Cytotoxicity and Oxidative Stress Induction by Root Canal Sealers in Human Periodontal Ligament Fibroblasts: An in vitro Study

Introduction: The aim of the study was to investigate the in vitro cytotoxicity, the profile of cell death, and the level of oxidative stress in human periodontal ligament fibroblasts (HPdLFs) after exposure to selected root canal sealers. Methods and Materials: Freshly mixed or set Endomethasone N (EN), RealSeal (RSEAL), Roeko Seal Automix (RSA), and Sealapex (SP) were incubated with HPdLFs. Fluorescein isothiocyanate (FITC)-annexin V (AnV) and propidium iodide (PI) staining followed by flow cytometry was used to identify the effects of the materials on cell viability and the profile of cell death. 2,7-dichlorodihydrofluorescein diacetate (DCFH-DA) with fluorescence-activated cell sorting was used to determine reactive oxygen species (ROS) formation in HPdLFs. Statistical analyses were performed using one-way ANOVA followed by post hoc tests, and significance was determined at P<0.05. Results: All materials reduced the viability of the cultured cells compared with the controls (P<0.05). Fresh SP and EN, and set RSA generated an increase of necrotic cells (P<0.05), whilst fresh RSEAL and RSA induced an elevation of apoptotic cells (P<0.001). Set RSEAL caused a rise in both apoptotic and necrotic cells compared with the controls (P<0.05). Fresh EN, RSEAL, and SP resulted in increased intracellular ROS generation compared with the negative control (P<0.001), whilst fresh RSA and all set materials were ineffective. Conclusions: This in vitro study showed us the materials tested were characterized by differentiated cytotoxic effects on HPdLFs. The fresh and set forms of sealers were capable of eliciting toxic action, inducing apoptosis and/or necrosis in HPdLFs. The toxic effects of fresh EN, RSEAL, and SP might have been due to the induction of oxidative stress in human periodontal fibroblasts. The cytotoxicity of RSA seemed to be related to the involvement of other mechanisms

Evaluation of cytotoxicity and pH changes generated by various dental pulp capping materials — an in vitro study

Introduction. Various materials are used in direct dental pulp capping method. Their biocompatibility and alkalizing abilities are of primary importance affecting therapeutic effects. The aim of this study was to evaluate and compare the cytotoxicity of various pulp-capping materials on human gingival fibroblasts and investigate the pH changes induced by these materials. Material and methods. Human gingival fibroblasts were cultured with nine direct pulp materials using culture plate inserts. The cytotoxic effects were recorded by using an MTT-based colorimetric assay after 3 and 24 h. In the second part of the experiment, the materials were inserted in dialysis tubes and transferred into plastic vials containing deionized water. The changes of the medium pH were measured after 3 and 24 h. Results. We showed differences in cell viability of gingival fibroblasts after varied time of exposition for the tested materials. Cell viability after 24 h increased for Dycal, Biopulp, and Calcipro, and decreased for Calcipulpe, Angelus, Angelus White, and ProRoot Regular. Cell viability for ProRoot and Life did not change. Non-setting calcium hydroxide preparations followed by the MTA group and setting calcium hydroxide materials produced the highest pH. All the tested materials significantly increased pH (p < 0.0001) at 24 h. Conclusions. Currently used pulp capping materials varied in their cytotoxicity relative to human gingival fibroblasts and their alkalizing capacities. Since most likely pH does not affect the viability of cultured cells, further investigations are required to determine physicochemical properties of these materials and the biological activity of the dental pulp

Cytotoxicity of Methacrylate Dental Resins to Human Gingival Fibroblasts

This study aimed to assess the acute and delayed cytotoxicity of three, popular light-cured methacrylate-based restorative resins (MRs): Charisma (C), Estelite (E), and Filtek (F), to human gingival fibroblasts in culture. Cells were grown for up to 24 h with light-cured (or pre-cured) resins. We evaluated resin cytotoxicity, redox imbalance, necrosis/apoptosis, miR-9, and heat shock protein 70 (HSP70). The role of resin-induced oxidative stress (damage) in HSP70-response (repair) was assessed using binary fluorescence labeling. All MRs decreased viable cell numbers and cell proliferation and damaged cell membranes, and their 24 h-delayed toxicity was lower (C), higher (F), or similar (E) to that induced by freshly-cured resins. Cell membrane damage induced by C and E decreased with time, while F produced a linear increase. All resins generated intracellular oxidative stress with the predominant necrotic outcome, and produced heterogeneous responses in miR-9 and HSP70. The double fluorescence (damage/repair) experiments pointed to common features of E and F but not C. In the subset of cells, the binary response induced by E and F was different from C, similar to each other, and positively interrelated. Experimental data show that selective MR cytotoxicity should be taken into account when considering repetitive use or massive reconstruction