BackgroundMicrobial 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.ResultsWe 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.ConclusionWe 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.

A method for reproducibly determining minimal inhibitory concentrations and minimal bactericidal concentrations of plant extracts towards fastidiously and facultatively anaerobic oral bacteria, predicated upon measurements of optical densities in microtitre plate wells, was devised. The antimicrobial properties of some botanical oils were surveyed; of these, Australian tea tree oil, peppermint oil, and sage oil proved to be the most potent essential oils, whereas thymol and eugenol were potent essential oil components.

Periodontal disease is an inflammatory condition that destroys the tooth-supporting tissues. The inflammation is initiated by oral bacteria in the form of multi-species biofilms, and is dominated by cytokines of the IL-1 family. IL-1 activation and processing is regulated by Caspase-1, within intracellular protein complexes, known as "inflammasomes". The present study employed culture supernatants of in vitro supragingival and subgingival biofilms, to challenge human GF cultures for 6h. The gene expression of inflammasome complex components was investigated by TaqMan qPCR. NLRP1 expression was not affected, whereas NLRP2 was not expressed. Supragingival biofilm challenge increased the expression of Caspase-1, the adaptor ASC, AIM2, as well as IL-1β and IL-18, but did not affect NLRP3 expression. Subgingival biofilm challenge enhanced Caspase-1, ASC, AIM2, IL-1β and IL-18 gene expression at lower concentrations, followed by their down-regulation at higher concentrations, which was also evident for NLRP3 expression. Hence, supragingival and subgingival biofilms differentially regulate the gene expressions of NLRP3 and AIM2 inflammasomes and their down-stream IL-1 targets. Increased inflammasome transcription in response to supragingival biofilms is commensurate with early inflammatory events in periodontal disease, whereas decreased transcription in response to subgingival biofilms corroborates the dampening of host immune responses, in favour of pathogen survival and persistence.

BACKGROUND:In melanoma, different classification systems have been proposed that predict overall survival (OS) and recurrence-free survival (RFS) based on findings in the sentinel lymph node (SLN). The authors of this report compared the RFS and OS of 697 melanoma patients as predicted by various classification systems. METHODS: The Rotterdam system (based on the greatest dimension of the largest tumor cell deposit), the Augsburg S-classification (based on tumor penetrative depth [TPD]), and the Hannover system (based on a combination of tumor load, TPD, and invasion of the capsule) were studied in 697 consecutive melanoma patients who underwent SLN biopsy at the authors' center. RESULTS: In univariate analyses, the Rotterdam and Hannover systems (but not the S-classification) identified 1 group of SLN-positive patients that had OS and RFS similar to the OS and RFS of SLN-negative patients. The intermediate groups from all classification systems did not differ significantly with regard to RFS and/or OS from the adjacent groups. In multivariate analysis using a Cox model, the greatest dimension of the largest tumor cell deposit (cutoff point, <0.1 mm vs 0.1 mm), the TPD (cutoff point, 2 mm vs >2 mm), and capsular involvement represented independent parameters for RFS; and TPD and capsular involvement also were independent parameters for OS. On the basis of these 3 parameters, a new scoring system for risk assessment in patients with melanoma is proposed that can distinguish 3 separate groups of patients that differed significantly in OS and RFS. CONCLUSIONS: Different parameters of independent prognostic significance were identified in SLNs from patients with melanoma. Combining these parameters, the prognosis of patients with melanoma was predicted more precisely by the new scoring system than by currently published classification systems. Cancer 2010;116:3178-88.