Research Article: An in vitro model to assess effects of a desensitising agent on bacterial biofilm formation

Date Published: December 24, 2019

Publisher: Taylor & Francis

Author(s): Jamie Coulter, Nicholas S. Jakubovics, Philip M. Preshaw, Matthew J. German.


Desensitising agents are added to dentifrices to occlude exposed dentine tubules and reduce pain associated with dentine hypersensitivity. In occluding the tubules these agents may alter the surface layer of the dentine and consequently affect bacterial biofilm formation. This research sought to examine the effects of desensitising agents on dentinal biofilms using an in vitro model. A constant depth film fermenter (CDFF) was selected to mimic the oral environment and human dentine with exposed tubules was analysed. Calcium sodium phosphosilicate (CSPS) was selected as a model desensitising agent. Dentine discs were treated with pumice or CSPS-containing dentifrices with or without fluoride, or left untreated (control). Dual-species biofilms of Streptococcus mutans and Streptococcus sobrinus were grown in artificial saliva and analysed by viable counts, polymerase chain reaction (PCR) and scanning electron microscopy (SEM). SEM images confirmed the presence of occluded tubules after CSPS application and demonstrated the formation of biofilms containing extracellular matrix material. Analysis of PCR and viable count data using a one-way ANOVA showed no significant differences for bacterial composition for any of the four treatments. There were, however, trends towards increased numbers of bacteria for the pumice and CSPS treated samples which was reversed by the addition of fluoride to CSPS. In conclusion, CSPS was not found to have a significant effect on biofilms and an in vitro model for testing desensitising agents has been developed, however, further work is required to improve the reproducibility of the biofilms formed and to explore the trends seen.

Partial Text

Dentine hypersensitivity occurs when a tooth with exposed dentine interprets relatively innocuous stimuli as noxious [1]. One explanation for dentine hypersensitivity is the hydrodynamic theory; this stipulates that a stimulus causes movement of fluid within dentine tubules resulting in nerve depolarisation and a painful stimulus. Thus, dentine tubules exposed to temperature changes or air pressure could result in fluid movement and pain [2]. Some treatments such as the desensitising agents added to dentifrices focus upon blocking or occluding the dentinal tubules in an attempt to reduce sensitivity. One product which works via this mechanism is calcium sodium phosphosilicate (CSPS). This forms a layer of carbonated hydroxyapatite crystals on dentine when in contact with an aqueous environment [3]. CSPS occludes significantly more dentine tubules and decreases dentine permeability significantly more than other control toothpastes [4,5].

A CDFF was selected to model cariogenic biofilms due to its similarities to the oral environment. Specifically, biofilms are grown on a solid surface and nutrients are provided in a thin film which is constantly replenished [15]. CDFFs have been used previously in dental research, for example to test the demineralisation of dentine [11] or how the roughness of denture material effects biofilm formation [16]. However, these studies have used bovine tooth tissue, synthetic hydroxyapatite or restorative materials which do not represent a dentinal surface with exposed tubules that would receive CSPS treatment. Creating dentine discs from human teeth has resolved this issue, and patent dentine tubules were clearly observed. CSPS-containing dentifrices were selected as an example of a desensitising agent and SEM images such as Figure 2 revealed tubule occlusion and deposition of a surface layer following its use, something which appears to remain during biofilm growth. These images agree with what has occurred in previous CSPS in vivo research [4] and so it appears that the model mimics the action of desensitising agents in the clinical environment.

Statistical analysis of the qPCR and viable count data found none of the treatments to be significantly different from each other, indicating that CSPS pastes cause no significant change to biofilm formation compared to control dentifrices. However, on examining the data trends were found towards increased numbers of S. mutans and S. sobrinus in some circumstances that are worthy of further exploration with larger sample numbers. Importantly, an in vitro model for testing the impact of desensitising agents on dentine biofilms has been developed using the CDFF to mimic the oral environment, dentine discs with exposed dentine tubules and Mutans streptococci to represent the cariogenic challenge. Future experiments will aim to improve the reproducibility of the biofilms and will compare the effects of different desensitising agents on oral biofilm formation in the CDFF and against other established models of biofilm formation such as those using samples mounted on intra-oral devices.




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