Comparison of cytotoxicity, genotoxicity and immunological inflammatory biomarker activity of several endodontic sealers against immortalized human pulp cells.

AIM: To establish an SV40 T-Ag-transfected cell line of human pulp-derived cells in order to compare the cytotoxicity, genotoxicity and to investigate the activities of immunological biomarkers of several endodontic sealers. METHODOLOGY: Primary human pulp cells and transfected cells were cultured. Cell morphology and proliferation were analysed, and the expression of cell-specific gene transcripts and proteins was detected by RT-PCR and immunohistochemistry. Transfection of human pulp-derived cells resulted in an immortalized cell line retaining phenotypic characteristics from the primarily cells tested. The SV40 T-Ag-transfected cells were cultured and stimulated by sealers (Apexit Plus, Real Seal, AH Plus, and EndoREZ) to evaluate the cytotoxicity and genotoxicity by MTT and MTN assays, respectively. Immunological inflammatory biomarkers (IL6, IL8 and TNF-α) were determined by ELISA assay. The differences between median values were statistically analysed using Kruskal-Wallis and Dunn's tests at 5% significance level. RESULTS: The cytotoxicity assay revealed that multimethacrylate (Real Seal) was the most cytotoxic sealer (P < 0.05) and exhibited the highest inflammatory potential against the SV40 T-Ag-transfected cells (P < 0.05). All root canal sealers tested were able to stimulate the immortalized pulp cells to produce IL-6, IL-8 and TNF-α, with differences in relation to the control group (P < 0.05). Higher levels of IL-6, IL-8 and TNF-α were found in cell supernatant after stimulation with multimethacrylate (Real Seal) compared to all other sealers tested (P < 0.05). No differences were found comparing epoxy resin-based sealer (AHPlus), single-methacrylate sealer (EndoREZ) and calcium hydroxide-based sealer (Apexit Plus), regardless of the cytokine investigated (all P > 0.05). CONCLUSIONS: A SV40 T-Ag-transfected cell line of human pulp-derived cells was established. The methacrylate resin-based sealer (Real Seal) exhibited the greatest cytoxicity and inflammatory potential against immortalized pulp cells compared to an epoxy resin-based sealer (AH Plus), a methacrylate-based sealer (EndoRez) and a calcium hydroxide-based sealer (Apexit).

Antimicrobial analysis of chlorhexidine gel and intracanal medicaments against microorganisms inoculated in root canals.

AIM: The aim of this paper was to evaluate the antimicrobial activity of 2% chlorhexidine gel (CLX) associated with various intracanal medicaments against Candida albicans and Enterococcus faecalis inoculated in root canals. METHODS: Thirty six human single-rooted teeth were contaminated with C.albicans and E.faecalis. The canals were instrumented using 2% CLX gel and were divided into three groups according to the intracanal medicaments (ICM) used. Group 1: calcium hydroxide paste [Ca(OH)2], Group 2: 2% chlorhexidine gel (CLX) and Group 3: 2% CLX gel + Ca(OH)2. The root canal collections were performed after 21 days of contamination (control collection), after instrumentation (1st collection), after 14 days of intracanal medicament (2nd collection) and 7 days after medicament removal (3rd collection). The microbiological samples were plated in culture media and incubated for 48 hours. The results were submitted to Kruskal-Wallis test (P ≤ 0.05). RESULTS: It was verified that the instrumentation with CLX reduced the number of CFU/ml significantly when compared with the confirmation collection (control). However, the use of the ICM was only capable to eliminate completely the microorganisms in the root canals without difference statistics between them. CONCLUSION: Although the use of 2% chlorherixidine gel reduces the number of microorganisms significantly, only the ICM calcium hydroxide and calcium hydroxide associated with chlorhexidine are able to eliminate these microorganisms completely.

Cytotoxicity and genotoxicity of pulp capping materials in two cell lines.

AIM: The aim of this study was to evaluate the cytotoxicity and genotoxicity of the new castor oil bean cement (COB) material in comparison to commonly used pulp capping materials. METHODOLOGY: Specimens of COB, calcium hydroxide (Hydro C), and mineral trioxide aggregate (white and gray MTA) were extracted in culture medium (91.6 mm(2) sample surface mL(-1)). Transfected human pulp cells (tHPCs) were exposed to dilutions of the extracts for 1 h, and the generation of reactive oxygen species (ROS) was determined by flow cytometry (FACS) using H(2)DCF-DA as a dye. Survival of tHPCs was measured photometrically using a crystal violet assay after a 24-h exposure period. Genotoxicity as indicated by the formation of micronuclei in V79 cells, and the modification of the normal cell cycle by extracts of the materials was analysed by FACS. RESULTS: Clear cytotoxic effects were detected only with extracts of Hydro C under the current experimental conditions. The two MTA preparations induced an insignificant reduction in the number of cells. In contrast, the extracts of COB slightly induced cell proliferation. Extracts of Hydro C caused a twofold increase in ROS production, whilst the other tested materials were ineffective. An increase in the number of micronuclei was not detected with any material tested; Hydro C slightly increased the number of cells in G1 and G2. CONCLUSIONS: The COB and the two MTA preparations did not negatively influence cell survival or ROS production and may thus be further considered for pulp capping studies.