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Sample records for cell lines ht-29

  1. Inhibitory Effects of Probiotic Lactobacillus on the Growth of Human Colonic Carcinoma Cell Line HT-29

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    Zhung-Yuan Chen

    2017-01-01

    Full Text Available This study was conducted to investigate the inhibitory effect of Lactobacillus cells and supernatants on the growth of the human colon cancer cell line HT-29. Our study results indicated that the PM153 strain exhibits the best adhesion ability and the highest survival in the gastrointestinal tract simulation experiment. Furthermore, after an 8-h co-culture of PM153 and HT-29 cells, the PM153 strain can induce the secretion of nitric oxide from the HT-29 cells. In addition, after the co-culture of the BCRC17010 strain (109 cfu/mL and HT-29 cells, the Bax/Bcl-2 ratio in the HT-29 cells was 1.19, which showed a significant difference from the other control and LAB groups (p < 0.05, which therefore led to the inference that the BCRC17010 strain exerts a pro-apoptotic effect on the HT-29 cells. Upon co-culture with HT-29 cells for 4, 8 and 12 h, the BCRC14625 strain (109 cfu/mL demonstrated a significant increase in lactate dehydrogenase (LDH activity (p < 0.05, causing harm to the HT-29 cell membrane; further, after an 8-h co-culture with the HT-29 cells, it induced the secretion of nitric oxide (NO from the HT-29 cells. Some lactic acid bacteria (LAB strains have ability to inhibit the growth of the colorectal cancer cell line HT-29 Bax/Bcl-2 pathway or NO production. In summary, we demonstrated that the BCRC17010 strain, good abilities of adhesion and increased LDH release, was the best probiotic potential for inhibition of HT-29 growth amongst the seven LAB strains tested in vitro.

  2. Inhibitory Effects of Probiotic Lactobacillus on the Growth of Human Colonic Carcinoma Cell Line HT-29.

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    Chen, Zhung-Yuan; Hsieh, You-Miin; Huang, Chun-Chih; Tsai, Cheng-Chih

    2017-01-10

    This study was conducted to investigate the inhibitory effect of Lactobacillus cells and supernatants on the growth of the human colon cancer cell line HT-29. Our study results indicated that the PM153 strain exhibits the best adhesion ability and the highest survival in the gastrointestinal tract simulation experiment. Furthermore, after an 8-h co-culture of PM153 and HT-29 cells, the PM153 strain can induce the secretion of nitric oxide from the HT-29 cells. In addition, after the co-culture of the BCRC17010 strain (10⁸ cfu/mL) and HT-29 cells, the Bax/Bcl-2 ratio in the HT-29 cells was 1.19, which showed a significant difference from the other control and LAB groups ( p strain exerts a pro-apoptotic effect on the HT-29 cells. Upon co-culture with HT-29 cells for 4, 8 and 12 h, the BCRC14625 strain (10⁸ cfu/mL) demonstrated a significant increase in lactate dehydrogenase (LDH) activity ( p strains have ability to inhibit the growth of the colorectal cancer cell line HT-29 Bax/Bcl-2 pathway or NO production. In summary, we demonstrated that the BCRC17010 strain, good abilities of adhesion and increased LDH release, was the best probiotic potential for inhibition of HT-29 growth amongst the seven LAB strains tested in vitro.

  3. In Vitro Study of Influence of Au Nanoparticles on HT29 and SPEV Cell Lines

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    Pavlovich, Elena; Volkova, Nataliia; Yakymchuk, Elena; Perepelitsyna, Olena; Sydorenko, Michail; Goltsev, Anatoliy

    2017-08-01

    Cell culture models are excellent tools for potential toxicity of nanoparticles and fundamental investigations in cancer research. Thus, information about AuNP potential toxicity and effects on human health is necessary for the use of nanomaterials in clinical settings. The aim of our research is to examine the effects of AuNPs on the epithelial origin cell lines: continuous and oncogenic. Embryonic porcine kidney epithelial inoculated (SPEV) cell line and colorectal carcinoma cell line (HT29) were used. In the test cultures, the cell proliferation, necrosis/apoptosis, and multicellular spheroids generation were evaluated. We demonstrated that AuNP concentrations of 6-12 μg/ml reduced the proliferation of SPEV and HT29 cells and increased the cell number at early and late stages of apoptosis and necrosis. It was shown that small concentrations of AuNPs (1-3 μg/ml) stimulate multicellular spheroid formation by HT29 and SPEV cells. However, higher AuNP concentrations (6-12 μg/ml) had both cytotoxic and anti-cohesive effects on cell in suspension. The large sensitiveness to the action of AuNPs was shown by the line of HT29 (6 μg/ml) as compared to the SPEV cells (12 μg/ml). This experimental study of the effect of AuNPs on SPEV and HT29 cell lines will justify their further application in AuNP-mediated anticancer treatment.

  4. Characterization of four clones derived from human adenocarcinoma cell line, HT29, and analysis of their response to sodium butyrate

    Czech Academy of Sciences Publication Activity Database

    Štokrová, Jitka; Šloncová, Eva; Sovová, Vlasta; Kučerová, Dana; Žíla, Vojtěch; Turečková, Jolana; Vojtěchová, Martina; Korb, Jan; Tuháčková, Zdena

    2006-01-01

    Roč. 28, č. 2 (2006), s. 559-565 ISSN 1019-6439 R&D Projects: GA AV ČR(CZ) KJB5052302 Institutional research plan: CEZ:AV0Z50520514 Keywords : adenocarcinoma * HT29 cell line * ultrastructure analysis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.556, year: 2006

  5. Lichen secondary metabolites are responsible for induction of apoptosis in HT-29 and A2780 human cancer cell lines.

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    Bačkorová, M; Jendželovský, R; Kello, M; Bačkor, M; Mikeš, J; Fedoročko, P

    2012-04-01

    Lichens are a known source of approximately 800 unique secondary metabolites, many of which play important ecological roles, including regulating the equilibrium between symbionts. However, only a few of these compounds have been assessed for their effectiveness against various in vitro cancer models. Moreover, the mechanisms of biological activity of lichen secondary metabolites on living cells (including cancer cells) are still almost entirely unknown. In the present study, we investigated the mechanisms of cytotoxicity of four lichen secondary metabolites (parietin, atranorin, usnic acid and gyrophoric acid) on A2780 and HT-29 cancer cell lines. We found that usnic acid and atranorin were more effective anti-cancer compounds when compared to parietin and gyrophoric acid. Usnic acid and atranorin were capable of inducing a massive loss in the mitochondrial membrane potential, along with caspase-3 activation (only in HT-29 cells) and phosphatidylserine externalization in both tested cell lines. Induction of both ROS and especially RNS may be responsible, at least in part, for the cytotoxic effects of the tested compounds. Based on the detection of protein expression (PARP, p53, Bcl-2/Bcl-xL, Bax, p38, pp38) we found that usnic acid and atranorin are activators of programmed cell death in A2780 and HT-29, probably through the mitochondrial pathway. Copyright © 2012 Elsevier Ltd. All rights reserved.

  6. Synergistic inhibitory effects of curcumin and 5-fluorouracil on the growth of the human colon cancer cell line HT-29.

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    Du, Boyu; Jiang, Liping; Xia, Quan; Zhong, Laifu

    2006-01-01

    The synergistic effect of combination treatment with COX-2 inhibitors and chemotherapy may be another promising therapy regimen in the future treatment of colorectal cancer. Curcumin, a major yellow pigment in turmeric which is used widely all over the world, inhibits the growth of human colon cancer cell line HT-29 significantly and specifically inhibits the expression of COX-2 protein. However, the worldwide exposure of populations to curcumin raised the question of whether this agent would enhance or inhibit the effects of chemotherapy. In this report, we evaluated the growth-inhibitory effect of curcumin and a traditional chemotherapy agent, 5-FU, against the proliferation of a human colon cancer cell line (HT-29). The combination effect was quantitatively determined using the method of median-effect principle and the combination index. The inhibition of COX-2 expression after treatment with the curcumin-5-FU combination was also evaluated by Western blot analysis. The IC(50) value in the HT-29 cells for curcumin was 15.9 +/- 1.96 microM and for 5-FU it was 17.3 +/- 1.85 microM. When curcumin and 5-FU were used concurrently, synergistic inhibition of growth was quantitatively demonstrated. The level of COX-2 protein expression was reduced almost 6-fold after the combination treatment. Our results demonstrate synergism between curcumin and 5-FU at higher doses against the human colon cancer cell line HT-29. This synergism was associated with the decreased expression of COX-2 protein. Copyright 2006 S. Karger AG, Basel.

  7. HT-29 and Caco-2 Reporter Cell Lines for Functional Studies of Nuclear Factor Kappa B Activation

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    Giuliana Mastropietro

    2015-01-01

    Full Text Available The NF-κB is a transcription factor which plays a key role in regulating biological processes. In response to signals, NF-κB activation occurs via phosphorylation of its inhibitor, which dissociates from the NF-κB dimer allowing the translocation to the nucleus, inducing gene expression. NF-κB activation has direct screening applications for drug discovery for several therapeutic indications. Thus, pathway-specific reporter cell systems appear as useful tools to screen and unravel the mode of action of probiotics and natural and synthetic compounds. Here, we describe the generation, characterization, and validation of human epithelial reporter cell lines for functional studies of NF-κB activation by different pro- and anti-inflammatory agents. Caco-2 and HT-29 cells were transfected with a pNF-κB-hrGFP plasmid which contains the GFP gene under the control of NF-κB binding elements. Three proinflammatory cytokines (TNF-α, IL-1β, and LPS were able to activate the reporter systems in a dose-response manner, which corresponds to the activation of the NF-κB signaling pathway. Finally, the reporter cell lines were validated using lactic acid bacteria and a natural compound. We have established robust Caco-2-NF-κB-hrGFP and HT-29-NF-κB-hrGFP reporter cell lines which represent a valuable tool for primary screening and identification of bacterial strains and compounds with a potential therapeutic interest.

  8. Cytotoxic Activity of Kenaf Seed Oils from Supercritical Carbon Dioxide Fluid Extraction towards Human Colorectal Cancer (HT29 Cell Lines

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    Siti Aisyah Abd Ghafar

    2013-01-01

    Full Text Available Kenaf (Hibiscus cannabinus from the family Malvaceae, is a valuable fiber plant native to India and Africa and is currently planted as the fourth commercial crop in Malaysia. Kenaf seed oil contains alpha-linolenic acid, phytosterol such as β-sitosterol, vitamin E, and other antioxidants with chemopreventive properties. Kenaf seeds oil (KSO was from supercritical carbon dioxide extraction fluid (SFE at 9 different permutations of parameters based on range of pressures from 200 to 600 bars and temperature from 40 to 80°C. They were 200/40, 200/60, 200/80, 400/40, 400/60, 400/80, 600/40, 600/60, and 600/80. Extraction from 9 parameters of KSO-SFE was screened for cytotoxicity towards human colorectal cancer cell lines (HT29 and mouse embryonic fibroblast (NIH/3T3 cell lines using MTS assay. KSO-SFE at 600/40 showed the strongest cytotoxicity towards HT29 with IC50 of 200 µg/mL. The IC50 for NIH/3T3 was not detected even at highest concentration employed. Cell cycle analysis showed a significant increase in the accumulation of KSO-SFE-treated cells at sub-G1 phase, indicating the induction of apoptosis by KSO-SFE. Further apoptosis induction was confirmed by Annexin V/PI and AO/PI staining.

  9. Effects of lactoferrin on the production of interferon-λ by the human intestinal epithelial cell line HT-29.

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    Shin, Kouichirou; Oda, Hirotsugu; Wakabayashi, Hiroyuki; Yamauchi, Koji; Abe, Fumiaki

    2017-02-01

    We examined the in-vitro effects of bovine lactoferrin (LF) on the production of interferon-λ (IFN-λ), an antiviral cytokine important for the defense of enterocytes, using the human intestinal epithelial cell line HT-29. HT-29 cell cultures were treated with LF for 1 h, and the cultures were stimulated with polyinosinic-polycytidylic acid (poly I:C). LF increased the concentration of IFN-λ in the culture supernatant after stimulation in a dose-dependent manner. A similar increase in the concentration of IFN-λ was observed in the supernatant of cells washed between treatment with LF and stimulation with poly I:C. At 6 and 24 h after stimulation with poly I:C (early and late phases, respectively) treated cultures contained significantly higher concentrations of IFN-λ1 in the culture supernatant, and significantly higher IFN-λ1 and IFN-λ2 mRNA levels, than controls. These results suggest that LF activates the innate cellular immunity of the enterocytes to double-stranded RNA and increases the production of IFN-λ.

  10. Cytotoxic activity of isolated constituents from leaves of Premna serratifolia on MCF-7 and HT-29 cell lines

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    Mahesh Biradi

    2015-03-01

    Full Text Available Premna serratifolia (Syn: Premna integrifolia is an important medicinal herb known as “Agnimantha” in Ayurveda and traditionally used for anticancer activity. The objective of present study was to isolate the cytotoxic phytoconstituents from the n-hexane soluble fraction of P. serratifolia leaf extract. Unsaponifiable portion of n-hexane soluble fraction was subjected to silica based column chromatography. The major constituents present in all the sub-fractions were identified by TLC and phytochemical tests. Two constituents were isolated and they were purified. Sub-fractions with isolates were tested for cytotoxic effect by BSL bioassay. Two isolates were found to be active and which were tested on cancer cell lines MCF-7 and HT-29 for their cytotoxicity. Among two isolates, one compound has shown significant cytotoxicity. From the results we conclude that the plant isolates showed cytotoxicity against selected human cancer cell lines.

  11. Use of HT-29, a cultured human colon cancer cell line, to study the effect of fermented milks on colon cancer cell growth and differentiation.

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    Baricault, L; Denariaz, G; Houri, J J; Bouley, C; Sapin, C; Trugnan, G

    1995-02-01

    Epidemiological and in vivo and in vitro experimental studies have suggested that fermented milks may interfere with the emergence and/or the development of colon cancer. The results, however, remain inconclusive. This prompted us to develop a new approach based on the use of HT-29, a cultured human colon cancer cell line, to study at the cellular level the effect of fermented milks on colon cancer cell growth and differentiation characteristics. Undifferentiated HT-29 cells have been grown in the continuous presence of milks fermented by one of the following bacterial populations: Lactobacillus helveticus, Bifidobacterium, L.acidophilus or a mix of Streptococcus thermophilus and L. bulgaricus. Penicillin G was added to the cell culture medium, resulting in a complete blockade of bacterial growth without significant effect on bacterial viability. One out of the four bacteria species studied, namely L.acidophilus, was without effect on both cell growth and differentiation. The three other bacterial strains induced a significant, although variable, reduction in the growth rate of HT-29 cells, which resulted in a 10-50% decrease in the cell number at steady-state (i.e. at cell confluency). The most efficient strains in lowering the HT-29 growth rate were L. helveticus and Bifidobacterium. Concomitantly, the specific activities of dipeptidyl peptidase IV (DPP IV), a sensitive and specific marker of HT-29 cell differentiation, and that of three other brush border enzymes (sucrase, aminopeptidase N and alkaline phosphatase) were significantly increased, thus suggesting that these cells may have entered a differentiation process. Altogether, these results indicate that the use of cultured colon cancer cells may be a useful tool to further study the effect of fermented milks on colon cancer and that bacterial strains may exert a different and specific effect on cancer cell growth and differentiation when used in fermented milk products.

  12. Sensitization to oxaliplatin in HCT116 and HT29 cell lines by metformin and ribavirin and differences in response to mitochondrial glutaminase inhibition

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    Silvina M Richard

    2015-01-01

    Full Text Available Aim of study: In the present study, we evaluated the effect of ribavirin and metformin on the sensitivity of oxaliplatin and 5-fluorouracil (5-FU on colon cancer. Materials and Methods: Cell viability of two commercially available colon cancer cell lines (HT29 and HCT116 were analyzed by sulforhodamine B (SRB assay. Results: A clinically achievable and nontoxic concentration of ribavirin and metformin showed a significant synergistic effect on oxaliplatin in HT29 and HCT116 cell lines. Ribavirin showed a synergistic effect on oxaliplatin in HT29 (R = 2.93, P < 0.001 and HCT116 (R = 1.71, P < 0.001, while only in HT29 metformin synergized with oxaliplatin by 2.66 (± 0.28, P < 0.01. In addition, both cell lines showed significant differences in response to Compound 968, inhibitor of mitochondrial glutaminase activity. Conclusion: The data suggested that these cell lines not only turn to metabolic different sustainability process after oxaliplatin treatment but that they also have different basal metabolic requirements of glutamine in vitro which can be exploits in the future for colorectal cancer (CRC treatment and further studies are required.

  13. Prodigiosin, the red pigment of Serratia marcescens, shows cytotoxic effects and apoptosis induction in HT-29 and T47D cancer cell lines.

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    Dalili, D; Fouladdel, Sh; Rastkari, N; Samadi, N; Ahmadkhaniha, R; Ardavan, A; Azizi, E

    2012-01-01

    In this study, a red pigment of Serratia marcescens PTCC 1111 was purified and identified for antiproliferative activities in HT-29 and T47D cancer cell lines. (1)H-NMR spectroscopy and LC/MS analysis confirmed prodigiosin structure. The antiproliferative effects of prodigiosin were determined by employing the MTT assay. The changes in cell cycle pattern were studied with 4',6-diamidino-2-phenylindole (DAPI) reagent using flow cytometry assay, and Annexin V-PI method was used for apoptotic analysis. Results of MTT assay showed that HT-29 cells were more sensitive to prodigiosin than T47D cells. Prodigiosin-treated HT-29 cells showed increase in S phase and decrease in G2/M, but treated T47D cells showed cell cycle pattern relatively similar to Roswell Park Memorial Institute medium (RPMI). Apoptotic effect of prodigiosin was higher than doxorubicin in HT-29 cells. The data reported here indicate that prodigiosin is a promising antineoplastic agent that triggers apoptosis in different cancer cell lines.

  14. Quercetin induces cell cycle arrest and apoptosis in CD133+ cancer stem cells of human colorectal HT29 cancer cell line and enhances anticancer effects of doxorubicin

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    Shekoufeh Atashpour

    2015-07-01

    Conclusion:The CSCs were a minor population with a significantly high level of drug resistance within the HT29 cancer cells. Quercetin alone exhibited significant cytotoxic effects on HT29 cells and also increased cytoxicity of Dox in combination therapy. Altogether, our data showed that adding quercetin to Dox chemotherapy is an effective strategy for treatment of both CSCs and bulk tumor cells.

  15. Extracellular synthesis of Selenium nanoparticles from an Actinomycetes Streptomyces griseoruber and evaluation of its cytotoxicity on HT-29 cell line.

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    Ranjitha, V R; Vittal, Ravishankar Rai

    2017-11-13

    In the present study, the extracellular synthesis of Selenium nanoparticles (SeNPs) was carried out by using culture supernatant of Streptomyces griseoruber, an Actinomycetes member isolated from the soil. Bioreduction of SeNPs was confirmed by UV-Visible spectrophotometer that showed the peak at 575 nm. XRD pattern confirmed the non-crystalline or amorphous nature of the synthesised SeNPs. FTIR spectra revealed the possible functional group that is responsible for mediating the SeNPs formation. Size and distribution of the biosynthesised SeNPs were analysed by HR-TEM that showed the formation of particle size in the range of 100-250nm. The synthesised SeNPs showed good cytotoxic activity against HT-29 cell lines where apoptosis was studied by different staining procedures. The study reports for the first time, the rapid and eco-friendly synthesis of SeNPs from Streptomyces griseoruber and opens up a new possibility of commercially producing the SeNPs from biorenewable source as a natural chemotherapeutic compound. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  16. CytotoxicEffect of Curcumin on Proliferation of HT_29 Cell Line

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    Mohamad Nabiuni

    2017-10-01

    Conclusion:According to molecular mechanisms of cell proliferation and curcumin ability in the induction of pro_apoptotic proteins and the inhibition of anti_apoptotic proteins as well as inhibition of as survival pathways,like NF_KB and AKT, this predisposition makes curcumin a good anticancer drug.

  17. Tratamiento y post-tratamiento con lonidamina en la línea celular de carcinoma colónico humano HT-29 Treatment and post-treatment with lonidamine in human colon carcinoma HT-29 cell line

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    Alicia G. Fuchs

    2008-02-01

    Full Text Available Lonidamina (1-[ 2,4-diclorofenil metil]-1H indazol-3-ácido carboxílico, (lnd, es una droga antineoplásica cuyo mecanismo de acción se ejerce sobre el metabolismo intermedio de la glucosa. Los efectos de la lnd sobre el crecimiento celular y el metabolismo celular se investigaron en las células HT- 29, línea celular de carcinoma colónico humano, que requiere altas concentraciones de glucosa para su crecimiento indiferenciado en cultivo. La lnd en dosis de 0.2 mM disminuyó significativamente el crecimiento celular y la formación de colonias en agar; con la interrupción del tratamiento se observó el restablecimiento del crecimiento celular en 24 horas. El tratamiento con lnd produce la redistribución de los glicoconjugados y el receptor de la manosa, sin afectar en forma drástica la síntesis de glucógeno ni la de proteínas. Estas posiblemente sean las causas de la rápida reversibilidad del tratamiento.Lonidamine (1-[ 2,4-dichlorophenyl methyl]-1H indazole-3-carboxylic acid, lnd, is an antitumoral drug acting on mitochondria and glucose metabolism. Cell growth and metabolic effects of lnd and drug post-treatment effect were investigated in undifferentiated HT-29 human colonic carcinoma cell line which requires high glucose medium concentration for growth. 0.2 mM lnd significantly decreased cell spreading and growth in monolayer or agar cell culture. After drug treatment cell growth was reestablished to control value within 24 h. Ind modified glycoconjugates and mannose-receptor distribution (analyzed by confocal microscopy, while glucose-glycogen and protein synthesis were not affected, these being the possible reasons for the fast reversible effect.

  18. Regulatory effect of evodiamine on the malignant biological behaviors and Wnt/β-catenin signaling pathway of colorectal cancer cell lines HT29

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    Yuan-Hui Wang

    2016-04-01

    Full Text Available Objective: To study the regulatory effect of evodiamine on the malignant biological behaviors and Wnt/β-catenin signaling pathway of colorectal cancer cell lines HT29. Methods: Colorectal cancer cell lines HT29 were cultured and divided into blank control group and evodiamine group, and after different treatment, cell viability, proportion of different cell cycle as well as the contents of VEGFA, VEGFB, VEGFC, MMP3, MMP14, Wnt and β-catenin were detected. Results: (1 Cell viability: MTT value of evodiamine group was significantly lower than that of blank control group; (2 Cell cycle: proportion of both S phase and G2/M phase of evodiamine group were lower than those of blank control group, and proportion of G0/ G1 phase was higher than that of blank control group; (3 VEGF and MMP contents: VEGFA, VEGFB, VEGFC, MMP3 and MMP14 contents of evodiamine group were lower than those of blank control group; (4 Wnt/β-catenin signaling pathway: Wnt and β-catenin contents of evodiamine group were lower than those of blank control group. Conclusion: Evodiamine can inhibit the proliferation of colorectal cancer cell lines HT29 and down-regulate the expression of VEGF and MMP, and the effect may be achieved by inhibiting the activation of Wnt/β-catenin signaling pathway.

  19. Anticancer effect of dentatin and dentatin-hydroxypropyl-β-cyclodextrin complex on human colon cancer (HT-29 cell line

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    AL-Abboodi AS

    2017-11-01

    Full Text Available Ashwaq Shakir AL-Abboodi,1,2 Abdullah Rasedee,3 Ahmad Bustamam Abdul,1,4 Yun Hin Taufiq-Yap,5 Wafaa Abd Alwahed Alkaby,6 Mostafa Saddam Ghaji,7 Peter M Waziri,1,8 Mothanna Sadiq Al-Qubaisi1 1MAKNA-UPM, Cancer Research Laboratory, Institute of Bioscience, University Putra Malaysia, Serdang, Malaysia; 2Basic Science Branch, Faculty of Dentistry, University of Al-Qadisiyah, Al Diwaniyah, Iraq; 3Department of Veterinary Laboratory Diagnosis, Faculty of Veterinary Medicine, University Putra Malaysia, Serdang, Malaysia; 4Department of Biomedical Science, Faculty of Medicine and Health Science, University Putra Malaysia, Serdang, Malaysia; 5Department of Chemistry, Faculty of Science, University Putra Malaysia, Serdang, Malaysia; 6Department of Biomedical, Faculty of Biotechnology, University of AL-Qadisiyah, Al Diwaniyah, Iraq; 7Department of Anatomy and Histology, Faculty of Veterinary Medicine, University of Basrah, Basrah, Iraq; 8Department of Biochemistry, Kaduna State University, Main Campus,  Kaduna, Nigeria Introduction: Dentatin (DEN (5-methoxy-2, 2-dimethyl-10-(1, 1-dimethyl-2propenyl dipyran-2-one, a natural compound present in the roots of Clausena excavata Burm f, possesses pro-apoptotic and antiproliferative effects in various cancer cells. Because of its hydrophobicity, it is believed that its complexation with hydroxy-β-cyclodextrin (HPβCD will make it a potent inhibitor of cancer cell growth. In the current work, the molecular mechanisms of apoptosis induced by DEN and DEN-HPβCD complex were demonstrated in human colon HT-29 cancer cells.Materials and methods: After the human colon HT-29 cancer cells were treated with DEN and DEN-HPβCD complex, their effects on the expression of apoptotic-regulated gene markers in mitochondria-mediated apoptotic and death receptor pathways were detected by Western blot analysis and reverse transcription polymerase chain reaction. These markers included caspases-9, 3, and 8, cytochrome c, poly (ADP

  20. Differential expression of sphingolipids in MRP1 overexpressing HT29 cells

    NARCIS (Netherlands)

    Kok, JW; Veldman, Robert; Klappe, K; Koning, H; Filipeanu, Catalin M.; Muller, Michael

    2000-01-01

    We have obtained a novel multidrug resistant cell line, derived from HT29 G(+) human colon carcinoma cells, by selection with gradually increasing concentrations of the anti-mitotic, microtubule-disrupting agent colchicine. This HT29(col) cell line displayed a 25-fold increase in colchicine

  1. In vitro cytotoxic effects of modified zinc oxide quantum dots on breast cancer cell lines (MCF7), colon cancer cell lines (HT29) and various fungi

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    Fakhroueian, Zahra; Dehshiri, Alireza Mozafari; Katouzian, Fatemeh; Esmaeilzadeh, Pegah

    2014-07-01

    An important ideal objective of this study was to perform surface functionalization of fine (1-3 nm) ZnO quantum dot nanoparticles (QD NPs) in order to inhibit decomposition and agglomeration of nanoparticles in aqueous media. Polymers, oily herbal fatty acids, PEG (polyethylene glycol), and organosilanes are the main reagents used in these reactions, because they are completely soluble in water, and can be used as biological probes in nanomedicine. Vegetable fatty acid-capped ZnO (QD NPs) was fabricated by dissolving at a suitable pH after sol-gel method in the presence of nonionic surfactants as efficient templates with a particular HLB (hydrophilic-lipophilic balance) value (9.7 and 8.2). In the present research, we focused on the cellular toxicity of fine zinc oxide QD NPs containing particular blue fluorescence for targeted delivery of MCF7 and HT29 cancer cell lines. The IC50 values were determined as 10.66 and 5.75 µg/ml for MCF7 and HT29, respectively. These findings showed that ZnO QDs have low toxicity in normal cells (MDBK) and can display potential application in cancer chemotherapy in the near future. These properties could result in the generation of a promising candidate in the field of nanobiomedicine. The robust-engineered ZnO QD NPs showed their antibacterial and antifungal activities against Bacillus anthracis, Staphylococcus aureus, Klebsiella pneumonia, and Staphylococcus epidermidis bacteria and also different fungi such as Microsporum gypseum, Microsporum canis, Trichophyton mentagrophytes, Candida albicans, and Candida tropicalis, compared with the standard antibiotic agents like Gentamicin and Clotrimazol.

  2. Biogenic Synthesis, Characterization and Evaluation of Silver Nanoparticles from Aspergillus niger JX556221 Against Human Colon Cancer Cell Line HT-29.

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    Chengzheng, Wang; Jiazhi, Wen; Shuangjiang, Chen; Swamy, Mallappa Kumara; Sinniah, Uma Rani; Akhtar, Mohd Sayeed; Umar, Ahmad

    2018-05-01

    Nanobiotechnology has emerged as a promising technology to develop new therapeutically active nanomaterials. The present study was aimed to biosynthesize AgNPs extracellularly using Aspergillus niger JX556221 fungal extract and to evaluate their anticancer potential against colon cancer cell line, HT-29. UV-visible spectral characterization of the synthesized AgNPs showed higher absorption peak at 440 nm wavelength. Transmission Electron Microscopy (TEM) analysis revealed the monodispersed nature of synthesized AgNPs occurring in spherical shape with a size in the range of 20-25 nm. Further, characterization using Energy Dispersive Spectroscopy (EDX) confirmed the face-centred cubic crystalline structure of metallic AgNPs. FTIR data revealed the occurrence of various phytochemicals in the cell free fungal extract which substantiated the fungal extract mediated AgNPs synthesis. The cytotoxic effect of AgNPs was studied by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The results evidenced the cytotoxic effect of AgNPs on HT-29 cell lines in a dose dependent manner. The highest activity was found at 100 μg/ml concentration after 24 h of incubation. Use of propidium iodide staining examination method confirmed the cytotoxic effect of AgNPs through inducing cell apoptosis. AgNPs cytotoxicity was found to be through elevating reactive oxygen species (ROS), and caspase-3 activation resulting in induced apoptosis. Therefore, this research finding provides an insight towards the development of novel anticancer agents using biological sources.

  3. Development of a serum-free co-culture of human intestinal epithelium cell-lines (Caco-2/HT29-5M21

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    Schneider Yves-Jacques

    2006-05-01

    Full Text Available Abstract Background The absorptive and goblet cells are the main cellular types encountered in the intestine epithelium. The cell lineage Caco-2 is a model commonly used to reproduce the features of the bowel epithelium. However, there is a strong debate regarding the value of Caco-2 cell culture to mimick in vivo situation. Indeed, some authors report in Caco-2 a low paracellular permeability and an ease of access of highly diffusible small molecules to the microvilli, due to an almost complete lack of mucus. The HT29-5M21 intestinal cell lineage is a mucin-secreting cellular population. A co-culture system carried out in a serum-free medium and comprising both Caco-2 and HT29-5M21 cells was developed. The systematic use of a co-culture system requires the characterization of the monolayer under a given experimental procedure. Results In this study, we investigated the activity and localization of the alkaline phosphatase and the expression of IAP and MUC5AC genes to determine a correlation between these markers and the cellular composition of a differentiated monolayer obtained from a mixture of Caco-2 and HT29-5M21 cells. We observed that the culture conditions used (serum-free medium did not change the phenotype of each cell type, and produced a reproducible model. The alkaline phosphatase expression characterizing Caco-2 cells was influenced by the presence of HT29-5M21 cells. Conclusion The culture formed by 75% Caco-2 and 25% HT29-5M21 produce a monolayer containing the two main cell types of human intestinal epithelium and characterized by a reduced permeability to macromolecules.

  4. Development of a serum-free co-culture of human intestinal epithelium cell-lines (Caco-2/HT29-5M21)

    Science.gov (United States)

    Nollevaux, Géraldine; Devillé, Christelle; El Moualij, Benaïssa; Zorzi, Willy; Deloyer, Patricia; Schneider, Yves-Jacques; Peulen, Olivier; Dandrifosse, Guy

    2006-01-01

    Background The absorptive and goblet cells are the main cellular types encountered in the intestine epithelium. The cell lineage Caco-2 is a model commonly used to reproduce the features of the bowel epithelium. However, there is a strong debate regarding the value of Caco-2 cell culture to mimick in vivo situation. Indeed, some authors report in Caco-2 a low paracellular permeability and an ease of access of highly diffusible small molecules to the microvilli, due to an almost complete lack of mucus. The HT29-5M21 intestinal cell lineage is a mucin-secreting cellular population. A co-culture system carried out in a serum-free medium and comprising both Caco-2 and HT29-5M21 cells was developed. The systematic use of a co-culture system requires the characterization of the monolayer under a given experimental procedure. Results In this study, we investigated the activity and localization of the alkaline phosphatase and the expression of IAP and MUC5AC genes to determine a correlation between these markers and the cellular composition of a differentiated monolayer obtained from a mixture of Caco-2 and HT29-5M21 cells. We observed that the culture conditions used (serum-free medium) did not change the phenotype of each cell type, and produced a reproducible model. The alkaline phosphatase expression characterizing Caco-2 cells was influenced by the presence of HT29-5M21 cells. Conclusion The culture formed by 75% Caco-2 and 25% HT29-5M21 produce a monolayer containing the two main cell types of human intestinal epithelium and characterized by a reduced permeability to macromolecules. PMID:16670004

  5. Biphasic increase of apical Cl- conductance by muscarinic stimulation of ht-29cl.19a human colon carcinoma cell line: Evidence for activation of different cl- conductances by carbachol and forskolin

    NARCIS (Netherlands)

    R.B. Bajnath (R.); K. Dekker (K.); A.B. Vaandrager (Arie); H.R. de Jonge (Hugo); J.A. Groot (J.)

    1992-01-01

    textabstractThe modulation of ion transport pathways in filtergrown monolayers of the Cl--secreting subclone (19A) of the human colon carcinoma cell line HT-29 by muscarinic stimulation was studied by combined Ussing chamber and microimpalement experiments. Basolateral addition of 10-4m carbachol

  6. Inhibition of Lipopolysaccharide-Induced Interleukin 8 in Human Adenocarcinoma Cell Line HT-29 by Spore Probiotics: B. coagulans and B. subtilis (natto).

    Science.gov (United States)

    Azimirad, Masoumeh; Alebouyeh, Masoud; Naji, Tahereh

    2017-03-01

    Probiotics are used as a treatment for different intestinal disorders. They confer health benefits by different ways. This study was aimed to investigate immunomodulatory effect of Bacillus probiotic spores on the production of lipopolysaccharide (LPS)-induced interleukin 8 (IL-8) in HT-29 intestinal epithelial cells. Differentiated intestinal epithelial cell line was used as a model for the study of colonization of purified spores (Bacillus subtilis (natto) and B. coagulans) and their anti-inflammatory effects. MTT assay and trypan blue staining were used for the detection of optimal concentration of the purified spores and LPS. Pre-treatment assay was done by treatment of the cells with the purified spores for 2 h, followed by challenges with LPS for 3 and 18 h. Post-treatment assay was done by initial treatment of the cells with LPS for 18 h, followed by the spores for 3 and 6 h. Levels of IL-8 secretion and its mRNA expression were measured by ELISA and relative Q real-time PCR. Our results showed similar rates of adherence to intestinal epithelial cells by the spore probiotics, while displaying no cytotoxic effect. In the pre-treatment assay, a significant decrease in IL-8, at both protein and mRNA levels, was measured for B. coagulans spores after the addition of LPS, which was higher than those observed for Bacillus subtilis (natto) spores. In the post-treatment assay, while Bacillus subtilis (but not B. coagulans) diminished the LPS-stimulated IL-8 levels after 3 h of incubation, the inhibitory effect was not constant. In conclusion, ability of Bacillus spore probiotics for adherence to intestinal epithelial cell and their anti-inflammatory effects, through interference with LPS/IL-8 signaling, was shown in this study. Further studies are needed to characterize responsible bacterial compounds associated with these effects.

  7. Purine-Metabolizing Ectoenzymes Control IL-8 Production in Human Colon HT-29 Cells

    Directory of Open Access Journals (Sweden)

    Fariborz Bahrami

    2014-01-01

    Full Text Available Interleukin-8 (IL-8 plays key roles in both chronic inflammatory diseases and tumor modulation. We previously observed that IL-8 secretion and function can be modulated by nucleotide (P2 receptors. Here we investigated whether IL-8 release by intestinal epithelial HT-29 cells, a cancer cell line, is modulated by extracellular nucleotide metabolism. We first identified that HT-29 cells regulated adenosine and adenine nucleotide concentration at their surface by the expression of the ectoenzymes NTPDase2, ecto-5′-nucleotidase, and adenylate kinase. The expression of the ectoenzymes was evaluated by RT-PCR, qPCR, and immunoblotting, and their activity was analyzed by RP-HPLC of the products and by detection of Pi produced from the hydrolysis of ATP, ADP, and AMP. In response to poly (I:C, with or without ATP and/or ADP, HT-29 cells released IL-8 and this secretion was modulated by the presence of NTPDase2 and adenylate kinase. Taken together, these results demonstrate the presence of 3 ectoenzymes at the surface of HT-29 cells that control nucleotide levels and adenosine production (NTPDase2, ecto-5′-nucleotidase and adenylate kinase and that P2 receptor-mediated signaling controls IL-8 release in HT-29 cells which is modulated by the presence of NTPDase2 and adenylate kinase.

  8. Evaluation of a possible direct effect by casein phosphopeptides on paracellular and vitamin D controlled transcellular calcium transport mechanisms in intestinal human HT-29 and Caco2 cell lines.

    Science.gov (United States)

    Colombini, Alessandra; Perego, Silvia; Ardoino, Ilaria; Marasco, Emiliano; Lombardi, Giovanni; Fiorilli, Amelia; Biganzoli, Elia; Tettamanti, Guido; Ferraretto, Anita

    2013-08-01

    Intestinal cells are continuously exposed to food whose components are able to modulate some of their physiological functions. Among the bioactive food derivatives are casein phosphopeptides (CPPs), coming from the in vitro or in vivo casein digestion, which display the ability to form aggregates with calcium ions and to increase the uptake of the minerals in differentiated intestinal human HT-29 and Caco2 cells. Since extracellular calcium is a known inactivator of the TRPV6 channel, which is also involved in the colon cancer progression, the present study aims to determine a possible modulation by CPPs of the molecular structures responsible for paracellular and/or transcellular calcium absorption in these two cell lines. The paracellular calcium transport was determined by TEER measurements in Caco2 cells and by Lucifer Yellow flow in HT-29 cells. The possible modulation of transcellular calcium absorption machinery by CPPs was investigated by determining the mRNA expression for both the TRPV6 calcium channel and the VDR receptor in 1,25(OH)₂D₃ pre-treated undifferentiated/differentiated cells. The results obtained point out that: (i) CPPs do not affect paracellular calcium absorption; (ii) 1,25(OH)₂D₃ increases the TRPV6 mRNA expression in both types of cells. In the case of HT-29 cells this is the first determination of the presence of the TRPV6 channel; (iii) CPPs per se are not able to affect the VDR and TRPV6 mRNA expression; (iv) CPP administration does not affect the TRPV6 mRNA expression in 1,25(OH)₂D₃ pre-treated HT-29 cells and Caco2 cells. Unlike peptides coming from the digestion of cheese whey protein digest, the digestion of milk casein produces peptides with no effects on TRPV6 calcium channel expression, though the same peptides are able to determine a calcium uptake by the intestinal cells.

  9. In vitro anti-proliferative activity on colon cancer cell line (HT-29) of Thai medicinal plants selected from Thai/Lanna medicinal plant recipe database "MANOSROI III".

    Science.gov (United States)

    Manosroi, Aranya; Akazawa, Hiroyuki; Akihisa, Toshihiro; Jantrawut, Pensak; Kitdamrongtham, Worapong; Manosroi, Worapaka; Manosroi, Jiradej

    2015-02-23

    Thai/Lanna region has its own folklore wisdoms including the traditional medicinal plant recipes. Thai/Lanna medicinal plant recipe database "MANOSROI III" has been developed by Prof. Dr. Jiradej Manosroi. It consists of over 200,000 recipes for all diseases including cancer. To investigate the anti-proliferative and apoptotic activities on human colon cancer cell line (HT-29) as well as the cancer cell selectivity of the methanolic extracts (MEs) and fractions of the 23 selected plants from the "MANOSROI III" database. The 23 selected plants were extracted with methanol under reflux and evaluated for their anti-proliferative activity by sulforhodamine B assay. The 5 plants (Gloriosa superba, Caesalpinia sappan, Fibraurea tinctoria, Ventilago denticulata and Psophocarpus tetragonolobus) with potent anti-proliferative activity were fractionated by liquid-liquid partition to give 4 fractions including each hexane (HF), methanol-water (MF), n-butanol (BF) and water (WF) fractions. They were tested for anti-proliferative activity and cancer cell selectivity. The ME and fractions of G. superba which showed potent anti-proliferative activity were further examined for morphological changes and apoptotic activities by acridine orange (AO)/ethidium bromide (EB) staining. The ME of G. superba root showed active with the highest anti-proliferative activity at 9.17 and 1.58 folds of cisplatin and doxorubicin, respectively. After liquid-liquid partition, HF of V. denticulata, MFs of F. tinctoria, V. denticulata and BF of P. tetragonolobus showed higher anti-proliferative activities than their MEs. The MF of G. superba indicated the highest anti-proliferative activity at 7.73 and 1.34 folds of cisplatin and doxorubicin, respectively, but only 0.86 fold of its ME. The ME and HF, MF and BF of G. superba and MF of F. tinctoria demonstrated high cancer cell selectivity. At 50 µg/ml, ME, HF, MF and BF of G. superba demonstrated higher apoptotic activities than the two standard drugs

  10. Milk fat globule membrane isolate induces apoptosis in HT-29 human colon cancer cells.

    Science.gov (United States)

    Zanabria, Romina; Tellez, Angela M; Griffiths, Mansel; Corredig, Milena

    2013-02-01

    A native milk fat globule membrane (MFGM) isolate obtained from raw milk was assessed for its anticarcinogenic capacity using a colon cancer cell line (HT-29). To prevent microbial contamination and eliminate the presence of lipopolysaccharide (LPS) in the milk used for MFGM isolation, the milk was obtained from the mammary glands of cows using a catheter. Cell proliferation assays demonstrated a reduction of exponentially growing cancer cells of up to 53%, expressed as DNA synthesis (BrdU test), after 72 h stimulation with 100 μg of MFGM protein per mL. Using a similar MFGM concentration, the sulforhodamine B assay resulted in 57% reduction of cell density after 48 h incubation. This bioactivity was comparable to that of known anticancer drugs, 0.1 mM melphalan and 20 μM C2-ceramide, which achieved a cell division reduction of 25 and 40%, respectively, under the same experimental conditions. The toxic effect of the MFGM extracts on HT-29 cells was confirmed by the significant reduction in lactate dehydrogenase enzyme (LDH) by the residual viable cells. An increase of caspase-3 activity (up to 26%) led to the conclusion that MFGM has an apoptotic effect on HT-29 cancer cells.

  11. Alisertib Induces Cell Cycle Arrest, Apoptosis, Autophagy and Suppresses EMT in HT29 and Caco-2 Cells

    Science.gov (United States)

    Ren, Bao-Jun; Zhou, Zhi-Wei; Zhu, Da-Jian; Ju, Yong-Le; Wu, Jin-Hao; Ouyang, Man-Zhao; Chen, Xiao-Wu; Zhou, Shu-Feng

    2015-01-01

    Colorectal cancer (CRC) is one of the most common malignancies worldwide with substantial mortality and morbidity. Alisertib (ALS) is a selective Aurora kinase A (AURKA) inhibitor with unclear effect and molecular interactome on CRC. This study aimed to evaluate the molecular interactome and anticancer effect of ALS and explore the underlying mechanisms in HT29 and Caco-2 cells. ALS markedly arrested cells in G2/M phase in both cell lines, accompanied by remarkable alterations in the expression level of key cell cycle regulators. ALS induced apoptosis in HT29 and Caco-2 cells through mitochondrial and death receptor pathways. ALS also induced autophagy in HT29 and Caco-2 cells, with the suppression of phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR), but activation of 5′ AMP-activated protein kinase (AMPK) signaling pathways. There was a differential modulating effect of ALS on p38 MAPK signaling pathway in both cell lines. Moreover, induction or inhibition of autophagy modulated basal and ALS-induced apoptosis in both cell lines. ALS potently suppressed epithelial to mesenchymal transition (EMT) in HT29 and Caco-2 cells. Collectively, it suggests that induction of cell cycle arrest, promotion of apoptosis and autophagy, and suppression of EMT involving mitochondrial, death receptor, PI3K/Akt/mTOR, p38 MAPK, and AMPK signaling pathways contribute to the cancer cell killing effect of ALS on CRC cells. PMID:26729093

  12. Alisertib Induces Cell Cycle Arrest, Apoptosis, Autophagy and Suppresses EMT in HT29 and Caco-2 Cells

    Directory of Open Access Journals (Sweden)

    Bao-Jun Ren

    2015-12-01

    Full Text Available Colorectal cancer (CRC is one of the most common malignancies worldwide with substantial mortality and morbidity. Alisertib (ALS is a selective Aurora kinase A (AURKA inhibitor with unclear effect and molecular interactome on CRC. This study aimed to evaluate the molecular interactome and anticancer effect of ALS and explore the underlying mechanisms in HT29 and Caco-2 cells. ALS markedly arrested cells in G2/M phase in both cell lines, accompanied by remarkable alterations in the expression level of key cell cycle regulators. ALS induced apoptosis in HT29 and Caco-2 cells through mitochondrial and death receptor pathways. ALS also induced autophagy in HT29 and Caco-2 cells, with the suppression of phosphoinositide 3-kinase (PI3K/protein kinase B (Akt/mammalian target of rapamycin (mTOR, but activation of 5′ AMP-activated protein kinase (AMPK signaling pathways. There was a differential modulating effect of ALS on p38 MAPK signaling pathway in both cell lines. Moreover, induction or inhibition of autophagy modulated basal and ALS-induced apoptosis in both cell lines. ALS potently suppressed epithelial to mesenchymal transition (EMT in HT29 and Caco-2 cells. Collectively, it suggests that induction of cell cycle arrest, promotion of apoptosis and autophagy, and suppression of EMT involving mitochondrial, death receptor, PI3K/Akt/mTOR, p38 MAPK, and AMPK signaling pathways contribute to the cancer cell killing effect of ALS on CRC cells.

  13. The effect of radio-adaptive doses on HT29 and GM637 cells

    Directory of Open Access Journals (Sweden)

    Hell Roswitha

    2008-04-01

    Full Text Available Abstract Background The shape of the dose-response curve at low doses differs from the linear quadratic model. The effect of a radio-adaptive response is the centre of many studies and well known inspite that the clinical applications are still rarely considered. Methods We studied the effect of a low-dose pre-irradiation (0.03 Gy – 0.1 Gy alone or followed by a 2.0 Gy challenging dose 4 h later on the survival of the HT29 cell line (human colorectal cancer cells and on the GM637 cell line (human fibroblasts. Results 0.03 Gy given alone did not have a significant effect on both cell lines, the other low doses alone significantly reduced the cell survival. Applied 4 h before the 2.0 Gy fraction, 0.03 Gy led to a significant induced radioresistance in GM637 cells, but not in HT29 cells, and 0.05 Gy led to a significant hyperradiosensitivity in HT29 cells, but not in GM637 cells. Conclusion A pre-irradiation with 0.03 Gy can protect normal fibroblasts, but not colorectal cancer cells, from damage induced by an irradiation of 2.0 Gy and the application of 0.05 Gy prior to the 2.0 Gy fraction can enhance the cell killing of colorectal cancer cells while not additionally damaging normal fibroblasts. If these findings prove to be true in vivo as well this may optimize the balance between local tumour control and injury to normal tissue in modern radiotherapy.

  14. Effects of Lidocaine on HT-29 and SW480 Colon Cancer CellsIn Vitro.

    Science.gov (United States)

    Bundscherer, Anika C; Malsy, Manuela; Bitzinger, Diane I; Wiese, Christoph H R; Gruber, Michael A; Graf, Bernhard M

    2017-04-01

    Evidence is growing that the risk of cancer dissemination may be enhanced during the perioperative period. Whether particular anesthetic techniques influence oncological outcome is still under discussion. For pain management, lidocaine can be administered perioperatively by intravenous, intraperitoneal or epidural infusion. Here we investigated the effect of lidocaine on colon carcinoma cell lines (HT-29 and SW480) in vitro. ELISA BrdU (Roche) for cell proliferation and FITC Annexin V detection kit (BD Pharming) for apoptosis analysis were applied. Cell-cycle profiles were investigated by flow cytometry. Cell-cycle arrest was induced in both cell lines by 1000 μM lidocaine, while no inhibition of cell proliferation was detected. Apoptosis decreased in SW480 but not in HT-29 cells. Lidocaine induces cell-cycle arrest in both colon carcinoma cell lines in vitro. The effective drug concentration can be obtained by local infiltration. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  15. Necrosis - the dominant form of cell death after phototoxicity impact of hypericin in colon adenocarcinoma cells HT29

    International Nuclear Information System (INIS)

    Mikes, J.; Kleban, J.; Sackova, V.; Solar, P.; Fedorocko, P.

    2006-01-01

    Photodynamic therapy (PDT) is a therapeutical approach for the treatment of malignant as well as non-malignant disorders based on administration of nontoxic/weakly toxic photosensitive compound and its activation with light. Hypericin, one of the promising photosensitizers, is known to induce apoptosis with high efficiency in various cell line models. However, here we report the prevalence of necrosis in colon adenocarcinoma HT-29 cells exposed to an extensive range of PDT doses evoked by variations in two variables - hypericin concentration and light dose. Necrosis was the principal mode of cell death despite different PDT doses and the absence of anti-apoptotic Bcl-2 expression. It is likely that the mutation in p53 plays a crucial role in cell death signaling in HT-29. Data indicating proliferation shifting in HT29 cells, incidence of cell death (apoptosis, necrosis and secondary necrosis) and comparison of cytotoxicity and caspase-3 activity of HT29 with HeLa cells are presented. (authors)

  16. Title of paper: the induction of P-53 independent programmed cell death (apoptosis) with ionizing radiation and 5-fluorouracil (5-FU) in the HT-29 human colon carcinoma cell line

    International Nuclear Information System (INIS)

    Blackstock, A. Wm.; Gill, Misha; Hess, Suzanne M.; Fisher, Robert W.; Leadon, Steven A.; Tepper, Joel E.

    1996-01-01

    Purpose/Objective: The role of programmed cell death (apoptosis) as a cellular response to cancer therapy such as radiation or chemotherapy is the subject of much study, and manipulation of the apoptotic response in tumor cells may be valuable in the treatment of a variety of cancers. Both p53 dependent and independent apoptotic pathways have been identified; p53 is mutated in at least 50 % of human cancers and a majority of radiation resistant tumors contain p53 mutations. This study is designed to examine the induction of programmed cell death in a human colon carcinoma cell line that possesses two mutated p53 alleles. Ionizing radiation alone, or in combination with the chemotherapeutic drug 5-fluorouracil (5-FU), were used to elicit the apoptotic response. This study will focus on whether these treatments can induce a significant apoptotic response in cells that have mutated p53 alleles. Materials and Methods: HT-29 cells were assessed for clonogenic survival after being plated at a variety of densities, and treated with single graded doses of radiation (0, 1, 2, 4, 6, 8, 10 Gy) either alone or immediately prior to a 24 hour exposure to 5-FU (2 ug/ml). The extent of radiation and 5-FU-induced apoptosis was determined in the HT-29 cell line after single doses of 0, 2, 5, and 10 Gy either alone or immediately prior to a 24 hour incubation in 5-FU (2 ug/ml). Three separate assays were used to evaluate the apoptotic response. Cells undergoing apoptosis undergo gross morphological changes including a condensation of chromatin, membrane blebbing, and an eventual release of membrane bound cytoplasmic fragments. Hematoxylin and eosin staining were used to visualize some of these morphological changes. Another characteristic of the apoptotic response is the activation of an endonuclease that cleaves DNA into specific fragments. Accordingly, an ELISA cell death assay (Boehringer Mannheim, Indianapolis IN) was used to quantitate cytoplasmic histone-associated DNA

  17. Modulation of carcinoembryonic antigen release by glucosylceramide : Implications for HT29 cell differentiation

    NARCIS (Netherlands)

    Babia, T; Hoekstra, D; Kok, JW; Veldman, Robert

    1998-01-01

    Previous work suggested that glucosylceramide (GlcCer) plays a role in the regulation of cell differentiation of HT29 human colon tumor cells [I]. In the present study, we investigated the role of GlcCer in the cellular release of carcinoembryonic antigen (CEA), a marker for cell differentiation.

  18. Mechanisms underlying apoptosis-inducing effects of Kaempferol in HT-29 human colon cancer cells.

    Science.gov (United States)

    Lee, Hyun Sook; Cho, Han Jin; Yu, Rina; Lee, Ki Won; Chun, Hyang Sook; Park, Jung Han Yoon

    2014-02-17

    We previously noted that kaempferol, a flavonol present in vegetables and fruits, reduced cell cycle progression of HT-29 cells. To examine whether kaempferol induces apoptosis of HT-29 cells and to explore the underlying molecular mechanisms, cells were treated with various concentrations (0-60 μmol/L) of kaempferol and analyzed by Hoechst staining, Annexin V staining, JC-1 labeling of the mitochondria, immunoprecipitation, in vitro kinase assays, Western blot analyses, and caspase-8 assays. Kaempferol increased chromatin condensation, DNA fragmentation and the number of early apoptotic cells in HT-29 cells in a dose-dependent manner. In addition, kaempferol increased the levels of cleaved caspase-9, caspase-3 and caspase-7 as well as those of cleaved poly (ADP-ribose) polymerase. Moreover, it increased mitochondrial membrane permeability and cytosolic cytochrome c concentrations. Further, kaempferol decreased the levels of Bcl-xL proteins, but increased those of Bik. It also induced a reduction in Akt activation and Akt activity and an increase in mitochondrial Bad. Additionally, kaempferol increased the levels of membrane-bound FAS ligand, decreased those of uncleaved caspase-8 and intact Bid and increased caspase-8 activity. These results indicate that kaempferol induces the apoptosis of HT-29 cells via events associated with the activation of cell surface death receptors and the mitochondrial pathway.

  19. Different responses of Fe transporters in Caco2/HT29-MTX cocultures than in independent Caco-2 cell cultures

    Science.gov (United States)

    The human intestinal epithelium is composed of several cell types; mainly enterocytes and globet (mucin-secreting) cells. This study compares the cellular response for Fe transporters in Caco-2, HT29-MTX, and Caco-2/HT29-MTX coculture models for Fe bioavailability studies. Under culture, Caco-2 cell...

  20. Alterations in mitochondria function and morphology in HT29 cells upon conditions inducing differentiation and apoptosis

    Czech Academy of Sciences Publication Activity Database

    Sovová, Vlasta; Štokrová, Jitka; Šloncová, Eva; Kučerová, Dana; Tuháčková, Zdena; Korb, Jan

    2003-01-01

    Roč. 23, č. 6 (2003), s. 1755-1760 ISSN 1019-6439 Institutional research plan: CEZ:AV0Z5052915 Keywords : differentiation-apoptosis-HT29cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.536, year: 2003

  1. Inhibition of bacterial adhesion to HT-29 cells by lipoteichoic acid ...

    African Journals Online (AJOL)

    The aim of this experiment was to study the effect of the lipoteichoic acid (LTA) extracted from Clostridium butyricum on the adhesion of C. butyricum and Escherichia coli to HT-29 human intestinal cells. The method of extraction of lipoteichoic acid form C. butyricum by TX114 was evaluated. The purification of the LTA by ...

  2. Identification of an anticancer compound against HT-29 cells from Phellinus linteus grown on germinated brown rice.

    Science.gov (United States)

    Jeon, Tae-Il; Jung, Chang-Hwa; Cho, Jeong-Yong; Park, Dong Ki; Moon, Jae-Hak

    2013-10-01

    To isolate and identify the anticancer compound against proliferation of human colon cancer cells from ethyl acetate (EtOAc) extract of Phellinus linteus grown on germinated brown rice (PB). EtOAc extract of PB was partitioned with n-hexane, EtOAc, and water-saturated n-butanol. Anticancer compound of n-hexane layer was isolated and identified by HPLC and NMR, respectively. Cytotoxicity against HT-29 cells was tested by SRB assay. The n-hexane layer obtained after solvent fractionation of PB EtOAc extracts showed a potent anticancer activity against the HT-29 cell line. Atractylenolide I, a eudesmane-type sesquiterpene lactone, a major anticancer substance of PB, was isolated from the n-hexane layer by silica gel column chromatography and preparative-HPLC. This structure was elucidated by one- and two-dimensional NMR spectroscopic data. Atractylenolide I has not been reported in mushrooms or rice as of yet. The isolated compound dose-dependently inhibited the growth of HT-29 human colon cancer cells. Atractylenolide I might contribute to the anticancer effect of PB. Copyright © 2013 Asian Pacific Tropical Biomedical Magazine. Published by Elsevier B.V. All rights reserved.

  3. The effect of five artificial sweeteners on Caco-2, HT-29 and HEK-293 cells.

    Science.gov (United States)

    van Eyk, Armorel Diane

    2015-01-01

    Artificial sweeteners (AS) have been associated with tumor development (including colon cancer) in both animals and humans although evidence has been conflicting. Additional research was thus conducted by studying the effects of 5 AS on the morphology, cell proliferation and DNA in cells by utilizing Caco-2, HT-29 (colon) and HEK-293 (kidney) cell lines. Cells were exposed to sodium cyclamate, sodium saccharin, sucralose and acesulfame-K (0-50 mM) and aspartame (0-35 mM) over 24, 48 and 72 hours. Morphological changes were presented photographically and % cell viability was determined by using the MTT cell viability assay. Possible DNA damage (comet assay) induced by the AS (0.1, 1 and 10 mM, treated for 24, 48 and 72 hours) was studied. The appearance of "comets" was scored from no damage to severe damage (0-4). Cells became flatter and less well defined at higher AS concentrations (>10 mM). At concentrations >10 mM, decreased cell viability was noted with both increasing concentration and increasing incubation time for all cell lines tested. In general, HEK-293 cells seemed to be less affected then the colon cancer cells. Sucralose and sodium saccharin seemed to elicit the greatest degree of DNA fragmentation of all the sweeteners tested in all the cell lines used. Morphological cell alterations, cell viability and DNA fragmentation seemed to be more in the colon cancer cells. Further studies have to be performed to clarify mechanisms involved causing these alterations in mammalian cells.

  4. NHERF-1 regulation of EGF and neurotensin signalling in HT-29 epithelial cells

    International Nuclear Information System (INIS)

    Kruger, Wade A.; Monteith, Gregory R.; Poronnik, Philip

    2013-01-01

    Highlights: ► NHERF-1 expression was abundant throughout HT-29 cells consistent with a cancerous phenotype. ► Knockdown of NHERF-1 lead to a significant reduction in cell proliferation. ► EGF and neurotensin-mediated proliferation was inhibited by knockdown of NHERF-1. ► Neurotensin-mediated Ca 2+ response was abolished by knockdown of NHERF-1. -- Abstract: Neurotensin receptors (NT-R) and the epidermal growth factor receptors (EGF-R) are commonly overexpressed in many epithelial origin tumours. In addition to their role as mitogenic mediators through specific cell signalling, recent studies indicate that the activity/expression of scaffold proteins responsible for the assembly and coordination of the signalling complexes may also have central roles in epithelial transformation. In particular, the “epithelial” PSD-95/Dlg/Zo-1 (PDZ) scaffold/adapter protein, Na + /H + exchanger regulatory factor isoform one (NHERF-1), has been identified as a potential regulator of cellular transformation. NHERF-1 is a known regulator of EGF-R function and plays numerous roles in G-protein-coupled receptor signalling. Because of the synergistic signalling between these two potent mitogens, we investigated a potential role for NHERF-1 in the molecular mechanism linking the aberrant proliferative phenotype initiated by some G-Protein-coupled receptor activators in the colon adenocarcinoma HT-29 cell line. Knockdown (80%) of endogenous NHERF-1 leads to significant reduction in proliferation rate; an effect that could not be recovered by exogenous application of either NT or EGF. Inhibition of the EGF-R with AG1487 also inhibited proliferation and this effect could not be recovered with NT. Knockdown of NHERF-1 significantly altered the expression of the EGF-R, and almost completely abolished the NT-mediated increases in intracellular free Ca 2+ . Knockdown of NHERF-1 also attenuated UTP-mediated purinergic Ca 2+ signalling. Taken together, these data suggest that NHERF-1

  5. Effect of Uncaria tomentosa Extract on Apoptosis Triggered by Oxaliplatin Exposure on HT29 Cells

    OpenAIRE

    de Oliveira, Liliane Z.; Farias, Iria Luiza G.; Rigo, Melânia L.; Glanzner, Werner G.; Gonçalves, Paulo Bayard D.; Cadoná, Francine C.; Cruz, Ivana B.; Farias, Júlia G.; Duarte, Marta M. M. F.; Franco, Luzia; Bertol, Gustavo; Colpo, Elisangela; Brites, Patricia C.; Rocha, João Batista T.; Leal, Daniela B. R.

    2014-01-01

    Background/Aim. The use of herbal products as a supplement to minimize the effects of chemotherapy for cancer treatment requires further attention with respect to the activity and toxicity of chemotherapy. Uncaria tomentosa extract, which contains oxindole alkaloids, is one of these herbal products. The objective of this study was to evaluate whether Uncaria tomentosa extract modulates apoptosis induced by chemotherapy exposure. Materials and Methods. Colorectal adenocarcinoma cells (HT29 cel...

  6. DNA Tetrahedron Delivery Enhances Doxorubicin-Induced Apoptosis of HT-29 Colon Cancer Cells

    Science.gov (United States)

    Zhang, Guiyu; Zhang, Zhiyong; Yang, Junen

    2017-08-01

    As a nano-sized drug carrier with the advantage of modifiability and proper biocompatibility, DNA tetrahedron (DNA tetra) delivery is hopeful to enhance the inhibitory efficiency of nontargeted anticancer drugs. In this investigation, doxorubicin (Dox) was assembled to a folic acid-modified DNA tetra via click chemistry to prepare a targeted antitumor agent. Cellular uptake efficiency was measured via fluorescent imaging. Cytotoxicity, inhibition efficiency, and corresponding mechanism on colon cancer cell line HT-29 were evaluated by MTT assay, cell proliferation curve, western blot, and flow cytometry. No cytotoxicity was induced by DNA tetra, but the cellular uptake ratio increased obviously resulting from the DNA tetra-facilitated penetration through cellular membrane. Accordingly, folic acid-DNA tetra-Dox markedly increased the antitumor efficiency with increased apoptosis levels. In details, 100 μM was the effective concentration and a 6-h incubation period was needed for apoptosis induction. In conclusion, nano-sized DNA tetrahedron was a safe and effective delivery system for Dox and correspondingly enhanced the anticancer efficiency.

  7. Glycosylation-related gene expression in HT29-MTX-E12 cells upon infection by Helicobacter pylori.

    Science.gov (United States)

    Cairns, Michael T; Gupta, Ananya; Naughton, Julie A; Kane, Marian; Clyne, Marguerite; Joshi, Lokesh

    2017-10-07

    To identify glycosylation-related genes in the HT29 derivative cell line, HT29-MTX-E12, showing differential expression on infection with Helicobacter pylori ( H. pylori ). Polarised HT29-MTX-E12 cells were infected for 24 h with H. pylori strain 26695. After infection RNA was isolated from both infected and non-infected host cells. Sufficient infections were carried out to provide triplicate samples for microarray analysis and for qRT-PCR analysis. RNA was isolated and hybridised to Affymetrix arrays. Analysis of microarray data identified genes significantly differentially expressed upon infection. Genes were grouped into gene ontology functional categories. Selected genes associated with host glycan structure (glycosyltransferases, hydrolases, lectins, mucins) were validated by real-time qRT-PCR analysis. Infection of host cells was confirmed by the isolation of live bacteria after 24 h incubation and by PCR amplification of bacteria-specific genes from the host cell RNA. H. pylori do not survive incubation under the adopted culture conditions unless they associate with the adherent mucus layer of the host cell. Microarray analysis identified a total of 276 genes that were significantly differentially expressed ( P < 0.05) upon H. pylori infection and where the fold change in expression was greater than 2. Six of these genes are involved in glycosylation-related processes. Real-time qRT-PCR demonstrated significant downregulation (1.8-fold, P < 0.05) of the mucin MUC20. REG4 was heavily expressed and significantly downregulated (3.1-fold, P < 0.05) upon infection. Gene ontology analysis was consistent with previous studies on H. pylori infection. Gene expression data suggest that infection with H. pylori causes a decrease in glycan synthesis, resulting in shorter and simpler glycan structures.

  8. Carcino-embryonic antigen in monitoring the growth of human colon adenocarcinoma tumour cells SK-CO-1 and HT-29 in vitro and in nude mice

    DEFF Research Database (Denmark)

    Sölétormos, G; Fogh, J M; Sehested-Hansen, B

    1997-01-01

    A set of experimental model systems were designed to investigate (a) the inter-relationship between growth of two human cancer cell lines (SK-CO-1, HT-29) and carcino-embryonic antigen (CEA) kinetics; and (b) whether neoplastic growth or CEA concentration is modulated by human growth hormone (h...

  9. Carcino-embryonic antigen in monitoring the growth of human colon adenocarcinoma tumour cells SK-CO-1 and HT-29 in vitro and in nude mice

    DEFF Research Database (Denmark)

    Sölétormos, G; Fogh, J M; Sehested-Hansen, B

    1997-01-01

    A set of experimental model systems were designed to investigate (a) the inter-relationship between growth of two human cancer cell lines (SK-CO-1, HT-29) and carcino-embryonic antigen (CEA) kinetics; and (b) whether neoplastic growth or CEA concentration is modulated by human growth hormone (hGH...

  10. Induction of apoptosis by tomato using space mutation breeding in human colon cancer SW480 and HT-29 cells.

    Science.gov (United States)

    Shi, Jiahui; Yang, Bin; Feng, Pan; Li, Duo; Zhu, Jiajin

    2010-03-15

    As far as we know, there have been no reports concerning the functional characteristics of tomatoes using space mutation breeding. The aim of this study was to evaluate the anti-colon cancer effect of tomatoes M1 and M2 using space mutation breeding. In the present study, obvious anti-cancer activity was shown with tomato juice of M1 and M2 and their parent CK treatment in colon cancer cell lines SW480 and HT-29 in cell growth inhibition. In addition, SW480 cells were more sensitive to M1 and M2 than HT-29 cells in cell apoptosis. Furthermore, M1 and M2 induced cell cycle arrest both in G0-G1 and G2/M phases. These data suggest that consumption of tomato using space mutation breeding may provide benefits to inhibit growth of colon cancer cells. Therefore, tomato production using space mutation breeding may be a good candidate for development as a dietary supplement in drug therapy for colon cancer.

  11. Cellular Homeostasis and Antioxidant Response in Epithelial HT29 Cells on Titania Nanotube Arrays Surface

    Directory of Open Access Journals (Sweden)

    Rabiatul Basria SMN Mydin

    2017-01-01

    Full Text Available Cell growth and proliferative activities on titania nanotube arrays (TNA have raised alerts on genotoxicity risk. Present toxicogenomic approach focused on epithelial HT29 cells with TNA surface. Fledgling cell-TNA interaction has triggered G0/G1 cell cycle arrests and initiates DNA damage surveillance checkpoint, which possibly indicated the cellular stress stimuli. A profound gene regulation was observed to be involved in cellular growth and survival signals such as p53 and AKT expressions. Interestingly, the activation of redox regulator pathways (antioxidant defense was observed through the cascade interactions of GADD45, MYC, CHECK1, and ATR genes. These mechanisms furnish to protect DNA during cellular division from an oxidative challenge, set in motion with XRRC5 and RAD50 genes for DNA damage and repair activities. The cell fate decision on TNA-nanoenvironment has been reported to possibly regulate proliferative activities via expression of p27 and BCL2 tumor suppressor proteins, cogent with SKP2 and BCL2 oncogenic proteins suppression. Findings suggested that epithelial HT29 cells on the surface of TNA may have a positive regulation via cell-homeostasis mechanisms: a careful circadian orchestration between cell proliferation, survival, and death. This nanomolecular knowledge could be beneficial for advanced medical applications such as in nanomedicine and nanotherapeutics.

  12. Antiproliferative effect of chitosan-added kimchi in HT-29 human colon carcinoma cells.

    Science.gov (United States)

    Kong, Chang-Suk; Bahn, Young-Eun; Kim, Boh-Kyung; Lee, Kang-Yoon; Park, Kun-Young

    2010-02-01

    The anticancer effects of chitosan-added kimchi were investigated by using an in vitro cellular system with HT-29 human colon carcinoma cells. Two different kinds of chitosan-soluble chitosan with a 90% degree of deacetylation and 3 cps viscosity and nonsoluble chitosan with a 95% degree of deacetylation and 22 cps viscosity-were used as sub-ingredients to increase anticancer effects of kimchi. The soluble chitosan-added kimchi (SK) and nonsoluble chitosan-added kimchi (NK) were stronger growth inhibitors in HT-29 cells than the control kimchi (CK) according to the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and the growth inhibition test. Treatment with SK and NK induced apoptosis, as determined by 4,6-diamidino-2-phenylindole staining, and resulted in the up-regulation of Bax expression and down-regulation of Bcl-2, cIAP-1, cellular inhibitor of apoptosis-2, cyclooxygenase-2, inhibitory nitric oxide synthase, and nuclear factor kappaB (NF-kappaB) expressions when compared to CK. The antiproliferative and anti-apoptotic effects appeared to be more pronounced in the cells treated with NK. The antiproliferative effects of the chitosan-added kimchi appeared to be associated with the induction of apoptosis through NF-kappaB or an NF-kappaB-dependent pathway. These results suggest that chitosan has potential to be a valuable active ingredient in functional kimchi products with anticancer effects.

  13. Adrenaline promotes cell proliferation and increases chemoresistance in colon cancer HT29 cells through induction of miR-155

    Energy Technology Data Exchange (ETDEWEB)

    Pu, Jun [Department of General Surgery, Tangdu Hospital of the Fourth Military Medical University, Xi' an 710038 (China); Bai, Danna [Department of Cardiology, 323 Hospital of PLA, Xi' an 710054 (China); Yang, Xia [Department of Teaching and Medical Administration, Tangdu Hospital of the Fourth Military Medical University, Xi' an 710038 (China); Lu, Xiaozhao [Department of Nephrology, The 323 Hospital of PLA, Xi' an 710054 (China); Xu, Lijuan, E-mail: 13609296272@163.com [Department of Nephrology, The 323 Hospital of PLA, Xi' an 710054 (China); Lu, Jianguo, E-mail: lujianguo029@yahoo.com.cn [Department of General Surgery, Tangdu Hospital of the Fourth Military Medical University, Xi' an 710038 (China)

    2012-11-16

    Highlights: Black-Right-Pointing-Pointer Adrenaline increases colon cancer cell proliferation and its resistance to cisplatin. Black-Right-Pointing-Pointer Adrenaline activates NF{kappa}B in a dose dependent manner. Black-Right-Pointing-Pointer NF{kappa}B-miR-155 pathway contributes to cell proliferation and resistance to cisplatin. -- Abstract: Recently, catecholamines have been described as being involved in the regulation of cancer genesis and progression. Here, we reported that adrenaline increased the cell proliferation and decreased the cisplatin induced apoptosis in HT29 cells. Further study found that adrenaline increased miR-155 expression in an NF{kappa}B dependent manner. HT29 cells overexpressing miR-155 had a higher cell growth rate and more resistance to cisplatin induced apoptosis. In contrast, HT29 cells overexpressing miR-155 inhibitor displayed decreased cell proliferation and sensitivity to cisplatin induced cell death. In summary, our study here revealed that adrenaline-NF{kappa}B-miR-155 pathway at least partially contributes to the psychological stress induced proliferation and chemoresistance in HT29 cells, shedding light on increasing the therapeutic strategies of cancer chemotherapy.

  14. Adrenaline promotes cell proliferation and increases chemoresistance in colon cancer HT29 cells through induction of miR-155

    International Nuclear Information System (INIS)

    Pu, Jun; Bai, Danna; Yang, Xia; Lu, Xiaozhao; Xu, Lijuan; Lu, Jianguo

    2012-01-01

    Highlights: ► Adrenaline increases colon cancer cell proliferation and its resistance to cisplatin. ► Adrenaline activates NFκB in a dose dependent manner. ► NFκB–miR-155 pathway contributes to cell proliferation and resistance to cisplatin. -- Abstract: Recently, catecholamines have been described as being involved in the regulation of cancer genesis and progression. Here, we reported that adrenaline increased the cell proliferation and decreased the cisplatin induced apoptosis in HT29 cells. Further study found that adrenaline increased miR-155 expression in an NFκB dependent manner. HT29 cells overexpressing miR-155 had a higher cell growth rate and more resistance to cisplatin induced apoptosis. In contrast, HT29 cells overexpressing miR-155 inhibitor displayed decreased cell proliferation and sensitivity to cisplatin induced cell death. In summary, our study here revealed that adrenaline–NFκB–miR-155 pathway at least partially contributes to the psychological stress induced proliferation and chemoresistance in HT29 cells, shedding light on increasing the therapeutic strategies of cancer chemotherapy.

  15. Profilin 2 promotes migration, invasion, and stemness of HT29 human colorectal cancer stem cells.

    Science.gov (United States)

    Kim, Min-Jung; Lee, Yoo-Sun; Han, Gi-Yeon; Lee, Han-Na; Ahn, Chiyoung; Kim, Chan-Wha

    2015-01-01

    We investigated the role of profilin 2 in the stemness, migration, and invasion of HT29 cancer stem cells (CSCs). Increased and decreased levels of profilin 2 significantly enhanced and suppressed the self-renewal, migration, and invasion ability of HT29 CSCs, respectively. Moreover, profilin 2 directly regulated the expression of stemness markers (CD133, SOX2, and β-catenin) and epithelial mesenchymal transition (EMT) markers (E-cadherin and snail). CD133 and β-catenin were up-regulated by overexpression of profilin 2 and down-regulated by depletion of profilin 2. SOX2 was decreased by profilin 2 depletion. E-cadherin was not influenced by profilin 2- overexpression but increased by profilin 2- knockdown. The expression of snail was suppressed by profilin 2- knockdown. We speculated that stemness and the EMT are closely linked through profilin 2-related pathways. Therefore, this study indicates that profilin 2 affects the metastatic potential and stemness of colorectal CSCs by regulating EMT- and stemness-related proteins.

  16. Effect of Uncaria tomentosa Extract on Apoptosis Triggered by Oxaliplatin Exposure on HT29 Cells.

    Science.gov (United States)

    de Oliveira, Liliane Z; Farias, Iria Luiza G; Rigo, Melânia L; Glanzner, Werner G; Gonçalves, Paulo Bayard D; Cadoná, Francine C; Cruz, Ivana B; Farias, Júlia G; Duarte, Marta M M F; Franco, Luzia; Bertol, Gustavo; Colpo, Elisangela; Brites, Patricia C; Rocha, João Batista T; Leal, Daniela B R

    2014-01-01

    Background/Aim. The use of herbal products as a supplement to minimize the effects of chemotherapy for cancer treatment requires further attention with respect to the activity and toxicity of chemotherapy. Uncaria tomentosa extract, which contains oxindole alkaloids, is one of these herbal products. The objective of this study was to evaluate whether Uncaria tomentosa extract modulates apoptosis induced by chemotherapy exposure. Materials and Methods. Colorectal adenocarcinoma cells (HT29 cells) were grown in the presence of oxaliplatin and/or Uncaria tomentosa extract. Results. The hydroalcoholic extract of Uncaria tomentosa enhanced chemotherapy-induced apoptosis, with an increase in the percentage of Annexin positive cells, an increase in caspase activities, and an increase of DNA fragments in culture of the neoplastic cells. Moreover, antioxidant activity may be related to apoptosis. Conclusion. Uncaria tomentosa extract has a role for cancer patients as a complementary therapy. Further studies evaluating these beneficial effects with other chemotherapy drugs are recommended.

  17. Effect of Uncaria tomentosa Extract on Apoptosis Triggered by Oxaliplatin Exposure on HT29 Cells

    Directory of Open Access Journals (Sweden)

    Liliane Z. de Oliveira

    2014-01-01

    Full Text Available Background/Aim. The use of herbal products as a supplement to minimize the effects of chemotherapy for cancer treatment requires further attention with respect to the activity and toxicity of chemotherapy. Uncaria tomentosa extract, which contains oxindole alkaloids, is one of these herbal products. The objective of this study was to evaluate whether Uncaria tomentosa extract modulates apoptosis induced by chemotherapy exposure. Materials and Methods. Colorectal adenocarcinoma cells (HT29 cells were grown in the presence of oxaliplatin and/or Uncaria tomentosa extract. Results. The hydroalcoholic extract of Uncaria tomentosa enhanced chemotherapy-induced apoptosis, with an increase in the percentage of Annexin positive cells, an increase in caspase activities, and an increase of DNA fragments in culture of the neoplastic cells. Moreover, antioxidant activity may be related to apoptosis. Conclusion. Uncaria tomentosa extract has a role for cancer patients as a complementary therapy. Further studies evaluating these beneficial effects with other chemotherapy drugs are recommended.

  18. Differentiation of colon adenocarcinoma HT29 cells potentates photodynamic therapy with hypericin

    International Nuclear Information System (INIS)

    Mikes, J.; Kleban, J.; Koval, J.; Fedorocko, P.; Hyzdalova, M.

    2006-01-01

    Photodynamic therapy (PDT) is a therapeutical approach for the treatment of malignant as well as non-malignant disorders based on administration of nontoxic/weakly toxic photosensitive compound and its activation with light. The phototoxicity of PDT depends on generation of superoxide radicals (Type-I reaction), which in turn might form peroxide and hydroxyl radicals, and production of singlet oxygen ( 1 O 2 ) (Type-II reaction) after irradiation with light of appropriate wavelength which properly overlaps the photosensitizer's absorbing spectra. Oxidative damage in the cell induced by reactive oxygen species depends on the intracellular localization and affects different cell organelles. In our study, effectiveness of PDT with hypericin in differentiated versus undifferentiated HT29 cells in the relation to differentiation status has been evaluated. (authors)

  19. Immunomodulatory activity of two potential probiotic strains in LPS-stimulated HT-29 cells.

    Science.gov (United States)

    Duary, Raj Kumar; Batish, Virender Kumar; Grover, Sunita

    2014-05-01

    The relative expression of mucin, pro- and anti-inflammatory genes besides other signaling molecules in HT-29 cells by two test probiotic strains of Lactobacillus plantarum Lp9 and Lp91 and the reference strain L. plantarum 5276 was evaluated by RT-qPCR using Relative Expression Software Tool qBase-Plus under in vitro simulated gut conditions. Ten house keeping genes were evaluated by using geNorm 3.4 excel based application. The most stable genes were RPL27, ACTB and B2M which were subsequently used for calculating the normalization factor. Under pretreatment conditions (4 h probiotic treatment, followed by lipopolysaccharide challenge for 3 h), all the three strains evoked downregulation of IL-8 expression by ~100 %, while in case of TNF-α, the downregulation of the relative gene expression was at the rate of 98.2, 93.8 and 98.0 % with Lp5276, Lp9 and Lp91, respectively, under the same set of conditions. Lp91 evoked maximum downregulation of IL12p35 and IFN-γ with corresponding fold reduction in relative expression of the two genes by 96.5 and 96.7 % during pre-treatment conditions. However, IL-10 and IFN-α were significantly upregulated to the extent of 8.13 ± 0.36 and 2.62 ± 0.14 fold by Lp91 under the same conditions. Lp9 and Lp91 were also quite effective in inducing the expression of Cox-1 and Cox-2 in HT-29 cells as can be reflected from their ratios, i.e., 5.90 and 6.50 (under pretreatment conditions); 3.79 and 4.36 (under co-culture conditions). Thus, the two putative indigenous L. plantarum strains Lp9 and Lp91 demonstrated immunomodulating functions in HT-29 cells at significant levels under different experimental conditions.

  20. Oleic Acid Uptake Reveals the Rescued Enterocyte Phenotype of Colon Cancer Caco-2 by HT29-MTX Cells in Co-Culture Mode

    Science.gov (United States)

    Berger, Emmanuelle; Nassra, Merian; Atgié, Claude; Plaisancié, Pascale; Géloën, Alain

    2017-01-01

    Gastrointestinal epithelium is the unique route for nutrients and for many pharmaceuticals to enter the body. The present study aimed to analyze precisely whether co-culture of two colon cancer cell lines, mucus-producing cells HT29-MTX and enterocyte-like Caco-2 cells, ameliorate differentiation into an in vitro intestinal barrier model and the signaling pathways involved. Differentiated Caco-2 cells gene datasets were compared first to intestinal or cancer phenotypes and second to signaling pathway gene datasets. Experimental validations were performed in real-time experiments, immunochemistry, and gene expression analyses on Caco-2 versus co-cultures of Caco-2 and HT29-MTX (10%) cells. Partial maintenance of cancer-cell phenotype in differentiated Caco-2 cells was confirmed and fatty acids merged as potential regulators of cancer signaling pathways. HT29-MTX cells induced morphological changes in Caco-2 cells, slightly increased their proliferation rate and profoundly modified gene transcription of phenotype markers, fatty acid receptors, intracellular transporters, and lipid droplet components as well as functional responses to oleic acid. In vitro, enterocyte phenotype was rescued partially by co-culture of cancer cells with goblet cells and completed through oleic acid interaction with signaling pathways dysregulated in cancer cells. PMID:28726765

  1. Data for comparative proteomics analysis of the antitumor effect of CIGB-552 peptide in HT-29 colon adenocarcinoma cells

    Directory of Open Access Journals (Sweden)

    Teresa Núñez de Villavicencio-Díaz

    2015-09-01

    Full Text Available CIGB-552 is a second generation antitumor peptide that displays potent cytotoxicity in lung and colon cancer cells. The nuclear subproteome of HT-29 colon adenocarcinoma cells treated with CIGB-552 peptide was identified and analyzed [1]. This data article provides supporting evidence for the above analysis.

  2. Wild-type p53 gene transfer into mutated p53 HT29 cells improves sensitivity to photodynamic therapy via induction of apoptosis.

    Science.gov (United States)

    Barberi-Heyob, Muriel; Védrine, Pierre-Olivier; Merlin, Jean-Louis; Millon, Régine; Abecassis, Joseph; Poupon, Marie-France; Guillemin, François

    2004-04-01

    Photodynamic therapy (PDT) is an effective local cancer treatment that induces cytotoxicity through the intracellular generation of reactive oxygen species. It is generally thought that p53 regulates chemotherapy and radiation therapy responsiveness via apoptosis induction control. The current study investigated whether cellular sensitivity to PDT is increased when a wild-type (wt) p53 status is restored by gene transfer in the established HT9blk Ala273-mutant p53 human colon cancer cell line. The photosensitizer accumulation was similar in both cell lines, and survival measurements using MTT test and clonogenic assays demonstrated that wt p53 transfected cells (HT29A4) were significantly more sensitive to chlorin e6-mediated PDT. P53 protein expression and its functionality as a transcription factor demonstrated through the induction of mdm2 transactivation, were not found to be directly involved in this differential photosensitivity. However, induction of caspase 3 activation (2.6-fold), leading to significant apoptosis induction 24-h after PDT was observed in HT29A4 cells. These results suggest that the introduction of wt p53 in HT29A4 potentiates the cell sensitivity to PDT through the induction of apoptosis in relation to p53 mutational status, but independently of p53 expression level and transcriptional activity.

  3. Carcino-embryonic antigen in monitoring the growth of human colon adenocarcinoma tumour cells SK-CO-1 and HT-29 in vitro and in nude mice

    DEFF Research Database (Denmark)

    Sölétormos, G; Fogh, J M; Sehested-Hansen, B

    1997-01-01

    A set of experimental model systems were designed to investigate (a) the inter-relationship between growth of two human cancer cell lines (SK-CO-1, HT-29) and carcino-embryonic antigen (CEA) kinetics; and (b) whether neoplastic growth or CEA concentration is modulated by human growth hormone (h......GH). We found that increasing CEA concentration depended on tumour burden. SK-CO-1 cells had the lowest growth rates but the highest rates of CEA production. The rate of CEA increase exceeded the growth rate of both SK-CO-1 and HT-29. hGH modulated neither neoplastic growth nor CEA production....... In conclusion, our results suggest that experimental models may be useful for investigating the role of serological markers as monitors of increasing tumour burden. It will be of interest to investigate the performance of those model systems in examining the effect of cytotoxic agents in neoplastic growth....

  4. Defining conditions for the co-culture of Caco-2 and HT29-MTX cells using Taguchi design.

    Science.gov (United States)

    Chen, Xiu-Min; Elisia, Ingrid; Kitts, David D

    2010-01-01

    The co-culture of Caco-2 and HT29 cells for testing intestinal drug and nutrient transport and metabolism provides the presence of both absorptive and goblet cells, both of which have different culture requirements for optimal growth and function. The research on the co-culture of Caco-2 and HT29 cells is very limited in respect to refining specific conditions that reduce intra- and inter-laboratory variations. In the present study we reported conditions that enable reproducible results to be obtained for drug permeability using in vitro co-culture of Caco-2 and HT29-MTX based on Taguchi experimental design. The selection of four factors that specified cell culture conditions, namely culture medium, seeding time, seeding density, and Caco-2:HT29-MTX ratio on TEER value and individual permeability coefficients of propranolol, ketoprofen and furosemide was established. Based on the selected conditions for co-culture, we also confirmed the functionality of the final chosen culture condition using nitric oxide as an indicator of intestinal inflammation. Choice of cell culture time and culture medium represented two of the most important factors that affected TEER values and the permeability coefficients of the model drugs. On the other hand, the seeding density and the Caco-2:HT29-MTX ratio exerted no significant influence on TEER values and the drug permeability coefficients. No absolute optimal cell culture condition could be obtained for all drugs; however subsequent confirmation experiments concluded that excellent precision for TEER values and drug permeability coefficients was obtained from the two operators using the following combination of conditions, namely an initial seeding density of 1 x 10(5) Caco-2 and HT29-MTX cells/cm(2) at a ratio of 9:1, followed by a 21day culture time in MEM medium. Finally, functionality of the co-culture model system using the above selected in vitro conditions resulted in comparable nitric oxide synthesis to that of a Caco-2

  5. Chemopreventive Potential of Powdered Red Wine Pomace Seasonings against Colorectal Cancer in HT-29 Cells.

    Science.gov (United States)

    Del Pino-García, Raquel; Rivero-Pérez, María D; González-SanJosé, María L; Ortega-Heras, Miriam; García Lomillo, Javier; Muñiz, Pilar

    2017-01-11

    This study evaluates the antiproliferative and antigenotoxic actions of powdered red wine pomace seasonings (Sk-S, seedless; W-S, whole; Sd-S, seeds). In vitro gastrointestinal digested and colonic fermented fractions of the seasonings were used as cell treatments. Phenolic acids from Sk-S showed the highest bioaccessibility in the small intestine, whereas polyphenols contained in Sd-S might be the most fermentable in the colon. Dietary fiber from Sk-S was the best substrate for short chain fatty acids production by gut microbiota. Colon cancerous (HT-29) cell viability was inhibited by 50% (IC 50 values) at treatment concentrations ranging from 845 (Sk-S) to 1085 (Sd-S) μg/mL prior digestion, but all digested fractions exhibited similar antiproliferative activities (mean IC 50 = 814 μg/mL). Oxidative DNA damage in cells was also attenuated by the treatments (200 μg/mL, 24 h preincubation), with all colonic fermented fractions displaying similar genoprotective action. These results suggest the potential of red wine pomace seasonings as chemopreventive agents in colorectal cancer.

  6. Genistein modulates the anti-tumor activity of cisplatin in MCF-7 breast and HT-29 colon cancer cells.

    Science.gov (United States)

    Hu, Xiao-Juan; Xie, Ming-Yong; Kluxen, Felix M; Diel, Patrick

    2014-03-01

    The function of genistein (GEN) on tumor prevention and tumor promotion is discussed controversially. A possible interference of GEN with chemotherapy has been only rarely addressed so far. In this study, effects of GEN on the anti-tumor activity of cisplatin (CIS) were investigated in the presence and absence of estradiol (10(-10) M) in MCF-7 breast and HT-29 colon cancer cells. Cells were treated with graded concentrations of GEN (10(-4)-10(-6) M), E2, CIS and combinations. Cell growth, proliferation and apoptosis were determined as well as the expression level of PCNA, Ki67 and BCL-2 family members. CIS and GEN 10(-4) M inhibited cell growth and induced apoptosis in MCF-7 and HT-29 cells in the presence and absence of E2. Co-treatment with CIS and 10(-4)M GEN resulted in additive effects. In concentrations of 10(-5) and 10(-6) M, GEN stimulated cell growth in MCF-7 cells. It promoted proliferation, inhibited apoptosis and counteracted the anti-tumor activity of CIS in MCF-7 and HT-29 cells. Particularly the ability of CIS to induce apoptosis was antagonized. In ER alpha-positive MCF-7 cells, but not in ER alpha-negative HT-29 cells, E2 was able to neutralize the anti-CIS effects of GEN. Our data provide evidence that GEN in the absence of E2, a situation which occurs in postmenopausal women, directly affects the anti-tumor activity of cytostatic drugs like CIS. The exact molecular mechanism has to be investigated in future studies.

  7. In vitro study of the effects of radio frequency generated for plasma in neoplastic cells HT-29; Estudo in vitro dos efeitos da radiofrequencia gerada por plasmas em celulas neoplasicas HT-29

    Energy Technology Data Exchange (ETDEWEB)

    Andrighetto, Daniela; Dornelles, Eduardo Bortoluzzi; Cruz, Ivana Beatrice Manica da; Lüdke, Everton, E-mail: daniela.andrighetto@hotmail.com, E-mail: dornellesedu@gmail.com, E-mail: ibmcruz@hotmail.com, E-mail: evertonludke@gmail.com [Universidade Federal de Santa Maria (UFSM), RS (BRazil)

    2014-07-01

    The goal of this study is to develop an in vitro irradiation cell system with controllable irradiation intensities of 27 MHz produced by an argon plasma column with variable amplitude modulation in the 100-700 kHz range. This paper presents and discusses a proposed experiment, with toxicity analysis (DNA Picogreen®) and cell viability (MTT assay) in the radiation-induced HT-29 cell line (colon adenocarcinoma). The data allow us to observe that cellular toxicity effects may occur with exposure to fields produced by argon plasma with intensities on the order of at least 3.2 W / cm2 and exposure times above 3.5 hours continuously. An analysis of cell populations for cell toxicity tests using the Student's t-test did not show significant changes (p <0.05) in the amount of DNA released by the action of radiofrequency, although it has been found that cell viability (MTT) is not significantly altered by long exposures to radiation induced plasma RF signals in 27 MHz (p> 0.34). Cytotoxic effects due to the destruction of cell wall by heating the samples were not detected in any of the tests.

  8. Modulation of pathogen-induced CCL20 secretion from HT-29 human intestinal epithelial cells by commensal bacteria.

    LENUS (Irish Health Repository)

    Sibartie, Shomik

    2009-01-01

    BACKGROUND: Human intestinal epithelial cells (IECs) secrete the chemokine CCL20 in response to infection by various enteropathogenic bacteria or exposure to bacterial flagellin. CCL20 recruits immature dendritic cells and lymphocytes to target sites. Here we investigated IEC responses to various pathogenic and commensal bacteria as well as the modulatory effects of commensal bacteria on pathogen-induced CCL20 secretion. HT-29 human IECs were incubated with commensal bacteria (Bifidobacterium infantis or Lactobacillus salivarius), or with Salmonella typhimurium, its flagellin, Clostridium difficile, Mycobacterium paratuberculosis, or Mycobacterium smegmatis for varying times. In some studies, HT-29 cells were pre-treated with a commensal strain for 2 hr prior to infection or flagellin stimulation. CCL20 and interleukin (IL)-8 secretion and nuclear factor (NF)-kappaB activation were measured using enzyme-linked immunosorbent assays. RESULTS: Compared to untreated cells, S. typhimurium, C. difficile, M. paratuberculosis, and flagellin activated NF-kappaB and stimulated significant secretion of CCL20 and IL-8 by HT-29 cells. Conversely, B. infantis, L. salivarius or M. smegmatis did not activate NF-kappaB or augment CCL20 or IL-8 production. Treatment with B. infantis, but not L. salivarius, dose-dependently inhibited the baseline secretion of CCL20. In cells pre-treated with B. infantis, C. difficile-, S. typhimurium-, and flagellin-induced CCL20 were significantly attenuated. B. infantis did not limit M. Paratuberculosis-induced CCL20 secretion. CONCLUSION: This study is the first to demonstrate that a commensal strain can attenuate CCL20 secretion in HT-29 IECs. Collectively, the data indicate that M. paratuberculosis may mediate mucosal damage and that B. infantis can exert immunomodulatory effects on IECs that mediate host responses to flagellin and flagellated enteric pathogens.

  9. Luteolin inhibits the colon cancer HT-29 cell proliferation, migration and epithelial-mesenchymal transition: an experimental study

    Directory of Open Access Journals (Sweden)

    Xin Meng

    2017-11-01

    Full Text Available Objective: To study the regulating effect of luteolin on colon cancer HT-29 cell proliferation, migration and epithelial-mesenchymal transition. Methods: Colon cancer HT-29 cells were cultured and randomly divided into two groups, control group were treated with serum-free medium without drugs and LUT group were treated with serum-free medium containing luteolin. After 24 h of treatment, cells were collected to extract RNA, and then fluorescent quantitative PCR method was used to determine the mRNA expression of proliferation genes, migration genes and epithelial-mesenchymal transition genes. Results: After 24 h of luteolin treatment, Lrig1, TSPYL5, Bim, SOX15 and DLC1 mRNA expression in LUT group were significantly higher than those in control group while RPS15a, Bad, TRPV5, TRPV6, PLD2, IBP, SphK1, FAK, Vimentin and N-cadherin mRNA expression were significantly lower than those in control group. Conclusion: Luteolin has inhibiting effect on colon cancer HT-29 cell proliferation, migration and epithelial-mesenchymal transition.

  10. Rottlerin inhibits ROS formation and prevents NFkappaB activation in MCF-7 and HT-29 cells

    Czech Academy of Sciences Publication Activity Database

    Maioli, E.; Greci, L.; Souček, Karel; Hýžďalová, Martina; Pecorelli, A.; Fortino, V.; Valacchi, G.

    -, - (2009), s. 1-7 ISSN 1110-7243 R&D Projects: GA ČR(CZ) GA204/07/0834 Grant - others:GA ČR(CZ) GA310/07/0961 Program:GA Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : Rottlerin * NFkappaB * HT-29 cells Subject RIV: BO - Biophysics Impact factor: 1.750, year: 2009

  11. Transcriptional response of HT-29 intestinal epithelial cells to human and bovine milk oligosaccharides.

    Science.gov (United States)

    Lane, Jonathan A; O'Callaghan, John; Carrington, Stephen D; Hickey, Rita M

    2013-12-01

    Human milk oligosaccharides (HMO) have been shown to interact directly with immune cells. However, large quantities of HMO are required for intervention or clinical studies, but these are unavailable in most cases. In this respect, bovine milk is potentially an excellent source of commercially viable analogues of these unique molecules. In the present study, we compared the transcriptional response of colonic epithelial cells (HT-29) to the entire pool of HMO and bovine colostrum oligosaccharides (BCO) to determine whether the oligosaccharides from bovine milk had effects on gene expression that were similar to those of their human counterparts. Gene set enrichment analysis of the transcriptional data revealed that there were a number of similar biological processes that may be influenced by both treatments including a response to stimulus, signalling, locomotion, and multicellular, developmental and immune system processes. For a more detailed insight into the effects of milk oligosaccharides, the effect on the expression of immune system-associated glycogenes was chosen as a case study when performing validation studies. Glycogenes in the current context are genes that are directly or indirectly regulated in the presence of glycans and/or glycoconjugates. RT-PCR analysis revealed that HMO and BCO influenced the expression of cytokines (IL-1β, IL-8, colony-stimulating factor 2 (granulocyte-macrophage) (GM-CSF2), IL-17C and platelet factor 4 (PF4)), chemokines (chemokine (C-X-C motif) ligand 1 (CXCL1), chemokine (C-X-C motif) ligand 3 (CXCL3), chemokine (C-C motif) ligand 20 (CCL20), chemokine (C-X-C motif) ligand 2 (CXCL2), chemokine (C-X-C motif) ligand 6 (CXCL6), chemokine (C-C motif) ligand 5 (CCL5), chemokine (C-X3-C motif) ligand 1 (CX3CL1) and CXCL2) and cell surface receptors (interferon γ receptor 1 (IFNGR1), intercellular adhesion molecule-1 (ICAM-1), intercellular adhesion molecule-2 (ICAM-2) and IL-10 receptor α (IL10RA)). The present study suggests

  12. The natural triterpene maslinic acid induces apoptosis in HT29 colon cancer cells by a JNK-p53-dependent mechanism

    International Nuclear Information System (INIS)

    Reyes-Zurita, Fernando J; Pachón-Peña, Gisela; Lizárraga, Daneida; Rufino-Palomares, Eva E; Cascante, Marta; Lupiáñez, José A

    2011-01-01

    Maslinic acid, a pentacyclic triterpene found in the protective wax-like coating of the leaves and fruit of Olea europaea L., is a promising agent for the prevention of colon cancer. We have shown elsewhere that maslinic acid inhibits cell proliferation to a significant extent and activates mitochondrial apoptosis in colon cancer cells. In our latest work we have investigated further this compound's apoptotic molecular mechanism. We used HT29 adenocarcinoma cells. Changes genotoxicity were analyzed by single-cell gel electrophoresis (comet assay). The cell cycle was determined by flow cytometry. Finally, changes in protein expression were examined by western blotting. Student's t-test was used for statistical comparison. HT29 cells treated with maslinic acid showed significant increases in genotoxicity and cell-cycle arrest during the G0/G1 phase after 72 hours' treatment and an apoptotic sub-G0/G1 peak after 96 hours. Nevertheless, the molecular mechanism for this cytotoxic effect of maslinic acid has never been properly explored. We show here that the anti-tumoral activity of maslinic acid might proceed via p53-mediated apoptosis by acting upon the main signaling components that lead to an increase in p53 activity and the induction of the rest of the factors that participate in the apoptotic pathway. We found that in HT29 cells maslinic acid activated the expression of c-Jun NH2-terminal kinase (JNK), thus inducing p53. Treatment of tumor cells with maslinic acid also resulted in an increase in the expression of Bid and Bax, repression of Bcl-2, release of cytochrome-c and an increase in the expression of caspases -9, -3, and -7. Moreover, maslinic acid produced belated caspase-8 activity, thus amplifying the initial mitochondrial apoptotic signaling. All these results suggest that maslinic acid induces apoptosis in human HT29 colon-cancer cells through the JNK-Bid-mediated mitochondrial apoptotic pathway via the activation of p53. Thus we propose

  13. Dual effects of human adipose tissue-derived mesenchymal stem cells in human lung adenocarcinoma A549 xenografts and colorectal adenocarcinoma HT-29 xenografts in mice.

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    Rhyu, Jung Joo; Yun, Jun-Won; Kwon, Euna; Che, Jeong-Hwan; Kang, Byeong-Cheol

    2015-10-01

    Human adipose tissue-derived mesenchymal stem cells (hATMSCs) have great potential as a therapy for various diseases. However, emerging evidence shows that there are conflicting results concerning effects of hATMSCs on tumor progression. Our objective was to determine whether and how hATMSCs modulate tumor growth. After cancer cell lines were subcutaneously inoculated into BALB/c-nude and hairless severe combined immunodeficient mice, hATMSCs were intratumorally injected into the mice. The growth of the A549 tumors was inhibited by hATMSCs, yet that of the HT-29 tumors was significantly promoted by hATMSCs in the in vivo xenograft models. In vitro study using a co-culture system of cancer cells and hATMSCs was consistent with the in vivo experiments. To reveal the molecular events induced by hATMSCs in the xenograft models, global gene expression profiles of the A549 and HT-29 tumors in the absence or presence of hATMSCs were determined. Significant numbers of genes involved in biological processes were altered in the hATMSC-treated A549 tumors, whereas no biological process was regulated by treatment with hATMSCs in the HT-29 tumors, reflecting the different effects of hATMSCs in the different types of cancer. Notably, histone cluster 1, H2aj and neuropeptide Y receptor Y4 were found to be expressed in direct or inverse proportion to tumor size in both xenograft models. In addition, nuclear factor κB (NF-κB) p65 was differentially phosphorylated by the hATMSCs dependent on the source of the cancer cells. In conclusion, the identified gene profiling and NF-κB signaling provide molecular evidence to explain the conflicting findings in tumor‑MSC studies, although further study is needed to confirm these findings using various types of cancer.

  14. Targeting miR-21 enhances the sensitivity of human colon cancer HT-29 cells to chemoradiotherapy in vitro

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    Deng, Jun; Lei, Wan; Fu, Jian-Chun; Zhang, Ling; Li, Jun-He; Xiong, Jian-Ping, E-mail: jpxiong@medmail.com.cn

    2014-01-17

    Highlight: •MiR-21 plays a significant role in 5-FU resistance. •This role might be attributed to targeting of hMSH2 as well as TP and DPD via miR-21 targeted hMSH2. •Indirectly targeted TP and DPD to influence 5-FU chemotherapy sensitivity. -- Abstract: 5-Fluorouracil (5-FU) is a classic chemotherapeutic drug that has been widely used for colorectal cancer treatment, but colorectal cancer cells are often resistant to primary or acquired 5-FU therapy. Several studies have shown that miR-21 is significantly elevated in colorectal cancer. This suggests that this miRNA might play a role in this resistance. In this study, we investigated this possibility and the possible mechanism underlying this role. We showed that forced expression of miR-21 significantly inhibited apoptosis, enhanced cell proliferation, invasion, and colony formation ability, promoted G1/S cell cycle transition and increased the resistance of tumor cells to 5-FU and X radiation in HT-29 colon cancer cells. Furthermore, knockdown of miR-21 reversed these effects on HT-29 cells and increased the sensitivity of HT-29/5-FU to 5-FU chemotherapy. Finally, we showed that miR-21 targeted the human mutS homolog2 (hMSH2), and indirectly regulated the expression of thymidine phosphorylase (TP) and dihydropyrimidine dehydrogenase (DPD). These results demonstrate that miR-21 may play an important role in the 5-FU resistance of colon cancer cells.

  15. Influence of in vitro supplementation with lipids from conventional and Alpine milk on fatty acid distribution and cell growth of HT-29 cells

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    Dänicke Sven

    2011-08-01

    Full Text Available Abstract Background To date, the influence of milk and dairy products on carcinogenesis remains controversial. However, lipids of ruminant origin such as conjugated linoleic acids (CLA are known to exhibit beneficial effects in vitro and in vivo. The aim of the present study was to determine the influence of milk lipids of different origin and varying quality presenting as free fatty acid (FFA solutions on cellular fatty acid distribution, cellular viability, and growth of human colon adenocarcinoma cells (HT-29. Methods FAME of conventional and Alpine milk lipids (MLcon, MLalp and cells treated with FFA derivatives of milk lipids were analyzed by means of GC-FID and Ag+-HPLC. Cellular viability and growth of the cells were determined by means of CellTiter-Blue®-assay and DAPI-assay (4',6-diamidino-2-phenylindole dihydrochloride, respectively. Results Supplementation with milk lipids significantly decreased viability and growth of HT-29 cells in a dose- and time-dependent manner. MLalp showed a lower SFA/MUFA ratio, a 8 fold increased CLA content, and different CLA profile compared to MLcon but did not demonstrate additional growth-inhibitory effects. In addition, total concentration and fatty acid distribution of cellular lipids were altered. In particular, treatment of the cells yielded highest amounts of two types of milk specific major fatty acids (μg FA/mg cellular protein after 8 h of incubation compared to 24 h; 200 μM of MLcon (C16:0, 206 ± 43, 200 μM of MLalp (C18:1 c9, (223 ± 19. Vaccenic acid (C18:1 t11 contained in milk lipids was converted to c9,t11-CLA in HT-29 cells. Notably, the ratio of t11,c13-CLA/t7,c9-CLA, a criterion for pasture feeding of the cows, was significantly changed after incubation for 8 h with lipids from MLalp (3.6 - 4.8, compared to lipids from MLcon (0.3 - 0.6. Conclusions Natural lipids from conventional and Alpine milk showed similar growth inhibitory effects. However, different changes in cellular

  16. Influence of in vitro supplementation with lipids from conventional and Alpine milk on fatty acid distribution and cell growth of HT-29 cells

    Science.gov (United States)

    2011-01-01

    Background To date, the influence of milk and dairy products on carcinogenesis remains controversial. However, lipids of ruminant origin such as conjugated linoleic acids (CLA) are known to exhibit beneficial effects in vitro and in vivo. The aim of the present study was to determine the influence of milk lipids of different origin and varying quality presenting as free fatty acid (FFA) solutions on cellular fatty acid distribution, cellular viability, and growth of human colon adenocarcinoma cells (HT-29). Methods FAME of conventional and Alpine milk lipids (MLcon, MLalp) and cells treated with FFA derivatives of milk lipids were analyzed by means of GC-FID and Ag+-HPLC. Cellular viability and growth of the cells were determined by means of CellTiter-Blue®-assay and DAPI-assay (4',6-diamidino-2-phenylindole dihydrochloride), respectively. Results Supplementation with milk lipids significantly decreased viability and growth of HT-29 cells in a dose- and time-dependent manner. MLalp showed a lower SFA/MUFA ratio, a 8 fold increased CLA content, and different CLA profile compared to MLcon but did not demonstrate additional growth-inhibitory effects. In addition, total concentration and fatty acid distribution of cellular lipids were altered. In particular, treatment of the cells yielded highest amounts of two types of milk specific major fatty acids (μg FA/mg cellular protein) after 8 h of incubation compared to 24 h; 200 μM of MLcon (C16:0, 206 ± 43), 200 μM of MLalp (C18:1 c9, (223 ± 19). Vaccenic acid (C18:1 t11) contained in milk lipids was converted to c9,t11-CLA in HT-29 cells. Notably, the ratio of t11,c13-CLA/t7,c9-CLA, a criterion for pasture feeding of the cows, was significantly changed after incubation for 8 h with lipids from MLalp (3.6 - 4.8), compared to lipids from MLcon (0.3 - 0.6). Conclusions Natural lipids from conventional and Alpine milk showed similar growth inhibitory effects. However, different changes in cellular lipid composition

  17. Chemopreventive activity of ellagitannins and their derivatives from black raspberry seeds on HT-29 colon cancer cells.

    Science.gov (United States)

    Cho, Hyunnho; Jung, Hana; Lee, Heejae; Yi, Hae Chang; Kwak, Ho-kyung; Hwang, Keum Taek

    2015-05-01

    Black raspberry (BRB) seeds are a major waste product after fruit processing. The seeds are abundant in ellagitannins (ET), a class of hydrolysable tannins, which are hydrolyzed to ellagic acid (EA) and further metabolized to urolithin A (UA) and urolithin B (UB), known to be bioavailable in the colon and the prostate. In this study, the anti-cancer activities of these compounds were evaluated on HT-29 colon cancer cells. ET, EA, UA and UB inhibited the proliferation of the cancer cells. EA caused a slight, but significant cell cycle arrest at the G1 phase, and urolithins caused cell cycle arrest at the G2/M phase and upregulated p21 expression. Apoptotic cells were detected by Annexin V-FITC/PI assay when treated with the compounds. Disruption in mitochondrial membrane potential and activation of caspases 8 and 9 suggest that both extrinsic and intrinsic apoptotic pathways may be involved. Activation of caspase 3 and cleavage of PARP further confirmed the induction of the apoptosis. ET, EA, UA and UB showed anti-cancer activity by arresting the cell cycle and inducing apoptosis on HT-29 human colon cancer cells. This study suggests that the BRB seeds could be a potential source of anti-cancer ET.

  18. A novel cycloartane triterpenoid from Cimicifuga induces apoptotic and autophagic cell death in human colon cancer HT-29 cells.

    Science.gov (United States)

    Dai, Xiaoli; Liu, Jing; Nian, Yin; Qiu, Ming-Hua; Luo, Ying; Zhang, Jihong

    2017-04-01

    The extract from Cimicifuga, a genus of flowering plants, has been demonstrated to have mainly therapeutic effects on menstrual and menopausal symptoms and also exhibits immunomodulatory, anti-inflammatory and antimicrobial activity. Moreover, the anticancer effects of Cimicifuga have been reported, but the underlying mechanism causing cancer cell death has been poorly described. The present study was designed to investigate the antitumor effects and underlying molecular mechanisms of cimigenol (KY17), a novel cycloartane triterpenoid from Cimicifuga. KY17-induced autophagy and apoptotic cell death in human colon cancer cells (HT-29) was investigated. KY17 treatment induced growth inhibition and apoptotic cell death in a concentration-dependent manner. The induction of apoptosis was confirmed by a change in cell morphology, and an increase in the G2/M phase, as well as increased protein levels of cleaved-caspase-8 and -3; cleavage of poly(ADP-ribose) polymerase (PARP) in the HT-29 cells following KY17 treatment. In addition, autophagy was evaluated by the accumulation of acridine orange, the appearance of green fluorescent protein-light-chain 3 (LC3) punctate structures and increased levels of LC3-II protein expression. Furthermore, combination treatment with the autophagy inhibitor bafilomycin A1 enhanced the induction of apoptosis by KY17. Taken together, the present study provides new insight into the role of KY17 as a potential antitumor compound. Combination of KY17 with an autophagy inhibitor may be a valuable strategy for the chemoprevention or treatment of colon cancer.

  19. Dietary compounds that induce cancer preventive phase 2 enzymes activate apoptosis at comparable doses in HT29 colon carcinoma cells.

    Science.gov (United States)

    Kirlin, W G; Cai, J; DeLong, M J; Patten, E J; Jones, D P

    1999-10-01

    Dietary agents that induce glutathione S-transferases and related detoxification systems (Phase 2 enzyme inducers) are thought to prevent cancer by enhancing elimination of chemical carcinogens. The present study shows that compounds of this group (benzyl isothiocyanate, allyl sulfide, dimethyl fumarate, butylated hydroxyanisole) activated apoptosis in human colon carcinoma (HT29) cells in culture over the same concentration ranges that elicited increases in enzyme activity (5-25, 25-100, 10-100, 15-60 micromol/L, respectively). Pretreatment of cells with sodium butyrate, an agent that induces HT29 cell differentiation, resulted in parallel increases in Phase 2 enzyme activities and induction of apoptosis in response to the inducers. Cell death characteristics included apoptotic morphological changes, appearance of cells at sub-G1 phase on flow cytometry, caspase activation, DNA fragmentation and TUNEL-positive staining. The results suggest that dietary Phase 2 inducers may protect against cancer by a mechanism distinct from and in addition to that associated with enhanced elimination of carcinogens. If this occurs in vivo, diets high in such compounds could eliminate precancerous cells by apoptosis at time points well after initial exposure to chemical mutagens and carcinogens.

  20. Morusin induces apoptosis and suppresses NF-κB activity in human colorectal cancer HT-29 cells

    International Nuclear Information System (INIS)

    Lee, J.-C.; Won, S.-J.; Chao, C.-L.; Wu, F.-L.; Liu, H.-S.; Ling Pin; Lin, C.-N.; Su, C.-L.

    2008-01-01

    Morusin is a pure compound isolated from root bark of Morusaustralis (Moraceae). In this study, we demonstrated that morusin significantly inhibited the growth and clonogenicity of human colorectal cancer HT-29 cells. Apoptosis induced by morusin was characterized by accumulation of cells at the sub-G 1 phase, fragmentation of DNA, and condensation of chromatin. Morusin also inhibited the phosphorylation of IKK-α, IKK-β and IκB-α, increased expression of IκB-α, and suppressed nuclear translocation of NF-κB and its DNA binding activity. Dephosphorylation of NF-κB upstream regulators PI3K, Akt and PDK1 was also displayed. In addition, activation of caspase-8, change of mitochondrial membrane potential, release of cytochrome c and Smac/DIABLO, and activation of caspase-9 and -3 were observed at the early time point. Downregulation in the expression of Ku70 and XIAP was exhibited afterward. Caspase-8 or wide-ranging caspase inhibitor suppressed morusin-induced apoptosis. Therefore, the antitumor mechanism of morusin in HT-29 cells may be via activation of caspases and inhibition of NF-κB

  1. Interaction of cruciferin-based nanoparticles with Caco-2 cells and Caco-2/HT29-MTX co-cultures.

    Science.gov (United States)

    Akbari, Ali; Lavasanifar, Afsaneh; Wu, Jianping

    2017-12-01

    The objective of this work was to assess the potential of Cruciferin/Calcium (Cru/Ca) and Cruciferin/Chitosan (Cru/Cs) nanoparticles for oral drug delivery. For this purpose, Cru/Ca and Cru/Cs nanoparticles were developed through cold gelation of Cruciferin, a major canola protein, and in interaction with calcium and chitosan, respectively. The extent and rate of particle uptake in Caco-2 cells and Caco-2/HT29 co-culture was then evaluated by fluorescence spectroscopy as well as flow cytometry. Through pre-incubation of Caco-2 cell monolayer with specific endocytosis inhibitors, the mechanism of cell uptake was investigated. Our results showed that the uptake of negatively-charged Cru/Ca particles to be ∼3 times higher than positively-charged Cru/Cs ones by Caco-2 cells. Presence of mucus secreted by HT29 cells in their co-culture with Caco-2 had negligible influence on the uptake and transport of both particles. In contrast to Cru/Ca particles which were dissociated in the simulated gastrointestinal conditions, digestion of Cru/Cs particles resulted in 6- and 2-fold increase in the cellular uptake and transport of encapsulated coumarin in the latter particles, respectively. While the presence of mucus in Caco-2/HT29 co-culture caused 40-50% decrease of cellular uptake and transport for coumarin encapsulated in digested Cru/Cs particles, it had no significant effect on the cell uptake and transport of coumarin associated with Cru/Ca particles after digestion. Energy-dependent mechanisms were the dominant mechanism for uptake of both undigested and digested particles. Therefore, in Caco-2/HT29 co-culture which closely simulated intestinal epithelial cells, undigested Cru/Ca and Cru/Cs particles had the ability to penetrate mucus layers, while digested Cru/Cs particles showed mucoadhesive property, and digested Cru/Ca particles were dissociated. Our results points to a potential for cruciferin based nanoparticles for oral drug delivery. The long-term objective of

  2. Influence of cell cycle phase on radiation-induced cytotoxicity and DNA damage in human colon cancer (HT29) and Chinese hamster ovary cells

    International Nuclear Information System (INIS)

    Tang, H.; Davis, M.A.; Strickfaden, S.M.; Maybaum, J.; Lawrence, T.S.

    1994-01-01

    We have previously shown that fluorodeoxyuridine (FdUrd) radiosensitizes HT29 human colon carcinoma cells. Since treatment with FdUrd arrests cells at the G 1 /S-phase interface, a condition associated with increased radiation sensitivity in some cells, it seemed possible that redistribution of cells in the phases of the cell cycle might account for FdUrd-mediated radiosensitization. To begin to test this, HT29 cells were separated by centrifugal elutriation according to cell cycle phase and assessed for radiosensitivity, using a clonogenic assay, and radiation-induced DNA damage, using pulsed-field gel electrophoresis. We found that all of the elutriated fractions (which contained cells enriched in G 1 , G 1 /early S, mid to late S or G 2 /M phase) had the same radiation sensitivity and expressed a similar extent of radiation-induced DNA damage. To determine if the techniques used in this study could detect differences between the radiation sensitivity of cells in different phases of the cell cycle, analogous experiments were carried out using Chinese hamster ovary (CHO) cells. In contrast with the results of experiments with HT29 cells, but in agreement with previous studies, CHO cells separated under the same conditions as were used for HT29 cells showed a marked dependence on cell age of both clonogenic survival and radiation-induced DNA damage. Thus, within the limitations of the purity of separation obtained using elutriation, the radiation sensitivity of HT29 cells does not vary substantially as a function of cell cycle phase. Therefore, it seems unlikely that cell cycle redistribution alone explains the radiation sensitivity produced by exposure to FdUrd. 20 refs., 4 figs., 1 tab

  3. Oral and fecal Campylobacter concisus strains perturb barrier function by apoptosis induction in HT-29/B6 intestinal epithelial cells.

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    Hans Linde Nielsen

    Full Text Available Campylobacter concisus infections of the gastrointestinal tract can be accompanied by diarrhea and inflammation, whereas colonization of the human oral cavity might have a commensal nature. We focus on the pathophysiology of C. concisus and the effects of different clinical oral and fecal C. concisus strains on human HT-29/B6 colon cells. Six oral and eight fecal strains of C. concisus were isolated. Mucus-producing HT-29/B6 epithelial monolayers were infected with the C. concisus strains. Transepithelial electrical resistance (R(t and tracer fluxes of different molecule size were measured in Ussing chambers. Tight junction (TJ protein expression was determined by Western blotting, and subcellular TJ distribution was analyzed by confocal laser-scanning microscopy. Apoptosis induction was examined by TUNEL-staining and Western blot of caspase-3 activation. All strains invaded confluent HT-29/B6 cells and impaired epithelial barrier function, characterized by a time- and dose-dependent decrease in R(t either after infection from the apical side but even more from the basolateral compartment. TJ protein expression changes were sparse, only in apoptotic areas of infected monolayers TJ proteins were redistributed. Solely the barrier-forming TJ protein claudin-5 showed a reduced expression level to 66±8% (P<0.05, by expression regulation from the gene. Concomitantly, Lactate dehydrogenase release was elevated to 3.1±0.3% versus 0.7±0.1% in control (P<0.001, suggesting cytotoxic effects. Furthermore, oral and fecal C. concisus strains elevated apoptotic events to 5-fold. C. concisus-infected monolayers revealed an increased permeability for 332 Da fluorescein (1.74±0.13 vs. 0.56±0.17 10(-6 cm/s in control, P<0.05 but showed no difference in permeability for 4 kDa FITC-dextran (FD-4. The same was true in camptothecin-exposed monolayers, where camptothecin was used for apoptosis induction.In conclusion, epithelial barrier dysfunction by oral and

  4. Acetylbritannilactone suppresses growth via upregulation of krüppel-like transcription factor 4 expression in HT-29 colorectal cancer cells.

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    Fang, Xin-Mei; Liu, Bin; Liu, Ya-Bin; Wang, Jun-Jie; Wen, Jin-Kun; Li, Bing-Hui; Han, Mei

    2011-11-01

    Acetylbritannilactone (ABL) is a new active compound isolated from Inula Britannica L, a traditional Chinese medicinal herb. It has been reported that ABL can inhibit the proliferation of vascular smooth muscle cells (VSMCs) and neointima formation after balloon injury in rats. ABL also shows chemopreventive properties by inducing cell apoptosis in breast and ovarian cancers, but the antitumor activity and the molecular targets of ABL in colon cancer cells have not been determined. In this study, we showed that ABL inhibits the growth in dose- and time-dependent manners by inducing cell cycle arrest in G0/G1 phase of HT-29 human colon cancer cells. This suppression was accompanied by a strong decrease of cyclin E and CDK4 protein levels, and an increase in p21 protein expression in HT-29 cells. We also show that ABL-induced growth inhibition is associated with the upregulation of KLF4 expression. The overexpression of KLF4 by infection with pAd-KLF4 resulted in growth inhibition, with decrease in the protein levels of cyclin E and CDK4, and increase in the expression of p21, similarly to the effects of ABL. Conversely, knockdown of KLF4 using a specific siRNA impaired the ABL-induced growth inhibition in HT-29 cells. These results suggest that KLF4 as an important cellular target of ABL mediates the growth inhibition of HT-29 cells induced by ABL via upregulation of p21 expression.

  5. Crataegus azarolus Leaves Induce Antiproliferative Activity, Cell Cycle Arrest, and Apoptosis in Human HT-29 and HCT-116 Colorectal Cancer Cells.

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    Mustapha, Nadia; Pinon, Aline; Limami, Youness; Simon, Alain; Ghedira, Kamel; Hennebelle, Thierry; Chekir-Ghedira, Leila

    2016-05-01

    Limited success has been achieved in extending the survival of patients with metastatic colorectal cancer (CRC). There is a strong need for novel agents in the treatment and prevention of CRC. Therefore, in the present study we evaluated the antiproliferative and pro-apoptotic potential of Crataegus azarolus ethyl acetate extract in HCT-116 and HT-29 human colorectal cancer cell lines. Moreover, we attempted to investigate the signaling pathways that should be involved in its cytotoxic effect. The Crataegus azarolus ethyl acetate extract-induced growth inhibitory effect was associated with DNA fragmentation, sub-G1 peak, loss of mitochondrial potential, and poly (ADP-ribose) polymerase (PARP) cleavage. In addition, ethyl acetate extract of Crataegus azarolus induced the cleavage of caspase-8. It has no effect on steady-state levels of total Bcl-2 protein. Whereas Bax levels decreased significantly in a dose-dependent manner in both tested cell lines. Taken together, these findings confirm the involvement of the extrinsic pathway of apoptosis. The apoptotic cell death induced by ethyl acetate extract of Crataegus azarolus was accompanied by an enhancement of the p21 expression but not through p53 activation in human colorectal cancer cells. The above-mentioned data provide insight into the molecular mechanisms of Crataegus azarolus ethyl acetate extract-induced apoptosis in CRC. Therefore, this compound should be a potential anticancer agent for the treatment of CRC. © 2015 Wiley Periodicals, Inc.

  6. Induction of G1 and G2/M cell cycle arrests by the dietary compound 3,3'-diindolylmethane in HT-29 human colon cancer cells

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    Choi Hyun

    2009-05-01

    Full Text Available Abstract Background 3,3'-Diindolylmethane (DIM, an indole derivative produced in the stomach after the consumption of broccoli and other cruciferous vegetables, has been demonstrated to exert anti-cancer effects in both in vivo and in vitro models. We have previously determined that DIM (0 – 30 μmol/L inhibited the growth of HT-29 human colon cancer cells in a concentration-dependent fashion. In this study, we evaluated the effects of DIM on cell cycle progression in HT-29 cells. Methods HT-29 cells were cultured with various concentrations of DIM (0 – 30 μmol/L and the DNA was stained with propidium iodide, followed by flow cytometric analysis. [3H]Thymidine incorporation assays, Western blot analyses, immunoprecipitation and in vitro kinase assays for cyclin-dependent kinase (CDK and cell division cycle (CDC2 were conducted. Results The percentages of cells in the G1 and G2/M phases were dose-dependently increased and the percentages of cells in S phase were reduced within 12 h in DIM-treated cells. DIM also reduced DNA synthesis in a dose-dependent fashion. DIM markedly reduced CDK2 activity and the levels of phosphorylated retinoblastoma proteins (Rb and E2F-1, and also increased the levels of hypophosphorylated Rb. DIM reduced the protein levels of cyclin A, D1, and CDK4. DIM also increased the protein levels of CDK inhibitors, p21CIP1/WAF1 and p27KIPI. In addition, DIM reduced the activity of CDC2 and the levels of CDC25C phosphatase and cyclin B1. Conclusion Here, we have demonstrated that DIM induces G1 and G2/M phase cell cycle arrest in HT-29 cells, and this effect may be mediated by reduced CDK activity.

  7. Nicotine promotes cell proliferation via α7-nicotinic acetylcholine receptor and catecholamine-synthesizing enzymes-mediated pathway in human colon adenocarcinoma HT-29 cells

    International Nuclear Information System (INIS)

    Wong, Helen Pui Shan; Yu Le; Lam, Emily Kai Yee; Tai, Emily Kin Ki; Wu, William Ka Kei; Cho, Chi Hin

    2007-01-01

    Cigarette smoking has been implicated in colon cancer. Nicotine is a major alkaloid in cigarette smoke. In the present study, we showed that nicotine stimulated HT-29 cell proliferation and adrenaline production in a dose-dependent manner. The stimulatory action of nicotine was reversed by atenolol and ICI 118,551, a β 1 - and β 2 -selective antagonist, respectively, suggesting the role of β-adrenoceptors in mediating the action. Nicotine also significantly upregulated the expression of the catecholamine-synthesizing enzymes [tyrosine hydroxylase (TH), dopamine-β-hydroxylase (DβH) and phenylethanolamine N-methyltransferase]. Inhibitor of TH, a rate-limiting enzyme in the catecholamine-biosynthesis pathway, reduced the actions of nicotine on cell proliferation and adrenaline production. Expression of α7-nicotinic acetylcholine receptor (α7-nAChR) was demonstrated in HT-29 cells. Methyllycaconitine, an α7-nAChR antagonist, reversed the stimulatory actions of nicotine on cell proliferation, TH and DβH expression as well as adrenaline production. Taken together, through the action on α7-nAChR nicotine stimulates HT-29 cell proliferation via the upregulation of the catecholamine-synthesis pathway and ultimately adrenaline production and β-adrenergic activation. These data reveal the contributory role α7-nAChR and β-adrenoceptors in the tumorigenesis of colon cancer cells and partly elucidate the carcinogenic action of cigarette smoke on colon cancer

  8. Zinc Sensing Receptor Signaling, Mediated by GPR39, Reduces Butyrate-Induced Cell Death in HT29 Colonocytes via Upregulation of Clusterin

    Science.gov (United States)

    Cohen, Limor; Azriel-Tamir, Hagit; Arotsker, Natan; Sekler, Israel; Hershfinkel, Michal

    2012-01-01

    Zinc enhances epithelial proliferation, protects the digestive epithelial layer and has profound antiulcerative and antidiarrheal roles in the colon. Despite the clinical significance of this ion, the mechanisms linking zinc to these cellular processes are poorly understood. We have previously identified an extracellular Zn2+ sensing G-protein coupled receptor (ZnR) that activates Ca2+ signaling in colonocytes, but its molecular identity as well as its effects on colonocytes' survival remained elusive. Here, we show that Zn2+, by activation of the ZnR, protects HT29 colonocytes from butyrate induced cell death. Silencing of the G-protein coupled receptor GPR39 expression abolished ZnR-dependent Ca2+ release and Zn2+-dependent survival of butyrate-treated colonocytes. Importantly, GPR39 also mediated ZnR-dependent upregulation of Na+/H+ exchange activity as this activity was found in native colon tissue but not in tissue obtained from GPR39 knock-out mice. Although ZnR-dependent upregulation of Na+/H+ exchange reduced the cellular acid load induced by butyrate, it did not rescue HT29 cells from butyrate induced cell death. ZnR/GPR39 activation however, increased the expression of the anti-apoptotic protein clusterin in butyrate-treated cells. Furthermore, silencing of clusterin abolished the Zn2+-dependent survival of HT29 cells. Altogether, our results demonstrate that extracellular Zn2+, acting through ZnR, regulates intracellular pH and clusterin expression thereby enhancing survival of HT29 colonocytes. Moreover, we identify GPR39 as the molecular moiety of ZnR in HT29 and native colonocytes. PMID:22545109

  9. Protective effect of Carnobacterium spp. against Listeria monocytogenes during host cell invasion using in vitro HT29 model

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    Tereza Pilchova

    2016-08-01

    Full Text Available The pathogenesis of listeriosis results mainly from the ability of Listeria monocytogenes to attach, invade, replicate and survive within various cell types in mammalian tissues. In this work, the effect of two bacteriocin-producing Carnobacterium (C. divergens V41 and C. maltaromaticum V1 and three non-bacteriocinogenic strains: (C. divergens V41C9, C. divergens 2763 and C. maltaromaticum 2762 was investigated on the reduction of L. monocytogenes Scott A plaque-forming during human infection using the HT-29 in vitro model. All Carnobacteria tested resulted in a reduction in the epithelial cell invasion caused by L. monocytogenes Scott A. To understand better the mechanism underlying the level of L. monocytogenes infection inhibition by Carnobacteria, infection assays from various pretreatments of Carnobacteria were assessed. The results revealed the influence of bacteriocin production combined with a passive mechanism of mammalian cell monolayers protection by Carnobacteria. These initial results showing a reduction in L. monocytogenes virulence on epithelial cells by Carnobacteria would be worthwhile analyzing further as a promising probiotic tool for human health.

  10. Annona muricata leaves induce G₁ cell cycle arrest and apoptosis through mitochondria-mediated pathway in human HCT-116 and HT-29 colon cancer cells.

    Science.gov (United States)

    Zorofchian Moghadamtousi, Soheil; Karimian, Hamed; Rouhollahi, Elham; Paydar, Mohammadjavad; Fadaeinasab, Mehran; Abdul Kadir, Habsah

    2014-10-28

    Annona muricata known as "the cancer killer" has been widely used in the traditional medicine for the treatment of cancer and tumors. The purpose of this study is to investigate the anticancer properties of ethyl acetate extract of Annona muricata leaves (EEAM) on HT-29 and HCT-116 colon cancer cells and the underlying mechanisms. The effect of EEAM on the cell proliferation of HT-29 and HCT-116 cells was analyzed by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium) assay. High content screening system (HCS) was applied to investigate the cell membrane permeability, mitochondrial membrane potential (MMP), nuclear condensation and cytochrome c translocation from mitochondria to cytosol. Reactive oxygen species (ROS) formation, lactate dehydrogenase (LDH) release and activation of caspase-3/7, -8 and -9 were measured while treatment. Flow cytometric analysis was used to determine the cell cycle distribution and phosphatidylserine externalization. The protein expression of Bax and Bcl-2 was determined using immunofluorescence analysis. In addition, the potential of EEAM to suppress the migration and invasion of colon cancer cells was also examined. EEAM exerted significant cytotoxic effects on HCT-116 and HT-29 cells as determined by MTT and LDH assays. After 24 h treatment, EEAM exhibited the IC₅₀ value of 11.43 ± 1.87 µg/ml and 8.98 ± 1.24 µg/ml against HT-29 and HCT-116 cells, respectively. Flow cytometric analysis demonstrated the cell cycle arrest at G1 phase and phosphatidylserine externalization confirming the induction of apoptosis. EEAM treatment caused excessive accumulation of ROS followed by disruption of MMP, cytochrome c leakage and activation of the initiator and executioner caspases in both colon cancer cells. Immunofluorescence analysis depicted the up-regulation of Bax and down-regulation of Bcl-2 proteins while treated with EEAM. Furthermore, EEAM conspicuously blocked the migration and invasion of HT-29 and HCT-116 cells. These

  11. An Hydroalcoholic Chamomile Extract Modulates Inflammatory and Immune Response in HT29 Cells and Isolated Rat Colon.

    Science.gov (United States)

    Menghini, Luigi; Ferrante, Claudio; Leporini, Lidia; Recinella, Lucia; Chiavaroli, Annalisa; Leone, Sheila; Pintore, Giorgio; Vacca, Michele; Orlando, Giustino; Brunetti, Luigi

    2016-09-01

    Inflammatory bowel diseases (IBDs) are chronic disorders characterized by disruption and ulceration of the colonic mucosa or of any part of the digestive tract (Crohn's disease). Antioxidant/anti-inflammatory herbal extract supplementation could represent an innovative approach to contrast IBDs. Clinical trials demonstrated the efficacy of natural formulas, containing chamomile, in patients with gastrointestinal disorders. This is consistent, albeit in part, with the antioxidant and anti-inflammatory properties of chamomile. The aim of the present study was to explore the possible protective role of a chamomile extract, on human colorectal adenocarcinoma HT29 cell, and rat colon specimens treated with lipopolysaccharide (LPS) to induce an inflammatory stimulus, a well established model of acute ulcerative colitis. In this context, the activities of different biomarkers of inflammation and lipid peroxidation such as ROS, myeloperoxidase (MPO), serotonin (5-HT), prostaglandin (PG)E2 , 8-iso-prostaglandin (8-iso-PG)F2α , NF-kB, tumor necrosis factor (TNF)α and interleukin (IL)-6 were assessed. We found that chamomile extract was as effective as sulfasalazine (2 mg/ml) in reducing the production of MPO, 5-HT, IL-6, NF-kB, TNFα, PGE2 and 8-iso-PGF2α , after inflammatory stimulus. The observed modulatory effects support a rationale use of chamomile supplementation as a promising pharmacological tool for the prevention and management of ulcerative colitis in humans. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  12. Luffa echinata Roxb. Induces Human Colon Cancer Cell (HT-29 Death by Triggering the Mitochondrial Apoptosis Pathway

    Directory of Open Access Journals (Sweden)

    Yan Yu

    2012-05-01

    Full Text Available The antiproliferative properties and cell death mechanism induced by the extract of the fruits of Luffa echinata Roxb. (LER were investigated. The methanolic extract of LER inhibited the proliferation of human colon cancer cells (HT-29 in both dose-dependent and time-dependent manners and caused a significant increase in the population of apoptotic cells. In addition, obvious shrinkage and destruction of the monolayer were observed in LER-treated cells, but not in untreated cells. Analysis of the cell cycle after treatment of HT-29 cells with various concentrations indicated that LER extracts inhibited the cellular proliferation of HT-29 cells via G2/M phase arrest of the cell cycle. The Reactive oxygen species (ROS level determination revealed that LER extracts induced apoptotic cell death via ROS generation. In addition, LER treatment led to a rapid drop in mitochondrial membrane potential (MMP as a decrease in fluorescence. The transcripts of several apoptosis-related genes were investigated by RT-PCR analysis. The caspase-3 transcripts of HT-29 cells significantly accumulated and the level of Bcl-XL mRNA was decreased after treatment with LER extract. Furthermore, the ratio of mitochondria-dependent apoptosis genes (Bax and Bcl-2 was sharply increased from 1.6 to 54.1. These experiments suggest that LER has anticancer properties via inducing the apoptosis in colon cancer cells, which provided the impetus for further studies on the therapeutic potential of LER against human colon carcinoma.

  13. Green synthesis of NiO nanoparticles using Moringa oleifera extract and their biomedical applications: Cytotoxicity effect of nanoparticles against HT-29 cancer cells.

    Science.gov (United States)

    Ezhilarasi, A Angel; Vijaya, J Judith; Kaviyarasu, K; Maaza, M; Ayeshamariam, A; Kennedy, L John

    2016-11-01

    Green protocols for the synthesis of nickel oxide nanoparticles using Moringa oleifera plant extract has been reported in the present study as they are cost effective and ecofriendly, moreover this paper records that the nickel oxide (NiO) nanoparticles prepared from green method shows better cytotoxicity and antibacterial activity. The NiO nanoparticles were characterized by X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), High resolution transmission electron microscopy (HRTEM), Energy dispersive X-ray analysis (EDX), and Photoluminescence spectroscopy (PL). The formation of a pure nickel oxide phase was confirmed by XRD and FTIR. The synthesized NiO nanoparticles was single crystalline having face centered cubic phase and has two intense photoluminescence emissions at 305.46nm and 410nm. The formation of nano- and micro-structures was confirmed by HRTEM. The in-vitro cytotoxicity and cell viability of human cancer cell HT-29 (Colon Carcinoma cell lines) and antibacterial studies against various bacterial strains were studied with various concentrations of nickel oxide nanoparticles prepared from Moringa oleifera plant extract. MTT assay measurements on cell viability and morphological studies proved that the synthesized NiO nanoparticles posses cytotoxic activity against human cancer cells and the various zones of inhibition (mm), obtained revealed the effective antibacterial activity of NiO nanoparticles against various Gram positive and Gram negative bacterial pathogens. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. TRAIL and docosahexaenoic acid cooperate to induce HT-29 colon cancer cell death

    Czech Academy of Sciences Publication Activity Database

    Vaculová, Alena; Hofmanová, Jiřina; Anděra, Ladislav; Kozubík, Alois

    2005-01-01

    Roč. 229, č. 1 (2005), s. 43-48 ISSN 0304-3835 R&D Projects: GA ČR(CZ) GA524/04/0895; GA ČR(CZ) GA524/03/0766 Institutional research plan: CEZ:AV0Z50040507 Keywords : TRAIL * DHA * cell death Subject RIV: BO - Biophysics Impact factor: 3.049, year: 2005

  15. Response of HT29 cells to butyrate treatment depends on time of exposure and glucose deprivation

    Czech Academy of Sciences Publication Activity Database

    Kučerová, Dana; Štokrová, Jitka; Korb, Jan; Šloncová, Eva; Tuháčková, Zdena; Sovová, Vlasta

    2002-01-01

    Roč. 10, č. 6 (2002), s. 779-784 ISSN 1107-3756 Institutional research plan: CEZ:AV0Z5052915 Keywords : colorectal carcinoma cells, butyrate treatment,glucose deprivation Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.063, year: 2002

  16. Oral and Fecal Campylobacter concisus Strains induce Barrier dysfunction by Apoptosis in HT-29/B6 Intestinal Epithelial Cells

    DEFF Research Database (Denmark)

    Nielsen, Hans Linde; Nielsen, Henrik Ib; Ejlertsen, Tove

    in Ussing chambers. Tight junction (TJ) protein expression was determined by Western blotting, and subcellular TJ distribution was analyzed by confocal laser-scanning microscopy. Apoptosis induction was examined by TUNEL-staining and Western blot of caspase-3 activation. All strains invaded confluent HT-29...

  17. LA-12 overcomes confluence-dependent resistance of HT-29 colon cancer cells to Pt (II) compounds

    Czech Academy of Sciences Publication Activity Database

    Šindlerová, Lenka; Foltinová, V.; Vaculová, Alena; Horváth, Viktor; Souček, Karel; Sova, P.; Hofmanová, Jiřina; Kozubík, Alois

    2010-01-01

    Roč. 30, č. 4 (2010), s. 1183-1188 ISSN 0250-7005 R&D Projects: GA ČR(CZ) GA301/07/1557 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : LA-12 * resistance * HT-29 Subject RIV: BO - Biophysics Impact factor: 1.656, year: 2010

  18. γ-Tocotrienol and 6-Gingerol in Combination Synergistically Induce Cytotoxicity and Apoptosis in HT-29 and SW837 Human Colorectal Cancer Cells

    Directory of Open Access Journals (Sweden)

    Khairunnisa' Md Yusof

    2015-06-01

    Full Text Available Numerous bioactive compounds have cytotoxic properties towards cancer cells. However, most studies have used single compounds when bioactives may target different pathways and exert greater cytotoxic effects when used in combination. Therefore, the objective of this study was to determine the anti-proliferative effect of γ-tocotrienol (γ-T3 and 6-gingerol (6G in combination by evaluating apoptosis and active caspase-3 in HT-29 and SW837 colorectal cancer cells. MTS assays were performed to determine the anti-proliferative and cytotoxicity effect of γ-T3 (0–150 µg/mL and 6G (0–300 µg/mL on the cells. The half maximal inhibitory concentration (IC50 value of 6G+ γ-T3 for HT-29 was 105 + 67 µg/mL and for SW837 it was 70 + 20 µg/mL. Apoptosis, active caspase-3 and annexin V FITC assays were performed after 24 h of treatment using flow cytometry. These bioactives in combination showed synergistic effect on HT-29 (CI: 0.89 ± 0.02, and SW837 (CI: 0.79 ± 0.10 apoptosis was increased by 21.2% in HT-29 and 55.4% in SW837 (p < 0.05 after 24 h treatment, while normal hepatic WRL-68 cells were unaffected. Increased apoptosis by the combined treatments was also observed morphologically, with effects like cell shrinkage and pyknosis. In conclusion, although further studies need to be done, γ-T3 and 6G when used in combination act synergistically increasing cytotoxicity and apoptosis in cancer cells.

  19. Effect of low frequency magnetic fields on the growth of MNP-treated HT29 colon cancer cells

    Science.gov (United States)

    Spyridopoulou, K.; Makridis, A.; Maniotis, N.; Karypidou, N.; Myrovali, E.; Samaras, T.; Angelakeris, M.; Chlichlia, K.; Kalogirou, O.

    2018-04-01

    Recent investigations have attempted to understand and exploit the impact of magnetic field-actuated internalized magnetic nanoparticles (MNPs) on the proliferation rate of cancer cells. Due to the complexity of the parameters governing magnetic field-exposure though, individual studies to date have raised contradictory results. In our approach we performed a comparative analysis of key parameters related to the cell exposure of cancer cells to magnetic field-actuated MNPs, and to the magnetic field, in order to better understand the factors affecting cellular responses to magnetic field-stimulated MNPs. We used magnetite MNPs with a hydrodynamic diameter of 100 nm and studied the proliferation rate of MNPs-treated versus untreated HT29 human colon cancer cells, exposed to either static or alternating low frequency magnetic fields with varying intensity (40-200 mT), frequency (0-8 Hz) and field gradient. All three parameters, field intensity, frequency, and field gradient affected the growth rate of cells, with or without internalized MNPs, as compared to control MNPs-untreated and magnetic field-untreated cells. We observed that the growth inhibitory effects induced by static and rotating magnetic fields were enhanced by pre-treating the cells with MNPs, while the growth promoting effects observed in alternating field-treated cells were weakened by MNPs. Compared to static, rotating magnetic fields of the same intensity induced a similar extend of cell growth inhibition, while alternating fields of varying intensity (70 or 100 mT) and frequency (0, 4 or 8 Hz) induced cell proliferation in a frequency-dependent manner. These results, highlighting the diverse effects of mode, intensity, and frequency of the magnetic field on cell growth, indicate that consistent and reproducible results can be achieved by controlling the complexity of the exposure of biological samples to MNPs and external magnetic fields, through monitoring crucial experimental parameters. We

  20. Microbiota source impact in vitro metabolite colonic production and anti-proliferative effect of spent coffee grounds on human colon cancer cells (HT-29).

    Science.gov (United States)

    Hernández-Arriaga, Angélica María; Dave Oomah, B; Campos-Vega, Rocio

    2017-07-01

    Human gut flora-mediated non-digestible fraction of spent coffee grounds (hgf-NDSCG) was evaluated for its chemopreventive effect and molecular mechanisms involved on human colon adenocarcinoma HT-29 cell survival using two different microbiota source [lean (L) and overweight (OW)]. The source of human gut flora (hgf) (L or OW) affected the pH of hgf-NDSCG only minimally, but linearly reduced those of hgf-inulin. The variability between lean and overweight microbiota was characterized by the metabolism and/or bioaccessibility of different phenolic metabolites, their intermediate and end products as well as by variable time courses. Apoptosis of colon cancer HT-29 cells depended on the microbiota source with the lean microbiota expressing a low lethal concentration 50 (LC 50 /L-hgf-NDSCG=13.5%). We demonstrate that NDSCG and its colonic metabolite from lean microbiota induced HT-29 cell apoptosis by reducing catalase and 8-iso-prostaglandin F2α as biomarkers of in vivo oxidative stress as the primary mechanism underlying its overall chemoprotection against colon cancer. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Inducing G2/M Cell Cycle Arrest and Apoptosis through Generation Reactive Oxygen Species (ROS)-Mediated Mitochondria Pathway in HT-29 Cells by Dentatin (DEN) and Dentatin Incorporated in Hydroxypropyl-β-Cyclodextrin (DEN-HPβCD).

    Science.gov (United States)

    Ashwaq, Al-Abboodi Shakir; Al-Qubaisi, Mothanna Sadiq; Rasedee, Abdullah; Abdul, Ahmad Bustamam; Taufiq-Yap, Yun Hin; Yeap, Swee Keong

    2016-10-18

    Dentatin (DEN), purified from the roots of Clausena excavata Burm f., has poor aqueous solubility that reduces its therapeutic application. The aim of this study was to assess the effects of DEN-HPβCD (hydroxypropyl-β-cyclodextrin) complex as an anticancer agent in HT29 cancer cell line and compare with a crystal DEN in dimethyl sulfoxide (DMSO). The exposure of the cancer cells to DEN or DEN-HPβCD complex leads to cell growth inhibition as determined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. To analyze the mechanism, in which DEN or DEN-HPβCD complex causes the death in human colon HT29 cancer cells, was evaluated by the enzyme-linked immunosorbent assay (ELIZA)-based assays for caspase-3, 8, 9, and reactive oxygen species (ROS). The findings showed that an anti-proliferative effect of DEN or DEN-HPβCD complex were via cell cycle arrest at the G2/M phase and eventually induced apoptosis through both mitochondrial and extrinsic pathways. The down-regulation of poly(ADP-ribose) polymerase (PARP) which leaded to apoptosis upon treatment, was investigated by Western-blotting. Hence, complexation between DEN and HPβCD did not diminish or eliminate the effective properties of DEN as anticancer agent. Therefore, it would be possible to resolve the conventional and current issues associated with the development and commercialization of antineoplastic agents in the future.

  2. Cytotoxic effects of Urtica dioica radix on human colon (HT29) and gastric (MKN45) cancer cells mediated through oxidative and apoptotic mechanisms.

    Science.gov (United States)

    Ghasemi, S; Moradzadeh, M; Mousavi, S H; Sadeghnia, H R

    2016-10-15

    Defects in the apoptotic pathways are responsible for both the colorectal cancer pathogenesis and resistance to therapy. In this study, we examined the level of cellular oxidants, cytotoxicity and apoptosis induced by hydroalcoholic extract of U. dioica radix (0-2000 µg/mL) and oxaliplatin (0-1000 µg/mL, as positive control) in human gastric (MKN45) and colon (HT29) cancer, as well as normal human foreskin fibroblast (HFF) cells. Exposure to U. dioica or oxaliplatin showed a concentration dependent suppression in cell survival with IC50 values of 24.7, 249.9 and 857.5 µg/mL for HT29, MKN45 and HFF cells after 72 h treatment, respectively. ROS formation and lipid peroxidation were also concentration-dependently increased following treatment with U. dioica, similar to oxaliplatin. In addition, the number of apoptotic cells significantly increased concomitantly with concentration of U. dioica as compared with control cells, which is similar to oxaliplatin and serum-deprived cancer cells. In conclusion, the present study demonstrated that U. dioica inhibited proliferation of gastric and colorectal cancer cells while posing no significant toxic effect on normal cells. U. dioica not only increased levels of oxidants, but also induced concomitant increase of apoptosis. The precise signaling pathway by which U. dioica induce apoptosis needs further research.

  3. Carcino-embryonic antigen in monitoring the growth of human colon adenocarcinoma tumour cells SK-CO-1 and HT-29 in vitro and in nude mice

    DEFF Research Database (Denmark)

    Sölétormos, G; Fogh, J M; Sehested-Hansen, B

    1997-01-01

    . In conclusion, our results suggest that experimental models may be useful for investigating the role of serological markers as monitors of increasing tumour burden. It will be of interest to investigate the performance of those model systems in examining the effect of cytotoxic agents in neoplastic growth.......GH). We found that increasing CEA concentration depended on tumour burden. SK-CO-1 cells had the lowest growth rates but the highest rates of CEA production. The rate of CEA increase exceeded the growth rate of both SK-CO-1 and HT-29. hGH modulated neither neoplastic growth nor CEA production...

  4. Poly-γ-Glutamic Acid Induces Apoptosis via Reduction of COX-2 Expression in TPA-Induced HT-29 Human Colorectal Cancer Cells

    Directory of Open Access Journals (Sweden)

    Eun Ju Shin

    2015-04-01

    Full Text Available Poly-γ-glutamic acid (PGA is one of the bioactive compounds found in cheonggukjang, a fast-fermented soybean paste widely utilized in Korean cooking. PGA is reported to have a number of beneficial health effects, and interestingly, it has been identified as a possible anti-cancer compound through its ability to promote apoptosis in cancer cells, although the precise molecular mechanisms remain unclear. Our findings demonstrate that PGA inhibits the pro-proliferative functions of the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA, a known chemical carcinogen in HT-29 human colorectal cancer cells. This inhibition was accompanied by hallmark apoptotic phenotypes, including DNA fragmentation and the cleavage of poly (ADP-ribose polymerase (PARP and caspase 3. In addition, PGA treatment reduced the expression of genes known to be overexpressed in colorectal cancer cells, including cyclooxygenase 2 (COX-2 and inducible nitric oxide synthase (iNOS. Lastly, PGA promoted activation of 5' adenosine monophosphate-activated protein (AMPK in HT-29 cells. Taken together, our results suggest that PGA treatment enhances apoptosis in colorectal cancer cells, in part by modulating the activity of the COX-2 and AMPK signaling pathways. These anti-cancer functions of PGA make it a promising compound for future study.

  5. Discerning apical and basolateral properties of HT-29/B6 and IPEC-J2 cell layers by impedance spectroscopy, mathematical modeling and machine learning.

    Directory of Open Access Journals (Sweden)

    Thomas Schmid

    Full Text Available Quantifying changes in partial resistances of epithelial barriers in vitro is a challenging and time-consuming task in physiology and pathophysiology. Here, we demonstrate that electrical properties of epithelial barriers can be estimated reliably by combining impedance spectroscopy measurements, mathematical modeling and machine learning algorithms. Conventional impedance spectroscopy is often used to estimate epithelial capacitance as well as epithelial and subepithelial resistance. Based on this, the more refined two-path impedance spectroscopy makes it possible to further distinguish transcellular and paracellular resistances. In a next step, transcellular properties may be further divided into their apical and basolateral components. The accuracy of these derived values, however, strongly depends on the accuracy of the initial estimates. To obtain adequate accuracy in estimating subepithelial and epithelial resistance, artificial neural networks were trained to estimate these parameters from model impedance spectra. Spectra that reflect behavior of either HT-29/B6 or IPEC-J2 cells as well as the data scatter intrinsic to the used experimental setup were created computationally. To prove the proposed approach, reliability of the estimations was assessed with both modeled and measured impedance spectra. Transcellular and paracellular resistances obtained by such neural network-enhanced two-path impedance spectroscopy are shown to be sufficiently reliable to derive the underlying apical and basolateral resistances and capacitances. As an exemplary perturbation of pathophysiological importance, the effect of forskolin on the apical resistance of HT-29/B6 cells was quantified.

  6. Role of Caveolin 1, E-Cadherin, Enolase 2 and PKCalpha on resistance to methotrexate in human HT29 colon cancer cells

    DEFF Research Database (Denmark)

    Selga, Elisabet; Morales Torres, Christina; Noé, Véronique

    2008-01-01

    ABSTRACT: BACKGROUND: Methotrexate is one of the earliest cytotoxic drugs used in cancer therapy, and despite the isolation of multiple other folate antagonists, methotrexate maintains its significant role as a treatment for different types of cancer and other disorders. The usefulness of treatment...... validations were performed either by siRNAs or by transfection of an expression plasmid. RESULTS: Genes adjacent to the dhfr locus and included in the 5q14 amplicon were overexpressed in HT29 MTX-resistant cells. Treatment with siRNAs against those genes caused a slight reduction in cell viability in both HT...... viability and caused a decreased MTX resistance capacity. Moreover, overexpression of E-cadherin, which was found underexpressed in MTX-resistant cells, also sensitized the cells toward the chemotherapeutic agent. Combined treatments targeting siRNA inhibition of caveolin 1 and overexpression of E...

  7. Recombinant Lactococcus lactis NZ9000 secretes a bioactive kisspeptin that inhibits proliferation and migration of human colon carcinoma HT-29 cells.

    Science.gov (United States)

    Zhang, Bo; Li, Angdi; Zuo, Fanglei; Yu, Rui; Zeng, Zhu; Ma, Huiqin; Chen, Shangwu

    2016-06-10

    Proteinaceous bioactive substances and pharmaceuticals are most conveniently administered orally. However, the facing problems are the side effects of proteolytic degradation and denaturation in the gastrointestinal tract. In recent years, lactic acid bacteria (LAB) have been verified to be a promising delivery vector for susceptible functional proteins and drugs. KiSS-1 peptide, a cancer suppressor, plays a critical role in inhibiting cancer metastasis and its activity has been confirmed by direct administration. However, whether this peptide can be functionally expressed in LAB and exert activity on cancer cells, thus providing a potential alternative administration manner in the future, has not been demonstrated. A recombinant Lactococcus lactis strain NZ9000-401-kiss1 harboring a plasmid containing the gene of the tumor metastasis-inhibiting peptide KiSS1 was constructed. After optimization of the nisin induction conditions, the recombinant strain efficiently secreted KiSS1 with a maximum detectable amount of 27.9 μg/ml in Dulbecco's Modified Eagle medium. The 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide and would healing assays, respectively, indicated that the secreted KiSS1 peptide remarkably inhibited HT-29 cell proliferation and migration. Furthermore, the expressed KiSS1 was shown to induce HT-29 cell morphological changes, apoptosis and reduce the expression of matrix metalloproteinase 9 (MMP-9) at both mRNA and protein levels. A recombinant L. lactis NZ9000-401-kiss1 successfully expressing the human kiss1 was constructed. The secreted KiSS1 peptide inhibited human HT-29 cells' proliferation and migration probably by invoking, or mediating the cell-apoptosis pathway and by down regulating MMP-9 expression, respectively. Our results suggest that L. lactis is an ideal cell factory for secretory expression of tumor metastasis-inhibiting peptide KiSS1, and the KiSS1-producing L. lactis strain may serve as a new tool for cancer therapy in

  8. Gc, gc-ms analysis of lipophilic fractions of aerial parts of fagonia indica burm.f. showing growth inhibitory effect on ht 29 colorectal cancer cells

    International Nuclear Information System (INIS)

    Farheen, R.; Mahmood, I.

    2016-01-01

    Fagonia indica Burm.f. is a small genus of herbs and under shrubs. The plant contains potentially active substances and has been used traditionally for the treatment of many illnesses including cancer. Many polar compounds have been reported from this plant but its non-polar constituents have only been rarely studied. In the present studies these constituents of aerial parts of Fagonia indica Burm.f. and its sub fractions showing growth inhibitory effect on HT 29 colorectal cancer cells were analyzed using flame ionization detector (GC-FID) and GC-EIMS analysis. The present studies exhibited the presence of free fatty acids and their esters along with structurally diverse constituents including triterpene, heterocyclic organic compound, aromatics, hydrocarbons, alcohols, lactone and sterols which may be responsible for this activity. The results suggest that the non-polar constituents of F. indica bear a potential of further studies. (author)

  9. Screening of Cytotoxic B. cereus on Differentiated Caco-2 Cells and in Co-Culture with Mucus-Secreting (HT29-MTX) Cells

    Science.gov (United States)

    Castiaux, Virginie; Laloux, Laurie; Schneider, Yves-Jacques; Mahillon, Jacques

    2016-01-01

    B. cereus is an opportunistic foodborne pathogen able to cause diarrhoea. However, the diarrhoeal potential of a B. cereus strain remains difficult to predict, because no simple correlation has yet been identified between the symptoms and a unique or a specific combination of virulence factors. In this study, 70 B. cereus strains with different origins (food poisonings, foods and environment) have been selected to assess their enterotoxicity. The B. cereus cell-free supernatants have been tested for their toxicity in vitro, on differentiated (21 day-old) Caco-2 cells, using their ATP content, LDH release and NR accumulation. The genetic determinants of the main potential enterotoxins and virulence factors (ces, cytK, entFM, entS, hbl, nhe, nprA, piplC and sph) have also been screened by PCR. This analysis showed that none of these genes was able to fully explain the enterotoxicity of B. cereus strains. Additionally, in order to assess a possible effect of the mucus layer in vitro, a cytotoxicity comparison between a monoculture (Caco-2 cells) and a co-culture (Caco-2 and HT29-MTX mucus-secreting cells) model has been performed with selected B. cereus supernatants. It appeared that, in these conditions, the mucus layer had no notable influence on the cytotoxicity of B. cereus supernatants. PMID:27827957

  10. The chemopotential effect of Annona muricata leaves against azoxymethane-induced colonic aberrant crypt foci in rats and the apoptotic effect of Acetogenin Annomuricin E in HT-29 cells: a bioassay-guided approach.

    Science.gov (United States)

    Zorofchian Moghadamtousi, Soheil; Rouhollahi, Elham; Karimian, Hamed; Fadaeinasab, Mehran; Firoozinia, Mohammad; Ameen Abdulla, Mahmood; Abdul Kadir, Habsah

    2015-01-01

    Annona muricata has been used in folk medicine for the treatment of cancer and tumors. This study evaluated the chemopreventive properties of an ethyl acetate extract of A. muricata leaves (EEAML) on azoxymethane-induced colonic aberrant crypt foci (ACF) in rats. Moreover, the cytotoxic compound of EEAML (Annomuricin E) was isolated, and its apoptosis-inducing effect was investigated against HT-29 colon cancer cell line using a bioassay-guided approach. This experiment was performed on five groups of rats: negative control, cancer control, EEAML (250 mg/kg), EEAML (500 mg/kg) and positive control (5-fluorouracil). Methylene blue staining of colorectal specimens showed that application of EEAML at both doses significantly reduced the colonic ACF formation compared with the cancer control group. Immunohistochemistry analysis showed the down-regulation of PCNA and Bcl-2 proteins and the up-regulation of Bax protein after administration of EEAML compared with the cancer control group. In addition, an increase in the levels of enzymatic antioxidants and a decrease in the malondialdehyde level of the colon tissue homogenates were observed, suggesting the suppression of lipid peroxidation. Annomuricin E inhibited the growth of HT-29 cells with an IC50 value of 1.62 ± 0.24 μg/ml after 48 h. The cytotoxic effect of annomuricin E was further substantiated by G1 cell cycle arrest and early apoptosis induction in HT-29 cells. Annomuricin E triggered mitochondria-initiated events, including the dissipation of the mitochondrial membrane potential and the leakage of cytochrome c from the mitochondria. Prior to these events, annomuricin E activated caspase 3/7 and caspase 9. Upstream, annomuricin E induced a time-dependent upregulation of Bax and downregulation of Bcl-2 at the mRNA and protein levels. In conclusion, these findings substantiate the usage of A. muricata leaves in ethnomedicine against cancer and highlight annomuricin E as one of the contributing compounds in the

  11. The chemopotential effect of Annona muricata leaves against azoxymethane-induced colonic aberrant crypt foci in rats and the apoptotic effect of Acetogenin Annomuricin E in HT-29 cells: a bioassay-guided approach.

    Directory of Open Access Journals (Sweden)

    Soheil Zorofchian Moghadamtousi

    Full Text Available Annona muricata has been used in folk medicine for the treatment of cancer and tumors. This study evaluated the chemopreventive properties of an ethyl acetate extract of A. muricata leaves (EEAML on azoxymethane-induced colonic aberrant crypt foci (ACF in rats. Moreover, the cytotoxic compound of EEAML (Annomuricin E was isolated, and its apoptosis-inducing effect was investigated against HT-29 colon cancer cell line using a bioassay-guided approach. This experiment was performed on five groups of rats: negative control, cancer control, EEAML (250 mg/kg, EEAML (500 mg/kg and positive control (5-fluorouracil. Methylene blue staining of colorectal specimens showed that application of EEAML at both doses significantly reduced the colonic ACF formation compared with the cancer control group. Immunohistochemistry analysis showed the down-regulation of PCNA and Bcl-2 proteins and the up-regulation of Bax protein after administration of EEAML compared with the cancer control group. In addition, an increase in the levels of enzymatic antioxidants and a decrease in the malondialdehyde level of the colon tissue homogenates were observed, suggesting the suppression of lipid peroxidation. Annomuricin E inhibited the growth of HT-29 cells with an IC50 value of 1.62 ± 0.24 μg/ml after 48 h. The cytotoxic effect of annomuricin E was further substantiated by G1 cell cycle arrest and early apoptosis induction in HT-29 cells. Annomuricin E triggered mitochondria-initiated events, including the dissipation of the mitochondrial membrane potential and the leakage of cytochrome c from the mitochondria. Prior to these events, annomuricin E activated caspase 3/7 and caspase 9. Upstream, annomuricin E induced a time-dependent upregulation of Bax and downregulation of Bcl-2 at the mRNA and protein levels. In conclusion, these findings substantiate the usage of A. muricata leaves in ethnomedicine against cancer and highlight annomuricin E as one of the contributing

  12. Extraction of Peptidoglycan from L. paracasei subp. Paracasei X12 and Its Preliminary Mechanisms of Inducing Immunogenic Cell Death in HT-29 Cells

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    Pei-Jun Tian

    2015-08-01

    Full Text Available L. paracasei subp. paracasei X12 was previously isolated from a Chinese traditional fermented cheese with anticancer activities and probiotic potential. Herein, the integral peptidoglycan (X12-PG was extracted by a modified trichloroacetic acid (TCA method. X12-PG contained the four representative amino acids Asp, Glu, Ala and Lys, and displayed the similar lysozyme sensitivity, UV-visible scanning spectrum and molecular weight as the peptidoglycan standard. X12-PG could induce the production of apoptotic bodies observed by transmission electron microscopy (TEM. X12-PG could significantly induced the translocation of calreticulin (CRT and the release of high mobility group box 1 protein (HMGB1, the two notable hallmarks of immunogenic cell death (ICD, with the endoplastic reticulum (ER damaged and subsequently intracellular [Ca2+] elevated. Our findings implied that X12-PG could induce the ICD of HT-29 cells through targeting at the ER. The present results may enlighten the prospect of probiotics in the prevention of colon cancer.

  13. Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins.

    Science.gov (United States)

    Phang, Chung-Weng; Karsani, Saiful Anuar; Abd Malek, Sri Nurestri

    2017-07-01

    Chalcones have been shown to exhibit anti-cancer properties by targeting multiple molecular pathways. It was, therefore, of interest to investigate flavokawain C (FKC), a naturally occurring chalcone, which can be isolated from Kava ( Piper methysticum Forst) root extract. The aim of this study was to investigate the inhibitory effect of FKC on the growth of HT-29 cells and its underlying mechanism of action. Cell viability of HT-29 cells was assessed by Sulforhodamine B assay after FKC treatment. Induction of apoptosis was examined by established morphological and biochemical assays. ROS generation was determined by dichlorofluorescein fluorescence staining, and superoxide dismutase activity was measured using the spectrophotometric method. Western blotting was used to examine the changes in the protein levels. FKC markedly decreased the cell viability of HT-29 cells and the cells showed dramatic changes in cellular and nuclear morphologies with typical apoptotic features. The induction of apoptosis correlated well with the externalization of phosphatidylserine, DNA fragmentation, decreased mitochondrial membrane potential, activation of caspases, and PARP cleavage. This was associated with an increase in reactive oxygen species and a decrease in SOD activity. The protein levels of XIAP, c-IAP1, and c-IAP2 were downregulated, whereas the GADD153 was upregulated after FKC treatment. FKC induced cell cycle arrest at the G 1 and G 2 /M phases via upregulation of p21 and p27 in a p53-independent manner. Our results provide evidence that FKC has the potential to be developed into chemotherapeutic drug for the treatment of colon adenocarcinoma. Flavokawain C inhibited the growth of HT-29 human colon adenocarcinoma cellsFlavokawain C induced apoptosis in HT-29 cells, associated with an increase in reactive oxygen species and a decrease in SOD activityFlavokawain C induced cell cycle arrest at the G 1 and G 2 /M phases via upregulation of p21 and p27 in HT-29 cellsHT-29

  14. Cordyceps militaris Grown on Germinated Soybean Induces G2/M Cell Cycle Arrest through Downregulation of Cyclin B1 and Cdc25c in Human Colon Cancer HT-29 Cells

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    Mohammad Lalmoddin Mollah

    2012-01-01

    Full Text Available Cordyceps militaris (CM is an insect-borne fungus that has been used in traditional Chinese medicine because of its wide range of pharmacological activities. In this paper, we studied CM grown on germinated soybean (GSC and investigated the possible mechanisms underlying antiproliferative effect of GSC on HT-29 human colon cancer cells. In comparison with CM extracts and germinated soybean (GS BuOH extracts, BuOH extracts of GSC showed remarkable inhibitory and antiproliferative effects on HT-29 colon cancer cells. After GSC treatment, HT-29 cells became smaller and irregular in shape. High G2/M phase cell populations were observed in the GSC-treated group. The levels of cyclin B1 and Cdc25 in the GSC-treated group were lower than those in the control group. These findings suggest that GSC BuOH extracts might act as an effective anti-proliferative agent by inducing G2/M cell cycle arrest in colon cancer cells.

  15. Protein expression profile of HT-29 human colon cancer cells after treatment with a cytotoxic daunorubicin-GnRH-III derivative bioconjugate.

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    Verena Natalie Schreier

    Full Text Available Targeted delivery of chemotherapeutic agents is a new approach for the treatment of cancer, which provides increased selectivity and decreased systemic toxicity. We have recently developed a promising drug delivery system, in which the anticancer drug daunorubicin (Dau was attached via oxime bond to a gonadotropin-releasing hormone-III (GnRH-III derivative used as a targeting moiety (Glp-His-Trp-Lys(Ac-His-Asp-Trp-Lys(Da  = Aoa-Pro-Gly-NH2; Glp = pyroglutamic acid, Ac = acetyl; Aoa = aminooxyacetyl. This bioconjugate exerted in vitro cytostatic/cytotoxic effect on human breast, prostate and colon cancer cells, as well as significant in vivo tumor growth inhibitory effect on colon carcinoma bearing mice. In our previous studies, H-Lys(Dau = Aoa-OH was identified as the smallest metabolite produced in the presence of rat liver lysosomal homogenate, which was able to bind to DNA in vitro. To get a deeper insight into the mechanism of action of the bioconjugate, changes in the protein expression profile of HT-29 human colon cancer cells after treatment with the bioconjugate or free daunorubicin were investigated by mass spectrometry-based proteomics. Our results indicate that several metabolism-related proteins, molecular chaperons and proteins involved in signaling are differently expressed after targeted chemotherapeutic treatment, leading to the conclusion that the bioconjugate exerts its cytotoxic action by interfering with multiple intracellular processes.

  16. Isolation and Characterization of Lectin from the Artist's Conk Medicinal Mushroom, Ganoderma applanatum (Agaricomycetes), and Evaluation of Its Antiproliferative Activity in HT-29 Colon Cancer Cells.

    Science.gov (United States)

    Kumaran, Sekar; Pandurangan, Ashok Kumar; Shenbhagaraman, Ramalingam; Esa, Norhaizan Mohd

    2017-01-01

    The growth and lectin production of Ganoderma applanatum, a white rot fungus, was optimized in broth cultures. The fungus was found to have a higher growth rate and higher lectin activity when grown in a medium adjusted to pH 6.5 at 26°C under stationary conditions. Expression of lectin activity started in 5-day-old mycelial culture; maximum activity was expressed after the 15th day of incubation. Among the various carbon and nitrogen sources tested, the carbon source sucrose and the nitrogen source yeast extract support maximum growth and lectin production. Lectin from G. applanatum was purified by ammonium sulfate precipitation and ion exchange chromatography. The purified fraction revealed a single band with a molecular weight of 35.0 kDa. Moreover, carbohydrates such as mannitol, glucose, sucrose, maltose, mannose, galactose, sorbose, and fructose were found to inhibit the hemagglutinating activity of the lectin. The purified lectins from G. applanatum contain cytotoxic and proapoptotic activities against HT-29 colon adenocarcinoma cells.

  17. Phenolic composition of selected herbal infusions and their anti-inflammatory effect on a colonic model in vitro in HT-29 cells

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    Elda Herrera-Carrera

    2015-12-01

    Full Text Available Some herbal infusions used in folk medicine in Mexico to treat gastrointestinal disorders were evaluated. Antioxidant activity and phenolic compounds were analyzed on the lyophilized aqueous crude extracts (LACE of arnica (Aster gymnocephalus, chamomile (Chamaemelum nobile, cumin (Cominum cyminum, desert resurrection plant (DRP (Selaginella lepidophylla, laurel (Listea glaucescens, marjoram (Origanum majorana, mint (Mentha spicata, salvilla (Buddleia scordioides and yerbaniz (Tagetes lucida. Total phenolic content ranged from 8.0 to 70.7 μg GAE/mg for DRP and laurel respectively. Major phenolic compounds were identified by gas chromatography–mass spectrometry and high-performance liquid chromatography. The IC50 determined by the degradation of the deoxy-d-ribose ranged from 2,452.53 to 5,097.11 μg/mL. The cytoprotective effect of the LACE alone and on indomethacin-induced oxidative stress in HT-29 cells was tested. The tetrazolium dye MTT assay was performed in concentrations of 0.125–10 mg/mL allowing choosing the lowest concentration for this experimentation. Inflammation markers were measured by Western blotting. None of the extracts inhibited COX-1 by themselves; however, it was observed that extracts have a modulation effect over COX-2, TNFα, NFκB, and IL-8. By the decrease in the expression of pro-inflammatory cytokines, it follows that salvilla, chamomile, and laurel show promising anti-inflammatory effects.

  18. TFF3-dependent resistance of human colorectal adenocarcinoma cells HT-29/B6 to apoptosis is mediated by miR-491-5p regulation of lncRNA PRINS

    OpenAIRE

    Hanisch, Carlos; Sharbati, Jutta; Kutz-Lohroff, Barbara; Huber, Otmar; Einspanier, Ralf; Sharbati, Soroush

    2017-01-01

    Tumour necrosis factor-? (TNF-?) is a double-edged cytokine associated with pathogenesis of inflammatory-related cancers being also able to induce cancer cell death. In the process of tumour development or metastasis, cancer cells can become resistant to TNF-?. In trefoil factor 3 (TFF3) overexpressing colorectal adenocarcinoma cells (HT-29/B6), we observed enhanced resistance against TNF-?/interferon gamma-induced apoptosis. TFF3 is a secreted small peptide that supports intestinal tissue re...

  19. Effect of Growth factors, estradiol 17-ß, and short chain fatty acids on the intestinal HT29-MTX cells

    DEFF Research Database (Denmark)

    Giromini, Carlotta; Baldi, Antonella; Fusi, Eleonora

    2015-01-01

    Peptides growth factors, hormones, and short chain fatty acids (SCFAs) are constantly in contact with the human bowel when secreted by gland or ingested by food, as milk and colostrum, or, as in the case of SCFAs, produced by fermentation processes. This study considers the effect of growth factors...... studies. The effect of insulin-like growth factors (IGF)-I, epidermal growth factors (EGF), transforming growth factor alpha (TGF-α), transforming growth factor beta (TGF-β), estradiol 17-β and butyrate, propionate, and acetate was assessed on metabolic activity and proliferation of E12 cells using Alamar...... of the cells. Further, a dose-dependent inhibition of cell metabolic activity was detected in the presence of all SCFAs tested. Butyrate showed to be the most active in the inhibition of E12 metabolic activity and its effect was enhanced by the presence of propionate and acetate. E12 cells, in undifferentiated...

  20. Comprehensive and Holistic Analysis of HT-29 Colorectal Cancer Cells and Tumor-Bearing Nude Mouse Model: Interactions Among Fractions Derived From the Chinese Medicine Formula Tian Xian Liquid in Effects on Human Colorectal Carcinoma.

    Science.gov (United States)

    Leigh, Annballaw Bridget; Cheung, Ho Pan; Lin, Li-Zhu; Ng, Tzi Bun; Lao, Lixing; Zhang, Yanbo; Zhang, Zhang-Jin; Tong, Yao; Sze, Stephen Cho Wing

    2017-09-01

    The Chinese medicine formula Tian Xian Liquid (TXL) has been used clinically for cancer therapy in China for more than 25 years. However, the comprehensive and holistic effects of its bioactive fractions for various antitumor therapeutic effects have not been unraveled. This is the first study to scientifically elucidate the holistic effect of Chinese medicine formula for treating colon cancer, hence allowing a better understanding of the essence of Chinese medicine formula, through the comparison of the actions of TXL and its functional constituent fractions, including ethyl acetate (EA), butanol (BU), and aqueous (WA) fractions. Tissue-specific proliferative/antiproliferative effects of these fractions on human colorectal carcinoma HT-29 cells and splenocytes were studied by using the MTT assay. Their modulations on the expression of markers of antiproliferation, antimetastasis, reversion of multidrug resistance in treated HT-29 cells were examined with real-time polymerase chain reaction and Western blot analysis, and their modulations in a xenografted nude mouse model were examined by Western blot analysis. Results revealed that EA fraction slightly inhibited the proliferation of HT-29 cells, but tissue-specifically exerted the most potent antiproliferative effect on splenocytes. On the contrary, only TXL and BU fraction tissue-specifically contributed to the proliferation of splenocytes, but inhibited the proliferation of HT-29 cells. WA fraction exerted the most potent antiproliferative effect on HT-29 cells and also the strongest inhibitory action on tumor size in the nude mouse model in our previous study. In the HT-29 model, TXL and WA fraction exerted the most pronounced effect on upregulation of p21 mRNA and protein; TXL, and EA and WA fractions exerted the effect on downregulation of G1 phase cell cycle protein, cyclin D1 mRNA and protein; EA and BU fractions exerted the most prominent anti-invasive effect on anti-invasion via downregulation of MMP-1 m

  1. Reparative properties of the traditional Chinese medicine Cordyceps sinensis (Chinese caterpillar mushroom) using HT29 cell culture and rat gastric damage models of injury.

    Science.gov (United States)

    Marchbank, Tania; Ojobo, Ehighale; Playford, Christopher J; Playford, Raymond J

    2011-05-01

    Cordyceps sinensis (CS) is a traditional Chinese medicine and health food used to support many organ systems. It is commercially produced by cultivation in a liquid medium or on a solid (grain/potato) phase. We tested the effects of hot water extracts of liquid-phase and solid-phase commercially grown CS on its ability to influence proliferation (using Alamar blue, an oxidation/reduction indicator), migration (serial-wounded monolayer photomicroscopy), invasion through collagen gel (fluorometric assay) and indomethacin-induced apoptosis (active caspase-3 colorimetric assay) of human colon cancer HT29 cells. An in vivo study used a rat gastric damage model (indomethacin 20 mg/kg and 4 h restraint with oral administration). The CS extract stimulated cell proliferation threefold when added at 10 μg/ml (P < 0·01). Cell migration increased by 69 % and invasion by 17 % when CS was added at 5 mg/ml (P < 0·01). The results also showed that 93 % of the pro-proliferative activity was soluble in ethanol, whereas pro-migratory activity was divided (61:49) into both ethanol-soluble and ethanol-insoluble sub-fractions. Indomethacin-induced apoptosis was not affected by the presence of CS. CS reduced the amount of gastric injury by 63 % when administered orally at 20 mg/ml (P < 0·01), the results being similar to using the potent cytoprotective agent epidermal growth factor at 25 μg/ml (83 % reduction). We conclude that both methods of cultivated CS possess biological activity when analysed using a variety of gut models of injury and repair. Functional foods, such as CS, could provide a novel approach for the prevention and treatment of injury to the bowel.

  2. A combination of indol-3-carbinol and genistein synergistically induces apoptosis in human colon cancer HT-29 cells by inhibiting Akt phosphorylation and progression of autophagy

    Directory of Open Access Journals (Sweden)

    Watanabe Hirotsuna

    2009-11-01

    Full Text Available Abstract Background The chemopreventive effects of dietary phytochemicals on malignant tumors have been studied extensively because of a relative lack of toxicity. To achieve desirable effects, however, treatment with a single agent mostly requires high doses. Therefore, studies on effective combinations of phytochemicals at relatively low concentrations might contribute to chemopreventive strategies. Results Here we found for the first time that co-treatment with I3C and genistein, derived from cruciferous vegetables and soy, respectively, synergistically suppressed the viability of human colon cancer HT-29 cells at concentrations at which each agent alone was ineffective. The suppression of cell viability was due to the induction of a caspase-dependent apoptosis. Moreover, the combination effectively inhibited phosphorylation of Akt followed by dephosphorylation of caspase-9 or down-regulation of XIAP and survivin, which contribute to the induction of apoptosis. In addition, the co-treatment also enhanced the induction of autophagy mediated by the dephosphorylation of mTOR, one of the downstream targets of Akt, whereas the maturation of autophagosomes was inhibited. These results give rise to the possibility that co-treatment with I3C and genistein induces apoptosis through the simultaneous inhibition of Akt activity and progression of the autophagic process. This possibility was examined using inhibitors of Akt combined with inhibitors of autophagy. The combination effectively induced apoptosis, whereas the Akt inhibitor alone did not. Conclusion Although in vivo study is further required to evaluate physiological efficacies and toxicity of the combination treatment, our findings might provide a new insight into the development of novel combination therapies/chemoprevention against malignant tumors using dietary phytochemicals.

  3. Knockdown of long non-coding RNA prostate cancer-associated ncRNA transcript 1 inhibits multidrug resistance and c-Myc-dependent aggressiveness in colorectal cancer Caco-2 and HT-29 cells.

    Science.gov (United States)

    Qiao, Lei; Liu, Xiangyu; Tang, Yichao; Zhao, Zheng; Zhang, Jilong; Liu, Huayan

    2018-04-01

    The long non-coding RNA (lncRNA) prostate cancer-associated ncRNA transcript 1 (PCAT-1) has been shown to promote prostate cancer cell proliferation through c-Myc and is associated with the poor prognosis of CRC patients. In the current study, it was hypothesized that the effect of PCAT-1 on the aggressiveness of CRC cells was dependent on the function of c-Myc. Human CRC cell lines Caco-2 and HT-29 were transfected with specific PCAT-1 shRNAs, and cell migration, invasiveness, and resistance to 5-fluorouracil were measured. To elucidate the role of c-Myc in PCAT-1 function, c-Myc was overexpressed in PCAT-1-silenced CRC cells and the effect of c-Myc overexpression on the aggressiveness of PCAT-1-silenced cells was detected. The results showed that knockdown of PCAT-1 in CRC cells suppressed cell motility and invasiveness, and sensitized the cells to 5-fluorouracil, as evidenced by the reduced viability and induced apoptosis in PCAT-1-silenced cells compared to the parental cells in response to 5-fluorouracil treatment. The expression of c-Myc in PCAT-1-silenced CRC cells was down-regulated, and forced expression of c-Myc partially restored the invasiveness in PCAT-1-silenced cells. In summary, the findings outlined in the current study suggest that PCAT-1 regulates the invasiveness and drug resistance in CRC cells and that PCAT-1 may promote CRC cell invasion by modulating the expression of c-Myc.

  4. Modulation of mRNA expression and activities of xenobiotic metabolizing enzymes, CYP1A1, CYP1A2, CYP2E1, GPx and GSTP1 by the Salicornia freitagii extract in HT-29 human colon cancer cells

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    Altay Ahmet

    2017-01-01

    Full Text Available Phase I-II detoxification and antioxidant enzymes are responsible for the detoxification and elimination of activated carcinogens, acting as important biomarkers for chemoprevention. Among them, cytochrome P450s plays a prominent role in the metabolic activation of xenobiotics. The herb Salicornia freitagii (SF (Amaranthaceae is known for its anticancer, antioxidant, antidiabetic and antiinflammatory activities. In this study, we determined the bioactive phenolics in the SF methanol extract and investigated its antiproliferative potential in HT-29 human colon cancer cells. We also investigated the modulation of some phase I and II enzyme (CYP 1A1, 1A2, 2E1, GSTP1 and GPx mRNA expression and enzymatic activities by the SF extract and its major bioactive phenolic compounds. LC/MS-MS analysis showed that the main phenolic compounds of the methanolic SF extract are vanillic acid (48 μg/100g and p-coumaric acid (10.8 μg/100g. SF extract, vanillic acid and p-coumaric acid exhibited high antiproliferative activities in HT-29 cells, with IC50 values of 81.79μg/mL, 98.8 μM and 221.6 μM, respectively. The mRNA expression levels of CYP1A2 and CYP2E1 were decreased, while those of GSTP1 and GPx in HT-29 cells were increased after application of either the SF extract or vanillic acid. The SF extract by itself also increased the activities of GPx and GSTP1 enzymes 1.68- and 1.49-fold, respectively. Our data indicate that the SF extract and its major bioactive compound, vanillic acid, could exert a modulatory effect on the expression of enzymes that are involved in xenobiotic activation and detoxification pathways in the gastrointestinal tract. For this reason, SF can be considered as a natural source of chemopreventive agents.

  5. Eupatorin and Salvigenin Potentiate Doxorubicin-Induced Apoptosis and Cell Cycle Arrest in HT-29 and SW948 Human Colon Cancer Cells

    Science.gov (United States)

    Namazi Sarvestani, Nazanin; Sepehri, Houri; Delphi, Ladan; Moridi Farimani, Mahdi

    2018-01-27

    Background: Cancer persists as one of the world’s most pressing maladies. Notable points about chemotherapy are drug side effects which are almost universally encountered. Emerging knowledge focusing on mechanisms of toxicity due to chemotherapy has led to characterization of novel methods, including the exploitation of natural compounds, in combination therapies. Flavonoids are natural polyphenolic compounds that play protective roles against tumor cell development. The focus of this study was apoptotic effects of two flavonoids, eupatorin and salvigenin, in combination with doxorubicin on a cellular model of colon cancer. Method: Upon establishing a non-effective dose of doxorubicin, and effective doses of eupatorin (100μM) and salvigenin (150μM) via MTT, morphological features of apoptosis were distinguished using DAPI staining and cell cycle blockage in the sub-G1 phase. Apoptosis was determined by annexin/ PI and western blotting. ROS levels and MMP were measured to show any role of mitochondria in apoptosis. Results: Co-administration of flavonoids with doxorubicin induced apoptosis via the mitochondrial pathway as mitochondrial membrane potential and ROS production were changed. Annexin/PI analysis demonstrated that apoptosis frequency was increased with the combination treatments in colon cancer cells. Finally, the combination of these flavonoids with doxorubicin increased the Bax/Bcl-2 ratio, caspase-3 expression and PARP cleavage. Conclusion: Combination of flavonoids with doxorubicin induces apoptosis and enhances effect on cancer cells which might allow amelioration of side effects by dose lowering. Creative Commons Attribution License

  6. Parallel comparison of Illumina RNA-Seq and Affymetrix microarray platforms on transcriptomic profiles generated from 5-aza-deoxy-cytidine treated HT-29 colon cancer cells and simulated datasets

    Science.gov (United States)

    2013-01-01

    Background High throughput parallel sequencing, RNA-Seq, has recently emerged as an appealing alternative to microarray in identifying differentially expressed genes (DEG) between biological groups. However, there still exists considerable discrepancy on gene expression measurements and DEG results between the two platforms. The objective of this study was to compare parallel paired-end RNA-Seq and microarray data generated on 5-azadeoxy-cytidine (5-Aza) treated HT-29 colon cancer cells with an additional simulation study. Methods We first performed general correlation analysis comparing gene expression profiles on both platforms. An Errors-In-Variables (EIV) regression model was subsequently applied to assess proportional and fixed biases between the two technologies. Then several existing algorithms, designed for DEG identification in RNA-Seq and microarray data, were applied to compare the cross-platform overlaps with respect to DEG lists, which were further validated using qRT-PCR assays on selected genes. Functional analyses were subsequently conducted using Ingenuity Pathway Analysis (IPA). Results Pearson and Spearman correlation coefficients between the RNA-Seq and microarray data each exceeded 0.80, with 66%~68% overlap of genes on both platforms. The EIV regression model indicated the existence of both fixed and proportional biases between the two platforms. The DESeq and baySeq algorithms (RNA-Seq) and the SAM and eBayes algorithms (microarray) achieved the highest cross-platform overlap rate in DEG results from both experimental and simulated datasets. DESeq method exhibited a better control on the false discovery rate than baySeq on the simulated dataset although it performed slightly inferior to baySeq in the sensitivity test. RNA-Seq and qRT-PCR, but not microarray data, confirmed the expected reversal of SPARC gene suppression after treating HT-29 cells with 5-Aza. Thirty-three IPA canonical pathways were identified by both microarray and RNA

  7. The effects of TNF-alpha and inhibitors of arachidonic acid metabolism on human colon HT-29 cells depend on differentiation status

    Czech Academy of Sciences Publication Activity Database

    Kovaříková, Martina; Hofmanová, Jiřina; Souček, Karel; Kozubík, Alois

    2004-01-01

    Roč. 72, č. 1 (2004), s. 23-31 ISSN 0301-4681 R&D Projects: GA ČR GA525/01/0419; GA ČR GP524/02/P051; GA AV ČR IBS5004009 Institutional research plan: CEZ:AV0Z5004920 Keywords : colon cancer * cell differentiation * TNF-alpha Subject RIV: BO - Biophysics Impact factor: 4.481, year: 2004

  8. TNF-alfa modulates the differentiation induced by butyrate in the HT-29 human colon adenocarcinoma cell line

    Czech Academy of Sciences Publication Activity Database

    Kovaříková, Martina; Pacherník, Jiří; Hofmanová, Jiřina; Zadák, Z.; Kozubík, Alois

    2000-01-01

    Roč. 36, - (2000), s. 1844-1852 ISSN 0959-8049 R&D Projects: GA ČR GA524/98/0190; GA MZd IZ4095; GA MZd NC6171; GA AV ČR KSK2039602 Institutional research plan: CEZ:AV0Z5004920 Subject RIV: BO - Biophysics Impact factor: 2.725, year: 2000

  9. Screening of bifidobacteria and lactobacilli able to antagonise the cytotoxic effect of Clostridium difficile upon intestinal epithelial HT29 monolayer

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    Lorena eValdés-Varela

    2016-04-01

    Full Text Available Clostridium difficile is an opportunistic pathogen inhabiting the human gut, often being the aetiological agent of infections after a microbiota dysbiosis following, for example, an antibiotic treatment. C. difficile infections (CDI constitute a growing health problem with increasing rates of morbidity and mortality at groups of risk, such as elderly and hospitalized patients, but also in populations traditionally considered low-risk. This could be related to the occurrence of virulent strains which, among other factors, have high-level of resistance to fluoroquinolones, more efficient sporulation and markedly high toxin production. Several novel intervention strategies against CDI are currently under study, such as the use of probiotics to counteract the growth and/or toxigenic activity of C. difficile.In this work, we have analysed the capability of twenty Bifidobacterium and Lactobacillus strains, from human intestinal origin, to counteract the toxic effect of C. difficile LMG21717 upon the human intestinal epithelial cell line HT29. For this purpose, we incubated the bacteria together with toxigenic supernatants obtained from C. difficile. After this co-incubation new supernatants were collected in order to quantify the remnant A and B toxins, as well as to determine their residual toxic effect upon HT29 monolayers. To this end, the real time cell analyser (RTCA model, recently developed in our group to monitor C. difficile toxic effect, was used. Results obtained showed that strains of Bifidobacterium longum and Bifidobacterium breve were able to reduce the toxic effect of the pathogen upon HT29, the RTCA normalized cell-index values being inversely correlated with the amount of remnant toxin in the supernatant. The strain B. longum IPLA20022 showed the highest ability to counteract the cytotoxic effect of C. difficile acting directly against the toxin, also having the highest capability for removing the toxins from the clostridial

  10. Probiotics promote endocytic allergen degradation in gut epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Song, Chun-Hua [Department of Epidemiology and Biostatistics, College of Public Health, Zhengzhou University, Zhengzhou (China); Liu, Zhi-Qiang [Department of Gastroenterology, The Second Hospital, Zhengzhou University, Zhengzhou (China); Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada); Huang, Shelly [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada); Zheng, Peng-Yuan, E-mail: medp7123@126.com [Department of Gastroenterology, The Second Hospital, Zhengzhou University, Zhengzhou (China); Yang, Ping-Chang, E-mail: yangp@mcmaster.ca [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada)

    2012-09-14

    Highlights: Black-Right-Pointing-Pointer Knockdown of A20 compromised the epithelial barrier function. Black-Right-Pointing-Pointer The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Black-Right-Pointing-Pointer Antigens transported across A20-deficient HT-29 monolayers conserved antigenicity. Black-Right-Pointing-Pointer Probiotic proteins increased the expression of A20 in HT-29 cells. -- Abstract: Background and aims: Epithelial barrier dysfunction plays a critical role in the pathogenesis of allergic diseases; the mechanism is to be further understood. The ubiquitin E3 ligase A20 (A20) plays a role in the endocytic protein degradation in the cells. This study aims to elucidate the role of A20 in the maintenance of gut epithelial barrier function. Methods: Gut epithelial cell line, HT-29 cell, was cultured into monolayers to evaluate the barrier function in transwells. RNA interference was employed to knock down the A20 gene in HT-29 cells to test the role of A20 in the maintenance of epithelial barrier function. Probiotic derived proteins were extracted from the culture supernatants using to enhance the expression of A20 in HT-29 cells. Results: The results showed that the knockdown of A20 compromised the epithelial barrier function in HT-29 monolayers, mainly increased the intracellular permeability. The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Allergens collected from the transwell basal chambers of A20-deficient HT-29 monolayers still conserved functional antigenicity. Treating with probiotic derived proteins increased the expression of A20 in HT-29 cells and promote the barrier function. Conclusion: A20 plays an important role in the maintenance of epithelial barrier function as shown by HT-29 monolayer. Probiotic derived protein increases the expression of A20 and promote the HT-29 monolayer barrier function.

  11. The 18-kDa translocator protein, formerly known as the peripheral-type benzodiazepine receptor, confers proapoptotic and antineoplastic effects in a human colorectal cancer cell line.

    Science.gov (United States)

    Shoukrun, Rami; Veenman, Leo; Shandalov, Yulia; Leschiner, Svetlana; Spanier, Ilana; Karry, Rachel; Katz, Yeshayahu; Weisinger, Gary; Weizman, Abraham; Gavish, Moshe

    2008-11-01

    The involvement of the 18-kDa translocator protein (TSPO), formerly known as the peripheral-type benzodiazepine receptor, in apoptosis regulation of HT29 colorectal cancer cells was studied in-vitro. In-vivo TSPO involvement in tumor growth of HT29 cells xenografted into SCID mice was studied. Knockdown of TSPO expression in the human HT29 cell line was established by stable transfection with vectors containing the TSPO gene in the antisense direction. Successful TSPO knockdown was characterized by reduction of 20% in TSPO RNA levels, 50% in protein expression of the TSPO, and 50% in binding with the TSPO ligand, [3H]PK 11195. Subsequently, in-vitro cell viability and proliferation assays were applied. In addition, transient transfecton with short interfering RNA (siRNA) directed against human TSPO was studied in this way. Furthermore, we also grafted HT29 cells subcutaneously into the right thighs of SCID mice to examine the effects of the putative TSPO agonist, FGIN-1-27, on tumor growth in-vivo. In-vitro TSPO knockdown established by stable transfection of TSPO antisense gene resulted in HT29 clones displaying significantly lower levels of cell death as determined with trypan blue (50% less), lower apoptotic rates (28% less), and higher proliferation rates (48% more one week after seeding and 27% more two weeks after seeding). Transient transfection with anti-human TSPO siRNA resulted in similar viability and antiapoptotic effects. In-vivo, the proapoptotic TSPO ligand, FGIN-1-27 significantly reduced the growth rate of grafted tumors (40% less), in comparison with vehicle-treated mice. TSPO knockdown by genetic manipulation transforms the human HT29 cancer line to a more malignant type in-vitro. In-vivo pharmacological treatment with the putative TSPO agonist FGIN-1-27 reduces tumor growth of the HT29 cell line. These data suggest that TSPO involvement in apoptosis provides a target for anticancer treatment.

  12. Interferon-gamma sensitizes colonic epithelial cell lines to physiological and therapeutic inducers of colonocyte apoptosis.

    LENUS (Irish Health Repository)

    O'Connell, J

    2012-02-03

    Homeostasis in the colonic epithelium is achieved by a continuous cycle of proliferation and apoptosis, in which imbalances are associated with disease. Inflammatory bowel disease (IBD) and colon cancer are associated with either excessive or insufficient apoptosis of colonic epithelial cells, respectively. By using two colonic epithelial cell lines, HT29 and SW620, we investigated how the epithelial cell\\'s sensitivity to apoptosis was regulated by the proinflammatory cytokine interferon-gamma (IFN-gamma). We found that IFN-gamma sensitized HT29 cells, and to a lesser extent SW620, to diverse inducers of apoptosis of physiologic or therapeutic relevance to the colon. These apoptosis inducers included Fas (CD95\\/APO-1) ligand (FasL), short-chain fatty acids, and chemotherapeutic drugs. The extent of IFN-gamma-mediated apoptosis sensitization in these two cell lines correlated well with the degree of IFN-gamma-mediated upregulation of the proapoptotic protease caspase-1. Although IFN-gamma alone effectively sensitized HT29 cells to apoptosis, inclusion of the protein synthesis inhibitor cyclohexamide (CHX) during apoptotic challenge was necessary for maximal sensitization of SW620. The requirement of CHX to sensitize SW620 cells to apoptosis implies a need to inhibit translation of antiapoptotic proteins absent from HT29. In particular, the antiapoptotic protein Bcl-2 was strongly expressed in SW620 cells but absent from HT29. Our results indicate that IFN-gamma increases the sensitivity of colonic epithelial cells to diverse apoptotic stimuli in concert, via upregulation of caspase-1. Our findings implicate caspase-1 and Bcl-2 as important central points of control determining the general sensitivity of colonic epithelial cells to apoptosis.

  13. Cytotoxicity and analysis of apoptosis gene expression in colon cancer cell line treated with cell extract of Lactobacillus casei as indigenous probiotic bacterium

    Directory of Open Access Journals (Sweden)

    Amir Mirzaie

    2017-03-01

    Full Text Available Background and aim: Nowadays, the probiotic bacteria such as lactobacilli are known as prevention factor for various disease especially cancer. The aim of this study was to investigate the cytotoxic effect of Lactobacillus casei PTCC 1608 cell extract as probiotic bacteria on colon cancer cell line (HT29 and analysis of Bax and Bcl2 apoptosis gene expression. Methods: In this experimental study, the cell extract of heat killed L. casei was prepared in 0.01, 0.1, 1, 10, 100 and 1000 µg/ml concentration and subsequently, the cytotoxicity of various cell extracts on HT29 and HEC293 cell lines were evaluated in 24 hours using MTT assay. Moreover, the Bax and Bcl2 apoptosis gene expression level in HT29 cell line was analyzed using Real Time PCR. The apoptotic effects of cell extract was determined using Flow-cytometry technique. Finally, the collected data were statistically analyzed using one-way anal­ysis of variance with the SPSS/18 software. Results: The results of MTT test show that cell extracts of L. casei is able to reduce the survival rate of HT29 cell line to 0.95±0.44, 73.45±0.21, 51.49±0.87, 39.5±0.45 and 19.7±0.55. In addition to, the Real Time PCR results indicated the expression level of Bax and Bcl2 was increased and decreased respectively, in HT29 cell line (2.76 ± 0.54 (P<0.05, 0.21 ± 0.43 (P< 0.05 in 24 h. Moreover, the flow cytometry results indicated the 35.62 % apoptosis in HT29 cell line treated with IC50 value. Conclusion: The results show that the cell extract of L. casei PTCC 1608 could induced the apoptosis in HT29 cell line and it had low toxicity on HEC293 cell line. Therefore, it seems that L. casei has potential uses as probiotic for pharmaceutical applications including prevention and treatment of colon cancer.

  14. Sensitization of colon cancer cell lines to butyrate-mediated proliferation inhibition by combined application of indomethacin and nordihydroguaiaretic acid.

    Science.gov (United States)

    Galfi, Peter; Neogrady, Zsuzsa; Amberger, Albert; Margreiter, Raimund; Csordas, Adam

    2005-01-01

    The aim of the study was to investigate the effects of nonsteroidal anti-inflammatory drugs (NSAIDs) on histone deacetylase-mediated proliferation inhibition. In the colon cancer cell line HT29 butyrate-mediated proliferation inhibition was enhanced by the additional presence of indomethacin (IM) and/or nordihydroguaiaretic acid (NDGA). Sensitisation to butyrate-mediated proliferation inhibition was abolished by the general caspase inhibitor Z-VAD-fmk, however, only IM-induced cell detachment was prevented by the caspase inhibitor but not that induced by NDGA or NDGA plus IM. In contrast to the parental cell line HT29, in the methotrexate-resistant sub-lines HT29-12 and HT29-21, IM counteracted butyrate-mediated proliferation inhibition, which was abrogated by NDGA. In all the investigated cell lines, proliferation inhibition was most effectively achieved under the combined application of butyrate with IM and NDGA, suggesting that inhibition of both cyclooxygenase (COX) and lipoxygenase (LOX) isoenzymes is needed for proliferation inhibition by NSAIDs in tumour cells.

  15. Investigation of the selenium metabolism in cancer cell lines

    DEFF Research Database (Denmark)

    Lunøe, Kristoffer; Gabel-Jensen, Charlotte; Stürup, Stefan

    2011-01-01

    incubated with cells for 24 h and the induction of cell death was measured using flow cytometry. The amounts of total selenium in cell medium, cell lysate and the insoluble fractions was determined by ICP-MS. Speciation analysis of cellular fractions was performed by reversed phase, anion exchange and size......The aim of this work was to compare different selenium species for their ability to induce cell death in different cancer cell lines, while investigating the underlying chemistry by speciation analysis. A prostate cancer cell line (PC-3), a colon cancer cell line (HT-29) and a leukaemia cell line...... (Jurkat E6-1) were incubated with five selenium compounds representing inorganic as well as organic Se compounds in different oxidation states. Selenomethionine (SeMet), Se-methylselenocysteine (MeSeCys), methylseleninic acid (MeSeA), selenite and selenate in the concentration range 5-100 mu M were...

  16. Transport Study of Egg-Derived Antihypertensive Peptides (LKP and IQW) Using Caco-2 and HT29 Coculture Monolayers.

    Science.gov (United States)

    Xu, Qingbiao; Fan, Hongbing; Yu, Wenlin; Hong, Hui; Wu, Jianping

    2017-08-30

    The objective of this study was to investigate the mechanisms of the transport of antihypertensive tripeptides LKP (Leu-Lys-Pro) and IQW (Ile-Gln-Trp) derived from egg white using a coculture system of Caco-2 and HT29 cell monolayers. The results revealed that LKP and IQW have no cytotoxicity to the cell viability after 2 h incubation, could be transported intact across coculture monolayers (apparent permeability coefficient: (18.11 ± 1.57) × 10 -8 and (13.21 ± 1.12) × 10 -8 cm/s, respectively), and were resistant to peptidase secreted by enterocytes. In addition, the transports were significantly inhibited by dipeptide Gly-Pro (P Caco-2/HT29 coculture monolayers.

  17. Modulation of cellular redox state underlies antagonism between oxaliplatin and cetuximab in human colorectal cancer cell lines.

    Science.gov (United States)

    Dahan, Laetitia; Sadok, Amine; Formento, Jean-Louis; Seitz, Jean François; Kovacic, Hervé

    2009-09-01

    Oxaliplatin is the first platinum-based compound effective in the treatment of colorectal cancer. Oxaliplatin combined with cetuximab for metastatic colorectal cancer is under evaluation. The preliminary results seem controversial, particularly for the use of cetuximab in K-Ras mutated patients. K-Ras mutation is known to affect redox homeostasis. Here we evaluated how the efficacy of oxaliplatin alone or combined with cetuximab varied according to the Ras mutation and redox status in a panel of colorectal tumour cell lines. Viability was evaluated by methylthiazoletetrazolium assay, reactive oxygen species production by DCFDA and lucigenin on HT29-D4, Caco-2, SW480 and SW620 cell lines. Combination of oxaliplatin and cetuximab was less cytotoxic than oxaliplatin alone in colorectal cells harbouring wild-type Ras and membrane expression of receptors for epidermal growth factor receptor (EGFR), such as HT29-D4 and Caco-2 cells. In contrast, cetuximab did not affect oxaliplatin efficiency in cells harbouring K-Ras(V12) mutation, irrespective of membrane EGFR expression (SW620 and SW480 cells). Transfection of HT29-D4 with K-Ras(V12) decreased oxaliplatin IC(50) and impaired cetuximab sensitivity, without affecting expression of membrane EGFR compared with HT29-D4 control. Oxaliplatin efficacy relies on endogenous production of H(2)O(2). Cetuximab inhibits H(2)O(2) production inhibiting the EGFR/Nox1 NADPH oxidase pathway. Oxaliplatin efficacy was impaired by short hairpin RNA for Nox1 and by catalase (H(2)O(2) scavenger). Cetuximab limited oxaliplatin efficiency by affecting the redox status of cancer cells through Nox1. Such combined therapy might be improved by controlling H(2)O(2) elimination.

  18. Uptake of Ga-curcumin derivatives in different cancer cell lines: Toward the development of new potential 68Ga-labelled curcuminoids-based radiotracers for tumour imaging.

    Science.gov (United States)

    Rubagotti, Sara; Croci, Stefania; Ferrari, Erika; Orteca, Giulia; Iori, Michele; Capponi, Pier C; Versari, Annibale; Asti, Mattia

    2017-08-01

    Thanks to the ability to suppress the proliferation and to kill tumour cells, several studies have shown the anti-cancer effects of curcumin (CUR) and its derivatives, i.e. diacetylcurcumin (DAC) and bis-dehydroxycurcumin (bDHC). This study is focused onto the development of curcuminoid complexes with gallium-68 employed as potential new radio-labelled probes to detect neoplastic tissues through imaging techniques such as positron emission tomography. To this purpose, the uptake of three Ga-curcuminoid complexes, namely Ga(CUR) 2 + , Ga(DAC) 2 + , Ga(bDHC) 2 + , by various tumour cell lines was compared with the uptake of the same compounds by normal human lymphocytes by flow cytometry using the intrinsic fluorescence of the curcuminoids. Ga(CUR) 2 + , and particularly Ga(DAC) 2 + , showed a higher uptake by colorectal carcinoma (HT29) and lymphoma (K562) cell lines than lymphocytes, while the uptake of Ga(bDHC) 2 + was higher in lymphocytes than in all the other cell lines. Based on the fluorescence data, Gallium-68 labelled complexes were then tested in HT29 cell line. 68 Ga(DAC) 2 + showed the highest uptake by HT29 cells (higher internalization with a lower externalization) and the highest affinity. The obtained results are promising and the findings foster further investigation on the development of curcumin-metal-based radiopharmaceuticals. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Cytotoxic effects of the newly-developed chemotherapeutic agents 17-AAG in combination with oxaliplatin and capecitabine in colorectal cancer cell lines.

    Science.gov (United States)

    Mohammadian, Mahshid; Zeynali, Shima; Azarbaijani, Anahita Fathi; Khadem Ansari, Mohammad Hassan; Kheradmand, Fatemeh

    2017-12-01

    The use of heat shock protein 90 inhibitors like 17-allylamino-17-demethoxy-geldanamycin (17-AAG) has been recently introduced as an attractive anticancer therapy. It has been shown that 17-AAG may potentiate the inhibitory effects of some classical anticolorectal cancer (CRC) agents. In this study, two panels of colorectal carcinoma cell lines were used to evaluate the effects of 17-AAG in combination with capecitabine and oxaliplatin as double and triple combination therapies on the proliferation of CRC cell lines. HT-29 and all HCT-116 cell lines were seeded in culture media in the presence of different doses of the mentioned drugs in single, double, and triple combinations. Water-soluble tetrazolium-1 (WST-1) assay was used to investigate cell proliferation 24 h after treatments. Then, dose-response curves were plotted using WST-1outputs, and IC 50 values were determined. For double and triple combinations respectively 0.5 × IC 50 and 0.25 × IC 50 were used. Data was analyzed with the software CompuSyn. Drug interactions were analyzed using Chou-Talalay method to calculate the combination index (CI).The data revealed that 17-AAG shows a potent synergistic interaction (CI 1) in HT-29 and a synergistic effect (CI AAG with oxaliplatin or capecitabine might be effective against HCT-116 and HT-29 cell lines. However, in triple combinations, positive results were seen only against HCT-116. Further investigation is suggested to confirm the effectiveness of these combinations in clinical trials.

  20. Essential Oil Content of the Rhizome of Curcuma purpurascens Bl. (Temu Tis and Its Antiproliferative Effect on Selected Human Carcinoma Cell Lines

    Directory of Open Access Journals (Sweden)

    Sok-Lai Hong

    2014-01-01

    Full Text Available Curcuma purpurascens Bl., belonging to the Zingiberaceae family, is known as temu tis in Yogyakarta, Indonesia. In this study, the hydrodistilled dried ground rhizome oil was investigated for its chemical content and antiproliferative activity against selected human carcinoma cell lines (MCF7, Ca Ski, A549, HT29, and HCT116 and a normal human lung fibroblast cell line (MRC5. Results from GC-MS and GC-FID analysis of the rhizome oil of temu tis showed turmerone as the major component, followed by germacrone, ar-turmerone, germacrene-B, and curlone. The rhizome oil of temu tis exhibited strong cytotoxicity against HT29 cells (IC50 value of 4.9 ± 0.4 μg/mL, weak cytotoxicity against A549, Ca Ski, and HCT116 cells (with IC50 values of 46.3 ± 0.7, 32.5 ± 1.1, and 35.0 ± 0.3 μg/mL, resp., and no inhibitory effect against MCF7 cells. It exhibited mild cytotoxicity against a noncancerous human lung fibroblast cell line (MRC5, with an IC50 value of 25.2 ± 2.7 μg/mL. This is the first report on the chemical composition of this rhizome’s oil and its selective antiproliferative effect on HT29. The obtained data provided a basis for further investigation of the mode of cell death.

  1. Essential Oil Content of the Rhizome of Curcuma purpurascens Bl. (Temu Tis) and Its Antiproliferative Effect on Selected Human Carcinoma Cell Lines

    Science.gov (United States)

    Hong, Sok-Lai; Lee, Guan-Serm; Ahmed Hamdi, Omer Abdalla; Awang, Khalijah; Aznam Nugroho, Nurfina

    2014-01-01

    Curcuma purpurascens Bl., belonging to the Zingiberaceae family, is known as temu tis in Yogyakarta, Indonesia. In this study, the hydrodistilled dried ground rhizome oil was investigated for its chemical content and antiproliferative activity against selected human carcinoma cell lines (MCF7, Ca Ski, A549, HT29, and HCT116) and a normal human lung fibroblast cell line (MRC5). Results from GC-MS and GC-FID analysis of the rhizome oil of temu tis showed turmerone as the major component, followed by germacrone, ar-turmerone, germacrene-B, and curlone. The rhizome oil of temu tis exhibited strong cytotoxicity against HT29 cells (IC50 value of 4.9 ± 0.4 μg/mL), weak cytotoxicity against A549, Ca Ski, and HCT116 cells (with IC50 values of 46.3 ± 0.7, 32.5 ± 1.1, and 35.0 ± 0.3 μg/mL, resp.), and no inhibitory effect against MCF7 cells. It exhibited mild cytotoxicity against a noncancerous human lung fibroblast cell line (MRC5), with an IC50 value of 25.2 ± 2.7 μg/mL. This is the first report on the chemical composition of this rhizome's oil and its selective antiproliferative effect on HT29. The obtained data provided a basis for further investigation of the mode of cell death. PMID:25177723

  2. Study on the Cytotoxic Activity of Drimane Sesquiterpenes and Nordrimane Compounds against Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Ivan Montenegro

    2014-11-01

    Full Text Available Twelve drimanes, including polygodial (1, isopolygodial (2, drimenol (3, confertifolin (4, and isodrimenin (5, were obtained from natural sources. Semi-synthetic derivatives 6–12 were obtained from 1 and 2, and cytotoxic activity was evaluated in vitro against cancer cell lines (HT-29, MDA-MB231, DHF, MCF-7, PC-3, DU-145, and CoN. IC50 values were determined at concentrations of 12.5–100 µM of each compound for 72 h. In addition, it was found that polygodial (1, 8, and 12 induced changes in mitochondrial membrane permeability in CoN, MCF-7, and PC-3 cells.

  3. Oxidative Stress Promotes Doxorubicin-Induced Pgp and BCRP Expression in Colon Cancer Cells Under Hypoxic Conditions.

    Science.gov (United States)

    Pinzón-Daza, Martha L; Cuellar-Saenz, Yenith; Nualart, Francisco; Ondo-Mendez, Alejandro; Del Riesgo, Lilia; Castillo-Rivera, Fabio; Garzón, Ruth

    2017-07-01

    P-glycoprotein (Pgp) and breast cancer resistance protein (BCRP) are ATP binding cassette (ABC) transporters that are overexpressed in different drug-resistant cancer cell lines. In this study, we investigated whether doxorubicin promotes Pgp and/or BCRP expression to induce drug resistance in colon cancer cells under hypoxic conditions. We analyzed HIF-1α activity via ELISA, Pgp, and BCRP expression by qRT-PCR and the relationship between doxorubicin uptake and ABC transporter expression via confocal microscopy in HT-29WT and HT-29 doxorubicin-resistant colon cancer cells (HT-29DxR). These cells were treated with doxorubicin and/or CoCl 2 (chemical hypoxia), and reactive oxygen species inductors. We found that the combination of chemically induced hypoxia and doxorubicin promoted Pgp mRNA expression within 24 h in HT-29WT and HT-29DxR cells. Both doxorubicin and CoCl 2 alone or in combination induced Pgp and BCRP expression, as demonstrated via confocal microscopy in each of the above two cell lines. Thus, we surmised that Pgp and BCRP expression may result from synergistic effects exerted by the combination of doxorubicin-induced ROS production and HIF-1α activity under hypoxic conditions. However, HIF-1α activity disruption via the administration of E3330, an APE-1 inhibitor, downregulated Pgp expression and increased doxorubicin delivery to HT-29 cells, where it served as a substrate for Pgp, indicating the existence of an indirect relationship between Pgp expression and doxorubicin accumulation. Thus, we concluded that Pgp and BCRP expression can be regulated via cross-talk between doxorubicin and hypoxia, promoting drug resistance in HT-29 WT, and HT-29DxR cells and that this process may be ROS dependent. J. Cell. Biochem. 118: 1868-1878, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  4. SILENCING THE NUCLEOCYTOPLASMIC O-GLCNAC TRANSFERASE REDUCES PROLIFERATION, ADHESION AND MIGRATION OF CANCER AND FETAL HUMAN COLON CELL LINES

    Directory of Open Access Journals (Sweden)

    AGATA eSTEENACKERS

    2016-05-01

    Full Text Available The post-translational modification of proteins by O-linked β-N-acetylglucosamine (O-GlcNAc is regulated by a unique couple of enzymes. O-GlcNAc transferase (OGT transfers the GlcNAc residue from UDP-GlcNAc, the final product of the hexosamine biosynthetic pathway (HBP, whereas O-GlcNAcase (OGA removes it. This study and others show that OGT and O-GlcNAcylation levels are increased in cancer cell lines. In that context we studied the effect of OGT silencing in the colon cancer cell lines HT29 and HCT116 and the primary colon cell line CCD841CoN. Herein we report that OGT silencing diminished proliferation, in vitro cell survival and adhesion of primary and cancer cell lines. SiOGT dramatically de-creased HT29 and CCD841CoN migration, CCD841CoN harboring high capabilities of mi-gration in Boyden chamber system when compared to HT29 and HCT116. The expression levels of actin and tubulin were unaffected by OGT knockdown but siOGT seemed to disor-ganize microfilament, microtubule and vinculin networks in CCD841CoN. While cancer cell lines harbor higher levels of OGT and O-GlcNAcylation to fulfill their proliferative and migra-tory properties, in agreement with their higher consumption of HBP main substrates glucose and glutamine, our data demonstrate that OGT expression is not only necessary for the biolog-ical properties of cancer cell lines but also for normal cells.

  5. O-naphthoquinone isolated from Capraria biflora L. induces selective cytotoxicity in tumor cell lines.

    Science.gov (United States)

    de S Wisintainer, G G N; Scola, G; Moura, S; Lemos, T L G; Pessoa, C; de Moraes, M O; Souza, L G S; Roesch-Ely, M; Henriques, J A P

    2015-12-21

    Biflorin is an o-naphthoquinone isolated from the roots of the plant Capraria biflora L. (Scrophulariaceae). In this study, the cytotoxic effects of biflorin were verified, and late apoptosis was detected in various cancer cell lines by in situ analysis. The cytotoxicity was further evaluated exclusively for 48 h of treatment in different tumor and non-tumor cell lines (Hep-2, HeLa, HT-29, A-375, and A-549, and HEK-293, respectively). The results indicated that biflorin induced selective cytotoxicity in tumor cells. HeLa cells were more susceptible to biflorin, followed by HT-29, A-549, A-375, and Hep-2 at all concentrations (range 5-50 μg/mL), and the highest half-maximal inhibitory concentration IC50 (56.01 ± 1.17 μg/mL) was observed in HEK-293 cells. Late apoptotic/necrotic events, observed by in situ immunostaining with Annexin V, varied with each cell line; an increase in late apoptotic events was observed corresponding to the increase in biflorin dosage. Hep-2 cells showed a greater percentage of late apoptotic events among the tumor cell lines when treated with higher concentrations of biflorin (69.63 ± 2.28%). The non-tumor HEK-293 line showed greater resistance to late apoptotic events, as well as a lower level of cytotoxicity (77.69 ± 6.68%) than the tested tumor lines. The data presented indicate that biflorin showed an important, possibly selective, cytotoxicity against tumor cell lines, thereby revealing a promising novel substance with potential anticancer activity for tumor therapy.

  6. In vitro anticancer effect of venom from Cuban scorpion Rhopalurus junceus against a panel of human cancer cell lines

    OpenAIRE

    D?az-Garc?a, Alexis; Morier-D?az, Luis; Fri?n-Herrera, Yahima; Rodr?guez-S?nchez, Hermis; Caballero-Lorenzo, Yamira; Mendoza-Llanes, Dianeya; Riquenes-Garlobo, Yanelis; Fraga-Castro, Jos? A

    2013-01-01

    In Cuba the endemic species of scorpion Rhopalurus junceus has been used in traditional medicine for cancer treatment. However, there is little scientific evidence about its potential in cancer therapy. The effect of a range of scorpion venom concentrations (0.1, 0.25, 0.5, 0.75 and 1mg/ml) against a panel of human tumor cell lines from epithelial (Hela, SiHa, Hep-2, NCI-H292, A549, MDA-MB-231, MDA-MB-468, HT-29), hematopoietic origins (U937, K562, Raji) and normal cells (MRC-5, MDCK, Vero) w...

  7. Cytokinetic effects of razoxane and relevance to radiopotentiation in a human adenocarcinoma cell line

    International Nuclear Information System (INIS)

    Norimah Yusof.

    1982-01-01

    Razoxane (+-1,2-bis (3,5-dioxopiperazin-1-yl)propane) or ICRF 159 has been shown to enhance the X-radiation sensitivity of a human colorectal adenocarcinoma cell line (HT29R) in culture if the cells were incubated with the drug for more than ten hours prior irradiation. Razoxane was shown not to interfere with repair of sublethal of potentially lethal damage of radiation. The radiosensitizing effect is therefore attributed to a reduced ability to accumulate such damage. As the effect was shown greater in exponential phase, which consists of greater proportion of dividing cells than plateau phase cells, it is believed that the drug (0.1 - 100 μg/ml) potentiates the effect of X-radiation in HT29R cells by pertubation in the cell cycle kinetics. Razoxane at low and high concentrations (5 and 100 μg/ml) was shown to delay the cell progression on cells entering mitosis. Chromosomes of drug-treated cells seemed to be unable to condense at the premitosis stage and cells with chromosomes resembling those in prophase were allowed to proceed through mitosis but progressed very slowly so that accumulation of mitotic cells were seen. Cells were than allowed to complete mitosis but frequently failed to undergo cytokinesis and subsequently produced many giant multinucleate cells. It is likely that the cells in mitosis and multinucleate cells are more radiosensitive. However, the mitotic delay was found to be reversible following progressive drug inactivation in the cells. Cells appeared to return to normal cell progression rate and division while the radiopotentiating effect was disappearing. The mechanism of drug action in inhibiting chromosome condensation was shown to involve the drug interference with histone H 1 phosphorylation. The inhibition of H 1 phosphorylation might reduce the cross-linking of the histone on DNA strands and thus preventing them from participating in supercoiling of the chromosome. (author)

  8. Antiproliferative effects of extracts from Iranian Artemisia species on cancer cell lines.

    Science.gov (United States)

    Taghizadeh Rabe, Shahrzad Zamanai; Mahmoudi, Mahmoud; Ahi, Ali; Emami, Seyed Ahmad

    2011-09-01

    Different species of Artemisia (Asteraceae) have shown to exhibit antitumor activity. The aim of this study was to identify the antiproliferative effect of some Artemisia species from Iran on cultured human cancer cells. Methanol, ethyl acetate, dichloromethane and n-hexane extracts from aerial parts of seven species of Artemisia were prepared and their antiproliferative effects on four cancer (AGS, HeLa, HT-29 and MCF-7) and normal cell line (L929) were determined. Different concentrations of extracts were added to cultured cells and incubated for 72 h. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay was employed to assess the cell viability. Different extracts exert various growth inhibitory effects. In case of AGS cells, dichloromethane and methanol extracts of A. ciniformis Krasch. & Popov ex Poljak. (IC(50) values: 35 and 60 µg/ml, respectively) showed the highest growth inhibitory effects. HeLa cells were more sensitive to both A. diffusa Krasch. ex Poljak. dichloromethane (IC(50) value: 71 µg/ml) and A. ciniformis ethylacetate (IC(50) value: 73 µg/ml) extracts. Dichloromethane extracts of A. diffusa, A. santolina Schrenk and A. ciniformis (IC(50) values: 42, 91 and 94 µg/ml, respectively) exhibited more inhibition on HT-29 cells in comparison to other extracts. MCF-7 cells were best inhibited by A. ciniformis dichloromethane (IC(50) value: 29 µg/ml) and A. vulgaris L. ethyl acetate (IC(50) value: 57 µg/ml) extracts. This study shows the antiproliferative effects of Artemisia extracts on malignant cell lines. Artemisia could be also considered as a promising chemotherapeutic agent in cancer treatment.

  9. Vitamin E analogue, D-alpha tocopherol succinate, enhances x-ray induced growth delay of human adenocarcinoma cancer cell line

    International Nuclear Information System (INIS)

    Jaworska, A.; Ottesen, T.E.

    2003-01-01

    The purpose of this study was to assess the effects of d-alpha Tocopherol succinate (alpha-TS) in modifying radiation-induced viability reduction and apoptosis occurrence in the model for normal and cancer cells. Our hypothesis was that alpha-TS enhances the growth-inhibitory effect of x-irradiation in cancer cells and that the effect is more pronounced in these cells than in normal cells. Murine NIH 3T3 Swiss albino embryonic cells and HT29 human Caucasian colon adenocarcinoma cells were used in the experiments. Alpha-TS was added to the cultures 1 h prior to irradiation with doses of 2 or 5Gy of x-ray. After irradiation cells were incubated for 73 h. Trypan blue exclusion viability test and estimation of apoptosis and necrosis were made. Apoptotic and necrotic cells were counted in fluorescence microscope using fluorescence dyes: propidium iodide and Hoechst 33342. For experiments with the dose of 5 Gy at least five series of experiments were performed. At lower doses (up to approximately 25μM/ml) treatment with alpha-TS alone enhanced growth of both cell lines. At higher doses treatment with alpha-TS alone delayed the growth of the cell cultures, accompanied by 20-25% necrosis. At the concentrations higher than 25μM/mL alpha-TS alone caused growth delay of both cell cultures, being much more pronounced for the cancer cell line HT29. At the concentrations of 50 μM/mL, responsible for about 30-60% of growth delay, there was observed a synergy effect for x-rays and alpha-TS for both cell lines. The effect was more pronounced for HT29 cells (DMF=0.48 for HT29 versus DMF=0.73 for NIH 3T3). These results may confirm the views of the literature reports suggesting that use of vitamin E together with radiation could be favorable for colon cancer treatment; however, more experiments using more advanced techniques are needed

  10. Cytotoxicity screening of essential oils in cancer cell lines

    Directory of Open Access Journals (Sweden)

    Pollyanna Francielli de Oliveira

    Full Text Available Abstract This study evaluated the cytotoxicity activity of the essential oils of Tagetes erecta L., Asteraceae (TE-OE, Tetradenia riparia (Hochst. Codd, Lamiaceae (TR-OE, Bidens sulphurea (Cav. Sch. Bip., Asteraceae (BS-OE, and Foeniculum vulgare Mill., Apiaceae (FV-OE, traditionally used in folk medicine, against the tumor cell lines murine melanoma (B16F10, human colon carcinoma (HT29, human breast adenocarcinoma (MCF-7, human cervical adenocarcinoma (HeLa, human hepatocellular liver carcinoma (HepG2, and human glioblastoma (MO59J, U343, and U251. Normal hamster lung fibroblasts (V79 cells were included as control. The cells were treated with essential oil concentrations ranging from 3.12 to 400 µg/ml for 24 h. The cytotoxic activity was evaluated using the XTT assay; results were expressed as IC50, and the selectivity index was calculated. The results were compared with those achieved for classic chemotherapeutic agents. TE-OE was the most promising among the evaluated oils: it afforded the lowest IC50 values for B16F10 cells (7.47 ± 1.08 µg/ml and HT29 cells (6.93 ± 0.77 µg/ml, as well as selectivity indices of 2.61 and 2.81, respectively. The major BS-EO, FV-EO and TE-EO chemical constituents were identified by gas chromatography mass spectrometry as being (E-caryophyllene (10.5%, germacrene D (35.0% and 2,6-di-tert-butyl-4-methylphenol (43.0% (BS-EO; limonene (21.3% and (E-anethole (70.2% (FV-EO; limonene (10.4%, dihydrotagetone (11.8%, α-terpinolene (18.1% and (E-ocimenone (13.0% (TE-EO; and fenchone (6.1%, dronabinol (11.0%, aromadendrene oxide (14.7% and (E,E–farnesol (15.0% (TR-EO. 2,6-di-tert-butyl-4-methylphenol (43.0%, (E-anethole (70.2% and α-terpinolene (18.1%, respectively. These results suggest that TE-OE may be used to treat cancer without affecting normal cells.

  11. The effect of some cosolvents and surfactants on viability of cancerous cell lines

    Directory of Open Access Journals (Sweden)

    M. Hamzeloo-Moghadam

    2014-06-01

    Full Text Available Improving the solubility of non-soluble herbal materials is an issue of interest in cell culture based experiments. Evaluating the biological activity of these materials could become possible with the aid of cosolvents/surfactants which obviously should have little or no cytotoxic activity. In the present study, the cytotoxic activity of six cosolvents/surfactants: ethanol, methanol, Tween 20 and 80, propylene glycol (PG and poly ethylene glycol 400 (PEG which are usually helpful in dissolving non-soluble herbal extracts, has been evaluated against HepG-2, MCF-7 and HT-29 cells by MTT assay. Among the investigated cosolvents/surfactants, Tween 20 and 80 demonstrated the highest and ethanol and methanol the lowest cytotoxicity to the evaluated cell lines, suggesting the two latter as proper aids for improving solubility in biological experiments.

  12. Effect of antigens from gram-positive and gram-negative bacteria, and of ŕ-gliadin on the intestinal epithelial cell lines HT-29, SW-480 and Caco-2

    Czech Academy of Sciences Publication Activity Database

    Funda, David P.; Farré, Maria Antionette; Horáková, Dana; Kolínská, Jiřina; Ceska, M.; Barot, R.; Tučková, Ludmila; Tlaskalová, Helena

    1997-01-01

    Roč. 75, č. 1 (1997), s. 52 ISSN 0818-9641. [International Congress of Mucosal Immunology /9./. 27.01.1997-31.01.1997, Sydney] R&D Projects: GA AV ČR IAA7020716; GA ČR GA310/96/1366; GA MZd IZ3761

  13. β1 integrin mediates colorectal cancer cell proliferation and migration through regulation of the Hedgehog pathway.

    Science.gov (United States)

    Song, Jia; Zhang, Jixiang; Wang, Jing; Wang, Jun; Guo, Xufeng; Dong, Weiguo

    2015-03-01

    β1 integrin (ITGB1) is the major expressed integrin protein of normal cells and tumor-associated cells. It is often up-regulated in human malignancies and is involved in many developmental processes, such as tumor progression and metastasis. However, little is known about the function of ITGB1 in colorectal cancer. We constructed lentiviral vectors expressing ITGB1 or ITGB1-specific RNA interference (RNAi) and an unrelated control vector. After infecting HT29 cells in vitro, proliferation and migration were evaluated by Cell Counting Kit 8 (CCK-8) assays, transwell invasion assays, and Western blots. The influence of lentivirus infection on the tumor development capacity of HT29 cells in vivo was examined by xenografting the tumor cells. The expression of ITGB1 in the xenografted tumor cells was analyzed by immunohistochemistry. The up-regulation of ITGB1 significantly increased the proliferation in HT29 cells in vitro. Moreover, we found that the overexpression of ITGB1 up-regulated sonic hedgehog (Shh) while down-regulating Gli1 and SuFu in HT29-ITGB1 cells compared to controls. Moreover, the levels of c-myc and cyclin D1 proteins were up-regulated. Transwell assays showed that the number of migrating HT29-RNAi cells was lower than that in the other cell groups, indicating that ITGB1 significantly enhances the invasive ability of HT29 cells. In addition to these in vitro results, ITGB1 was found to be a significantly effective growth factor in a xenografted tumor mouse model. These results suggest that ITGB1 induces growth and invasion in a human colorectal cancer cell line through the hedgehog (Hh) signaling pathway in vitro and in vivo.

  14. Cell line provenance.

    Science.gov (United States)

    Freshney, R Ian

    2002-07-01

    Cultured cell lines have become an extremely valuable resource, both in academic research and in industrial biotechnology. However, their value is frequently compromised by misidentification and undetected microbial contamination. As detailed elsewhere in this volume, the technology, both simple and sophisticated, is available to remedy the problems of misidentification and contamination, given the will to apply it. Combined with proper records of the origin and history of the cell line, assays for authentication and contamination contribute to the provenance of the cell line. Detailed records should start from the initiation or receipt of the cell line, and should incorporate data on the donor as well as the tissue from which the cell line was derived, should continue with details of maintenance, and include any accidental as well as deliberate deviations from normal maintenance. Records should also contain details of authentication and regular checks for contamination. With this information, preferably stored in a database, and suitable backed up, the provenance of the cell line so created makes the cell line a much more valuable resource, fit for validation in industrial applications and more likely to provide reproducible experimental results when disseminated for research in other laboratories.

  15. Dihydrochalcone Compounds Isolated from Crabapple Leaves Showed Anticancer Effects on Human Cancer Cell Lines

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    Xiaoxiao Qin

    2015-11-01

    Full Text Available Seven dihydrochalcone compounds were isolated from the leaves of Malus crabapples, cv. “Radiant”, and their chemical structures were elucidated by UV, IR, ESI-MS, 1H-NMR and 13C-NMR analyses. These compounds, which include trilobatin (A1, phloretin (A2, 3-hydroxyphloretin (A3, phloretin rutinoside (A4, phlorizin (A5, 6′′-O-coumaroyl-4′-O-glucopyranosylphloretin (A6, and 3′′′-methoxy-6′′-O-feruloy-4′-O-glucopyranosyl-phloretin (A7, all belong to the phloretin class and its derivatives. Compounds A6 and A7 are two new rare dihydrochalcone compounds. The results of a MTT cancer cell growth inhibition assay demonstrated that phloretin and these derivatives showed significant positive anticancer activities against several human cancer cell lines, including the A549 human lung cancer cell line, Bel 7402 liver cancer cell line, HepG2 human ileocecal cancer cell line, and HT-29 human colon cancer cell line. A7 had significant effects on all cancer cell lines, suggesting potential applications for phloretin and its derivatives. Adding a methoxyl group to phloretin dramatically increases phloretin’s anticancer activity.

  16. Protein and cell patterning in closed polymer channels by photoimmobilizing proteins on photografted poly(ethylene glycol) diacrylate

    DEFF Research Database (Denmark)

    Larsen, Esben Kjær Unmack; Mikkelsen, Morten Bo Lindholm; Larsen, Niels Bent

    2014-01-01

    on uncoated polymer surfaces. The coating could also efficiently suppress the adhesion of mammalian cells as demonstrated using the HT-29 cancer cell line. In a subsequent equivalent process step, protein in aqueous solution could be anchored onto the PEGDA coating in spatially defined patterns...

  17. Cytotoxic Activity of Selected Iranian Traditional Medicinal Plants on Colon, Colorectal and Breast Cancer Cell Lines

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    Leila Mohammad Taghizadeh Kashani

    2014-11-01

    Full Text Available Background: Many natural products from plants have been recognized to exert anticancer activity. In this study, ethanolic extracts of selected medicinal herbs from Iranian flora including Alyssum homolocarpum Fisch. (from seeds, Urtica dioica L. (from aerial parts, Cichorium intybus L. (from roots and Solanum nigrum L. (from fruits, were evaluated for their cytotoxic effect on different cell lines.Methods: Cytotoxic effect of these extracts was studied on three different cancer cell lines; colon carcinoma (HT-29, colorectal adenocarcinoma (Caco-2 and breast ductal carcinoma (T47D. In addition, Swiss mouse embryo fibroblasts (NIH 3T3 were used as normal nonmalignant cells. MTT assay (3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide was utilized for calculating the cytotoxicity of extracts on cell lines.Results: Results showed the potent cytotoxic activity of U. dioica ethanolic extract against T47D cell line with IC50 value of 46.14±4.55 µg/ml. Other extracts showed poor activity with IC50>100 µg/ml.Conclusions: Cytotoxic activity recorded in the present study revealed high potential antiproliferative activity of U. dioica ethanolic extract against T47D cell line. The real IC50 values of this extract may be considerably lower than the IC50 measured in our study if its pharmacological active compounds become pure. The results emphasize the importance of studies on U. dioica ethanolic extract to characterize potential components as cytotoxic natural medicines.

  18. Arctigenin induces apoptosis in colon cancer cells through ROS/p38MAPK pathway.

    Science.gov (United States)

    Li, Qing-chun; Liang, Yun; Tian, Yuan; Hu, Guang-rui

    2016-01-01

    In the current study the antiproliferative effect of arctigenin, plant lignin, was evaluated on human colon cancer cell line HT-29. Furthermore, attempts were made to explore the signaling mechanism which may be responsible for its effect. Cell growth inhibition was assessed by MTT and LDH assays. Flow cytometric analysis was performed to determine cell arrest in the cell cycle phase and apoptosis. Furthermore, to confirm the apoptotic activity of arctigenin, caspase-9 and -3 activities analysis was performed. The levels of reactive oxygen species (ROS) and p38 mitogen activated protein kinase (MAPK) were investigated to determine their role in inducing apoptosis in arctigenin-treated HT-29 colon cancer cell line. MTT and LDH results demonstrated significant cell growth inhibitory effect of arctigenin on HT-29 cells in a dose-dependent manner. Furthermore, increase in cell number arrested at G2/M phase was observed in flow cytometric analysis upon arctigenin treatment. In addition, arctigenin increased the apoptotic ratio in a dose-dependent manner. The involvement of intrinsic apoptotic pathway was indicated by the activation of caspase-9 and -3. Moreover, increased ROS production, activation of p38 MAPK and changes in mitochondrial membrane potential (ΔΨm) also revealed the role of intrinsic apoptotic signaling pathway in cell growth inhibition after arctigenin exposure. Arctigenin induces apoptosis in HT-29 colon cancer cells by regulating ROS and p38 MAPK pathways.

  19. Cell diameter measurements obtained with a handheld cell counter could be used as a surrogate marker of G2/M arrest and apoptosis in colon cancer cell lines exposed to SN-38

    International Nuclear Information System (INIS)

    Tahara, Makiko; Inoue, Takeshi; Miyakura, Yasuyuki; Horie, Hisanaga; Yasuda, Yoshikazu; Fujii, Hirofumi; Kotake, Kenjiro; Sugano, Kokichi

    2013-01-01

    Highlights: •Chemo-sensitivity to SN-38 was assayed by the automated cell counter. •Colon cancer cell line, HCT116 cells were more sensitive to SN-38 than HT29 cells. •Increase of cell size reflects G2/M arrest. •Appearance of small particles indicates cell apoptosis. -- Abstract: In vitro assessment of chemosensitivity are important for experiments evaluating cancer therapies. The Scepter 2.0 cell counter, an automated handheld device based on the Coulter principle of impedance-based particle detection, enables the accurate discrimination of cell populations according to cell size and volume. In this study, the effects of SN-38, the active metabolite of irinotecan, on the colon cancer cell lines HCT116 and HT29 were evaluated using this device. The cell count data obtained with the Scepter counter were compared with those obtained with the 3 H-thymidine uptake assay, which has been used to measure cell proliferation in many previous studies. In addition, we examined whether the changes in the size distributions of these cells reflected alterations in the frequency of cell cycle arrest and/or apoptosis induced by SN-38 treatment. In our experiments using the Scepter 2.0 cell counter, the cell counts were demonstrated to be accurate and reproducible measure and alterations of cell diameter reflected G2/M cell cycle arrest and apoptosis. Our data show that easy-to-use cell counting tools can be utilized to evaluate the cell-killing effects of novel treatments on cancer cells in vitro

  20. Autonomous inhibition of apoptosis correlates with responsiveness of colon carcinoma cell lines to ciglitazone.

    Directory of Open Access Journals (Sweden)

    David M Baron

    Full Text Available Colorectal cancer is a leading cause of mortality worldwide. Resistance to therapy is common and often results in patients succumbing to the disease. The mechanisms of resistance are poorly understood. Cells basically have two possibilities to survive a treatment with potentially apoptosis-inducing substances. They can make use of their existing proteins to counteract the induced reactions or quickly upregulate protective factors to evade the apoptotic signal. To identify protein patterns involved in resistance to apoptosis, we studied two colorectal adenocarcinoma cell lines with different growth responses to low-molar concentrations of the thiazolidinedione Ciglitazone: HT29 cells underwent apoptosis, whereas SW480 cells increased cell number. Fluorescence detection and autoradiography scans of 2D-PAGE gels were performed in both cell lines to assess protein synthesis and turnover, respectively. To verify the data we performed shotgun analysis using the same treatment procedure as in 2D-experiments. Biological functions of the identified proteins were mainly associated with apoptosis regulation, chaperoning, intrinsic inflammation, and DNA repair. The present study suggests that different growth response of two colorectal carcinoma cell lines after treatment with Ciglitazone results from cell-specific protein synthesis and differences in protein regulation.

  1. Heterogeneity in cancer cells: variation in drug response in different primary and secondary colorectal cancer cell lines in vitro.

    Science.gov (United States)

    Arul, Melanie; Roslani, April Camilla; Cheah, Swee Hung

    2017-05-01

    Tumor heterogeneity may give rise to differential responses to chemotherapy drugs. Therefore, unraveling tumor heterogeneity has an implication for biomarker discovery and cancer therapeutics. To test this phenomenon, we investigated the differential responses of three secondary colorectal cancer cell lines of different origins (HCT116, HT29, and SW620 cells) and four novel primary cell lines obtained from different colorectal cancer patients to 5-fluorouracil (5-FU) and oxaliplatin (L-OHP) and explored the differences in gene expression among the primary cell lines in response to exposure to cytotoxic drugs. Cells were exposed to different doses of 5-FU and L-OHP separately or in combinations of equitoxic drug or equimolar drug ratios (median effect of Chou-Talalay principle). Cell viability was assessed using MTT assay and the respective IC 50 values were determined. Changes in gene expression in primary cell lines after exposure to the same drug doses were compared using real-time PCR array. The sensitivities (IC 50 ) of different cell lines, both secondary and primary, to 5-FU and L-OHP were significantly different, whether in monotherapy or combined treatment. Primary cell lines needed higher doses to reach IC 50 . There were variations in gene expression among the primary cell lines of different chemosensitivities to the challenge of the same combined dose of 5-FU and L-OHP. The results confirm the heterogeneous nature of colorectal cancer cells from different patient tumors. Studies using primary cancer cells established from patient's tumors rather than secondary cell lines will more closely reflect the actual character of the disease.

  2. The effects of caffeic, coumaric and ferulic acids on proliferation, superoxide production, adhesion and migration of human tumor cells in vitro.

    Science.gov (United States)

    Nasr Bouzaiene, Nouha; Kilani Jaziri, Soumaya; Kovacic, Hervé; Chekir-Ghedira, Leila; Ghedira, Kamel; Luis, José

    2015-11-05

    Reactive oxygen species are well-known mediators of various biological responses. In this study, we examined the effect of three phenolic acids, caffeic, coumaric and ferulic acids, on superoxide anion production, adhesion and migration of human lung (A549) and colon adenocarcinoma (HT29-D4) cancer cell lines. Proliferation of both tumor cells was inhibited by phenolic acids. Caffeic, coumaric and ferulic acids also significantly inhibited superoxide production in A549 and HT29-D4 cells. Superoxide anion production decreased by 92% and 77% at the highest tested concentration (200 µM) of caffeic acid in A549 and HT29-D4 cell lines respectively. Furthermore, A549 and HT29-D4 cell adhesion was reduced by 77.9% and 79.8% respectively at the higher tested concentration of ferulic acid (200 µM). Migration assay performed towards A549 cell line, revealed that tested compounds reduced significantly cell migration. At the highest concentration tested (200 µM), the covered surface was 7.7%, 9.5% and 35% for caffeic, coumaric or ferulic acids, respectively. These results demonstrate that caffeic, coumaric and ferulic acids may participate as active ingredients in anticancer agents against lung and colon cancer development, at adhesion and migration steps of tumor progression. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Cell Penetrating Capacity and Internalization Mechanisms Used by the Synthetic Peptide CIGB-552 and Its Relationship with Tumor Cell Line Sensitivity.

    Science.gov (United States)

    Astrada, Soledad; Fernández Massó, Julio Raúl; Vallespí, Maribel G; Bollati-Fogolín, Mariela

    2018-03-30

    CIGB-552 is a twenty-amino-acid novel synthetic peptide that has proven to be effective in reducing tumor size and increasing lifespan in tumor-bearing mice. Such capability is conferred by its cell-penetrating peptide character, which allows it to enter cells and elicit a pro-apoptotic effect through its major mediator, COMMD1 protein. Cell-penetrating peptides are able to use different internalization mechanisms, such as endocytosis or direct transduction through the plasma membrane. Although CIGB-552 cytotoxicity has been evaluated in several non-tumor- and tumor-derived cell lines, no data regarding the relationship between cell line sensitivity, cell penetrating capacity, the internalization mechanisms involved, COMMD1 expression levels, or its subcellular localization has yet been produced. Here, we present the results obtained from a comparative analysis of CIGB-552 sensitivity, internalization capacity and the mechanisms involved in three human tumor-derived cell lines from different origins: mammary gland, colon and lung (MCF-7, HT-29 and H460, respectively). Furthermore, cell surface markers relevant for internalization processes such as phosphatidylserine, as well as CIGB-552 target COMMD1 expression/localization, were also evaluated. We found that both endocytosis and transduction are involved in CIGB-552 internalization in the three cell lines evaluated. However, CIGB-552 incorporation efficiency and contribution of each mechanism is cell-line dependent. Finally, sensitivity was directly correlated with high internalization capacity in those cell lines where endocytosis had a major contribution on CIGB-552 internalization.

  4. Three lysophosphatidic acids with a distinct long chain moiety differently affect cell differentiation of human colon epithelial cells to goblet cells.

    Science.gov (United States)

    Hidaka, Mayumi; Nishihara, Mai; Tokumura, Akira

    2018-03-15

    The intestinal mucus layer helps maintain intestinal homeostasis. In this study, we investigated the effects of lysophosphatidic acids (LPA) on differentiation of human colon carcinoma cell line, HT-29, to goblet cells with and without sodium butyrate, a known differentiation factor for intestinal cells. Number and average size of cells with goblet-like morphology in five photographs per dish were measured for assessment of differentiation of HT-29 cells to goblet cells as well as their relative portion of surface of to whole surface area of the photograph. Our results revealed that 18:1 LPA enhanced butyrate-induced differentiation of HT-29 cells. Because increased mRNA expression of LPA 5 and decreased mRNA expression of LPA 6 were observed in HT-29 cells after treatment with butyrate, we explored the effects of alkyl LPA and 20:4 LPA, which show preferentially higher affinities to LPA 5 and LPA 6 , respectively. As a result, the cell differentiation to goblet cell was increased by alkyl LPA but decreased by 20:4 LPA. Further, alkyl LPA and 18:1 LPA, but not 20:4 LPA, were found to reduce the numbers of cells surviving after incubation in a standard culture medium containing 10% fetal calf serum. We suggest that the three LPAs positively and negatively affect the differentiation of HT-29 cells to goblet cells, which may be associated with their reduced survival through the activation of distinct LPA receptor(s). Copyright © 2018 Elsevier Inc. All rights reserved.

  5. Antrodia camphorata grown on germinated brown rice inhibits HT-29 human colon carcinoma proliferation through inducing G0/G1 phase arrest and apoptosis by targeting the β-catenin signaling.

    Science.gov (United States)

    Park, Dong Ki; Lim, Yoong Ho; Park, Hye-Jin

    2013-08-01

    Antrodia camphorata (AC) has been used as a traditional medicine to treat food and drug intoxication, diarrhea, abdominal pain, hypertension, pruritis (skin itch), and liver cancer in East Asia. In this study, we investigated anticancer activities of AC grown on germinated brown rice (CBR) in HT-29 human colon cancer cells. We found that the inhibitory efficacy of CBR 80% ethanol (EtOH) extract on HT-29 and CT-26 cell proliferation was more effective than ordinary AC EtOH 80% extract. Next, 80% EtOH extract of CBR was further separated into four fractions; hexane, ethyl acetate (EtOAc), butanol (BuOH), and water. Among them, CBR EtOAc fraction showed the strongest inhibitory activity against HT-29 cell proliferation. Therefore, CBR EtOAc fraction was chosen for further studies. Annexin V-fluorescein isothiocyanate staining data indicated that CBR EtOAc fraction induced apoptosis. Induction of G0/G1 cell cycle arrest on human colon carcinoma cell was observed in CBR EtOAc fraction-treated cells. We found that CBR decreased the level of proteins involved in G0/G1 cell cycle arrest and apoptosis. CBR EtOAc fraction inhibited the β-catenin signaling pathway, supporting its suppressive activity on the level of cyclin D1. High performance liquid chromatography analysis data indicated that CBR EtOAc fraction contained adenosine. This is the first investigation that CBR has a greater potential as a novel chemopreventive agent than AC against colon cancer. These data suggest that CBR might be useful as a chemopreventive agent against colorectal cancer.

  6. Smoothened antagonist GDC-0449 (Vismodegib) inhibits proliferation and triggers apoptosis in colon cancer cell lines.

    Science.gov (United States)

    Wu, Chuanqing; Hu, Shaobo; Cheng, Ji; Wang, Guobin; Tao, Kaixiong

    2017-05-01

    The sonic hedgehog (Shh) pathway has been proven to be involved in embryonic development and cancer growth. GDC-0449, an antagonist of the hedgehog signaling receptor Smoothened (Smo), was recently approved by the US Food and Drug Administration as a prescription for skin basal cell carcinoma. However, the efficacy of GDC-0449 in the treatment of colon cancer and other malignancies, such as basal cell carcinoma and pancreatic cancer, has remained to be proven. The present study assessed the effect of GDC-0449 on the colon cancer cell lines Caco-2 and Ht-29. A Cell Counting Kit-8 assay was applied to assess the cell proliferation rate and apoptosis was tested by flow cytometry. Reverse-transcription quantitative PCR and western blot analysis were used for analyzing expression levels of target genes. Cell proliferation was inhibited, while apoptosis was increased by GDC-0449, whereas the expression of B-cell lymphoma 2 (Bcl-2), a downstream target of Shh signaling, was decreased. Consistent with the inhibition of Gli1 expression, the cancer stem cell markers CD44 and ALDH were decreased in the presence of GDC-0449. In conclusion, GDC-0449 was shown to inhibit the replication of colon cancer cells and trigger apoptosis through downregulating Bcl-2. This may also influence the stemness of cancer stem cells as indicated by the decreased stem cell surface markers.

  7. Carboxylated nanodiamonds are neither cytotoxic nor genotoxic on liver, kidney, intestine and lung human cell lines.

    Science.gov (United States)

    Paget, V; Sergent, J A; Grall, R; Altmeyer-Morel, S; Girard, H A; Petit, T; Gesset, C; Mermoux, M; Bergonzo, P; Arnault, J C; Chevillard, S

    2014-08-01

    Although nanodiamonds (NDs) appear as one of the most promising nanocarbon materials available so far for biomedical applications, their risk for human health remains unknown. Our work was aimed at defining the cytotoxicity and genotoxicity of two sets of commercial carboxylated NDs with diameters below 20 and 100 nm, on six human cell lines chosen as representative of potential target organs: HepG2 and Hep3B (liver), Caki-1 and Hek-293 (kidney), HT29 (intestine) and A549 (lung). Cytotoxicity of NDs was assessed by measuring cell impedance (xCELLigence® system) and cell survival/death by flow cytometry while genotoxicity was assessed by γ-H2Ax foci detection, which is considered the most sensitive technique for studying DNA double-strand breaks. To validate and check the sensitivity of the techniques, aminated polystyrene nanobeads were used as positive control in all assays. Cell incorporation of NDs was also studied by flow cytometry and luminescent N-V center photoluminescence (confirmed by Raman microscopy), to ensure that nanoparticles entered the cells. Overall, we show that NDs effectively entered the cells but NDs do not induce any significant cytotoxic or genotoxic effects on the six cell lines up to an exposure dose of 250 µg/mL. Taken together these results strongly support the huge potential of NDs for human nanomedicine but also their potential as negative control in nanotoxicology studies.

  8. DNA microarray profiling of genes differentially regulated by the histone deacetylase inhibitors vorinostat and LBH589 in colon cancer cell lines

    Directory of Open Access Journals (Sweden)

    Lenz Heinz-Josef

    2009-11-01

    Full Text Available Abstract Background Despite the significant progress made in colon cancer chemotherapy, advanced disease remains largely incurable and novel efficacious chemotherapies are urgently needed. Histone deacetylase inhibitors (HDACi represent a novel class of agents which have demonstrated promising preclinical activity and are undergoing clinical evaluation in colon cancer. The goal of this study was to identify genes in colon cancer cells that are differentially regulated by two clinically advanced hydroxamic acid HDACi, vorinostat and LBH589 to provide rationale for novel drug combination partners and identify a core set of HDACi-regulated genes. Methods HCT116 and HT29 colon cancer cells were treated with LBH589 or vorinostat and growth inhibition, acetylation status and apoptosis were analyzed in response to treatment using MTS, Western blotting and flow cytometric analyses. In addition, gene expression was analyzed using the Illumina Human-6 V2 BeadChip array and Ingenuity® Pathway Analysis. Results Treatment with either vorinostat or LBH589 rapidly induced histone acetylation, cell cycle arrest and inhibited the growth of both HCT116 and HT29 cells. Bioinformatic analysis of the microarray profiling revealed significant similarity in the genes altered in expression following treatment with the two HDACi tested within each cell line. However, analysis of genes that were altered in expression in the HCT116 and HT29 cells revealed cell-line-specific responses to HDACi treatment. In addition a core cassette of 11 genes modulated by both vorinostat and LBH589 were identified in both colon cancer cell lines analyzed. Conclusion This study identified HDACi-induced alterations in critical genes involved in nucleotide metabolism, angiogenesis, mitosis and cell survival which may represent potential intervention points for novel therapeutic combinations in colon cancer. This information will assist in the identification of novel pathways and targets

  9. Screening Antitumor Bioactive Fraction from Sauromatum giganteum (Engl. Cusimano & Hett and Sensitive Cell Lines with the Serum Pharmacology Method and Identification by UPLC-TOF-MS

    Directory of Open Access Journals (Sweden)

    Shi-Yong Gao

    2015-03-01

    Full Text Available Sauromatum giganteum (Engl. Cusimano & Hett Tuber are used in Chinese folklore medicine for treatment of neoplasms. However, the claim has not been scientifically validated. The aim of the study is to screen the antitumor bioactive fraction of Sauromatum giganteum (Engl. Cusimano & Hett Tuber and sensitive tumor cell lines using a cytotoxicity assay in vitro and tumor transplantation method in vivo, to support its use in folk medicine. The petroleum ether fraction, chloroform fraction, ethyl acetate fraction, n-butanol fraction and water fraction were successively extracted by turn by the maceration under reflux assay. Screening of antitumor bioactive fraction and sensitive cell lines were measured by MTT assay and the serum pharmacology method, and in vivo the antitumor activities of the active fraction was evaluated by using S180 or H22 tumor-bearing mice model and Kunming mice. The active constituents of ethyl acetate fraction of Sauromatum giganteum (Engl. Cusimano & Hett were characterized by UPLC-TOF-MS. Compared with control groups, mice serum containing ethyl acetate fraction had a inhibition effect on SMMC-7721 cell, SGC-7901 cell, MCF-7 cell, HeLa cell, A549 cell, HT-29, and MDA-MB-231, respectively, but mice serum containing other four fractions had no different with that of control group. The inhibition capabilities of mice serum containing ethyl acetate fraction on the seven cell lines in descending order is SGC-7901 > SMMC-7721 > MCF-7 > HT-29 > A549 > HeLa > MDA-MB-231. In vivo the inhibition rate of 106, 318, 954 mg/kg·d ethyl acetate fraction dry extract to sarcoma S180 is 15.22%, 26.15% and 40.24%, respectively, and life prolonging rate to hepatoma H22 is 33.61%, 40.16% and 55.74%. A total of 14 compounds were identified in the ethyl acetate fraction of Sauromatum giganteum (Engl. Cusimano & Hett. The results of the experimental studies proved the antitumor activity of Sauromatum giganteum (Engl. Cusimano & Hett and supported

  10. Radiosensitivity of mesothelioma cell lines

    International Nuclear Information System (INIS)

    Haekkinen, A.M.; Laasonen, A.; Linnainmaa, K.; Mattson, K.; Pyrhoenen, S.

    1996-01-01

    The present study was carried out in order to examine the radiosensitivity of malignant pleural mesothelioma cell lines. Cell kinetics, radiation-induced delay of the cell cycle and DNA ploidy of the cell lines were also determined. For comparison an HeLa and a human foetal fibroblast cell line were simultaneously explored. Six previously cytogenetically and histologically characterized mesothelioma tumor cell lines were applied. A rapid tiazolyl blue microtiter (MTT) assay was used to analyze radiosensitivity and cell kinetics and DNA ploidy of the cultured cells were determined by flow cytometry. The survival fraction after a dose of 2 Gy (SF2), parameters α and β of the linear quadratic model (LQ-model) and mean inactivation dose (D MID ) were also estimated. The DNA index of four cell lines equaled 1.0 and two cell lines equaled 1.5 and 1.6. Different mesothelioma cell lines showed a great variation in radiosensitivity. Mean survival fraction after a radiation dose of 2 Gy (SF2) was 0.60 and ranged from 0.36 to 0.81 and mean α value was 0.26 (range 0.48-0.083). The SF2 of the most sensitive diploid mesothelioma cell line was 0.36: Less than that of the foetal fibroblast cell line (0.49). The survival fractions (0.81 and 0.74) of the two most resistant cell lines, which also were aneuploid, were equal to that of the HeLa cell line (0.78). The α/β ratios of the most sensitive cell lines were almost an order of magnitude greater than those of the two most resistant cell lines. Radiation-induced delay of the most resistant aneuploid cell line was similar to that of HeLa cells but in the most sensitive (diploid cells) there was practically no entry into the G1 phase following the 2 Gy radiation dose during 36 h. (orig.)

  11. CLO : The cell line ontology

    NARCIS (Netherlands)

    Sarntivijai, Sirarat; Lin, Yu; Xiang, Zuoshuang; Meehan, Terrence F.; Diehl, Alexander D.; Vempati, Uma D.; Schuerer, Stephan C.; Pang, Chao; Malone, James; Parkinson, Helen; Liu, Yue; Takatsuki, Terue; Saijo, Kaoru; Masuya, Hiroshi; Nakamura, Yukio; Brush, Matthew H.; Haendel, Melissa A.; Zheng, Jie; Stoeckert, Christian J.; Peters, Bjoern; Mungall, Christopher J.; Carey, Thomas E.; States, David J.; Athey, Brian D.; He, Yongqun

    2014-01-01

    Background: Cell lines have been widely used in biomedical research. The community-based Cell Line Ontology (CLO) is a member of the OBO Foundry library that covers the domain of cell lines. Since its publication two years ago, significant updates have been made, including new groups joining the CLO

  12. New insights into the molecular mechanism of Boletus edulis ribonucleic acid fraction (BE3) concerning antiproliferative activity on human colon cancer cells.

    Science.gov (United States)

    Lemieszek, Marta Kinga; Ribeiro, Miguel; Marques, Guilhermina; Nunes, Fernando Milheiro; Pożarowski, Piotr; Rzeski, Wojciech

    2017-05-24

    One of the relatively new and promising strategies of cancer treatment is chemoprevention, which involves the use of natural or synthetic compounds to block, inhibit or reverse carcinogenesis. A valuable and still untapped source of chemopreventive compounds seems to be edible mushrooms belonging to higher Basidiomycetes. Boletus edulis biopolymers extracted with hot water and purified by anion-exchange chromatography showed antiproliferative activity in colon cancer cells, but only fraction BE3, mostly composed of ribonucleic acids, was able to inhibit DNA synthesis in HT-29 cells. The present work aims to elucidate the molecular mechanism of this Boletus edulis ribonucleic acid fraction and in this sense flow cytometry and western blotting were applied to cell cycle analysis in HT-29 cells. We found that the antiproliferative ability of fraction BE3 observed in HT-29 cells was associated with the modulation of expression of cell cycle regulatory proteins (Cyclin D1, Cyclin A, p21 and p27) leading to cell accumulation in the S phase of the cell cycle. Furthermore, the BE3 fraction showed effective silencing of the signal transduction in an MAPK/Erk pathway in HT-29 and LS180 colon cancer cell lines. Thus, the previously and currently obtained results indicate that the BE3 fraction from Boletus edulis has great potential and needs to be further exploited through animal and clinical studies in order to develop a new efficient and safe therapeutic strategy for people who have been threatened by or suffered from colon cancer.

  13. Polyunsaturated fatty acids sensitize human colon adenocarcinoma HT-29 cells to death receptor-mediated apoptosis

    Czech Academy of Sciences Publication Activity Database

    Hofmanová, Jiřina; Vaculová, Alena; Kozubík, Alois

    2005-01-01

    Roč. 218, - (2005), s. 33-41 ISSN 0304-3835 R&D Projects: GA ČR(CZ) GA524/04/0895 Institutional research plan: CEZ:AV0Z50040507 Keywords : colon cancer * diet * butyrate Subject RIV: BO - Biophysics Impact factor: 3.049, year: 2005

  14. Induction of oncogene addiction shift to NF-κB by camptothecin in solid tumor cells

    International Nuclear Information System (INIS)

    Togano, Tomiteru; Sasaki, Masataka; Watanabe, Mariko; Nakashima, Makoto; Tsuruo, Takashi; Umezawa, Kazuo; Higashihara, Masaaki; Watanabe, Toshiki; Horie, Ryouichi

    2009-01-01

    The biological basis of the resistance of solid tumor cells to chemotherapy is not well understood. While addressing this problem, we found that gastric cancer cell line St-4/CPT, lung cancer cell line A549/CPT, and colon cancer cell line HT-29/CPT, all of which are resistant to camptothecin (CPT), showed strong and constitutive nuclear factor (NF)-κB activity driven by IκB kinase compared with their parental cell lines St-4, A549, and HT-29. A new NF-κB inhibitor, dehydroxymethylepoxyquinomicin (DHMEQ), reduced viability and induced apoptosis in St-4/CPT, A549/CPT, and HT-29/CPT cell lines, while their parental cell lines were resistant to DHMEQ. The results in this study present an example of the shift in signals that support the survival of solid tumor cells to NF-κB during the acquisition of resistance to CPT. The results also indicate that solid tumor cells that become resistant to chemotherapy may be more easily treated by NF-κB inhibitors.

  15. Induction of oncogene addiction shift to NF-{kappa}B by camptothecin in solid tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Togano, Tomiteru; Sasaki, Masataka; Watanabe, Mariko; Nakashima, Makoto [Department of Hematology, School of Medicine, Kitasato University, 1-15-1 Kitasato, Sagamihara, Kanagawa 228-8555 (Japan); Tsuruo, Takashi [Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, 3-10-6 Ariake, Koto-ku, Tokyo 135-8550 (Japan); Umezawa, Kazuo [Department of Applied Chemistry, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, Kanagawa 223-0061 (Japan); Higashihara, Masaaki [Department of Hematology, School of Medicine, Kitasato University, 1-15-1 Kitasato, Sagamihara, Kanagawa 228-8555 (Japan); Watanabe, Toshiki [Laboratory of Tumor Cell Biology, Department of Medical Genome Sciences, Graduate School of Frontier Sciences, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639 (Japan); Horie, Ryouichi, E-mail: rhorie@med.kitasato-u.ac.jp [Department of Hematology, School of Medicine, Kitasato University, 1-15-1 Kitasato, Sagamihara, Kanagawa 228-8555 (Japan)

    2009-12-04

    The biological basis of the resistance of solid tumor cells to chemotherapy is not well understood. While addressing this problem, we found that gastric cancer cell line St-4/CPT, lung cancer cell line A549/CPT, and colon cancer cell line HT-29/CPT, all of which are resistant to camptothecin (CPT), showed strong and constitutive nuclear factor (NF)-{kappa}B activity driven by I{kappa}B kinase compared with their parental cell lines St-4, A549, and HT-29. A new NF-{kappa}B inhibitor, dehydroxymethylepoxyquinomicin (DHMEQ), reduced viability and induced apoptosis in St-4/CPT, A549/CPT, and HT-29/CPT cell lines, while their parental cell lines were resistant to DHMEQ. The results in this study present an example of the shift in signals that support the survival of solid tumor cells to NF-{kappa}B during the acquisition of resistance to CPT. The results also indicate that solid tumor cells that become resistant to chemotherapy may be more easily treated by NF-{kappa}B inhibitors.

  16. Cytotoxic activity of some marine brown algae against cancer cell lines

    Directory of Open Access Journals (Sweden)

    MAHNAZ KHANAVI

    2010-01-01

    Full Text Available The aim of this study was to investigate the in vitro cytotoxic activity of total extract of MeOH (70% and partition fractions of hexan, chloroform (CHCL3, ethylacetate (EtOAc and MeOH-H2O of brown algae species (Sargassum swartzii, Cystoseira myrica, Colpomenia sinuosa found in the Persian Gulf against in different cell lines including HT-29, Caco-2, T47D, MDA-MB468 and NIH 3T3 cell lines by MTT and AnnexinV-PI assay. The hexan fraction of S. swartzii and C. myrica showed selective cytotoxicity against proliferation of Caco-2 cells (IC50<100 μg/ml T47D cell line (IC50<100 μg/ml, respectively. S. swartzii and C. myrica were also observed for increasing apoptosis in Caco-2 and T47D cells. Total extract and fractions of C. sinuosa did not show any significant cytotoxicity against the studied cell lines. MDA-MB468 cells were more sensitive to C. myrica than was T47D (IC50 99.9±8.11 vs. 56.50‘± 0.88. This reflects an estrogen receptor independent mechanism for cytotoxicity of the extract. The IC50 of the hexan fraction of C. myrica on T47D parent cells was lower than it was on T47D-TR cells (IC50 99.9±8.11 vs. 143.15 ± 7.80. This finding suggests a role for the MDR-1 in the development of possible future tolerance to the extract.

  17. Radiosensitivity of mesothelioma cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Haekkinen, A.M. [Dept. of Oncology, Univ. Central Hospital, Helsinki (Finland); Laasonen, A. [Dept. of Pathology, Central Hospital of Etelae-Pohjanmaa, Seinaejoki (Finland); Linnainmaa, K. [Dept. of Industrial Hygiene and Toxicology, Inst. of Occupational Health, Helsinki (Finland); Mattson, K. [Dept. Pulmonary Medicine, Univ. Central Hospital, Helsinki (Finland); Pyrhoenen, S. [Dept. of Oncology, Univ. Central Hospital, Helsinki (Finland)

    1996-10-01

    The present study was carried out in order to examine the radiosensitivity of malignant pleural mesothelioma cell lines. Cell kinetics, radiation-induced delay of the cell cycle and DNA ploidy of the cell lines were also determined. For comparison an HeLa and a human foetal fibroblast cell line were simultaneously explored. Six previously cytogenetically and histologically characterized mesothelioma tumor cell lines were applied. A rapid tiazolyl blue microtiter (MTT) assay was used to analyze radiosensitivity and cell kinetics and DNA ploidy of the cultured cells were determined by flow cytometry. The survival fraction after a dose of 2 Gy (SF2), parameters {alpha} and {beta} of the linear quadratic model (LQ-model) and mean inactivation dose (D{sub MID}) were also estimated. The DNA index of four cell lines equaled 1.0 and two cell lines equaled 1.5 and 1.6. Different mesothelioma cell lines showed a great variation in radiosensitivity. Mean survival fraction after a radiation dose of 2 Gy (SF2) was 0.60 and ranged from 0.36 to 0.81 and mean {alpha} value was 0.26 (range 0.48-0.083). The SF2 of the most sensitive diploid mesothelioma cell line was 0.36: Less than that of the foetal fibroblast cell line (0.49). The survival fractions (0.81 and 0.74) of the two most resistant cell lines, which also were aneuploid, were equal to that of the HeLa cell line (0.78). The {alpha}/{beta} ratios of the most sensitive cell lines were almost an order of magnitude greater than those of the two most resistant cell lines. Radiation-induced delay of the most resistant aneuploid cell line was similar to that of HeLa cells but in the most sensitive (diploid cells) there was practically no entry into the G1 phase following the 2 Gy radiation dose during 36 h. (orig.).

  18. The Acetone Extract of Sclerocarya birrea (Anacardiaceae) Possesses Antiproliferative and Apoptotic Potential against Human Breast Cancer Cell Lines (MCF-7)

    Science.gov (United States)

    Tanih, Nicoline Fri; Ndip, Roland Ndip

    2013-01-01

    Interesting antimicrobial data from the stem bark of Sclerocarya birrea, which support its use in traditional medicine for the treatment of many diseases, have been delineated. The current study was aimed to further study some pharmacological and toxicological properties of the plant to scientifically justify its use. Anticancer activity of water and acetone extracts of S. birrea was evaluated on three different cell lines, HT-29, HeLa, and MCF-7 using the cell titre blue viability assay in 96-well plates. Apoptosis was evaluated using the acridine orange and propidium iodide staining method, while morphological structure of treated cells was examined using SEM. The acetone extract exhibited remarkable antiproliferative activities on MCF-7 cell lines at dose- and time-dependent manners (24 h and 48 h of incubation). The extract also exerted apoptotic programmed cell death in MCF-7 cells with significant effect on the DNA. Morphological examination also displayed apoptotic characteristics in the treated cells, including clumping, condensation, and culminating to budding of the cells to produce membrane-bound fragmentation, as well as formation of apoptotic bodies. The acetone extract of S. birrea possesses antiproliferative and apoptotic potential against MCF-7-treated cells and could be further exploited as a potential lead in anticancer therapy. PMID:23576913

  19. Evaluation of drug permeation under fed state conditions using mucus-covered Caco-2 cell epithelium

    DEFF Research Database (Denmark)

    Birch, Ditlev; Diedrichsen, Ragna G; Christophersen, Philip C

    2018-01-01

    The absence of a surface-lining mucus layer is a major pitfall for the Caco-2 epithelial model. However, this can be alleviated by applying biosimilar mucus (BM) to the apical surface of the cell monolayer, thereby constructing a mucosa mimicking in vivo conditions. This study aims to elucidate...... the influence of BM as a barrier towards exogenic compounds such as permeation enhancers, and components of fed state simulated intestinal fluid (FeSSIF). Caco-2 cell monolayers surface-lined with BM were exposed to several compounds with distinct physicochemical properties, and the cell viability...... and permeability of the cell monolayer was compared to that of cell monolayers without BM and well-established mucus-secreting epithelial models (HT29 monolayers and HT29/Caco-2 co-culture monolayers). Exposure of BM-covered cells to constituents from FeSSIF revealed that it comprised a strong, hydrophilic barrier...

  20. Low-dose effects of Bisphenol A on human primary vascular endothelial cells and colon cancer cells

    OpenAIRE

    Varandas, Edna Soraia Gregório Ribeiro Varandas

    2014-01-01

    Doutoramento em Biologia - Instituto Superior de Agronomia Bisphenol A (BPA) is an extensively utilized endocrine disruptor for which human exposure is considered generalized through ingestion. Information regarding BPA effects on vascular and digestive tract tissues is scarce. Therefore, in this work primary Human Umbilical Vein Endothelial Cells (HUVEC) and human colon adenocarcinona cell line HT29 were used to evaluate BPA effects at two distinct low-dose concentrations r...

  1. Revisiting the thiosemicarbazonecopper(II) reaction with glutathione. Activity against colorectal carcinoma cell lines.

    Science.gov (United States)

    García-Tojal, Javier; Gil-García, Rubén; Fouz, Víctor Ivo; Madariaga, Gotzon; Lezama, Luis; Galletero, María S; Borrás, Joaquín; Nollmann, Friederike I; García-Girón, Carlos; Alcaraz, Raquel; Cavia-Saiz, Mónica; Muñiz, Pilar; Palacios, Òscar; Samper, Katia G; Rojo, Teófilo

    2018-03-01

    Thiosemicarbazones (TSCs), and their copper derivatives, have been extensively studied mainly due to the potential applications as antitumor compounds. A part of the biological activity of the TSC-Cu II complexes rests on their reactivity against cell reductants, as glutathione (GSH). The present paper describes the structure of the [Cu(PTSC)(ONO 2 )] n compound (1) (HPTSC=pyridine-2-carbaldehyde thiosemicarbazone) and its spectroscopic and magnetic properties. ESI studies performed on the reaction of GSH with 1 and the analogous [{Cu(PTSC*)(ONO 2 )} 2 ] derivative (2, HPTSC*=pyridine-2-carbaldehyde 4N-methylthiosemicarbazone) show the absence of peaks related with TSC-Cu-GSH species. However GSH-Cu ones are detected, in good agreement with the release of Cu I ions after reduction in the experimental conditions. The reactivity of 1 and 2 with cytochrome c and myoglobin and their activities against HT-29 and SW-480 colon carcinoma cell lines are compared with those shown by the free HPTSC and HPTSC* ligands. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Loratadine dysregulates cell cycle progression and enhances the effect of radiation in human tumor cell lines

    International Nuclear Information System (INIS)

    Soule, Benjamin P; Simone, Nicole L; DeGraff, William G; Choudhuri, Rajani; Cook, John A; Mitchell, James B

    2010-01-01

    The histamine receptor-1 (H1)-antagonist, loratadine has been shown to inhibit growth of human colon cancer xenografts in part due to cell cycle arrest in G2/M. Since this is a radiation sensitive phase of the cell cycle, we sought to determine if loratadine modifies radiosensitivity in several human tumor cell lines with emphasis on human colon carcinoma (HT29). Cells were treated with several doses of loratadine at several time points before and after exposure to radiation. Radiation dose modifying factors (DMF) were determined using full radiation dose response survival curves. Cell cycle phase was determined by flow cytometry and the expression of the cell cycle-associated proteins Chk1, pChk1 ser345 , and Cyclin B was analyzed by western blot. Loratadine pre-treatment of exponentially growing cells (75 μM, 24 hours) increased radiation-induced cytotoxicity yielding a radiation DMF of 1.95. However, treatment of plateau phase cells also yielded a DMF of 1.3 suggesting that mechanisms other than cell cycle arrest also contribute to loratadine-mediated radiation modification. Like irradiation, loratadine initially induced G2/M arrest and activation of the cell-cycle associated protein Chk1 to pChk1 ser345 , however a subsequent decrease in expression of total Chk1 and Cyclin B correlated with abrogation of the G2/M checkpoint. Analysis of DNA repair enzyme expression and DNA fragmentation revealed a distinct pattern of DNA damage in loratadine-treated cells in addition to enhanced radiation-induced damage. Taken together, these data suggest that the observed effects of loratadine are multifactorial in that loratadine 1) directly damages DNA, 2) activates Chk1 thereby promoting G2/M arrest making cells more susceptible to radiation-induced DNA damage and, 3) downregulates total Chk1 and Cyclin B abrogating the radiation-induced G2/M checkpoint and allowing cells to re-enter the cell cycle despite the persistence of damaged DNA. Given this unique possible

  3. Loratadine dysregulates cell cycle progression and enhances the effect of radiation in human tumor cell lines

    Directory of Open Access Journals (Sweden)

    Cook John A

    2010-02-01

    Full Text Available Abstract Background The histamine receptor-1 (H1-antagonist, loratadine has been shown to inhibit growth of human colon cancer xenografts in part due to cell cycle arrest in G2/M. Since this is a radiation sensitive phase of the cell cycle, we sought to determine if loratadine modifies radiosensitivity in several human tumor cell lines with emphasis on human colon carcinoma (HT29. Methods Cells were treated with several doses of loratadine at several time points before and after exposure to radiation. Radiation dose modifying factors (DMF were determined using full radiation dose response survival curves. Cell cycle phase was determined by flow cytometry and the expression of the cell cycle-associated proteins Chk1, pChk1ser345, and Cyclin B was analyzed by western blot. Results Loratadine pre-treatment of exponentially growing cells (75 μM, 24 hours increased radiation-induced cytotoxicity yielding a radiation DMF of 1.95. However, treatment of plateau phase cells also yielded a DMF of 1.3 suggesting that mechanisms other than cell cycle arrest also contribute to loratadine-mediated radiation modification. Like irradiation, loratadine initially induced G2/M arrest and activation of the cell-cycle associated protein Chk1 to pChk1ser345, however a subsequent decrease in expression of total Chk1 and Cyclin B correlated with abrogation of the G2/M checkpoint. Analysis of DNA repair enzyme expression and DNA fragmentation revealed a distinct pattern of DNA damage in loratadine-treated cells in addition to enhanced radiation-induced damage. Taken together, these data suggest that the observed effects of loratadine are multifactorial in that loratadine 1 directly damages DNA, 2 activates Chk1 thereby promoting G2/M arrest making cells more susceptible to radiation-induced DNA damage and, 3 downregulates total Chk1 and Cyclin B abrogating the radiation-induced G2/M checkpoint and allowing cells to re-enter the cell cycle despite the persistence of

  4. Differential expression of nanog1 and nanogp8 in colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Ishiguro, Tatsuya; Sato, Ai; Ohata, Hirokazu; Sakai, Hiroaki [Division of Cancer Differentiation, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Nakagama, Hitoshi, E-mail: hnakagam@ncc.go.jp [Division of Cancer Development System, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Okamoto, Koji, E-mail: kojokamo@ncc.go.jo [Division of Cancer Differentiation, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan)

    2012-02-10

    Highlights: Black-Right-Pointing-Pointer Nanog is expressed in a majority of colon cancer cell lines examined. Black-Right-Pointing-Pointer Both nanog1 and nanogp8 are expressed in colon cancer cells with varying ratios. Black-Right-Pointing-Pointer Nanog mediates cell proliferation of colon cancer cells. Black-Right-Pointing-Pointer Nanog predominantly localizes in cytoplasm of colon cancer cells. -- Abstract: Nanog, a homeodomain transcription factor, is an essential regulator for promotion of self-renewal of embryonic stem cells and inhibition of their differentiation. It has been demonstrated that nanog1 as well as nanogp8, a retrogene of nanog1, is preferentially expressed in advanced stages of several types of cancer, suggesting their involvement during cancer progression. Here, we investigated the expression of Nanog in well-characterized colon cancer cell lines. Expression of Nanog was detectable in 5 (HCT116, HT29, RKO, SW48, SW620) out of seven cell lines examined. RNA expression analyses of nanog1 and nanogp8 indicated that, while nanog1 was a major form in SW620 as well as in teratoma cells Tera-2, nanogp8 was preferentially expressed in HT29 and HCT116. In accordance with this, shRNA-mediated knockdown of nanog1 caused the reduction of Nanog in SW620 but not in HT29. Inhibition of Nanog in SW620 cells negatively affected cell proliferation and tumor formation in mouse xenograft. Biochemical subcellular fractionation and immunostaining analyses revealed predominant localization of Nanog in cytoplasm in SW620 and HT29, while it was mainly localized in nucleus in Tera-2. Our data indicate that nanog1 and nanogp8 are differentially expressed in colon cancer cells, and suggest that their expression contributes to proliferation of colon cancer cells.

  5. Differential expression of nanog1 and nanogp8 in colon cancer cells

    International Nuclear Information System (INIS)

    Ishiguro, Tatsuya; Sato, Ai; Ohata, Hirokazu; Sakai, Hiroaki; Nakagama, Hitoshi; Okamoto, Koji

    2012-01-01

    Highlights: ► Nanog is expressed in a majority of colon cancer cell lines examined. ► Both nanog1 and nanogp8 are expressed in colon cancer cells with varying ratios. ► Nanog mediates cell proliferation of colon cancer cells. ► Nanog predominantly localizes in cytoplasm of colon cancer cells. -- Abstract: Nanog, a homeodomain transcription factor, is an essential regulator for promotion of self-renewal of embryonic stem cells and inhibition of their differentiation. It has been demonstrated that nanog1 as well as nanogp8, a retrogene of nanog1, is preferentially expressed in advanced stages of several types of cancer, suggesting their involvement during cancer progression. Here, we investigated the expression of Nanog in well-characterized colon cancer cell lines. Expression of Nanog was detectable in 5 (HCT116, HT29, RKO, SW48, SW620) out of seven cell lines examined. RNA expression analyses of nanog1 and nanogp8 indicated that, while nanog1 was a major form in SW620 as well as in teratoma cells Tera-2, nanogp8 was preferentially expressed in HT29 and HCT116. In accordance with this, shRNA-mediated knockdown of nanog1 caused the reduction of Nanog in SW620 but not in HT29. Inhibition of Nanog in SW620 cells negatively affected cell proliferation and tumor formation in mouse xenograft. Biochemical subcellular fractionation and immunostaining analyses revealed predominant localization of Nanog in cytoplasm in SW620 and HT29, while it was mainly localized in nucleus in Tera-2. Our data indicate that nanog1 and nanogp8 are differentially expressed in colon cancer cells, and suggest that their expression contributes to proliferation of colon cancer cells.

  6. New polyhydroxylated sterols from Palythoa tuberculosa and their apoptotic activity in cancer cells.

    Science.gov (United States)

    Elbagory, Abdulrahman M; Meyer, Mervin; Ali, Abdel-Hamid A M; Ameer, Farouk; Parker-Nance, Shirley; Benito, Maria Teresa; Doyagüez, Elisa Garcia; Jimeno, Maria Luisa; Hussein, Ahmed A

    2015-09-01

    The chemical study on the total extract of the zoanthid Palythoa tuberculosa, collected from the Red Sea, resulted in the isolation of seven polyhydroxylated sterols (1-7), six of which, palysterols A-F (2-7), are new. Their chemical structures were elucidated on the basis of extensive analysis of their 1-, 2D NMR and MS spectroscopic data. This is the first chemical investigation on the species collected from Red Sea. We studied the cytotoxic effects of the total extract and some of the new polyhydroxylated sterols in three human cancer cell lines (MCF-7, HeLa, and HT-29) and one non-cancerous human cell line (KMST-6). Palysterol F (7), in particular, was able to selectively induce high levels of apoptosis (>75%) in breast adenocarcinoma (MCF-7) cells but not HeLa, HT-29 and KMST-6 cells. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Peroxisome proliferator-activated receptor gamma overexpression suppresses proliferation of human colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Tsukahara, Tamotsu, E-mail: ttamotsu@shinshu-u.ac.jp [Department of Integrative Physiology and Bio-System Control, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 (Japan); Haniu, Hisao [Department of Orthopaedic Surgery, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 (Japan)

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer We examined the correlation between PPAR{gamma} expression and cell proliferation. Black-Right-Pointing-Pointer PPAR{gamma} overexpression reduces cell viability. Black-Right-Pointing-Pointer We show the synergistic effect of cell growth inhibition by a PPAR{gamma} agonist. -- Abstract: Peroxisome proliferator-activated receptor gamma (PPAR{gamma}) plays an important role in the differentiation of intestinal cells and tissues. Our previous reports indicate that PPAR{gamma} is expressed at considerable levels in human colon cancer cells. This suggests that PPAR{gamma} expression may be an important factor for cell growth regulation in colon cancer. In this study, we investigated PPAR{gamma} expression in 4 human colon cancer cell lines, HT-29, LOVO, DLD-1, and Caco-2. Real-time polymerase chain reaction (PCR) and Western blot analysis revealed that the relative levels of PPAR{gamma} mRNA and protein in these cells were in the order HT-29 > LOVO > Caco-2 > DLD-1. We also found that PPAR{gamma} overexpression promoted cell growth inhibition in PPAR{gamma} lower-expressing cell lines (Caco-2 and DLD-1), but not in higher-expressing cells (HT-29 and LOVO). We observed a correlation between the level of PPAR{gamma} expression and the cells' sensitivity for proliferation.

  8. Synergistic cytotoxicity against human tumor cell lines by oncolytic adenovirus dl1520 (ONYX-015) and melphalan.

    Science.gov (United States)

    Ferguson, Peter J; Sykelyk, Alexander; Figueredo, Rene; Koropatnick, James

    2016-01-01

    In light of the need for more selective anticancer therapy, much work has been directed at developing compounds or biological agents that target functions specific to cancer cells. To this end, numerous viruses have been engineered to exploit the dependence of cancer cells on particular anomalies that contribute to their rogue proliferative activity, such as dysfunctional p53, overactive mitogenic signaling, or a defective interferon response. The oncolytic human adenovirus dl1520 (ONYX-015) was engineered to propagate specifically in p53-deficient tumors, which comprise over half of all tumors. Based on successes in clinical trials, the full potential of dl1520 and other oncolytic viruses may be even better realized by using them in combination with conventional chemotherapy drugs. As a model system in which to test this potential, representative cell lines from 2 common cancer types, oral squamous cell carcinoma (HN-5a) and colon adenocarcinoma (HT-29), were chosen, as well as platinum-drug-resistant variants of each. Following preliminary screening of virus and drug combinations, dl1520 and melphalan were found to synergistically inhibit proliferation of all the cancer cell lines. Melphalan pretreatment or cotreatment with dl1520 enhanced inhibition of proliferation by dl1520 by up to 60% and increased apoptosis by up to 25%. The tight-junction protein CAR (coxsackie and adenovirus receptor), via which adenovirus enters cells, was not upregulated by treatment with melphalan, suggesting that other mechanisms contribute to synergy. The synergy between melphalan and dl1520 suggests that tumor-selective cell killing by oncolytic viruses may be augmented by combining with cytotoxic drugs.

  9. Molecular Mechanism of Enhanced Anticancer Effect of Nanoparticle Formulated LY2835219 via p16-CDK4/6-pRb Pathway in Colorectal Carcinoma Cell Line

    Directory of Open Access Journals (Sweden)

    Xu Tang

    2016-01-01

    Full Text Available LY2835219 is a dual inhibitor to CDK4 and CDK6. This study was to prepare LY2835219-loaded chitosan nanoparticles (CNP/LY and LY2835219-loaded hyaluronic acid-conjugated chitosan nanoparticles (HACNP/LY and revealed their anticancer effect and influence on p16-CDK4/6-pRb pathway against colon cell line. The nanoparticle sizes of CNP/LY and HACNP/LY were approximately 195±39.6 nm and 217±31.1 nm, respectively. The zeta potentials of CNP/LY and HACNP/LY were 37.3±1.5 mV and 30.3±2.2 mV, respectively. And the preparation process showed considerable drug encapsulation efficiency and loading efficiency. LY2835219, CNP/LY, and HACNP/LY inhibited HT29 cell proliferation with 0.68, 0.54, and 0.30 μM of IC50, respectively. G1 phase was arrested by LY2835219 and its formulations. Furthermore, inhibition of CDK4/6 by LY2835219 formulations induced CDK4, CDK6, cyclin D1, and pRb decrease and p16 increase at both protein and mRNA levels. Overall, nanoparticle formulated LY2835219 could enhance the cytotoxicity and cell cycle arrest, and HACNP/LY strengthened the trend furtherly compared to CNP/LY. It is the first time to demonstrate the anticancer effect and mechanism against HT29 by LY2835219 and its nanoparticles. The drug and its nanoparticle formulations delay the cell growth and arrest cell cycle through p16-CDK4/6-pRb pathway, while the nanoparticle formulated LY2835219 could strengthen the process.

  10. [Construction of γ-synuclein eukaryotic expression vector and its effect on invasion and metastasis of colon cancer cell line SW1116 in vitro].

    Science.gov (United States)

    Ye, Qing; Huang, Feng; Zheng, Qiuhong; Wang, Xiaoying; Xu, Yangmei; Gong, Fusheng; Huang, Lijie

    2014-01-01

    To construct γ-synuclein gene eukaryotic expression vector, and to study its effect on the invasion of colon cancer cell line SW1116 and the adhesion between SW1116 and human umbilical vein endothelial cells(HUVECs) in vitro. Total RNA was extracted from colon cancer cell line HT29 and the cDNA of γ-synuclein was amplified using RT-PCR. The digested fragment of cDNA coding sequence was linked to the eukaryotic expression vector pEGFP-C1 containing the GFP gene. After identification by sequence analysis, the recombinant plasmid was transfected into colon cancer cell line SW1116 via lipofectamine. The stable cell line was selected with G-418. The invasion in vitro was tested by Transwell invasion chamber assay. HUVECs were previously seeded onto 96-well plates before SW1116 cells seeded, and fluorescence intensity of GFP was detected to represent the amount of adhesion cells by ELISA. Human γ-synuclein eukaryotic expression vector was successfully constructed, which was stably expressed in SW1116 cells and could translate the GFP-γ-synuclein protein in vitro. γ-synuclein facilitated SW1116 cell passing through matrigel and filter membrane(198.4±20.7 vs. 98.8±13.2, Pcells to HUVECs(3.08±0.36 vs. 1.22±0.21, Pcell SW1116 potentiality of invasion and metastasis in vitro.

  11. Murine endothelioma cell lines transformed by polyoma middle T oncogene as target for and producers of cytokines.

    Science.gov (United States)

    Bussolino, F; De Rossi, M; Sica, A; Colotta, F; Wang, J M; Bocchietto, E; Padura, I M; Bosia, A; DeJana, E; Mantovani, A

    1991-10-01

    We studied cytokine-related functional properties of four mouse endotheliomas from different anatomical sites obtained by transformation with middle T oncogene. We examined mRNA expression of IL-6, IL-1 alpha, macrophage-CSF, granulocyte/macrophage-CSF, and two members of an emerging super-family of chemotactic cytokines (JE/monocyte chemoattractant protein-1 (MCP-1) and KC). Exposure to IL-1 augmented or induced cytokine gene transcripts in three endothelioma lines (eEnd.1, sEnd.1, and tEnd) with maximal expression in tEnd.1 cells. Endothelioma cells also responded to TNF-alpha and LPS. Levels of IL-6 and monocyte chemotactic activity (a JE/MCP activity) correlated with mRNA expression. IL-1 also induced production of procoagulant activity and platelet-activating factor in endothelioma cells, with heterogeneity in the levels of response among individuals lines. Murine melanoma B16-F1, human colon carcinoma HT29 cells, CB33MT lymphoblastoid cells, and monocytes adhered to endothelioma monolayers and the adhesive properties of these cell lines were modulated by IL-1 beta, with marked differences among themselves. Murine EC derived from brain capillaries, used as control, shared several properties with bEnd.4 line. Endothelioma lines cause tumors by recruiting host cells. The capacity to produce cytokines that directly or indirectly attract host vascular cells, may play an important role in hemangioma induction in vivo. Murine endothelioma lines, generated by transformation with the polyoma middle T oncogene, retain functional properties of normal endothelium, and may represent an invaluable tool for analysis of the immunobiology and heterogeneity of EC in different tissues.

  12. Synsepalum dulcificum extracts exhibit cytotoxic activity on human colorectal cancer cells and upregulate c-fos and c-jun early apoptotic gene expression

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    Jichang Seong

    2018-01-01

    Full Text Available Objective: To explore cytotoxicity of Synsepalum dulcificum (S. dulcificum Daniell (Sapotaceae on human colon cancer (HCT-116 and HT-29, human monocytic leukemia (THP-1 and normal (HDFn cell lines, and its effect on the expression of early apoptotic genes, c-fos and c-jun. Methods: Leaf, stem and berry of S. dulcificum were separately extracted by using 2 solvents, 10% ethanol (EtOH and 80% methanol (MeOH. PrestoBlue® cell viability assay and qRT-PCR assay were conducted to examine the above objectives respectively. Results: Stem MeOH, stem EtOH, and berry EtOH extracts of S. dulcificum were cytotoxic to HCT-116 and HT-29 human colon cancer cells. For HCT-116, IC50 values of these 3 extracts were not significantly different (P>0.05 from that of the positive control bleomycin (IC50 of 33.57 μg/mL, while for HT-29, IC50 values of these 3 extracts were significantly lower (P<0.05 than that of bleomycin (IC50 of 25.24 μg/mL. None of the extracts were cytotoxic to the THP-1 monocytic leukemia cells and HDFn normal human dermal fibroblasts. For both HCT-116 and HT-29, these extracts significantly up-regulated (P<0.05 the expression of c-fos and c-jun compared to the untreated negative control. Conclusions: The results of this study suggest that cytotoxicity of stem MeOH, stem EtOH, and berry EtOH extracts of S. dulcificum on HCT-116 and HT-29 colon cancer cells is due to the induced apoptosis which is caused by the up-regulation of the expression of early apoptotic genes, c-fos and c-jun.

  13. Evaluation of chemical composition, antioxidant, antibacterial, cytotoxic and apoptotic effects of Aloysia citrodora extract on colon cancer cell line

    Directory of Open Access Journals (Sweden)

    Amir Mirzaie

    2016-06-01

    Full Text Available Background: Aloysia citrodora belongs to the Verbenaceae family of plants, a well-known herbal medicine in Iran. The aim of the present study was to investigate the chemical composition, antioxidant, antibacterial, cytotoxic and apoptotic effect of A. citrodora extract against human colon cancer (HT29 cells by using real-time polymerase chain reaction and flow-cytometry methods. Methods: This experimental study was carried out in Islamic Azad University, East Tehran Branch, from March to September of 2014. At first, the A. citrodora chemical constituents were analyzed by gas chromatography-mass spectrometry (GC-MS technique. In addition, antioxidant assay, antibacterial and anti-cancer effect was performed using 1,1-diphenyl-2-picrylhydrazyl (DPPH, disk diffusion and 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT methods, respectively. The half maximal inhibitory concentration (IC50 value was calculated. We extracted total RNA molecules by using RNX solution, after which cDNA was synthesized. Finally, the pro-apoptotic (Bax and anti-apoptotic (Bcl2 gene expression was performed by real-time polymerase chain reaction and apoptotic effects were analyzed using Flow-cytometry method. Results: GC-MS analysis of Aloysia citrodora extract was shown 37 major components and the most frequent component was belonged to Spathulenol (17.57% and Caryophyllene oxide (15.15% The antioxidant activity of the extract was IC50= 0.6±0.03 mg/ml. The maximum and minimum antibacterial effects of extract were belonged to gram-negative and gram-positive bacteria, respectively. Cytotoxic results revealed that the A.citrodora extract have IC50= 20.1±0.78 mg/ml against colon cancer (HT29 cell line and real-time polymerase chain reaction results showed the expression level of Bax and Bcl2 was increased and decreased respectively in colon cancer cell line (3.470±0.72 (P< 0.05, 0.43±0.35 (P< 0.05. In addition, the flow-cytometry results indicated the 38

  14. Matrigel Basement Membrane Matrix influences expression of microRNAs in cancer cell lines

    International Nuclear Information System (INIS)

    Price, Karina J.; Tsykin, Anna; Giles, Keith M.; Sladic, Rosemary T.; Epis, Michael R.; Ganss, Ruth; Goodall, Gregory J.; Leedman, Peter J.

    2012-01-01

    Highlights: ► Matrigel alters cancer cell line miRNA expression relative to culture on plastic. ► Many identified Matrigel-regulated miRNAs are implicated in cancer. ► miR-1290, -210, -32 and -29b represent a Matrigel-induced miRNA signature. ► miR-32 down-regulates Integrin alpha 5 (ITGA5) mRNA. -- Abstract: Matrigel is a medium rich in extracellular matrix (ECM) components used for three-dimensional cell culture and is known to alter cellular phenotypes and gene expression. microRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression and have roles in cancer. While miRNA profiles of numerous cell lines cultured on plastic have been reported, the influence of Matrigel-based culture on cancer cell miRNA expression is largely unknown. This study investigated the influence of Matrigel on the expression of miRNAs that might facilitate ECM-associated cancer cell growth. We performed miRNA profiling by microarray using two colon cancer cell lines (SW480 and SW620), identifying significant differential expression of miRNAs between cells cultured in Matrigel and on plastic. Many of these miRNAs have previously been implicated in cancer-related processes. A common Matrigel-induced miRNA signature comprised of up-regulated miR-1290 and miR-210 and down-regulated miR-29b and miR-32 was identified using RT-qPCR across five epithelial cancer cell lines (SW480, SW620, HT-29, A549 and MDA-MB-231). Experimental modulation of these miRNAs altered expression of their known target mRNAs involved in cell adhesion, proliferation and invasion, in colon cancer cell lines. Furthermore, ITGA5 was identified as a novel putative target of miR-32 that may facilitate cancer cell interactions with the ECM. We propose that culture of cancer cell lines in Matrigel more accurately recapitulates miRNA expression and function in cancer than culture on plastic and thus is a valuable approach to the in vitro study of miRNAs.

  15. The novel anthraquinone derivative IMP1338 induces death of human cancer cells by p53-independent S and G2/M cell cycle arrest.

    Science.gov (United States)

    Choi, Hyun Kyung; Ryu, Hwani; Son, A-Rang; Seo, Bitna; Hwang, Sang-Gu; Song, Jie-Young; Ahn, Jiyeon

    2016-04-01

    To identify novel small molecules that induce selective cancer cell death, we screened a chemical library containing 1040 compounds in HT29 colon cancer and CCD18-Co normal colon cells, using a phenotypic cell-based viability assay system with the Cell Counting Kit-8 (CCK-8). We discovered a novel anthraquinone derivative, N-(4-[{(9,10-dioxo-9,10-dihydro-1-anthracenyl)sulfonyl}amino]phenyl)-N-methylacetamide (IMP1338), which was cytotoxic against the human colon cancer cells tested. The MTT cell viability assay showed that treatment with IMP1338 selectively inhibited HCT116, HCT116 p53(-/-), HT29, and A549 cancer cell proliferation compared to that of Beas2B normal epithelial cells. To elucidate the cellular mechanism underlying the cytotoxicity of IMP1338, we examined the effect of IMP1338 on the cell cycle distribution and death of cancer cells. IMP1338 treatment significantly arrested the cell cycle at S and G2/M phases by DNA damage and led to apoptotic cell death, which was determined using FACS analysis with Annexin V/PI double staining. Furthermore, IMP1338 increased caspase-3 cleavage in wild-type p53, p53 knockout HCT116, and HT29 cells as determined using immunoblotting. In addition, IMP1338 markedly induced the phosphorylation of histone H2AX and Chk1 in both cell lines while the combination of 5-fluorouracil (5-FU) and radiation inhibited the viability of HCT116, HCT116 p53(-/-), and HT29 cells compared to 5-FU or radiation alone. Our findings indicated that IMP1338 induced p53-independent cell death through S and G2/M phase arrest as well as DNA damage. These results provide a basis for future investigations assessing the promising anticancer properties of IMP1338. Copyright © 2016. Published by Elsevier Masson SAS.

  16. Calorimetric signatures of human cancer cells and their nuclei

    Energy Technology Data Exchange (ETDEWEB)

    Todinova, S. [Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, Sofia 1113 (Bulgaria); Stoyanova, E. [Department of Molecular Immunology, Institute of Biology and Immunology of Reproduction, Bulgarian Academy of Sciences, Tzarigradsko shose Blvd. 73, Sofia 1113 (Bulgaria); Krumova, S., E-mail: sakrumo@gmail.com [Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, Sofia 1113 (Bulgaria); Iliev, I. [Institute of Experimental Morphology, Pathology and Anthropology with Museum, Acad. G. Bonchev Str., Bl. 25, Sofia 1113 (Bulgaria); Taneva, S.G. [Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, Sofia 1113 (Bulgaria)

    2016-01-10

    Graphical abstract: - Highlights: • Two temperature ranges are distinguished in the thermograms of cells/nuclei. • Different thermodynamic properties of cancer and normal human cells/nuclei. • Dramatic reduction of the enthalpy of the low-temperature range in cancer cells. • Oxaliplatin and 5-FU affect the nuclear matrix proteins and the DNA stability. - Abstract: The human cancer cell lines HeLa, JEG-3, Hep G2, SSC-9, PC-3, HT-29, MCF7 and their isolated nuclei were characterized by differential scanning calorimetry. The calorimetric profiles differed from normal human fibroblast (BJ) cells in the two well distinguished temperature ranges—the high-temperature range (H{sub T}, due to DNA-containing structures) and the low-temperature range (L{sub T}, assigned to the nuclear matrix and cellular proteins). The enthalpy of the L{sub T} range, and, respectively the ratio of the enthalpies of the L{sub T}- vs. H{sub T}-range, ΔH{sub L}/ΔH{sub H}, is strongly reduced for all cancer cells compared to normal fibroblasts. On the contrary, for most of the cancer nuclei this ratio is higher compared to normal nuclei. The HT-29 human colorectal cancer cells/nuclei differed most drastically from normal human fibroblast cells/nuclei. Our data also reveal that the treatment of HT-29 cancer cells with cytostatic drugs affects not only the DNA replication but also the cellular proteome.

  17. Azidothymidine and cisplatin increase p14ARF expression in OVCAR-3 ovarian cancer cell line

    International Nuclear Information System (INIS)

    Vaskivuo, Liisa; Rysae, Jaana; Koivuperae, Johanna; Myllynen, Paeivi; Vaskivuo, Tommi; Chvalova, Katerina; Serpi, Raisa; Savolainen, Eeva-Riitta; Puistola, Ulla; Vaehaekangas, Kirsi

    2006-01-01

    p14 ARF tumor suppressor protein regulates p53 by interfering with mdm2-p53 interaction. p14 ARF is activated in response to oncogenic stimuli but little is known of the responses of endogenous p14 ARF to different types of cellular stress or DNA damage. Azidothymidine (AZT) is being tested in several clinical trials as an enhancer of anticancer chemotherapy. However, the knowledge of the relationship between AZT and cellular pathways, e.g. p53 pathway, is very limited. In this study, we show that AZT, cisplatin (CDDP) and docetaxel (DTX) all induce unique molecular responses in OVCAR-3 ovarian carcinoma cells carrying a mutated p53, while in A2780, ovarian carcinoma and MCF-7 breast carcinoma cells with wild type p53, all of these drugs cause similar p53 responses. We found that endogenous p14 ARF protein in OVCAR-3 cells is down-regulated by DTX but induced by AZT and a short CDDP pulse treatment. In HT-29 colon carcinoma cells with a mutated p53, all treatments down-regulated p14 ARF protein. Both CDDP and AZT increased the expression of p14ARF mRNA in OVCAR-3 cells. Differences in cell death induced by these drugs did not explain the differences in protein and mRNA expressions. No increase in the level of either c-Myc or H-ras oncoproteins was seen in OVCAR-3 cells after AZT or CDDP-treatment. These results suggest that p14 ARF can respond to DNA damage without oncogene activation in cell lines without functional p53

  18. Thyroid cell lines in research on goitrogenesis.

    Science.gov (United States)

    Gerber, H; Peter, H J; Asmis, L; Studer, H

    1991-12-01

    Thyroid cell lines have contributed a lot to the understanding of goitrogenesis. The cell lines mostly used in thyroid research are briefly discussed, namely the rat thyroid cell lines FRTL and FRTL-5, the porcine thyroid cell lines PORTHOS and ARTHOS, The sheep thyroid cell lines OVNIS 5H and 6H, the cat thyroid cell lines PETCAT 1 to 4 and ROMCAT, and the human thyroid cell lines FTC-133 and HTh 74. Chinese hamster ovary (CHO) cells and COS-7 cells, stably transfected with TSH receptor cDNA and expressing a functional TSH receptor, are discussed as examples for non-thyroidal cells, transfected with thyroid genes.

  19. Different effects of two newly-isolated probiotic Lactobacillus plantarum 15HN and Lactococcus lactis subsp. Lactis 44Lac strains from traditional dairy products on cancer cell lines.

    Science.gov (United States)

    Haghshenas, Babak; Abdullah, Norhafizah; Nami, Yousef; Radiah, Dayang; Rosli, Rozita; Khosroushahi, Ahmad Yari

    2014-12-01

    Lactobacillus and Lactococcus strains isolated from food products can be introduced as probiotics because of their health-promoting characteristics and non-pathogenic nature. This study aims to perform the isolation, molecular identification, and probiotic characterization of Lactobacillus and Lactococcus strains from traditional Iranian dairy products. Primary probiotic assessments indicated high tolerance to low pH and high bile salt conditions, high anti-pathogenic activities, and susceptibility to high consumption antibiotics, thus proving that both strains possess probiotic potential. Cytotoxicity assessments were used to analyze the effects of the secreted metabolite on different cancer cell lines, including HT29, AGS, MCF-7, and HeLa, as well as a normal human cell line (HUVEC). Results showed acceptable cytotoxic properties for secreted metabolites (40 μg/ml dry weight) of Lactococcus lactis subsp. Lactis 44Lac. Such performance was similar to that of Taxol against all of the treated cancer cell lines; however, the strain exhibited no toxicity on the normal cell line. Cytotoxic assessments through flow cytometry and fluorescent microscopy demonstrated that apoptosis is the main cytotoxic mechanism for secreted metabolites of L. lactis subsp. Lactis 44Lac. By contrast, the effects of protease-treated metabolites on the AGS cell line verified the protein nature of anti-cancer metabolites. However, precise characterizations and in vitro/in vivo investigations on purified proteins should be conducted before these metabolites are introduced as potential anti-cancer therapeutics. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. Toxicity assessment of metoprolol and its photodegradation mixtures obtained by using different type of TiO{sub 2} catalysts in the mammalian cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Četojević-Simin, Dragana D., E-mail: ddaaggeerr@gmail.com [University of Novi Sad, Faculty of Medicine, Oncology Institute of Vojvodina, Dr Goldmana 4, 21204 Sremska Kamenica (Serbia); Armaković, Sanja J., E-mail: sanja.armakovic@dh.uns.ac.rs [University of Novi Sad, Faculty of Sciences, Department of Chemistry, Biochemistry and Environmental Protection, Trg D. Obradovića 3, 21000 Novi Sad (Serbia); Šojić, Daniela V., E-mail: daniela.sojic@dh.uns.ac.rs [University of Novi Sad, Faculty of Sciences, Department of Chemistry, Biochemistry and Environmental Protection, Trg D. Obradovića 3, 21000 Novi Sad (Serbia); Abramović, Biljana F., E-mail: biljana.abramovic@dh.uns.ac.rs [University of Novi Sad, Faculty of Sciences, Department of Chemistry, Biochemistry and Environmental Protection, Trg D. Obradovića 3, 21000 Novi Sad (Serbia)

    2013-10-01

    Toxicity of metoprolol (MET) alone and in mixtures with its photocatalytic degradation intermediates obtained by using TiO{sub 2} Wackherr and Degussa P25 under UV irradiation in the presence of O{sub 2} was evaluated in vitro in a panel of three histologically different cell lines: rat hepatoma (H-4-II-E), human colon adenocarcinoma (HT-29) and human fetal lung (MRC-5). Both catalysts promoted a time-dependent increase in the toxicity of the photodegradation products, and those obtained using Degussa P25 photocatalyst were more toxic. The most pronounced and selective toxic action of MET and products of its photodegradation was observed in the hepatic cell line. The higher toxicity of the mixtures obtained using Degussa P25 catalyst could be explained by a different mechanism of MET degradation, i.e. by the presence or higher concentrations of some intermediates. Although the concentrations of intermediates obtained using TiO{sub 2} Wackherr catalyst were higher, they did not affect significantly the growth of the examined cell lines, indicating their lower toxicity. This suggests that a treatment aiming at complete mineralization should be performed bearing in mind that the type of catalyst, the concentration of target molecule, and the duration of the process are significant factors that determine the nature and toxicity of the resulting mixtures. Although the EC{sub 50} values of MET obtained in mammalian cell lines were higher compared to the bioassays for lower trophic levels, the time-dependent promotion of toxicity of degradation mixtures should be attributed to the higher sensitivity of mammalian cell bioassays. - Highlights: • Toxicity study of metoprolol and its photocatalytic degradation mixtures • Toxicity evaluation in vitro in H-4-II-E, HT-29 and MRC-5 cell lines • TiO{sub 2} Wackherr and Degussa P25 promoted a time-dependent increase in toxicity. • The higher toxicity of degradation mixtures obtained using Degussa P25 • Most pronounced and

  1. Molecular mechanisms of apoptosis and cell selectivity of zinc dithiocarbamates functionalized with hydroxyethyl substituents.

    Science.gov (United States)

    Tan, Yee Seng; Ooi, Kah Kooi; Ang, Kok Pian; Akim, Abdah Md; Cheah, Yoke-Kqueen; Halim, Siti Nadiah Abdul; Seng, Hoi-Ling; Tiekink, Edward R T

    2015-09-01

    In the solid state each of three binuclear zinc dithiocarbamates bearing hydroxyethyl groups, {Zn[S2CN(R)CH2CH2OH]2}2 for R = iPr (1), CH2CH2OH (2), and Me (3), and an all alkyl species, [Zn(S2CNEt2)2]2 (4), features a centrosymmetric {ZnSCS}2 core with a step topology; both 1 and 3 were isolated as monohydrates. All compounds were broadly cytotoxic, specifically against human cancer cell lines compared with normal cells, with greater potency than cisplatin. Notably, some selectivity were indicated with 2 being the most potent against human ovarian carcinoma cells (cisA2780), and 4 being more cytotoxic toward multidrug resistant human breast carcinoma cells (MCF-7R), human colon adenocarcinoma cells (HT-29), and human lung adenocarcinoma epithelial cells (A549). Based on human apoptosis PCR-array analysis, caspase activities, DNA fragmentation, cell apoptotic assays, intracellular reactive oxygen species (ROS) measurements and human topoisomerase I inhibition, induction of apoptosis in HT-29 cells is demonstrated via both extrinsic and intrinsic pathways. Compounds 2-4 activate the p53 gene while 1 activates both p53 and p73. Cell cycle arrest at the S and G2/M phases correlates with inhibition of HT-29 cell growth. Cell invasion is also inhibited by 1-4 which is correlated with down-regulation of NF-κB. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. Protein kinase Cγ in colon cancer cells: Expression, Thr514 phophorylation and sensitivity to butyrate-mediated upregulation as related to the degree of differentiation

    Science.gov (United States)

    Garczarczyk, Dorota; Szeker, Krisztina; Galfi, Peter; Csordas, Adam; Hofmann, Johann

    2010-01-01

    Protein kinase C (PKC) isoenzymes are expressed and activated in a cell type-specific manner, and play an essential role in tissue-specific signal transduction. The presence of butyrate at millimolar concentrations in the colon raises the question of whether it affects the expression of PKC isoenzymes in the different cell types of the colonic epithelium. We investigated the protein expression levels of PKCγ, Thr514-phosphorylated PKCγ (pPKCγ-Thr514), and their subcellular distribution as affected by butyrate in a set of colon cancer cell lines. Thr514-phosphorylation of de novo synthesized PKCγ is the first step in priming of the inactive PKCγ before its release into the cytoplasm. For immunoblot analysis, we employed three antibodies, one against an unmodified sequence, mapping within 50 amino acids at its C-terminus, a second against pPKCγ-Thr514, and a third against pPKCγ-pan-Thr514. The antibody against an unmodified C-terminal peptide epitope did not recognize pPKCγ-Thr514, suggesting that phosphorylation at this site interferes with the binding of the antibody to the C-terminus. Marked butyrate-induced upregulation of PKCγ occurred in HT29 cells (colonocyte stem cells) and HT29-derived cell lines. However, in Caco2 and IEC-18 cells (differentiated intestinal epithelial cells), PKCγ was insensitive to upregulation, and present exclusively as pPKCγ-Thr514. Lovo and SW480 expressed higher levels of PKCγ. In HT29 cells, butyrate-induced upregulation of the non-phosphorylated PKCγ was observed in both the membrane and cytosolic fraction. In Caco2 cells, the Thr514-phosphorylated form was present at high levels in both fractions. The presence of unphosphorylated PKCγ in HT29 cells, and its complete absence in Caco2 cells demonstrates a cell type-dependent differential coupling of Thr514-phosphorylation with de novo synthesis of PKCγ in colon cancer cells. PMID:20188713

  3. Study of Alpha Tocopherol, Celecoxib Induced Apoptosis in Human Colorectal Carcinoma Cell Line

    Directory of Open Access Journals (Sweden)

    Solgui R

    2010-03-01

    Full Text Available Background and Objectives: Chronic, unbridled oxidative damages have been known as the culprits behind many chronic diseases, including cancers, atherosclerosis, diabetes and Alzheimer’s. Cyclooxygenase-2 (COX-2-the main enzyme involved in inducing these processes- plays an important role in tumor development and progression. COX-2 inhibitors should be used at high doses for a long time in order to bring about chemoprevention and induction of anti-tumor effects. For example, celecoxib prevents colorectal tumor growth and induces apoptosis in both in vitro and in vivo models. Disregulation of COX-2 expression coincides with the development of gastrointestinal malignancy in humans and in animal models of colorectal cancer. Increased COX-2 expression in human colorectal adeno-carcinomas has been elucidated when compared with normal adjacent colonic mucosa. The capacity of vitamin E, particularly in α form, to quench free radical damage, induces apoptosis and impact expression of oncogenes makes it an appropriate choice for chemotherapeutic strategies. Studies have shown that carcinogenesis and DNA damage due to UV are inhibited by vitamin E. The goal of this study was to investigate alpha tocopherol and celecoxib induced apoptosis in human colorectal carcinoma cell line.Methods: In this study, HT29 cells were exposed to different concentrations of tocopherol (5, 10, and 20µM and celecoxib (25, 50, 75, 100µM followed by DNA extraction and fragmentation for demonstrating cell death process. Results: The results indicated that celecoxib at lower doses (25, and 50µM could not induce cell death, but at higher doses (75, and 100 µM, DNA fragmentation results typically resembled programmed cell death.Conclusion: ocopherol (5, 10, and 20µM in combination with celecoxib improved the impact of celecoxib on cell death induction and made it the rational notion to be combined with vitamin E in clinical practice.

  4. Resveratrol suppresses human colon cancer cell proliferation and induces apoptosis via targeting the pentose phosphate and the talin-FAK signaling pathways-A proteomic approach

    Directory of Open Access Journals (Sweden)

    Reddivari Lavanya

    2011-08-01

    Full Text Available Abstract Background We and others have previously reported that resveratrol (RSV suppresses colon cancer cell proliferation and elevates apoptosis in vitro and/or in vivo, however molecular mechanisms are not fully elucidated. Particularly, little information is available on RSV's effects on metabolic pathways and the cell-extra cellular matrix (ECM communication that are critical for cancer cell growth. To identify important targets of RSV, we analyzed whole protein fractions from HT-29 advanced human colon cancer cell line treated with solvent control, IGF-1 (10 nM and RSV (150 μM using LC/MS/MS-Mud PIT (Multidimensional Protein Identification Technology. Results Pentose phosphate pathway (PPP, a vital metabolic pathway for cell cycle progression, was elevated and suppressed by IGF-1 and RSV, respectively in the HT-29 cell line. Enzymatic assays confirmed RSV suppression of glucose-6 phosphate dehydrogenase (rate limiting and transketolase, key enzymes of the PPP. RSV (150 μM suppressed, whereas IGF-1 (10 nM elevated focal adhesion complex (FAC proteins, talin and pFAK, critical for the cell-ECM communication. Western blotting analyses confirmed the suppression or elevation of these proteins in HT-29 cancer cells treated with RSV or IGF-1, respectively. Conclusions Proteomic analysis enabled us to establish PPP and the talin-pFAK as targets of RSV which suppress cancer cell proliferation and induce apoptosis in the colon cancer cell line HT-29. RSV (150 μM suppressed these pathways in the presence and absence of IGF-1, suggesting its role as a chemo-preventive agent even in obese condition.

  5. Chromatin status of apoptosis genes correlates with sensitivity to chemo-, immune- and radiation therapy in colorectal cancer cell lines.

    Science.gov (United States)

    Benard, Anne; Janssen, Connie M; van den Elsen, Peter J; van Eggermond, Marja C J A; Hoon, Dave S B; van de Velde, Cornelis J H; Kuppen, Peter J K

    2014-12-01

    The apoptosis pathway of programmed cell death is frequently deregulated in cancer. An intact apoptosis pathway is required for proper response to anti-cancer treatment. We investigated the chromatin status of key apoptosis genes in the apoptosis pathway in colorectal cancer cell lines in relation to apoptosis induced by chemo-, immune- or radiation therapy. Using chromatin immunoprecipitation (ChIP), we measured the presence of transcription-activating histone modifications H3Ac and H3K4me3 and silencing modifications H3K9me3 and H3K27me3 at the gene promoter regions of key apoptosis genes Bax, Bcl2, Caspase-9, Fas (CD95) and p53. Cell lines DLD1, SW620, Colo320, Caco2, Lovo and HT29 were treated with cisplatin, anti-Fas or radiation. The apoptotic response was measured by flow cytometry using propidium iodide and annexin V-FITC. The chromatin status of the apoptosis genes reflected the activation status of the intrinsic (Bax, Bcl2, Caspase-9 and p53) and extrinsic (Fas) pathways. An active intrinsic apoptotic pathway corresponded to sensitivity to cisplatin and radiation treatment of cell lines DLD1, SW620 and Colo320. An active Fas promoter corresponded to an active extrinsic apoptotic pathway in cell line DLD1. mRNA expression data correlated with the chromatin status of the apoptosis genes as measured by ChIP. In conclusion, the results presented in this study indicate that the balance between activating and silencing histone modifications, reflecting the chromatin status of apoptosis genes, can be used to predict the response of tumor cells to different anti-cancer therapies and could provide a novel target to sensitize tumors to obtain adequate treatment responses.

  6. Brucella invasion of human intestinal epithelial cells elicits a weak proinflammatory response but a significant CCL20 secretion.

    Science.gov (United States)

    Ferrero, Mariana C; Fossati, Carlos A; Rumbo, Martín; Baldi, Pablo C

    2012-10-01

    In spite of the frequent acquisition of Brucella infection by the oral route in humans, the interaction of the bacterium with cells of the intestinal mucosa has been poorly studied. Here, we show that different Brucella species can invade human colonic epithelial cell lines (Caco-2 and HT-29), in which only smooth species can replicate efficiently. Infection with smooth strains did not produce a significant cytotoxicity, while the rough strain RB51 was more cytotoxic. Infection of Caco-2 cells or HT-29 cells with either smooth or rough strains of Brucella did not result in an increased secretion of TNF-α, IL-1β, MCP-1, IL-10 or TGF-β as compared with uninfected controls, whereas all the infections induced the secretion of IL-8 and CCL20 by both cell types. The MCP-1 response to flagellin from Salmonella typhimurium was similar in Brucella-infected or uninfected cells, ruling out a bacterial inhibitory mechanism as a reason for the weak proinflammatory response. Infection did not modify ICAM-1 expression levels in Caco-2 cells, but increased them in HT-29 cells. These results suggest that Brucella induces only a weak proinflammatory response in gut epithelial cells, but produces a significant CCL20 secretion. The latter may be important for bacterial dissemination given the known ability of Brucella to survive in dendritic cells. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  7. The use of EEM fluorescence data and OPLS/UPLS-DA algorithm to discriminate between normal and cancer cell lines: a feasibility study.

    Science.gov (United States)

    Neves, Ana Carolina de Oliveira; Fernandes de Araújo, Raimundo; Luiza Cabral de Sá Leitão Oliveira, Ana; Antunes de Araújo, Aurigena; de Lima, Kássio Michell Gomes

    2014-05-21

    Excitation emission matrix (EEM) fluorescence spectroscopy combined with the OPLS method has been investigated as a promising tool to discriminate between normal and cancer cell lines in two datasets: (i) using several types of normal and cancer cells (including 3T3, ARPE, HEK, HepG2, HeLa, HT-29 and 786-0 cells); (ii) considering the expression of matrix metalloproteinase-2 and -9 (MMP-2 and MMP-9) in suspensions of HEK and 786-0 cell lines. Partial Least Squares-Discriminant Analysis (PLS-DA) using the score matrix from PARAFAC (Parallel Factor Analysis), UPLS-DA (Unfolded Partial Least Squares with Discriminant Analysis) and orthogonal projection to latent structures (OPLS) were used as the bases for the discrimination models. UPLS-DA presented relevant performance for cancer cells in both datasets, with 100% and 66.7% correct prediction for first and second cases, respectively, and poor discrimination relative to normal cells in the first dataset (25%). By using the OPLS, we achieved 75% correct prediction for normal cells and maintained 100% concordance for cancer objects. On applying OPLS to the second dataset, we obtained 100% correct prediction in both classes (normal and cancer) for calibration and prediction sets. These results suggest that EEM fluorescence spectroscopy combined with chemometrics could be used as a clinical tool for cancer cell detection based on intrinsic biomolecular signatures.

  8. (2Alpha,3beta)-2,3-dihydroxyolean-12-en-28-oic acid, a new natural triterpene from Olea europea, induces caspase dependent apoptosis selectively in colon adenocarcinoma cells.

    Science.gov (United States)

    Reyes, Fernando J; Centelles, Josep J; Lupiáñez, José A; Cascante, Marta

    2006-11-27

    Triterpenoids are known to induce apoptosis and to be anti-tumoural. Maslinic acid, a pentacyclic triterpene, is present in high concentrations in olive pomace. This study examines the response of HT29 and Caco-2 colon-cancer cell lines to maslinic-acid treatment. At concentrations inhibiting cell growth by 50-80% (IC50HT29=61+/-1 microM, IC80HT29=76+/-1 microM and IC50Caco-2=85+/-5 microM, IC80Caco-2=116+/-5 microM), maslinic acid induced strong G0/G1 cell-cycle arrest and DNA fragmentation, and increased caspase-3 activity. However, maslinic acid did not alter the cell cycle or induce apoptosis in the non-tumoural intestine cell lines IEC-6 and IEC-18. Moreover, maslinic acid induced cell differentiation in colon adenocarcinoma cells. These findings support a role for maslinic acid as a tumour suppressant and as a possible new therapeutic tool for aberrant cell proliferation in the colon. In this report, we demonstrate for the first time that, in tumoural cancer cells, maslinic acid exerts a significant anti-proliferation effect by inducing an apoptotic process characterized by caspase-3 activation by a p53-independent mechanism, which occurs via mitochondrial disturbances and cytochrome c release.

  9. In vitro anticancer effect of venom from Cuban scorpion Rhopalurus junceus against a panel of human cancer cell lines.

    Science.gov (United States)

    Díaz-García, Alexis; Morier-Díaz, Luis; Frión-Herrera, Yahima; Rodríguez-Sánchez, Hermis; Caballero-Lorenzo, Yamira; Mendoza-Llanes, Dianeya; Riquenes-Garlobo, Yanelis; Fraga-Castro, José A

    2013-01-01

    In Cuba the endemic species of scorpion Rhopalurus junceus has been used in traditional medicine for cancer treatment. However, there is little scientific evidence about its potential in cancer therapy. The effect of a range of scorpion venom concentrations (0.1, 0.25, 0.5, 0.75 and 1mg/ml) against a panel of human tumor cell lines from epithelial (Hela, SiHa, Hep-2, NCI-H292, A549, MDA-MB-231, MDA-MB-468, HT-29), hematopoietic origins (U937, K562, Raji) and normal cells (MRC-5, MDCK, Vero) was determined by the MTT assay. Additionally, the effect of venom on tumor cell death was assayed by Fluorescence microscopy, RT-PCR and western blot. Only the epithelial cancer cells showed significant cell viability reduction, with medium cytotoxic concentration (IC50) ranging from 0.6-1mg/ml, in a concentration-dependent manner. There was no effect on either normal or hematopoietic tumor cells. Scorpion venom demonstrated to induce apoptosis in less sensitive tumor cells (Hela) as evidenced by chromatin condensation, over expression of p53 and bax mRNA, down expression of bcl-2 mRNA and increase of activated caspases 3, 8, 9. In most sensitive tumor cells (A549), scorpion venom induced necrosis evidenced by acridine orange/ethidium bromide fluorescent dyes and down-expression of apoptosis-related genes. We concluded the scorpion venom from R. junceus possessed a selective and differential toxicity against epithelial cancer cells. This is the first report related to biological effect of R. junceus venom against a panel of tumor cells lines. All these results make R. junceus venom as a promise natural product for cancer treatment.

  10. Evaluation of chemical composition, antioxidant, antibacterial, cytotoxic and apoptotic effects of Aloysia citrodora extract on colon cancer cell line

    OpenAIRE

    Amir Mirzaie; Seyed Ataollah Sadat Shandiz; Hassan Noorbazargan; Elahe Ali Asgary

    2016-01-01

    Background: Aloysia citrodora belongs to the Verbenaceae family of plants, a well-known herbal medicine in Iran. The aim of the present study was to investigate the chemical composition, antioxidant, antibacterial, cytotoxic and apoptotic effect of A. citrodora extract against human colon cancer (HT29) cells by using real-time polymerase chain reaction and flow-cytometry methods. Methods: This experimental study was carried out in Islamic Azad University, East Tehran Branch, from March to...

  11. Synthesis of Tolmetin Hydrazide-Hydrazones and Discovery of a Potent Apoptosis Inducer in Colon Cancer Cells.

    Science.gov (United States)

    Küçükgüzel, Ş Güniz; Koç, Derya; Çıkla-Süzgün, Pelin; Özsavcı, Derya; Bingöl-Özakpınar, Özlem; Mega-Tiber, Pınar; Orun, Oya; Erzincan, Pınar; Sağ-Erdem, Safiye; Şahin, Fikrettin

    2015-10-01

    Tolmetin hydrazide and a novel series of tolmetin hydrazide-hydrazones 4a-l were synthesized in this study. The structures of the new compounds were determined by spectral (FT-IR, (1)H NMR) methods. N'-[(2,6-Dichlorophenyl)methylidene]-2-[1-methyl-5-(4-methylbenzoyl)-1H-pyrrol-2-yl]acetohydrazide (4g) was evaluated in vitro using the MTT colorimetric method against the colon cancer cell lines HCT-116 (ATCC, CCL-247) and HT-29 (ATCC, HTB-38) to determine growth inhibition and cell viability at different doses. Compound 4g exhibited anti-cancer activity with an IC50 value of 76 μM against colon cancer line HT-29 (ATCC, HTB-38) and did not display cytotoxicity toward control NIH3T3 mouse embryonic fibroblast cells compared to tolmetin. In addition, this compound was evaluated for caspase-3, caspase-8, caspase-9, and annexin-V activation in the apoptotic pathway, which plays a key role in the treatment of cancer. We demonstrated that the anti-cancer activity of this compound was due to the activation of caspase-8 and caspase-9 involved in the apoptotic pathway. In addition, in this study, we investigated the catalytical effect of COX on the HT-29 cancer line, the apoptotic mechanism, and the moleculer binding of tolmetin and compound 4g on the COX enzyme active site. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Portulaca oleracea extract can inhibit nodule formation of colon cancer stem cells by regulating gene expression of the Notch signal transduction pathway.

    Science.gov (United States)

    Jin, Heiying; Chen, Li; Wang, Shuiming; Chao, Deng

    2017-07-01

    To investigate whether Portulaca oleracea extract affects tumor formation in colon cancer stem cells and its chemotherapy sensitivity. In addition, to analyze associated genetic changes within the Notch signal transduction pathway. Serum-free cultures of colon cancer cells (HT-29) and HT-29 cancer stem cells were treated with the chemotherapeutic drug 5-fluorouracil to assess sensitivity. Injections of the stem cells were also given to BALB/c mice to confirm tumor growth and note its characteristics. In addition, the effect of different concentrations of P. oleracea extract was tested on the growth of HT-29 colon cancer cells and HT-29 cancer stem cells, as determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. The effects of P. oleracea extract on the expression of β-catenin, Notch1, and Notch2 in the HT-29 cells were studied using reverse transcription polymerase chain reaction and Western blotting. The tumor volume of the HT29 cells was two times larger than that of HT29 cancer stem cells. Treatment with P. oleracea extract inhibited the proliferation of both HT-29 cancer cells and HT-29 cancer stem cells at doses from 0.07 to 2.25 µg/mL. Apoptosis of HT-29 cancer cells and HT-29 cancer stem cells was assessed by flow cytometry; it was enhanced by the addition of P. oleracea extract. Finally, treatment with P. oleracea extract significantly downregulated the expression of the Notch1 and β-catenin genes in both cell types. The results of this study show that P. oleracea extract inhibits the growth of colon cancer stem cells in a dose-dependent manner. Furthermore, it inhibits the expression of the Notch1 and β-catenin genes. Taken together, this suggests that it may elicit its effects through regulatory and target genes that mediate the Notch signal transduction pathway.

  13. Galactose conjugated platinum(II) complex targeting the Warburg effect for treatment of non-small cell lung cancer and colon cancer.

    Science.gov (United States)

    Wu, Meng; Li, Hong; Liu, Ran; Gao, Xiangqian; Zhang, Menghua; Liu, Pengxing; Fu, Zheng; Yang, Jinna; Zhang-Negrerie, Daisy; Gao, Qingzhi

    2016-03-03

    Malignant neoplasms exhibit a higher rate of glycolysis than normal cells; this is known as the Warburg effect. To target it, a galactose-conjugated (trans-R,R-cyclohexane-1,2-diamine)-2-chloromalonato-platinum(II) complex (Gal-Pt) was designed, synthesized, and evaluated in five human cancer cell lines and against two different xenograft tumour models. Gal-Pt exhibits much higher aqueous solubility (over 25 times) and improved cytotoxicity than oxaliplatin, especially in human colon (HT29) and lung (H460) cancer cell lines. The safety profile of Gal-Pt was investigated in vivo by exploring the maximum tolerated dose (MTD) and animal mortality rate. The ratios of the animal lethal dosage values to the cytotoxicity in HT29 (LD50/IC50) showed that Gal-Pt was associated with an increased therapeutic index by over 30-fold compared to cisplatin and oxaliplatin. We evaluated in vivo antitumor activity by single agent intravenous treatment comparison studies of Gal-Pt (50 mg/kg as 65% MTD) and cisplatin (3 mg/kg, as 80% MTD) in a H460 lung cancer xenograft model, and with oxaliplatin (7 mg/kg, as 90% MTD) in a HT29 colon cancer xenograft model. The results show that Gal-Pt was more efficacious against H460 than cisplatin, and had superior potency in HT29 cells compared to oxaliplatin under nontoxic dosage conditions. The dependency between cytotoxicity of Gal-Pt and glucose transporters (GLUTs) was investigated by using quercetin as an inhibitor of GLUTs in HT29 cells. The cytotoxic potency of Gal-Pt was highly reduced by the inhibitor, suggesting that the uptake of Gal-Pt was regulated by glucose transporters. The GLUT mediated transportability and cellular uptake of Gal-Pt was also demonstrated using a fluorescent glucose bioprobe in HT29 competition assay. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  14. Selective toxicity of a quinoxaline 1,4-di-N-oxide derivative in human tumour cell lines.

    Science.gov (United States)

    Amaia, Azquetaa; Pachón, Gisela; Cascante, Marta; Creppy, Edmond E; Monge, Antonio; López de Cerain, Adela

    2005-01-01

    The presence of hypoxic cells in human solid tumours is one of the causes of tumour resistance to conventional therapy, and is also associated with processes that promote the tumour progression. Different chemical agents have been designed in order to take advantage of the particular metabolic characteristics of hypoxic regions. These drugs, called bioreductive agents, are activated inside the hypoxic cells to give active species that, in the presence of oxygen, are oxidised back to the non-toxic parent compound. Several quinoxaline 1,4-di-N-oxides have been described as potential bioreductive agents, and among them, 7-cloro-3-[[(N,N-dimethylamino)propy]amino]-2-quinoxalinecarbonitrile 1,4-di-N-oxide hydrocloride (Q-85 HCl) appeared to be the most promising one. In the present work, the selective cytotoxicity of Q-85 HCl was studied in several human tumour cell lines of different origin (Caco2, MCF-7, HT-29 and Tk-10). Cell viability was calculated after 2 h treatment under hypoxic and well-oxygenated conditions. The potency (the concentration that gives 1% of cell survival) in hypoxia and hypoxia cytotoxicity ratio (HCR = potency in oxygenated conditions/potency in hypoxia) were calculated after a 14-day clonogenic assay. Q-85 HCl was more toxic in hypoxia than in well-oxygenated cells in all the tumour cell lines. The best profile of potency in hypoxia (0.4 micromol/L) and selectivity (HCR=155) was found in CaCo-2 cells. Altogether, these results suggest an in vitro biological profile for Q-85 HCl that makes it an interesting candidate for the development as a bioreductive agent.

  15. Potential anti-inflammatory effects of the hydrophilic fraction of pomegranate (Punica granatum L.) seed oil on breast cancer cell lines.

    Science.gov (United States)

    Costantini, Susan; Rusolo, Fabiola; De Vito, Valentina; Moccia, Stefania; Picariello, Gianluca; Capone, Francesca; Guerriero, Eliana; Castello, Giuseppe; Volpe, Maria Grazia

    2014-06-24

    In this work, we characterized conjugated linolenic acids (e.g., punicic acid) as the major components of the hydrophilic fraction (80% aqueous methanol extract) from pomegranate (Punica granatum L.) seed oil (PSO) and evaluated their anti-inflammatory potential on some human colon (HT29 and HCT116), liver (HepG2 and Huh7), breast (MCF-7 and MDA-MB-231) and prostate (DU145) cancer lines. Our results demonstrated that punicic acid and its congeners induce a significant decrease of cell viability for two breast cell lines with a related increase of the cell cycle G0/G1 phase respect to untreated cells. Moreover, the evaluation of a great panel of cytokines expressed by MCF-7 and MDA-MB-231 cells showed that the levels of VEGF and nine pro-inflammatory cytokines (IL-2, IL-6, IL-12, IL-17, IP-10, MIP-1α, MIP-1β, MCP-1 and TNF-α) decreased in a dose dependent way with increasing amounts of the hydrophilic extracts of PSO, supporting the evidence of an anti-inflammatory effect. Taken together, the data herein suggest a potential synergistic cytotoxic, anti-inflammatory and anti-oxidant role of the polar compounds from PSO.

  16. Potential Anti-Inflammatory Effects of the Hydrophilic Fraction of Pomegranate (Punica granatum L. Seed Oil on Breast Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Susan Costantini

    2014-06-01

    Full Text Available In this work, we characterized conjugated linolenic acids (e.g., punicic acid as the major components of the hydrophilic fraction (80% aqueous methanol extract from pomegranate (Punica granatum L. seed oil (PSO and evaluated their anti-inflammatory potential on some human colon (HT29 and HCT116, liver (HepG2 and Huh7, breast (MCF-7 and MDA-MB-231 and prostate (DU145 cancer lines. Our results demonstrated that punicic acid and its congeners induce a significant decrease of cell viability for two breast cell lines with a related increase of the cell cycle G0/G1 phase respect to untreated cells. Moreover, the evaluation of a great panel of cytokines expressed by MCF-7 and MDA-MB-231 cells showed that the levels of VEGF and nine pro-inflammatory cytokines (IL-2, IL-6, IL-12, IL-17, IP-10, MIP-1α, MIP-1β, MCP-1 and TNF-α decreased in a dose dependent way with increasing amounts of the hydrophilic extracts of PSO, supporting the evidence of an anti-inflammatory effect. Taken together, the data herein suggest a potential synergistic cytotoxic, anti-inflammatory and anti-oxidant role of the polar compounds from PSO.

  17. Sesquiterpene lactones from Inula britannica and their cytotoxic and apoptotic effects on human cancer cell lines.

    Science.gov (United States)

    Bai, Naisheng; Lai, Ching-Shu; He, Kan; Zhou, Zhu; Zhang, Li; Quan, Zheng; Zhu, Nanqun; Zheng, Qun Yi; Pan, Min-Hsiung; Ho, Chi-Tang

    2006-04-01

    Three new sesquiterpenes (1-3), together with four known sesquiterpene lactones, were isolated from the flowers of Inula britannica var. chinensis. Structures were established on the basis of high-field 1D and 2D NMR methods supported by HRMS. All sesquiterpene lactones were tested for cytotoxicity as well as apoptotic ratio in human COLO 205, HT 29, HL-60, and AGS cancer cells. Compounds 3 and 4, two alpha-methylene gamma-lactone-bearing sesquiterpenes, were modestly active in these assays.

  18. Antioxidant activity and cytotoxic profie of Chuquiraga spinosa Lessing on human tumor cell lines: A promissory plant from Peruvian flra

    Directory of Open Access Journals (Sweden)

    Oscar Herrera-Calderon

    2017-05-01

    Full Text Available Objective: To determine the phytochemical content, antioxidant activity in vitro and cytotoxicity of crude ethanol extract (CEE, n-hexane fraction (NHF, petroleum ether fraction (PEF, chloroform fraction (CLF and ethyl acetate fraction (EAF of aerial parts of Chuquiraga spinosa (C. spinosa Lessing. Methods: Phytochemical screening was developed by color and precipitated formation. The evaluation of antioxidant activity was assessed using hydroxyl and nitric oxide radical. Total phenolic content (TPC and total flavonoids content (TFC were measured by using standard methods by spectrophotometry. The cytotoxic effect was determined on human tumor cell lines including MCF-7, H-460, HT-29, M-14, HUTU-80, K-562 and DU-145. Results: Phytochemical analysis confirmed the presence of phenols, flavonoids in crude extract and its all fractions. The CEE showed the highest antioxidant activity, for OH and NO radical scavenging tests (IC50 = 15.16 ± 3.45 μg/mL and IC50 = 18.91 ± 1.13 μg/mL, respectively. TPC was found to be the highest in the CEE (121.36 mg of gallic acid equivalent/g of dried extract compared to other fractions. The ranking order of NHF, PEF, CLF, EAF and CEE for TFC was 21.17 < 35.20 < 62.19 < 70.25 < 78.25 mg quercetin equivalent/g of dried extract. The crude ethanolic extract (μg/mL showed a high cytotoxicity on MCF-7 (IC50 = 9.25 ± 0.81, K-562 (IC50 = 7.34 ± 1.00, HT-29 (IC50 = 8.52 ± 2.69, H-460 (IC50 = 5.32 ± 1.05, M-14 (IC50 = 8.30 ± 0.60, DU-145 (IC50 = 7.09 ± 0.09, HUTU-80 (IC50 = 6.20 ± 0.50. Conclusions: The study showed that CEE of the aerial parts of C. spinosa can be measured as a natural source of antioxidant which might be effective towards preventing or slowing oxidative stress related to chronic diseases as well as cytotoxic agent.

  19. Maslinic Acid, a Natural Triterpene, Induces a Death Receptor-Mediated Apoptotic Mechanism in Caco-2 p53-Deficient Colon Adenocarcinoma Cells

    Science.gov (United States)

    Reyes-Zurita, Fernando J.; Rufino-Palomares, Eva E.; García-Salguero, Leticia; Peragón, Juan; Medina, Pedro P.; Parra, Andrés; Cascante, Marta; Lupiáñez, José A.

    2016-01-01

    Maslinic acid (MA) is a natural triterpene present in high concentrations in the waxy skin of olives. We have previously reported that MA induces apoptotic cell death via the mitochondrial apoptotic pathway in HT29 colon cancer cells. Here, we show that MA induces apoptosis in Caco-2 colon cancer cells via the extrinsic apoptotic pathway in a dose-dependent manner. MA triggered a series of effects associated with apoptosis, including the cleavage of caspases -8 and -3, and increased the levels of t-Bid within a few hours of its addition to the culture medium. MA had no effect on the expression of the Bax protein, release of cytochrome-c or on the mitochondrial membrane potential. This suggests that MA triggered the extrinsic apoptotic pathway in this cell type, as opposed to the intrinsic pathway found in the HT29 colon-cancer cell line. Our results suggest that the apoptotic mechanism induced in Caco-2 may be different from that found in HT29 colon-cancer cells, and that in Caco-2 cells MA seems to work independently of p53. Natural antitumoral agents capable of activating both the extrinsic and intrinsic apoptotic pathways could be of great use in treating colon-cancer of whatever origin. PMID:26751572

  20. Response to Multiple Radiation Doses of Human Colorectal Carcinoma Cells Infected With Recombinant Adenovirus Containing Dominant-Negative Ku70 Fragment

    International Nuclear Information System (INIS)

    Urano, Muneyasu; He Fuqiu; Minami, Akiko; Ling, C. Clifton; Li, Gloria C.

    2010-01-01

    Purpose: To investigate the effect of recombinant replication-defective adenovirus containing dominant-negative Ku70 fragment on the response of tumor cells to multiple small radiation doses. Our ultimate goal is to demonstrate the feasibility of using this virus in gene-radiotherapy to enhance the radiation response of tumor cells. Methods and Materials: Human colorectal HCT8 and HT29 carcinoma cells were plated in glass tubes, infected with virus (25 multiplicity of infection), and irradiated with a single dose or zero to five doses of 3 Gy each at 6-h intervals. Hypoxia was induced by flushing with 100% nitrogen gas. The cells were trypsinized 0 or 6 h after the final irradiation, and cell survival was determined by colony formation. The survival data were fitted to linear-quadratic model or exponential line. Results: Virus infection enhanced the radiation response of the HCT8 and HT29 cells. The virus enhancement ratio for single-dose irradiation at a surviving fraction of 0.1 was ∼1.3 for oxic and hypoxic HCT8 and 1.4 and 1.1 for oxic and hypoxic HT29, respectively. A similar virus enhancement ratio of 1.2-1.3 was observed for both oxic and hypoxic cells irradiated with multiple doses; however, these values were smaller than the values found for dominant-negative Ku70-transfected Rat-1 cells. This difference has been discussed. The oxygen enhancement ratio for HCT8 and HT29 receiving fractionated doses was 1.2 and 2.0, respectively, and virus infection altered them slightly. Conclusion: Infection of recombinant replication-defective adenovirus containing dominant-negative Ku70 fragment enhanced the response of human colorectal cancer cells to single and multiple radiation doses.

  1. Autophagy mediates cytotoxicity of human colorectal cancer cells treated with garcinielliptone FC.

    Science.gov (United States)

    Won, Shen-Jeu; Yen, Cheng-Hsin; Lin, Ting-Yu; Jiang-Shieh, Ya-Fen; Lin, Chun-Nan; Chen, Jyun-Ti; Su, Chun-Li

    2018-01-01

    The tautomeric pair of garcinielliptone FC (GFC) is a novel tautomeric pair of polyprenyl benzophenonoid isolated from the pericarps of Garcinia subelliptica Merr. (G. subelliptica, Clusiaceae), a tree with abundant sources of polyphenols. Our previous report demonstrated that GFC induced apoptosis on various types of human cancer cell lines including chemoresistant human colorectal cancer HT-29 cells. In the present study, we observed that many autophagy-related genes in GFC-treated HT-29 cells were up- and down-regulated using a cDNA microarray containing oncogenes and kinase genes. GFC-induced autophagy of HT-29 cells was confirmed by observing the formation of acidic vesicular organelles, LC3 puncta, and double-membrane autophagic vesicles using flow cytometry, confocal microscopy, and transmission electron microscopy, respectively. Inhibition of AKT/mTOR/P70S6K signaling as well as formation of Atg5-Atg12 and PI3K/Beclin-1 complexes were observed using Western blot. Administration of autophagy inhibitor (3-methyladenine and shRNA Atg5) and apoptosis inhibitor Z-VAD showed that the GFC-induced autophagy was cytotoxic form and GFC-induced apoptosis enhanced GFC-induced autophagy. Our data suggest the involvement of autophagy and apoptosis in GFC-induced anticancer mechanisms of human colorectal cancer. © 2017 Wiley Periodicals, Inc.

  2. In vitro anticancer activity of ethanolic extracts of Piper nigrum against colorectal carcinoma cell lines

    Science.gov (United States)

    Prashant, Akila; Rangaswamy, Chandini; Yadav, Anshu Kumar; Reddy, Varun; Sowmya, MN; Madhunapantula, Subbarao

    2017-01-01

    Background: Piper nigrum (PN) is well known for its cytotoxic and pharmacological benefits. However, there is minimal documented evidence about its cytotoxic efficacy against colorectal carcinoma. We therefore sought to procure a precisely quantitative and qualitative result, pertaining the efficacy of an ethanolic extract of PN (EEPN) against colorectal carcinoma. Materials and Methods: EEPN was prepared by subjecting dried PN seeds to gradient ethanol fractionation. The total phenol content (TPC), antioxidant activity (AOA), and anti-inflammatory activity (AIA) were determined using Folin–Ciocalteu assay, ferric reducing ability of plasma and 2, 2-diphenyl-1-picrylhydrazyl methods, and human red blood cells membrane stabilizing assay, respectively. Colorectal carcinoma cell lines (HCT-116, HCT-15, and HT-29) were procured from National Centre for Cell Science, Pune, and were cultured in Dulbecco's modified eagle media supplemented with 10% fetal bovine serum and 1 mM L-glutamine. Cells were seeded into a 96-well plate, followed by treatment with increasing concentrations of EEPN. The cytotoxic efficacy was evaluated based on percentage inhibition of cells, using sulforhodamine-B assay. The IC-50 values were calculated using Prism software (Prism from GraphPad software, Inc. CA, USA). Results: Biochemical analysis revealed that 50% EEPN exhibited higher TPC, AOA, and AIA when compared to 70% and 100% EEPN at any given concentration (P = 0.041). Cytotoxic analysis revealed a dose-dependent response with maximum cellular inhibition at TPC of 6 and 3 μg/ml, using 50% EEPN. However, 50% inhibition of cellular growth using 50% EEPN was seen with TPC of 3.2, 2.9, and 1.9 μg/ml at 24, 48, and 72 h, respectively, in HCT-15 cells. Hence, time- and dose-dependent increase in the cytotoxic efficacy of 50% EEPN against colorectal carcinoma cell lines were noted (P < 0.001). Conclusion: Given the significantly positive correlations exhibited between the biochemical and the

  3. Ciprofloxacin Affects Host Cells by Suppressing Expression of the Endogenous Antimicrobial Peptides Cathelicidins and Beta-Defensin-3 in Colon Epithelia

    Directory of Open Access Journals (Sweden)

    Protim Sarker

    2014-07-01

    Full Text Available Antibiotics exert several effects on host cells including regulation of immune components. Antimicrobial peptides (AMPs, e.g., cathelicidins and defensins display multiple functions in innate immunity. In colonic mucosa, cathelicidins are induced by butyrate, a bacterial fermentation product. Here, we investigated the effect of antibiotics on butyrate-induced expression of cathelicidins and beta-defensins in colon epithelial cells. Real-time PCR analysis revealed that ciprofloxacin and clindamycin reduce butyrate-induced transcription of the human cathelicidin LL-37 in the colonic epithelial cell line HT-29. Suppression of LL-37 peptide/protein by ciprofloxacin was confirmed by Western blot analysis. Immunohistochemical analysis demonstrated that ciprofloxacin suppresses the rabbit cathelicidin CAP-18 in rectal epithelia of healthy and butyrate-treated Shigella-infected rabbits. Ciprofloxacin also down-regulated butyrate-induced transcription of the human beta-defensin-3 in HT-29 cells. Microarray analysis of HT-29 cells revealed upregulation by butyrate with subsequent down-regulation by ciprofloxacin of additional genes encoding immune factors. Dephosphorylation of histone H3, an epigenetic event provided a possible mechanism of the suppressive effect of ciprofloxacin. Furthermore, LL-37 peptide inhibited Clostridium difficile growth in vitro. In conclusion, ciprofloxacin and clindamycin exert immunomodulatory function by down-regulating AMPs and other immune components in colonic epithelial cells. Suppression of AMPs may contribute to the overgrowth of C. difficile, causing antibiotic-associated diarrhea.

  4. Defective interleukin-4/Stat6 activity correlates with increased constitutive expression of negative regulators SOCS-3, SOCS-7, and CISH in colon cancer cells.

    Science.gov (United States)

    Liu, Xiao Hong; Xu, Shuang Bing; Yuan, Jia; Li, Ben Hui; Zhang, Yan; Yuan, Qin; Li, Pin Dong; Li, Feng; Zhang, Wen Jie

    2009-12-01

    Interleukin-4 (IL-4)-induced Stat6 activities (phenotypes) vary among human cancer cells, of which the HT-29 cell line carries an active Stat6(high) phenotype, while Caco-2 carries a defective Stat6(null) phenotype, respectively. Cancer cells with Stat6(high) show resistance to apoptosis and exaggerated metastasis, suggesting the clinical significance of Stat6 phenotypes. We previously showed that Stat6(high) HT-29 cells exhibited low constitutive expression of Stat6-negative regulators SOCS-1 and SHP-1 because of gene hypermethylation. This study further examined the constitutive expression of other closely related SOCS family numbers including SOCS-3, SOCS-5, SOCS-7, and CISH using RT-PCR. Similar to SOCS-1 and SHP-1, Stat6(high) HT-29 cells expressed low constitutive mRNA of SOCS-3, SOCS-7, and CISH than Stat6(null) Caco-2 cells. Interestingly, DNA demethylation using 5-aza-2'-deoxycytidine in HT-29 cells up-regulated mRNA expression of the above genes, indicating a hypermethylation status, which was confirmed by methylation-specific sequencing in selected SOCS-3 gene. Furthermore, defective Stat6(null) Caco-2 exhibited impaired phosphorylation of Stat6 after IL-4 stimulation by flow cytometry, in keeping with the notion of an over-performed negative regulation. The findings that IL-4/Stat6 phenotypes show differential expression of multiple negative regulators suggest a model that a collective force of powerful negative regulators, directly and indirectly, acts on Stat6 activation, which may result in differential Stat6 phenotypes.

  5. Interaction of polyunsaturated fatty acids and sodium butyrate during apoptosis in HT-29 human colon adenocarcinoma cells

    Czech Academy of Sciences Publication Activity Database

    Hofmanová, Jiřina; Vaculová, Alena; Lojek, Antonín; Kozubík, Alois

    2005-01-01

    Roč. 44, č. 1 (2005), s. 40-51 ISSN 1436-6207 R&D Projects: GA ČR(CZ) GA525/01/0419 Institutional research plan: CEZ:AV0Z50040507 Keywords : colon cancer * diet * butyrate Subject RIV: BO - Biophysics Impact factor: 2.257, year: 2005

  6. Structure-function relationships of anthocyanins from various anthocyanin-rich extracts on the inhibition of colon cancer cell growth.

    Science.gov (United States)

    Jing, Pu; Bomser, Joshua A; Schwartz, Steven J; He, Jian; Magnuson, Bernadene A; Giusti, M Mónica

    2008-10-22

    Anthocyanins are potent antioxidants and may be chemoprotective. However, the structure-function relationships are not well understood. The objectives of this study were to compare the chemoprotective properties of anthocyanin-rich extracts (AREs) with variable anthocyanin profiles to understand the relationship between anthocyanin chemical structure and chemoprotective activity, measured as inhibition of colon cancer cell proliferation. Additionally, the chemoprotective interaction of anthocyanins and other phenolics was investigated. AREs with different anthocyanin profiles from purple corn, chokeberry, bilberry, purple carrot, grape, radish, and elderberry were tested for growth inhibition (GI 50) using a human colorectal adenocarcinoma (HT29) cell line. All AREs suppressed HT29 cell growth to various degrees as follows: purple corn (GI 50 approximately 14 microg of cy-3-glu equiv/mL) > chokeberry and bilberry > purple carrot and grape > radish and elderberry (GI 50 > 100 microg of cy-3-glu equiv/mL). Anthocyanins played a major role in AREs' chemoprotection and exerted an additive interaction with the other phenolics present. Statistical analyses suggested that anthocyanin chemical structure affected chemoprotection, with nonacylated monoglycosylated anthocyanins having greater inhibitory effect on HT-29 cell proliferation, whereas anthocyanins with pelargonidin, triglycoside, and/or acylation with cinnamic acid exerted the least effect. These findings should be considered for crop selection and the development of anthocyanin-rich functional foods.

  7. Cytotoxicity of probiotics from Philippine commercial dairy products on cancer cells and the effect on expression of cfos and cjun early apoptotic-promoting genes and Interleukin-1 β and Tumor Necrosis Factor-α proinflammatory cytokine genes.

    Science.gov (United States)

    Shyu, Peter T; Oyong, Glenn G; Cabrera, Esperanza C

    2014-01-01

    This study determined cytotoxicity of probiotic Lactobacillus spp. from Philippine dairy products on cancer cells and normal fibroblasts and their effects on expression of early apoptotic-promoting cfos, cjun and proinflammatory cytokine IL-1β, TNF-α genes. Cultures were from Yakult, Bear Brand Probiotic Drink, Nido3+ Powdered Milk. Filter-sterilized supernatants from cultures of Lactobacillus spp. were evaluated for cytotoxicity to colon cancer cells (HT-29 and HCT116), leukemia cells (THP-1), and normal human dermal fibroblasts (HDFn) using PrestoBlue. Bleomycin was the positive control. Absolute quantification of transcript levels was conducted using qRT-PCR. Cytotoxicity index profiles on HDFn, THP-1 of all probiotic supernatants and negative controls suggest nontoxicity to the cells when compared to bleomycin, whereas all probiotic supernatants were found to be cytotoxic to HT-29 and HCT-116 colon cancer cell lines. Expression of cfos, cjun transcripts was significantly upregulated in HT-29 and HCT116 cells treated with probiotic supernatants compared to untreated baseline levels (P probiotic supernatants compared to those exposed to MRS medium (P < 0.05). Results provide strong support for the role of Lactobacillus spp. studied in anticancer therapy and in prevention of inflammation that may act as precursor to carcinogenesis.

  8. Expression of matrix metalloprotease-2, -7 and -9 on human colon, liver and bile duct cell lines by enteric and gastric Helicobacter species.

    Science.gov (United States)

    Yanagisawa, Naoko; Geironson, Linda; Al-Soud, Waleed Abu; Ljungh, Sa

    2005-05-01

    Gastric and enteric Helicobacter species have been associated with malignant and inflammatory diseases of the stomach, liver, gall bladder and intestine. Matrix metalloproteinases (MMPs) participate in degradation of extracellular matrix, which allows bacteria to come in contact with and interact with the cells. Enhanced level of MMPs facilitates metastasis and cell invasion of tumor cells by removal of physical barriers, as well as modulation of biologic activities of the proteins residing in the extracellular matrix. The aim of this study was to evaluate the effect of gastric and enteric Helicobacter on induction of MMPs in hepatocytes and epithelial cells of gall bladder and colon. Human hepatocytes HepG2, gall bladder epithelial cells TFK-1, and colon epithelial cells HT29 were infected with strains of H. pylori cagA+, cagE+, H. pylori cagA-, cagE-, H. pullorum, H. cholecystus, H. bilis and H. hepaticus. Protein levels of MMPs were analyzed by enzyme-linked immunosorbent assay and immunohistochemistry. Reverse transcription-quantitative polymerase chain reaction was used to study mRNA levels. Increased expression of MMP-2 and MMP-9 was observed on HepG2, TFK-1 and HT29 infected with H. pylori cagA+, cagE+ and H. cholecystus strains. H. pylori cagA+, cagE+, H. cholecystus, H. pullorum, H. bilis and H. hepaticus strains increased expression of MMP-7 on HT29, compared to uninfected control cells. The effect of MMP upregulation on HepG2, TFK-1 and HT29 was bacterial dose dependent. H. pylori cagA-, cagE- strain did not increase expression of MMPs. Inducible MMPs on colon and bile duct epithelial cells as well as hepatocytes may play an important role in facilitating invasion and progression of cancer by Helicobacter species colonizing the hepatobiliary and gastrointestinal tract.

  9. Fas ligand expression in human and mouse cancer cell lines; a caveat on over-reliance on mRNA data

    Directory of Open Access Journals (Sweden)

    Ryan Aideen E

    2006-02-01

    Full Text Available Abstract Background During carcinogenesis, tumors develop multiple mechanisms for evading the immune response, including upregulation of Fas ligand (FasL/CD95L expression. Expression of FasL may help to maintain tumor cells in a state of immune privilege by inducing apoptosis of anti-tumor immune effector cells. Recently this idea has been challenged by studies reporting that tumor cells of varying origin do not express FasL. In the present study, we aimed to comprehensively characterize FasL expression in tumors of both murine and human origin over a 72 hour time period. Methods RNA and protein was extracted from six human (SW620, HT29, SW480, KM12SM, HCT116, Jurkat and three mouse (CMT93, CT26, B16F10 cancer cell lines at regular time intervals over a 72 hour time period. FasL expression was detected at the mRNA level by RT-PCR, using intron spanning primers, and at the protein level by Western Blotting and immunofluorescence, using a polyclonal FasL- specific antibody. Results Expression of FasL mRNA and protein was observed in all cell lines analysed. However, expression of FasL mRNA varied dramatically over time, with cells negative for FasL mRNA at many time points. In contrast, 8 of the 9 cell lines constitutively expressed FasL protein. Thus, cells can abundantly express FasL protein at times when FasL mRNA is absent. Conclusion These findings demonstrate the importance of complete analysis of FasL expression by tumor cells in order to fully characterize its biological function and may help to resolve the discrepancies present in the literature regarding FasL expression and tumor immune privilege.

  10. Study of Alpha Tocopherol, Celecoxib Induced Apoptosis in Human Colorectal Carcinoma Cell Line

    Directory of Open Access Journals (Sweden)

    R Solgui

    2012-05-01

    Full Text Available

    Background and Objectives: Chronic, unbridled oxidative damages have been known as the culprits behind many chronic diseases, including cancers, atherosclerosis, diabetes and Alzheimer’s. Cyclooxygenase-2 (COX-2-the main enzyme involved in inducing these processes- plays an important role in tumor development and progression. COX-2 inhibitors should be used at high doses for a long time in order to bring about chemoprevention and induction of anti-tumor effects. For example, celecoxib prevents colorectal tumor growth and induces apoptosis in both in vitro and in vivo models. Disregulation of COX-2 expression coincides with the development of gastrointestinal malignancy in humans and in animal models of colorectal cancer. Increased COX-2 expression in human colorectal adeno-carcinomas has been elucidated when compared with normal adjacent colonic mucosa. The capacity of vitamin E, particularly in α form, to quench free radical damage, induces apoptosis and impact expression of oncogenes makes it an appropriate choice for chemotherapeutic strategies. Studies have shown that carcinogenesis and DNA damage due to UV are inhibited by vitamin E. The goal of this study was to investigate alpha tocopherol and celecoxib induced apoptosis in human colorectal carcinoma cell line.

    Methods: In this study, HT29 cells were exposed to different concentrations of tocopherol (5, 10, and 20µM and celecoxib (25, 50, 75, 100µM followed by DNA extraction and fragmentation for demonstrating cell death process.

    Results: The results indicated that celecoxib at lower doses (25, and 50µM could not induce cell death, but at higher doses (75, and 100 µM, DNA fragmentation results typically resembled programmed cell death.

  11. Piceatannol promotes apoptosis via up-regulation of microRNA-129 expression in colorectal cancer cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Haogang [Department of General Surgery, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150081 (China); Jia, Ruichun [Department of Blood Transfusion, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150081 (China); Wang, Chunjing; Hu, Tianming [Department of General Surgery, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150081 (China); Wang, Fujing, E-mail: wangfujing-hyd@163.com [Department of General Surgery, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150081 (China)

    2014-09-26

    Highlights: • Piceatannol induces apoptosis in cultured CRC cells. • Piceatannol promotes expression of miR-129. • miR-129 mediates proapoptotic effects of piceatannol. - Abstract: Piceatannol, a naturally occurring analog of resveratrol, has been confirmed as an antitumor agent by inhibiting proliferation, migration, and metastasis in diverse cancer. However, the effect and mechanisms of piceatannol on colorectal cancer (CRC) have not been well understood. This study aimed to test whether piceatannol could inhibit growth of CRC cells and reveal its underlying molecular mechanism. MTT assay was used to detect the cell viability in HCT116 and HT29 cells. Flow cytometry analysis was employed to measure apoptosis of CRC cells. Bcl-2, Bax and caspase-3 levels were analyzed by Western blot and miR-129 levels were determined by real-time RT-PCR. Our study showed that piceatannol inhibited HCT116 and HT29 cells growth in a concentration- and time-dependent manner. Piceatannol induced apoptosis by promoting expression of miR-129, and then inhibiting expression of Bcl-2, an known target for miR-129. Moreover, knock down of miR-129 could reverse the reduction of cell viability induced by piceatannol in HCT116 and HT29 cells. Taken together, our study unraveled the ability of piceatannol to suppress colorectal cancer growth and elucidated the participation of miR-129 in the anti-cancer action of piceatannol. Our findings suggest that piceatannol can be considered to be a promising anticancer agent for CRC.

  12. Fraction against Human Cancer Cell Lines

    African Journals Online (AJOL)

    kidney carcinoma cell lines of hamsters (BSR). [11]. While, in another article, A. sieberia unrefined extract exhibited dose dependent antiproliferative activity against several cancer cell lines (human bladder carcinoma RT112, human laryngeal carcinoma and human myelogenous leukaemia K562), with IC50. = 81.59, 59.05 ...

  13. Difference in membrane repair capacity between cancer cell lines and a normal cell line

    DEFF Research Database (Denmark)

    Frandsen, Stine Krog; McNeil, Anna K.; Novak, Ivana

    2016-01-01

    repair was investigated by disrupting the plasma membrane using laser followed by monitoring fluorescent dye entry over time in seven cancer cell lines, an immortalized cell line, and a normal primary cell line. The kinetics of repair in living cells can be directly recorded using this technique......, providing a sensitive index of repair capacity. The normal primary cell line of all tested cell lines exhibited the slowest rate of dye entry after laser disruption and lowest level of dye uptake. Significantly, more rapid dye uptake and a higher total level of dye uptake occurred in six of the seven tested...

  14. Biophysical Profiling of Tumor Cell Lines

    Directory of Open Access Journals (Sweden)

    Frederick Coffman

    2011-01-01

    Full Text Available Despite significant differences in genetic profiles, cancer cells share common phenotypic properties, including membrane-associated changes that facilitate invasion and metastasis. The Corning Epic® optical biosensor was used to monitor dynamic mass rearrangements within and proximal to the cell membrane in tumor cell lines derived from cancers of the colon, bone, cervix, lung and breast. Data was collected in real time and required no exogenously added signaling moiety (signal-free technology. Cell lines displayed unique profiles over the time-courses: the time-courses all displayed initial signal increases to maximal values, but the rate of increase to those maxima and the value of those maxima were distinct for each cell line. The rate of decline following the maxima also differed among cell lines. There were correlations between the signal maxima and the observed metastatic behavior of the cells in xenograft experiments; for most cell types the cells that were more highly metastatic in mice had lower time-course maxima values, however the reverse was seen in breast cancer cells. The unique profiles of these cell lines and the correlation of at least one profile characteristic with metastatic behavior demonstrate the potential utility of biophysical tumor cell profiling in the study of cancer biology.

  15. Growth-inhibitory effects of the chemopreventive agent indole-3-carbinol are increased in combination with the polyamine putrescine in the SW480 colon tumour cell line

    Science.gov (United States)

    Hudson, E Ann; Howells, Lynne M; Gallacher-Horley, Barbara; Fox, Louise H; Gescher, Andreas; Manson, Margaret M

    2003-01-01

    Background Many tumours undergo disregulation of polyamine homeostasis and upregulation of ornithine decarboxylase (ODC) activity, which can promote carcinogenesis. In animal models of colon carcinogenesis, inhibition of ODC activity by difluoromethylornithine (DFMO) has been shown to reduce the number and size of colon adenomas and carcinomas. Indole-3-carbinol (I3C) has shown promising chemopreventive activity against a range of human tumour cell types, but little is known about the effect of this agent on colon cell lines. Here, we investigated whether inhibition of ODC by I3C could contribute to a chemopreventive effect in colon cell lines. Methods Cell cycle progression and induction of apoptosis were assessed by flow cytometry. Ornithine decarboxylase activity was determined by liberation of CO2 from 14C-labelled substrate, and polyamine levels were measured by HPLC. Results I3C inhibited proliferation of the human colon tumour cell lines HT29 and SW480, and of the normal tissue-derived HCEC line, and at higher concentrations induced apoptosis in SW480 cells. The agent also caused a decrease in ODC activity in a dose-dependent manner. While administration of exogenous putrescine reversed the growth-inhibitory effect of DFMO, it did not reverse the growth-inhibition following an I3C treatment, and in the case of the SW480 cell line, the effect was actually enhanced. In this cell line, combination treatment caused a slight increase in the proportion of cells in the G2/M phase of the cell cycle, and increased the proportion of cells undergoing necrosis, but did not predispose cells to apoptosis. Indole-3-carbinol also caused an increase in intracellular spermine levels, which was not modulated by putrescine co-administration. Conclusion While indole-3-carbinol decreased ornithine decarboxylase activity in the colon cell lines, it appears unlikely that this constitutes a major mechanism by which the agent exerts its antiproliferative effect, although accumulation

  16. Growth-inhibitory effects of the chemopreventive agent indole-3-carbinol are increased in combination with the polyamine putrescine in the SW480 colon tumour cell line

    Directory of Open Access Journals (Sweden)

    Gescher Andreas

    2003-01-01

    Full Text Available Abstract Background Many tumours undergo disregulation of polyamine homeostasis and upregulation of ornithine decarboxylase (ODC activity, which can promote carcinogenesis. In animal models of colon carcinogenesis, inhibition of ODC activity by difluoromethylornithine (DFMO has been shown to reduce the number and size of colon adenomas and carcinomas. Indole-3-carbinol (I3C has shown promising chemopreventive activity against a range of human tumour cell types, but little is known about the effect of this agent on colon cell lines. Here, we investigated whether inhibition of ODC by I3C could contribute to a chemopreventive effect in colon cell lines. Methods Cell cycle progression and induction of apoptosis were assessed by flow cytometry. Ornithine decarboxylase activity was determined by liberation of CO2 from 14C-labelled substrate, and polyamine levels were measured by HPLC. Results I3C inhibited proliferation of the human colon tumour cell lines HT29 and SW480, and of the normal tissue-derived HCEC line, and at higher concentrations induced apoptosis in SW480 cells. The agent also caused a decrease in ODC activity in a dose-dependent manner. While administration of exogenous putrescine reversed the growth-inhibitory effect of DFMO, it did not reverse the growth-inhibition following an I3C treatment, and in the case of the SW480 cell line, the effect was actually enhanced. In this cell line, combination treatment caused a slight increase in the proportion of cells in the G2/M phase of the cell cycle, and increased the proportion of cells undergoing necrosis, but did not predispose cells to apoptosis. Indole-3-carbinol also caused an increase in intracellular spermine levels, which was not modulated by putrescine co-administration. Conclusion While indole-3-carbinol decreased ornithine decarboxylase activity in the colon cell lines, it appears unlikely that this constitutes a major mechanism by which the agent exerts its antiproliferative

  17. Establishment of cell lines with rat spermatogonial stem cell characteristics

    NARCIS (Netherlands)

    van Pelt, Ans M. M.; Roepers-Gajadien, Hermien L.; Gademan, Iris S.; Creemers, Laura B.; de Rooij, Dirk G.; van Dissel-Emiliani, Federica M. F.

    2002-01-01

    Spermatogonial cell lines were established by transfecting a mixed population of purified rat A(s) (stem cells), A(pr) and A(al) spermatogonia with SV40 large T antigen. Two cell lines were characterized and found to express Hsp90alpha and oct-4, specific markers for germ cells and A spermatogonia,

  18. Comparative effects of RRR-alpha- and RRR-gamma-tocopherol on proliferation and apoptosis in human colon cancer cell lines

    Directory of Open Access Journals (Sweden)

    Sherman Devin

    2006-01-01

    Full Text Available Abstract Background Mediterranean societies, with diets rich in vitamin E isoforms, have a lower risk for colon cancer than those of northern Europe and the Americas. Vitamin E rich diets may neutralize free radicals generated by fecal bacteria in the gut and prevent DNA damage, but signal transduction activities can occur independent of the antioxidant function. The term vitamin E represents eight structurally related compounds, each differing in their potency and mechanisms of chemoprevention. The RRR-γ-tocopherol isoform is found primarily in the US diet, while RRR-α-tocopherol is highest in the plasma. Methods The effectiveness of RRR-α- and RRR-γ-tocopherol at inhibiting cell growth and inducing apoptosis in colon cancer cell lines with varying molecular characteristics (SW480, HCT-15, HCT-116 and HT-29 and primary colon cells (CCD-112CoN, nontransformed normal phenotype was studied. Colon cells were treated with and without RRR-α- or RRR-γ-tocopherol using varying tocopherol concentrations and time intervals. Cell proliferation and apoptosis were measured using the trypan blue assay, annexin V staining, DNA laddering and caspase activation. Results Treatment with RRR-γ-tocopherol resulted in significant cell death for all cancer cell lines tested, while RRR-α-tocopherol did not. Further, RRR-γ-tocopherol treatment showed no cytotoxicity to normal colon cells CCD-112CoN at the highest concentration and time point tested. RRR-γ-tocopherol treatment resulted in cleavage of PARP, caspase 3, 7, and 8, but not caspase 9. Differences in the percentage cell death and apoptosis were observed in different cell lines suggesting that molecular differences in these cell lines may influence the ability of RRR-γ-tocopherol to induce cell death. Conclusion This is the first study to demonstrate that multiple colon cancer cell lines containing varying genetic alterations will under go growth reduction and apoptosis in the presence of RRR

  19. Comparative effects of RRR-alpha- and RRR-gamma-tocopherol on proliferation and apoptosis in human colon cancer cell lines.

    Science.gov (United States)

    Campbell, Sharon E; Stone, William L; Lee, Steven; Whaley, Sarah; Yang, Hongsong; Qui, Min; Goforth, Paige; Sherman, Devin; McHaffie, Derek; Krishnan, Koyamangalath

    2006-01-17

    Mediterranean societies, with diets rich in vitamin E isoforms, have a lower risk for colon cancer than those of northern Europe and the Americas. Vitamin E rich diets may neutralize free radicals generated by fecal bacteria in the gut and prevent DNA damage, but signal transduction activities can occur independent of the antioxidant function. The term vitamin E represents eight structurally related compounds, each differing in their potency and mechanisms of chemoprevention. The RRR-gamma-tocopherol isoform is found primarily in the US diet, while RRR-alpha-tocopherol is highest in the plasma. The effectiveness of RRR-alpha- and RRR-gamma-tocopherol at inhibiting cell growth and inducing apoptosis in colon cancer cell lines with varying molecular characteristics (SW480, HCT-15, HCT-116 and HT-29) and primary colon cells (CCD-112CoN, nontransformed normal phenotype) was studied. Colon cells were treated with and without RRR-alpha- or RRR-gamma-tocopherol using varying tocopherol concentrations and time intervals. Cell proliferation and apoptosis were measured using the trypan blue assay, annexin V staining, DNA laddering and caspase activation. Treatment with RRR-gamma-tocopherol resulted in significant cell death for all cancer cell lines tested, while RRR-alpha-tocopherol did not. Further, RRR-gamma-tocopherol treatment showed no cytotoxicity to normal colon cells CCD-112CoN at the highest concentration and time point tested. RRR-gamma-tocopherol treatment resulted in cleavage of PARP, caspase 3, 7, and 8, but not caspase 9. Differences in the percentage cell death and apoptosis were observed in different cell lines suggesting that molecular differences in these cell lines may influence the ability of RRR-gamma-tocopherol to induce cell death. This is the first study to demonstrate that multiple colon cancer cell lines containing varying genetic alterations will under go growth reduction and apoptosis in the presence of RRR-gamma-tocopherol without damage to

  20. Comparative effects of RRR-alpha- and RRR-gamma-tocopherol on proliferation and apoptosis in human colon cancer cell lines

    International Nuclear Information System (INIS)

    Campbell, Sharon E; Krishnan, Koyamangalath; Stone, William L; Lee, Steven; Whaley, Sarah; Yang, Hongsong; Qui, Min; Goforth, Paige; Sherman, Devin; McHaffie, Derek

    2006-01-01

    Mediterranean societies, with diets rich in vitamin E isoforms, have a lower risk for colon cancer than those of northern Europe and the Americas. Vitamin E rich diets may neutralize free radicals generated by fecal bacteria in the gut and prevent DNA damage, but signal transduction activities can occur independent of the antioxidant function. The term vitamin E represents eight structurally related compounds, each differing in their potency and mechanisms of chemoprevention. The RRR-γ-tocopherol isoform is found primarily in the US diet, while RRR-α-tocopherol is highest in the plasma. The effectiveness of RRR-α- and RRR-γ-tocopherol at inhibiting cell growth and inducing apoptosis in colon cancer cell lines with varying molecular characteristics (SW480, HCT-15, HCT-116 and HT-29) and primary colon cells (CCD-112CoN, nontransformed normal phenotype) was studied. Colon cells were treated with and without RRR-α- or RRR-γ-tocopherol using varying tocopherol concentrations and time intervals. Cell proliferation and apoptosis were measured using the trypan blue assay, annexin V staining, DNA laddering and caspase activation. Treatment with RRR-γ-tocopherol resulted in significant cell death for all cancer cell lines tested, while RRR-α-tocopherol did not. Further, RRR-γ-tocopherol treatment showed no cytotoxicity to normal colon cells CCD-112CoN at the highest concentration and time point tested. RRR-γ-tocopherol treatment resulted in cleavage of PARP, caspase 3, 7, and 8, but not caspase 9. Differences in the percentage cell death and apoptosis were observed in different cell lines suggesting that molecular differences in these cell lines may influence the ability of RRR-γ-tocopherol to induce cell death. This is the first study to demonstrate that multiple colon cancer cell lines containing varying genetic alterations will under go growth reduction and apoptosis in the presence of RRR-γ-tocopherol without damage to normal colon cells. The amount growth

  1. BHD Tumor Cell Line and Renal Cell Carcinoma Line | NCI Technology Transfer Center | TTC

    Science.gov (United States)

    Scientists at the National Cancer Institute  have developed a novel renal cell carcinoma (RCC) cell line designated UOK257, which was derived from the surgical kidney tissue of a patient with hereditary Birt-Hogg-Dube''''(BHD) syndrome and companion cell line UOK257-2 in which FLCN expression has been restored by lentivirus infection. The NCI Urologic Oncology Branch seeks parties interested in licensing or collaborative research to co-develop, evaluate, or commercialize kidney cancer tumor cell lines.

  2. Different molecular organization of two carotenoids, lutein and zeaxanthin, in human colon epithelial cells and colon adenocarcinoma cells

    Science.gov (United States)

    Grudzinski, Wojciech; Piet, Mateusz; Luchowski, Rafal; Reszczynska, Emilia; Welc, Renata; Paduch, Roman; Gruszecki, Wieslaw I.

    2018-01-01

    Two cell lines, human normal colon epithelial cells (CCD 841 CoTr) and human colon adenocarcinoma cells (HT-29) were cultured in the presence of exogenous carotenoids, either zeaxanthin or lutein. Both carotenoids demonstrated cytotoxicity with respect to cancer cells but not to normal cells. Cells from both the cell lines were analyzed with application of fluorescence lifetime imaging microscopy and Raman scattering microscopy. Both imaging techniques show effective incorporation of carotenoid molecules into growing cells. Comparison of the Raman scattering and fluorescence lifetime characteristics reveals different molecular organization of carotenoids in the carcinoma and normal cells. The main difference consists in a carotenoid aggregation level which is substantially lower in the carcinoma cells as compared to the normal cells. Different molecular organization of carotenoids was interpreted in terms of a different metabolism of normal and carcinoma cells and has been concluded to provide a possibility of cancer diagnosis based on spectroscopic analyses.

  3. RNAi-mediated knock-down of arylamine N-acetyltransferase-1 expression induces E-cadherin up-regulation and cell-cell contact growth inhibition.

    Directory of Open Access Journals (Sweden)

    Jacky M Tiang

    Full Text Available Arylamine N-acetyltransferase-1 (NAT1 is an enzyme that catalyzes the biotransformation of arylamine and hydrazine substrates. It also has a role in the catabolism of the folate metabolite p-aminobenzoyl glutamate. Recent bioinformatics studies have correlated NAT1 expression with various cancer subtypes. However, a direct role for NAT1 in cell biology has not been established. In this study, we have knocked down NAT1 in the colon adenocarcinoma cell-line HT-29 and found a marked change in cell morphology that was accompanied by an increase in cell-cell contact growth inhibition and a loss of cell viability at confluence. NAT1 knock-down also led to attenuation in anchorage independent growth in soft agar. Loss of NAT1 led to the up-regulation of E-cadherin mRNA and protein levels. This change in E-cadherin was not attributed to RNAi off-target effects and was also observed in the prostate cancer cell-line 22Rv1. In vivo, NAT1 knock-down cells grew with a longer doubling time compared to cells stably transfected with a scrambled RNAi or to parental HT-29 cells. This study has shown that NAT1 affects cell growth and morphology. In addition, it suggests that NAT1 may be a novel drug target for cancer therapeutics.

  4. Colorectal cancer cell lines made resistant to SN38-and Oxaliplatin: Roles of altered ion transporter function in resistance?

    DEFF Research Database (Denmark)

    Sandra, Christensen; Jensen, Niels Frank; Stoeckel, Johanne Danmark

    2013-01-01

    Colorectal cancer (CRC) is the 3rd most common cancer globally, with 5year survival rates of ~50%. Response rates to standard treatments (irinotecan (SN38) or Oxaliplatin (Oxp)) are 31–56% and drug resistance is a major problem. Thus, we established in vitro CRC models to investigate SN38 and Oxp...... resistance in HCT-116, HT-29 and LoVo cells. Microarray analysis and qPCR validation showed that mRNA expression of glutamate transporters SLC1A1 and SLC1A3 were markedly altered in resistant cells. Remarkably, mRNA levels of SLC1A3 were increased by ~40-and ~2500-fold in SN38-and Oxp-resistant HT29 cells......, respectively. Studies are ongoing to assess glutamate uptake in parental and resistant CRC cells and the effect of inhibition/knockdown of SLC1A1 and -3 on SN38- and Oxp resistance. In conclusion, SN38-and Oxp-resistance in CRC cells is associated with SLC1A1 and -3 dysregulation. As these transporters have...

  5. Evaluation of drug permeation under fed state conditions using mucus-covered Caco-2 cell epithelium.

    Science.gov (United States)

    Birch, Ditlev; Diedrichsen, Ragna G; Christophersen, Philip C; Mu, Huiling; Nielsen, Hanne M

    2018-03-07

    The absence of a surface-lining mucus layer is a major pitfall for the Caco-2 epithelial model. However, this can be alleviated by applying biosimilar mucus (BM) to the apical surface of the cell monolayer, thereby constructing a mucosa mimicking in vivo conditions. This study aims to elucidate the influence of BM as a barrier towards exogenic compounds such as permeation enhancers, and components of fed state simulated intestinal fluid (FeSSIF). Caco-2 cell monolayers surface-lined with BM were exposed to several compounds with distinct physicochemical properties, and the cell viability and permeability of the cell monolayer was compared to that of cell monolayers without BM and well-established mucus-secreting epithelial models (HT29 monolayers and HT29/Caco-2 co-culture monolayers). Exposure of BM-covered cells to constituents from FeSSIF revealed that it comprised a strong, hydrophilic barrier effect as 90% of BM-covered cells remained viable for >4 h, and the permeation rate of hydrophobic drugs was reduced. In contrast, the permeation rate of hydrophilic drugs was largely unaffected. Control monolayers displayed a loss of barrier function and Caco-2 cell monolayers surface-lined with BM constitute a valuable in vitro model that makes it possible to mimic intestinal fed state conditions when studying drug permeation. Copyright © 2017. Published by Elsevier B.V.

  6. Contextual niche signals towards colorectal tumor progression by mesenchymal stem cell in the mouse xenograft model.

    Science.gov (United States)

    Nakagaki, Suguru; Arimura, Yoshiaki; Nagaishi, Kanna; Isshiki, Hiroyuki; Nasuno, Masanao; Watanabe, Shuhei; Idogawa, Masashi; Yamashita, Kentaro; Naishiro, Yasuyoshi; Adachi, Yasushi; Suzuki, Hiromu; Fujimiya, Mineko; Imai, Kohzoh; Shinomura, Yasuhisa

    2015-09-01

    The role of mesenchymal stem/stromal cells (MSCs) in tumorigenesis remains controversial. This study aimed to determine whether heterotypic interactions between MSCs and colon cancer cells can supply contextual signals towards tumor progression. Xenografts consisting of co-implanted human colorectal cancer cells with rat MSCs in immunodeficient mice were evaluated by tumor progression, angiogenic profiles, and MSC fate. Furthermore, we investigated how MSCs function as a cancer cell niche by co-culture experiments in vitro. Tumor growth progressed in two ways, either independent of or dependent on MSCs. Such cell line-specific dependency could not be explained by host immune competency. COLO 320 xenograft angiogenesis was MSC-dependent, but less dependent on vascular endothelial growth factor (VEGF), whereas HT-29 angiogenesis was not MSC-dependent, but was VEGF-dependent. MSCs and COLO 320 cells established a functional positive feedback loop that triggered formation of a cancer cell niche, leading to AKT activation. Subsequently, MSCs differentiated into pericytes that enhanced angiogenesis as a perivascular niche. In contrast, the MSC niche conferred an anti-proliferative property to HT-29 cells, through mesenchymal-epithelial transition resulting in p38 activation. In conclusion, MSCs demonstrate pleiotropic capabilities as a cancer cell or perivascular niche to modulate colorectal cancer cell fate in a cell line-dependent manner in a xenogeneic context.

  7. Antioxidant effectiveness of coffee extracts and selected constituents in cell-free systems and human colon cell lines.

    Science.gov (United States)

    Bakuradze, Tamara; Lang, Roman; Hofmann, Thomas; Stiebitz, Herbert; Bytof, Gerhard; Lantz, Ingo; Baum, Matthias; Eisenbrand, Gerhard; Janzowski, Christine

    2010-12-01

    Epidemiological studies suggest that coffee can reduce the risk of degenerative diseases such as diabetes type 2, cardiovascular disease and cancer. These beneficial effects have partly been attributed to the antioxidant activity of coffee. We determined composition and antioxidant potential of differentially roasted coffee extracts and investigated the impact of selected original constituents and roast products. Parameters studied were direct antioxidant activity (trolox equivalent antioxidant capacity/oxygen radical absorbing capacity), cellular reactive oxygen species (ROS) level, DNA damage and protein expression of NAD(P)H: quinone oxidoreductase, γ-glutamylcysteine ligase and glutathione reductase in HT-29/Caco-2 cells at 24-h incubation. All extracts showed distinct direct antioxidant activity: medium roasts>light roast AB1 (caffeoylquinic acid (CQA)-rich Arabica Brazil extract); dark roast AB2 (N-methylpyridinium (NMP)-rich Arabica Brazil extract), and diminished t-butylhydroperoxide-induced ROS level in HT-29 cells (AB2>medium roasts>AB1). NAD(P)H:quinone oxidoreductase 1 expression and γ-glutamylcysteine ligase expression were distinctly induced by AB1 and 5-CQA, but not by AB2 and NMP. 5-CQA and caffeic acid exhibited highest trolox equivalent antioxidant capacity/oxygen radical absorbing capacity values (5-CQA: 1.3/3.5 mM and caffeic acid: 1.3/3.9 mM trolox); ROS level was distinctly diminished by 5-CQA (≥3 μM), catechol (30 μM) and trigonelline (≥30 μM), whereas menadione-induced DNA damage in Caco-2 cells was reduced by NMP compounds (1-30 μM). The results emphasize that both original constituents and roast products contribute to the cellular antioxidant effectiveness of coffee. Copyright © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Genomic characterisation of acral melanoma cell lines.

    Science.gov (United States)

    Furney, Simon J; Turajlic, Samra; Fenwick, Kerry; Lambros, Maryou B; MacKay, Alan; Ricken, Gerda; Mitsopoulos, Costas; Kozarewa, Iwanka; Hakas, Jarle; Zvelebil, Marketa; Lord, Christopher J; Ashworth, Alan; Reis-Filho, Jorge S; Herlyn, Meenhard; Murata, Hiroshi; Marais, Richard

    2012-07-01

    Acral melanoma is a rare melanoma subtype with distinct epidemiological, clinical and genetic features. To determine if acral melanoma cell lines are representative of this melanoma subtype, six lines were analysed by whole-exome sequencing and array comparative genomic hybridisation. We demonstrate that the cell lines display a mutation rate that is comparable to that of published primary and metastatic acral melanomas and observe a mutational signature suggestive of UV-induced mutagenesis in two of the cell lines. Mutations were identified in oncogenes and tumour suppressors previously linked to melanoma including BRAF, NRAS, KIT, PTEN and TP53, in cancer genes not previously linked to melanoma and in genes linked to DNA repair such as BRCA1 and BRCA2. Our findings provide strong circumstantial evidence to suggest that acral melanoma cell lines and acral tumours share genetic features in common and that these cells are therefore valuable tools to investigate the biology of this aggressive melanoma subtype. Data are available at: http://rock.icr.ac.uk/collaborations/Furney_et_al_2012/. © 2012 John Wiley & Sons A/S.

  9. Metronidazole affects breast cancer cell lines.

    Science.gov (United States)

    Sadowska, A; Prokopiuk, S; Miltyk, W; Surażyński, A; Konończuk, J; Sawicka, D; Car, H

    2013-01-01

    The aim of our study was to evaluate the impact of metronidazole (MTZ) on cytotoxicity and DNA synthesis in MCF-7 (estrogen receptor positive) and MDA-MB-231 (estrogen receptor negative) breast cancer cell lines. Toxicity of MTZ was determined by MTT test. MCF-7 and MDA-MB-231 cells were incubated with metronidazole used in different concentrations for 24, 48 and 72 hours. The effect of MTZ on DNA synthesis was measured as [3H]-thymidine incorporation. We showed that MTZ in concentration 250 μg/ml significantly increases the growth of MCF-7 cell lines after 24 hours of incubation, but it reduces cell viability in concentrations 1 and 10 μg/ml 72 hours after the drug application. Significant increase of MDA-MB-231 cell viability was obtained in MTZ concentration of 250 μg/ml after 24 and 72 hours. The increase of [3H]-thymidine incorporation in MCF-7 cell line treated with MTZ in concentration 250 μg/ml was statistically significant after 24 hours. Great suppression of cell proliferation was obtained in MDA-MB-231 breast cell line after application of the following concentrations of MTZ: 0.1 μg/ml (after 24 hours) and 0.1, 10, 50, 250 μg/ml (after 72h). We found that metronidazole exerts different dose- and time- dependent effects on human breast cancer cell lines characterized by presence or absence of estrogen receptors. We suggest that these discrepancies may be influenced by the estrogen signaling.

  10. 5-Fluorouracil modulation of radiosensitivity in cultured human carcinoma cells

    International Nuclear Information System (INIS)

    Smalley, S.R.; Kimler, B.F.; Evans, R.G.

    1991-01-01

    We evaluated conventional pulse exposure versus continuous exposure models of 5-fluorouracil (5-FU) radiosensitization in HT-29 (human colon adenocarcinoma) and DU-145 (human prostate cancer adenocarcinoma) cell lines. Cell survival following treatment with drug and/or radiation was determined by colony formation assays. Radiation was delivered either by itself, approximately midway through a 1-hr exposure to 5-FU (10 micrograms/ml), or at various times following initiation of exposure to 5-FU (0.5 microgram/ml) present throughout the entire period of incubation. Drug concentrations were selected to approximate those achieved in vivo in humans. HT-29 cells showed a plating efficiency of 87% and similar cytotoxicity (survival reduced to 0.57-0.71) for all 5-FU conditions. The Do's of the radiation survival curves were not different for 1 hr of 5-FU exposure versus radiation alone. However, continuous exposure conditions demonstrated statistically significantly different Do's from radiation alone and pulse 5-FU exposure. DU-145 cells displayed a plating efficiency of 17% and cytotoxicities of 0.10-0.91 for the 5-FU conditions. DU-145 cells showed different radiation 5-FU interactions: 5-FU produced statistically significant changes in Do well as the differences between cell lines insofar as their radiosensitization by 5-FU underscore the caution required in extrapolating these radiobiologic models to the clinical setting

  11. Cell Line Data Base: structure and recent improvements towards molecular authentication of human cell lines.

    Science.gov (United States)

    Romano, Paolo; Manniello, Assunta; Aresu, Ottavia; Armento, Massimiliano; Cesaro, Michela; Parodi, Barbara

    2009-01-01

    The Cell Line Data Base (CLDB) is a well-known reference information source on human and animal cell lines including information on more than 6000 cell lines. Main biological features are coded according to controlled vocabularies derived from international lists and taxonomies. HyperCLDB (http://bioinformatics.istge.it/hypercldb/) is a hypertext version of CLDB that improves data accessibility by also allowing information retrieval through web spiders. Access to HyperCLDB is provided through indexes of biological characteristics and navigation in the hypertext is granted by many internal links. HyperCLDB also includes links to external resources. Recently, an interest was raised for a reference nomenclature for cell lines and CLDB was seen as an authoritative system. Furthermore, to overcome the cell line misidentification problem, molecular authentication methods, such as fingerprinting, single-locus short tandem repeat (STR) profile and single nucleotide polymorphisms validation, were proposed. Since this data is distributed, a reference portal on authentication of human cell lines is needed. We present here the architecture and contents of CLDB, its recent enhancements and perspectives. We also present a new related database, the Cell Line Integrated Molecular Authentication (CLIMA) database (http://bioinformatics.istge.it/clima/), that allows to link authentication data to actual cell lines.

  12. Bisphenol A Disrupts Transcription and Decreases Viability in Aging Vascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Edna Ribeiro-Varandas

    2014-09-01

    Full Text Available Bisphenol A (BPA is a widely utilized endocrine disruptor capable of mimicking endogenous hormones, employed in the manufacture of numerous consumer products, thereby interfering with physiological cellular functions. Recent research has shown that BPA alters epigenetic cellular mechanisms in mammals and may be correlated to enhanced cellular senescence. Here, the effects of BPA at 10 ng/mL and 1 µg/mL, concentrations found in human samples, were analyzed on HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR transcriptional analysis of the Long Interspersed Element-1 (LINE-1 retroelement showed that BPA induces global transcription deregulation in both cell lines, although with more pronounced effects in HUVEC cells. Whereas there was an increase in global transcription in HT29 exclusively after 24 h of exposure, this chemical had prolonged effects on HUVEC. Immunoblotting revealed that this was not accompanied by alterations in the overall content of H3K9me2 and H3K4me3 epigenetic marks. Importantly, cell viability assays and transcriptional analysis indicated that prolonged BPA exposure affects aging processes in senescent HUVEC. To our knowledge this is the first report that BPA interferes with senescence in primary vascular endothelial cells, therefore, suggesting its association to the etiology of age-related human pathologies, such as atherosclerosis.

  13. Bisphenol A Disrupts Transcription and Decreases Viability in Aging Vascular Endothelial Cells

    Science.gov (United States)

    Ribeiro-Varandas, Edna; Pereira, H. Sofia; Monteiro, Sara; Neves, Elsa; Brito, Luísa; Boavida Ferreira, Ricardo; Viegas, Wanda; Delgado, Margarida

    2014-01-01

    Bisphenol A (BPA) is a widely utilized endocrine disruptor capable of mimicking endogenous hormones, employed in the manufacture of numerous consumer products, thereby interfering with physiological cellular functions. Recent research has shown that BPA alters epigenetic cellular mechanisms in mammals and may be correlated to enhanced cellular senescence. Here, the effects of BPA at 10 ng/mL and 1 µg/mL, concentrations found in human samples, were analyzed on HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC). Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) transcriptional analysis of the Long Interspersed Element-1 (LINE-1) retroelement showed that BPA induces global transcription deregulation in both cell lines, although with more pronounced effects in HUVEC cells. Whereas there was an increase in global transcription in HT29 exclusively after 24 h of exposure, this chemical had prolonged effects on HUVEC. Immunoblotting revealed that this was not accompanied by alterations in the overall content of H3K9me2 and H3K4me3 epigenetic marks. Importantly, cell viability assays and transcriptional analysis indicated that prolonged BPA exposure affects aging processes in senescent HUVEC. To our knowledge this is the first report that BPA interferes with senescence in primary vascular endothelial cells, therefore, suggesting its association to the etiology of age-related human pathologies, such as atherosclerosis. PMID:25207595

  14. Epigenetic mechanisms involved in differential MDR1 mRNA expression between gastric and colon cancer cell lines and rationales for clinical chemotherapy

    Directory of Open Access Journals (Sweden)

    Kim Kyung-Jong

    2008-08-01

    Full Text Available Abstract Background The membrane transporters such as P-glycoprotein (Pgp, the MDR1 gene product, are one of causes of treatment failure in cancer patients. In this study, the epigenetic mechanisms involved in differential MDR1 mRNA expression were compared between 10 gastric and 9 colon cancer cell lines. Methods The MDR1 mRNA levels were determined using PCR and real-time PCR assays after reverse transcription. Cytotoxicity was performed using the MTT assay. Methylation status was explored by quantification PCR-based methylation and bisulfite DNA sequencing analyses. Results The MDR1 mRNA levels obtained by 35 cycles of RT-PCR in gastric cancer cells were just comparable to those obtained by 22 cycles of RT-PCR in colon cancer cells. Real-time RT-PCR analysis revealed that MDR1 mRNA was not detected in the 10 gastric cancer cell lines but variable MDR1 mRNA levels in 7 of 9 colon cancer cell lines except the SNU-C5 and HT-29 cells. MTT assay showed that Pgp inhibitors such as cyclosporine A, verapamil and PSC833 sensitized Colo320HSR (colon, highest MDR1 expression but not SNU-668 (gastric, highest and SNU-C5 (gastric, no expression to paclitaxel. Quantification PCR-based methylation analysis revealed that 90% of gastric cancer cells, and 33% of colon cancer cells were methylated, which were completely matched with the results obtained by bisulfite DNA sequencing analysis. 5-aza-2'-deoxcytidine (5AC, a DNA methyltransferase inhibitor increased the MDR1 mRNA levels in 60% of gastric cells, and in 11% of colon cancer cells. Trichostatin A (TSA, histone deacetylase inhibitor increased the MDR1 mRNA levels in 70% of gastric cancer cells and 55% of colon cancer cells. The combined treatment of 5AC with TSA increased the MDR1 mRNA levels additively in 20% of gastric cancer cells, but synergistically in 40% of gastric and 11% of colon cancer cells. Conclusion These results indicate that the MDR1 mRNA levels in gastric cancer cells are significantly

  15. Subcloning of ovarian cancer cell lines.

    Science.gov (United States)

    Grunt, T W

    2001-01-01

    Cellular heterogeneity of malignant tissues is a well-known phenomenon (1). Intralineal/intraclonal diversity may be explained in part by proposing the concept of a hierarchically ordered, differentiating and self-renewing stem cell system for transformed cell populations (2). However, in many solid tumors, the stem cells are not easily accessible to phenotypic identification. In the past, density gradient centrifugation was successfully used to separate cells from tumors and from cell lines into distinct subpopulations (3-5). Using Percoll density gradients, we isolated undifferentiated clonogenic tumor stem-cell fractions from HOC-7 human ovarian adenocarcinoma cells. In addition, we also identified a low-density cell subpopulation formed by large, vacuolated, slowly growing, adenoid differentiated cells with very low clonogenic activity (6-11). Further characterization of these cell fractions in terms of stability of the isolated phenotypes is essential for the assessment of their biological significance. Subcloning of the isolated cell fractions by limiting dilution culture (12) followed by long-term culture yielded three permanent monoclonal sublines, which reveal a stable adenoid differentiated phenotype, and three subclones representing undifferentiated, clonogenic tumor stem cells (13). These data demonstrate that the isolated phenotypes represent distinct cell entities reflecting specific stages of ovarian surface epithelial cell differentiation.

  16. Human cytomegalovirus infection enhances cell proliferation, migration and upregulation of EMT markers in colorectal cancer-derived stem cell-like cells.

    Science.gov (United States)

    Teo, Wan Huai; Chen, Hsin-Pai; Huang, Jason C; Chan, Yu-Jiun

    2017-11-01

    Increasing evidence suggests a link between persistent human cytomegalovirus (HCMV) infection and cancer. Although the role of HCMV in cancer is still elusive, recent studies revealed the presence of HCMV nucleic acids and proteins in different cancer types such as glioblastoma, colorectal, breast, and prostate cancers, and neuroblastoma. Although HCMV may not be directly associated with the neoplastic transformation, the presence of HCMV DNA in the tumorous tissue has been associated with altered clinical outcomes in cancer patients. However, the mechanisms involved in the association between colorectal cancer (CRC) and HCMV are unclear. In this study, we investigated the influence of HCMV infection on CRC or their derived cells. Proliferation and migration assays revealed a high infection efficiency in CRC-derived HT29 and SW480 'stem‑like' cells. After 24, 48 and 72 h of HCMV infection, both HT29 and SW480 parental and stem‑like cells showed a significant increase in cell proliferation and viability (pcell migration. These results demonstrate a significant phenotypic alteration in the CRC cell line upon HCMV infection. Using epithelial to mesenchymal transition (EMT) assays, we demonstrated that the EMT markers and driver genes were upregulated during the virus infection. The WNT signaling pathway, which is associated with the proliferation and migration of CRC cells, was upregulated (6-fold) in HCMV-infected cells as compared to the non‑infected cells at day 7 from infection.

  17. SN38 conjugated hyaluronic acid gold nanoparticles as a novel system against metastatic colon cancer cells.

    Science.gov (United States)

    Hosseinzadeh, Hosniyeh; Atyabi, Fatemeh; Varnamkhasti, Behrang Shiri; Hosseinzadeh, Reza; Ostad, Seyed Nasser; Ghahremani, Mohammad Hossein; Dinarvand, Rassoul

    2017-06-30

    Combination of chemotherapy and photothermal therapy has been proposed for better treatment of metastatic colon cancer. In this study SN38, a highly potent cytotoxic agent, was conjugated to negatively charged hyaluronic acid (HA), which was deposited on the surface of the positively charged gold nanoparticles via electrostatic interaction. The drug conjugation and its interaction with gold nanoparticles were verified by 1 H NMR and UV-vis spectroscopies, respectively. The prepared SN38-HA gold NPs are negatively charged spherical nanoparticles with an average size of 75±10nm. In vitro release study revealed that drug release in acidic conditions (pH 5.2) was faster than that in physiological pH. Red light emitting diode (LED, 630nm, 30mW) was used as a light source for photothermal experiments. The drug release in acidic conditions was increased up to 30% using red LED illumination (6min) in comparison with experiment carried out indark. The cytotoxicity study on MUC1 positive HT29, SW480 colon cancer cells and MUC1 negative CHO cells, showed higher toxicity of the nanoparticles on HT29 and SW480 cell lines compared to CHO cells. Confocal microscopy images along with flow cytometry analysis confirm the cytotoxicity results. The incubation time for reaching IC50 decreases from 48h to 24h by LED illumination after nanoparticle treatment. Migratory potential of the HT29 and SW480 cell lines was reduced by co-application of SN38-HA gold NPs and LED radiation. Also anti-proliferative study indicates that LED radiation has increased the cytotoxicity of the nanoparticles and this effect is remained up to 8days. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Hiwi Promotes the Proliferation of Colorectal Cancer Cells via Upregulating Global DNA Methylation

    Directory of Open Access Journals (Sweden)

    Lin Yang

    2015-01-01

    Full Text Available Hiwi is well known for its role in stem cell renewal, maintaining the resting stage, and downregulating cell cycle of stem cells via RNA silencing. And Hiwi overexpression has been recognized in several types of cancers. In the present study, we examined the Hiwi expression in colorectal cancer (CRC specimens in both mRNA and protein levels via real-time quantitative PCR, western blot assay, and immunohistochemical staining. Then we explored the role of Hiwi in the cancer cell proliferation and in the DNA methylation in human CRC Caro-2 and HT-29 cell lines. Results demonstrated that both mRNA and protein levels of Hiwi were significantly higher in 38 CRC tissues than in 38 peritumor tissues. Moreover, the Hiwi overexpression with an adenovirus vector significantly promoted the proliferation of Caro-2 and HT-29 cells, associated with significant increase in the global DNA methylation levels. And the chemical inhibition of DNA methylation significantly restrained such proliferation promotion. In summary, we confirmed that Hiwi was overexpressed in CRC tissues and that the forced Hiwi overexpression promoted the proliferation and global DNA methylation of CRC cell lines. Our results imply for the first time that Hiwi promotes the proliferation of CRC cells via promoting global DNA methylation.

  19. (Asteraceae) Fraction against Human Cancer Cell Lines

    African Journals Online (AJOL)

    Purpose: To investigate the anti-proliferative and apoptotic activity of crude and dichloromethane fraction of A. sieberi against seven cancer cell lines (Colo20, HCT116, DLD, MCF7, Jurkat, HepG2 and L929). Methods: A. sieberi was extracted with methanol and further purification was carried out using liquidliquid extraction ...

  20. Breast cancer cell lines: friend or foe?

    International Nuclear Information System (INIS)

    Burdall, Sarah E; Hanby, Andrew M; Lansdown, Mark RJ; Speirs, Valerie

    2003-01-01

    The majority of breast cancer research is conducted using established breast cancer cell lines as in vitro models. An alternative is to use cultures established from primary breast tumours. Here, we discuss the pros and cons of using both of these models in translational breast cancer research

  1. Efecto citotóxico del extracto metanólico de tres ecotipos de Lepidium peruvianum Chacón sobre líneas celulares HeLa y HT-29

    Directory of Open Access Journals (Sweden)

    Libertad Alzamora

    2013-05-01

    Full Text Available La búsqueda de compuestos naturales con actividad citotóxica y antitumoral es una de las prioridades actuales de la lucha contra el cáncer; motivo por el cual el objetivo del presente trabajo fue evaluar la actividad citotóxica de los extractos metanólicos (EM de los ecotipos negro, morado y amarillo de Lepidium peruvianum, Chacón (conocida también como Lepidium meyenii Walp. (maca sobre las líneas celulares HeLa (Human Epithelial Carcinoma y HT-29 (Human Colon Adenocarcinoma. Se determinó que la concentración inhibitoria del 50% del crecimiento celular (IC50 para la línea celular HT-29, con los ecotipos negro, morado y amarillo fue de 8,32 mg/ml, 9,28 mg/ml y 0,487 mg/ml respectivamente, mientras que para la línea celular HeLa fue de 2,4 mg/ml, 1,93 mg/ml y 0,66 mg/ml respectivamente. Adicionalmente, se evaluó un EM del ecotipo amarillo con dos años de almacenamiento (10 ºC determinándose como IC50 4,29 mg/ml para HT-29 y 4,17 mg/ml para HeLa. Se concluye que el efecto citotóxico del ecotipo amarillo sobre HT-29 y HeLa fue superior al mostrado por los ecotipos negro y morado; que la línea celular más sensible a los ecotipos amarillo, negro y morado es HeLa, y que el EM del ecotipo amarillo conservó sus propiedades citotóxicas pese al tiempo de almacenamiento, aunque éstas disminuyeron.

  2. Cytotoxicity of withasteroids: withametelin induces cell cycle arrest at G2/M phase and mitochondria-mediated apoptosis in non-small cell lung cancer A549 cells.

    Science.gov (United States)

    Rao, Poorna Chandra; Begum, Sajeli; Jahromi, Mohammad Ali Farboodniay; Jahromi, Zahra Hosseini; Sriram, Saketh; Sahai, Mahendra

    2016-09-01

    Considerable interest has been gained by withasteroids because of their structural uniqueness and wide spectrum of biological activities. However, limited systematic studies for proving their cytotoxic potential have so far been reported. Hence, an attempt was made to test the cytotoxicity of six withasteroids viz., withametelin (WM), withaphysalin D, withaphysalin E, 12-deoxywithastramonolide, Withaperuvin B, and physalolactone against A549, HT-29, and MDA-MB-231 cancer cell lines. Significant cytotoxic effect of WM against A549 cells (IC 50 value of 6.0 μM), MDA-MB-231 cells (IC 50 value of 7.6 μM), and HT-29 cells (IC 50 value of 8.2 μM) was observed. Withaperuvin B and physalolactone were found to be effective against MDA-MB-231 cells. The significantly active WM arrested the A549 cells at G2/M phase and downregulated the expression of G2/M regulatory proteins such as cdc2, cyclin B1, and cdc25C. Apoptosis induced by WM in A549 cells was associated with the generation of ROS and depletion of MMP. Furthermore, WM treatment resulted in Bax upregulation, Bcl-2 downregulation, translocation of cytochrome c to mitochondria, activation of caspase-9 and -3, and PARP cleavage corroborating the apoptosis induction through intrinsic apoptotic pathway. Thus, WM possessing broader cytotoxic effect is a promising lead molecule which has the potential to be developed as a new therapeutic agent for NSCLC.

  3. Boletus edulis ribonucleic acid - a potent apoptosis inducer in human colon adenocarcinoma cells.

    Science.gov (United States)

    Lemieszek, Marta Kinga; Ribeiro, Miguel; Guichard Alves, Helena; Marques, Guilhermina; Nunes, Fernando Milheiro; Rzeski, Wojciech

    2016-07-13

    Despite the large popularity of the Boletus edulis mushroom, little is known about its influence on human health and the possibilities of its therapeutic use. Nevertheless, several reports revealed the usefulness of biopolymers isolated from it in cancer treatment. Our previous studies have shown that B. edulis water soluble biopolymers are not toxic against normal colon epithelial cells (CCD841 CoTr) and at the same concentration range elicited a very prominent antiproliferative effect in colon cancer cells (LS180) which was accompanied with cell cycle arrest in the G0/G1 phase. The purpose of the present study was to verify the proapoptotic properties of a selected fraction from B. edulis - BE3, as well as determine its chemical nature. The BE3 fraction was extracted with hot water and purified by anion-exchange chromatography. Further chemical examinations revealed that BE3 consists mainly of ribonucleic acid (59.1%). The ability of BE3 to induce programmed cell death was examined in human colon cancer cell lines LS180 and HT-29 by measuring caspase activation, DNA fragmentation and expression of BAX, BCL2, TP53 and CDKN1A genes. The sensitivity of colon cancer cells with silenced BAX, TP53 and CDKN1A expression to BE3 treatment was also evaluated. We have demonstrated for the first time that the BE3 fraction is a potent apoptosis inducer in human colon cancer cells. The revealed mechanism of apoptosis triggering was dependent on the presence of functional p53 and consequently was a little different in investigated cell lines. Our results indicated that BE3 stimulated proapoptotic genes BAX (LS180, HT-29), TP53 (LS180) and CDKN1A (HT-29) while at the same time silenced the expression of the key prosurvival gene BCL2 (LS180, HT-29). The obtained results indicate the high therapeutic potential of the BE3 fraction against colon cancer, yet it is necessary to further confirm fraction efficacy and safety in animal and clinical studies.

  4. CRF2 signaling is a novel regulator of cellular adhesion and migration in colorectal cancer cells.

    Science.gov (United States)

    Ducarouge, Benjamin; Pelissier-Rota, Marjolaine; Lainé, Michèle; Cristina, Nadine; Vachez, Yvan; Scoazec, Jean-Yves; Bonaz, Bruno; Jacquier-Sarlin, Muriel

    2013-01-01

    Stress has been proposed to be a tumor promoting factor through the secretion of specific neuromediators, such as Urocortin2 and 3 (Ucn2/3), however its role in colorectal cancer (CRC) remains elusive. We observed that Ucn2/3 and their receptor the Corticotropin Releasing Factor receptor 2 (CRF2) were up-regulated in high grade and poorly differentiated CRC. This suggests a role for CRF2 in the loss of cellular organization and tumor progression. Using HT-29 and SW620 cells, two CRC cell lines differing in their abilities to perform cell-cell contacts, we found that CRF2 signals through Src/ERK pathway to induce the alteration of cell-cell junctions and the shuttle of p120ctn and Kaiso in the nucleus. In HT-29 cells, this signaling pathway also leads to the remodeling of cell adhesion by i) the phosphorylation of Focal Adhesion Kinase and ii) a modification of actin cytoskeleton and focal adhesion complexes. These events stimulate cell migration and invasion. In conclusion, our findings indicate that CRF2 signaling controls cellular organization and may promote metastatic potential of human CRC cells through an epithelial-mesenchymal transition like process. This contributes to the comprehension of the tumor-promoting effects of stress molecules and designates Ucn2/3-CRF2 tandem as a target to prevent CRC progression and aggressiveness.

  5. Antiproliferative effects of β-blockers on human colorectal cancer cells.

    Science.gov (United States)

    Coelho, M; Moz, M; Correia, G; Teixeira, A; Medeiros, R; Ribeiro, L

    2015-05-01

    Colon cancer is the fourth and third most common cancer, respectively in men and women worldwide and its incidence is on the increase. Stress response has been associated with the incidence and development of cancer. The catecholamines (CA), adrenaline (AD) and noradrenaline (NA), are crucial mediators of stress response, exerting their effects through interaction with α- and β-adrenergic receptors (AR). Colon cancer cells express β-AR, and their activation has been implicated in carcinogenesis and tumor progression. Interest concerning the efficacy of β-AR blockers as possible additions to cancer treatment has increased. The aim of this study was to investigate the effect of several AR agonists and β-blockers following cell proliferation of HT-29 cells, a human colon adenocarcinoma cell line. For this purpose, HT-29 cells were incubated in the absence (control) or in the presence of the AR-agonists, AD, NA and isoprenaline (ISO) (0.1-100 µM) for 12 or 24 h. The tested AR agonists revealed proliferative effects on HT-29 cells. In order to study the effect of several β-blockers following proliferation induced by AR activation, the cells were treated with propranolol (PRO; 50 µM), carvedilol (CAR; 5 µM), atenolol (ATE; 50 µM), or ICI 118,551 (ICI; 5 µM) for 45 min prior, and simultaneously, to incubation with each of the AR agonists, AD and ISO, both at 1 and 10 µM. The results suggested that adrenergic activation plays an important role in colon cancer cell proliferation, most probably through β-AR. The β-blockers under study were able to reverse the proliferation induced by AD and ISO, and some of these blockers significantly decreased the proliferation of HT-29 cells. The elucidation of the intracellular pathways involved in CA-induced proliferation of colon cancer cells, and in the reversion of this effect by β-blockers, may contribute to identifying promising strategies in cancer treatment.

  6. Cellular radiosensitivity of small-cell lung cancer cell lines

    DEFF Research Database (Denmark)

    Krarup, M; Poulsen, H S; Spang-Thomsen, M

    1997-01-01

    PURPOSE: The objective of this study was to determine the radiobiological characteristics of a panel of small-cell lung cancer (SCLC) cell lines by use of a clonogenic assay. In addition, we tested whether comparable results could be obtained by employing a growth extrapolation method based...

  7. Cellular radiosensitivity of small-cell lung cancer cell lines

    DEFF Research Database (Denmark)

    Krarup, M; Poulsen, H S; Spang-Thomsen, M

    1997-01-01

    PURPOSE: The objective of this study was to determine the radiobiological characteristics of a panel of small-cell lung cancer (SCLC) cell lines by use of a clonogenic assay. In addition, we tested whether comparable results could be obtained by employing a growth extrapolation method based...... on the construction of continuous exponential growth curves. METHODS AND MATERIALS: Fifteen SCLC cell lines were studied, applying a slightly modified clonogenic assay and a growth extrapolation method. A dose-survival curve was obtained for each experiment and used for calculating several survival parameters...... to calculate the surviving fraction after 2-Gy irradiation (SF2). RESULTS: In our investigation, the extrapolation method proved to be inappropriate for the study of in vitro cellular radiosensitivity due to lack of reproducibility. The results obtained by the clonogenic assay showed that the cell lines...

  8. Impact of mesenchymal stem cell secreted PAI-1 on colon cancer cell migration and proliferation.

    Science.gov (United States)

    Hogan, Niamh M; Joyce, Myles R; Murphy, J Mary; Barry, Frank P; O'Brien, Timothy; Kerin, Michael J; Dwyer, Roisin M

    2013-06-14

    Mesenchymal Stem Cells are known to engraft and integrate into the architecture of colorectal tumours, with little known regarding their fate following engraftment. This study aimed to investigate mediators of Mesenchymal Stem Cell (MSC) and colon cancer cell (CCC) interactions. Mesenchymal Stem Cells and colon cancer cells (HT29 and HCT-116) were cultured individually or in co-culture on 3-dimensional scaffolds. Conditioned media containing all secreted factors was harvested at day 1, 3 and 7. Chemokine secretion and expression were analyzed by Chemi-array, ELISA (Macrophage migration inhibitory factor (MIF), plasminogen activator inhibitor type 1 (PAI-1)) and RQ-PCR. Colon cancer cell migration and proliferation in response to recombinant PAI-1, MSCs and MSCs+antibody to PAI-1 was analyzed using Transwell inserts and an MTS proliferation assay respectively. Chemi-array revealed secretion of a wide range of factors by each cell population, including PAI-1 and MIF. ELISA analysis revealed Mesenchymal Stem Cells to secrete the highest levels of PAI-1 (MSC mean 10.6 ng/mL, CCC mean 1.01 ng/mL), while colon cancer cells were the principal source of MIF. MSC-secreted PAI-1 stimulated significant migration of both CCC lines, with an antibody to the chemokine shown to block this effect (67-88% blocking,). A cell-line dependant effect on CCC proliferation was shown for Mesenchymal Stem Cell-secreted PAI-1 with HCT-116 cells showing decreased proliferation at all concentrations, and HT29 cells showing increased proliferation in the presence of higher PAI-1 levels. This is the first study to identify PAI-1 as an important mediator of Mesenchymal Stem Cell/colon cancer cell interactions and highlights the significant functional impact of Mesenchymal Stem Cell-secreted PAI-1 on colon cancer cells. Copyright © 2013 Elsevier Inc. All rights reserved.

  9. Induced pluripotent stem cell lines derived from human somatic cells.

    Science.gov (United States)

    Yu, Junying; Vodyanik, Maxim A; Smuga-Otto, Kim; Antosiewicz-Bourget, Jessica; Frane, Jennifer L; Tian, Shulan; Nie, Jeff; Jonsdottir, Gudrun A; Ruotti, Victor; Stewart, Ron; Slukvin, Igor I; Thomson, James A

    2007-12-21

    Somatic cell nuclear transfer allows trans-acting factors present in the mammalian oocyte to reprogram somatic cell nuclei to an undifferentiated state. We show that four factors (OCT4, SOX2, NANOG, and LIN28) are sufficient to reprogram human somatic cells to pluripotent stem cells that exhibit the essential characteristics of embryonic stem (ES) cells. These induced pluripotent human stem cells have normal karyotypes, express telomerase activity, express cell surface markers and genes that characterize human ES cells, and maintain the developmental potential to differentiate into advanced derivatives of all three primary germ layers. Such induced pluripotent human cell lines should be useful in the production of new disease models and in drug development, as well as for applications in transplantation medicine, once technical limitations (for example, mutation through viral integration) are eliminated.

  10. Positional isomerism markedly affects the growth inhibition of colon cancer cells by NOSH-aspirin: COX inhibition and modeling.

    Science.gov (United States)

    Vannini, Federica; Chattopadhyay, Mitali; Kodela, Ravinder; Rao, Praveen P N; Kashfi, Khosrow

    2015-12-01

    We recently reported the synthesis of NOSH-aspirin, a novel hybrid that releases both nitric oxide (NO) and hydrogen sulfide (H2S). In NOSH-aspirin, the two moieties that release NO and H2S are covalently linked at the 1, 2 positions of acetyl salicylic acid, i.e. ortho-NOSH-aspirin (o-NOSH-aspirin). In the present study, we compared the effects of the positional isomers of NOSH-ASA (o-NOSH-aspirin, m-NOSH-aspirin and p-NOSH-aspirin) to that of aspirin on growth of HT-29 and HCT 15 colon cancer cells, belonging to the same histological subtype, but with different expression of cyclooxygenase (COX) enzymes; HT-29 express both COX-1 and COX-2, whereas HCT 15 is COX-null. We also analyzed the effect of these compounds on proliferation and apoptosis in HT-29 cells. Since the parent compound aspirin, inhibits both COX-1 and COX-2, we also evaluated the effects of these compounds on COX-1 and COX-2 enzyme activities and also performed modeling of the interactions between the positional isomers of NOSH-aspirin and COX-1 and COX-2 enzymes. We observed that the three positional isomers of NOSH aspirin inhibited the growth of both colon cancer cell lines with IC50s in the nano-molar range. In particular in HT-29 cells the IC50s for growth inhibition were: o-NOSH-ASA, 0.04±0.011 µM; m-NOSH-ASA, 0.24±0.11 µM; p-NOSH-ASA, 0.46±0.17 µM; and in HCT 15 cells the IC50s for o-NOSH-ASA, m-NOSH-ASA, and p-NOSH-ASA were 0.062 ±0.006 µM, 0.092±0.004 µM, and 0.37±0.04 µM, respectively. The IC50 for aspirin in both cell lines was >5mM at 24h. The reduction of cell growth appeared to be mediated through inhibition of proliferation, and induction of apoptosis. All 3 positional isomers of NOSH-aspirin preferentially inhibited COX-1 over COX-2. These results suggest that the three positional isomers of NOSH-aspirin have the same biological actions, but that o-NOSH-ASA displayed the strongest anti-neoplastic potential. Copyright © 2015 The Authors. Published by Elsevier B.V. All

  11. Prolonged exposure of colon cancer cells to 5-fluorouracil nanoparticles improves its anticancer activity

    Directory of Open Access Journals (Sweden)

    Essam Tawfik

    2017-02-01

    Full Text Available In this study, we aimed to improve the anticancer effect of 5-FU on human colon cancer cell lines by incorporating in poly(d,l lactic-co-glycolic acid (PLGA nanoparticles (NPs. The 5-FU-PLGA NPs were prepared by nanoprecipitation technique. Prepared NPs were moderately dispersed with an average diameter of 133 ± 25.19 nm. Scanning Electron Microscope (SEM images revealed spherical structures with subtle surface irregularity. Free 5-FU dose–response curves were constructed (12.5–2000 μM using MTT assay on HCT 116 and HT-29 cell lines for 1, 3, and 5 days. The calculated IC50 on HCT 116 were 185 μM after 1 day, 11.3 μM after 3 days, and 1.48 μM after 5 days. On HT-29, IC50 was only reached after 5 days of 5-FU treatment (11.25 μM. The HCT 116 viability following treatment with 100 μM 5-FU in free or NPs forms for 3 days was 38.8% and 18.6%, respectively. Similarly, when 250 μM was applied, HCT 116 viability was 17.03% and 14.6% after treatment with free and NPs forms of 5-FU, respectively. Moreover, HT-29 cell viability after 250 μM 5-FU treatment in free or NPs forms was 55.45% and 34.01%, respectively. We also noticed that HCT 116 cells were more sensitive to 5-FU-PLGA NPs as compared to HT-29 cells. Overall, our data indicate that 5-FU activity is time dependent and the prolonged effects created by PLGA NPs may contribute, at least in part, to the noticed enhancement of the anticancer activity of 5-FU drug.

  12. Susceptibility testing of fish cell lines for virus isolation

    DEFF Research Database (Denmark)

    Ariel, Ellen; Skall, Helle Frank; Olesen, Niels Jørgen

    2009-01-01

    compare susceptibility between cell lines and between lineages within a laboratory and between laboratories (Inter-laboratory Proficiency Test). The objective being that the most sensitive cell line and lineages are routinely selected for diagnostic purposes.In comparing cell lines, we simulated "non......-cell-culture-adapted" virus by propagating the virus in heterologous cell lines to the one tested. A stock of test virus was produced and stored at - 80 °C and tests were conducted biannually. This procedure becomes complicated when several cell lines are in use and does not account for variation among lineages. In comparing...... cell lineages, we increased the number of isolates of each virus, propagated stocks in a given cell line and tested all lineages of that line in use in the laboratory. Testing of relative cell line susceptibility between laboratories is carried out annually via the Inter-laboratory Proficiency Test...

  13. Butyrate may enhance toxicological defence in primary, adenoma and tumor human colon cells by favourably modulating expression of glutathione S-transferases genes, an approach in nutrigenomics.

    Science.gov (United States)

    Pool-Zobel, Beatrice Louise; Selvaraju, Veeriah; Sauer, Julia; Kautenburger, Tanja; Kiefer, Jeannette; Richter, Konrad Klaus; Soom, Malle; Wölfl, Stefan

    2005-06-01

    Butyrate, formed by bacterial fermentation of plant foods, has been suggested to reduce colon cancer risks by suppressing the proliferation of tumor cells. In addition, butyrate has been shown to induce glutathione S-transferases (GSTs) in tumor cell lines, which may contribute to the detoxification of dietary carcinogens. We hypothesize that butyrate also affects biotransformation in non-transformed colon cells. Thus, we have investigated the gene expression of drug metabolism genes in primary human colon tissue, premalignant LT97 adenoma and HT29 tumor cells cultured in an appropriate medium+/-butyrate. A total of 96 drug metabolism genes (including 12 GSTs) spotted on cDNA macroarrays (Superarray; n = 3) were hybridized with biotin-labeled cDNA probes. To validate the expression detected with Superarray, samples of LT97 cells were also analyzed with high density microarrays (Affymetrix U133A), which include biotransformation genes that overlap with the set of genes represented on the Superarray. Relative expression levels were compared across colon samples and for each colon sample+/-butyrate. Compared with fresh tissue, 13 genes were downregulated in primary cells cultivated ex vivo, whereas 8 genes were upregulated. Several genes were less expressed in LT97 (40 genes) or in HT29 (41 and 17 genes, grown for 72 and 48 h, respectively) compared with primary colon tissue. Butyrate induced GSTP1, GSTM2, and GSTA4 in HT29 as previously confirmed by other methods (northern blot/qPCR). We detected an upregulation of GSTs (GSTA2, GSTT2) that are known to be involved in the defence against oxidative stress in primary cells upon incubation with butyrate. The changes in expression detected in LT97 by Superarray and Affymetrix were similar, confirming the validity of the results. We conclude that low GST expression levels were favourably altered by butyrate. An induction of the toxicological defence system possibly contributes to reported chemopreventive properties of

  14. Engineered cell lines for fish health research.

    Science.gov (United States)

    Collet, Bertrand; Collins, Catherine; Lester, Katherine

    2018-03-01

    As fish farming continues to increase worldwide, the related research areas of fish disease and immunology are also expanding, aided by the revolution in access to genomic information and molecular technology. The genomes of most fish species of economic importance are now available and annotation based on sequence homology with characterised genomes is underway. However, while useful, functional homology is more difficult to determine, there being a lack of widely distributed and well characterised reagents such as monoclonal antibodies, traditionally used in mammalian studies, to help with confirming functions and cellular interactions of fish molecules. In this context, fish cell lines and the possibility of their genetic engineering offer good prospects for studying functional genomics with respect to fish diseases. In this review, we will give an overview of available permanently genetically engineered fish cell lines, as cell-based reporter systems or platforms for expression of endogenous immune or pathogen genes, to investigate interactions and function. The advantages of such systems and the technical challenge for their development will be discussed. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

  15. Radiosensitivity of Human Melanoma Cell Lines

    Energy Technology Data Exchange (ETDEWEB)

    Bergoc, R. M.; Medina, V.; Cricco, G.; Mohamed, N.; Martin, G.; Nunez, M.; Croci, M.; Crescenti, E. J.; Rivera, E. S.

    2004-07-01

    Cutaneous melanoma is a skin cancer resulting from the malign transformation of skin-pigment cells, the melanocytes. The radiotherapy, alone or in combination with other treatment, is an important therapy for this disease. the objective of this paper was to determine in vitro the radiosensitivity of two human melanoma cell lines with different metastatic capability: WM35 and MI/15, and to study the effect of drugs on radiobiological parameters. The Survival Curves were adjusted to the mathematical Linear-quadratic model using GrapsPad Prism software. Cells were seeded in RPMI medium (3000-3500 cells/flask), in triplicate and irradiated 24 h later. The irradiation was performed using an IBL 437C H Type equipment (189 TBq, 7.7 Gy/min) calibrated with a TLD 700 dosimeter. The range of Doses covered from 0 to 10 Gy and the colonies formed were counted at day 7th post-irradiation. Results obtained were: for WM35, {alpha}=0.37{+-}0.07 Gy''-1 and {beta}=0.06{+-}0.02 Gy''-2, for M1/15m {alpha}=0.47{+-}0.03 Gy''-1 and {beta}=0.06{+-}0.01 Gy''-2. The {alpha}/{beta} values WM35: {alpha}/{beta} values WM35: {alpha}/{beta}=6.07 Gy and M1/15: {alpha}/{beta}{sub 7}.33 Gy were similar, independently of their metastatic capabillity and indicate that both lines exhibit high radioresistance. Microscopic observation of irradiated cells showed multinuclear cells with few morphologic changes non-compatible with apoptosis. By means of specific fluorescent dyes and flow cytometry analysis we determined the intracellular levels of the radicals superoxide and hydrogen peroxide and their modulation in response to ionizing radiation. The results showed a marked decreased in H{sub 2}O{sub 2} intracellular levels with a simultaneous increase in superoxide that will be part of a mechanism responsible for induction of cell radioresistance. This response triggered by irradiated cells could not be abrogated by different treatments like histamine or the

  16. Histone signature of metanephric mesenchyme cell lines.

    Science.gov (United States)

    McLaughlin, Nathan; Yao, Xiao; Li, Yuwen; Saifudeen, Zubaida; El-Dahr, Samir S

    2013-09-01

    The metanephric mesenchyme (MM) gives rise to nephrons, the filtering units of the mature kidney. The MM is composed of uninduced (Six2(high)/Lhx1(low)) and induced (Wnt-stimulated, Six2(low)/Lhx1(high)) cells. The global epigenetic state of MM cells is unknown, partly due to technical difficulty in isolating sufficient numbers of homogenous cell populations. We therefore took advantage of two mouse clonal cell lines representing the uninduced (mK3) and induced (mK4) metanephric mesenchyme (based on gene expression profiles and ability to induce branching of ureteric bud). ChIP-Seq revealed that whereas H3K4me3 active region "peaks" are enriched in metabolic genes, H3K27me3 peaks decorate mesenchyme and epithelial cell fate commitment genes. In uninduced mK3 cells, promoters of "stemness" genes (e.g., Six2, Osr1) are enriched with H3K4me3 peaks; these are lost in induced mK4 cells. ChIP-qPCR confirmed this finding and further demonstrated that G9a/H3K9me2 occupy the promoter region of Six2 in induced cells, consistent with the inactive state of transcription. Conversely, genes that mark the induced epithelialized state (e.g., Lhx1, Pax8), transition from a non-permissive to an active chromatin signature in mK3 vs. mK4 cells, respectively. Importantly, stimulation of Wnt signaling in uninduced mK3 cells provokes an active chromatin state (high H3K4me3, low H3K27me3), recruitment of β-catenin, and loss of pre-bound histone methyltransferase Ezh2 in silent induced genes followed by activation of transcription. We conclude that the chromatin signature of uninduced and induced cells correlates strongly with their gene expression states, suggesting a role of chromatin-based mechanisms in MM cell fate.

  17. Menadione inhibits MIBG uptake in two neuroendocrine cell lines

    NARCIS (Netherlands)

    Cornelissen, J.; Tytgat, G. A.; van den Brug, M.; van Kuilenburg, A. B.; Voûte, P. A.; van Gennip, A. H.

    1997-01-01

    In this paper we report on our studies of the effect of menadione on the uptake of MIBG in the neuroendocrine cell lines PC12 and SK-N-SH. Menadione inhibits the uptake of MIBG in both cell lines in a dose-dependent manner. Inhibition of MIBG uptake is most pronounced in the PC12 cell line.

  18. 77 FR 5489 - Identification of Human Cell Lines Project

    Science.gov (United States)

    2012-02-03

    ... selection of this technology over other possible candidates for this project include: (i) The ability to... cell line, whether the cell line is misidentified, cross- contaminated, or genetically unstable... database. Submission Process: Submitters should contact Margaret Kline with a list of proposed cell lines...

  19. Mechanisms involved in the cell cycle and apoptosis of HT-29 cells pre-treated with MK-886 prior to photodynamic therapy with hypericin

    Czech Academy of Sciences Publication Activity Database

    Kleban, J.; Mikeš, J.; Horváth, Viktor; Sačková, V.; Hofmanová, Jiřina; Kozubík, Alois; Fedoročko, P.

    2008-01-01

    Roč. 93, č. 2 (2008), s. 108-118 ISSN 1011-1344 R&D Projects: GA AV ČR(CZ) 1QS500040507 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : hypericin * MK-886 * ROS Subject RIV: BO - Biophysics Impact factor: 1.838, year: 2008

  20. Necrosis predominates in the cell death of human colon adenocarcinoma HT-29 cells treated under variable conditions of photodynamic therapy with hypericin

    Czech Academy of Sciences Publication Activity Database

    Mikeš, J.; Kleban, J.; Sačková, V.; Horváth, Viktor; Jamborová, E.; Vaculová, Alena; Kozubík, Alois; Hofmanová, Jiřina; Fedoročko, P.

    2007-01-01

    Roč. 6, č. 7 (2007), s. 758-766 ISSN 1474-905X R&D Projects: GA AV ČR(CZ) 1QS500040507 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : PDT * hypericin * p53 Subject RIV: BO - Biophysics Impact factor: 2.208, year: 2007

  1. Pre-treatment of HT-29 cells with 5-LOX inhibitor (MK-886) induces changes in cell cycle and increases apoptosis after photodynamic therapy with hypericin

    Czech Academy of Sciences Publication Activity Database

    Kleban, J.; Szilárdiová, B.; Mikeš, J.; Horváth, Viktor; Sačková, V.; Brezáni, P.; Hofmanová, Jiřina; Kozubík, Alois; Fedoročko, P.

    2006-01-01

    Roč. 84, č. 2 (2006), s. 79-88 ISSN 1011-1344 Institutional research plan: CEZ:AV0Z50040507 Keywords : photocytotoxicity * hypericin * lipoxygenase inhibitor MK-886 Subject RIV: BO - Biophysics Impact factor: 1.909, year: 2006

  2. Lower sensitivity of FHC fetal colon epithelial cells to photodynamic therapy compared to HT-29 colon adenocarcinoma cells despite higher intracellular accumulation of hypericin

    Czech Academy of Sciences Publication Activity Database

    Mikeš, J.; Hýžďalová, Martina; Kočí, Lenka; Jendželovský, R.; Koval, J.; Vaculová, Alena; Hofmanová, Jiřina; Kozubík, Alois; Fedoročko, P.

    2011-01-01

    Roč. 10, č. 4 (2011), s. 626-632 ISSN 1474-905X R&D Projects: GA ČR(CZ) GA301/07/1557 Grant - others:GA ČR(CZ) GA305/09/1526 Program:GA Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : photodynamic therapy * hypericin * colon cancer Subject RIV: BO - Biophysics Impact factor: 2.584, year: 2011

  3. The integrin-linked kinase gene up-regulated by hypoxia plays its pro-survival role in colorectal cancer cells.

    Science.gov (United States)

    Xiao, Lei; Yue, Xiaolong; Ming, Xiaodong; Xu, Lishan; Ding, Mingfeng; Xu, Jun; Liu, Qian

    2014-02-01

    Colorectal cancer (CRC) is a leading cause of cancer death in recent years. It is believed that there are hypoxic regions in both early and advanced stage of tumor and hypoxia is able to reinforce the aggressiveness of tumor cells and accelerate the progression of cancer. Until now the mechanisms by which hypoxia promotes the progression of CRC are far from well understood. Integrin-linked kinase (ILK) is a crucial mediator and over-expressed in CRC patients. But whether ILK is involved in the process that hypoxia promotes CRC cells growth and silencing the ILK gene results in CRC cells apoptosis is not clear. Lentivirus transfection, invasion assay, TUNEL assay, Bromodeoxyuridine incorporation and mitochondrial function assay were applied to demonstrate our hypothesis. In this study, we found that hypoxia induced the expression of ILK in a time-dependent manner, and after knocking down ILK expression with ILK shRNA, the cells proliferation promoted by hypoxia was inhibited in HT29 cell line. Moreover, blocking the ILK pathway led to caspase-3 and caspase-9 activations, the decrease of mitochondrial membrane potential, and cells apoptosis. And the inhibitory effects of hypoxia on cells apoptosis were mediated by the ILK pathway. In addition, hypoxia promoted HT29 cells metastasis and invasion through the ILK pathway. Therefore, we conclude that the CRC cells survival and invasion enhanced by hypoxia are mediated by ILK, and ILK may be an important potential therapeutic target for CRC.

  4. Isolation of a potential anticancer agent with protein phosphatase ...

    African Journals Online (AJOL)

    Results: It was found that a colorectal cancer cell line, HT-29, was susceptible to Fraction S1 and Citrinin 9318. A propidium iodide (PI)-incorporated DNA assay was used to show that there was G2/M arrest in HT-29 by Citrinin 9318. Conclusion: Citrinin 9318 inhibits the viability of HT-29 via mitotic block. The results suggest ...

  5. Autumn Royal and Ribier Grape Juice Extracts Reduced Viability and Metastatic Potential of Colon Cancer Cells

    Science.gov (United States)

    Valenzuela, Manuel; Bastias, Lorena; Montenegro, Iván; Werner, Enrique; Madrid, Alejandro; Godoy, Patricio

    2018-01-01

    Antioxidants are known to be beneficial to health. This paper evaluates the potential chemopreventive and anticancer properties of phenolic compounds present in grape juice extracts (GJE) from Autumn Royal and Ribier varieties. The effects of these GJE on viability (SRB day assay) and metastatic potential (migration and invasion parameters) of colon cancer cell lines HT-29 and SW-480 were evaluated. The effects of GJE on two matrix metalloproteinase gene expressions (MMP2 and MMP9) were also evaluated via qRT-PCR. In the former, GJE reduced cell viability in both cell lines in a dose-dependent manner. GJE treatment also reduced cell migration and invasion. Moreover, MMP-2 and MMP-9 gene expression diminished depending on extract and on cell type. Conclusions. These results provide novel information concerning anticancer properties of selected GJE by revealing selective cytotoxicity and the ability to reduce invasiveness of colon cancer cells. PMID:29552079

  6. A method for high purity intestinal epithelial cell culture from adult human and murine tissues for the investigation of innate immune function.

    Science.gov (United States)

    Graves, Christina L; Harden, Scott W; LaPato, Melissa; Nelson, Michael; Amador, Byron; Sorenson, Heather; Frazier, Charles J; Wallet, Shannon M

    2014-12-01

    Intestinal epithelial cells (IECs) serve as an important physiologic barrier between environmental antigens and the host intestinal immune system. Thus, IECs serve as a first line of defense and may act as sentinel cells during inflammatory insults. Despite recent renewed interest in IEC contributions to host immune function, the study of primary IEC has been hindered by lack of a robust culture technique, particularly for small intestinal and adult tissues. Here, a novel adaptation for culture of primary IEC is described for human duodenal organ donor tissue as well as duodenum and colon of adult mice. These epithelial cell cultures display characteristic phenotypes and are of high purity. In addition, the innate immune function of human primary IEC, specifically with regard to Toll-like receptor (TLR) expression and microbial ligand responsiveness, is contrasted with a commonly used intestinal epithelial cell line (HT-29). Specifically, TLR expression at the mRNA level and production of cytokine (IFNγ and TNFα) in response to TLR agonist stimulation is assessed. Differential expression of TLRs as well as innate immune responses to ligand stimulation is observed in human-derived cultures compared to that of HT-29. Thus, use of this adapted method to culture primary epithelial cells from adult human donors and from adult mice will allow for more appropriate studies of IECs as innate immune effectors. Published by Elsevier B.V.

  7. Microanalytical method development for Fe, Cu and Zn determination in colorectal cancer cells.

    Science.gov (United States)

    Polgári, Zsófia; Ajtony, Zsolt; Kregsamer, Peter; Streli, Christina; Mihucz, Victor G; Réti, Andrea; Budai, Barna; Kralovánszky, Judit; Szoboszlai, Norbert; Záray, Gyula

    2011-09-30

    Microanalytical methods suitable for the determination of Fe, Cu in HT-29 (human colon adenocarcinoma) cells treated with different iron compounds (Fe(II) sulfate, Fe(III) chloride, Fe(III) citrate and Fe(III) transferrin) and cultured in medium supplemented or not with 10% (v/v) fetal calf serum (FCS) by total reflection X-ray fluorescence spectrometry (TXRF) and simultaneous graphite furnace atomic absorption spectrometry (GF-AAS) were developed. The developed TXRF method was also suitable for Zn determination in the samples. The main advantage of the proposed methods is the execution of all sample preparation steps following incubation and prior to the elemental analysis in the same Eppendorf tubes. Sample preparation was performed at microscale (115 μL sample volume) with 65% nitric acid and 30% hydrogen peroxide. According to scanning electron microscopic measurements, the organic matrix of the cell samples could be eliminated to the extent that accurate results were obtained for Cu and Fe by analyzing the same samples by TXRF and GF-AAS. Concerning the iron uptake, HT-29 cells incubated in FCS-free medium contained Fe in cca. 5-50 times higher amounts compared to cells cultured in FCS supplemented medium. Pronounced differences in the iron uptake compared to the iron supply (inorganic vs. organic chelated as well as iron(II) vs. iron(III)) were observed in the case of cell lines incubated in FCS-free medium. Copyright © 2011 Elsevier B.V. All rights reserved.

  8. Pleiotropic effects of cancer cells' secreted factors on human stromal (mesenchymal) stem cells

    DEFF Research Database (Denmark)

    Al-toub, Mashael; Almusa, Abdulaziz; Almajed, Mohammed

    2013-01-01

    cells' secreted factors as represented by a panel of human cancer cell lines (breast (MCF7 and MDA-MB-231); prostate (PC-3); lung (NCI-H522); colon (HT-29) and head & neck (FaDu)) on the biological characteristics of MSCs. METHODS: Morphological changes were assessed using fluorescence microscopy......INTRODUCTION: Studying cancer tumors' microenvironment may reveal a novel role in driving cancer progression and metastasis. The biological interaction between stromal (mesenchymal) stem cells (MSCs) and cancer cells remains incompletely understood. Herein, we investigated the effects of tumor...... exposed to tumor CM, which was found to be positively regulated by FAK and MAPK signaling and negatively regulated by TGFβ signaling. Thus, our data support a model where MSCs could promote cancer progression through becoming pro-inflammatory cells within the cancer stroma....

  9. Nutraceutical properties of cumin residue generated from Ayurvedic industries using cell line models.

    Science.gov (United States)

    Arun, K B; Aswathi, U; Venugopal, V V; Madhavankutty, T S; Nisha, P

    2016-10-01

    Spent cumin (SC), generated from Ayurvedic industry, was evaluated for its nutraceutical potential in terms of antioxidant, antidiabetic and anticancer properties, and compared with that of the raw cumin (RC). SC and RC seeds were extracted with ethyl acetate (E) and methanol (M). SCM (methanol extract) were rich in p-coumaric acid, ferulic acid, ellagic acid and cinnamic acid (6.4445, 5.8286, 2.1519, 4.3085 mg/g dry extract). SCM reduced Fe 2+ ion (89.68 µM AA/g dry weight), scavenged DPPH radical (IC 50 -238.6 µg/mL), better α-amylase inhibition (IC 50 -337.22 µg/mL) and glucose uptake activity in 30.7% of L6 cells. SCM inhibited viability, retarded migration area up to 41.02%, arrested cell cycle at S phase and induced apoptosis in 2.45% of HT29 colon cancer cells. The results indicated that dietary interventions using nutraceutical food formulation made out of SC can play a significant role in the prevention and management of degenerative diseases.

  10. Libidibia ferrea presents antiproliferative, apoptotic and antioxidant effects in a colorectal cancer cell line.

    Science.gov (United States)

    Guerra, Andreza Conceição Véras de Aguiar; Soares, Luiz Alberto Lira; Ferreira, Magda Rhayanny Assunção; Araújo, Aurigena Antunes de; Rocha, Hugo Alexandre de Oliveira; Medeiros, Juliana Silva de; Cavalcante, Rômulo Dos Santos; Júnior, Raimundo Fernandes de Araújo

    2017-08-01

    Colorectal cancer is noted for being one of the most frequent of tumors, with expressive morbidity and mortality rates. In new drug discovery, plants stand out as a source capable of yielding safe and high-efficiency products. Well known in Brazilian popular medicine, Libidibia ferrea (Mart. Ex Tul.) L.P. Queiroz var. ferrea (better known as "ironwood" or "jucá"), has been used to treat a wide spectrum of conditions and to prevent cancer. Using methodologies that involved flow cytometry, spectrophotometry and RT-qPCR assays, crude extracts of the fruits of L. ferrea (20T, 40T, 60T and 80T) were evaluated at 24h and/or 48h for: their ability to inhibit cell proliferation; induce apoptosis through Bcl-2, caspase-3 and Apaf-1; their antioxidant activity and effects on important targets related to cell proliferation (EGFR and AKT) in the HT-29 human colorectal cancer lineage. The results revealed high antiproliferative potential as compared to the controls, induction of apoptosis through the intrinsic pathway, and probable tumor inhibition activity under the mediation of important targets in tumorigenesis. In addition, L. ferrea revealed antioxidant, lipid peroxidation and chemoprotective effects in healthy cells. Thus, L. ferrea derivatives have important anticancer effects, and may be considered promising candidate for colorectal cancer therapy. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  11. Cellular radiosensitivity of small-cell lung cancer cell lines

    International Nuclear Information System (INIS)

    Krarup, Marianne; Poulsen, Hans Skovgaard; Spang-Thomsen, Mogens

    1997-01-01

    Purpose: The objective of this study was to determine the radiobiological characteristics of a panel of small-cell lung cancer (SCLC) cell lines by use of a clonogenic assay. In addition, we tested whether comparable results could be obtained by employing a growth extrapolation method based on the construction of continuous exponential growth curves. Methods and Materials: Fifteen SCLC cell lines were studied, applying a slightly modified clonogenic assay and a growth extrapolation method. A dose-survival curve was obtained for each experiment and used for calculating several survival parameters. The multitarget single hit model was applied to calculate the cellular radiosensitivity (D 0 ), the capacity for sublethal damage repair (D q ), and the extrapolation number (n). Values for α and β were determined from best-fit curves according to the linear-quadratic model and these values were applied to calculate the surviving fraction after 2-Gy irradiation (SF 2 ). Results: In our investigation, the extrapolation method proved to be inappropriate for the study of in vitro cellular radiosensitivity due to lack of reproducibility. The results obtained by the clonogenic assay showed that the cell lines studied were radiobiologically heterogeneous with no discrete features of the examined parameters including the repair capacity. Conclusion: The results indicate that SCLC tumors per se are not generally candidates for hyperfractionated radiotherapy

  12. In vitro radiosensitivity of human leukemia cell lines

    International Nuclear Information System (INIS)

    Weichselbaum, R.R.; Greenberger, J.S.; Schmidt, A.; Karpas, A.; Moloney, W.C.; Little, J.B.

    1981-01-01

    The in vitro radiobiologic survival values (anti n, D 0 ) of four tumor lines derived from human hematopoietic tumors were studied. These cell lines were HL60 promyelocytic leukemia; K562 erythroleukemia; 45 acute lymphocytic leukemia; and 176 acute monomyelogenous leukemia. More cell lines must be examined before the exact relationship between in vitro radiosensitivity and clinical radiocurability is firmly established

  13. Green synthesis of gold nanoparticles using brown algae Cystoseira baccata: Its activity in colon cancer cells.

    Science.gov (United States)

    González-Ballesteros, N; Prado-López, S; Rodríguez-González, J B; Lastra, M; Rodríguez-Argüelles, M C

    2017-05-01

    This study is the first dealt with the use of brown macroalgae Cystoseira baccata (CB) extracts in obtaining gold nanoparticles (Au@CB) through an eco-friendly, fast, one-pot synthetic route. The formation of spherical, stable, polycrystalline nanoparticles with mean diameter of 8.4±2.2nm was demonstrated by UV-vis spectroscopy, TEM, HRTEM, STEM and zeta potential measurements. The extract appears to act as a protective agent where the particles are embedded, keeping them separated, avoiding aggregation and coalescence. The EELS and EDS analyses confirmed the elemental composition of the extract and nanoparticles. Moreover, the functional group of biomolecules present in CB and Au@CB were characterized by FTIR. The effects of CB extract and Au@CB were tested in vitro on the colon cancer cell lines HT-29 and Caco-2, as well as on normal primary neonatal dermal fibroblast cell line PCS-201-010. Results show a stronger cytotoxic effect against HT-29 than that on Caco-2; interestingly, a lack of toxicity on PCS-201-010 was obtained. Finally, the apoptotic activity was determined; Au@CB is able to induce apoptosis activation by the extrinsic and mitochondrial pathway in our CRC in vitro model. These encouraging results suggest that Au@CB has a significant potential for the treatment of colon rectal cancer. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Angiotensin II activates MAP kinase and NF-kappaB through angiotensin II type I receptor in human pancreatic cancer cells.

    Science.gov (United States)

    Amaya, Koji; Ohta, Tetsuo; Kitagawa, Hirohisa; Kayahara, Masato; Takamura, Hiroyuki; Fujimura, Takashi; Nishimura, Gen-Ichi; Shimizu, Koichi; Miwa, Koichi

    2004-10-01

    Pancreatic ductal cancer has higher angiotensin II concentrations compared with normal pancreas or other solid tumors. This study examined angiotensin II type 1 (AT1) receptor expression and the role of angiotensin II in proliferation and survival of human pancreatic cancer cells. All three pancreatic cancer cell lines studied, from well to poorly-differentiated types, HPAF-II, AsPC-1, and Panc-1, showed strong expression of AT1 receptor. In contrast, HT-29 human colon cancer cells showed extremely weak expression. Angiotensin II stimulated the growth of pancreatic cancer cells through MAP kinase activation but had no significant effect on proliferation of HT-29 colon cancer cells. In addition, angiotensin II significantly prevented cisplatin (CDDP)-induced apoptosis through NF-kappaB activation and the subsequent production of anti-apoptotic molecules, including survivin and Bcl-XL, in pancreatic cancer cells. These findings suggest that angiotensin II plays a role in the growth and chemoresistance of AT1-positive pancreatic cancer cells through its action as a potent mitogen and anti-apoptotic molecule.

  15. Tumour-Derived Interleukin-1 Beta Induces Pro-inflammatory Response in Human Mesenchymal Stem Cells

    DEFF Research Database (Denmark)

    Alajez, Nehad M; Al-toub, Mashael; Almusa, Abdulaziz

    ’ secreted factors as represented by a panel of human cancer cell lines (breast (MCF7 and MDA-MB-231); prostate (PC-3); lung (NCI-H522); colon (HT-29) and head & neck (FaDu)) on the biological characteristics of MSCs. Background Over the past several years, significant amount of research has emerged......, the goal of this study was to assess the cellular and molecular changes in MSCs in response to secreted factors present in conditioned media (CM) from a panel of human tumor cell lines covering a spectrum of human cancers (Breast, Prostate, Lung, colon, and head and neck). Research Morphological changes......Problem Studying cancer tumors microenvironment may reveal a novel role in driving cancer progression and metastasis. The biological interaction between stromal (mesenchymal) stem cells (MSCs) and cancer cells remains incompletely understood. Herein, we investigated the effects of tumor cells...

  16. Analysis of renal cancer cell lines from two major resources enables genomics-guided cell line selection

    Science.gov (United States)

    Sinha, Rileen; Winer, Andrew G.; Chevinsky, Michael; Jakubowski, Christopher; Chen, Ying-Bei; Dong, Yiyu; Tickoo, Satish K.; Reuter, Victor E.; Russo, Paul; Coleman, Jonathan A.; Sander, Chris; Hsieh, James J.; Hakimi, A. Ari

    2017-05-01

    The utility of cancer cell lines is affected by the similarity to endogenous tumour cells. Here we compare genomic data from 65 kidney-derived cell lines from the Cancer Cell Line Encyclopedia and the COSMIC Cell Lines Project to three renal cancer subtypes from The Cancer Genome Atlas: clear cell renal cell carcinoma (ccRCC, also known as kidney renal clear cell carcinoma), papillary (pRCC, also known as kidney papillary) and chromophobe (chRCC, also known as kidney chromophobe) renal cell carcinoma. Clustering copy number alterations shows that most cell lines resemble ccRCC, a few (including some often used as models of ccRCC) resemble pRCC, and none resemble chRCC. Human ccRCC tumours clustering with cell lines display clinical and genomic features of more aggressive disease, suggesting that cell lines best represent aggressive tumours. We stratify mutations and copy number alterations for important kidney cancer genes by the consistency between databases, and classify cell lines into established gene expression-based indolent and aggressive subtypes. Our results could aid investigators in analysing appropriate renal cancer cell lines.

  17. Novel carcinoembryonic-antigen-(CEA)-specific pretargeting system to assess tumor cell viability after irradiation of colorectal cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Meller, Birgit [Univ. Medical Center, Goettingen Univ. (Germany). Dept. of Nuclear Medicine; Halle Univ. (Germany). Dept. of Nuclear Medicine; Rave-Fraenck, Margarete [Univ. Medical Center, Goettingen Univ. (Germany). Dept. of Radiotherapy and Radiation Oncology; Breunig, Christian [Luebeck Univ. (Germany). Dept. of Radiology and Nuclear Medicine; Schirmer, Markus [Univ. Medical Center, Goettingen Univ. (Germany). Dept. of Clinical Pharmacology; Baehre, Manfred [Halle Univ. (Germany). Dept. of Nuclear Medicine; Nadrowitz, Roger [Luebeck Univ. (Germany). Dept. of Radiotherapy; Liersch, Torsten [Univ. Medical Center, Goettingen Univ. (Germany). Dept. of General Surgery; Meller, Johannes [Univ. Medical Center, Goettingen Univ. (Germany). Dept. of Nuclear Medicine

    2011-02-15

    Purpose: To date, no valid imaging modality exists for early response prediction to neoadjuvant radiochemotherapy in carcinoembryonic-antigen-(CEA)-expressing rectal cancers (UICC stages II and III). It is hypothesized that the uptake of an anti-CEA antibody is directly related to the number of viable tumor cells and may be quantified by immuno-positron emission tomography (immuno-PET). Therefore, we evaluated a novel pretargeting system using TF2, a humanized bispecific trivalent monoclonal antibody (mAb), directed against CEA and the IMP-288-peptide, a hapten for binding radiometals for imaging. Uptake and kinetics of the pretargeting system were investigated in vitro prior to and after irradiation. Methods: TF2 was labeled with {sup 131}I and IMP-288 with {sup 111}InCl{sub 3}. The colorectal cancer cell lines HT29, SW480, and T84 with known varying CEA expression were incubated ({<=} 72 hours) with {sup 131}I-TF2 or the TF2-{sup 111}In-IMP-288 pretargeting system. Parallel cultures were irradiated with 2-10 Gy high-energy photons. Tracer uptake, proliferation, apoptosis, and CEA-RNA expression of cancer cells were investigated. Results: The uptake of tracers was dependent on CEA expression and cell count of the cell lines (uptake/106 cells: 0.3% in HT29, 1.5% in SW480, and 14% in T84, p < 0.001). The TF2-{sup 111}In-IMP-288 pretargeting system showed a higher uptake after 4 and 72 hours compared to {sup 131}I-TF2 in parallel cultures. Only in one cell line (SW480) an increased apoptosis after irradiation could be detected. Irradiation increased dose dependently both the specific uptake of {sup 131}I-TF2 and of the TF2-{sup 111}In-IMP-288 system (4-fold in HT29 and T84 after 10 Gy (72 hours), p < 0.001). These results were CEA-mRNA independent. Conclusion: This novel pretargeting system allows the quantitative analysis of CEA-expressing colorectal cancer cells and represents a promising tool for evaluation of tumor cell viability after irradiation. (orig.)

  18. MODERATE CYTOTOXICITY OF PROANTHOCYANIDINS TO HUMAN TUMOR-CELL LINES

    NARCIS (Netherlands)

    KOLODZIEJ, H; HABERLAND, C; WOERDENBAG, HJ; KONINGS, AWT

    In the present study the cytotoxicity of 16 proanthocyanidins was evaluated in GLC(4), a human small cell lung carcinoma cell line, and in COLO 320, a human colorectal cancer cell line, using the microculture tetrazolium (MTT) assay. With IC50 values ranging from 18 to >200 mu m following continuous

  19. Modeling Adenovirus Latency in Human Lymphocyte Cell Lines ▿ †

    OpenAIRE

    Zhang, Yange; Huang, Wen; Ornelles, David A.; Gooding, Linda R.

    2010-01-01

    Species C adenovirus establishes a latent infection in lymphocytes of the tonsils and adenoids. To understand how this lytic virus is maintained in these cells, four human lymphocytic cell lines that support the entire virus life cycle were examined. The T-cell line Jurkat ceased proliferation and died shortly after virus infection. BJAB, Ramos (B cells), and KE37 (T cells) continued to divide at nearly normal rates while replicating the virus genome. Viral genome numbers peaked and then decl...

  20. Modulation of hypericin photodynamic therapy by pretreatment with 12 various inhibitors of arachidonic acid metabolism in colon adenocarcinoma HT-29 cells

    Czech Academy of Sciences Publication Activity Database

    Kleban, J.; Mikeš, J.; Szilárdiová, B.; Koval, J.; Sačková, V.; Solár, P.; Horváth, Viktor; Hofmanová, Jiřina; Kozubík, Alois; Fedoročko, P.

    2007-01-01

    Roč. 83, č. 5 (2007), s. 1174-1185 ISSN 0031-8655 R&D Projects: GA AV ČR(CZ) 1QS500040507 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : hypericin * photodynamic therapy * arachidonic acid inhibitors Subject RIV: BO - Biophysics Impact factor: 2.172, year: 2007

  1. Activity of glycogen synthase kinase-3á is down-regulated during transient differentiation of human colon cancer HT-29 cells

    Czech Academy of Sciences Publication Activity Database

    Tuháčková, Zdena; Šloncová, Eva; Hlaváček, Jan; Sovová, Vlasta; Velek, Jiří

    1999-01-01

    Roč. 6, č. 4 (1999), s. 827-832 ISSN 1021-335X R&D Projects: GA MZd IZ3517; GA ČR GV312/96/K205; GA ČR GA312/97/1188; GA AV ČR IPP2020702 Institutional research plan: CEZ:AV0Z5052915 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.659, year: 1999

  2. Drug efflux transporters, MRP1 and BCRP, affect the outcome of hypericin-mediated photodynamic therapy in HT-29 adenocarcinoma cells

    Czech Academy of Sciences Publication Activity Database

    Jendželovský, R.; Mikeš, J.; Koval, J.; Souček, Karel; Procházková, Jiřina; Kello, M.; Sačková, V.; Hofmanová, Jiřina; Kozubík, Alois; Fedoročko, P.

    2009-01-01

    Roč. 8, č. 12 (2009), s. 1716-1723 ISSN 1474-905X Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : hypericin * photodynamic therapy * ABC transporters Subject RIV: BO - Biophysics Impact factor: 2.708, year: 2009

  3. NF-kappaB is not directly responsible for photoresistance induced by fractionated light delivery in HT-29 colon adenocarcinoma cells

    Czech Academy of Sciences Publication Activity Database

    Kuliková, L.; Mikeš, J.; Hýžďalová, Martina; Palumbo, G.; Fedoročko, P.

    2010-01-01

    Roč. 86, č. 6 (2010), s. 1285-1293 ISSN 0031-8655 R&D Projects: GA ČR(CZ) GA301/07/1557 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : photodynamic therapy * hypericin * NF-kappaB Subject RIV: BO - Biophysics Impact factor: 2.679, year: 2010

  4. Pectic oligosaccharide structure-function relationships: prebiotics, inhibitors of Escherichia coli O157:H7 adhesion and reduction of Shiga toxin cytotoxicity in HT29 cells

    Science.gov (United States)

    Shiga toxin (Stx)-producing, food-contaminating Escherichia coli (STEC) is a major health concern. Plant-derived pectin and pectic-oligosaccharides (POS) that are abundant in biomass have been considered as prebiotics and for the protection of humans from Stx intoxication. Five structurally differ...

  5. Inhibition of Various Cancer Cells Proliferation of Bornyl Acetate and Essential Oil from Inula graveolens (Linnaeus Desf

    Directory of Open Access Journals (Sweden)

    Tunay Karan

    2018-05-01

    Full Text Available Inula species are medicinal and aromatic plants used for folk medicine extensively. In this work, hydrodistilled essential oil of Inula graveolens (Linnaeus Desf. analyzed by GC-MS which revealed that bornyl acetate was the major product (69.15%. Camphene was the second major compound (11.11%. Antiproliferative activity of the essential oil and bornyl acetate was investigated on HeLa (human cervix carcinoma, HT29 (human colon carcinoma , A549 (human lung carcinoma, MCF7 ( human breast adenocarcinoma cancer cells and FL (human amnion cells normal cells. The cytotoxicity was executed by a Lactate Dehydrogenase (LDH Cytotoxicity Detection Kit. Essential oil and bornyl acetate displayed the outstanding activities on HeLa (IC 50, 64.1, 72.0 µg/mL, HT29 (IC 50, 24.6, 60.5 µg/mL, A549 (IC 50, 28.3, 44.1 µg/mL, MCF-7 (IC 50, 66.5, 85.6 µg/mL, and FL (IC 50, 42.1, 50.6 µg/mL cell lines respectively.

  6. Isolation of a Wheat Cell Line with Altered Membrane Properties

    Science.gov (United States)

    Erdei, László; Vigh, László; Dudits, Dénes

    1982-01-01

    A spontaneous dimethylsulfoxide (DMSO)-tolerant cell line was isolated from a cell culture of wheat (Triticum monococcum L.). The tolerant cells were able to grow in the presence of 4% DMSO. Cells formed from protoplasts of the tolerant line required DMSO for division in culture medium of high osmotic value. Fatty acid composition and the molar ratio of phospholipids/sterols suggest a more ordered membrane structure in the tolerant line. Accordingly, a lower K+ influx rate was detected in the tolerant cells in comparison with the original line. These characteristics were maintained after 6 months' cultivation of the cells in DMSO-free growth medium. This suggested that genetic changes could be responsible for differences between the two cell lines. PMID:16662251

  7. The pursuit of ES cell lines of domesticated ungulates

    Science.gov (United States)

    In contrast to differentiated cells, embryonic stem cells (ESC) maintain an undifferentiated state, have the ability to self-renew, and exhibit pluripotency, i.e., they can give rise to most if not all somatic cell types and to the germ cells, egg and sperm. These characteristics make ES cell lines...

  8. Rab27A mediated by NF-κB promotes the stemness of colon cancer cells via up-regulation of cytokine secretion.

    Science.gov (United States)

    Feng, Feixue; Jiang, Yinghao; Lu, Huanyu; Lu, Xiaozhao; Wang, Shan; Wang, Lifeng; Wei, Mengying; Lu, Wei; Du, Zhichao; Ye, Zichen; Yang, Guodong; Yuan, Fang; Ma, Yanxia; Lei, Xiaoying; Lu, Zifan

    2016-09-27

    Recent evidences have unveiled critical roles of cancer stem cells (CSCs) in tumorigenicity, but how interactions between CSC and tumor environments help maintain CSC initiation remains obscure. The small GTPases Rab27A regulates autocrine and paracrine cytokines by monitoring exocytosis of extracellular vesicles, and is reported to promote certain tumor progression. We observe that overexpression of Rab27A increased sphere formation efficiency (SFE) by increasing the proportion of CD44+ and PKH26high cells in HT29 cell lines, and accelerating the growth of colosphere with higher percentage of cells at S phase. Mechanism study revealed that the supernatant derived from HT29 sphere after Rab27A overexpression was able to expand sphere numbers with elevated secretion of VEGF and TGF-β. In tumor implanting nude mice model, tumor initiation rates and tumor sizes were enhanced by Rab27A with obvious angiogenesis. As a contrast, knocking down Rab27A impaired the above effects. More importantly, the correlation between higher p65 level and Rab27A in colon sphere was detected, p65 was sufficient to induce up-regulation of Rab27A and a functional NF-κB binding site in the Rab27A promoter was demonstrated. Altogether, our findings reveal a unique mechanism that tumor environment related NF-κB signaling promotes various colon cancer stem cells (cCSCs) properties via an amplified paracrine mechanism regulated by higher Rab27A level.

  9. VOLIN and KJON-Two novel hyperdiploid myeloma cell lines.

    Science.gov (United States)

    Våtsveen, Thea Kristin; Børset, Magne; Dikic, Aida; Tian, Erming; Micci, Francesca; Lid, Ana H B; Meza-Zepeda, Leonardo A; Coward, Eivind; Waage, Anders; Sundan, Anders; Kuehl, W Michael; Holien, Toril

    2016-11-01

    Multiple myeloma can be divided into two distinct genetic subgroups: hyperdiploid (HRD) or nonhyperdiploid (NHRD) myeloma. Myeloma cell lines are important tools to study myeloma cell biology and are commonly used for preclinical screening and testing of new drugs. With few exceptions human myeloma cell lines are derived from NHRD patients, even though about half of the patients have HRD myeloma. Thus, there is a need for cell lines of HRD origin to enable more representative preclinical studies. Here, we present two novel myeloma cell lines, VOLIN and KJON. Both of them were derived from patients with HRD disease and shared the same genotype as their corresponding primary tumors. The cell lines' chromosomal content, genetic aberrations, gene expression, immunophenotype as well as some of their growth characteristics are described. Neither of the cell lines was found to harbor immunoglobulin heavy chain translocations. The VOLIN cell line was established from a bone marrow aspirate and KJON from peripheral blood. We propose that these unique cell lines may be used as tools to increase our understanding of myeloma cell biology. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  10. Expression of ICAM-1 in colon epithelial cells

    DEFF Research Database (Denmark)

    Vainer, Ben; Sørensen, Susanne; Seidelin, Jakob

    2003-01-01

    on monolayers of cancer cells. Conflicting results exist on epithelial ICAM-1 expression, and the aim of this study was to compare the expression in various models of colonic epithelium. MATERIALS AND METHODS: Colonic biopsies from four UC patients and four controls were examined by cryoimmuno......-electron microscopy using ICAM-1-antibodies. In four other controls, the epithelium was isolated from colonic biopsies, embedded in collagen, and evaluated similarly. Isolated crypts and cultured cancer cells were stimulated with interferon-gamma (IFN-gamma) or tumor necrosis factor-alpha (TNF-alpha). RESULTS: ICAM-1......, both colonocytes and HT29 cells were capable of expressing ICAM-1 on their apical membranes in response to supraphysiologic cytokine concentrations. These observations question the justification of extrapolating observations from colon cancer cell lines to in vivo inflammatory conditions....

  11. Poly(C)-binding protein 1 mediates drug resistance in colorectal cancer.

    Science.gov (United States)

    Guo, Jiani; Zhu, Changli; Yang, Kangqun; Li, Jin; Du, Nan; Zong, Mingzhu; Zhou, Jianwei; He, Jingdong

    2017-02-21

    Oxaliplatin (L-OHP) is standard treatment for colorectal cancer. However, resistance to L-OHP often leads to treatment failure or cancer relapse. Understanding of the mechanism underlying L-OHP resistance is important to overcome the resistance and improve colorectal cancer treatment. This study aimed to identify new proteins that mediates L-OHP resistance in colorectal cancer and elucidate their mode of function. HT-29 cells were exposed to gradually increased concentration of L-OHP to select L-OHP resistant HT-29/L-OHP cell line. Proteomic analysis of HT-29 and HT-29/L-OHP cells were performed to identify differentially expressed proteins, including Poly(C)-binding protein 1 (PCBP1). PCBP1 expression level in 20 cases of L-OHP sensitive patients and 20 cases of L-OHP refractory patients was analyzed by immunohistochemistry. Chemoresistance and Akt activation in HT-29 and HT-29/L-OHP cells were analyzed by MTT assay and Western blot analysis. We identified 37 proteins showing differential expression in HT-29/L-OHP and HT-29 cells. In particular, PCBP1 protein level increased 15.6 fold in HT-29/L-OHP cells compared to HT-29 cells. Knockdown of PCBP1 sensitized HT-29/L-OHP and HT-29 cells to L-OHP, while overexpression of PCBP1 increased L-OHP resistance in HT-29 cells. In addition, PCBP1 expression was significantly higher in tumor samples from L-OHP refractory patients than in those from L-OHP responsive patients. Furthermore, we found that knockdown of PCBP1 inhibited the activation of Akt in HT-29/L-OHP and HT-29 cells. In conclusion, our findings suggest that PCBP1 is a molecular marker of L-OHP resistance in colorectal cancer and a promising target for colorectal cancer therapy.

  12. Authentication of M14 melanoma cell line proves misidentification of MDA-MB-435 breast cancer cell line.

    Science.gov (United States)

    Korch, Christopher; Hall, Erin M; Dirks, Wilhelm G; Ewing, Margaret; Faries, Mark; Varella-Garcia, Marileila; Robinson, Steven; Storts, Douglas; Turner, Jacqueline A; Wang, Ying; Burnett, Edward C; Healy, Lyn; Kniss, Douglas; Neve, Richard M; Nims, Raymond W; Reid, Yvonne A; Robinson, William A; Capes-Davis, Amanda

    2018-02-01

    A variety of analytical approaches have indicated that melanoma cell line UCLA-SO-M14 (M14) and breast carcinoma cell line MDA-MB-435 originate from a common donor. This indicates that at some point in the past, one of these cell lines became misidentified, meaning that it ceased to correspond to the reported donor and instead became falsely identified (through cross-contamination or other means) as a cell line from a different donor. Initial studies concluded that MDA-MB-435 was the misidentified cell line and M14 was the authentic cell line, although contradictory evidence has been published, resulting in further confusion. To address this question, we obtained early samples of the melanoma cell line (M14), a lymphoblastoid cell line from the same donor (ML14), and donor serum preserved at the originator's institution. M14 samples were cryopreserved in December 1975, before MDA-MB-435 cells were established in culture. Through a series of molecular characterizations, including short tandem repeat (STR) profiling and cytogenetic analysis, we demonstrated that later samples of M14 and MDA-MB-435 correspond to samples of M14 frozen in 1975, to the lymphoblastoid cell line ML14, and to the melanoma donor's STR profile, sex and blood type. This work demonstrates conclusively that M14 is the authentic cell line and MDA-MB-435 is misidentified. With clear provenance information and authentication testing of early samples, it is possible to resolve debates regarding the origins of problematic cell lines that are widely used in cancer research. © 2017 The Authors International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.

  13. Authentication of M14 melanoma cell line proves misidentification of MDA‐MB‐435 breast cancer cell line

    Science.gov (United States)

    Korch, Christopher; Hall, Erin M.; Dirks, Wilhelm G.; Ewing, Margaret; Faries, Mark; Varella‐Garcia, Marileila; Robinson, Steven; Storts, Douglas; Turner, Jacqueline A.; Wang, Ying; Burnett, Edward C.; Healy, Lyn; Kniss, Douglas; Neve, Richard M.; Nims, Raymond W.; Reid, Yvonne A.; Robinson, William A.

    2017-01-01

    A variety of analytical approaches have indicated that melanoma cell line UCLA‐SO‐M14 (M14) and breast carcinoma cell line MDA‐MB‐435 originate from a common donor. This indicates that at some point in the past, one of these cell lines became misidentified, meaning that it ceased to correspond to the reported donor and instead became falsely identified (through cross‐contamination or other means) as a cell line from a different donor. Initial studies concluded that MDA‐MB‐435 was the misidentified cell line and M14 was the authentic cell line, although contradictory evidence has been published, resulting in further confusion. To address this question, we obtained early samples of the melanoma cell line (M14), a lymphoblastoid cell line from the same donor (ML14), and donor serum preserved at the originator's institution. M14 samples were cryopreserved in December 1975, before MDA‐MB‐435 cells were established in culture. Through a series of molecular characterizations, including short tandem repeat (STR) profiling and cytogenetic analysis, we demonstrated that later samples of M14 and MDA‐MB‐435 correspond to samples of M14 frozen in 1975, to the lymphoblastoid cell line ML14, and to the melanoma donor's STR profile, sex and blood type. This work demonstrates conclusively that M14 is the authentic cell line and MDA‐MB‐435 is misidentified. With clear provenance information and authentication testing of early samples, it is possible to resolve debates regarding the origins of problematic cell lines that are widely used in cancer research. PMID:28940260

  14. Improved replication efficiency of echovirus 5 after transfection of colon cancer cells using an authentic 5' RNA genome end methodology.

    Science.gov (United States)

    Israelsson, S; Sävneby, A; Ekström, J-O; Jonsson, N; Edman, K; Lindberg, A M

    2014-12-01

    Oncolytic virotherapy is a promising novel form of cancer treatment, but the therapeutic efficiency needs improvement. A potential strategy to enhance the therapeutic effect of oncolytic viruses is to use infectious nucleic acid as therapeutic agent to initiate an oncolytic infection, without administrating infectious viral particles. Here we demonstrate improved viral replication activation efficiency when transfecting cells with 5' end authentic in vitro transcribed enterovirus RNA as compared to genomic RNA with additional non-genomic 5' nucleotides generated by conventional cloning methods. We used echovirus 5 (E5) as an oncolytoc model virus due to its ability to replicate in and completely destroy five out of six colon cancer cell lines and kill artificial colon cancer tumors (HT29 spheroids), as shown here. An E5 infectious cDNA clone including a hammerhead ribozyme sequence was used to generate in vitro transcripts with native 5' genome ends. In HT29 cells, activation of virus replication is approximately 20-fold more efficient for virus genome transcripts with native 5' genome ends compared to E5 transcripts generated from a standard cDNA clone. This replication advantage remains when viral progeny release starts by cellular lysis 22 h post transfection. Hence, a native 5' genomic end improves infection activation efficacy of infectious nucleic acid, potentially enhancing its therapeutic effect when used for cancer treatment. The clone design with a hammerhead ribozyme is likely to be applicable to a variety of oncolytic positive sense RNA viruses for the purpose of improving the efficacy of oncolytic virotherapy.

  15. A factor converting viable but nonculturable Vibrio cholerae to a culturable state in eukaryotic cells is a human catalase.

    Science.gov (United States)

    Senoh, Mitsutoshi; Hamabata, Takashi; Takeda, Yoshifumi

    2015-08-01

    In our previous work, we demonstrated that viable but nonculturable (VBNC) Vibrio cholerae O1 and O139 were converted to culturable by coculture with eukaryotic cells. Furthermore, we isolated a factor converting VBNC V. cholerae to culturable (FCVC) from a eukaryotic cell line, HT-29. In this study, we purified FCVC by successive column chromatographies comprising UNO Q-6 anion exchange, Bio-Scale CHT2-1 hydroxyapatite, and Superdex 200 10/300 GL. Homogeneity of the purified FCVC was demonstrated by SDS-PAGE. Nano-LC MS/MS analysis showed that the purified FCVC was a human catalase. An experiment of RNAi knockdown of catalase mRNA from HT-29 cells and treatment of the purified FCVC with a catalase inhibitor, 3-amino-1,2,4-triazole confirmed that the FCVC was a catalase. A possible role of the catalase in converting a VBNC V. cholerae to a culturable state in the human intestine is discussed. © 2015 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  16. Derivation of the human embryonic stem cell line RCM1

    Directory of Open Access Journals (Sweden)

    P.A. De Sousa

    2016-03-01

    Full Text Available The human embryonic stem cell line RCM-1 was derived from a failed to fertilise egg undergoing parthenogenetic stimulation. The cell line shows normal pluripotency marker expression and differentiation to three germ layers in vitro and in vivo. It has a normal 46XX female karyotype and microsatellite PCR identity, HLA and blood group typing data is available.

  17. Beryllium-stimulated apoptosis in macrophage cell lines.

    Science.gov (United States)

    Sawyer, R T; Fadok, V A; Kittle, L A; Maier, L A; Newman, L S

    2000-08-21

    In vitro stimulation of bronchoalveolar lavage cells from patients with chronic beryllium disease (CBD) induces the production of TNF-alpha. We tested the hypothesis that beryllium (Be)-stimulated TNF-alpha might induce apoptosis in mouse and human macrophage cell lines. These cell lines were selected because they produce a range of Be-stimulated TNF-alpha. The mouse macrophage cell line H36.12j produces high levels of Be-stimulated TNF-alpha. The mouse macrophage cell line P388D.1 produces low, constitutive, levels of TNF-alpha and does not up-regulate Be-stimulated TNF-alpha production. The DEOHS-1 human CBD macrophage cell line does not produce constitutive or Be-stimulated TNF-alpha. Apoptosis was determined by microscopic observation of propidium iodide stained fragmented nuclei in unstimulated and BeSO(4)-stimulated macrophage cell lines. BeSO(4) induced apoptosis in all macrophage cell lines tested. Beryllium-stimulated apoptosis was dose-responsive and maximal after 24 h of exposure to 100 microM BeSO(4). In contrast, unstimulated and Al(2)(SO(4))(3)-stimulated macrophage cell lines did not undergo apoptosis. The general caspase inhibitor BD-fmk inhibited Be-stimulated macrophage cell line apoptosis at concentrations above 50 microM. Our data show that Be-stimulated macrophage cell line apoptosis was caspase-dependent and not solely dependent on Be-stimulated TNF-alpha levels. We speculate that the release of Be-antigen from apoptotic macrophages may serve to re-introduce Be material back into the lung microenvironment, make it available for uptake by new macrophages, and thereby amplify Be-stimulated cytokine production, promoting ongoing inflammation and granuloma maintenance in CBD.

  18. Mycophenolate mofetil modulates adhesion receptors of the beta1 integrin family on tumor cells: impact on tumor recurrence and malignancy

    International Nuclear Information System (INIS)

    Engl, Tobias; Makarević, Jasmina; Relja, Borna; Natsheh, Iyad; Müller, Iris; Beecken, Wolf-Dietrich; Jonas, Dietger; Blaheta, Roman A

    2005-01-01

    Tumor development remains one of the major obstacles following organ transplantation. Immunosuppressive drugs such as cyclosporine and tacrolimus directly contribute to enhanced malignancy, whereas the influence of the novel compound mycophenolate mofetil (MMF) on tumor cell dissemination has not been explored. We therefore investigated the adhesion capacity of colon, pancreas, prostate and kidney carcinoma cell lines to endothelium, as well as their beta1 integrin expression profile before and after MMF treatment. Tumor cell adhesion to endothelial cell monolayers was evaluated in the presence of 0.1 and 1 μM MMF and compared to unstimulated controls. beta1 integrin analysis included alpha1beta1 (CD49a), alpha2beta1 (CD49b), alpha3beta1 (CD49c), alpha4beta1 (CD49d), alpha5beta1 (CD49e), and alpha6beta1 (CD49f) receptors, and was carried out by reverse transcriptase-polymerase chain reaction, confocal microscopy and flow cytometry. Adhesion of the colon carcinoma cell line HT-29 was strongly reduced in the presence of 0.1 μM MMF. This effect was accompanied by down-regulation of alpha3beta1 and alpha6beta1 surface expression and of alpha3beta1 and alpha6beta1 coding mRNA. Adhesion of the prostate tumor cell line DU-145 was blocked dose-dependently by MMF. In contrast to MMF's effects on HT-29 cells, MMF dose-dependently up-regulated alpha1beta1, alpha2beta1, alpha3beta1, and alpha5beta1 on DU-145 tumor cell membranes. We conclude that MMF possesses distinct anti-tumoral properties, particularly in colon and prostate carcinoma cells. Adhesion blockage of HT-29 cells was due to the loss of alpha3beta1 and alpha6beta1 surface expression, which might contribute to a reduced invasive behaviour of this tumor entity. The enhancement of integrin beta1 subtypes observed in DU-145 cells possibly causes re-differentiation towards a low-invasive phenotype

  19. Mycophenolate mofetil modulates adhesion receptors of the beta1 integrin family on tumor cells: impact on tumor recurrence and malignancy

    Directory of Open Access Journals (Sweden)

    Beecken Wolf-Dietrich

    2005-01-01

    Full Text Available Abstract Background Tumor development remains one of the major obstacles following organ transplantation. Immunosuppressive drugs such as cyclosporine and tacrolimus directly contribute to enhanced malignancy, whereas the influence of the novel compound mycophenolate mofetil (MMF on tumor cell dissemination has not been explored. We therefore investigated the adhesion capacity of colon, pancreas, prostate and kidney carcinoma cell lines to endothelium, as well as their beta1 integrin expression profile before and after MMF treatment. Methods Tumor cell adhesion to endothelial cell monolayers was evaluated in the presence of 0.1 and 1 μM MMF and compared to unstimulated controls. beta1 integrin analysis included alpha1beta1 (CD49a, alpha2beta1 (CD49b, alpha3beta1 (CD49c, alpha4beta1 (CD49d, alpha5beta1 (CD49e, and alpha6beta1 (CD49f receptors, and was carried out by reverse transcriptase-polymerase chain reaction, confocal microscopy and flow cytometry. Results Adhesion of the colon carcinoma cell line HT-29 was strongly reduced in the presence of 0.1 μM MMF. This effect was accompanied by down-regulation of alpha3beta1 and alpha6beta1 surface expression and of alpha3beta1 and alpha6beta1 coding mRNA. Adhesion of the prostate tumor cell line DU-145 was blocked dose-dependently by MMF. In contrast to MMF's effects on HT-29 cells, MMF dose-dependently up-regulated alpha1beta1, alpha2beta1, alpha3beta1, and alpha5beta1 on DU-145 tumor cell membranes. Conclusion We conclude that MMF possesses distinct anti-tumoral properties, particularly in colon and prostate carcinoma cells. Adhesion blockage of HT-29 cells was due to the loss of alpha3beta1 and alpha6beta1 surface expression, which might contribute to a reduced invasive behaviour of this tumor entity. The enhancement of integrin beta1 subtypes observed in DU-145 cells possibly causes re-differentiation towards a low-invasive phenotype.

  20. Sodium Butyrate Upregulates miR-203 Expression to Exert Anti-Proliferation Effect on Colorectal Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ruirui Han

    2016-10-01

    Full Text Available Background: As the end product of the bacterial fermentation of dietary fiber in the colonic lumen, sodium butyrate (NaBt has been reported to exert antitumor effects on colorectal cancer (CRC. In addition to functioning as a histone deacetylase (HDAC inhibitor, NaBt also regulates the expression of microRNAs (miRNAs to inhibit CRC cell proliferation. Yet, the mechanisms involved are not completely understood. Here we investigate whether NaBt regulates miR-203 to inhibit CRC growth and explore the promising target gene of miR-203 in CRC cells. Methods: We conducted qRT-PCR and Western blotting assays to evaluate the effects of NaBt on the expression of miR-203 and NEDD9 in HT-29 and Caco-2 cell lines. The promising target gene of miR-203 was predicted by miRNA target prediction and dual luciferase reporter assay. CRC Cell proliferation, colony formation, cell apoptosis and cell invasion assays were performed to explore the effect of NaBt, miR-203 and NEDD9 on HT-29 and Caco-2 cell lines. Results: The results showed that NaBt increased the expression of miR-203 to induce CRC cell apoptosis as well as inhibit cell proliferation, colony formation and cell invasion. Moreover, we determined that the NEDD9 was a target gene of miR-203. NEDD9 partially overcame the inhibitory effects of miR-203 on CRC cell colony formation and invasion. Conclusions: NaBt could induce CRC cell apoptosis, inhibit CRC cell proliferation, colony formation and invasion through miR-203/NEDD9 cascade. The present study may enrich the mechanisms underlying the process that NaBt exerts anti-tumor effects on CRC cells.

  1. Susceptibility of various cell lines to Neospora caninum tachyzoites cultivation

    Directory of Open Access Journals (Sweden)

    Khordadmehr, M.,

    2014-05-01

    Full Text Available Neospora caninum is a coccidian protozoan parasite which is a major cause of bovine abortions and neonatal mortality in cattle, sheep, goat and horse. Occasionally, cultured cells are used for isolation and multiplication of the agent in vitro with several purposes. In this study the tachyzoite yields of N. caninum were compared in various cell cultures as the host cell lines. Among the cell cultures tested, two presented good susceptibility to the agent: cell lines Vero and MA-104. SW742 and TLI (in vitro suspension culture of lymphoid cells infected with Theileria lestoquardi showed moderate sensitivity. No viable tachyzoite were detected in the culture of MDCK and McCoy cell lines. These results demonstrate that MA-104 and SW742 cells present adequate susceptibility to N. caninum compared to Vero cells, which have been largely used to multiply the parasite in vitro. Moreover, these have easy manipulation, fast multiplication and relatively low nutritional requirements. In addition, the result of this study showed that TLI cell line as a suspension cell culture is susceptible to Nc-1 tachyzoites infection and could be used as an alternative host cell line for tachyzoites culture in vitro studies.

  2. Comparative Study of Green Sub- and Supercritical Processes to Obtain Carnosic Acid and Carnosol-Enriched Rosemary Extracts with in Vitro Anti-Proliferative Activity on Colon Cancer Cells

    Directory of Open Access Journals (Sweden)

    Andrea del Pilar Sánchez-Camargo

    2016-12-01

    Full Text Available In the present work, four green processes have been compared to evaluate their potential to obtain rosemary extracts with in vitro anti-proliferative activity against two colon cancer cell lines (HT-29 and HCT116. The processes, carried out under optimal conditions, were: (1 pressurized liquid extraction (PLE, using an hydroalcoholic mixture as solvent at lab-scale; (2 Single-step supercritical fluid extraction (SFE at pilot scale; (3 Intensified two-step sequential SFE at pilot scale; (4 Integrated PLE plus supercritical antisolvent fractionation (SAF at pilot scale. Although higher extraction yields were achieved by using PLE (38.46% dry weight, this extract provided the lowest anti-proliferative activity with no observed cytotoxic effects at the assayed concentrations. On the other hand, extracts obtained using the PLE + SAF process provided the most active rosemary extracts against both colon cancer cell lines, with LC50 ranging from 11.2 to 12.4 µg/mL and from 21.8 to 31.9 µg/mL for HCT116 and HT-29, respectively. In general, active rosemary extracts were characterized by containing carnosic acid (CA and carnosol (CS at concentrations above 263.7 and 33.9 mg/g extract, respectively. Some distinct compounds have been identified in the SAF extracts (rosmaridiphenol and safficinolide, suggesting their possible role as additional contributors to the observed strong anti-proliferative activity of CA and CS in SAF extracts.

  3. Natural killer cells for immunotherapy – Advantages of cell lines over blood NK cells

    Directory of Open Access Journals (Sweden)

    Hans eKlingemann

    2016-03-01

    Full Text Available Natural killer cells are potent cytotoxic effector cells for cancer therapy and potentially for severe viral infections. However, there are technical challenges to obtain sufficient numbers of functionally active NK cells form a patient’s blood since they represent only 10% of the lymphocytes. Especially, cancer patients are known to have dysfunctional NK cells. The alternative is to obtain cells from a healthy donor, which requires depletion of the allogeneic T-cells. Establishing cell lines from donor blood NK cells have not been successful, in contrast to blood NK cells obtained from patients with a clonal NK cell lymphoma. Those cells can be expanded in culture in the presence of IL-2. However, except for the NK-92 cell line none of the other six known cell lines has consistent and reproducibly high anti-tumor cytotoxicity, nor can they be easily genetically manipulated to recognize specific tumor antigens or to augment monoclonal antibody activity through ADCC. NK-92 is also the only cell line product that has been widely given to patients with advanced cancer with demonstrated efficiency and minimal side effects.

  4. Radiation sensitivity of human lung cancer cell lines

    International Nuclear Information System (INIS)

    Carmichael, J.; Degraff, W.G.; Gamson, J.; Russo, G.; Mitchell, J.B.; Gazdar, A.F.; Minna, J.D.; Levitt, M.L.

    1989-01-01

    X-Ray survival curves were determined using a panel of 17 human lung cancer cell lines, with emphasis on non-small cell lung cancer (NSCLC). In contrast to classic small cell lung cancer (SCLC) cell lines, NSCLC cell lines were generally less sensitive to radiation as evidenced by higher radiation survival curve extrapolation numbers, surviving fraction values following a 2Gy dose (SF2) and the mean inactivation dose values (D) values. The spectrum of in vitro radiation responses observed was similar to that expected in clinical practice, although mesothelioma was unexpectedly sensitive in vitro. Differences in radiosensitivity were best distinguished by comparison of SF2 values. Some NSCLC lines were relatively sensitive, and in view of this demonstrable variability in radiation sensitivity, the SF2 value may be useful for in vitro predictive assay testing of clinical specimens. (author)

  5. Automated counting of mammalian cell colonies.

    Science.gov (United States)

    Barber, P R; Vojnovic, B; Kelly, J; Mayes, C R; Boulton, P; Woodcock, M; Joiner, M C

    2001-01-01

    Investigating the effect of low-dose radiation exposure on cells using assays of colony-forming ability requires large cell samples to maintain statistical accuracy. Manually counting the resulting colonies is a laborious task in which consistent objectivity is hard to achieve. This is true especially with some mammalian cell lines which form poorly defined or 'fuzzy' colonies, typified by glioma or fibroblast cell lines. A computer-vision-based automated colony counter is presented in this paper. It utilizes novel imaging and image-processing methods involving a modified form of the Hough transform. The automated counter is able to identify less-discrete cell colonies typical of these cell lines. The results of automated colony counting are compared with those from four manual (human) colony counts for the cell lines HT29, A172, U118 and IN1265. The results from the automated counts fall well within the distribution of the manual counts for all four cell lines with respect to surviving fraction (SF) versus dose curves, SF values at 2 Gy (SF2) and total area under the SF curve (Dbar). From the variation in the counts, it is shown that the automated counts are generally more consistent than the manual counts.

  6. Establishment and characterization of rat portal myofibroblast cell lines.

    Directory of Open Access Journals (Sweden)

    Michel Fausther

    Full Text Available The major sources of scar-forming myofibroblasts during liver fibrosis are activated hepatic stellate cells (HSC and portal fibroblasts (PF. In contrast to well-characterized HSC, PF remain understudied and poorly defined. This is largely due to the facts that isolation of rodent PF for functional studies is technically challenging and that PF cell lines had not been established. To address this, we have generated two polyclonal portal myofibroblast cell lines, RGF and RGF-N2. RGF and RGF-N2 were established from primary PF isolated from adult rat livers that underwent culture activation and subsequent SV40-mediated immortalization. Specifically, Ntpdase2/Cd39l1-sorted primary PF were used to generate the RGF-N2 cell line. Both cell lines were functionally characterized by RT-PCR, immunofluorescence, immunoblot and bromodeoxyuridine-based proliferation assay. First, immortalized RGF and RGF-N2 cells are positive for phenotypic myofibroblast markers alpha smooth muscle actin, type I collagen alpha-1, tissue inhibitor of metalloproteinases-1, PF-specific markers elastin, type XV collagen alpha-1 and Ntpdase2/Cd39l1, and mesenchymal cell marker ecto-5'-nucleotidase/Cd73, while negative for HSC-specific markers desmin and lecithin retinol acyltransferase. Second, both RGF and RGF-N2 cell lines are readily transfectable using standard methods. Finally, RGF and RGF-N2 cells attenuate the growth of Mz-ChA-1 cholangiocarcinoma cells in co-culture, as previously demonstrated for primary PF. Immortalized rat portal myofibroblast RGF and RGF-N2 cell lines express typical markers of activated PF-derived myofibroblasts, are suitable for DNA transfection, and can effectively inhibit cholangiocyte proliferation. Both RGF and RGF-N2 cell lines represent novel in vitro cellular models for the functional studies of portal (myofibroblasts and their contribution to the progression of liver fibrosis.

  7. UCI-VULV-1, a vulvar squamous carcinoma cell line.

    Science.gov (United States)

    Carpenter, P M; Gamboa-Vujicic, G; Mascarello, J T; Wilczynski, S; Bhaumik, M; Dorion, G; Manetta, A

    1995-05-01

    Squamous carcinoma of the vulva (SCV) is an uncommon neoplasm of uncertain etiology. There is evidence that there are two subgroups of SCV, one associated with human papilloma virus (HPV) and a second HPV-negative group. The UCI-VULV-1 cell line, obtained from a lymph node metastasis of an SCV, grows with a population doubling time of approximately 60 hr. The saturation density is 10(5) cells/cm2. The cell line does not exhibit anchorage independence and is weakly tumorigenic. The cells range in appearance from an abundant spindle cell to a less common larger, flat cell. All of the cells are immunoreactive for high-molecular-weight keratin, but only the flat cells, which form squamous pearls in vivo, are immunoreactive for low-molecular-weight keratin. The cell line expresses epidermal growth factor (EGF), transforming growth factor-alpha, the EGF receptor, and p53 protein. Polymerase chain reaction revealed no HPV DNA within the cells. Early passage cells exhibited karyotypic heterogeneity with few similarities to previous described SCV karyotypes. The cells display sensitivity to cis-platinum in concentrations toxic to many ovarian and cervical carcinoma lines. UCI-VULV-1 may be helpful for studying the properties of the HPV-negative form of SCV.

  8. Differential effects of bisphosphonates on breast cancer cell lines

    NARCIS (Netherlands)

    Verdijk, R.; Franke, H.R.; Wolbers, F.; Vermes, I.

    2007-01-01

    Bisphosphonates may induce direct anti-tumor effects in breast cancers cells in virtro. In this study, six bisphosphonates were administered to three breast caner cell lines. Cell proliferation was measured by quantification of th expressio of Cyclin D1 mRNA. Apoptosis was determined by flow

  9. A stromal myoid cell line provokes thymic erythropoiesis between ...

    African Journals Online (AJOL)

    Background: The thymus provides an optimal cellular and humoral microenvironment for cell line committed differentiation of haematopoietic stem cells. The immigration process requires the secretion of at least one peptide called thymotaxine by cells of the reticulo-epithelial (RE) network of the thymic stromal cellular ...

  10. Cytotoxicity against MCF-7 breast cancer cell line and interaction ...

    African Journals Online (AJOL)

    N6-furfuryladenine (kinetin) is a cytokinin growth factor with several biological effects observed in human cells and fruit flies. Kinetin exists naturally in the DNA of almost all organisms tested so far, including human cells and various plants. The cytotoxicity effect of kinetin on MCF-7 breast cancer cell lines was measured by ...

  11. Susceptibilities of medaka (Oryzias latipes cell lines to a betanodavirus

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    Adachi Kei

    2010-07-01

    Full Text Available Abstract Background Betanodaviruses, members of the family Nodaviridae, have bipartite, positive-sense RNA genomes and are the causal agents of viral nervous necrosis in many marine fish species. Recently, the viruses were shown to infect a few freshwater fish species including a model fish medaka (Oryzias latipes. Although virological study using cultured medaka cells would provide a lot of insight into virus-fish interactions in molecular aspects, no such cells have yet been tested for virus susceptibility. Results We tested ten medaka cell lines for susceptibilities to redspotted grouper nervous necrosis virus (RGNNV. Although the viral coat protein was detected in all the cell lines inoculated, the levels of cytopathic effect development and viral propagation were quite different among the cell lines. Those levels were especially high in OLHNI-1 and OLHNI-2 cells, but were extremely low in OLME-104 cells. Some cell lines entered into antiviral state after RGNNV infections probably because of inducing an antiviral system. This is the first report to examine the susceptibilities of cultured medaka cells against a virus. Conclusion OLHNI-1 and OLHNI-2 cells are candidates of new standard cells for betanodavirus study because of their high susceptibilities to the virus and their several advantages as model fish cells.

  12. Establishment and characterization of a chicken mononuclear cell line.

    Science.gov (United States)

    Qureshi, M A; Miller, L; Lillehoj, H S; Ficken, M D

    1990-11-01

    A new chicken mononuclear cell line (MQ-NCSU) has been established. The starting material used to initiate this cell line was a transformed spleen from a female Dekalb XL chicken which had been experimentally challenged with the JM/102W strain of the Marek's disease virus. After homogenization, a single cell suspension of splenic cells was cultured using L.M. Hahn medium supplemented with 10 microM 2-mercaptoethanol. Under these culture conditions, a rapidly proliferating cell was observed and then expanded after performing limiting dilution cultures. These cells were moderately adherent and phagocytic for sheep red blood cells and Salmonella typhimurium. When tested against a panel of monoclonal antibodies (mAb) using the flow cytometry, MQ-NCSU cells stained readily with anti-chicken monocyte specific (K-1) mAb but did not stain with mAb detecting T-helper, T-cytotoxic/suppressor, and NK cells. MQ-NCSU cells expressed very high levels of Ia antigens and transferrin receptors. In addition, cell-free supernatant obtained from MQ-NCSU culture contained a factor which exhibited cytolytic activity against tumor cell targets. Based on their cultural, morphological, and functional characteristics and mAb reactivity profile, we conclude that MQ-NCSU cell line represents a malignantly-transformed cell which shares features characteristic of cells of the mononuclear phagocyte lineage.

  13. Global Conservation of Protein Status between Cell Lines and Xenografts

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    Julian Biau

    2016-08-01

    Full Text Available Common preclinical models for testing anticancer treatment include cultured human tumor cell lines in monolayer, and xenografts derived from these cell lines in immunodeficient mice. Our goal was to determine how similar the xenografts are compared with their original cell line and to determine whether it is possible to predict the stability of a xenograft model beforehand. We studied a selection of 89 protein markers of interest in 14 human cell cultures and respective subcutaneous xenografts using the reverse-phase protein array technology. We specifically focused on proteins and posttranslational modifications involved in DNA repair, PI3K pathway, apoptosis, tyrosine kinase signaling, stress, cell cycle, MAPK/ERK signaling, SAPK/JNK signaling, NFκB signaling, and adhesion/cytoskeleton. Using hierarchical clustering, most cell culture-xenograft pairs cluster together, suggesting a global conservation of protein signature. Particularly, Akt, NFkB, EGFR, and Vimentin showed very stable protein expression and phosphorylation levels highlighting that 4 of 10 pathways were highly correlated whatever the model. Other proteins were heterogeneously conserved depending on the cell line. Finally, cell line models with low Akt pathway activation and low levels of Vimentin gave rise to more reliable xenograft models. These results may be useful for the extrapolation of cell culture experiments to in vivo models in novel targeted drug discovery.

  14. Pathway-specific differences between tumor cell lines and normal and tumor tissue cells

    Directory of Open Access Journals (Sweden)

    Tozeren Aydin

    2006-11-01

    Full Text Available Abstract Background Cell lines are used in experimental investigation of cancer but their capacity to represent tumor cells has yet to be quantified. The aim of the study was to identify significant alterations in pathway usage in cell lines in comparison with normal and tumor tissue. Methods This study utilized a pathway-specific enrichment analysis of publicly accessible microarray data and quantified the gene expression differences between cell lines, tumor, and normal tissue cells for six different tissue types. KEGG pathways that are significantly different between cell lines and tumors, cell lines and normal tissues and tumor and normal tissue were identified through enrichment tests on gene lists obtained using Significance Analysis of Microarrays (SAM. Results Cellular pathways that were significantly upregulated in cell lines compared to tumor cells and normal cells of the same tissue type included ATP synthesis, cell communication, cell cycle, oxidative phosphorylation, purine, pyrimidine and pyruvate metabolism, and proteasome. Results on metabolic pathways suggested an increase in the velocity nucleotide metabolism and RNA production. Pathways that were downregulated in cell lines compared to tumor and normal tissue included cell communication, cell adhesion molecules (CAMs, and ECM-receptor interaction. Only a fraction of the significantly altered genes in tumor-to-normal comparison had similar expressions in cancer cell lines and tumor cells. These genes were tissue-specific and were distributed sparsely among multiple pathways. Conclusion Significantly altered genes in tumors compared to normal tissue were largely tissue specific. Among these genes downregulation was a major trend. In contrast, cell lines contained large sets of significantly upregulated genes that were common to multiple tissue types. Pathway upregulation in cell lines was most pronounced over metabolic pathways including cell nucleotide metabolism and oxidative

  15. Induction of apoptosis by opium in some tumor cell lines.

    Science.gov (United States)

    Khaleghi, M; Farsinejad, A; Dabiri, S; Asadikaram, G

    2016-09-30

    The current study is aimed at investigation of the opium effects on the apoptosis of different cell lines in culture medium and compares such effects with one another. The study is carried out on over 8 cell lines (AA8, AGS, Hela, HepG2, MCF7, N2a, PC12, WEHI). A 2.86 x 10-4 g/ml opium concentration was prepared and added to the culture medium of the cell lines for 48 hours. Cytotoxicity was tested by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. The apoptotic effect of opium on the cell lines was analyzed by Annexin-PI test. Opium with concentration of 2.86 x 10-4 g/ml in 48 hours significantly induces apoptosis in certain cell lines (i.e. AA8, N2a, WEHI), apoptosis and necrosis in some others (i.e. Hela, HepG2, MCF7, and PC12), and also solely necrosis in the AGS cell line. One could infer that the usage of opium with different levels in different tissues leads to certain disorders in some tissues and may have therapeutic effects under distinctive conditions (i.e. unchecked growth of cells) as confirmed by the results.

  16. Fish cell lines as a tool in aquatic toxicology.

    Science.gov (United States)

    Segner, H

    1998-01-01

    In aquatic toxicology, cytotoxicity tests using continuous fish cell lines have been suggested as a tool for (1) screening or toxicity ranking of anthropogenic chemicals, compound mixtures and environmental samples, (2) establishment of structure-activity relationships, and (3) replacement or supplementation of in vivo animal tests. Due to the small sample volumes necessary for cytotoxicity tests, they appear to be particularly suited for use in chemical fractionation studies. The present contribution reviews the existing literature on cytotoxicity studies with fish cells and considers the influence of cell line and cytotoxicity endpoint selection on the test results. Furthermore, in vitro/in vivo correlations between fish cell lines and intact fish are discussed. During recent years, fish cell lines have been increasingly used for purposes beyond their meanwhile established role for cytotoxicity measurements. They have been successfully introduced for detection of genotoxic effects, and cell lines are now applied for investigations on toxic mechanisms and on biomarkers such as cytochrome P4501A. The development of recombinant fish cell lines may further support their role as a bioanalytical tool in environmental diagnostics.

  17. Curcumin analog EF24 induces apoptosis via ROS-dependent mitochondrial dysfunction in human colorectal cancer cells.

    Science.gov (United States)

    He, Guodong; Feng, Chen; Vinothkumar, Rajamanickam; Chen, Weiqian; Dai, Xuanxuan; Chen, Xi; Ye, Qingqing; Qiu, Chenyu; Zhou, Huiping; Wang, Yi; Liang, Guang; Xie, Yubo; Wu, Wei

    2016-12-01

    Colorectal cancer is the most commonly diagnosed malignancy with high mortality rates worldwide. Improved therapeutic strategies with minimal adverse side effects are urgently needed. In this study, the anti-tumor effects of EF24, a novel analog of the natural compound curcumin, were evaluated in colorectal cancer cells. The anti-tumor activity of EF24 on human colon cancer lines (HCT-116, SW-620, and HT-29) was determined by measures of cell cycle arrest, apoptosis, and mitochondrial function. The contribution of ROS in the EF24-induced anti-tumor activity was evaluated by measures of H 2 O 2 and pretreatment with an ROS scavenger, NAC. The findings indicated that EF24 treatment dose-dependently inhibited cell viability and caused cell cycle arrest at G2/M phase in all the tested colon cancer cell lines. Furthermore, we demonstrated that EF24 treatment induced apoptosis effectively via enhancing intracellular accumulation of ROS in both HCT-116 and SW-620 cells, but with moderate effects in HT-29 cells. We found that EF24 treatment decreased the mitochondrial membrane potential in the colon cancer cells, leading to the release of mitochondrial cytochrome c. Also, EF24 induced activation of caspases 9 and 3, causing decreased Bcl-2 protein expression and Bcl-2/Bax ratio. Pretreatment with NAC, a ROS scavenger, abrogated the EF24-induced cell death, apoptosis, cell cycle arrest, and mitochondrial dysfunction, suggesting an upstream ROS generation which was responsible for the anticancer effects of EF24. Our findings support an anticancer mechanism by which EF24 enhanced ROS accumulation in colon cancer cells, thereby resulting in mitochondrial membrane collapse and activated intrinsic apoptotic signaling. Thus, EF24 could be a potential candidate for therapeutic application of colon cancer.

  18. Lining cells on normal human vertebral bone surfaces

    International Nuclear Information System (INIS)

    Henning, C.B.; Lloyd, E.L.

    1982-01-01

    Thoracic vertebrae from two individuals with no bone disease were studied with the electron microscope to determine cell morphology in relation to bone mineral. The work was undertaken to determine if cell morphology or spatial relationships between the bone lining cells and bone mineral could account for the relative infrequency of bone tumors which arise at this site following radium intake, when compared with other sites, such as the head of the femur. Cells lining the vertebral mineral were found to be generally rounded in appearance with varied numbers of cytoplasmic granules, and they appeared to have a high density per unit of surface area. These features contrasted with the single layer of flattened cells characteristic of the bone lining cells of the femur. A tentative discussion of the reasons for the relative infrequency of tumors in the vertebrae following radium acquisition is presented

  19. MORPHOMETRIC SUBTYPING FOR A PANEL OF BREAST CANCER CELL LINES

    Energy Technology Data Exchange (ETDEWEB)

    Han, Ju; Chang, Hang; Fontenay, Gerald; Wang, Nicholas J.; Gray, Joe W.; Parvin, Bahram

    2009-05-08

    A panel of cell lines of diverse molecular background offers an improved model system for high-content screening, comparative analysis, and cell systems biology. A computational pipeline has been developed to collect images from cell-based assays, segment individual cells and colonies, represent segmented objects in a multidimensional space, and cluster them for identifying distinct subpopulations. While each segmentation strategy can vary for different imaging assays, representation and subpopulation analysis share a common thread. Application of this pipeline to a library of 41 breast cancer cell lines is demonstrated. These cell lines are grown in 2D and imaged through immunofluorescence microscopy. Subpopulations in this panel are identified and shown to correlate with previous subtyping literature that was derived from transcript data.

  20. DNA fingerprinting of the NCI-60 cell line panel.

    Science.gov (United States)

    Lorenzi, Philip L; Reinhold, William C; Varma, Sudhir; Hutchinson, Amy A; Pommier, Yves; Chanock, Stephen J; Weinstein, John N

    2009-04-01

    The National Cancer Institute's NCI-60 cell line panel, the most extensively characterized set of cells in existence and a public resource, is frequently used as a screening tool for drug discovery. Because many laboratories around the world rely on data from the NCI-60 cells, confirmation of their genetic identities represents an essential step in validating results from them. Given the consequences of cell line contamination or misidentification, quality control measures should routinely include DNA fingerprinting. We have, therefore, used standard DNA microsatellite short tandem repeats to profile the NCI-60, and the resulting DNA fingerprints are provided here as a reference. Consistent with previous reports, the fingerprints suggest that several NCI-60 lines have common origins: the melanoma lines MDA-MB-435, MDA-N, and M14; the central nervous system lines U251 and SNB-19; the ovarian lines OVCAR-8 and OVCAR-8/ADR (also called NCI/ADR); and the prostate lines DU-145, DU-145 (ATCC), and RC0.1. Those lines also show that the ability to connect two fingerprints to the same origin is not affected by stable transfection or by the development of multidrug resistance. As expected, DNA fingerprints were not able to distinguish different tissues-of-origin. The fingerprints serve principally as a barcodes.

  1. Characterization of newly established colorectal cancer cell lines ...

    Indian Academy of Sciences (India)

    Unknown

    2000-12-19

    Gastroenterology Service,. Department of Medicine, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021, USA. Abstract. We have established a series of 20 colorectal cancer cell lines and performed ...

  2. Gamma-Mangostin, a Micronutrient of Mangosteen Fruit, Induces Apoptosis in Human Colon Cancer Cells

    Directory of Open Access Journals (Sweden)

    Hui-Fang Chang

    2012-07-01

    Full Text Available Recently colorectal cancer rates have increased rapidly in Taiwan. The treatment of colorectal cancer includes surgery, radiation therapy and chemotherapy. Mangosteen (Garcinia mangostana is a famous Asian tropical fruit. γ-Mangostin is a xanthone derivative isolated from the fruit hull. In previous studies, we found evidence of anti-inflammatory and anti-brain tumor activities in γ-mangostin. In this study, we performed further studies to assess the apoptotic effects of γ-mangostin on colorectal adenocarcinoma cells HT29. γ-Mangostin showed concentration and time-dependent cytotoxic effects on HT29 cells. Microscopic observation under Giemsa staining showed that γ-mangostin induced cellular swelling and the appearance of apoptotic bodies, characteristic of apoptosis in HT29 cells. In addition, flow cytometry analysis showed an increase of hypodiploid cells in γ-mangostin-treated HT29 cells, while enhancement of intracellular peroxide production was detected in the same γ-mangostin-treated cells by DCHDA assay and DiOC6(3 staining. In view of the above results, γ-mangostin has demonstrated anticancer activity and induces apoptosis in HT29 colorectal adenocarcinoma cells. The evidence suggests that γ-mangostin could serve as a micronutrient for colon cancer prevention and is a potential lead compound for the development of anti-colon cancer agents.

  3. Isolation of two chloroethylnitrosourea-sensitive Chinese hamster cell lines

    International Nuclear Information System (INIS)

    Hata, H.; Numata, M.; Tohda, H.; Yasui, A.; Oikawa, A.

    1991-01-01

    1-[(4-Amino-2-methylpyrimidin-5-yl)methyl]-3-(2-chloroethyl)-3- nitrosourea hydrochloride (ACNU), a cancer chemotherapeutic bifunctional alkylating agent, causes chloroethylation of DNA and subsequent DNA strand cross-linking through an ethylene bridge. We isolated and characterized two ACNU-sensitive mutants from mutagenized Chinese hamster ovary cells and found them to be new drug-sensitive recessive Chinese hamster mutants. Both mutants were sensitive to various monofunctional alkylating agents in a way similar to that of the parental cell lines CHO9. One mutant (UVS1) was cross-sensitive to UV and complemented the UV sensitivity of all Chinese hamster cell lines of 7 established complementation groups. Since UV-induced unscheduled DNA synthesis was very low, a new locus related to excision repair is thought to be defective in this cell line. Another ACNU-sensitive mutant, CNU1, was slightly more sensitive to UV than the parent cell line. CNU1 was cross-sensitive to 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea and slightly more sensitive to mitomycin C. No increased accumulation of ACNU and a low level of UV-induced unscheduled DNA synthesis in this cell as compared with the parental cell line suggest that there is abnormality in a repair response of this mutant cell to some types of DNA cross-links

  4. In vitro Rb-1 gene transfer to retinoblastoma cell lines

    International Nuclear Information System (INIS)

    Choi, Sang Wook; Ham, Yong Hoh; Kim, Mee Heui

    1994-04-01

    After transfection of Rb-vector to packaging cell line (CRIP) by Ca-P precipitation method, we could select nineteen colonies of G-418 resistant clone by ring cloning. Each colony was transduced to NIH3T3 cells to select the one which produces high titer virus. After NIH3T3 cells transduction, we could get 28 colony counts for the high, 127 for the middle, and 6 for the low viral titer. With the supernatant of the high viral titer colony (CRIPRb 2-5). We transduct retinoblastoma cell lines. 5 figs, 11 refs. (Author)

  5. Assessment of cancer cell line representativeness using microarrays for Merkel cell carcinoma.

    Science.gov (United States)

    Daily, Kenneth; Coxon, Amy; Williams, Jonathan S; Lee, Chyi-Chia R; Coit, Daniel G; Busam, Klaus J; Brownell, Isaac

    2015-04-01

    When using cell lines to study cancer, phenotypic similarity to the original tumor is paramount. Yet, little has been done to characterize how closely Merkel cell carcinoma (MCC) cell lines model native tumors. To determine their similarity to MCC tumor samples, we characterized MCC cell lines via gene expression microarrays. Using whole transcriptome gene expression signatures and a computational bioinformatic approach, we identified significant differences between variant cell lines (UISO, MCC13, and MCC26) and fresh frozen MCC tumors. Conversely, the classic WaGa and Mkl-1 cell lines more closely represented the global transcriptome of MCC tumors. When compared with publicly available cancer lines, WaGa and Mkl-1 cells were similar to other neuroendocrine tumors, but the variant cell lines were not. WaGa and Mkl-1 cells grown as xenografts in mice had histological and immunophenotypical features consistent with MCC, whereas UISO xenograft tumors were atypical for MCC. Spectral karyotyping and short tandem repeat analysis of the UISO cells matched the original cell line's description, ruling out contamination. Our results validate the use of transcriptome analysis to assess the cancer cell line representativeness and indicate that UISO, MCC13, and MCC26 cell lines are not representative of MCC tumors, whereas WaGa and Mkl-1 more closely model MCC.

  6. Establishment of cell lines from adult T-cell leukemia cells dependent on negatively charged polymers.

    Science.gov (United States)

    Kagami, Yoshitoyo; Uchiyama, Susumu; Kato, Harumi; Okada, Yasutaka; Seto, Masao; Kinoshita, Tomohiro

    2017-07-05

    Growing adult T-cell leukemia/lymphoma (ATLL) cells in vitro is difficult. Here, we examined the effects of static electricity in the culture medium on the proliferation of ATLL cells. Six out of 10 ATLL cells did not proliferate in vitro and thus had to be cultured in a medium containing negatively charged polymers. In the presence of poly-γ-glutamic acid (PGA) or chondroitin sulfate (CDR), cell lines (HKOX3-PGA, HKOX3-CDR) were established from the same single ATLL case using interleukin (IL)-2, IL-4, and feeder cells expressing OX40L (OX40L + HK). Dextran sulfate inhibited growth in both HKOX3 cell lines. Both PGA and OX40L + HK were indispensable for HKOX3-PGA growth, but HKOX3-CDR could proliferate in the presence of CDR or OX40L + HK alone. Thus, the specific action of each negatively charged polymer promoted the growth of specific ATLL cells in vitro.

  7. Rosiglitazone and AS601245 decrease cell adhesion and migration through modulation of specific gene expression in human colon cancer cells.

    Directory of Open Access Journals (Sweden)

    Angelo Cerbone

    Full Text Available PPARs are nuclear receptors activated by ligands. Activation of PPARγ leads to a reduction of adhesion and motility in some cancer models. PPARγ transcriptional activity can be negatively regulated by JNK-mediated phosphorylation. We postulated that the use of agents able to inhibit JNK activity could increase the effectiveness of PPARγ ligands. We analysed the effects of rosiglitazone (PPARγ ligand and AS601245 (a selective JNK inhibitor alone or in association on adhesion and migration of CaCo-2, HT29, and SW480 human colon cancer cells and investigated, through microarray analysis, the genes involved in these processes. Cell adhesion and migration was strongly inhibited by rosiglitazone and AS601245. Combined treatment with the two compounds resulted in a greater reduction of the adhesion and migration capacity. Affymetrix analysis in CaCo-2 cells revealed that some genes which were highly modulated by the combined treatment could be involved in these biological responses. Rosiglitazone, AS601245 and combined treatment down-regulated the expression of fibrinogen chains in all three cell lines. Moreover, rosiglitazone, alone or in association with AS601245, caused a decrease in the fibrinogen release. ARHGEF7/β-PIX gene was highly down-regulated by combined treatment, and western blot analysis revealed that β-PIX protein is down-modulated in CaCo-2, HT29 and SW480 cells, also. Transfection of cells with β-PIX gene completely abrogated the inhibitory effect on cell migration, determined by rosiglitazone, AS601245 and combined treatment. Results demonstrated that β-PIX protein is involved in the inhibition of cell migration and sustaining the positive interaction between PPARγ ligands and anti-inflammatory agents in humans.

  8. Antitumor Activity of Propolis on Differantiated Cancer Cell Lines

    OpenAIRE

    , Neşe Ersöz Gülçelik, Dilara Zeybek, Fige

    2012-01-01

    Propolis is a natural bee product with several pharmacological activities. Nowadays, it is also investigated for its antitumor properties. There are contraversies on the antitumor activity of propolis, not all tumour cells seem to respond to propolis treatment. The aim of our study is to evaluate the activity of propolis on differantiated thyroid cancer cell lines. Tyripan blue test and MTT assay were performed to evaluate the cell viability of B-CPAP cells after propolis treatment and compar...

  9. Lactobacillus casei Exerts Anti-Proliferative Effects Accompanied by Apoptotic Cell Death and Up-Regulation of TRAIL in Colon Carcinoma Cells

    Science.gov (United States)

    Santarmaki, Valentina; Aindelis, Georgios; Tompoulidou, Evgenia; Lamprianidou, Eleftheria E.; Saxami, Georgia; Ypsilantis, Petros; Lampri, Evangeli S.; Simopoulos, Constantinos; Kotsianidis, Ioannis; Galanis, Alex; Kourkoutas, Yiannis; Dimitrellou, Dimitra; Chlichlia, Katerina

    2016-01-01

    Probiotic microorganisms such as lactic acid bacteria (LAB) exert a number of strain-specific health-promoting activities attributed to their immunomodulatory, anti-inflammatory and anti-carcinogenic properties. Despite recent attention, our understanding of the biological processes involved in the beneficial effects of LAB strains is still limited. To this end, the present study investigated the growth-inhibitory effects of Lactobacillus casei ATCC 393 against experimental colon cancer. Administration of live Lactobacillus casei (as well as bacterial components thereof) on murine (CT26) and human (HT29) colon carcinoma cell lines raised a significant concentration- and time-dependent anti-proliferative effect, determined by cell viability assays. Specifically, a dramatic decrease in viability of colon cancer cells co-incubated with 109 CFU/mL L. casei for 24 hours was detected (78% for HT29 and 52% for CT26 cells). In addition, live L. casei induced apoptotic cell death in both cell lines as revealed by annexin V and propidium iodide staining. The significance of the in vitro anti-proliferative effects was further confirmed in an experimental tumor model. Oral daily administration of 109 CFU live L. casei for 13 days significantly inhibited in vivo growth of colon carcinoma cells, resulting in approximately 80% reduction in tumor volume of treated mice. Tumor growth inhibition was accompanied by L. casei-driven up-regulation of the TNF-related apoptosis-inducing ligand TRAIL and down-regulation of Survivin. Taken together, these findings provide evidence for beneficial tumor-inhibitory, anti-proliferative and pro-apoptotic effects driven by this probiotic LAB strain. PMID:26849051

  10. Lactobacillus casei Exerts Anti-Proliferative Effects Accompanied by Apoptotic Cell Death and Up-Regulation of TRAIL in Colon Carcinoma Cells.

    Directory of Open Access Journals (Sweden)

    Angeliki Tiptiri-Kourpeti

    Full Text Available Probiotic microorganisms such as lactic acid bacteria (LAB exert a number of strain-specific health-promoting activities attributed to their immunomodulatory, anti-inflammatory and anti-carcinogenic properties. Despite recent attention, our understanding of the biological processes involved in the beneficial effects of LAB strains is still limited. To this end, the present study investigated the growth-inhibitory effects of Lactobacillus casei ATCC 393 against experimental colon cancer. Administration of live Lactobacillus casei (as well as bacterial components thereof on murine (CT26 and human (HT29 colon carcinoma cell lines raised a significant concentration- and time-dependent anti-proliferative effect, determined by cell viability assays. Specifically, a dramatic decrease in viability of colon cancer cells co-incubated with 10(9 CFU/mL L. casei for 24 hours was detected (78% for HT29 and 52% for CT26 cells. In addition, live L. casei induced apoptotic cell death in both cell lines as revealed by annexin V and propidium iodide staining. The significance of the in vitro anti-proliferative effects was further confirmed in an experimental tumor model. Oral daily administration of 10(9 CFU live L. casei for 13 days significantly inhibited in vivo growth of colon carcinoma cells, resulting in approximately 80% reduction in tumor volume of treated mice. Tumor growth inhibition was accompanied by L. casei-driven up-regulation of the TNF-related apoptosis-inducing ligand TRAIL and down-regulation of Survivin. Taken together, these findings provide evidence for beneficial tumor-inhibitory, anti-proliferative and pro-apoptotic effects driven by this probiotic LAB strain.

  11. Lactobacillus casei Exerts Anti-Proliferative Effects Accompanied by Apoptotic Cell Death and Up-Regulation of TRAIL in Colon Carcinoma Cells.

    Science.gov (United States)

    Tiptiri-Kourpeti, Angeliki; Spyridopoulou, Katerina; Santarmaki, Valentina; Aindelis, Georgios; Tompoulidou, Evgenia; Lamprianidou, Eleftheria E; Saxami, Georgia; Ypsilantis, Petros; Lampri, Evangeli S; Simopoulos, Constantinos; Kotsianidis, Ioannis; Galanis, Alex; Kourkoutas, Yiannis; Dimitrellou, Dimitra; Chlichlia, Katerina

    2016-01-01

    Probiotic microorganisms such as lactic acid bacteria (LAB) exert a number of strain-specific health-promoting activities attributed to their immunomodulatory, anti-inflammatory and anti-carcinogenic properties. Despite recent attention, our understanding of the biological processes involved in the beneficial effects of LAB strains is still limited. To this end, the present study investigated the growth-inhibitory effects of Lactobacillus casei ATCC 393 against experimental colon cancer. Administration of live Lactobacillus casei (as well as bacterial components thereof) on murine (CT26) and human (HT29) colon carcinoma cell lines raised a significant concentration- and time-dependent anti-proliferative effect, determined by cell viability assays. Specifically, a dramatic decrease in viability of colon cancer cells co-incubated with 10(9) CFU/mL L. casei for 24 hours was detected (78% for HT29 and 52% for CT26 cells). In addition, live L. casei induced apoptotic cell death in both cell lines as revealed by annexin V and propidium iodide staining. The significance of the in vitro anti-proliferative effects was further confirmed in an experimental tumor model. Oral daily administration of 10(9) CFU live L. casei for 13 days significantly inhibited in vivo growth of colon carcinoma cells, resulting in approximately 80% reduction in tumor volume of treated mice. Tumor growth inhibition was accompanied by L. casei-driven up-regulation of the TNF-related apoptosis-inducing ligand TRAIL and down-regulation of Survivin. Taken together, these findings provide evidence for beneficial tumor-inhibitory, anti-proliferative and pro-apoptotic effects driven by this probiotic LAB strain.

  12. Drug/Cell-line Browser: interactive canvas visualization of cancer drug/cell-line viability assay datasets.

    Science.gov (United States)

    Duan, Qiaonan; Wang, Zichen; Fernandez, Nicolas F; Rouillard, Andrew D; Tan, Christopher M; Benes, Cyril H; Ma'ayan, Avi

    2014-11-15

    Recently, several high profile studies collected cell viability data from panels of cancer cell lines treated with many drugs applied at different concentrations. Such drug sensitivity data for cancer cell lines provide suggestive treatments for different types and subtypes of cancer. Visualization of these datasets can reveal patterns that may not be obvious by examining the data without such efforts. Here we introduce Drug/Cell-line Browser (DCB), an online interactive HTML5 data visualization tool for interacting with three of the recently published datasets of cancer cell lines/drug-viability studies. DCB uses clustering and canvas visualization of the drugs and the cell lines, as well as a bar graph that summarizes drug effectiveness for the tissue of origin or the cancer subtypes for single or multiple drugs. DCB can help in understanding drug response patterns and prioritizing drug/cancer cell line interactions by tissue of origin or cancer subtype. DCB is an open source Web-based tool that is freely available at: http://www.maayanlab.net/LINCS/DCB CONTACT: avi.maayan@mssm.edu Supplementary data are available at Bioinformatics online. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  13. Effect of failures and repairs on multiple cell production lines

    Energy Technology Data Exchange (ETDEWEB)

    Legato, P.; Bobbio, A.; Roberti, L.

    1989-01-01

    This paper examines a production line composed of multiple stages, or cells, which are passed in sequential order to arrive to the final product. Two possible coordination disciplines are considered, namely: the classical tandem arrangement of sequential working centers with input buffer and the kanban scheme, considered the Japanese shop floor realization of the Just-In-Time (JIT) manifacturing approach. The production line is modelled and analysed by means of Stochastic Petri Nets (SPN). Finally an analysis is made of the possibility that the working cells can incur failure/repair cycles perturbing the production flow of the line and thus reduce performance indices.

  14. Neurohypophysial Receptor Gene Expression by Thymic T Cell Subsets and Thymic T Cell Lymphoma Cell Lines

    Directory of Open Access Journals (Sweden)

    I. Hansenne

    2004-01-01

    transcribed in thymic epithelium, while immature T lymphocytes express functional neurohypophysial receptors. Neurohypophysial receptors belong to the G protein-linked seven-transmembrane receptor superfamily and are encoded by four distinct genes, OTR, V1R, V2R and V3R. The objective of this study was to identify the nature of neurohypophysial receptor in thymic T cell subsets purified by immunomagnetic selection, as well as in murine thymic lymphoma cell lines RL12-NP and BW5147. OTR is transcribed in all thymic T cell subsets and T cell lines, while V3R transcription is restricted to CD4+ CD8+ and CD8+ thymic cells. Neither V1R nor V2R transcripts are detected in any kind of T cells. The OTR protein was identified by immunocytochemistry on thymocytes freshly isolated from C57BL/6 mice. In murine fetal thymic organ cultures, a specific OTR antagonist does not modify the percentage of T cell subsets, but increases late T cell apoptosis further evidencing the involvement of OT/OTR signaling in the control of T cell proliferation and survival. According to these data, OTR and V3R are differentially expressed during T cell ontogeny. Moreover, the restriction of OTR transcription to T cell lines derived from thymic lymphomas may be important in the context of T cell leukemia pathogenesis and treatment.

  15. Reversible effect of dextran sodium sulfate on mucus secreting intestinal epithelial cells

    DEFF Research Database (Denmark)

    Nielsen, Ditte Søvsø Gundelund; Fredborg, Marlene; Andersen, V

    2016-01-01

    investigated effects of increasing doses of DSS on viability and integrity of these intestinal epithelial cells. For cell viability studies, cells were treated with DSS solutions for 24 or 48 h and viability was measured fluorometrically by PicoGreen double-stranded DNA quantitation. HT29-MTX-E12 cells were...... provide valuable insight into a possible mechanism for dextran sodium sulfate (DSS)–induced colitis of importance for the design of subsequent in vivo studies. To develop a new in vitro IBD model with DSS-induced inflammation in human mucus-secreting intestinal epithelial cells (HT29-MTX-E12), we first...... affects the viability and disrupts the intestinal barrier function of HT29-MTX-E12 monolayers, a main feature observed in IBD. Furthermore, the harmful effect of DSS is reversible, suggesting that recovery of intestinal integrity after DSS treatment by potential therapeutic drugs can be studied in vitro....

  16. Membrane lipidome of an epithelial cell line

    DEFF Research Database (Denmark)

    Sampaio, Julio L; Gerl, Mathias J; Klose, Christian

    2011-01-01

    Tissue differentiation is an important process that involves major cellular membrane remodeling. We used Madin-Darby canine kidney cells as a model for epithelium formation and investigated the remodeling of the total cell membrane lipidome during the transition from a nonpolarized morphology...... to an epithelial morphology and vice versa. To achieve this, we developed a shotgun-based lipidomics workflow that enabled the absolute quantification of mammalian membrane lipidomes with minimal sample processing from low sample amounts. Epithelial morphogenesis was accompanied by a major shift from sphingomyelin...... to glycosphingolipid, together with an increase in plasmalogen, phosphatidylethanolamine, and cholesterol content, whereas the opposite changes took place during an epithelial-to-mesenchymal transition. Moreover, during polarization, the sphingolipids became longer, more saturated, and more hydroxylated as required...

  17. Solid Oxide Fuel Cell Systems PVL Line

    International Nuclear Information System (INIS)

    Shearer, Susan; Rush, Gregory

    2012-01-01

    In July 2010, Stark State College (SSC), received Grant DE-EE0003229 from the U.S. Department of Energy (DOE), Golden Field Office, for the development of the electrical and control systems, and mechanical commissioning of a unique 20kW scale high-pressure, high temperature, natural gas fueled Stack Block Test System (SBTS). SSC worked closely with subcontractor, Rolls-Royce Fuel Cell Systems (US) Inc. (RRFCS) over a 13 month period to successfully complete the project activities. This system will be utilized by RRFCS for pre-commercial technology development and training of SSC student interns. In the longer term, when RRFCS is producing commercial products, SSC will utilize the equipment for workforce training. In addition to DOE Hydrogen, Fuel Cells, and Infrastructure Technologies program funding, RRFCS internal funds, funds from the state of Ohio, and funding from the DOE Solid State Energy Conversion Alliance (SECA) program have been utilized to design, develop and commission this equipment. Construction of the SBTS (mechanical components) was performed under a Grant from the State of Ohio through Ohio's Third Frontier program (Grant TECH 08-053). This Ohio program supported development of a system that uses natural gas as a fuel. Funding was provided under the Department of Energy (DOE) Solid-state Energy Conversion Alliance (SECA) program for modifications required to test on coal synthesis gas. The subject DOE program provided funding for the electrical build, control system development and mechanical commissioning. Performance testing, which includes electrical commissioning, was subsequently performed under the DOE SECA program. Rolls-Royce Fuel Cell Systems is developing a megawatt-scale solid oxide fuel cell (SOFC) stationary power generation system. This system, based on RRFCS proprietary technology, is fueled with natural gas, and operates at elevated pressure. A critical success factor for development of the full scale system is the capability to

  18. Antiproliferative effects on colon adenocarcinoma cells induced by co-administration of vitamin K1 and Lactobacillus rhamnosus GG.

    Science.gov (United States)

    Orlando, Antonella; Linsalata, Michele; Russo, Francesco

    2016-06-01

    Vitamin K (VK), an essential nutrient associated with the clotting cascade, has also been demonstrated to have anticancer properties in various cancer cells including colon cancer cells. Also probiotics have gained interest as potential anticancer agents. Among them, Lactobacillus rhamnosus GG (L.GG) has been shown to inhibit cell proliferation and polyamine biosynthesis as well as to induce apoptosis in different human gastrointestinal cancer cells. Nevertheless, the exact mechanisms involved in these actions are not completely elucidated. Therefore, the aims of the present study were to evaluate in three differently graded human colon cancer cells (namely Caco-2, HT-29 and SW480) the effects of increasing VK1 concentrations, administered alone or in combination with viable L.GG, on the cell proliferation evaluated by MTT test, apoptosis investigated by Bax/Bcl-2 ratio and the percentage of the apoptotic cells, and the cell cycle evaluated by MUSE cell analyzer. Both VK1 and L.GG administered alone up to 72 h, caused inhibition of proliferation, induction of apoptosis and the cell cycle arrest in all the tested colon cancer cells. When VK1 and L.GG were co-administered, the addition of increasing VK1 concentrations potentiated the probiotic antiproliferative effect in a dose-dependent manner, being also related to the individual features of each cell line. The effect was more evident in Caco-2 and HT-29 cells compared to the less differentiated SW480. The enhanced antiproliferative efficacy due to co-administration of L.GG and VK1 could represent a suitable option in a functional food strategy for cancer growth inhibition and chemoprevention.

  19. Novel human multiple myeloma cell line UHKT-893

    Czech Academy of Sciences Publication Activity Database

    Uherková, L.; Vančurová, I.; Vyhlídalová, I.; Pleschnerová, M.; Špička, I.; Mihalová, R.; Březinová, J.; Hodný, Zdeněk; Čermáková, K.; Polanská, V.; Marinov, I.; Jedelský, P.L.; Kuželová, K.; Stöckbauer, P.

    2013-01-01

    Roč. 37, č. 3 (2013), s. 320-326 ISSN 0145-2126 Institutional support: RVO:68378050 Keywords : human myeloma cell line * human multiple myeloma * plasma cell * IL-6 dependence * immunoglobulin * free light chain Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.692, year: 2013

  20. a stromal myoid cell line provokes thymic erythropoiesis between

    African Journals Online (AJOL)

    hi-tech

    81 No. 2 February 2004. A STROMAL MYOID CELL LINE PROVOKES THYMIC ERYTHROPOIESIS BETWEEN 16TH TO 20TH WEEKS OF INTRAUTERINE LIFE ... proliferation and differentiation in different stages of development: the stromal myoid cells. Design: ... human myasthenia gravis (MG) has been suggested(3).

  1. Frequency and distribution of Notch mutations in tumor cell lines

    International Nuclear Information System (INIS)

    Mutvei, Anders Peter; Fredlund, Erik; Lendahl, Urban

    2015-01-01

    Deregulated Notch signaling is linked to a variety of tumors and it is therefore important to learn more about the frequency and distribution of Notch mutations in a tumor context. In this report, we use data from the recently developed Cancer Cell Line Encyclopedia to assess the frequency and distribution of Notch mutations in a large panel of cancer cell lines in silico. Our results show that the mutation frequency of Notch receptor and ligand genes is at par with that for established oncogenes and higher than for a set of house-keeping genes. Mutations were found across all four Notch receptor genes, but with notable differences between protein domains, mutations were for example more prevalent in the regions encoding the LNR and PEST domains in the Notch intracellular domain. Furthermore, an in silico estimation of functional impact showed that deleterious mutations cluster to the ligand-binding and the intracellular domains of NOTCH1. For most cell line groups, the mutation frequency of Notch genes is higher than in associated primary tumors. Our results shed new light on the spectrum of Notch mutations after in vitro culturing of tumor cells. The higher mutation frequency in tumor cell lines indicates that Notch mutations are associated with a growth advantage in vitro, and thus may be considered to be driver mutations in a tumor cell line context. The online version of this article (doi:10.1186/s12885-015-1278-x) contains supplementary material, which is available to authorized users

  2. Clonogenic cell line survival of a human liver cancer cell line SMMC-7721 after carbon ion irradiation with different LET

    International Nuclear Information System (INIS)

    Lei Suwen; Su Xu; Wang Jifang; Li Wenjian

    2003-01-01

    Objective: To investigate the survival fraction of a human liver cancer cell line SMMC-7721 following irradiation with carbon ions with different LET. Methods: cells of the human liver cancer cell line SMMC-7721 were irradiated with carbon ions (LET=30 and 70 keV/μm). The survival fraction was determined with clonogenic assay after 9 days incubation in a 5% CO 2 incubator at 37 degree C. Results: When the survival fractions of 70 keV/μm were D s = 0.1 and D s=0.01 absorption dose were 2.94 and 5.88 Gy respectively, and those of 30 keV/μm were 4.00 and 8.00 Gy respectively. Conclusion: For the SMMC-7721 cell line, 70 keV/μm is more effective for cell killing than 30 keV/μm

  3. Guidelines for the use of cell lines in biomedical research.

    Science.gov (United States)

    Geraghty, R J; Capes-Davis, A; Davis, J M; Downward, J; Freshney, R I; Knezevic, I; Lovell-Badge, R; Masters, J R W; Meredith, J; Stacey, G N; Thraves, P; Vias, M

    2014-09-09

    Cell-line misidentification and contamination with microorganisms, such as mycoplasma, together with instability, both genetic and phenotypic, are among the problems that continue to affect cell culture. Many of these problems are avoidable with the necessary foresight, and these Guidelines have been prepared to provide those new to the field and others engaged in teaching and instruction with the information necessary to increase their awareness of the problems and to enable them to deal with them effectively. The Guidelines cover areas such as development, acquisition, authentication, cryopreservation, transfer of cell lines between laboratories, microbial contamination, characterisation, instability and misidentification. Advice is also given on complying with current legal and ethical requirements when deriving cell lines from human and animal tissues, the selection and maintenance of equipment and how to deal with problems that may arise.

  4. Guidelines for the use of cell lines in biomedical research

    Science.gov (United States)

    Geraghty, R J; Capes-Davis, A; Davis, J M; Downward, J; Freshney, R I; Knezevic, I; Lovell-Badge, R; Masters, J R W; Meredith, J; Stacey, G N; Thraves, P; Vias, M

    2014-01-01

    Cell-line misidentification and contamination with microorganisms, such as mycoplasma, together with instability, both genetic and phenotypic, are among the problems that continue to affect cell culture. Many of these problems are avoidable with the necessary foresight, and these Guidelines have been prepared to provide those new to the field and others engaged in teaching and instruction with the information necessary to increase their awareness of the problems and to enable them to deal with them effectively. The Guidelines cover areas such as development, acquisition, authentication, cryopreservation, transfer of cell lines between laboratories, microbial contamination, characterisation, instability and misidentification. Advice is also given on complying with current legal and ethical requirements when deriving cell lines from human and animal tissues, the selection and maintenance of equipment and how to deal with problems that may arise. PMID:25117809

  5. Establishment of mesenchymal cell line derived from human developing odontoma.

    Science.gov (United States)

    Hatano, H; Kudo, Y; Ogawa, I; Shimasue, H; Shigeishi, H; Ohta, K; Higashikawa, K; Takechi, M; Takata, T; Kamata, N

    2012-11-01

    An odontoma, which shows proliferating odontogenic epithelium and mesenchymal tissue, is one of the most common odontogenic tumors encountered. These are commonly found in tooth-bearing regions, although the etiology remains unknown. There are no previous reports of an established line of immortalized human odontoma cells. Using odontoma fragments obtained from a girl treated at our department, we established an immortalized human odontoma cell line and investigated cell morphology, dynamic proliferation, the presence of contamination, and karyotype. Moreover, cell characterization was examined using osteogenic and odontogenic markers. We successfully established a mesenchymal odontoma cell (mOd cells). The cells were found to be fibroblastic and had a high level of telomerase activity. Cell growth was confirmed after more than 200 population doublings without significant growth retardation. mOd cells expressed mRNA for differentiation markers, including collagen type I (COLI), alkaline phosphatase, bone sialoprotein, osteopontin, osteocalcin, cementum-derived protein (CP-23), dentin sialophosphoprotein (DSPP), and distal-less homeobox 3 (DLX3), as well as bone morphogenetic proteins (BMPs). In addition, they showed a high level of calcified nodule formation activity in vitro. We successfully established a cell line that may be useful for investigating the mechanisms of normal odontogenesis as well as characteristics of odontoma tumors. © 2012 John Wiley & Sons A/S.

  6. Isolation of Oct4-expressing extraembryonic endoderm precursor cell lines.

    Directory of Open Access Journals (Sweden)

    Bisrat G Debeb

    Full Text Available BACKGROUND: The extraembryonic endoderm (ExEn defines the yolk sac, a set of membranes that provide essential support for mammalian embryos. Recent findings suggest that the committed ExEn precursor is present already in the embryonic Inner Cell Mass (ICM as a group of cells that intermingles with the closely related epiblast precursor. All ICM cells contain Oct4, a key transcription factor that is first expressed at the morula stage. In vitro, the epiblast precursor is most closely represented by the well-characterized embryonic stem (ES cell lines that maintain the expression of Oct4, but analogous ExEn precursor cell lines are not known and it is unclear if they would express Oct4. METHODOLOGY/PRINCIPAL FINDINGS: Here we report the isolation and characterization of permanently proliferating Oct4-expressing rat cell lines ("XEN-P cell lines", which closely resemble the ExEn precursor. We isolated the XEN-P cell lines from blastocysts and characterized them by plating and gene expression assays as well as by injection into embryos. Like ES cells, the XEN-P cells express Oct4 and SSEA1 at high levels and their growth is stimulated by leukemia inhibitory factor, but instead of the epiblast determinant Nanog, they express the ExEn determinants Gata6 and Gata4. Further, they lack markers characteristic of the more differentiated primitive/visceral and parietal ExEn stages, but exclusively differentiate into these stages in vitro and contribute to them in vivo. CONCLUSIONS/SIGNIFICANCE: Our findings (i suggest strongly that the ExEn precursor is a self-renewable entity, (ii indicate that active Oct4 gene expression (transcription plus translation is part of its molecular identity, and (iii provide an in vitro model of early ExEn differentiation.

  7. Establishment, immortalisation and characterisation of pteropid bat cell lines.

    Directory of Open Access Journals (Sweden)

    Gary Crameri

    Full Text Available BACKGROUND: Bats are the suspected natural reservoir hosts for a number of new and emerging zoonotic viruses including Nipah virus, Hendra virus, severe acute respiratory syndrome coronavirus and Ebola virus. Since the discovery of SARS-like coronaviruses in Chinese horseshoe bats, attempts to isolate a SL-CoV from bats have failed and attempts to isolate other bat-borne viruses in various mammalian cell lines have been similarly unsuccessful. New stable bat cell lines are needed to help with these investigations and as tools to assist in the study of bat immunology and virus-host interactions. METHODOLOGY/FINDINGS: Black flying foxes (Pteropus alecto were captured from the wild and transported live to the laboratory for primary cell culture preparation using a variety of different methods and culture media. Primary cells were successfully cultured from 20 different organs. Cell immortalisation can occur spontaneously, however we used a retroviral system to immortalise cells via the transfer and stable production of the Simian virus 40 Large T antigen and the human telomerase reverse transcriptase protein. Initial infection experiments with both cloned and uncloned cell lines using Hendra and Nipah viruses demonstrated varying degrees of infection efficiency between the different cell lines, although it was possible to infect cells in all tissue types. CONCLUSIONS/SIGNIFICANCE: The approaches developed and optimised in this study should be applicable to bats of other species. We are in the process of generating further cell lines from a number of different bat species using the methodology established in this study.

  8. Non-targeted radiation effects in vertebrate cell lines

    Science.gov (United States)

    Ryan, Lorna

    Radiation effects, such as bystander effects, hyper radiosensitivity/induced radioresistance (HRS/IRR) and adaptive response that are not related to direct DNA damage are now accepted. However the inter-relationship between them and the possible impact on the scientific basis for radiation protection are highly controversial. This thesis attempts to elucidate the mechanisms of some of these well known but little understood effects. Each paper examines some aspect of bystander effects, adaptive responses and HRS/IRR in an effort to understand how they vary with cell type, dose and time of exposure to single or multiple doses. All the effects involve non-linear dose effect curves and are mainly evident following low doses. Overall findings of the thesis include (1) A clear difference was observed between radioresistant, tumorigenic cell lines with mutant p53 gene expression, and radiosensitive, more normal, cell lines with wild type p53. In general death inducing bystander responses are induced in normal cell populations exposed to low doses of radiation while survival inducing IRR and adaptive responses are seen in the radioresistant tumorigenic cell lines. (2) A cohort of fish cell lines which demonstrated survival promoting bystander effects, also did not show a protective adaptive responses. (3) Adaptive responses traditionally occur when a large challenge dose is given 4--6hrs following low (10--100mGy) priming doses but this thesis shows that for the epithelial cell lines tested, the size of the priming dose (range 0.1--2Gy) does not appear to alter the size of the recovery response. Additionally increased survival could be detected in some cases when the challenge dose was given within one hour of the priming dose. The overall conclusion is that cell lines induce either a bystander response or a protective/adaptive response depending on genetic background and other factors. Care is needed in the interpretation of data generated from only one or two cell lines

  9. Characterization of a human ovarian carcinoma cell line: UCI 101.

    Science.gov (United States)

    Fuchtner, C; Emma, D A; Manetta, A; Gamboa, G; Bernstein, R; Liao, S Y

    1993-02-01

    A new epithelial ovarian carcinoma cell line (UCI 101) has been established from the ascitic fluids and solid tumor of a patient with progressive papillary adenocarcinoma of the ovary shown previously to be refractory to combination chemotherapy consisting of cyclophosphamide, Adriamycin, and cisplatin as well as single-agent chemotherapy of taxol and high-dose cisplatin. The UCI 101 cell line grows well with an in vitro doubling time of 24 hr. The cell line expresses the B 72.3 (Tag 72), CA125, MH99 (ESA), and E29 (EMA) cell surface antigens and AE1/AE3 cytokeratins. This cell line overexpresses (as determined by immunocytochemistry) both p-glycoprotein and the epidermal growth factor receptor. The in vitro drug response to single agents including Adriamycin, cisplatin, dequalinium chloride, etoposide, 5-fluorouracil, taxol, and tumor necrosis factor was examined. Intraperitoneal transplantation of the cells into athymic mice resulted in foci of tumor on all peritoneal surfaces including the viscera and diaphragm ultimately leading to solid bulky disease with massive production of ascites. High levels of CA125 (> 500 units/ml) were detected in the serum of tumor-bearing mice. Cytogenetic analysis of cultured cells shows several marker chromosomes containing deletions, duplications, and translocations. Cytologic and histologic evaluation of the xenograft revealed morphological characteristics identical to those of the original tumor.

  10. SENSORY HAIR CELL REGENERATION IN THE ZEBRAFISH LATERAL LINE

    Science.gov (United States)

    Lush, Mark E.; Piotrowski, Tatjana

    2014-01-01

    Damage or destruction of sensory hair cells in the inner ear leads to hearing or balance deficits that can be debilitating, especially in older adults. Unfortunately, the damage is permanent, as regeneration of the inner ear sensory epithelia does not occur in mammals. Zebrafish and other non-mammalian vertebrates have the remarkable ability to regenerate sensory hair cells and understanding the molecular and cellular basis for this regenerative ability will hopefully aid us in designing therapies to induce regeneration in mammals. Zebrafish not only possess hair cells in the ear but also in the sensory lateral line system. Hair cells in both organs are functionally analogous to hair cells in the inner ear of mammals. The lateral line is a mechanosensory system found in most aquatic vertebrates that detects water motion and aids in predator avoidance, prey capture, schooling and mating. Although hair cell regeneration occurs in both the ear and lateral line, most research to date has focused on the lateral line due to its relatively simple structure and accessibility. Here we review the recent discoveries made during the characterization of hair cell regeneration in zebrafish. PMID:25045019

  11. Nestin expression in the cell lines derived from glioblastoma multiforme

    International Nuclear Information System (INIS)

    Veselska, Renata; Kuglik, Petr; Cejpek, Pavel; Svachova, Hana; Neradil, Jakub; Loja, Tomas; Relichova, Jirina

    2006-01-01

    Nestin is a protein belonging to class VI of intermediate filaments that is produced in stem/progenitor cells in the mammalian CNS during development and is consecutively replaced by other intermediate filament proteins (neurofilaments, GFAP). Down-regulated nestin may be re-expressed in the adult organism under certain pathological conditions (brain injury, ischemia, inflammation, neoplastic transformation). Our work focused on a detailed study of the nestin cytoskeleton in cell lines derived from glioblastoma multiforme, because re-expression of nestin together with down-regulation of GFAP has been previously reported in this type of brain tumor. Two cell lines were derived from the tumor tissue of patients treated for glioblastoma multiforme. Nestin and other cytoskeletal proteins were visualized using imunocytochemical methods: indirect immunofluorescence and immunogold-labelling. Using epifluorescence and confocal microscopy, we described the morphology of nestin-positive intermediate filaments in glioblastoma cells of both primary cultures and the derived cell lines, as well as the reorganization of nestin during mitosis. Our most important result came through transmission electron microscopy and provided clear evidence that nestin is present in the cell nucleus. Detailed information concerning the pattern of the nestin cytoskeleton in glioblastoma cell lines and especially the demonstration of nestin in the nucleus represent an important background for further studies of nestin re-expression in relationship to tumor malignancy and invasive potential

  12. Pluronic polyols in human lymphocyte cell line cultures.

    Science.gov (United States)

    Mizrahi, A

    1975-01-01

    Pluronic polyols markedly improved the growth of two human lymphocyte cell lines when added to the growth medium in concentrations of 0.05 to 0.1%. The results of the current studies suggest that, in addition to the protective effect of polyols against mechanical damage of mammalian cells in submerged cultures, the pluronic compounds may also, by lowering surface tension, facilitate transport of metabolites into cells and thus increase the growth rate. PMID:1063740

  13. Lactobacillaceae and cell adhesion: genomic and functional screening.

    Directory of Open Access Journals (Sweden)

    Williams Turpin

    Full Text Available The analysis of collections of lactic acid bacteria (LAB from traditional fermented plant foods in tropical countries may enable the detection of LAB with interesting properties. Binding capacity is often the main criterion used to investigate the probiotic characteristics of bacteria. In this study, we focused on a collection of 163 Lactobacillaceace comprising 156 bacteria isolated from traditional amylaceous fermented foods and seven strains taken from a collection and used as controls. The collection had a series of analyses to assess binding potential for the selection of new probiotic candidates. The presence/absence of 14 genes involved in binding to the gastrointestinal tract was assessed. This enabled the detection of all the housekeeping genes (ef-Tu, eno, gap, groEl and srtA in the entire collection, of some of the other genes (apf, cnb, fpbA, mapA, mub in 86% to 100% of LAB, and of the other genes (cbsA, gtf, msa, slpA in 0% to 8% of LAB. Most of the bacteria isolated from traditional fermented foods exhibited a genetic profile favorable for their binding to the gastrointestinal tract. We selected 30 strains with different genetic profiles to test their binding ability to non-mucus (HT29 and mucus secreting (HT29-MTX cell lines as well as their ability to degrade mucus. Assays on both lines revealed high variability in binding properties among the LAB, depending on the cell model used. Finally, we investigated if their binding ability was linked to tighter cross-talk between bacteria and eukaryotic cells by measuring the expression of bacterial genes and of the eukaryotic MUC2 gene. Results showed that wild LAB from tropical amylaceous fermented food had a much higher binding capacity than the two LAB currently known to be probiotics. However their adhesion was not linked to any particular genetic equipment.

  14. Human squamous cell carcinoma. Establishment and characterization of new permanent cell lines.

    Science.gov (United States)

    Krause, C J; Carey, T E; Ott, R W; Hurbis, C; McClatchey, K D; Regezi, J A

    1981-11-01

    Squamous cell carcinoma is the most common of human cancers, and yet because it is poorly represented by cultured cell lines, little is known about the characteristic cell biology and the cell-surface antigenic phenotypes of such tumors. To develop a continuously available source of squamous cell carcinoma for repeated and reproducible serologic analysis and for better understanding of its biologic characteristics, tissue culture methods and nude mice were used to establish new cell lines of squamous carcinoma. Special media, serum supplements from several sources, and methods of handling fresh tissue specimens were all examined as a means of improving the survival of tumor cell lines. Several new cell lines were established. Features characteristic of a squamous cell origin, eg, microvilli, desmosomes, tonofilaments, and the squamous cell differentiation antigen (pemphigus antigen), were found. The clinical course of disease in individual donor patients has been examined.

  15. ERK controls epithelial cell death receptor signalling and cellular FLICE-like inhibitory protein (c-FLIP) in ulcerative colitis

    DEFF Research Database (Denmark)

    Seidelin, Jakob Benedict; Coskun, Mehmet; Vainer, Ben

    2013-01-01

    Intestinal epithelial cell (IEC) death signalling through the Fas receptor is impaired in active ulcerative colitis (UC). This is possibly due to the activation of cytoprotective pathways resulting in limitation of the tissue injury secondary to inflammation. We hypothesized that inflammatory...... in IECs in active UC as well as IECs exposed to pro-inflammatory cytokines in vitro. Similarly, the short form of c-FLIP (c-FLIPS) was found to be upregulated in IECs from patients with active UC. c-FLIPS was the main splice variant found in both HT-29 cells and primary human IECs. Both splice variants....... Similarly, ERK - but not NF-κB - inhibited Fas ligand and TNF-α-mediated apoptosis responses in both cell line experiments and primary IECs. The present study identifies the MEK-ERK pathway as a major regulator of apoptosis in IECs during flares of UC and an inducer of c-FLIPS. The results explain...

  16. NADPH oxidase 1 supports proliferation of colon cancer cells by modulating reactive oxygen species-dependent signal transduction.

    Science.gov (United States)

    Juhasz, Agnes; Markel, Susan; Gaur, Shikha; Liu, Han; Lu, Jiamo; Jiang, Guojian; Wu, Xiwei; Antony, Smitha; Wu, Yongzhong; Melillo, Giovanni; Meitzler, Jennifer L; Haines, Diana C; Butcher, Donna; Roy, Krishnendu; Doroshow, James H

    2017-05-12

    Reactive oxygen species (ROS) play a critical role in cell signaling and proliferation. NADPH oxidase 1 (NOX1), a membrane-bound flavin dehydrogenase that generates O 2 ̇̄ , is highly expressed in colon cancer. To investigate the role that NOX1 plays in colon cancer growth, we used shRNA to decrease NOX1 expression stably in HT-29 human colon cancer cells. The 80-90% decrease in NOX1 expression achieved by RNAi produced a significant decline in ROS production and a G 1 /S block that translated into a 2-3-fold increase in tumor cell doubling time without increased apoptosis. The block at the G 1 /S checkpoint was associated with a significant decrease in cyclin D 1 expression and profound inhibition of mitogen-activated protein kinase (MAPK) signaling. Decreased steady-state MAPK phosphorylation occurred concomitant with a significant increase in protein phosphatase activity for two colon cancer cell lines in which NOX1 expression was knocked down by RNAi. Diminished NOX1 expression also contributed to decreased growth, blood vessel density, and VEGF and hypoxia-inducible factor 1α (HIF-1α) expression in HT-29 xenografts initiated from NOX1 knockdown cells. Microarray analysis, supplemented by real-time PCR and Western blotting, revealed that the expression of critical regulators of cell proliferation and angiogenesis, including c-MYC, c-MYB, and VEGF, were down-regulated in association with a decline in hypoxic HIF-1α protein expression downstream of silenced NOX1 in both colon cancer cell lines and xenografts. These studies suggest a role for NOX1 in maintaining the proliferative phenotype of some colon cancers and the potential of NOX1 as a therapeutic target in this disease. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Distinct metabolic responses of an ovarian cancer stem cell line.

    Science.gov (United States)

    Vermeersch, Kathleen A; Wang, Lijuan; McDonald, John F; Styczynski, Mark P

    2014-12-18

    Cancer metabolism is emerging as an important focus area in cancer research. However, the in vitro cell culture conditions under which much cellular metabolism research is performed differ drastically from in vivo tumor conditions, which are characterized by variations in the levels of oxygen, nutrients like glucose, and other molecules like chemotherapeutics. Moreover, it is important to know how the diverse cell types in a tumor, including cancer stem cells that are believed to be a major cause of cancer recurrence, respond to these variations. Here, in vitro environmental perturbations designed to mimic different aspects of the in vivo environment were used to characterize how an ovarian cancer cell line and its derived, isogenic cancer stem cells metabolically respond to environmental cues. Mass spectrometry was used to profile metabolite levels in response to in vitro environmental perturbations. Docetaxel, the chemotherapeutic used for this experiment, caused significant metabolic changes in amino acid and carbohydrate metabolism in ovarian cancer cells, but had virtually no metabolic effect on isogenic ovarian cancer stem cells. Glucose deprivation, hypoxia, and the combination thereof altered ovarian cancer cell and cancer stem cell metabolism to varying extents for the two cell types. Hypoxia had a much larger effect on ovarian cancer cell metabolism, while glucose deprivation had a greater effect on ovarian cancer stem cell metabolism. Core metabolites and pathways affected by these perturbations were identified, along with pathways that were unique to cell types or perturbations. The metabolic responses of an ovarian cancer cell line and its derived isogenic cancer stem cells differ greatly under most conditions, suggesting that these two cell types may behave quite differently in an in vivo tumor microenvironment. While cancer metabolism and cancer stem cells are each promising potential therapeutic targets, such varied behaviors in vivo would need to

  18. Modeling adenovirus latency in human lymphocyte cell lines.

    Science.gov (United States)

    Zhang, Yange; Huang, Wen; Ornelles, David A; Gooding, Linda R

    2010-09-01

    Species C adenovirus establishes a latent infection in lymphocytes of the tonsils and adenoids. To understand how this lytic virus is maintained in these cells, four human lymphocytic cell lines that support the entire virus life cycle were examined. The T-cell line Jurkat ceased proliferation and died shortly after virus infection. BJAB, Ramos (B cells), and KE37 (T cells) continued to divide at nearly normal rates while replicating the virus genome. Viral genome numbers peaked and then declined in BJAB cells below one genome per cell at 130 to 150 days postinfection. Ramos and KE37 cells maintained the virus genome at over 100 copies per cell over a comparable period of time. BJAB cells maintained the viral DNA as a monomeric episome. All three persistently infected cells lost expression of the cell surface coxsackie and adenovirus receptor (CAR) within 24 h postinfection, and CAR expression remained low for at least 340 days postinfection. CAR loss proceeded via a two-stage process. First, an initial loss of cell surface staining for CAR required virus late gene expression and a CAR-binding fiber protein even while CAR protein and mRNA levels remained high. Second, CAR mRNA disappeared at around 30 days postinfection and remained low even after virus DNA was lost from the cells. At late times postinfection (day 180), BJAB cells could not be reinfected with adenovirus, even when CAR was reintroduced to the cells via retroviral transduction, suggesting that the expression of multiple genes had been stably altered in these cells following infection.

  19. Differential heat shock response of primary human cell cultures and established cell lines

    DEFF Research Database (Denmark)

    Richter, W W; Issinger, O G

    1986-01-01

    degrees C treatment, whereas in immortalized cell lines usually 90% of the cells were found in suspension. Enhanced expression of the major heat shock protein (hsp 70) was found in all heat-treated cells. In contrast to the primary cell cultures, established and transformed cell lines synthesized...... a protein with an apparent molecular mass of 70 kDa and an isoelectric pH of 7.0 as early as 3 h after the initial hyperthermal treatment....

  20. Dipeptidyl peptidase IV in two human glioma cell lines

    Directory of Open Access Journals (Sweden)

    A Sedo

    2009-12-01

    Full Text Available There is growing evidence that dipeptidyl peptidase IV [DPP-IV, EC 3.4.14.5] takes part in the metabolism of biologically active peptides participating in the regulation of growth and transformation of glial cells. However, the knowledge on the DPP-IV expression in human glial and glioma cells is still very limited. In this study, using histochemical and biochemical techniques, the DPP-IV activity was demonstrated in two commercially available human glioma cell lines of different transformation degree, as represented by U373 astrocytoma (Grade III and U87 glioblastoma multiforme (Grade IV lines. Higher total activity of the enzyme, as well as its preferential localisation in the plasma membrane, was observed in U87 cells. Compared to U373 population, U87 cells were morphologically more pleiomorphic, they were cycling at lower rate and expressing less Glial Fibrillary Acidic Protein. The data revealed positive correlation between the degree of transformation of cells and activity of DPP-IV. Great difference in expression of this enzyme, together with the phenotypic differences of cells, makes these lines a suitable standard model for further 57 studies of function of this enzyme in human glioma cells.

  1. Effect of Predatory Bacteria on Human Cell Lines.

    Directory of Open Access Journals (Sweden)

    Shilpi Gupta

    Full Text Available Predatory bacteria are Gram-negative bacteria that prey on other Gram-negative bacteria and have been considered as potential therapeutic agents against multi-drug resistant pathogens. In vivo animal models have demonstrated that predatory bacteria are non-toxic and non-immunogenic in rodents. In order to consider the use of predatory bacteria as live antibiotics, it is important to investigate their effect on human cells. The aim of this study was to determine the effect of Bdellovibrio bacteriovorus strains 109J and HD100, and Micavibrio aeruginosavorus strain ARL-13 on cell viability and inflammatory responses of five human cell lines, representative of clinically relevant tissues. We found that the predators were not cytotoxic to any of the human cell lines tested. Microscopic imaging showed no signs of cell detachment, as compared to predator-free cells. In comparison to an E. coli control, exposure to higher concentrations of the predators did not trigger a significant elevation of pro-inflammatory cytokines in four of the five human cell lines tested. Our work underlines the non-pathogenic attributes of predatory bacteria on human cells and highlights their potential use as live antibiotics against human pathogens.

  2. Pheochromocytoma cell lines from heterozygous neurofibromatosis knockout mice.

    Science.gov (United States)

    Powers, J F; Evinger, M J; Tsokas, P; Bedri, S; Alroy, J; Shahsavari, M; Tischler, A S

    2000-12-01

    Transplantable tumors and cell lines have been developed from pheochromocytomas arising in mice with a heterozygous knockout mutation of the neurofibromatosis gene, Nf1. Nf1 encodes a ras-GTPase-activating protein, neurofibromin, and mouse pheochromocytoma (MPC) cells in primary cultures typically show extensive spontaneous neuronal differentiation that may result from the loss of the remaining wild-type allele and defective regulation of ras signaling. However, all MPC cell lines express neurofibromin, suggesting that preservation of the wild-type allele may be required to permit the propagation of MPC cells in vitro. MPC lines differ from PC12 cells in that they express both endogenous phenylethanolamine N-methyltransferase (PNMT) and full-length PNMT reporter constructs. PNMT expression is increased by dexamethasone and by cell-cell contact in suspension cultures. Mouse pheochromocytomas are a new tool for studying genes and signaling pathways that regulate cell growth and differentiation in adrenal medullary neoplasms and are a unique model for studying the regulation of PNMT expression.

  3. Graphene Oxide Nanoribbons Induce Autophagic Vacuoles in Neuroblastoma Cell Lines

    Directory of Open Access Journals (Sweden)

    Emanuela Mari

    2016-11-01

    Full Text Available Since graphene nanoparticles are attracting increasing interest in relation to medical applications, it is important to understand their potential effects on humans. In the present study, we prepared graphene oxide (GO nanoribbons by oxidative unzipping of single-wall carbon nanotubes (SWCNTs and analyzed their toxicity in two human neuroblastoma cell lines. Neuroblastoma is the most common solid neoplasia in children. The hallmark of these tumors is the high number of different clinical variables, ranging from highly metastatic, rapid progression and resistance to therapy to spontaneous regression or change into benign ganglioneuromas. Patients with neuroblastoma are grouped into different risk groups that are characterized by different prognosis and different clinical behavior. Relapse and mortality in high risk patients is very high in spite of new advances in chemotherapy. Cell lines, obtained from neuroblastomas have different genotypic and phenotypic features. The cell lines SK-N-BE(2 and SH-SY5Y have different genetic mutations and tumorigenicity. Cells were exposed to low doses of GO for different times in order to investigate whether GO was a good vehicle for biological molecules delivering individualized therapy. Cytotoxicity in both cell lines was studied by measuring cellular oxidative stress (ROS, mitochondria membrane potential, expression of lysosomial proteins and cell growth. GO uptake and cytoplasmic distribution of particles were studied by Transmission Electron Microscopy (TEM for up to 72 h. The results show that GO at low concentrations increased ROS production and induced autophagy in both neuroblastoma cell lines within a few hours of exposure, events that, however, are not followed by growth arrest or death. For this reason, we suggest that the GO nanoparticle can be used for therapeutic delivery to the brain tissue with minimal effects on healthy cells.

  4. External Qi of Yan Xin Qigong Inhibits Activation of Akt, Erk1/2 and NF-ĸB and Induces Cell Cycle Arrest and Apoptosis in Colorectal Cancer Cells

    Directory of Open Access Journals (Sweden)

    Xin Yan

    2013-01-01

    Full Text Available Background/Aims: Colorectal cancer (CRC is the second leading cause of cancer death in the Western countries. Novel approaches of treatment are needed for CRC. The purpose of the present study was to investigate cytotoxic effect of external Qi of Yan Xin Qigong (YXQ-EQ on human colorectal cancer cells. Methods: The effect of YXQ-EQ on viability, cell cycle progression and apoptosis in colorectal cancer HT-29 cells was investigated. Phosphorylation of Akt and Erk1/2, activation of NF-ĸB and the expression of proteins involved in regulation of cell cycle and apoptosis were examined by Western blot analysis. Results: YXQ-EQ markedly decreased viability and blocked colony formation of HT-29 cells. YXQ-EQ downregulated cyclin D1 expression and increased accumulation of cyclin-dependent kinase inhibitors p21Cip1 and p27Kip1, resulting in G1 cell cycle arrest. YXQ-EQ induced apoptosis in HT-29 cells in association with decreased expression of antiapoptotic proteins Bcl-xL, XIAP, survivin and Mcl-1 and elevated expression of proapoptotic protein Bax. YXQ-EQ significantly repressed phosphorylation of Akt and Erk1/2 and NF-ĸB activation in HT-29 cells, suggesting that YXQ-EQ may exert cytotoxic effect through regulating signaling pathways critical for cell proliferation and survival. Furthermore, YXQ-EQ treated PBS and an YXQ-EQ treated plant extract induced apoptosis in HT-29 cells. Conclusion: These findings show that YXQ-EQ has potent cytotoxic effect on HT-29 cells and suggest that YXQ-EQ could be potentially used for colorectal cancer treatment either directly or indirectly via carriers.

  5. Antiproliferative effect of Tualang honey on oral squamous cell carcinoma and osteosarcoma cell lines

    Directory of Open Access Journals (Sweden)

    Ismail Noorliza M

    2010-09-01

    Full Text Available Abstract Background The treatment of oral squamous cell carcinomas (OSCC and human osteosarcoma (HOS includes surgery and/or radiotherapy which often lead to reduced quality of life. This study was aimed to study the antiproliferative activity of local honey (Tualang on OSCC and HOS cell lines. Methods Several concentrations of Tualang honey (1% - 20% were applied on OSCC and HOS cell lines for 3, 6, 12, 24, 48 and 72 hours. Morphological characteristics were observed under light and fluorescent microscope. Cell viability was assessed using MTT assay and the optical density for absorbance values in each experiment was measured at 570 nm by an ELISA reader. Detection of cellular apoptosis was done using the Annexin V-FITC Apoptosis Detection Kit. Results Morphological appearance showed apoptotic cellular changes like becoming rounded, reduction in cell number, blebbed membrane and apoptotic nuclear changes like nuclear shrinkage, chromatin condensation and fragmented nucleus on OSCC and HOS cell lines. Cell viability assay showed a time and dose-dependent inhibitory effect of honey on both cell lines. The 50% inhibitory concentration (IC50 for OSCC and HOS cell lines was found to be 4% and 3.5% respectively. The maximum inhibition of cell growth of ≥80% was obtained at 15% for both cell lines. Early apoptosis was evident by flow cytometry where percentage of early apoptotic cells increased in dose and time dependent manner. Conclusion Tualang honey showed antiproliferative effect on OSCC and HOS cell lines by inducing early apoptosis.

  6. Characterization of cloned cells from an immortalized fetal pulmonary type II cell line

    Energy Technology Data Exchange (ETDEWEB)

    Henderson, R.F.; Waide, J.J.; Lechner, J.F.

    1995-12-01

    A cultured cell line that maintained expression of pulmonary type II cell markers of differentiation would be advantageous to generate a large number of homogenous cells in which to study the biochemical functions of type II cells. Type II epithelial cells are the source of pulmonary surfactant and a cell of origin for pulmonary adenomas. Last year our laboratory reported the induction of expression of two phenotypic markers of pulmonary type II cells (alkaline phosphatase activity and surfactant lipid synthesis) in cultured fetal rat lung epithelial (FRLE) cells, a spontaneously immortalized cell line of fetal rat lung type II cell origin. Subsequently, the induction of the ability to synthesize surfactant lipid became difficult to repeat. We hypothesized that the cell line was heterogenuous and some cells were more like type II cells than others. The purpose of this study was to test this hypothesis and to obtain a cultured cell line with type II cell phenotypic markers by cloning several FRLE cells and characterizing them for phenotypic markers of type II cells (alkaline phosphatase activity and presence of surfactant lipids). Thirty cloned cell lines were analyzed for induced alkaline phosphatase activity (on x-axis) and for percent of phospholipids that were disaturated (i.e., surfactant).

  7. Expression of the epidermal growth factor receptor in human small cell lung cancer cell lines

    DEFF Research Database (Denmark)

    Damstrup, L; Rygaard, K; Spang-Thomsen, M

    1992-01-01

    Epidermal growth factor (EGF) receptor expression was evaluated in a panel of 21 small cell lung cancer cell lines with radioreceptor assay, affinity labeling, and Northern blotting. We found high-affinity receptors to be expressed in 10 cell lines. Scatchard analysis of the binding data...... lung cancer cell lines express the EGF receptor....... of EGF receptor mRNA in all 10 cell lines that were found to be EGF receptor-positive and in one cell line that was found to be EGF receptor-negative in the radioreceptor assay and affinity labeling. Our results provide, for the first time, evidence that a large proportion of a broad panel of small cell...

  8. Opposing effects of low versus high concentrations of water soluble vitamins/dietary ingredients Vitamin C and niacin on colon cancer stem cells (CSCs).

    Science.gov (United States)

    Sen, Utsav; Shenoy P, Sudheer; Bose, Bipasha

    2017-10-01

    Colorectal cancer is one of the global causes of cancer deaths. Cancer stem cells (CSCs) inside the tumour niche responsible for metastasis and relapses, and hence need to be targeted for cancer therapeutics. Although dietary fibre and lifestyle changes have been recommended as measures for colorectal cancer prevention, no such recommendations are available for using water soluble vitamins as prophylaxis measure for colorectal cancers. High dose of Vitamin C has been proven to selectively kill colon cancer cells having BRAF and KRAS mutations by inducing oxidative stress. In this study, we show for the first time the opposing effects of the low and high dose of Vitamin C and vitamin B3 on colon CSCs isolated from HT-29 and HCT-15 colorectal carcinoma cell lines. At small doses, both of these vitamins exerted a cell proliferative effect only on CSCs, while there was no change in the proliferation status of non-stem cancer cells and wild-type (WT) populations. On the other hand, the death effects induced by high doses of Vitamin C and B3 were of the order of 50-60% and ∼30% on CSCs from HT-29 and HCT15, respectively. Interestingly, the control fibroblast cell line (NIH3T3) was highly refractory all the tested concentrations of Vitamin C and B3, except for the highest dose - 10,000 μg of Vitamin C that induced only 15% of cell death. Hence, these results indicate the future scope of use of therapeutic doses of Vitamin C and B3 especially in patients with advanced colorectal cancer. © 2017 International Federation for Cell Biology.

  9. TKTL1 expression in human malign and benign cell lines.

    Science.gov (United States)

    Kämmerer, Ulrike; Gires, Olivier; Pfetzer, Nadja; Wiegering, Armin; Klement, Rainer Johannes; Otto, Christoph

    2015-06-10

    Overexpression of transketolase-like 1 protein TKTL1 in cancer cells has been reported to correlate with enhanced glycolysis and lactic acid production. Furthermore, enhanced TKTL1 expression was put into context with resistance to chemotherapy and ionizing radiation. Here, a panel of human malign and benign cells, which cover a broad range of chemotherapy and radiation resistance as well as reliance on glucose metabolism, was analyzed in vitro for TKTL1 expression. 17 malign and three benign cell lines were characterized according to their expression of TKTL1 on the protein level with three commercially available anti-TKTL1 antibodies utilizing immunohistochemistry and Western blot, as well as on mRNA level with three published primer pairs for RT-qPCR. Furthermore, sensitivities to paclitaxel, cisplatin and ionizing radiation were assessed in cell survival assays. Glucose consumption and lactate production were quantified as surrogates for the "Warburg effect". Considerable amounts of tktl1 mRNA and TKTL1 protein were detected only upon stable transfection of the human embryonic kidney cell line HEK293 with an expression plasmid for human TKTL1. Beyond that, weak expression of endogenous tktl1 mRNA was measured in the cell lines JAR and U251. Western blot analysis of JAR and U251 cells did not detect TKTL1 at the expected size of 65 kDa with all three antibodies specific for TKTL1 protein and immunohistochemical staining was observed with antibody JFC12T10 only. All other cell lines tested here revealed expression of tktl1 mRNA below detection limits and were negative for TKTL1 protein. However, in all cell lines including TKTL1-negative HEK293-control cells, antibody JFC12T10 detected multiple proteins with different molecular weights. Importantly, JAR and U251 did neither demonstrate an outstanding production of lactic acid nor increased resistance against chemotherapeutics or to ionizing radiation, respectively. Using RT-qPCR and three different antibodies we

  10. TKTL1 expression in human malign and benign cell lines

    International Nuclear Information System (INIS)

    Kämmerer, Ulrike; Gires, Olivier; Pfetzer, Nadja; Wiegering, Armin; Klement, Rainer Johannes; Otto, Christoph

    2015-01-01

    Overexpression of transketolase-like 1 protein TKTL1 in cancer cells has been reported to correlate with enhanced glycolysis and lactic acid production. Furthermore, enhanced TKTL1 expression was put into context with resistance to chemotherapy and ionizing radiation. Here, a panel of human malign and benign cells, which cover a broad range of chemotherapy and radiation resistance as well as reliance on glucose metabolism, was analyzed in vitro for TKTL1 expression. 17 malign and three benign cell lines were characterized according to their expression of TKTL1 on the protein level with three commercially available anti-TKTL1 antibodies utilizing immunohistochemistry and Western blot, as well as on mRNA level with three published primer pairs for RT-qPCR. Furthermore, sensitivities to paclitaxel, cisplatin and ionizing radiation were assessed in cell survival assays. Glucose consumption and lactate production were quantified as surrogates for the “Warburg effect”. Considerable amounts of tktl1 mRNA and TKTL1 protein were detected only upon stable transfection of the human embryonic kidney cell line HEK293 with an expression plasmid for human TKTL1. Beyond that, weak expression of endogenous tktl1 mRNA was measured in the cell lines JAR and U251. Western blot analysis of JAR and U251 cells did not detect TKTL1 at the expected size of 65 kDa with all three antibodies specific for TKTL1 protein and immunohistochemical staining was observed with antibody JFC12T10 only. All other cell lines tested here revealed expression of tktl1 mRNA below detection limits and were negative for TKTL1 protein. However, in all cell lines including TKTL1-negative HEK293-control cells, antibody JFC12T10 detected multiple proteins with different molecular weights. Importantly, JAR and U251 did neither demonstrate an outstanding production of lactic acid nor increased resistance against chemotherapeutics or to ionizing radiation, respectively. Using RT-qPCR and three different antibodies

  11. Maslinic acid inhibits proliferation of renal cell carcinoma cell lines and suppresses angiogenesis of endothelial cells

    Directory of Open Access Journals (Sweden)

    Parth Thakor

    2017-03-01

    Full Text Available Despite the introduction of many novel therapeutics in clinical practice, metastatic renal cell carcinoma (RCC remains a treatment-re-sistant cancer. As red and processed meat are considered risk factors for RCC, and a vegetable-rich diet is thought to reduce this risk, research into plant-based therapeutics may provide valuable complementary or alternative therapeutics for the management of RCC. Herein, we present the antiproliferative and antiangiogenic effects of maslinic acid, which occurs naturally in edible plants, particularly in olive fruits, and also in a variety of medicinal plants. Human RCC cell lines (ACHN, Caki-1, and SN12K1, endothelial cells (human umbilical vein endothelial cell line [HUVEC], and primary cultures of kidney proximal tubular epithelial cells (PTEC were treated with maslinic acid. Maslinic acid was relatively less toxic to PTEC when compared with RCC under similar experimental conditions. In RCC cell lines, maslinic acid induced a significant reduction in proliferation, proliferating cell nuclear antigen, and colony formation. In HUVEC, maslinic acid induced a significant reduction in capillary tube formation in vitro and vascular endothelial growth factor. This study provides a rationale for incorporating a maslinic acid–rich diet either to reduce the risk of developing kidney cancer or as an adjunct to existing antiangiogenic therapy to improve efficacy.

  12. Characterization of stem-like cells in a new astroblastoma cell line

    Energy Technology Data Exchange (ETDEWEB)

    Coban, Esra Aydemir; Kasikci, Ezgi [Department of Genetics and Bioengineering, Faculty of Engineering and Architecture, Yeditepe University, Istanbul (Turkey); Karatas, Omer Faruk [Molecular Biology and Genetics Department, Erzurum Technical University, Erzurum (Turkey); Suakar, Oznur; Kuskucu, Aysegul [Department of Medical Genetics, Yeditepe University Medical School and Yeditepe University Hospital, Istanbul (Turkey); Altunbek, Mine [Department of Genetics and Bioengineering, Faculty of Engineering and Architecture, Yeditepe University, Istanbul (Turkey); Türe, Uğur [Department of Neurosurgery, Yeditepe University School of Medicine, Istanbul (Turkey); Sahin, Fikrettin [Department of Genetics and Bioengineering, Faculty of Engineering and Architecture, Yeditepe University, Istanbul (Turkey); Bayrak, Omer Faruk, E-mail: ofbayrak@yeditepe.edu.tr [Department of Medical Genetics, Yeditepe University Medical School and Yeditepe University Hospital, Istanbul (Turkey)

    2017-03-15

    Cell lines established from tumors are the most commonly used models in cancer research, and their use in recent years has enabled a greater understanding of the biology of cancer and the means to develop effective treatment strategies. Astroblastomas are uncommon neuroepithelial tumors of glial origin, predominantly affecting young people, mainly teenagers and children, predominantly females. To date, only a single study has reported that astroblastomas contain a large number of neural stem-like cells, which had only a partial proliferation capacity and differentiation. Our objective was to establish an astroblastoma cell line to investigate the presence of astroblastic cells and cancer stem-like cells. The migratory and invasion abilities of the cells were quantified with invasion and migration assays and compared to a glioblastoma cell line. The presence of stem cells was detected with surface-marker analysis by using flow cytometry, and measuring the differentiation ability with a differentiation assay and the self-renewal capacity with a sphere-forming assay. These characteristics may determine whether this novel cell line is a model for astroblastomas that may have stem-cell characteristics. With this novel cell line, scientists can investigate the molecular pathways underlying astroblastomas and develop new therapeutic strategies for patients with these tumors. - Highlights: • An establishment of a novel astroblastoma cell line was proposed. • The presence of astroblastic cells and cancer stem-like cells was investigated. • The molecular pathways underlying astroblastomas may be investigated. • New therapeutic strategies for patients with astroblastoma may be developed.

  13. In vitro invasion of small-cell lung cancer cell lines correlates with expression of epidermal growth factor receptor

    DEFF Research Database (Denmark)

    Damstrup, L; Rude Voldborg, B; Spang-Thomsen, M

    1998-01-01

    receptor (EGFR) in a panel of 21 small-cell lung cancer (SCLC) cell lines. We have previously reported that ten of these cell lines expressed EGFR protein detected by radioreceptor and affinity labelling assays. In 11 small-cell lung cancer (SCLC) cell lines, EGFR mRNA was detected by Northern blot...... analysis. In vitro invasion in a Boyden chamber assay was found in all EGFR-positive cell lines, whereas no invasion was detected in the EGFR-negative cell lines. Quantification of the in vitro invasion in 12 selected SCLC cell lines demonstrated that, in the EGFR-positive cell lines, between 5% and 16......-PCR). However, in vitro invasive SCLC cell lines could not be distinguished from non-invasive cell lines based on the expression pattern of these molecules. In six SCLC cell lines, in vitro invasion was also determined in the presence of the EGFR-neutralizing monoclonal antibody mAb528. The addition...

  14. Changes in Chromosome Counts and Patterns in CHO Cell Lines upon Generation of Recombinant Cell Lines and Subcloning.

    Science.gov (United States)

    Vcelar, Sabine; Melcher, Michael; Auer, Norbert; Hrdina, Astrid; Puklowski, Anja; Leisch, Friedrich; Jadhav, Vaibhav; Wenger, Till; Baumann, Martina; Borth, Nicole

    2018-03-01

    Chinese hamster ovary (CHO) cells are the number one production system for therapeutic proteins. A pre-requirement for their use in industrial production of biopharmaceuticals is to be clonal, thus originating from a single cell in order to be phenotypically and genomically identical. In the present study it was evaluated whether standard procedures, such as the generation of a recombinant cell line in combination with selection for a specific and stable phenotype (expression of the recombinant product) or subcloning have any impact on karyotype stability or homogeneity in CHO cells. Analyses used were the distribution of chromosome counts per cell as well as chromosome painting to identify specific karyotype patterns within a population. Results indicate that subclones both of the host and the recombinant cell line are of comparable heterogeneity and (in)stability as the original pool. In contrast, the rigorous selection for a stably expressing phenotype generated cell lines with fewer variation and more stable karyotypes, both at the level of the sorted pool and derivative subclones. We conclude that the process of subcloning itself does not contribute to an improved karyotypic homogeneity of a population, while the selection for a specific cell property inherently can provide evolutionary pressure that may lead to improved chromosomal stability as well as to a more homogenous population. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Derivation of Ethnically Diverse Human Induced Pluripotent Stem Cell Lines.

    Science.gov (United States)

    Chang, Eun Ah; Tomov, Martin L; Suhr, Steven T; Luo, Jiesi; Olmsted, Zachary T; Paluh, Janet L; Cibelli, Jose

    2015-10-20

    The human genome with all its ethnic variations contributes to differences in human development, aging, disease, repair, and response to medical treatments and is an exciting area of research and clinical study. The availability of well-characterized ethnically diverse stem cell lines is limited and has not kept pace with other advances in stem cell research. Here we derived xenofree ethnically diverse-human induced pluripotent stem cell (ED-iPSC) lines from fibroblasts obtained from individuals of African American, Hispanic-Latino, Asian, and Caucasian ethnic origin and have characterized the lines under a uniform platform for comparative analysis. Derived ED-iPSC lines are low passage number and evaluated in vivo by teratoma formation and in vitro by high throughput microarray analysis of EB formation and early differentiation for tri-lineage commitment to endoderm, ectoderm and mesoderm. These new xenofree ED-iPSC lines represent a well-characterized valuable resource with potential for use in future research in drug discovery or clinical investigations.

  16. Impact of Mesenchymal Stem Cell secreted PAI-1 on colon cancer cell migration and proliferation

    International Nuclear Information System (INIS)

    Hogan, Niamh M.; Joyce, Myles R.; Murphy, J. Mary; Barry, Frank P.; O’Brien, Timothy; Kerin, Michael J.; Dwyer, Roisin M.

    2013-01-01

    Highlights: •MSCs were directly co-cultured with colorectal cancer (CRC) cells on 3D scaffolds. •MSCs influence CRC protein/gene expression, proliferation and migration. •We report a significant functional role of MSC-secreted PAI-1 in colon cancer. -- Abstract: Mesenchymal Stem Cells are known to engraft and integrate into the architecture of colorectal tumours, with little known regarding their fate following engraftment. This study aimed to investigate mediators of Mesenchymal Stem Cell (MSC) and colon cancer cell (CCC) interactions. Mesenchymal Stem Cells and colon cancer cells (HT29 and HCT-116) were cultured individually or in co-culture on 3-dimensional scaffolds. Conditioned media containing all secreted factors was harvested at day 1, 3 and 7. Chemokine secretion and expression were analyzed by Chemi-array, ELISA (Macrophage migration inhibitory factor (MIF), plasminogen activator inhibitor type 1 (PAI-1)) and RQ-PCR. Colon cancer cell migration and proliferation in response to recombinant PAI-1, MSCs and MSCs + antibody to PAI-1 was analyzed using Transwell inserts and an MTS proliferation assay respectively. Chemi-array revealed secretion of a wide range of factors by each cell population, including PAI-1and MIF. ELISA analysis revealed Mesenchymal Stem Cells to secrete the highest levels of PAI-1 (MSC mean 10.6 ng/mL, CCC mean 1.01 ng/mL), while colon cancer cells were the principal source of MIF. MSC-secreted PAI-1 stimulated significant migration of both CCC lines, with an antibody to the chemokine shown to block this effect (67–88% blocking,). A cell-line dependant effect on CCC proliferation was shown for Mesenchymal Stem Cell-secreted PAI-1 with HCT-116 cells showing decreased proliferation at all concentrations, and HT29 cells showing increased proliferation in the presence of higher PAI-1 levels. This is the first study to identify PAI-1 as an important mediator of Mesenchymal Stem Cell/colon cancer cell interactions and highlights the

  17. Dipeptidyl peptidase IV in two human glioma cell lines

    Czech Academy of Sciences Publication Activity Database

    Šedo, A.; Malík, Radek; Drbal, K.; Lisá, Věra; Vlašicová, K.; Mareš, Vladislav

    2000-01-01

    Roč. 44, č. 1 (2000), s. 57-63 ISSN 1121-760X Grant - others:GA UK(XC) 58/1999/C; GA UK(XC) 206019-2 Institutional research plan: CEZ:AV0Z5011922 Keywords : dipeptidyl peptidase IV * glioma cell lines * cell proliferation and differentiation Subject RIV: FH - Neurology Impact factor: 1.039, year: 2000

  18. Cysteine modified polyaniline films improve biocompatibility for two cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Yslas, Edith I., E-mail: eyslas@exa.unrc.edu.ar [Departamento de Biología Molecular, Universidad Nacional de Río Cuarto, Agencia Postal Nro3, X580BYA Río Cuarto (Argentina); Cavallo, Pablo; Acevedo, Diego F.; Barbero, César A. [Departamento de Química, Universidad Nacional de Río Cuarto, Agencia Postal Nro3, X580BYA Río Cuarto (Argentina); Rivarola, Viviana A. [Departamento de Biología Molecular, Universidad Nacional de Río Cuarto, Agencia Postal Nro3, X580BYA Río Cuarto (Argentina)

    2015-06-01

    This work focuses on one of the most exciting application areas of conjugated conducting polymers, which is cell culture and tissue engineering. To improve the biocompatibility of conducting polymers we present an easy method that involves the modification of the polymer backbone using L-cysteine. In this publication, we show the synthesis of polyaniline (PANI) films supported onto Polyethylene terephthalate (PET) films, and modified using cysteine (PANI-Cys) in order to generate a biocompatible substrate for cell culture. The PANI-Cys films are characterized by Fourier Transform infrared and UV–visible spectroscopy. The changes in the hydrophilicity of the polymer films after and before the modification were tested using contact angle measurements. After modification the contact angle changes from 86° ± 1 to 90° ± 1, suggesting a more hydrophylic surface. The adhesion properties of LM2 and HaCaT cell lines on the surface of PANI-Cys films in comparison with tissue culture plastic (TCP) are studied. The PANI-Cys film shows better biocompatibility than PANI film for both cell lines. The cell morphologies on the TCP and PANI-Cys film were examined by florescence and Atomic Force Microscopy (AFM). Microscopic observations show normal cellular behavior when PANI-Cys is used as a substrate of both cell lines (HaCaT and LM2) as when they are cultured on TCP. The ability of these PANI-Cys films to support cell attachment and growth indicates their potential use as biocompatible surfaces and in tissue engineering. - Highlights: • A new surface PANI-Cys was produced on films of polyethylene terephthalate. • The relationship between surface characteristics and biocompatibility is analyzed. • The PANI-Cys film presents good biocompatibility for two cell lines.

  19. Establishment of clinically relevant radioresistant cell lines and their characteristics

    International Nuclear Information System (INIS)

    Fukumoto, Manabu; Kuwahara, Yoshikazu; Suzuki, Masatoshi

    2014-01-01

    Although radiotherapy is one of the major therapeutic modalities for eradicating malignant tumors, the existence of radioresistant cells remains one of the most critical obstacles. Standard radiotherapy consists of fractionated radiation (FR) of 2-Gy X-rays once a day, 5 days a week, over 60 Gy in total. To understand the characteristics of radioresistant cells and to develop more effective radiotherapy, we have established novel radioresistant cell lines by long-term (> 5 years) exposure to moderate doses of fractionated X-rays. While all the parental human cancer cells ceased, their radioresistant derivatives continue to proliferate with daily exposure to 2-Gy FR for more than 30 days. We have coined those cells as 'clinically relevant radioresistant' (CRR) cells. Transplanted tumors into nude mice were also CRR, indicating that CRR cell lines are powerful tools to improve cancer radiotherapy. We have shown that the suppression of autophagic cell death but not apoptosis was mainly involved in cellular radioresistance. An inhibitor of the mTOR pathway which enhances autophagy was effective to overcome CRR tumors induced in nude mice. But the underlined mechanism was not through the inhibition of autophagy. Guanine nucleotide-binding protein 1 (GBP1) over expression was necessary for maintaining the CRR phenotype, but radioresistant cells were not necessarily cancer stem cells (CSCs). Targeting GBP1 positive cancer cells may be a more efficient method in conquering cancer than targeting CSCs. Slight but significant radioresistance was acquired by 0.5 Gy/12 hrs of long-term FR exposures to parental cells for more than 31 days in accordance with cyclinD1 over expression. This acquired radioresistance (ARR) was stably maintained in the tumor cells even on 31 days after the cessation of 0.5-Gy FR. Present observations give a mechanistic insight for ARR of tumor cells through long-term FR exposure, and provide novel therapeutic targets for radiosensitization

  20. Differential proteome analysis of human embryonic kidney cell line (HEK-293 following mycophenolic acid treatment

    Directory of Open Access Journals (Sweden)

    Rahman Hazir

    2011-09-01

    Full Text Available Abstract Background Mycophenolic acid (MPA is widely used as a post transplantation medicine to prevent acute organ rejection. In the present study we used proteomics approach to identify proteome alterations in human embryonic kidney cells (HEK-293 after treatment with therapeutic dose of MPA. Following 72 hours MPA treatment, total protein lysates were prepared, resolved by two dimensional gel electrophoresis and differentially expressed proteins were identified by QTOF-MS/MS analysis. Expressional regulations of selected proteins were further validated by real time PCR and Western blotting. Results The proliferation assay demonstrated that therapeutic MPA concentration causes a dose dependent inhibition of HEK-293 cell proliferation. A significant apoptosis was observed after MPA treatment, as revealed by caspase 3 activity. Proteome analysis showed a total of 12 protein spots exhibiting differential expression after incubation with MPA, of which 7 proteins (complement component 1 Q subcomponent-binding protein, electron transfer flavoprotein subunit beta, cytochrome b-c1 complex subunit, peroxiredoxin 1, thioredoxin domain-containing protein 12, myosin regulatory light chain 2, and profilin 1 showed significant increase in their expression. The expression of 5 proteins (protein SET, stathmin, 40S ribosomal protein S12, histone H2B type 1 A, and histone H2B type 1-C/E/F/G/I were down-regulated. MPA mainly altered the proteins associated with the cytoskeleton (26%, chromatin structure/dynamics (17% and energy production/conversion (17%. Both real time PCR and Western blotting confirmed the regulation of myosin regulatory light chain 2 and peroxiredoxin 1 by MPA treatment. Furthermore, HT-29 cells treated with MPA and total kidney cell lysate from MMF treated rats showed similar increased expression of myosin regulatory light chain 2. Conclusion The emerging use of MPA in diverse pathophysiological conditions demands in-depth studies to

  1. Myelinating cocultures of rodent stem cell line-derived neurons and immortalized Schwann cells.

    Science.gov (United States)

    Ishii, Tomohiro; Kawakami, Emiko; Endo, Kentaro; Misawa, Hidemi; Watabe, Kazuhiko

    2017-10-01

    Myelination is one of the most remarkable biological events in the neuron-glia interactions for the development of the mammalian nervous system. To elucidate molecular mechanisms of cell-to-cell interactions in myelin synthesis in vitro, establishment of the myelinating system in cocultures of continuous neuronal and glial cell lines are desirable. In the present study, we performed co-culture experiments using rat neural stem cell-derived neurons or mouse embryonic stem (ES) cell-derived motoneurons with immortalized rat IFRS1 Schwann cells to establish myelinating cultures between these cell lines. Differentiated neurons derived from an adult rat neural stem cell line 1464R or motoneurons derived from a mouse ES cell line NCH4.3, were mixed with IFRS1 Schwann cells, plated, and maintained in serum-free F12 medium with B27 supplement, ascorbic acid, and glial cell line-derived neurotrophic factor. Myelin formation was demonstrated by electron microscopy at 4 weeks in cocultures of 1464R-derived neurons or NCH4.3-derived motoneurons with IFRS1 Schwann cells. These in vitro coculture systems utilizing the rodent stable stem and Schwann cell lines can be useful in studies of peripheral nerve development and regeneration. © 2017 Japanese Society of Neuropathology.

  2. CellMinerHCC: a microarray-based expression database for hepatocellular carcinoma cell lines.

    Science.gov (United States)

    Staib, Frank; Krupp, Markus; Maass, Thorsten; Itzel, Timo; Weinmann, Arndt; Lee, Ju-Seog; Schmidt, Bertil; Müller, Martina; Thorgeirsson, Snorri S; Galle, Peter R; Teufel, Andreas

    2014-04-01

    Therapeutic options for hepatocellular carcinoma (HCC) still remain limited. Development of gene targeted therapies is a promising option. A better understanding of the underlying molecular biology is gained in in vitro experiments. However, even with targeted manipulation of gene expression varying treatment responses were observed in diverse HCC cell lines. Therefore, information on gene expression profiles of various HCC cell lines may be crucial to experimental designs. To generate a publicly available database containing microarray expression profiles of diverse HCC cell lines. Microarray data were analyzed using an individually scripted R program package. Data were stored in a PostgreSQL database with a PHP written web interface. Evaluation and comparison of individual cell line expression profiles are supported via public web interface. This database allows evaluation of gene expression profiles of 18 HCC cell lines and comparison of differential gene expression between multiple cell lines. Analysis of commonly regulated genes for signaling pathway enrichment and interactions demonstrates a liver tumor phenotype with enrichment of major cancer related KEGG signatures like 'cancer' and 'inflammatory response'. Further molecular associations of strong scientific interest, e.g. 'lipid metabolism', were also identified. We have generated CellMinerHCC (http://www.medicalgenomics.org/cellminerhcc), a publicly available database containing gene expression data of 18 HCC cell lines. This database will aid in the design of in vitro experiments in HCC research, because the genetic specificities of various HCC cell lines will be considered. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. Cell lines radiosensitization of thyroid cancer by histone deacetylase inhibitors

    International Nuclear Information System (INIS)

    Perona, M; Dagrosa, M A; Rossich, L; Casal, M; Pisarev, M A; Thomasz, L; Juvenal G J

    2012-01-01

    Introduction: Thyroid cancer is the most common endocrine neoplasia. Surgical resection and radioactive iodine is an effective treatment for well-differentiated tumors. Histone deacetylase inhibitors (HDAC-I) are agents that cause hyperacetylation of histone proteins and as a consequence remodeling of chromatin structure. They can induce growth arrest, differentiation and apoptotic cell death in different tumor cells. The use of HDAC-I agents could be of utility to enhance the response to external radiation therapy of those thyroid cancers that are refractory to most conventional therapeutic treatments. Objective: To study the effect of HDAC-I as radiosensitizers for the treatment of thyroid cancer and their ability to induce differentiation of thyroid cancer cells. Materials and methods: The human thyroid follicular (WRO) and papillary (TPC-1) carcinoma cell lines were seeded and incubated with increasing doses (0, 0.3, 0.5, 1 and 1.5 mM) of the HDAC-I sodium butirate (NaB) and valproic acid (VA) to evaluate cell proliferation and iodide uptake. Cells were irradiated with a 60 Co γ-ray source (1 ± 5% Gy/min) and postirradiation survival was quantified with the colony formation assay. Survival fraction at 2 Gy (SF2) was calculated for each cell line. Cell cycle and cell death were evaluated at a dose of 3 Gy. Iodide uptake, PCR analysis and transient transfection studies were performed. Results: Cell proliferation was not significantly suppressed after 24 hours of incubation with both drugs at all assayed doses. Iodide uptake was not modified after incubation with HDAC-I of both cell lines. SF2 was reduced from 68 ± 1.6 % in the control WRO cells to 42 ± 3.8 % (P<0.001) in NaB-treated cells. In TPC-1 SF2 was reduced from 32 ± 1.1 % in the control cells to 24 ± 0.8 % (P<0.01). In VA-treated cells SF2 was reduced from 69 ± 0.02 % in control WRO cells to 56 ± 0.01 % (P<0.01) and from 31 ± 2 % in control TPC-1 cells to 11 ± 1 % (P<0.01). There was an arrest

  4. Detection of immunotoxicity using T-cell based cytokine reporter cell lines ('Cell Chip')

    International Nuclear Information System (INIS)

    Ringerike, Tove; Ulleraas, Erik; Voelker, Rene; Verlaan, Bert; Eikeset, Aase; Trzaska, Dominika; Adamczewska, Violetta; Olszewski, Maciej; Walczak-Drzewiecka, Aurelia; Arkusz, Joanna; Loveren, Henk van; Nilsson, Gunnar; Lovik, Martinus; Dastych, Jaroslaw; Vandebriel, Rob J.

    2005-01-01

    Safety assessment of chemicals and drugs is an important regulatory issue. The evaluation of potential adverse effects of compounds on the immune system depends today on animal experiments. An increasing demand, however, exists for in vitro alternatives. Cytokine measurement is a promising tool to evaluate chemical exposure effects on the immune system. Fortunately, this type of measurement can be performed in conjunction with in vitro exposure models. We have taken these considerations as the starting point to develop an in vitro method to efficiently screen compounds for potential immunotoxicity. The T-cell lymphoma cell line EL-4 was transfected with the regulatory sequences of interleukin (IL)-2, IL-4, IL-10, interferon (IFN)-γ or actin fused to the gene for enhanced green fluorescent protein (EGFP) in either a stabile or a destabilised form. Consequently, changes in fluorescence intensity represent changes in cytokine expression with one cell line per cytokine. We used this prototype 'Cell Chip' to test, by means of flow cytometry, the immunomodulatory potential of 13 substances and were able to detect changes in cytokine expression in 12 cases (successful for cyclosporine, rapamycin, pentamidine, thalidomide, bis(tri-n-butyltin)oxide, house dust mite allergen (Der p I), 1-chloro-2,4-dinitrobenzene, benzocaine, tolylene 2,4-diisocyanate, potassium tetrachloroplatinate, sodium dodecyl sulphate and mercuric chloride; unsuccessful for penicillin G). In conclusion, this approach seems promising for in vitro screening for potential immunotoxicity, especially when additional cell lines besides T-cells are included

  5. DNA methylation and sensitivity to antimetabolites in cancer cell lines.

    Science.gov (United States)

    Sasaki, Shin; Kobunai, Takashi; Kitayama, Joji; Nagawa, Hirokazu

    2008-02-01

    The prediction of the cellular direction of metabolic pathways toward either DNA synthesis or DNA methylation is crucial for determining the susceptibility of cancers to anti-metabolites such as fluorouracil (5-FU). We genotyped the methylenetetrahydrofolate reductase (MTHFR) gene in NCI-60 cancer cell lines, and identified the methylation status of 24 tumor suppressor genes using methylation-specific multiplex ligation-dependent probe amplification. The susceptibility of the cancer cell lines to seven antimetabolites was then determined. Cells homozygous for CC at MTHFR-A1298C were significantly more sensitive to cyclocytidine, cytarabine (AraC) and floxuridine than those with AA or AC (p=0.0215, p=0.0166, and p=0.0323, respectively), and carried more methylated tumor suppressor genes (p=0.0313). Among the 12 tumor suppressor genes which were methylated in >25% of cancer cell lines, the methylation status of TIMP3, APC and IGSF4 significantly correlated with sensitivity to pyrimidine synthesis inhibitors. In particular, cells with methylated TIMP3 had reduced mRNA levels and were significantly more sensitive to aphidicolin-glycinate, AraC and 5-FU than cells with unmethylated TIMP3. We speculate that MTHFR-A1298C homozygous CC might direct the methylation rather than the synthesis of DNA, and result in the methylation of several tumor suppressor genes such as TIMP3. These genes could be useful biological markers for predicting the efficacy of antimetabolites.

  6. Cryopreservation of specialized chicken lines using cultured primordial germ cells.

    Science.gov (United States)

    Nandi, S; Whyte, J; Taylor, L; Sherman, A; Nair, V; Kaiser, P; McGrew, M J

    2016-08-01

    Biosecurity and sustainability in poultry production requires reliable germplasm conservation. Germplasm conservation in poultry is more challenging in comparison to other livestock species. Embryo cryopreservation is not feasible for egg-laying animals, and chicken semen conservation has variable success for different chicken breeds. A potential solution is the cryopreservation of the committed diploid stem cell precursors to the gametes, the primordial germ cells ( PGCS: ). Primordial germ cells are the lineage-restricted cells found at early embryonic stages in birds and form the sperm and eggs. We demonstrate here, using flocks of partially inbred, lower-fertility, major histocompatibility complex- ( MHC-: ) restricted lines of chicken, that we can easily derive and cryopreserve a sufficient number of independent lines of male and female PGCs that would be sufficient to reconstitute a poultry breed. We demonstrate that germ-line transmission can be attained from these PGCs using a commercial layer line of chickens as a surrogate host. This research is a major step in developing and demonstrating that cryopreserved PGCs could be used for the biobanking of specialized flocks of birds used in research settings. The prospective application of this technology to poultry production will further increase sustainability to meet current and future production needs. © The Author 2016. Published by Oxford University Press on behalf of Poultry Science Association.

  7. Antibacterial and anti-breast cancer cell line activities of ...

    African Journals Online (AJOL)

    Purpose: To evaluate the activity of extracts of Sanghuangporus sp.1 fungus against pathogenic bacteria and a breast cancer cell line. Methods: The wild fruiting body and mycelium of Sanghuangporus sp.1 were extracted with water and ethanol by ultrasonication extraction. The activity of the extracts against pathogenic ...

  8. Characterization of newly established colorectal cancer cell lines

    Indian Academy of Sciences (India)

    We have established a series of 20 colorectal cancer cell lines and performed cytogenetic and RFLP analyses to show that the recurrent genetic abnormalities of chromosomes 1, 5, 17 and 18 associated with multistep tumorigenesis in colorectal cancer, and frequently detected as recurrent abnormalities in primary tumours, ...

  9. Apoptosis induction of epifriedelinol on human cervical cancer cell line

    African Journals Online (AJOL)

    Background: Present investigation evaluates the antitumor activity of epifriedelinol for the management of cervical cancer by inducing process of apoptosis. Methods: Human Cervical Cancer Cell Line, C33A and HeLa were selected for study and treated with epifriedelinol at a concentration of (50-1000 μg/ml). Cytotoxicity of ...

  10. Characterization of newly established colorectal cancer cell lines ...

    Indian Academy of Sciences (India)

    We have established a series of 20 colorectal cancer cell lines and performed cytogenetic and RFLP analyses to show that the recurrent genetic abnormalities of chromosomes 1, 5, 17 and 18 associated with multistep tumorigenesis in colorectal cancer, and frequently detected as recurrent abnormalities in primary tumours, ...

  11. AAVS1-Targeted Plasmid Integration in AAV Producer Cell Lines.

    Science.gov (United States)

    Luo, Yuxia; Frederick, Amy; Martin, John M; Scaria, Abraham; Cheng, Seng H; Armentano, Donna; Wadsworth, Samuel C; Vincent, Karen A

    2017-06-01

    Adeno-associated virus (AAV) producer cell lines are created via transfection of HeLaS3 cells with a single plasmid containing three components (the vector sequence, the AAV rep and cap genes, and a selectable marker gene). As this plasmid contains both the cis (Rep binding sites) and trans (Rep protein encoded by the rep gene) elements required for site-specific integration, it was predicted that plasmid integration might occur within the AAVS1 locus on human chromosome 19 (chr19). The objective of this study was to investigate whether integration in AAVS1 might be correlated with vector yield. Plasmid integration sites within several independent cell lines were assessed via Southern, fluorescence in situ hybridization (FISH) and PCR analyses. In the Southern analyses, the presence of fragments detected by both rep- and AAVS1-specific probes suggested that for several mid- and high-producing lines, plasmid DNA had integrated into the AAVS1 locus. Analysis with puroR and AAVS1-specific probes suggested that integration in AAVS1 was a more widespread phenomenon. High-producing AAV2-secreted alkaline phosphatase (SEAP) lines (masterwell 82 [MW82] and MW278) were evaluated via FISH using probes specific for the plasmid, AAVS1, and a chr19 marker. FISH analysis detected two plasmid integration sites in MW278 (neither in AAVS1), while a total of three sites were identified in MW82 (two in AAVS1). An inverse PCR assay confirmed integration within AAVS1 for several mid- and high-producing lines. In summary, the FISH, Southern, and PCR data provide evidence of site-specific integration of the plasmid within AAVS1 in several AAV producer cell lines. The data also suggest that integration in AAVS1 is a general phenomenon that is not necessarily restricted to high producers. The results also suggest that plasmid integration within the AAVS1 locus is not an absolute requirement for a high vector yield.

  12. Sphere-forming cell subpopulations with cancer stem cell properties in human hepatoma cell lines

    Directory of Open Access Journals (Sweden)

    Chen Lei

    2011-06-01

    Full Text Available Abstract Background Cancer stem cells (CSCs are regarded as the cause of tumor formation and recurrence. The isolation and identification of CSCs could help to develop novel therapeutic strategies specifically targeting CSCs. Methods Human hepatoma cell lines were plated in stem cell conditioned culture system allowed for sphere forming. To evaluate the stemness characteristics of spheres, the self-renewal, proliferation, chemoresistance, tumorigenicity of the PLC/PRF/5 sphere-forming cells, and the expression levels of stem cell related proteins in the PLC/PRF/5 sphere-forming cells were assessed, comparing with the parental cells. The stem cell RT-PCR array was performed to further explore the biological properties of liver CSCs. Results The PLC/PRF/5, MHCC97H and HepG2 cells could form clonal nonadherent 3-D spheres and be serially passaged. The PLC/PRF/5 sphere-forming cells possessed a key criteria that define CSCs: persistent self-renewal, extensive proliferation, drug resistance, overexpression of liver CSCs related proteins (Oct3/4, OV6, EpCAM, CD133 and CD44. Even 500 sphere-forming cells were able to form tumors in NOD/SCID mice, and the tumor initiating capability was not decreased when spheres were passaged. Besides, downstream proteins DTX1 and Ep300 of the CSL (CBF1 in humans, Suppressor of hairless in Drosophila and LAG1 in C. elegans -independent Notch signaling pathway were highly expressed in the spheres, and a gamma-secretase inhibitor MRK003 could significantly inhibit the sphere formation ability. Conclusions Nonadherent tumor spheres from hepatoma cell lines cultured in stem cell conditioned medium possess liver CSC properties, and the CSL-independent Notch signaling pathway may play a role in liver CSCs.

  13. Thymol inhibits cell migration and invasion by downregulating the ...

    African Journals Online (AJOL)

    The supernatants were separated by 10 % SDS-PAGE containing 1 %. (m/v) gelatin. The gels were visualized after staining with Coomassie blue and then photographed. Western blotting. Cell lysates from HT29 cells were extracted in. NP40 lysis buffer, separated by 10 % SDS-. PAGE gel and further transferred to PVDF.

  14. Effects of hypoxia on human cancer cell line chemosensitivity

    Science.gov (United States)

    2013-01-01

    Background Environment inside even a small tumor is characterized by total (anoxia) or partial oxygen deprivation, (hypoxia). It has been shown that radiotherapy and some conventional chemotherapies may be less effective in hypoxia, and therefore it is important to investigate how different drugs act in different microenvironments. In this study we perform a large screening of the effects of 19 clinically used or experimental chemotherapeutic drugs on five different cell lines in conditions of normoxia, hypoxia and anoxia. Methods A panel of 19 commercially available drugs: 5-fluorouracil, acriflavine, bortezomib, cisplatin, digitoxin, digoxin, docetaxel, doxorubicin, etoposide, gemcitabine, irinotecan, melphalan, mitomycin c, rapamycin, sorafenib, thalidomide, tirapazamine, topotecan and vincristine were tested for cytotoxic activity on the cancer cell lines A2780 (ovarian), ACHN (renal), MCF-7 (breast), H69 (SCLC) and U-937 (lymphoma). Parallel aliquots of the cells were grown at different oxygen pressures and after 72 hours of drug exposure viability was measured with the fluorometric microculture cytotoxicity assay (FMCA). Results Sorafenib, irinotecan and docetaxel were in general more effective in an oxygenated environment, while cisplatin, mitomycin c and tirapazamine were more effective in a low oxygen environment. Surprisingly, hypoxia in H69 and MCF-7 cells mostly rendered higher drug sensitivity. In contrast ACHN appeared more sensitive to hypoxia, giving slower proliferating cells, and consequently, was more resistant to most drugs. Conclusions A panel of standard cytotoxic agents was tested against five different human cancer cell lines cultivated at normoxic, hypoxic and anoxic conditions. Results show that impaired chemosensitivity is not universal, in contrast different cell lines behave different and some drugs appear even less effective in normoxia than hypoxia. PMID:23829203

  15. Crude subcellular fractionation of cultured mammalian cell lines

    Directory of Open Access Journals (Sweden)

    Holden Paul

    2009-12-01

    Full Text Available Abstract Background The expression and study of recombinant proteins in mammalian culture systems can be complicated during the cell lysis procedure by contaminating proteins from cellular compartments distinct from those within which the protein of interest resides and also by solubility issues that may arise from the use of a single lysis buffer. Partial subcellular fractionation using buffers of increasing stringency, rather than whole cell lysis is one way in which to avoid or reduce this contamination and ensure complete recovery of the target protein. Currently published protocols involve time consuming centrifugation steps which may require expensive equipment and commercially available kits can be prohibitively expensive when handling large or multiple samples. Findings We have established a protocol to sequentially extract proteins from cultured mammalian cells in fractions enriched for cytosolic, membrane bound organellar, nuclear and insoluble proteins. All of the buffers used can be made inexpensively and easily and the protocol requires no costly equipment. While the method was optimized for a specific cell type, we demonstrate that the protocol can be applied to a variety of commonly used cell lines and anticipate that it can be applied to any cell line via simple optimization of the primary extraction step. Conclusion We describe a protocol for the crude subcellular fractionation of cultured mammalian cells that is both straightforward and cost effective and may facilitate the more accurate study of recombinant proteins and the generation of purer preparations of said proteins from cell extracts.

  16. Glucocorticoid inhibition of cellular proliferation in rat hepatoma cell lines

    International Nuclear Information System (INIS)

    Cook, P.W.

    1987-01-01

    Glucocorticoids were shown to inhibit the growth rate of Fu5 rat hepatoma cells cultured in the presence or absence of serum and thus, induced a more stringent dependence on serum for growth in this cell line. Fu5 cells, made quiescent at low cell density by continuous exposure to glucocorticoid in the absence of serum, were induced with serum and insulin, which subsequently caused a rapid reinitiation of cellular proliferation. Analysis of total RNA isolated from hormone treated Fu5 cells undergoing serum/insulin induction of DNA synthesis revealed a sequential expression of cellular proto-oncogene products in the absence of any immediate changes in intracellular Ca ++ levels. Introduction of functional glucocorticoid receptor genes into both classes of dexamethasone resistant variants restored glucocorticoid responsiveness and suppression of cell growth. The BDS1 rat hepatoma cell line, an Fu5 derived subclone hypersensitive to the antiprofliferation effects of glucocorticoid, was observed to externalize a glucocorticoid suppressible mitogen (GSM) activity capable of mimicking EGF and insulin induced stimulation of [ 3 H]thymidine incorporation into serum starved, competant Balb/c 3T3 cells

  17. Differences in radiosensitivity between three HER2 overexpressing cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Steffen, Ann-Charlott; Tolmachev, Vladimir; Stenerloew, Bo [Uppsala University, Unit of Biomedical Radiation Sciences, Department of Oncology, Radiology and Clinical Immunology, Rudbeck Laboratory, Uppsala (Sweden); Goestring, Lovisa [Affibody AB, Bromma (Sweden); Palm, Stig [Sahlgrenska Academy at Goeteborg University, Department of Radiation Physics, Goeteborg (Sweden); Carlsson, Joergen [Uppsala University, Unit of Biomedical Radiation Sciences, Department of Oncology, Radiology and Clinical Immunology, Rudbeck Laboratory, Uppsala (Sweden); Rudbeck Laboratory, Biomedical Radiation Sciences, Uppsala (Sweden)

    2008-06-15

    HER2 is a potential target for radionuclide therapy, especially when HER2 overexpressing breast cancer cells are resistant to Herceptin {sup registered} treatment. Therefore, it is of interest to analyse whether HER2 overexpressing tumour cells have different inherent radiosensitivity. The radiosensitivity of three often used HER2 overexpressing cell lines, SKOV-3, SKBR-3 and BT-474, was analysed. The cells were exposed to conventional photon irradiation, low linear energy transfer (LET), to characterise their inherent radiosensitivity. The analysis was made with clonogenic survival and growth extrapolation assays. The cells were also exposed to alpha particles, high LET, from {sup 211}At decays using the HER2-binding affibody molecule {sup 211}At-(Z{sub HER2:4}){sub 2} as targeting agent. Assays for studies of internalisation of the affibody molecule were applied. SKOV-3 cells were most radioresistant, SKBR-3 cells were intermediate and BT-474 cells were most sensitive as measured with the clonogenic and growth extrapolation assays after photon irradiation. The HER2 dependent cellular uptake of {sup 211}At was qualitatively similar for all three cell lines. However, the sensitivity to the alpha particles from {sup 211}At differed; SKOV-3 was most resistant, SKBR-3 intermediate and BT-474 most sensitive. These differences were unexpected because it is assumed that all types of cells should have similar sensitivity to high-LET radiation. The sensitivity to alpha particle exposure correlated with internalisation of the affibody molecule and with size of the cell nucleus. There can be differences in radiosensitivity, which, if they also exist between patient breast cancer cells, are important to consider for both conventional radiotherapy and for HER2-targeted radionuclide therapy. (orig.)

  18. Radiosensitization of colorectal carcinoma cell lines by histone deacetylase inhibition

    International Nuclear Information System (INIS)

    Flatmark, Kjersti; Nome, Ragnhild V; Folkvord, Sigurd; Bratland, Åse; Rasmussen, Heidi; Ellefsen, Mali Strand; Fodstad, Øystein; Ree, Anne Hansen

    2006-01-01

    The tumor response to preoperative radiotherapy of locally advanced rectal cancer varies greatly, warranting the use of experimental models to assay the efficacy of molecular targeting agents in rectal cancer radiosensitization. Histone deacetylase (HDAC) inhibitors, agents that cause hyperacetylation of histone proteins and thereby remodeling of chromatin structure, may override cell cycle checkpoint responses to DNA damage and amplify radiation-induced tumor cell death. Human colorectal carcinoma cell lines were exposed to ionizing radiation and HDAC inhibitors, and cell cycle profiles and regulatory factors, as well as clonogenicity, were analyzed. In addition to G 2 /M phase arrest following irradiation, the cell lines displayed cell cycle responses typical for either intact or defective p53 function (the presence or absence, respectively, of radiation-induced expression of the cell cycle inhibitor p21 and subsequent accumulation of G 1 phase cells). In contrast, histone acetylation was associated with complete depletion of the G 1 population of cells with functional p53 but accumulation of both G 1 and G 2 /M populations of cells with defective p53. The cellular phenotypes upon HDAC inhibition were consistent with the observed repression of Polo-like kinase-1, a regulatory G 2 /M phase kinase. Following pre-treatment with HDAC inhibitors currently undergoing clinical investigation, the inhibitory effect of ionizing radiation on clonogenicity was significantly amplified. In these experimental models, HDAC inhibition sensitized the tumor cells to ionizing radiation, which is in accordance with the concept of increased probability of tumor cell death when chromatin structure is modified

  19. Differences in radiosensitivity between three HER2 overexpressing cell lines

    International Nuclear Information System (INIS)

    Steffen, Ann-Charlott; Tolmachev, Vladimir; Stenerloew, Bo; Goestring, Lovisa; Palm, Stig; Carlsson, Joergen

    2008-01-01

    HER2 is a potential target for radionuclide therapy, especially when HER2 overexpressing breast cancer cells are resistant to Herceptin registered treatment. Therefore, it is of interest to analyse whether HER2 overexpressing tumour cells have different inherent radiosensitivity. The radiosensitivity of three often used HER2 overexpressing cell lines, SKOV-3, SKBR-3 and BT-474, was analysed. The cells were exposed to conventional photon irradiation, low linear energy transfer (LET), to characterise their inherent radiosensitivity. The analysis was made with clonogenic survival and growth extrapolation assays. The cells were also exposed to alpha particles, high LET, from 211 At decays using the HER2-binding affibody molecule 211 At-(Z HER2:4 ) 2 as targeting agent. Assays for studies of internalisation of the affibody molecule were applied. SKOV-3 cells were most radioresistant, SKBR-3 cells were intermediate and BT-474 cells were most sensitive as measured with the clonogenic and growth extrapolation assays after photon irradiation. The HER2 dependent cellular uptake of 211 At was qualitatively similar for all three cell lines. However, the sensitivity to the alpha particles from 211 At differed; SKOV-3 was most resistant, SKBR-3 intermediate and BT-474 most sensitive. These differences were unexpected because it is assumed that all types of cells should have similar sensitivity to high-LET radiation. The sensitivity to alpha particle exposure correlated with internalisation of the affibody molecule and with size of the cell nucleus. There can be differences in radiosensitivity, which, if they also exist between patient breast cancer cells, are important to consider for both conventional radiotherapy and for HER2-targeted radionuclide therapy. (orig.)

  20. Plasmids and packaging cell lines for use in phage display

    Science.gov (United States)

    Bradbury, Andrew M.

    2012-07-24

    The invention relates to a novel phagemid display system for packaging phagemid DNA into phagemid particles which completely avoids the use of helper phage. The system of the invention incorporates the use of bacterial packaging cell lines which have been transformed with helper plasmids containing all required phage proteins but not the packaging signals. The absence of packaging signals in these helper plasmids prevents their DNA from being packaged in the bacterial cell, which provides a number of significant advantages over the use of both standard and modified helper phage. Packaged phagemids expressing a protein or peptide of interest, in fusion with a phage coat protein such as g3p, are generated simply by transfecting phagemid into the packaging cell line.

  1. Preferential radiosensitization of human prostatic carcinoma cells by mild hyperthermia

    International Nuclear Information System (INIS)

    Ryu, Samuel; Brown, Stephen L.; Kim, Sang-Hie; Khil, Mark S.; Kim, Jae Ho

    1996-01-01

    Purpose: Recent cell culture studies by us and others suggest that some human carcinoma cells are more sensitive to heat than are rodent cells following mild hyperthermia. In studying the cellular mechanism of enhanced thermosensitivity of human tumor cells to hyperthermia, prostatic carcinoma cells of human origin were found to be more sensitive to mild hyperthermia than other human cancer cells. The present study was designed to determine the magnitude of radiosensitization of human prostatic carcinoma cells by mild hyperthermia and to examine whether the thermal radiosensitization is related to the intrinsic thermosensitivity of cancer cells. Methods and Materials: Two human prostatic carcinoma cell lines (DU-145 and PC-3) and other carcinoma cells of human origin, in particular, colon (HT-29), breast (MCF-7), lung (A-549), and brain (U-251) were exposed to temperatures of 40-41 deg. C. Single acute dose rate radiation and fractionated radiation were combined with mild hyperthermia to determine thermal radiosensitization. The end point of the study was the colony-forming ability of single-plated cells. Results: DU-145 and PC-3 cells were found to be exceedingly thermosensitive to 41 deg. C for 24 h, relative to other cancer cell lines. Ninety percent of the prostatic cancer cells were killed by a 24 h heat exposure. Prostatic carcinoma cells exposed to a short duration of heating at 41 deg. C for 2 h resulted in a substantial enhancement of radiation-induced cytotoxicity. The thermal enhancement ratios (TERs) of single acute dose radiation following heat treatment 41 deg. C for 2 h were 2.0 in DU-145 cells and 1.4 in PC-3 cells. The TERs of fractionated irradiation combined with continuous heating at 40 deg. C were similarly in the range of 2.1 to 1.4 in prostate carcinoma cells. No significant radiosensitization was observed in MCF-7 and HT-29 cells under the same conditions. Conclusion: The present data suggest that a significant radiosensitization of

  2. Subcloning of three osteoblastic cell lines with distinct differentiation phenotypes from the mouse osteoblastic cell line KS-4.

    Science.gov (United States)

    Yamashita, T; Ishii, H; Shimoda, K; Sampath, T K; Katagiri, T; Wada, M; Osawa, T; Suda, T

    1996-11-01

    Three distinct osteoblastic cell lines (KS418, KS460, and KS483) were subcloned from the mouse osteoblastic KS-4 cells, which possessed the abilities not only to differentiate into mature osteoblasts, but also to support osteoclast differentiation in coculture with spleen cells. The order of the magnitude of the basal alkaline phosphatase (ALP) activity was KS483 > KS418 > KS460. KS483 cells were also more differentiated than KS418 and KS460 in terms of ALP activity and osteocalcin production, when cultured in growth medium containing 10% fetal bovine serum. In long-term culture, KS418 and KS483 apparently differentiated into mature osteoblasts and formed calcified nodules without addition of beta-glycerophosphate. Electron microscopic analysis demonstrated that calcification occurring in the nodules was initiated in the matrix vesicles as observed in bone formation in vivo. Nodule formation and mineral deposition occurred simultaneously in the presence of beta-glycerophosphate, but the former always preceded the latter without addition of beta-glycerophosphate. In contrast, KS460 cells did not show time-dependent increases of ALP activity, type I collagen expression and osteocalcin production, which were induced by treatment with recombinant osteogenic protein-1 (OP-1). The three cell lines similarly supported osteoclast differentiation in coculture with spleen