Marginal microleakage of composite resin restorations bonded by desensitizing one step self etch adhesive

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ScienceDirect Tanta Dental Journal 11 (2014) 180e188 www.elsevier.com/locate/tdj

Marginal microleakage of composite resin restorations bonded by desensitizing one step self etch adhesive H.Y. El Sayed*, A.I. Abdalla, M.E. Shalby Faculty of Dentistry, Operative Dentistry, Tanta University, Egypt Received 20 April 2014; revised 30 September 2014; accepted 10 October 2014 Available online 19 December 2014

Abstract Aim: The aim of this study was to investigate the effect of desensitizing agents on microleakage of composite resin restorations bonded by one step desensitized self etch adhesive. Materials and methods: Class V cavities (3
4
2 mm) were prepared on the buccal surfaces of 30 human molars, Teeth were classified into three groups according to the type of adhesive used (n ¼ 10). Group 1: Adper Single Bond2. Group 2: l-bond. Group 3: is a control group of composite resin restoration without bonding agent. All cavities were restored with Filtek Z250 composite resin, stored in distilled water at 37  C for 24 h and thermal cycled for 500 cycles. The root apices were sealed with utility wax, and all the surfaces, except for the restorations and 1 mm from the margins, were coated with two layers of nail varnish. The teeth were immersed in a 3% methylene blue dye solution for 24 h, and then rinsed in running water, blot-dried and sectioned longitudinally through the center of restorations from the facial to lingual surface. The sections were blindly assessed for microleakage of dye penetration by two independent evaluators using a stereomicroscope at 30
magnification for both the occlusal and gingival margins. Data were collected and statistically analyzed. Results: A ManneWhitney test demonstrated the highest significantly dye penetration rate for group 3 versus tested groups comparing the occlusal and gingival scores for each group, Wilcoxon Rank test showed significant difference for all groups. Etch and rinse adper single bond2 adhesive record a lower significant scores of dye penetration than self etch adhesive I-bond at both enamel and dentin margins. Conclusion: All adhesive systems exhibited dye penetration at both occlusal and gingival margins. Etch and rinse system is still considered a gold standard adhesive. Adper single bond2 revealed significantly less leakage compared to I-Bond. © 2014, Hosting by Elsevier B.V. on behalf of the Faculty of Dentistry, Tanta University.

Keywords: Dentin hypersensitivity; Self etch desensitized adhesive; Marginal leakage; Composite resin; Thermocycling

1. Introduction * Corresponding author. E-mail addresses: [email protected], hussien. [email protected] (H.Y. El Sayed), ali_abdalla79@yahoo. com, [email protected] (A.I. Abdalla), magda.shalabi@ dent.tanta.edu.eg (M.E. Shalby). Peer review under the responsibility of the Faculty of Dentistry, Tanta University.

Dentin hypersensitivity due to dentin exposure, or after tooth restoration, has been effectively explained as the hydrodynamic theory of pain, attributed to fluid flow in the dentinal tubules which stimulates the nerve fibers within the dentinal tubules, resulting in pain [1].

http://dx.doi.org/10.1016/j.tdj.2014.10.001 1687-8574/© 2014, Hosting by Elsevier B.V. on behalf of the Faculty of Dentistry, Tanta University.

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The principles of adhesive dentistry date back to 1955 when Buonocore, using techniques of industrial bonding, postulated that acids could be used as a surface treatment before application of the resins [2]. As time went on, variations in duration of the acid-etching procedure and concentration of the phosphoric acid, along with alternative acids, were tested for the etching of enamel [3]. The current thinking is that a 30 to 40 percent phosphoric acid etch of 15 s is acceptable. Bonding to etched enamel was considered a safe and reliable procedure for many years. Because of the inorganic composition of the enamel, the acid attack produces inter-prismatic and prismatic dissolution, creating irregularities into which the resin can flow and, after polymerization, create a mechanical interlocking. The demineralization of the enamel depends on the low pH of the acid and on the etching time. The pH and the etching time must be sufficient to provide adequate enamel retention without the need for additional steps. The morphological studies made on the first five generations of bonding systems, in which phosphoric acid was used to etch enamel, showed a uniform etch pattern. When phosphoric acid was not used or when self-etching primers (fifth and sixth generations) were applied, the bonding mechanism of adhesives to enamel was less effective. The bonding mechanism to dentin was effective and predictable when the smear layer was completely dissolved, intertubular and peritubular dentin were dissolved, collagen fibers exposed and, after infiltration of resin monomers, a hybrid layer formed. This bonding mechanism was evident from fourth to sixth generations of enamel dentin bonding systems [4e6]. In fact, the mechanical properties of the bonding mechanism achieved with hybrid layer and resin tag formation can be greater than the forces of polymerization contraction. Also, bonding systems are indicated in any direct esthetic restorations [7]. The ability of composite to bond reliably to enamel is now well-accepted, but adhesion of restorative materials to dentin has proved to be more elusive [8]. Early attempts to bond to dentin resulted in poor bond strengths [9]. This is not surprising given the fact that while enamel contains little protein, dentin is 17% collagen by volume. This collagen is inaccessible due to surrounding hydroxyapatite crystals [10]. The dentinal tubules are the only pores available for micromechanical retention. These tubules contain fluid, which would be an impediment to bonding. The number of tubules available for bond also varies depending on location, with deep dentin having more tubules than superficial dentin. Other factors such as age of teeth, direction of tubules, and type of

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dentin can affect dentin bonding [11]. The bond strength between restorative material and dentine plays an important role in the success of a restoration. This bond is influenced by the surface structure of the restorative material and dentine. Many factors, including the nature of the prepared dentine surface, physical and chemical properties of the restorative material, the operating limitations of the restorative materials, intra-oral environment and the occlusal forces, need to be considered for long-term restorative applications [12e14]. Both primer and adhesive agent have to penetrate dentine to form a micromechanical bond with the microporosities in the dentinal tubules and collagen fibers [15]. When using a conventional acid-etching method, total demineralization of the etched dentinal surfaces opens up dentinal tubules. In class V cavities with deeper carious lesions, phosphoric acid may cause postoperative sensitivity from the damage to the pulp [16]. Using aggressive acids to provide acid etching can denature collagen fibers, and demineralize a thicker layer of dentine, into which resins cannot reach to the same depth [17,18]. A self-etching system uses a weak acid to etch dentine surface, which hardly damages to the pulp and prevents sensitivity [12,19]. In addition, this system reduces the number of application steps and gives rise to adequate bond strength [18]. To reduce postoperative sensitivity, dentists increasingly use desensitizers based on hydroxyethyl metacrylate (HEMA), fluoride, and chlorhexidine gluconate after tooth preparation for restorations. The function of fluoride present in dentin desensitizers is to seal the dentinal tubules with incorporation of mainly HEMA, which increases the infiltration ability of primers [20]. Chlorhexidine is an antiseptic with a wide spectrum of action that has been used over the past two decades for the chemical control of bacterial plaque and the prevention of dental caries [23]. It is the most effective antimicrobial agent [24], and it has a proven ability to delay bond degradation [71]. For this reason, chlorhexidine has been added to the desensitizers in recent years. Although there is no information concerning the effects of chlorhexidine based desensitizers on the bonding performance of composites to tooth tissues, previous studies have shown that application of chlorhexidine-containing cavity disinfectants before or after acid-etching procedures does not have a negative effect on the shear bond strength; in fact, this procedure may increase bond strength and durability [21,22]. In 2007, report stated that, increasing the strength of the dentin matrix by using cross-linking agents may improve both the strength and durability of resin-dentin bonds, and showing that glutaraldehyde is a collagen

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Table 1 Tested materials. Tested material

Composition

Manufacture

Manufacturer's instruction for use

Scotch bond Etchant (gel) (pH 0.6) Adper Single Bond2 (pH 4.7) [two step, etch& rinse adhesive system] Nanofillers

35% phosphoric acid, colloidal silica Water, ethanol, HEMA, dimethacrylates, methacrylate functional copolymers of polyacrylic and politaconic acids, (Bis-GMA), silica nanofillers and photo initiator

3M ESPE, St. paul, MN, USA

I-Bond Self etch (pH 2e2.2) [one step, non etch& rinse desensitizing adhesive system) No fillers

-

Heraeus Kulzer, Inc., New York

Filtek Z250 Micro-hybrid resin Composite

BIS-GMA and Low-viscosity resin TEGMA (Tri ethylene glycol dimethacrylate). Blend of UDMA (urethane dimethacrylate) and BisEMA (Bisphenol A polyethylene glycol diether dimethacrylate) in place of TEGDMA. . Z250 is filled to 60% by volume with zirconia/silica particles having a size range of 0.01 e3.5 microns and an average size of 0.6 micron.

- Etching: Apply Etchant, wait 15 s. Rinse for 10 s. Blot excess water using a mini-sponge. Surface should appear glistening without pooling of water. - Immediately after blotting, apply 2 e3 consecutive coats of adhesive for 15 s with gentle agitation using a fully saturated applicator. Gently air thin for 5 s to evaporate solvents. Light cure for 10 s. - Saturate microbrush with i-Bond liquid from bottle, Apply three consecutive coats of i-Bond followed by gentle rubbing for 30 s. - Use gentle air pressure to remove acetone and water solvent and Cure for 20 s with a dental curing light of at least 500 mW/C2. Application of composite in layers less than 2 mm in thickness and cure with light cure unit for 20 s according to Manufacturer's instructions.

UDMA, 4-Meta Acetone and water. Camphorquinone. Glutaraldehyde. Stabilizers.

cross linker, which increases the stiffness of the demineralized dentin [25]. Since dentin is inherently wet, the development of hydrophilic primers dissolved in inorganic solvents (acetone/alcohol) was an important achievement in dentin bonding-embedding the exposed collagen fibrils with resin. HEMA is a component of many hydrophilic adhesives because of HEMA's ability to facilitate and increase the adhesion to dentin collagen due to its hydrophilic nature [26,27]. Because these glutaraldehyde/HEMA products also contain water, they act as rewetting agents, and in the case of total-etch bonding procedures, the “rewetting after the acid etch acts to expand the demineralized collagen and increase its surface energy, facilitating the diffusion of the hydrophilic resin monomer into the etched dentin [28], and facilitating adhesion translate into higher and more durable bond strengths. Glutaraldehyde is a very effective fixative agent having the ability to create a coagulation plug inside the dentinal tubules. It has been accepted extremely well by dentists as a material that reduces or totally eliminates tooth sensitivity [29,30].

3M ESPE, St. paul, MN, USA

2. Aim of the study The aim of the present study was to investigate the effect of desensitizing agents on micro-leakage of composite resin bonded to dentin by two step etch and rinse adhesive system and one step desensitized self etch adhesive. The null hypothesis tested was that the application of simplified desensitizing adhesive system on enamel and dentin would have no effect on the microleakage of resin composite restorations. 3. Material and methods Materials that have been used in this study illustrated in Table 1 including the following: 1- Two step, etch and rinse adhesive system: Adper Single Bond21

1

3M ESPE, St. Paul, MN, USA.

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2- One step, non-water rinse self-etch desensitizing adhesive system: I-Bond Self etch2 3- Filtek Z250 Micro hybrid Composite resin restoration3

3.1. Specimen preparation Thirty extracted human third molars free from crakes, restorations or caries were used in this study. A written consent was taken from patients after the study was approved by the Ethics Committee of Tanta University to ensure their agreement to use their teeth in the current study. Immediately after extraction, the teeth were scraped of any residual tissue tags, pumiced, and washed under running tap water. The teeth were stored in distilled water at þ4  C until required, as the freezing process maintain the dentinal collagen as closest as the in vivo condition thus avoiding its denaturation or fixation which occurs in the conventional methods of storage (water, saline solution, formalin, among others) [31,32]. Standardized, Class V cavities with butt-jointed margins were prepared on the buccal aspect of each tooth with round internal angles, 1 mm below the CEJ with occlusal margins in enamel and cervical margins in dentin, and with following dimensions: 3 mm gingival-occlusally, 4 mm mesio-distally and 2 mm deep, using a water-cooled high-speed diamond bur #330.4 Used burs were replaced by new ones after each five preparations. After preparation, the teeth were randomly divided into three groups 10 specimens each. The desensitizer tested in this study was I-Bond self-etch one step, non-water rinse adhesive system. In Group 1, applying composite resin bonded with Adper Single Bond2 (two step, etch and rinse adhesive system) (n ¼ 10). In Group 2, applying composite resin bonded with I-Bond self-etch one step, non-water rinse desensitizing adhesive system (n ¼ 10). In Group 3, control group applying composite resin without any adhesive system as negative group (n ¼ 10). All the cavities were restored with a shade (3) micro-hybrid resin composite, Filtek Z250, in two increments and each increment was polymerized for 40 s

Fig. 1. Diagram showing the 0e3 point evaluation scales for composite resin micro-leakage.

using a conventional halogen light-curing unit.5 The restorations were finished and polished with OptiDisk.6 The teeth were stored in distilled water at room temperature for 24 h before being subjected to 500 thermal cycles between 5  C and 55  C water baths with a 30-second dwell time and a 15-second transfer time. The root apices were sealed with utility wax, and all the surfaces, except for the restorations and 1 mm from the margins, were coated with two layers of nail varnish. The teeth were immersed in a 3% methylene blue dye solution for 24 h. They were then rinsed in running water, blot-dried and sectioned longitudinally through the center of the restorations from the facial to lingual surface with a water-cooled diamond wheel saw.7 The sections (n ¼ 20 for each group) were blindly assessed for dye penetration by two independent evaluators using a stereomi-croscope8at 30
magnification. Dye penetration at the composite/tooth interface was scored for both the occlusal and gingival margins from 0 to 3 (Fig. 1): 0 ¼ no leakage visible, 1 ¼ penetration of dye along the cavity wall, but less than ½ of the cavity depth. 2 ¼ penetration of dye along the cavity wall, more than ½ of the cavity depth. 3 ¼ penetration of dye spreading to and along the axial wall. Data were collected and statistically analyzed using the KruskaleWallis and ManneWhitney U-tests. A comparison of the occlusal and gingival margins of the groups was performed.

2

Heraeus Kulzer, Inc., New York.. 3M ESPE, St. Paul, MN, USA. 4 KG Sorensen, Barueri, SP, Brazil. 5 Coltolux 75, Colt
ene/Whaldent AG, Alst€atten, Switzerland, 500 mW/cm2. 3

6 7 8

Kerr Corporation. Isomet, Buehler, Lake Bluff, IL, USA. Olympus Co., Tokyo, Japan.

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Table 2 Dye Penetration Scores, means and standard deviations for the Tested Materials at the occlusal and gingival walls. Microleakage scores of Z250 composite bonded with I-bond

Microleakage scores of Z250 composite bonded with adper single bond2

Microleakage scores of Z250 composite (control group)

Occlusal

Occlusal

Cervical

Occlusal

Cervical

N ¼ 20 Mean ¼ 0.15 s.d. ¼ ±0.3663 3.01 (0.0075)**

N ¼ 20 Mean ¼ 0.8 s.d. ¼ ±0.8944

N ¼ 20 Mean ¼ 2.15 s.d. ¼ ±0.8751 3.64 (0.0019)**

N ¼ 20 Mean ¼ 2.8 s.d. ¼ ±0.4104

Cervical

N ¼ 20 N ¼ 20 Mean ¼ 0.45 Mean ¼ 1.3 s.d. ¼ ±0.5104 s.d. ¼ ±0.9787 T(P): 3.52 (0.0024)** F (P). Occlusal margins ¼ 59.58(0.000) **. Cervical margins ¼ 35.61(0.000) **. T ¼ T-test of paired samples. *Significant at P < 0.001. **Highly significant at P < 0.001.

4. Results Statistical analysis was performed utilizing the KruskaleWallis one-way ANOVA followed by a ManneWhitney test. The difference between the occlusal and gingival dye penetration scores for each group was analyzed by the Wilcoxon Signed Rank test. Statistical analysis was performed using the following computer program: Statistical Package for the Social Sciences.9 In the current study, dye penetration mean scores for the occlusal and gingival walls are presented in Tables 2 and 3 and Fig. 2. KruskaleWallis one-way ANOVA indicated significant differences between adhesive groups for occlusal and gingival mean scores, where the enamel margins subjecting to a lower dye penetration than dentin margins (mean scores at enamel walls were 0.45, 0.15, 2.15 of I-bond, Adper single bond2 and composite without bonding respectively while at dentin walls were 1.3, 0.8 and 2.8 respectively). On the other hand, non-adhesive group subject to a well significant higher dye penetration scores at both enamel and dentin margins than the adhesive groups. The ManneWhitney test was performed to evaluate significant differences of occlusal mean scores between groups. The results demonstrated a significant high leakage differences between control group (composite resin eno bonding) and two other bonding groups (I-Bond and Adper single bond2) at p > 0.05, and i-Bond had recorded a significantly more dye penetration score when compared to Adper single bond2 (0.45, 0.15 respectively, p < 0.001).

9

SPSS Inc, Chicago, IL, USA.

Gingival mean scores between groups also showed a high significant differences of dye penetration dentin leakage between control group (composite resin eno bonding) and two other bonding groups (I-Bond and Adper single bond2) at p > 0.05: and i-Bond showed significantly more marginal dentin dye penetration when compared to Adper single bond2 (1.3, 0.8 respectively, p < 0.001). When comparing the mean values of occlusal and gingival mean scores for each group, the Wilcoxon Rank test showed that, occlusal enamel margins recorded a lower significant difference of dye penetration scores than gingival dentin walls for I-Bond (0.45, 1.3) and Adper single bond adhesives (0.15, 0.8) at (p < 0.001). 5. Discussion Adhesives are necessary to prevent micro-leakage at resin composite restoration/tooth interface while dental composites are not able to bond to dental tissues. However, clinical microleakage remains the major cause for composite restoration failures implying postoperative sensibility, margin colorations, secondary decay, or pulpal inflammation. When composite resin restorations used without bonding, the results of the current study where dye penetration leakage scores at the enamel and dentin walls are significantly higher than composite resin with either self etch or total etch adhesives and this result in agreement with reports indicated by several studies [33e35]. Therefore, manufacturers have proposed many different adhesives involving different adhesion strategies, the Etch and Rinse (ER) adhesive systems (in three or two clinical steps), the Self-Etch (SE) adhesive systems (in two or one clinical step(s), and the glass ionomer adhesives [36,37]).

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Table 3 Dye Penetration Scores for the Tested Materials at occlusal and gingival walls. Occlusal Group/score Composite bonded with I-bond. n ¼ 20 (*SE one step adhesive non-etch & rinse) Composite bonded with Adper single bond2. n ¼ 20 (*ER two step adhesive) Composite no bonding (Control group) n ¼ 20

Gingival

0 11

1 9

2 0

3 0

0 5

1 6

2 7

3 2

17

3

0

0

10

4

6

0

1

3

8

8

0

0

4

16

*SE ¼ self etch adhesive, * ER ¼ etch and rinse adhesive.

The major goal of using dentin bonding agents is to enhance the bonding strength between the resin and the tooth structure, increase the retention of restoration, reduce the micro-leakage across dentin-resin interface [38]. The introduction of the total etches technique and recent developments in the chemistry of dentin adhesives have made resin based composite restorative materials nearly free of micro-leakage with bond strengths approaching those of enamel bonding [39,40]. Generally, lower bond strength of dental adhesives to dentin might be attributed to various factors: First, dentin is less mineralized tissue and has a higher water content compared to enamel. Second, presence of the smear layer. Third, fluid filled channels in dentin that are under slight but constant out ward pressure from the pulp decrease the stability and durability of the composite resin-dentin bond [41]. Clinically an adhesive that can bond to any surface is preferred, as can mild self etch adhesives [42]. The current study, evaluated the microleakage of different adhesives comparing self-etch and total-etch and rinse systems, all of which demonstrated dye penetration scores (leakage) at both the enamel and dentin margins. According to the Wilcoxon signed

Fig. 2. Represent mean dye penetration scores of I-Bond and Adper Single Bond2 adhesive systems at both occlusal and cervical margins of composite restorations.

rank test, significantly lesser dye penetration score leakage was recorded at the enamel margins compared to the dentin margins of the adhesive groups. These results are in accordance with several studies that evaluated self-etch and total-etch and rinse adhesive systems, less microleakage was also reported at the enamel margins compared to the dentin margins [43e46]. It was stated that, self etch-adhesive systems can hybridize the dental hard tissues through one or two operatory-steps. In part depending on pH, the self etchadhesives may be classified according to the interaction depth at dentin into ultramild e (pH > 2.5 “nanointeraction”), mild (pH z 2 e interaction depth about 1mm), intermediate strong (pH 1e2, interaction depth between 1 and 2mm), and strong (pH  1 e interaction of several micrometers' e depth). The latter e are able to produce an interfacial ultra morphology resembling that typically produced by etch and rinse adhesives [47]. Nevertheless, several in vitro studies have shown that despite the reasonable bonding potential of the strong self etchadhesives, reduced bond strength [49,50], and increased interfacial Nanoleakage are recorded in comparison to the multistep adhesives, especially regarding the strong simplified adhesives rich in HEMA [48]. In fact, these less-favorable e in vitro results were confirmed by the inferior clinical performance of the strong one stepselfetchadhesives [51]. Analysis of the data from this study revealed significantly lower mean values of microleakage with a total-etch and rinse adhesive (Adper Single bond2primer þ bonding agent pH 4.7/etchant pH 0.6) compared to the other self etch one step adhesive IBond (pH 2e2.2) at the enamel and dentin walls. This finding was in agreement with studies reporting decreased leakage associated with total-etch especially at the enamel margins, compared to self-etch adhesive systems [52e54]. Also, a reduced microleakage scores has been reported when using filled adhesives [55,56]. As Adper

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single bond2 in comparison to I-bond where no fillers are present. However, some contradictory results have been reported by other researchers [44e46], who reported that I-Bond have no significant difference of leakage at dentin margins in comparable to total etch and rinse adhesive. Microleakage of restorations using self-etch no etch and rinse adhesives could have resulted from incomplete etching of enamel and dentin surfaces by acidic monomers, allowing for higher leakage values than the total-etch systems using a separate phosphoric acid etchant. Scanning electron microscopy (SEM) studies have shown that the use of phosphoric acid as an enamel and dentin etchant improves penetration and the subsequent attachment of adhesive monomers [57,58]. On the other hand, mild self-etch adhesives produce hybrid layers (HL) thinner than total-etch systems. As dentin demineralization is less pronounced, smear plugs occlude the orifice of the dentinal tubules, which are partially infiltrated by resin; a reduced resin tag formation occurs with these systems [59,60]. It was suggested that the bonding effectiveness of mild selfetch adhesives may result from a combined micromechanical and chemical interaction with tooth substrate [37,61]. The chemical component may be able to compensate for the reduced bonding effectiveness from decreased micromechanical interlocking. Also in the current study, Adper single bond2 is etch-rinse and alcohol-based solvent system and record a highly significantly lower mean scores of dye penetration at enamel and dentin margins than I-Bond desensitizing self etch which is acetone-based solvent and containing Glutaraldehyde/HEMA solution as a cross-linking agent. This result is disagreed with AlAmmar et al. [62] who found that “the application of selective collagen cross-linkers during adhesive restorative procedures may be a new approach to improve dentin bond strength properties” and showed that the chemical modification to the dentin matrix promoted by glutaraldehyde increased bond strengths. It has also been shown that a Glutaraldehyde/HEMA solution does not interfere with the bonding procedure to dentin when either an acetone primer or an alcoholbased primer is used [63]. Authors Reported advantages of self-etch systems include “simple” application procedures and a reduction and/or elimination of post-treatment sensitivity; however, because of numerous uncontrolled variables encountered during patient treatment, perceived advantages can potentially become disadvantages [64,65].

In the present study, the microleakage of adhesively bonded composite resin was evaluated in Class V cavities. The reason for studying Class V cavities was that (1) Class V cavities have unfavorable C-factors, resulting in high contraction scores within an adhesively fixed resin material, (2) Class V restoration margins are located in enamel as well as in dentin, (3) preparation and restoration of Class V lesions are minimal and relatively easy, thereby somewhat reducing practitioner variability, and (4) it is easier to standardize the preparation of Class V cavities than Class II cavities [66,67]. The dye penetration method used in the current study is a gross assessment of the quality of the interface. Although microleakage increases as dentin permeability rises [68], a reduction in dentin permeability does not necessarily result in a drop in microleakage. Any interference with hybrid layer formation and adaptation/bonding to the composite would adversely affect marginal microleakage. A significant role of hybridization on marginal leakage and bond strength has previously been reported [69]. On this base, in the current study, highest significant scores of dye penetration were observed in the negative control group composite resin without any adhesive application followed by using I-Bond self etch adhesive, while Adper single Bond2 record a significant lowest score of mean dye penetration. Since the hybrid layer morphology was not evaluated microscopically in this study, the specific nature of restoration failure (microleakage) for each adhesive system is unknown, although anecdotally four factors are strongly suspected: inefficiency of acidic monomers in alteration of the smear layer for classic hybrid layer formation, cavity C-factor, orientation of dentinal tubules to the CEJ and post-treatment stresses caused by polymerization contraction [70,71]. 6. Conclusions Within the limitations of this in vitro study, the following conclusions were reached: 1- The null hypothesis that desensitizer do not adversely affect micro-seal of composite resin restorations is rejected 2- All adhesive system groups exhibited dye penetration (leakage) at both occlusal (enamel) and gingival (dentin) margins. 3- An inter-group comparison revealed highly significantly lower leakage at the enamel margin versus the dentin margin of all adhesive groups.

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