In-Vitro Safety Evaluation of Sodium Hypochlorite (NaOCI) as Part of Step 2 and Maintenance Therapy Protocols in Patients with Periodontitis Stages III-IV

Purpose: Since NaOCl acts as a strong oxidizing agent and presents potential toxicity, this study was adressed to evaluate the in-vitro safety of NaOCl solutions at concentrations below the limit of patient tolerance, i.e. >= 0.5%. Materials and Methods: First, an in-silico evaluation was conducted to predict the potential toxicity of NaOCl in terms of mutagenic, tumorigenic, irritant, and reproductive risks, as well as some drug-like properties of the molecule. The in-vitro experiments were based on 2D and 3D models. For the 2D approach, two selected cell lines -HaCaT (human skin kerati-nocytes) and HGF (human gingival fibroblasts) -were exposed to NaOCl at five concentrations (0.05-0.5%) for 10, 30, and 60s to simulate possible clinical administration. The irritative potential of NaOCl 0.05% and 0.25% was assessed in a 3D in-vitro model (EpiDerm, reconstructed human epidermis). Statistical significance was set at p<0.05. Results: The main findings suggest that NaOCl exerts cytotoxicity towards HaCaT immortalised keratinocytes and HGF primary gingival fibroblasts in a cell type-, dose-and time-dependent manner, with the most prominent effect being re-corded in HaCaT cells after 60s of treatment with NaOCl 0.5%. However, NaOCl was computationally predicted as free of mutagenic, tumorigenic, irritant, and reproductive toxicity, and showed no irritative potential in 3D reconstructed epider-mis at concentrations of 0.05% and 0.25%. Conclusion: Further clinical and histological studies are required to confirm these results, as well as elucidate the poten-tial cytotoxic mechanism induced by NaOCl in HaCaT and HGF cells at the tested concentrations.