Effect of irrigants on the survival of human stem cells of the apical papilla in a platelet-rich plasma scaffold in human root tips

Introduction: Intracanal disinfection is a crucial step in regenerative endodontic procedures. However, this novel endodontic treatment lacks standardization, and numerous treatment protocols have been reported without knowledge of the effect of disinfection protocols on the survival of stem cells. The aim of this study was to test the hypothesis that different root canal irrigation protocols alter survival of stem cells from the apical papilla (SCAP). Methods: SCAP were isolated from immature human third molars, and a subpopulation of STRO-1 expressing cells was selected and expanded in vitro. Standardized human root segments (n = 5/group) were irrigated with 1 of 4 protocols: (1) 17% ethylenediaminetetraacetic acid (EDTA), (2) 6% NaOCl/17% EDTA/6% NaOCl, (3) 17% EDTA/2% chlorhexidine (CHX), or (4) 6% NaOCl/17% EDTA/6% NaOCl/isopropyl alcohol/2% CHX. Subsequently, STRO-1-enriched SCAP were mixed with platelet-rich-plasma, seeded into the root tips, and cultured for 21 days. Roots were then decalcified, processed for immunohistochemistry, and stained for vimentin and TO-PRO-3. The proportion of viable (vimentin-positive) cells was calculated on the basis of the total cell counts (TO-PRO-3) for each group. Results: Irrigation with 17% EDTA best supported cell survival (89% viability; P <.001 versus all other groups), followed by irrigation with 6% NaOCl/17% EDTA/6% NaOCl (74%; P <.001 versus the 2 groups containing 2% CHX). Conversely, protocols that included 2% CHX lacked any viable cells. Conclusions: Collectively, the results suggest that irrigants alone greatly affect the survivability of STRO-1-enriched SCAP within the root canal environment and that inclusion of EDTA in irrigation protocols might be beneficial in regenerative procedures.