Original Article

Set9, NF-κB, and microRNA-21 mediate berberine-induced apoptosis of human multiple myeloma cells

Authors: Hai-yan Hu, Kun-peng Li, Xiu-ju Wang, Yuan Liu, Zhi-gang Lu, Rui-hong Dong, Hong-bo Guo, Mei-xia Zhang
DOI: 10.1038/aps.2012.161

Abstract

Hai-yan HU1, #, Kun-peng LI4, #, Xiu-ju WANG2, #, Yuan LIU3, Zhi-gang LU3, Rui-hong DONG3, Hong-bo GUO3, *, Mei-xia ZHANG3, *

1Oncology Department of Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai 200233, China; 2Hematology Department of Sun Yat-Sen University, Guangzhou 501170, China; 3Hematology Department of Zhujiang Hospital of Southern Medical University, Guangzhou 510282, China; 4School of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, China

Aim: To investigate the mechanisms by which berberine suppressed the proliferation of human multiple myeloma cells.

Methods: Human U266 multiple myeloma cell line was tested. Cell proliferation, apoptosis, ultramicrostructure and secretion function were examined using Cell Counting Kit-8 (CCK8), flow cytometry (FCM), electron and fluorescence microscopy, as well as ELISA assay. The microRNAs (miRs) and transcription factors in U266 cells were detected using arrays and verified by qRT-PCR. EMSA and luciferase assays were used to verify the p65-dependent transactivation of miR-21 gene.

Results: Treatment of U266 cells with berberine (40−160 µmol/L) suppressed cell proliferation and IL-6 secretion in dose- and time-dependent manners. Meanwhile, berberine dose-dependently induced ROS generation, G2/M phase arrest and apoptosis in U266 cells, and decreased the levels of miR-21 and Bcl-2. Overexpression of miR-21 counteracted berberine-induced suppression of cell proliferation and IL-6 secretion. In U266 cells treated with berberine (80 µmol/L), the activity of NF-κB was decreased by approximately 50%, followed by significant reduction of miR-21 level. berberine (80−160 µmol/L) increased the level of Set9 (lysine methyltransferase) by more than 2-fold, caused methylation of the RelA subunit, which inhibited NF-κB nuclear translocation and miR-21 transcription. In U266 cells treated with berberine (80 µmol/L), knockdown of Set9 with siRNAs significantly increased NF-κB protein level accompanying with a partial recovery of proliferation.

Conclusion: In U266 cells, berberine suppresses NF-κB nuclear translocation via Set9-mediated lysine methylation, leads to decrease in the levels miR21 and Bcl-2, which induces ROS generation and apoptosis.


Keywords: multiple myeloma; berberine; apoptosis; NF-κB; Set9; methylation; RelA; microRNA-21; Bcl-2; ROS

This paper was supported by Guangdong Science and Technology Plan (No 2012A030400012).
# The first three authors contributed equally to this work.
* To whom correspondence should be addressed.
E-mail guohongbo7272@126.com (Hong-bo GUO); meixiaz88@yahoo.com.cn (Mei-xia ZHANG)
Received 2012-07-08 Accepted 2012-10-22
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