Side Population Increase after Simulated Transient Ischemia in Human Dental Pulp Cell

Abstract 

Introduction

Dental pulp is often exposed to ischemia in case of injury or inflammation because of narrow vascular openings at the apex and poor blood circulation in dental pulp tissue. Resident stem cell populations are thought to contribute to the postischemic regeneration process. The aim of this study was to investigate the influence of simulated ischemia (serum deprivation and hypoxia) on side population (SP) stem cells of human dental pulp cells in order to provide a better understanding of the postischemic tissue repair and regeneration process.

Methods

The proliferation of dental pulp cells (DPCs) after exposure to ischemic culture conditions (2% O₂, 2% serum) for 24 hours and 48 hours was investigated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay. The SP fraction was detected by Hoechst 33342 fluorescence flow cytometry, and the expression of SP marker ABCG2 was investigated by immunofluorescence. ABCG2 and OCT4 messenger RNA levels before and after transient ischemia were determined by real-time polymerase chain reaction.

Results

Proliferation rate of DPCs was lower in 24- and 48-hour ischemic groups than control from day 5 to day 7. SP proportion was significantly higher 24 and 48 hours after simulated ischemic treatment, and immunofluorescence staining of ABCG2 also verified the increasing trend of side population. ABCG2 and OCT4 messenger RNA levels increased more than three folds in 48 hours ischemic group compared with control group.

Conclusions

Side population in dental pulp cells increase notably after transient simulated ischemic culture, suggesting that SP may participate in post-ischemic repair and regeneration process of dental pulp.

Key Words: ABCG2, dental pulp cell, ischemia, side population