1

Introduction

The search for dental materials that are biocompatible with the dentin–pulp complex, in addition to having adequate physical, mechanical and esthetic properties, has guided the development of several commercial products. The availability of materials with adhesion to dental hard tissues has enabled the modern approach to operative dentistry involving minimal intervention and very conservative preparations to preserve sound tooth structure.

Direct and indirect pulp capping are clinical procedures that aim to preserve pulp vitality . Several materials are indicated for indirect pulp capping, including resin-modified glass ionomer cements (RMGICs) . As cavity liners RMGICs provide adequate protection to the dental pulp, preventing the occurrence of postoperative sensitivity. This characteristic is due in part to their self-adhesive nature and their capacity for reduction of stress generated by composite resin polymerization shrinkage . Adhesion of RMGICs to dental substrates combined with their ability to release fluoride, which may act to prevent secondary caries, makes these materials an excellent option for use in a wide array of clinical procedures .

In addition to hydrosoluble polyacrylic acid ion-leachable glass, which is also part of the makeup of conventional glass ionomer cements (GICs), RMGIC formulations include organic monomers (e.g. HEMA) associated with a photoinitiator system . This composition gives RMGICs a dual polymerization reaction (chemical cure and photoactivation) leading to higher flexural and diametral strengths, modulus of elasticity and wear resistance than conventional GICs . On the other hand, the presence of these organic monomers may produce adverse biological reactions, such as local and systemic toxicity, pulpal reactions and allergic and estrogenic effects . Regarding pulpal reactions, it is known that due to its low molecular weight and high hydrophilicity, HEMA may diffuse through dentin and reach the pulp . In addition to HEMA, other monomers have been incorporated into RMGICs, such as Bis-GMA, which has higher cytotoxic potential than HEMA and other monomers contained in different resin-based dental materials .

The potential for RMGICs to have a cytotoxic effect on fibroblast and odontoblast-like cell cultures has been extensively documented . Their toxic effects as direct and indirect pulp capping agents have also been investigated . However, these studies have been conducted with RMGIC materials that were available in powder/liquid form with components that were hand mixed immediately before clinical use. More recently, paste/liquid versions of RMGIC liners have been developed and introduced to the market in convenient dispensing devices with the aim of facilitating handling, offering an improved component mixing ratio, and shortening the clinical time. However, little information is available in the literature regarding their performance, and the biological effects of these new materials have not yet been investigated. Among these new paste/liquid RMGIC formulations, Vitrebond Plus Light Cure Glass Ionomer Liner/Base (VBP) is available in a proprietary ‘clicker’ device, which is claimed to give simpler and more consistent mixing. Although some of its characteristics such as adhesion to dentin have been investigated , there are no research-based data on the effects of this new ionomeric liner when used close to the pulp. The aim of this study was to evaluate the in vivo biocompatibility of VBP compared to the conventional powder/liquid formulation of this material (Vitrebond Light Cure Glass Ionomer Liner/Base – VB) and calcium hydroxide cement (Dycal – DY – Dentsply, Milford, DE, USA) in deep cavities prepared in sound human teeth.

2

Materials and methods

Thirty-eight caries-free human premolars in functional occlusion and scheduled to be extracted for orthodontic reasons were selected from young patients. The mean age of the patients was around 14 years. After receiving all necessary explanations about the research protocol, experimental rationale, clinical procedures and possible risks, the parents/guardians and volunteers read and signed an informed consent form previously approved by the Institutional Human Subjects Ethics Committee.

The radiographs taken for orthodontic treatment were used to evaluate the possible presence of proximal caries or potential periapical pathology. As a common diagnostic procedure for tooth extraction, periapical radiographs were taken immediately before the extraction of each tooth.

Thirty-four teeth were divided into 3 experimental groups in which the cavity floor was lined with VBP, VB or DY ( Table 1 ). Four sound teeth with no cavity preparation were included as an intact control group. Manufacturer information and the chemical composition of the liners are described in Table 1 .

In the experimental groups, asepsis of the oral cavity was performed with a 0.12% chlorhexidine rinse prior to administration of local anesthesia. After cleaning the teeth with rubber cups and pumice slurry, buccal Class V cavities were prepared using a high-speed handpiece with copious water spray cooling. In order to standardize the cavity to a preset depth, a slightly tapered diamond bur, with its cutting area previously limited to 2.5 mm by means of a resin cap was used. The bur was replaced after every fourth cavity preparation to avoid overheating. The final dimensions of the buccal cavities were 3.0 mm long, 2.5 mm deep and 1.5 mm wide, with no undercuts . All liners were prepared and applied strictly following the manufacturers’ instructions for clinical use in cavity lining procedures. Then, enamel and the lateral cavity walls were conditioned with 35% phosphoric acid for 15 s. After etching, the cavities were water rinsed thoroughly for 30 s to remove the acidic agent, and enamel was air dried and the cavity walls (dentin) were gently dried with sterile absorbent paper. Two coats of Adper Single Bond 2 adhesive system (3M ESPE) were applied to enamel and dentin, air-thinned and light-cured for 20 s (XL1000 Curing Light, 3 M ESPE). All cavities were restored to the enamel cavosurface margin with Filtek Z350 composite resin (3M ESPE) applied in increments of 1.5 mm or less. Each increment was light-cured for 40 s. A radiometer (Demetron/Kerr – Model 100P/N 10503, Danbury, CT, USA) was used to check the curing light intensity immediately prior to each clinical procedure. The light intensity was standardized at 420 mW/cm ² during the experiment. When necessary, any excess material at the cervical margin was mechanically removed using a fine grit diamond bur at high speed. After 24 h, the restorations were finished and polished using fine diamond burs and mounted abrasive stones. The clinical steps for each one of the groups are summarized in Table 2 .

At 7 or 30–60 days after the clinical procedures, a new radiograph was taken and the study teeth were extracted under local or regional anesthesia. The roots were immediately sectioned midway between the cementoenamel junction and the root tip with a high-speed handpiece under water spray. The teeth were stored for 48 h in formalin fixative solution at pH 7.2, decalcified in buffered Morse’s solution, dehydrated, vacuum-infiltrated with wax paraffin and finally embedded in paraffin. Six-μm-thick serial sections were obtained, mounted on glass slides, and stained with hematoxylin and eosin (H/E) and Masson’s Trichrome. The presence of bacteria was checked using the Brown & Brenn staining technique. All sections were evaluated blind for five histological features according to predefined criteria ( Table 3 ), as previously reported . The pulpal response was evaluated by light microscopy (Carl Zeiss 62774, Oberköchen, West Germany).

The remaining dentin thickness (RDT) between the cavity floor and the pulp chamber was measured for each tooth using a light microscope (Carl Zeiss) connected to a video camera (Samsung Digital Camera – SSC/131, Samsung Electronics Co. Ltd., Korea). The images were loaded into a computer and processed using standard software (ImageLab, Softium Informática, São Paulo, SP, Brazil). Data were analyzed statistically by two-way ANOVA ( material and period ) using the preset level of significance of 5% ( α = 0.05).

2

Materials and methods

Thirty-eight caries-free human premolars in functional occlusion and scheduled to be extracted for orthodontic reasons were selected from young patients. The mean age of the patients was around 14 years. After receiving all necessary explanations about the research protocol, experimental rationale, clinical procedures and possible risks, the parents/guardians and volunteers read and signed an informed consent form previously approved by the Institutional Human Subjects Ethics Committee.

The radiographs taken for orthodontic treatment were used to evaluate the possible presence of proximal caries or potential periapical pathology. As a common diagnostic procedure for tooth extraction, periapical radiographs were taken immediately before the extraction of each tooth.

Thirty-four teeth were divided into 3 experimental groups in which the cavity floor was lined with VBP, VB or DY ( Table 1 ). Four sound teeth with no cavity preparation were included as an intact control group. Manufacturer information and the chemical composition of the liners are described in Table 1 .

In the experimental groups, asepsis of the oral cavity was performed with a 0.12% chlorhexidine rinse prior to administration of local anesthesia. After cleaning the teeth with rubber cups and pumice slurry, buccal Class V cavities were prepared using a high-speed handpiece with copious water spray cooling. In order to standardize the cavity to a preset depth, a slightly tapered diamond bur, with its cutting area previously limited to 2.5 mm by means of a resin cap was used. The bur was replaced after every fourth cavity preparation to avoid overheating. The final dimensions of the buccal cavities were 3.0 mm long, 2.5 mm deep and 1.5 mm wide, with no undercuts . All liners were prepared and applied strictly following the manufacturers’ instructions for clinical use in cavity lining procedures. Then, enamel and the lateral cavity walls were conditioned with 35% phosphoric acid for 15 s. After etching, the cavities were water rinsed thoroughly for 30 s to remove the acidic agent, and enamel was air dried and the cavity walls (dentin) were gently dried with sterile absorbent paper. Two coats of Adper Single Bond 2 adhesive system (3M ESPE) were applied to enamel and dentin, air-thinned and light-cured for 20 s (XL1000 Curing Light, 3 M ESPE). All cavities were restored to the enamel cavosurface margin with Filtek Z350 composite resin (3M ESPE) applied in increments of 1.5 mm or less. Each increment was light-cured for 40 s. A radiometer (Demetron/Kerr – Model 100P/N 10503, Danbury, CT, USA) was used to check the curing light intensity immediately prior to each clinical procedure. The light intensity was standardized at 420 mW/cm ² during the experiment. When necessary, any excess material at the cervical margin was mechanically removed using a fine grit diamond bur at high speed. After 24 h, the restorations were finished and polished using fine diamond burs and mounted abrasive stones. The clinical steps for each one of the groups are summarized in Table 2 .

Table 2

Clinical steps for each one of the groups.

Group	Clinical steps
Vitrebond™ Plus (Group 1)	1. Fully depress clicker lever to dispense one ‘click’ of liquid and paste onto the mixing pad. One to two ‘clicks’ will be sufficient for most restorations
	2. Using a small spatula rapidly mix the liquid and paste together for 10–15 s.
	3. Apply a thin layer of the mixed liner to the cavity floor using a ball applicator. Light-cure for 20 s.
	4. Apply 35% phosphoric acid etchant to enamel and dentin, wait 15 s, rinse for 10 s
	5. Blot excess water leaving tooth moist
	6. Using a fully saturated brush tip for each coat, apply two consecutive coats of Adper Single Bond 2 adhesive to etched enamel and dentin with agitation for 15 s
	7. Dry gently for 2–5 s
	8. Light cure for 10 s
	9. Cavity is restored with Filtek Z350 composite resin.
	10. Select shade and place in increments of 2 mm. Light cure each increment 20 s
	11. Finish and polish
Vitrebond™ (Group 2)	1. Fluff Vitrebond powder in jar by shaking. Dispense one level scoop of powder and one drop of liquid onto the mixing pad.
	2. Using a small spatula mix the powder and liquid together for 10 s.
	3. Apply a thin layer of the mixed liner to the cavity floor using a ball applicator. Light-cure for 30 s.
	The following procedures are the same as described for Group 1.
Dycal (Group 3)	1. Dispense equal amounts of catalyst and base pastes onto the mixing pad.
	2. Using a small spatula mix the pastes together for 10 s.
	3. Apply a thin layer of the mixed liner to the cavity floor using a ball applicator.
	The following procedures are the same as described for Group 1.

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