Cytotoxicity evaluation of a new fast set highly viscous conventional glass ionomer cement with L929 fibroblast cell line

Abstract

Aim:

This study aims to evaluate the cytotoxicity of a new fast set highly viscous conventional glass ionomer cement (GIC) with L929 fibroblasts.

Materials and Methods:

The cement capsule was mixed and introduced into a paraffin wax mould. After setting, the cement was incubated in Dulbecco's Modified Eagle's Medium. Six replicates of the material extract were added to the culture medium in 96-well plates. L929 mouse fibroblast cells were added into the wells and then incubated for 48 h. Dimethylthiazol diphenyltetrazolium bromide test was performed for cytotoxicity evaluation.

Results:

The results showed that this GIC brand did not yield a half-maximal inhibitory concentration value, IC50, as the cell viability was above 50% at all concentrations. Cell viability over 90% was observed at the concentrations of 3.125 and 1.5625 mg/ml. Maximum concentration of the material showed cell viability of 59.4%.

Conclusions:

This new fast set highly viscous conventional GIC showed low cytotoxicity to mouse fibroblast cells, and it can be suggested as a substitute for dental cements exhibiting a long setting time.

INTRODUCTION

Since 1960, the idea of adhesion to dental hard tissues resulted in the invention of many dental cements including glass ionomer cements (GICs).[1] GIC is basically a product of acid base reaction between ion leachable glass and aqueous solution of polyacrylic acid.[2] This cement exhibits several desirable properties including good biocompatibility, physicochemical bonding to enamel and dentin, low coefficient of thermal expansion and anticariogenic activity.[3,4] Following many improvements, the GICs became one of the most successful dental cements in clinical dentistry.[4]

Beside all the developments and evolution of resin-modified GICs and compomers, conventional GICs have been subjected to many substantial modifications in their composition and powder/liquid ratio in order to overcome their compromised physical and mechanical properties.[5–12] These improved formulations, provided with fast set applications, would significantly reduce the probability of moisture contamination. In addition, the chair time will be reduced thus making the material more acceptable for both the patient and the clinician.[13]

Fast set, highly viscous conventional GICs were also introduced to extend the indication for occlusal restorations, which require high-strength materials. Even though they did not show an improvement in the mechanical and wear properties in the initial stages, the long term wear resistance and hardness was found to be high, and was suggested to compete with that of composite resins.[14,15]

As a result of this considerable heterogeneity in the composition of new conventional GICs, it is essential to evaluate their cytotoxic activity and host tissue response.[10,16] This preliminary study aims to investigate the cytotoxicity of a new fast set highly viscous radiopaque conventional GIC with the L929 mouse fibroblast cell line.

MATERIALS AND METHODS

A Paraffin wax mould, with a diameter 1.5 cm and depth 2 mm, supported by sterile stainless steel mould was prepared as mentioned by Min et al.[17] (with modification) [Figure 1a]. The fast set, highly viscous radiopaque conventional glass ionomer capsule (Ionofil Molar AC Quick, Voco, Germany) was activated and mixed immediately in a capsule mixer for 10 s as recommended by the manufacturer. The mix was then introduced into the paraffin wax mould [Figure 1b].

Figure 1.

(a) Paraffin wax mould supported by sterile stainless steel mould. (b) Glass ionomer cement after application. (c) Sterilization of the set cement by ultraviolet radiation

After setting of the cement (setting time 2.5 min), it was sterilized using ultraviolet radiation for 30 min [Figure 1c]. The cement was weighed and then introduced into sterile glass bottles containing Dulbecco's Modified Eagle's Medium (200 mg/ml). Following this, it was incubated for 72 h at 37°C.

After incubation, the material extract was passed through a filter (20 μm) into another sterile glass bottle. Six replicates of the material extract were added into 96-well plate. L929 mouse fibroblast cell culture was applied into the wells containing full and diluted concentrations of the material extract, and then incubated for 48 h at 37°C and 5% CO₂. Dimethylthiazol diphenyltetrazolium bromide was added to each well for 4 h. The wells were evacuated and Dimethyl sulphoxide was then introduced into each well and the optical density (wavelength 570 nm) was determined using an enzyme-linked immunosorbent assay reader.

RESULTS

The results showed that this GIC brand did not yield a half-maximal inhibitory concentration (IC50) as the cell viability was above 50% at all concentrations [Figure 2]. Cell viability over 90% was observed at the concentrations of 3.125 and 1.5625 mg/ml. At the concentration of 6.25, 12.5 and 25 mg/ml, the cell viability was above 80%. At the concentration 50 mg/ml, the cell viability was 68%. Maximum concentration of the material showed cell viability of 59.4%.

Figure 2.

Cell viability at different concentrations of the material

DISCUSSION

GIC is widely used in clinical dentistry as a luting cement, cavity base or liner and for the restoration of primary and permanent teeth.[18] In endodontic treatment, it can be used for root end filling, repair of perforation areas, filling of resorption defects and temporary filling during endodontic therapy.[4] Despite these wide clinical indications, the imperfect physical and mechanical properties of conventional GIC were the main cause for the resistance to its adoption in wider clinical applications.[3,4]

Resin-modified GICs were introduced in the late 1980s as an attempt to improve the physical and mechanical properties while maintaining the basic features of the conventional GIC.[4,7] However, many studies found them to be cytotoxic.[19,20] Furthermore, they also showed strong genotoxic effects.[21] These unfavorable properties are mainly attributed to their ability to release the monomer HEMA (2 hydroxethyl methacrylate) which can lead to a variety of adverse biological effects.[20]

The low cytotoxic activity observed in this fast set, highly viscous conventional GIC brand is consistent with other conventional types that show good biocompatibility due to their rapid neutralization, release of generally benign ions from the set cement and low setting exothermic reaction.[20]

Some clinical situations, like root end surgery, warrant the use of fast set dental cement, as this will prevent its solubility, disintegration and dislodgement thus, maintaining its dimensional stability.[22,23] The presence of these fast set conventional GICs in pre-proportioned, mechanically mixed capsules will provide an easier procedure with a consistent mix, which can be effectively applied to the area of interest.

The concurrent existence of low cytotoxicity along with favorable physical properties including simpler handling properties, suitable working and setting time, is highly advantageous when the material is going to be introduced in areas where the moisture is difficult to control, and it will also be in direct contact with vital tissues, like the periapical tissues.[24] This would appreciably reduce the host tissue response and maintain the dimensional stability of the cement.

CONCLUSIONS

The new fast set highly viscous conventional GIC showed low cytotoxicity to mouse fibroblast cells and, in some clinical situations, it can be suggested as a substitute for dental cements exhibiting long setting time