Orostachys japonicus induces apoptosis and cell cycle arrest through the mitochondria-dependent apoptotic pathway in AGS human gastric cancer cells

Ryu,Deok-Seon; Kim,Seon-Hee; Kwon,Ji-Hae; Lee,Dong-Seok

doi:10.3892/ijo.2014.2404

Orostachys japonicus induces apoptosis and cell cycle arrest through the mitochondria-dependent apoptotic pathway in AGS human gastric cancer cells

Authors:
- Deok-Seon Ryu
- Seon-Hee Kim
- Ji-Hae Kwon
- Dong-Seok Lee
View Affiliations

Affiliations: Department of Biomedical Laboratory Science, Inje University, Gimhae, Gyungnam 621-749, Republic of Korea, Department of Smart Foods and Drugs, Graduate School of Inje University, Gimhae, Gyungnam 621-749, Republic of Korea
Published online on: April 28, 2014 https://doi.org/10.3892/ijo.2014.2404
Pages: 459-469

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

We investigated the anticancer mechanisms of the ethylacetate (EtOAc) fraction from Orostachys japonicus in human gastric cancer (AGS) cells. Flow cytometric analysis revealed that the number of total apoptotic cells following treatment with the EtOAc fraction increased in a dose-dependent manner. In the cell cycle analyses, the EtOAc fraction increased the peak in the sub-G1, indicating apoptosis, and in the G2/M phases in a dose-dependent manner. In the RT-PCR analysis, the expression of cyclin-dependent kinase 1 (CDK 1) and cyclin B1 decreased in a dose- and time-dependent manner. The results of western blotting revealed that the protein levels of p53, cytochrome c, and cleaved caspase-3, -8 and -9 proteins increased and those of B cell lymphoma-2 (bcl-2) and pro-caspase-3, -8 and -9 proteins decreased in a dose- and time-dependent manner, whereas the levels of bcl-2-associated x protein (bax) remained unchanged. Furthermore, the changes in the levels of pro-caspase-3, -8 and -9 and cleaved caspase-3, -8 and -9 were abolished by the pan-caspase inhibitor Z-VAD-FMK. In addition, phosphorylation of p38 and JNK increased in a time-dependent manner. These results, for the first time, provide an understanding of the potential anticancer activity of the O. japonicus, which functions through the induction of apoptosis and cell cycle arrest.

View Figures

View References

1.	Kelloff GJ: Perspectives on cancer chemoprevention research and drug development. Adv in Cancer Res. 78:199–334. 2000. View Article : Google Scholar : PubMed/NCBI
2.	Ryu DS, Baek GO, Kim EY, Kim KH and Lee DS: Effects of polysaccharides derived from Orostachys japonicus on induction of cell cycle arrest and apoptotic cell death in human colon cancer cells. BMB Rep. 43:750–755. 2010.PubMed/NCBI
3.	Choi SY, Chung MJ, Seo WD, Shin JH, Shon MY and Sung NJ: Inhibitory effects of Orostachys japonicus extracts on the formation of N-nitrosodimethylamine. J Agric Food Chem. 54:6075–6078. 2006.
4.	Jung HJ, Choi J, Nam JH and Park HJ: Anti-ulcerogenic effects of the flavonoid-rich fraction from the extract of Orostachys japonicus in mice. J Med Food. 10:702–706. 2007. View Article : Google Scholar : PubMed/NCBI
5.	Lee HS, Ryu DS, Lee GS and Lee DS: Anti-inflammatory effects of dichloromethane fraction from Orostachys japonicus in RAW 264.7 cells: Suppression of NF-κB activation and MAPK signaling. J Ethnopharmacol. 140:271–276. 2012.PubMed/NCBI
6.	Je Ma C, Jung WJ, Lee KY, Kim YC and Sung SH: Calpain inhibitory flavonoids isolated from Orostachys japonicus. J Enzyme Inhib Med Chem. 24:676–679. 2009.PubMed/NCBI
7.	Park HJ, Young HS, Park KY, Rhee SH, Chung HY and Choi JS: Flavonoids from the whole plants of Orostachys japonicus. Arch Pharm Res. 14:167–171. 1991. View Article : Google Scholar
8.	Ryu DS, Lee HS, Lee GS and Lee DS: Effect 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. 35:660–665. 2012.
9.	Singh SV, Herman-Antosiewicz A, Singh AV, Lew KL, Srivastava SK, Kamath R, Brown KD, Zhang L and Baskarah R: Sulforaphane-induced G2/M phase cell cycle arrest involves checkpoint kinase 2-mediated phosphorylation of cell division cycle 25C. J Biol Chem. 279:25813–25822. 2004. View Article : Google Scholar : PubMed/NCBI
10.	Harris SL and Levin AJ: The p53 pathway: positive and negative feedback loops. Oncogene. 24:2899–2908. 2005. View Article : Google Scholar : PubMed/NCBI
11.	Vazquez A, Bond EE, Levine AJ and Bond GL: The genetics of the p53 pathway, apoptosis and cancer therapy. Nat Rev Drug Discov. 7:979–987. 2008. View Article : Google Scholar : PubMed/NCBI
12.	Shiozaki EN and Shi Y: Caspases, IAPs and Smac/DIABLO: mechanisms from structural biology. Trends Biochem Sci. 29:486–494. 2004. View Article : Google Scholar : PubMed/NCBI
13.	Tsujimoto Y: Bcl-2 family of proteins: life-or-death switch in mitochondria. Biosci Rep. 22:47–58. 2002. View Article : Google Scholar : PubMed/NCBI
14.	Dhanalakshmi S, Agarwal P, Globe L and Agarwal R: Silibinin sensitizes human prostate carcinoma DU145 cells to cisplatinand carboplatin-induced growth inhibition and apoptotic death. Int J Cancer. 106:699–705. 2003. View Article : Google Scholar : PubMed/NCBI
15.	Brooks G: Cyclin, cyclin-dependent kinases, and cyclin-dependent kinase inhibitors: detection methods and activity measurements. Methods Mol Biol. 296:291–298. 2005.PubMed/NCBI
16.	Vermeulen K, Berneman ZN and Van Bockstaele DR: Cell cycle and apoptosis. Cell Prolif. 36:165–175. 2003. View Article : Google Scholar
17.	Sebolt-Leopold JS: Development of anticancer drugs targeting the MAP kinase pathway. Oncogene. 19:6594–6599. 2000. View Article : Google Scholar : PubMed/NCBI
18.	Sebolt-Leopold JS and Herrera R: Targeting the mitogen-activated protein kinase cascade to treat cancer. Nat Rev Cancer. 4:937–947. 2004. View Article : Google Scholar : PubMed/NCBI
19.	Jeong JH, Ryu DS, Suk DH and Lee DS: Anti-inflammatory effects of ethanol extract from Orostachys japonicus on modulation of signal pathways in LPS-stimulated RAW 264.7 cells. BMB Rep. 44:399–404. 2011.PubMed/NCBI
20.	Lee HS, Bilehal D, Lee GS, Ryu DS, Kim HK, Suk DH and Lee DS: Anti-inflammatory effect of the hexane fraction from Orostachys japonicus in RAW 264.7 cells by suppression of NF-κB and PI3K-Akt signaling. J Funct Foods. 5:1217–1225. 2013.
21.	Kaufmann SH and Henqartner MO: Programmed cell death: alive and well in the new millennium. Trends Cell Biol. 11:526–534. 2001. View Article : Google Scholar : PubMed/NCBI
22.	Ryu DS, Kim SH and Lee DS: Anti-proliferative effect of polysaccharides from Salicornia herbacea on induction of G2/M arrest and apoptosis in human colon cancer cells. J Microbiol Biotechnol. 19:1482–1489. 2009.PubMed/NCBI
23.	Nakanishi M, Shimada N and Niida H: Genetic instability in cancer cells by impaired cell cycle checkpoints. Cancer Sci. 97:984–989. 2006. View Article : Google Scholar : PubMed/NCBI
24.	Elledge SJ and Harpae JW: CDK inhibitors; on the threshold of checkpoints and development. Curr Opin Cell Biol. 6:847–852. 1994. View Article : Google Scholar : PubMed/NCBI
25.	Vousden KH: Apoptosis. p53 and PUMA: a deadly duo. Science. 309:1685–1686. 2005. View Article : Google Scholar : PubMed/NCBI
26.	Lavin MF and Gueven N: The complexity of p53 stabilization and activation. Cell Death Differ. 13:941–950. 2006. View Article : Google Scholar : PubMed/NCBI
27.	Ai Z, Lu W and Qin X: Arsenic trioxide induces gallbladder carcinoma cell apoptosis via down regulation of bcl-2. Biochem Biophys Res Commun. 348:1075–1081. 2006. View Article : Google Scholar : PubMed/NCBI
28.	Wang JB, QI LL, Zheng SD and Wu TX: Curcumin induces apoptosis through the mitochondria-mediated apoptotic pathway in HT-29 cells. J Zhejizng Univ Sci B. 10:93–102. 2009. View Article : Google Scholar : PubMed/NCBI
29.	Lavrik I, Golks A and Krammer PH: Death receptor signaling. J Cell Sci. 118:265–267. 2005. View Article : Google Scholar
30.	Fujita E, Egashira J, Urase K, Kuida K and Momoi T: Caspase-9 processing by caspase-3 via a feedback amplification loop in vivo. Cell Death Differ. 8:335–344. 2001. View Article : Google Scholar : PubMed/NCBI
31.	Seger R and Krebs EG: The MAPK signaling cascade. FASEB J. 9:726–735. 1995.PubMed/NCBI
32.	Meier P, Finch A and Evan G: Apoptosis in development. Nature. 407:796–801. 2000. View Article : Google Scholar
33.	Evan GI and Vousden KH: Proliferation, cell cycle and apoptosis in cancer. Nature. 411:342–348. 2001. View Article : Google Scholar : PubMed/NCBI
34.	Schwartz GK and Shah MA: Targeting the cell cycle: a new approach to cancer therapy. J Clin Oncol. 23:9408–9421. 2005. View Article : Google Scholar : PubMed/NCBI
35.	Koff A, Giordano A, Desai D, Yamashita K, Harper JW, Elledge S, Nishimoto T, Morgan DO, Franza BR and Roberts JM: Formation and activation of a cyclin E-cdk2 complex during the G1 phase of the human cell cycle. Science. 257:1689–1694. 1992. View Article : Google Scholar : PubMed/NCBI
36.	Fisher DE: The p53 tumor suppressor: critical regulator of life & death in cancer. Apoptosis. 6:7–15. 2001. View Article : Google Scholar
37.	Moll UM and Zaika A: Nuclear and mitochondrial apoptotic pathways of p53. FEBS Lett. 493:65–69. 2001. View Article : Google Scholar : PubMed/NCBI
38.	Erster S, Mihara M, Kim RH, Petrenko O and Moll UM: In vivo mitochondrial p53 translocation triggers a rapid first wave of cell death in response to DNA damage that can precede p53 target gene activation. Mol Cell Biol. 24:6728–6741. 2004. View Article : Google Scholar : PubMed/NCBI
39.	Jiang X and Wang X: Cytochrome c-mediated apoptosis. Annu Rev Biochem. 73:87–106. 2004. View Article : Google Scholar
40.	Nagata S: Fas ligand-induced apoptosis. Annu Rev Genet. 33:29–55. 1999. View Article : Google Scholar
41.	Willis SN and Adams JM: Life in the balance: how BH3-only proteins induce apoptosis. Curr Opin Cell Biol. 17:617–625. 2005. View Article : Google Scholar : PubMed/NCBI
42.	Pop C, Timmer J, Sperandio S and Salvesen GS: The apoptosome activates caspase-9 by dimerization. Mol Cell. 22:269–275. 2006. View Article : Google Scholar : PubMed/NCBI
43.	Zhou Z, Sun X and Kang YJ: Ethanol-induced apoptosis in mouse liver: Fas- and cytochrome c-mediated caspase-3 activation pathway. Am J Pathol. 159:329–338. 2001. View Article : Google Scholar : PubMed/NCBI
44.	Jin UH, Song KH, Motomura M, Suzuki I, Gu YH, Kang YJ, Moon TC and Kim CH: Caffeic acid phenethyl ester induces mitochondria-mediated apoptosis in human myeloid leukemia U937 cells. Mol Cell Biochem. 310:43–48. 2008. View Article : Google Scholar : PubMed/NCBI
45.	Zheng Y, Zhou M, Ye A, Li Q, Bai Y and Zhang Q: The conformation change of Bcl-2 is involved in arsenic trioxide-induced apoptosis and inhibition of proliferation in SGC7901 human gastric cancer cells. World J Sur Oncol. 8:312010. View Article : Google Scholar : PubMed/NCBI
46.	Cho SH, Chung KS, Choi JH, Kim DH and Lee KT: Compound K, a metabolite of ginseng saponin, induces apoptosis via caspase-8-dependent pathway in HL-60 human leukemia cells. BMC Cancer. 9:4492009. View Article : Google Scholar : PubMed/NCBI
47.	Martin GS: Cell signaling and cancer. Cancer Cell. 4:167–174. 2003. View Article : Google Scholar
48.	Freeman SM and Whartenby KA: The role of the mitogen-activated protein kinase cellular signaling pathway in tumor cell survival and apoptosis. Drug News Perspect. 17:237–242. 2004. View Article : Google Scholar : PubMed/NCBI
49.	Lin A and Dibling B: The true face of JNK activation in apoptosis. Aging Cell. 1:112–116. 2002. View Article : Google Scholar : PubMed/NCBI
50.	Lunghi P, Giuliani N, Mazzera L, Lombardi G, Ricca M, Corradi A, Cantoni AM, Salvatore L, Riccioni R and Costanzo A: Targeting MEK/MAPK signal transduction module potentiates ATO-induced apoptosis in multiple myeloma cells through multiple signaling pathways. Blood. 112:2450–2462. 2008. View Article : Google Scholar
51.	Mansouri A, Ridgway LD, Korapati AL, Zhang Q, Tian L, Wang Y, Siddik ZH, Mills GB and Claret FX: Sustained activation of JNK/p38 MAPK pathways in response to cisplatin leads to Fas ligand induction and cell death in ovarian carcinoma cells. J Biol Chem. 278:19245–19256. 2003. View Article : Google Scholar : PubMed/NCBI