis-associated’ and ‘periodontitis-associated’ bacterial biofilms were co-cultured with the human gingival epithelium (HGE) to investigate the host inflammatory response to different biofilms. Immune cells (peripheral blood mononuclear cells (PBMCs) and CD14+ monocytes) were also cultured with or without un-stimulated and stimulated HGE. Epithelial/immune cell gene expression and protein release were assessed by quantitative PCR and ELISA. The morphology and integrity of the epithelium was evaluated by histological staining; the associated biofilms were visualised by scanning/transmission electron microscopy (SEM/TEM) and confocal laser scanning microscope (CLSM) imaging. Results: HGE used in the study formed 6-8 layers, which histologically demonstrated characteristics of gingival epithelium. The different multi-species oral biofilms containing “health-associated”, “gingivitis-associated” and “periodontitis-associated” microbiota possessed unique composition, architecture and morphologies as assessed by quantitative PCR and SEM, TEM and CLSM imaging. We observed unique inflammatory responses in HGE cultured with multi-species oral biofilms. The “health-associated” biofilm had minimal impact on the epithelium whilst “gingivitis-associated” and “periodontitis-associated” biofilms induced a pro-inflammatory response, including elevated interleukin-8 gene expression and increased protein release. Biofilm-stimulated HGE differentially regulated inflammatory responses in co-cultured PBMCs and CD14+ monocytes. Conclusions: Our results show 3D epithelial models may offer a substantial advantage for in vitro study of host-pathogen interactions. These tissue models recapitulate similar inflammatory responses and cell signalling interactions observed at the oral mucosa in certain diseases like gingivitis and PD. This platform allows in vitro dissection of the impact of oral biofilms cultured with epithelial tissue and immune cells. Findings from this 3D model may contribute to our understanding of the pathogenesis of oral inflammatory diseases in vivo.

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