The aim of this study was to investigate the effects of casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) application after interproximal stripping on enamel surface structures in vivo.
Fifteen patients with a mean age of 15.8 years participated in this study. For each patient, the extraction of 4 first premolars was part of the orthodontic treatment plan. The patients were randomly divided into 5 groups of 3 patients. With the exception of group 1, the mesial and distal surfaces of all first premolars were stripped with a stripping disc (Komet; Gebr Brasseler, Lemgo, Germany) under air cooling and then polished with Sof-Lex polishing discs (3M Dental Products, St Paul, Minn). In group 1, no stripping was performed, and the teeth were removed immediately. In group 2, the teeth were removed immediately after the stripping. In group 3, the stripped teeth were extracted after exposure to oral conditions for 3 months. In groups 4 and 5, CPP-ACP (Recaldent Tooth Mousse; GC Europe, Leuven, Belgium) or fluoride varnish (Bifluoride 12; Voco, Cuxhaven, Germany) was applied to the stripped surfaces for 3 months, respectively, before the teeth were extracted. Surface roughness and microhardness values were evaluated with 1-way analysis of variance and Tukey HSD tests.
The CPP-ACP and the fluoride varnish applications increased the surface roughness and microhardness values that had been decreased by stripping. No statistically significant differences were found between groups 3, 4, and 5 for microhardness or between groups 4 and 5 for surface roughness ( P > 0.5).
The saliva and saliva plus remineralizing agents (fluoride varnish and CPP-ACP) increased the microhardness and surface roughness values of stripped enamel surfaces that had been decreased by stripping.
Stripped enamel surfaces after remineralizing agent application were examined in vivo.
Interproximal enamel stripping decreased the microhardness of enamel surfaces.
Saliva, fluoride varnish, and CPP-ACP increased the microhardness of stripped enamel.
Smoother enamel surfaces were produced than intact enamel with subsequent polishing.
Exposure to saliva + remineralizing agents increased surface roughness.
Interproximal enamel stripping is defined as clinically removing part of the dental enamel from an interproximal contact area by grinding. This approach has been applied in orthodontics for many years to obtain more space to align crowded teeth or to correct a Bolton tooth-size discrepancy. By means of enamel reduction, approximal contacts can also be reshaped to correct morphologic anomalies and to camouflage interdental gingival papilla retraction. In clinical orthodontics, grinding of dental enamel can be achieved with handheld or motor-driven abrasive strips but also with discs or burs mounted on a hand piece.
Several studies have investigated the detrimental effects of enamel stripping due to loss of the protective superficial enamel layer by interdental stripping. Some studies have claimed that iatrogenic injuries to the integrity of the proximal enamel surface by stripping can be predisposing factors for caries and periodontal disease. Qualitative scanning electron microscope (SEM) evaluations have shown that all stripping methods affect enamel surface morphology, leaving furrows and scratches. These surface irregularities might facilitate bacteria adherence and plaque accumulation.
On the other hand, contradictory results also have been reported in previous studies evaluating the relationship between the stripping and caries and periodontal problems. Long-term results of interdental stripping showed no iatrogenic damage: eg, dental caries, gingival problems, or increased alveolar bone loss. It is still controversial whether a significant clinical relationship exists between stripping procedures and increased susceptibility to caries or periodontal disease. However, the use of polishing discs and some agents (fluoride products and sealants) has been recommended to prevent the undesirable side effects of interdental stripping by producing smoother enamel surfaces and enhancing remineralization.
During the last decade, bioactive agents based on milk products have been developed that enhance the remineralization of enamel and dentin by means of releasing active ions under cariogenic conditions. This agent is based on a nano-complex of the milk protein casein-phosphopeptide (CPP) with amorphous calcium phosphate (ACP). CPP-ACP serves as a reservoir source for calcium and phosphate ions on the tooth surface, thus helping to depress demineralization, enhance remineralization, and increase the microhardness of softened enamel.
Although many studies are focusing on the use of CPP-ACP for white spot lesion prevention and caries prophylaxis before bracket bonding procedures, only 2 studies were identified that evaluated the effects of CPP-ACP paste on stripped enamel in orthodontic practice. Giulio et al evaluated in vitro the effect of CPP-ACP on enamel surfaces after interdental stripping and reported that CPP-ACP is effective in promoting enamel remineralization. However, it is a well-recognized fact that in-vitro studies cannot exactly simulate clinical situations for reasons such as differences in the mineral content of enamel, plaque formation, oral hygiene, and diet of patients. Also, saliva is an important factor for the remineralization of enamel. In a recent in-vivo study, the changes of morphology and composition of stripped enamel surfaces after exposure to saliva and CPP-ACP with sodium fluoride were investigated by Paganelli et al. They concluded that the effects of saliva and CPP-ACP with sodium fluoride on stripped enamel in vivo showed no difference after 30 days.
The aims of our in-vivo study were to investigate and to compare the effects of a commercial paste based on CPP-ACP complex and a fluoride varnish application after interproximal stripping of enamel surfaces. The research hypotheses were that the remineralizing agent application (1) will not alter the roughness and (2) will not increase the microhardness of the stripped enamel surfaces.
Material and methods
A power analysis was performed by G*Power software (version 3.0.10; Franz Faul Universitat, Kiel, Germany). Based on a 1:1 ratio between groups, a sample size of 24 surfaces in each group would give more than 80% power to detect significant differences with a 0.40 effect size at a significance level of α = 0.05.
Fifteen patients (6 boys, 9 girls) with skeletal Class I malocclusion participated in the study. The average age was 15.8 years, with a range of 13.5 to 18.7 years. These patients were to receive routine orthodontic treatment with fixed appliances. For each patient, the extraction of 4 first premolars and the need for moderate anchorage mechanics were parts of the treatment plan. Before the study, information about the study design was given to the subjects, and informed consent was obtained from all adult patients and the parents of those under 18 years of age. This study protocol was approved at the Karadeniz Technical University by the ethical committee of Trabzon Clinical Researches.
Before this study, all patients received full-mouth clinical and radiographic caries assessments by an examiner (C.Y.). For this purpose, posterior bitewing radiographs were taken by using the long-cone technique. Selection criteria called for no cracks, hypoplasia, caries, fillings, or exposure to chemical agents (ie, bleaches) at the first premolars.
All patients were given oral hygiene instructions and monitored for 2 weeks. They were instructed by the orthodontist to brush their teeth for 3 minutes using the toothpaste (containing 1450 ppm of fluoride) supplied for daily use throughout the study. They were told not to use any other oral agents, including oral irrigators or antimicrobial mouth rinses.
The patients were randomly divided into 5 groups of 3 patients (12 premolars, 24 surfaces) according to the following procedures.
Group 1: No stripping was performed, and the first premolars were removed immediately.
Group 2: Approximal surfaces of the first premolars were stripped with a stripping disc (Komet; Gebr Brasseler, Lemgo, Germany) and then polished with extra-thin Sof-Lex discs (3M Dental Products, St Paul, Minn). The teeth were removed immediately after the stripping.
Group 3: Approximal surfaces of the first premolars were stripped with a stripping disc (Komet) and then polished with extra-thin Sof-Lex discs (3M Dental Products). The teeth were removed after exposure to oral conditions for 3 months.
Group 4: Approximal surfaces of the first premolars were stripped with a stripping disc (Komet) and then polished with extra-thin Sof-Lex discs (3M Dental Products). CPP-ACP (Recaldent Tooth Mousse; GC Europe, Leuven, Belgium) was applied to the stripped surfaces for 3 months. The teeth were then extracted for evaluation.
Group 5: Approximal surfaces of the first premolars were stripped with a stripping disc (Komet) and then polished with extra-thin Sof-Lex discs (3M Dental Products). Fluoride varnish (6% sodium fluoride and 6% calcium fluoride, Bifluoride 12; Voco, Cuxhaven, Germany) was applied to the stripped surfaces for 3 months. The teeth were then extracted for evaluation.
The interproximal enamel stripping was performed on both mesial and distal surfaces of the first premolars in the stripping groups. Before the stripping procedure, some precautions to prevent risk of damage to adjacent teeth were taken. In all subjects, elastic separators were placed on the mesial and distal contacts of the first premolars for 3 days. During the stripping and polishing, a metal separator (Hager & Werken, Duisburg, Germany) was used to improve access to the interproximal surfaces. Additionally, metal strips were applied to adjacent teeth to prevent any enamel damage.
The teeth were stripped with a stripping disc (Komet) under air cooling, followed by extra-thin medium, fine, and superfine Sof-Lex polishing discs (3M Dental Products); 10 and 20 strokes were made on the proximal surfaces of each tooth with the stripping disc and the Sof-Lex discs, respectively. One operator (M.B.) performed all stripping procedures. A new Sof-Lex disc per surface and a new stripping disc per patient were used (4 teeth, 8 surfaces).
All of the remineralization agent application protocol was performed by 1 clinician (A.K.) immediately after the stripping. In group 4, CCP-ACP application to the stripped enamel surfaces was demonstrated to the patients by the clinician at the first appointment. Patients were told to treat the stripped surfaces with CCP-ACP after brushing their teeth at home according to the instructions given by the examiner for 3 minutes once a day for 3 months. A handle and brush tips (3M Unitek, Monrovia, Calif) were given to the patients for this application. They were advised not to eat or drink for 30 minutes after application.
In group 5, after cotton roll isolation had been achieved, the stripped surfaces of the teeth were gently dried with air. In accordance with the manufacturer’s instructions, the fluoride varnish was applied to the stripped surfaces by using a handle and brush tip once a month for 3 months by the same clinician. The varnish was then air-dried for 10 to 20 seconds, and the treated surface was protected from salivary contamination for a further 20 to 25 seconds. The patients were told not to wash with water, not to eat or drink anything for 3 hours, not to brush their teeth for 24 hours, and not to use floss or a toothpick for a week.
In all subjects who participated in this study, the orthodontic treatment began after the extraction of the first premolars. After removal, all teeth (n = 60) were collected and stored in deionized water until needed.
After the root portion of the crown was removed, each tooth crown was sectioned into 2 halves in the buccolingual direction using a diamond saw under water lubrication (Isomed-Buhler, Lake Bluff, Ill). Thus, 120 enamel slices were obtained. The samples were then ultrasonically cleaned for 10 minutes in deionized water. To carry out the microhardness and surface roughness tests, the teeth were embedded in self-curing acrylic resin, leaving the proximal enamel surfaces uncovered. Surface roughness parameters were assessed with a profilometer (Surf Test SJ 201 P/M; Mitutoyo, Takatsu-ku, Japan). The mean arithmetic roughness was used to assess surface changes. For each specimen, 3 measurements were made with the contact stylus for each registration in the perpendicular direction. The mean value of these measurements on 1 specimen was used as the mean arithmetic roughness of that specimen.
Vickers hardness was tested with a Micro Hardness Tester (Duramin-3; Struers, Ballerup, Denmark) by applying a square-base pyramidal diamond indenter (Vickers pyramid with angles of 136°) on the stripped surface of the tooth sections under a load of 200 g for 10 seconds. The samples were stabilized parallel to the base of the hardness measurement device by pressing the sample over a thin layer of a plasticizing material (Plasticine; Beuhlers, Princeton, Ind). For each specimen, 3 indentations were made, and the average was calculated. The determined values were averaged to represent the Vickers hardness value of that specimen.
For the SEM evaluation, 5 specimens (1 sample from each group) were prepared to evaluate the enamel surfaces qualitatively. The photomicrographs were taken with a SEM (JSM-5600; Jeol, Tokyo, Japan) with 500-times magnification for visual inspection.
Data analyses were performed by using Statistical Package for the Social Sciences software (version 13.0; SPSS, Chicago, Ill). The Shapiro-Wilk normality test and the Levene variance homogeneity test were applied to the microhardness and surface roughness data. The data showed normal distributions, and there was homogeneity of variances between the groups. One-way analysis of variance (ANOVA) was performed to examine the effects of the remineralization agent application on the stripped enamel surface. The Tukey HSD test was used for post hoc comparisons of the groups. The results were evaluated with a 95% confidence interval. The significance level was set at P < 0.05.
The descriptive statistics, including means, standard deviations, minimum and maximum values, and the results of the Tukey HSD post hoc test for the surface roughness and microhardness data are shown in Tables I and II , respectively.