Triptolide induces cell-cycle arrest and apoptosis of human multiple myeloma cells in vitro via altering expression of histone demethylase LSD1 and JMJD2B
Abstract
Aim: To elucidate the relationship between triptolide-induced changes in histone methylation and its antitumor effect on human multiple myeloma (MM) cells in vitro.
Methods: Human multiple myeloma cell line RPMI8226 was used. Apoptosis was evaluated using Annexin-V-FITC/PI-labeled flow cytometry, Hoechst 33258 staining, and transmission electron microscopy. Flow cytometry was used to detect the cell cycle distribution of the apoptotic cells. The presence of the LSD1, JMJD2B, H3K4me2, H3K9me2, and H3K36me2 proteins was verified by Western blot analysis. Semi-quantitative real-time PCR was performed to examine the expression of LSD1 and JMJD2B.
Results: Triptolide (10–160 nmol/L) suppressed the proliferation of MM cells in a dose- and time-dependent manner with an IC50 value of 99.2±9.0 nmol/L at 24 h. Triptolide (50 nmol/L) induced G0/G1 cell cycle arrest in MM cells. The agent (50–150 nmol/L) induced apoptosis of MM cells in a dose-dependent manner. The same concentrations of triptolide suppressed the expression of dimethylated H3K4, dimethylated H3K9 and dimethylated H3K36 by altering the expression of histone demethylase LSD1 and JMJD2B without affecting the expression of histone demethylase LSD1.
Conclusion: Triptolide potently inhibits the growth of MM cells via regulating the expression of histone demethylase LSD1 and JMJD2B, which lead to abnormal histone methylation.
Keywords:
Methods: Human multiple myeloma cell line RPMI8226 was used. Apoptosis was evaluated using Annexin-V-FITC/PI-labeled flow cytometry, Hoechst 33258 staining, and transmission electron microscopy. Flow cytometry was used to detect the cell cycle distribution of the apoptotic cells. The presence of the LSD1, JMJD2B, H3K4me2, H3K9me2, and H3K36me2 proteins was verified by Western blot analysis. Semi-quantitative real-time PCR was performed to examine the expression of LSD1 and JMJD2B.
Results: Triptolide (10–160 nmol/L) suppressed the proliferation of MM cells in a dose- and time-dependent manner with an IC50 value of 99.2±9.0 nmol/L at 24 h. Triptolide (50 nmol/L) induced G0/G1 cell cycle arrest in MM cells. The agent (50–150 nmol/L) induced apoptosis of MM cells in a dose-dependent manner. The same concentrations of triptolide suppressed the expression of dimethylated H3K4, dimethylated H3K9 and dimethylated H3K36 by altering the expression of histone demethylase LSD1 and JMJD2B without affecting the expression of histone demethylase LSD1.
Conclusion: Triptolide potently inhibits the growth of MM cells via regulating the expression of histone demethylase LSD1 and JMJD2B, which lead to abnormal histone methylation.