# In Vitro Biocompatibility of Calcium Silicate-Based Materials for Retrograde Endodontic Treatment Under Different Setting Conditions

## TL;DR

This study examines how different setting conditions affect the biocompatibility of calcium silicate-based dental materials using human fibroblasts.

## Contribution

The study introduces a novel in vitro model to assess how setting environments influence the biocompatibility of retrograde endodontic materials.

## Key findings

- Cell viability varied depending on the setting environment and eluate maturation time.
- Biodentine showed the most consistent and stable cellular response across all conditions.
- Materials set in blood had cell viability comparable to those set in PBS or dry conditions.

## Abstract

Background: Calcium silicate-based materials are widely used in retrograde endodontic treatment due to their bioactivity and favorable biological properties. The environmental conditions during setting and the time-dependent release of soluble components may influence cellular responses; however, these factors remain insufficiently investigated. Aim: This in vitro study evaluated the cellular response to three calcium silicate-based materials—MTA+, Biodentine, and NeoPUTTY—after setting under different environmental conditions. Materials and Methods: Cylindrical specimens were allowed to set under three conditions: dry environment, phosphate-buffered saline (PBS), and human blood. Eluates obtained after 1, 3, and 5 days were applied to human BJ fibroblasts. Cell viability, based on metabolic activity measured using the AlamarBlue assay, was evaluated at 48 and 96 h. Biocompatibility was inferred from cell viability, reflecting eluate-mediated effects rather than direct material–cell contact. Results: Cell viability was influenced by both the setting environment and eluate maturation time. PBS-set materials showed variable cellular responses, with high viability at early time points but marked decreases at 96 h for some MTA+ and NeoPUTTY groups. Biodentine demonstrated the most stable cellular response across all conditions. Materials set in blood produced cellular responses comparable to those observed for PBS and dry conditions, with no statistically significant overall reduction in cell viability. Conclusions: Within the limitations of this in vitro eluate-based model, blood exposure during setting had a minimal influence on the cell viability to the tested materials. Among the evaluated materials, Biodentine exhibited the most stable biological profile. These findings reflect time-dependent, eluate-mediated cellular effects and should be interpreted with caution when extrapolating to clinical conditions.