Advertisement

Reversine induces caspase-dependent apoptosis of human osteosarcoma cells through extrinsic and intrinsic apoptotic signaling pathways

  • Jae-Sung Kim
  • In-A Cho
  • Kyeong-Rok Kang
  • HyangI Lim
  • Tae-Hyeon Kim
  • Sun-Kyoung Yu
  • Heung-Joong Kim
  • Seul Ah Lee
  • Sung Min Moon
  • Hong Sung Chun
  • Chun Sung Kim
  • Do Kyung KimEmail author
Research Article
  • 28 Downloads

Abstract

Background

The 2-(4-morpholinoanilino)-6-cyclohexylaminopurine (reversine) acts as a chemopreventive agent and induces apoptotic cell death in various cancer cells. However, the anticancer effects of reversine on osteosarcoma cells are not clearly established.

Objective

The purpose of this study was to investigate the effect of reversine on cell proliferation and induction of apoptosis in human osteosarcoma cells.

Methods

Cell viability assay, histological analysis, DAPI staining, caspase activation analysis, flow cytometric analysis and immunoblotting were carried out in MG-63 osteosarcoma cells.

Results

Reversine inhibited the growth of cells in a dose-dependent manner and induced nuclear condensation and fragmentation. Reversine-treated cells showed caspase-3/7 activation and increased apoptosis versus control cells. FasL, a death ligand associated with extrinsic apoptotic signaling pathways, was significantly up-regulated by reversine treatment. Moreover, the caspase-8, a part of the extrinsic apoptotic pathway, was activated by reversine treatments. Expressions of anti-apoptotic factors such as Bcl-2 and Bcl-xL, components of the mitochondria dependent intrinsic apoptosis pathway, significantly decreased following reversine treatment. The expressions of pro-apoptotic factors such as BAX, BAD and caspase-9 increased by reversine treatments. In addition, reversine activated caspase-3 and Poly (ADP-ribose) polymerase (PARP) to induce cell death. The Z-VAD-fmk significantly inhibited cell death through the suppression of caspase-3 expression in MG-63 cells treated with reversine.

Conclusion

These results suggest that the reversine may inhibit cell proliferation and induce apoptotic cell death in MG-63 osteosarcoma cells through both the mitochondria-mediated intrinsic pathway and the death receptor-mediated extrinsic pathway, and may have potential properties for the discovery of anti-cancer agents.

Keywords

Cell death Apoptosis Osteosarcoma Anticancer activity Caspase Reversine 

Notes

Compliance with ethical standards

Conflict of interest

Jae-Sung Kim, In-A Cho, Kyeong-Rok Kang, HyangI Lim, Tae-Hyeon Kim, Sun-Kyoung Yu, Heung-Joong Kim, Seul Ah Lee, Sung Min Moon, Hong Sung Chun, Chun Sung Kim and Do Kyung Kim declare that they have no conflict of interest.

Informed consent

Informed consent was obtained from all individual participants included in the study.

Research involving human and animal rights

This article does not contain any studies with human subjects or animals performed by any of the authors.

References

  1. Bae YH, Shin JM, Park HJ, Jang HO, Bae MK, Bae SK (2014) Gain-of-function mutant p53-R280K mediates survival of breast cancer cells. Genes Genom 36:171–178CrossRefGoogle Scholar
  2. Cheng L, Wang H, Guo K, Wang Z, Zhang Z, Shen C, Chen L, Lin J (2018) Reversine, a substituted purine, exerts an inhibitive effect on human renal carcinoma cells via induction of cell apoptosis and polyploidy. Onco Targets Ther 11:1025–1035CrossRefGoogle Scholar
  3. D’Alise AM, Amabile G, Iovino M, Di Giorgio FP, Bartiromo M, Sessa F, Villa F, Musacchio A, Cortese R (2008) Reversine, a novel Aurora kinases inhibitor, inhibits colony formation of human acute myeloid leukemia cells. Mol Cancer Ther 7:1140–1149CrossRefGoogle Scholar
  4. Daiy H, Huangy Y, Li Y, Meng G, Wang Y, Guo QN (2012) TSSC3 overexpression associates with growth inhibition, apoptosis induction and enhances chemotherapeutic effects in human osteosarcoma. Carcinogenesis 33:30–40CrossRefGoogle Scholar
  5. Fisher DE (2001) The p53 tumor suppressor: critical regulator of life and death in cancer. Apoptosis 6:7–15CrossRefGoogle Scholar
  6. Fuchs B, Pritchard DJ (2002) Etiology of osteosarcoma. Clin Orthop Relat Res 397:40–52CrossRefGoogle Scholar
  7. Gibbs CP Jr, Weber K, Scarborough MT (2002) Malignant bone tumors. Instr Course Lect 51:413–428Google Scholar
  8. Heath-Engel HM, Chang NC, Shore GC (2008) The endoplasmic reticulum in apoptosis and autophagy: role of the BCL-2 protein family. Oncogene 27:6419–6433CrossRefGoogle Scholar
  9. Herrnring C, Reimer T, Jeschke U, Makovitzky J, Krüger K, Gerber B, Kabelitz D, Friese K (2000) Expression of the apoptosis-inducing ligands FasL and TRAIL in malignant and benign human breast tumors. Histochem Cell Biol 113:189–194CrossRefGoogle Scholar
  10. Hsieh TC, Traganos F, Darzynkiewicz Z, Wu JM (2007) The 2,6-disubstituted purine reversine induces growth arrest and polyploidy in human cancer cells. Int J Oncol 31:1293–1300Google Scholar
  11. Kaufmann SH, Earnshaw WC (2000) Induction of apoptosis by cancer chemotherapy. Exp Cell Res 256:42–49CrossRefGoogle Scholar
  12. Kaufmann SH, Hengartner MO (2001) Programmed cell death: alive and well in the new millennium. Trends Cell Biol 11:526–534CrossRefGoogle Scholar
  13. Kim YK, Choi HY, Kim NH, Lee W, Seo DW, Kang DW, Lee HY, Han JW, Park SW, Kim SN (2007) Reversine stimulates adipocyte differentiation and downregulates Akt and p70(s6k) signaling pathways in 3T3-L1 cells. Biochem Biophys Res Commun 358:553–558CrossRefGoogle Scholar
  14. Kluck RM, Bossy-Wetzel E, Green DR, Newmeyer DD (1997) The release of cytochrome c from mitochondria: a primary site for Bcl-2 regulation of apoptosis. Science 275:1132–1136CrossRefGoogle Scholar
  15. Kluck RM, Esposti MD, Perkins G, Renken C, Kuwana T, Bossy-Wetzel E, Goldberg M, Allen T, Barber MJ, Green DR et al (1999) The pro-apoptotic proteins, Bid and Bax, cause a limited permeabilization of the mitochondrial outer membrane that is enhanced by cytosol. J Cell Biol 147:809–822CrossRefGoogle Scholar
  16. Kuo CH, Lu YC, Tseng YS, Shi CS, Chen SH, Chen PT, Wu FL, Chang YP, Lee YR (2014) Reversine induces cell cycle arrest, polyploidy, and apoptosis in human breast cancer cells. Breast Cancer 21:358–369CrossRefGoogle Scholar
  17. Lavin MF, Gueven N (2006) The complexity of p53 stabilization and activation. Cell Death Differ 13:941–950CrossRefGoogle Scholar
  18. Lee YR, Wu WC, Ji WT, Chen JP, Cheng YP, Chiang MK, Chen HR (2012) Reversine suppresses oral squamous cell carcinoma via cell cycle arrest and concomitantly apoptosis and autophagy. J Biomed Sci 19:9CrossRefGoogle Scholar
  19. Li HJ, Wang CY, Mi Y, Du CG, Cao GF, Sun XC, Liu DJ, Shorgan B (2013) FasL-induced apoptosis in bovine oocytes via the Bax signal. Theriogenology 80:248–255CrossRefGoogle Scholar
  20. Lu CH, Liu TW, Hua SC, Yu HI, Chang YP, Lee YR (2012) Autophagy induction of reversine on human follicular thyroid cancer cells. Biomed Pharmacother 66:642–647CrossRefGoogle Scholar
  21. McMillin DW, Delmore J, Weisberg E, Negri JM, Geer DC, Klippel S, Mitsiades N, Schlossman RN, Munshi NC, Kung AL et al (2010) Tumor cell-specific bioluminescence platform to identify stroma-induced changes to anticancer drug activity. Nat Med 16:483–489CrossRefGoogle Scholar
  22. Oltvai ZN, Milliman CL, Korsmeyer SJ (1993) Bcl-2 heterodimerizes in vivo with a conserved homolog, Bax, that accelerates programmed cell death. Cell 74:609–619CrossRefGoogle Scholar
  23. Park ZB, Lee JS, Cho BP, Rhee KJ, Baik SK, Kim J, Kang SJ, Park DJ, Oh JE, Shin HC et al (2015) Adipose tissue-derived mesenchymal stem cells cultured at high cell density express brain-derived neurotrophic factor and exert neuroprotective effects in a 6-hydroxydopamine rat model of Parkinson’s disease. Genes Genom 37:213–221CrossRefGoogle Scholar
  24. Prokhorova EA, Zamaraev AV, Kopeina GS, Zhivotovsky B, Lavrik IN (2015) Role of the nucleus in apoptosis: signaling and execution. Cell Mol Life Sci 72:4593–4612CrossRefGoogle Scholar
  25. Qin HX, Yang J, Cui HK, Li SP, Zhang W, Ding XL, Xia YH (2013) Synergistic antitumor activity of reversine combined with aspirin in cervical carcinoma in vitro and in vivo. Cytotechnology 65:643–653CrossRefGoogle Scholar
  26. Reed JC (2001) Apoptosis-regulating proteins as targets for drug discovery. Trends Mol Med 7:314–319CrossRefGoogle Scholar
  27. Ryu DS, Lee HS, Lee GS, Lee DS (2012) Effects of the ethylacetate extract of Orostachys japonicus on induction of apoptosis through the p53-mediated signaling pathway in human gastric cancer cells. Biol Pharm Bull 35:660–665CrossRefGoogle Scholar
  28. Saraiya M, Nasser R, Zeng Y, Addya S, Ponnappan RK, Fortina P, Anderson DG, Albert TJ, Shapiro IM, Risbud MV (2010) Reversine enhances generation of progenitor-like cells by dedifferentiation of annulus fibrosus cells. Tissue Eng Part A 16:1443–1455CrossRefGoogle Scholar
  29. Yang SJ, Lee SA, Park MG, Kim JS, Yu SK, Kim CS, Kim SG, Kim JS, Oh JS, Kim HJ et al (2014) Induction of apoptosis by diphenyldifluoroketone in osteogenic sarcoma cells is associated with activation of caspases. Oncol Rep 31:2286–2292CrossRefGoogle Scholar
  30. Yu CS, Huang AC, Lai KC, Huang YP, Lin MW, Yang JS, Chung JG (2012) Diallyl trisulfide induces apoptosis in human primary colorectal cancer cells. Oncol Rep 28:949–954CrossRefGoogle Scholar
  31. Yu SJ, Cho IA, Kang KR, Jung YR, Cho SS, Yoon G, Oh JS, You JS, Seo YS, Lee GJ et al (2017) Licochalcone-E induces caspase-dependent death of human pharyngeal squamous carcinoma cells through the extrinsic and intrinsic apoptotic signaling pathways. Oncol Lett 13:3662–3668CrossRefGoogle Scholar
  32. Zamaraev AV, Kopeina GS, Zhivotovsky B, Lavrik IN (2015) Cell death controlling complexes and their potential therapeutic role. Cell Mol Life Sci 72:505–517CrossRefGoogle Scholar
  33. Zao Q, Wang C, Zhu J, Wang L, Dong S, Zhang G, Tian J (2011) RNAi-mediated knockdown of cyclooxygenase2 inhibits the growth, invasion and migration of Saos2 human osteosarcoma cells: a case control study. J Exp Clin Cancer Res 30:26CrossRefGoogle Scholar

Copyright information

© The Genetics Society of Korea 2019

Authors and Affiliations

  • Jae-Sung Kim
    • 1
  • In-A Cho
    • 1
  • Kyeong-Rok Kang
    • 1
  • HyangI Lim
    • 1
  • Tae-Hyeon Kim
    • 1
  • Sun-Kyoung Yu
    • 1
  • Heung-Joong Kim
    • 1
  • Seul Ah Lee
    • 1
  • Sung Min Moon
    • 1
  • Hong Sung Chun
    • 2
  • Chun Sung Kim
    • 1
  • Do Kyung Kim
    • 1
    • 3
    Email author
  1. 1.Institute of Dental ScienceChosun UniversityGwangjuRepublic of Korea
  2. 2.Department of Biomedical ScienceChosun UniversityGwangjuRepublic of Korea
  3. 3.Department of Oral Physiology, School of DentistryChosun UniversityGwangjuRepublic of Korea

Personalised recommendations