Effect of different concentrations of hydrogen peroxide on the expression of autophagy-related genes in human dental pulp stem cells

Abstract

Autophagy is a vital pathway required for cell survival during starvation through autophagy and destruction of cell organelles. Considering the role of hydrogen peroxide in the autophagy process, the present study investigated the effect of H2O2 as an oxidative agent in initiating the autophagy process in dental pulp stem cells.

Materials and Method: Dental pulp cells with cell line DPS_7 and cell bank code IBRCC10371 were purchased from the Genetic Reserve Center. Stem cells were treated with different concentrations of H2O2 and their survival percentage was checked using MTT test. In order to measure the expression level of genes related to autophagy, the cells were treated for 24 hours with doses lower than the IC50 of hydrogen peroxide, and the total RNA of the cells was extracted using the RNA extraction kit based on the protocol of the manufacturer of the kit. Then RNA was converted into complementary DNA (cDNA) using cDNA synthesis kit. Finally, the expression level of autophagy genes (LC3, Beclin-1, p62) was investigated using Real time PCR technique. All tests were repeated three times and the group that did not receive H2O2 was considered as the control group. All the results obtained from the treated groups were analyzed with the control group by SPSS 24 software.

Results: The viability of pulp stem cells was significantly changed by increasing the concentration of hydrogen peroxide, so that one molar concentration of hydrogen peroxide with 22.5% cell viability was considered as a toxic concentration. Increasing hydrogen peroxide concentration decreased the expression of P62 gene and increased the expression of Becline-1 and LC3 genes. Conclusion:

H2O2, as an oxidative agent, stimulated the autophagy process in pulp stem cells.