The anti-tumor effects of catechins and gallic acid (GA) in-vitro was investigated in this paper. Fluo-3AM, Calcium-AM (Ca-AM), 2′, 7′ -dichlorofluorescein-diacetate(DCFH-DA), 4′, 6-diamidino-2- phenylindole (DAPI) and Ca-AM plus colbat were used to characterize intracellular calcium, labile iron pool (LIP), reactive oxygen species (ROS), nuclei morphology and mitochondrial permeability transition pore (mPTP) opening, respectively. High performance liquid chromatography (HPLC) was used to quantitate catechins and GA in the cultural medium. The results indicated that each of them showed dose response inhibition of cell growth, provoking nuclei condensation, intracellular calcium elevation, mPTP opening, LIP reduction, and cytochrome c (Cyt-C) to release into cytosol. The caspase inhibitors, 2-aminoethoxydiphenol borate (APB) or Fe3+ could inhibit lethal effects of GA and (-)-epigallocatechin (EGC), but failed to affect (-)-epigallocatechin gallate (EGCG) and (-)-epicatechin gallate (ECG). Level of ROS presented negative growth while their concentration decreased in the medium. In conclusion, our findings suggest that viability of RKO decreased because of their good correlation with elevation of calcium and loss of LIP and ROS in cytosol.
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Ahmedin J, Freddie B, Melissa M C, et al. Global Cancer statistics [J]. CA: A Cancer Journal for Clinicians, 2011, 61(2): 69–90.
Rebecca S, Elizabeth W, Otis B, et al. Cancer Statistics, 2011: The impact of eliminating socioeconomic and racial disparities on premature cancer deaths [J]. CA: A Cancer Journal for Clinicians, 2011, 61(4): 212–236.
Watson A J M, Collins P D. Colon Cancer: A civilization disorder [J]. Digestive Diseases, 2011, 29(2):222–228.
Cunningham D, Atkin W, Lenz H J, et al. Colorectal cancer [J]. The Lancet, 2010, 375(9719):1030–1047.
Ravishankar D, Rajora A K, Greco F, et al. Flavonoids as prospective compounds for anti-cancer therapy [J]. The International Journal of Biochemistry & Cell Biology, 2013, 45(12):2821–2831.
Mascitelli L, Goldstein M R. Inhibition of iron absorption by polyphenols as an anti-cancer mechanism [J]. The Quarterly journal of medicine, 2011, 104(5):459–461.
Lin J K, Liang Y C, Lin-Shiau S Y. Cancer chemoprevention by tea polyphenols through mitotic signal transduction blockade [J]. Biochemical Pharmacology, 1999, 58(6):911–915.
Lea M A, Xiao Q, Sadhukhan A K, et al. Inhibitory effects of tea extracts and (-)-epigallocatechin gallate on DNA synthesis and proliferation of hepatoma and erythroleukemia cells [J]. Cancer Letters, 1993, 68(2–3): 231–236.
Valcic S, Timmermann B N, Alberts D S, et al. Ihibitory effect of six green tea catechins and caffeine on the growth of four selected human tumor cell lines [J]. Anti-Cancer Drugs, 1996, 7(4):461–468.
Yang C S, Wang Z Y. Tea and Cancer [J]. Journal of the National Cancer Institute, 1993, 85(13): 1038–1049.
Yuan J M. Cancer prevention by green tea: Evidence from epidemiologic studies [J]. The American Journal of Clinical Nutrition, 2013, 98(6):1676S–1681S.
Lambert J D. Does tea prevent cancer? Evidence from laboratory and human intervention studies [J]. The American Journal of Clinical Nutrition, 2013, 98(6):1667S–1675S.
Khan N, Mukhtar H. Multitargeted therapy of cancer by green tea polyphenols [J]. Cancer Letters, 2008, 269(2): 269–280.
Ramos S. Effects of dietary flavonoids on apoptotic pathways related to cancer chemoprevention [J]. The Journal of Nutritional Biochemistry, 2007, 18(7):427–442.
Baek S J, Kim J S, Jackson F R, et al. Epicatechin gallate-induced expression of NAG-1 is associated with growth inhibition and apoptosis in colon cancer cells [J]. Carcinogenesis, 2004, 25(12):2425–2432.
Hong J, Smith T J, Ho C T, et al. Effects of purified green and black tea polyphenols on cyclooxygenase and lipoxygenase–dependent metabolism of arachidonic acid in human colon mucosa and colon tumor tissues [J]. Biochemical Pharmacology, 2001, 62(9):1175–1183.
Larsen C A, Bisson W H, Dashwood R H. Tea catechins inhibit hepatocyte growth factor receptor (MET kinase) activity in human colon cancer cells: Kinetic and molecular docking studies [J]. Journal of Medicinal Chemistry, 2009, 52(21):6543–6545.
Berger N A, Petzold S J. Identification of minimal size requirements of DNA for activation of poly(ADP-ribose) polymerase [J]. Biochemistry, 1985, 24(16):4352–4355.
Hou Z, Lambert J D, Chin K V, et al. Effects of tea polyphenols on signal transduction pathways related to cancer chemoprevention [J]. Mutation Research, 2004, 555(1–2):3–19.
Baumgartner H K, Gerasimenko J V, Thorne C, et al. Calcium elevation in mitochondria is the main Ca2+ requirement for mitochondrial permeability transition pore (mPTP) opening [J]. The Journal of Biological Chemistry, 2009, 284(31):20796–20803.
Xiao J X, Huang G Q, Chi Y S, et al. Soy isoflavones induced apoptosis, cell cycle arrest and [Ca2+]i elevation in hela cells [J]. Acta Nutrimenta Sinica, 2012, 34(4):373–378 (Ch).
Liu X P, Wen X L, Zou S N, et al. Induction of apoptosis by Epigallocatechin-3-gallate via activating mitochondrial signaling in human gastric cancer cells [J]. Journal of Nanhua University (Medical Edition), 2007, 35(4): 499–502 (Ch).
Chitambar C R, Narasimhan J. Targeting iron-dependent DNA synthesis with gallium and transferrin-gallium [J]. Pathobiology, 1991, 59(1):3–10.
Muñoz M, Villar I, García-Erce J A. An update on iron physiology [J]. World Journal of Gastroenterology, 2009, 15(37):4617–4626.
Yang S F, Xia H Y, Zhou D, et al. Recent progress on mitochondrial iron metabolism and human diseases [J]. Chinese Bulletin of Life Sciences, 2012, 24(8): 742–752 (Ch).
Ramiro-Cortés Y, Morán J. Role of oxidative stress and JNK pathway in apoptotic death induced by potassium deprivation and staurosporine in cerebellar granule neurons [J]. Neuro–chemistry International, 2009, 55(7):581–592.
Lipinski P, Drapier J C, Oliveira L, et al. Intracellular iron status as a hallmark of mammalian cell susceptibility to oxidative stress: a study of L5178Y mouse lymphoma cell lines differentially sensitive to H2O2 [J]. Blood, 2000, 95(9):2960–2966.
Epsztejn S, Kakhlon O, Glickstein H, et al. Fluorescence analysis of the Labile Iron Pool of mammalian cells [J]. Analytical Biochemistry, 1997, 248(1):31–40.
Berridge M J. Inositol trisphosphate and calcium signalling mechanisms [J]. Biochimica et Biophysica Acta, 2009, 1793(6):933–940.
Berridge M J, Bootman M D, Lipp P. Calcium—A life and death signal [J]. Nature, 1998, 395(6703): 645–648.
Gaido M L, Cidlowski J A. Identification, purification, and characterization of a calcium-dependent endonuclease (NUC18) from apoptotic rat thymocytes. NUC18 is not histone H2B [J]. The Journal of Biological Chemistry, 1991, 266(28):18580–18585.
Cohen J J, Duke R C. Glucocorticoid activation of a calciumdependent endonuclease in thymocyte nuclei leads to cell death [J]. The Journal of Immunology, 1984, 132(1): 38–42.
Zhivotovsky B, Nicotera P, Bellomo G, et al. Ca2+ and Endonuclease Activation in Radiation-Induced Lymphoid Cell Death [J]. Experimental Cell Research, 1993, 207(1): 163–170.
Bootman M D, Collins T J, Mackenzie L, et al. 2-aminoethoxydiphenyl borate (2-APB) is a reliable blocker of store-operated Ca2+ entry but an inconsistent inhibitor of InsP3-induced Ca2+ release [J]. The FASEB Journal, 2002, 16(10):1145–1150.
Nicholson D W, Ali A, Thornberry N A, et al. Identification and inhibition of the ICE/CED-3 protease necessary for mammalian apoptosis [J]. Nature, 1995, 376(6535):37–43.
Tewari M, Quan L T, O'Rourke K, et al. Yama/CPP32 beta, a mammalian homolog of CED-3, is a CrmA-inhibitable protease that cleaves the death substrate poly(ADP-ribose) polymerase [J]. Cell, 1995, 81(5):801–809.
Le R Y, Kirkland J B, Shah G M. Cellular responses to DNA damage in the absence of poly(ADP-ribose) polymerase [J]. Biochemical and Biophysical Research Communications, 1998, 245(1):1–10.
Nie G, Chen G, Sheftel A D, et al. In vivo tumor growth is inhibited by cytosolic iron deprivation caused by the expression of mitochondrial ferritin [J]. Blood, 2006, 108(7):2428–2434.
López-Lázaro M. Dual role of hydrogen peroxide in cancer: possible relevance to cancer chemoprevention and therapy [J]. Cancer Letters, 2007, 252(1):1–8.
Elizabeth A V, Alison M D, Brain A M. Hydrogen peroxide sensing and Signaling [J]. Molecular Cell, 2007, 26(1): 1–14.
Hancock J T, Desikan R, Neill S J. Role of reactive oxygen species in cell signalling pathways [J]. Biochemical Society Transactions, 2001, 29(2):345–350.
Mitchell C A. The colorimetric estimation of pyrogallol, gallotannin and gallic acid[J]. Analyst, 1923, 562(48): 2–15.
Srichairatanakool S, Kulprachakarn K, Pangjit K, et al. Green tea extract and epigallocatechin 3-gallate reduced Labile Iron Pool and protected oxidative stress in iron-loaded cultured hepatocytes [J]. Advances in Bioscience and Biotechnology, 2012, 3(8): 1140–1150.
Foundation item: Supported by Zhejiang Provincial Natural Science Foundation of China (Y3100683)
Biography: TU Yunfei, male, M.D., research direction: active components of tea and their anti-cancer activity.
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Tu, Y. Apoptosis of RKO induced by catechins and GA through Ca2+ and LIP. Wuhan Univ. J. Nat. Sci. 19, 341–349 (2014). https://doi.org/10.1007/s11859-014-1023-3
- tea polyphenols
- labile iron pool
- reactive oxygen species