Chang Xiao-Gang, Ji Ou, Yao Hao, Zhuang Yun, Dong Wen, Lin Lin, Shen Qun
Department of Hematology, Affiliated Hospital, Nanjing University of Traditional Chinese Medicine, Nanjing 210029, Jiangsu Province, China.
Department of Hematology, Affiliated Hospital, Nanjing University of Traditional Chinese Medicine, Nanjing 210029, Jiangsu Province, China,E-mail:
Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2019 Apr;27(2):458-463. doi: 10.19746/j.cnki.issn.1009-2137.2019.02.024.
To investigate the effects of oridonin (ORI) on the proliferation and apoptosis of human multiple myeloma cell line H929 and its possible mechanism.
H929 cells were exposed to ORI 0、4、8、12、16、20、24、28、32 μmol/L for 12, 24 and 36 hours respectively. The prolifcration inhibitory effect of ORI on H929 cells was determined by MTT assay and then the working concentrations of ORI were determined. The morphological changes and apoptosis of H929 cells were observed by TUNEL (TdT-mediated dUTP Nick-End Labeling) and fluorescence microscopy. The apoptosis rate of H929 cells was detected by flow cytometry with Annexin V-FITC/PI staining. The protein expressions of pro-caspase-3, BCL-2,p-PI3K, p-Akt, BAX, Cleaved PARP and p-JNK, p-ERK and p-p38 in H929 cells were detected by Western blot.
Compared with the control group, the proliferation of H929 cells treated with the ORI of 8-16 μmol/L was significantly inhibited and the apoptosis of H929 cells was obviously increased in dose- and time-dependent manners. As for morphological changes, the characteristics of apoptotic cells were presented in H929 cells treated with ORI for 24 hours. The protein levels of pro-caspase-3, BCL-2,p-PI3K, p-Akt were down-regulated with increasing of ORI concentration(r=0.9861, r=0.9725, r=0.9413, r=0.9373), while the BAX, Cleaved PARP and p-JNK, p-ERK and p-p38 were up-regulated(r=0.9178, r=0.8877, r=0.882, r=0.9645, r=0.8623).
The ORI possesses anti-myeloma effects, can inhibit the proliferation and induce the apoptosis of H929 cell line in vitro. Its potential mechanism may be related with up-regulating the MAPK and down-regulating the PI3K/Akt signal pathways.
探讨冬凌草甲素(ORI)对人多发性骨髓瘤细胞系H929增殖和凋亡的影响及其可能机制。
将H929细胞分别暴露于0、4、8、12、16、20、24、28、32 μmol/L的ORI中12、24和36小时。采用MTT法检测ORI对H929细胞的增殖抑制作用,进而确定ORI的工作浓度。通过TUNEL(末端脱氧核苷酸转移酶介导的dUTP缺口末端标记法)和荧光显微镜观察H929细胞的形态变化和凋亡情况。采用Annexin V-FITC/PI染色,通过流式细胞术检测H929细胞的凋亡率。采用蛋白质印迹法检测H929细胞中procaspase-3、BCL-2、p-PI3K、p-Akt、BAX、Cleaved PARP以及p-JNK、p-ERK和p-p38的蛋白表达。
与对照组相比,8-16 μmol/L的ORI处理后,H929细胞的增殖受到显著抑制,且H929细胞的凋亡以剂量和时间依赖性方式明显增加。就形态学变化而言,用ORI处理24小时的H929细胞呈现出凋亡细胞的特征。随着ORI浓度升高,procaspase-3、BCL-2、p-PI3K、p-Akt的蛋白水平下调(r = 0.9861,r = 0.9725,r = 0.9413,r = 0.9373),而BAX、Cleaved PARP以及p-JNK、p-ERK和p-p38上调(r = 0.9178,r = 0.8877,r = 0.882,r = 0.9645,r = 0.8623)。
冬凌草甲素具有抗骨髓瘤作用,能在体外抑制H929细胞系的增殖并诱导其凋亡。其潜在机制可能与上调MAPK和下调PI3K/Akt信号通路有关。