Article Options


Advanced Search

This service is provided on D[e]nt Publishing standard Terms and Conditions. Please read our Privacy Policy. To enquire about a licence to reproduce material from and/or JofER, click here.
This website is published by D[e]nt Publishing Ltd, Phoenix AZ, US.
D[e]nt Publishing is part of the specialist publishing group Oral Science & Business Media Inc.

Creative Commons License

Recent Articles RSS:
Subscribe to recent articles RSS
or Subscribe to Email.

Blog RSS:
Subscribe to blog RSS
or Subscribe to Email.

Azerbaycan Saytlari

 »  Home  »  Endodontic Articles 1  »  In vitro cytotoxicity of a composite resin and compomer
In vitro cytotoxicity of a composite resin and compomer
Discussion - References.

A dentine barrier, simulating the clinical situation, was used to determine whether the two dental restorative materials were cytotoxic in vitro. The dentine disc acted as a physical barrier between the restorative material and the underlying culture medium and cells. However, a dentine thickness of only 70 microns was chosen, because a barrier of increased thickness may mask the material’s potential cytotoxicity. Earlier work by Tyas (1977) using dentine chips showed the toxicity of zinc phosphate cement to be substantially reduced by such a barrier. Although it is not possible to make a direct comparison between two different cell lines, this would appear consistent with the more rapid appearance of toxicity in the HL-60 cells incubated in the absence of the dentine barrier, where both materials resulted in substantial cell death after 12 h, as compared to the adherent ECV-304 cell-line in the presence of the barrier, where substantial toxicity was not observed until after 2 or 3 days of incubation. To mimic the clinical situation more closely, primer was placed on the dentine prior to placement of the restorative material. This layer forms a thin barrier between the restorative and the pulp. However, the effectiveness of this barrier, in terms of limiting ingress from the restorative material, is not known.
Whereas the MTT assay is a good indicator of cell viability, the LDH assay is an indicator of cell lysis. The results of these two assays revealed that, in the presence of the 70 m dentine barrier, Dyract® AP had no effect upon cell lysis until the third day of incubation, whereas the toxic effects of Spectrum® were evident after only 2 days’ incubation. Thus the cytotoxicity exerted through dentine slices by the Dyract® AP restorative material was less than that of Spectrum®, which was extremely cytotoxic and more likely to evoke an adverse pulpal reaction.
During cavity preparation, prior to placement of the restorative material, the mechanical action of the bur may cause microclefts in the dentine. In such cases, the restorative material would be in direct contact with the pulpal tissue, thus magnifying the potential to evoke an adverse pulpal response (Swift et al. 1995). In the present work a suspensory cell-line (HL-60) was used, in the absence of the dentine barrier, for studies on such direct toxicity and on the mechanisms underlying the toxicity of these restorative materials.
Microscopic examination of the stained cells revealed both restorative materials cause cell death, with Spectrum® being more toxic than Dyract® AP. When fully cured materials (40 s light-curing) were used, the predominant form of cell death was apoptosis. In contrast, the main form of cell death following exposure to partially cured materials (4 s light-curing) was necrosis.
Fully cured samples of Spectrum® and Dyract® AP caused activation of caspase-3. Thus, in agreement with the morphological staining data, the fully cured samples induce apoptotic cell death. However, exposure to the partially cured (1 and 4 s) materials resulted in less caspase activation than the fully cured materials. This contrasts with the morphological staining results that showed the partially cured samples to be more toxic than the fully cured material. This would indicate that whilst the partially cured samples cause some apoptosis, the materials provide such an overwhelming insult that the majority of the cells die rapidly by necrosis, and the less cured the material is, the greater the toxicity. Apoptosis and necrosis have been shown to occur together in some other systems. For example, in myocardial infarction cell death is associated with rapid necrosis immediately around the central ischaemic zone, whereas outside the central zone cells die more slowly by apoptosis (Cohen 1993).
Even in fully set restorative materials substantial amounts of short-chain polymers remain unbound, with the result that there is possible elution of leachable toxic components toward the pulp (Ferracane 1994). This may be compounded by inadequate light-curing. In some instances the light-curing unit may emit light that is not at the optimum wavelength (490 nm) for complete setting of the material (Miyazaki et al. 1998). As a result, the restoration might appear solid to probing but would in fact, be incompletely set and capable of initiating adverse pulpal reactions. Poor awareness of the need for regular maintenance of the light intensity of light-curing units amongst dentists has been reported (Martin 1998). In the clinical situation necrotic cell death would result in an inflammatory response and possibly secondary tissue damage to the patient. The present findings indicate that the degree of light-cure is an important factor in the toxic potential of a material, where an inverse relationship exists between the degree of light-curing and restoration cytotoxicity.


Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of proteindye binding. Annals of Biochemistry 72, 248-54.
Brannstrom M (1996) Reducing the risk of sensitivity and pulpal complications after the placement of crowns and fixed partial dentures. Quintessence International 27, 673-80.
Cohen JJ (1993) Apoptosis: the physiologic pathway of cell death. Hospital Practitioner 28, 35-43.
Cotter TG, Martin SJ (1995) Techniques in Apoptosis - A User’s Guide, 1st edn. London, UK: Portland Press Inc.
EscargueilBlanc I, AndrieuAbadie N, CasparBauguil S et al. (1998) Apoptosis and activation of the sphingomyelinceramide pathway induced by oxidized low density lipoproteins are not causally related in ECV304 endothelial cell. Journal of Biological Chemistry 273, 89-95.
Ferracane JL (1994) Elution of leachable components from composites. Journal of Oral Rehabilitation 21, 441-52.
Geurtsen W, Spahl W, Leyhausen G (1998) Residual monomer/additive release and variability in cytotoxicity of lightcuring glassionomer cements and compomers. Journal of Dental Research 77, 2012-9.
Gerzina TM, Hume WR (1994) Effect of dentine on release of TEGDMA from resin composites in vitro. Journal of Oral Rehabilitation 21, 463-8.
Hanks CT, Wataha JC, Parsell RR, Strawn SE, Fat JC (1994) Permeability of biological and synthetic molecules through dentine. Journal of Oral Rehabilitation 21, 475-87.
Jarvis WD, Turner AJ, Povirk LF, Traylor RS (1994) Induction of apoptotic DNA fragmentation and cell death in HL60 human promyelocytic leukemia cells by pharmacological inhibitors of protein kinase c. Cancer Research 54, 1707-14.
Kaushal GP, Ueda N, Shah SV (1997) Role of caspases (ICE/CED 3 proteases) in DNA damage and cell death in response to a mitochondrial inhibitor, antimycin A. Kidney International 52, 438-45.
Kawahara H, Yamagami A, Nakamura M (1968) Biological testing of dental materials by means of cell culture. International Dental Journal 18, 443-67.
Korzeniewski C, Callewaert DM (1983) An enzymerelease assay for natural cytotoxicity. Journal of Immunological Methods 64, 313-20.
Martin FE (1998) A survey of the efficiency of visible light curing units. Journal of Dentistry 26, 239-43.
Meryon SD, Riches DWH (1982) A comparison of the in vitro cytotoxicity of four restorative materials assessed by changes in enzyme levels in two cell types. Journal of Biomedical Materials Research 16, 519-28.
Miyazaki M, Hattori T, Ichiishi Y, Kondo M, Onose H, Moore BK (1998) Evaluation of curing units used in private dental offices. Operative Dentistry 23, 50-4.
Mosmann T (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. Journal of Immunological Methods 56, 55-63.
Muller J, Bruckner G, Kraft E, Horz W (1990) Reaction of cultured pulp cells to eight different cements based on glass ionomers. Dental Materials 6, 172-7.
Pashley DH (1996) Dynamics of the pulpodentin complex. Critical Review Oral Biology and Medicine 7, 104-33.
Pashley DH, Michelich V, Kehl T (1981) Dentin permeability: effects of smear layer removal. Journal of Prosthetic Dentistry 46, 531-7.
Powis DR, Prosser HJ, Wilson AD (1988) Longterm monitoring of microleakage of dental cements by radiochemical diffusion. Journal of Prosthetic Dentistry 59, 651-7.
Rathbun MA, Craig RG, Hanks CT, Filisko FE (1991) Cytotoxicity of a BisGMA dental composite before and after leaching in organic solvents. Journal of Biomedical Materials Research 25, 443-57.
Ritke MK, Rusnak JM, Lazo JS et al. (1994) Differential induction of etoposidemediated apoptosis in human leukemia HL60 and K562 cells. Molecular Pharmacology 46, 605-11.
Rueggeberg FA, Craig RG (1988) Correlation of parameters used to estimate monomer conversion in a lightcured composite. Journal of Dental Research 67, 932-7.
Swift EJ, Perdigao J, Heymann HO (1995) Bonding to enamel and dentine: a brief history and state of the art. Quintessence International 26, 95-110.
Thompson CB (1995) Apoptosis in the pathogenesis and treatment of disease. Science 267, 1456-62.
Tyas MJ (1977) A method for in vitro toxicity testing of dental restorative materials. Journal of Dental Research 56, 1285-90.
Villa P, Kaufmann SH, Earnshaw WC (1997) Caspases and caspase inhibitors. Trends in Biochemical Sciences 22, 388-92.
Vistica DT, Skehan P, Scudiero D, Monks A, Pittman A, Boyd MR (1991) Tetrazoliumbased assays for cellular viability: a critical examination of selected parameters affecting formazan production. Cancer Research 51, 2515-20.
Wataha JC, Hanks CT, Strawn SE, Fat JC (1994) Cytotoxicity of components of resins and other dental restorative materials. Journal of Oral Rehabilitation 21, 453-62.