X. Zhang
2015
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Journal
Academic Journal of Chinese PLA Medical School
Abstract
Objective To investigate the suppression of mibefradil(a kind of T-type calcium channels retardant) on the proliferation of neural stem cell in vitro and clarify the role of T-type calcium channels in the proliferation of neural stem cells indirectly. Methods Adult mice brain injury model was established and neural stem cell was separated from subventricular zone(SVZ). The proliferation and growth after adding 0, 1.25, 2.5, 5, 10 and 20μmol/L of mibefradil in culture medium were observed and nestin staining was performed. The cells were subcultured and differentiated, and NSE and GFAP immunofluorescence staining were performed in differentiated cells. The effect of mibefradil on cell proliferation was tested by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-Htetrazolium bromide(MTT) and neural stem cell sphere counting. Then 50% inhibiting concentration(IC50) was calculated and IC50 of mibefradil was added in culture medium and neural stem cells were divided into two groups: control group and experimental group. The expression of Cyclin A was detected by western blot. Results The number of neural stem cell sphere decreased after adding mibefradil. MTT showed that the proliferation of cells could be inhibited significantly when mibefradil was added with the concentration over 5μmol/L. The OD value in 5μmol/L group, 10μmol/L group and 20μmol/L was significantly lower than that in control group. The IC50 rate of mibefradil to neural stem cells was 8.93μmol/L. The expression of Cyclin A protein in experimental group decreased significantly compared to control group. Conclusion Mibefradil can suppress the proliferation of neural stem cell. The mechanism is related to inhibit cell cycle progression.