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Homeopathy 2015; 104(01): 36-47
DOI: 10.1016/j.homp.2014.10.001
Original Paper
Copyright © The Faculty of Homeopathy 2014

DNA fragmentation and cell cycle arrest: a hallmark of apoptosis induced by Ruta graveolens in human colon cancer cells

Shagun Arora
,
Simran Tandon
Subject Editor:
Further Information

Publication History

Received21 May 2014
revised15 October 2014

accepted20 October 2014

Publication Date:
19 December 2017 (online)

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In the present study, we investigated the anti-cancer effect of various potencies of Ruta graveolens (Ruta) on COLO-205 cell line, as evidenced by cytotoxicity, migration, clonogenecity, morphological and biochemical changes and modification in the levels of genes associated with apoptosis and cell cycle. On treatment of COLO-205 cells maximal effects were seen with mother tincture (MT) and 30C potencies, wherein decrease in cell viability along with reduced clonogenecity and migration capabilities were noted. In addition morphological and biochemical alterations such as nuclear changes (fragmented nuclei with condensed chromatin) and DNA ladder-like pattern (increased amount of fragmented DNA) in COLO-205 cells indicating apoptotic related cell death were seen. The expression of apoptosis and cell-cycle related regulatory genes assessed by reverse transcriptase-PCR revealed an up-regulation of caspase 9, caspase-3, Bax, p21 and p27 expression and down-regulation of Bcl-2 expression in treated cells. The mode of cell death was suggestive of intrinsic apoptotic pathway along with cell cycle arrest at the G2/M of the cell cycle. Our findings indicate that phytochemicals present in Ruta showed potential for natural therapeutic product development for colon carcinoma.

Keywords

Ruta - Apoptosis - Cell cycle arrest - GC–MS
 

  • References

  • 1 Globocan colorectal cancer estimated incidence, mortality and prevalence worldwide in 2012. http://globocan.iarc.fr/Pages/fact_sheets_cancer.aspx.
    PubMed
  • 2 Moongkarndi P., Kosem N., Kaslungka S., Luanratana O., Pongpan N. Antiproliferation, antioxidation and induction of apoptosis by Garcinia Mangostona (Mangosteen) On Skbr3 Human Breast Cancer Cell Line. J Ethnopharmacol 2004; 90: 161-166.
    CrossrefPubMedGoogle Scholar
  • 3 Hahnemann S. Organon of medicine. 6th edn JP Tarcher, Inc.; Los Angeles: 1982.
    Google Scholar
  • 4 Downer S.M., Cody M.M., McCluskey P. et al. Pursuit and practice of complementary therapies by cancer patients receiving conventional treatment. Br Med J 1994; 309: 86-89.
    CrossrefPubMedGoogle Scholar
  • 5 Banerji P., Campbell D.R., Banerji P. Cancer patients treated with the Banerji protocols utilising homoeopathic medicine: a best case series program of the National Cancer Institute USA. Oncol Reports 2008; 20 (01) 69-74.
    PubMedGoogle Scholar
  • 6 Parray S.A., Bhat J.U., Ahmad G., Jahan N., Sofi G., Iqbal S.M. Ruta graveolens: from traditional system of medicine to modern pharmacology: an overview. Am J PharmTech Res 2012; 2 (02) 1-14.
    PubMedGoogle Scholar
  • 7 Diwan R., Shinde A., Malpathak N. Phytochemical composition and antioxidant potential of Ruta graveolens L. In Vitro culture lines. J Bot 2012; 2012: 1-7.
    PubMedGoogle Scholar
  • 8 Pathak S., Multani A.S., Banerji P., Banerji P. Ruta 6 selectively induces cell death in brain cancer cells but proliferation in normal peripheral blood lymphocytes: a novel treatment for human brain cancer. Int J Oncol 2003; 2003 (23) 975-982.
    PubMedGoogle Scholar
  • 9 Preethi K., Kuttan G., Kuttan R. Anti-tumour activity of Ruta Graveolens Extract. Asian Pac J Cancer Prev 2006; 7: 439-443.
    PubMedGoogle Scholar
  • 10 Kumar K.B., Sunila E.S., Kuttan G., Preethi K.C., Venugopal C.M., Kuttan R. Inhibition of chemically induced carcinogenesis by drugs used in homeopathic medicine. Asian Pac J Cancer Prev 2007; 8: 98-102.
    PubMedGoogle Scholar
  • 11 Preethi K.C., Nair C.K., Kuttan R. Clastogenic potential of Ruta graveolens extract and a homeopathic preparation in mouse bone marrow cells. Asian Pac J Cancer Prev 2008; 9: 763-769.
    PubMedGoogle Scholar
  • 12 Fadlalla K., Watson A., Yehualaeshet T., Turner T., Samuel T. Ruta Graveolens extract induces dna damage pathways and blocks Akt activation to inhibit cancer cell proliferation and survival. Anticancer Res 2011; 31 (01) 233-241.
    PubMedGoogle Scholar
  • 13 Réthy B., Zupkó I., Minorics R., Hohmann J., Ocsovszki I., Falkay G. Investigation of cytotoxic activity on human cancer cell lines of arborinine and furanoacridones isolated from Ruta graveolens . Planta Medica 2007; 73: 41-48.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 14 Arora S., Aggarwal A., Singla P., Jyoti S., Tandon S. Anti-proliferative effects of homeopathic medicines on human kidney, colon and breast cancer cells. Homeopathy 102 2013; 274-282.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 15 Arora S., Tandon S., Tandon C. Evaluation of the cytotoxic effects of CAM therapies: An in vitro study in normal kidney cell line. World Sci J 2014; 2014: 1-11.
    PubMedGoogle Scholar
  • 16 Roudkenar M., Halabian R., Roushandeh A. et al. Lipocalin 2 regulation by thermal stresses: protective role of Lcn2/NGAL against cold and heat stresses. Exp Cell Res 2009; 315: 3140-3151.
    CrossrefPubMedGoogle Scholar
  • 17 Monga J., Pandit S., Chauhan C., Sharma M. Cytotoxicity and apoptosis induction in human breast adenocarcinoma MCF-7cells by (+)-cyanidan-3-ol. Exp Toxicol Pathology 2013; 65: 1-19.
    PubMedGoogle Scholar
  • 18 Manoj J., Kumar T., Varghese S., Murugan K. The effect of methanol and water extract of Leucobryum bowringii Mitt on growth, migration and invasion of MCF-7 human breast cancer cell lines in vitro”. Indian J Exp Biol 2012; 50: 602-611.
    PubMedGoogle Scholar
  • 19 Kim A., Im M., Yim N., Jung Y., Ma J. Aqueous extract of Bambusae Caulis in taeniam inhibits PMA-induced tumor cell invasion and pulmonary metastasis: suppression of NF-kB activation through ROS signaling. Plosone 2013; 8: 1-13.
    PubMedGoogle Scholar
  • 20 Franken N., Rodermond H., Stap J., Haveman J., Bree C. Clonogenic assay of cells: in vitro. Nat Protoc 2006; 1 (05) 2315.
    CrossrefPubMedGoogle Scholar
  • 21 Allen J., Sotos M., Sylte J., Czuprynsk C.J. Use of Hoechst 33342 staining to detect apoptotic changes in bovine mononuclear phagocytes infected with Mycobacterium Avium subsp. paratuberculosis. Clin Diagnostic Laboratory Immunol 2001; 8 (02) 460-464.
    PubMedGoogle Scholar
  • 22 Silva M.S., Filho J.M., Anazetti M.C. et al. Xylodiol from Xylopia langsdorfiana induces apoptosis in HL60 cells. Braz J Pharmacogn 2011; 21 (06) 1035-1042.
    CrossrefPubMedGoogle Scholar
  • 23 Zinadah O.A., Khalil W.K., Ashmaoui H.M., Abdul F., Alsoud M.E. Evaluation of the anti-genotoxicity and growth performance impacts of green algae on Mugil cephalus . Life Sci J 2013; 10 (03) 1543-1554.
    PubMedGoogle Scholar
  • 24 Azizi E., Abdolmohammadi M.H., Fouladdel S., Shafiee A., Amin G., Ghaffari S. Evaluation of p53 and Bcl-2 genes and proteins expression in human breast cancer T47D cells treated with extracts of Astrodaucus persicus (Boiss.) drude in comparison to Tamoxifen. Daru 2009; 17 (03) 181-186.
    PubMedGoogle Scholar
  • 25 Choi S.C., Kim K.D., Kim J.T. et al. Expression of human NDRG2 by myeloid dendritic cells inhibits down-regulation of activated leukocyte cell adhesion molecule (ALCAM) and contributes to maintenance of T cell stimulatory activity. J Leukoc Biol 2008; 83 (01) 89-98.
    CrossrefPubMedGoogle Scholar
  • 26 Tsuan H.Y., Lu C., Yang J.S. et al. Berberine induced apoptosis via promoting the expression of caspase-8, -9 and -3, apoptosis-inducing factor and endonuclease G in SCC-4 human tongue squamous carcinoma cancer cells. Anticancer Res 2009; 29: 4063-4070.
    PubMedGoogle Scholar
  • 27 Shang L.H., Mei Li C., Yang Yang Z., Che D.H., Cao J.Y., Yu Y. Luffa echinata Roxb induces human colon cancer cell (HT-29) death by triggering the mitochondrial apoptosis pathway. Molecules 2012; 17: 5780-5794.
    CrossrefPubMedGoogle Scholar
  • 28 Ling Q., Xu X., Wei X. et al. Oxymatrine induces human pancreatic cancer PANC-1 cells apoptosis via regulating expression of Bcl-2 and IAP families, and releasing of cytochrome C. J Exp Clin Cancer Res 2013; 30: 66.
    PubMedGoogle Scholar
  • 29 Pozarowski P and Darzynkiewicz Z. Methods in Molecular Biology: Checkpoint Controls and Cancer, Activation and Regulation Protocols”. Humana Press Inc., 281(2):303.
    PubMed
  • 30 Guzman M. Cannabinoids: potential anticancer agents. Nat Rev Cancer 2003; 3: 745-755.
    CrossrefPubMedGoogle Scholar
  • 31 Naser B., Bodinet C., Tegtmeier M., Lindequist U. Thuja occidentalis (Arbor vitae): a review of its pharmaceutical, pharmacological and clinical properties. Evidence Based Complementary Altern Med 2005; 2: 69-78.
    PubMedGoogle Scholar
  • 32 Moghde A., Gangal S., Shilkar S. In vitro cytotoxicity of bryonia laciniosa. Naud. on human cancer cells. Indian J Nat Prod Resour 2011; 2: 322-329.
    PubMedGoogle Scholar
  • 33 Biswas R., Mandal S., Dutta S. et al. Thujone-rich fraction of Thuja occidentalis demonstrates major anti-cancer potentials: evidences from In Vitro studies on A375 cells. Evidence Based Complementary Altern Med 2011; 2011: 1-16.
    PubMedGoogle Scholar
  • 34 Yang H., Chen C., Chang W. et al. Growth inhibition and induction of apoptosis in MCF-7 breast cancer cells by Antrodia camphorata . Cancer Lett 2006; 23: 215-227.
    PubMedGoogle Scholar
  • 35 Shukla Y., Singh M. Cancer preventive properties of ginger: a brief review. Food Chem Toxicol 2007; 45: 683-690.
    CrossrefPubMedGoogle Scholar
  • 36 Ghosh S., Bishayee K., Khuda-Bukhsh A.R. Graveoline isolated from ethanolic extract of Ruta graveolens triggers apoptosis and autophagy in skin melanoma cells: a novel apoptosis-independent autophagic signaling pathway. Phytotherapy Res 2013; 2013: 1-10.
    PubMedGoogle Scholar
  • 37 Cheng E., Kirsch D., Clem R.J. et al. Conversion of Bcl-2 to a Bax-like death effector by caspases. Science 1997; 278: 1966-1968.
    CrossrefPubMedGoogle Scholar
  • 38 Leung L.K., Wang T.T. Differential effects of chemotherapeutic agents on the Bcl-2/Bax apoptosis pathway in human breast cancer cell line MCF-7. Breast Cancer Res Treat 1999; 55: 73-83.
    CrossrefPubMedGoogle Scholar
  • 39 Hanahan D., Weinberg R.A. Hallmarks of cancer: the next generation. Cell 2011; 144: 646-674.
    CrossrefPubMedGoogle Scholar
  • 40 Hoyt M.A. A new checkpoint takes shape. Nat Cell Biol 2004; 6: 801-803.
    CrossrefPubMedGoogle Scholar
  • 41 Wang Y, Ji P, Liu J, et al. Centrosome-associated regulators of the G (2)/M checkpoint as targets for cancer therapy. Mol Cancer 2009; 8(8).
    PubMedGoogle Scholar
  • 42 Coqueret O. New roles for p21 and p27 cell-cycle inhibitors: a function for each cell compartment?. Trends Cell Biol 2003; 13: 65-70.
    CrossrefPubMedGoogle Scholar
  • 43 Wang Y., Hong C., Zhou C., Xu D., Qu H. Screening antitumor compounds psoralen and isopsoralen from Psoralea corylifolia L. Seeds. Evidence Based Complementary Altern Med 2011; 2011: 1-7.
    PubMedGoogle Scholar
  • 44 Gallo M.B., Sarachine M.J. Biological activities of Lupeol. Int J Biomed Pharm Sci 2009; 3: 46-66.
    PubMedGoogle Scholar
  • 45 Licchetta A., Correale P., Migali C. et al. Oral metronomic chemo-hormonal-therapy of metastatic breast cancer with cyclophosphamide and megestrol acetate. J Chemother 2010; 22: 201-204.
    CrossrefPubMedGoogle Scholar