Effects of Aralia continentalis Root Extract on Cell Proliferation and Apoptosis in Human Promyelocytic Leukemia HL-60 Cells

  • Lim Hae-Young (Department of Medical Genetics, College of Medicine, Hanyang University) ;
  • Oh Ha-Lim (Department of Medical Genetics, College of Medicine, Hanyang University) ;
  • Lee Chul-Hoon (Department of Medical Genetics, College of Medicine, Hanyang University)
  • Published : 2006.09.01

Abstract

The roots of Aralia continentalis (AC) have been used traditionally in Korean as a folk medicine for anti-inflammation and as an anti-rheumatic. In this study, we report that the ethyl acetate-soluble traction (ACE) of the methanolic extract of AC root inhibited the cell growth of various human cancer cell lines and induced apoptosis of HL-60, human promyelocytic leukemia cells. Its $IC_{50}$ values on growth inhibition were estimated to be $56.3{\mu}g/ml$ on HL-60, $87.2{\mu}g/ml$ on HepG2, $93.2{\mu}g/ml$ on HeLa, $135.5{\mu}g/ml$ on DU-145, and $135.8{\mu}g/ml$ on HT-29 cells. Interestingly, ACE showed no antiproliferative effect on normal lymphocyte cells used as control. Furthermore, nuclear DAPI staining revealed the typical nuclear features of apoptosis in the HL-60 cells exposed to $80{\mu}g/ml$ ACE, and a flow cytometric analysis of the HL-60 cells using propidium iodide showed that the apoptotic cell population increased gradually from 5% at 0 h to 16% at 12 h and 20% at 24 h after treated with $50{\mu}g/ml$ of ACE. TUNEL assay also revealed the apoptotic induction of the HL-60 cells treated with ACE. To obtain further information on the ACE-induced apoptosis, the expression level of certain apoptosis-associated proteins was examined using a Western blot analysis. Treatment of the HL-60 cells with ACE resulted in the activation of caspase-3, and subsequent proteolytic cleavage of poly(ADP-ribose) polymerase (PARP). The above results confirmed that the apoptosis in the HL-60 cells was induced by ACE, and that caspase-3-mediated PARP cleavage was involved in the process.

Keywords

Aralia continentalis;Dokwhal;cell growth inhibition;apoptosis

References

  1. Enari, M., H. Sakahira, H. Yokoyama, K. Okawa, A. Iwamatsu, and S. Nagata. 1998. A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD. Nature 391: 43-50 https://doi.org/10.1038/34112
  2. Han, B. H., E. R. Woo, M. H. Park, and Y. N. Han. 1985. Studies on the anti-inflammatory activity of Aralia continentalis (III). Arch. Pharm. Res. 8: 59-65 https://doi.org/10.1007/BF02912223
  3. Los, M., C. J. Burek, C. Strol, K. Bnedyk, H. Hug, and A. Mackiewicz. 2003. Anticancer drugs of tomorrow: Apoptotic pathways as targets for drug design. Drug Discov. Today 8: 67-77 https://doi.org/10.1016/S1359-6446(02)02563-1
  4. Nunez, G, M. A. Benedict, Y. Hu, and N. Inohara. 1998. Caspases: The proteases of the apoptotic pathway. Oncogene 17: 3237-3245 https://doi.org/10.1038/sj.onc.1202581
  5. Reed, J. C. 2001. Apoptosis regulatory proteins as targets for drug discovery. Trends Mol. Med. 7: 314-318 https://doi.org/10.1016/S1471-4914(01)02026-3
  6. Kerr, J. F. R., A. H. Wyllie, and A. R. Currie. 1972. Apoptosis: A basic biological phenomenon with wide-ranging implication in tissue kinetics. Brit. J. Cancer 26: 239-257 https://doi.org/10.1038/bjc.1972.33
  7. Lim, H., M. Kim, Y. Cho, J. Kim, Y. Lim, and C.-H. Lee. 2004. Inhibition of cell cycle progression and induction of apoptosis in HeLa cells by HY558-1, a novel CDK inhibitor isolated from Penicillium minioluteum F558. J. Microbiol. Biotechnol. 14: 978-984
  8. Suh, N., L. Luyengi, H. H. Fong, A. D. Kinghorn, and J. M. Pezzuto. 1995. Discovery of natural product chemopreventive agents utilizing HL-60 cell differentiation as a model. Anticancer Res. 15: 233-239
  9. Lee, H.-K., I.-H. Lee, J.-S. Yim, Y.-H. Kim, S.-H. Lee, K.-S. Lee, Y.-M. Koo, S.-J. Kim, and B.-C. Jeong. 2005. Purification and characterization of CDMHK, a growth inhibitory molecule against cancer cell lines, from Myxobacterium sp. HK1 isolated from Korean soil. J. Microbiol. Biotechnol. 15: 734-739
  10. Han, B. H., Y. N. Han, K. A. Han, M. H. Park, and E. O. Lee. 1983. Studies on the anti-inflammatory activity of Aralia continentalis (I). Arch. Pharm. Res. 6: 17-23 https://doi.org/10.1007/BF02855697
  11. Kaufman, S. H. and W. C. Earnshaw. 2000. Induction of apoptosis by cancer chemotherapy. Exp. Cell Res. 256: 42-49 https://doi.org/10.1006/excr.2000.4838
  12. Brown, J. M. and B. G. Wouters. 1999. Apoptosis, p53, and tumor cell sensitivity to anticancer agents. Cancer Res. 59: 1391-1399
  13. Lim, T. Y 1987. Comparative study on the pharmacological effect of kinds of Dokhwal. J. Kor. Orient. Med. 8: 97-98
  14. Sebbagh, M., C. Renvoize, J. Hamelin, N. Riche, J. Bertoglio, and J. Breard. 2001. Caspase-3-mediated cleavage of ROCK I induces MLC phosphorylation and apoptotic membrane blebbing. Nat. Cell Biol. 3: 346-352 https://doi.org/10.1038/35070019
  15. Hengartner, M. O. 2000. The biochemistry of apoptosis. Nature 407: 770-776 https://doi.org/10.1038/35037710
  16. Studzinski, G P. 1999. An overview of apoptosis. In G P. Studzinski (ed.), Apoptosis: A Practical Approach. Oxford University Press, Oxford, U.K
  17. Sahara, S., M. Aoto, Y. Eguchi, N. Imamoto, Y. Yoneda, and Y. Tsujimoto. 1999. Acinus is a caspase-3-activated protein required for apoptotic chromatin condensation. Nature 401: 168-173 https://doi.org/10.1038/43678
  18. Kim, M., C. Shin, H. Yang, S. Kim, H. Lim, C.-H. Lee, M. Kim, and Y Lim. 2004. Naltriben analogues as peptide anticancer drugs. J. Microbiol. Biotechnol. 14: 881-884
  19. Dantzer, R., V. Schreiber, C. Niedergang, C. Trucco, E. Flatter, G De La Rubia, J. Oliver, V. Rolli, J. Menissier-de Murcia, and G. de Murcia. 1999. Involvement of poly(ADP-ribose) polymerase in base excision repair. Biochimie 81: 69-75 https://doi.org/10.1016/S0300-9084(99)80040-6
  20. Mosmann, T. 1983. Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays. J. Immunol. Methods 65: 55-63 https://doi.org/10.1016/0022-1759(83)90303-4
  21. Coleman, M. L., E. A. Sahai, M. Yeo, M. Bosch, A. Dewar, and M. F. Olson. 2001. Membrane blebbing during apoptosis results from caspase-mediated activation of ROCK I. Nat. Cell Biol. 3: 339-345 https://doi.org/10.1038/35070009
  22. Kaufman, S. H. 1989. Induction of endonucleolytic DNA cleavage in human acute myelogenous leukemia cells by etoposide, camptothecin and other cytotoxic anticancer drugs: A cautionary note. Cancer Res. 49: 5870-5878
  23. Lim, H., M. Kim, Y. Lim, Y. Cho, and C.-H. Lee. 2006. Inhibition of cell-cycle progression in HeLa cells by HY52, a novel cyclin-dependent kinase inhibitor isolated from Bauhiniaforficata. Cancer Lett. 233: 89-97 https://doi.org/10.1016/j.canlet.2005.03.002
  24. Lee, C.-H., H. Lim, M. Kim, Y. Cho, D. Oh, C. Kim, and Y. Lim. 2002. Isolation and biological properties of novel cell cycle inhibitor, HY558, isolated from Penicillium minioluteum F558. J. Microbiol. Biotechnol. 12: 470-475