Skip to main content

Diosgenin induces cell cycle arrest and apoptosis in human leukemia K562 cells with the disruption of Ca2+ homeostasis

Cancer Chemotherapy and Pharmacology volume 55, pages 79–90 (2005)Cite this article

Abstract

Purpose

Diosgenin is a steroidal sapogenin with estrogenic and antitumor properties. In order to elucidate the mechanism of its antiproliferative activity, we investigated its effects on the cell cycle and apoptosis in human chronic myelogenous leukemia K562 cells.

Methods

Cell viability was assessed via an MTT assay. Apoptosis was investigated in terms of nuclear morphology, DNA fragmentation, and phosphatidylserine externalization. Cell cycle analysis was performed via PI staining and flow cytometry (FCM). Western blotting and immunofluorescence methods were used to determine the levels of p53, cell cycle-related proteins and Bcl-2 family members. FCM was also used to estimate the changes in mitochondrial membrane potential (MMP), intracellular Ca2+ concentration and reactive oxygen species (ROS) generation.

Results

Cell cycle analysis showed that diosgenin caused G2/M arrest independently of p53. The levels of cyclin B1 and p21Cip1/Waf1 were decreased, whereas cdc2 levels were increased. Subsequent apoptosis was demonstrated with the dramatic activation of caspase-3. A dramatic decline in intracellular Ca2+ concentration was observed as an initiating event in the process of cell cycle arrest and apoptosis, which was followed by the hyperpolarization and depolarization of MMP. Generation of ROS was observed in the progression of apoptosis. The antiapoptotic Bcl-2 and Bcl-xL proteins were downregulated, whereas the proapoptotic Bax was upregulated.

Conclusions

Diosgenin inhibits K562 cell proliferation via cell cycle G2/M arrest and apoptosis, with disruption of Ca2+ homeostasis and mitochondrial dysfunction playing vital roles.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

43,34 €

Price includes VAT (Finland)
Tax calculation will be finalised during checkout.

Fig. 1
Fig. 2a, b
Fig. 3a–f
Fig. 4a, b
Fig. 5a–c
Fig. 6a, b
Fig. 7
Fig. 8
Fig. 9a, b
Fig. 10a, b

References

  1. Aradhana, Rao AR, Kale RK (1992) Diosgenin—a growth stimulator of mammary gland of ovariectomized mouse. Indian J Exp Biol 30:367

    CAS  PubMed  Google Scholar 

  2. Arion D, Meijer L, Brizuela L, Beach D (1988) cdc2 is a component of the M phase-specific histone H1 kinase: evidence for identity with MPF. Cell 55:371

    CAS  PubMed  Google Scholar 

  3. Attele AS, Wu JA, Yuan CS (1999) Ginseng pharmacology: multiple constituents and multiple actions. Biochem Pharmacol 58:1685

    CAS  PubMed  Google Scholar 

  4. Beneytout JL, Nappez C, Leboutet MJ, Malinvaud G (1995) A plant steroid, diosgenin, a new megakaryocytic differentiation inducer of HEL cells. Biochem Biophys Res Commun 207:398

    CAS  PubMed  Google Scholar 

  5. Corbiere C, Liagre B, Bianchi A, Bordji K, Dauca M, Netter P, Beneytout JL (2003) Different contribution of apoptosis to the antiproliferative effects of diosgenin and other plant steroids, hecogenin and tigogenin, on human 1547 osteosarcoma cells. Int J Oncol 22:899

    CAS  PubMed  Google Scholar 

  6. Cory S, Adams JM (2002) The Bcl-2 family: regulators of the cellular life-or-death switch. Nat Rev Cancer 2:647

    CAS  PubMed  Google Scholar 

  7. Crompton M (1999) The mitochondrial permeability transition pore and its role in cell death. Biochem J 34:1233

    Google Scholar 

  8. Desagher S, Martinou JC (2000) Mitochondria as the central control point of apoptosis. Trends Cell Biol 10:369

    CAS  PubMed  Google Scholar 

  9. Donato NJ, Perez M (1998) Tumor necrosis factor-induced apoptosis stimulates p53 accumulation and p21WAF1 proteolysis in ME-180 cells. J Biol Chem 273:5067

    CAS  PubMed  Google Scholar 

  10. Duchen MR (1999) Contributions of mitochondria to animal physiology: from homeostatic sensor to calcium signaling and cell death. J Physiol 516:1

    CAS  PubMed  Google Scholar 

  11. Elledge SJ (1996) Cell cycle checkpoints: preventing an identity crisis. Science 274:1664

    Article  CAS  PubMed  Google Scholar 

  12. Gervais JL, Seth P, Zhang H (1998) Cleavage of CDK inhibitor p21(Cip1/Waf1) by caspases is an early event during DNA damage-induced apoptosis. J Biol Chem 273:19207

    CAS  PubMed  Google Scholar 

  13. Green DR, Reed JC (1998) Mitochondria and apoptosis. Science 281:1309

    Article  CAS  PubMed  Google Scholar 

  14. Gross A, McDonnell JM, Korsmeyer SJ (1999) BCL-2 family members and the mitochondria in apoptosis. Genes Dev 13:1899

    CAS  PubMed  Google Scholar 

  15. Hu K, Dong A, Yao X, Kobayashi H, Iwasaki S (1996) Antineoplastic agents. I. Three spirostanol glycosides from rhizomes of Dioscorea collettii var. hypoglauca. Planta Med 62:573

    CAS  PubMed  Google Scholar 

  16. Innocente SA, Abrahamson JL, Cogswell JP, Lee JM (1999) p53 regulates a G2 checkpoint through cyclin B1. Proc Natl Acad Sci U S A 96:2147

    CAS  PubMed  Google Scholar 

  17. Jiang X, Wang X (2000) Cytochrome c promotes caspase-9 activation by inducing nucleotide binding to Apaf-1. J Biol Chem 275:31199

    CAS  PubMed  Google Scholar 

  18. Johnstone RW, Ruefli AA, Lowe SW (2002) Apoptosis: a link between cancer genetics and chemotherapy. Cell 108:153

    CAS  PubMed  Google Scholar 

  19. Jordan MA, Wendell K, Gardiner S, Derry WB, Copp H, Wilson L (1996) Mitotic block induced in HeLa cells by low concentrations of paclitaxel (Taxol) results in abnormal mitotic exit and apoptotic cell death. Cancer Res 56:816

    CAS  PubMed  Google Scholar 

  20. Kroemer G, Dallaporta B, Resche-Rigon M (1998) The mitochondrial death/life regulation of apoptosis and necrosis. Annu Rev Physiol 60:619

    Article  CAS  PubMed  Google Scholar 

  21. Li PF, Dietz R, von Harsdorf R (1999) p53 regulates mitochondrial membrane potential through reactive oxygen species and induces cytochrome c independent apoptosis blocked by bcl-2. EMBO J 18:6027

    Article  CAS  PubMed  Google Scholar 

  22. Lubbert M, Miller CW, Crawford L, Koeffler HP (1988) p53 in chronic myelogenous leukemia, study of mechanisms of differential expression. J Exp Med 167:873

    Article  CAS  PubMed  Google Scholar 

  23. Matsuyama S, Lopis J, Deveraux QL, Tsien RY, Reed JC (2000) Changes in intramitochondrial and cytosolic pH: early events that modulate caspase activation during apoptosis. Nat Cell Biol 2:318

    Article  CAS  PubMed  Google Scholar 

  24. Minn AJ, Boise LH, Thompson CB (1996) Expression of bcl-x(L) and loss of p53 can cooperate to overcome a cell cycle checkpoint induced by mitotic spindle damage. Genes Dev 10:12621

    Google Scholar 

  25. Moalic S, Liagre B, Corbiere C, Bianchi A, Dauca M, Bordji K, Beneytout JL (2001) A plant steroid, diosgenin, induces apoptosis, cell cycle arrest and COX activity in osteosarcoma cells. FEBS Lett 506:225

    Article  CAS  PubMed  Google Scholar 

  26. Nappez C, Liagre B, Beneytout JL (1995) Changes in lipoxygenase activities in human erythroleukemia (HEL) cells during diosgenin-induced differentiation. Cancer Lett 96:133

    Article  CAS  PubMed  Google Scholar 

  27. Nutt LK, Pataer A, Pahler J, Fang B, Roth J, McConkey DJ, Swisher SG (2002) Bax and Bak promote apoptosis by modulating endoplasmic reticular and mitochondrial Ca2+ stores. J Biol Chem 277:9219

    Article  CAS  PubMed  Google Scholar 

  28. Reed JC, Jurgensmeier JM, Matsuyama S (1998) Bcl-2 family proteins and mitochondria. Biochim Biophys Acta 1366:127

    Article  CAS  PubMed  Google Scholar 

  29. Roman ID, Thewles A, Coleman R (1995) Fractionation of livers following diosgenin treatment to elevate biliary cholesterol. Biochim Biophys Acta 1255:77

    Article  CAS  PubMed  Google Scholar 

  30. Taylor WR, Stark GR (2001) Regulation of the G2/M transition by p53. Oncogene 20:1803

    Article  CAS  PubMed  Google Scholar 

  31. Thornberry NA, Rano TA, Peterson EP, Rasper DM, Timkey T, Garcia-Calvo M, Houtzager VM, Nordstrom PA, Roy S, Vaillancourt JP, Chapman KT, Nicholson DW (1997) A combinatorial approach defines specificities of members of the caspase family and granzyme B. Functional relationships established for key mediators of apoptosis. J Biol Chem 272:17907

    Article  CAS  PubMed  Google Scholar 

  32. Vander Heiden MG, Chandel NS, Schumacker PT, Thompson CB (1999) Bcl-xL prevents cell death following growth factor withdrawal by facilitating mitochondrial ATP/ADP exchange. Mol Cell 3:159

    Article  PubMed  Google Scholar 

  33. Wallace KB, Eells JT, Madeira VMC, Cortopassi G, Jones DP (1997) Mitochondria-mediated cell injury. Fundam Appl Toxicol 38:23

    Article  CAS  PubMed  Google Scholar 

  34. Zhang Y, Fujita N, Tsuruo T (1999) Caspase-mediated cleavage of p21Waf1/Cip1 converts cancer cells from growth arrest to undergoing apoptosis. Oncogene 18:1131

    Article  CAS  PubMed  Google Scholar 

  35. Zhu L, Ling S, Yu XD, Venkatesh LK, Subramanian T, Chinnadurai G, Kuo TH (1999) Modulation of mitochondrial Ca2+ homeostasis by Bcl-2. J Biol Chem 274:33267

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported in part by grants from the Tsinghua University, Hong Kong Baptist University Joint Institute for Research of Chinese Medicine, and Tsinghua University’s 985 Project.

Author information

Affiliations

  1. Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing, People’s Republic of China

    Ming-Jie Liu, Zhao Wang & Qing-Yu Wu

  2. Institute of Biomedicine, Tsinghua University, Beijing, People’s Republic of China

    Zhao Wang

  3. Key Laboratory of Bioorganic Phosphorus Chemistry, Department of Chemistry, Tsinghua University, Beijing, People’s Republic of China

    Yong Ju

  4. Department of Biology, Hong Kong Baptist University, Hong Kong, People’s Republic of China

    Ricky Ngok-Shun Wong

Authors
  1. Ming-Jie Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Zhao Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Yong Ju
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ricky Ngok-Shun Wong
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Qing-Yu Wu
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Zhao Wang.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Liu, MJ., Wang, Z., Ju, Y. et al. Diosgenin induces cell cycle arrest and apoptosis in human leukemia K562 cells with the disruption of Ca2+ homeostasis. Cancer Chemother Pharmacol 55, 79–90 (2005). https://doi.org/10.1007/s00280-004-0849-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00280-004-0849-3

Keywords

  • Diosgenin
  • Apoptosis
  • Cell cycle arrest
  • Ca2+ homeostasis
  • Mitochondrial hyperpolarization