Hygiene Tribune Middle East & Africa No. 1, 2015
A clinical assessment of the efficacy of a Stannous - containing Sodium Fluoride Dentifrice on Dentinal Hypersensitivity / Removal of interproximal dental biofilms by high-velocity Water Microdrops
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Dental Tribune Middle East & Africa Edition | January-February 2015
hygiene tribune 1B
A clinical assessment of the efficacy
of a Stannous - containing Sodium Fluoride
Dentifrice on Dentinal Hypersensitivity
Originally published in the Journal of Contemporary Dental Practice, 11, No. 1, January 1, 2010
By Trevor N. Day, PhD;
Johannes Einwag, Prof. Dr.
med. dent.; Joachim S. Hermann, PD Dr. med. dent.; Tao
He, DDS, PhD; Mary Kay Anastasia, BA; Matthew Barker,
PhD; Yuqing Zhang, MS
jects with (medium) Oral-B Advantage 40 toothbrushes (The
Procter & Gamble Company,
Cincinnati, OH, USA). The test
products were supplied in kits
containing the assigned toothpaste, toothbrush, and written
usage instructions. The dentifrice in both kits was supplied
blinded in white tubes.
A
im: To measure the desensitizing benefits of an
experimental stannouscontaining sodium fluoride dentifrice versus a regular sodium
fluoride negative control.
Methods and Materials: This
study was a randomized, doubleblind, parallel group, fourweek
clinical trial. Subjects reporting
dentinal hypersensitivity were
enrolled and randomized to the
experimental dentifrice or the
control dentifrice to use twice
daily for four weeks.
Efficacy assessments (Air Blast)
were performed at baseline and
weeks two and four. Separate
analyses were performed for the
two most sensitive teeth at baseline and for all 12 teeth. Results
for weeks two and four combined also were analyzed.
Results: Thirty-one subjects
were included in the analyses.
For the two most sensitive teeth,
the experimental dentifrice
showed statistically significantly
less sensitivity (p<0.05) versus
the control at weeks two and
four and for weeks two and four
combined. The sensitivity reduction ranged from 24.9% to
28.4% over the control. For all
12 teeth, the experimental group
had statistically significantly
(p<0.03) lower sensitivity scores
versus the control group at week
two and weeks two and four
combined.
Conclusion: The experimental
dentifrice demonstrated significant desensitizing advantages
versus the control.
Clinical Significance: This stannouscontaining sodium fluoride
dentifrice provides an effective treatment for patients with
dentinal hypersensitivity, significantly reducing sensitivity
versus a negative control in this
four-week trial. Keywords: Stannous, dentifrice, sodium fluoride, sensitivity, clinical trial.
Introduction
Dentinal hypersensitivity is a
highly prevalent condition reported to affect from 4% to 57%
of the population.1,2 The causes
of sensitivity are well characterized as exposed dentinal tubules
most commonly resulting from
gingival recession followed by
loss of cementum. The mechanism by which nerves are triggered to result in the pain associated with hypersensitivity is
now widely accepted as that of
the Brännström hydrodynamic
theory.3 This postulates that
changing physical conditions on
the dentin surface such as heat,
pressure, or osmotic potential
give rise to fluid movement in
the tubules.4–6 The consequent
pressure change stimulates the
nerves giving rise to the pain.
The mechanism of action of
stannous ions7 in reducing dentinal hypersensitivity has been
found to be the precipitation of
stannous compounds occluding the dentinal tubules and
thus preventing stimulation of
the nerves in the pulp cavity. In
vitro studies using various techniques, such as scanning electron microscopy, electron probe
microanalysis, and Vickers surface microhardness, demonstrate deposition of tin and fluoride on the surface and covering
of the dentinal tubules.8,9 One
laboratory evaluation showed
that while both zinc and tin covered or obturated tubules, zinc
was largely removed by washing whereas tin remained covering the tubules.10 Another study
showed specimens treated with
stannous fluoride (Crest® ProHealth®, The Procter & Gamble
Company, Cincinnati, OH, USA)
appeared to resist acid solubilization.9
A number of clinical studies
also have been conducted to
investigate the effectiveness of
stannous-containing oral care
products upon dentinal hypersensitivity. Most of the early
studies focused on gels containing 0.4% stannous fluoride,11
whereas the majority of contemporary trials have evaluated
stannous-containing dentifrice
formulations.12–18 The collective
findings demonstrate the effectiveness of numerous stannouscontaining products in reducing
sensitivity.
Recently,
a
new
stannous-
containing sodium fluoride
dentifrice was developed. This
clinical trial was conducted to
evaluate the effectiveness of this
formulation relative to a negative control for the treatment of
dentinal hypersensitivity.
Methods and Materials
Study Design
This was a randomized, parallel
group, doubleblind, four-week
clinical trial to assess changes
in subject perceived tooth hypersensitivity via air blast induced examiner grade assessment among subjects using a
stannous-containing
sodium
fluoride dentifrice compared to
those using a negative control
dentifrice. Measurements were
conducted at baseline, week
two, and week four visits.
Entrance Criteria
Following Ethics Committee
approval, at least 30 generally
healthy adults age 18–70 reporting tooth sensitivity were sought.
Subjects had to agree to refrain
from using anti-hypersensitivity products or having elective
dental procedures (including
prophylaxis) performed during
the study.
Subjects who were currently
using an antisensitivity toothpaste or another anti- sensitivity
product or who had used such a
product in the previous month
were excluded. Subjects with
carious teeth or with any other
condition that the investigator
considered may compromise
the results also were excluded.
Subjects taking daily doses of
anticonvulsants, sedatives, tranquilizers, or other mood-altering
drugs were excluded as well as
subjects with a history of significant adverse effects following
the use of oral hygiene products
such as toothpaste and mouth
rinse.
Test Dentifrices, Assignment
to Treatment Sequence
The two treatments used in this
study were:
1. An experimental stannouscontaining sodium fluoride dentifrice with 1450 ppm F- sodium
fluoride and stannous chloride
as a key excipient (Procter &
Gamble UK, Surrey, United
Kingdom)
2. Crest® Decay Protection (UK)
with 1450 ppm F- sodium fluoride (Procter & Gamble UK, Surrey, United Kingdom)
Both were supplied to the sub-
Subjects were stratified at baseline into one of four strata depending on their gender (female
or male) and the baseline selfreported tooth hypersensitivity
(low or high). Within strata, subjects were randomly assigned to
one of the two treatment groups
using an encoded program. Subjects residing in the same house-
> Page 2B
[2] =>
2B hygiene tribune
Dental Tribune Middle East & Africa Edition | January-February 2015
< Page 1B
hold were assigned to the same
treatment group.
Treatment Regimens
Subjects used the assigned products for the first time under supervision at the clinical site. Subjects used the products at home
in place of their normal toothbrush and dentifrice for a period
of four weeks. Subjects were instructed to brush twice daily for
two minutes each time. Subjects
were instructed not to alter their
other oral hygiene habits (e.g.,
flossing) with the exception that
no anti–tooth hypersensitivity
products should be used.
Air Blast Tooth Specific Sensitivity Assessment
The thermal sensitivity perceived by the subject was measured by the examiner by directing an air blast individually at
each of the premolars and canines at baseline, week two, and
week four visits. Each tooth was
isolated with cotton rolls and
the air blast was delivered from
a distance of 1.0 centimeter for
1 second. The following scale19
was used by the examiner to assess the level of hypersensitivity
for each of the 12 teeth examined:
• 0 – Absence of pain, but perceiving stimulus
• 1 – Slight pain
• 2 – Pain during application of
stimulus
• 3 – Pain during application
of stimulus and immediately
thereafter
Statistical Methods
For air blast–induced hypersensitivity scores, separate analyses were performed for the two
teeth with the most sensitivity
at baseline and for all 12 teeth
combined. Analysis of covariance (ANCOVA) with treatment
as a factor and the baseline score
and age as the covariates were
used to assess treatment differences in hypersensitivity at the
post-baseline visits. Also for the
hypersensitivity scores, separate
repeated measures models were
used to investigate the overall
relationship between the treatment groups and the post-baseline visits (weeks two and four)
with statistical testing for the interaction and overall treatment
effects using a two-sided 5% significance level. In this study, the
interaction between treatment
and week was not statistically
significant (p>0.45) for each hypersensitivity score, and the interaction was removed from the
repeated measures model.
Results
Thirty-one subjects were enrolled at the baseline visit, received product, and completed
the study through week four.
Subjects ranged in age from 23
to 65 years with an average of 42
years, and 68% of the subjects
were female. The treatment
groups were balanced (p>0.86)
for all demographic characteristics. Mean baseline scores
were not significantly differ-
Table 1. Between treatment comparison of air blast score for two most
sensitive teeth
Figure 1. Overall results pooling
weeks two and four from the
repeated measures analysis.
ent (p>0.56) between groups at
baseline for either the two most
sensitive teeth or for all 12 teeth
combined.
Efficacy Results
At the week two and week four
post-baseline visits and combining weeks two and four, the
experimental group had mean
air blast scores for two most
sensitive teeth that were 28.4%,
24.9%, and 27% lower, respectively, than the control group
(p<0.05, Figure 1). At week two,
mean scores for the experimental and control groups were 1.51
and 2.11, respectively (Table 1).
At week four, the experimental
group had a mean score of 1.42
compared to 1.89 for the control
group. The weeks two and four
combined mean score was 1.46
for the experimental group and
2.00 for the control group.
At week two and combining
weeks two and four, the experimental group provided significantly (p<0.03) lower mean air
blast scores for all 12 teeth relative to the control group (Table
2).
The experimental group had
a mean score of 1.18 at week
two while the mean score for
the control group was 1.52. The
weeks two and four combined
score was 1.17 for the experimental group and 1.46 for the
control. At the week four visit,
the experimental group provided directionally (p=0.07)
lower mean air blast scores for
all 12 teeth relative to the control
group (1.16 and 1.40, respectively). This difference was not
statistically significant using a
two-sided 5% significance level.
The desensitizing benefit of the
experimental dentifrice over the
control was 22.4% at week two,
17.1% at week four, and 19.9%
for the combined weeks two and
four visits (Figure 1).
Safety Results
One subject in the experimental
group had a possible related adverse event (desquamation) observed by the examiner that was
mild in nature. Another subject
in the experimental group had
a nontreatment-related adverse
event (herpetic lesion) reported
and observed that was mild in
nature.
Table 2. Between treatment comparison of air blast score for all 12
teeth.
Discussion
In this clinical trial, the experimental group exhibited a significantly greater reduction in
tooth sensitivity via air blast
measurements than the control among the two most sensitive teeth (p<0.05) at both postbaseline measurements and the
combined weeks two and four
visits. The experimental group
also demonstrated significantly
greater reductions than the
control in tooth sensitivity via
air blast measurements among
all 12 teeth on post-baseline
measurements at week two and
the combined results for weeks
two and four (p<0.03). The assessment of all 12 teeth was included in this trial for comparative purposes but is not a widely
used measure in sensitivity trials since the condition typically
does not affect each tooth.
These results are aligned with
other studies evaluating stannous-containing
dentifrices.
Five trials in the literature evaluated the desensitizing effects of a
combination of 0.454% stannous
fluoride and 5% potassium nitrate relative to other products;
four trials compared the dentifrice to a positive control sensitivity toothpaste containing 5%
potassium nitrate and the fifth
study was versus a commercial
nondesensitizing control dentifrice.12–16 The effectiveness of the
product containing the combination of stannous fluoride with
potassium nitrate was found
to be greater than that of the
sodium fluoride product with
potassium nitrate12–15 and the
nondesensitizing control.16 Two
published randomized parallel
group studies were conducted
on a dentifrice containing stannous fluoride compared to a sodium fluoride negative control
toothpaste.17,18 In both studies,
the sensitivity scores of the stannous fluoride group after four
and eight weeks of product usage were statistically significantly lower than the control toothpaste group, demonstrating the
effectiveness of the stannous
fluoride toothpaste in reducing
dentinal hypersensitivity.
In addition, the significant reductions in dentinal hypersensitivity demonstrated by the stannouscontaining sodium fluoride
dentifrice, now marketed in
parts of Europe and China, are
consistent with outcomes of
other research on this particular formulation. An eight-week,
randomized, parallel group, two
treatment, double-blind study
was conducted among generally healthy adults with moderate thermal and tactile dentinal
hypersensitivity.20 Subjects were
stratified at baseline according
to age, gender, and thermal dentinal sensitivity scores and randomly assigned to one of the two
treatments: the new stannouscontaining sodium fluoride dentifrice or a marketed potassium
nitrate control (Crest® Sensitivity Protection, The Procter &
Gamble Company, Cincinnati,
OH, USA) for twice daily usage.
Hypersensitivity was assessed
via Yeaple Probe and cold Air
Blast/Schiff Air Index for tactile and thermal assessments
respectively, at baseline, week
four, and week eight. Fifty-eight
subjects completed all evaluations. Both treatments produced
significant (p<0.05) reductions
in hypersensitivity compared to
baseline at both week four and
week eight time points. There
were no significant differences
between the two treatments at
either week four or week eight
(p>0.534) for either assessment.
One advantage of this stannouscontaining sodium fluoride dentifrice formulation relative to
other desensitizing treatments
is its effectiveness against other
common oral conditions. A recent study by He and colleagues
evaluated its plaque prevention efficacy relative to a positive (Colgate®Total®, ColgatePalmolive, New York, NY, USA)
and negative (Crest® Cavity
Protection, The Procter & Gamble Company, Cincinnati, OH,
USA) control dentifrice.21 Following a dental polish, subjects
brushed lingual surfaces only,
then swished with a slurry of the
dentifrice over the entire dentition twice per day over a fourday period. At baseline and after
four days, plaque levels were
assessed by the Turesky Modification of the Quigley–Hein
plaque index (TMQHPI). The
whole mouth TMQHPI plaque
scores after treatment for both
the experimental and the positive control dentifrices were statistically significantly lower than
those for the negative control by
11.4% and 8.4%, respectively
(p<0.0001).
Another recent study showed
the benefit of this stannouscontaining sodium fluoride
dentifrice against extrinsic
stain.22 While many stannous
luoride products can produce
minor extrinsic tooth stain, this
formulation uses a polychelation technology to stabilize the
stannous-fluoride
complex
and prevent stain. The study
also included an experimental
stannous-containing dentifrice,
a nonstaining marketed dentifrice (Colgate® Total®), and
a stannous fluoride dentifrice
(Crest® Gum Care, The Procter
& Gamble Company, Cincinnati,
OH, USA) previously established
to induce extrinsic stain. Following a baseline Lobene stain
examination, subjects received
a prophylaxis on the 12 anterior
teeth to remove extrinsic stain
and tartar. Subjects were randomly assigned based on baseline stain scores to receive one
of the four treatments and to use
them twice daily over a fiveweek period. Results showed
that there was significantly less
stain after product use in the
new stannous-containing sodium fluoride dentifrice group,
the experimental stannous-containing sodium fluoride dentifrice group, and the Colgate Total group compared to the Crest
Gum Care group (p<0.0001).
There were no other statistically
significant treatment differences
between the two stannous-containing sodium fluoride groups
and the Colgate Total group at
either time point for any Lobene
measures (p>0.145).
Further research is warranted
on this formulation to demonstrate the full breadth, as well as
magnitude, of benefits.
Conclusion
The stannous-containing sodium fluoride dentifrice provides
statistically significant benefits
for dentinal hypersensitivity and
should be considered as a home
care option for patients who experience this condition.
Clinical Significance
This stannous-containing sodium fluoride dentifrice provides
an effective treatment for patients with dentinal hypersensitivity.
References
1. Rees JS, Addy M. A cross-sectional study of buccal cervical
sensitivity in UK general dental
practice and a summary review
of prevalence studies. Int J Dent
Hyg. 2004; 2(2):64-9.
2. Irwin CR, McCusker P. Prevalence of dentine hypersensitivity
in a general dental population. J
Ir Dent Assoc. 1997; 43(1):7-9.
3. Jacobsen PL, Bruce G. Clinical dentin hypersensitivity: understanding the causes and prescribing a treatment. J Contemp
Dent Pract. 2001; 2(1):1-12.
4. Rosenthal MW. Historic review of the management of
tooth hypersensitivity. Dent Clin
North Am. 1990; 34(3):403-27.
5. Brännström M, Aström A.
The hydrodynamics of the dentine; its possible relationship to
dentinal pain. Int Dent J. 1972;
22(2):219-27.
6. Kramer IRH. The relationship
between dentine sensitivity and
movements in the contents of
the dentinal tubules. Br Dent J.
1955; 98: 391-2.
7. Miller S, Truong T, Heu R,
Stranick M, Bouchard D, Gaffar
A. Recent advances in stannous
fluoride technology: antibacterial efficacy and mechanism of
action towards hypersensitivity. Int Dent J. 1994; 44(1 Suppl
1):83-98.
8. Ellingsen JE, Rölla G. Treatment of dentin with stannous
fluoride—SEM and electron
microprobe study. Scand J Dent
Res. 1987; 95(4):281-6.
Full list of references available
from the publisher.
Article “Oral Probiotics - Overwiew” published in Hygiene
Tribune Nov-Dec 2014 edition is
written by Dr. JJ Smith, Dr. John
Nosti, Shirley Gutkowski and
Victoria Wilson.
[3] =>
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4B hygiene tribune
Dental Tribune Middle East & Africa Edition | January-February 2015
Removal of interproximal dental biofilms
by high-velocity Water Microdrops
By A. Rmaile, D. Carugo, L.
Capretto, M. Aspiras, M. De
Jager, M. Ward and P. Stoodley
A
bstract
The influence of the impact of a high-velocity
water microdrop on the detachment of Streptococcus mutans
UA159 biofilms from the interproximal (IP) space of teeth in
a training typodont was studied
experimentally and computationally. Twelve-day-old S. mutans biofilms in the IP space
were exposed to a prototype
AirFloss delivering 115 μL water at a maximum exit velocity
of 60 m/sec in a 30-msec burst.
Using confocal microscopy and
image analysis, we obtained
quantitative measurements of
the percentage removal of biofilms from different locations in
the IP space.
The 3D geometry of the typodont
and the IP spaces was obtained
by micro-computed tomography
(μ-CT) imaging. We performed
computational fluid dynamics
(CFD) simulations to calculate
the wall shear stress (τw) distribution caused by the drops on
the tooth surface. A qualitative
agreement and a quantitative relationship between experiments
and simulations were achieved.
The wall shear stress (τw) generated by the prototype AirFloss
and its spatial distribution on the
teeth surface played a key role in
dictating the efficacy of biofilm
removal in the IP space.
Key words: oral hygiene, Streptococcus mutans, micro-computed tomography, microscopy,
interproximal cleaning, dental
plaque.
Introduction
Good oral hygiene practice
maintains a healthy oral cavity,
controls the progress of dental
plaque biofilms (ten Cate, 2006)
and calculus, and prevents further complications such as gum
diseases and tooth decay (Costerton et al., 1999; Jakubovics
and Kolenbrander, 2010; Bjarnsholt et al., 2011; Marsh et al.,
2011).
The challenge of dental care
products is to efficiently and
quickly remove plaque from the
interproximal (IP) space. Mechanical removal of IP plaque
by traditional dental flossing
products has been accompanied
with bleeding, stuck or shredded floss, and prolonged flossing
time (Darby, 2003). Fluid shear
stress is an alternative mechanical approach for controlling
biofilm build-up (Stewart, 2012).
Previous studies have demonstrated that if sufficiently high
fluid shear stress can be generated, this alone can stimulate
biofilm detachment (Rutter and
Vincent, 1988; Hope et al., 2003;
Sharma et al., 2005a; Paramonova et al., 2009). High-velocity
water droplets (Cense et al.,
2006) and entrained air bubbles
(Parini et al., 2005; Sharma et al.,
2005b) have also been shown to
be able to remove bacteria and
biofilms from surfaces utilizing
the additional effect of generating a “surface-tension force”
way from the surface by the passage of an air/water interface
(Gómez-Suárez et al., 2001). An
advantage of using fluid forces
to remove biofilms is that mechanical forces can be projected
beyond the device itself, by generating currents in the fluid surrounding the teeth by powered
brushing (Adams et al., 2002) or
through the generation of water jets by oral irrigation (Lyle,
2011). However, continuous water jets have a disadvantage of
requiring large reservoirs and
can be messy to use because
of the large volumes of water
involved. More recently, the
Sonicare™ AirFloss device has
been introduced for removing
IP plaque. The AirFloss shoots a
microdrop volume of water and
entrained air at a high velocity
into the IP space in a discrete
burst, thus creating high wall
shear stress (τw) and high-impact pressure over short periods
of time, minimizing water volume and cleaning times.
We previously reported the influence of high-velocity water
microdrop impact on the detachment of artificial plaque
from the IP spaces, to demonstrate how a real biofilm might
detach (Rmaile et al., 2013).
Here, we go on to use the same
in vitro model to look at bacterial
biofilm removal and apply computational fluid dynamics (CFD)
numerical techniques to model
and predict the spatial distribution of fluid wall shear stress (τw)
required to remove the biofilm.
This paper reports the results of
an experimental and numerical
study on the influence of a highvelocity water microdrop impact
on the detachment of Streptococcus mutans biofilms from the
IP spaces of a typodont model.
Materials & Methods
Bacteria and Growth Media
Biofilms were grown from S.
mutans UA159 (ATCC 700610).
Stock cultures of S. mutans were
stored at -80o C in 10% glycerol
in physiological buffered saline
(PBS). Biofilms were cultured
with sucrose (2% w/v) supplemented brain heart infusion
(BHI+S) medium (Sigma-Aldrich, Dorset, UK) and incubated at 37o C and 5% CO2.
Figure 2. Representative CLSM images of S. mutans biofilm of 5 different locations (A, B, C, D, E) across the IP space at the level of the prototype AirFloss
tip from the proximo-labial to the proximo-palatal side of a maxillary central incisor (the 5 locations are identified clearly in Fig. 3). A1, B1, C1, D1,
and E1 are the images of the biofilm before the burst (on the untreated tooth),
and A2, B2, C2, D2, and E2 are the corresponding images after thresholding with ImageJ (the biofilm is in black in these images, while the white
areas are biofilm-free regions). Meanwhile, A3, B3, C3, D3, and E3 are the
images of the biofilm on the treated tooth after the burst, and A4, B4, C4, D4,
and E4 are the corresponding thresholded images. The untreated samples
(columns 1 and 2) and treated samples (columns 3 and 4) are not from the
same specimens. We calculated the % removal by subtracting the amount of
biofilm that remained from the original amount of biofilm.
Typodont Model and Microburst
To recreate a realistic geometry associated with the IP
space, we grew biofilms on the
2 upper central incisors (teeth
8 and 9) removed from a training typodont (A-PZ periodontal
model 4030025, Frasaco GmbH,
Tettnang, Germany) (Fig. 1A). A
prototype AirFloss was used to
generate a microburst of 115 μL
Figure 1. Digitization process of the training typodont. (A) Photograph
showing the typodont used in the study. (B) Micro-CT image of the typodont
(maxillary dental arch). (C) CAD-based 3D rendering of the IP space used
in the study. (D) The 3D meshwork showing the geometry of the tooth surface that was used for the computational simulations. The sketch (right)
shows the mesial view of a maxillary left central incisor, and the dashed
square shows the region of interest used in the study.
(±50; n = 30) over a time period
of approximately 0.033 sec (Appendix I).
CLSM Microscopy and Image
Analysis
The amount of biofilm on the
IP surfaces of the typodont teeth
was measured with a Leica TCS
SP2 AOBS (Leica Microsystems,
Nanterre, France) confocal laser
scanning microscope (Fig. 2; Appendix II).
Micro-computed Tomography
( μ-CT) Geometry
Reconstruction of the Typodont Model
μ-CT was used to image the typodont in 3D and construct a
model of the IP space to be used
in subsequent CFD modeling
(Fig. 1B; Appendix III).
Streptococcus mutans Biofilms inside
Microfluidic Channels
To estimate a critical hydrodynamic shear stress required for
S. mutans biofilm detachment,
which could be used as a model
input parameter for predicting
the spatial distribution of biofilm
removal, we used a BioFluxTM
1000 device (Fluxion Biosciences, South San Francisco, CA,
USA) (Appendix IV).
Computational Fluid Dynamics Simulations
To model the dynamic behavior
of the microburst created within
the IP space, the tomography obtained from μ-CT was converted
to a 3D computer-aided design
(CAD) file geometry with Amira
software (Mercury Computer
Systems, Fürth, Germany). The
computational domain, represented by the IP space, was discretized with software Gambit
2.4.6 (Symetrix Inc., Mountlake
Terrace, WA, USA) and using a
tetrahedral meshing scheme. A
cell size of ~155 μm was chosen,
which led to a total number of
143,985 mesh tetrahedral cells.
Since the IP space was symmetrical, only half of it was modeled,
reducing computational cost
and time.
CFD simulations were performed with ANSYS Fluent 12.1.4
software (ANSYS Inc., Canonsburg, PA, USA), which allowed
for the determination of the flow
field within the IP space and τw
generated on the tooth surface
(Appendix V).
Statistical Analysis
Statistical comparisons were
made by one-way analysis of
variance (ANOVA) (Excel 2003,
Microsoft). Differences were reported as statistically significant
for p < .05.
Results
3D Imaging of Typodont Model
High-resolution 3D images detailing the micro-architecture of
the typodont were obtained by
μ-CT (Fig. 1B). This allowed us
to computationally disassemble
the typodont, maintaining the
relevant juxtaposition between
the individual teeth, and to create computational meshes of the
teeth without interference from
the other typodont materials.
Quantification of Biofilm Removal
With confocal microscopy, S.
mutans biofilms grown in the IP
space showed bacterial cells aggregating and forming complex
cell cluster colonies consisting
of ‘tower’-, ‘mushroom’-, and
‘mound’-shaped structures. The
thickness of the resulting biofilm on each tooth surface was
approximately 200 to 300 μm.
After the microburst, the images
taken for the proximal surface of
the teeth showed almost no biofilm close to the nozzle tip of the
prototype AirFloss. Image analysis showed 95% removal close to
the tip, 62% removal at approximately half the labio-palatal distance from the tip to the back of
the teeth, and 8% removal at the
back of the teeth (Fig. 3). The
percentage removal values were
plotted vs. the distance from the
nozzle tip to the midpoint of
the palatal surface of the teeth
(Fig. 3A). The resulting curve
was compared with the values
obtained from the numerical
simulations for τw at the same
locations (Figs. 3C, 3D).
Critical Shear Stress for Biofilmaggregate Detachment
The morphology of the biofilms
in the BioFluxTM 1000 microfluidic channels varied markedly
between one channel and the
> Page 6B
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6B hygiene tribune
Dental Tribune Middle East & Africa Edition | January-February 2015
< Page 4B
Figure 3. Biofilm removal as a function of shear stress and distance from
the front of the tooth. (A) Percentage removal of the biofilm quantified from
the CLSM images at 5 different locations on the tooth surface in the IP space.
Three individual runs are shown by different symbols. The error bars represent standard deviations of the mean from 5 CLSM images. Solid line and
heavy bars are the mean of the individual means (n = 3), which have been
slightly offset for clarity. The schematic inset shows the proximal view of the
upper central incisor, where the black squares represent the different locations where the CLSM images were taken. (B) Contour map showing the
spatial distribution of τw on the tooth surface, as calculated from numerical
simulations (circular nozzle tip; z/H = 0.5). The color bar is a linear scale
showing the shear stress (Pa). (C) τw on the tooth surface (in kPa) at different
y-positions along the tooth (i.e., from labial to palatal side), at a fixed zposition (gingivo-incisal), as also calculated from numerical simulations.
Empty squares correspond to the measurement points (squares) in 3B. On
the secondary y-axis, the mean percentage removal measured experimentally in 3A (i.e., solid line) is plotted, with the 5 empty circles (denoted as A, B,
C, D, and E) corresponding to the same positions as in 3A. (D) Relationship
between percentage removal (determined experimentally) and τw on the
tooth surface (determined computationally). Datapoints were interpolated
with a linear trend (red line).
gates detachment shear stress”
(CDSSagg) of 1.7 Pa for the computational modeling. Above 2 Pa,
the smaller biofilm clusters still
appeared to be firmly attached
to the substrate, and remained
attached even after the shear
stress was increased to 3 Pa.
Numerical Simulations
Mesh Independence Study
A mesh independence study
was performed, and a cell size
of 0.155 mm was selected for
further numerical studies (Appendix V).
Figure 4. Effect of nozzle tip z-position
(z/H) on fluid τw spatial distribution over
the tooth surface. Tip cross-section is circular, and z/H was varied between 0.17 and
0.83 (a-c). The blue arrow indicates the
flow direction. The red area corresponds
to the tooth surface area where the shear
stress is lower than CDSSagg = 1.7 Pa.
other, and also within the same
channel. Structural heterogeneity is a common feature of
biofilms. Nevertheless, common
features could be noted, as seen
by the microscopic images (Appendix Fig. 2): (i) the presence
of individual bacteria and (ii) the
presence of biofilm clusters of
different sizes.
When τw was increased from 0 to
2 Pa, there was a slight increase
in the overall detachment rate
of the biofilm (Appendix Fig. 2),
seemingly caused by adhesive
failure (von Fraunhofer, 2012).
The bacterial cells as well as the
biofilmaggregates appeared to
slide along the surface before
coming off. There was minimal
detachment of the individual
bacteria and the small aggregates over the applied elevated
shear stress; but the larger biofilm clusters (with diameters
over ~50 μm) detached when
the shear stress ranged from 0.3
to 1.7 Pa. We extrapolated a conservative “critical biofilm-aggre-
Quantification of Wall Shear
Stress Distribution
A representative contour plot of
the fluid τw spatial distribution
on the tooth surface is shown in
Fig. 3B. This simulation corresponded to a velocity inlet of 60
m/sec, with the circular nozzle
tip located at z/H = 0.5, gingivo-incisally, where z (mm) is a
spatial coordinate from the supragingival base of the tooth perpendicular to the tip of the tooth,
and H (mm) is the supragingival
height of the tooth. Thus, z/H
= 0.5 equates to halfway up the
tooth. The simulation showed
the predicted fluid τw distribution on the proximal surface of
the tooth, starting from the labial
side of the IP space close to the
nozzle tip (τw ~2.7 kPa), to the
midpoint of the palatal surface
of the tooth (τw ~0.3 kPa).
Computational Prediction of
Shear Stress and Experimental
Biofilm Removal
The τw distribution obtained
computationally was compared
with experimentally measured
removal of biofilms. A linear
correlation of % removal as a
function of τw was found according to:
Percent removal = kτw (r2 = 0.94)
(1) where τw is wall shear stress
(in Pa), and k (in Pa-1) is the
slope of the interpolating function (Figs. 3C, 3D).
Effect of the Nozzle z-position
on Wall Shear Stress Distribution
Contours of fluid τw on the tooth
surface at 5 nozzle tip zpositions
were obtained to investigate the
effect of tip positioning on the
device’s hydrodynamic performance. Fig. 4 shows the tooth
surface area where τw is lower
than the critical value of 1.7 Pa.
Computational results predicted
that the maximum % of biofilm
removal would take place when
the nozzle tip is placed at z/H =
0.5 or z/H = 0.66, while the efficacy of biofilm removal would
be significantly reduced at extreme z/H positions, namely, z/H
= 0.17 (i.e., close to the gum line)
or z/H = 0.83 (i.e., close to the incisal edge).
Discussion
In the flow cell experiments, S.
mutans biofilms were successfully grown inside microchannels under gravitational flow
conditions. Under transmitted
light microscopy (Appendix Fig.
2), the biofilm size and morphology showed resemblance to
previously reported data (Costerton et al., 1999; Heersink et
al., 2003). When the biofilm was
subjected to an increased shear
stress from 0 to 2 Pa, the large
aggregates resisted movement
from the surface until the wall
shear stress reached a critical
value, or CDSSagg, which ranged
between 0.3 and 1.7 Pa, at which
they detached. However, even
at this critical value, the smaller
biofilm patches and individual
bacterial cells remained attached. Generally, detachment
of biofilm fragments (erosion),
or even of the entire biofilm
(sloughing), is caused by high
flow shear stress levels that exceed the adhesion strength of
the biofilm (Ohashi and Harada,
1994, 1996). The detachment of
the large aggregates occurred at
a relatively low shear stress (~1
Pa), while the smaller patches
remained firmly attached, even
after the flow increased up to 3
Pa. The S. mutans biofilms were
grown under static or low shear
conditions, thus leading to the
formation of large cell aggregates, which tend to be approximately circular compared with
the streamers that usually form
under dynamic conditions. The
streamlined shape has a significant effect on reducing the fluid
drag on the elongated biofilms
(Stoodley et al., 1998, 1999).
Streamers develop viscoelastic
flexible bodies which oscillate
rapidly when exposed to the flow
forces, thus resisting detachment better than the large circular biofilm-aggregates grown
at low laminar flow conditions.
The large circular aggregates
show different behavior under
flow, with less ability to flex, resulting in detachment at lower
fluid shear stress. This explains
the experimentally observed detachment of the large aggregates
at a relatively low shear stress
(~1 Pa). So, the CDSSagg estimated here describes the detachment involving large aggregates
only and not the total biofilm,
which requires a higher critical shear stress for detachment,
which was beyond the range of
the microfluidics system under
our operating conditions. The
critical shear stress value of 1.7
Pa is close to the range of previously reported values of shear
stress (5-12 Pa) required for
detachment of non-dental biofilms (Ohashi and Harada, 1994;
Stoodley et al., 2002).
The exit velocity of the microdrops from the prototype AirFloss was 60 m/sec, and, based
on earlier experiments, the flow
was a steady stream (Rmaile
et al., 2013). Even though the
shearing force was applied over
very short periods of 30 msec,
the generated fluid τw proved to
be effective in removing the attached biofilm by both adhesive
and cohesive failure (Rmaile et
al., 2013). However, fractions
of the biofilm remained on the
back of the teeth, due to tooth
architecture and the fluid flow
behavior in these regions, i.e.,
the inability of the fluid to flow
around the anatomical curvature and undercuts associated
with the palatal surface of the
upper central incisors. These
observations were predicted by
the computational simulations
in which τw on the proximal surface of the teeth was observed to
decrease gradually in the labiopalatal direction.
The simulations predicted τw
distribution on the tooth surface
caused by the microburst to be
in the kPa range within the IP
space, except in areas on the
palatal side of the tooth, where
τw became significantly lower
(~200 Pa). The maximum computational values for the fluid τw
were ~1,000 times higher than
the CDSSagg obtained from the
flow-cell experiments, and ~200
times higher than the estimated
shear stress, reported in the
literature, for biofilm detachment (Ohashi and Harada, 1994;
Stoodley et al., 2002). Thus, the
simulations predict that a significant percentage area of the
tooth is subjected to τw values
capable of removing the plaque
from the IP spaces. The large
difference in adhesive strength
between the 2 systems illustrates
the importance of the physical growth conditions and surface type on adhesion strength.
It was beyond the scope of this
study to determine the influence
of surface or hydrodynamics on
adhesion strength. In mechanical testing, properties reports
for the same species commonly
vary by 3 orders of magnitude
or greater (Shaw et al., 2004).
Whether this variability is true at
different locations in the mouth
or between patients is unknown,
but measurements of the adhesion strength of real oral biofilm
plaques would be useful in developing relevant in vitro models which look at mechanically
induced detachment.
The 3D simulations for predicting τw were consistent with the
experimental results obtained.
As might be expected, the biofilm survived the burst at areas
of low τw, but was flushed away
at areas where τw was higher.
A linear relationship was found
between the predicted fluid τw
and the amount of detached
biofilm obtained experimentally
(Eq. 1). This relationship could
be used to predict the efficacy of
oral health care devices that use
shear forces to remove plaque.
The computational model developed allowed for prediction
of the effect of changing the
position of the nozzle tip in the
z-direction
(inciso-gingivally)
on biofilm removal efficacy.
The numerical simulations predicted that placing the nozzle tip
in or close to the middle of the
inciso-gingival height (z/H = 0.5
or 0.67) provides more effective
biofilm removal, in comparison
with placing the tip closer to either the incisal edge or the gum
line (Fig. 4). To the best of our
knowledge, this is the first time
that CFD has been used to calculate the wall shear stress distribution, caused by water drops
generated from an oral hygiene
device, on the tooth surface.
In this study, an experimental
set-up was built and a methodology was developed to characterize, visualize, and quantify the
efficacy of biofilm detachment
by high-velocity water droplets,
which prevents the accumulation of biofilm and automatically
translates into prevention of
dental caries formation at these
sites.
Acknowledgments
The use of both the IRIDIS HighPerformance Computing Facility, and μ-VIS (CT centre), and
associated support services at
the University of Southampton is
sincerely acknowledged.
The authors also acknowledge
Dr. Phil Preshaw from Newcastle University for helping with
the development of the typodont model, Dr. Suraj Patel from
labtech for helping with the BioFlux experiments, Dr. Philipp
Thurner from the University of
Southhampton for advice with
μ-CT, and the late Dr. Hansjürgen Schuppe for helping with
the CLSM images. This work
was financially supported by
Philips Oral Healthcare, Bothell, WA, USA. M. Aspiras and M.
Ward are employed by Philips
Oral Healthcare, Bothell, WA,
USA. The other authors declare
no potential conflicts of interest
with respect to the authorship
and/or publication of this article.
About the Authors
A. Rmaile1*, D. Carugo2, L. Capretto2, M. Aspiras3, M. De Jager4, M.
Ward3, and P. Stoodley1,5
nCATS, Faculty of Engineering and the Environment (FEE),
University of Southampton, UK;
2
Bioengineering Group, Faculty of
Engineering and the Environment
(FEE), University of Southampton,
UK; 3Philips Oral Healthcare Inc.
(POH), Bothell, WA, USA; 4Philips
Oral Healthcare, Philips Research,
Eindhoven, The Netherlands; and
5
Center for Microbial Interface
Biology, Departments of Microbial Infection and Immunity, and
Orthopaedics, The Ohio State University, Columbus, OH, USA; *corresponding author, ar1a09@soton.
ac.uk
1
J Dent Res 93(1):68-73, 2014
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