Biological Advantages of Calcium Hypochlorite Solution Over Sodium Hypochlorite for Regenerative Endodontic Procedures: An Ex Vivo and In Vitro Study on Human Apical Papilla

Abstract

AIM: Calcium hypochlorite [Ca(OCl)₂] has been proposed as an alternative to sodium hypochlorite (NaOCl) for use as an irrigant. This study aimed to assess morphological changes in human apical papilla (AP) ex vivo, and to evaluate viability, proliferation, chemotaxis and osteogenic differentiation of human apical papilla cells (hAPCs) in vitro following exposure to Ca(OCl)₂, in comparison to NaOCl. METHODOLOGY: Ex vivo, three AP samples per group were exposed to 1.5% Ca(OCl)₂, 1.5% NaOCl or control solutions [17% ethylenediaminetetraacetic acid (EDTA) and saline] for 3 min. The AP sections were stained with haematoxylin and eosin (H&E), Masson's trichrome and Alcian Blue for morphological analysis. In vitro, hAPCs were exposed to Ca(OCl)₂, NaOCl, EDTA or culture medium. Cell viability was assessed with the methyl-thiazole-tetrazolium (MTT) assay; proliferation by bromodeoxyuridine incorporation; chemotaxis by transwell assay; and mineralised nodule formation by alizarin red staining. Data were analysed by two-way ANOVA followed by Tukey's test or by the Kruskal-Wallis and Dunn tests (α = 0.05). RESULTS: Ex vivo, marked loss of both cells and extracellular matrix components was observed in the outer layer of AP samples, particularly in the NaOCl and Ca(OCl)₂ groups, with more severe damage found in the NaOCl samples. Samples treated with EDTA exhibited structural organisation similar to those treated with saline. In vitro, Ca(OCl)₂ induced less cytotoxicity, resulted in the highest proliferation (p < 0.05), but promoted lower chemotaxis than the other irrigants (p < 0.05). EDTA and Ca(OCl)₂ led to greater mineralised nodule formation than the other solutions (p < 0.05). CONCLUSIONS: Ca(OCl)₂ at 1.5% caused less structural damage to AP than NaOCl at the same concentration and had a more favourable influence on the viability, proliferation and osteogenic differentiation of hAPCs. Moreover, it did not impair cell chemotaxis. These findings suggest that Ca(OCl)₂ may offer biological advantages in regenerative endodontic procedures.