NAC Rescues Dental Pulp Microtissues Affected by Toxic Resin Monomer

Objectives: Dental pulp is exposed to resin monomers leaching from adhesive capping materials. Toxic doses of the monomer triethylene glycol dimethacrylate (TEGDMA) impact cell structure, enhance inflammatory and oxidative stress responses in the tissue and, ultimately, lead to tissue necrosis and tooth loss. An anti-inflammatory and antioxidative agent is required to rescue the affected cells and modulate their responses. Objective of this study was to assess the functionality of N-Acetyl Cysteine (NAC) as tissue rescue agent by employing our 3D dental pulp micro-tissue platform.
Methods: 3D immortalized/primary dental pulp microtissues developed and matured in 14 d in extracellular matrix (ECM) to resemble native conditions. TEGDMA was introduced at 0 (control), 1.5 or 2.5 mmol/L. NAC (10 mmol/L) was administered simultaneously with TEGDMA, and 7 d before or after TEGDMA addition during the development and after the maturation stages of the microtissue. Cultures without NAC were used as a control. Spatial growth was validated by confocal microscopy and image processing. Levels of inflammatory (Cox2, NLRP3, IL-8) and oxidative stress (GSH) markers were quantified by immunocytochemistry and ELISA (n=3/experimental group).
Results: NAC treatments, in parallel with TEGDMA challenge or 7d post-challenge, enhanced growth of the underdeveloped microtissues and increased their thicknesses (2.5 and 2.0 folds, respectively) and protected mature microtissues from deterioration (42 µm decrease in thickness). Pre-treatment with NAC was ineffective. Recovery effects correlated with the modulation of both responses. Levels of Cox2, NLRP3, IL-8 and GSH decreased 6, 4.7, 6.4 and 16 fold, respectively. Secreted levels of IL-8 decreased by more than 130 pg/mL.
Conclusions: Our 3D/ECM-based dental pulp platform is an efficient tool for drug rescue screening process. NAC supported microtissues development by immunomodulating and maintaining the oxidative balance. This study encourages the involvement of NAC in endodontic therapy and its integration into dental restoratives to maintain pulp vitality.