Chalcone-Induced Apoptosis through Caspase-Dependent Intrinsic Pathways in Human Hepatocellular Carcinoma Cells
"> Figure 1
<p>Expression of hepatic markers by RT-PCR and Western blot: (<b>a</b>) Markers of hepatic mRNAs were amplified. Electrophoresis agarose seen in the amplified fragments of 310, 210 and 190 bp, corresponding to albumin, transferrin and HNF-4α, respectively. The fragment was generated from the cDNA obtained from total RNA Huh-7, HepG2 and HEPM cells. Huh-7 and HepG2 cells were used as positive control. HFK cells were used as negative control. Reverse transcriptase in the absence of product is not detected (lanes 2, 4, 6, 8). M corresponds to the marking of 100 bp molecular size; (<b>b</b>) Constitutively expressed gene 18S rRNA was used as an internal control; (<b>c</b>) Expression of the protein αSMA. By Western blot, the protein expression of αSMA was determined in the hepatic (Huh-7, HepG2 and HepM) and fibroblast cells (Fb (+)). Constitutively expressed calnexin was used as an internal control.</p> "> Figure 2
<p>Effect of the chalcones CH1 and CH2 on the growth of human hepatocellular carcinoma (HepG2 and HuH-7) and normal mouse hepatocyte (HepM). Structures of CH1 and CH2 (<b>a</b>); HepG2 (<b>b</b>,<b>c</b>); HuH-7 (<b>d</b>) and HepM (<b>e</b>) were treated with various concentrations of the compounds CH1 (Black bars) and CH2 (White bars) for 24 (<b>b</b>) and 48 (<b>c</b>–<b>e</b>) h. Chalcones combined (CH1:CH2) were used in HepG2 (<b>f</b>) and HuH-7 (<b>g</b>) cells for 48 h. Cell viability was measured using the MTT assay. Data are expressed as the mean ± SEM from three independent experiments, each performed in triplicate. Statistical differences were assessed by a one-way ANOVA (Kruskal–Wallis) followed by Dunn’s <span class="html-italic">post hoc</span> test. * <span class="html-italic">p</span> < 0.05, ** <span class="html-italic">p</span> < 0.01, *** <span class="html-italic">p</span> < 0.001 compared with the condition 0 μM, NS, Non-significant.</p> "> Figure 3
<p>HepG2 and HepM cells exposed to chalcones CH1 and CH2. HepG2 and HepM cells were grown for 24 h, and then treated with each chalcone, independently. Figures show representative phase-contrast images from at least three separates experiments of HepG2 (<b>a</b>–<b>c</b>) and HepM (<b>d</b>–<b>f</b>) exposed to vehicle (control) (<b>a</b>,<b>d</b>); 50 μM CH1 (green) (<b>b</b>,<b>e</b>); and 50 μM CH2 (green) (<b>c</b>,<b>f</b>). Bar scale represents 10 μm.</p> "> Figure 4
<p>Chalcones induced laddering and nuclear condensation of HepG2 cells a process that requires caspase activity. (<b>a</b>) Cells were treated with CH1 and CH2 (50 μM) for 24 h and preincubated with zVAD (50 μM) for 2 h (case +), when the cells were incubated in the absence of inhibitor (case -), typical laddering indicative of apoptosis was observed. DMSO was used as control positive of laddering; (<b>b</b>) HepG2 (<b>i</b>–<b>iii</b>) and HepM (<b>iv</b>–<b>vi</b>) cells were treated with vehicle (<b>i</b>,<b>iv</b>), CH1 (<b>ii</b>,<b>v</b>) and CH2 (<b>iii</b>,<b>vi</b>) for 24 h. Cells were stained with DAPI (blue) and the arrow indicated nuclear condensation. Bar scale represents 10 μm.</p> "> Figure 5
<p>Chalcone-induced apoptosis in HepG2 cells as assayed by flow cytometry. (<b>a</b>) HepG2 cells were treated with CH1 and CH2 (50 μM) for 24 h. The cells were then harvested and stained with Annexin V and PI and flow cytometric analysis was performed to analyze the apoptosis; (<b>b</b>) Summary of the apoptosis data in histogram form. Data are expressed as the mean ± standard error of the mean (SEM) from at least three independent experiments. * <span class="html-italic">p</span> < 0.05 <span class="html-italic">vs.</span> the untreated (UT) group; ** <span class="html-italic">p</span> < 0.01 <span class="html-italic">vs.</span> the untreated group.</p> "> Figure 6
<p>Chalcones do not induce caspase extrinsic pathway in HepG2 cells. (<b>a</b>–<b>d</b>) Cells were treated with CH1 and CH2 (50 μM, each) and protein expression was analyzed. (<b>a</b>,<b>c</b>) Representative images from Western blot experiments performed for the detection of caspase-8; (<b>b</b>,<b>d</b>) Densitometric analyses of the experiments shown in (<b>a</b>,<b>c</b>), respectively. Protein levels were normalized against HSP70 and data are expressed relative to the UT (untreated) condition. Data are expressed as the mean ± standard error of the mean (SEM) from at least three independent experiments. Statistical differences were assessed by a one-way ANOVA (Kruskal–Wallis) followed by Dunn’s <span class="html-italic">post hoc</span> test. NS: Not significant.</p> "> Figure 7
<p>Chalcones-induced caspase intrinsic pathway in HepG2 cells. (<b>a</b>–<b>h</b>) Cells were treated with CH1 and CH2 (50 μM, each) and protein expression was analyzed. (<b>a</b>,<b>c</b>,<b>e</b>,<b>g</b>) Representative images from Western blot experiments performed for the detection of C-caspase-9 and C-caspase-3; (<b>b</b>,<b>d</b>,<b>f</b>,<b>h</b>) Densitometric analyses of the experiments shown in (<b>a</b>,<b>c</b>,<b>e</b>,<b>g</b>), respectively. Protein levels were normalized against HSP70 and data are expressed relative to the UT (untreated) condition. Data are expressed as the mean ± standard error of the mean (SEM) from at least three independent experiments. Statistical differences were assessed by a one-way ANOVA (Kruskal–Wallis) followed by Dunn’s <span class="html-italic">post hoc</span> test. * <span class="html-italic">p</span> < 0.05; ** <span class="html-italic">p</span> < 0.01 <span class="html-italic">vs.</span> UT group.</p> "> Figure 8
<p>Chalcones-induced increase ratio Bax/Bcl-2 in HepG2 cells. (<b>a</b>–<b>h</b>) Cells were treated with CH1 and CH2 (50 μM, each) and protein expression was analyzed; (<b>a</b>,<b>c</b>,<b>e</b>,<b>g</b>) Representative images from Western blot experiments performed for the detection of Bax and Bcl-2; (<b>b</b>,<b>d</b>,<b>f</b>,<b>h</b>) Densitometric analyses of the experiments shown in (<b>a</b>,<b>c</b>,<b>e</b>,<b>g</b>), respectively. (<b>i</b>,<b>j</b>) the data were presented in the bar graphs as Bax/Bcl-2 ratio. Protein levels were normalized against HSP70 and data were expressed relative to the UT (untreated) condition. Data are expressed as the mean ± standard error of the mean (SEM) from at least three independent experiments. Statistical differences were assessed by a one-way ANOVA (Kruskal–Wallis) followed by Dunn’s <span class="html-italic">post hoc</span> test. * <span class="html-italic">p</span> < 0.05; ** <span class="html-italic">p</span> < 0.01 <span class="html-italic">vs.</span> UT group.</p> "> Figure 9
<p>Protein expression of caspase-3 and Bcl-2 in HepG2 cells as shown by immunocytochemical analysis. HepG2 cells were treated with 50 μM of CH1 for 24 h, and then immunocytochemical analyses were performed as described in Materials and Methods. Cells were fixed and immunostained with anti-tubulin antibody (green), anti-caspase-3 (red) and cell nuclei were counterstained with DAPI reagent (blue). (<b>a</b>) The results revealed that CH1 increased caspase-3 expression in HepG2 cells; (<b>b</b>) CH1 was shown to decrease Bcl-2 expression in HerpG2 cells. These findings also suggest that CH1 induces the apoptosis of HepG2 cells <span class="html-italic">in vitro</span>. Tubulin was used as expression control. Bar scale represents 20 μm.</p> "> Figure 10
<p>Protein expression of caspase-3 and Bcl-2 in HepG2 cells as shown by immunocytochemical analysis. Cells were treated with 50 μM of CH2 for 24 h, and then immunocytochemical analyses were performed as described in Materials and Methods. Cells were fixed and immunostained with anti-tubulin antibody (green), anti-caspase-3 (red) and cell nuclei were counterstained with DAPI reagent (blue). (<b>a</b>) The results revealed that CH2 increased caspase-3 expression in HepG2 cells; (<b>b</b>) CH2 was shown to decrease Bcl-2 expression in HerpG2 cells. These findings also suggest that CH2 induces the apoptosis of HepG2 cells <span class="html-italic">in vitro</span>. Tubulin was used as expression control. Bar scale represents 20 μm.</p> "> Figure 11
<p>Chalcone effects on the HepG2 cells; Intercelular reactive oxygen species generation. CH1 and CH2 enhanced cellular ROS level. Cells were exposed to CH1 and CH2 at 50 μM for 4, 8 and 24 h. (<b>a</b>) Stained cells with DCFDA and analyzed by fluorescence in a plate reader (Tecan infinite<sup>®</sup> m200pro, Grodig, Austria); (<b>b</b>) Stained cells with DAPI (blue) and DCFDA (green), and analyzed under fluorescent microscopy EVOS<sup>®</sup> FLoid<sup>®</sup> cell (Life Tehnologies, CA, USA), Black bars represents CH1 and White bars represents CH2. Data are expressed as the mean ± standard error of the mean (SEM) from three independent experiments, each performed in triplicate. Statistical differences were assessed by a one-way ANOVA (Kruskal–Wallis) followed by Dunn’s <span class="html-italic">post hoc</span> test. * <span class="html-italic">p</span> < 0.05; ** <span class="html-italic">p</span> < 0.01; *** <span class="html-italic">p</span> < 0.001 <span class="html-italic">vs.</span> Untreated (UT) group. Bar scale represents 20 μm.</p> "> Figure 12
<p>Proposed model for Chalcone-mediated apoptosis in human hepatoma cells. Solid arrow represent activation.</p> ">
Abstract
:1. Introduction
2. Results
2.1. Cytotoxic Effect of Chalcones in Human Hepatoma Cells (HepG2) and Normal Mouse Hepatocytes (HepM)
2.2. Both Chalcones Induces Apoptosis in Human Hepatoma through Caspase-Dependent Pathways
2.3. Chalcones Induces Cell Death through an Intrinsic Apoptotic Pathway
2.4. Chalcones Lead to Increased ROS
3. Discussion
4. Materials and Methods
4.1. Synthesis of Chalcone
4.2. Cell Culture
4.3. Ethics Approval
4.4. MTT Assay
4.5. Western Blot
4.6. Fluorescence Microscopy
4.7. Determination of Intracellular ROS
4.8. Annexin-V Apoptosis Assay
4.9. DNA Laddering Experiments
4.10. RT-PCR Amplification
4.11. Data Analysis
5. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
- Ferlay, J.; Shin, H.R.; Bray, F.; Forman, D.; Mathers, C.; Parkin, D.M. Estimates of worldwide burden of cancer in 2008: GLOBOCAN 2008. Int. J. Cancer 2010, 127, 2893–2917. [Google Scholar] [CrossRef] [PubMed]
- Witjes, C.D.M.; Karim-Kos, H.E.; Visser, O.; van den Akker, S.A.W.; de Vries, E.; Ijzermans, J.N.M.; de Man, R.A.; Coebergh, J.W.W.; Verhoef, C. Hepatocellular carcinoma in a low-endemic area: Rising incidence and improved survival. Eur. J. Gastroenterol. Hepatol. 2012, 24, 450–457. [Google Scholar] [CrossRef] [PubMed]
- Shen, D.W.; Lu, Y.G.; Chin, K.V.; Pastan, I.; Gottesman, M.M. Human hepatocellular carcinoma cell lines exhibit multidrug resistance unrelated to MRD1 gene expression. J. Cell Sci. 1991, 98, 317–322. [Google Scholar] [PubMed]
- Honda, K.; Sbisa, E.; Tullo, A.; Papeo, P.A.; Saccone, C.; Poole, S.; Pignatelli, M.; Mitry, R.R.; Ding, S.; Isla, A.; et al. p53 mutation is a poor prognostic indicator for survival in patients with hepatocellular carcinoma undergoing surgical tumour ablation. Br. J. Cancer 1998, 77, 776–782. [Google Scholar] [CrossRef] [PubMed]
- Mor, G.; Montagna, M.K.; Alvero, A.B. Modulation of apoptosis to reverse chemoresistance. Methods Mol. Biol. 2008, 414, 1–12. [Google Scholar] [PubMed]
- Giménez-Bonafé, P.; Tortosa, A.; Pérez-Tomás, R. Overcoming drug resistance by enhancing apoptosis of tumor cells. Curr. Cancer Drug Targets 2009, 9, 320–340. [Google Scholar] [CrossRef] [PubMed]
- Forner, A.; Llovet, J.M.; Bruix, J. Hepatocellular carcinoma. Lancet 2012, 379, 1245–1255. [Google Scholar] [CrossRef]
- Bosch, F.X.; Ribes, J.; Borràs, J. Epidemiology of primary liver cancer. Semin. Liver Dis. 1999, 19, 271–285. [Google Scholar] [CrossRef] [PubMed]
- Gridelli, C.; Rossi, A.; Maione, P.; Ferrara, M.L.; Castaldo, V.; Sacco, P.C. Vaccines for the treatment of non-small cell lung cancer: A renewed anticancer strategy. Oncologist 2009, 14, 909–920. [Google Scholar] [CrossRef] [PubMed]
- Connolly, E.C.; Saunier, E.F.; Quigley, D.; Luu, M.T.; De Sapio, A.; Hann, B.; Yingling, J.M.; Akhurst, R.J. Outgrowth of drug-resistant carcinomas expressing markers of tumor aggression after long-term TβRI/II kinase inhibition with LY2109761. Cancer Res. 2011, 71, 2339–2349. [Google Scholar] [CrossRef] [PubMed]
- Li, Y.; Tan, B.B.; Zhao, Q.; Fan, L.-Q.; Liu, Y.; Hao, Y.-J.; Zhao, X.-F. Tumor chemosensitivity is correlated with expression of multidrug resistance associated factors in variously differentiated gastric carcinoma tissues. Hepatogastroenterology 2013, 60, 213–216. [Google Scholar] [CrossRef] [PubMed]
- Efferth, T.; Giaisi, M.; Merling, A.; Krammer, P.H.; Li-Weber, M. Artesunate induces ROS-mediated apoptosis in doxorubicin-resistant T leukemia cells. PLoS ONE 2007, 2, e693. [Google Scholar] [CrossRef] [PubMed]
- Bechtel, W.; Bauer, G. Catalase protects tumor cells from apoptosis induction by intercellular ROS signaling. Anticancer Res. 2009, 29, 4541–4557. [Google Scholar] [PubMed]
- Wiseman, H.; Halliwell, B. Damage to DNA by reactive oxygen and nitrogen species: Role in inflammatory disease and progression to cancer. Biochem. J. 1996, 313, 17–29. [Google Scholar] [CrossRef] [PubMed]
- Hassan, S.K.; Mousa, A.M.; Eshak, M.G.; Farrag, A.E.R.H.; Badawi, A.E.F.M. Therapeutic and chemopreventive effects of nano curcumin against diethylnitrosamine induced hepatocellular carcinoma in rats. Int. J. Pharm. Pharm. Sci. 2014, 6, 54–62. [Google Scholar]
- Chen, B.; Ning, M.; Yang, G. Effect of paeonol on antioxidant and immune regulatory activity in hepatocellular carcinoma rats. Molecules 2012, 17, 4672–4683. [Google Scholar] [CrossRef] [PubMed]
- Zhang, C.L.; Zeng, T.; Zhao, X.L.; Yu, L.H.; Zhu, Z.P.; Xie, K.Q. Protective effects of garlic oil on hepatocarcinoma induced by N-nitrosodiethylamine in rats. Int. J. Biol. Sci. 2012, 8, 363–374. [Google Scholar] [CrossRef] [PubMed]
- Dimmock, J.R.; Elias, D.W.; Beazely, M.A.; Kandepu, N.M. Bioactivities of chalcones. Curr. Med. Chem. 1999, 6, 1125–1149. [Google Scholar] [PubMed]
- Echeverria, C.; Santibañez, J.F.; Donoso-Tauda, O.; Escobar, C.A.; Ramirez-Tagle, R. Structural antitumoral activity relationships of synthetic chalcones. Int. J. Mol. Sci. 2009, 10, 221–231. [Google Scholar] [CrossRef] [PubMed]
- Katsori, A.-M.; Hadjipavlou-Litina, D. Chalcones in cancer: Understanding their role in terms of QSAR. Curr. Med. Chem. 2009, 16, 1062–1081. [Google Scholar] [CrossRef] [PubMed]
- Shah, A.; Khan, A.M.; Qureshi, R.; Ansari, F.L.; Nazar, M.F.; Shah, S.S. Redox behavior of anticancer chalcone on a glassy carbon electrode and evaluation of its interaction parameters with DNA. Int. J. Mol. Sci. 2008, 9, 1424–1434. [Google Scholar] [CrossRef] [PubMed]
- Miranda, C.L.; Stevens, J.F.; Helmrich, A.; Henderson, M.C.; Rodriguez, R.J.; Yang, Y.H.; Deinzer, M.L.; Barnes, D.W.; Buhler, D.R. Antiproliferative and cytotoxic effects of prenylated flavonoids from hops (Humulus lupulus) in human cancer cell lines. Food Chem. Toxicol. 1999, 37, 271–285. [Google Scholar] [CrossRef]
- Nowakowska, Z. A review of anti-infective and anti-inflammatory chalcones. Eur. J. Med. Chem. 2007, 42, 125–137. [Google Scholar] [CrossRef] [PubMed]
- Zamule, S.M.; Coslo, D.M.; Chen, F.; Omiecinski, C.J. Differentiation of human embryonic stem cells along a hepatic lineage. Chem. Biol. Interact. 2011, 190, 62–72. [Google Scholar] [CrossRef] [PubMed]
- Bonzo, J.A.; Ferry, C.H.; Matsubara, T.; Kim, J.; Gonzalez, F.J. Suppression of hepatocyte proliferation by hepatocyte nuclear factor 4α in adult mice. J. Biol. Chem. 2012, 287, 7345–7356. [Google Scholar] [CrossRef] [PubMed]
- Alarcón, J.; Alderete, J.; Escobar, C.; Araya, R.; Cespedes, C.L. Aspergillus niger catalyzes the synthesis of flavonoids from chalcones. Biocatal. Biotransform. 2013, 31, 160–167. [Google Scholar] [CrossRef]
- Escobar, C.A.; Trujillo, A.; Howard, J.A.K.; Fuentealba, M. (E)-1-(3-Bromo-phen-yl)-3-(3,4-dimeth-oxy-phen-yl)prop-2-en-1-one. Acta Crystallogr. Sect. E. Struct. Rep. Online 2012, 68, o887. [Google Scholar] [CrossRef] [PubMed]
- Koiri, R.K.; Trigun, S.K. Dimethyl sulfoxide activates tumor necrosis factorα-p53 mediated apoptosis and down regulates D-fructose-6-phosphate-2-kinase and lactate dehydrogenase-5 in Dalton’s lymphoma in vivo. Leuk. Res. 2011, 35, 950–956. [Google Scholar] [CrossRef] [PubMed]
- Winter, E.; Chiaradia, L.D.; Silva, A.H.; Nunes, R.J.; Yunes, R.A.; Creczynski-Pasa, T.B. Involvement of extrinsic and intrinsic apoptotic pathways together with endoplasmic reticulum stress in cell death induced by naphthylchalcones in a leukemic cell line: Advantages of multi-target action. Toxicol. In Vitro 2014, 28, 769–777. [Google Scholar] [CrossRef] [PubMed]
- Wong, R.S.Y. Apoptosis in cancer: From pathogenesis to treatment. J. Exp. Clin. Cancer Res. 2011, 30, 87. [Google Scholar] [CrossRef] [PubMed]
- Elmore, S. Apoptosis: A review of programmed cell death. Toxicol. Pathol. 2007, 35, 495–516. [Google Scholar] [CrossRef] [PubMed]
- Conradt, B. Genetic control of programmed cell death during animal development. Annu. Rev. Genet. 2009, 43, 493–523. [Google Scholar] [CrossRef] [PubMed]
- Pilatova, M.; Varinska, L.; Perjesi, P.; Sarissky, M.; Mirossay, L.; Solar, P.; Ostro, A.; Mojzis, J. In vitro antiproliferative and antiangiogenic effects of synthetic chalcone analogues. Toxicol. Vitr. 2010, 24, 1347–1355. [Google Scholar] [CrossRef] [PubMed]
- Sashidhara, K.V.; Kumar, A.; Kumar, M.; Sarkar, J.; Sinha, S. Synthesis and in vitro evaluation of novel coumarin-chalcone hybrids as potential anticancer agents. Bioorg. Med. Chem. Lett. 2010, 20, 7205–7211. [Google Scholar] [CrossRef] [PubMed]
- Kumar, S.K.; Hager, E.; Pettit, C.; Gurulingappa, H.; Davidson, N.E.; Khan, S.R. Design, synthesis, and evaluation of novel boronic-chalcone derivatives as antitumor agents. J. Med. Chem. 2003, 46, 2813–2815. [Google Scholar] [CrossRef] [PubMed]
- Shen, K.-H.; Chang, J.-K.; Hsu, Y.-L.; Kuo, P.-L. Chalcone arrests cell cycle progression and induces apoptosis through induction of mitochondrial pathway and inhibition of nuclear factor κB signalling in human bladder cancer cells. Basic Clin. Pharmacol. Toxicol. 2007, 101, 254–261. [Google Scholar] [CrossRef] [PubMed]
- Winter, E.; Chiaradia, L.D.; de Cordova, C.A.S.; Nunes, R.J.; Yunes, R.A.; Creczynski-Pasa, T.B. Naphthylchalcones induce apoptosis and caspase activation in a leukemia cell line: The relationship between mitochondrial damage, oxidative stress, and cell death. Bioorg. Med. Chem. 2010, 18, 8026–8034. [Google Scholar] [CrossRef] [PubMed]
- Pereira, C.; Calhelha, R.C.; Barros, L.; Ferreira, I.C.F.R. Antioxidant properties, anti-hepatocellular carcinoma activity and hepatotoxicity of artichoke, milk thistle and borututu. Ind. Crops Prod. 2013, 49, 61–65. [Google Scholar] [CrossRef]
- Sikander, M.; Malik, S.; Yadav, D.; Biswas, S.; Katare, D.P.; Jain, S.K. Cytoprotective activity of a trans-chalcone against hydrogen peroxide induced toxicity in hepatocellular carcinoma (HepG2) cells. Asian Pac. J. Cancer Prev. 2011, 12, 2513–2516. [Google Scholar] [PubMed]
- Tsai, J.P.; Hsiao, P.C.; Yang, S.F.; Hsieh, S.C.; Bau, D.T.; Ling, C.L.; Pai, C.L.; Hsieh, Y.H. Licochalcone a suppresses migration and invasion of human hepatocellular carcinoma cells through downregulation of MKK4/JNK via NF-κB mediated urokinase plasminogen activator expression. PLoS ONE 2014, 9. [Google Scholar] [CrossRef] [PubMed]
- Wu, W.; Ye, H.; Wan, L.; Han, X.; Wang, G.; Hu, J.; Tang, M.; Duan, X.; Fan, Y.; He, S.; et al. Millepachine, a novel chalcone, induces G2/M arrest by inhibiting CDK1 activity and causing apoptosis via ROS-mitochondrial apoptotic pathway in human hepatocarcinoma cells in vitro and in vivo. Carcinogenesis 2013, 34, 1636–1643. [Google Scholar] [CrossRef] [PubMed]
- Miccadei, S.; di Venere, D.; Cardinali, A.; Romano, F.; Durazzo, A.; Foddai, M.S.; Fraioli, R.; Mobarhan, S.; Maiani, G. Antioxidative and apoptotic properties of polyphenolic extracts from edible part of artichoke (Cynara scolymus L.) on cultured rat hepatocytes and on human hepatoma cells. Nutr. Cancer 2008, 60, 276–283. [Google Scholar] [CrossRef] [PubMed]
- Ghosh, D.; Choudhury, S.T.; Ghosh, S.; Mandal, A.K.; Sarkar, S.; Ghosh, A.; Saha, K.D.; Das, N. Nanocapsulated curcumin: Oral chemopreventive formulation against diethylnitrosamine induced hepatocellular carcinoma in rat. Chem. Biol. Interact. 2012, 195, 206–214. [Google Scholar] [CrossRef] [PubMed]
- Park, C.-S.; Ahn, Y.; Lee, D.; Moon, S.W.; Kim, K.H.; Yamabe, N.; Hwang, G.S.; Jang, H.J.; Lee, H.; Kang, K.S.; et al. Synthesis of apoptotic chalcone analogues in HepG2 human hepatocellular carcinoma cells. Bioorg. Med. Chem. Lett. 2015, 25, 5705–5707. [Google Scholar] [CrossRef] [PubMed]
- Abu, N.; Akhtar, M.N.; Yeap, S.K.; Lim, K.L.; Ho, W.Y.; Zulfadli, A.J.; Omar, A.R.; Sulaiman, M.R.; Abdullah, M.P.; Alitheen, N.B. Flavokawain a induces apoptosis in MCF-7 and MDA-MB231 and inhibits the metastatic process in vitro. PLoS ONE 2014, 9, e105244. [Google Scholar] [CrossRef] [PubMed]
- Deb Majumdar, I.; Devanabanda, A.; Fox, B.; Schwartzman, J.; Cong, H.; Porco, J.A.; Weber, H.C. Synthetic cyclohexenyl chalcone natural products possess cytotoxic activities against prostate cancer cells and inhibit cysteine cathepsins in vitro. Biochem. Biophys. Res. Commun. 2011, 416, 397–402. [Google Scholar] [CrossRef] [PubMed]
- Tang, Y.; Simoneau, A.R.; Xie, J.; Shahandeh, B.; Zi, X. Effects of the kava chalcone flavokawain A differ in bladder cancer cells with wild-type versus mutant p53. Cancer Prev. Res. 2008, 1, 439–451. [Google Scholar] [CrossRef] [PubMed]
- Arianingrum, R.; Sunarminingsih, R.; Meiyanto, E.; Mubarika, S. Potential of a Chalcone Derivate Compound as Cancer Chemoprevention in Breast Cancer. In Proceedings of the 2012 3rd International Conference on Chemistry and Chemical Engineering IPCBEE, Singapore, 29–30 June 2012; Volume 38, pp. 41–45.
- Singh, N.; Sarkar, J.; Sashidhara, K.V.; Ali, S.; Sinha, S. Anti-tumour activity of a novel coumarin-chalcone hybrid is mediated through intrinsic apoptotic pathway by inducing PUMA and altering Bax/Bcl-2 ratio. Apoptosis 2014, 19, 1017–1028. [Google Scholar] [CrossRef] [PubMed]
- Kamada, S.; Kikkawa, U.; Tsujimoto, Y.; Hunter, T. Nuclear translocation of caspase-3 is dependent on its proteolytic activation and recognition of a substrate-like protein(s). J. Biol. Chem. 2005, 280, 857–860. [Google Scholar] [CrossRef] [PubMed]
- Raisova, M.; Hossini, A.M.; Eberle, J.; Riebeling, C.; Wieder, T.; Sturm, I.; Daniel, P.T.; Orfanos, C.E.; Geilen, C.C. The Bax/Bcl-2 ratio determines the susceptibility of human melanoma cells to CD95/Fas-mediated apoptosis. J. Investig. Dermatol. 2001, 117, 333–340. [Google Scholar] [CrossRef] [PubMed]
- Yang, E.; Zha, J.; Jockel, J.; Boise, L.H.; Thompson, C.B.; Korsmeyer, S.J. Bad, a heterodimeric partner for Bcl-XL and Bcl-2, displaces Bax and promotes cell death. Cell 1995, 80, 285–291. [Google Scholar] [CrossRef]
- Tamm, I.; Schriever, F.; Dörken, B. Apoptosis: Implications of basic research for clinical oncology. Lancet Oncol. 2001, 2, 33–42. [Google Scholar] [CrossRef]
- Liu, G. Effects of apoptosis-related proteins caspase-3, Bax and Bcl-2 on cerebral ischemia rats. Biomed. Rep. 2013, 1, 861–867. [Google Scholar] [CrossRef] [PubMed]
- Yang, B.; Johnson, T.S.; Thomas, G.L.; Watson, P.F.; Wagner, B.; Furness, P.N.; El Nahas, A.M. A shift in the Bax/Bcl-2 balance may activate caspase-3 and modulate apoptosis in experimental glomerulonephritis. Kidney Int. 2002, 62, 1301–1313. [Google Scholar] [CrossRef] [PubMed]
- Handayani, T.; Sakinah, S.; Nallappan, M.; Pihie, A.H.L. Regulation of p53-, Bcl-2- and caspase-dependent signaling pathway in xanthorrhizol-induced apoptosis of HepG2 hepatoma cells. Anticancer Res. 2007, 27, 965–971. [Google Scholar] [PubMed]
- Azad, M.B.; Chen, Y.; Gibson, S.B. Regulation of autophagy by reactive oxygen species (ROS): Implications for cancer progression and treatment. Antioxid. Redox Signal. 2009, 11, 777–790. [Google Scholar] [CrossRef] [PubMed]
- Simon, H.U.; Haj-Yehia, A.; Levi-Schaffer, F. Role of reactive oxygen species (ROS) in apoptosis induction. Apoptosis 2000, 5, 415–418. [Google Scholar] [CrossRef] [PubMed]
- Syam, S.; Abdelwahab, S.I.; Al-Mamary, M.A.; Mohan, S. Synthesis of chalcones with anticancer activities. Molecules 2012, 17, 6179–6195. [Google Scholar] [CrossRef] [PubMed]
- Belsare, D.P.; Pal, S.C.; Kazi, A.A.; Kankate, R.S.; Vanjari, S.S. Evaluation of antioxidant activity of chalcones and flavonoids. Int. J. ChemTech Res. 2010, 2, 1080–1089. [Google Scholar]
- Singh, S.; Sharma, P.K.; Kumar, N.; Dudhe, R. Anti-oxidant Activity of 2-hydroxyacetophenone Chalcone. J. Adv. Sci. Res. 2011, 2, 37–41. [Google Scholar]
- Anto, R.J.; Sukumaran, K.; Kuttan, G.; Rao, M.N.; Subbaraju, V.; Kuttan, R. Anticancer and antioxidant activity of synthetic chalcones and related compounds. Cancer Lett. 1995, 97, 33–37. [Google Scholar] [CrossRef]
- Shenvi, S.; Kumar, K.; Hatti, K.S.; Rijesh, K.; Diwakar, L.; Reddy, G.C. Synthesis, anticancer and antioxidant activities of 2,4,5-trimethoxy chalcones and analogues from asaronaldehyde: Structure-activity relationship. Eur. J. Med. Chem. 2013, 62, 435–442. [Google Scholar] [CrossRef] [PubMed]
- Clippinger, A.J.; Bouchard, M.J. Hepatitis B virus HBx protein localizes to mitochondria in primary rat hepatocytes and modulates mitochondrial membrane potential. J. Virol. 2008, 82, 6798–6811. [Google Scholar] [CrossRef] [PubMed]
- Institute of Laboratory Animal Resources Committee. Guide for the Care and Use of Laboratory Animals; The National Academies Press: Washington, DC, USA, 2011. [Google Scholar]
- Hempel, S.L.; Buettner, G.R.; O’Malley, Y.Q.; Wessels, D.A.; Flaherty, D.M. Dihydrofluorescein diacetate is superior for detecting intracellular oxidants: Comparison with 2′,7′-dichlorodihydrofluorescein diacetate, 5(and 6)-carboxy-2′,7′-dichlorodihydrofluorescein diacetate, and dihydrorhodamine 123. Free Radic. Biol. Med. 1999, 27, 146–159. [Google Scholar] [CrossRef]
- Echeverría, C.; Montorfano, I.; Cabello-Verrugio, C.; Armisén, R.; Varela, D.; Simon, F. Suppression of transient receptor potential melastatin 4 expression promotes conversion of endothelial cells into fibroblasts via transforming growth factor/activin receptor-like kinase 5 pathway. J. Hypertens. 2015, 33, 981–992. [Google Scholar] [CrossRef] [PubMed]
- Nuñez-Villena, F.; Becerra, A.; Echeverría, C.; Briceño, N.; Porras, O.; Armisén, R.; Varela, D.; Montorfano, I.; Sarmiento, D.; Simon, F. Increased expression of the transient receptor potential melastatin 7 channel is critically involved in lipopolysaccharide-induced reactive oxygen species-mediated neuronal death. Antioxid. Redox Signal. 2011, 15, 2425–2438. [Google Scholar] [CrossRef] [PubMed]
- Echeverría, C.; Becerra, A.; Nuñez-Villena, F.; Muñoz-Castro, A.; Stehberg, J.; Zheng, Z.; Arratia-Perez, R.; Simon, F.; Ramírez-Tagle, R. The paramagnetic and luminescent [Re6Se8I6]3− cluster. Its potential use as an antitumoral and biomarker agent. New J. Chem. 2012, 36, 927. [Google Scholar] [CrossRef]
- Villota, C.; Campos, A.; Vidaurre, S.; Oliveira-Cruz, L.; Boccardo, E.; Burzio, V.A.; Varas, M.; Villegas, J.; Villa, L.L.; Valenzuela, P.D.T.; et al. Expression of mitochondrial non-coding RNAs (ncRNAs) is modulated by high risk human papillomavirus (HPV) oncogenes. J. Biol. Chem. 2012, 287, 21303–21315. [Google Scholar] [CrossRef] [PubMed]
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Ramirez-Tagle, R.; Escobar, C.A.; Romero, V.; Montorfano, I.; Armisén, R.; Borgna, V.; Jeldes, E.; Pizarro, L.; Simon, F.; Echeverria, C. Chalcone-Induced Apoptosis through Caspase-Dependent Intrinsic Pathways in Human Hepatocellular Carcinoma Cells. Int. J. Mol. Sci. 2016, 17, 260. https://doi.org/10.3390/ijms17020260
Ramirez-Tagle R, Escobar CA, Romero V, Montorfano I, Armisén R, Borgna V, Jeldes E, Pizarro L, Simon F, Echeverria C. Chalcone-Induced Apoptosis through Caspase-Dependent Intrinsic Pathways in Human Hepatocellular Carcinoma Cells. International Journal of Molecular Sciences. 2016; 17(2):260. https://doi.org/10.3390/ijms17020260
Chicago/Turabian StyleRamirez-Tagle, Rodrigo, Carlos A. Escobar, Valentina Romero, Ignacio Montorfano, Ricardo Armisén, Vincenzo Borgna, Emanuel Jeldes, Luis Pizarro, Felipe Simon, and Cesar Echeverria. 2016. "Chalcone-Induced Apoptosis through Caspase-Dependent Intrinsic Pathways in Human Hepatocellular Carcinoma Cells" International Journal of Molecular Sciences 17, no. 2: 260. https://doi.org/10.3390/ijms17020260