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Journal of Chinese Integrative Medicine ›› 2010, Vol. 8 ›› Issue (6): 554-561.doi: 10.3736/jcim20100608

• Original Experimental Research • Previous Articles     Next Articles

Effects of extract of Polygonum multiflorum on cell cycle arrest and apoptosis of human liver cell line L02

Rui-chen Zhanga,Bin Liub,Zhen-xiao Suna,Dong-yan Xub   

  1. a Department of Biopharmaceutics, School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing 100102, China
    b Department of Traditional Chinese Medicine Chemistry, School of Chinese Materia Medica, Beijing University of Chinese Medicine,Beijing 100102,China
  • Received:2010-03-01 Accepted:2010-04-12 Online:2010-06-20 Published:2010-06-15
  • Contact: Zhen-xiao Sun E-mail:sunzxcn@hotmail.com

Objective

To analyze the chemical constituents of Polygonum multiflorum extract which may cause human liver cell damage and to explore the mechanism.
Methods

Raw and processed Polygonum multiflorum were extracted by 70% ethanol, then raw and processed Polygonum multiflorum water-eluted material (RW and PW), 50% ethanol-eluted material (R50 and P50) and 95% ethanol-eluted material (R95 and P95) were obtained by absorbing through AB-8 macroporous resin, followed by water, 50% ethanol and 95% ethanol elution in order. The water extracts of raw and processed Polygonum multiflorum (RWE or PWE) were obtained by boiling them in water as usual. Normal human liver L02 cells were treated by different concentrations of eluted Polygonum multiflorum materials for different time, and the cell growth inhibition of each group was determined by methylthiazolyldiphenyl-tetrazolium bromide method. The chemical constituents which had a significant cytotoxicity to L02 cells were analyzed by high-performance liquid chromatography (HPLC). Morphological changes of L02 cells were observed by Giemsa staining and cell cycle distribution was observed by flow cytometry.
Results

It was found that 95% ethanol-eluted extracts of raw and processed Polygonum multiflorum showed significant growth inhibition on normal human liver L02 cells, while the other components showed no significant inhibition on cell growth. HPLC analysis showed that the main component in 95% ethanol-eluted extract of raw and processed Polygonum multiflorum was emodin at content of (18.53±2.96)% and (10.28±1.34)% respectively. Cell cycle analysis showed that 95% ethanol-eluted material of Polygonum multiflorum and emodin had a similar significant effect of S phase arrest and all could induce L02 cell apoptosis.
Conclusion

The main part of Polygonum multiflorum causing liver cell damage is the 95% ethanol-eluted extract, and emodin is one of the important chemical constituents leading to liver cell damage.

Key words: Polygonum multiflorum, Hepatocytes, Cytotoxicity, Liver damage, Chemical constituents, Anthra- quinone, Emodin, Cell cycle

"

Figure 1

Components of R95 and P95 observed by HPLCA: Reference substances; B: R95; C: P95. ①: Aloe-emodin; ②: Rhein; ③: Emodin; ④: Chrysophanol; ⑤: Physcion."

"

Figure 2

Morphological changes of L02 cells after 48-h culture in different groups (Inverted microscopy, ×200) A: Control group; B: 100 μg/mL R95 group; C: 100 μg/mL P95 group; D: 50 μg/mL emodin group. Cells in group A show adjacent cells connecting, all in good shape with uniform refraction of cytoplasm and obvious nucleus. Cells in group B, C and D show margination of nuclear substances around the nuclear envelope and some vacuole-like structure appearing around the nucleus(→), cells shrinking from the surrounding cells(△), cell apoptosis and apoptotic bodies appearing(▲)."

Figure 3

Morphological changes of L02 cells after 48-h culture in different groups observed by Giemsa staining (Inverted microscopy, ×200) A: Control group; B: 100 μg/mL R95 group; C: 100 μg/mL P95 group; D: 50 μg/mL emodin group. Cells are intact in group A with large and clear outline of nucleus, and uniform cytoplasm. Morphology of some L02 cells and their nucleus is abnormal in group B, C and D, showing chromatin margination (→), apoptotic cells(△) and apoptotic bodies(▲)."

Figure 4

Cell cycle distributions of L02 cells after R95, P95 and emodin treatment for 0, 24 and 48 h detected by flow cytometry"

[1] Zhang L, Yang XH, Sun ZX, Qu Y . Retrospective study of adverse events of Polygonum multiflorum and risk control[J]. Zhongguo Zhong Yao Za Zhi, 2009,34(13):1724-1729
张力, 杨晓晖, 孙震晓, 曲毅 . 何首乌临床不良事件回顾性研究及风险控制措施探讨[J]. 中国中药杂志, 2009,34(13):1724-1729
[2] Zhang L, Yang XH, Deng YY . Evaluation and consideration on safety information abroad of Polygonum multiflorum and its preparations[J]. Zhongguo Zhong Yao Za Zhi, 2009,34(18):2414-2418
张力, 杨晓晖, 邓媛瑗 . 何首乌及其制剂国外安全性信息的评价与思考[J]. 中国中药杂志, 2009,34(18):2414-2418
[3] Sun ZX, Zhang L . Liver damage related to Polygonum multiflorum and its preparations: domestic literature review and analysis[J]. Yao Wu Bu Liang Fan Ying Za Zhi, 2010,12(1):26-30
孙震晓, 张力 . 何首乌相关肝损害文献回顾性分析[J]. 药物不良反应杂志, 2010,12(1):26-30
[4] Guo ZY, Jiao Q, Xu MH, Wang ST . An experimental study on the injury of human normal hepatocyte cell line L02 by LPS[J]. Yi Xue Fen Zi Sheng Wu Xue Za Zhi, 2008,5(4):323-327
郭竹英, 矫强, 徐芒华, 王世婷 . 脂多糖对人正常肝细胞株L02损伤的实验研究[J]. 医学分子生物学杂志, 2008,5(4):323-327
[5] Wen JY, Zhang JR, Zhou Y, Li P, Nie Q, Kang JB . Effects of NADH on DENA-induced H-ras gene mutation and expression of c-erbB-2 gene in L02 cells[J]. Zhong Liu, 2009,29(3):210-214
温居一, 张积仁, 周媛, 李鹏, 聂青, 康静波 . NADH对DENA所致L02细胞H-ras基因突变及c-erbB-2表达的影响[J]. 肿瘤, 2009,29(3):210-214
[6] Sun ZX, Zhao TD, Wei YL, Li JS . Study on apoptosis of human hepatoma cells(BEL-7402) induced by norcantharidin[J]. Jie Pou Xue Bao, 1999,30(1):65-68
孙震晓, 赵天德, 魏育林, 李家实 . 去甲斑蝥素诱导人肝癌BEL-7402细胞凋亡的研究[J]. 解剖学报, 1999,30(1):65-68
[7] Zhang ZG, Lu TS, Yao QQ . Studies on anthraquinones in Polygonum multiflorum[J]. Zhong Cao Yao, 2006,37(9):1311-1313
张志国, 吕泰省, 姚庆强 . 何首乌蒽醌类化学成分研究[J]. 中草药, 2006,37(9):1311-1313
[8] Yang DH, Cai SQ, Wang X, Wei LX . Determination of emodin and physcion in capsules of tuber fleeceflower(Polygonum multiflorum) and its complex prescription with cordycep by HPLC[J]. Zhong Cao Yao, 2000,31(3):177-179
杨东辉, 蔡少青, 王璇, 魏璐雪 . HPLC法测定何首乌及复方虫草胶囊中大黄素、大黄素甲醚的含量[J]. 中草药, 2000,31(3):177-179
[9] Sun ZX, Wei YL, Zhao TD, Li JS . Studies on apoptotic cytology of K562 human myeloid leukemia cells induced by cantharidin and norcantharidin[J]. Jie Pou Xue Bao, 2000,3(1):56-60
孙震晓, 魏育林, 赵天德, 李家实 . 斑蝥素及去甲斑蝥素诱导人红白血病K562细胞凋亡的细胞学研究[J]. 解剖学报, 2000,3(1):56-60
[10] Zhan YT, Li DG, Wei HS, Wang ZR, Huang X, Li ZD, Xu QF, Lu HM . Effect of emodin on development of hepatic fibrosis in rats[J]. Zhongguo Zhong Xi Yi Jie He Za Zhi, 2000,20(4):276-278
展玉涛, 李定国, 魏红山, 王志荣, 黄新, 李兆东, 徐芹芳, 陆汉明 . 大黄素对大鼠肝纤维化形成的影响[J]. 中国中西医结合杂志, 2000,20(4):276-278
[11] Zhang X, Dang SS, Cheng YA, Jia XL, Chen MX, Liu EQ . Effects of emodin and Astragalus polysaccharides on hepatic ultrastructure in rat hepatic injury[J]. Xi’an Jiao Tong Da Xue Xue Bao Yi Xue Ban, 2009,30(4):502-505
张欣, 党双锁, 程延安, 贾晓黎, 陈明霞, 刘恩岐 . 大黄素和黄芪多糖对实验性肝损伤大鼠肝脏超微结构的影响[J]. 西安交通大学学报(医学版), 2009,30(4):502-505
[12] Oshima T, Hirayama F, Saitou F, Sagara K, Wang K, Tong YY . The analysis of anthraquinones in Polygonum multiflorum by HPLC[J]. Yao Wu Fen Xi Za Zhi, 1996,16(4):219-222
大岛俊幸, 平山总良, 齐藤文孝, 相乐和彦, 王珂, 童玉懿 . 高效液相色谱法测定何首乌和夜交藤中蒽醌类成分的含量[J]. 药物分析杂志, 1996,16(4):219-222
[13] Ren LJ, Jin Y, Yin SY . Influence of process methods on contents of chemical component in Radix Polygoni Multiflori[J]. Shi Zhen Guo Yi Guo Yao, 2007,18(3):637-638
任丽建, 金勇, 尹寿玉 . 炮制方法对何首乌有效成分影响的研究[J]. 时珍国医国药, 2007,18(3):637-638
[14] Wang XH, Zhen YS . Inhibitory effect of emodin on metastasis associated properties of PG cells from a human highly metastatic giant lung carcinoma cell line[J]. Ai Zheng, 2001,20(8):789-793
王心华, 甄永苏 . 大黄素抑制人高转移巨细胞肺癌PG细胞的肿瘤转移相关性质[J]. 癌症, 2001,20(8):789-793
[15] Liao ZJ, Song YC, Chen XQ . Study on the effect of emodin on the cell proliferation periods of human lung adenocarcinoma cell line A-549[J]. Zhonghua Zhong Liu Fang Zhi Za Zhi, 2007,14(5):342-344
廖子君, 宋永春, 陈晓泉 . 大黄素对人肺腺癌A-549细胞增殖周期影响的实验研究[J]. 中华肿瘤防治杂志, 2007,14(5):342-344
[16] Zhang L, Lau YK, Xi L, Hong RL, Kim DS, Chen CF, Hortobagyi GN, Chang C, Hung MC . Tyrosine kinase inhibitors, emodin and its derivative repress HER-2/neu-induced cellular transformation and metastasis-associated properties[J]. Oncogene, 1998,16(22):2855-2863
doi: 10.1038/sj.onc.1201813
[17] Battistutta R, Sarno S, De Moliner E, Papinutto E, Zanotti G, Pinna LA . The replacement of ATP by the competitive inhibitor emodin induces conformational modifications in the catalytic site of protein kinase CK2[J]. J Biol Chem, 2000,275(38):29618-29622
doi: 10.1074/jbc.M004257200
[18] Kumar A, Dhawan S, Aggarwal BB . Emodin(3-methyl-1,6-trihydroxyanthraquinone) inhibits TNF-induced NF-kappaB activation, IkappaB degradation, and expression of cell surface adhesion proteins in human vascular endothelial cells[J]. Oncogene, 1998,17(7):913-918
doi: 10.1038/sj.onc.1201998
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