The computer-controlled artificial mouth used in this study is an advanced model system used for caries research . The model mimics real mouth conditions in terms of temperature, humidity, sucrose supply, and saliva flow rate. This artificial mouth model could provide useful information to predict clinical outcome. Based on the findings we got from this study, subsequent study could be performed to assess the extent of the data extrapolation. Nevertheless, the artificial mouth model system creates its own stable environment that differs from in vivo situations. The limitations of such an in vitro study must be taken into account when considering the findings of this study. It is noteworthy that dental caries is a polymicrobial infection process, and the mouth had over 700 species of oral bacteria . Five common cariogenic bacteria have been used to form cariogenic biofilm in this study. It is however still far from and is incomparable with the natural biofilm causing caries. The results cannot be extrapolated directly to the in vivo situation, and caution should be exercised in their interpretation.
The early stage of bacterial invasion in the caries process involves Streptococci, Lactobacilli, and Actinomycetes . Streptococci mutans and Streptococci sobrinus are two of the most important odontopathogens involved in the initiation and progression of caries . Caries can be artificially induced in vitro through inoculation onto tooth enamel. Lactobacilli rhamnosus and Lactobacilli acidophilus are the two most abundant species frequently found in both superficial and deep carious lesions, where the pH tends to be acidic . Actinomycetes naeslundii has the potential to invade dentinal tubules and is associated with root caries . For the foregoing reasons, these five cariogenic species from the three specific genera were chosen to form a multi-species cariogenic biofilm.
SDF has been used to manage caries in particular in patients with high caries prevalence . It’s simplicity of application enables it’s use on young children . The inherent disadvantage of using SDF to arrest caries is that the lesions will be stained black. It has been suggested that when carious dentin is treated with SDF, silver phosphate is formed and precipitated . 38% SDF is a colorless solution containing 44,800 ppm fluoride ions. Its high fluoride concentration can be toxic when swallowed in large doses .
This study demonstrated SDF-inhibited cariogenic biofilm formation. The inhibition was obvious in the first 7 days after SDF application based on a very low CFU counting, and the CFU increased over time. Since the amount of bacteria in the biofilm after 7 days was small, the variation between samples was relatively large (Table 1). In this study, computer software was used to differentiate different color and area to make quantitative analysis of CLSM . The results however might vary due to uneven distribution of the amount bacteria in different thickness of the biofilm. Furthermore, the quality of images could be affected by conditions such as brightness, white balance and contrast. Therefore, findings of the CLSM were used support the conclusion.
Both the CFU and CLSM results corroborate the manufacturer’s suggestion that repeated SDF application after one week will enhance its effectiveness in caries arrest. The 38% SDF solution contains high concentrations of silver (253,870 ppm) and fluoride (44,800 ppm) ions. Both the silver and fluoride ions released from the SDF appeared to have inhibited the cariogenic biofilms. Silver ions are bactericidal metal cations that inhibit biofilm formation  by inactivating and interfering with the bacterial synthesis of cellular polysaccharides through the inactivation of the glycosyltransferase enzymes responsible for the synthesis of soluble and insoluble glucan. Glucan not only contributes to the bulk of biofilms, but also plays an essential role in the sucrose-dependent adhesion of organisms to tooth surfaces . Fluoride in high concentration also can inhibit biofilm formation . Fluoride ions can bind to bacterial cell constituents and influence enzymes, such as enolase and proton-extruding adenosine triphosphatase (ATPase). The latter enzymes are considered to effectively inhibit the carbohydrate metabolism of acidogenic oral bacteria, as well as their sugar uptake .
Studies have shown that changes in the microhardness of dentin are directly related to its mineral content [19, 28]. Therefore, measuring the microhardness is a reasonable, indirect method of examining the mineral content of dentin. Studies reported that SDF treatment increased the microhardness of carious dentin . Another study reported that less soluble or virtually insoluble calcium fluoride, silver phosphate, and silver protein were formed and precipitated on the dentin surface when SDF was applied . This formed an insoluble protective layer that decreased calcium and phosphorous loss from the carious lesions. In this study, precipitates containing high silver and phosphorus content (measured by EDX, data not shown) were observed occluding the tubule orifices after SDF application.
The organic dentin matrix mainly consists of collagen, which is the structural backbone holding the hydroxyapatite together . Collagen fibers are exposed when hydroxyapatite is dissolved by acid. This profoundly increases the surface area and exposes more carbonyl groups. As a result, there is an increase in the FTIR signal on demineralized dentin . In this study, the log [amide I: HPO42-] of the control group increased over time, suggesting that more collagen I was exposed and less HPO42- remained in the dentin surface. Therefore, the demineralization of dentin progressed over time. In contrast, a lower log [amide I: HPO42-] value was found in the test group, indicating that SDF could protect collagen I from further exposure, and that a regain of HPO42- might also occur . Our recent laboratory study indicated that SDF has an inhibitory effect on matrix metalloproteinases (MMPs) . MMPs play a crucial role in collagen breakdown in carious lesions . SDF’s inhibition of MMP activities can protect against collagen degradation.