White-Spot Lesions in Orthodontics: Incidence and Prevention

The most common negative effect of orthodontic treatment with fixed appliances is the development of incipient carious lesions around brackets. The objectives of this chapter are to present some of the results of two studies aiming: 1) to evaluate patients treated with comprehensive orthodontics to determine the incidence of new carious lesions during treatment; and 2) to investigate the potential of ACP-containing resin cement and other treatments (fluoride varnish, resin sealer, MI Paste) to prevent incipient carious lesions on bracketed teeth. In the first study, 350 orthodontic patients were selected randomly. The preand post-treatment photographs of the patients were examined to determine lesion development. The labial surface of each tooth was scored with a standardized system based on the International Caries Determination and Assessment System II. The independent variables were collected by chart abstraction. In the second study, 100 extracted human premolars were allocated randomly to five groups (N = 20). Brackets were bonded with ACP-cement (Aegis-Ortho), Transbond


Introduction
The most common negative effect of orthodontic treatment with fixed appliances is the development of incipient carious lesions around brackets. The objectives of this chapter are to present some of the results of two studies aiming: 1) to evaluate patients treated with comprehensive orthodontics to determine the incidence of new carious lesions during treatment; and 2) to investigate the potential of ACP-containing resin cement and other treatments (fluoride varnish, resin sealer, MI Paste) to prevent incipient carious lesions on bracketed teeth. In the first study, 350 orthodontic patients were selected randomly. The preand post-treatment photographs of the patients were examined to determine lesion development. The labial surface of each tooth was scored with a standardized system based on the International Caries Determination and Assessment System II. The independent variables were collected by chart abstraction. In the second study, 100 extracted human premolars were allocated randomly to five groups (N = 20). Brackets were bonded with ACP-cement (Aegis-Ortho), Transbond XT (Control), Transbond XT followed by application of fluoride varnish (Vanish), resin sealer (Pro-seal) and CPP-ACP paste (MI Paste). All teeth were pH cycled for 15 days in demineralization solution and artificial saliva. The extent of demineralization in each group was assessed using Quantified Light-induced Fluorescence (QLF) and Confocal Laser Scanning Microscopy (CLSM). The incidence of patients who developed at least one new white-spot lesion during treatment was 73%. Treatment length was associated significantly with new white-spot lesion development. The independent variables of gender, age and extraction/non-extraction were not associated with lesion development. Fluorescence loss and lesion depth measurements demonstrated that the Pro-seal and Vanish groups had the least amount of demineralization. The control group showed the most demineralization. Although the MI Paste and Aegis-Ortho groups experienced less demineralization than controls, neither was significant statistically. Only the Pro-seal and Vanish groups had significantly smaller lesions than the control group for both QLF and CLSM. Thus, the development of new lesions appeared to be related to treatment duration and, to a lesser degree, to initial oral hygiene score. Light-cured filled sealer (Pro-seal) and the fluoride varnish (Vanish) have the potential to prevent enamel demineralization adjacent to orthodontic brackets exposed to cariogenic conditions.

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Contemporary Approach to Dental Caries 314

White-spot lesions
One of the most common negative side effects of orthodontic treatment with fixed appliances is the development of incipient caries lesions around brackets and bands, particularly in cases with poor oral hygiene (Fig. 1). Caries lesions typically form around the bracket interface, usually near the gingival margin (Gorelick et al., 1982). Certain bacterial groups such as mutans streoptococci and lactobacilli ferment sugars to create an acidic environment that over time might lead to the development of dental caries. Since orthodontic appliances make plaque removal more difficult, patients are more susceptible to carious lesions. The irregular surfaces of brackets, bands, wires, and other attachments also limit naturally occurring self-cleaning mechanisms, such as movement of the oral musculature and saliva (Rosenbloom and Tinanoff, 1991). Incipient lesions are characterized by their opacity, mineral loss, and decrease of fluorescence radiance when compared to healthy enamel surfaces. Many incipient enamel lesions have a white appearance due to an optical phenomenon caused by mineral loss in the surface and sub-surface that alters the refractive index and increases the scattering of light in the affected area, all resulting in greater visual enamel opacity.
Studies have shown that white spot lesions can take only one month to develop (Øgaard et al., 1988;O'Reilly and Featherstone, 1987;Gorton and Featherstone, 2003). A clinical study reported the prevalence at 50% (Gorelick et al., 1982), while recent investigations put the incidence of white spot lesions in the orthodontic populations studied at 73-95% (Richter et al., 2009;Lovrov et al, 2007). Orthodontists and patients will notice these lesions after removal of the fixed appliances, especially since the white spots tend to form in the maxillary esthetic zone (Gorelick et al., 1982;Banks and Richmond, 1994). While some studies have reported a decrease in the display of white spot lesions over time post-orthodontic treatment, these unesthetic spots tend to remain unless they are resolved with more aggressive treatment, such as minimally invasive or even full restorative dentistry (Øgaard, 1989;Årtun and Thylstrup, 1989).

Oral hygiene
The first line of defense against the development of incipient caries lesions has traditionally been patient education, with a special emphasis on optimal oral hygiene. The advocacy organization for orthodontists in the United States known as the American Association of Orthodontists (AAO) has developed patient manuals and a website to provide recommendations for patients undergoing orthodontic treatment (AAO, 2009). Specifically, the website suggests extra time for toothbrushing, specialized tips to get in between the braces, floss threaders, oral irrigators, and over-the-counter mouthrinses. Additionally, the AAO sponsored informed consent form emphasizes the need for excellent oral hygiene and routine visits to the general dentist (AAO, 2005). It also warns that inadequate oral hygiene could result in caries, discolored teeth, and periodontal disease. Finally, the form explains that the aforementioned problems may be aggravated if the patient has not had the benefit of fluoridated water. In many cases, patient education will also include an emphasis on proper diet with reduced intake of sugars. Despite these efforts by the orthodontist and staff members, many patients will still be non-compliant with oral hygiene instructions. Unfortunately, most orthodontists have a limited background in the behavioral basis of compliance (Mehra et al., 1998). Thus, patient non-compliance presents a unique challenge to orthodontic practices.

Fluoride during orthodontic treatment (rinses, etc)
In addition to reinforced oral hygiene instructions, orthodontists have turned to various products and preventive measures to reduce this problem. Dental professionals have employed fluoride for years to prevent caries and remineralize enamel in patients. A systematic review found a reduced level of caries and adolescents who have regular supervised rinsing with a fluoride mouthwash (Marinho, 2004). Daily fluoride rinses have shown promising results, and a significant reduction in enamel lesions can be achieved during orthodontic therapy through the daily use of a 10 mL neutral 0.05% sodium fluoride rinse. However, typical patient compliance rates with this protocol have been relatively low (Geiger et al., 1992).

Fluoride varnish
Preventive measures that do not require patient compliance would seem to make more sense for the typical orthodontic patient population of adolescents. For some patients, professional fluoride varnish application by orthodontic auxiliaries at routine appointments can in part address this compliance issue (Vivaldi-Rodrigues et al., 2006). On the other hand, each application requires over five minutes of chair-time, and whether or not today's high efficiency/high volume orthodontic practice will devote the time and resources to apply this protocol is debatable. Generally however, fluoride varnishes have a proven track record in caries reduction when applied properly. Vanish (3M/Omni) is a very popular 5% NaF white varnish used for prevention of dental caries. The manufacturer advertises the ease of use, lack of an unesthetic yellow color found in other varnishes, enhanced flow characteristics, and its fluoride delivery of 22,600 ppm. Its name comes from an alleged ability to disappear after application. Data gathered by the manufacturer declare greater fluoride release over a 48-hour period in comparison to other fluoride products. To date, Vanish in particular has not been tested in any of the in-vitro or in-vivo trials in the literature.

Resin sealer
Just as sealants have been shown to prevent caries in molars with deep fissures, resin-based sealers have been applied on facial surfaces of bracketed teeth to prevent enamel caries. In addition to the increased chair-time for this procedure, earlier generations of resin sealers have been found to have very low wear resistance. Previous studies have proven that most of the chemically cured sealants (Zachrisson et al., 1979) do not effectively seal smooth enamel surfaces, because of oxygen inhibition of polymerization when the sealant is in contact with the air in a thin layer. Instead, only "islands" of cured sealant remain where resin pooling occurs. Even light-cured sealants (Banks and Richmond, 1994) that were unfilled or lightly filled could not provide any more protection than the chemically cured sealants. A more recent developed product Pro-seal (Reliance, Itasca, IL) has been marketed as a sealer that is more resistant to toothbrush abrasion than earlier generations, since it is a highly filled resin. In patients with poor oral hygiene, Pro-seal can be added before bracket bonding or after bonding. Additionally, the manufacturer claims that Pro-seal releases fluoride, which further enhances its anticariogenic properties.

ACP/CPP-ACP
Recently, there has been increased interest and development in calcium phosphate-based remineralization technology (Reynolds and del Rio, 1984;Rosen et al., 1984). One of the newest modalities in preventive dentistry is the introduction of amorphous calcium phosphate (ACP) into methacrylate composites, gum, pastes, and other dental products. Casein is the predominant phosphoprotein in bovine milk and accounts for almost 80 percent of its total protein, primarily as calcium phosphate stabilized micellular complexes (Aimutis, 2004). Several laboratory and animal experiments have investigated the low cariogenic potential and the possible cario-static activity of dairy products (milk, casein, caseinates and cheeses). The use of casein as an anticariogenic additive to food, toothpaste or drinking water has not been implemented because of its adverse organoleptic properties and the large amount required for efficacy (Reynolds, 1998).
Casein phosphopeptide (CPP) contains the cluster sequence of -Ser (P)-Ser (P)-Ser (P)-Glu-Glu from casein (Iijima et al., 2004). CPP does not have the limitations of casein, has the potential for specific anticariogenic activity, and is at least 10 times greater on a weight basis than it is for casein (so not as much is needed for it to be effective). CPP can remarkably stabilize calcium phosphate (which usually is highly insoluble) in a state-forming CPPamorphous calcium phosphate (ACP) complex. There is no conclusive evidence that ACP is an integral mineral component in hard tissues. Its advocates theorize that it likely plays a special role as a precursor to bioapatite and as a transient phase in biomineralization. In solutions, ACP is converted readily to stable crystalline phases such as octacalcium phosphate or apatitic products (Mathew and Takagi, 2001). Reynolds and colleagues have proposed that under acidic conditions, localized CPP-ACP buffers the free calcium and phosphate ions, substantially increasing the level of calcium phosphate in plaque and, therefore, maintaining a state of supersaturation that inhibits enamel demineralization and enhances remineralization (Reynolds et al., 1999). Rose conducted a laboratory experiment in which he showed that CPP-ACP binds well to dental plaque, providing a large calcium reservoir that may inhibit demineralization and assist in subsequent remineralization (Rose, 2000). This technology has entered the orthodontic marketplace in two different forms: resin bracket bonding cement containing ACP and topical paste containing the CPP-ACP complex. Aegis-Ortho, an ACP-including resin bonding cement, has been marketed by Bosworth (Skokie, IL) as a substitute for ordinary bracket bonding cement, with the added benefit of caries prevention. The manufacturer claims that the acidic challenge (pH at or below 5.8) to the surrounding bracket area will trigger the release of calcium and phosphate from the cement, and a supersaturated calcium phosphate matrix will not only inhibit demineralization, but also remineralize the enamel. ACP-filled composite resins have been shown to recover 71% of the lost mineral content of demineralized teeth (Skrtic et al., 1996).
A similar chemical process is manifested with MI paste (GC America, Alsip, IL). Instead of residing in the resin cement, the casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) is applied topically in the mouth to affected areas. The manufacturer implicates this product not only lesion prevention (applied twice daily after brushing throughout orthodontic treatment), but also claims the patient can expect the complete reversal of such lesions after three months use post-debonding. Additionally, the manufacturer has recommended MI paste for dental patients with xerostomia, dental sensitivity, gastric reflux, fluorosis, exposed root surfaces, and as an adjunct to tooth bleaching.

Fluoride varnish
A myriad of in-vivo and in-vitro studies have been carried out to study the efficacy of preventive measures against white spot lesion formation during orthodontic treatment. Fluoride varnish has by far the strongest evidence base. The potential of fluoride varnish has been evaluated in-vitro (Adriens et al., 1990;van der Linden and Dermaut, 1998;Todd et al., 1999;Demito et al., 2004) as well as in-vivo (Vivaldi-Rodrigues et al., 2006;Øgaard et al., 2001). Generally, investigations carried out in-vitro indicate a moderate to strong beneficial effect of the tested varnishes on enamel demineralization. Two in-vivo studies have emerged. In a split-mouth prospective study, there was 44.3% less demineralization noted for teeth that had been treated every 12 weeks with fluoride varnish during orthodontic treatment (Vivaldi-Rodrigues et al., 2006). In a double-blinded randomized placebo-controlled trial, Stecksén-Blicks et al. reported that although fluoride varnish did not totally prevent white spot lesion formation, the incidence was significantly reduced in the fluoride varnish group. In addition to differences in study design, the frequency of fluoride application also varied among the studies. Stecksén et al. applied the fluoride varnish at six week intervals, the typical appointment interval for most orthodontic patients.

Resin sealer
After less successful earlier sealers, findings about the application of a filled-resin sealer (Pro-seal) have been published in the literature. One in-vitro study using an acid challenge found that demineralization was significantly less with Pro-seal treatment, compared to an untreated enamel surface (Hu and Featherstone, 2005). In fact, the demineralization levels established by microhardness profiles showed that the Pro-seal group had 98% less demineralization than the control group. This study also featured a group of teeth treated with fluoride varnish. While both the Pro-seal and fluoride varnish had significantly less demineralization than the control group, the sealer had significantly less demineralization than the varnish. Furthermore, the study also found that Pro-seal can stand up to acid challenge and toothbrush abrasion in a laboratory environment. These outcomes were corroborated by another in-vitro study, that also found that the filled-resin sealer (Pro-seal) provided significantly more protection than either fluoride varnish or an unfilled resin sealer, with a 92% reduction in lesion depth compared with the controls using polarized light microscopy (Buren et al., 2008). In looking at its supposed fluoride release, one study found that Pro-seal released fluoride ions in a sustained way -with significantly decreasing amounts over a 17-week period, though this release was measured to be sub-ppm (Soliman et al., 2006). Despite some favorable results with in-vitro models, no in-vivo trials with Proseal have been published in the literature.

CPP-ACP paste
Due to the early stages of this technology, published independent research on the ACP products like MI Paste is limited. Generally, the studies on caries prevention with CPP-ACP consist of in-situ caries models with gums, mouthrinses, or lozenges (Iijima et al., 2004;Reynolds et al., 2003). In addition, the vast majority of these studies were carried out by the same group that first isolated CPP-ACP at the University of Melbourne, Australia. For example, using topical applications of CPP-ACP via sugar-free chewing gum and mouthrinse, Reynolds et al. showed that CPP-ACP incorporated into dental plaque can significantly increase the levels of plaque calcium and phosphate ions (Reynolds et al., 2003). Conversely, an in-vitro study carried out by an American group found that while fluoride 5000 ppm paste had a statistically significant protective effect against demineralization on enamel sections, MI Paste had no effect (Pulido et al., 2008).There are two published studies that examine the role of CPP-ACP paste in orthodontics. In an in-vitro study that assessed the demineralization around bonded molar tubes on extracted third molars, a mild decrease in demineralization was found with the application of CPP-ACP (Sudjalim et al., 2007). On the other hand, the authors of this very article recommended combining CPP-ACP with a fluoride gel to enhance the treatment effect. For the most part, clinicians loyal to the CPP-ACP protocol apply it without a fluoride gel, and the brochures by the manufacturer make no mention of additional rinses or gels. Andersson et al. conducted an in-vivo postorthodontic treatment study, in which they compared the remineralization capabilities of 0.05% Sodium Fluoride mouthwash and the application of Topacal (CPP-ACP topical cream) on patients with white spot lesions. The study found significant remineralization with both protocols, and found no significant differences between the groups over time.
Still, the authors pointed out that the remineralization that occurred with CPP-ACP treatment was generally more esthetic than with the fluoride rinse.

ACP-containing bonding cement
As for ACP bonding cement products like Aegis-Ortho, there are currently no published comparative studies on its ability to prevent white spot lesions in the peer-reviewed literature. Two reports detail the questionable bond strength of ACP-containing cement. In spite of its potential benefits, frequent bond failures with ACP-cement have been reported. An in-vitro study with an earlier generation of Aegis-Ortho showed that orthodontic brackets bonded to teeth with an ACP-containing composite material failed at significantly lower forces than brackets bonded to teeth with a conventional resin-based composite orthodontic cement (Dunn, 2007). Another in-vitro study found that brackets bonded with the conventional Transbond XT had more than two times the shear bond strength in comparison to brackets bonded with Aegis-Ortho cement (Foster et al., 2008). In evaluating the current evidence base for ACP and its various products, the number of published in-vitro and in-vivo trials is clearly underwhelming. In a 2008 systematic literature review published in the Journal of the American Dental Association (JADA), the authors concluded that there is insufficient clinical trial evidence to make a recommendation regarding the long-term effectiveness of casein derivatives, specifically CPP-ACP, in preventing caries in-vivo (Azarpazhooh and Limeback, 2008).

Summary of evidence
Of all the treatments for incipient caries lesions during orthodontic treatment, agents with fluoride including varnish have the highest level of evidence. Multiple laboratory and clinical studies have demonstrated its efficacy. Highly-filled resin sealers like Pro-seal are relatively new, although the results from a few in vitro studies have demonstrated impressive results. On the other hand, a clinical study on its demineralization prevention has not yet surfaced. At this juncture, the evidence level for ACP products like CPP-ACP paste or ACP resin bonding cement is low. There is a clear need for more independent research of casein derivatives like CPP-ACP to make conclusions about its efficacy in caries prevention.

Clinical status quo
In terms of the clinical status quo for prevention of incipient caries lesions during orthodontic treatment, one has to first reference the AAO sponsored informed consent form, in which there is an emphasis on excellent oral hygiene, regular visits to the general dentist, and access to fluoridated water (AAO, 2009). In looking at practice trends, a recent survey by the Journal of Clinical Orthodontics does provide some information about the usage of some of the preventive measures previously outlined (Keim et al., 2008). Despite its proven efficacy, only 9.3% of orthodontists deliver fluoride varnish to their patients. The article also mentions that only 7.4% of orthodontists employ the fluoride-releasing glass ionomer adhesive for bracket bonding, which is understandable given its questionable physical properties. There were no data in the article detailing the usage of ACP products or resin sealers. In any event, the fact that more than half of orthodontic patients develop incipient caries lesions (Gorelick et al., 1982;Richter et al., 2009) and that only 9.3% of orthodontists give their patients fluoride varnish (Keim et al., 2008), raises questions about emphasis of preventive care in today's orthodontic practice.

Need for investigation
Due to high caries incidence, low patient compliance, and low usage of fluoride varnish by orthodontists, there appears to be a need for a better treatment modality for patients undergoing orthodontic treatment. Regardless of the exact prevalence rate for white spot lesion development, most dental professionals would agree that it is currently far too high. While adjuncts to treatment such as fluoride rinse can potentially reduce the incidence of white spot lesions, the required compliance of high caries-risk patients is dubious. Equally troublesome, available non-compliant and proven treatments like professionally applied fluoride varnish have failed to catch the attention of practicing orthodontists. The resin sealer (Pro-seal) seems to address the patient compliance issue, but if orthodontists have neither the time nor the interest to deliver fluoride varnish, their likelihood of investing the resources and chair time to etch and light cure Pro-seal on twenty teeth is probably low. Aegis-Ortho bracket cement containing ACP seems to address all these issues. It does not require patient compliance, and it does not require any additional chair time in the office, since time allotted for orthodontic bracket bonding is already a part of the treatment plan. In spite of some reports, which document a low bond strength of its earlier generations, if ameliorated, this product holds immense potential for preventive care during orthodontic treatment. First however, there is a need to test and document the preventive properties of ACP-containing bracket cement with in-vitro studies.

Methods to assess demineralization
With a heightened interest in evidenced-based dentistry, the dental research community has over the years employed various modes of technology to quantify extent of enamel demineralization. The ideal method of assessment should be simple, noninvasive, reproducible, and precise. The following is brief description of four commonly employed techniques (TMR, PLM, QLF, and CLSM).

Microradiography
Transverse Microradiography (TMR) or contact-microradiography is one of the most widely accepted methods used to assess demineralization and remineralization in dental hard tissues in in-situ and in-vitro studies. It is a highly sensitive method to measure the morphology of and the change in mineral content of enamel and dentin samples (Arends and Ten Bosch, 1992). In TMR the tooth sample to be investigated is cut into thin slices (about 80 µm and 200 µm for dentine samples). A microradiographic image is made on high resolution film by X-ray exposure of the sections together with a calibration stepwedge. The microradiogram is digitized by a video camera or photomultiplier. The mineral can be automatically calculated from the gray levels of the images of section and stepwedge using a custom-made software. In examining the reliability of TMR, Exterkate et al. found that repeated microradiographs of the same thin enamel sections resulted in a negligible spread in mineral loss among them (Exterkate et al., 1993). Such reliability and the more recent application of computer imaging make microradiography a standard method used in caries research for the assessment of lesion profiles.

Polarized light microscopy
Polarized light evaluations of enamel sections have been useful in describing the early caries lesion and alterations in structure upon further demineralization or remineralization. Generally, it provides information on absorption color and boundaries between minerals of differing refraction indices. Materials such as enamel act as beam splitters and divide light rays into two parts. Polarized Light microscopy (PLM) in turn exploits the interference of split light rays, as they are reunited along the same optical path to extract information about materials. Essentially, polarized light microscopy allows the visualization of areas with different porosities. The histologic features seen under a polarized light microscope allow the examiner to distinguish carious and non-carious enamel by their respective distribution of pores (Gwinnett, 1966). Polarized light examination of enamel specimens is a wellestablished procedure in which it is customary to view quinoline-imbibed sections orientated so that normal enamel is blue/green in color (Gilmour and Edmunds, 1998).

Quantitative light-induced fluorescence
Quantitative Light-induced Fluorescence (QLF) is one method of assessing levels of enamel demineralization. With QLF, real-time fluorescent images are captured into a computer and stored in an image database. Optional quantitative analysis tools enable the user to quantify parameters like mineral loss, lesion depth, lesion size, stain size and severity with high precision and repeatability. The QLF method is based on the auto-fluorescence of teeth. When teeth are illuminated with high intensity blue light they will start to emit light in the green part of the spectrum. When enamel demineralization takes place, minerals are replaced mainly by water from saliva, causing a decrease in the light path in the tooth substance. This results in less light absorption by enamel. Because fluorescence is a result of light absorption, the intensity of fluorescence decreases in demineralized regions of the enamel, which appear darker than sound tooth structures (de Josselin et al., 1995;al-Khateeb et al., 1998;Rousseau et al., 2002). Thus, the fluorescence of the dental tissue has a direct relation with the mineral content of the enamel. The effectiveness of QLF for measurement of enamel demineralization has been demonstrated in several studies. The use of QLF allows for quantitative analysis has been reported to be well correlated (0.73-0.83) with the degree of mineral loss from early enamel lesions in-vitro when measured by longitudinal microradiography. (Hafstrom-Bjorkman et al., 1992;Emami et al., 1996;Lagerweij et al., 1996). The use of QLF as a method of following caries development during orthodontic treatment has been suggested and encouraged by the results of several in-vitro studies. (Benson et al., 2003 andPretty et al., 2003). Recent studies also indicate that QLF is suitable for in-vivo monitoring of mineral changes in incipient enamel lesions (Van der Veen et al., 2000 andAl Khateeb et al., 2002).

Confocal laser scanning ,icroscopy
Confocal Laser Scanning Microscopy (CLSM) is yet another method of assessing enamel demineralization. This technique accelerates and simplifies the measuring of mineral loss. The enamel specimens are sectioned in half, stained with fluorescent dye, and analyzed www.intechopen.com Contemporary Approach to Dental Caries 322 using a CLSM system (Fontana et al., 1996). The major advantage of this method is that it enables quantitative analysis of thick samples without the problems of thin section preparation required for microradiography or polarized light microscopy. Essentially, CLSM allows a subsurface examination since the scattered, reflected, and fluorescent light from planes out of focus is eliminated -providing a subsurface image only from a thin layer upon which it is focused. This processed digital image can be used to determine surface features, area and volume analysis of given structures, and views of the total structure from any angle in three dimensions. In terms of efficacy, a statistically significant high correlation was found between mineral changes measured using microradiography and the changes in lesion parameters analyzed by confocal microscopy (González-Cabezas et al., 1998) With all the treatment modalities flooding the marketplace, the orthodontist might find it difficult to sort out what works best and why when oral hygiene deteriorates. The objectives of this chapter are to highlight the results of two recent studies that investigated: 1. The incidence of new WSLs before and after orthodontic treatment using photographic records; and 2. The potential of ACP-containing resin cement and other treatments (fluoride varnish, resin sealer, MI Paste) to prevent incipient caries lesions next to bracketed teeth.

Selection of subjects
From a population of 2,296 patients treated in the graduate orthodontic clinic at the University of Michigan School of Dentistry (UMSD) between 1997 and 2004, 350 patient records were selected randomly using a random number sequence. Inclusion criteria for record selection consisted of patients who: 1. Underwent comprehensive orthodontic treatment utilizing full fixed appliances on labial tooth surfaces; 2. Had complete initial and final series of intraoral photographs; and 3. Had complete treatment log information within their chart.

Chart abstraction
Data collection from de-identified patient charts included gender and age at initiation of orthodontic treatment, and treatment variables such as extraction therapy and comprehensive treatment time. Comprehensive treatment time was defined as the period between initiation of full fixed appliance therapy and removal of all active fixed appliances. Initial oral hygiene score, frequency of oral hygiene discussion, oral hygiene instruction and fluoride application and/or rinse were recorded from progress notes in the chart.

Photography
Intraoral pre-treatment (initial) and post-treatment (final) photographs of each patient were taken as part of standard orthodontic recordkeeping procedures. All photographs, stored as 35 mm slides, were taken in the Clinical Photography Department at the UMSD by two professional photographers utilizing a standardized intraoral photography procedure.
Individual slides were scanned into digital format using a Nikon Slide Feeder SF-200 (S) and Super Coolscan 4000 ED scanner. Scanned images were enlarged 325% and imported into an individual Microsoft PowerPoint presentation for each patient.

Dental caries determination
Images were evaluated by trained investigators using a scoring system specifically adapted for use with photographed images (International Caries Detection and Assessment System II; Ismail, 2005). Visible labial surfaces examined included maxillary and mandibular central and lateral incisors, canines, first and second premolars, and first molars. The evaluators scored each visible labial tooth surface before and after orthodontic treatment. The scores were combined to determine the labial caries incidence for each patient. Teeth were examined and scored from first molar to first molar, maxilla and mandible (Fig. 2).

Results: Part I
The overall incidence of patients who developed at least one WSL during orthodontic treatment was 72.9% (N = 255; Table 1 and Fig. 3), while for newly developed cavitated lesions that were unrestored on the final record was 2.3%. Of the eight patients that developed cavitated lesions during orthodontic treatment, four (1.1%) developed one new cavitated lesion, three (0.9%) developed two new cavitated lesions and one (0.3%) developed four new cavitated lesions. Of the maximum 24 surfaces investigated per patient, on average 4.2 surfaces in each patient showed new WSL. The average of surfaces with new cavitations was only 0.04 and 0.05 with restorations. Even though infrequently, some early WSL regressed to sound (0.07 per patient). Demographic variables of gender and age at initiation of treatment were not related significantly to development of new decalcified or cavitated lesions. There was a significant relationship between increased treatment length and number of newly developed lesions (P = 0.03; Table 2). The mean number of labial surfaces per patient that developed new WSL was 3.01 for patients with a treatment length of less than 22 months. This increased to 5.28 teeth for patients with therapy longer than 33 months. The number of new cavitations, however, showed only a nonsignificant trend (P = 0.08) with increased treatment time. In addition, the number of newly developed lesions (both WSL and cavitations) showed no significant association with extraction or nonextraction treatment protocols (Table 3). Although no relationship was demonstrated between pretreatment oral hygiene scores and lesion development, the recorded number of oral hygiene discussions between provider and patient were associated significantly with development of both white-spot (P <0.0001) and cavitated (P = 0.0006) lesions. The mean number of new lesions for patients with whom oral hygiene discussions had never been noted in the chart was 3.08, while the mean number of decalcified lesions for patients who were given oral hygiene instruction on three or more occasions increased to 7.78. A similar increase was exhibited for the mean number of cavitated lesions for patients given three or more oral hygiene discussions (mean = 0.20) vs. those with whom oral hygiene was not discussed after initial instruction (mean = 0.01). Age group (P = 0.03), treatment length (P = 0.01) and number of oral hygiene discussions (P < 0.0001) were associated with development of WSL. There was a decrease in WSLs associated with increasing age group (regression coefficient = -0.59). An increase in WSLs was associated with both increased treatment time (regression coefficient = 0.07) and increased number of oral hygiene discussions (regression coefficient = 1.88).

Sample preparation
One hundred human premolar teeth were collected from various oral surgery practices located in southeast Michigan. Only premolars presenting a healthy facial enamel surface were included. All teeth were assigned randomly to five equal groups of 20 teeth. One of the groups had brackets bonded with Aegis-Ortho resin cement while the remaining groups were bonded with Transbond XT. Of the four Transbond XT groups, one served as a control, another received Vanish (3M, Espe, MN) fluoride varnish, another received MI Paste and the final received a coat of Pro-seal as adjunctive treatments.

Demineralization protocol
Teeth were exposed to a pH cycling system to develop caries-like lesions. Each day teeth were incubated in demineralization solution (lactic acid and Carbopol [pH = 5.0], 50% saturated with hydroxyapatite) for eight hours, rinsed with de-ionized water and placed in artificial saliva for 30 minutes, followed by two seconds of brushing with a powerbrush (Sonicare, Philips) and fluoridated dentifrice (NaF, 1,100 ppm F), rinsed again and placed back in artificial saliva until next demineralization period (next day).

Results: Part II
Demineralization assessed by QLF is shown in Table 4. The Proseal group had the least amount of fluorescence loss followed by the Vanish group. Aegis-Ortho group, MI Paste group and the control group (Transbond) had the most fluorescence loss and were not different significantly. Demineralization assessed by CLSM is shown in Table 5. No detectable lesion depth was seen in any of the specimens of Pro-seal and Vanish groups. The greatest lesion depth was found in the control group (Transbond), but it was not different significantly from Aegis-Ortho and MI Paste.

Discussion
The use of intraoral photographs for caries determination in orthodontic patients is a wellaccepted method. Standardized photographs taken before and after appliance placement are available readily as a standard procedure in orthodontic care. Color photography as a means of recording prevalence of enamel opacity is a powerful method (Ellwood, 1993). Studies have shown that assessment of enamel demineralization from color images appears to be more reproducible than direct clinical observation utilizing only the naked eye (Benson et al., 1998). Moreover, photographic records provide an efficient means to capture the appearance of enamel and provide a permanent record at a given time point. It allows an examiner, therefore, to assess the caries experience of a patient blindly and randomly. Based on pre-and post-orthodontic treatment photographic patient records, this study showed a high incidence of new WSLs (72.9%) in patients treated with comprehensive orthodontics, while the incidence of new cavitated lesions in this population was 2.3%. Gender, age and oral hygiene at start of treatment were not associated with lesion development, while a significant association was evidenced with treatment duration. Patients in treatment for less than 22 months developed on average three WSLs, while patients in treatment for 33 months or longer developed on average more than five lesions. Linear regression analysis suggested that as the duration of fixed appliances increased one month, 0.08 new WSLs were developed. The in vitro study sought to test four different treatments, which comprise much of the currently available therapies to prevent WSLs. The four experimental groups differed in their application, chemistry and physical properties. The Aegis-Ortho cement serves as a replacement for a typical bracket bonding cement. This ACP-containing material supposedly reduces the incidence of enamel demineralization with the release of calcium and phosphate ions -not only to reduce demineralization, but also to promote the remineralization of enamel. The fluoride varnish group received the same bonding cement as the control plus an application of Vanish, a popular fluoride varnish used for caries prevention. Unlike fluoride rinses that require patient compliance, the delivery of Vanish takes place in the dental chair and could be applied at the monthly orthodontic appointment. The CPP-ACP group teeth received an adjunctive daily application of MI Paste, whose chemical mechanism of action resembles that of the ACP cement. Instead of having ACP just residing in the bracket cement, the preventive protocol for MI Paste demands a daily application and, thus, a certain degree of patient compliance. MI Paste is claimed to have the ability to prevent WSLs during orthodontic treatment. Teeth in the final group received a light cured filled sealant as adjunctive treatment. Though it claims to offer some fluoride release, Pro-seal at its core functions as a protective physical barrier against the acid attacks.
Compared with the control group, the Pro-seal group had a statistically significant difference in regard to both outcome measures (i.e., lesion depth and fluorescence loss). The CLSM results indicated that there was no demineralization on any of the specimens in this group. Similarly, the QLF test demonstrated that teeth treated with Pro-seal had the least amount of fluorescence loss by far. The findings of this study confirmed that the Pro-seal functions as a protective barrier that is impermeable to the daily acid challenge. This impressive display of demineralization prevention under in vitro cariogenic conditions also has been observed in other studies (Hu and Featherstone, 2005;Buren et al., 2008).
When interpreting the results of the current study, it is important to examine the experimental methods used. Obviously, the oral cavity of the typical teenager presents a much more dynamic and abrasive environment than those used in this in vitro study. However, it has been shown that Pro-seal sealant also displays physical properties when subjected to abrasion (Hu and Featherstone, 2005). Pro-seal prevented enamel demineralization convincingly and, thus, seems to be a reasonable treatment option that requires zero patient compliance.
The results from this study also indicated that teeth treated with the fluoride varnish had less enamel demineralization than the control and the ACP groups. Although it had a statistically significant difference in both lesion depth and fluorescence loss when compared to the control group, the difference was not nearly as dramatic in the QLF test. Currently, there are no other in vitro studies in the literature that examine fluoride varnish around orthodontic brackets with both CLSM and QLF.
In that there was zero demineralization measured with the CLSM but some degree of fluorescence loss found with the QLF raises questions. In spite having the specimens brushed daily, for the most part the fluoride varnish remained unexpectedly on the tooth surface throughout the experiment and had to be removed with a plastic scaler at the end of the experiment. Therefore, its mechanism of action must be considered. In addition to the anti-cariogenic properties of fluoride as rationale for use, the fluoride may not have been the only mechanism of action in this in vitro experiment in that the varnish formed a physical barrier to the acid challenge.
In this study, the Aegis-Ortho group and the MI Paste group showed less demineralization numerically than the control group for both the CLSM and QLF test, though neither had statistical significance. Thus, both Aegis-Ortho and MI Paste were not different from the control group. The similar numerical levels of effectiveness for Aegis-Ortho and MI Paste are not surprising, given their similar mode of action. In analyzing these two treatments, the obvious disadvantage for the MI Paste group is that it requires daily application, whereas the ACP in Aegis-Ortho simply resides in the bracket bonding cement.
While the results of this study help us better understand the prevention potential of these products, in vitro experimental conditions cannot encapsulate all the complexities of a living oral cariogenic environment.
The ultimate answer on efficacy of these products has to come from well-designed controlled clinical trials. An in vivo randomized controlled trial study that employs proven methods for clinical evaluation of incipient lesions around brackets and also includes the patient compliance factor would provide the highest level of evidence with respect to the preventive treatment modalities discussed.

Conclusions
The incidence of WSLs in patients treated with comprehensive orthodontics was very high, suggesting that any preventive therapy provided appeared to be ineffective. This widespread problem poses an alarming concern and warrants significant attention from both patients and providers that should result in greatly increased emphasis on effective caries prevention. Results from this study suggest that both the lightcured filled sealer (Proseal) and the fluoride varnish (Vanish) have the potential to prevent enamel demineralization next to orthodontic brackets exposed to cariogenic conditions.