Mehmet SARIMAHMUT, Ümit YIRTICI, Aysun ERGENE, Fatih GÖGER

Cytotoxic and apoptotic effects of endemic Centaurea fenzlii Reichardt on the MCF-7 breast cancer cell line

The main purpose of this study was to analyze the cytotoxic activity of an extract obtained from Centaurea fenzlii Reichardt, and the fractions eluted from this extract, in breast cancer cells. After isolation and structural analysis of the fractions were conducted, a meaningful cytotoxic effect was indicated. The goal of the analysis was to reveal the mechanism by which this effect occurs through researching the apoptotic side of these fractions and determining the amount of several proteins that are the products of the genes. Test substances were applied to breast cancer cells and the inhibitory concentration value 50 (IC50) that caused a cytotoxic effect was determined using MTT and ATP assays. The Centaurea fenzlii Reichardt dichloromethane extracts-ethyl acetate fractions (CFDCMEAF) exhibited a stronger growth-inhibitory effect on MCF-7 cells (45.771 µg/mL). The apoptotic effect was studied using double staining and flow cytometry. The death rate in the cells treated with the CFDCM-EAF IC50 dose was approximately 90%: 9.2% living cells, 22.8% necrotic cells, 62.3% late apoptotic cells, and 5.8% early apoptotic cells. Structural analysis of the CFDCM-EAF, which indicated significant cytotoxic effects, was performed using chromatographic methods. Hispidulin was the major component of the CFDCM-EAF by LC-APCI-MS/MS analysis.

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Ahmed SA, Kamel EM (2014). Cytotoxic activities of flavonoids from Centaurea scoparia. Sci World J 2014: 274207.
Andreotti PE, Cree IA, Kurbacher CM, Hartmann DM, Linder D, Harel G, Gleiberman I, Caruso PA, Ricks SH, Untch M et al. (1995). Chemosensitivity testing of human tumors using a microplate adenosine-triphosphate luminescence assay - clinical correlation for cisplatin resistance of ovarian carcinoma. Cancer Res 55: 5276-5282.
Ari F, Ikitimur EI, Ulukaya E (2010). The ATP assay, but not the MTT assay, detects further cytotoxicity of the combination of anthracycline-based therapy with histone deacetylase inhibitor (valproic acid) in breast cancer cells. Turk J Biochem 35: 293- 299.
Beltagy AM (2015). Chemical composition and cytotoxic activity of Centaurea scoparea Sieb against four human cell Lines. J Pharmacol Sci Res 7: 103-107.
Candan F, Uysal T, Tugay O, Bozkurt M, Ertugrul K, Demirelma H (2016). The examinations of achene ultrastructural features of section Acrolophus (Centaurea, Asteraceae) via scanning electron microscopy. Turk J Bot 40: 147-163.
Csupor-Loffler B, Hajdu Z, Rethy B, Zupko I, Mathe I, Redei T, Falkay G, Hohmann J (2009). Antiproliferative activity of Hungarian Asteraceae species against human cancer cell lines. Part II. Phytother Res 23: 1109-1115.
Csupor D, Widowitz U, Blazsó G, Laczkó-Zöld E, Tatsimo JS, Balogh A, Boros K, Dankó B, Bauer R, Hohmann J. (2013). Antiinflammatory activities of eleven Centaurea species occurring in the Carpathian Basin. Phytother Res 27: 540-544.
Devika PT, Stanely Mainzen Prince P (2008). (-)Epigallocatechin-gallate (EGCG) prevents mitochondrial damage in isoproterenol-induced cardiac toxicity in albino Wistar rats: a transmission electron microscopic and in vitro study. Pharmacol Res 57: 351-357.
Dural H, Bagci Y, Ertugrul K, Demirelma H, Flamini G, Cioni PL, Morelli I (2003). Essential oil composition of two endemic Centaurea species from Turkey, Centaurea mucronifera and Centaurea chrysantha, collected in the same habitat. Biochem Syst Ecol 31: 1417-1425.
Erel SB, Demir S, Nalbantsoy A, Ballar P, Khan S, Yavasoglu NU, Karaalp C (2014). Bioactivity screening of five Centaurea species and in vivo anti-inf
Erel SB, Karaalp C, Bedir E, Kaehlig H, Glasl S, Khan S, Krenn L (2011). Secondary metabolites of Centaurea calolepis and evaluation of cnicin for anti-inflammatory, antioxidant, and cytotoxic activities. Pharm Biol 49: 840-849.
Forgo P, Zupko I, Molnar J, Vasas A, Dombi G, Hohmann J (2012). Bioactivity-guided isolation of antiproliferative compounds from Centaurea jacea L. Fitoterapia 83: 921-925.
Garbacki N, Gloaguen V, Damas J, Bodart P, Tits M, Angenot L (1999). Anti-inflammatory and immunological effects of Centaurea cyanus flower-heads. J Ethnopharmacol 68: 235- 241.
Ghantous A, Tayyoun AA, Lteif GA, Saliba NA, Gali-Muhtasib H, El-Sabban M, Darwiche N (2008). Purified salograviolide A isolated from Centaurea ainetensis causes growth inhibition and apoptosis in neoplastic epidermal cells. Int J Oncol 32: 841-849.
Hammoud L, Seghiri R, Benayache S, Mosset P, Lobstein A, Chaabi M, León F, Brouard I, Bermejo J, Benayache F (2012). A new flavonoid and other constituents from Centaurea nicaeensis All. var. walliana M. Nat Prod Res 26:203-208.
Hossain MB, Rai DK, Brunton NP, Martin-Diana AB, BarryRyan C (2010). Characterization of phenolic composition in Lamiaceae spices by LC-ESI-MS/MS. J Agric Food Chem 58: 10576-10581.
Hsu S, Bollag WB, Lewis J, Huang Q, Singh B, Sharawy M, Yamamoto T, Schuster G (2003). Green tea polyphenols induce differentiation and proliferation in epidermal keratinocytes. J Pharmacol Exp Ther 306: 29-34.
Karamenderes C, Konyalioglu S, Khan S, Khan IA (2007). Total phenolic contents, free radical scavenging activities and inhibitory effects on the activation of NF-kappa B of eight Centaurea L. species. Phytother Res 21: 488-491.
Kaufmann SH, Desnoyers S, Ottaviano Y, Davidson NE, Poirier GG (1993). Specific proteolytic cleavage of poly(ADP-ribose) polymerase: an early marker of chemotherapy-induced apoptosis. Cancer Res 53: 3976-3985.
Kitouni R, Benayache F, Benayache S (2015). Flavonoids of the exudate of Centaurea calcitrapa. Chem Nat Compd 51: 762- 763.
Kolli E, Leon F, Benayache F, Estevez S, Quintana J, Estevez F, Brouard I, Bermejo J, Benayache S (2012). Cytotoxic sesquiterpene lactones and other constituents of Centaurea omphalotricha. J Brazil Chem Soc 23: 977-983.
Kontogianni VG, Tomic G, Nikolic I, Nerantzaki AA, Sayyad N, Stosic-Grujicic S, Stojanovic I, Gerothanassis IP, Tzakos AG (2013). Phytochemical profile of Rosmarinus officinalis and Salvia officinalis extracts and correlation to their antioxidant and anti-proliferative activity. Food Chem 136: 120-129.
Mosmann T (1983). Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 65: 55-63.
Pirvu L, Dragomir C, Schiopu S, Mihul SC (2012). Vegetal extracts with gastroprotective activity. Part. I. Extracts obtained from Centaurea cyanus L. raw material. Rom Biotech Lett 17: 7169- 7176.
Radic S, Prolic M, Pavlica M, Pevalek-Kozlina B (2005). Cytogenetic stability of Centaurea ragusina long-term culture. Plant Cell Tiss Org 82: 343-348.
Seghiri R, Boumaza O, Mekkiou R, Benayacheb S, Mosset P, Quintanae J, Estévez F, León F, Bermejo J, Benayach F (2009). A flavonoid with cytotoxic activity and other constituents from Centaurea africana. Phytochem Lett 2: 114-118.
Shoeb M, Jaspars M, MacManus SM, Celik S, Nahar L, KongThoo-Lin P, Sarker SD (2006). Anti-colon cancer potential of phenolic compounds from the aerial parts of Centaurea gigantea (Asteraceae). J Nat Med 61: 164-169.
Shoeb M, MacManus SM, Jaspars M, Trevidu J, Nahar L, Kong-ThooLin P, Sarker SD (2006). Montamine, a unique dimeric indole alkaloid, from the seeds of Centaurea montana (Asteraceae), and its in vitro cytotoxic activity against the CaCo-2 colon cancer cells. Tetrahedron 62: 11172-11177.
Tewari M, Quan LT, ORourke K, Desnoyers S, Zeng Z, Beidler DR, Poirier GG, Salvesen GS, Dixit VM (1995). Yama/CPP32β, a mammalian homolog of CED-3, is a CrmA-inhibitable protease that cleaves the death substrate poly (ADP-ribose) polymerase. Cell 81: 801-809.
Torre LA, Bray F, Siegel RL, Ferlay J, Lortet-Tieulent J, Jemal A (2015). Global cancer statistics, 2012. CA Cancer J Clin 65: 87-108.
Ulukaya E, Kurt A, Wood EJ (2001). 4-(N-hydroxyphenyl) retinamide can selectively induce apoptosis in human epidermoid carcinoma cells but not in normal dermal fibroblasts. Cancer Invest 19: 145-154.
Ulukaya E, Ozdikicioglu F, Oral AY, Demirci M (2008). The MTT assay yields a relatively lower result of growth inhibition than the ATP assay depending on the chemotherapeutic drugs tested. Toxicol In Vitro 22: 232-239.
Vermes I, Haanen C, Steffensnakken H, Reutelingsperger C (1995). A novel assay for apoptosis - flow cytometric detection of phosphatidylserine expression on early apoptotic cells using fluorescein-labeled annexin-V. J Immunol Methods 184: 39-51.
Wang P, Henning SM, Heber D (2010). Limitations of MTT and MTS-based assays for measurement of antiproliferative activity of green tea polyphenols. PLoS One 5:e10202.