White mineral trioxide aggregate induces migration and proliferation of stem cells from the apical papilla

Abstract

Introduction: Regenerative endodontic protocols recommend white mineral trioxide aggregate (WMTA) as a capping material because of its osteoinductive properties. Stem cells from the apical papilla (SCAP) are presumed to be involved in this regenerative process, but the effects of WMTA on SCAP are largely unknown. Our hypothesis was that WMTA induces proliferation and migration of SCAP.

Methods: Here we used an unsorted population of SCAP (passages 3-5) characterized by high CD24, CD146, and Stro-1 expression. The effect of WMTA on SCAP migration was assessed by using transwells, and its effect on proliferation was determined by the WST-1 assay. Fetal bovine serum (FBS) and calcium chloride-enriched medium were used as positive controls.

Results: The SCAP analyzed here showed a low percentage of STRO-1+ and CD24+ cells. Both set and unset WMTA significantly increased the short-term migration of SCAP after 6 hours (P < .05), whereas calcium chloride-enriched medium did after 24 hours of exposure. Set WMTA significantly increased proliferation on days 1-5, whereas calcium-enriched medium showed a significant increase on day 7, with a significant reduction on proliferation afterwards. SCAP migration and proliferation were significantly and steadily induced by the presence of 2% and 10% FBS.

Conclusions: Collectively, these data demonstrate that WMTA induced an early short-term migration and proliferation of a mixed population of stem cells from apical papilla as compared with a later and longer-term induction by calcium chloride or FBS.

Keywords: Calcium; MTA; SCAP; chemotaxis; dental; stem cells.

Conflict of interest statement

The authors declare no conflicts of interest related to this study.

Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Figures

Figure 1. Morphology, proliferation and characterization of SCAP

(A) Microphotographs showing SCAP at passage 4 after 7 days of culture in plain α-MEM, 2% FBS α-MEM plus 24 h set WMTA, 2% and 10% FBS α-MEM. Phase contrast microphotographs at 100X. FBS induces proliferation. (B) 2% and 10% FBS had a significant effect on SCAP proliferation as analyzed by WST-1. Stem cell markers decrease on SCAP after passage 4.(C) Flow-cytometry data expressed in percentage of the total cell population expressing CD24, CD146, and STRO-1 markers at passage 2, 4, and 10 in culture with 10% FBS α-MEM.

Figure 2. WMTA induces short-term migration of SCAP

(A) SCAP migration in plain α-MEM. The SCAP migration started after 3 hours and got a maximum level and significance at 6 h for all groups exposed to WMTA as compared to the plain α-MEM group (*P≤ 0.05). The migration was then gradually reduced from 12 to 48 h of exposure to WMTA and significantly induced in the calcium-chloride (0.3 and 0.03mmol) enriched groups (*P≤ 0.05). (B) SCAP migration in α- MEM with 2% or 10% FBS. The migration was induced after 12 h of exposure to α-MEM with 2% or 10% FBS and 1 or 24 h set WMTA in 2% FBS as compared with plain α-MEM groups (* P≤ 0.05). (C) Migration at 6 hours. (D) Migration at 24 hours. There was statistically significant difference when compared to the plain α-MEM group (* P≤ 0.05).

Figure 3. WMTA induces short-term proliferation of SCAP

(A) SCAP Proliferation in plain α-MEM from 1 to 14 days analyzed by WST-1 assay. Cells growing in plain α-MEM did no show any significant increase in proliferation after 14 days. 24 h set WMTA in plain α-MEM induced proliferation up to 5 days of exposure, but gradually was reduced to the level of plain α-MEM groups. 1 h set WMTA only induced proliferation after the first day and then the proliferation was below the levels of the plain α-MEM group. The calcium-chloride (0.3 and 0.03mmol) enriched groups induced proliferation at 7 days and gradually reduced to the level of the plain α-MEM group (* P≤ 0.05). (B) SCAP proliferation in plain α-MEM at 1, 5 and 7 days. There was SS difference in proliferation when the groups were compared to the plain α-MEM (* P≤ 0.05). (C) WMTA did not induce significant proliferation on SCAP grown in α-MEM with 2% FBS. There is SS difference when the cells were exposed to any of the conditions in 2% or 10% FBS when compared to plain α-MEM (* P≤ 0.05). Cells grown in WMTA in 2% FBS did no show induction in proliferation as compared to the 2% FBS α-MEM group. (D) SCAP proliferation in α-MEM with 2% or 10% FBS at 1, 5, 7 days. There was SS difference in proliferation when the groups were compared to the plain α-MEM groups (* P≤ 0.05).