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Sample records for apoptosis increases chemosensitivity

  1. Fatty acid synthase regulates the chemosensitivity of breast cancer cells to cisplatin-induced apoptosis.

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    Al-Bahlani, Shadia; Al-Lawati, Hanaa; Al-Adawi, Moza; Al-Abri, Nadia; Al-Dhahli, Buthaina; Al-Adawi, Kawther

    2017-06-01

    Fatty acid synthase (FASN) is a key enzyme in fat biosynthesis that is over-expressed in advanced breast cancer stages. Cisplatin (CDDP) is a platinum-based drug used in the treatment of certain types of this disease. Although it was shown that FASN inhibition induced apoptosis by enhancing the cytotoxicity of certain drugs in breast cancer, its role in regulating the chemosensitivity of different types of breast cancer cells to CDDP-induced apoptosis is not established yet. Therefore, two different breast cancer cell lines; triple negative breast cancer (TNBC; MDA-MB-231) and triple positive breast cancer (TPBC; BT-474) cells were used to examine such role. We show that TNBC cells had naturally less fat content than TPBC cells. Subsequently, the fat content increased in both cells when treated with Palmitate rather than Oleate, whereas both fatty acids produced apoptotic ultra-structural effects and attenuated FASN expression. However, Oleate increased FASN expression in TPBC cells. CDDP decreased FASN expression and increased apoptosis in TNBC cells. These effects were further enhanced by combining CDDP with fatty acids. We also illustrate that the inhibition of FASN by either siRNA or exogenous inhibitor decreased CDDP-induced apoptosis in TPBC cells suggesting its role as an apoptotic factor, while an opposite finding was observed in TNBC cells when siRNA and fatty acids were used, suggesting its role as a survival factor. To our knowledge, we are the first to demonstrate a dual role of FASN in CDDP-induced apoptosis in breast cancer cells and how it can modulate their chemosensitivity.

  2. Upregulation of AIOLOS induces apoptosis and enhances etoposide chemosensitivity in Jurkat leukemia cells.

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    Zhuang, Yong; Lu, Yuanyuan; Li, Dong; Sun, Nianzheng; Ju, Xiuli

    2015-03-01

    T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive neoplastic disorder of immature hematopoietic precursors committed to T-cell lineage. T-ALL accounts for ~15% of pediatric ALL cases and is prone to early relapse. With new and improved treatment protocols, the prognosis of T-ALL has improved particularly in children; however, the outcome of relapsed T-ALL cases remains poor. The AIOLOS gene is necessary to control lymphocyte differentiation and may be a potential target of T-ALL therapy. In the present study, Jurkat cells were divided into three groups: untransfected (UT) control, lentiviral vector control (Lenti-Mock) and AIOLOS-overexpressing (Lenti-AIOLOS) groups. Lenti-AIOLOS Jurkat cells were constructed by lentiviral transduction; cell cycle analysis, apoptosis and cytotoxicity assays were then performed to evaluate the effects of AIOLOS on cell cycle distribution, apoptosis and cell chemosensitivity to etoposide of Jurkat cells in vitro. Moreover, the expression levels of genes associated with apoptosis and cell cycle were investigated by quantitative reverse transcription-polymerase chain reaction. Results showed that the percentage of Jurkat cells in the G0/G1 phase increased from 71.5 (UT) to 85.4% (Lenti-AIOLOS; PJurkat cells (21.93%) compared with this percentage in the Lenti-Mock (13.35%) or the UT group (13.30%; PJurkat cells induced cell apoptosis, arrested the cell cycle at the G0/G1 phase, and synergistically increased the sensitivity of Jurkat cells to etoposide by inhibiting NF-κB activity.

  3. Use of the microculture kinetic assay of apoptosis to determine chemosensitivities of leukemias.

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    Kravtsov, V D; Greer, J P; Whitlock, J A; Koury, M J

    1998-08-01

    Chemotherapeutic agents exert their antitumor effects by inducing apoptosis. The microculture kinetic (MiCK) assay provides an automated, continuous means of monitoring apoptosis in a cell population. We used the MiCK assay to determine the chemosensitivities of the human promyelocytic HL-60 and lymphoblastic CEM cell lines and leukemia cells freshly isolated from patients with acute nonlymphocytic (ANLL) or acute lymphocytic (ALL) leukemias. Continuous monitoring of apoptosis in the MiCK assay permits determination of the time to the maximum apoptosis (Tm) and its two components which are initiation time (Ti) and development time (Td). Duration of the three timing components of apoptosis varies from hours to days depending on the drug, drug concentration, and type of target cells. In the MiCK assay, the extent of apoptosis is reported in kinetic units of apoptosis. Kinetic units are determined by the slope of the curve created when optical density caused by cell blebbing is plotted as a function of time. Using the leukemia cell lines, we define the relationship between kinetic units determined by the MiCK assay and the percentage of morphologically apoptotic cells in the culture. Flow cytometry analysis of apoptosis in Annexin-V-fluorescein isothiocyanate-labeled preparations of HL-60 and CEM cells was also used to compare with data obtained by the MiCK assay. The feasibility of the MiCK assay of apoptosis as a chemosensitivity test was confirmed by its comparison with a 3H-thymidine incorporation assay. We show that samples from 10 ANLL and ALL patients patients tested for sensitivity to various doses of idarubicin (IDR), daunorubicin (DNR), or mitoxantrone (MTA) gave the same percentages of apoptotic cells when calculated by the MiCK assay as when determined by morphological analysis. The MiCK assay was used for dose-response analyses of the sensitivities to IDR, DNR, and MTA of leukemia cells from 4 other patients (2 ANLL and 2 ALL). The results from both cell

  4. Docosahexaenoic Acid Induces Oxidative DNA Damage and Apoptosis, and Enhances the Chemosensitivity of Cancer Cells

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    Eun Ah Song

    2016-08-01

    Full Text Available The human diet contains low amounts of ω-3 polyunsaturated fatty acids (PUFAs and high amounts of ω-6 PUFAs, which has been reported to contribute to the incidence of cancer. Epidemiological studies have shown that a high consumption of fish oil or ω-3 PUFAs reduced the risk of colon, pancreatic, and endometrial cancers. The ω-3 PUFA, docosahexaenoic acid (DHA, shows anticancer activity by inducing apoptosis of some human cancer cells without toxicity against normal cells. DHA induces oxidative stress and oxidative DNA adduct formation by depleting intracellular glutathione (GSH and decreasing the mitochondrial function of cancer cells. Oxidative DNA damage and DNA strand breaks activate DNA damage responses to repair the damaged DNA. However, excessive DNA damage beyond the capacity of the DNA repair processes may initiate apoptotic signaling pathways and cell cycle arrest in cancer cells. DHA shows a variable inhibitory effect on cancer cell growth depending on the cells’ molecular properties and degree of malignancy. It has been shown to affect DNA repair processes including DNA-dependent protein kinases and mismatch repair in cancer cells. Moreover, DHA enhanced the efficacy of anticancer drugs by increasing drug uptake and suppressing survival pathways in cancer cells. In this review, DHA-induced oxidative DNA damage, apoptotic signaling, and enhancement of chemosensitivity in cancer cells will be discussed based on recent studies.

  5. Docosahexaenoic Acid Induces Oxidative DNA Damage and Apoptosis, and Enhances the Chemosensitivity of Cancer Cells.

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    Song, Eun Ah; Kim, Hyeyoung

    2016-08-03

    The human diet contains low amounts of ω-3 polyunsaturated fatty acids (PUFAs) and high amounts of ω-6 PUFAs, which has been reported to contribute to the incidence of cancer. Epidemiological studies have shown that a high consumption of fish oil or ω-3 PUFAs reduced the risk of colon, pancreatic, and endometrial cancers. The ω-3 PUFA, docosahexaenoic acid (DHA), shows anticancer activity by inducing apoptosis of some human cancer cells without toxicity against normal cells. DHA induces oxidative stress and oxidative DNA adduct formation by depleting intracellular glutathione (GSH) and decreasing the mitochondrial function of cancer cells. Oxidative DNA damage and DNA strand breaks activate DNA damage responses to repair the damaged DNA. However, excessive DNA damage beyond the capacity of the DNA repair processes may initiate apoptotic signaling pathways and cell cycle arrest in cancer cells. DHA shows a variable inhibitory effect on cancer cell growth depending on the cells' molecular properties and degree of malignancy. It has been shown to affect DNA repair processes including DNA-dependent protein kinases and mismatch repair in cancer cells. Moreover, DHA enhanced the efficacy of anticancer drugs by increasing drug uptake and suppressing survival pathways in cancer cells. In this review, DHA-induced oxidative DNA damage, apoptotic signaling, and enhancement of chemosensitivity in cancer cells will be discussed based on recent studies.

  6. Chemosensitizing effects of carbon-based nanomaterials in cancer cells: enhanced apoptosis and inhibition of proliferation as underlying mechanisms

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    Erdmann, Kati; Ringel, Jessica; Rieger, Christiane; Huebner, Doreen; Wirth, Manfred P; Fuessel, Susanne; Hampel, Silke

    2014-01-01

    Recent studies have shown that carbon nanomaterials such as carbon nanofibres (CNFs) and multi-walled carbon nanotubes (CNTs) can exert antitumor activities themselves and sensitize cancer cells to conventional chemotherapeutics such as carboplatin and cisplatin. In the present study, the chemosensitizing effect of CNFs and CNTs on cancer cells of urological origin was investigated regarding the underlying mechanisms. Prostate cancer (DU-145, PC-3) and bladder cancer (EJ28) cells were treated with carbon nanomaterials (CNFs, CNTs) and chemotherapeutics (carboplatin, cisplatin) alone as well as in combination for 24 h. Forty-eight (EJ28) or 72 h (DU-145, PC-3) after the end of treatment the effects on cellular proliferation, clonogenic survival, cell death rate and cell cycle distribution were evaluated. Depending on the cell line, simultaneous administration of chemotherapeutics and carbon nanomaterials produced an additional inhibition of cellular proliferation and clonogenic survival of up to 77% and 98%, respectively, compared to the inhibitory effects of the chemotherapeutics alone. These strongly enhanced antiproliferative effects were accompanied by an elevated cell death rate, which was predominantly mediated via apoptosis and not by necrosis. The antitumor effects of combinations with CNTs were less pronounced than those with CNFs. The enhanced effects of the combinatory treatments on cellular function were mostly of additive to partly synergistic nature. Furthermore, cell cycle analysis demonstrated an arrest at the G2/M phase mediated by a monotreatment with chemotherapeutics. Following combinatory treatments, mostly less than or nearly additive increases of cell fractions in the G2/M phase could be observed. In conclusion, the pronounced chemosensitizing effects of CNFs and CNTs were mediated by an enhanced apoptosis and inhibition of proliferation. The combination of carbon-based nanomaterials and conventional chemotherapeutics represents a novel

  7. BRCA1 Expression is an Important Biomarker for Chemosensitivity: Suppression of BRCA1 Increases the Apoptosis via Up-regulation of p53 and p21 During Cisplatin Treatment in Ovarian Cancer Cells

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    Ikuo Konishi

    2006-01-01

    Full Text Available BRCA1 is a tumor suppressor which plays a crucial role in the repair of DNA double-strand breaks, and its abnormality is responsible for hereditary ovarian cancer syndrome. It has recently been reported that reduced expression of BRCA1 is also common in sporadic ovarian carcinoma via its promoter hypermethylation, and that ovarian carcinoma patients negative for BRCA1 expression showed favorable prognosis. To address if BRCA1 expression plays a role in the chemotherapeutic response, we analyzed the effect of BRCA1 suppression on the sensitivity to cisplatin and paclitaxel in ovarian cancer cells. Specific siRNA for BRCA1 gene was transfected into 3 ovarian cancer cell lines with various p53 status. Reduced expression of BRCA1 by transfection of BRCA1-siRNA resulted in a 5.3-fold increase in sensitivity to cisplatin in p53-wild A2780 cells, but not in p53-mutated A2780/CDDP and p53-deleted SKOV3 cells. Regarding the sensitivity to paclitaxel, BRCA1 suppression caused no significant changes in all the 3 cell lines. For ionizing radiation sensitivity, BRCA1 suppression also showed a significant higher sensitivity in A2780 cells. Growth curve and cell cycle analyses showed no signifi cant differences between BRCA1-siRNA-transfected A2780 cells and control cells. However, cisplatin treatment under suppression of BRCA1 showed a significantly increased apoptosis along with up-regulation of p53 and p21 in A2780 cells. Accordingly, reduced expression of BRCA1 enhances the cisplatin sensitivity and apoptosis via up-regulation of p53 and p21, but does not affect the paclitaxel sensitivity. Expression of BRCA1 might be an important biomarker for cisplatin resistance in ovarian carcinoma.

  8. Reactive oxygen species-mediated apoptosis contributes to chemosensitization effect of saikosaponins on cisplatin-induced cytotoxicity in cancer cells

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    He Fan

    2010-12-01

    Full Text Available Abstract Background Saikosaponin-a and -d, two naturally occurring compounds derived from Bupleurum radix, have been shown to exert anti-cancer activity in several cancer cell lines. However, the effect of combination of saikosaponins with chemotherapeutic drugs has never been addressed. Thus, we investigated whether these two saikosaponins have chemosensitization effect on cisplatin-induced cancer cell cytotoxicity. Methods Two cervical cancer cell lines, HeLa and Siha, an ovarian cancer cell line, SKOV3, and a non-small cell lung cancer cell line, A549, were treated with saikosaponins or cisplatin individually or in combination. Cell death was quantitatively detected by the release of lactate dehydrogenase (LDH using a cytotoxicity detection kit. Cellular ROS was analyzed by flow cytometry. Apoptosis was evaluated by AO/EB staining, flow cytometry after Anexin V and PI staining, and Western blot for caspase activation. ROS scavengers and caspase inhibitor were used to determine the roles of ROS and apoptosis in the effects of saikosaponins on cisplatin-induced cell death. Results Both saikosaponin-a and -d sensitized cancer cells to cisplatin-induced cell death in a dose-dependent manner, which was accompanied with induction of reactive oxygen species (ROS accumulation. The dead cells showed typical apoptotic morphologies. Both early apoptotic and late apoptotic cells detected by flow cytometry were increased in saikosaponins and cisplatin cotreated cells, accompanied by activation of the caspase pathway. The pan-caspase inhibitor z-VAD and ROS scanvengers butylated hydroxyanisole (BHA and N-acetyl-L-cysteine (NAC dramatically suppressed the potentiated cytotoxicity achieved by combination of saikosaponin-a or -d and cisplatin. Conclusions These results suggest that saikosaponins sensitize cancer cells to cisplatin through ROS-mediated apoptosis, and the combination of saikosaponins with cisplatin could be an effective therapeutic strategy.

  9. Knockdown of Eag1 Expression by RNA Interference Increases Chemosensitivity to Cisplatin in Ovarian Cancer Cells.

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    Hui, Chen; Lan, Zhang; Yue-li, Lin; Li-lin, Hong; Li-lin, Huang

    2015-12-01

    Ether á go-go 1 (Eag1) is frequently highly expressed in various malignant cancers and its excessive expression is correlated with poor prognosis in various cancers. However, the relationship of Eag1 expression with the clinical outcome of patients having ovarian cancer treated with cisplatin-based adjuvant chemotherapy is still unknown. In this study, we measured the expression of Eag1 in ovarian cancer and investigated the association between cisplatin chemosensitivity of ovarian cancer cells and Eag1 expression level. We demonstrate that decreased expression of Eag1 correlates with favorable prognosis in patients treated with cisplatin-based adjuvant chemotherapy and predicts higher cisplatin sensitivity in ovarian cancer cells. In vitro, knockdown of Eag1 by small interfering RNA facilitated the sensitivity of ovarian cancer cells (SKOV3 and TYK) to cisplatin-induced apoptosis via nuclear factor κ-light chain-enhancer of activated B cells (NF-κB) pathway. Furthermore, knockdown of Eag1 expression was associated with decreased expression of the P-glycoprotein without affecting multidrug resistance-associated protein 1 expression. Taken together, Eag1 may serve as a potential indicator to predict Eag1 chemosensitivity, and silencing Eag1 may represent a potential therapeutic strategy for ovarian cancer. © The Author(s) 2015.

  10. Salinomycin increases chemosensitivity to the effects of doxorubicin in soft tissue sarcomas

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    Liffers, Sven-T; Tischoff, Iris; Mirmohammadsadegh, Alireza; Tilkorn, Daniel J; Stricker, Ingo; Junge, Christoph Günter; Al-Benna, Sammy; Vogt, Markus; Verdoodt, Berlinda; Steinau, Hans-U; Tannapfel, Andrea

    2013-01-01

    Chemotherapy for soft tissue sarcomas remains unsatisfactory due to their low chemosensitivity. Even the first line chemotherapeutic agent doxorubicin only yields a response rate of 18-29%. The antibiotic salinomycin, a potassium ionophore, has recently been shown to be a potent compound to deplete chemoresistant cells like cancer stem like cells (CSC) in adenocarcinomas. Here, we evaluated the effect of salinomycin on sarcoma cell lines, whereby salinomycin mono- and combination treatment with doxorubicin regimens were analyzed. To evaluate the effect of salinomycin on fibrosarcoma, rhabdomyosarcoma and liposarcoma cell lines, cells were drug exposed in single and combined treatments, respectively. The effects of the corresponding treatments were monitored by cell viability assays, cell cycle analysis, caspase 3/7 and 9 activity assays. Further we analyzed NF-κB activity; p53, p21 and PUMA transcription levels, together with p53 expression and serine 15 phosphorylation. The combination of salinomycin with doxorubicin enhanced caspase activation and increased the sub-G1 fraction. The combined treatment yielded higher NF-κB activity, and p53, p21 and PUMA transcription, whereas the salinomycin monotreatment did not cause any significant changes. Salinomycin increases the chemosensitivity of sarcoma cell lines - even at sub-lethal concentrations - to the cytostatic drug doxorubicin. These findings support a strategy to decrease the doxorubicin concentration in combination with salinomycin in order to reduce toxic side effects

  11. A synthetic cryptochrome inhibitor induces anti-proliferative effects and increases chemosensitivity in human breast cancer cells

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    Chun, Sung Kook [Department of Brain & Cognitive Sciences, Daegu-Gyeongbuk Institute of Science & Technology, Daegu, 711-873 (Korea, Republic of); Department of Biological Sciences, Seoul National University, Seoul, 151-747 (Korea, Republic of); Department of Brain & Cognitive Sciences, Seoul National University, Seoul, 151-747 (Korea, Republic of); Chung, Sooyoung [Department of Biological Sciences, Seoul National University, Seoul, 151-747 (Korea, Republic of); Department of Biomedical Sciences, College of Medicine, Korea University, Seoul, 136-705 (Korea, Republic of); Kim, Hee-Dae [Department of Biological Sciences, Seoul National University, Seoul, 151-747 (Korea, Republic of); Lee, Ju Hyung [Department of Systems Biology, Yonsei University College of Life Science and Biotechnology, Seoul 120-749 (Korea, Republic of); Jang, Jaebong [College of Pharmacy, Seoul National University, Seoul, 151-742 (Korea, Republic of); Kim, Jeongah; Kim, Doyeon [Department of Brain & Cognitive Sciences, Daegu-Gyeongbuk Institute of Science & Technology, Daegu, 711-873 (Korea, Republic of); Department of Biological Sciences, Seoul National University, Seoul, 151-747 (Korea, Republic of); Department of Brain & Cognitive Sciences, Seoul National University, Seoul, 151-747 (Korea, Republic of); Son, Gi Hoon [Department of Biomedical Sciences, College of Medicine, Korea University, Seoul, 136-705 (Korea, Republic of); Oh, Young J. [Department of Systems Biology, Yonsei University College of Life Science and Biotechnology, Seoul 120-749 (Korea, Republic of); Suh, Young-Ger [College of Pharmacy, Seoul National University, Seoul, 151-742 (Korea, Republic of); Lee, Cheol Soon [Gachon Clinical Trials Center, Gachon University, Incheon, 417-842 (Korea, Republic of); and others

    2015-11-13

    Disruption of circadian rhythm is a major cause of breast cancer in humans. Cryptochrome (CRY), a circadian transcription factor, is a risk factor for initiation of breast cancer, and it is differentially expressed between normal and breast cancer tissues. Here, we evaluated the anti-proliferative and pro-apoptotic activity of KS15, a recently discovered small-molecule inhibitor of CRY, in human breast cancer cells. First, we investigated whether KS15 treatment could promote E-box-mediated transcription by inhibiting the activity of CRY in MCF-7 human breast cancer cells. Protein and mRNA levels of regulators of cell cycle and apoptosis, as well as core clock genes, were differentially modulated in response to KS15. Next, we investigated whether KS15 could inhibit proliferation and increase sensitivity to anti-tumor drugs in MCF-7 cells. We found that KS15 decreased the speed of cell growth and increased the chemosensitivity of MCF-7 cells to doxorubicin and tamoxifen, but had no effect on MCF-10A cells. These findings suggested that pharmacological inhibition of CRY by KS15 exerts an anti-proliferative effect and increases sensitivity to anti-tumor drugs in a specific type of breast cancer. - Highlights: • Cryptochrome inhibitor (KS15) has anti-tumor activity to human breast cancer cells. • KS15 induces differential changes in cell cycle regulators and pro-apoptotic genes. • KS15 inhibits MCF-7 cell growth and enhances susceptibility to anti-tumor drugs.

  12. A synthetic cryptochrome inhibitor induces anti-proliferative effects and increases chemosensitivity in human breast cancer cells

    International Nuclear Information System (INIS)

    Chun, Sung Kook; Chung, Sooyoung; Kim, Hee-Dae; Lee, Ju Hyung; Jang, Jaebong; Kim, Jeongah; Kim, Doyeon; Son, Gi Hoon; Oh, Young J.; Suh, Young-Ger; Lee, Cheol Soon

    2015-01-01

    Disruption of circadian rhythm is a major cause of breast cancer in humans. Cryptochrome (CRY), a circadian transcription factor, is a risk factor for initiation of breast cancer, and it is differentially expressed between normal and breast cancer tissues. Here, we evaluated the anti-proliferative and pro-apoptotic activity of KS15, a recently discovered small-molecule inhibitor of CRY, in human breast cancer cells. First, we investigated whether KS15 treatment could promote E-box-mediated transcription by inhibiting the activity of CRY in MCF-7 human breast cancer cells. Protein and mRNA levels of regulators of cell cycle and apoptosis, as well as core clock genes, were differentially modulated in response to KS15. Next, we investigated whether KS15 could inhibit proliferation and increase sensitivity to anti-tumor drugs in MCF-7 cells. We found that KS15 decreased the speed of cell growth and increased the chemosensitivity of MCF-7 cells to doxorubicin and tamoxifen, but had no effect on MCF-10A cells. These findings suggested that pharmacological inhibition of CRY by KS15 exerts an anti-proliferative effect and increases sensitivity to anti-tumor drugs in a specific type of breast cancer. - Highlights: • Cryptochrome inhibitor (KS15) has anti-tumor activity to human breast cancer cells. • KS15 induces differential changes in cell cycle regulators and pro-apoptotic genes. • KS15 inhibits MCF-7 cell growth and enhances susceptibility to anti-tumor drugs.

  13. Altered features and increased chemosensitivity of human breast cancer cells mediated by adipose tissue-derived mesenchymal stromal cells.

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    Kucerova, Lucia; Skolekova, Svetlana; Matuskova, Miroslava; Bohac, Martin; Kozovska, Zuzana

    2013-11-09

    Mesenchymal stromal cells (MSCs) represent heterogeneous cell population suitable for cell therapies in regenerative medicine. MSCs can also substantially affect tumor biology due to their ability to be recruited to the tumor stroma and interact with malignant cells via direct contacts and paracrine signaling. The aim of our study was to characterize molecular changes dictated by adipose tissue-derived mesenchymal stromal cells (AT-MSCs) and the effects on drug responses in human breast cancer cells SKBR3. The tumor cells were either directly cocultured with AT-MSCs or exposed to MSCs-conditioned medium (MSC-CM). Changes in cell biology were evaluated by kinetic live cell imaging, fluorescent microscopy, scratch wound assay, expression analysis, cytokine secretion profiling, ATP-based viability and apoptosis assays. The efficiency of cytotoxic treatment in the presence of AT-MSCs or MSCs-CM was analyzed. The AT-MSCs altered tumor cell morphology, induced epithelial-to-mesenchymal transition, increased mammosphere formation, cell confluence and migration of SKBR3. These features were attributed to molecular changes induced by MSCs-secreted cytokines and chemokines in breast cancer cells. AT-MSCs significantly inhibited the proliferation of SKBR3 cells in direct cocultures which was shown to be dependent on the SDF-1α/CXCR4 signaling axis. MSC-CM-exposed SKBR3 or SKBR3 in direct coculture with AT-MSCs exhibited increased chemosensitivity and induction of apoptosis in response to doxorubicin and 5-fluorouracil. Our work further highlights the multi-level nature of tumor-stromal cell interplay and demonstrates the capability of AT-MSCs and MSC-secreted factors to alter the anti-tumor drug responses.

  14. Altered features and increased chemosensitivity of human breast cancer cells mediated by adipose tissue-derived mesenchymal stromal cells

    International Nuclear Information System (INIS)

    Kucerova, Lucia; Skolekova, Svetlana; Matuskova, Miroslava; Bohac, Martin; Kozovska, Zuzana

    2013-01-01

    Mesenchymal stromal cells (MSCs) represent heterogeneous cell population suitable for cell therapies in regenerative medicine. MSCs can also substantially affect tumor biology due to their ability to be recruited to the tumor stroma and interact with malignant cells via direct contacts and paracrine signaling. The aim of our study was to characterize molecular changes dictated by adipose tissue-derived mesenchymal stromal cells (AT-MSCs) and the effects on drug responses in human breast cancer cells SKBR3. The tumor cells were either directly cocultured with AT-MSCs or exposed to MSCs-conditioned medium (MSC-CM). Changes in cell biology were evaluated by kinetic live cell imaging, fluorescent microscopy, scratch wound assay, expression analysis, cytokine secretion profiling, ATP-based viability and apoptosis assays. The efficiency of cytotoxic treatment in the presence of AT-MSCs or MSCs-CM was analyzed. The AT-MSCs altered tumor cell morphology, induced epithelial-to-mesenchymal transition, increased mammosphere formation, cell confluence and migration of SKBR3. These features were attributed to molecular changes induced by MSCs-secreted cytokines and chemokines in breast cancer cells. AT-MSCs significantly inhibited the proliferation of SKBR3 cells in direct cocultures which was shown to be dependent on the SDF-1α/CXCR4 signaling axis. MSC-CM-exposed SKBR3 or SKBR3 in direct coculture with AT-MSCs exhibited increased chemosensitivity and induction of apoptosis in response to doxorubicin and 5-fluorouracil. Our work further highlights the multi-level nature of tumor-stromal cell interplay and demonstrates the capability of AT-MSCs and MSC-secreted factors to alter the anti-tumor drug responses

  15. Growth Factor Receptors and Apoptosis Regulators: Signaling Pathways, Prognosis, Chemosensitivity and Treatment Outcomes of Breast Cancer

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    Siddik Sarkar

    2009-01-01

    Full Text Available Biomarkers of breast cancer are necessary for prognosis and prediction to chemotherapy. Prognostic biomarkers provide information regarding outcome irrespective of therapy, while predictive biomarkers provide information regarding response to therapy. Candidate prognostic biomarkers for breast cancers are growth factor receptors, steroid receptors, Ki-67, cyclins, urokinase plasminogen activator, p53, p21, pro- and anti-apoptotic factors, BRCA1 and BRCA2. But currently, the predictive markers are Estrogen and Progesterone receptors responding to endocrine therapy, and HER-2 responding to herceptin. But there are numerous breast cancer cases, where tamoxifen is ineffective even after estrogen receptor positivity. This lead to search of new prognostic and predictive markers and the number of potential markers is constantly increasing due to proteomics and genomics studies. However, most biomarkers individually have poor sensitivity or specificity, or other clinical value. It can be resolved by studying various biomarkers simultaneously, which will help in better prognosis and increasing sensitivity for chemotherapeutic agents. This review is focusing on growth factor receptors, apoptosis markers, signaling cascades, and their correlation with other associated biomarkers in breast cancers. As our knowledge regarding molecular biomarkers for breast cancer increases, prognostic indices will be developed that combine the predictive power of individual molecular biomarkers with specific clinical and pathologic factors. Rigorous comparison of these existing as well as emerging markers with current treatment selection is likely to see an escalation in an era of personalized medicines to ensure the breast cancer patients receive optimal treatment. This will also solve the treatment modalities and complications related to chemotherapeutic regimens.

  16. Endometrial carcinoma in vitro chemosensitivity testing of single and combination chemotherapy regimens using the novel microculture kinetic apoptosis assay: implications for endometrial cancer treatment.

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    Ballard, Karen S; Homesley, Howard D; Hodson, Charles; Presant, Cary A; Rutledge, James; Hallquist, Allan; Perree, Mathieu

    2010-03-01

    The in vitro microculture kinetic (MiCK) apoptosis assay has been used to predict single or combination chemotherapy response in leukemia patients. This feasibility study addressed MiCK in endometrial cancer specimens. Endometrial cancer specimens from total abdominal hysterectomies were processed at a central laboratory. Single cell suspensions of viable endometrial cancer cells were plated in individual wells. Single and combination regimens were tested: combinations of doxorubicin, cisplatin, and paclitaxel and carboplatin and paclitaxel (Gynecologic Oncology Group [GOG] 209 endometrial cancer phase III trial arms) as well as single agent testing with paclitaxel, carboplatin, doxorubicin, cisplatin, ifosfamide, and vincristine (active agents in GOG trials). Apoptosis was measured continuously over 48 hours. Fifteen of nineteen patients had successful assays. The highest mean chemo sensitivity was noted in the combination of cisplatin, doxorubicin, and paclitaxel with lower mean chemosensitivity for carboplatin and paclitaxel. Combination chemotherapy had higher chemosensitivity than single drug chemotherapy. However, in 25% of patients a single drug had higher chemosensitivity than combination chemotherapy. As single agents, ifosfamide, cisplatin, and paclitaxel had the highest kinetic unit values. Using a panel of agents simulating clinical dose regimens, the MiCK assay was feasible in evaluating in vitro chemosensitivity of endometrial cancer. MiCK assay results correlated with GOG clinical trial results. However, 25% of patients might be best treated with single agent chemotherapy selected by MiCK. Ifosfamide, cisplatin, and paclitaxel appear to have high activity as single agents. MiCK may be useful in future new drug testing and individualizing endometrial cancer patient's chemotherapy management.

  17. Mus81 knockdown improves chemosensitivity of hepatocellular carcinoma cells by inducing S-phase arrest and promoting apoptosis through CHK1 pathway

    International Nuclear Information System (INIS)

    Wu, Fan; Chen, Wei-Jia; Yan, Lun; Tan, Guo-Qian; Li, Wei-Tao; Zhu, Xuan-Jin; Ge, Xiao-Chuan; Liu, Jian-Wei; Wang, Bai-Lin

    2015-01-01

    As a critical endonuclease in DNA repair, Mus81 is traditionally regarded as a tumor suppressor, but recently correlated with the sensitivity of mitomycin C and 5-fluorouracil in colon cancer and breast cancer cells. However, its role in chemosensitivity of other human malignancies still remains unknown. This study therefore aims to investigate the effects of Mus81 knockdown on the chemosensitivity of hepatocellular carcinoma (HCC), a usually chemorefractory tumor, and explore the underlying mechanisms. Mus81 expression in HepG2 and Bel-7402 HCC cell lines was depleted by lentivirus-mediated short hairpin RNA and the elevated sensitivity of these Mus81-inhibited HCC cells to therapeutic agents, especially to epirubicin (EPI), was evidenced by MTT assay and an HCC chemotherapy mouse model. Flow cytometric analysis also showed that Mus81 knockdown lead to an obvious S-phase arrest and an elevated apoptosis in EPI-treated HepG2 and Bel-7402 cells, which could be rescued by CHK1 inhibition. The activation of CHK1/CDC25A/CDK2 pathway was also demonstrated in Mus81-inhibited HepG2 cells and xenograft mouse tumors under EPI treatment. Meanwhile, the apoptosis of HepG2 cells in response to EPI was remarkably promoted by Mus81 knockdown through activating p53/Bax/Caspase-3 pathway under the controlling of CHK1. In addition, CHK2 inhibition slightly raised CHK1 activity, thereby enhancing the S-phase arrest and apoptosis induced by EPI in Mus81-suppressed HCC cells. In conclusion, Mus81 knockdown improves the chemosensitivity of HCC cells by inducing S-phase arrest and promoting apoptosis through CHK1 pathway, suggesting Mus81 as a novel therapeutic target for HCC

  18. Growth inhibition and chemosensitization of exogenous nitric oxide released from NONOates in glioma cells in vitro.

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    Weyerbrock, Astrid; Baumer, Brunhilde; Papazoglou, Anna

    2009-01-01

    Exogenous nitric oxide (NO) from NO donors has cytotoxic, chemosensitizing, and radiosensitizing effects, and increases vascular permeability and blood flow in tumors. Yet little is known about whether these cytotoxic and chemosensitizing effects can be observed in glioma cells at doses that alter tumor physiological characteristics in vivo and whether these effects are tumor selective. The effect of NO released from proline NONOate, diethylamine NONOate, spermine NONOate, and sodium nitrite on cell proliferation, apoptosis, and chemosensitivity to carboplatin of cultured glioma cells was studied in C6, U87 glioma cells, human glioblastoma cells, and human astrocytes and fibroblasts. Although proline NONOate failed to induce cell death, the other NO donors induced growth arrest when present in high concentrations (10(-2) M) in all cell lines. Chemosensitization was observed after concomitant incubation with spermine NONOate and carboplatin in C6 and human glioblastoma cells. There is strong evidence that cell death occurs primarily by necrosis and to a lesser degree by apoptosis. The NO doses, which altered tumor physiology in vivo, were not cytotoxic, indicating that NO alters vascular permeability and cell viability in vivo by different mechanisms. The authors found that NO-generating agents at high concentrations are potent growth inhibitors and might also be useful as chemosensitizers in glioma cells. These data corroborate the theory that the use of NOgenerating agents may play a role in the multimodal treatment of malignant gliomas but that the NO release must be targeted more specifically to tumor cells to improve selectivity and efficacy.

  19. Overexpression of human sperm protein 17 increases migration and decreases the chemosensitivity of human epithelial ovarian cancer cells

    International Nuclear Information System (INIS)

    Li, Fang-qiu; Han, Yan-ling; Liu, Qun; Wu, Bo; Huang, Wen-bin; Zeng, Su-yun

    2009-01-01

    Most deaths from ovarian cancer are due to metastases that are resistant to conventional therapies. But the factors that regulate the metastatic process and chemoresistance of ovarian cancer are poorly understood. In the current study, we investigated the aberrant expression of human sperm protein 17 (HSp17) in human epithelial ovarian cancer cells and tried to analyze its influences on the cell behaviors like migration and chemoresistance. Immunohistochemistry and immunocytochemistry were used to identify HSp17 in paraffin embedded ovarian malignant tumor specimens and peritoneal metastatic malignant cells. Then we examined the effect of HSp17 overexpression on the proliferation, migration, and chemoresistance of ovarian cancer cells to carboplatin and cisplatin in a human ovarian carcinoma cell line, HO8910. We found that HSp17 was aberrantly expressed in 43% (30/70) of the patients with primary epithelial ovarian carcinomas, and in all of the metastatic cancer cells of ascites from 8 patients. The Sp17 expression was also detected in the metastatic lesions the same as in ovarian lesions. None of the 7 non-epithelial tumors primarily developed in the ovaries was immunopositive for HSp17. Overexpression of HSp17 increased the migration but decreased the chemosensitivity of ovarian carcinoma cells to carboplatin and cisplatin. HSp17 is aberrantly expressed in a significant proportion of epithelial ovarian carcinomas. Our results strongly suggest that HSp17 plays a role in metastatic disease and resistance of epithelial ovarian carcinoma to chemotherapy

  20. RNAi-mediated silencing of CD147 inhibits tumor cell proliferation, invasion and increases chemosensitivity to cisplatin in SGC7901 cells in vitro

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    Zhu Chan

    2010-06-01

    Full Text Available Abstract Background CD147 is a widely distributed cell surface glycoprotein that belongs to the Ig superfamily. CD147 has been implicated in numerous physiological and pathological activities. Enriched on the surface of many tumor cells, CD147 promotes tumor growth, invasion, metastasis and angiogenesis and confers resistance to some chemotherapeutic drugs. In this study, we investigated the possible role of CD147 in the progression of gastric cancer. Methods Short hairpin RNA (shRNA expressing vectors targeting CD147 were constructed and transfected into human gastric cancer cells SGC7901 and CD147 expression was monitored by quantitative realtime RT-PCR and Western blot. Cell proliferation, the activities of MMP-2 and MMP-9, the invasive potential and chemosensitivity to cisplatin of SGC7901 cells were determined by MTT, gelatin zymography, Transwell invasion assay and MTT, respectively. Results Down-regulation of CD147 by RNAi approach led to decreased cell proliferation, MMP-2 and MMP-9 activities and invasive potential of SGC7901 cells as well as increased chemosensitivity to cisplatin. Conclusion CD147 involves in proliferation, invasion and chemosensitivity of human gastric cancer cell line SGC7901, indicating that CD147 may be a promising therapeutic target for gastric cancer.

  1. Apollon modulates chemosensitivity in human esophageal squamous cell carcinoma

    Science.gov (United States)

    Weng, Shuqiang; Liu, Xijun; Rao, Benqiang; Gu, Jianxin; Chen, She; Wang, Qun; Shen, Xizhong; Xue, Ruyi; Dong, Ling

    2014-01-01

    Patients with esophageal squamous cell carcinoma (ESCC) are often diagnosed with advanced diseases that respond poorly to chemotherapy. Here we reported that Apollon, a membrane-associated inhibitor of apoptosis protein, was overexpressed in ESCC cell lines and clinical ESCC tissues, and Apollon overexpression clinically correlated with poor response to chemotherapy (P = 0.001), and short overall survival (P = 0.021). Apollon knockdown increased cisplatin/docetaxel-induced apoptosis, mitochondrial dysfunction and cytochrome c release in two ESCC cell lines. Apollon knockdown potentiated cisplatin/docetaxel-induced long-term cell growth inhibition, and enhanced chemosensitivity of ESCC cells to cisplatin/docetaxel in xenograft tumor models. Apollon knockdown also enhanced cisplatin/docetaxel-induced activation of caspase-8 (extrinsic pathway) and caspase-9 (intrinsic pathway) in ESCC cells and xenograft tumor models. Mechanism studies revealed that the effect of Apollon on chemosensitivity is mainly mediated by Smac. Apollon expression strongly and negatively correlated with Smac expression in clinical ESCC tissues (P = 0.001). Apollon targeted Smac for degradation in ESCC cells. The effect of Apollon on chemosensitivity was reversed by Smac knockdown in ESCC cells. Taken together, our data show association of Apollon expression with chemotherapeutic response in ESCC, and provide a strong rationale for combining Apollon antagonism with chemotherapy to treat ESCC. PMID:25216531

  2. Expression of the apoptosis-related genes BCL-2 and BAD in human breast carcinoma and their associated relationship with chemosensitivity

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    Fan Yuan-ming

    2010-08-01

    Full Text Available Abstract Objective To evaluate the expression of BCL-2 and BAD genes in tissues of breast carcinoma and investigate the relationship between the expression of BCL-2 and BAD in breast cancer cells with chemosensitivity. Methods Immunohistochemical technique was used to detect the expression of BCL-2, BAD in 10 normal breast tissues, 10 breast fibroadenoma tissues, 40 youth human breast carcinoma tissues, 40 menopause human breast carcinoma tissues. And to detect the expression of ER, PR in 80 human breast carcinoma tissues. 20 Surgical samples of breast cancer, diagnosed by pathology, were obtained from The First Affiliated Hospital of Chongqing Medical University. The cancer sample cells were cultured separately in the incubator at 37°C, 5% CO2 in vitro. The rate of inhibition of cancer cells in 4 kinds of anticancer drugs-- Epirubicin Adriamycin (EADM,5-Fluorouracil (5-Fu, Navelbine(NVB and Diaminedichloroplatinum (DDP, were assayed by MTT method. Results The expression of BCL-2, BAD genes in young human breast carcinoma tissues were lower than that in menopause human breast carcinoma tissues (P . There was a negative correlation between the positive expression rate of BCL-2 and histologic grade or the lymph node metastasis (P . There was a positive correlation between the expression rates of BCL-2 and of ER, PR (P . The expression of BAD had no relationship with the expression of ER, PR, histologic grade and the lymph node metastasis(P = NS. Sensitivity rates of 20 breast cancer cells in 0.1 × PPC within 48 h in vitro were 30% EADM,20% 5-Fu,45% NVB and 25% DDP. Respectively, the rate of inhibition of EADM,5- Fu, NVB and DDP were significantly higher in the BCL-2 negative cancer cells than in the BCL-2 positive cancer cells. A negative correlation was found between expression of BCL-2 and chemosensitivity for all the 4 anticancer drugs. The inhibition rates of EADM and NVB were significantly lower in the BAD negative cancer cells than in the

  3. MicroRNA-203 Is a Prognostic Indicator in Bladder Cancer and Enhances Chemosensitivity to Cisplatin via Apoptosis by Targeting Bcl-w and Survivin.

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    Xin Zhang

    Full Text Available Resistance to cisplatin-based chemotherapy is a major cause of treatment failure in advanced bladder cancer (BC patients. There is increasing evidence that microRNAs are involved in the development and progression of BC. However, little is known about the function of microRNAs in predicting the effect of adjuvant chemotherapy on BC survival and regulating response to cisplatin. To address this issue, we employed RT-qPCR to evaluate the clinical significance of miR-203 expression in 108 tissues of BC patients receiving cisplatin-based adjuvant chemotherapy, and performed in vitro studies to explore chemotherapeutic sensitivity to cisplatin in miR-203 overexpressing BC cells. We found miR-203 levels were significantly lower in BC progression group than non-progression group (P<0.001. ROC curve analysis illustrated miR-203 could significantly distinguish progressed patients from those without progression (P<0.001, yielding an area under the ROC curve of 0.839 (95% CI, 0.756-0.903. Moreover, low miR-203 expression correlated with shortened progression free survival (PFS and overall survival (OS of BC patients, and was an independent prognostic factor. Overexpression of miR-203 in 5637 and T24 BC cells could decrease cell viability, enhance cisplatin cytotoxicity, and promote apoptosis. Western blotting and luciferase reporter assay showed Bcl-w and Survivin were direct downstream targets of miR-203. There was also a significant inverse association between miR-203 and Bcl-w or Survivin expression in BC tissues (r = -0.781, -0.740, both P<0.001. In conclusion, decreased miR-203 predicts progression and poor prognosis for BC patients treated with cisplatin-based chemotherapy while miR-203 overexpression can enhance cisplatin sensitization by promoting apoptosis via directly targeting Bcl-w and Survivin.

  4. In Vitro Chemosensitization of Plasmodium falciparum to Antimalarials by Verapamil and Probenecid▿

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    Masseno, Victor; Muriithi, Steven; Nzila, Alexis

    2009-01-01

    We tested the effect of probenecid and verapamil in chemosensitizing Plasmodium falciparum to 14 antimalarials using the multidrug-resistant strain V1S and the drug-sensitive 3D7. Verapamil chemosensitizes V1S to quinine and chloroquine. Interestingly, probenecid profoundly chemosensitizes V1S to piperaquine. Thus, probenecid could be used to increase piperaquine efficacy in vivo.

  5. In vitro chemosensitization of Plasmodium falciparum to antimalarials by verapamil and probenecid.

    Science.gov (United States)

    Masseno, Victor; Muriithi, Steven; Nzila, Alexis

    2009-07-01

    We tested the effect of probenecid and verapamil in chemosensitizing Plasmodium falciparum to 14 antimalarials using the multidrug-resistant strain V1S and the drug-sensitive 3D7. Verapamil chemosensitizes V1S to quinine and chloroquine. Interestingly, probenecid profoundly chemosensitizes V1S to piperaquine. Thus, probenecid could be used to increase piperaquine efficacy in vivo.

  6. Spaceflight alters microtubules and increases apoptosis in human lymphocytes (Jurkat)

    Science.gov (United States)

    Lewis, M. L.; Reynolds, J. L.; Cubano, L. A.; Hatton, J. P.; Lawless, B. D.; Piepmeier, E. H.

    1998-01-01

    Alteration in cytoskeletal organization appears to underlie mechanisms of gravity sensitivity in space-flown cells. Human T lymphoblastoid cells (Jurkat) were flown on the Space Shuttle to test the hypothesis that growth responsiveness is associated with microtubule anomalies and mediated by apoptosis. Cell growth was stimulated in microgravity by increasing serum concentration. After 4 and 48 h, cells filtered from medium were fixed with formalin. Post-flight, confocal microscopy revealed diffuse, shortened microtubules extending from poorly defined microtubule organizing centers (MTOCs). In comparable ground controls, discrete microtubule filaments radiated from organized MTOCs and branched toward the cell membrane. At 4 h, 30% of flown, compared to 17% of ground, cells showed DNA condensation characteristic of apoptosis. Time-dependent increase of the apoptosis-associated Fas/ APO-1 protein in static flown, but not the in-flight 1 g centrifuged or ground controls, confirmed microgravity-associated apoptosis. By 48 h, ground cultures had increased by 40%. Flown populations did not increase, though some cells were cycling and actively metabolizing glucose. We conclude that cytoskeletal alteration, growth retardation, and metabolic changes in space-flown lymphocytes are concomitant with increased apoptosis and time-dependent elevation of Fas/APO-1 protein. We suggest that reduced growth response in lymphocytes during spaceflight is linked to apoptosis.

  7. Syringic acid from Tamarix aucheriana possesses antimitogenic and chemo-sensitizing activities in human colorectal cancer cells.

    Science.gov (United States)

    Abaza, Mohamed-Salah; Al-Attiyah, Raja'a; Bhardwaj, Radhika; Abbadi, Ghaneim; Koyippally, Mathew; Afzal, Mohammad

    2013-09-01

    For its variety of biological activities, Tamarix aucheriana (Decne.) Baum. (Tamaricaceae) has an extensive history as a traditional Arab medicine. Antimitogenic and chemo-sensitizing activities of syringic acid (SA) were studied against human colorectal cancer. Chromatographic and spectral data were used for the isolation and identification of SA. MTT, flow cytometry, in vitro invasion and angiogenesis assays, fluoremetry, ELISA and Real Time qPCR were used to test antimitogenic and chemo-sensitizing activities of SA, cell cycle, apoptosis, proteasome and NFκB-DNA-binding activities, cancer cell invasion and angiogenesis, and expression of cell cycle/apoptosis-related genes. SA showed a time- and dose-dependent (IC₅₀ = 0.95-1.2 mg mL⁻¹) antimitogenic effect against cancer cells with little cytotoxicity on normal fibroblasts (≤20%). SA-altered cell cycle (S/G2-M or G1/G2-M phases) in a time-dependent manner, induced apoptosis, inhibited DNA-binding activity of NFκB (p ≤ 0.0001), chymotrypsin-like/PGPH (peptidyl-glutamyl peptide-hydrolyzing) (p ≤ 0.0001) and the trypsin-like (p ≤ 0.002) activities of 26S proteasome and angiogenesis. SA also differentially sensitized cancer cells to standard chemotherapies with a marked increase in their sensitivity to camptothecin (500-fold), 5FU (20,000-fold), doxorubicin (210-fold), taxol (3134-fold), vinblastine (1000-fold), vincristine (130-fold) and amsacrine (107-fold) compared to standard drugs alone. SA exerted its chemotherapeutic and chemo-sensitizing effects through an array of mechanisms including cell-cycle arrest, apoptosis induction, inhibition of cell proliferation, cell migration, angiogenesis, NFκB DNA-binding and proteasome activities. These results demonstrate the potential of SA as an antimitogenic and chemo-sensitizing agent for human colorectal cancer.

  8. SiRNA knockdown of the DEK nuclear protein mRNA enhances apoptosis and chemosensitivity of canine transitional cell carcinoma cells.

    Science.gov (United States)

    Yamazaki, Hiroki; Iwano, Tomomi; Otsuka, Saori; Kagawa, Yumiko; Hoshino, Yuki; Hosoya, Kenji; Okumura, Masahiro; Takagi, Satoshi

    2015-04-01

    Transitional cell carcinoma (TCC) in dogs is an aggressive malignant neoplasm, originating in the epithelium of the urinary bladder. The DEK nuclear protein is overexpressed in several types of human bladder cancer, where it is involved in chromatin reconstruction, gene transcription and apoptosis. Since DEK represents a potential therapeutic target for canine TCC, this study was designed to investigate DEK expression in canine TCC and to determine the effects of DEK mRNA silencing on TCC cells in vitro. The gene expression profiles of seven selected cancer-associated genes was assessed in four canine TCC cell lines and expression of DEK protein was evaluated in bladder tissue biopsies from healthy dogs and those affected with cystitis or TCC. After transfection of four canine TCC cell lines with DEK-specific or scrambled siRNA, annexin V staining was performed to evaluate apoptosis, and methylthiazole tetrazolium assays were performed to assess both cell viability and sensitivity to carboplatin. DEK mRNA expression was relatively high in canine TCC cells and expression of the DEK protein was significantly greater in TCC tumours compared with the other tissue samples. After transfection with DEK-specific siRNA, apoptosis, cell growth inhibition, and enhanced sensitivity to carboplatin were observed in all TCC cells assessed. These research findings suggest that DEK could be a potential therapeutic target for canine TCC. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Nanoparticle delivery of chemosensitizers improve chemotherapy efficacy without incurring additional toxicity.

    Science.gov (United States)

    Caster, Joseph M; Sethi, Manish; Kowalczyk, Sonya; Wang, Edina; Tian, Xi; Nabeel Hyder, Sayed; Wagner, Kyle T; Zhang, Ying-Ao; Kapadia, Chintan; Man Au, Kin; Wang, Andrew Z

    2015-02-14

    Chemosensitizers can improve the therapeutic index of chemotherapy and overcome treatment resistance. Successful translation of chemosensitizers depends on the development of strategies that can preferentially deliver chemosensitizers to tumors while avoiding normal tissue. We hypothesized that nanoparticle (NP) formulation of chemosensitizers can improve their delivery to tumors which can in turn improve their therapeutic index. To demonstrate the proof of principle of this approach, we engineered NP formulations of two chemosensitizers, the PI3-kindase inhibitor wortmanin (Wtmn) and the PARP inhibitor olaparib. NP Wtmn and NP olaparib were evaluated as chemosensitizers using lung cancer cells and breast cancer cells respectively. We found Wtmn to be an efficient chemosensitizer in all tested lung-cancer cell lines reducing tumor cell growth between 20 and 60% compared to drug alone. NP formulation did not decrease its efficacy in vitro. Olaparib showed less consistent chemosensitization as a free drug or in NP formulation. NP Wtmn was further evaluated as a chemosensitizer using mouse models of lung cancer. We found that NP Wtmn is an effective chemosensitizer and more effective than free Wtmn showing a 32% reduction in tumor growth compared to free Wtmn when given with etoposide. Importantly, NP Wtmn was able to sensitize the multi-drug resistant H69AR cells to etoposide. Additionally, the combination of NP Wtmn and etoposide chemotherapy did not significantly increase toxicity. The present study demonstrates the proof of principle of using NP formulation of chemosensitizing drugs to improve the therapeutic index of chemotherapy.

  10. The Coexistence of Hypertension and Ovariectomy Additively Increases Cardiac Apoptosis

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    Yi-Yuan Lin

    2016-12-01

    Full Text Available To investigate whether the coexistence of hypertension and ovariectomy will increase cardiac Fas receptor and mitochondrial-dependent apoptotic pathways, histopathological analysis, the TUNEL assay and Western blotting were performed on the excised hearts from three groups of female spontaneously hypertensive rats (SHR, which were divided into a sham-operated group (SHR-Sham, bilaterally ovariectomized group (SHR-OVX and normotensive Wistar Kyoto rats (WKY. Compared with the WKY group, the SHR-Sham group exhibited decreased protein levels of ERα, ERβ, p-Akt/Akt, Bcl-2, Bcl-xL and p-Bad and decreased further in the SHR-OVX group, as well as protein levels of t-Bid, Bak, Bad, Bax, cytochrome c, activated caspase-9 and activated caspase-3 (mitochondria-dependent apoptosis increased in the SHR-Sham group and increased further in the SHR-OVX group. Compared with the WKY group, protein levels of Fas ligand, TNF-α, Fas death receptors, TNFR1, FADD and activated caspase-8 (Fas receptor-dependent apoptosis increased in the SHR-Sham group, but did not increase in the SHR-OVX group, except Fas ligand and TNF-α. The coexistence of hypertension and ovariectomy attenuated the estrogen receptor survival pathway and appeared to additively increase the cardiac mitochondria-dependent, but not the Fas receptor-dependent apoptosis pathway, which might provide one possible mechanism for the development of cardiac abnormalities in hypertensive postmenopausal women.

  11. Inhibition of MerTK increases chemosensitivity and decreases oncogenic potential in T-cell acute lymphoblastic leukemia

    International Nuclear Information System (INIS)

    Brandao, L N; Winges, A; Christoph, S; Sather, S; Migdall-Wilson, J; Schlegel, J; McGranahan, A; Gao, D; Liang, X; DeRyckere, D; Graham, D K

    2013-01-01

    Pediatric leukemia survival rates have improved dramatically over the past decades. However, current treatment protocols are still largely ineffective in cases of relapsed leukemia and are associated with a significant rate of chronic health conditions. Thus, there is a continued need for new therapeutic options. Here, we show that mer receptor tyrosine kinase (MerTK) was abnormally expressed in approximately one half of pediatric T-cell leukemia patient samples and T-cell acute lymphoblastic leukemia (T-ALL) cell lines. Stimulation of MerTK by the ligand Gas6 led to activation of the prosurvival proteins Erk 1/2 and Stat5, and MerTK-dependent activation of the STAT pathway in leukemia represents a novel finding. Furthermore, inhibition of MerTK expression increased the sensitivity of T-ALL cells to treatment with chemotherapeutic agents and decreased the oncogenic potential of the Jurkat T-ALL cell line in a methylcellulose colony-forming assay. Lastly, inhibition of MerTK expression significantly increased median survival in a xenograft mouse model of leukemia (30.5 days vs 60 days, P<0.0001). These results suggest that inhibition of MerTK is a promising therapeutic strategy for the treatment of leukemia and may allow for dose reduction of currently used chemotherapeutics resulting in decreased rates of therapy-associated toxicities

  12. Nicotine Induces Podocyte Apoptosis through Increasing Oxidative Stress.

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    Xiqian Lan

    Full Text Available Cigarette smoking plays an important role in the progression of chronic kidney disease (CKD. Nicotine, one of the major components of cigarette smoking, has been demonstrated to increase proliferation of renal mesangial cells. In this study, we examined the effect of nicotine on podocyte injury.To determine the expression of nicotinic acetylcholine receptors (nAChR subunits in podocytes, cDNAs and cell lysate of cultured human podocytes were used for the expression of nAChR mRNAs and proteins, respectively; and mouse renal cortical sections were subjected to immunofluorescant staining. We also studied the effect of nicotine on podocyte nephrin expression, reactive oxygen species (ROS generation (via DCFDA loading followed by fluorometric analysis, proliferation, and apoptosis (morphologic assays. We evaluated the effect of nicotine on podocyte downstream signaling including phosphorylation of ERK1/2, JNK, and p38 and established causal relationships by using respective inhibitors. We used nAChR antagonists to confirm the role of nicotine on podocyte injury.Human podocytes displayed robust mRNA and protein expression of nAChR in vitro studies. In vivo studies, mice renal cortical sections revealed co-localization of nAChRs along with synaptopodin. In vitro studies, nephrin expression in podocyte was decreased by nicotine. Nicotine stimulated podocyte ROS generation; nonetheless, antioxidants such as N-acetyl cysteine (NAC and TEMPOL (superoxide dismutase mimetic agent inhibited this effect of nicotine. Nicotine did not modulate proliferation but promoted apoptosis in podocytes. Nicotine enhanced podocyte phosphorylation of ERK1/2, JNK, and p38, and their specific inhibitors attenuated nicotine-induced apoptosis. nAChR antagonists significantly suppressed the effects of nicotine on podocyte.Nicotine induces podocyte apoptosis through ROS generation and associated downstream MAPKs signaling. The present study provides insight into molecular

  13. Peiminine serves as an adriamycin chemosensitizer in gastric cancer by modulating the EGFR/FAK pathway.

    Science.gov (United States)

    Tang, Qianqian; Wang, Yunfei; Ma, Lanjing; Ding, Meiling; Li, Tingyu; Nie, Yongzhan; Gu, Zhengyi

    2018-03-01

    Gastric cancer (GC) is one of the most common malignancies of the digestive tract. Adriamycin (ADR) has been widely utilized in various chemotherapy regimens for treating GC, yet its long-term application may increase drug resistance resulting in treatment failure. Increasing evidence shows that bioactive natural products can be used as chemotherapeutic sensitizers that can significantly improve chemotherapy sensitivity. Peiminine (PMI) is a biologically active component extracted from Fritillaria walujewii Regel. Thus, in the present study, we aimed to investigate whether peiminine (PMI) alters the chemosensitivity of GC to adriamycin (ADR). GC cells were treated with ADR with or without PMI. MTT assay, flow cytometry and a nude mouse tumor xenograft model of SGC7901 cells were used to evaluate the chemosensitization activity of PMI combined with ADR. Western blotting was used to examine the expression of cyclin D1 and cleaved PARP. The RayBio® Human RTK phosphorylation antibody array kit was used to test the differential protein expression. Compared with the ADR group, PMI combined with ADR significantly suppressed cell proliferation and induced cell apoptosis in vitro. The growth curve and tumor weight of the tumor xenografts were significantly decreased in mice treated with the combination of PMI and ADR. However, the organs showed no obvious abnormality after treatment with PMI plus ADR. The expression of cyclin D1 was decreased and the level of cleaved PARP was increased after treatment with PMI and ADR. The expression of p-EGFR and p-FAK was downregulated in cells treated with PMI and ADR, and the validation of p-EGFR and p-FAK was in accordance with the result of the phosphorylation antibody array kit. PMI may serve as a new chemosensitizer by inhibiting the proliferation and inducing the apoptosis to enhance the chemotherapeutic drug sensitivity of ADR in GC.

  14. Baicalin prevents Candida albicans infections via increasing its apoptosis rate

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    Yang, Shulong; Fu, Yingyuan, E-mail: yingyuanfu@126.com; Wu, Xiuzhen; Zhou, Zhixing; Xu, Jing; Zeng, Xiaoping; Kuang, Nanzhen; Zeng, Yurong

    2014-08-15

    Highlights: • Baicalin increases the ratio of the G0/G1 stages and C. albicans apoptosis. • Baicalin decreases the proliferation index of C. albicans. • Baicalin inhibits the biosynthesis of DNA, RNA and protein in C. albicans. • Baicalin depresses Succinate Dehydrogenase and Ca{sup 2+}–Mg{sup 2+} ATPase in C. albicans. • Baicalin increases the endocytic free Ca{sup 2+} concentration in C. albicans. - Abstract: Background: These experiments were employed to explore the mechanisms underlying baicalin action on Candida albicans. Methodology and principal findings: We detected the baicalin inhibition effects on three isotope-labeled precursors of {sup 3}H-UdR, {sup 3}H-TdR and {sup 3}H-leucine incorporation into C. albicans using the isotope incorporation technology. The activities of Succinate Dehydrogenase (SDH), cytochrome oxidase (CCO) and Ca{sup 2+}–Mg{sup 2+} ATPase, cytosolic Ca{sup 2+} concentration, the cell cycle and apoptosis, as well as the ultrastructure of C.albicans were also tested. We found that baicalin inhibited {sup 3}H-UdR, {sup 3}H-TdR and {sup 3}H-leucine incorporation into C.albicans (P < 0.005). The activities of the SDH and Ca{sup 2+}–Mg{sup 2+} ATPase of C.albicans in baicalin groups were lower than those in control group (P < 0.05). Ca{sup 2+} concentrations of C. albicans in baicalin groups were much higher than those in control group (P < 0.05). The ratio of C.albicans at the G0/G1 stage increased in baicalin groups in dose dependent manner (P < 0.01). There were a significant differences in the apoptosis rate of C.albicans between baicalin and control groups (P < 0.01). After 12–48 h incubation with baicalin (1 mg/ml), C. albicans shown to be markedly damaged under transmission electron micrographs. Innovation and significance: Baicalin can increase the apoptosis rate of C. albicans. These effects of Baicalin may involved in its inhibiting the activities of the SDH and Ca{sup 2+}–Mg{sup 2+} ATPase, increasing

  15. Baicalin prevents Candida albicans infections via increasing its apoptosis rate

    International Nuclear Information System (INIS)

    Yang, Shulong; Fu, Yingyuan; Wu, Xiuzhen; Zhou, Zhixing; Xu, Jing; Zeng, Xiaoping; Kuang, Nanzhen; Zeng, Yurong

    2014-01-01

    Highlights: • Baicalin increases the ratio of the G0/G1 stages and C. albicans apoptosis. • Baicalin decreases the proliferation index of C. albicans. • Baicalin inhibits the biosynthesis of DNA, RNA and protein in C. albicans. • Baicalin depresses Succinate Dehydrogenase and Ca 2+ –Mg 2+ ATPase in C. albicans. • Baicalin increases the endocytic free Ca 2+ concentration in C. albicans. - Abstract: Background: These experiments were employed to explore the mechanisms underlying baicalin action on Candida albicans. Methodology and principal findings: We detected the baicalin inhibition effects on three isotope-labeled precursors of 3 H-UdR, 3 H-TdR and 3 H-leucine incorporation into C. albicans using the isotope incorporation technology. The activities of Succinate Dehydrogenase (SDH), cytochrome oxidase (CCO) and Ca 2+ –Mg 2+ ATPase, cytosolic Ca 2+ concentration, the cell cycle and apoptosis, as well as the ultrastructure of C.albicans were also tested. We found that baicalin inhibited 3 H-UdR, 3 H-TdR and 3 H-leucine incorporation into C.albicans (P < 0.005). The activities of the SDH and Ca 2+ –Mg 2+ ATPase of C.albicans in baicalin groups were lower than those in control group (P < 0.05). Ca 2+ concentrations of C. albicans in baicalin groups were much higher than those in control group (P < 0.05). The ratio of C.albicans at the G0/G1 stage increased in baicalin groups in dose dependent manner (P < 0.01). There were a significant differences in the apoptosis rate of C.albicans between baicalin and control groups (P < 0.01). After 12–48 h incubation with baicalin (1 mg/ml), C. albicans shown to be markedly damaged under transmission electron micrographs. Innovation and significance: Baicalin can increase the apoptosis rate of C. albicans. These effects of Baicalin may involved in its inhibiting the activities of the SDH and Ca 2+ –Mg 2+ ATPase, increasing cytosolic Ca 2+ content and damaging the ultrastructure of C. albicans

  16. Wip1 gene silencing enhances the chemosensitivity of human colon cancer cells.

    Science.gov (United States)

    Xia, Zhong-Sheng; Wu, Di; Zhong, Wa; Lu, Xi-Ji; Yu, Tao; Chen, Qi-Kui

    2017-08-01

    Colon cancer is one of the most common cancers in the world. Multidrug resistance is one of the main reasons for failure of therapy in patients with advanced colon cancer. In previous studies, multiple methods were investigated to reverse the multidrug resistance of colon cancer cells. However, to date, no clinical method has been identified to be satisfactory. Therefore, successful reversal of drug resistance in colon cancer cells still requires new therapeutic strategies or pharmaceuticals. Wild-type p53-induced phosphatase (Wip1), a member of the 2C type serine/threonine protein phosphatase family, is closely associated with the p53 gene, which is the most important tumor-suppressor gene. Wip1 was reported to be associated with the chemosensitivity of breast cancer cells. However, the correlation between the expression of Wip1 gene and the chemosensitivity of colon cancer cells has not been reported yet. In the present study, Wip1-811 small interfering RNA (siRNA) targeting Wip1 was investigated to reverse the multidrug resistance of colon cancer cells. The siRNA duplexes were transfected into RKO colon cancer cells. The messenger RNA (mRNA) expression of Wip1 was measured by reverse transcription-quantitative polymerase chain reaction. The protein level of Wip1 was detected by western blotting. The cell viability was measured by MTS assay. The cell apoptosis and cell cycle were analyzed by flow cytometry. Intracellular adriamycin cumulative concentration was determined using flow cytometry. Wip1-811 siRNA efficiently inhibited the expression of Wip1 at the mRNA and protein levels, and enhanced the sensitivity of RKO colon cancer cells towards chemotherapy, which was accompanied by increased cell apoptosis, following the inhibition of Wip1 gene expression. These results indicate that Wip1 gene silencing could enhance the chemosensitivity of colon cancer cells, which may provide a new potential approach for the reversal of multidrug resistance in colon cancer

  17. Vorinostat enhances chemosensitivity to arsenic trioxide in K562 cell line

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    Nainong Li

    2015-05-01

    Full Text Available Objective. This study aimed to investigate the chemosensitive augmentation effect and mechanism of HDAC inhibitor Vorinostat (SAHA in combination with arsenic trioxide (ATO on proliferation and apoptosis of K562 cells.Methods. The CCK-8 assay was used to compare proliferation of the cells. Annexin-V and PI staining by flow cytometry and acridine orange/ethidium bromide stains were used to detect and quantify apoptosis. Western blot was used to detect expression of p21, Akt, pAkt, p210, Acetyl-Histone H3, and Acetyl-Histone H4 proteins.Results. SAHA and ATO inhibited proliferation of K562 cells in an additive and time- and dose-dependent manner. SAHA in combination with ATO showed significant apoptosis of K562 cells in comparison to the single drugs alone (p < 0.01. Both SAHA and ATO alone and in combination showed lower levels of p210 expression. SAHA and SAHA and ATO combined treatment showed increased levels of Acetyl-Histone H3 and Acetyl-Histone H4 protein expression. SAHA alone showed increased expression of p21, while ATO alone and in combination with SAHA showed no significant change. SAHA and ATO combined therapy showed lower levels of Akt and pAkt protein expression than SAHA or ATO alone.Conclusion. SAHA and ATO combined treatment inhibited proliferation, induced apoptosis, and showed a chemosensitive augmentation effect on K562 cells. The mechanism might be associated with increasing histone acetylation levels as well as regulating the Akt signaling pathway.

  18. THE LOCUS COERULEUS AND CENTRAL CHEMOSENSITIVITY

    Science.gov (United States)

    Gargaglioni, Luciane H.; Hartzler, Lynn K.; Putnam, Robert W.

    2010-01-01

    The locus coeruleus (LC) lies in the dorsal pons and supplies noradrenergic (NA) input to many regions of the brain, including respiratory control areas. The LC may provide tonic input for basal respiratory drive and is involved in central chemosensitivity since focal acidosis of the region stimulates ventilation and ablation reduces CO2-induced increased ventilation. The output of LC is modulated by both serotonergic and glutamatergic inputs. A large percentage of LC neurons are intrinsically activated by hypercapnia. This percentage and the magnitude of their response are highest in young neonates and decrease dramatically after postnatal day P10. The cellular bases for intrinsic chemosensitivity of LC neurons are comprised of multiple factors, primary among them being reduced extracellular and intracellular pH, which inhibit inwardly rectifying and voltage-gated K+ channels, and activate L-type Ca2+ channels. Activation of KCa channels in LC neurons may limit their ultimate response to hypercapnia. Finally, the LC mediates central chemosensitivity and contains pH-sensitive neurons in amphibians, suggesting that the LC has a long-standing phylogenetic role in respiratory control. PMID:20435170

  19. The marine toxin, Yessotoxin, induces apoptosis and increases mitochondrial activity

    Directory of Open Access Journals (Sweden)

    Andrea Fernandez-Araujo

    2014-06-01

    Discussion: Colorimetric MTT assay is widely used as a viability measurement method (McHale and L., 1988;Chiba et al., 1998. But after YTX treatment, MTT assay had shown problems to detect a cell viability decrease. In this sense, in primary cardiac cell cultures, a false increment of the proliferation rate opposite to Sulforhodamine B assay (SRB results was reported after YTX treatment (Dell'Ovo et al., 2008. Also the same effect was obtained in different cancer cell lines after assaying anticancer therapies (Ulukaya et al., 2004. In our study, an increase in cell viability using MTT was observed when the number of cells was high, while by using the LDH assay a significant viability decrease was measured. In these conditions, YTX is activating extrinsic apoptosis cell death by increasing caspase 8 activity and caspase 3 levels. The explanation for this increase was found when the mitochondrial activity was quantified cell by cell in a cytometer. In these conditions a significant increment of mitochondrial activity was detected. Since the cell population is too high, the increase in mitochondrial activity that detects the MTT test disguised the decrease of signal due to the cell death and point to a false proliferation increase. In this sense, a mitochondrial activity decrease was observed after 48 hours YTX treatment in BE(2-M17 neuroblastoma cell line (Leira et al., 2002. However, this study was done in a microplate reader with a small number of cells (Leira et al., 2002. Therefore, to measure the viability by MTT assay is very important to take into account the number of cells per condition when the experiment is designed. Alternative assays, such as LDH test, independently of the direct mitochondrial activity, can be used.

  20. Cellular mechanisms involved in CO(2) and acid signaling in chemosensitive neurons.

    Science.gov (United States)

    Putnam, Robert W; Filosa, Jessica A; Ritucci, Nicola A

    2004-12-01

    An increase in CO(2)/H(+) is a major stimulus for increased ventilation and is sensed by specialized brain stem neurons called central chemosensitive neurons. These neurons appear to be spread among numerous brain stem regions, and neurons from different regions have different levels of chemosensitivity. Early studies implicated changes of pH as playing a role in chemosensitive signaling, most likely by inhibiting a K(+) channel, depolarizing chemosensitive neurons, and thereby increasing their firing rate. Considerable progress has been made over the past decade in understanding the cellular mechanisms of chemosensitive signaling using reduced preparations. Recent evidence has pointed to an important role of changes of intracellular pH in the response of central chemosensitive neurons to increased CO(2)/H(+) levels. The signaling mechanisms for chemosensitivity may also involve changes of extracellular pH, intracellular Ca(2+), gap junctions, oxidative stress, glial cells, bicarbonate, CO(2), and neurotransmitters. The normal target for these signals is generally believed to be a K(+) channel, although it is likely that many K(+) channels as well as Ca(2+) channels are involved as targets of chemosensitive signals. The results of studies of cellular signaling in central chemosensitive neurons are compared with results in other CO(2)- and/or H(+)-sensitive cells, including peripheral chemoreceptors (carotid body glomus cells), invertebrate central chemoreceptors, avian intrapulmonary chemoreceptors, acid-sensitive taste receptor cells on the tongue, and pain-sensitive nociceptors. A multiple factors model is proposed for central chemosensitive neurons in which multiple signals that affect multiple ion channel targets result in the final neuronal response to changes in CO(2)/H(+).

  1. Chemosensitization of Human Renal Cell Cancer Using Antisense Oligonucleotides Targeting the Antiapoptotic Gene Clusterin

    Directory of Open Access Journals (Sweden)

    Tobias Zellweger

    2001-01-01

    administration of clusterin ASO plus paclitaxel acted synergistically to increase apoptosis and significantly delay Caki-2 tumor growth, compared to mismatch control oligonucleotide plus paclitaxel. In addition, TUNEL staining revealed increased apoptotic cells in tumors treated with clusterin ASO plus paclitaxel compared to treatment with either clusterin ASO or paclitaxel alone. CONCLUSION: These findings confirm that the use of clusterin ASO may be a feasible strategy to enhance chemosensitivity for patients with advanced RCC.

  2. N-acetylphytosphingosine enhances the radiosensitivity of tumor cells by increasing apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Han, Y.; Kim, Y.; Yun, Y.; Jeon, S.; Kim, K.; Song, J. [Lab. of Immunology, Korea Inst. of Radiological and Medical Sciences, KAERI, Seoul (Korea); Hong, S.H. [Lab. of Experimental therapeutics, Korea Inst. of Radiological and Medical Sciences, KAERI, Seoul (Korea); Park, C. [Doosan Biotech BU, Yongin-City, Kyonggi-Do (Korea)

    2005-07-01

    Ceramides are well-known second messengers which mediate apoptosis, proliferation, differentiation in mammalian cells, but the physiological roles of phytosphingosines are poorly understood. We hypothesized that one of the phytosphingosine derivatives, N-acetylphytosphingosine (NAPS) can induce apoptosis in human leukemia Jurkat cell line and increase apoptosis in irradiated MDA-MB-231 cells. We first examined the effect of NAPS on apoptosis of Jurkat cells. NAPS had a more rapid and stronger apoptotic effect than C{sub 2}-ceramide in Jurkat cells and significant increase of apoptosis was observed at 3 h after treatment. In contrast, the apoptosis induced by C2-ceramide was observed only after 16 h of treatment. NAPS induced apoptosis was mediated by caspase 3 and 8 activation and inhibited by z-VAD-fmk. Ceramide plays a pivotal role in radiation induced apoptosis. We postulated that exogenous treatment of NAPS sensitizes tumor cells to ionizing radiation, since NAPS might be used as a more effective alternative to C2-ceramide. As expected, NAPS decreased clonogenic survival of irradiated MDA-MB-231 cells dose dependently, and apoptosis of irradiated cells in the presence of NAPS was increased through the caspase activation. Taken together, NAPS is an effective apoptosis-inducing agent, which can be readily synthesized from yeast sources, and is a potent alternative to ceramide for the further study of ceramide associated signaling and the development of radiosensitizing agent. (orig.)

  3. Increased apoptosis skull of pups born to Toxoplasma gondii ...

    African Journals Online (AJOL)

    Background: Toxoplasma gondii is an intracellular obligate protozoan parasite that infects most warm-blooded animals including humans. It can cause congenital infection with clinical symptoms ranging from mild to severe including microcephaly. At the cellular level, infection T. gondii causes apoptosis in some tissues and ...

  4. Increased radiosensitivity and radiation-induced apoptosis in SRC-3 knockout mice

    International Nuclear Information System (INIS)

    Jin Jie; Wang Yu; Xu Yang; Chen Shilei; Wang Junping; Ran Xinze; Su Yongping; Wang Jin

    2014-01-01

    Steroid receptor coactivator-3 (SRC-3), a multifunctional transcriptional coactivator, plays an important role in regulation of cell apoptosis in chemoresistant cancer cells. However, its role in radiation-induced apoptosis in hematopoietic cells is still unclear. In this study, we used SRC-3 knockout (SRC-3 -/- ) mice to assess the role of SRC-3 in radiation-induced hematopoietic injury in vivo. After a range of doses of irradiation, SRC-3 -/- mice exhibited lower counts of peripheral blood cells and bone marrow (BM) mononuclear cells and excessive BM depression, which resulted in a significantly higher mortality compared with wildtype mice. Moreover, BM mononuclear cells obtained from SRC-3 -/- mice showed a remarkable increase in radiation-induced apoptosis. Collectively, our data demonstrate that SRC-3 plays a role in radiation-induced apoptosis of BM hematopoietic cells. Regulation of SRC-3 might influence the radiosensitivity of hematopoietic cells, which highlights a potential therapeutic target for radiation-induced hematopoietic injury. (author)

  5. TIMP-1 gene deficiency increases tumour cell sensitivity to chemotherapy-induced apoptosis

    DEFF Research Database (Denmark)

    Davidsen, Marie Louise; Würts, S.Ø.; Rømer, Maria Unni Koefoed

    2006-01-01

    in cancer. In this regard, several studies have demonstrated an antiapoptotic effect of TIMP-1 in a number of different cell types. Since chemotherapy works by inducing apoptosis in cancer cells, we raised the hypothesis that TIMP-1 promotes resistance against chemotherapeutic drugs. In order to investigate...... this hypothesis, we have established TIMP-1 gene-deficient and TIMP-1 wild-type fibrosarcoma cells from mouse lung tissue. We have characterised these cells with regard to TIMP-1 genotype, TIMP-1 expression, malignant transformation and sensitivity to chemotherapy-induced apoptosis. We show that TIMP-1 gene...... deficiency increases the response to chemotherapy considerably, confirming that TIMP-1 protects the cells from apoptosis. This is to our knowledge the first study investigating TIMP-1 and chemotherapy-induced apoptosis employing a powerful model system comprising TIMP-1 gene-deficient cells...

  6. Effect of GEN1 interference on the chemosensitivity of the breast cancer MCF-7 and SKBR3 cell lines.

    Science.gov (United States)

    Wu, Yunlu; Qian, Ying; Zhou, Guozhong; Lv, Juan; Yan, Qiuyue; Dong, Xuejun

    2016-06-01

    Chemotherapy is a notable method for the treatment of breast cancer. Numerous genes associated with the sensitivity of cancer to chemotherapy have been found. In recent years, evidence has suggested that a particular structure termed Holliday junction (HJ) plays a crucial role in cancer chemosensitivity. Targeting HJ resolvases, such as structure-specific endonuclease subunit SLX4 (Slx4) and MUS81 structure-specific endonuclease subunit (Mus81), significantly increases the chemosensitivity of tumor cells. Flap endonuclease GEN homolog 1 (GEN1) is a HJ resolvase that belongs to the Rad2/xeroderma pigmentosum complementation group G nuclease family. Whether GEN1 affects the chemosensitivity of tumor cells in a similar manner to Slx4 and Mus81 remains unknown. The aim of the present study was to determine the effect of GEN1 interference on the chemosensitivity of breast cancer cell lines. The investigation of the function of GEN1 was performed using MCF-7 and SKBR3 cells. Short hairpin RNA was used to suppress the expression of GEN1, and western blot analysis and reverse transcription-quantitative polymerase chain reaction were used to detect gene expression. In addition, a cell counting kit-8 assay was performed to detect the viability of cells and flow cytometry was performed to test apoptosis levels. Suppression of GEN1 in SKBR3 cells effectively increased the sensitivity to the chemotherapeutic drug 5-fluorouracil (5-FU), while MCF-7 cells showed no significant change in sensitivity following GEN1 suppression. However, when GEN1 was targeted in addition to Mus81, the MCF-7 cells also demonstrated a significantly increased sensitivity to 5-FU. In addition, when the level of Mus81 was low, GEN1 expression was increased under a low concentration of 5-FU. The present results suggest that GEN1 may play different roles in different breast cancer cell lines. The function of GEN1 may be affected by the level of Mus81 in the cell line. In addition, GEN1 interference may

  7. Characterized mechanism of alpha-mangostin-induced cell death: caspase-independent apoptosis with release of endonuclease-G from mitochondria and increased miR-143 expression in human colorectal cancer DLD-1 cells.

    Science.gov (United States)

    Nakagawa, Yoshihito; Iinuma, Munekazu; Naoe, Tomoki; Nozawa, Yoshinori; Akao, Yukihiro

    2007-08-15

    alpha-Mangostin, a xanthone from the pericarps of mangosteen (Garcinia mangostana Linn.), was evaluated for in vitro cytotoxicity against human colon cancer DLD-1 cells. The number of viable cells was consistently decreased by the treatment with alpha-mangostin at more than 20 microM. The cytotoxic effect of 20 microM alpha-mangostin was found to be mainly due to apoptosis, as indicated by morphological findings. Western blotting, the results of an apoptosis inhibition assay using caspase inhibitors, and the examination of caspase activity did not demonstrate the activation of any of the caspases tested. However, endonuclease-G released from mitochondria with the decreased mitochondrial membrane potential was shown. The levels of phospho-Erk1/2 were increased in the early phase until 1h after the start of treatment and thereafter decreased, and increased again in the late phase. On the other hand, the level of phospho-Akt was sharply reduced with the process of apoptosis after 6h of treatment. Interestingly, the level of microRNA-143, which negatively regulates Erk5 at translation, gradually increased until 24h following the start of treatment. We also examined the synergistic growth suppression in DLD-1 cells by the combined treatment of the cells with alpha-mangostin and 5-FU which is one of the most effective chemotherapeutic agents for colorectal adenocarcinoma. The co-treatment with alpha-mangostin and 5-FU, both at 2.5 microM, augmented growth inhibition compared with the treatment with 5 microM of alpha-mangostin or 5 microM 5-FU alone. These findings indicate unique mechanisms of alpha-mangostin-induced apoptosis and its action as an effective chemosensitizer.

  8. NFAT2 mediates high glucose-induced glomerular podocyte apoptosis through increased Bax expression

    Energy Technology Data Exchange (ETDEWEB)

    Li, Ruizhao, E-mail: liruizhao1979@126.com [Department of Nephrology, Guangdong General Hospital, Guangdong Academy of Medical Sciences, 106 Zhongshan No. 2 Road, Guangzhou, 510080 (China); Zhang, Li, E-mail: Zhanglichangde@163.com [Department of Nephrology, Guangdong General Hospital, Guangdong Academy of Medical Sciences, 106 Zhongshan No. 2 Road, Guangzhou, 510080 (China); Southern Medical University, Guangzhou, Guangdong (China); Shi, Wei, E-mail: shiwei.gd@139.com [Department of Nephrology, Guangdong General Hospital, Guangdong Academy of Medical Sciences, 106 Zhongshan No. 2 Road, Guangzhou, 510080 (China); Zhang, Bin, E-mail: zhangbinyes@yahoo.com.cn [Department of Nephrology, Guangdong General Hospital, Guangdong Academy of Medical Sciences, 106 Zhongshan No. 2 Road, Guangzhou, 510080 (China); Liang, Xinling, E-mail: xinlingliang@yahoo.com [Department of Nephrology, Guangdong General Hospital, Guangdong Academy of Medical Sciences, 106 Zhongshan No. 2 Road, Guangzhou, 510080 (China); Liu, Shuangxin, E-mail: mplsxi@yahoo.com.cn [Department of Nephrology, Guangdong General Hospital, Guangdong Academy of Medical Sciences, 106 Zhongshan No. 2 Road, Guangzhou, 510080 (China); Wang, Wenjian, E-mail: wwjph@yahoo.com [Department of Nephrology, Guangdong General Hospital, Guangdong Academy of Medical Sciences, 106 Zhongshan No. 2 Road, Guangzhou, 510080 (China)

    2013-04-15

    Background: Hyperglycemia promotes podocyte apoptosis and plays a key role in the pathogenesis of diabetic nephropathy. However, the mechanisms that mediate hyperglycemia-induced podocyte apoptosis is still far from being fully understood. Recent studies reported that high glucose activate nuclear factor of activated T cells (NFAT) in vascular smooth muscle or pancreatic β-cells. Here, we sought to determine if hyperglycemia activates NFAT2 in cultured podocyte and whether this leads to podocyte apoptosis. Meanwhile, we also further explore the mechanisms of NFAT2 activation and NFAT2 mediates high glucose-induced podocyte apoptosis. Methods: Immortalized mouse podocytes were cultured in media containing normal glucose (NG), or high glucose (HG) or HG plus cyclosporine A (a pharmacological inhibitor of calcinerin) or 11R-VIVIT (a special inhibitor of NFAT2). The activation of NFAT2 in podocytes was detected by western blotting and immunofluorescence assay. The role of NFAT2 in hyperglycemia-induced podocyte apoptosis was further evaluated by observing the inhibition of NFAT2 activation by 11R-VIVIT using flow cytometer. Intracellular Ca{sup 2+} was monitored in HG-treated podcocytes using Fluo-3/AM. The mRNA and protein expression of apoptosis gene Bax were measured by real time-qPCR and western blotting. Results: HG stimulation activated NFAT2 in a time- and dose-dependent manner in cultured podocytes. Pretreatment with cyclosporine A (500 nM) or 11R-VIVIT (100 nM) completely blocked NFAT2 nuclear accumulation. Meanwhile, the apoptosis effects induced by HG were also abrogated by concomitant treatment with 11R-VIVIT in cultured podocytes. We further found that HG also increased [Ca{sup 2+}]i, leading to activation of calcineurin, and subsequent increased nuclear accumulation of NFAT2 and Bax expression in cultured podocytes. Conclusion: Our results identify a new finding that HG-induced podocyte apoptosis is mediated by calcineurin/NFAT2/Bax signaling pathway

  9. Curcumin chemosensitizes 5-fluorouracil resistant MMR-deficient human colon cancer cells in high density cultures.

    Directory of Open Access Journals (Sweden)

    Mehdi Shakibaei

    Full Text Available OBJECTIVE: Treatment of colorectal cancer (CRC remains a clinical challenge, as more than 15% of patients are resistant to 5-Fluorouracil (5-FU-based chemotherapeutic regimens, and tumor recurrence rates can be as high as 50-60%. Cancer stem cells (CSC are capable of surviving conventional chemotherapies that permits regeneration of original tumors. Therefore, we investigated the effectiveness of 5-FU and plant polyphenol (curcumin in context of DNA mismatch repair (MMR status and CSC activity in 3D cultures of CRC cells. METHODS: High density 3D cultures of CRC cell lines HCT116, HCT116+ch3 (complemented with chromosome 3 and their corresponding isogenic 5-FU-chemo-resistant derivative clones (HCT116R, HCT116+ch3R were treated with 5-FU either without or with curcumin in time- and dose-dependent assays. RESULTS: Pre-treatment with curcumin significantly enhanced the effect of 5-FU on HCT116R and HCR116+ch3R cells, in contrast to 5-FU alone as evidenced by increased disintegration of colonospheres, enhanced apoptosis and by inhibiting their growth. Curcumin and/or 5-FU strongly affected MMR-deficient CRC cells in high density cultures, however MMR-proficient CRC cells were more sensitive. These effects of curcumin in enhancing chemosensitivity to 5-FU were further supported by its ability to effectively suppress CSC pools as evidenced by decreased number of CSC marker positive cells, highlighting the suitability of this 3D culture model for evaluating CSC marker expression in a close to vivo setting. CONCLUSION: Our results illustrate novel and previously unrecognized effects of curcumin in enhancing chemosensitization to 5-FU-based chemotherapy on DNA MMR-deficient and their chemo-resistant counterparts by targeting the CSC sub-population. (246 words in abstract.

  10. Curcumin chemosensitizes 5-fluorouracil resistant MMR-deficient human colon cancer cells in high density cultures.

    Science.gov (United States)

    Shakibaei, Mehdi; Buhrmann, Constanze; Kraehe, Patricia; Shayan, Parviz; Lueders, Cora; Goel, Ajay

    2014-01-01

    Treatment of colorectal cancer (CRC) remains a clinical challenge, as more than 15% of patients are resistant to 5-Fluorouracil (5-FU)-based chemotherapeutic regimens, and tumor recurrence rates can be as high as 50-60%. Cancer stem cells (CSC) are capable of surviving conventional chemotherapies that permits regeneration of original tumors. Therefore, we investigated the effectiveness of 5-FU and plant polyphenol (curcumin) in context of DNA mismatch repair (MMR) status and CSC activity in 3D cultures of CRC cells. High density 3D cultures of CRC cell lines HCT116, HCT116+ch3 (complemented with chromosome 3) and their corresponding isogenic 5-FU-chemo-resistant derivative clones (HCT116R, HCT116+ch3R) were treated with 5-FU either without or with curcumin in time- and dose-dependent assays. Pre-treatment with curcumin significantly enhanced the effect of 5-FU on HCT116R and HCR116+ch3R cells, in contrast to 5-FU alone as evidenced by increased disintegration of colonospheres, enhanced apoptosis and by inhibiting their growth. Curcumin and/or 5-FU strongly affected MMR-deficient CRC cells in high density cultures, however MMR-proficient CRC cells were more sensitive. These effects of curcumin in enhancing chemosensitivity to 5-FU were further supported by its ability to effectively suppress CSC pools as evidenced by decreased number of CSC marker positive cells, highlighting the suitability of this 3D culture model for evaluating CSC marker expression in a close to vivo setting. Our results illustrate novel and previously unrecognized effects of curcumin in enhancing chemosensitization to 5-FU-based chemotherapy on DNA MMR-deficient and their chemo-resistant counterparts by targeting the CSC sub-population. (246 words in abstract).

  11. Increased enterocyte apoptosis and Fas-Fas ligand system in celiac disease.

    Science.gov (United States)

    Ciccocioppo, R; Di Sabatino, A; Parroni, R; Muzi, P; D'Alò, S; Ventura, T; Pistoia, M A; Cifone, M G; Corazza, G R

    2001-04-01

    Our aim was to evaluate whether increased enterocyte apoptosis was responsible for mucosal flattening in celiac disease (CD), and, since the mechanisms responsible for tissue injury in this condition are unknown, we studied the possibility that the Fas-Fas ligand (FasL) system may be involved. Endoscopic duodenal biopsy specimens from 12 patients with untreated and 12 with treated CD and 12 control subjects were evaluated for enterocyte apoptosis by the terminal deoxynucleotidyl transferase-mediated digoxigenin-deoxyuridine triphosphate nick-end labeling assay and for Fas and FasL expression by immunohistochemistry. A coculture of isolated enterocytes (targets) and purified lamina propria mononuclear cells (LPMCs) (effectors) was performed in the absence or presence of an antagonistic ZB4 anti-Fas antibody. We found a significant correlation between the degree of villous atrophy, morphometrically evaluated, and the level of enterocyte apoptosis, suggesting that mucosal flattening is a consequence of exaggerated epithelial cell death. Most celiac enterocytes express Fas, and LPMCs express FasL. The abolishment of enterocyte apoptosis observed in the presence of ZB4 antibody suggests that enterocytes are potential targets of lymphocyte infiltrate. These results directly demonstrate that FasL-mediated apoptosis is a major mechanism responsible for enterocyte death in CD.

  12. Inhibition ofSTAT3with the Generation 2.5 Antisense Oligonucleotide, AZD9150, Decreases Neuroblastoma Tumorigenicity and Increases Chemosensitivity.

    Science.gov (United States)

    Odate, Seiichi; Veschi, Veronica; Yan, Shuang; Lam, Norris; Woessner, Richard; Thiele, Carol J

    2017-04-01

    Purpose: Neuroblastoma is a pediatric tumor of peripheral sympathoadrenal neuroblasts. The long-term event-free survival of children with high-risk neuroblastoma is still poor despite the improvements with current multimodality treatment protocols. Activated JAK/STAT3 pathway plays an important role in many human cancers, suggesting that targeting STAT3 is a promising strategy for treating high-risk neuroblastoma. Experimental Design: To evaluate the biologic consequences of specific targeting of STAT3 in neuroblastoma, we assessed the effect of tetracycline (Tet)-inducible STAT3 shRNA and the generation 2.5 antisense oligonucleotide AZD9150 which targets STAT3 in three representative neuroblastoma cell line models (AS, NGP, and IMR32). Results: Our data indicated that Tet-inducible STAT3 shRNA and AZD9150 inhibited endogenous STAT3 and STAT3 target genes. Tet-inducible STAT3 shRNA and AZD9150 decreased cell growth and tumorigenicity. In vivo , STAT3 inhibition by Tet-inducible STAT3 shRNA or AZD9150 alone had little effect on growth of established tumors. However, when treated xenograft tumor cells were reimplanted into mice, there was a significant decrease in secondary tumors in the mice receiving AZD9150-treated tumor cells compared with the mice receiving ntASO-treated tumor cells. This indicates that inhibition of STAT3 decreases the tumor-initiating potential of neuroblastoma cells. Furthermore, inhibition of STAT3 significantly increased neuroblastoma cell sensitivity to cisplatin and decreased tumor growth and increased the survival of tumor-bearing mice in vivo Conclusions: Our study supports the development of strategies targeting STAT3 inhibition in combination with conventional chemotherapy for patients with high-risk neuroblastoma. Clin Cancer Res; 23(7); 1771-84. ©2016 AACR . ©2016 American Association for Cancer Research.

  13. Chemosensitization of plant pathogenic fungi to agricultural fungicides.

    Directory of Open Access Journals (Sweden)

    Vitaly eDzhavakhiya

    2012-03-01

    Full Text Available A common consequence of using agricultural fungicides is the development of resistance by fungal pathogens, which undermines reliability of fungicidal effectiveness. A potentially new strategy to aid in overcoming or minimizing this problem is enhancement of pathogen sensitivity to fungicides, or chemosensitization. Chemosensitization can be accomplished by combining a commercial fungicide with a certain non- or marginally fungicidal substance at levels where, alone, neither compound would be effective. Chemosensitization decreases the probability of the pathogen developing resistance, reduces the toxic impact on the environment by lowering effective dosage levels of toxic fungicides, and improves efficacy of antifungal agents. The present study shows that the antifungal activity of azole and strobilurin fungicides can be significantly enhanced through their co-application with certain natural or synthetic products against several economically important plant pathogenic fungi. Quadris (azoxystrobin combined with thymol at a non-fungitoxic concentration produced much higher growth inhibition of Bipolaris sorokiniana, Phoma glomerata, Alternaria sp. and Stagonospora nodorum than the fungicide alone. The effect of Dividend (difenoconazole applied with thymol significantly enhanced antifungal activity against B. sorokiniana and S. nodorum. Folicur (tebuconazole combined with 4-hydroxybenzaldehyde (4-HBA, 2,3-dihydroxybenzaldehyde or thymol significantly inhibited growth of A. alternata, at a much greater level than the fungicide alone. In addition, co-application of Folicur and 4-HBA resulted in a similar enhancement of antifungal activity against Fusarium culmorum. Lastly, we discovered that metabolites in the culture liquid of F. sambucinum biocontrol isolate FS-94 also had chemosensitizing activity, increasing S. nodorum sensitivity to Folicur and Dividend.

  14. Targeted regulation of MiR-98 on E2F1 increases chemosensitivity of leukemia cells K562/A02

    Directory of Open Access Journals (Sweden)

    Huang YD

    2017-06-01

    Full Text Available Yingdan Huang,1,2 Xiuli Hong,2 Jiasheng Hu,2 Quanyi Lu2 1Department of Blood Transfusion, The First Affiliated Hospital of Xiamen University, 2Department of Hematology, Zhongshan Hospital of Xiamen University, Xiamen, People’s Republic of China Background: miRNA is a microRNA that negatively regulates protein expression at post-transcriptional or translational level. It is widely involved in the pathogenesis of tumors. miR-98 belongs to the let-7 family, and its overexpression can increase the sensitivity to drugs in solid cancer cells. However, the function of miR-98 in leukemia is still unclear. In this study, the effect of miR-98 on drug resistance and proliferation of leukemia cells were investigated. Methods: Real-time quantitative polymerase chain reaction analyzed the expression difference between miR-98 and E2F1 in leukemia cell lines, K562 and K562/A02. The downstream target gene of miR-98 was predicted by TargetScan; K562/A02 was transiently transfected with miR-98 mimic to upregulate the expression of miR-98; real-time quantitative polymerase chain reaction and Western blot were used to analyze the expression alterations of E2F1; cell counting kit-8 was used to evaluate the influence on K562/A02 proliferation and sensitivity to chemotherapeutic drugs; meanwhile, Western blot was used to analyze the expression of p21, Bax, matrix metalloproteinase 9 and ABCG2 proteins. Results: E2F1 is one of the target genes of miR-98 proved by bioinformatics. Compared with the K562, the level of miRNA-98 expression was decreased in K562/A02, but the level of E2F1 expression was upregulated. Leukemia cell line K562/A02 was transfected with miR-98 mimic to upregulate the expression of miR-98, the expression of E2F1 was significantly decreased. After upregulating the miR-98 expression in K562/A02, the proliferation was weakened, and the sensitivity to chemotherapy was increased. Western blot showed that upregulated miR-98 expression increased the

  15. Mechanisms of decreased intestinal epithelial proliferation and increased apoptosis in murine acute lung injury.

    Science.gov (United States)

    Husain, Kareem D; Stromberg, Paul E; Woolsey, Cheryl A; Turnbull, Isaiah R; Dunne, W Michael; Javadi, Pardis; Buchman, Timothy G; Karl, Irene E; Hotchkiss, Richard S; Coopersmith, Craig M

    2005-10-01

    The aim of this study was to determine the effects of acute lung injury on the gut epithelium and examine mechanisms underlying changes in crypt proliferation and apoptosis. The relationship between severity and timing of lung injury to intestinal pathology was also examined. Randomized, controlled study. University research laboratory. Genetically inbred mice. Following induction of acute lung injury, gut epithelial proliferation and apoptosis were assessed in a) C3H/HeN wild-type and C3H/HeJ mice, which lack functional Toll-like receptor 4 (n = 17); b) C57Bl/6 mice that received monoclonal anti-tumor necrosis factor-alpha or control antibody (n = 22); and c) C57Bl/6 wild-type and transgenic mice that overexpress Bcl-2 in their gut epithelium (n = 21). Intestinal epithelial proliferation and death were also examined in animals with differing degrees of lung inflammation (n = 24) as well as in a time course analysis following a fixed injury (n = 18). Acute lung injury caused decreased proliferation and increased apoptosis in crypt epithelial cells in all animals studied. C3H/HeJ mice had higher levels of proliferation than C3H/HeN animals without additional changes in apoptosis. Anti-tumor necrosis factor-alpha antibody had no effect on gut epithelial proliferation or death. Overexpression of Bcl-2 did not change proliferation despite decreasing gut apoptosis. Proliferation and apoptosis were not correlated to severity of lung injury, as gut alterations were lost in mice with more severe acute lung injury. Changes in both gut epithelial proliferation and death were apparent within 12 hrs, but proliferation was decreased 36 hrs following acute lung injury while apoptosis returned to normal. Acute lung injury causes disparate effects on crypt proliferation and apoptosis, which occur, at least in part, through differing mechanisms involving Toll-like receptor 4 and Bcl-2. Severity of lung injury does not correlate with perturbations in proliferation or death in the

  16. Smac modulates chemosensitivity in head and neck cancer cells through the mitochondrial apoptotic pathway.

    Science.gov (United States)

    Sun, Quanhong; Zheng, Xingnan; Zhang, Lin; Yu, Jian

    2011-04-15

    Overexpression of inhibitors of apoptosis proteins (IAP) contributes to therapeutic resistance. Second mitochondria-derived activator of caspase (Smac) promotes caspase activation by binding to IAPs upon release from the mitochondria. IAP antagonists, also called SMAC mimetics, are promising anticancer agents modeled after this mechanism. We investigated the role and mechanisms of Smac- and Smac mimetic-mediated chemosensitization in head and neck squamous cell carcinoma (HNSCC) cells. The effects of SMAC knockdown, SMAC overexpression, and a small molecule Smac mimetic on the chemosensitivities of HNSCC cells were determined. The mechanisms of Smac- and Smac mimetic-mediated chemosensitization were investigated by analyzing growth suppression, the mitochondrial apoptotic pathway, caspase activation, and IAP proteins. The therapeutic responses of HNSCC cells with different levels of Smac were compared in xenograft models. We found that Smac mediates apoptosis induced by several classes of therapeutic agents through the mitochondrial pathway. SMAC knockdown led to impaired caspase activation, mitochondrial membrane depolarization, and release of cytochrome c. A small molecule Smac mimetic, at nanomolar concentrations, significantly sensitized HNSCC cells to gemcitabine-induced apoptosis and restored gemcitabine sensitivity in SMAC knockdown cells, through caspase activation, X-linked IAP dissociation, and mitochondria-associated events, but not the TNF-α pathway. Furthermore, Smac levels modulated the therapeutic response of HNSCC cells to gemcitabine in xenograft models. Our results establish a critical role of Smac in mediating therapeutic responses of HNSCC cells and provide a strong rationale for combining Smac mimetics with other anticancer agents to treat HNSCC. ©2011 AACR.

  17. Cancer causes increased mortality and is associated with altered apoptosis in murine sepsis.

    Science.gov (United States)

    Fox, Amy C; Robertson, Charles M; Belt, Brian; Clark, Andrew T; Chang, Katherine C; Leathersich, Ann M; Dominguez, Jessica A; Perrone, Erin E; Dunne, W Michael; Hotchkiss, Richard S; Buchman, Timothy G; Linehan, David C; Coopersmith, Craig M

    2010-03-01

    Whereas most septic patients have an underlying comorbidity, most animal models of sepsis use mice that were healthy before the onset of infection. Malignancy is the most common comorbidity associated with sepsis. The purpose of this study was to determine whether mice with cancer have a different response to sepsis than healthy animals. Prospective, randomized controlled study. Animal laboratory in a university medical center. C57Bl/6 mice. Animals received a subcutaneous injection of either 250,000 cells of the transplantable pancreatic adenocarcinoma cell line Pan02 (cancer) or phosphate-buffered saline (healthy). Three weeks later, mice given Pan02 cells had reproducible, nonmetastatic tumors. Both groups of mice then underwent intratracheal injection of either Pseudomonas aeruginosa (septic) or 0.9% NaCl (sham). Animals were killed 24 hrs postoperatively or followed-up 7 days for survival. Mice with cancer and healthy mice appeared similar when subjected to sham operation, although cancer animals had lower levels of T- and B-lymphocyte apoptosis. Septic mice with cancer had increased mortality compared to previously healthy septic mice subjected to the identical injury (52% vs. 28%; p = .04). This was associated with increased bacteremia but no difference in local pulmonary infection. Septic mice with cancer also had increased intestinal epithelial apoptosis. Although sepsis induced an increase in T- and B-lymphocyte apoptosis in all animals, septic mice with cancer had decreased T- and B-lymphocyte apoptosis compared to previously healthy septic mice. Serum and pulmonary cytokines, lung histology, complete blood counts, and intestinal proliferation were similar between septic mice with cancer and previously healthy septic mice. When subjected to the same septic insult, mice with cancer have increased mortality compared to previously healthy animals. Decreased systemic bacterial clearance and alterations in intestinal epithelial and lymphocyte apoptosis may

  18. Increased sensitivity of HPV-positive head and neck cancer cell lines to x-irradiation ± Cisplatin due to decreased expression of E6 and E7 oncoproteins and enhanced apoptosis.

    Science.gov (United States)

    Ziemann, Frank; Arenz, Andrea; Preising, Stefanie; Wittekindt, Claus; Klussmann, Jens P; Engenhart-Cabillic, Rita; Wittig, Andrea

    2015-01-01

    Squamous cell carcinoma of the head and neck region (HNSCC), which is related to an infection with human papilloma virus (HPV), responds better to simultaneous radio-chemotherapy with Cisplatin based regimens than HPV-negative tumors. The underlying molecular mechanisms for this clinical observation are not fully understood. Therefore, the response of four HPV-positive (HPV+) (UM-SCC-47, UM-SCC-104, 93-VU-147T, UPCI:SCC152) and four HPV-negative (HPV-) (UD-SCC-1, UM-SCC-6, UM-SCC-11b, UT-SCC-33) HNSCC cell lines to x-irradiation ± Cisplatin incubation in terms of clonogenic survival, cell cycle progression, protein expression (cyclin A2, cyclin E2, E6, E7, p53) and induction of apoptosis, was investigated. HPV+ cells were more radio- and chemosensitive and were more effectively sensitized to x-irradiation by simultaneous Cisplatin incubation than HPV- cell lines. HPV+ cell lines revealed an increased and prolonged G2/M arrest after irradiation, whereas Cisplatin induced a blockage of cells in S phase. In comparison to irradiation only, addition of Cisplatin significantly enhanced apoptosis especially in HPV+ cell lines. While irradiation alone increased the amount of HPV E6 and E7 proteins, both were down-regulated by Cisplatin incubation either alone or in combination with x-rays, which however did not increase the expression of endogenous p53. Our results demonstrate that cell cycle deregulation together with downregulation of HPV E6 and E7 proteins facilitating apoptosis after Cisplatin incubation promote the enhanced sensitivity of HPV+ HNSCC cells to simultaneous radio-chemotherapy. Combined effects of irradiation and Cisplatin appear to be relevant in mediating the enhanced therapeutic response of HPV-related HNSCC and are indicative of the benefit of combined modality approaches in future treatment optimization strategies.

  19. Gambogic Acid Lysinate Induces Apoptosis in Breast Cancer MCF-7 Cells by Increasing Reactive Oxygen Species

    Directory of Open Access Journals (Sweden)

    Yong-Zhan Zhen

    2015-01-01

    Full Text Available Gambogic acid (GA inhibits the proliferation of various human cancer cells. However, because of its water insolubility, the antitumor efficacy of GA is limited. Objectives. To investigate the antitumor activity of gambogic acid lysinate (GAL and its mechanism. Methods. Inhibition of cell proliferation was determined by MTT assay; intracellular ROS level was detected by staining cells with DCFH-DA; cell apoptosis was determined by flow cytometer and the mechanism of GAL was investigated by Western blot. Results. GAL inhibited the proliferation of MCF-7 cells with IC50 values 1.46 μmol/L comparable with GA (IC50, 1.16 μmol/L. GAL promoted the production of ROS; however NAC could remove ROS and block the effect of GAL. GAL inhibited the expression of SIRT1 but increased the phosphorylation of FOXO3a and the expression of p27Kip1. At knockdown of FOXO3a, cell apoptosis induced by GAL can be partly blocked. In addition it also enhanced the cleavage of caspase-3. Conclusions. GAL inhibited MCF-7 cell proliferation and induced MCF-7 cell apoptosis by increasing ROS level which could induce cell apoptosis by both SIRT1/FOXO3a/p27Kip1 and caspase-3 signal pathway. These results suggested that GAL might be useful as a modulation agent in cancer chemotherapy.

  20. Fish oil increases mitochondrial phospholipid unsaturation, upregulating reactive oxygen species and apoptosis in rat colonocytes

    Science.gov (United States)

    Hong, Mee Young; Chapkin, Robert S.; Barhoumi, Rola; Burghardt, Robert C.; Turner, Nancy D.; Henderson, Cara E.; Sanders, Lisa M.; Fan, Yang-Yi; Davidson, Laurie A.; Murphy, Mary E.; hide

    2002-01-01

    We have shown that a combination of fish oil (high in n-3 fatty acids) with the butyrate-producing fiber pectin, upregulates apoptosis in colon cells exposed to the carcinogen azoxymethane, protecting against colon tumor development. We now hypothesize that n-3 fatty acids prime the colonocytes such that butyrate can initiate apoptosis. To test this, 30 Sprague-Dawley rats were provided with diets differing in the fatty acid composition (corn oil, fish oil or a purified fatty acid ethyl ester diet). Intact colon crypts were exposed ex vivo to butyrate, and analyzed for reactive oxygen species (ROS), mitochondrial membrane potential (MMP), translocation of cytochrome C to the cytosol, and caspase-3 activity (early events in apoptosis). The fatty acid composition of the three major mitochondrial phospholipids was also determined, and an unsaturation index calculated. The unsaturation index in cardiolipin was correlated with ROS levels (R = 0.99; P = 0.02). When colon crypts from fish oil and FAEE-fed rats were exposed to butyrate, MMP decreased (P = 0.041); and translocation of cytochrome C to the cytosol (P = 0.037) and caspase-3 activation increased (P = 0.032). The data suggest that fish oil may prime the colonocytes for butyrate-induced apoptosis by enhancing the unsaturation of mitochondrial phospholipids, especially cardiolipin, resulting in an increase in ROS and initiating apoptotic cascade.

  1. Activation of Wnt/β-catenin signaling increases apoptosis in melanoma cells treated with trail.

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    Zachary F Zimmerman

    Full Text Available While the TRAIL pathway represents a promising therapeutic target in melanoma, resistance to TRAIL-mediated apoptosis remains a barrier to its successful adoption. Since the Wnt/β-catenin pathway has been implicated in facilitating melanoma cell apoptosis, we investigated the effect of Wnt/β-catenin signaling on regulating the responses of melanoma cells to TRAIL. Co-treatment of melanoma cell lines with WNT3A-conditioned media and recombinant TRAIL significantly enhanced apoptosis compared to treatment with TRAIL alone. This apoptosis correlates with increased abundance of the pro-apoptotic proteins BCL2L11 and BBC3, and with decreased abundance of the anti-apoptotic regulator Mcl1. We then confirmed the involvement of the Wnt/β-catenin signaling pathway by demonstrating that siRNA-mediated knockdown of an intracellular β-catenin antagonist, AXIN1, or treating cells with an inhibitor of GSK-3 also enhanced melanoma cell sensitivity to TRAIL. These studies describe a novel regulation of TRAIL sensitivity in melanoma by Wnt/β-catenin signaling, and suggest that strategies to enhance Wnt/β-catenin signaling in combination with TRAIL agonists warrant further investigation.

  2. Characterization of the chemosensitive response of individual solitary complex neurons from adult rats

    Science.gov (United States)

    Nichols, Nicole L.; Mulkey, Daniel K.; Wilkinson, Katherine A.; Powell, Frank L.; Dean, Jay B.; Putnam, Robert W.

    2009-01-01

    We studied the CO2/H+-chemosensitive responses of individual solitary complex (SC) neurons from adult rats by simultaneously measuring the intracellular pH (pHi) and electrical responses to hypercapnic acidosis (HA). SC neurons were recorded using the blind whole cell patch-clamp technique and loading the soma with the pH-sensitive dye pyranine through the patch pipette. We found that SC neurons from adult rats have a lower steady-state pHi than SC neurons from neonatal rats. In the presence of chemical and electrical synaptic blockade, adult SC neurons have firing rate responses to HA (percentage of neurons activated or inhibited and the magnitude of response as determined by the chemosensitivity index) that are similar to SC neurons from neonatal rats. They also have a typical response to isohydric hypercapnia, including decreased ΔpHi, followed by pHi recovery, and increased firing rate. Thus, the chemosensitive response of SC neurons from adults is similar to the chemosensitive response of SC neurons from neonatal rats. Because our findings for adults are similar to previously reported values for neurons from neonatal rats, we conclude that intrinsic chemosensitivity is established early in development for SC neurons and is maintained throughout adulthood. PMID:19144749

  3. Prevention of lymphocyte apoptosis in septic mice with cancer increases mortality.

    Science.gov (United States)

    Fox, Amy C; Breed, Elise R; Liang, Zhe; Clark, Andrew T; Zee-Cheng, Brendan R; Chang, Katherine C; Dominguez, Jessica A; Jung, Enjae; Dunne, W Michael; Burd, Eileen M; Farris, Alton B; Linehan, David C; Coopersmith, Craig M

    2011-08-15

    Lymphocyte apoptosis is thought to have a major role in the pathophysiology of sepsis. However, there is a disconnect between animal models of sepsis and patients with the disease, because the former use subjects that were healthy prior to the onset of infection while most patients have underlying comorbidities. The purpose of this study was to determine whether lymphocyte apoptosis prevention is effective in preventing mortality in septic mice with preexisting cancer. Mice with lymphocyte Bcl-2 overexpression (Bcl-2-Ig) and wild type (WT) mice were injected with a transplantable pancreatic adenocarcinoma cell line. Three weeks later, after development of palpable tumors, all animals received an intratracheal injection of Pseudomonas aeruginosa. Despite having decreased sepsis-induced T and B lymphocyte apoptosis, Bcl-2-Ig mice had markedly increased mortality compared with WT mice following P. aeruginosa pneumonia (85 versus 44% 7-d mortality; p = 0.004). The worsened survival in Bcl-2-Ig mice was associated with increases in Th1 cytokines TNF-α and IFN-γ in bronchoalveolar lavage fluid and decreased production of the Th2 cytokine IL-10 in stimulated splenocytes. There were no differences in tumor size or pulmonary pathology between Bcl-2-Ig and WT mice. To verify that the mortality difference was not specific to Bcl-2 overexpression, similar experiments were performed in Bim(-/-) mice. Septic Bim(-/-) mice with cancer also had increased mortality compared with septic WT mice with cancer. These data demonstrate that, despite overwhelming evidence that prevention of lymphocyte apoptosis is beneficial in septic hosts without comorbidities, the same strategy worsens survival in mice with cancer that are given pneumonia.

  4. Increased autophagy and apoptosis contribute to muscle atrophy in a myotonic dystrophy type 1 Drosophila model

    Directory of Open Access Journals (Sweden)

    Ariadna Bargiela

    2015-07-01

    Full Text Available Muscle mass wasting is one of the most debilitating symptoms of myotonic dystrophy type 1 (DM1 disease, ultimately leading to immobility, respiratory defects, dysarthria, dysphagia and death in advanced stages of the disease. In order to study the molecular mechanisms leading to the degenerative loss of adult muscle tissue in DM1, we generated an inducible Drosophila model of expanded CTG trinucleotide repeat toxicity that resembles an adult-onset form of the disease. Heat-shock induced expression of 480 CUG repeats in adult flies resulted in a reduction in the area of the indirect flight muscles. In these model flies, reduction of muscle area was concomitant with increased apoptosis and autophagy. Inhibition of apoptosis or autophagy mediated by the overexpression of DIAP1, mTOR (also known as Tor or muscleblind, or by RNA interference (RNAi-mediated silencing of autophagy regulatory genes, achieved a rescue of the muscle-loss phenotype. In fact, mTOR overexpression rescued muscle size to a size comparable to that in control flies. These results were validated in skeletal muscle biopsies from DM1 patients in which we found downregulated autophagy and apoptosis repressor genes, and also in DM1 myoblasts where we found increased autophagy. These findings provide new insights into the signaling pathways involved in DM1 disease pathogenesis.

  5. Cinnabar-Induced Subchronic Renal Injury Is Associated with Increased Apoptosis in Rats

    Directory of Open Access Journals (Sweden)

    Ying Wang

    2015-01-01

    Full Text Available The aim of this study was to explore the role of apoptosis in cinnabar-induced renal injury in rats. To test this role, rats were dosed orally with cinnabar (1 g/kg/day for 8 weeks or 12 weeks, and the control rats were treated with 5% carboxymethylcellulose solution. Levels of urinary mercury (UHg, renal mercury (RHg, serum creatinine (SCr, and urine kidney injury molecule 1 (KIM-1 were assessed, and renal pathology was analyzed. Apoptotic cells were identified and the apoptotic index was calculated. A rat antibody array was used to analyze expression of cytokines associated with apoptosis. Results from these analyses showed that UHg, RHg, and urine KIM-1, but not SCr, levels were significantly increased in cinnabar-treated rats. Renal pathological changes in cinnabar-treated rats included vacuolization of tubular cells, formation of protein casts, infiltration of inflammatory cells, and increase in the number of apoptotic tubular cells. In comparison to the control group, expression of FasL, Fas, TNF-α, TRAIL, activin A, and adiponectin was upregulated in the cinnabar-treated group. Collectively, our results suggest that prolonged use of cinnabar results in kidney damage due to accumulation of mercury and that the underlying mechanism involves apoptosis of tubular cells via a death receptor-mediated pathway.

  6. Depletion of cdc-25.3, a Caenorhabditis elegans orthologue of cdc25, increases physiological germline apoptosis.

    Science.gov (United States)

    Sung, Minhee; Kawasaki, Ichiro; Shim, Yhong-Hee

    2017-07-01

    In Caenorhabditis elegans hermaphrodites, physiological germline apoptosis is higher in cdc-25.3 mutants than in wild-type. The elevated germline apoptosis in cdc-25.3 mutants seems to be induced by accumulation of double-stranded DNA breaks (DSBs). Both DNA damage and synapsis checkpoint genes are required to increase the germline apoptosis. Notably, the number of germ cells that lose P-granule components, PGL-1 and PGL-3, increase in cdc-25.3 mutants, and the increase in germline apoptosis requires the activity of SIR-2.1, a Sirtuin orthologue. These results suggest that elevation of germline apoptosis in cdc-25.3 mutants is induced by accumulation of DSBs, leading to a loss of PGL-1 and PGL-3 in germ cells, which promotes cytoplasmic translocation of SIR-2.1, and finally activates the core apoptotic machinery. © 2017 Federation of European Biochemical Societies.

  7. Murine Lung Cancer Increases CD4+ T Cell Apoptosis and Decreases Gut Proliferative Capacity in Sepsis.

    Science.gov (United States)

    Lyons, John D; Mittal, Rohit; Fay, Katherine T; Chen, Ching-Wen; Liang, Zhe; Margoles, Lindsay M; Burd, Eileen M; Farris, Alton B; Ford, Mandy L; Coopersmith, Craig M

    2016-01-01

    Mortality is significantly higher in septic patients with cancer than in septic patients without a history of cancer. We have previously described a model of pancreatic cancer followed by sepsis from Pseudomonas aeruginosa pneumonia in which cancer septic mice have higher mortality than previously healthy septic mice, associated with increased gut epithelial apoptosis and decreased T cell apoptosis. The purpose of this study was to determine whether this represents a common host response by creating a new model in which both the type of cancer and the model of sepsis are altered. C57Bl/6 mice received an injection of 250,000 cells of the lung cancer line LLC-1 into their right thigh and were followed three weeks for development of palpable tumors. Mice with cancer and mice without cancer were then subjected to cecal ligation and puncture and sacrificed 24 hours after the onset of sepsis or followed 7 days for survival. Cancer septic mice had a higher mortality than previously healthy septic mice (60% vs. 18%, p = 0.003). Cancer septic mice had decreased number and frequency of splenic CD4+ lymphocytes secondary to increased apoptosis without changes in splenic CD8+ numbers. Intestinal proliferation was also decreased in cancer septic mice. Cancer septic mice had a higher bacterial burden in the peritoneal cavity, but this was not associated with alterations in local cytokine, neutrophil or dendritic cell responses. Cancer septic mice had biochemical evidence of worsened renal function, but there was no histologic evidence of renal injury. Animals with cancer have a significantly higher mortality than previously healthy animals following sepsis. The potential mechanisms associated with this elevated mortality differ significantly based upon the model of cancer and sepsis utilized. While lymphocyte apoptosis and intestinal integrity are both altered by the combination of cancer and sepsis, the patterns of these alterations vary greatly depending on the models used.

  8. Murine Lung Cancer Increases CD4+ T Cell Apoptosis and Decreases Gut Proliferative Capacity in Sepsis.

    Directory of Open Access Journals (Sweden)

    John D Lyons

    Full Text Available Mortality is significantly higher in septic patients with cancer than in septic patients without a history of cancer. We have previously described a model of pancreatic cancer followed by sepsis from Pseudomonas aeruginosa pneumonia in which cancer septic mice have higher mortality than previously healthy septic mice, associated with increased gut epithelial apoptosis and decreased T cell apoptosis. The purpose of this study was to determine whether this represents a common host response by creating a new model in which both the type of cancer and the model of sepsis are altered.C57Bl/6 mice received an injection of 250,000 cells of the lung cancer line LLC-1 into their right thigh and were followed three weeks for development of palpable tumors. Mice with cancer and mice without cancer were then subjected to cecal ligation and puncture and sacrificed 24 hours after the onset of sepsis or followed 7 days for survival.Cancer septic mice had a higher mortality than previously healthy septic mice (60% vs. 18%, p = 0.003. Cancer septic mice had decreased number and frequency of splenic CD4+ lymphocytes secondary to increased apoptosis without changes in splenic CD8+ numbers. Intestinal proliferation was also decreased in cancer septic mice. Cancer septic mice had a higher bacterial burden in the peritoneal cavity, but this was not associated with alterations in local cytokine, neutrophil or dendritic cell responses. Cancer septic mice had biochemical evidence of worsened renal function, but there was no histologic evidence of renal injury.Animals with cancer have a significantly higher mortality than previously healthy animals following sepsis. The potential mechanisms associated with this elevated mortality differ significantly based upon the model of cancer and sepsis utilized. While lymphocyte apoptosis and intestinal integrity are both altered by the combination of cancer and sepsis, the patterns of these alterations vary greatly depending on

  9. Knockdown of UbcH10 enhances the chemosensitivity of dual drug resistant breast cancer cells to epirubicin and docetaxel.

    Science.gov (United States)

    Wang, Cheng; Pan, Yun-Hao; Shan, Ming; Xu, Ming; Bao, Jia-Lin; Zhao, Li-Ming

    2015-03-02

    Breast cancer is one of the most common and lethal cancers in women. As a hub gene involved in a diversity of tumors, the ubiquitin-conjugating enzyme H10 (UbcH10), may also play some roles in the genesis and development of breast cancer. In the current study, we found that the expression of UbcH10 was up-regulated in some breast cancer tissues and five cell lines. We established a dual drug resistant cell line MCF-7/EPB (epirubicin)/TXT (docetaxel) and a lentiviral system expressing UbcH10 shRNA to investigate the effects of UbcH10 knockdown on the chemosensitivity of MCF-7/EPB/TXT cells to epirubicin and docetaxel. The knockdown of UbcH10 inhibited the proliferation of both MCF-7 and MCF-7/EPB/TXT cells, due to the G1 phase arrest in cell cycle. Furthermore, UbcH10 knockdown increased the sensitivity of MCF-7/EPB/TXT cells to epirubicin and docetaxel and promoted the apoptosis induced by these two drugs. Protein detection showed that, in addition to inhibiting the expression of Ki67 and cyclin D1, UbcH10 RNAi also impaired the increased BCL-2 and MDR-1 expression levels in MCF-7/EPB/TXT cells, which may contribute to abating the drug resistance in the breast cancer cells. Our research in the current study demonstrated that up-regulation of UbcH10 was involved in breast cancer and its knockdown can inhibit the growth of cancer cells and increase the chemosensitivity of the dual drug resistant breast cancer cells to epirubicin and docetaxel, suggesting that UbcH10 may be a promising target for the therapy of breast cancer.

  10. Knockdown of UbcH10 Enhances the Chemosensitivity of Dual Drug Resistant Breast Cancer Cells to Epirubicin and Docetaxel

    Directory of Open Access Journals (Sweden)

    Cheng Wang

    2015-03-01

    Full Text Available Breast cancer is one of the most common and lethal cancers in women. As a hub gene involved in a diversity of tumors, the ubiquitin-conjugating enzyme H10 (UbcH10, may also play some roles in the genesis and development of breast cancer. In the current study, we found that the expression of UbcH10 was up-regulated in some breast cancer tissues and five cell lines. We established a dual drug resistant cell line MCF-7/EPB (epirubicin/TXT (docetaxel and a lentiviral system expressing UbcH10 shRNA to investigate the effects of UbcH10 knockdown on the chemosensitivity of MCF-7/EPB/TXT cells to epirubicin and docetaxel. The knockdown of UbcH10 inhibited the proliferation of both MCF-7 and MCF-7/EPB/TXT cells, due to the G1 phase arrest in cell cycle. Furthermore, UbcH10 knockdown increased the sensitivity of MCF-7/EPB/TXT cells to epirubicin and docetaxel and promoted the apoptosis induced by these two drugs. Protein detection showed that, in addition to inhibiting the expression of Ki67 and cyclin D1, UbcH10 RNAi also impaired the increased BCL-2 and MDR-1 expression levels in MCF-7/EPB/TXT cells, which may contribute to abating the drug resistance in the breast cancer cells. Our research in the current study demonstrated that up-regulation of UbcH10 was involved in breast cancer and its knockdown can inhibit the growth of cancer cells and increase the chemosensitivity of the dual drug resistant breast cancer cells to epirubicin and docetaxel, suggesting that UbcH10 may be a promising target for the therapy of breast cancer.

  11. Nicotine-induced embryonic malformations mediated by apoptosis from increasing intracellular calcium and oxidative stress.

    Science.gov (United States)

    Zhao, Zhiyong; Reece, E Albert

    2005-10-01

    Tobacco smoking by women during pregnancy increases the risk of congenital birth defects in the infants. Among the smoke products, nicotine is believed to be the major teratogenic factor that perturbs embryonic development. However, the role of nicotine in embryonic malformations has not been addressed, and the mechanisms by which nicotine affects embryonic development remain to be delineated. To investigate the effects of nicotine on early embryogenesis, murine embryos at embryonic day (E) 8.5 were dissected out of the uteri, cultured in a roller bottle system, and treated with nicotine (0.6-6 microM) or vehicle. Embryonic morphogenesis and growth were examined in terms of structural morphology and crown/rump length, respectively. Programmed cell death (apoptosis) was assessed using LysoTracker Red staining of whole mount embryos and TUNEL assay of tissue sections. Changes in intracellular calcium concentration ([Ca2+]i) and reactive oxygen species (ROS) production were assessed using fluorescent dyes (Flu-4, AM; H2DCFDA, respectively) under a confocal microscope. To further investigate the role of intracellular calcium and ROS in nicotine-induced embryopathy, embryos were treated with BAPTA-AM (2 microM) to inhibit [Ca2+]i elevation and ascorbic acid (vitamin C; 100 microg/ml) to scavenge ROS in presence of nicotine (6 microM). The embryos treated with nicotine in 3-6 microM were smaller than those treated with vehicle. Most of the embryos had open neural tube in the anterior (brain) regions. The embryos treated with 6 microM nicotine also exhibited severe defects in the posterior trunk, resembling caudal dysplasia. Excessive apoptosis was observed in the deformed structures. Significant increases in [Ca2+]i and ROS were seen in the tissues that had higher levels of apoptosis. Furthermore, inhibition of [Ca2+]i and scavenging of ROS significantly reduced embryonic malformation and apoptotic rates in the embryos. Nicotine affects embryonic development in a

  12. MicroRNA-31 Regulates Chemosensitivity in Malignant Pleural Mesothelioma

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    Hannah L. Moody

    2017-09-01

    Full Text Available Malignant pleural mesothelioma (MPM is associated with an extremely poor prognosis, and most patients initially are or rapidly become unresponsive to platinum-based chemotherapy. MicroRNA-31 (miR-31 is encoded on a genomic fragile site, 9p21.3, which is reportedly lost in many MPM tumors. Based on previous findings in a variety of other cancers, we hypothesized that miR-31 alters chemosensitivity and that miR-31 reconstitution may influence sensitivity to chemotherapeutics in MPM. Reintroduction of miR-31 into miR-31 null NCI-H2452 cells significantly enhanced clonogenic resistance to cisplatin and carboplatin. Although miR-31 re-expression increased chemoresistance, paradoxically, a higher relative intracellular accumulation of platinum was detected. This was coupled to a significantly decreased intranuclear concentration of platinum. Linked with a downregulation of OCT1, a bipotential transcriptional regulator with multiple miR-31 target binding sites, we subsequently identified an indirect miR-31-mediated upregulation of ABCB9, a transporter associated with drug accumulation in lysosomes, and increased uptake of platinum to lysosomes. However, when overexpressed directly, ABCB9 promoted cellular chemosensitivity, suggesting that miR-31 promotes chemoresistance largely via an ABCB9-independent mechanism. Overall, our data suggest that miR-31 loss from MPM tumors promotes chemosensitivity and may be prognostically beneficial in the context of therapeutic sensitivity. Keywords: malignant pleural mesothelioma, microRNA-31, chemoresistance, cisplatin, ABCB9

  13. Paclitaxel stimulates chromosomal fusion and instability in cells with dysfunctional telomeres: Implication in multinucleation and chemosensitization

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jeong-Eun [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Woo, Seon Rang [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Department of Biochemistry, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Kang, Chang-Mo [Laboratory of Cytogenetics and Tissue Regeneration, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Juhn, Kyoung-Mi; Ju, Yeun-Jin; Shin, Hyun-Jin; Joo, Hyun-Yoo; Park, Eun Ran; Park, In-chul; Hong, Sung Hee; Hwang, Sang-Gu [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Lee, Jung-Kee [Department of Life Science and Genetic Engineering, Paichai University, Daejeon 302-735 (Korea, Republic of); Kim, Hae Kwon [Department of Biotechnology, Seoul Woman' s University, Seoul 139-774 (Korea, Republic of); Cho, Myung-Haing [Laboratory of Toxicology, College of Veterinary Medicine, Seoul National University, Seoul 151-74-2 (Korea, Republic of); Park, Gil Hong [Department of Biochemistry, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Lee, Kee-Ho, E-mail: khlee@kirams.re.kr [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of)

    2011-01-14

    Research highlights: {yields} Paclitaxel serves as a stimulator of chromosomal fusion in cells in which telomeres are dysfunctional. {yields} Typical fusions involve p-arms, but paclitaxel-induced fusions occur between both q- and p-arms. {yields} Paclitaxel-stimulated fusions in cells in which telomeres are dysfunctional evoke prolonged G2/M cell cycle arrest and delay multinucleation. {yields} Upon telomere erosion, paclitaxel promotes chromosomal instability and subsequent apoptosis. {yields} Chromosomal fusion enhances paclitaxel chemosensitivity under telomere dysfunction. -- Abstract: The anticancer effect of paclitaxel is attributable principally to irreversible promotion of microtubule stabilization and is hampered upon development of chemoresistance by tumor cells. Telomere shortening, and eventual telomere erosion, evoke chromosomal instability, resulting in particular cellular responses. Using telomerase-deficient cells derived from mTREC-/-p53-/- mice, here we show that, upon telomere erosion, paclitaxel propagates chromosomal instability by stimulating chromosomal end-to-end fusions and delaying the development of multinucleation. The end-to-end fusions involve both the p- and q-arms in cells in which telomeres are dysfunctional. Paclitaxel-induced chromosomal fusions were accompanied by prolonged G2/M cell cycle arrest, delayed multinucleation, and apoptosis. Telomere dysfunctional cells with mutlinucleation eventually underwent apoptosis. Thus, as telomere erosion proceeds, paclitaxel stimulates chromosomal fusion and instability, and both apoptosis and chemosensitization eventually develop.

  14. Recombinant human erythropoietin increases survival and reduces neuronal apoptosis in a murine model of cerebral malaria

    DEFF Research Database (Denmark)

    Wiese, Lothar; Hempel, Casper; Penkowa, Milena

    2008-01-01

    BACKGROUND: Cerebral malaria (CM) is an acute encephalopathy with increased pro-inflammatory cytokines, sequestration of parasitized erythrocytes and localized ischaemia. In children CM induces cognitive impairment in about 10% of the survivors. Erythropoietin (Epo) has - besides of its well known...... with recombinant human Epo (rhEpo; 50-5000 U/kg/OD, i.p.) at different time points. The effect on survival was measured. Brain pathology was investigated by TUNEL (Terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-digoxigenin nick end labelling), as a marker of apoptosis. Gene...

  15. Differential role of RIP1 in Smac mimetic-mediated chemosensitization of neuroblastoma cells.

    Science.gov (United States)

    Czaplinski, Sebastian; Abhari, Behnaz Ahangarian; Torkov, Alica; Seggewiß, Dominik; Hugle, Manuela; Fulda, Simone

    2015-12-08

    We explored the potential of Smac mimetics, which antagonize Inhibitor of Apoptosis (IAP) proteins, for chemosensitization of neuroblastoma (NB). Here, we report that Smac mimetics, e.g. BV6, prime NB cells for chemotherapeutics including the topoisomerase II inhibitor doxorubicin (DOX) and vinca alkaloids such as Vincristine (VCR), Vinblastine (VBL) and Vinorelbine (VNR). Additionally, BV6 acts in concert with DOX or VCR to suppress long-term clonogenic growth. While BV6 causes rapid downregulation of cellular IAP (cIAP)1 protein and nuclear factor-kappaB (NF-κB) activation, DOX/BV6- or VCR/BV6-induced apoptosis occurs independently of NF-κB or TNFα signaling, since overexpression of dominant-negative IκBα superrepressor or the Tumor Necrosis Factor (TNF)α-blocking antibody Enbrel fail to block cell death. Mechanistic studies reveal that Receptor-interacting protein (RIP)1 is required for DOX/BV6-, but not for VCR/BV6-induced apoptosis, since transient or stable knockdown of RIP1 or the pharmacological RIP1 inhibitor necrostatin-1 significantly reduce apoptosis. By comparison, VCR/BV6-mediated apoptosis critically depends on the mitochondrial pathway. VCR/BV6 cotreatment causes phosphorylation of BCL-2 during mitotic arrest, enhanced activation of BAX and BAK and loss of mitochondrial membrane potential (MMP). Additionally, overexpression of BCL-2 profoundly suppresses VCR/BV6-induced apoptosis. Thus, BV6 sensitizes NB cells to chemotherapy-induced apoptosis via distinct initial signaling mechanisms depending on the chemotherapeutic drug. These findings provide novel mechanistic insights into Smac mimetic-mediated chemosensitization of NB.

  16. Kaempferol increases apoptosis in human acute promyelocytic leukemia cells and inhibits multidrug resistance genes.

    Science.gov (United States)

    Moradzadeh, Maliheh; Tabarraei, Alijan; Sadeghnia, Hamid Reza; Ghorbani, Ahmad; Mohamadkhani, Ashraf; Erfanian, Saiedeh; Sahebkar, Amirhossein

    2018-02-01

    Acute promyelocytic leukemia (APL) is one of the most life-threatening hematological malignancies. Defects in the cell growth and apoptotic pathways are responsible for both disease pathogenesis and treatment resistance. Therefore, pro-apoptotic agents are potential candidates for APL treatment. Kaempferol is a flavonoid with antioxidant and anti-tumor properties. This study was designed to investigate the cytotoxic, pro-apoptotic, and differentiation-inducing effects of kaempferol on HL-60 and NB4 leukemia cells. Resazurin assay was used to determine cell viability following treatment with kaempferol (12.5-100 μM) and all-trans retinoic acid (ATRA; 10 μM; used as a positive control). Apoptosis and differentiation were also detected using propidium iodide and NBT staining techniques, respectively. Furthermore, the expression levels of genes involved in apoptosis (PI3 K, AKT, BCL2, BAX, p53, p21, PTEN, CASP3, CASP8, and CASP9), differentiation (PML-RAR and HDAC1), and multi-drug resistance (ABCB1 and ABCC1) were determined using quantitative real-time PCR. The protein expressions of Bax/Bcl2 and casp3 were confirmed using Western blot. The results showed that kaempferol decreased cell viability and increased subG1 population in the tested leukemic cells. This effect was associated with decreased expression of Akt, BCL2, ABCB1, and ABCC1 genes, while the expression of CASP3 and BAX/BCL-2 ratio were significantly increased at both gene and protein levels. Kaempferol promoted apoptosis and inhibited multidrug resistance in a concentration-dependent manner, without any differential effect on leukemic cells. In conclusion, this study suggested that kaempferol may be utilized as an appropriate alternative for ATRA in APL patients. © 2017 Wiley Periodicals, Inc.

  17. Increased cell surface metallopeptidase activity in cells undergoing UV-induced apoptosis

    International Nuclear Information System (INIS)

    Piva, T.J.; Davern, C.M.; Ellem, K.A.O.

    1999-01-01

    Full text: We have previously shown that UVC irradiation activated a range of cell surface peptidases (CSP) in HeLa cell monolayer cultures 20 h post-irradiation (1). In cells undergoing apoptosis there is an increase in CSP activity compared to control viable cells in cultures which have been treated by a wide range of agents including UV-irradiation (2). In order to further understand the mechanism involved in this process, we induced apoptosis in HeLa cells using 500 Jm -2 UVB. The separation of viable, apoptotic and necrotic cells of irradiated HeLa cell cultures was made by FACS analysis and sorting. The three populations were distinguished by their staining with PI and Hoechst 33342 dyes. CSP activity was measured using the P9 assay developed in this laboratory (1-3). The viable fraction of the irradiated cells had a higher level of CSP activity compared to unirradiated controls. The level of CSP activity in the apoptotic fraction was higher than that of the viable fraction, however that of the necrotic fraction was significantly lower. This finding agreed with that seen in UVC-irradiated (50 Jm -2 ) cultures (2). In order to elucidate the mechanism by which CSP activity was increased in UVB-irradiated cells undergoing apoptosis, the cultures were treated with the following agents: bestatin, aminopeptidase inhibitor, DEVD, caspase 3 inhibitor, and 3-aminobenzamide (3AB), PARP activation inhibitor. Bestatin and DEVD did not affect the level of CSP activity in the different cell subpopulations following UVB-irradiation. Treatment with 3AB abolished the increased CSP activity seen in the viable and apoptotic fraction following UVB-irradiation. All treated cells had the same morphology as observed under EM. The degree of phosphatidylserine eversion on the cell membrane was similar as were the cleavage profiles of PARP and actin. Only DEVD-treated cells had reduced caspase 3 activity which confirmed that the activation of CSP activity in apoptotic cells is

  18. Macroautophagy-generated increase of lysosomal amyloid β-protein mediates oxidant-induced apoptosis of cultured neuroblastoma cells

    DEFF Research Database (Denmark)

    Zheng, Lin; Terman, Alexei; Hallbeck, Martin

    2011-01-01

    and accumulation of Aβ within lysosomes, induced apoptosis in differentiated SH-SY5Y neuroblastoma cells. Cells under hyperoxia showed: (1) increased numbers of autophagic vacuoles that contained amyloid precursor protein (APP) as well as Aβ monomers and oligomers, (2) increased reactive oxygen species production......, and (3) enhanced apoptosis. Oxidant-induced apoptosis positively correlated with cellular Aβ production, being the highest in cells that were stably transfected with APP Swedish KM670/671NL double mutation. Inhibition of γ-secretase, prior and/or in parallel to hyperoxia, suggested that the increase...... and resulting lysosomal Aβ accumulation are essential for oxidant-induced apoptosis in cultured neuroblastoma cells and provide additional support for the interactive role of oxidative stress and the lysosomal system in AD-related neurodegeneration....

  19. Dienogest reduces proliferation, aromatase expression and angiogenesis, and increases apoptosis in human endometriosis.

    Science.gov (United States)

    Miyashita, Mariko; Koga, Kaori; Takamura, Masashi; Izumi, Gentaro; Nagai, Miwako; Harada, Miyuki; Hirata, Tetsuya; Hirota, Yasushi; Fujii, Tomoyuki; Osuga, Yutaka

    2014-09-01

    Dienogest is a novel progestin that is highly selective for progesterone receptors and inhibits endometriosis. However, it remains unknown how the administration of dienogest to patients with endometriosis impacts on their lesion tissues. The aim of this study was to evaluate the in vivo effect of dienogest on endometriosis tissue. We collected endometrioma tissues from patients treated with dienogest (N = 7) or not treated (N = 11, controls). Cell proliferation, aromatase expression and blood vessel density were evaluated by staining for Ki67, aromatase and the von Willebrand factor, respectively. Apoptosis was detected using the TUNEL assay. The proportion of Ki67 and aromatase positive epithelial cells was significantly lower in the dienogest group than in controls (p dienogest group (p dienogest group compared with controls (p = 0.20). Our study demonstrates that endometrioma taken from patients treated with dienogest show remarkable histological features such as reduction of proliferation, aromatase expression and angiogenesis, and increase of apoptosis. This study clarified the impact of dienogest on local histological events that explain its therapeutic effect on endometriosis.

  20. Inhibition of apoptosis signal-regulating kinase 1 enhances endochondral bone formation by increasing chondrocyte survival

    Science.gov (United States)

    Eaton, G J; Zhang, Q-S; Diallo, C; Matsuzawa, A; Ichijo, H; Steinbeck, M J; Freeman, T A

    2014-01-01

    Endochondral ossification is the result of chondrocyte differentiation, hypertrophy, death and replacement by bone. The careful timing and progression of this process is important for normal skeletal bone growth and development, as well as fracture repair. Apoptosis Signal-Regulating Kinase 1 (ASK1) is a mitogen-activated protein kinase (MAPK), which is activated by reactive oxygen species and other cellular stress events. Activation of ASK1 initiates a signaling cascade known to regulate diverse cellular events including cytokine and growth factor signaling, cell cycle regulation, cellular differentiation, hypertrophy, survival and apoptosis. ASK1 is highly expressed in hypertrophic chondrocytes, but the role of ASK1 in skeletal tissues has not been investigated. Herein, we report that ASK1 knockout (KO) mice display alterations in normal growth plate morphology, which include a shorter proliferative zone and a lengthened hypertrophic zone. These changes in growth plate dynamics result in accelerated long bone mineralization and an increased formation of trabecular bone, which can be attributed to an increased resistance of terminally differentiated chondrocytes to undergo cell death. Interestingly, under normal cell culture conditions, mouse embryonic fibroblasts (MEFs) derived from ASK1 KO mice show no differences in either MAPK signaling or osteogenic or chondrogenic differentiation when compared with wild-type (WT) MEFs. However, when cultured with stress activators, H2O2 or staurosporine, the KO cells show enhanced survival, an associated decrease in the activation of proteins involved in death signaling pathways and a reduction in markers of terminal differentiation. Furthermore, in both WT mice treated with the ASK1 inhibitor, NQDI-1, and ASK1 KO mice endochondral bone formation was increased in an ectopic ossification model. These findings highlight a previously unrealized role for ASK1 in regulating endochondral bone formation. Inhibition of ASK1 has

  1. SOMATIC VERSUS DENDRITIC RESPONSES TO HYPERCAPNIA IN CHEMOSENSITIVE LOCUS COERULEUS NEURONS FROM NEONATAL RATS

    Science.gov (United States)

    Ritucci, Nick A.; Dean, Jay B.; Putnam, Robert W.

    2005-01-01

    Cardiorespiratory control is mediated in part by central chemosensitive neurons that respond to increased CO2 (hypercapnia). Activation of these neurons is believed to involve hypercapnia-induced decreases in intracellular pH (pHi). All previous measurements of hypercapnia-induced pHi changes in chemosensitive neurons have been made from the soma, but chemosensitive signaling could be initiated in the dendrites of these neurons. In this study, membrane potential (Vm) and pHi were measured simultaneously in chemosensitive locus coeruleus (LC) neurons from neonatal rat brainstem slices using whole-cell pipettes and the pH-sensitive fluorescent dye pyranine. We measured pHi from the soma as well as from primary dendrites to a distance of 160 μm from the edge of the soma. Hypercapnia (15% CO2, pHo 7.00; control: 5% CO2, pHo 7.45) resulted in an acidification of similar magnitude in dendrites and soma (about 0.26 pH unit), but that was faster in the more distal regions of the dendrites. Neither the dendrites nor the soma exhibited pHi recovery during hypercapnia-induced acidification, but both regions contain pH-regulating transporters since they exhibit pHi recovery from an NH4Cl prepulse-induced acidification (at constant pHo 7.45). Exposing a portion of the dendrites to hypercapnic solution did not increase firing rate, but exposing the soma to hypercapnic solution resulted in a near maximal increase in firing rate. These data show that while the pHi response to hypercapnia is similar in the dendrites and soma, somatic exposure to hypercapnia plays a major role in the activation of chemosensitive LC neurons from neonatal rats. PMID:16014703

  2. Transient p53 suppression increases reprogramming of human fibroblasts without affecting apoptosis and DNA damage

    DEFF Research Database (Denmark)

    Rasmussen, Mikkel Aabech; Holst, Bjørn; Tümer, Zeynep

    2014-01-01

    The discovery of human-induced pluripotent stem cells (iPSCs) has sparked great interest in the potential treatment of patients with their own in vitro differentiated cells. Recently, knockout of the Tumor Protein 53 (p53) gene was reported to facilitate reprogramming but unfortunately also led...... to genomic instability. Here, we report that transient suppression of p53 during nonintegrative reprogramming of human fibroblasts leads to a significant increase in expression of pluripotency markers and overall number of iPSC colonies, due to downstream suppression of p21, without affecting apoptosis...... and DNA damage. Stable iPSC lines generated with or without p53 suppression showed comparable expression of pluripotency markers and methylation patterns, displayed normal karyotypes, contained between 0 and 5 genomic copy number variations and produced functional neurons in vitro. In conclusion...

  3. Spaceflight and clinorotation cause cytoskeleton and mitochondria changes and increases in apoptosis in cultured cells

    Science.gov (United States)

    Schatten, H.; Lewis, M. L.; Chakrabarti, A.

    2001-01-01

    The cytoskeleton is a complex network of fibers that is sensitive to environmental factors including microgravity and altered gravitational forces. Cellular functions such as transport of cell organelles depend on cytoskeletal integrity; regulation of cytoskeletal activity plays a role in cell maintenance, cell division, and apoptosis. Here we report cytoskeletal and mitochondria alterations in cultured human lymphocyte (Jurkat) cells after exposure to spaceflight and in insect cells of Drosophila melanogaster (Schneider S-1) after exposure to conditions created by clinostat rotation. Jurkat cells were flown on the space shuttle in Biorack cassettes while Schneider S-1 cells were exposed to altered gravity forces as produced by clinostat rotation. The effects of both treatments were similar in the different cell types. Fifty percent of cells displayed effects on the microtubule network in both cell lines. Under these experimental conditions mitochondria clustering and morphological alterations of mitochondrial cristae was observed to various degrees after 4 and 48 hours of culture. Jurkat cells underwent cell divisions during exposure to spaceflight but a large number of apoptotic cells was also observed. Similar results were obtained in Schneider S-1 cells cultured under clinostat rotation. Both cell lines displayed mitochondria abnormalities and mitochondria clustering toward one side of the cells which is interpreted to be the result of microtubule disruption and failure of mitochondria transport along microtubules. The number of mitochondria was increased in cells exposed to altered gravity while cristae morphology was severely affected indicating altered mitochondria function. These results show that spaceflight as well as altered gravity produced by clinostat rotation affects microtubule and mitochondria organization and results in increases in apoptosis. Grant numbers: NAG 10-0224, NAG2-985. c 2001. Elsevier Science Ltd. All rights reserved.

  4. Disruption of the Cx43/miR21 pathway leads to osteocyte apoptosis and increased osteoclastogenesis with aging.

    Science.gov (United States)

    Davis, Hannah M; Pacheco-Costa, Rafael; Atkinson, Emily G; Brun, Lucas R; Gortazar, Arancha R; Harris, Julia; Hiasa, Masahiro; Bolarinwa, Surajudeen A; Yoneda, Toshiyuki; Ivan, Mircea; Bruzzaniti, Angela; Bellido, Teresita; Plotkin, Lilian I

    2017-06-01

    Skeletal aging results in apoptosis of osteocytes, cells embedded in bone that control the generation/function of bone forming and resorbing cells. Aging also decreases connexin43 (Cx43) expression in bone; and osteocytic Cx43 deletion partially mimics the skeletal phenotype of old mice. Particularly, aging and Cx43 deletion increase osteocyte apoptosis, and osteoclast number and bone resorption on endocortical bone surfaces. We examined herein the molecular signaling events responsible for osteocyte apoptosis and osteoclast recruitment triggered by aging and Cx43 deficiency. Cx43-silenced MLO-Y4 osteocytic (Cx43 def ) cells undergo spontaneous cell death in culture through caspase-3 activation and exhibit increased levels of apoptosis-related genes, and only transfection of Cx43 constructs able to form gap junction channels reverses Cx43 def cell death. Cx43 def cells and bones from old mice exhibit reduced levels of the pro-survival microRNA miR21 and, consistently, increased levels of the miR21 target phosphatase and tensin homolog (PTEN) and reduced phosphorylated Akt, whereas PTEN inhibition reduces Cx43 def cell apoptosis. miR21 reduction is sufficient to induce apoptosis of Cx43-expressing cells and miR21 deletion in miR21 fl/fl bones increases apoptosis-related gene expression, whereas a miR21 mimic prevents Cx43 def cell apoptosis, demonstrating that miR21 lies downstream of Cx43. Cx43 def cells release more osteoclastogenic cytokines [receptor activator of NFκB ligand (RANKL)/high-mobility group box-1 (HMGB1)], and caspase-3 inhibition prevents RANKL/HMGB1 release and the increased osteoclastogenesis induced by conditioned media from Cx43 def cells, which is blocked by antagonizing HMGB1-RAGE interaction. These findings identify a novel Cx43/miR21/HMGB1/RANKL pathway involved in preventing osteocyte apoptosis that also controls osteoclast formation/recruitment and is impaired with aging. © 2017 The Authors. Aging Cell published by the Anatomical Society

  5. Increased p53 and decreased p21 accompany apoptosis induced by ultraviolet radiation in the nervous system of a crustacean

    Energy Technology Data Exchange (ETDEWEB)

    Hollmann, Gabriela, E-mail: gabrielahollmann@biof.ufrj.br [Programa de Pós Graduação em Ciências Biológicas-Fisiologia, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro-UFRJ, Rio de Janeiro, RJ 21941-590 (Brazil); Linden, Rafael, E-mail: rlinden@biof.ufrj.br [Programa de Pós Graduação em Ciências Biológicas-Fisiologia, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro-UFRJ, Rio de Janeiro, RJ 21941-590 (Brazil); Giangrande, Angela, E-mail: angela.giangrande@igbmc.fr [Institut de Génétique et de Biologie Moléculaire et Cellulaire-IGBMC, INSERM, Strasbourg (France); Allodi, Silvana, E-mail: sallodi@biof.ufrj.br [Programa de Pós Graduação em Ciências Biológicas-Fisiologia, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro-UFRJ, Rio de Janeiro, RJ 21941-590 (Brazil)

    2016-04-15

    Highlights: • The paper characterizes molecular pathways of cell responses to environmental doses of UV in brain tissue of a crab species. • The UV radiation changes levels of proteins which trigger apoptotic or cell cycle arrest pathways and also it changes neurotrophins which lead to apoptosis of neural cell in the central nervous system (CNS) of the crab Ucides cordatus. • The UVB wavelengths in the solar simulator damaged the DNA, either directly or indirectly, by increasing ROS, and induced the increase of p53 and AKT, which blocked p21 and increased the expression of activated caspase-3, triggering apoptosis. The signs of death increased the expression of neurotrophins (BDNF and GDNF), which continued to stimulate the apoptosis signaling mediated by caspase-3. • In the brain of the crab U. cordatus, p53/p21 relationship in response to UV radiation is different from that of most mammals. - Abstract: Ultraviolet (UV) radiation can produce biological damage, leading the cell to apoptosis by the p53 pathway. This study evaluated some molecular markers of the apoptosis pathway induced by UVA, UVB and UVA+ UVB (Solar Simulator, SIM) in environmental doses, during five consecutive days of exposure, in the brain of the crab Ucides cordatus. We evaluated the central nervous system (CNS) by immunoblotting the content of proteins p53, p21, phosphorylated AKT, BDNF, GDNF, activated caspase-3 (C3) and phosphohistone H3 (PH3); and by immunohistochemical tests of the cells labeled for PH3 and C3. After the fifth day of exposure, UVB radiation and SIM increased the protein content of p53, increasing the content of AKT and, somehow, blocking p21, increasing the content of activated caspase-3, which led the cells to apoptosis. The signs of death affected the increase in neurotrophins, such as BDNF and GDNF, stimulating the apoptotic cascade of events. Immunohistochemical assays and immunoblotting showed that apoptosis was present in the brains of all UV groups, while

  6. Increase in intracellular PGE2 induces apoptosis in Bax-expressing colon cancer cell

    International Nuclear Information System (INIS)

    Lalier, Lisenn; Pedelaborde, François; Braud, Christophe; Menanteau, Jean; M Vallette, François; Olivier, Christophe

    2011-01-01

    NSAIDs exhibit protective properties towards some cancers, especially colon cancer. Yet, it is not clear how they play their protective role. PGE 2 is generally shown as the only target of the NSAIDs anticancerous activity. However, PGE 2 known targets become more and more manifold, considering both the molecular pathways involved and the target cells in the tumour. The role of PGE 2 in tumour progression thus appears complex and multipurpose. To gain understanding into the role of PGE 2 in colon cancer, we focused on the activity of PGE 2 in apoptosis in colon cancer cell lines. We observed that an increase in intracellular PGE 2 induced an apoptotic cell death, which was dependent on the expression of the proapoptotic protein Bax. This increase was induced by increasing PGE 2 intracellular concentration, either by PGE 2 microinjection or by the pharmacological inhibition of PGE 2 exportation and enzymatic degradation. We present here a new sight onto PGE 2 in colon cancer cells opening the way to a new prospective therapeutic strategy in cancer, alternative to NSAIDs

  7. Results of in vitro chemosensitivity assays

    International Nuclear Information System (INIS)

    Tanigawa, Nobuhiko; Morimoto, Hideki; Akita, Toshiaki; Inoue, Hiroshi; Tanaka, Takeo.

    1986-01-01

    The authors reviewed their experiences to date with chemosensitivity testing of 629 tumors by human tumor clonogenic assay (HTCA) and of 199 tumors by scintillation assay (SA). HTCA and SA were both performed using a double-layer-soft-agar system with continuous exposure of cells to one concentration of standard anticancer drugs. Overall, 60 % of specimens in HTCA and 58 % in SA produced significant growth in vitro. HTCA was 52 % (13/25) reliable for predicting in vivo sensitivity, and 95 % (36/38) reliable for in vivo resistance, whereas SA was 40 % (8/20) reliable for in vivo sensitivity and 88 % (21/24) for in vivo resistance. In vitro success rates were variable, depending on the tumor histology. In vitro growth of gastric cancer specimens was characteristically lower than that of colon cancer specimens (48 % and 60 % in HTCA, and 46 % and 68 % in SA, respectively). (p < 0.005). Optimal in vitro-in vivo drug concentrations and culture conditions are still being defined. Correlation studies of in vitro-in vivo responses of gastrointestinal cancers suggested that in vitro concentrations of 5-fluorouracil and mitomycin C used in this study were considerably higher than their optimal doses. Tumor cell heterogeneity poses significant problems in the clinical use of chemosensitivity assays. In this last study, we sought evidence of tumor heterogeneity by comparing chemosensitivity responses between : 1) different portions of a single tumor, 2) a primary and a metastatic biopsy taken from a patient on the same day, and 3) different metastases from a patient taken on the same day. The results demonstrated the presence of considerable heterogeneity of response to chemotherapy among different tumors from the same patient, and even within the same tumor. The reported discrepancies of in vitro and in vivo sensitivity may be due to such therapeutic heterogeneity among tumors. (J.P.N.)

  8. Decorin in human oral cancer: A promising predictive biomarker of S-1 neoadjuvant chemosensitivity

    Energy Technology Data Exchange (ETDEWEB)

    Kasamatsu, Atsushi, E-mail: kasamatsua@faculty.chiba-u.jp [Department of Oral Science, Graduate School of Medicine, Chiba University, Chiba 260-8670 (Japan); Department of Dentistry and Oral–Maxillofacial Surgery, Chiba University Hospital, Chiba 260-8670 (Japan); Uzawa, Katsuhiro, E-mail: uzawak@faculty.chiba-u.jp [Department of Oral Science, Graduate School of Medicine, Chiba University, Chiba 260-8670 (Japan); Department of Dentistry and Oral–Maxillofacial Surgery, Chiba University Hospital, Chiba 260-8670 (Japan); Minakawa, Yasuyuki; Ishige, Shunsaku; Kasama, Hiroki [Department of Oral Science, Graduate School of Medicine, Chiba University, Chiba 260-8670 (Japan); Endo-Sakamoto, Yosuke; Ogawara, Katsunori [Department of Dentistry and Oral–Maxillofacial Surgery, Chiba University Hospital, Chiba 260-8670 (Japan); Shiiba, Masashi; Takiguchi, Yuichi [Medical Oncology, Graduate School of Medicine, Chiba University, Chiba 260-8670 (Japan); Tanzawa, Hideki [Department of Oral Science, Graduate School of Medicine, Chiba University, Chiba 260-8670 (Japan); Department of Dentistry and Oral–Maxillofacial Surgery, Chiba University Hospital, Chiba 260-8670 (Japan)

    2015-01-30

    Highlights: • DCN is significantly up-regulated in chemoresistant cancer cell lines. • DCN is a key regulator for chemoresistant mechanisms in vitro and in vivo. • DCN predicts the clinical responses to S-1 NAC for patients with oral cancer. - Abstract: We reported previously that decorin (DCN) is significantly up-regulated in chemoresistant cancer cell lines. DCN is a small leucine-rich proteoglycan that exists and functions in stromal and epithelial cells. Accumulating evidence suggests that DCN affects the biology of several types of cancer by directly/indirectly targeting the signaling molecules involved in cell growth, survival, metastasis, and angiogenesis, however, the molecular mechanisms of DCN in chemoresistance and its clinical relevance are still unknown. Here we assumed that DCN silencing cells increase chemosusceptibility to S-1, consisted of tegafur, prodrug of 5-fluorouracil. We first established DCN knockdown transfectants derived from oral cancer cells for following experiments including chemosusceptibility assay to S-1. In addition to the in vitro data, DCN knockdown zenografting tumors in nude mice demonstrate decreasing cell proliferation and increasing apoptosis with dephosphorylation of AKT after S-1 chemotherapy. We also investigated whether DCN expression predicts the clinical responses of neoadjuvant chemotherapy (NAC) using S-1 (S-1 NAC) for oral cancer patients. Immunohistochemistry data in the preoperative biopsy samples was analyzed to determine the cut-off point for status of DCN expression by receiver operating curve analysis. Interestingly, low DCN expression was observed in five (83%) of six cases with complete responses to S-1 NAC, and in one (10%) case of 10 cases with stable/progressive disease, indicating that S-1 chemosensitivity is dramatically effective in oral cancer patients with low DCN expression compared with high DCN expression. Our findings suggest that DCN is a key regulator for chemoresistant mechanisms, and

  9. Overendocytosis of gold nanoparticles increases autophagy and apoptosis in hypoxic human renal proximal tubular cells

    Directory of Open Access Journals (Sweden)

    Ding F

    2014-09-01

    production of reactive oxygen species, the loss of mitochondrial membrane potential (ΔΨM, and an increase in apoptosis and autophagic cell death. Conclusion/significance: Our results demonstrate that renal tubular epithelial cells presented different responses under normoxic and hypoxic environments, which provide an important basis for understanding the risks associated with GNP use–especially for the potential GNP-related therapies in chronic kidney disease patients. Keywords: gold nanoparticles (GNPs, toxicity, autophagy, apoptosis 

  10. [Influence of human epidermal growth factor receptor-2 siRNA on chemosensitivity to cisplatin of human ovarian carcinoma cells: an in vitro experiment].

    Science.gov (United States)

    Lu, Yan-Ming; Zhang, Shu-Lan; Meng, Li-Rong; Zhao, Yan-Yan

    2008-04-01

    To investigate the effects of RNA interference (RNAi) targeting human epidermal growth factor receptor 2 (HER-2) gene on the change of chemosensitivity to cisplatin of ovarian carcinoma cells. Methods Three kinds of HER-2 gene targeting siRNA, HER-2 siRNA I-III, were synthesized and the best one (HER-2 siRNA III) was screened. Human ovarian carcinoma cells of the line SKOV3 were cultured randomly divided into 3 groups: HER-2 siRNA III group, transfected with HER-2 siRNA III, non-specific siRNA group, transfected with non-specific siRNA III, and control group, without transfection. Cisplatin of the concentrations of 0, 0.05, 0.2. 0.4, 0.8, 1.0, 2.0, 10, and 20 microg/ml was added into the culture fluid for 24 h. MTT method was used to detect the proliferation rate of the SKOV3 cells. Other SKOV3 cells were divided into 3 groups: siRNA group, transfected with HER-2 siRNA III, cisplatin group, exposed to cisplatin, and HER-2 siRNA III and exposed to cisplatin. Annexin V method and flow cytometry were used to detect the apoptosis of the SKOV3 cells. The HER-2 gene expression was assessed by Western blotting. The chemosensitivity of transfected cells to cisplatin was measured by MTT. Western blotting was used to detect the protein expression of apoptosis related proteins: Bcl-2, surviving, XIAP, and Smac. After exposed to cisplatin, the cell survival rate decreased as the dose of cisplatin increased. The proliferation rate of the SKOV3 cells transfected with HER-2 siRNA III and exposed to 1 microg/ml cisplatin was (58 +/- 5)%, significantly lower than those of the nonspecific siRNA III transfection group [(65 +/- 6)%] and the control group [(68 +/- 3)%, both P 0.05). The apoptosis rates at different time point of the HER-2 siRNA III + cisplatin group were all higher than those of the other 2 groups (all P cisplatin group were significantly lower than those of the cisplatin group, and the protein expression level of the apoptotic protein Smac of the HER-2 siRNA III

  11. Chemosensitization of Plasmodium falciparum by Probenecid In Vitro

    Science.gov (United States)

    Nzila, Alexis; Mberu, Eddy; Bray, Pat; Kokwaro, Gilbert; Winstanley, Peter; Marsh, Kevin; Ward, Steve

    2003-01-01

    Resistance to drugs can result from changes in drug transport, and this resistance can sometimes be overcome by a second drug that modifies the transport mechanisms of the cell. This strategy has been exploited to partly reverse resistance to chloroquine in Plasmodium falciparum. Studies with human tumor cells have shown that probenecid can reverse resistance to the antifolate methotrexate, but the potential for reversal of antifolate resistance has not been studied in P. falciparum. In the present study we tested the ability of probenecid to reverse antifolate resistance in P. falciparum in vitro. Probenecid, at concentrations that had no effect on parasite viability alone (50 μM), was shown to increase the sensitivity of a highly resistant parasite isolate to the antifolates pyrimethamine, sulfadoxine, chlorcycloguanil, and dapsone by seven-, five-, three-, and threefold, respectively. The equivalent effects against an antifolate-sensitive isolate were activity enhancements of approximately 3-, 6-, 1.2-, and 19-fold, respectively. Probenecid decreased the level of uptake of radiolabeled folic acid, suggesting a transport-based mechanism linked to folate salvage. When probenecid was tested with chloroquine, it chemosensitized the resistant isolate to chloroquine (i.e., enhanced the activity of chloroquine). This enhancement of activity was associated with increased levels of chloroquine accumulation. In conclusion, we have shown that probenecid can chemosensitize malaria parasites to antifolate compounds via a mechanism linked to reduced folate uptake. Notably, this effect is observed in both folate-sensitive and -resistant parasites. In contrast to the activities of antifolate compounds, the effect of probenecid on chloroquine sensitivity was selective for chloroquine-resistant parasites (patent P407595GB [W. P. Thompson & Co., Liverpool, United Kingdom] has been filed to protect this intellectual property). PMID:12821454

  12. Association of telomerase activity with radio- and chemosensitivity of neuroblastomas

    Directory of Open Access Journals (Sweden)

    Willich Normann

    2010-07-01

    Full Text Available Abstract Background Telomerase activity compensates shortening of telomeres during cell division and enables cancer cells to escape senescent processes. It is also supposed, that telomerase is associated with radio- and chemoresistance. In the here described study we systematically investigated the influence of telomerase activity (TA and telomere length on the outcome of radio- and chemotherapy in neuroblastoma. Methods We studied the effects on dominant negative (DN mutant, wild type (WT of the telomerase catalytic unit (hTERT using neuroblastoma cell lines. The cells were irradiated with 60Co and treated with doxorubicin, etoposide, cisplatin and ifosfamide, respectively. Viability was determined by MTS/MTT-test and the GI50 was calculated. Telomere length was measured by southernblot analysis and TA by Trap-Assay. Results Compared to the hTERT expressing cells the dominant negative cells showed increased radiosensitivity with decreased telomere length. Independent of telomere length, telomerase negative cells are significantly more sensitive to irradiation. The effect of TA knock-down or overexpression on chemosensitivity were dependent on TA, the anticancer drug, and the chemosensitivity of the maternal cell line. Conclusions Our results supported the concept of telomerase inhibition as an antiproliferative treatment approach in neuroblastomas. Telomerase inhibition increases the outcome of radiotherapy while in combination with chemotherapy the outcome depends on drug- and cell line and can be additive/synergistic or antagonistic. High telomerase activity is one distinct cancer stem cell feature and the here described cellular constructs in combination with stem cell markers like CD133, Aldehyddehydrogenase-1 (ALDH-1 or Side population (SP may help to investigate the impact of telomerase activity on cancer stem cell survival under therapy.

  13. Nitric oxide coating polypropylene mesh increases angiogenesis and reduces inflammatory response and apoptosis.

    Science.gov (United States)

    Prudente, Alessandro; Favaro, Wágner José; Reis, Leonardo Oliveira; Riccetto, Cássio Luis Zanettini

    2017-04-01

    To evaluate the effect of implanted S-nitrosoglutathione (GSNO) coating polypropylene mesh in foreign-body response of rats. Thirty female rats underwent to subcutaneous implant of five polypropylene (PP) fragments: uncoated PP (control); PP polyvinylalcohol (PVA) coated and PP PVA + GSNO (1, 10 and 70 mMol) coated. After euthanasia (4 and 30 days), eight slides were prepared from each animal: hematoxylin-eosin (inflammatory response); unstained (birefringence collagen evaluation); TUNEL technique (apoptosis); and five for immunohistochemical processing: CD-31 (angiogenesis), IL-1 and TNF-α (proinflammatory cytokynes), iNOS (NO synthesis) and MMP-2 (collagen metabolism). The inflammation area, birefringence index, apoptotic index, immunoreactivity and vessel density were objectively measured. Inflammatory reaction area at 4 days was 11.3, 15.2, 25.1, 17.1 and 19.3% of pure PP, PVA, GSNO 1, 10 and 70 mM, respectively, p = 0.0006 (PP × Others). At 30 days lower inflammatory area was observed in GSNO 10 and 70 mM compared to pure PP (5.3, 5.2 and 11.1%, respectively, p = 0.0001). Vessel density was higher for GSNO 1 mM (25.5%) compared to pure PP (19.6%) at 30 days only, p = 0.0081. Apoptotic index at 4 days was lower for GSNO 1 mM (49.3%) than pure PVA (60.6%), p = 0.0124. GSNO 10 and 70 mM reduced their apoptotic index at 30 days compared to 4 days (49.9 vs. 36.9 and 59.1 vs. 47.5%, respectively, p = 0.0397). Birefringence index, IL-1, TNF, MMP-2 and iNOS were not different. Depending on concentrations, GSNO can increase angiogenesis, reduce inflammation and apoptosis compared to pure PP, without impact on cytokine, collagen organization/metabolism and endogenous NO synthesis.

  14. Increased levels of apoptosis in the prefusion neural folds underlie the craniofacial disorder, Treacher Collins syndrome.

    Science.gov (United States)

    Dixon, J; Brakebusch, C; Fässler, R; Dixon, M J

    2000-06-12

    Treacher Collins syndrome (TCS) is an autosomal dominant disorder of human craniofacial development that results from loss-of-function mutations in the gene TCOF1. Although this gene has been demonstrated to encode the nucleolar phosphoprotein treacle, the developmental mechanism underlying TCS remains elusive, particularly as expression studies have shown that the murine orthologue, Tcof1, is widely expressed. To investigate the molecular pathogenesis of TCS, we replaced exon 1 of Tcof1 with a neomycin-resistance cassette via homologous recombination in embryonic stem cells. Tcof1 heterozygous mice die perinatally as a result of severe craniofacial anomalies that include agenesis of the nasal passages, abnormal development of the maxilla, exencephaly and anophthalmia. These defects arise due to a massive increase in the levels of apoptosis in the prefusion neural folds, which are the site of the highest levels of Tcof1 expression. Our results demonstrate that TCS arises from haploinsufficiency of a protein that plays a crucial role in craniofacial development and indicate that correct dosage of treacle is essential for survival of cephalic neural crest cells.

  15. Pneumococcal infections in humans are associated with increased apoptosis and trafficking of type 1 cytokine-producing T cells

    DEFF Research Database (Denmark)

    Kemp, Kåre; Bruunsgaard, Helle; Skinhøj, Peter

    2002-01-01

    , little is known regarding the T-cell response during in vivo infections in humans. The purpose of this study was to test the hypothesis that activated T cells producing type 1 cytokines were engaged in the host response to pneumococcal infections. The phenotype and function of T cells were studied in 22...... tissues and/or apoptosis. In fact, increased activation-induced apoptosis in the remaining peripheral lymphocytes and elevated levels of soluble Fas ligand were detected at admission to the hospital. In conclusion, these data suggest that activated T lymphocytes with a type 1 cytokine profile are highly...

  16. Increased apoptosis and hypomyelination in cerebral white matter of macular mutant mouse brain

    Directory of Open Access Journals (Sweden)

    Shoichi Takikita

    2015-09-01

    Full Text Available Hypomyelination in developing brain is often accompanied by congenital metabolic disorders. Menkes kinky hair disease is an X-linked neurodegenerative disease of impaired copper transport, resulting from a mutation of the Menkes disease gene, a transmembrane copper-transporting p-type ATPase gene (ATP7A. In a macular mutant mouse model, the murine ortholog of Menkes gene (mottled gene is mutated, and widespread neurodegeneration and subsequent death are observed. Although some biochemical analysis of myelin protein in macular mouse has been reported, detailed histological study of myelination in this mouse model is currently lacking. Since myelin abnormality is one of the neuropathologic findings of human Menkes disease, in this study early myelination in macular mouse brain was evaluated by immunohistochemistry. Two-week-old macular mice and normal littermates were perfused with 4% paraformaldehyde. Immunohistochemical staining of paraffin embedded and vibratome sections was performed using antibodies against either CNPase, cleaved caspase-3 or O4 (marker of immature oligodendrocytes. This staining showed that cerebral myelination in macular mouse was generally hypoplastic and that hypomyelination was remarkable in internal capsule, corpus callosum, and cingulate cortex. In addition, an increased number of cleaved caspase-3 positive cells were observed in corpus callosum and internal capsule. Copper deficiency induced by low copper diet has been reported to induce oligodendrocyte dysfunction and leads to hypomyelination in this mouse model. Taken together, hypomyelination observed in this study in a mouse model of Menkes disease is assumed to be induced by increased apoptosis of immature oligodendrocytes in developing cerebrum, through deficient intracellular copper metabolism.

  17. CYP24A1 exacerbated activity during diabetes contributes to kidney tubular apoptosis via caspase-3 increased expression and activation.

    Directory of Open Access Journals (Sweden)

    Alexandre Tourigny

    Full Text Available Decreases in circulating 25,hydroxyl-vitamin D3 (25 OH D3 and 1,25,dihydroxyl-vitamin D3 (1,25 (OH2 D3 have been extensively documented in patients with type 2 diabetes. Nevertheless, the molecular reasons behind this drop, and whether it is a cause or an effect of disease progression is still poorly understood. With the skin and the liver, the kidney is one of the most important sites for vitamin D metabolism. Previous studies have also shown that CYP24A1 (an enzyme implicated in vitamin D metabolism, might play an important role in furthering the progression of kidney lesions during diabetic nephropathy. In this study we show a link between CYP24A1 increase and senescence followed by apoptosis induction in the renal proximal tubules of diabetic kidneys. We show that CYP24A1 expression was increased during diabetic nephropathy progression. This increase derived from protein kinase C activation and increased H(2O(2 cellular production. CYP24A1 increase had a major impact on cellular phenotype, by pushing cells into senescence, and later into apoptosis. Our data suggest that control of CYP24A1 increase during diabetes has a beneficial effect on senescence induction and caspase-3 increased expression. We concluded that diabetes induces an increase in CYP24A1 expression, destabilizing vitamin D metabolism in the renal proximal tubules, leading to cellular instability and apoptosis, and thereby accelerating tubular injury progression during diabetic nephropathy.

  18. Progesterone Increases Apoptosis and Inversely Decreases Autophagy in Human Hepatoma HA22T/VGH Cells Treated with Epirubicin

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    Wen-Tsan Chang

    2014-01-01

    Full Text Available Hepatocellular carcinoma (HCC is the leading cause of cancer-related deaths worldwide. Epirubicin can induce intracellular reactive oxygen species and is widely used to treat unresectable HCC. Progesterone has been found to inhibit the proliferation of hepatoma cells. This study was designed to test the combined effects of epirubicin and progesterone on human hepatoma cell line, HA22T/VGH. These cells were treated with different concentrations of epirubicin with or without the coaddition of 30 μM progesterone and then analyzed for apoptosis, autophagy, and expressions of apoptotic-related proteins and multidrug-resistant gene. Epirubicin treatment dose-dependently inhibited the growth of HA22T/VGH cells. Addition of 30 μM progesterone, which was inactive alone, augmented the effect of epirubicin on the inhibition of growth of HA22T/VGH cells. Cotreatment with progesterone enhanced epirubicin-induced apoptosis, as evidenced by greater increase in caspase-3 activity and in the ratio of the apoptosis-regulating protein, Bax/Bcl-XL. The combination also caused a decrease in autophagy and in the expression of multidrug resistance-related protein 1 mRNA compared to epirubicin alone. This study shows the epirubicin/progesterone combination was more effective in increasing apoptosis and inversely decreasing autophagy on HA22T/VGH cells treated with epirubicin alone, suggesting that this combination can potentially be used to treat HCC.

  19. Lens epithelial cell apoptosis and intracellular Ca2+ increase in the presence of xanthurenic acid

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    Richter Christoph

    2002-04-01

    Full Text Available Abstract Background Xanthurenic acid is an endogenous product of tryptophan degradation by indoleamine 2,3-dioxygenase (IDO. We have previously reported that IDO is present in mammalian lenses, and xanthurenic acid is accumulated in the lenses with aging. Here, we studied the involvement of xanthurenic acid in the human lens epithelial cell physiology. Methods Human lens epithelial cells primary cultures were used. Control cells, and cells in the presence of xanthurenic acid grow in the dark. Western blot analysis and immunofluorescence studies were performed. Results In the presence of xanthurenic acid human lens epithelial cells undergo apoptosis-like cell death. In the control cells gelsolin stained the perinuclear region, whereas in the presence of 10 μM xanthurenic acid gelsolin is translocated to the cytoskeleton, but does not lead to cytoskeleton breakdown. In the same condition caspase-3 activation, and DNA fragmentation was observed. At low (5 to 10 μM of xanthurenic acid concentration, the elongation of the cytoskeleton was associated with migration of mitochondria and cytochrome c release. At higher concentrations xanthurenic acid (20 μM and 40 μM damaged mitochondria were observed in the perinuclear region, and nuclear DNA cleavage was observed. We observed an induction of calpain Lp 82 and an increase of free Ca2+ in the cells in a xanthurenic acid concentration-dependent manner. Conclusions The results show that xanthurenic acid accumulation in human lens epithelial cells disturbs the normal cell physiology and leads to a cascade of pathological events. Xanthurenic acid induces calpain Lp82 and caspases in the cells growing in the dark and can be involved in senile cataract development.

  20. Increased oxidative stress is associated with balanced increases in hepatocyte apoptosis and proliferation in glycerol-3-phosphate acyltransferase-1 deficient mice

    Science.gov (United States)

    Hammond, Linda E.; Albright, Craig D.; He, Lihua; Rusyn, Ivan; Watkins, Steven M.; Doughman, Scott D.; Lemasters, John J.; Coleman, Rosalind A.

    2007-01-01

    The absence of mouse mitochondrial glycerol-3-phosphate acyltransferase-1 (Gpat1-/-) increases the amount of arachidonate in liver phospholipids and increases β-hydroxybutyrate and acyl-carnitines, suggesting an elevated rate of liver fatty acid oxidation. We asked whether these alterations might increase reactive oxygen species (ROS), apoptosis, or hepatocyte proliferation. Compared to wildtype controls, liver mitochondria from Gpat1-/- mice showed a 20% increase in the rate of ROS production and a markedly increased sensitivity to the induction of the mitochondrial permeability transition. Mitochondrial phosphatidylethanolamine and phosphatidylcholine from Gpat1-/- liver contained 21% and 67% more arachidonate, respectively, than wildtype controls, and higher amounts of 4-hydroxynonenal, a product of arachidonate peroxidation. Oxidative stress was associated with an increase in apoptosis, and with 3-fold and 15-fold higher TUNEL positive cells in liver from young and old Gpat1-/- mice, respectively, compared to age-matched controls. Compared to controls, bromodeoxyuridine labeling was 50% and 7-fold higher in livers from young and old Gpat1-/- mice, respectively, but fewer glutathione-S-transferase positive cells were present. Thus, Gpat1-/- liver exhibits increased oxidative stress and sensitivity of the mitochondrial permeability transition pore, and a balanced increase in apoptosis and proliferation. PMID:17258706

  1. Coenzyme Q blocks biochemical but not receptor-mediated apoptosis by increasing mitochondrial antioxidant protection

    Czech Academy of Sciences Publication Activity Database

    Alleva, R.; Tomasetti, M.; Anděra, Ladislav; Gellert, N.; Borghi, B.; Weber, C.; Murphy, M. P.; Neužil, J.

    2001-01-01

    Roč. 503, č. 1 (2001), s. 46-50 ISSN 0014-5793 R&D Projects: GA ČR GA301/99/0350 Institutional research plan: CEZ:AV0Z5052915 Keywords : coenzyme Q * apoptosis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.644, year: 2001

  2. Tributyltin (TBT) increases TNFα mRNA expression and induces apoptosis in the murine macrophage cell line in vitro.

    Science.gov (United States)

    Nakano, Ken; Tsunoda, Masashi; Konno, Nobuhiro

    2004-11-01

    Tributyltin (TBT) compounds have been widely used as antifouling agents for shipbottom paint. The immune system is a target of TBT intoxication. We evaluated the effects of TBT chloride in macrophages, which have critical roles in the immune system, using a murine macrophage lineage cell line, J774.1,in vitro. We examined tumor necrosis factor α (TNFα), interleukin-1β (IL-1β) andc-jun mRNA expression in J774.1 cells. The effects of TBT on the apoptosis of J774.1 cells were examined by determining the percentage of TUNEL-positive cells and caspase-3 activity. The mean values of the viabilities of J774.1 cells exposed to TBT decreased dose-dependently. The relative mRNA expression of TNFα increased dose-dependently, however, that of IL-1β was not significantly different among the groups. The mean percentage of TUNEL-positive cells increased dose-dependently. Increases in the caspase-3 activities of J774.1 cells were observed in the groups exposed to higher concentrations of TBT. The mean value of relative mRNA expression of c-Jun transcription factor increased dose-dependently. The increases in the percentage of TUNEL-positive cells and in caspase-3 activity suggested that exposure to TBT induces apoptosis of J774.1 cells. The increases in the mRNA expression of TNFα andc-jun by TBT may be related to apoptosis in macrophages.

  3. Apoptosis and necrosis increase antigenicity of proteins recognized by antinuclear antibodies

    Directory of Open Access Journals (Sweden)

    J.J. Bollain-y-Goitia

    2011-09-01

    Full Text Available Obiettivo. Lo scopo di questo studio è quello di indagare se l’apoptosis e la necrosi aumentano l’antigenicità delle proteine riconosciute da anticorpi antinucleo. Materiale e metodi. Cellule HEp-2 sono state coltivate in condizioni standard; l’apoptosis è stata indotta con camptecina e la necrosi con cloruro di mercurio. L’antigenicità delle proteine estratte dalle cellule è stato testata su membrane di nitrocellulosa e sondata con sieri positivi o negativi per anticorpi antinucleo utilizzando un sistema ELISA a luminescenza (luminescent. Risultati. Le alterazioni apoptotiche nelle cellule HEp-2 sono apparse entro 24 ore dall’esposizione alla camptoicina, mentre i segni di necrosi si sono evidenziati più precocemente. La luminescenza si è dimostrata significativamente superiore nei sieri ANA positivi che nei controlli ANA negativi. Gli antcorpi antinucleari sieirici riconoscono meglio gli antigeni da cellule apoptotiche e necrotiche rispetto ai controlli che non hanno subito trattamenti chimici. Conclusioni. L’apoptosi e la necrosi incrementano la capacità legante degli ANA attraverso una migliore disponibilità di antigeni intracellulari o svelando epitopi criptici.

  4. Melatonin increases survival of HaCaT keratinocytes by suppressing UV-induced apoptosis.

    Science.gov (United States)

    Fischer, T W; Zbytek, B; Sayre, R M; Apostolov, E O; Basnakian, A G; Sweatman, T W; Wortsman, J; Elsner, P; Slominski, A

    2006-01-01

    Melatonin is a potent antioxidant and direct radical scavenger. As keratinocytes represent the major population in the skin and UV light causes damage to these cells, the possible protective effects of melatonin against UV-induced cell damage in HaCaT keratinocytes were investigated in vitro. Cells were preincubated with melatonin at graded concentrations from 10(-9) to 10(-3) m for 30 min prior to UV irradiation at doses of 25 and 50 mJ/cm2. Biological markers of cellular viability such as DNA synthesis and colony-forming efficiency as well as molecular markers of apoptosis were measured. DNA synthesis was determined by [3H]-thymidine incorporation into insoluble cellular fraction, clonogenicity through plating efficiency experiments and apoptosis by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. DNA synthesis experiments showed a strong protective effect by preincubation with melatonin at concentrations of 10(-4) m (P UV incubation protective effect. These results indicate that preincubation is a requirement for melatonin to exert its protective effects. The mechanism of melatonin's protective effect (10(-6) to 10(-3) m) includes inhibition of apoptosis as measured by TUNEL assay. Moreover, the biological significance of these effects is supported by clonogenic studies showing a significantly higher number of colonies in cultures treated with melatonin compared to controls. Thus, pretreatment with melatonin led to strong protection against UVB-induced damage in keratinocytes.

  5. Development of Chemosensitivity in Neurons from the Nucleus Tractus Solitarii (NTS) of Neonatal Rats

    Science.gov (United States)

    Conrad, Susan C.; Nichols, Nicole L.; Ritucci, Nick A.; Dean, Jay B.; Putnam, Robert W.

    2009-01-01

    We studied the development of chemosensitivity during the neonatal period in rat Nucleus tractus solitarii (NTS) neurons. We determined the percentage of neurons activated by hypercapnia (15% CO2) and assessed the magnitude of the response by calculating the chemosensitivity index (CI). There were no differences in the percentage of neurons that were inhibited (9%) or activated (44.8%) by hypercapnia or in the magnitude of the activated response (CI 164±4.9%) in NTS neurons from neonatal rats of all ages. To assess the degree of intrinsic chemosensitivity in these neurons we used chemical synaptic block medium and the gap junction blocker carbenoxolone. Chemical synaptic block medium slightly decreased basal firing rate but did not affect the percentage of NTS neurons that responded to hypercapnia at any neonatal age. However, in neonates aged increased CI. Carbenoxolone did not significantly alter the number of NTS neurons activated by hypercapnia in neonatal rats of any age. In summary, the response of NTS neurons from neonatal rats appears to be intrinsic and largely unchanged throughout early development. In young neonates (

  6. Postnatal High-Fat Diet Increases Liver Steatosis and Apoptosis Threatened by Prenatal Dexamethasone through the Oxidative Effect

    Directory of Open Access Journals (Sweden)

    Ying-Hsien Huang

    2016-03-01

    Full Text Available The objective of this study was to investigate cellular apoptosis in prenatal glucocorticoid overexposure and a postnatal high fat diet in rats. Pregnant Sprague-Dawley rats at gestational days 14 to 21 were administered saline (vehicle or dexamethasone and weaned onto either a normal fat diet or a high fat diet for 180 days; in total four experimental groups were designated, i.e., vehicle treated group (VEH, dexamethasone treated group (DEX, vehicle treated plus high-fat diet (VHF, and dexamethasone treated plus high-fat diet (DHF. Chronic effects of prenatal liver programming were assessed at postnatal day 180. The apoptotic pathways involved proteins were analyzed by Western blotting for their expressions. Apoptosis and liver steatosis were also examined by histology. We found that liver steatosis and apoptosis were increased in the DHF, DEX, and VHF treated groups, and that the DHF treated group was increased at higher levels than the DEX and VHF treated groups. The expression of leptin was decreased more in the DHF treated group than in the DEX and VHF treated groups. Decreased peroxisome proliferator-activated receptor-gamma coactivator 1α, phosphoinositide-3-kinase, manganese superoxide dismutase and increased malondialdehyde expression levels were seen in DHF treated group relative to the DEX treated group. The DHF treated group exhibited higher levels of oxidative stress, apoptosis and liver steatosis than the DEX treated group. These results indicate that the environment of high-fat diet plays an important role in the development of liver injury after prenatal stress.

  7. Troxerutin Reduces Kidney Damage against BDE-47-Induced Apoptosis via Inhibiting NOX2 Activity and Increasing Nrf2 Activity

    Directory of Open Access Journals (Sweden)

    Qun Shan

    2017-01-01

    Full Text Available 2,2,4,4-Tetrabromodiphenyl ether (BDE-47, one of the persistent organic pollutants, seriously influences the quality of life; however, its pathological mechanism remains unclear. Troxerutin is a flavonoid with pharmacological activity of antioxidation and anti-inflammation. In the present study, we investigated troxerutin against BDE-47-induced kidney cell apoptosis and explored the underlying mechanism. The results show that troxerutin reduced renal cell apoptosis and urinary protein secretion in BDE-47-treated mice. Western blot analysis shows that troxerutin supplement enhanced the ratio of Bcl-2/Bax; inhibited the release of cytochrome c from mitochondria, the activation of procaspase-9 and procaspase-3, and the cleavage of PARP; and reduced FAS, FASL, and caspase-8 levels induced by BDE-47. In addition, troxerutin decreased the production of reactive oxygen species (ROS and increased the activities of antioxidative enzymes. Furthermore, troxerutin blunted Nrf2 ubiquitylation, enhanced the activity of Nrf2, decreased the activity of NOX2, and ameliorated kidney oxidant status of BDE-47-treated mice. Together, these results confirm that troxerutin could alleviate the cytotoxicity of BDE-47 through antioxidation and antiapoptosis, which suggests that its protective mechanism is involved in the inhibition of apoptosis via suppressing NOX2 activity and increasing Nrf2 signaling pathway.

  8. Extracellular vesicle-mediated transfer of long non-coding RNA ROR modulates chemosensitivity in human hepatocellular cancer☆

    Science.gov (United States)

    Takahashi, Kenji; Yan, Irene K.; Kogure, Takayuki; Haga, Hiroaki; Patel, Tushar

    2014-01-01

    Hepatocellular cancers (HCC) are highly resistant to chemotherapy. TGFβ has been associated with chemoresistance in some human cancers but the mechanisms involved are unknown. We explored how TGFβ might contribute to altered responses to therapy by assessing the involvement and mechanistic contribution of extracellular vesicle long non-coding RNA (lncRNA) in mediating TGFβ-dependent chemoresistance. TGFβ reduced the sensitivity of HCC cells to sorafenib or doxorubicin and altered the release of both extracellular vesicles and of selected lncRNA within these vesicles. Amongst these, lincRNA-ROR (linc-ROR), a stress-responsive lncRNA was highly expressed in HCC cells and enriched within extracellular vesicles derived from tumor cells. Incubation with HCC-derived extracellular vesicles increased linc-ROR expression and reduced chemotherapy-induced cell death in recipient cells. Sorafenib increased linc-ROR expression in both tumor cells and extracellular vesicles, whereas siRNA to linc-ROR increased chemotherapy-induced apoptosis and cytotoxicity. Tumor-initiating cells that express CD133 have an increased resistance to therapy. TGFβ increased expression of CD133+ cells and colony growth in limiting dilution assays, both of which were attenuated by linc-ROR knockdown. These data provide mechanistic insights into primary chemoresistance in HCC by showing that: (a) TGFβ selectively enriches linc-RoR within extracellular vesicles, which has a potential role in intercellular signaling in response to TGFβ; (b) expression and enrichment of linc-ROR during chemotherapeutic stress plays a functional role in chemoresistance; and (c) the effects of TGFβ on chemoresistance in HCC may involve linc-RoR-dependent effects on tumor-initiating cells. These findings implicate extracellular vesicle lncRNA as mediators of the chemotherapeutic response, and support targeting linc-ROR to enhance chemosensitivity in HCC. PMID:24918061

  9. Extracellular vesicle-mediated transfer of long non-coding RNA ROR modulates chemosensitivity in human hepatocellular cancer.

    Science.gov (United States)

    Takahashi, Kenji; Yan, Irene K; Kogure, Takayuki; Haga, Hiroaki; Patel, Tushar

    2014-01-01

    Hepatocellular cancers (HCC) are highly resistant to chemotherapy. TGFβ has been associated with chemoresistance in some human cancers but the mechanisms involved are unknown. We explored how TGFβ might contribute to altered responses to therapy by assessing the involvement and mechanistic contribution of extracellular vesicle long non-coding RNA (lncRNA) in mediating TGFβ-dependent chemoresistance. TGFβ reduced the sensitivity of HCC cells to sorafenib or doxorubicin and altered the release of both extracellular vesicles and of selected lncRNA within these vesicles. Amongst these, lincRNA-ROR (linc-ROR), a stress-responsive lncRNA was highly expressed in HCC cells and enriched within extracellular vesicles derived from tumor cells. Incubation with HCC-derived extracellular vesicles increased linc-ROR expression and reduced chemotherapy-induced cell death in recipient cells. Sorafenib increased linc-ROR expression in both tumor cells and extracellular vesicles, whereas siRNA to linc-ROR increased chemotherapy-induced apoptosis and cytotoxicity. Tumor-initiating cells that express CD133 have an increased resistance to therapy. TGFβ increased expression of CD133+ cells and colony growth in limiting dilution assays, both of which were attenuated by linc-ROR knockdown. These data provide mechanistic insights into primary chemoresistance in HCC by showing that: (a) TGFβ selectively enriches linc-RoR within extracellular vesicles, which has a potential role in intercellular signaling in response to TGFβ; (b) expression and enrichment of linc-ROR during chemotherapeutic stress plays a functional role in chemoresistance; and (c) the effects of TGFβ on chemoresistance in HCC may involve linc-RoR-dependent effects on tumor-initiating cells. These findings implicate extracellular vesicle lncRNA as mediators of the chemotherapeutic response, and support targeting linc-ROR to enhance chemosensitivity in HCC.

  10. Microinjection manipulation resulted in the increased apoptosis of spermatocytes in testes from intracytoplasmic sperm injection (ICSI derived mice.

    Directory of Open Access Journals (Sweden)

    Yang Yu

    Full Text Available The invention of intracytoplasmic sperm injection (ICSI has possibly been the most important development in reproductive medicine, one that has given hope to thousands of infertile couples worldwide. However, concerns remain regarding the safety of this method since it is a more invasive procedure than in vitro fertilization (IVF, since a spermatozoon is injected into the oocyte cytoplasm. Using mice derived from IVF technology as a control, we assessed the influence of invasive microinjection in the process of transferring sperm into oocyte cytoplasm in ICSI procedure on the development and physiologic function of resultant offspring. Our results demonstrated that mice produced from ICSI and IVF had no significant difference in phenotypic indices including body weight, forelimb physiology, and learning and memory ability. However, increased spermatocyte apoptosis was observed in the testis of adult ICSI mice, when compared with IVF mice. And, decreased testis weight and marked damage of spermatogenic epithelia were found in aged ICSI mice. Furthermore, proteomic analysis verified that most of the differentiated proteins in testes between adult ICSI and IVF mice were those involved in regulation of apoptosis pathways. Our results demonstrated that the microinjection manipulation used in the ICSI procedure might pose potential risks to the fertility of male offspring. The changed expression of a series of proteins relating to apoptosis or proliferation might contribute to it. Further studies are necessary to better understand all the risks of ICSI.

  11. Suture compression induced midpalatal suture chondrocyte apoptosis with increased caspase-3, caspase-9, Bad, Bak, Bax and Bid expression.

    Science.gov (United States)

    Lan, Tingting; Zhao, Hanchi; Xiang, Bilu; Wang, Jun; Liu, Yang

    2017-07-22

    Previous studies found bone resorption and chondrocytes loss in mouse models of mid-palatal suture when given continuous compressive force, although chondrocytes response remained unknown. Herein, we design this study to determine how continuous compression force induces chondrocytes apoptosis. Thirty C57BL/6 male mice (aged 6 weeks) were randomly assigned into controls (not ligated to a spring), blank controls (ligated with no compression) and the compression group (ligated with 20-g compression). After 4 d, palatal tissues were sampled and stained by TB and safranin-O. Tunel staining measured the percentage of apoptotic chondrocytes, and immunohistochemistry was performed to label apoptosis-associated proteins (e.g., Bcl-2, Bcl-xl, Bax, Bak, Bid, Bad, caspase-3, caspase-8 and caspase-9). Intergroup comparison was made by the rank sum test, and P compression group was significantly decreased, while the control group remained largely unaltered. Tunel staining showed that apoptotic cell numbers in the mid-palatal suture were significantly higher than the control group. Immunohistochemistry showed that mice in the compression group had significantly increased expression of caspase-3, caspase-9, Bad, Bak, Bax and Bid; However, caspase-8 remained unaltered. No expression of Bcl-2 and Bcl-xl was detected. Continuous compression force induces chondrocytes apoptosis in the mid-palatal suture. This process might be associated with the mitochondrial pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Conditional Deletion of PDK1 in the Forebrain Causes Neuron Loss and Increased Apoptosis during Cortical Development

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    Congyu Xu

    2017-10-01

    Full Text Available Decreased expression but increased activity of PDK1 has been observed in neurodegenerative disease. To study in vivo function of PDK1 in neuron survival during cortical development, we generate forebrain-specific PDK1 conditional knockout (cKO mice. We demonstrate that PDK1 cKO mice display striking neuron loss and increased apoptosis. We report that PDK1 cKO mice exhibit deficits on several behavioral tasks. Moreover, PDK1 cKO mice show decreased activities for Akt and mTOR. These results highlight an essential role of endogenous PDK1 in the maintenance of neuronal survival during cortical development.

  13. Type 2 diabetes mellitus induces congenital heart defects in murine embryos by increasing oxidative stress, endoplasmic reticulum stress, and apoptosis.

    Science.gov (United States)

    Wu, Yanqing; Reece, E Albert; Zhong, Jianxiang; Dong, Daoyin; Shen, Wei-Bin; Harman, Christopher R; Yang, Peixin

    2016-09-01

    Maternal type 1 and 2 diabetes mellitus are strongly associated with high rates of severe structural birth defects, including congenital heart defects. Studies in type 1 diabetic embryopathy animal models have demonstrated that cellular stress-induced apoptosis mediates the teratogenicity of maternal diabetes leading to congenital heart defect formation. However, the mechanisms underlying maternal type 2 diabetes mellitus-induced congenital heart defects remain largely unknown. We aim to determine whether oxidative stress, endoplasmic reticulum stress, and excessive apoptosis are the intracellular molecular mechanisms underlying maternal type 2 diabetes mellitus-induced congenital heart defects. A mouse model of maternal type 2 diabetes mellitus was established by feeding female mice a high-fat diet (60% fat). After 15 weeks on the high-fat diet, the mice showed characteristics of maternal type 2 diabetes mellitus. Control dams were either fed a normal diet (10% fat) or the high-fat diet during pregnancy only. Female mice from the high-fat diet group and the 2 control groups were mated with male mice that were fed a normal diet. At E12.5, embryonic hearts were harvested to determine the levels of lipid peroxides and superoxide, endoplasmic reticulum stress markers, cleaved caspase 3 and 8, and apoptosis. E17.5 embryonic hearts were harvested for the detection of congenital heart defect formation using India ink vessel patterning and histological examination. Maternal type 2 diabetes mellitus significantly induced ventricular septal defects and persistent truncus arteriosus in the developing heart, along with increasing oxidative stress markers, including superoxide and lipid peroxidation; endoplasmic reticulum stress markers, including protein levels of phosphorylated-protein kinase RNA-like endoplasmic reticulum kinase, phosphorylated-IRE1α, phosphorylated-eIF2α, C/EBP homologous protein, and binding immunoglobulin protein; endoplasmic reticulum chaperone gene

  14. Enterolactone: A novel radiosensitizer for human breast cancer cell lines through impaired DNA repair and increased apoptosis

    International Nuclear Information System (INIS)

    Bigdeli, Bahareh; Goliaei, Bahram; Masoudi-Khoram, Nastaran; Jooyan, Najmeh; Nikoofar, Alireza; Rouhani, Maryam; Haghparast, Abbas; Mamashli, Fatemeh

    2016-01-01

    Introduction: Radiotherapy is a potent treatment against breast cancer, which is the most commonly diagnosed cancer among women. However, the emergence of radioresistance due to increased DNA repair leads to radiotherapeutic failure. Applying polyphenols combined with radiation is a more promising method leading to better survival. Enterolactone, a phytoestrogenic polyphenol, has been reported to inhibit an important radioresistance signaling pathway, therefore we conjectured that enterolactone could enhance radiosensitivity in breast cancer. To assess this hypothesis, radiation response of enterolactone treated MDA-MB-231 and T47D cell lines and corresponding cellular mechanisms were investigated. Methods: Cytotoxicity of enterolactone was measured via MTT assay. Cells were treated with enterolactone before X-irradiation, and clonogenic assay was used to evaluate radiosensitivity. Cell cycle distribution and apoptosis were measured by flow cytometric analysis. In addition, DNA damages and corresponding repair, chromosomal damages, and aberrations were assessed by comet, micronucleus, and cytogenetic assays, respectively. Results: Enterolactone decreased the viability of cells in a concentration- and time dependent manner. Enterolactone significantly enhanced radiosensitivity of cells by abrogating G2/M arrest, impairing DNA repair, and increasing radiation-induced apoptosis. Furthermore, increased chromosomal damages and aberrations were detected in cells treated with enterolactone combined with X-rays than X-ray alone. These effects were more prominent in T47D than MDA-MB-231 cells. Discussion: To our knowledge, this is the first report that enterolactone is a novel radiosensitizer for breast cancer irrespective of estrogen receptor status. Authors propose enterolactone as a candidate for combined therapy to decrease the radiation dose delivered to patients and subsequent side effects. - Highlights: • Enterolactone is proposed to be a novel radiosensitizer for

  15. Disruption of IGF-1R signaling increases TRAIL-induced apoptosis: A new potential therapy for the treatment of melanoma

    International Nuclear Information System (INIS)

    Karasic, Thomas B.; Hei, Tom K.; Ivanov, Vladimir N.

    2010-01-01

    Resistance of cancer cells to apoptosis is dependent on a balance of multiple genetic and epigenetic mechanisms, which up-regulate efficacy of the surviving growth factor-receptor signaling pathways and suppress death-receptor signaling pathways. The Insulin-like Growth Factor-1 Receptor (IGF-1R) signaling pathway is highly active in metastatic melanoma cells by mediating downstream activation of PI3K-AKT and MAPK pathways and controlling general cell survival and proliferation. In the present study, we used human melanoma lines with established genotypes that represented different phases of cancer development: radial-growth-phase WM35, vertical-growth-phase WM793, metastatic LU1205 and WM9 [1]. All these lines have normal NRAS. WM35, WM793, LU1205 and WM9 cells have mutated BRAF (V600E). WM35 and WM9 cells express normal PTEN, while in WM793 cells PTEN expression is down-regulated; finally, in LU1205 cells PTEN is inactivated by mutation. Cyclolignan picropodophyllin (PPP), a specific inhibitor of IGF-1R kinase activity, strongly down-regulated the basal levels of AKT activity in WM9 and in WM793 cells, modestly does so in LU1205, but has no effect on AKT activity in the early stage WM35 cells that are deficient in IGF-1R. In addition, PPP partially down-regulated the basal levels of active ERK1/2 in all lines used, highlighting the role of an alternative, non-BRAF pathway in MAPK activation. The final result of PPP treatment was an induction of apoptosis in WM793, WM9 and LU1205 melanoma cells. On the other hand, dose-dependent inhibition of IGF-1R kinase activity by PPP at a relatively narrow dose range (near 500 nM) has different effects on melanoma cells versus normal cells, inducing apoptosis in cancer cells and G2/M arrest of fibroblasts. To further enhance the pro-apoptotic effects of PPP on melanoma cells, we used a combined treatment of TNF-Related Apoptosis-Inducing Ligand (TRAIL) and PPP. This combination substantially increased death by apoptosis for

  16. Disruption of IGF-1R signaling increases TRAIL-induced apoptosis: A new potential therapy for the treatment of melanoma

    Energy Technology Data Exchange (ETDEWEB)

    Karasic, Thomas B.; Hei, Tom K. [Center for Radiological Research, Department of Radiation Oncology, College of Physicians and Surgeons, Columbia University, New York, NY 10032 (United States); Ivanov, Vladimir N., E-mail: vni3@columbia.edu [Center for Radiological Research, Department of Radiation Oncology, College of Physicians and Surgeons, Columbia University, New York, NY 10032 (United States)

    2010-07-15

    Resistance of cancer cells to apoptosis is dependent on a balance of multiple genetic and epigenetic mechanisms, which up-regulate efficacy of the surviving growth factor-receptor signaling pathways and suppress death-receptor signaling pathways. The Insulin-like Growth Factor-1 Receptor (IGF-1R) signaling pathway is highly active in metastatic melanoma cells by mediating downstream activation of PI3K-AKT and MAPK pathways and controlling general cell survival and proliferation. In the present study, we used human melanoma lines with established genotypes that represented different phases of cancer development: radial-growth-phase WM35, vertical-growth-phase WM793, metastatic LU1205 and WM9 [1]. All these lines have normal NRAS. WM35, WM793, LU1205 and WM9 cells have mutated BRAF (V600E). WM35 and WM9 cells express normal PTEN, while in WM793 cells PTEN expression is down-regulated; finally, in LU1205 cells PTEN is inactivated by mutation. Cyclolignan picropodophyllin (PPP), a specific inhibitor of IGF-1R kinase activity, strongly down-regulated the basal levels of AKT activity in WM9 and in WM793 cells, modestly does so in LU1205, but has no effect on AKT activity in the early stage WM35 cells that are deficient in IGF-1R. In addition, PPP partially down-regulated the basal levels of active ERK1/2 in all lines used, highlighting the role of an alternative, non-BRAF pathway in MAPK activation. The final result of PPP treatment was an induction of apoptosis in WM793, WM9 and LU1205 melanoma cells. On the other hand, dose-dependent inhibition of IGF-1R kinase activity by PPP at a relatively narrow dose range (near 500 nM) has different effects on melanoma cells versus normal cells, inducing apoptosis in cancer cells and G2/M arrest of fibroblasts. To further enhance the pro-apoptotic effects of PPP on melanoma cells, we used a combined treatment of TNF-Related Apoptosis-Inducing Ligand (TRAIL) and PPP. This combination substantially increased death by apoptosis for

  17. Increased sensitivity of African American triple negative breast cancer cells to nitric oxide-induced mitochondria-mediated apoptosis

    International Nuclear Information System (INIS)

    Martinez, Luis; Thames, Easter; Kim, Jinna; Chaudhuri, Gautam; Singh, Rajan; Pervin, Shehla

    2016-01-01

    Breast cancer is a complex heterogeneous disease where many distinct subtypes are found. Younger African American (AA) women often present themselves with aggressive form of breast cancer with unique biology which is very difficult to treat. Better understanding the biology of AA breast tumors could lead to development of effective treatment strategies. Our previous studies indicate that AA but not Caucasian (CA) triple negative (TN) breast cancer cells were sensitive to nitrosative stress-induced cell death. In this study, we elucidate possible mechanisms that contribute to nitric oxide (NO)-induced apoptosis in AA TN breast cancer cells. Breast cancer cells were treated with various concentrations of long-acting NO donor, DETA-NONOate and cell viability was determined by trypan blue exclusion assay. Apoptosis was determined by TUNEL and caspase 3 activity as well as changes in mitochondrial membrane potential. Caspase 3 and Bax cleavage, levels of Cu/Zn superoxide dismutase (SOD) and Mn SOD was assessed by immunoblot analysis. Inhibition of Bax cleavage by Calpain inhibitor, and levels of reactive oxygen species (ROS) as well as SOD activity was measured in NO-induced apoptosis. In vitro and in vivo effect of NO treatment on mammary cancer stem cells (MCSCs) was assessed. NO induced mitocondria-mediated apoptosis in all AA but not in CA TN breast cancer cells. We found significant TUNEL-positive cells, cleavage of Bax and caspase-3 activation as well as depolarization mitochondrial membrane potential only in AA TN breast cancer cells exposed to NO. Inhibition of Bax cleavage and quenching of ROS partially inhibited NO-induced apoptosis in AA TN cells. Increase in ROS coincided with reduction in SOD activity in AA TN breast cancer cells. Furthermore, NO treatment of AA TN breast cancer cells dramatically reduced aldehyde dehydrogenase1 (ALDH1) expressing MCSCs and xenograft formation but not in breast cancer cells from CA origin. Ethnic differences in breast

  18. Cyclooxygenase inhibitors induce apoptosis in oral cavity cancer cells by increased expression of nonsteroidal anti-inflammatory drug-activated gene

    International Nuclear Information System (INIS)

    Kim, Kyung-Su; Yoon, Joo-Heon; Kim, Jin Kook; Baek, Seung Joon; Eling, Thomas E.; Lee, Won Jae; Ryu, Ji-Hwan; Lee, Jeung Gweon; Lee, Joo-Hwan; Yoo, Jong-Bum

    2004-01-01

    We have investigated whether NAG-1 is induced in oral cavity cancer cells by various NSAIDs and if apoptosis induced by NSAIDs can be linked directly with the induction of NAG-1. NAG-1 expression was increased by diclofenac, aceclofenac, indomethacin, ibuprofen, and sulindac sulfide, in the order of NAG-1 induction, but not by acetaminophen, piroxicam or NS-398. Diclofenac was the most effective NAG-1 inducer. Incubation with diclofenac inhibited cell proliferation and induced apoptosis. The expression of NAG-1 was observed in advance of the induction of apoptosis. Conditioned medium from NAG-1-overexpressing Drosophila cells inhibited SCC 1483 cells proliferation and induced apoptosis. In summary, some NSAIDs induce NAG-1 expression in oral cavity cancer cells and the induced NAG-1 protein appears to mediate apoptosis. Therefore, NSAIDs may be considered as a possible chemopreventive agent against oral cavity cancer

  19. Cellular prion protein contributes to LS 174T colon cancer cell carcinogenesis by increasing invasiveness and resistance against doxorubicin-induced apoptosis.

    Science.gov (United States)

    Chieng, Cornelius Kwang-Lee; Say, Yee-How

    2015-09-01

    As the cellular prion protein (PrP(C)) has been implicated in carcinogenesis, we aimed to investigate the effects of cancer cell-specific PrP(C) overexpression from the invasion, metastasis, and apoptosis aspects, by performing cell motility assays, cell proliferation assays under anchorage-dependent and anchorage-independent conditions, and apoptosis evasion when subjected to multiple anti-cancer drugs. Overexpression of PrP(C) in LS 174T was achieved by stable transfection. PrP(C) overexpression was shown to increase cell proliferation in anchorage-dependent and anchorage-independent manners, as shown by more viable cells in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, more colonies formed in soft agar assay and increased resistance to anoikis in poly-2-hydroxyethyl methacrylate-coated surface. PrP(C) overexpression also increased cell motility and invasiveness of LS 174T. Cell adhesion to extracellular matrix using collagen- and fibronectin-coated surfaces revealed increased cell attachment in LS 174T cells overexpressing PrP(C). Analysis of apoptotic and necrotic cells by propidium iodide/annexin V-fluorescein isothiocyanate microscopy and 7-amino-actinomycin D/annexin V-phycoerythrin flow cytometry revealed that PrP(C) overexpression attenuated doxorubicin-induced apoptosis. Human apoptosis antibody array with 35 apoptosis-related proteins revealed that three inhibitor of apoptosis proteins (IAPs)-survivin, X-linked inhibitor of apoptosis protein (XIAP), and cellular inhibitor of apoptosis protein-1 (cIAP-1)-were upregulated in LS 174T cells overexpressing PrP(C) in doxorubicin-induced apoptosis. In conclusion, the overexpression of PrP(C) could enhance the invasiveness and survival of LS 174T colorectal cancer cells, indicating that PrP(C) plays a role in colorectal cancer biology.

  20. SU-E-T-245: MR Guided Focused Ultrasound Increased PARP Related Apoptosis On Prostate Cancer in Vivo

    Energy Technology Data Exchange (ETDEWEB)

    Chen, L; Chen, X; Cvetkovic, D; Gupta, R; Yang, D; Ma, C [Fox Chase Cancer Center, Philadelphia, PA (United States)

    2014-06-01

    Purpose: Our previous study demonstrated that significant tumor growth delay was observed in the mice treated with pulsed high intensity focused ultrasound (pHIFU). The purpose of this study is to understand the cell killing mechanisms of pHIFU. Methods: Prostate cancer cells (LNCaP), were grown orthotopically in 17 nude mice. Tumor-bearing mice were treated using pHIFU with an acoustic power of 25W, pulse width 100msec and 300 pulses in one sonication under MR guidance. Mutiple sonications were used to cover the whole tumor volume. Temperature (less than 40 degree centigrade in the focal spot) was monitored using MR thermometry. Animals were euthanized at pre-determined time points (n=2) after treatment: 0 hours; 6 hrs; 24 hrs; 48 hrs; 4 days and 7 days. Two tumorbearing mice were used as control. Three tumor-bearing mice were treated with radiation (RT, 2 Gy) using 6 MV photon beams. RT treated mice were euthanized at 0 hr, 6 hrs and 24 hrs. The tumors were processed for immunohistochemical (IHC) staining for PARP (a surrogate of apoptosis). A multispectral imaging analysis system was used to quantify the expression of PARP staining. Cell apoptosis was calculated based on the PARP expression level, which is the intensity of the DAB reaction. Results: Our data showed that PARP related apoptosis peaked at 48 hrs and 7 days in pHIFU treated mice, which is comparable to that for the RT group at 24 hrs. The preliminary results from this study were consistent with our previous study on tumor growth delay using pHIFU. Conclusion: Our results demonstrated that non-thermal pHIFU increased apoptotic tumor cell death through the PARP related pathway. MR guided pHIFU may have a great potential as a safe, noninvasive treatment modality for cancer therapy. This treatment modality might be able to synergize with PARP inhibitors to achieve better result.

  1. Perinatal iodine deficiency and hypothyroidism increase cell apoptosis and alter doublecortin and reelin protein expressions in rat cerebellum.

    Science.gov (United States)

    Wang, Yi; Zhong, Jiapeng; Xu, Hongde; Wei, Wei; Dong, Jing; Yu, Fei; Wang, Yuan; Gong, Jian; Shan, Zhongyan; Teng, Weiping; Chen, Jie

    2012-05-01

    Adequate thyroid hormone is critical for cerebellar development. Developmental hypothyroidism induced by iodine deficiency during the perinatal period results in permanent impairments of cerebellar development with an unclear mechanism. In the present study we investigated effects of perinatal iodine deficiency and hypothyroidism on cerebellar cell apoptosis, doublecortin (Dcx) and reelin. Apoptosis is an essential part of neural development. Dcx and reelin are two important molecules involved in neuronal migration, structure, and development in cerebellum. Two developmental rat models were created by administering dam rats with either iodine-deficient diet or propylthiouracil (PTU, 5 ppm or 15 ppm)-added drinking water from gestational day (GD) 6 until postnatal day (PND) 28. TUNEL assay and protein levels of Dcx and reelin in cerebella were assessed on PND14, 21 and 28. Apoptotic cells were increased in the iodine-deficient and PTU-treated rats. Dcx protein levels in the cerebella of iodine-deficient and PTU-treated rats were significantly downregulated on PND14. Interestingly, iodine deficiency and PTU treatment upregulated the levels of Dcx protein on PND21 and 28. Reelin expressions in iodine-deficient and PTU-treated rats were significantly decreased on PND14 and 21. On PND28, reelin expressions of three treated groups were still lower than control group, although without significant difference. These findings may implicate alterations in cell apoptosis and levels of Dcx and reelin in the impairments of cerebellar development induced by developmental iodine deficiency and hypothyroidism. Copyright © 2012 IMSS. Published by Elsevier Inc. All rights reserved.

  2. Increased lymphocyte apoptosis in mouse models of colitis upon ABT-737 treatment is dependent upon BIM expression.

    Science.gov (United States)

    Lutz, C; Mozaffari, M; Tosevski, V; Caj, M; Cippà, P; McRae, B L; Graff, C L; Rogler, G; Fried, M; Hausmann, M

    2015-08-01

    Exaggerated activation of lymphocytes contributes to the pathogenesis of inflammatory bowel disease (IBD). Medical therapies are linked to the BCL-2 family-mediated apoptosis. Imbalance in BCL-2 family proteins may cause failure in therapeutic responses. We investigated the role of BCL-2 inhibitor ABT-737 for lymphocyte apoptosis in mice under inflammatory conditions. B.6129P2-interleukin (IL)-10(tm1Cgn) /J (IL-10(-/-) ) weighing 25-30 g with ongoing colitis were used. Fifty mg/kg/day ABT-737 was injected intraperitoneally (i.p.). Haematological analyses were performed with an ADVIA 2120 flow cytometer and mass cytometry with a CyTOF 2. Following i.p. administration, ABT-737 was detected in both spontaneous and acute colitis in peripheral blood (PBL) and colon tissue. Treatment led to lymphopenia. CD4(+) CD44(+) CD62L(+) central memory and CD8(+) , CD44(+) CD62L(-) central memory T cells were decreased in PBL upon ABT-737 compared to vehicle-receiving controls. Increased apoptosis upon ABT-737 was determined in blood lymphocytes, splenocytes and Peyer's patches and was accompanied by a decrease in TNF and IL-1B. ABT-737 positively altered the colonic mucosa and ameliorated inflammation, as shown by colonoscopy, histology and colon length. A decreased BIM/BCL-2 ratio or absence of BIM in both Bim(-) (/) (-) and Il10(-) (/) (-) × Bim(-) (/) (-) impeded the protective effect of ABT-737. The BIM/BCL-2 ratio decreased with age and during the course of treatment. Thus, long-term treatment resulted in adapted TNF levels and macroscopic mucosal damage. ABT-737 was efficacious in diminishing lymphocytes and ameliorating colitis in a BIM-dependent manner. Regulation of inappropriate survival of lymphocytes by ABT-737 may provide a therapeutic strategy in IBD. © 2015 British Society for Immunology.

  3. SU-E-T-245: MR Guided Focused Ultrasound Increased PARP Related Apoptosis On Prostate Cancer in Vivo

    International Nuclear Information System (INIS)

    Chen, L; Chen, X; Cvetkovic, D; Gupta, R; Yang, D; Ma, C

    2014-01-01

    Purpose: Our previous study demonstrated that significant tumor growth delay was observed in the mice treated with pulsed high intensity focused ultrasound (pHIFU). The purpose of this study is to understand the cell killing mechanisms of pHIFU. Methods: Prostate cancer cells (LNCaP), were grown orthotopically in 17 nude mice. Tumor-bearing mice were treated using pHIFU with an acoustic power of 25W, pulse width 100msec and 300 pulses in one sonication under MR guidance. Mutiple sonications were used to cover the whole tumor volume. Temperature (less than 40 degree centigrade in the focal spot) was monitored using MR thermometry. Animals were euthanized at pre-determined time points (n=2) after treatment: 0 hours; 6 hrs; 24 hrs; 48 hrs; 4 days and 7 days. Two tumorbearing mice were used as control. Three tumor-bearing mice were treated with radiation (RT, 2 Gy) using 6 MV photon beams. RT treated mice were euthanized at 0 hr, 6 hrs and 24 hrs. The tumors were processed for immunohistochemical (IHC) staining for PARP (a surrogate of apoptosis). A multispectral imaging analysis system was used to quantify the expression of PARP staining. Cell apoptosis was calculated based on the PARP expression level, which is the intensity of the DAB reaction. Results: Our data showed that PARP related apoptosis peaked at 48 hrs and 7 days in pHIFU treated mice, which is comparable to that for the RT group at 24 hrs. The preliminary results from this study were consistent with our previous study on tumor growth delay using pHIFU. Conclusion: Our results demonstrated that non-thermal pHIFU increased apoptotic tumor cell death through the PARP related pathway. MR guided pHIFU may have a great potential as a safe, noninvasive treatment modality for cancer therapy. This treatment modality might be able to synergize with PARP inhibitors to achieve better result

  4. Increased oxidative stress and apoptosis in the hypothalamus of diabetic male mice in the insulin receptor substrate-2 knockout model.

    Science.gov (United States)

    Baquedano, Eva; Burgos-Ramos, Emma; Canelles, Sandra; González-Rodríguez, Agueda; Chowen, Julie A; Argente, Jesús; Barrios, Vicente; Valverde, Angela M; Frago, Laura M

    2016-05-01

    Insulin receptor substrate-2-deficient (IRS2(-/-)) mice are considered a good model to study the development of diabetes because IRS proteins mediate the pleiotropic effects of insulin-like growth factor-I (IGF-I) and insulin on metabolism, mitogenesis and cell survival. The hypothalamus might play a key role in the early onset of diabetes, owing to its involvement in the control of glucose homeostasis and energy balance. Because some inflammatory markers are elevated in the hypothalamus of diabetic IRS2(-/-) mice, our aim was to analyze whether the diabetes associated with the absence of IRS2 results in hypothalamic injury and to analyze the intracellular mechanisms involved. Only diabetic IRS2(-/-) mice showed increased cell death and activation of caspase-8 and -3 in the hypothalamus. Regulators of apoptosis such as FADD, Bcl-2, Bcl-xL and p53 were also increased, whereas p-IκB and c-FLIPL were decreased. This was accompanied by increased levels of Nox-4 and catalase, enzymes involved in oxidative stress. In summary, the hypothalamus of diabetic IRS2(-/-) mice showed an increase in oxidative stress and inflammatory markers that finally resulted in cell death via substantial activation of the extrinsic apoptotic pathway. Conversely, non-diabetic IRS2(-/-) mice did not show cell death in the hypothalamus, possibly owing to an increase in the levels of circulating IGF-I and in the enhanced hypothalamic IGF-IR phosphorylation that would lead to the stimulation of survival pathways. In conclusion, diabetes in IRS2-deficient male mice is associated with increased oxidative stress and apoptosis in the hypothalamus. © 2016. Published by The Company of Biologists Ltd.

  5. Apoptosis is increased and cell proliferation is decreased in out-of-phase endometria from infertile and recurrent abortion patients

    Directory of Open Access Journals (Sweden)

    Irigoyen Marcela

    2010-10-01

    Full Text Available Abstract Background Various endometrial abnormalities have been associated with luteal phase deficiency: a significant dyssynchrony in the maturation of the glandular epithelium and the stroma and a prevalence of out-of-phase endometrial biopsy specimens. Out-of phase endometrium is a controversial disorder related to failed implantation, infertility and early pregnancy loss. Given that the regulation of the apoptotic process in endometrium of luteal phase deficiency is still unknown, the aim of this study was to evaluate cell proliferation, apoptosis and the levels of the main effector caspase, caspase-3 in the luteal in-phase and out-of-phase endometrium. Methods Thirty-seven endometrial samples from sterile or recurrent abortion patients were included in this study: 21 in-phase samples (controls and 16 samples with out-of-phase endometrium. Biopsy specimens of eutopic endometrium were obtained from all subjects during days 21-25 of the menstrual cycle. The endometrium with endometrial maturity of cycle day 25 or less at the time of menstruation was considered out-of phase. Endometrial tissues were fixed in 10% buffered formaldehyde. For apoptosis quantification, sections were processed for in situ immunohistochemical localization of nuclei exhibiting DNA fragmentation, by the terminal deoxynucleotidyl transferase (TdT-mediated dUTP digoxygenin nick-end labeling (TUNEL technique. Expressions of Proliferating Cell Nuclear Antigen (PCNA as a marker of cell proliferation, and of cleaved caspase-3 as a marker of apoptosis, were assessed by immunohistochemistry in the luteal in-phase and out-of-phase endometrium from infertile and recurrent abortion patients. Results Luteal out-of-phase endometrium had increased apoptosis levels compared to in-phase endometrium (p Conclusions this study represents the first report describing variations at the cell proliferation and cell death levels in the out-of-phase endometrium in comparison with in

  6. Theoretical perspectives on central chemosensitivity: CO2/H+-sensitive neurons in the locus coeruleus.

    Directory of Open Access Journals (Sweden)

    Maria C Quintero

    2017-12-01

    Full Text Available Central chemoreceptors are highly sensitive neurons that respond to changes in pH and CO2 levels. An increase in CO2/H+ typically reflects a rise in the firing rate of these neurons, which stimulates an increase in ventilation. Here, we present an ionic current model that reproduces the basic electrophysiological activity of individual CO2/H+-sensitive neurons from the locus coeruleus (LC. We used this model to explore chemoreceptor discharge patterns in response to electrical and chemical stimuli. The modeled neurons showed both stimulus-evoked activity and spontaneous activity under physiological parameters. Neuronal responses to electrical and chemical stimulation showed specific firing patterns of spike frequency adaptation, postinhibitory rebound, and post-stimulation recovery. Conversely, the response to chemical stimulation alone (based on physiological CO2/H+ changes, in the absence of external depolarizing stimulation, showed no signs of postinhibitory rebound or post-stimulation recovery, and no depolarizing sag. A sensitivity analysis for the firing-rate response to the different stimuli revealed that the contribution of an applied stimulus current exceeded that of the chemical signals. The firing-rate response increased indefinitely with injected depolarizing current, but reached saturation with chemical stimuli. Our computational model reproduced the regular pacemaker-like spiking pattern, action potential shape, and most of the membrane properties that characterize CO2/H+-sensitive neurons from the locus coeruleus. This validates the model and highlights its potential as a tool for studying the cellular mechanisms underlying the altered central chemosensitivity present in a variety of disorders such as sudden infant death syndrome, depression, and anxiety. In addition, the model results suggest that small external electrical signals play a greater role in determining the chemosensitive response to changes in CO2/H+ than previously

  7. Targeting MCL-1 sensitizes human esophageal squamous cell carcinoma cells to cisplatin-induced apoptosis.

    Science.gov (United States)

    Yu, Xinfang; Li, Wei; Xia, Zhenkun; Xie, Li; Ma, Xiaolong; Liang, Qi; Liu, Lijun; Wang, Jian; Zhou, Xinmin; Yang, Yifeng; Liu, Haidan

    2017-06-28

    Esophageal squamous cell carcinoma (ESCC) is one of the most lethal malignancies in China and is an exceptionally drug-resistant tumor with a 5-year survival rate less than 15%. Cisplatin is the most commonly used conventional chemotherapeutic drug for the treatment of ESCC, but some patients have a poor response to cisplatin-based chemotherapy. New strategies that could enhance chemosensitivity to cisplatin are needed. We used reverse transcription-RCR (RT-PCR), immunoblot, immunohistochemical (IHC) staining, anchorage-dependent and -independent growth assays, co-immunoprecipitation (Co-IP) assay, RNA interference and in vivo tumor growth assay to study the expression of MCL-1 in ESCCs and the response of ESCC cells to cisplatin. The present study showed that MCL-1 expression was significantly increased in ESCC tissues compared to normal adjacent tissues and was associated with depth of invasion and lymph node metastasis. Knockdown of MCL-1 produced significant chemosensitization to cisplatin in association with caspase-3 activation and PARP cleavage in KYSE150 and KYSE510 cells. The selective MCL-1 inhibitor UMI-77 caused dissociation of MCL-1 from the proapoptotic protein BAX and BAK, and enhanced KYSE150 and KYSE510 cells to cisplatin-induced apoptosis accompanied by caspase-3 activation and PARP cleavage. The current study suggests that MCL-1 contributes to the development of ESCC and is a promising therapeutic target for chemosensitization of ESCC cells to cisplatin. This might provide a scientific basis for developing effective approaches to treat the subset of ESCCs patients with MCL-1 overexpression.

  8. Platelet rich plasma (PRP) induces chondroprotection via increasing autophagy, anti-inflammatory markers, and decreasing apoptosis in human osteoarthritic cartilage

    Energy Technology Data Exchange (ETDEWEB)

    Moussa, Mayssam, E-mail: Moussa-mayssam@hotmail.com [Regenerative medicine and inflammation Laboratory, Faculty of Medicine, Saint-Joseph University, Beirut (Lebanon); Lajeunesse, Daniel, E-mail: daniel.lajeunesse@umontreal.ca [Research Centre in Osteoarthritis, Research Centre in Monteral University (Canada); Hilal, George, E-mail: George2266@gmail.com [Cancer and metabolism Laboratory, Faculty of Medicine, Saint-Joseph University, Beirut (Lebanon); El Atat, Oula, E-mail: oulaatat@hotmail.com [Regenerative medicine and inflammation Laboratory, Faculty of Medicine, Saint-Joseph University, Beirut (Lebanon); Haykal, Gaby, E-mail: Gaby.haykal@hdf.usj.edu.lb [Hotel Dieu de France, Faculty of Medicine, Saint-Joseph University, Beirut (Lebanon); Serhal, Rim, E-mail: rim.basbous@gmail.com [Regenerative medicine and inflammation Laboratory, Faculty of Medicine, Saint-Joseph University, Beirut (Lebanon); Chalhoub, Antonio, E-mail: Mava.o@hotmail.com [Carantina Hospital, Beirut (Lebanon); Khalil, Charbel, E-mail: charbelk3@hotmail.com [Regenerative medicine and inflammation Laboratory, Faculty of Medicine, Saint-Joseph University, Beirut (Lebanon); Alaaeddine, Nada, E-mail: Nada.aladdin@gmail.com [Regenerative medicine and inflammation Laboratory, Faculty of Medicine, Saint-Joseph University, Beirut (Lebanon)

    2017-03-01

    Objectives: Autophagy constitutes a defense mechanism to overcome aging and apoptosis in osteoarthritic cartilage. Several cytokines and transcription factors are linked to autophagy and play an important role in the degradative cascade in osteoarthritis (OA). Cell therapy such as platelet rich plasma (PRP) has recently emerged as a promising therapeutic tool for many diseases including OA. However, its mechanism of action on improving cartilage repair remains to be determined. The purpose of this study is to investigate the effect of PRP on osteoarthritic chondrocytes and to elucidate the mechanism by which PRP contributes to cartilage regeneration. Methods: Osteoarthritic chondrocytes were co-cultured with an increasing concentration of PRP obtained from healthy donors. The effect of PRP on the proliferation of chondrocytes was performed using cell counting and WST8 proliferation assays. Autophagy, apoptosis and intracellular level of IL-4, IL-10, and IL-13 were determined using flow cytometry analyses. Autophagy markers BECLIN and LC3II were also determined using quantitative polymerase chain reaction (qPCR). qPCR and ELISA were used to measure the expression of ADAMDTS-5, MMP3, MMP13, TIMP-1–2–3, aggregan, Collagen type 2, TGF-β, Cox-2, Il-6, FOXO1, FOXO3, and HIF-1 in tissues and co-cultured media. Results: PRP increased significantly the proliferation of chondrocytes, decreased apoptosis and increased autophagy and its markers along with its regulators FOXO1, FOXO3 and HIF-1 in osteoarthritic chondrocytes. Furthermore, PRP caused a dose-dependent significant decrease in MMP3, MMP13, and ADAMTS-5, IL-6 and COX-2 while increasing TGF-β, aggregan, and collagen type 2, TIMPs and intracellular IL-4, IL-10, IL-13. Conclusion: These results suggest that PRP could be a potential therapeutic tool for the treatment of OA. - Highlights: • Platelet Rich Plasma is suggested as a new treatment for osteoarthritis. • The proposed therapeutic effect is

  9. Knockdown of MACC1 expression increases cisplatin sensitivity in cisplatin-resistant epithelial ovarian cancer cells.

    Science.gov (United States)

    Zhang, Ruitao; Shi, Huirong; Ren, Fang; Li, Xia; Zhang, Minghui; Feng, Wei; Jia, Yanyan

    2016-04-01

    Abnormal expression of metastasis-associated in colon cancer 1 (MACC1) was found to be closely associated with several types of malignant tumors. The present study aimed to verify the relationship between MACC1 and cisplatin resistance in ovarian cancer cells and the possible mechanisms, which was implemented by inhibition of the expression of MACC1 in cisplatin-resistant human ovarian cancer cell lines A2780/DDP and COC1/DDP. MACC1 shRNA eukaryotic plasmids and negative control plasmids were transfected into A2780/DDP and COC1/DDP cells, respectively, while A2780/DDP and COC1/DDP cells were used as blank controls. Western blotting and sqRT-PCR were used to detect the expression of MACC1 in the different cell groups. Different concentrations of cispaltin (0, 10, 20, 30, 40, 50 and 60 µmol/l) were used to treat the cell groups, respectively, and then the chemosensitivity of cisplatin and cell apoptosis were examined by MTT and flow cytometry, respectively. The activity of caspase-3 was determined by spectrophotometry. Expression levels of p-ERK1/2, permeability glycoprotein (P-gp), B-cell lymphoma 2 (Bcl-2), Bcl-XL, Bax and Bad protein were detected in the different ovarian cancer cells by western blotting. After MACC1 knockdown, the chemosensitivity of cisplatin in the ovarian cancer cells was enhanced, and the cell growth inhibition and apoptosis rates were increased. The expression levels of Bax and Bad were upregulated, the activity of caspase-3 was increased, while the expression levels of p-ERK1/2, P-gp, Bcl-2 and Bcl-XL were downregulated as a result of MACC1 inhibition. These results indicate that inhibition of MACC1 improves the chemosensitivity of cisplatin in epithelial ovarian cancer cells, through the regulation of the ERK1/2 signaling pathway on P-gp and its downstream apoptosis proteins.

  10. Histamine deficiency exacerbates myocardial injury in acute myocardial infarction through impaired macrophage infiltration and increased cardiomyocyte apoptosis

    Science.gov (United States)

    Deng, Long; Hong, Tao; Lin, Jinyi; Ding, Suling; Huang, Zheyong; Chen, Jinmiao; Jia, Jianguo; Zou, Yunzeng; Wang, Timothy C.; Yang, Xiangdong; Ge, Junbo

    2015-01-01

    Histamine is a biogenic amine that is widely distributed and has multiple functions, but the role it plays in acute myocardial infarction (AMI) remains unclear. In this study, we investigated the origin and contribution of endogenous histamine to AMI. Histidine decarboxylase (HDC) is the unique enzyme responsible for histamine generation. Using HDC-EGFP bacterial artificial chromosome (BAC) transgenic mice in which EGFP expression is controlled by the HDC promoter, we identified HDC expression primarily in CD11b+Gr-1+ immature myeloid cells (IMCs) that markedly increase in the early stages of AMI. Deficiency of histamine in HDC knockout mice (HDC−/−) reduced cardiac function and exacerbated the injury of infarcted heart. Furthermore, administering either an H1 receptor antagonist (pyrilamine) or an H2 receptor antagonist (cimetidine) demonstrated a protective effect of histamine against myocardial injury. The results of in vivo and in vitro assays showed that histamine deficiency promotes the apoptosis of cardiomyocytes and inhibits macrophage infiltration. In conclusion, CD11b+Gr-1+ IMCs are the predominant HDC-expressing sites in AMI, and histamine plays a protective role in the process of AMI through inhibition of cardiomyocyte apoptosis and facilitation of macrophage infiltration. PMID:26278136

  11. Histamine deficiency exacerbates myocardial injury in acute myocardial infarction through impaired macrophage infiltration and increased cardiomyocyte apoptosis.

    Science.gov (United States)

    Deng, Long; Hong, Tao; Lin, Jinyi; Ding, Suling; Huang, Zheyong; Chen, Jinmiao; Jia, Jianguo; Zou, Yunzeng; Wang, Timothy C; Yang, Xiangdong; Ge, Junbo

    2015-08-17

    Histamine is a biogenic amine that is widely distributed and has multiple functions, but the role it plays in acute myocardial infarction (AMI) remains unclear. In this study, we investigated the origin and contribution of endogenous histamine to AMI. Histidine decarboxylase (HDC) is the unique enzyme responsible for histamine generation. Using HDC-EGFP bacterial artificial chromosome (BAC) transgenic mice in which EGFP expression is controlled by the HDC promoter, we identified HDC expression primarily in CD11b(+)Gr-1(+) immature myeloid cells (IMCs) that markedly increase in the early stages of AMI. Deficiency of histamine in HDC knockout mice (HDC(-/-)) reduced cardiac function and exacerbated the injury of infarcted heart. Furthermore, administering either an H1 receptor antagonist (pyrilamine) or an H2 receptor antagonist (cimetidine) demonstrated a protective effect of histamine against myocardial injury. The results of in vivo and in vitro assays showed that histamine deficiency promotes the apoptosis of cardiomyocytes and inhibits macrophage infiltration. In conclusion, CD11b(+)Gr-1(+) IMCs are the predominant HDC-expressing sites in AMI, and histamine plays a protective role in the process of AMI through inhibition of cardiomyocyte apoptosis and facilitation of macrophage infiltration.

  12. The potent anti-inflammatory agent escin does not increase corticosterone secretion and immune cell apoptosis in mice.

    Science.gov (United States)

    Zhang, Leiming; Wang, Hongsheng; Fan, Huaying; Wang, Tian; Jiang, Na; Yu, Pengfei; Fu, Fenghua

    2011-09-01

    Escin exerts potent glucocorticoid-like anti-inflammatory effects. The aim of this study was to investigate whether the anti-inflammatory effect of escin is through the up-regulation of glucocorticoids and if escin induces pathological changes in immune organs. Mice were administrated with escin intravenously for 7 days before observing the relevant parameters. The results showed that escin exhibits a potent anti-inflammatory effect, but does not increase corticosterone secretion in mice, and does not increase immune cell apoptosis in the spleen and thymus of mice. These findings suggest that the anti-inflammatory effect of escin is not dependent on the release of corticosterone. Copyright © 2011 Elsevier B.V. All rights reserved.

  13. Forskolin protects keratinocytes from ultraviolet (UV) B-induced apoptosis and increases DNA repair independent of its effects on melanogenesis

    Science.gov (United States)

    Passeron, Thierry; Namiki, Takeshi; Passeron, Hélène; Le Pape, Elodie; Hearing, Vincent J.

    2009-01-01

    Melanin pigments provide efficient protection against ultraviolet (UV) B radiation but DNA repair also plays a key role in eliminating UV-induced damage and preventing the development of skin cancers. In this study, we demonstrate that forskolin, an agent that increases intracellular levels of cAMP, protects keratinocytes from UVB-induced apoptosis independently from the amount of melanin in the skin. Forskolin enhances the removal of the two major types of UVB-induced DNA damage, cyclobutane pyrimidine dimers and 6,4-photoproducts, by facilitating DNA repair. These findings suggest new preventive approaches with topical formulations of forskolin or other bioactive agents that could be applied to the skin before sun exposure to increase its ability to repair DNA damage. PMID:18580960

  14. Forskolin protects keratinocytes from UVB-induced apoptosis and increases DNA repair independent of its effects on melanogenesis.

    Science.gov (United States)

    Passeron, Thierry; Namiki, Takeshi; Passeron, Hélène J; Le Pape, Elodie; Hearing, Vincent J

    2009-01-01

    Melanin pigments provide efficient protection against ultraviolet B (UVB) radiation but DNA repair also plays a key role in eliminating UV-induced damage and preventing the development of skin cancers. In this study, we demonstrate that forskolin (FSK), an agent that increases intracellular levels of cAMP, protects keratinocytes from UVB-induced apoptosis independently from the amount of melanin in the skin. FSK enhances the removal of the two major types of UVB-induced DNA damage, cyclobutane pyrimidine dimers and 6,4-photoproducts, by facilitating DNA repair. These findings suggest new preventive approaches with topical formulations of FSK or other bioactive agents that could be applied to the skin before sun exposure to increase its ability to repair DNA damage.

  15. Salinomycin sensitizes antimitotic drugs-treated cancer cells by increasing apoptosis via the prevention of G2 arrest

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Ju-Hwa; Yoo, Hye-In; Kang, Han Sung; Ro, Jungsil [Research Institute, National Cancer Center, Ilsan-gu, Goyang-si, Gyeonggi-do (Korea, Republic of); Yoon, Sungpil, E-mail: yoons@ncc.re.kr [Research Institute, National Cancer Center, Ilsan-gu, Goyang-si, Gyeonggi-do (Korea, Republic of)

    2012-02-03

    Highlights: Black-Right-Pointing-Pointer Sal sensitizes antimitotic drugs-treated cancer cells. Black-Right-Pointing-Pointer Sal sensitizes them by prevention of G2 arrest and reduced cyclin D1 levels. Black-Right-Pointing-Pointer Sal also sensitizes them by increasing DNA damage and reducing p21 level. Black-Right-Pointing-Pointer A low concentration of Sal effectively sensitized the cancer cells to antimitotic drugs. -- Abstract: Here, we investigated whether Sal could sensitize cancer cells to antimitotic drugs. We demonstrated that Sal sensitized paclitaxcel (PAC)-, docetaxcel (DOC)-, vinblastin (VIN)-, or colchicine (COL)-treated cancer cell lines, suggesting that Sal has the ability to sensitize the cells to any form of microtubule-targeting drugs. Sensitization to the antimitotic drugs could be achieved with very low concentrations of Sal, suggesting that there is a possibility to minimize Sal toxicity associated with human cancer patient treatments. Sensitization by Sal increased apoptosis, which was observed by C-PARP production. Sal sensitized the cancer cells to antimitotic drugs by preventing G2 arrest, suggesting that Sal contributes to the induction of mitotic catastrophe. Sal generally reduced cyclin D1 levels in PAC-, DOC-, and VIN-treated cells. In addition, Sal treatment increased pH2AX levels and reduced p21 levels in antimitotic drugs-treated cells. These observations suggest that the mechanisms underlying Sal sensitization to DNA-damaging compounds, radiation, and microtubule-targeting drugs are similar. Our data demonstrated that Sal sensitizes cancer cells to antimitotic drugs by increasing apoptosis through the prevention of G2 arrest via conserved Sal-sensitization mechanisms. These results may contribute to the development of Sal-based chemotherapy for cancer patients treated with antimitotic drugs.

  16. Heterogeneity of tumor chemosensitivity in ovarian epithelial cancer revealed using the adenosine triphosphate-tumor chemosensitivity assay

    Science.gov (United States)

    ZHANG, JIN; LI, HONGXIA

    2015-01-01

    Ovarian cancer has a poor prognosis, primarily due to the heterogeneity in chemosensitivity among patients. In the present study, this heterogeneity was evaluated in ovarian epithelial cancer (OEC) using an in vitro adenosine triphosphate tumor chemosensitivity assay (ATP-TCA). Specimens were collected from 80 patients who underwent cytoreductive surgery. Viable ovarian cancer cells obtained from malignant tissues were tested for sensitivity to paclitaxel (PTX), carboplatin (CBP), topotecan (TPT), gemcitabine (GEM), docetaxel (TXT), etoposide, bleomycin and 4-hydroperoxycyclophosphamide using ATP-TCA. The sensitivity, specificity, positive predictive value and negative predictive value for the clinical chemotherapy sensitivity of OEC were 88.6, 77.8, 83 and 84.8%, respectively. PTX demonstrated the highest sensitivity of all agents tested (82.5% in all specimens, 85.7% in recurrent specimens), followed by CBP (58.8 and 60.7%, respectively). The sensitivities to PTX and docetaxel (P<0.001) were correlated, in addition to those of CBP, TPT and GEM (P<0.001). Early-stage (I/II) and high- to mildly-differentiated OEC specimens revealed a lower chemosensitivity than advanced-stage (III) or low-differentiated specimens, respectively. The present study indicated that ATP-TCA is an effective method for guiding the choice of chemotherapy drugs. Notable heterogeneity of chemosensitivity was observed in the OEC specimens. PMID:26137074

  17. TLQP-21 protects human umbilical vein endothelial cells against high-glucose-induced apoptosis by increasing G6PD expression.

    Directory of Open Access Journals (Sweden)

    Wei Zhang

    Full Text Available Hyperglycemia causes oxidative stress that could damage vascular endothelial cells, leading to cardiovascular complications. The Vgf gene was identified as a nerve growth factor-responsive gene, and its protein product, VGF, is characterized by the presence of partially cleaved products. One of the VGF-derived peptides is TLQP-21, which is composed of 21 amino acids (residues 556-576. Past studies have reported that TLQP-21 could stimulate insulin secretion in pancreatic cells and protect these cells from apoptosis, which suggests that TLQP-21 has a potential function in diabetes therapy. Here, we explore the protective role of TLQP-21 against the high glucose-mediated injury of vascular endothelial cells. Using human umbilical vascular endothelial cells (HUVECs, we demonstrated that TLQP-21 (10 or 50 nM dose-dependently prevented apoptosis under high-glucose (30 mmol/L conditions (the normal glucose concentration is 5.6 mmol/L. TLQP-21 enhanced the expression of NAPDH, resulting in upregulation of glutathione (GSH and a reduction in the levels of reactive oxygen species (ROS. TLQP-21 also upregulated the expression of glucose-6-phosphate dehydrogenase (G6PD, which is known as the main source of NADPH. Knockdown of G6PD almost completely blocked the increase of NADPH induced by TLQP-21, indicating that TLQP-21 functions mainly through G6PD to promote NADPH generation. In conclusion, TLQP-21 could increase G6PD expression, which in turn may increase the synthesis of NADPH and GSH, thereby partially restoring the redox status of vascular endothelial cells under high glucose injury. We propose that TLQP-21 is a promising drug for diabetes therapy.

  18. Chemosensitization of cancer in vitro and in vivo by nitric oxide signaling.

    Science.gov (United States)

    Frederiksen, Lisa J; Sullivan, Richard; Maxwell, Lori R; Macdonald-Goodfellow, Shannyn K; Adams, Michael A; Bennett, Brian M; Siemens, D Robert; Graham, Charles H

    2007-04-01

    Hypoxia contributes to drug resistance in solid cancers, and studies have revealed that low concentrations of nitric oxide (NO) mimetics attenuate hypoxia-induced drug resistance in tumor cells in vitro. Classic NO signaling involves activation of soluble guanylyl cyclase, generation of cyclic GMP (cGMP), and activation of cGMP-dependent protein kinase. Here, we determined whether chemosensitization by NO mimetics requires cGMP-dependent signaling and whether low concentrations of NO mimetics can chemosensitize tumors in vivo. Survival of human prostate and breast cancer cells was assessed by clonogenic assays following exposure to chemotherapeutic agents. The effect of NO mimetics on tumor chemosensitivity in vivo was determined using a mouse xenograft model of human prostate cancer. Drug efflux in vitro was assessed by measuring intracellular doxorubicin-associated fluorescence. Low concentrations of the NO mimetics glyceryl trinitrate (GTN) and isosorbide dinitrate attenuated hypoxia-induced resistance to doxorubicin and paclitaxel. Similar to hypoxia-induced drug resistance, inhibition of various components of the NO signaling pathway increased resistance to doxorubicin, whereas activation of the pathway with 8-bromo-cGMP attenuated hypoxia-induced resistance. Drug efflux was unaffected by hypoxia and inhibitors of drug efflux did not significantly attenuate hypoxia-induced chemoresistance. Compared with mice treated with doxorubicin alone, tumor growth was decreased in mice treated with doxorubicin and a transdermal GTN patch. The presence of GTN and GTN metabolites in plasma samples was confirmed by gas chromatography. Tumor hypoxia induces resistance to anticancer drugs by interfering with endogenous NO signaling and reactivation of NO signaling represents a novel approach to enhance chemotherapy.

  19. Impaired chemosensitivity of mouse dorsal raphe serotonergic neurons overexpressing serotonin 1A (Htr1a) receptors.

    Science.gov (United States)

    Baccini, Gilda; Mlinar, Boris; Audero, Enrica; Gross, Cornelius Thilo; Corradetti, Renato

    2012-01-01

    Serotonergic system participates in a wide range of physiological processes and behaviors, but its role is generally considered as modulatory and noncrucial, especially concerning life-sustaining functions. We recently created a transgenic mouse line in which a functional deficit in serotonin homeostasis due to excessive serotonin autoinhibition was produced by inducing serotonin 1A receptor (Htr1a) overexpression selectively in serotonergic neurons (Htr1a raphe-overexpressing or Htr1a(RO) mice). Htr1a(RO) mice exhibit episodes of autonomic dysregulation, cardiovascular crises and death, resembling those of sudden infant death syndrome (SIDS) and revealing a life-supporting role of serotonergic system in autonomic control. Since midbrain serotonergic neurons are chemosensitive and are implicated in arousal we hypothesized that their chemosensitivity might be impaired in Htr1a(RO) mice. Loose-seal cell-attached recordings in brainstem slices revealed that serotonergic neurons in dorsal raphe nucleus of Htr1a(RO) mice have dramatically reduced responses to hypercapnic challenge as compared with control littermates. In control mice, application of 9% CO(2) produced an increase in firing rate of serotonergic neurons (0.260 ± 0.041 Hz, n=20, p=0.0001) and application of 3% CO(2) decreased their firing rate (-0.142 ± 0.025 Hz, n=17, p=0.0008). In contrast, in Htr1a(RO) mice, firing rate of serotonergic neurons was not significantly changed by 9% CO(2) (0.021 ± 0.034 Hz, n=16, p=0.49) and by 3% CO(2) (0.012 ± 0.046 Hz, n=12, p=0.97). Our findings support the hypothesis that chemosensitivity of midbrain serotonergic neurons provides a physiological mechanism for arousal responses to life-threatening episodes of hypercapnia and that functional impairment, such as excessive autoinhibition, of midbrain serotonergic neuron responses to hypercapnia may contribute to sudden death.

  20. Time-Dependent Increases in Protease Activities for Neuronal Apoptosis in Spinal Cords of Lewis Rats During Development of Acute Experimental Autoimmune Encephalomyelitis

    Science.gov (United States)

    Das, Arabinda; Guyton, M. Kelly; Matzelle, Denise D.; Ray, Swapan K.; Banik, Naren L.

    2008-01-01

    Multiple sclerosis (MS) is characterized by axonal demyelination and neurodegeneration, the latter having been inadequately explored in the MS animal model experimental autoimmune encephalomyelitis (EAE). The purpose of this study was to examine the time-dependent correlation between increased calpain and caspase activities and neurodegeneration in spinal cord tissues from Lewis rats with acute EAE. An increase in TUNEL-positive neurons and internucleosomal DNA fragmentation in EAE spinal cords suggested that neuronal death was a result of apoptosis on days 8–10 following induction of EAE. Increases in calpain expression in EAE correlated with activation of pro-apoptotic proteases, leading to apoptotic cell death beginning on day 8 of EAE, which occurred before the appearance of visible clinical symptoms. Increases in calcineurin expression and decreases in phospho-Bad (p-Bad) suggested Bad activation in apoptosis during acute EAE. Increases in the Bax:Bcl-2 ratio and activation of caspase-9 showed the involvement of mitochondria in apoptosis. Further, caspase-8 activation suggested induction of the death receptor–mediated pathway for apoptosis. Endoplasmic reticulum stress leading to caspase-3 activation was also observed, indicating that multiple apoptotic pathways were activated following EAE induction. In contrast, cell death was mostly a result of necrosis on the later day (day 11), when EAE entered a severe stage. From these findings, we conclude that increases in calpain and caspase activities play crucial roles in neuronal apoptosis during the development of acute EAE. PMID:18521931

  1. The small molecule survivin inhibitor YM155 may be an effective treatment modality for colon cancer through increasing apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Li, Wan Lu, E-mail: lvvlchina@msn.cn [Department of Pathology, Yonsei University Wonju College of Medicine, Wonju (Korea, Republic of); Lee, Mi-Ra, E-mail: mira1125@yonsei.ac.kr [Department of Pathology, Yonsei University Wonju College of Medicine, Wonju (Korea, Republic of); Cho, Mee-Yon, E-mail: meeyon@yonsei.ac.kr [Department of Pathology, Yonsei University Wonju College of Medicine, Wonju (Korea, Republic of); Institute of Genomic Cohort, Yonsei University Wonju College of Medicine, Wonju (Korea, Republic of)

    2016-03-04

    Survivin has a known beneficial role in the survival of both cancer cells and normal cells. Therapies targeting survivin have been proposed as an alternative treatment modality for various tumors; however, finding the proper indication for this toxic therapy is critical for reducing unavoidable side effects. We recently observed that high survivin expression in CD133{sup +} cells is related to chemoresistance in Caco-2 colon cancer cells. However, the effect of survivin-targeted therapy on CD133{sup +} colon cancer is unknown. In this study, we investigated the roles of CD133 and survivin expression in colon cancer biology in vitro and comparatively analyzed the anticancer effects of survivin inhibitor on CD133{sup +} cells (ctrl-siRNA group) and small interfering RNA (siRNA)-induced CD133{sup −} cells (CD133-siRNA group) obtained from a single colon cancer cell line. CD133 knockdown via siRNA transfection did not change the tumorigenicity of cells, although in vitro survivin expression levels in CD133{sup +} cells were higher than those in siRNA-induced CD133{sup −} cells. The transfection procedure seemed to induce survivin expression. Notably, a significant number of CD133{sup −} cells (33.8%) was found in the cell colonies of the CD133-siRNA group. In the cell proliferation assay after treatment, YM155 and a combination of YM155 and 5-fluorouracil (5-FU) proved to be far more effective than 5-FU alone. A significantly increased level of apoptosis was observed with increasing doses of YM155 in all groups. However, significant differences in therapeutic effect and apoptosis among the mock, ctrl-siRNA, and CD133-siRNA groups were not detected. Survivin inhibitor is an effective treatment modality for colon cancer; however, the role of CD133 and the use of survivin expression as a biomarker for this targeted therapy must be verified.

  2. Chemosensitizing indomethacin-conjugated chitosan oligosaccharide nanoparticles for tumor-targeted drug delivery.

    Science.gov (United States)

    Lee, Jae-Young; Termsarasab, Ubonvan; Lee, Mee Yeon; Kim, Dong-Hwan; Lee, Song Yi; Kim, Jung Sun; Cho, Hyun-Jong; Kim, Dae-Duk

    2017-07-15

    A chitosan oligosaccharide (CSO)-indomethacin (IDM) conjugate (CI) was synthesized to fabricate chemosensitizing nanoparticles (NPs) for tumor-targeted drug delivery. IDM was conjugated to a CSO backbone via amide bond formation, of which successful synthesis was confirmed by proton-nuclear magnetic resonance analyses. Doxorubicin (DOX)-loaded CI (CI10/DOX; CI:DOX=10:1 [w/w]) NPs with IDM increased in the presence of A549 cell lysates. In A549 cells (human lung carcinoma cells), more efficient cellular uptake of CI10/DOX NPs than that of free DOX was observed by using confocal laser scanning microscopy and flow cytometry. The in vitro cytotoxicity of CI10/DOX NPs in A549 cells was higher than those of free DOX and CI NPs with free DOX groups. In vivo pharmacokinetic studies after intravenous administration in rats showed significantly lower clearance of DOX from NPs compared with the free DOX group. Tumor targetability of the developed CI NPs was also verified by a real-time optical imaging study. In summary, the chemosensitizing CI/DOX NP with enhanced anticancer activity, prolonged blood circulation, and passive tumor targeting can be a promising anticancer drug delivery system for tumor-targeted therapy. Chemosensitizing nanoparticles (NPs) based on amphiphilic chitosan oligosaccharide-indomethacin (CSO-IDM; CI) conjugate were developed for tumor-targeted delivery of doxorubicin (DOX). IDM was introduced to the CSO backbone as a hydrophobic residue to synthesize an amphiphilic conjugate and a chemosenstizer of DOX for improving antitumor efficacies. IDM, conjugated to CSO, may inhibit the efflux of cellular uptaken DOX via multidrug resistance-associated protein (MRP) and subsequently augment the anti-proliferation potentials of DOX in A549 cells (MRP-expressed human lung cancer cells). Chemosensitizing properties of developed CI NPs were assessed in cell culture models and the tumor targetability of CI/DOX NPs was demonstrated in A549 tumor-xenografted mouse

  3. Implication from thyroid function decreasing during chemotherapy in breast cancer patients: chemosensitization role of triiodothyronine.

    Science.gov (United States)

    Huang, Jianbo; Jin, Liangbin; Ji, Guangyan; Xing, Lei; Xu, Chaobo; Xiong, Xiong; Li, Hongyuan; Wu, Kainan; Ren, Guosheng; Kong, Lingquan

    2013-07-06

    Thyroid hormones have been shown to regulate breast cancer cells growth, the absence or reduction of thyroid hormones in cells could provoke a proliferation arrest in G0-G1 or weak mitochondrial activity, which makes cells insensitive to therapies for cancers through transforming into low metabolism status. This biological phenomenon may help explain why treatment efficacy and prognosis vary among breast cancer patients having hypothyroid, hyperthyroid and normal function. Nevertheless, the abnormal thyroid function in breast cancer patients has been considered being mainly caused by thyroid diseases, few studied influence of chemotherapy on thyroid function and whether its alteration during chemotherapy can influence the respose to chemotherapy is still unclear. So, we aimed to find the alterations of thyroid function and non-thyroidal illness syndrome (NTIS) prevalence druing chemotherapy in breast cancer patients, and investigate the influence of thyroid hormones on chemotherapeutic efficacy. Thyroid hormones and NTIS prevalence at initial diagnosis and during chemotherapy were analyzed in 685 breast diseases patients (369 breast cancer, 316 breast benign lesions). The influence of thyroid hormones on chemotherapeutic efficacy was evaluated by chemosensitization test, to compare chemotherapeutic efficacy between breast cancer cells with chemotherapeutics plus triiodothyronine (T3) and chemotherapeutics only. In breast cancer, NTIS prevalence at the initial diagnosis was higher and increased during chemotherapy, but declined before the next chemotherapeutic course. Thyroid hormones decreased signigicantly during chemotherapy. T3 can enhance the chemosensitivity of MCF-7 to 5-Fu and taxol, with progression from G0-G1 phase to S phase. The similar chemosensitization role of T3 were found in MDA-MB-231. We compared chemotherapeutic efficacy among groups with different usage modes of T3, finding pretreatment with lower dose of T3, using higher dose of T3 together

  4. Chronic ethanol exposure induces SK-N-SH cell apoptosis by increasing N-methyl-D-aspartic acid receptor expression and intracellular calcium.

    Science.gov (United States)

    Wang, Hongbo; Wang, Xiaolong; Li, Yan; Yu, Hao; Wang, Changliang; Feng, Chunmei; Xu, Guohui; Chen, Jiajun; You, Jiabin; Wang, Pengfei; Wu, Xu; Zhao, Rui; Zhang, Guohua

    2018-04-01

    It has been identified that chronic ethanol exposure damages the nervous system, particularly neurons. There is scientific evidence suggesting that neuronal loss caused by chronic ethanol exposure has an association with neuron apoptosis and intracellular calcium oscillation is one of the primary inducers of apoptosis. Therefore, the present study aimed to investigate the inductive effects of intracellular calcium oscillation on apoptosis in SK-N-SH human neuroblastoma cells and the protective effects of the N-methyl-D-aspartic acid receptor (NMDAR) antagonist, memantine, on SK-N-SH cell apoptosis caused by chronic ethanol exposure. SK-N-SH cells were treated with 100 mM ethanol and memantine (4 µM) for 2 days. Protein expression of NR1 was downregulated by RNA interference (RNAi). Apoptosis was detected by Annexin V/propidium iodide (PI) double-staining and flow cytometry and cell viability was detected using an MTS kit. Fluorescence dual wavelength spectrophotometry was used to determine the intracellular calcium concentration and the levels of NR1 and caspase-3 were detected using western blotting. NR1 mRNA levels were also detected using qPCR. It was found that chronic ethanol exposure reduced neuronal cell viability and caused apoptosis of SK-N-SH cells, and the extent of damage in SK-N-SH cells was associated with ethanol exposure concentration and time. In addition, chronic ethanol exposure increased the concentration of intracellular calcium in SK-N-SH cells by inducing the expression of NMDAR, resulting in apoptosis, and memantine treatment reduced ethanol-induced cell apoptosis. The results of the present study indicate that the application of memantine may provide a novel strategy for the treatment of alcoholic dementia.

  5. Elevated pressure, a novel cancer therapeutic tool for sensitizing cisplatin-mediated apoptosis in A549

    International Nuclear Information System (INIS)

    Oh, Sangnam; Kim, Yanghee; Kim, Joonhee; Kwon, Daeho; Lee, Eunil

    2010-01-01

    Research highlights: → Sensitized apoptosis in cancer cells stimulated by EP precondition with p53 dependence. → EP attenuates several CDDP-resistance mechanisms. → No harmful effect of EP on normal fibroblasts. -- Abstract: Intensive cancer therapy strategies have thus far focused on sensitizing cancer cells to anticancer drug-mediated apoptosis to overcome drug resistance, and this strategy has led to more effective cancer therapeutics. Cisplatin (cis-diamminedichloroplatinum(II), CDDP) is an effective anticancer drug used to treat many types of cancer, including non-small cell lung carcinoma (NSCLC), and can be used in combination with various chemicals to enhance cancer cell apoptosis. Here, we introduce the use of elevated pressure (EP) in combination with CDDP for cancer treatment and explore the effects of EP on CDDP-mediated apoptosis in NSCLC cells. Our findings demonstrate that preconditioning NSCLC cells with EP sensitizes cells for CDDP-induced apoptosis. Enhanced apoptosis was dependent on p53 and HO-1 expression, and was associated with increased DNA damage and down-regulation of genes involved in nucleotide excision repair. The transcriptional levels of transporter proteins indicated that the mechanism by which EP-induced CDDP sensitization was intracellular drug accumulation. The protein levels of some antioxidants, such as hemeoxygenase-1 (HO-1), glutathione (GSH) and glutathione peroxidase (Gpx), were decreased in A549 cells exposed to EP via the down-regulation of the transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nrf-2). Furthermore, normal human fibroblasts were resistant to EP treatment, with no elevated DNA damage or apoptosis. Collectively, these data show that administration of EP is a potential adjuvant tool for CDDP-based chemosensitivity of lung cancer cells that may reduce drug resistance.

  6. Increased cell surface Fas expression is necessary and sufficient to sensitize lung fibroblasts to Fas ligation-induced apoptosis: implications for fibroblast accumulation in idiopathic pulmonary fibrosis.

    Science.gov (United States)

    Wynes, Murry W; Edelman, Benjamin L; Kostyk, Amanda G; Edwards, Michael G; Coldren, Christopher; Groshong, Steve D; Cosgrove, Gregory P; Redente, Elizabeth F; Bamberg, Alison; Brown, Kevin K; Reisdorph, Nichole; Keith, Rebecca C; Frankel, Stephen K; Riches, David W H

    2011-07-01

    Idiopathic pulmonary fibrosis (IPF) is associated with the accumulation of collagen-secreting fibroblasts and myofibroblasts in the lung parenchyma. Many mechanisms contribute to their accumulation, including resistance to apoptosis. In previous work, we showed that exposure to the proinflammatory cytokines TNF-α and IFN-γ reverses the resistance of lung fibroblasts to apoptosis. In this study, we investigate the underlying mechanisms. Based on an interrogation of the transcriptomes of unstimulated and TNF-α- and IFN-γ-stimulated primary lung fibroblasts and the lung fibroblast cell line MRC5, we show that among Fas-signaling pathway molecules, Fas expression was increased ∼6-fold in an NF-κB- and p38(mapk)-dependent fashion. Prevention of the increase in Fas expression using Fas small interfering RNAs blocked the ability of TNF-α and IFN-γ to sensitize fibroblasts to Fas ligation-induced apoptosis, whereas enforced adenovirus-mediated Fas overexpression was sufficient to overcome basal resistance to Fas-induced apoptosis. Examination of lung tissues from IPF patients revealed low to absent staining of Fas in fibroblastic cells of fibroblast foci. Collectively, these findings suggest that increased expression of Fas is necessary and sufficient to overcome the resistance of lung fibroblasts to Fas-induced apoptosis. Our findings also suggest that approaches aimed at increasing Fas expression by lung fibroblasts and myofibroblasts may be therapeutically relevant in IPF.

  7. Increased levels of circulating IL-16 and apoptosis markers are related to the activity of Whipple's disease.

    Directory of Open Access Journals (Sweden)

    Marie Benoit

    Full Text Available BACKGROUND: Whipple's disease (WD is an infectious disease caused by Tropheryma whipplei, which replicates in macrophages and induces the release of interleukin (IL-16, a substrate of caspase 3, and macrophage apoptosis. The disease is characterized by intestinal, cardiac or neurological manifestations; its diagnosis is based on invasive analysis requiring tissue biopsies or cerebrospinal fluid puncture. The disease progression is slow and often complicated by relapses despite empirical antibiotic treatment. METHODOLOGY/PRINCIPAL FINDINGS: We monitored circulating levels of IL-16 and nucleosomes in 36 French patients with WD; among them, some patients were enrolled in a longitudinal follow-up. As compared to control subjects, the circulating levels of both IL-16 and nucleosomes were increased in untreated patients with WD presenting as intestinal, cardiac or neurological manifestations. This finding was specific to WD since the circulating levels of IL-16 and nucleosomes were not increased in patients with unrelated inflammatory diseases such as inflammatory bowel disease or Q fever endocarditis. We also found that increased levels of IL-16 and nucleosomes were related to the activity of the disease. Indeed, successful antibiotic treatment decreased those levels down to those of control subjects. In contrast, patients who suffered from relapses exhibited circulating levels of IL-16 and nucleosomes as high as those of untreated patients. CONCLUSIONS/SIGNIFICANCE: Circulating levels of both IL-16 and nucleosomes were increased in WD. This finding provides simple and non-invasive tools for the diagnosis and the prognosis of WD.

  8. CytoregR inhibits growth and proliferation of human adenocarcinoma cells via induction of apoptosis

    Directory of Open Access Journals (Sweden)

    Hassanhi M

    2006-01-01

    Full Text Available Abstract Background Cancer is one of the devastating neovascular diseases that incapacitate so many people the world over. Recent reports from the National Cancer Institute indicate some significant gain therapy and cancer management as seen in the increase in the 5-year survival rate over the past two decades. Although near-perfect cure rate have been reported in the early-stage disease, these data reveal high recurrence rate and serious side effects including second malignancies and fatalities. Most of the currently used anticancer agents are only effective against proliferating cancer cells. Thus attention has been focused on potential anti-cancer agents capable of killing cancer cells independent of the cell cycle state, to ensure effective elimination of most cancer cells. The objective of this study was to test the chemosensitivity and potential mechanism of action of a novel cancer drug, CytoregR, in a panel of human cancer cells. Methods the study was performed using a series of bioassays including Trypan blue exclusion, MTS Growth inhibition, LDH-cytotoxicity, TUNEL-Terminal DNA fragmentation Apoptosis Assay, and the Caspase protease CPP32 activity assays. Results CytoregR induced significant dose- and time-dependent inhibition of growth in all the cells; with significant differences in chemosensitivity (P < 0.05 between the target cells becoming more apparent at 48 hr exposure. CytoregR showed no significant effect on normal cells relative to the tumor cells. Growth inhibition in all the cells was due to induction of apoptosis at lower concentrations of cytoregR (> 1:300. CytoregR-induced caspase protease-3 (CPP32 activation significantly and positively correlated with apoptosis induction and growth inhibition; thus implicating CPP32 as the principal death pathway in cytoregR-induced apoptosis. Conclusion CytoregR exerted a dose-and time-dependent growth inhibitory effect in all the target cells through induction of apoptosis via the

  9. Augmenting the activity of antifungal agents against aspergilli using structural analogues of benzoic acid as chemosensitizing agents.

    Science.gov (United States)

    Kim, Jong H; Campbell, Bruce C; Mahoney, Noreen; Chan, Kathleen L; Molyneux, Russell J; Balajee, Arunmozhi

    2010-10-01

    A number of benzoic acid analogues showed antifungal activity against strains of Aspergillus flavus, Aspergillus fumigatus and Aspergillus terreus, causative agents of human aspergillosis, in in vitro bioassays. Structure-activity analysis revealed that antifungal activities of benzoic and gallic acids were increased by addition of a methyl, methoxyl or chloro group at position 4 of the aromatic ring, or by esterification of the carboxylic acid with an alkyl group, respectively. Thymol, a natural phenolic compound, was a potent chemosensitizing agent when co-applied with the antifungal azole drugs fluconazole and ketoconazole. The thymol-azole drug combination demonstrated complete inhibition of fungal growth at dosages far lower than the drugs alone. Co-application of thymol with amphotericin B had an additive effect on all strains of aspergilli tested with the exception of two of three strains of A. terreus, where there was an antagonistic effect. Use of two mitogen-activated protein kinase (MAPK) mutants of A. fumigatus, sakAΔ and mpkCΔ, having gene deletions in the oxidative stress response pathway, indicated antifungal and/or chemosensitization activity of the benzo analogues was by disruption of the oxidative stress response system. Results showed that both these genes play overlapping roles in the MAPK system in this fungus. The potential of safe, natural compounds or analogues to serve as chemosensitizing agents to enhance efficacy of commercial antifungal agents is discussed. Published by Elsevier Ltd.

  10. Euglycemia in Diabetic Rats Leads to Reduced Liver Weight via Increased Autophagy and Apoptosis through Increased AMPK and Caspase-3 and Decreased mTOR Activities

    Directory of Open Access Journals (Sweden)

    Jun-Ho Lee

    2015-01-01

    Full Text Available Euglycemia is the ultimate goal in diabetes care to prevent complications. However, the benefits of euglycemia in type 2 diabetes are controversial because near-euglycemic subjects show higher mortality than moderately hyperglycemic subjects. We previously reported that euglycemic-diabetic rats on calorie-control lose a critical liver weight (LW compared with hyperglycemic rats. Here, we elucidated the molecular mechanisms underlying the loss of LW in euglycemic-diabetic rats and identified a potential risk in achieving euglycemia by calorie-control. Sprague-Dawley diabetic rats generated by subtotal-pancreatectomy were fed a calorie-controlled diet for 7 weeks to achieve euglycemia using 19 kcal% (19R or 6 kcal% (6R protein-containing chow or fed ad libitum (19AL. The diet in both R groups was isocaloric/kg body weight to the sham-operated group (19S. Compared with 19S and hyperglycemic 19AL, both euglycemic R groups showed lower LWs, increased autophagy, and increased AMPK and caspase-3 and decreased mTOR activities. Though degree of insulin deficiency was similar among the diabetic rats, Akt activity was lower, and PTEN activity was higher in both R groups than in 19AL whose signaling patterns were similar to 19S. In conclusion, euglycemia achieved by calorie-control is deleterious in insulin deficiency due to increased autophagy and apoptosis in the liver via AMPK and caspase-3 activation.

  11. Increased μ-Calpain Activity in Blasts of Common B-Precursor Childhood Acute Lymphoblastic Leukemia Correlates with Their Lower Susceptibility to Apoptosis.

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    Anna Mikosik

    Full Text Available Childhood acute lymphoblastic leukemia (ALL blasts are characterized by inhibited apoptosis promoting fast disease progress. It is known that in chronic lymphocytic and acute myeloid leukemias the reduced apoptosis is strongly related with the activity of calpain-calpastatin system (CCS composed of cytoplasmic proteases--calpains--performing the modulatory proteolysis of key proteins involved in cell proliferation and apoptosis, and of their endogenous inhibitor--calpastatin. Here, the CCS protein abundance and activity was for the first time studied in childhood ALL blasts and in control bone marrow CD19+ B cells by semi-quantitative flow cytometry and western blotting of calpastatin fragments resulting from endogenous calpain activity. Significantly higher μ-calpain (CAPN1 gene transcription, protein amounts and activity (but not those of m-calpain, with calpastatin amount and transcription of its gene (CAST greatly varying were observed in CD19(+ ALL blasts compared to control cells. Significant inverse relation between the amount/activity of calpain and spontaneous apoptosis was noted. Patients older than 10 years (considered at higher risk displayed increased amounts and activities of blast calpain. Finally, treatment of blasts with the tripeptide calpain inhibitors II and IV significantly and in dose-dependent fashion increased the percentage of blasts entering apoptosis. Together, these findings make the CCS a potential new predictive tool and therapeutic target in childhood ALL.

  12. Knockdown of peroxisome proliferator-activated receptor gamma coactivator-1 alpha increased apoptosis of human endometrial cancer HEC-1A cells

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    Yang H

    2016-08-01

    Full Text Available Hui Yang, Rui Yang, Hao Liu, Zhongqian Ren, Cuicui Wang, Da Li, Xiaoxin Ma Department of Obstetrics and Gynecology, Shengjing Hospital of China Medical University, Shenyang, Liaoning, People’s Republic of China Background: Peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α coactivates multiple transcription factors and regulates several metabolic processes. In this study, we focused on the roles of PGC-1α in the apoptosis of endometrial cancer HEC-1A cells. Materials and methods: PGC-1α expression in the HEC-1A cells was detected with real-time polymerase chain reaction and Western blot. Small interfering RNA directed against PGC-1α was designed and synthesized, and RNA interference technology was used to knock down PGC-1α mRNA and protein expression. Cell apoptosis, cell cycle, and mitochondrial membrane potential were then analyzed using flow cytometry. The expression of apoptotic proteins, Bcl-2 and Bax, was detected with Western blot. Results: The specific downregulation of PGC-1α expression in the HEC-1A cells increased their apoptosis through the mitochondrial apoptotic pathway by reducing the expression of Bcl-2 and increasing the expression of Bax. Conclusion: These results suggest that PGC-1α influences the apoptosis of HEC-1A cells and also provides a molecular basis for further investigation of the apoptotic mechanism in human endometrial cancer. Keywords: endometrial cancer, PGC-1α, apoptosis, Bcl-2, Bax

  13. Inhibition of autophagy initiation potentiates chemosensitivity in mesothelioma.

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    Follo, Carlo; Cheng, Yao; Richards, William G; Bueno, Raphael; Broaddus, Virginia Courtney

    2018-03-01

    The benefits of inhibiting autophagy in cancer are still controversial, with differences in outcome based on the type of tumor, the context and the particular stage of inhibition. Here, we investigated the impact of inhibiting autophagy at different stages on chemosensitivity using 3-dimensional (3D) models of mesothelioma, including ex vivo human tumor fragment spheroids. As shown by LC3B accumulation, we successfully inhibited autophagy using either an early stage ULK1/2 inhibitor (MRT 68921) or a late stage inhibitor (hydroxychloroquine). We found that inhibition of autophagy at the early stage, but not at late stage, potentiated chemosensitivity. This effect was seen only in those spheroids with high autophagy and active initiation at steady state. Inhibition of autophagy alone, at either early or late stage, did not cause cell death, showing that the inhibitors were non-toxic and that mesothelioma did not depend on autophagy at baseline, at least over 24 h. Using ATG13 puncta analysis, we found that autophagy initiation identified tumors that are more chemosensitive at baseline and after autophagy inhibition. Our results highlight a potential role of autophagy initiation in supporting mesothelioma cells during chemotherapy. Our work also highlights the importance of testing the inhibition of different stages in order to uncover the role of autophagy and the potential of its modulation in the treatment of cancer. © 2017 Wiley Periodicals, Inc.

  14. The flavonoid luteolin increases the resistance of normal, but not malignant keratinocytes, against UVB-induced apoptosis.

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    Verschooten, Lien; Smaers, Katrien; Van Kelst, Sofie; Proby, Charlotte; Maes, Daniel; Declercq, Lieve; Agostinis, Patrizia; Garmyn, Marjan

    2010-09-01

    Adequate protection of skin against the carcinogenic effects of UVB irradiation is essential. Flavonoids may have a conspicuous role in cancer prevention because of their antioxidant, anti-inflammatory, and growth-inhibitory effects. Therefore, we tested the effects of the flavone luteolin (LUT) on selected parameters of the sunburn response in normal human keratinocytes, exposed to physiological doses of UVB. LUT attenuated UVB-induced cell death through delay and inhibition of intrinsic apoptotic signaling. Moreover, LUT not only predominantly affected the mitochondrial apoptosis pathway through its antioxidant capacity, but also changed the balance of Bcl2 (B-cell leukemia/lymphoma 2)-family members. Furthermore, LUT had inhibitory effects on the UVB-induced release of the inflammatory mediators, IL-1alpha and prostaglandin-E(2). Using different cell lines derived from squamous cell carcinomas, we showed that LUT did not increase the resistance of malignant keratinocytes to UVB. Our data suggest that LUT inhibits different aspects of the sunburn response, which results ultimately in an increased survival of normal keratinocytes, whereas the sensitivity of malignant cells to UVB remain unchanged. Hence, LUT might have value in new photoprotective applications or improve existing ones.

  15. Cellular inhibitors of apoptosis (cIAP) 1 and 2 are increased in placenta from obese pregnant women.

    Science.gov (United States)

    Lappas, M

    2014-10-01

    Independent of their role in apoptosis, cellular inhibitors of apoptosis (cIAP) 1 and 2, have emerged as regulators of inflammation. Obesity in pregnancy is characterised by maternal and placental inflammation. Thus, the aim of this study was to determine the effect of maternal obesity and pro-inflammatory mediators on cIAP expression in human placenta. The expression of cIAP was assessed in human placenta from lean (n = 15) and obese (n = 14) patients by qRT-PCR and Western blotting. Primary trophoblast cells were used to determine the effect of pro-inflammatory cytokines on cIAP expression, and the effect of cIAP siRNA on pro-inflammatory cytokines. cIAP1 and cIAP2, gene and protein expression were significantly higher in placenta from women with pre-existing maternal obesity compared to placenta form lean women. Additionally, bacterial endotoxin LPS and the pro-inflammatory cytokines tumour necrosis factor (TNF)-α and interleukin (IL)-1β significantly increased the expression of both cIAP1 and cIAP2 in primary trophoblast cells isolated from human term placenta. Knockdown of cIAP1 or cIAP2 in human primary trophoblast cells significantly decreased TNF-α induced expression and secretion of pro-inflammatory cytokines IL-6 and IL-8 and of matrix metalloproteinase (MMP)-9. cIAP1 and cIAP2 expression is increased in placenta from women with pre-existing maternal obesity and in response to treatment with pro-inflammatory cytokines. Functional studies in placental trophoblast cells revealed that cIAPs are involved in TNF-α induced-expression of pro-inflammatory cytokines. Given the central role of pro-inflammatory cytokines in placental nutrient transport, this data suggest that cIAP1 and cIAP2 may play a role in fetal growth and development. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. Type 2 porcine reproductive and respiratory syndrome virus infection increases apoptosis at the maternal-fetal interface in late gestation pregnant gilts

    Science.gov (United States)

    Harding, John C. S.; Al-Dissi, Ahmad N; Detmer, Susan E.

    2017-01-01

    The pathogenesis of fetal death associated with porcine reproductive and respiratory syndrome (PRRS) is hypothesized to be a consequence of PRRS virus-induced apoptosis at the maternal-fetal interface (MFI). The objectives of this study were to evaluate distribution and degree of apoptosis in the uterine and fetal placental tissues during the experimental type 2 PRRS virus (PRRSV) infection and determine associations between apoptosis at the MFI, PRRSV RNA concentration and antigen staining intensity, PRRSV-induced microscopic lesions, and fetal preservation status. A total of 114 naïve, high-health pregnant gilts were inoculated with type 2 PRRSV on gestation day 85±1 with euthanasia 21 days later; 19 sham-inoculated gilts served as controls. Two hundred and fifty samples of uterine tissue with fetal placenta were selected based on negative, low PRRSV RNA, and high PRRSV RNA concentration (0, 2.7 log10 copies/mg, respectively). TUNEL assay was used to detect apoptosis in the endometrium and at the MFI. PRRSV RNA concentration and numbers of PRRSV immunopositive cells in uterine and placental tissue were positively associated with the severity of apoptosis in the endometrium and the MFI (P<0.001, P<0.05 and P<0.001, respectively). The number of TUNEL positive cells at the MFI was also positively associated with the severity (P<0.001) of vasculitis, but not total numbers of inflammatory cells in the endometrium. Increased numbers of TUNEL positive cells at the MFI were associated with PRRSV load in the fetal thymus, and greater odds of meconium staining of the fetus at 21 days post infection (P<0.001 for both). These findings suggest an important role of apoptosis in the pathogenesis of uterine epithelial and trophoblastic cell death at the MFI. Moreover, apoptosis at the MFI is significantly associated with fetal demise during in utero type 2 PRRSV infection. PMID:28253336

  17. Increased spontaneous apoptosis, but not survivin expression, is associated with histomorphologic response to neoadjuvant chemoradiation in rectal cancer.

    LENUS (Irish Health Repository)

    McDowell, Dermot T

    2009-11-01

    Survivin has been shown to be an important mediator of cellular radioresistance in vitro. This study aims to compare survivin expression and apoptosis to histomorphologic responses to neoadjuvant radiochemotherapy (RCT) in rectal cancer.

  18. Heterogeneity of tumor chemosensitivity in ovarian epithelial cancer revealed using the adenosine triphosphate-tumor chemosensitivity assay.

    Science.gov (United States)

    Zhang, Jin; Li, Hongxia

    2015-05-01

    Ovarian cancer has a poor prognosis, primarily due to the heterogeneity in chemosensitivity among patients. In the present study, this heterogeneity was evaluated in ovarian epithelial cancer (OEC) using an in vitro adenosine triphosphate tumor chemosensitivity assay (ATP-TCA). Specimens were collected from 80 patients who underwent cytoreductive surgery. Viable ovarian cancer cells obtained from malignant tissues were tested for sensitivity to paclitaxel (PTX), carboplatin (CBP), topotecan (TPT), gemcitabine (GEM), docetaxel (TXT), etoposide, bleomycin and 4-hydroperoxycyclophosphamide using ATP-TCA. The sensitivity, specificity, positive predictive value and negative predictive value for the clinical chemotherapy sensitivity of OEC were 88.6, 77.8, 83 and 84.8%, respectively. PTX demonstrated the highest sensitivity of all agents tested (82.5% in all specimens, 85.7% in recurrent specimens), followed by CBP (58.8 and 60.7%, respectively). The sensitivities to PTX and docetaxel (PIII) or low-differentiated specimens, respectively. The present study indicated that ATP-TCA is an effective method for guiding the choice of chemotherapy drugs. Notable heterogeneity of chemosensitivity was observed in the OEC specimens.

  19. RAGE Deletion Confers Renoprotection by Reducing Responsiveness to Transforming Growth Factor-β and Increasing Resistance to Apoptosis.

    Science.gov (United States)

    Hagiwara, Shinji; Sourris, Karly; Ziemann, Mark; Tieqiao, Wu; Mohan, Muthukumar; McClelland, Aaron D; Brennan, Eoin; Forbes, Josephine; Coughlan, Melinda; Harcourt, Brooke; Penfold, Sally; Wang, Bo; Higgins, Gavin; Pickering, Raelene; El-Osta, Assam; Thomas, Merlin C; Cooper, Mark E; Kantharidis, Phillip

    2018-02-15

    Signalling via the receptor of advanced glycation end-products (RAGE) although complex and not fully elucidated in the setting of diabetes, is considered a key injurious pathway in the development of diabetic nephropathy (DN). We report here that RAGE deletion resulted in increased expression of fibrotic (collagen I and IV, fibronectin) and the inflammatory marker, MCP-1 in primary mouse mesangial cells (MC) and in kidney cortex. RNA-seq analysis in MCs from RAGE -/- and wild type mice confirmed these observations. Nevertheless, despite these gene expression changes a decreased responsiveness to transforming growth factor-β was identified in RAGE -/- mice. Furthermore, RAGE deletion conferred a more proliferative phenotype in MCs and reduced susceptibility to staurosporine-induced apoptosis. RAGE restoration experiments in RAGE -/- MCs largely reversed these gene expression changes resulting in reduced expression of fibrotic and inflammatory markers. This study highlights that protection against DN in RAGE KO mice is likely in part to be due the result of decreased responsiveness to growth factor stimulation and an anti-apoptotic phenotype in mesangial cells. Furthermore, it extends our understanding of the role of RAGE in the progression of DN since RAGE appears to play a key role in modulating the sensitivity of the kidney to injurious stimuli such as prosclerotic cytokines. © 2018 by the American Diabetes Association.

  20. Thymoquinone inhibits the CXCL12-induced chemotaxis of multiple myeloma cells and increases their susceptibility to Fas-mediated apoptosis.

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    Gamal Badr

    Full Text Available In multiple myeloma (MM, malignant plasma cells reside in the bone marrow, where they accumulate in close contact with stromal cells. The mechanisms responsible for the chemotaxis of malignant plasma cells are still poorly understood. Thus, we investigated the mechanisms involved in the chemotaxis of MDN and XG2 MM cell lines. Both cell lines strongly expressed CCR9, CXCR3 and CXCR4 chemokine receptors but only migrated toward CXCL12. Activation of CXCR4 by CXCL12 resulted in the association of CXCR4 with CD45 and activation of PLCβ3, AKT, RhoA, IκBα and ERK1/2. Using siRNA-silencing techniques, we showed CD45/CXCR4 association is essential for CXCL12-induced migration of MM cells. Thymoquinone (TQ, the major active component of the medicinal herb Nigella sativa Linn, has been described as a chemopreventive and chemotherapeutic compound. TQ treatment strongly inhibited CXCL12-mediated chemotaxis in MM cell lines as well as primary cells isolated from MM patients, but not normal PBMCs. Moreover, TQ significantly down-regulated CXCR4 expression and CXCL12-mediated CXCR4/CD45 association in MM cells. Finally, TQ also induced the relocalization of cytoplasmic Fas/CD95 to the membrane of MM cells and increased CD95-mediated apoptosis by 80%. In conclusion, we demonstrate the potent anti-myeloma activity of TQ, providing a rationale for further clinical evaluation.

  1. DMS triggers apoptosis associated with the inhibition of SPHK1/NF-κB activation and increase in intracellular Ca2+ concentration in human cancer cells.

    Science.gov (United States)

    Chen, Kan; Pan, Qiuwei; Gao, Ying; Yang, Xinyan; Wang, Shibing; Peppelenbosch, Maikel P; Kong, Xiangdong

    2014-01-01

    N,N-Dimethyl-D-erythro-sphingosine (DMS) is known to induce cell apoptosis by specifically inhibiting sphingosine kinase 1 (SPHK1) and modulating the activity of cellular ceramide levels. The present study investigated the effects and the mechanism(s) of action of DMS in human lung cancer cells. We found that DMS dose-dependently suppressed cell proliferation and induced cell apoptosis in the human lung cancer cell line, A549. Mechanistically, treatment with DMS suppressed the activation of SPHK1 and nuclear factor-κB (NF-κB) p65, but increased intracellular [Ca2+]i in A549 cells. This study demonstrates that DMS triggers the apoptosis of human lung cancer cells through the modulation of SPHK1, NF-κB and calcium signaling. These molecules may represent targets for anticancer drug design.

  2. miR-1182 inhibits growth and mediates the chemosensitivity of bladder cancer by targeting hTERT

    International Nuclear Information System (INIS)

    Zhou, Jun; Dai, Wenbin; Song, Jianming

    2016-01-01

    microRNAs (miRNAs) have been demonstrated to contribute to tumor progression and metastasis and proposed to be key regulators of diverse biological processes. In this study, we report that miR-1182 is deregulated in bladder cancer tissues and cell lines. To characterize the role of miR-1182 in bladder cancer cells, we performed functional assays. The overexpression of miR-1182 significantly inhibits bladder cancer cell proliferation, colony formation, and invasion. Moreover, its up-regulation induced cell cycle arrest and apoptosis and mediated chemosensitivity to cisplatin in bladder cancer. Furthermore, a luciferase reporter assay and a rescue experiment indicated that miR-1182 directly targets hTERT by binding its 3′UTR. In conclusion, these results demonstrate that miR-1182 acts as a tumor suppressor and may be a potential biomarker for bladder cancer diagnosis and treatment.

  3. miR-1182 inhibits growth and mediates the chemosensitivity of bladder cancer by targeting hTERT

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Jun [Department of Urology, Huadong Hospital Affiliated to Fudan University, 221 Yan An Road(w), Shanghai 200040 (China); Dai, Wenbin, E-mail: daiwenbin271@163.com [Department of Urology, Huadong Hospital Affiliated to Fudan University, 221 Yan An Road(w), Shanghai 200040 (China); Song, Jianming [School of Medicine, Oregon Health & Science University, No.3181 S.W. Sam Jackson Park Road, Portland 97239-3098, OR (United States)

    2016-02-05

    microRNAs (miRNAs) have been demonstrated to contribute to tumor progression and metastasis and proposed to be key regulators of diverse biological processes. In this study, we report that miR-1182 is deregulated in bladder cancer tissues and cell lines. To characterize the role of miR-1182 in bladder cancer cells, we performed functional assays. The overexpression of miR-1182 significantly inhibits bladder cancer cell proliferation, colony formation, and invasion. Moreover, its up-regulation induced cell cycle arrest and apoptosis and mediated chemosensitivity to cisplatin in bladder cancer. Furthermore, a luciferase reporter assay and a rescue experiment indicated that miR-1182 directly targets hTERT by binding its 3′UTR. In conclusion, these results demonstrate that miR-1182 acts as a tumor suppressor and may be a potential biomarker for bladder cancer diagnosis and treatment.

  4. Depletion of the C. elegans NAC engages the unfolded protein response, resulting in increased chaperone expression and apoptosis.

    Directory of Open Access Journals (Sweden)

    Paul T Arsenovic

    Full Text Available The nascent polypeptide-associated complex (NAC is a highly conserved heterodimer important for metazoan development, but its molecular function is not well understood. Recent evidence suggests the NAC is a component of the cytosolic chaperone network that interacts with ribosomal complexes and their emerging nascent peptides, such that the loss of the NAC in chaperone-depleted cells results in an increase in misfolded protein stress. We tested whether the NAC functions similarly in Caeonorhabditis (C. elegans and found that its homologous NAC subunits, i.e. ICD-1 and -2, have chaperone-like characteristics. Loss of the NAC appears to induce misfolded protein stress in the ER triggering the unfolded protein response (UPR. Depletion of the NAC altered the response to heat stress, and led to an up-regulation of hsp-4, a homologue of the human chaperone and ER stress sensor GRP78/BiP. Worms lacking both ICD-1 and the UPR transcription factor XBP-1 generated a higher proportion of defective embryos, showed increased embryonic apoptosis and had a diminished survival rate relative to ICD-1-depleted animals with an intact UPR. Up-regulation of hsp-4 in NAC-depleted animals was specific to certain regions of the embryo; in embryos lacking ICD-1, the posterior region of the embryo showed strong up-regulation of hsp-4, while the anterior region did not. Furthermore, loss of ICD-1 produced prominent lysosomes in the gut region of adults and embryos putatively containing lipofuscins, lipid/protein aggregates associated with cellular aging. These results are the first set of evidence consistent with a role for C. elegans NAC in protein folding and localization during translation. Further, these findings confirm C. elegans as a valuable model for studying organismal and cell-type specific responses to misfolded protein stress.

  5. Snake venom causes apoptosis by increasing the reactive oxygen species in colorectal and breast cancer cell lines

    Directory of Open Access Journals (Sweden)

    Al-Asmari AK

    2016-10-01

    Full Text Available Abdulrahman Khazim Al-Asmari,1 Anvarbatcha Riyasdeen,1 Mohammad Hamed Al-Shahrani,2 Mozaffarul Islam1 1Research Center, 2Pediatric Hematology/Oncology and Bone Marrow Transplant Unit, Prince Sultan Military Medical City, Riyadh, Kingdom of Saudi Arabia Abstract: Snake venom possesses various kinds of proteins and neurotoxic polypeptides, which can negatively interfere with the neurotransmitter signaling cascade. This phenomenon occurs mainly due to the blocking of ion channels in the body system. Envenomation prevents or severely interrupts nerve impulses from being transmitted, inhibition of adenosine triphosphate synthesis, and proper functioning of the cardiac muscles. However, some beneficial properties of venoms have also been reported. The aim of this study was to examine the snake venom as an anticancer agent due to its inhibitory effects on cancer progression such as cell motility, cell invasion, and colony formation. In this study, the effect of venoms on phenotypic changes and the change on molecular level in colorectal and breast cancer cell lines were examined. A reduction of 60%–90% in cell motility, colony formation, and cell invasion was observed when these cell lines were treated with different concentrations of snake venom. In addition, the increase in oxidative stress that results in an increase in the number of apoptotic cancer cells was significantly higher in the venom-treated cell lines. Further analysis showed that there was a decrease in the expression of pro-inflammatory cytokines and signaling proteins, strongly suggesting a promising role for snake venom against breast and colorectal cancer cell progression. In conclusion, the snake venoms used in this study showed significant anticancer properties against colorectal and breast cancer cell lines. Keywords: colorectal cancer, breast cancer, cell motility, colony formation, oxidative stress, apoptosis, IL-8, IL-6, RhoC, p-Erk1/2

  6. Selective mitochondrial depletion, apoptosis resistance, and increased mitophagy in human Charcot-Marie-Tooth 2A motor neurons.

    Science.gov (United States)

    Rizzo, Federica; Ronchi, Dario; Salani, Sabrina; Nizzardo, Monica; Fortunato, Francesco; Bordoni, Andreina; Stuppia, Giulia; Del Bo, Roberto; Piga, Daniela; Fato, Romana; Bresolin, Nereo; Comi, Giacomo P; Corti, Stefania

    2016-10-01

    Charcot-Marie-Tooth 2A (CMT2A) is an inherited peripheral neuropathy caused by mutations in MFN2, which encodes a mitochondrial membrane protein involved in mitochondrial network homeostasis. Because MFN2 is expressed ubiquitously, the reason for selective motor neuron (MN) involvement in CMT2A is unclear. To address this question, we generated MNs from induced pluripotent stem cells (iPSCs) obtained from the patients with CMT2A as an in vitro disease model. CMT2A iPSC-derived MNs (CMT2A-MNs) exhibited a global reduction in mitochondrial content and altered mitochondrial positioning without significant differences in survival and axon elongation. RNA sequencing profiles and protein studies of key components of the apoptotic executioner program (i.e. p53, BAX, caspase 8, cleaved caspase 3, and the anti-apoptotic marker Bcl2) demonstrated that CMT2A-MNs are more resistant to apoptosis than wild-type MNs. Exploring the balance between mitochondrial biogenesis and the regulation of autophagy-lysosome transcription, we observed an increased autophagic flux in CMT2A-MNs that was associated with increased expression of PINK1, PARK2, BNIP3, and a splice variant of BECN1 that was recently demonstrated to be a trigger for mitochondrial autophagic removal. Taken together, these data suggest that the striking reduction in mitochondria in MNs expressing mutant MFN2 is not the result of impaired biogenesis, but more likely the consequence of enhanced mitophagy. Thus, these pathways represent possible novel molecular therapeutic targets for the development of an effective cure for this disease. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  7. Methylferulate from Tamarix aucheriana inhibits growth and enhances chemosensitivity of human colorectal cancer cells: possible mechanism of action.

    Science.gov (United States)

    Abaza, Mohamed Salah I; Afzal, Mohammad; Al-Attiyah, Raja'a J; Guleri, Radhika

    2016-10-01

    Natural products are valuable sources for anticancer agents. In the present study, methylferulate (MF) was identified for the first time from Tamarix aucheriana. Spectral data were used for identification of MF. The potential of MF to control cell growth, cell cycle, apoptosis, generation of reactive oxygen species (ROS), cancer cell invasion, nuclear factor kappa B (NFkB) DNA-binding activity and proteasomal activities, as well as the enhancement of chemosensitivity in human colorectal cancer cells, were evaluated. The possible molecular mechanism of MF's therapeutic efficacy was also assessed. Column chromatography and spectral data were used for isolation and identification of MF. MTT, immunofluorescence, flow cytometry, in vitro invasion, fluoremetry, EIA and Real time qPCR were used to measure antiproliferative, chemo-sensitizing effects and other biochemical parameters. MF showed a dose-dependent anti-proliferative effect on colorectal cancer cells (IC 50  = 1.73 - 1.9 mM) with a nonsignificant cytotoxicity toward normal human fibroblast. Colony formation inhibition (P ≤ 0.001, 0.0001) confirmed the growth inhibition by MF. MF arrested cell cycle progression in the S and G2/M phases; induced apoptosis and ROS generation; and inhibited NF-kB DNA-binding activity, proteasomal activities and cell invasion in colorectal cancer cells. MF up-regulated cyclin-dependent kinase inhibitors (p19 INK4D , p21 WAF1/CIP1 , p27 KIP1 ), pro-apoptotic gene expression (Bax, Bad, Apaf1, Bid, Bim, Smac) and caspases (caspase 2, 3, 6, 7, 8, 9). Moreover, MF down-regulated cyclin-dependent kinases (Cdk1, Cdk2) and anti-apoptotic gene expression (c-IAP-1, c-IAP-2, Bcl2,FLIP). In addition, MF differentially potentiated the sensitivity of colorectal cancer cells to standard chemotherapeutic drugs. MF showed a multifaceted anti-proliferative and chemosensitizing effects. These results suggest the chemotherapeutic and co-adjuvant potential of MF.

  8. The effects of benzene exposure on apoptosis in epithelial lung cells: localization by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) and the immunocytochemical localization of apoptosis-related gene products.

    Science.gov (United States)

    Weaver, C V; Liu, S-P; Lu, J-F; Lin, B-S

    2007-05-01

    Although benzene, a well-known human carcinogen, has been shown to induce apoptosis in vitro, no studies have been carried out to confirm and characterize its role in activating apoptosis in vivo. The present study investigated the effects of benzene inhalation on the epithelial cells lining the respiratory tract including bronchioles, terminal bronchioles, respiratory bronchioles and alveoli of male Sprague-Dawley rats. Inhalation of benzene 300 ppm for 7 days induced apoptotic changes in the parenchymal components in the lung that significantly exceeded the events of programmed cell death in normal control tissues. Apoptosis was confirmed by the electrophoretic analysis of internucleosomal DNA fragmentation of benzene-exposed lung tissues, which exhibited 180-200 bp laddering subunits indicative of genomic DNA degradation. Furthermore, semi-quantitative analysis of intracellular localization of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling TUNEL) showed a significant (p respiratory bronchiolar 60.8% segmental epithelial components as well as alveolar (55%) epithelia. Analysis of immunohistochemical expression of apoptosis-related gene products also supported the hypothesis that benzene can induce apoptosis in chemosensitive target cells in the lung parenchyma. Quantitative immunhistochemistry showed a statistically significant increase p respiratory system that demonstrates that benzene inhalation induces lung cell apoptosis as confirmed by DNA electrophoresis, in situ nick end labeling, and the upregulation of apoptosis-related gene products that facilitate caspase-cleaved enzymes which lead to cell degradation via programmed cell death. These responses may represent an important defense mechanism within the parenchymal cells of the respiratory system that reduce mutational hazard and the potential carcinogenic effects of benzene-initiated pathogenesis.

  9. Increase of autophagy and attenuation of apoptosis by Salvigenin promote survival of SH-SY5Y cells following treatment with H₂O₂.

    Science.gov (United States)

    Rafatian, Ghazaleh; Khodagholi, Fariba; Farimani, Mahdi Moridi; Abraki, Shahnaz Babaei; Gardaneh, Mossa

    2012-12-01

    Oxidative stress is a major component of harmful cascades activated in neurodegenerative disorders. Here, we tried to elucidate the possible neuroprotective effect of Salvigenin, a natural polyphenolic compound, on oxidative stress-induced apoptosis and autophagy in human neuroblastoma SH-SY5Y cells. We measured cell viability by MTT test and found that 25 μM is the best protective concentration of Salvigenin. GSH and SOD assays suggested that Salvigenin activates antioxidant factors. At the same time, measurement of ER stress-associated proteins including calpain and caspase-12 showed the ability of Salvigenin to decrease ER stress. We found that Salvigenin could decrease the apoptotic factors. Salvigenin inhibited H(2)O(2)-induced caspase-3 which is a hallmark of apoptosis in addition to reducing Bax\\Bcl-2 ratio by 1.45 fold. Additionally, Salvigenin increased the levels of autophagic factors. Our results showed an increase in LC3-II/LC3-I ratio, Atg7, and Atg12 in the presence of 25 μM of Salvigenin by about 1.28, 1.25, and 1.54 folds, respectively, compared to H(2)O(2)-treated cells. So it seems that H(2)O(2) cytotoxicity mainly results from apoptosis. Besides, Salvigenin helps cells to survive by inhibiting apoptosis and enhancing autophagy that opens a new horizon for the future experiments.

  10. Increased Susceptibility to Apoptosis and Growth Arrest of Human Breast Cancer Cells Treated by a Snake Venom-Loaded Silica Nanoparticles

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    Gamal Badr

    2014-11-01

    Full Text Available Background: The development of effective treatments against metastatic cancers, including breast cancer, is among the most important challenges in current experimental and clinical cancer research. We recently demonstrated that Walterinnesia aegyptia venom (WEV, either alone or in combination with silica nanoparticles (WEV+NP, resulted in the growth arrest and apoptosis of different cancer cell lines. Aims: In the present study, we evaluated the impact of WEV alone and WEV+NP on human breast cancer cells isolated from cancer biopsies. Methods: The potential effects of WEV alone and WEV+NP on the proliferation, induction of apoptosis and generation of free radicals in breast cancer cells isolated from 80 patients clinically diagnosed with breast cancer were evaluated by flow cytometry and ELISA. Results: WEV alone and WEV+NP inhibited the proliferation, altered the cell cycle and enhanced the induction of apoptosis of the breast cancer cells by increasing the activities of caspase-3, caspase-8 and caspase-9. In addition, the combination of WEV and NP robustly sensitized the breast cancer cells to growth arrest and apoptosis by increasing the generation of free radicals, including reactive oxygen species (ROS, hydroperoxide and nitric oxide. The combination of WEV with NP significantly enhanced the anti-tumor effect of WEV in breast cancer cells. Conclusion: Our data indicate the therapeutic potential of the nanoparticle-sustained delivery of snake venom for the treatment of breast cancer.

  11. Reduced proliferation and increased apoptosis of the SGC‑7901 gastric cancer cell line on exposure to GDC‑0449.

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    Wu, Chuanqing; Cheng, Ji; Hu, Shaobo; Deng, Rui; Muangu, Yamba Willy; Shi, Liang; Wu, Ke; Zhang, Peng; Chang, Weilong; Wang, Guobin; Tao, Kaixiong

    2016-02-01

    The sonic hedgehog (Shh) pathway is known to be vital in embryonic development and cancer propagation due to its irreplaceable role in cell proliferation and differentiation. GDC‑0449, a basal cell skin cancer target drug approved by the Food and Drugs Administration, is a smoothened (Smo)-specific antagonist. Although it has been clinically verified as a valid drug for the treatment of skin and pancreatic cancer, the application of GDC‑0449 in gastric cancer requires further investigation. In the present study, high-glucose Dulbecco's modified Eagle's medium with 10% fetal bovine serum was used for routine SGC‑7901 cell line culture. A Cell Counting Kit‑8 assay was employed for determination of the reproductive rate of the cells. Flow cytometry was performed to determine the apoptosis status of the SGC‑7901 cell line through Q4 analysis. Reverse transcription-quantitative polymerase chain reaction and Western blot analyses were used as target molecule detection vehicles. As expected, GDC‑0449 reduced the expression levels of Shh‑associated molecules, including Smo and gli1, compared with the blank group. The rate of cell proliferation was markedly limited and was accompanied by an increase in the apoptotic rate following GDC‑0449 exposure. In addition, further investigations confirmed B cell lymphoma‑2 (Bcl‑2) as the downstream molecular mechanism of GDC‑0449 efficacy. Of note, representatives of the cancer stem cell (CSC) surface marker, CD44 and CD133, demonstrated a similar trend to the Smo restriction observed. By repressing the expression of Bcl‑2, GDC‑0449 inhibited the normal proliferation of SGC‑7901 cells, and accelerated the apoptotic rate of the cells. It may also alter CSC properties due to the reduction in the expression of surface markers.

  12. Radotinib Induces Apoptosis of CD11b+ Cells Differentiated from Acute Myeloid Leukemia Cells.

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    Sook-Kyoung Heo

    Full Text Available Radotinib, developed as a BCR/ABL tyrosine kinase inhibitor (TKI, is approved for the second-line treatment of chronic myeloid leukemia (CML in South Korea. However, therapeutic effects of radotinib in acute myeloid leukemia (AML are unknown. In the present study, we demonstrate that radotinib significantly decreases the viability of AML cells in a dose-dependent manner. Kasumi-1 cells were more sensitive to radotinib than NB4, HL60, or THP-1 cell lines. Furthermore, radotinib induced CD11b expression in NB4, THP-1, and Kasumi-1 cells either in presence or absence of all trans-retinoic acid (ATRA. We found that radotinib promoted differentiation and induced CD11b expression in AML cells by downregulating LYN. However, CD11b expression induced by ATRA in HL60 cells was decreased by radotinib through upregulation of LYN. Furthermore, radotinib mainly induced apoptosis of CD11b+ cells in the total population of AML cells. Radotinib also increased apoptosis of CD11b+ HL60 cells when they were differentiated by ATRA/dasatinib treatment. We show that radotinib induced apoptosis via caspase-3 activation and the loss of mitochondrial membrane potential (ΔΨm in CD11b+ cells differentiated from AML cells. Our results suggest that radotinib may be used as a candidate drug in AML or a chemosensitizer for treatment of AML by other therapeutics.

  13. Regorafenib inhibits colorectal tumor growth through PUMA-mediated apoptosis

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    Chen, Dongshi; Wei, Liang; Yu, Jian; Zhang, Lin

    2014-01-01

    Purpose Regorafenib, a multi-kinase inhibitor targeting the Ras/Raf/MEK/ERK pathway, has recently been approved for the treatment of metastatic colorectal cancer (CRC). However, the mechanisms of action of regorafenib in CRC cells have been unclear. We investigated how regorafenib suppresses CRC cell growth and potentiates effects of other chemotherapeutic drugs. Experimental Design We determined whether and how regorafenib induces the expression of PUMA, a p53 target and a critical mediator of apoptosis in CRC cells. We also investigated whether PUMA is necessary for the killing and chemosensitization effects of regorafenib in CRC cells. Furthermore, xenograft tumors were used to test if PUMA mediates the in vivo antitumor, antiangiogenic and chemosensitization effects of regorafenib. Results We found that regorafenib treatment induces PUMA in CRC cells irrespective of p53 status through the NF-κB pathway following ERK inhibition and glycogen synthase kinase 3β (GSK3β) activation. Upregulation of PUMA is correlated with apoptosis induction in different CRC cell lines. PUMA is necessary for regorafenib-induced apoptosis in CRC cells. Chemosensitization by regorafenib is mediated by enhanced PUMA induction through different pathways. Furthermore, deficiency in PUMA abrogates the in vivo antitumor, antiangiogenic and chemosensitization effects of regorafenib. Conclusions Our results demonstrate a key role of PUMA in mediating the anticancer effects of regorafenib in CRC cells. They suggest that PUMA induction can be used as an indicator of regorafenib sensitivity, and also provide a rationale for manipulating the apoptotic machinery to improve the therapeutic efficacy of regorafenib and other targeted drugs. PMID:24763611

  14. Feasibility of chemosensitivity testing in soft tissue sarcomas

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    Steinstraesser Lars

    2005-04-01

    Full Text Available Abstract Background Soft tissue sarcomas comprise less than 1% of all solid malignancies. The presentation and behavior of these tumors differs depending on location and histological characteristics. Standard therapy consists of complete surgical resection in combination with adjuvant radiotherapy. The role of chemotherapy is not clearly defined and is largely restricted to clinical trials. Only a limited number of agents have proved to be effective in soft tissue sarcomas. The use of doxorubicin, epirubicin and ifosfamide allowed response rates of more than 20%. In addition, recent chemotherapy trials did not demonstrate any significant differences in efficacy for various histological subtypes. Methods The objective of this study was to gain additional information about the chemosensitivity of soft tissue sarcomas to seven 7 different chemotherapy agents as single drugs and 4 combinations. Therefore we used an established ATP based in-vitro testing system and examined 50 soft tissue sarcomas. Chemosensitivity was assessed using a luciferin-luciferase-based luminescence assay providing individual chemosensitivity indices for each agent tested. Results The sensitivity varied widely according to the histological subtypes. The tumors state of cellular dedifferentiation played a crucial role for the efficiency of the chemotherapeutic agents. The sensitivity also depended on the presentation of the sarcoma as a primary or recurrent tumor. The highest sensitivity was demonstrated for actinomycin D as a single agent, with 74% of the tumor samples exhibiting a high-grade sensitivity (20% low sensitivity, no resistance. The combination of actinomycin D and ifosfamide yielded a high sensitivity in 76% (2% resistance. Doxorubicin as a mono-therapy or in combination with ifosfamide achieved high sensitivity in 70% and 72%, respectively, and resistance in 6% of the samples. Conclusion Chemosensitivity testing is feasible in soft tissue sarcomas. It can be

  15. Cystogenesis in ARPKD results from increased apoptosis in collecting duct epithelial cells of Pkhd1 mutant kidneys

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    Hu, Bo; He, Xiusheng; Li, Ao; Qiu, Qingchao; Li, Cunxi; Liang, Dan; Zhao, Ping; Ma, Jie; Coffey, Robert J.; Zhan, Qimin; Wu, Guanqing

    2011-01-01

    Mutations in the PKHD1 gene result in autosomal recessive polycystic kidney disease (ARPKD) in humans. To determine the molecular mechanism of the cystogenesis in ARPKD, we recently generated a mouse model for ARPKD that carries a targeted mutation in the mouse orthologue of human PKHD1. The homozygous mutant mice display hepatorenal cysts whose phenotypes are similar to those of human ARPKD patients. By littermates of this mouse, we developed two immortalized renal collecting duct cell lines with Pkhd1 and two without. Under nonpermissive culture conditions, the Pkhd1 -/- renal cells displayed aberrant cell-cell contacts and tubulomorphogenesis. The Pkhd1 -/- cells also showed significantly reduced cell proliferation and elevated apoptosis. To validate this finding in vivo, we examined proliferation and apoptosis in the kidneys of Pkhd1 -/- mice and their wildtype littermates. Using proliferation (PCNA and Histone-3) and apoptosis (TUNEL and caspase-3) markers, similar results were obtained in the Pkhd1 -/- kidney tissues as in the cells. To identify the molecular basis of these findings, we analyzed the effect of Pkhd1 loss on multiple putative signaling regulators. We demonstrated that the loss of Pkhd1 disrupts multiple major phosphorylations of focal adhesion kinase (FAK), and these disruptions either inhibit the Ras/C-Raf pathways to suppress MEK/ERK activity and ultimately reduce cell proliferation, or suppress PDK1/AKT to upregulate Bax/caspase-9/caspase-3 and promote apoptosis. Our findings indicate that apoptosis may be a major player in the cyst formation in ARPKD, which may lead to new therapeutic strategies for human ARPKD.

  16. Inhibition of phospholipaseD2 increases hypoxia-induced human colon cancer cell apoptosis through inactivating of the PI3K/AKT signaling pathway.

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    Liu, Maoxi; Fu, Zhongxue; Wu, Xingye; Du, Kunli; Zhang, Shouru; Zeng, Li

    2016-05-01

    Hypoxia is a common feature of solid tumor, and is a direct stress that triggers apoptosis in many human cell types. As one of solid cancer, hypoxia exists in the whole course of colon cancer occurrence and progression. Our previous studies shown that hypoxia induce high expression of phospholipase D2 (PLD2) and survivin in colon cancer cells. However, the correlation between PLD2 and survivin in hypoxic colon cancer cells remains unknown. In this study, we observed significantly elevated PLD2 and survivin expression levels in colon cancer tissues and cells. This is a positive correlation between of them, and co-expression of PLD2 and survivin has a positive correlation with the clinicpatholic features including tumor size, TNM stage, and lymph node metastasis. We also found that hypoxia induced the activity of PLD increased significant mainly caused by PLD2 in colon cancer cells. However, inhibition the activity of PLD2 induced by hypoxia promotes the apoptosis of human colon cancer cells, as well as decreased the expression of apoptosis markers including survivin and bcl2. Moreover, the pharmacological inhibition of PI3K/AKT supported the hypothesis that promotes the apoptosis of hypoxic colon cancer cells by PLD2 activity inhibition may through inactivation of the PI3K/AKT signaling pathway. Furthermore, interference the PLD2 gene expression leaded to the apoptosis of hypoxic colon cancer cells increased and also decreased the expression level of survivin and bcl2 may through inactivation of PI3K/AKT signaling pathway. These results indicated that PLD2 play antiapoptotic role in colon cancer under hypoxic conditions, inhibition of the activity, or interference of PLD2 gene expression will benefit for the treatment of colon cancer patients.

  17. Ginsenoside Rh2 and Rg3 inhibit cell proliferation and induce apoptosis by increasing mitochondrial reactive oxygen species in human leukemia Jurkat cells.

    Science.gov (United States)

    Xia, Ting; Wang, Ying-Nan; Zhou, Chuan-Xin; Wu, Li-Mei; Liu, Yong; Zeng, Qian-Hong; Zhang, Xiang-Long; Yao, Jia-Hui; Wang, Min; Fang, Jian-Pei

    2017-06-01

    Ginsenoside Rh2 (GRh2) and ginsenoside Rg3 (GRg3) are primary bioactive components in Panax ginseng. The present study aimed to investigate the underlying mechanisms of apoptotic cell‑death induced by GRh2 and GRg3 in human leukemia Jurkat cells. The Cell Counting kit‑8 assay was used to determine cell proliferation. Apoptosis was detected by nuclear morphologic observation by Hoechst 33342 staining and Annexin V-allophycocyanin and 7-amino-actinomycin D assay. mitoTEMPO, a mitochondrial reactive oxygen species (ROS) scavenger, was used to examine the effects of mitochondrial ROS on cell viability and mitochondrial membrane potential (MMP). Finally, the expression levels of numerous mitochondrial‑associated apoptosis proteins were assessed by western blot analysis. These results demonstrated that GRh2 and GRg3 inhibited cell growth and induced apoptosis, and that GRh2 had greater cytotoxicity than GRg3. GRh2 induced generation of more mitochondrial ROS compared with GRg3 in Jurkat cells; however, this effect was ameliorated by subsequent treatment with mitoTEMPO. Furthermore, excess mitochondrial ROS induced by GRh2 was more potent than GRg3 in inhibiting cell proliferation and reducing MMP. In addition, expression levels of apoptosis‑associated proteins were significantly increased in Jurkat cells treated with GRh2 than GRg3. In conclusion, these findings suggested that GRh2 and GRg3 induce mitochondrial-associated apoptosis by increasing mitochondrial ROS in human leukemia Jurkat cells. GRh2 may more effectively inhibit cell growth and accelerate apoptosis than GRg3. This study provides a potential novel strategy for the treatment of acute lymphoblastic leukemia.

  18. Scorpion Venom Causes Apoptosis by Increasing Reactive Oxygen Species and Cell Cycle Arrest in MDA-MB-231 and HCT-8 Cancer Cell Lines.

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    Al-Asmari, Abdulrahman Khazim; Riyasdeen, Anvarbatcha; Islam, Mozaffarul

    2018-01-01

    The objective of this study was to examine the effect of scorpion venoms on cancer cell progression, apoptosis, and cell cycle arrest. Scorpion venoms are known to possess numerous bioactive compounds that act against cancer progression by inducing apoptosis. In this study, we have taken the venoms from the following 2 species of scorpion- Androctonus crassicauda and Leiurus quinquestriatus-and tested the anticancer properties of the venom against breast and colorectal cancer cell lines. Milking of scorpion venom and culturing the breast and colorectal cancer cell lines were done according to the standard procedure. The venom cytotoxicity was assessed by MTT methods, and the cellular and nuclear changes were studied with phase contrast and propidium iodide staining, respectively. The cell cycle arrest and accumulation of reactive oxygen species were analyzed on a Muse cell analyzer. The venoms exerted cytotoxic effects on breast and colorectal cell lines in a dose- and time-dependent manner. Enhanced apoptotic cells, increase in reactive oxygen species, and cell cycle arrest were observed after challenging these cell lines with scorpion venoms. Scorpion venom induces apoptosis in breast and colorectal cell lines as reflected by the changes in the cell morphology and cell cycle studies. Furthermore, a high percentage of total reactive oxygen species as well as apoptotic cells also contribute to cell death as observed after venom treatments. To the best of authors' knowledge, this is the first scientific evidence demonstrating the induction of apoptosis and cell cycle arrest by these species of scorpion venoms.

  19. Radiation-induced apoptosis in human myeloma cell line increases BCL-2/BAX dimer formation and does not result in BAX/BAX homodimerization.

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    Filippovich, I V; Sorokina, N I; Lisbona, A; Chérel, M; Chatal, J F

    2001-06-01

    A popular model of BCL-2 and BAX involvement in apoptosis suggests that upon apoptosis induction cytosolic BAX translocates to the mitochondria, where it displays the pro-apoptotic function, which involves its homodimerization. BCL-2 exerts anti-apoptotic function by forming heterodimers with BAX, thus neutralizing the pro-apoptotic activity of the latter. We have shown that irradiation of the human myeloma cell line RPMI-8226 induced apoptosis as determined by DNA degradation, cytochrome c release into cytoplasm and BCL-2 caspase-mediated cleavage. BCL-2 protein was present only in the membrane fraction, whereas BAX was found both in cytosol and membranes isolated from non-irradiated cells. Radiation induced moderate redistribution of BAX from cytosol to membranes with a concomitant increase in BCL-2/BAX heterodimer formation. Rapid and transient BCL-2 phosphorylation in membrane fractions of irradiated cells did not affect BCL-2/BAX heterodimerization. We failed to detect any BAX/BAX homodimers in apoptotic cells. Our findings show that in irradiated RPMI-8226 cells the formation of BCL-2/BAX heterodimers correlates with apoptosis. We conclude that BCL-2/BAX heterodimers are negative regulators of death protection, and our data agree with those who propose that BCL-2 does not require BAX to exert its survival function. Copyright 2001 Wiley-Liss, Inc.

  20. Impaired nuclear factor erythroid 2-related factor 2 expression increases apoptosis of airway epithelial cells in patients with chronic obstructive pulmonary disease due to cigarette smoking.

    Science.gov (United States)

    Yamada, Kazuhiro; Asai, Kazuhisa; Nagayasu, Fumihiro; Sato, Kanako; Ijiri, Naoki; Yoshii, Naoko; Imahashi, Yumiko; Watanabe, Tetsuya; Tochino, Yoshihiro; Kanazawa, Hiroshi; Hirata, Kazuto

    2016-02-09

    Cigarette smoking-induced oxidative stress is known to be a key mechanism in COPD pathogenesis. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a central transcription factor that regulates the antioxidant defense system. The aim of this study was to compare Nrf2 expression in COPD subjects and control subjects, and to determine the role of Nrf2 in protecting against oxidative stress-induced apoptosis. We enrolled 8 COPD subjects and 7 control subjects in this study. We performed bronchial brushing by bronchoscopy and obtained bronchial epithelial cells from the airways. Nrf2 expression in bronchial epithelial cells was evaluated by real-time PCR and Western blotting. We examined the effect of 10 or 15 % cigarette smoke extract (CSE) induced A549 cells apoptosis using a time-lapse cell imaging assay with caspase-3/7 activation detecting reagent and performed Terminal deoxynucleotidyltransferase-mediated dUTP nick end labelling assay for confirming A549 cells apoptosis. We also examined the effects of Nrf2 knockdown and, 0.1, 0.5, and 1.0 mM N-acetyl cysteine on CSE-induced apoptosis. Statistical analyses were performed using t-test, paired t-test or an analysis of variance followed by the Tukey-Kramer method. Nrf2 mRNA expression in COPD subjects was significantly lower than that in control subjects and Nrf2 mRNA were negatively correlated with pack year. Nrf2 protein in COPD subjects was significantly lower than that in control subjects. CSE-induced A549 cells apoptosis was increased in a time-, concentration-dependent manner, and was significantly increased by Nrf2 knockdown. N-acetyl cysteine significantly ameliorated CSE-induced apoptosis. Nrf2 expression was lower in COPD patients than in control subjects. Nrf2 might have a protective role against apoptosis caused by CSE-induced oxidative stress. These results suggest an involvement of Nrf2 in COPD and administration of antioxidants to patients with COPD might be a basic therapeutic option.

  1. Impaired chemosensitivity of mouse dorsal raphe serotonergic neurons overexpressing serotonin 1A (Htr1a receptors.

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    Gilda Baccini

    Full Text Available BACKGROUND: Serotonergic system participates in a wide range of physiological processes and behaviors, but its role is generally considered as modulatory and noncrucial, especially concerning life-sustaining functions. We recently created a transgenic mouse line in which a functional deficit in serotonin homeostasis due to excessive serotonin autoinhibition was produced by inducing serotonin 1A receptor (Htr1a overexpression selectively in serotonergic neurons (Htr1a raphe-overexpressing or Htr1a(RO mice. Htr1a(RO mice exhibit episodes of autonomic dysregulation, cardiovascular crises and death, resembling those of sudden infant death syndrome (SIDS and revealing a life-supporting role of serotonergic system in autonomic control. Since midbrain serotonergic neurons are chemosensitive and are implicated in arousal we hypothesized that their chemosensitivity might be impaired in Htr1a(RO mice. PRINCIPAL FINDINGS: Loose-seal cell-attached recordings in brainstem slices revealed that serotonergic neurons in dorsal raphe nucleus of Htr1a(RO mice have dramatically reduced responses to hypercapnic challenge as compared with control littermates. In control mice, application of 9% CO(2 produced an increase in firing rate of serotonergic neurons (0.260 ± 0.041 Hz, n=20, p=0.0001 and application of 3% CO(2 decreased their firing rate (-0.142 ± 0.025 Hz, n=17, p=0.0008. In contrast, in Htr1a(RO mice, firing rate of serotonergic neurons was not significantly changed by 9% CO(2 (0.021 ± 0.034 Hz, n=16, p=0.49 and by 3% CO(2 (0.012 ± 0.046 Hz, n=12, p=0.97. CONCLUSIONS: Our findings support the hypothesis that chemosensitivity of midbrain serotonergic neurons provides a physiological mechanism for arousal responses to life-threatening episodes of hypercapnia and that functional impairment, such as excessive autoinhibition, of midbrain serotonergic neuron responses to hypercapnia may contribute to sudden death.

  2. Deoxyribonucleoside triphosphate pools and thymidine chemosensitization in human T-cell leukemia.

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    Cohen, J D; Robins, H I; Katz, T B; Miller, E M; Kuzminsky, S R; Javid, M J

    1993-02-01

    Thymidine kills cells by depleting dCTP stores. The present experiments tested whether deoxycytidine, by replenishing dCTP pools, could prevent thymidine cytotoxicity and thymidine's enhancement of carboplatin killing in two human T-cell acute leukemia cell lines. MOLT3 and JM cells were exposed to combinations of thymidine, deoxycytidine, and carboplatin and then assessed for survival, the magnitude of thymidine-carboplatin chemosensitization, and changes in deoxyribonucleoside triphosphate pools. For both cell lines, deoxycytidine (up to 144.5 micrograms/ml x 24 h) completely restored dCTP pools but only partially protected against thymidine cytotoxicity (100-1000 micrograms/ml x 24 h) and thymidine-carboplatin sensitization (up to 60 micrograms carboplatin/ml during the last hour of thymidine). This contrasts with complete protection in prior studies using other cell types. Thymidine alone markedly increased dTTP and dGTP pools and decreased dCTP; dATP pools underwent a sharp decline which has not been observed before in any cell line. In subsequent studies 0.0336-137.3 micrograms deoxyadenosine/ml partially prevented cytotoxicity and carboplatin sensitization by 300 micrograms thymidine/ml. Together, deoxycytidine and deoxyadenosine completely prevented thymidine-carboplatin sensitization even though dATP and dCTP pools were not entirely returned to normal. These findings are discussed in regard to the unusual sensitivity of T-cell malignancies to thymidine toxicity, mechanisms of cytotoxicity and chemosensitization by thymidine, and the possibility of thymidine selectively sensitizing T-cell malignancies to killing by alkylating agents.

  3. Multiparametric Profiling for Identification of Chemosensitizers against Gram-Negative Bacteria

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    Vincent Lôme

    2018-02-01

    Full Text Available Antibiotic resistance is now a worldwide therapeutic problem. Since the beginning of anti-infectious treatment bacteria have rapidly shown an incredible ability to develop and transfer resistance mechanisms. In the last decades, the design variation of pioneer bioactive molecules has strongly improved their activity and the pharmaceutical companies partly won the race against the clock. Since the 1980s, the new classes of antibiotics that emerged were mainly directed to Gram-positive bacteria. Thus, we are now facing to multidrug-resistant Gram-negative bacteria, with no therapeutic options to deal with them. These bacteria are mainly resistant because of their double membrane that conjointly impairs antibiotic accumulation and extrudes these molecules when entered. The main challenge is to allow antibiotics to cross the impermeable envelope and reach their targets. One promising solution would be to associate, in a combination therapy, a usual antibiotic with a non-antibiotic chemosensitizer. Nevertheless, for effective drug discovery, there is a prominent lack of tools required to understand the rules of permeation and accumulation into Gram-negative bacteria. By the use of a multidrug-resistant enterobacteria, we introduce a high-content screening procedure for chemosensitizers discovery by quantitative assessment of drug accumulation, alteration of barriers, and deduction of their activity profile. We assembled and analyzed a control chemicals library to perform the proof of concept. The analysis was based on real-time monitoring of the efflux alteration and measure of the influx increase in the presence of studied compounds in an automatized bio-assay. Then, synergistic activity of compounds with an antibiotic was studied and kinetic data reduction was performed which led to the calculation of a score for each barrier to be altered.

  4. Targeting Survivin Enhances Chemosensitivity in Retinoblastoma Cells and Orthotopic Tumors.

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    Angela Ferrario

    Full Text Available Treatments for retinoblastoma (Rb vary depending on the size and location of the intraocular lesions and include chemotherapy and radiation therapy. We examined whether agents used to treat Rb induce a pro-survival phenotype associated with increased expression of survivin, a member of the inhibitor of apoptosis family of proteins. We document that exposure to carboplatin, topotecan or radiation resulted in elevated expression of survivin in two human Rb cell lines but not in normal retinal pigmented epithelial (RPE cells. Cellular levels of survivin were attenuated in Rb cells exposed to an imidazolium-based survivin suppressant, Sepantronium bromide (YM155. Protein expression patterns of survivin in RPE cells were not altered following treatment protocols involving exposure to YM155. Including YM155 with chemotherapy or radiation increased levels of apoptosis in Rb cells but not in RPE cells. Intraocular luciferase expressing Rb tumors were generated from the Rb cell lines and used to evaluate the effects of carboplatin and YM155 on in-vivo survivin expression and tumor growth. Carboplatin induced expression of survivin while carboplatin combined with YM155 reduced survivin expression in tumor bearing eyes. The combination protocol was also most effective in reducing the rate of tumor regrowth. These results indicate that targeted inhibition of the anti-apoptotic protein survivin provides a therapeutic advantage for Rb cells and tumors treated with chemotherapy.

  5. Camalexin induces apoptosis in T-leukemia Jurkat cells by increased concentration of reactive oxygen species and activation of caspase-8 and caspase-9.

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    Mezencev, Roman; Updegrove, Taylor; Kutschy, Peter; Repovská, Mária; McDonald, John F

    2011-07-01

    Camalexin, a major indole phytoalexin of Arabidopsis thaliana, accumulates in various cruciferous plants in response to environmental stress and reportedly displays antimicrobial activities against various plant pathogens. However, its cytotoxicity against eukaryotic cells and potential as a prospective drug for human diseases has been examined only in a limited context. Our data demonstrate the time- and concentration-dependent cytotoxicity of camalexin on human T-leukemia Jurkat cells in the micromolar range, and the lower potency of cytotoxic effects on human lymphoblasts and primary fibroblasts. Cytotoxicity of camalexin is enhanced by the glutathione-depleting agent buthionine sulfoximine and completely blocked by pan-caspase inhibitor Z-VAD-FMK. Treatment of Jurkat cells with camalexin resulted in activation of caspase-8, caspase-9, caspases-3/7, and apoptosis that was detected by the presence of a sub-G1 population of cells, externalization of phosphatidyl serine and decreased mitochondrial membrane potential. Staining with 2',7'-dichlorodihydrofluorescein diacetate and dihydroethidium bromide displayed increased concentration of reactive oxygen species (ROS) early in camalexin-treated Jurkat cells, prior to the onset of apoptosis, while staining with MitoSOX(™) dye identified mitochondria as a source of increased ROS. Our data suggest that this phytochemical, which has a wide range of predicted pharmacological activities, induces apoptosis in Jurkat leukemia cells through increased ROS followed by dissipation of mitochondrial membrane potential and execution of caspase-9- and caspase-8-initiated apoptosis. This is, to the best of our knowledge, the first report on antileukemic activity and mode of action of this unique indole phytoalexin.

  6. Kaempferol increases apoptosis in human cervical cancer HeLa cells via PI3K/AKT and telomerase pathways.

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    Kashafi, Elham; Moradzadeh, Maliheh; Mohamadkhani, Ashraf; Erfanian, Saiedeh

    2017-05-01

    Cervical cancer is one of the most frequent cancers in women worldwide. Defects in the apoptotic pathways are responsible for both the disease pathogenesis and its therapy resistance. It is thus a good candidate for treatment by pro-apoptotic agents. Kaempferol as a flavonoid has antioxidant and anti-tumor properties. Kaempferol has been shown to induce apoptosis and cell death in cancer cells. However, due to the problems in the treatment of cervical cancer, this study is designed to investigate the molecular mechanism by which kaempferol suppresses the growth of cervical cancer HeLa cell as compared with HFF cells (normal cells). Cells treated with kaempferol (12-100μM) and 5-FU (1-10μM), as the positive control, up to 72h. Cell viability was determined by MTT assay and real time PCR was used to investigate apoptosis and telomerase genes expression. The results showed that kaempferol decreased cell viability as concentration- and time-dependently. IC 50 values were 10.48μM for HeLa and 707.00μM for HFF cells, as compared with 1.40μM and 16.38μM for 5-FU after 72h treatment, respectively. Also, kaempferol induced cellular apoptosis and aging through down-regulating the PI3K/AKT and hTERT pathways. This study suggests that kaempferol may be a useful adjuvant therapeutic agent in the treatment of cervical cancer. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  7. Increasing the endogenous NO level causes catalase inactivation and reactivation of intercellular apoptosis signaling specifically in tumor cells.

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    Bauer, Georg

    2015-12-01

    Tumor cells generate extracellular superoxide anions and are protected against intercellular apoptosis-inducing HOCl- and NO/peroxynitrite signaling through the expression of membrane-associated catalase. This enzyme decomposes H2O2 and thus prevents HOCl synthesis. It efficiently interferes with NO/peroxynitrite signaling through oxidation of NO and decomposition of peroxynitrite. The regulatory potential of catalase at the crosspoint of ROS and RNS chemical biology, as well as its high local concentration on the outside of the cell membrane of tumor cells, establish tight control of intercellular signaling and thus prevent tumor cell apoptosis. Therefore, inhibition of catalase or its inactivation by singlet oxygen reactivate intercellular apoptosis-inducing signaling. Nitric oxide and peroxynitrite are connected with catalase in multiple and meaningful ways, as (i) NO can be oxidated by compound I of catalase, (ii) NO can reversibly inhibit catalase, (iii) peroxynitrite can be decomposed by catalase and (iv) the interaction between peroxynitrite and H2O2 leads to the generation of singlet oxygen that inactivates catalase. Therefore, modulation of the concentration of free NO through addition of arginine, inhibition of arginase, induction of NOS expression or inhibition of NO dioxygenase triggers an autoamplificatory biochemical cascade that is based on initial formation of singlet oxygen, amplification of superoxide anion/H2O2 and NO generation through singlet oxygen dependent stimulation of the FAS receptor and caspase-8. Finally, singlet oxygen is generated at sufficiently high concentration to inactivate protective catalase and to reactivate intercellular apoptosis-inducing ROS signaling. This regulatory network allows to establish several pathways for synergistic interactions, like the combination of modulators of NO metabolism with enhancers of superoxide anion generation, modulators of NO metabolism that act at different targets and between modulators of

  8. Increasing the endogenous NO level causes catalase inactivation and reactivation of intercellular apoptosis signaling specifically in tumor cells

    Science.gov (United States)

    Bauer, Georg

    2015-01-01

    Tumor cells generate extracellular superoxide anions and are protected against intercellular apoptosis-inducing HOCl- and NO/peroxynitrite signaling through the expression of membrane-associated catalase. This enzyme decomposes H2O2 and thus prevents HOCl synthesis. It efficiently interferes with NO/peroxynitrite signaling through oxidation of NO and decomposition of peroxynitrite. The regulatory potential of catalase at the crosspoint of ROS and RNS chemical biology, as well as its high local concentration on the outside of the cell membrane of tumor cells, establish tight control of intercellular signaling and thus prevent tumor cell apoptosis. Therefore, inhibition of catalase or its inactivation by singlet oxygen reactivate intercellular apoptosis-inducing signaling. Nitric oxide and peroxynitrite are connected with catalase in multiple and meaningful ways, as (i) NO can be oxidated by compound I of catalase, (ii) NO can reversibly inhibit catalase, (iii) peroxynitrite can be decomposed by catalase and (iv) the interaction between peroxynitrite and H2O2 leads to the generation of singlet oxygen that inactivates catalase. Therefore, modulation of the concentration of free NO through addition of arginine, inhibition of arginase, induction of NOS expression or inhibition of NO dioxygenase triggers an autoamplificatory biochemical cascade that is based on initial formation of singlet oxygen, amplification of superoxide anion/H2O2 and NO generation through singlet oxygen dependent stimulation of the FAS receptor and caspase-8. Finally, singlet oxygen is generated at sufficiently high concentration to inactivate protective catalase and to reactivate intercellular apoptosis-inducing ROS signaling. This regulatory network allows to establish several pathways for synergistic interactions, like the combination of modulators of NO metabolism with enhancers of superoxide anion generation, modulators of NO metabolism that act at different targets and between modulators of

  9. Functional link between the hypocretin and serotonin systems in the neural control of breathing and central chemosensitivity.

    Science.gov (United States)

    Corcoran, Andrea E; Richerson, George B; Harris, Michael B

    2015-07-01

    Serotonin (5-HT)-synthesizing neurons of the medullary raphe are putative central chemoreceptors, proposed to be one of potentially multiple brain stem chemosensitive cell types and loci interacting to produce the respiratory chemoreflex. Hypocretin-synthesizing neurons of the lateral hypothalamus are important contributors to arousal state, thermoregulation, and feeding behavior and are also reportedly involved in the hypercapnic ventilatory response. Recently, a functional interaction was found between the hypocretin system and 5-HT neurons of the dorsal raphe. The validity and potential significance of hypocretin modulation of medullary raphe 5-HT neurons, however, is unknown. As such, the purpose of this study was to explore functional interactions between the hypocretin system and 5-HT system of the medullary raphe on baseline respiratory output and central chemosensitivity. To explore such interactions, we used the neonatal in vitro medullary slice preparation derived from wild-type (WT) mice (normal 5-HT function) and a knockout strain lacking all central 5-HT neurons (Lmx1b(f/f/p) mice). We examined effects of acidosis, hypocretin-1, a hypocretin receptor antagonist (SB-408124), and the effect of the antagonist on the response to acidosis. We confirmed the critical role of 5-HT neurons in central chemosensitivity given that the increased hypoglossal burst frequency with acidosis, characteristic of WT mice, was absent in preparations derived from Lmx1b(f/f/p) mice. We also found that hypocretin facilitated baseline neural ventilatory output in part through 5-HT neurons. Although the impact of hypocretin on 5-HT neuronal sensitivity to acidosis is still unclear, hypocretins did appear to mediate the burst duration response to acidosis via serotonergic mechanisms.

  10. Knockdown of MALAT1 enhances chemosensitivity of ovarian cancer cells to cisplatin through inhibiting the Notch1 signaling pathway.

    Science.gov (United States)

    Bai, Lin; Wang, Aihua; Zhang, Yali; Xu, Xiaofeng; Zhang, Xiao

    2018-05-15

    Long non-coding RNAs (lncRNAs) are critical regulators in chemoresistance of various tumors including ovarian cancer. Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) has been reported to be upregulated and contributed to ovarian cancer tumorigenesis. The aim of this study was to explore the roles of MALAT1 and the underlying molecular regulatory mechanism in the chemoresistance of ovarian cancer cells. Our data demonstrated that MALAT1 and Notch1 mRNA were upregulated in ovarian cancer tissues, as well as cisplatin (CDDP)-resistant ovarian cancer cells. A positive correlation between MALAT1 and Notch1 mRNA expression was observed. MALAT1 knockdown significantly attenuated CDDP resistance, and enhanced CDDP-induced apoptosis in CDDP-resistant ovarian cancer cells. MALAT1 knockdown enhanced CDDP-induced apoptosis in vivo, as indicated by upregulation of Bax protein expression and downregulation of Bcl-2 protein expression. Additionally, MALAT1 knockdown inhibited the Notch1 pathway and ABCC1 expression in CDDP-resistant ovarian cancer cells. MALAT1 was demonstrated to interact with Notch1. Notch1 knockdown attenuated CDDP resistance, and downregulated the protein expression of ABCC1 in ovarian cancer cells. Taken together, our findings suggested that knockdown of MALAT-1 enhanced chemosensitivity of ovarian cancer cells to CDDP through inhibiting Notch1 signaling pathway. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. Interference with Activator Protein-2 transcription factors leads to induction of apoptosis and an increase in chemo- and radiation-sensitivity in breast cancer cells

    International Nuclear Information System (INIS)

    Thewes, Verena; Orso, Francesca; Jäger, Richard; Eckert, Dawid; Schäfer, Sabine; Kirfel, Gregor; Garbe, Stephan; Taverna, Daniela; Schorle, Hubert

    2010-01-01

    Activator Protein-2 (AP-2) transcription factors are critically involved in a variety of fundamental cellular processes such as proliferation, differentiation and apoptosis and have also been implicated in carcinogenesis. Expression of the family members AP-2α and AP-2γ is particularly well documented in malignancies of the female breast. Despite increasing evaluation of single AP-2 isoforms in mammary tumors the functional role of concerted expression of multiple AP-2 isoforms in breast cancer remains to be elucidated. AP-2 proteins can form homo- or heterodimers, and there is growing evidence that the net effect whether a cell will proliferate, undergo apoptosis or differentiate is partly dependent on the balance between different AP-2 isoforms. We simultaneously interfered with all AP-2 isoforms expressed in ErbB-2-positive murine N202.1A breast cancer cells by conditionally over-expressing a dominant-negative AP-2 mutant. We show that interference with AP-2 protein function lead to reduced cell number, induced apoptosis and increased chemo- and radiation-sensitivity. Analysis of global gene expression changes upon interference with AP-2 proteins identified 139 modulated genes (90 up-regulated, 49 down-regulated) compared with control cells. Gene Ontology (GO) investigations for these genes revealed Cell Death and Cell Adhesion and Migration as the main functional categories including 25 and 12 genes, respectively. By using information obtained from Ingenuity Pathway Analysis Systems we were able to present proven or potential connections between AP-2 regulated genes involved in cell death and response to chemo- and radiation therapy, (i.e. Ctgf, Nrp1, Tnfaip3, Gsta3) and AP-2 and other main apoptosis players and to create a unique network. Expression of AP-2 transcription factors in breast cancer cells supports proliferation and contributes to chemo- and radiation-resistance of tumor cells by impairing the ability to induce apoptosis. Therefore, interference

  12. Interference with Activator Protein-2 transcription factors leads to induction of apoptosis and an increase in chemo- and radiation- sensitivity in breast cancer cells

    LENUS (Irish Health Repository)

    Thewes, Verena

    2010-05-11

    Abstract Background Activator Protein-2 (AP-2) transcription factors are critically involved in a variety of fundamental cellular processes such as proliferation, differentiation and apoptosis and have also been implicated in carcinogenesis. Expression of the family members AP-2α and AP-2γ is particularly well documented in malignancies of the female breast. Despite increasing evaluation of single AP-2 isoforms in mammary tumors the functional role of concerted expression of multiple AP-2 isoforms in breast cancer remains to be elucidated. AP-2 proteins can form homo- or heterodimers, and there is growing evidence that the net effect whether a cell will proliferate, undergo apoptosis or differentiate is partly dependent on the balance between different AP-2 isoforms. Methods We simultaneously interfered with all AP-2 isoforms expressed in ErbB-2-positive murine N202.1A breast cancer cells by conditionally over-expressing a dominant-negative AP-2 mutant. Results We show that interference with AP-2 protein function lead to reduced cell number, induced apoptosis and increased chemo- and radiation-sensitivity. Analysis of global gene expression changes upon interference with AP-2 proteins identified 139 modulated genes (90 up-regulated, 49 down-regulated) compared with control cells. Gene Ontology (GO) investigations for these genes revealed Cell Death and Cell Adhesion and Migration as the main functional categories including 25 and 12 genes, respectively. By using information obtained from Ingenuity Pathway Analysis Systems we were able to present proven or potential connections between AP-2 regulated genes involved in cell death and response to chemo- and radiation therapy, (i.e. Ctgf, Nrp1, Tnfaip3, Gsta3) and AP-2 and other main apoptosis players and to create a unique network. Conclusions Expression of AP-2 transcription factors in breast cancer cells supports proliferation and contributes to chemo- and radiation-resistance of tumor cells by impairing the

  13. Resistance to ursodeoxycholic acid-induced growth arrest can also result in resistance to deoxycholic acid-induced apoptosis and increased tumorgenicity

    International Nuclear Information System (INIS)

    Powell, Ashley A; Akare, Sandeep; Qi, Wenqing; Herzer, Pascal; Jean-Louis, Samira; Feldman, Rebecca A; Martinez, Jesse D

    2006-01-01

    There is a large body of evidence which suggests that bile acids increase the risk of colon cancer and act as tumor promoters, however, the mechanism(s) of bile acids mediated tumorigenesis is not clear. Previously we showed that deoxycholic acid (DCA), a tumorogenic bile acid, and ursodeoxycholic acid (UDCA), a putative chemopreventive agent, exhibited distinct biological effects, yet appeared to act on some of the same signaling molecules. The present study was carried out to determine whether there is overlap in signaling pathways activated by tumorogenic bile acid DCA and chemopreventive bile acid UDCA. To determine whether there was an overlap in activation of signaling pathways by DCA and UDCA, we mutagenized HCT116 cells and then isolated cell lines resistant to UDCA induced growth arrest. These lines were then tested for their response to DCA induced apoptosis. We found that a majority of the cell lines resistant to UDCA-induced growth arrest were also resistant to DCA-induced apoptosis, implying an overlap in DCA and UDCA mediated signaling. Moreover, the cell lines which were the most resistant to DCA-induced apoptosis also exhibited a greater capacity for anchorage independent growth. We conclude that UDCA and DCA have overlapping signaling activities and that disregulation of these pathways can lead to a more advanced neoplastic phenotype

  14. Ubiquitination in apoptosis signaling

    NARCIS (Netherlands)

    van de Kooij, L.W.

    2014-01-01

    The work described in this thesis focuses on ubiquitination and protein degradation, with an emphasis on how these processes regulate apoptosis signaling. More specifically, our aims were: 1. To increase the understanding of ubiquitin-mediated regulation of apoptosis signaling. 2. To identify the E3

  15. Activation of Akt is increased in the dysplasia-carcinoma sequence in Barrett's oesophagus and contributes to increased proliferation and inhibition of apoptosis: a histopathological and functional study

    International Nuclear Information System (INIS)

    Beales, Ian LP; Ogunwobi, Olorunseun; Cameron, Ewen; El-Amin, Khalid; Mutungi, Gabriel; Wilkinson, Mark

    2007-01-01

    The incidence of oesophageal adenocarcinoma is increasing rapidly in the developed world. The serine-threonine protein kinase and proto-oncogene Akt has been reported to regulate proliferation and apoptosis in several tissues but there are no data on the involvement of Akt in oesophageal carcinogenesis. Therefore we have examined the activation of Akt in Barrett's oesophagus and oesophageal adenocarcinoma and the functional effects of Akt activation in vitro. Expression of total and active (phosphorylated) Akt were determined in endoscopic biopsies and surgical resection specimens using immunohistochemistry. The functional effects of Akt were examined using Barrett's adenocarcinoma cells in culture. In normal squamous oesophagus, erosive oesophagitis and non-dysplastic Barrett's oesophagus, phospho-Akt was limited to the basal 1/3 of the mucosa. Image analysis confirmed that Akt activation was significantly increased in non-dysplastic Barrett's oesophagus compared to squamous epithelium and further significantly increased in high-grade dysplasia and adenocarcinoma. In all cases of high grade dysplasia and adenocarcinoma Akt was activated in the luminal 1/3 of the epithelium. Transient acid exposure and the obesity hormone leptin activated Akt, stimulated proliferation and inhibited apoptosis: the combination of acid and leptin was synergistic. Inhibition of Akt phosphorylation with LY294002 increased apoptosis and blocked the effects of acid and leptin both alone and in combination. Activation of Akt was associated with downstream phosphorylation and deactivation of the pro-apoptotic protein Bad and phosphorylation of the Forkhead family transcription factor FOXO1. Akt is abnormally activated in Barrett's oesophagus, high grade dysplasia and adenocarcinoma. Akt activation promotes proliferation and inhibits apoptosis in Barrett's adenocarcinoma cells and both transient acid exposure and leptin stimulate Akt phosphorylation. Downstream

  16. ADAM12 redistributes and activates MMP-14, resulting in gelatin degradation, reduced apoptosis and increased tumor growth

    DEFF Research Database (Denmark)

    Albrechtsen, Reidar; Kveiborg, Marie; Hansen, Dorte Stautz

    2013-01-01

    Matrix metalloproteinases (MMPs), in particular MMP-2, MMP-9 and MMP-14, play a key role in various aspects of cancer pathology. Likewise, ADAMs (a disintegrin and metalloproteinases), including ADAM12, are upregulated in malignant tumors and contribute to the pathology of cancers. Here, we show...... degradation was stimulated and tumor cell apoptosis was decreased, with reduced expression of the pro-apoptotic proteins BCL2L11 and BIK. The effect on gelatin degradation was abrogated by inhibition of the MMP-14 activity and appeared to be dependent on cell surface αVβ3 integrin localization, but neither...... that there is a positive correlation between MMP-14 and ADAM12 expression in human breast cancer. We demonstrated that in 293-VnR and human breast cancer cells expressing ADAM12 at the cell surface, endogenous MMP-14 was recruited to the cell surface, resulting in its activation. Subsequent to this activation, gelatin...

  17. Increased topoisomerase IIalpha expression in colorectal cancer is associated with advanced disease and chemotherapeutic resistance via inhibition of apoptosis.

    LENUS (Irish Health Repository)

    Coss, Alan

    2012-02-01

    Topoisomerase IIalpha is a nuclear enzyme that regulates the tertiary structure of DNA. The influence of topoisomerase IIalpha gene (TOP2A) or protein alterations on disease progression and treatment response in colorectal cancer (CRC) is unknown. The study investigated the clinical relevance of topoisomerase IIalpha in CRC using in vivo and in vitro models. Differentially expressed genes in early and late-stage CRC were identified by array comparative genomic hybridization (CGH). Cellular location of gene amplifications was determined by fluorescence in situ hybridization (FISH). Topoisomerase IIalpha levels, proliferation index, and HER2 expression were examined in 228 colorectal tumors by immunohistochemistry. Overexpression of topoisomerase IIalpha in vitro was achieved by liposome-based transfection. Cell growth inhibition and apoptosis were quantified using the crystal violet assay and flow cytometry, respectively, in response to drug treatment. Amplification of TOP2A was identified in 3 (7.7%) tumors using array CGH and confirmed using FISH. At the protein level, topoisomerase IIalpha staining was observed in 157 (69%) tumors, and both staining and intensity levels were associated with an aggressive tumor phenotype (p values 0.04 and 0.005, respectively). Using logistic regression analysis, topoisomerase IIalpha remained significantly associated with advanced tumor stage when corrected for tumor proliferation (p=0.007) and differentiation (p=0.001). No association was identified between topoisomerase IIalpha and HER2. In vitro, overexpression of topoisomerase IIalpha was associated with resistance to irinotecan (p=0.001) and etoposide chemotherapy (p=0.03), an effect mediated by inhibition of apoptosis. Topoisomerase IIalpha overexpression is significantly associated with alterations in tumor behavior and response to drug treatment in CRC. Our results suggest that gene amplification may represent an important mechanism underlying these changes.

  18. Activation of the LKB1‑SIK1 signaling pathway inhibits the TGF‑β‑mediated epithelial‑mesenchymal transition and apoptosis resistance of ovarian carcinoma cells.

    Science.gov (United States)

    Hong, Bo; Zhang, Jianmei; Yang, Wenlan

    2018-02-01

    Ovarian cancer is the most common and lethal type of gynecological malignancy, due to its invasiveness. The present study aimed to analyze the molecular mechanism underlying chemoresistance in ovarian carcinoma cells, which may lead to local migration toward adjacent tissues and long‑distance metastasis to other organs. A total of 12 patients with ovarian fibroma were used to evaluate chemoresistance and chemosensitivity. The sensitivity and resistance of ovarian carcinoma cells was measured using apoptosis analysis, morphological observation, survival rate analysis, immunohistochemistry and immunostaining. The mechanism underlying the interaction between the epithelial‑mesenchymal transition (EMT) and liver kinase B1 (LKB1)‑salt‑inducible kinase 1 (SIK1) signaling pathways was additionally investigated in ovarian carcinoma. The results of the present study demonstrated that ovarian carcinoma cells isolated from patients exhibited apoptosis resistance. Inhibition of TGF‑β expression led to an inhibition of growth, migration and invasion, in addition to a promotion of apoptosis, in ovarian carcinoma cells treated with paclitaxel. Studies have indicated that the LKB1‑SIK1 signaling pathway may be suppressed in ovarian carcinoma cells compared with normal ovarian cells, leading to activation of the EMT signaling pathway. The results of the present study demonstrated that upregulation of LKB1 promoted SIK1 expression and markedly suppressed the growth and aggressiveness of ovarian cancer cells. Upregulation of LKB1 additionally promoted apoptosis in ovarian carcinoma cells. In addition, the results of the present study demonstrated that the knockdown of LKB1 further promoted the expression of transforming growth factor‑β and EMT, which downregulated the chemosensitivity of ovarian carcinoma cells. Additionally, overexpression of LKB1 in ovarian carcinoma cells increased chemosensitivity, resulting in a significant inhibition of migration and

  19. Increased expression of bHLH transcription factor E2A (TCF3) in prostate cancer promotes proliferation and confers resistance to doxorubicin induced apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Patel, Divya [Center for Cancer Research and Therapeutics Development, Clark Atlanta University, 223 James P. Brawley Dr. SW, Atlanta, GA 30314 (United States); Chaudhary, Jaideep, E-mail: jchaudhary@cau.edu [Center for Cancer Research and Therapeutics Development, Clark Atlanta University, 223 James P. Brawley Dr. SW, Atlanta, GA 30314 (United States); Dept. of Biological Sciences, Clark Atlanta University, 223 James P. Brawley Dr. SW, Atlanta, GA 30314 (United States)

    2012-05-25

    Highlights: Black-Right-Pointing-Pointer E2A, considered as a tumor suppressor is highly expressed in prostate cancer. Black-Right-Pointing-Pointer Silencing of E2A attenuates cell proliferation and promotes apoptosis. Black-Right-Pointing-Pointer E2A regulates c-myc, Id1, Id3 and CDKN1A expression. Black-Right-Pointing-Pointer Loss of E2A promotes doxorubicin dependent apoptosis in prostate cancer cells. Black-Right-Pointing-Pointer Results suggest that E2A acts as a tumor promoter at least in prostate cancer. -- Abstract: E2A (TCF3) is a multifunctional basic helix loop helix (bHLH), transcription factor. E2A regulates transcription of target genes by homo- or heterodimerization with cell specific bHLH proteins. In general, E2A promotes cell differentiation, acts as a negative regulator of cell proliferation in normal cells and cancer cell lines and is required for normal B-cell development. Given the diverse biological pathways regulated/influenced by E2A little is known about its expression in cancer. In this study we investigated the expression of E2A in prostate cancer. Unexpectedly, E2A immuno-histochemistry demonstrated increased E2A expression in prostate cancer as compared to normal prostate. Silencing of E2A in prostate cancer cells DU145 and PC3 led to a significant reduction in proliferation due to G1 arrest that was in part mediated by increased CDKN1A(p21) and decreased Id1, Id3 and c-myc. E2A silencing in prostate cancer cell lines also resulted in increased apoptosis due to increased mitochondrial permeability and caspase 3/7 activation. Moreover, silencing of E2A increased sensitivity to doxorubicin induced apoptosis. Based on our results, we propose that E2A could be an upstream regulator of Id1 and c-Myc which are highly expressed in prostate cancer. These results for the first time demonstrate that E2A could in fact acts as a tumor promoter at least in prostate cancer.

  20. Alteronol induces cell cycle arrest and apoptosis via increased reactive oxygen species production in human breast cancer T47D cells.

    Science.gov (United States)

    Ren, Boxue; Li, Defang; Si, Lingling; Ding, Yangfang; Han, Jichun; Chen, Xiaoyu; Zheng, Qiusheng

    2018-04-01

    Emerging evidence showed that alteronol has a potential antitumour effect in several tumour cells. However, the antitumour effect of alteronol on breast cancer has not been reported. This study investigated the mechanisms of alteronol-induced cell proliferation inhibition in human breast cancer T47D cells. After treatment with alteronol, T47D cell proliferation was examined by MTT assay. The cell cycle distribution, cell apoptosis, reactive oxygen species level and mitochondrial membrane potential were evaluated via flow cytometry. Next, the protein levels of cyclin B1, cdc2, p21, p-cyclin B1, p-cdc2, p53, Bax, Bcl-2 and cytochrome c were analysed using Western blot analysis. Meanwhile, the mRNA levels of cyclin B1, cdc2, p21 and p53 were examined by qRT-PCR. Our data showed that alteronol inhibited the proliferation of T47D cells via inducing G2-phase arrest and cell apoptosis. Compared with control group, alteronol significantly increased ROS level and triggered mitochondrial dysfunction in alteronol-treated T47D cells. Further studies showed that the mRNA and protein levels of cdc2 and cyclin B1 were downregulated, while the mRNA and protein levels of p21, p53, p-cyclin B1, p-cdc2 and cytochrome c were upregulated. In addition, the expression level of Bax was increased, and the expression level of Bcl-2 was decreased. Alteronol induced T47D cell cycle arrest and cell apoptosis through increasing ROS production and triggering mitochondrial dysfunction, and subsequently inhibiting T47D cell proliferation. © 2018 Royal Pharmaceutical Society.

  1. Mycoplasma hyopneumoniae-derived lipid-associated membrane proteins induce apoptosis in porcine alveolar macrophage via increasing nitric oxide production, oxidative stress, and caspase-3 activation.

    Science.gov (United States)

    Bai, Fangfang; Ni, Bo; Liu, Maojun; Feng, Zhixin; Xiong, Qiyan; Xiao, Shaobo; Shao, Guoqing

    2013-09-15

    Mycoplasma hyopneumoniae is the primary etiological agent of enzootic pneumonia in swine. Lipid-associated membrane proteins (LAMP) of mycoplasma are the main pathogenicity factors in mycoplasma diseases. In this study, we investigated the effects of M. hyopneumoniae LAMP on porcine alveolar macrophage (PAM) 3D4/21 cell line. Apoptotic features, such as chromatin condensation and apoptotic bodies, were observed in LAMP-treated PAM 3D4/21 cells. Moreover, LAMP significantly increased the number of TUNEL positive apoptotic cells in PAM 3D4/21 cells compared with the untreated control. In addition, flow cytometric analysis using dual staining with annexin-V-FITC and propidium iodide (PI) showed that LAMP of M. hyopneumoniae induced a time-dependent apoptosis in PAM 3D4/21 cells. Moreover, increased levels of superoxide anion production and activated caspase-3 in PAM 3D4/21 cells were observed after exposure to LAMP. Increased production of nitric oxide (NO) was also confirmed in the cell supernatants. Besides, apoptotic rates increase and caspase-3 activation were suppressed by NOS inhibitor or antioxidant. It is suggested that LAMP of M. hyopneumoniae induced apoptosis in porcine alveolar macrophage via NO production, superoxide anion production, and caspase-3 activation. Copyright © 2013 Elsevier B.V. All rights reserved.

  2. The Japanese Angelica acutiloba root and yokukansan increase hippocampal acetylcholine level, prevent apoptosis and improve memory in a rat model of repeated cerebral ischemia.

    Science.gov (United States)

    Nogami-Hara, Ai; Nagao, Masaki; Takasaki, Kotaro; Egashira, Nobuaki; Fujikawa, Risako; Kubota, Kaori; Watanabe, Takuya; Katsurabayashi, Shutaro; Hatip, Funda Bolukbasi; Hatip-Al-Khatib, Izzettin; Iwasaki, Katsunori

    2018-03-25

    Japanese Angelica acutiloba root (Angelica root) is included in several Kampo medicines including Yokukansan (YKS). Angelica root and YKS are used for the treatment of a variety of psychological and neurodegenerative disorders. Development of safe and effective therapeutic agents against cerebrovascular disorders will improve the treatment of patients with dementia. The effect of Angelica root and YKS on ischemia-impaired memory has not yet been fully investigated. The present study investigated whether Angelica root is also involved in memory improving and neuroprotective effect of YKS in a model of cerebrovascular ischemia. Male Wistar rats grouped into sham rats received saline, and other three groups subjected to repeated cerebral ischemia induced by 4-vessel occlusion (4-VO), received a 7-day oral administration of either saline, Angelica root or YKS. Memory was evaluated by eight-arm radial maze task. Acetylcholine release (ACh) in the dorsal hippocampus was investigated by microdialysis-HPLC. Apoptosis was determined by terminal deoxynucleotidyl transferase (TdT)-mediated fluorescein-deoxyuridine triphosphate (dUTP) nick-end labeling. Ischemia induced apoptosis, reduced release of ACh, and impaired the memory (increased error choices and decreased correct choices). Angelica root and YKS improved the memory deficits, upregulated the release of ACh and prevented 4-VO-induced hippocampal apoptosis. The dual ACh-increasing and neuroprotective effect of Angelica root could make it a promising therapeutic agent useful for the treatment of symptoms of cerebrovascular dementia. Angelica root could be one of the components contributing to the memory-improving and neuroprotective effects of YKS. Copyright © 2017. Published by Elsevier B.V.

  3. Theory of gastric CO2 ventilation and its control during respiratory acidosis: implications for central chemosensitivity, pH regulation, and diseases causing chronic CO2 retention.

    Science.gov (United States)

    Dean, Jay B

    2011-02-15

    The theory of gastric CO(2) ventilation describes a previously unrecognized reflex mechanism controlled by neurons in the caudal solitary complex (cSC) for non-alveolar elimination of systemic CO(2) during respiratory acidosis. Neurons in the cSC, which is a site of CO(2) chemosensitivity for cardiorespiratory control, also control various gastroesophageal reflexes that remove CO(2) from blood. CO(2) is consumed in the production of gastric acid and bicarbonate in the gastric epithelium and then reconstituted as CO(2) in the stomach lumen from the reaction between H(+) and HCO(3)(-). Respiratory acidosis and gastric CO(2) distension induce cSC/vagovagal mediated transient relaxations of the lower esophageal sphincter to vent gastric CO(2) upwards by bulk flow along an abdominal-to-esophageal (=intrapleural) pressure gradient the magnitude of which increases during abdominal (gastric) compression caused by increased contractions of respiratory muscles. Esophageal distension induces cSC/nucleus ambiguus/vagovagal reflex relaxation of the upper esophageal sphincter and CO(2) is vented into the pharynx and mixed with pulmonary gas during expiration or, alternatively, during eructation. It is proposed that gastric CO(2) ventilation provides explanations for (1) the postprandial increase in expired CO(2) and (2) the negative P(blood - expired)CO₂difference that occurs with increased inspired CO(2). Furthermore, it is postulated that gastric CO(2) ventilation and alveolar CO(2) ventilation are coordinated under dual control by CO(2) chemosensitive neurons in the cSC. This new theory, therefore, presupposes a level of neural control and coordination between two previously presumed dissimilar organ systems and supports the notion that different sites of CO(2) chemosensitivity address different aspects of whole body pH regulation. Consequently, not all sites of central chemosensitivity are equal regarding the mechanism(s) activated for CO(2) elimination. A distributed CO(2

  4. Decreased transcription-coupled nucleotide excision repair capacity is associated with increased p53- and MLH1-independent apoptosis in response to cisplatin

    International Nuclear Information System (INIS)

    Stubbert, Lawton J; Smith, Jennifer M; McKay, Bruce C

    2010-01-01

    One of the most commonly used classes of anti-cancer drugs presently in clinical practice is the platinum-based drugs, including cisplatin. The efficacy of cisplatin therapy is often limited by the emergence of resistant tumours following treatment. Cisplatin resistance is multi-factorial but can be associated with increased DNA repair capacity, mutations in p53 or loss of DNA mismatch repair capacity. RNA interference (RNAi) was used to reduce the transcription-coupled nucleotide excision repair (TC-NER) capacity of several prostate and colorectal carcinoma cell lines with specific defects in p53 and/or DNA mismatch repair. The effect of small inhibitory RNAs designed to target the CSB (Cockayne syndrome group B) transcript on TC-NER and the sensitivity of cells to cisplatin-induced apoptosis was determined. These prostate and colon cancer cell lines were initially TC-NER proficient and RNAi against CSB significantly reduced their DNA repair capacity. Decreased TC-NER capacity was associated with an increase in the sensitivity of tumour cells to cisplatin-induced apoptosis, even in p53 null and DNA mismatch repair-deficient cell lines. The present work indicates that CSB and TC-NER play a prominent role in determining the sensitivity of tumour cells to cisplatin even in the absence of p53 and DNA mismatch repair. These results further suggest that CSB represents a potential target for cancer therapy that may be important to overcome resistance to cisplatin in the clinic

  5. Reduced Nrf2 activation in PI3K phosphorylation-impaired vitiliginous keratinocytes increases susceptibility to ROS-generating chemical-induced apoptosis.

    Science.gov (United States)

    Kim, Hyangmi; Park, Chang Seo; Lee, Ai-Young

    2017-12-01

    Keratinocytes in affected epidermis of vitiligo patients are known to have impaired activation of the PI3K/AKT pathway. Based on critical roles of keratinocytes and oxidative stress in vitiligo development, this study examined whether keratinocytes with impaired PI3K activation were more vulnerable to apoptosis caused by oxidative stress from phenolic compounds, p-tert-butylphenol (4-TBP) and hydroquinone (HQ). Cell viability assay, FACS analysis, ELISA for TNF-α or IL-1a, ROS assay, Western blot analysis for Nrf2 expression, and confocal microscopy with anti-Nrf2 and phospho-PI3K antibodies were done on primary cultured normal human keratinocytes with or without PI3K knockdown in the presence or absence of chemical treatment or antioxidant. Immunofluorescence staining using anti-Nrf2, phospho-PI3K, TNF-ɑ, and IL-1ɑ antibodies, ROS assay using dihydroethidium, and TUNEL assay were performed on sets of depigmented and normally pigmented skin from vitiligo patients. Results showed that 4-TBP or HQ treatment increased apoptosis and the expression levels of TNF-ɑ, IL-1ɑ, and ROS in PI3K-knockdown keratinocytes which reduced Nrf2 nuclear translocation compared to control keratinocytes. These changes were significantly recovered by an antioxidant treatment. Depigmented epidermis of vitiligo patients also showed lower levels of Nrf2 and phospho-PI3K but higher levels of ROS, TNF-ɑ, IL-1ɑ, and ROS with more TUNEL-positive cells. Therefore, impaired PI3K activation in keratinocytes in depigmented epidermis of vitiligo patients are vulnerable to apoptosis caused by ROS-generating chemicals due to reduced Nrf2 activation. © 2017 Wiley Periodicals, Inc.

  6. Increased MiR-221 expression in hepatocellular carcinoma tissues and its role in enhancing cell growth and inhibiting apoptosis in vitro

    International Nuclear Information System (INIS)

    Rong, Minhua; Chen, Gang; Dang, Yiwu

    2013-01-01

    MiR-221 is over-expressed in human hepatocellular carcinoma (HCC), but its clinical significance and function in HCC remains uncertain. The aim of the study was to investigate the relationship between miR-221 overexpression and clinicopathological parameters in HCC formalin-fixed paraffin-embedded (FFPE) tissues, and the effect of miR-221 inhibitor and mimic on different HCC cell lines in vitro. MiR-221 expression was detected using real time RT-qPCR in FFPE HCC and the adjacent noncancerous liver tissues. The relationship between miR-221 level and clinicopathological features was also analyzed. Furthermore, miR-221 inhibitor and mimic were transfected into HCC cell lines HepB3, HepG2 and SNU449. The effects of miR-221 on cell growth, cell cycle, caspase activity and apoptosis were also investigated by spectrophotometry, fluorimetry, fluorescence microscopy and flow cytometry, respectively. The relative expression of miR-221 in clinical TNM stages III and IV was significantly higher than that in the stages I and II. The miR-221 level was also upregulated in the metastatic group compared to the nonmetastatic group. Furthermore, miR-221 over-expression was related to the status of tumor capsular infiltration in HCC clinical samples. Functionally, cell growth was inhibited, cell cycle was arrested in G1/S-phase and apoptosis was increased by miR-221 inhibitor in vitro. Likewise, miR-221 mimic accelerated the cell growth. Expression of miR-221 in FFPE tissues could provide predictive significance for prognosis of HCC patients. Moreover, miR-221 inhibitor could be useful to suppress proliferation and induce apoptosis in HCC cells. Thus miR-221 might be a critical targeted therapy strategy for HCC

  7. Chemosensitivity of MCF-7 cells to eugenol: release of cytochrome-c and lactate dehydrogenase

    Science.gov (United States)

    Al Wafai, Rana; El-Rabih, Warde; Katerji, Meghri; Safi, Remi; El Sabban, Marwan; El-Rifai, Omar; Usta, Julnar

    2017-01-01

    Phytochemicals have been extensively researched for their potential anticancer effects. In previous study, direct exposure of rat liver mitochondria to eugenol main ingredient of clove, uncoupled mitochondria and increased F0F1ATPase activity. In the present study, we further investigated the effects of eugenol on MCF-7 cells in culture. Eugenol demonstrated: a dose-dependent decrease in viability (MTT assay), and proliferation (real time cell analysis) of MCF-7 cells, (EC50: 0.9 mM); an increase in reactive oxygen species; a decrease in ATP level and mitochondrial membrane potential (MitoPT JC-1 assay); and a release of cytochrome-c and lactate dehydrogenase (Cytotoxicity Detection Kit PLUS) into culture media at eugenol concentration >EC50. Pretreatment with the antioxidants Trolox and N-acetyl cysteine partially restored cell viability and decreased ROS, with Trolox being more potent. Expression levels of both anti- and pro-apoptotic markers (Bcl-2 and Bax, respectively) decreased with increasing eugenol concentration, with no variation in their relative ratios. Eugenol-treated MCF-7 cells overexpressing Bcl-2 exhibited results similar to those of MCF-7. Our findings indicate that eugenol toxicity is non-apoptotic Bcl-2 independent, affecting mitochondrial function and plasma membrane integrity with no effect on migration or invasion. We report here the chemo-sensitivity of MCF-7 cells to eugenol, a phytochemical with anticancer potential. PMID:28272477

  8. Sodium orthovanadate associated with pharmacological doses of ascorbate causes an increased generation of ROS in tumor cells that inhibits proliferation and triggers apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Günther, T-hat nia Mara Fischer; Kviecinski, Maicon Roberto; Baron, Carla Cristine; Felipe, Karina Bettega; Farias, Mirelle Sifroni; Ourique da Silva, Fabiana; Bücker, Nádia Cristina Falcão [Departamento de Bioquímica, Universidade Federal de Santa Catarina, Florianópolis (Brazil); Pich, Claus Tröger [Campus de Araranguá, Universidade Federal de Santa Catarina, Araranguá (Brazil); Ferreira, Eduardo Antonio [Universidade de Brasília, Faculdade de Ceilândia, DF (Brazil); Filho, Danilo Wilhelm [Departamento de Ecologia e Zoologia, Universidade Federal de Santa Catarina, Florianópolis (Brazil); Verrax, Julien; Calderon, Pedro Buc [Toxicology and Cancer Biology Research Group, Louvain Drug Research Institute, Université Catholique de Louvain, Brussels (Belgium); Pedrosa, Rozangela Curi, E-mail: rozangelapedrosa@gmail.com [Departamento de Bioquímica, Universidade Federal de Santa Catarina, Florianópolis (Brazil)

    2013-01-18

    Graphical abstract: -- Abstract: Pharmacological doses of ascorbate were evaluated for its ability to potentiate the toxicity of sodium orthovanadate (Na{sub 3}VO{sub 4}) in tumor cells. Cytotoxicity, inhibition of cell proliferation, generation of ROS and DNA fragmentation were assessed in T24 cells. Na{sub 3}VO{sub 4} was cytotoxic against T24 cells (EC{sub 50} = 5.8 μM at 24 h), but in the presence of ascorbate (100 μM) the EC{sub 50} fell to 3.3 μM. Na{sub 3}VO{sub 4} plus ascorbate caused a strong inhibition of cell proliferation (up to 20%) and increased the generation of ROS (4-fold). Na{sub 3}VO{sub 4} did not directly cleave plasmid DNA, at this aspect no synergism was found occurring between Na{sub 3}VO{sub 4} and ascorbate once the resulting action of the combination was no greater than that of both substances administered separately. Cells from Ehrlich ascites carcinoma-bearing mice were used to determine the activity of antioxidant enzymes, the extent of the oxidative damage and the type of cell death. Na{sub 3}VO{sub 4} alone, or combined with ascorbate, increased catalase activity, but only Na{sub 3}VO{sub 4} plus ascorbate increased superoxide dismutase activity (up to 4-fold). Oxidative damage on proteins and lipids was higher due to the treatment done with Na{sub 3}VO{sub 4} plus ascorbate (2–3-fold). Ascorbate potentiated apoptosis in tumor cells from mice treated with Na{sub 3}VO{sub 4}. The results indicate that pharmacological doses of ascorbate enhance the generation of ROS induced by Na{sub 3}VO{sub 4} in tumor cells causing inhibition of proliferation and apoptosis. Apoptosis induced by orthovanadate and ascorbate is closer related to inhibition on Bcl-xL and activation of Bax. Our data apparently rule out a mechanism of cell demise p53-dependent or related to Cdk2 impairment.

  9. Knockdown of astrocyte elevated gene-1 inhibits proliferation and enhancing chemo-sensitivity to cisplatin or doxorubicin in neuroblastoma cells

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    Xie Li

    2009-02-01

    Full Text Available Abstract Background Astrocyte elevated gene-1 (AEG-1 was originally characterized as a HIV-1-inducible gene in primary human fetal astrocyte. Recent studies highlight a potential role of AEG-1 in promoting tumor progression and metastasis. The aim of this study was to investigate if AEG-1 serves as a potential therapeutic target of human neuroblastoma. Methods We employed RNA interference to reduce AEG-1 expression in human neuroblastoma cell lines and analyzed their phenotypic changes. Results We found that the knockdown of AEG-1 expression in human neuroblastoma cells significantly inhibited cell proliferation and apoptosis. The specific downregulation induced cell arrest in the G0/G1 phase of cell cycle. In the present study, we also observed a significant enhancement of chemo-sensitivity to cisplatin and doxorubicin by knockdown of AEG-1. Conclusion Our study suggests that overexpressed AEG-1 enhance the tumorogenic properties of neuroblastoma cells. The inhibition of AEG-1 expression could be a new adjuvant therapy for neuroblastoma.

  10. PNMA2 mediates heterodimeric interactions and antagonizes chemo-sensitizing activities mediated by members of PNMA family.

    Science.gov (United States)

    Lee, Yong Hoi; Pang, Siew Wai; Tan, Kuan Onn

    2016-04-22

    PNMA2, a member of the Paraneoplastic Ma Family (PNMA), was identified through expression cloning by using anti-sera from patients with paraneoplastic disorder. Tissue expression studies showed that PNMA2 was predominantly expressed in normal human brain; however, the protein was shown to exhibit abnormal expression profile as it was found to be expressed in a number of tumour tissues obtained from paraneopalstic patients. The abnormal expression profile of PNMA2 suggests that it might play an important role in tumorigenesis; however, apart from protein expression and immunological studies, the physiological role of PNMA2 remains unclear. In order to determine potential role of PNMA2 in tumorigenesis, and its functional relationship with PNMA family members, MOAP-1 (PNMA4) and PNMA1, expression constructs encoding the respective proteins were generated for both in vitro and in vivo studies. Our investigations showed that over-expressed MOAP-1 and PNMA1 promoted apoptosis and chemo-sensitization in MCF-7 cells as evidenced by condensed nuclei and Annexin-V positive MCF-7 cells; however, the effects mediated by these proteins were significantly inhibited or abolished when co-expressed with PNMA2 in MCF-7 cells. Furthermore, co-immunoprecipitation study showed that PNMA1 and MOAP-1 failed to associate with each other but readily formed respective heterodimer with PNMA2, suggesting that PNMA2 functions as antagonist of MOAP-1 and PNMA1 through heterodimeric interaction. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Signalling and chemosensitivity assays in melanoma: is mutated status a prerequisite for targeted therapy?

    Science.gov (United States)

    Passeron, Thierry; Lacour, Jean-Philippe; Allegra, Maryline; Ségalen, Coralie; Deville, Anne; Thyss, Antoine; Giacchero, Damien; Ortonne, Jean-Paul; Bertolotto, Corine; Ballotti, Robert; Bahadoran, Philippe

    2011-12-01

    Selection for targeted therapies in melanoma is currently based on the search for mutations in selected genes. We aimed at evaluating the interest of signalling and chemosensitivity studies in addition to genotyping for assessing the best suitable treatment in an individual patient. We extracted genomic DNA and melanoma cells from tumor tissue of a skin metastasis of a 17-year-old woman with stage IV melanoma progressing despite three successive lines of treatment. Despite the absence of mutation in BRAF, NRAS cKIT, the MAPK pathway was activated and a significant response to sorafenib, a mitogen-activated protein kinase (MAPK)/RAF inhibitor, was found in signalling and chemosensitivity assays. A treatment combining sorafenib and dacarbazine produced a partial response for 9 months, with marked necrosis in some lesions. Chemosensitivity assays and signalling pathway studies could be of great value in addition to genotyping for assessing the most appropriate treatment in melanoma. © 2011 John Wiley & Sons A/S.

  12. Depletion of intrinsic expression of Interleukin-8 in prostate cancer cells causes cell cycle arrest, spontaneous apoptosis and increases the efficacy of chemotherapeutic drugs

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    Lokeshwar Bal L

    2009-07-01

    Full Text Available Abstract Background The progression of all cancers is characterized by increased-cell proliferation and decreased-apoptosis. The androgen-independent prostate cancer (AIPC is the terminal stage of the disease. Many chemokines and cytokines are suspects to cause this increased tumor cell survival that ultimately leads to resistance to therapy and demise of the host. The AIPC cells, but not androgen-responsive cells, constitutively express abundant amount of the pro-inflammatory chemokine, Interleukin-8 (IL-8. The mechanism of IL-8 mediated survival and therapeutic resistance in AIPC cells is unclear at present. The purpose of this report is to show the pervasive role of IL-8 in malignant progression of androgen-independent prostate cancer (AIPC and to provide a potential new therapeutic avenue, using RNA interference. Results The functional consequence of IL-8 depletion in AIPC cells was investigated by RNA interference in two IL-8 secreting AIPC cell lines, PC-3 and DU145. The non-IL-8 secreting LNCaP and LAPC-4 cells served as controls. Cells were transfected with RISC-free siRNA (control or validated-pool of IL-8 siRNA. Transfection with 50 nM IL-8 siRNA caused >95% depletion of IL-8 mRNA and >92% decrease in IL-8 protein. This reduction in IL-8 led to cell cycle arrest at G1/S boundary and decreases in cell cycle-regulated proteins: Cyclin D1 and Cyclin B1 (both decreased >50% and inhibition of ERK1/2 activity by >50%. Further, the spontaneous apoptosis was increased by >43% in IL-8 depleted cells, evidenced by increases in caspase-9 activation and cleaved-PARP. IL-8 depletion caused significant decreases in anti-apoptotic proteins, BCL-2, BCL-xL due to decrease in both mRNA and post-translational stability, and increased levels of pro-apoptotic BAX and BAD proteins. More significantly, depletion of intracellular IL-8 increased the cytotoxic activity of multiple chemotherapeutic drugs. Specifically, the cytotoxicity of Docetaxel

  13. 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

  14. Nrdp1 Increases Ischemia Induced Primary Rat Cerebral Cortical Neurons and Pheochromocytoma Cells Apoptosis Via Downregulation of HIF-1α Protein

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    Yuan Zhang

    2017-09-01

    Full Text Available Neuregulin receptor degradation protein-1 (Nrdp1 is an E3 ubiquitin ligase that targets proteins for degradation and regulates cell growth, apoptosis and oxidative stress in various cell types. We have previously shown that Nrdp1 is implicated in ischemic cardiomyocyte death. In this study, we investigated the change of Nrdp1 expression in ischemic neurons and its role in ischemic neuronal injury. Primary rat cerebral cortical neurons and pheochromocytoma (PC12 cells were infected with adenoviral constructs expressing Nrdp1 gene or its siRNA before exposing to oxygen-glucose deprivation (OGD treatment. Our data showed that Nrdp1 was upregulated in ischemic brain tissue 3 h after middle cerebral artery occlusion (MCAO and in OGD-treated neurons. Of note, Nrdp1 overexpression by Ad-Nrdp1 enhanced OGD-induced neuron apoptosis, while knockdown of Nrdp1 with siRNA attenuated this effect, implicating a role of Nrdp1 in ischemic neuron injury. Moreover, Nrdp1 upregulation is accompanied by increased protein ubiquitylation and decreased protein levels of ubiquitin-specific protease 8 (USP8 in OGD-treated neurons, which led to a suppressed interaction between USP8 and HIF-1α and subsequently a reduction in HIF-1α protein accumulation in neurons under OGD conditions. In conclusion, our data support an important role of Nrdp1 upregulation in ischemic neuronal death, and suppressing the interaction between USP8 and HIF-1α and consequently the hypoxic adaptive response of neurons may account for this detrimental effect.

  15. Concurrent Inhibition of Pim and FLT3 Kinases Enhances Apoptosis of FLT3-ITD Acute Myeloid Leukemia Cells through Increased Mcl-1 Proteasomal Degradation.

    Science.gov (United States)

    Kapoor, Shivani; Natarajan, Karthika; Baldwin, Patrick R; Doshi, Kshama A; Lapidus, Rena G; Mathias, Trevor J; Scarpa, Mario; Trotta, Rossana; Davila, Eduardo; Kraus, Manfred; Huszar, Dennis; Tron, Adriana E; Perrotti, Danilo; Baer, Maria R

    2018-01-01

    Purpose: fms -like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) is present in 30% of acute myeloid leukemia (AML), and these patients have short disease-free survival. FLT3 inhibitors have limited and transient clinical activity, and concurrent treatment with inhibitors of parallel or downstream signaling may improve responses. The oncogenic serine/threonine kinase Pim-1 is upregulated downstream of FLT3-ITD and also promotes its signaling in a positive feedback loop, suggesting benefit of combined Pim and FLT3 inhibition. Experimental Design: Combinations of clinically active Pim and FLT3 inhibitors were studied in vitro and in vivo Results: Concurrent treatment with the pan-Pim inhibitor AZD1208 and FLT3 inhibitors at clinically applicable concentrations abrogated in vitro growth of FLT3-ITD, but not wild-type FLT3 (FLT3-WT), cell lines. AZD1208 cotreatment increased FLT3 inhibitor-induced apoptosis of FLT3-ITD, but not FLT3-WT, cells measured by sub-G 1 fraction, annexin V labeling, mitochondrial membrane potential, and PARP and caspase-3 cleavage. Concurrent treatment with AZD1208 and the FLT3 inhibitor quizartinib decreased growth of MV4-11 cells, with FLT3-ITD, in mouse xenografts, and prolonged survival, enhanced apoptosis of FLT3-ITD primary AML blasts, but not FLT3-WT blasts or remission marrow cells, and decreased FLT3-ITD AML blast colony formation. Mechanistically, AZD1208 and quizartinib cotreatment decreased expression of the antiapoptotic protein Mcl-1. Decrease in Mcl-1 protein expression was abrogated by treatment with the proteasome inhibitor MG132, and was preceded by downregulation of the Mcl-1 deubiquitinase USP9X, a novel mechanism of Mcl-1 regulation in AML. Conclusions: The data support clinical testing of Pim and FLT3 inhibitor combination therapy for FLT3-ITD AML. Clin Cancer Res; 24(1); 234-47. ©2017 AACR . ©2017 American Association for Cancer Research.

  16. Anaplasma phagocytophilum increases the levels of histone modifying enzymes to inhibit cell apoptosis and facilitate pathogen infection in the tick vector Ixodes scapularis.

    Science.gov (United States)

    Cabezas-Cruz, Alejandro; Alberdi, Pilar; Ayllón, Nieves; Valdés, James J; Pierce, Raymond; Villar, Margarita; de la Fuente, José

    2016-04-02

    Epigenetic mechanisms have not been characterized in ticks despite their importance as vectors of human and animal diseases worldwide. The objective of this study was to characterize the histones and histone modifying enzymes (HMEs) of the tick vector Ixodes scapularis and their role during Anaplasma phagocytophilum infection. We first identified 5 histones and 34 HMEs in I. scapularis in comparison with similar proteins in model organisms. Then, we used transcriptomic and proteomic data to analyze the mRNA and protein levels of I. scapularis histones and HMEs in response to A. phagocytophilum infection of tick tissues and cultured cells. Finally, selected HMEs were functionally characterized by pharmacological studies in cultured tick cells. The results suggest that A. phagocytophilum manipulates tick cell epigenetics to increase I. scapularis p300/CBP, histone deacetylase, and Sirtuin levels, resulting in an inhibition of cell apoptosis that in turn facilitates pathogen infection and multiplication. These results also suggest that a compensatory mechanism might exist by which A. phagocytophilum manipulates tick HMEs to regulate transcription and apoptosis in a tissue-specific manner to facilitate infection, but preserving tick fitness to guarantee survival of both pathogens and ticks. Our study also indicates that the pathogen manipulates arthropod and vertebrate cell epigenetics in similar ways to inhibit the host response to infection. Epigenetic regulation of tick biological processes is an essential element of the infection by A. phagocytophilum and the study of the mechanisms and principal actors involved is likely to provide clues for the development of anti-tick drugs and vaccines.

  17. Increases in apoptosis, caspase activity and expression of p53 and bax, and the transition between two types of mitochondrion-rich cells, in the gills of the climbing perch, Anabas testudineus, during a progressive acclimation from freshwater to seawater

    Science.gov (United States)

    Ching, Biyun; Chen, Xiu L.; Yong, Jing H. A.; Wilson, Jonathan M.; Hiong, Kum C.; Sim, Eugene W. L.; Wong, Wai P.; Lam, Siew H.; Chew, Shit F.; Ip, Yuen K.

    2013-01-01

    This study aimed to test the hypothesis that branchial osmoregulatory acclimation involved increased apoptosis and replacement of mitochdonrion-rich cells (MRCs) in the climbing perch, Anabas testudineus, during a progressive acclimation from freshwater to seawater. A significant increase in branchial caspase-3/-7 activity was observed on day 4 (salinity 20), and an extensive TUNEL-positive apoptosis was detected on day 5 (salinity 25), indicating salinity-induced apoptosis had occurred. This was further supported by an up-regulation of branchial mRNA expression of p53, a key regulator of cell cycle arrest and apoptosis, between day 2 (salinity 10) and day 6 (seawater), and an increase in branchial p53 protein abundance on day 6. Seawater acclimation apparently activated both the extrinsic and intrinsic pathways, as reflected by significant increases in branchial caspase-8 and caspase-9 activities. The involvement of the intrinsic pathway was confirmed by the significant increase in branchial mRNA expression of bax between day 4 (salinity 20) and day 6 (seawater). Western blotting results revealed the presence of a freshwater Na+/K+-ATPase (Nka) α-isoform, Nka α1a, and a seawater isoform, Nka α1b, the protein abundance of which decreased and increased, respectively, during seawater acclimation. Immunofluorescence microscopy revealed the presence of two types of MRCs distinctly different in sizes, and confirmed that the reduction in Nka α1a expression, and the prominent increases in expression of Nka α1b, Na+:K+:2Cl− cotransporter 1, and cystic fibrosis transmembrane conductance regulator Cl− channel coincided with the salinity-induced apoptotic event. Since modulation of existing MRCs alone could not have led to extensive salinity-induced apoptosis, it is probable that some, if not all, freshwater-type MRCs could have been removed through increased apoptosis and subsequently replaced by seawater-type MRCs in the gills of A. testudineus during seawater

  18. Elevated Markers of Death Receptor-Activated Apoptosis are Associated with Increased Risk for Development of Diabetes and Cardiovascular Disease

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    Ingrid Yao Mattisson

    2017-12-01

    Conclusion: The present study demonstrates an association between several cardiovascular risk factors and elevated levels of circulating markers of apoptotic cell death. It also shows that subjects with high levels of these biomarkers have increased risk of diabetes and CVD. This implies that soluble death receptors are markers of β-cell and vascular injury and potentially could be used as surrogate markers of therapeutic efficiency in risk factor interventions.

  19. Khz (fusion of Ganoderma lucidum and Polyporus umbellatus mycelia induces apoptosis by increasing intracellular calcium levels and activating JNK and NADPH oxidase-dependent generation of reactive oxygen species.

    Directory of Open Access Journals (Sweden)

    Tae Hwan Kim

    Full Text Available Khz is a compound derived from the fusion of Ganoderma lucidum and Polyporus umbellatus mycelia that inhibits the growth of cancer cells. The results of the present study show that Khz induced apoptosis preferentially in transformed cells and had only minimal effects on non-transformed cells. Furthermore, Khz induced apoptosis by increasing the intracellular Ca(2+ concentration ([Ca(2+](i and activating JNK to generate reactive oxygen species (ROS via NADPH oxidase and the mitochondria. Khz-induced apoptosis was caspase-dependent and occurred via a mitochondrial pathway. ROS generation by NADPH oxidase was critical for Khz-induced apoptosis, and although mitochondrial ROS production was also required, it appeared to occur secondary to ROS generation by NADPH oxidase. Activation of NADPH oxidase was demonstrated by the translocation of regulatory subunits p47(phox and p67(phox to the cell membrane and was necessary for ROS generation by Khz. Khz triggered a rapid and sustained increase in [Ca(2+](i, which activated JNK. JNK plays a key role in the activation of NADPH oxidase because inhibition of its expression or activity abrogated membrane translocation of the p47(phox and p67(phox subunits and ROS generation. In summary, these data indicate that Khz preferentially induces apoptosis in cancer cells, and the signaling mechanisms involve an increase in [Ca(2+](i, JNK activation, and ROS generation via NADPH oxidase and mitochondria.

  20. Targeting the Mitochondrial Respiratory Chain of Cryptococcus through Antifungal Chemosensitization: A Model for Control of Non-Fermentative Pathogens

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    Kathleen L. Chan

    2013-07-01

    Full Text Available Enhanced control of species of Cryptococcus, non-fermentative yeast pathogens, was achieved by chemosensitization through co-application of certain compounds with a conventional antimicrobial drug. The species of Cryptococcus tested showed higher sensitivity to mitochondrial respiratory chain (MRC inhibition compared to species of Candida. This higher sensitivity results from the inability of Cryptococcus to generate cellular energy through fermentation. To heighten disruption of cellular MRC, octyl gallate (OG or 2,3-dihydroxybenzaldehyde (2,3-DHBA, phenolic compounds inhibiting mitochondrial functions, were selected as chemosensitizers to pyraclostrobin (PCS; an inhibitor of complex III of MRC. The cryptococci were more susceptible to the chemosensitization (i.e., PCS + OG or 2,3-DHBA than the Candida with all Cryptococcus strains tested being sensitive to this chemosensitization. Alternatively, only few of the Candida strains showed sensitivity. OG possessed higher chemosensitizing potency than 2,3-DHBA, where the concentration of OG required with the drug to achieve chemosensitizing synergism was much lower than that required of 2,3-DHBA. Bioassays with gene deletion mutants of the model yeast Saccharomyces cerevisiae showed that OG or 2,3-DHBA affect different cellular targets. These assays revealed mitochondrial superoxide dismutase or glutathione homeostasis plays a relatively greater role in fungal tolerance to 2,3-DHBA or OG, respectively. These findings show that application of chemosensitizing compounds that augment MRC debilitation is a promising strategy to antifungal control against yeast pathogens.

  1. LIMITATIONS OF THE FAST GREEN ASSAY FOR CHEMOSENSITIVITY TESTING IN HUMAN LUNG-CANCER

    NARCIS (Netherlands)

    SMIT, EF; DEVRIES, EGE; MEIJER, C; MULDER, NH; POSTMUS, PE

    1991-01-01

    Selection of patients with lung cancer who are most likely to benefit from chemotherapeutic treatment would be a substantial step forward. Therefore, a prospective study in predictive chemosensitivity testing in vitro using the fast green assay (FGA) as developed by Weisenthal et al was carried out.

  2. Resistance gene expression determines the in vitro chemosensitivity of non-small cell lung cancer (NSCLC

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    Amer Khalid

    2009-08-01

    Full Text Available Abstract Background NSCLC exhibits considerable heterogeneity in its sensitivity to chemotherapy and similar heterogeneity is noted in vitro in a variety of model systems. This study has tested the hypothesis that the molecular basis of the observed in vitro chemosensitivity of NSCLC lies within the known resistance mechanisms inherent to these patients' tumors. Methods The chemosensitivity of a series of 49 NSCLC tumors was assessed using the ATP-based tumor chemosensitivity assay (ATP-TCA and compared with quantitative expression of resistance genes measured by RT-PCR in a Taqman Array™ following extraction of RNA from formalin-fixed paraffin-embedded (FFPE tissue. Results There was considerable heterogeneity between tumors within the ATP-TCA, and while this showed no direct correlation with individual gene expression, there was strong correlation of multi-gene signatures for many of the single agents and combinations tested. For instance, docetaxel activity showed some dependence on the expression of drug pumps, while cisplatin activity showed some dependence on DNA repair enzyme expression. Activity of both drugs was influenced more strongly still by the expression of anti- and pro-apoptotic genes by the tumor for both docetaxel and cisplatin. The doublet combinations of cisplatin with gemcitabine and cisplatin with docetaxel showed gene expression signatures incorporating resistance mechanisms for both agents. Conclusion Genes predicted to be involved in known mechanisms drug sensitivity and resistance correlate well with in vitro chemosensitivity and may allow the definition of predictive signatures to guide individualized chemotherapy in lung cancer.

  3. Sulforaphane increases the efficacy of doxorubicin in mouse fibroblasts characterized by p53 mutations

    Energy Technology Data Exchange (ETDEWEB)

    Fimognari, Carmela [Department of Pharmacology, University of Bologna, Bologna (Italy)]. E-mail: carmela.fimognari@unibo.it; Nuesse, Michael [GSF-Flow Cytometry Group, Neuherberg (Germany); Lenzi, Monia [Department of Pharmacology, University of Bologna, Bologna (Italy); Sciuscio, Davide [Department of Pharmacology, University of Bologna, Bologna (Italy); Cantelli-Forti, Giorgio [Department of Pharmacology, University of Bologna, Bologna (Italy); Hrelia, Patrizia [Department of Pharmacology, University of Bologna, Bologna (Italy)

    2006-10-10

    One novel strategy for increasing cancer chemotherapy efficacy and reversing chemoresistance involves co-administration of natural chemopreventive compounds alongside standard chemotherapeutic protocols. Sulforaphane is a particularly promising chemopreventive agent, which has been shown to exert proapoptotic effects on tumor cells containing p53 mutations. The p53{sup Ser220} mutation has been implicated in reduced efficacy and drug resistance in the context of osteosarcomas and breast tumors treated with doxorubicin-based protocols. We investigated the effects of a combination of doxorubicin and sulforaphane on cell viability and apoptosis induction in fibroblasts characterized by different p53 status (p53 wild-type, p53 knock-out, and p53{sup Ser220} mutation), and identified some of the molecular pathways triggered by the drug combination. Very high concentrations of doxorubicin were necessary to decrease the viability of p53{sup Ser220} and p53 knock-out (but not wild-type) cells. Treatment of p53{sup Ser220} and p53 knock-out cells with doxorubicin did not induce apoptosis, also at very high concentrations (10 {mu}M). Sulforaphane restored chemosensitivity and induced apoptosis in doxorubicin-resistant p53{sup Ser220} and p53 knock-out cells, irrespective of p53 status. The induction of apoptosis was caspase-3 dependent and caspase-8 independent. Bongkrekic acid, a mitochondrial membrane stabilizer, partially prevented the effects of doxorubicin plus sulforaphane on mitochondrial permeability but was unable to prevent the induction of apoptosis. N-acetyl-cysteine, a glutathione precursor, blocked the induction of apoptosis by doxorubicin plus sulforaphane. Considering the negligible safety profile of sulforaphane, our findings could prompt innovative clinical studies designed to investigate whether its coadministration can enhance the efficacy of doxorubicin-based regimens.

  4. Chemosensitization of Fusarium graminearum to Chemical Fungicides Using Cyclic Lipopeptides Produced by Bacillus amyloliquefaciens Strain JCK-12

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    Kihyun Kim

    2017-11-01

    Full Text Available Fusarium head blight (FHB caused by infection with Fusarium graminearum leads to enormous losses to crop growers, and may contaminate grains with a number of Fusarium mycotoxins that pose serious risks to human and animal health. Antagonistic bacteria that are used to prevent FHB offer attractive alternatives or supplements to synthetic fungicides for controlling FHB without the negative effects of chemical management. Out of 500 bacterial strains isolated from soil, Bacillus amyloliquefaciens JCK-12 showed strong antifungal activity and was considered a potential source for control strategies to reduce FHB. B. amyloliquefaciens JCK-12 produces several cyclic lipopeptides (CLPs including iturin A, fengycin, and surfactin. Iturin A inhibits spore germination of F. graminearum. Fengycin or surfactin alone did not display any inhibitory activity against spore germination at concentrations less than 30 μg/ml, but a mixture of iturin A, fengycin, and surfactin showed a remarkable synergistic inhibitory effect on F. graminearum spore germination. The fermentation broth and formulation of B. amyloliquefaciens JCK-12 strain reduced the disease incidence of FHB in wheat. Furthermore, co-application of B. amyloliquefaciens JCK-12 and chemical fungicides resulted in synergistic in vitro antifungal effects and significant disease control efficacy against FHB under greenhouse and field conditions, suggesting that B. amyloliquefaciens JCK-12 has a strong chemosensitizing effect. The synergistic antifungal effect of B. amyloliquefaciens JCK-12 and chemical fungicides in combination may result from the cell wall damage and altered cell membrane permeability in the phytopathogenic fungi caused by the CLP mixtures and subsequent increased sensitivity of F. graminearum to fungicides. In addition, B. amyloliquefaciens JCK-12 showed the potential to reduce trichothecenes mycotoxin production. The results of this study indicate that B. amyloliquefaciens JCK-12 could

  5. Chemosensitization ofFusarium graminearumto Chemical Fungicides Using Cyclic Lipopeptides Produced byBacillus amyloliquefaciensStrain JCK-12.

    Science.gov (United States)

    Kim, Kihyun; Lee, Yoonji; Ha, Areum; Kim, Ji-In; Park, Ae Ran; Yu, Nan Hee; Son, Hokyoung; Choi, Gyung Ja; Park, Hae Woong; Lee, Chul Won; Lee, Theresa; Lee, Yin-Won; Kim, Jin-Cheol

    2017-01-01

    Fusarium head blight (FHB) caused by infection with Fusarium graminearum leads to enormous losses to crop growers, and may contaminate grains with a number of Fusarium mycotoxins that pose serious risks to human and animal health. Antagonistic bacteria that are used to prevent FHB offer attractive alternatives or supplements to synthetic fungicides for controlling FHB without the negative effects of chemical management. Out of 500 bacterial strains isolated from soil, Bacillus amyloliquefaciens JCK-12 showed strong antifungal activity and was considered a potential source for control strategies to reduce FHB. B. amyloliquefaciens JCK-12 produces several cyclic lipopeptides (CLPs) including iturin A, fengycin, and surfactin. Iturin A inhibits spore germination of F. graminearum. Fengycin or surfactin alone did not display any inhibitory activity against spore germination at concentrations less than 30 μg/ml, but a mixture of iturin A, fengycin, and surfactin showed a remarkable synergistic inhibitory effect on F. graminearum spore germination. The fermentation broth and formulation of B. amyloliquefaciens JCK-12 strain reduced the disease incidence of FHB in wheat. Furthermore, co-application of B. amyloliquefaciens JCK-12 and chemical fungicides resulted in synergistic in vitro antifungal effects and significant disease control efficacy against FHB under greenhouse and field conditions, suggesting that B. amyloliquefaciens JCK-12 has a strong chemosensitizing effect. The synergistic antifungal effect of B. amyloliquefaciens JCK-12 and chemical fungicides in combination may result from the cell wall damage and altered cell membrane permeability in the phytopathogenic fungi caused by the CLP mixtures and subsequent increased sensitivity of F. graminearum to fungicides. In addition, B. amyloliquefaciens JCK-12 showed the potential to reduce trichothecenes mycotoxin production. The results of this study indicate that B. amyloliquefaciens JCK-12 could be used as an

  6. miR-212 increases tumor necrosis factor-related apoptosis-inducing ligand sensitivity in non-small cell lung cancer by targeting the antiapoptotic protein PED.

    Science.gov (United States)

    Incoronato, Mariarosaria; Garofalo, Michela; Urso, Loredana; Romano, Giulia; Quintavalle, Cristina; Zanca, Ciro; Iaboni, Margherita; Nuovo, Gerald; Croce, Carlo Maria; Condorelli, Gerolama

    2010-05-01

    PED/PEA-15 (PED) is a death effector domain family member of 15 kDa with a broad antiapoptotic function found overexpressed in a number of different human tumors, including lung cancer. To date, the mechanisms that regulate PED expression are unknown. Therefore, we address this point by the identification of microRNAs that in non-small cell lung cancer (NSCLC) modulate PED levels. In this work, we identify miR-212 as a negative regulator of PED expression. We also show that ectopic expression of this miR increases tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced cell death in NSCLC cells. In contrast, inhibition of endogenous miR-212 by use of antago-miR results in increase of PED protein expression and resistance to TRAIL treatment. Besides, in NSCLC, we show both in vitro and in vivo that PED and miR-212 expressions are inversely correlated, that is, PED is upregulated and miR-212 is rarely expressed. In conclusion, these findings suggest that miR-212 should be considered as a tumor suppressor because it negatively regulates the antiapoptotic protein PED and regulates TRAIL sensitivity. (c)2010 AACR.

  7. Expression of miR-200c in claudin-low breast cancer alters stem cell functionality, enhances chemosensitivity and reduces metastatic potential

    Science.gov (United States)

    Knezevic, Jana; Pfefferle, Adam D.; Petrovic, Ivana; Greene, Stephanie B.; Perou, Charles M.; Rosen, Jeffrey M.

    2015-01-01

    Claudin-low tumors are a highly aggressive breast cancer subtype with no targeted treatments and a clinically documented resistance to chemotherapy. They are significantly enriched in cancer stem cells (CSCs), which makes claudin-low tumor models particularly attractive for studying CSC behavior and developing novel approaches to minimize CSC therapy resistance. One proposed mechanism by which CSCs arise is via an epithelial-mesenchymal transition (EMT), and reversal of this process may provide a potential therapeutic approach for increasing tumor chemosensitivity. Therefore, we investigated the role of known EMT regulators, miR-200 family of microRNAs in controlling the epithelial state, stem-like properties, and therapeutic response in an in vivo primary, syngeneic p53null claudin-low tumor model that is normally deficient in miR-200 expression. Using an inducible lentiviral approach, we expressed the miR-200c cluster in this model and found that it changed the epithelial state, and consequently, impeded CSC behavior in these mesenchymal tumors. Moreover, these state changes were accompanied by a decrease in proliferation and an increase in the differentiation status. miR-200c expression also forced a significant reorganization of tumor architecture, affecting important cellular processes involved in cell-cell contact, cell adhesion, and motility. Accordingly, induced miR200c expression significantly enhanced the chemosensitivity and decreased the metastatic potential of this p53null claudin-low tumor model. Collectively, our data suggest that miR-200c expression in claudin-low tumors offers a potential therapeutic application to disrupt the EMT program on multiple fronts in this mesenchymal tumor subtype, by altering tumor growth, chemosensitivity, and metastatic potential in vivo. PMID:25746005

  8. Caloric restriction shortens lifespan through an increase in lipid peroxidation, inflammation and apoptosis in the G93A mouse, an animal model of ALS.

    Directory of Open Access Journals (Sweden)

    Barkha P Patel

    2010-02-01

    Full Text Available Caloric restriction (CR extends lifespan through a reduction in oxidative stress, delays the onset of morbidity and prolongs lifespan. We previously reported that long-term CR hastened clinical onset, disease progression and shortened lifespan, while transiently improving motor performance in G93A mice, a model of amyotrophic lateral sclerosis (ALS that shows increased free radical production. To investigate the long-term CR-induced pathology in G93A mice, we assessed the mitochondrial bioenergetic efficiency and oxidative capacity (CS--citrate synthase content and activity, cytochrome c oxidase--COX activity and protein content of COX subunit-I and IV and UCP3-uncoupling protein 3, oxidative damage (MDA--malondialdehyde and PC--protein carbonyls, antioxidant enzyme capacity (Mn-SOD, Cu/Zn-SOD and catalase, inflammation (TNF-alpha, stress response (Hsp70 and markers of apoptosis (Bax, Bcl-2, caspase 9, cleaved caspase 9 in their skeletal muscle. At age 40 days, G93A mice were divided into two groups: Ad libitum (AL; n = 14; 7 females or CR (n = 13; 6 females, with a diet equal to 60% of AL. COX/CS enzyme activity was lower in CR vs. AL male quadriceps (35%, despite a 2.3-fold higher COX-IV/CS protein content. UCP3 was higher in CR vs. AL females only. MnSOD and Cu/Zn-SOD were higher in CR vs. AL mice and CR vs. AL females. MDA was higher (83% in CR vs. AL red gastrocnemius. Conversely, PC was lower in CR vs. AL red (62% and white (30% gastrocnemius. TNF-alpha was higher (52% in CR vs. AL mice and Hsp70 was lower (62% in CR vs. AL quadriceps. Bax was higher in CR vs. AL mice (41% and CR vs. AL females (52%. Catalase, Bcl-2 and caspases did not differ. We conclude that CR increases lipid peroxidation, inflammation and apoptosis, while decreasing mitochondrial bioenergetic efficiency, protein oxidation and stress response in G93A mice.

  9. Increased Levels of Type 1 Interferon in a Type 1 Diabetic Mouse Model Induce the Elimination of B Cells from the Periphery by Apoptosis and Increase their Retention in the Spleen

    Directory of Open Access Journals (Sweden)

    Badr Mohamed Badr

    2015-01-01

    Full Text Available Background: The autoimmune disease type 1 diabetes mellitus (T1D is associated with a defect in the immune response, which increases susceptibility to infection. We recently demonstrated that prolonged elevated levels of type 1 interferon (IFN induce lymphocyte exhaustion during T1D. Aims: In the present study, we further investigated the effect of blocking the type I IFN receptor signaling pathway on diabetic dyslipidemia, in which an abnormal lipid profile leads to the exhaustion of B cells and alteration of their distribution and functions. Methods: T1D was induced in a mouse model by an intraperitoneal injection of a single dose (60 mg/kg of streptozotocin (STZ. Three groups of mice were examined: a non-diabetic control group, a diabetic group and a diabetic group treated with an anti-IFN (alpha, beta and omega receptor 1 (IFNAR1 blocking antibody to block type I IFN signaling. Results: We observed that induction of T1D was accompanied by a marked destruction of β cells and a reduction in the insulin levels in the diabetic group. Diabetic mice exhibited many changes, including alterations in their lipid profiles, expansion of splenic B cells, increased caspase-3, -8 and -9 activity, and apoptosis in peripheral B cells. Blocking type 1 IFN signaling in diabetic mice significantly returned the insulin and lipid profiles to normal levels, subsequently restored the B cell distribution, and rescued the peripheral B cells from apoptosis. Conclusion: Our data suggest the potential role of type I IFN in mediating diabetic dyslipidemia and an exhausted state of B cells during T1D.

  10. Loss of GCN5 leads to increased neuronal apoptosis by upregulating E2F1- and Egr-1-dependent BH3-only protein Bim.

    Science.gov (United States)

    Wu, Yanna; Ma, Shanshan; Xia, Yong; Lu, Yangpeng; Xiao, Shiyin; Cao, Yali; Zhuang, Sidian; Tan, Xiangpeng; Fu, Qiang; Xie, Longchang; Li, Zhiming; Yuan, Zhongmin

    2017-01-26

    Cellular acetylation homeostasis is a kinetic balance precisely controlled by histone acetyl-transferase (HAT) and histone deacetylase (HDAC) activities. The loss of the counterbalancing function of basal HAT activity alters the precious HAT:HDAC balance towards enhanced histone deacetylation, resulting in a loss of acetylation homeostasis, which is closely associated with neuronal apoptosis. However, the critical HAT member whose activity loss contributes to neuronal apoptosis remains to be identified. In this study, we found that inactivation of GCN5 by either pharmacological inhibitors, such as CPTH2 and MB-3, or by inactivation with siRNAs leads to a typical apoptosis in cultured cerebellar granule neurons. Mechanistically, the BH3-only protein Bim is transcriptionally upregulated by activated Egr-1 and E2F1 and mediates apoptosis following GCN5 inhibition. Furthermore, in the activity withdrawal- or glutamate-evoked neuronal apoptosis models, GCN5 loses its activity, in contrast to Bim induction. Adenovirus-mediated overexpression of GCN5 suppresses Bim induction and apoptosis. Interestingly, the loss of GCN5 activity and the induction of Egr-1, E2F1 and Bim are involved in the early brain injury (EBI) following subarachnoid haemorrhage (SAH) in rats. HDAC inhibition not only significantly rescues Bim expression and apoptosis induced by either potassium deprivation or GCN5 inactivation but also ameliorates these events and EBI in SAH rats. Taken together, our results highlight a new mechanism by which the loss of GCN5 activity promotes neuronal apoptosis through the transcriptional upregulation of Bim, which is probably a critical event in triggering neuronal death when cellular acetylation homeostasis is impaired.

  11. The significance of intestinal apoptosis

    International Nuclear Information System (INIS)

    Potten, C.S.

    1997-01-01

    Apoptosis occurs at a low level spontaneously in the small intestine (SI). The levels can be raised by a variety of cytotoxic agents including radiation. The apoptosis induced by radiation, and some drugs and the spontaneous apoptosis, show some specificity for the stem cells in the small intestinal crypt. In the colon, these agents target transit cells in the mid crypt. p53 expression is elevated at the same time as apoptosis in the SI but not in the cells undergoing apoptosis. The expression of bcl-2, a survival gene, is largely absent in the SI, but is expressed, albeit weakly, in the stem cells in the colon. Spontaneous apoptosis is observed in p53 null mice which also develop normally suggesting that spontaneous and developmental apoptosis are p53 independent and that spontaneous apoptosis is part of the homeostatic mechanisms maintaining stem cell numbers. Radiation induced apoptosis is completely absent at these early times post-irradiation in p53 nulls. In bel-2 null mice, the levels of spontaneous and radiation induced apoptosis are elevated in the colon. Bax, a death gene, is expressed on the villus and inter-crypt table in the colon suggesting that cells at the end of their lifespan initiate apoptosis. It has been suggested that apoptosis in the SI is a protective mechanism against carcinogenesis in the stem cells of the SI which rarely develops cancer. Cells that possess genetic damage detected. In the large bowel, this mechanism is not effective due to the action of bcl-2. Thus stem cells may persist in this tissue with genetic damage resulting in a higher cancer risk. Furthermore, the lack of spontaneous apoptosis in the colon may result in a gradual increase of the stem cells with time resulting in more ells at risk. (author)

  12. Chemosensitization of Prostate Carcinoma Cells with a Receptor-directed Smac Conjugate.

    Science.gov (United States)

    Sturzu, Alexander; Sheikh, Sumbla; Echner, Hartmut; Deeg, Martin; Nägele, Thomas; Weidenmaier, Christopher; Schwentner, Christian; Horger, Marius; Ernemann, Ulrike; Heckl, Stefan

    2016-01-01

    Second mitochondrial activator of caspase (Smac) is a short mitochondrial peptide. When released from the mitochondria into the cytoplasm, it binds to inhibitor of apoptotic proteins (IAPs) within the cytoplasm and prevents them from inhibiting apoptosis. Delivery of external synthetic Smac peptide into the cytoplasm of malignant cells could greatly improve the efficiency of apoptosis-inducing chemotherapeutic agents. In our study different conjugates based on the seven N-terminal amino acids AVPIAQK of Smac (SmacN7) were produced to obtain a cytoplasm-directed Smac variant. SmacN7 and a point mutant (AVPKAQK) were coupled either to rhodamine alone or to both rhodamine and undecylic aldehyde, which is an antagonist of the Lily-of-the-valley fragrance receptor. The fifth conjugate consisted of rhodamine coupled only to undecylic aldehyde, without SmacN7. The uptake of these five conjugates into three different human cell lines was characterized and quantified by confocal laser scanning microscopy and flow cytometry. A caspase apoptosis assay was performed for cells incubated with the five different conjugates after induction of apoptosis. The coupling of undecylic aldehyde to SmacN7 increased the cellular uptake of the correct and mutant conjugates. Caspase 3/7 apoptosis tests after induction of apoptosis with staurosporine or UV irradiation showed that the coupling of SmacN7 with undecylic aldehyde resulted in a greatly increased adjuvant pro-apoptotic effect compared to the separate components and a mutant SmacN7 peptide sequence in the LNCaP prostate carcinoma cells compared to the benign prostate hyperplasia (BPH) cells and the human embryonal kidney (HEK) cells.

  13. Increase of RhoB in {gamma}-radiation-induced apoptosis is regulated by c-Jun N-terminal kinase in Jurkat T cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Chun-Ho [Laboratory of Cytogenetics and Tissue Regeneration, KIRAMS, Seoul 139-706 (Korea, Republic of); Won, Misun; Choi, Chung-Hae; Ahn, Jiwon; Kim, Bo-Kyung [Genome Research Center, KRIBB, Daejeon 305-806 (Korea, Republic of); Song, Kyung-Bin [Department of Food Science and Technology, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Kang, Chang-Mo, E-mail: kangcm@kcch.re.kr [Laboratory of Cytogenetics and Tissue Regeneration, KIRAMS, Seoul 139-706 (Korea, Republic of); Chung, Kyung-Sook, E-mail: kschung@kribb.re.kr [Genome Research Center, KRIBB, Daejeon 305-806 (Korea, Republic of)

    2010-01-08

    The Ras-related small GTP-binding protein RhoB is known to be a pro-apoptotic protein and immediate-early inducible by genotoxic stresses. In addition, JNK activation is known to function in {gamma}-radiation-induced apoptosis. However, it is unclear how JNK activation and {gamma}-radiation-dependent RhoB induction are related. Here we verified the relationship between JNK activation and RhoB induction. RhoB induction by {gamma}-radiation occurred at the transcriptional level and transcriptional activation of RhoB was concomitant with an increase in RhoB protein. {gamma}-Radiation-induced RhoB expression was markedly attenuated by pretreatment with a JNK-specific inhibitor, SP600125, but not by a p38 MAPK inhibitor, SB203580. Inhibition of JNK caused a decrease in early apoptotic cell death that correlated with RhoB expression. However, PI3K inhibition had no significant effects, indicating that the AKT survival pathway was not involved. The siRNA knockdown of JNK resulted in a decrease in RhoB expression and the siRNA knockdown of RhoB restored cell growth even in the {gamma}-irradiated cells. These results suggest that RhoB regulation involves the JNK pathway and contributes to the early apoptotic response of Jurkat T cells to {gamma}-radiation.

  14. Polycystic Ovary Syndrome and Increased Soluble Tumor Necrosis Factor Like Weak Inducer of Apoptosis Levels Are Independent Predictors of Dyslipidemia in Youth.

    Science.gov (United States)

    Erkenekli, Kudret; Oztas, Efser; Kuscu, Elif; Keskin, Uğur; Kurt, Yasemin Gulcan; Tas, Ahmet; Yilmaz, Nafiye

    2017-01-01

    Dyslipidemia is common in women with polycystic ovary syndrome (PCOS) irrespective of age. Our aim was to investigate soluble tumor necrosis factor like weak inducer of apoptosis (sTWEAK), a cardiovascular risk marker in PCOS, and to determine if it is associated with dyslipidemia in youth. A prospective-observational study was carried out including 35 PCOS patients and 35 healthy controls. Serum sTWEAK levels were measured using commercially available kits. Multiple logistic regression analysis was then performed to verify the statistically significant differences in the possible predictors of dyslipidemia. Serum sTWEAK levels and the percentage of women with dyslipidemia were significantly higher in the PCOS group (p = 0.024 and p dyslipidemia. The percentage of women with PCOS was significantly higher in the dyslipidemic group when compared with controls; 70.7 vs. 20.7%, respectively (p 693 pg/ml; OR 3.810, 95% CI 1.075-13.501, p = 0.038) were independently associated with dyslipidemia. Increased levels of both sTWEAK and PCOS were found to be independently associated with dyslipidemia in youth. © 2016 S. Karger AG, Basel.

  15. Adrenalectomy promotes a permanent decrease of plasma corticoid levels and a transient increase of apoptosis and the expression of Transforming Growth Factor β1 (TGF-β1 in hippocampus: effect of a TGF-β1 oligo-antisense

    Directory of Open Access Journals (Sweden)

    Lara Hernán E

    2006-05-01

    Full Text Available Abstract Background Corticosterone reduction produced by adrenalectomy (ADX induces apoptosis in dentate gyrus (DG of the hippocampus, an effect related to an increase in the expression of the pro-apoptotic gene bax. However it has been reported that there is also an increase of the anti-apoptotic gene bcl-2, suggesting the promotion of a neuroprotective phenomenon, perhaps related to the expression of transforming growth factor β1 (TGF-β1. Thus, we have investigated whether TGF-β1 levels are induced by ADX, and whether apoptosis is increased by blocking the expression of TGF-β1 with an antisense oligonucleotide (ASO administered intracerebrally in corticosterone depleted rats. Results It was observed an increase of apoptosis in DG, 2 and 5 days after ADX, in agreement with a reduction of corticosterone levels. However, the effect of ADX on the number of apoptotic positive cells in DG was decreased 5 days after the lesion. In CA1–CA3 regions, the effect was only observed 2 days after ADX. TGF-β1 mRNA levels were increased 2 days after ADX. The sustained intracerebro-ventricular administration of a TGF-β1 ASO via an osmotic mini pump increased apoptosis levels in CA and DG regions 5 days after ADX as well as sham-operated control animals. No significant effect was observed following a scrambled-oligodeoxynucleotide treatment. Conclusion The changes in both the pattern and the magnitude of apoptotic-cell morphology observed 2 and 5 days after ADX suggest that, as a consequence of the reduction of corticosteroids, some trophic mechanisms restricting cell death to a particular time window are elicited. Sustained intracerebral administration of TGF-β1 ASO increased the apoptosis promoted by ADX, suggesting that TGF-β1 plays an anti-apoptotic role in vivo in hippocampus.

  16. Cytotoxic hydrogen bridged ruthenium quinaldamide complexes showing induced cancer cell death by apoptosis.

    Science.gov (United States)

    Lord, Rianne M; Allison, Simon J; Rafferty, Karen; Ghandhi, Laura; Pask, Christopher M; McGowan, Patrick C

    2016-08-16

    This report presents the first known p-cymene ruthenium quinaldamide complexes which are stabilised by a hydrogen-bridging atom, [{(p-cym)Ru(II)X(N,N)}{H(+)}{(N,N)XRu(II)(p-cym)}][PF6] (N,N = functionalised quinaldamide and X = Cl or Br). These complexes are formed by a reaction of [p-cymRu(μ-X)2]2 with a functionalised quinaldamide ligand. When filtered over NH4PF6, and under aerobic conditions the equilibrium of NH4PF6 ⇔ NH3 + HPF6 enables incorporation of HPF6 and the stabilisation of two monomeric ruthenium complexes by a bridging H(+), which are counter-balanced by a PF6 counterion. X-ray crystallographic analysis is presented for six new structures with OO distances of 2.420(4)-2.448(15) Å, which is significant for strong hydrogen bonds. Chemosensitivity studies against HCT116, A2780 and cisplatin-resistant A2780cis human cancer cells showed the ruthenium complexes with a bromide ancillary ligand to be more potent than those with a chloride ligand. The 4'-fluoro compounds show a reduction in potency for both chloride and bromide complexes against all cell lines, but an increase in selectivity towards cancer cells compared to non-cancer ARPE-19 cells, with a selectivity index >1. Mechanistic studies showed a clear correlation between IC50 values and induction of cell death by apoptosis.

  17. Central respiratory chemosensitivity and cerebrovascular CO2 reactivity: a rebreathing demonstration illustrating integrative human physiology.

    Science.gov (United States)

    MacKay, Christina M; Skow, Rachel J; Tymko, Michael M; Boulet, Lindsey M; Davenport, Margie H; Steinback, Craig D; Ainslie, Philip N; Lemieux, Chantelle C M; Day, Trevor A

    2016-03-01

    One of the most effective ways of engaging students of physiology and medicine is through laboratory demonstrations and case studies that combine 1) the use of equipment, 2) problem solving, 3) visual representations, and 4) manipulation and interpretation of data. Depending on the measurements made and the type of test, laboratory demonstrations have the added benefit of being able to show multiple organ system integration. Many research techniques can also serve as effective demonstrations of integrative human physiology. The "Duffin" hyperoxic rebreathing test is often used in research settings as a test of central respiratory chemosensitivity and cerebrovascular reactivity to CO2. We aimed to demonstrate the utility of the hyperoxic rebreathing test for both respiratory and cerebrovascular responses to increases in CO2 and illustrate the integration of the respiratory and cerebrovascular systems. In the present article, methods such as spirometry, respiratory gas analysis, and transcranial Doppler ultrasound are described, and raw data traces can be adopted for discussion in a tutorial setting. If educators have these instruments available, instructions on how to carry out the test are provided so students can collect their own data. In either case, data analysis and quantification are discussed, including principles of linear regression, calculation of slope, the coefficient of determination (R(2)), and differences between plotting absolute versus normalized data. Using the hyperoxic rebreathing test as a demonstration of the complex interaction and integration between the respiratory and cerebrovascular systems provides senior undergraduate, graduate, and medical students with an advanced understanding of the integrative nature of human physiology. Copyright © 2016 The American Physiological Society.

  18. Disulfiram suppresses growth of the malignant pleural mesothelioma cells in part by inducing apoptosis.

    Directory of Open Access Journals (Sweden)

    Vino T Cheriyan

    Full Text Available Dithiocarbamate compound Disulfiram (DSF that binds with copper and functions as an inhibitor of aldehyde dehydrogenase is a Food and Drug Administration approved agent for treatment of alcoholism. Copper complexed DSF (DSF-Cu also possesses anti-tumor and chemosensitizing properties; however, its molecular mechanisms of action remain unclear. Here we investigated malignant pleural mesothelioma (MPM suppressive effects of DSF-Cu and the molecular mechanisms involved. DSF-Cu inhibited growth of the murine as well as human MPM cells in part by increasing levels of ubiquitinated proteins. DSF-Cu exposure stimulated apoptosis in MPM cells that involved activation of stress-activated protein kinases (SAPKs p38 and JNK1/2, caspase-3, and cleavage of poly-(ADP-ribose-polymerase, as well as increased expression of sulfatase 1 and apoptosis transducing CARP-1/CCAR1 protein. Gene-array based analyses revealed that DSF-Cu suppressed cell growth and metastasis-promoting genes including matrix metallopeptidase 3 and 10. DSF inhibited MPM cell growth and survival by upregulating cell cycle inhibitor p27Kip1, IGFBP7, and inhibitors of NF-κB such as ABIN 1 and 2 and Inhibitory κB (IκBα and β proteins. DSF-Cu promoted cleavage of vimentin, as well as serine-phosphorylation and lysine-63 linked ubiquitination of podoplanin. Administration of 50 mg/kg DSF-Cu by daily i.p injections inhibited growth of murine MPM cell-derived tumors in vivo. Although podoplanin expression often correlates with metastatic disease and poor prognosis, phosphorylation of serines in cytoplasmic domain of podoplanin has recently been shown to interfere with cellular motility and migration signaling. Post-translational modification of podoplanin and cleavage of vimentin by DSF-Cu underscore a metastasis inhibitory property of this agent and together with our in vivo studies underscore its potential as an anti-MPM agent.

  19. Arctigenin enhances chemosensitivity to cisplatin in human nonsmall lung cancer H460 cells through downregulation of survivin expression.

    Science.gov (United States)

    Wang, Huan-qin; Jin, Jian-jun; Wang, Jing

    2014-01-01

    Arctigenin, a dibenzylbutyrolactone lignan, enhances cisplatin-mediated cell apoptosis in cancer cells. Here, we sought to investigate the effects of arctigenin on cisplatin-treated non-small-cell lung cancer (NSCLC) H460 cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and annexin-V/propidium iodide staining were performed to analyze the proliferation and apoptosis of H460 cells. Arctigenin dose-dependently suppressed cell proliferation and potentiated cell apoptosis, coupled with increased cleavage of caspase-3 and poly(ADP-ribose) polymerase. Moreover, arctigenin sensitized H460 cells to cisplatin-induced proliferation inhibition and apoptosis. Arctigenin alone or in combination with cisplatin had a significantly lower amount of survivin. Ectopic expression of survivin decreased cell apoptosis induced by arctigenin (P arctigenin (P arctigenin has a therapeutic potential in combina-tion with chemotherapeutic agents for NSLC. © 2013 Wiley Periodicals, Inc.

  20. Inhibition of the epidermal growth factor receptor in bladder cancer cells treated with the DNA-damaging drug etoposide markedly increases apoptosis

    DEFF Research Database (Denmark)

    Munk, Mathias; Memon, Ashfaque Ahmed; Nexo, Ebba

    2007-01-01

    : The bladder cancer cell lines RT4 and T24, representing low- and high-malignancy grades respectively, were treated with VP16 (10 or 50 microM) and the level of apoptosis determined using a commercial kit. EGFR receptor activity was determined by western blotting using antibodies against phosphorylated EGFR...... of apoptotic cells with VP16 alone. This was found in both T24 and RT4 cells. Gefitinib used alone (1 and 5 microM) generated no apoptosis in the cells. Treatment of T24 cells with Z-VAD showed that apoptosis induced by both VP16 alone and VP16 with gefitinib was caspase-mediated. CONCLUSION: These results...

  1. Conditional Macrophage Depletion Increases Inflammation and Does Not Inhibit the Development of Osteoarthritis in Obese Macrophage Fas-Induced Apoptosis-Transgenic Mice.

    Science.gov (United States)

    Wu, Chia-Lung; McNeill, Jenna; Goon, Kelsey; Little, Dianne; Kimmerling, Kelly; Huebner, Janet; Kraus, Virginia; Guilak, Farshid

    2017-09-01

    To investigate whether short-term, systemic depletion of macrophages can mitigate osteoarthritis (OA) following injury in the setting of obesity. CSF-1R-GFP+ macrophage Fas-induced apoptosis (MaFIA)-transgenic mice that allow conditional depletion of macrophages were placed on a high-fat diet and underwent surgery to induce knee OA. A small molecule (AP20187) was administrated to deplete macrophages in MaFIA mice. The effects of macrophage depletion on acute joint inflammation, OA severity, and arthritic bone changes were evaluated using histology and micro-computed tomography. Immunohistochemical analysis was performed to identify various immune cells. The levels of serum and synovial fluid cytokines were also measured. Macrophage-depleted mice had significantly fewer M1 and M2 macrophages in the surgically operated joints relative to controls and exhibited decreased osteophyte formation immediately following depletion. Surprisingly, macrophage depletion did not attenuate the severity of OA in obese mice; instead, it induced systemic inflammation and led to a massive infiltration of CD3+ T cells and particularly neutrophils, but not B cells, into the injured joints. Macrophage-depleted mice also demonstrated a markedly increased number of proinflammatory cytokines including granulocyte colony-stimulating factor, interleukin-1β (IL-1β), IL-6, IL-8, and tumor necrosis factor in both serum and joint synovial fluid, although the mice showed a trend toward decreased levels of insulin and leptin in serum after macrophage depletion. Our findings indicate that macrophages are vital for modulating homeostasis of immune cells in the setting of obesity and suggest that more targeted approaches of depleting specific macrophage subtypes may be necessary to mitigate inflammation and OA in the setting of obesity. © 2017, American College of Rheumatology.

  2. Light-Emitting Diode (LED) therapy improves occipital cortex damage by decreasing apoptosis and increasing BDNF-expressing cells in methanol-induced toxicity in rats.

    Science.gov (United States)

    Ghanbari, Amir; Ghareghani, Majid; Zibara, Kazem; Delaviz, Hamdallah; Ebadi, Elham; Jahantab, Mohammad Hossein

    2017-05-01

    Methanol-induced retinal toxicity, frequently associated with elevated free radicals and cell edema, is characterized by progressive retinal ganglion cell (RGC) death and vision loss. Previous studies investigated the effect of photomodulation on RGCs, but not the visual cortex. In this study, the effect of 670nm Light-Emitting Diode (LED) therapy on RGCs and visual cortex recovery was investigated in a seven-day methanol-induced retinal toxicity protocol in rats. Methanol administration showed a reduction in the number of RGCs, loss of neurons (neuronal nuclear antigen, NeuN+), activation of glial fibrillary acidic protein (GFAP+) expressing cells, suppression of brain-derived neurotrophic factor (BDNF+) positive cells, increase in apoptosis (caspase 3+) and enhancement of nitric oxide (NO) release in serum and brain. On the other hand, LED therapy significantly reduced RGC death, in comparison to the methanol group. In addition, the number of BDNF positive cells was significantly higher in the visual cortex of LED-treated group, in comparison to methanol-intoxicated and control groups. Moreover, LED therapy caused a significant decrease in cell death (caspase 3+ cells) and a significant reduction in the NO levels, both in serum and brain tissue, in comparison to methanol-intoxicated rats. Overall, LED therapy demonstrated a number of beneficial effects in decreasing oxidative stress and in functional recovery of RGCs and visual cortex. Our data suggest that LED therapy could be a potential condidate as a non-invasive approach for treatment of retinal damage, which needs further clinicl studies. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  3. Chronic Cigarette Smoking Impairs Erectile Function through Increased Oxidative Stress and Apoptosis, Decreased nNOS, Endothelial and Smooth Muscle Contents in a Rat Model.

    Directory of Open Access Journals (Sweden)

    Yun-Ching Huang

    Full Text Available Cigarette use is an independent risk factor for the development of erectile dysfunction (ED. While the association between chronic smoking and ED is well established, the fundamental mechanism(s of cigarette-related ED are incompletely understood, partly due to no reliable animal model of smoking-induced ED. The present study was designed to validate an in vivo rat model of chronic cigarette-induced ED. Forty 12-week old male Sprague-Dawley rats were divided into 4 groups. Ten rats served as control group and were exposed only to room air. The remaining 30 rats were passively exposed to cigarette smoke (CS for 4 weeks (n = 10, 12 weeks (n = 10, and 24 weeks (n = 10. At the 24-week time point all rats were assessed with intracavernous pressure (ICP during cavernous nerve electrostimulation. Blood and urine were collected to measure serum testosterone and oxidative stress, respectively. Corporal tissue was assessed by Western blot for neuronal nitric oxide synthase (nNOS. Penile tissues were subjected to immunohistochemistry for endothelial, smooth muscle, and apoptotic content. Mean arterial pressure (MAP was significantly higher in 24-week cigarette exposed animals compared to the control animals. Mean ICP/MAP ratio and cavernosal smooth muscle/endothelial contents were significantly lower in the 12- and 24-week rats compared to control animals. Oxidative stress was significantly higher in the 24-week cigarette exposed group compared to control animals. Mean nNOS expression was significantly lower, and apoptotic index significantly higher, in CS-exposed animals compared to control animals. These findings indicate that the rat model exposure to CS increases apoptosis and oxidative stress and decreases nNOS, endothelial and smooth muscle contents, and ICP in a dose dependent fashion. The rat model is a useful tool for further study of the molecular and cellular mechanisms of CS-related ED.

  4. Influence of p53 and bcl-2 on chemosensitivity in benign and malignant prostatic cell lines.

    Science.gov (United States)

    Serafin, Antonio M; Bohm, Lothar

    2005-01-01

    The administration of cancer chemotherapeutic agents results in an increase in the apoptotic cells in the tumor: therefore, it has been assumed that anticancer drugs exhibit their cytotoxic effects via apoptotic signaling pathways. Characteristics that confer sensitivity to drug-induced apoptosis are, a functional p53 protein and expression of the apoptosis-promoting protein, bax. The role of p53 and bax/bcl-2 in drug-induced apoptosis was assessed in six prostate cell lines, 1532T, 1535T, 1542T, 1542N, BPH-1 and LNCaP using TD(50) concentrations of etoposide, vinblastine and estramustine. Cell death was monitored morphologically by fluorescent microscopy, and by flow cytometry (Annexin-V assay). Apoptotic morphology was rather low and ranged from 0.1% to 12.1%, 3.0% to 6.0% and 0.1% to 8.5% for etoposide, estramustine and vinblastine, respectively. Annexin-V binding and flow cytometry indicated apoptotic propensities of 0% to 4%, 0% to 3% and 0% to 5%, respectively. The percentage of cells responding to drug-induced apoptosis was, on average, higher in the tumor cell lines than in the normal cell lines, but showed no correlation with p53 status. The percentage of cells showing necrosis, assessed by Annexin binding and Propidium Iodide permeability in aqueous medium, tended to be much higher, and was found to be at the level of 5% to 30%. Immunoblotting demonstrated that bax and bcl-2 proteins were expressed at a basal level in all cell lines, but did not increase after exposure to TD(50) doses of the three drugs. The ratio of bax and bcl-2, measured by laser scanning densitometry, was not altered by the drug-induced DNA damage. The results suggest that apoptosis is not a major mechanism of drug-induced cell death in prostate cell lines and appears to be independent of p53 status and bax/bcl-2 expression.

  5. ABC transporters as multidrug resistance mechanisms and the development of chemosensitizers for their reversal

    Directory of Open Access Journals (Sweden)

    Choi Cheol-Hee

    2005-10-01

    Full Text Available Abstract One of the major problems related with anticancer chemotherapy is resistance against anticancer drugs. The ATP-binding cassette (ABC transporters are a family of transporter proteins that are responsible for drug resistance and a low bioavailability of drugs by pumping a variety of drugs out cells at the expense of ATP hydrolysis. One strategy for reversal of the resistance of tumor cells expressing ABC transporters is combined use of anticancer drugs with chemosensitizers. In this review, the physiological functions and structures of ABC transporters, and the development of chemosensitizers are described focusing on well-known proteins including P-glycoprotein, multidrug resistance associated protein, and breast cancer resistance protein.

  6. Chemosensitivity testing of primary human renal cell carcinoma by a tetrazolium based microculture assay (MTT).

    Science.gov (United States)

    Mickisch, G; Fajta, S; Keilhauer, G; Schlick, E; Tschada, R; Alken, P

    1990-01-01

    MTT staining procedures have been used in chemosensitivity testing of established cell lines of human and other sources as well as of human leukaemias, but only limited information on its application in primary solid human tumors is presently available. We have evaluated MTT staining in primary human Renal Cell Carcinomas (RCCs), studied various factors interfering with the optimal use, and finally applied it in subsequent chemosensitivity testing. The method depends on the conversion of a water-soluble tetrazolium salt (MTT) to a purple colored formazan precipitate, a reaction effected by enzymes active only in living cells. Single cell suspensions of RCCs were obtained either by enzymatic dispersion or by mechanical dissagregation, filtered through gauze, and purified by Ficoll density centrifugation. Tests were carried out in 96-well microculture plates. 10(4) viable tumor cells per well at 4 h incubation time with 20 micrograms MTT/100 microliters total medium volume yielded best results. Formazan crystals were dissolved with DMSO, and the plates were immediately measured on a microculture plate reader at 540 nm. Under these criteria, linearity of the system could be demonstrated. For chemosensitivity testing, cells were continuously exposed to a number of drugs prior to the MTT staining procedure. Reproducibility of results was assessed and confirmed by culturing RCCs in flasks additionally, resubmitting them after 1, 2, and 4 weeks to the MTT assay. We conclude that the semiautomated MTT assay offers a valid, rapid, reliable and simple method to determine the degree of chemoresistance in primary human RCCs.

  7. Selected aspects of apoptosis in psoriasis

    Directory of Open Access Journals (Sweden)

    Hanna Myśliwiec

    2017-03-01

    Full Text Available Apoptosis is a physiological mechanism of programmed cell death, in contrast to necrosis, without eliciting an inflammatory response. It plays the key role in the functioning of different tissues and organs. The balance between proliferation and apoptosis of keratinocytes is important for epidermal maintenance of homeostasis. Dysfunctional apoptosis plays an important role in the development of several skin disorders. Diseases with an increase of apoptosis are usually acute, while those with inhibited apoptosis tend to be chronic. Psoriasis is an immune-mediated chronic inflammatory skin disease. It is characterized by keratinocyte hyperproliferation and abnormal differentiation. Psoriatic keratinocytes are resistant to apoptosis and this phenomenon can be the key event in psoriatic hyperplasia. Apoptosis disturbances can also affect immune cells involved in the pathogenesis of psoriasis. In this review, we describe the basic concept of apoptosis and its relevance in psoriatic pathogenesis.

  8. Evaluation of the in vitro Chemosensitivity and Correlation with Clinical Outcomes in Lung Cancer using the ATP-TCA.

    Science.gov (United States)

    Chen, Zhiyao; Zhang, Shichao; Ma, Sheng; Li, Chang; Xu, Chun; Shen, Yinfang; Zhao, Jun; Miao, Liyan

    2018-01-01

    Multiple drug resistance (MDR) to chemotherapeutic agents often leads to a failure to respond to chemotherapy. We utilized an in vitro chemosensitivity test to identify sensitive and effective chemotherapeutic drugs and further elucidated the correlation between the in vivo chemosensitivity and clinical outcomes. Here, we evaluated the in vitro chemosensitivity and MDR of 120 lung cancer patients to eight singledrug chemotherapies and of 291 lung cancer patients to seven chemotherapy regimens using an ATP-based tumor chemosensitivity assay (ATP-TCA). Additionally, the chemosensitivity profiles of lung adenocarcinoma patients (284 cases) and lung squamous cell carcinoma patients (90 cases) to these single-drug and chemotherapy regimens were compared. Furthermore, the correlations between the chemosensitivity and clinical outcomes were investigated in 16 stage III squamous cell carcinoma patients. PTX (51.7%), TXT (43.3%), GEM (12.5%), PTX+DDP (62.5%), TXT+L-OHP (54.3%) and VP-16+DDP (16.2%) had the highest in vitro chemosensitivity rates. Approximately 31.7% of patients developed resistance to all eight single-drug chemotherapies, and 25.8% of patients displayed resistance to all seven chemotherapy regimens. In addition, lung squamous cell carcinoma was significantly more sensitive to GEM and MTA+DDP than lung adenocarcinoma (P<0.05). Further analysis showed that patients with higher drug sensitivity tended to have longer disease-free survival (18 months vs. 8.5 months) than patients displaying drug resistance (P<0.05). These results suggest that the implementation of in vitro drug susceptibility testing before chemotherapy can effectively prevent the occurrence of primary drug resistance and inappropriate drug treatment. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  9. 4-Acetyl-12,13-epoxyl-9-trichothecene-3, 15-diol from Isaria japonica mediates apoptosis of rat bladder carcinoma NBT-II cells by decreasing anti-apoptotic Bcl-2 expression and increasing pro-apoptotic Bax expression.

    Science.gov (United States)

    Kim, Hyung-Jin; Jang, Seon Il; Kim, Young-Jun; Pae, Hyun-Ock; Won, Hae-Young; Hong, Kyung-Hwan; Oh, Hyuncheol; Kwon, Tae-Oh; Chung, Hun-Taeg

    2004-01-01

    We studied the effect of 4-acetyl-12,13-epoxyl-9-trichothecene-3, 15-diol (AETD) isolated from Isaria japonica, one of the most popular Chinese fungal medicines, on the induction of apoptosis in rat bladder carcinoma NBT-II cells. AETD was cytotoxic to NBT-II cells, and this cytotoxic effect appears to be attributed to its induction of apoptotic cell death, as AETD induced nuclear morphological changes and internucleosomal DNA fragmentation, and increased the proportion of hypodiploid cells and activity of caspase-3. AETD treatment also decreased the expression of the anti-apoptotic protein Bcl-2 and increased the expression of the pro-apoptotic protein Bax. These results provide important information in understanding the mechanism(s) of AETD-induced apoptosis.

  10. Serum levels of pancreatic stone protein (PSP)/reg1A as an indicator of beta-cell apoptosis suggest an increased apoptosis rate in hepatocyte nuclear factor 1 alpha (HNF1A-MODY) carriers from the third decade of life onward

    LENUS (Irish Health Repository)

    Bacon, Siobhan

    2012-07-18

    AbstractBackgroundMutations in the transcription factor hepatocyte nuclear factor-1-alpha (HNF1A) result in the commonest type of maturity onset diabetes of the young (MODY). HNF1A-MODY carriers have reduced pancreatic beta cell mass, partially due to an increased rate of apoptosis. To date, it has not been possible to determine when apoptosis is occurring in HNF1A-MODY.We have recently demonstrated that beta cell apoptosis stimulates the expression of the pancreatic stone protein\\/regenerating (PSP\\/reg) gene in surviving neighbour cells, and that PSP\\/reg1A protein is subsequently secreted from these cells. The objective of this study was to determine whether serum levels of PSP\\/reg1A are elevated during disease progression in HNF1A-MODY carriers, and whether it may provide information regarding the onset of beta-cell apoptosis.MethodsWe analysed serum PSP\\/reg1A levels and correlated with clinical and biochemical parameters in subjects with HNF1A-MODY, glucokinase (GCK-MODY), and type 1 diabetes mellitus. A control group of normoglycaemic subjects was also analysed.ResultsPSP\\/reg1A serum levels were significantly elevated in HNF1A-MODY (n = 37) subjects compared to controls (n = 60) (median = 12.50 ng\\/ml, IQR = 10.61-17.87 ng\\/ml versus median = 10.72 ng\\/ml, IQR = 8.94-12.54 ng\\/ml, p = 0.0008). PSP\\/reg1A correlated negatively with insulin levels during OGTT, (rho = −0.40, p = 0.02). Interestingly we noted a significant positive correlation of PSP\\/reg1A with age of the HNF1A-MODY carriers (rho = 0.40 p = 0.02) with an age of 25 years separating carriers with low and high PSP\\/reg1A levels. Patients with type 1 diabetes mellitus also had elevated serum levels of PSP\\/reg1A compared to controls, however this was independent of the duration of diabetes.ConclusionOur data suggest that beta cell apoptosis contributes increasingly to the pathophysiology of HNF1A-MODY in patients 25 years and

  11. Predictors of Chemosensitivity in Triple Negative Breast Cancer: An Integrated Genomic Analysis.

    Directory of Open Access Journals (Sweden)

    Tingting Jiang

    2016-12-01

    .The comprehensive molecular analysis presented in this study directly links BRCA deficiency with increased clonal mutation burden and significantly enhanced chemosensitivity in TNBC and suggests that functional RNA-based BRCA deficiency needs to be further examined in TNBC.

  12. Tubulin binding cofactor C (TBCC) suppresses tumor growth and enhances chemosensitivity in human breast cancer cells

    International Nuclear Information System (INIS)

    Hage-Sleiman, Rouba; Herveau, Stéphanie; Matera, Eva-Laure; Laurier, Jean-Fabien; Dumontet, Charles

    2010-01-01

    Microtubules are considered major therapeutic targets in patients with breast cancer. In spite of their essential role in biological functions including cell motility, cell division and intracellular transport, microtubules have not yet been considered as critical actors influencing tumor cell aggressivity. To evaluate the impact of microtubule mass and dynamics on the phenotype and sensitivity of breast cancer cells, we have targeted tubulin binding cofactor C (TBCC), a crucial protein for the proper folding of α and β tubulins into polymerization-competent tubulin heterodimers. We developed variants of human breast cancer cells with increased content of TBCC. Analysis of proliferation, cell cycle distribution and mitotic durations were assayed to investigate the influence of TBCC on the cell phenotype. In vivo growth of tumors was monitored in mice xenografted with breast cancer cells. The microtubule dynamics and the different fractions of tubulins were studied by time-lapse microscopy and lysate fractionation, respectively. In vitro sensitivity to antimicrotubule agents was studied by flow cytometry. In vivo chemosensitivity was assayed by treatment of mice implanted with tumor cells. TBCC overexpression influenced tubulin fraction distribution, with higher content of nonpolymerizable tubulins and lower content of polymerizable dimers and microtubules. Microtubule dynamicity was reduced in cells overexpressing TBCC. Cell cycle distribution was altered in cells containing larger amounts of TBCC with higher percentage of cells in G2-M phase and lower percentage in S-phase, along with slower passage into mitosis. While increased content of TBCC had little effect on cell proliferation in vitro, we observed a significant delay in tumor growth with respect to controls when TBCC overexpressing cells were implanted as xenografts in vivo. TBCC overexpressing variants displayed enhanced sensitivity to antimicrotubule agents both in vitro and in xenografts. These

  13. Differential effects of arsenic trioxide on chemosensitization in human hepatic tumor and stellate cell lines

    International Nuclear Information System (INIS)

    Rangwala, Fatima; Williams, Kevin P; Smith, Ginger R; Thomas, Zainab; Allensworth, Jennifer L; Lyerly, H Kim; Diehl, Anna Mae; Morse, Michael A; Devi, Gayathri R

    2012-01-01

    Crosstalk between malignant hepatocytes and the surrounding peritumoral stroma is a key modulator of hepatocarcinogenesis and therapeutic resistance. To examine the chemotherapy resistance of these two cellular compartments in vitro, we evaluated a well-established hepatic tumor cell line, HepG2, and an adult hepatic stellate cell line, LX2. The aim was to compare the chemosensitization potential of arsenic trioxide (ATO) in combination with sorafenib or fluorouracil (5-FU), in both hepatic tumor cells and stromal cells. Cytotoxicity of ATO, 5-FU, and sorafenib, alone and in combination against HepG2 cells and LX2 cells was measured by an automated high throughput cell-based proliferation assay. Changes in survival and apoptotic signaling pathways were analyzed by flow cytometry and western blot. Gene expression of the 5-FU metabolic enzyme, thymidylate synthase, was analyzed by real time PCR. Both HepG2 and LX2 cell lines were susceptible to single agent sorafenib and ATO at 24 hr (ATO IC 50 : 5.3 μM in LX2; 32.7 μM in HepG2; Sorafenib IC 50 : 11.8 μM in LX2; 9.9 μM in HepG2). In contrast, 5-FU cytotoxicity required higher concentrations and prolonged (48–72 hr) drug exposure. Concurrent ATO and 5-FU treatment of HepG2 cells was synergistic, leading to increased cytotoxicity due in part to modulation of thymidylate synthase levels by ATO. Concurrent ATO and sorafenib treatment showed a trend towards increased HepG2 cytotoxicity, possibly due to a significant decrease in MAPK activation in comparison to treatment with ATO alone. ATO differentially sensitizes hepatic tumor cells and adult hepatic stellate cells to 5-FU and sorafenib. Given the importance of both of these cell types in hepatocarcinogenesis, these data have implications for the rational development of anti-cancer therapy combinations for the treatment of hepatocellular carcinoma (HCC)

  14. Differential effects of arsenic trioxide on chemosensitization in human hepatic tumor and stellate cell lines

    Directory of Open Access Journals (Sweden)

    Rangwala Fatima

    2012-09-01

    Full Text Available Abstract Background Crosstalk between malignant hepatocytes and the surrounding peritumoral stroma is a key modulator of hepatocarcinogenesis and therapeutic resistance. To examine the chemotherapy resistance of these two cellular compartments in vitro, we evaluated a well-established hepatic tumor cell line, HepG2, and an adult hepatic stellate cell line, LX2. The aim was to compare the chemosensitization potential of arsenic trioxide (ATO in combination with sorafenib or fluorouracil (5-FU, in both hepatic tumor cells and stromal cells. Methods Cytotoxicity of ATO, 5-FU, and sorafenib, alone and in combination against HepG2 cells and LX2 cells was measured by an automated high throughput cell-based proliferation assay. Changes in survival and apoptotic signaling pathways were analyzed by flow cytometry and western blot. Gene expression of the 5-FU metabolic enzyme, thymidylate synthase, was analyzed by real time PCR. Results Both HepG2 and LX2 cell lines were susceptible to single agent sorafenib and ATO at 24 hr (ATO IC50: 5.3 μM in LX2; 32.7 μM in HepG2; Sorafenib IC50: 11.8 μM in LX2; 9.9 μM in HepG2. In contrast, 5-FU cytotoxicity required higher concentrations and prolonged (48–72 hr drug exposure. Concurrent ATO and 5-FU treatment of HepG2 cells was synergistic, leading to increased cytotoxicity due in part to modulation of thymidylate synthase levels by ATO. Concurrent ATO and sorafenib treatment showed a trend towards increased HepG2 cytotoxicity, possibly due to a significant decrease in MAPK activation in comparison to treatment with ATO alone. Conclusions ATO differentially sensitizes hepatic tumor cells and adult hepatic stellate cells to 5-FU and sorafenib. Given the importance of both of these cell types in hepatocarcinogenesis, these data have implications for the rational development of anti-cancer therapy combinations for the treatment of hepatocellular carcinoma (HCC.

  15. [Forkhead domain inhibitor-6 (FDI-6) increases apoptosis and inhibits invasion and migration of laryngeal carcinoma cells by down-regulating nuclear FoxM1].

    Science.gov (United States)

    Liu, Yanan; Zhu, Lin; Wen, Taoyu; Wan, Jie; Lei, Yue; Chen, Hongyan

    2017-05-01

    Objective To study the effects of new small molecular inhibitor, forkhead domain inhibitor-6 (FDI-6), on proliferation, apoptosis, invasion and migration in human laryngeal carcinoma Hep-2 cell line and the related mechanism. Methods MTT assay was used to test the proliferation rate of Hep-2 cells before and 12, 24 hours after treated with (5, 10, 20, 40, 80) μmol/L of FDI-6. Flow cytometry (FCM) and Transwell TM chamber assay were respectively carried out to detect the apoptosis rate, cell invasion and migration in Hep-2 cells after treated by 10, 20 μmol/L FDI-6 for 24 hours. Real-time quantitative PCR (qRT-PCR) and Western blotting were performed to determine the mRNA and protein levels of FoxM1, Bcl-2 and BAX, respectively. Results Cell proliferation rate was inhibited by FDI-6 in a dose- and time-dependent manner. Twenty-four hours after 10, 20 μmol/L FDI-6 treatment, the apoptosis rate in Hep-2 cells was elevated and the ability of cell invasion and migration was inhibited in a dose-dependent manner. The qRT-PCR showed that there was no significant change in FoxM1 mRNA expression with or without FDI-6 treatment. Western blotting showed that the total protein level of FoxM1 was not obviously changed, but Bcl-2 was down-regulated, BAX was up-regulated. However, in the nuclear FoxM1 protein level decreased along with the ascent of FDI-6 concentration. Conclusion FDI-6 could induce cell apoptosis and inhibit cell proliferation, invasion and migration in Hep-2 cells. This may be related to the down-regulation of FoxM1 in the nucleus.

  16. Pre-Treatment with Either L-Carnitine or Piracetam Increases Ultrasound-Mediated Gene Transfection by Reducing Sonoporation-Associated Apoptosis.

    Science.gov (United States)

    Liao, Wei-Hao; Wu, Chueh-Hung; Chen, Wen-Shiang

    2018-03-14

    Sonoporation, the use of ultrasound to alter the permeability of cell membranes, is a non-viral technique used to facilitate gene delivery, possibly by opening transient pores in the cell membrane. However, sonoporation may have negative bio-effects on cells, such as causing apoptosis, which limits its efficacy in gene delivery. In this study, we investigated whether pre-treatment with either L-carnitine or piracetam could protect cells from undergoing apoptosis after sonoporation and the possible mechanisms. We found that either L-carnitine or piracetam can promote gene transfection without reducing cell viability, possibly by reducing cavitation-induced reactive oxygen species generation, reversing alterations of mitochondrial membrane potential, preventing caspase-3/7 activity and facilitating mitochondrial ATP production. In conclusion, pre-treatment with either L-carnitine or piracetam could protect cells from sonoporation-associated apoptosis by preserving mitochondrial function. Copyright © 2018 World Federation for Ultrasound in Medicine and Biology. Published by Elsevier Inc. All rights reserved.

  17. Correlation of the microculture-kinetic drug-induced apoptosis assay with patient outcomes in initial treatment of adult acute myelocytic leukemia.

    Science.gov (United States)

    Strickland, Stephen A; Raptis, Anastasios; Hallquist, Allan; Rutledge, James; Chernick, Michael; Perree, Mathieu; Talbott, Mahsa S; Presant, Cary A

    2013-03-01

    Overall survival (OS) with acute myeloid leukemia (AML) remains poor. Determining prognostic factors will help in selecting patients for appropriate treatments. Our aim was to determine whether the level of drug-induced apoptosis (chemosensitivity) demonstrated by the microculture-kinetic drug-induced apoptosis (MiCK) assay significantly predicted outcomes after standard AML induction therapy. A total of 109 patients with untreated AML had blood and/or bone marrow aspirate samples analyzed for anthracycline-induced apoptosis using the MiCK assay. The amount of apoptosis observed over 48 h was determined and expressed as kinetic units of apoptosis (KU). Complete remission (CR) was significantly higher (72%) in patients with high idarubicin-induced apoptosis >3 KU compared to patients with apoptosis ≤ 3 KU (p = 0.01). Multivariate analysis showed the only significant variables to be idarubicin-induced apoptosis and karyotype. Median overall survival of patients with idarubicin-induced apoptosis >3 KU was 16.1 months compared to 4.5 months in patients with apoptosis ≤ 3 KU (p = 0.004). Multivariate analysis showed the only significant variable to be idarubicin-induced apoptosis. Chemotherapy-induced apoptosis measured by the MiCK assay demonstrated significant correlation with outcomes and appears predictive of complete remission and overall survival for patients receiving standard induction chemotherapy.

  18. Increased radiosensitivity of HPV-positive head and neck cancer cell lines due to cell cycle dysregulation and induction of apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Arenz, Andrea; Ziemann, Frank; Wittig, Andrea; Preising, Stefanie; Engenhart-Cabillic, Rita [Philipps-University, Department of Radiotherapy and Radiooncology, BMFZ - Biomedical Research Center, Marburg (Germany); Mayer, Christina; Wagner, Steffen; Klussmann, Jens-Peter; Wittekindt, Claus [Justus Liebig University, Department of Otorhinolaryngology and Head and Neck Surgery, Giessen (Germany); Dreffke, Kirstin [Philipps-University, Institute for Radiobiology and Molecular Radiooncology, Marburg (Germany)

    2014-09-15

    Human Papillomavirus (HPV)-related head and neck squamous cell carcinoma (HNSCC) respond favourably to radiotherapy as compared to HPV-unrelated HNSCC. We investigated DNA damage response in HPV-positive and HPV-negative HNSCC cell lines aiming to identify mechanisms, which illustrate reasons for the increased sensitivity of HPV-positive cancers of the oropharynx. Radiation response including clonogenic survival, apoptosis, DNA double-strand break (DSB) repair, and cell cycle redistribution in four HPV-positive (UM-SCC-47, UM-SCC-104, 93-VU-147T, UPCI:SCC152) and four HPV-negative (UD-SCC-1, UM-SCC-6, UM-SCC-11b, UT-SCC-33) cell lines was evaluated. HPV-positive cells were more radiosensitive (mean SF2: 0.198 range: 0.22-0.18) than HPV-negative cells (mean SF2: 0.34, range: 0.45-0.27; p = 0.010). Irradiated HPV-positive cell lines progressed faster through S-phase showing a more distinct accumulation in G2/M. The abnormal cell cycle checkpoint activation was accompanied by a more pronounced increase of cell death after x-irradiation and a higher number of residual and unreleased DSBs. The enhanced responsiveness of HPV-related HNSCC to radiotherapy might be caused by a higher cellular radiosensitivity due to cell cycle dysregulation and impaired DNA DSB repair. (orig.) [German] Fuer Patienten mit HPV-assoziierten Kopf-Hals-Tumoren (HNSCC) ist im Vergleich zu Patienten mit nicht-HPV-assoziierten Tumoren ein besseres Ueberleben nach Radiotherapie gesichert. Ziel der Untersuchung war die Identifizierung von Unterschieden in der zellulaeren DNA-Schadensantwort von HPV-positiven und HPV-negativen Zelllinien, wodurch die bereits in Erprobung stehende Deeskalation einer Radiotherapie bei Patienten mit HPV-assoziierten HNSCC durch experimentelle Daten abgesichert werden koennte. Klonogenes Ueberleben, Induktion von Apoptose, DNA-Doppelstrang-Reparatur und Zellzyklusverhalten wurden in vier HPV-positiven (UM-SCC-47, UM-SCC-104, 93-VU-147T, UPCI:SCC152) und vier HPV

  19. HBXIP regulates etoposide-induced cell cycle checkpoints and apoptosis in MCF-7 human breast carcinoma cells.

    Science.gov (United States)

    Fei, Hong-Rong; Li, Zhao-Jun; Ying-Zhang; Yue-Liu; Wang, Feng-Ze

    2018-03-20

    Etoposide, an anticancer DNA topoisomerase II poison, plays an important role in the therapy for human cancers. Unfortunately, many cancers develop etoposide resistance and do not respond to chemotherapy, leading to difficulty in treatment and poor prognosis. In this study, we investigate the effects of HBXIP gene silencing on etoposide chemosensitivity in MCF-7 human breast cancer cells. We find that etoposide increases HBXIP expression and promotes mobilization of HBXIP to the nucleus in MCF-7 cells. Knockdown of HBXIP alleviates etoposide-induced G2/M or S phase arrest. Upregulation of p53 and p21 upon etoposide treatment is attenuated in HBXIP knock-down cells. Moreover, HBXIP gene silencing sensitizes etoposide-induced cell apoptosis and cleavage of caspase-9 and PARP in MCF-7 cells. Knockdown of HBXIP expression by RNAi abrogates the etoposide-activated ERK and Akt. These results indicate that HBXIP can modulate the etoposide sensitivity of MCF-7 cell lines and further implicate HBXIP as a target for human breast cancer. Copyright © 2018 Elsevier B.V. All rights reserved.

  20. Epithelial apoptosis: cause or consequence of ulcerative colitis?

    DEFF Research Database (Denmark)

    Seidelin, Jakob Benedict; Nielsen, Ole Haagen

    2009-01-01

    OBJECTIVE: Epithelial apoptosis rates are increased in ulcerative colitis (UC). The increased apoptosis rate could expose mucosal cells to luminal pathogens and thereby be regarded as a primary pathogenic factor in UC. On the other hand, the local inflammatory reaction could cause epithelial...... apoptosis secondary to the release of cytotoxic mediators. If apoptosis is a primary defect, apoptosis rates could influence the degree of spreading of inflammation and the clinical course of UC. If apoptosis is a side effect of local inflammation, apoptosis rates would be expected only to correlate...

  1. miR-216b promotes cell growth and enhances chemosensitivity of colorectal cancer by suppressing PDZ-binding kinase.

    Science.gov (United States)

    Zou, Jun; Kuang, Weihua; Hu, Jilong; Rao, Huamin

    2017-06-24

    PDZ-binding kinase (PBK/TOPK) acts as oncogene in various cancers and correlates with drug response. However, few studies have examined the expression and roles of PBK in colonrectal cancer (CRC). In this study, we found a significant increase in the expression of PBK in CRC tissues and cell lines. While overexpression of PBK promoted cell growth and decreased the toxicity effect of oxaliplation (OXA), targeting PBK with short hairpin RNA (shRNA) or novel PBK inhibitor HI-TOPK-032 effectively suppressed tumor growth and potentiated chemosensitivity in vitro and in vivo. Furthermore, there was a significant inverse correlation between the expressions of miR-216b and PBK. Further found that miR-216b could down-regulate PBK levels by binding to the 3' untranslated region (3'UTR) of PBK. Notably, while miR-216b decreased cell proliferation and enhanced sensitivity of CRC cells to oxaliplation, re-expression of PBK dramatically reversed these events. Collectively, our data indicated that miR-216b may function as a tumor suppressor though regulating PBK expression, which provided promising targets and possible therapeutic strategies for CRC treatment. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. VEGFR3 Inhibition Chemosensitizes Ovarian Cancer Stemlike Cells through Down-Regulation of BRCA1 and BRCA2

    Directory of Open Access Journals (Sweden)

    Jaeyoung Lim

    2014-04-01

    Full Text Available In ovarian cancer, loss of BRCA gene expression in tumors is associated with improved response to chemotherapy and increased survival. A means to pharmacologically downregulate BRCA gene expression could improve the outcomes of patients with BRCA wild-type tumors. We report that vascular endothelial growth factor receptor 3 (VEGFR3 inhibition in ovarian cancer cells is associated with decreased levels of both BRCA1 and BRCA2. Inhibition of VEGFR3 in ovarian tumor cells was associated with growth arrest. CD133+ ovarian cancer stemlike cells were preferentially susceptible to VEGFR3-mediated growth inhibition. VEGFR3 inhibition–mediated down-regulation of BRCA gene expression reversed chemotherapy resistance and restored chemosensitivity in resistant cell lines in which a BRCA2 mutation had reverted to wild type. Finally, we demonstrate that tumor-associated macrophages are a primary source of VEGF-C in the tumor microenvironment. Our studies suggest that VEGFR3 inhibition may be a pharmacologic means to downregulate BRCA genes and improve the outcomes of patients with BRCA wild-type tumors.

  3. Conditioned medium from MCF-7 cell line induces myofibroblast differentiation, decreased cell proliferation, and increased apoptosis in cultured normal fibroblasts but not in fibroblasts from malignant breast tissue.

    Science.gov (United States)

    Valenti, M T; Azzarello, G; Balducci, E; Sartore, S; Sandri, M; Manconi, R; Sicari, U; Bari, M; Vinante, O

    2001-01-01

    We studied the effect of conditioned medium (CM) obtained from cultures of oestrogen-receptor positive breast cancer MCF7 cell line on the differentiation, proliferation and apoptosis patterns of cultured breast fibroblasts from normal interstitial and malignant stromal tissue. Fibroblasts were grown in the presence or absence of CM and examined for the differentiation pattern by immunofluorescence and Western blotting procedures, for proliferation profile by Ki67 expression, and for apoptosis by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling technique. Monoclonal antibodies specific for non-muscle (NM), smooth muscle (SM) lineage and differentiation markers were applied to these cultures. CM is able to induce a SM-like differentiation in interstitial fibroblasts, i.e., essentially myofibroblast formation. Fibroblasts from tumour stroma showed the presence of a small number of smooth muscle cells (SMC) along with a large number of myofibroblasts. Treatment of these cultures with CM was unable to change this pattern. Only normal fibroblasts were responsive to the proliferation/apoptotic-inhibitory effect of the CM. These data suggest that structural and functional differences exist between stromal fibroblasts from normal breast and breast cancer with respect to the responsiveness to soluble factors present in the CM. We hypothesize that the lack of in vitro sensitivity to CM shown by 'tumour' fibroblasts is the result of an in vivo inherent and stable phenotypic change on the fibroblasts surrounding breast tumour cells occurring via a paracrine mechanism.

  4. Isololiolide, a carotenoid metabolite isolated from the brown alga Cystoseira tamariscifolia, is cytotoxic and able to induce apoptosis in hepatocarcinoma cells through caspase-3 activation, decreased Bcl-2 levels, increased p53 expression and PARP cleavage.

    Science.gov (United States)

    Vizetto-Duarte, Catarina; Custódio, Luísa; Gangadhar, Katkam N; Lago, João Henrique G; Dias, Catarina; Matos, Ana Marta; Neng, Nuno; Nogueira, José Manuel Florêncio; Barreira, Luísa; Albericio, Fernando; Rauter, Amelia P; Varela, João

    2016-05-15

    Brown macroalgae have attracted attention because they display a wide range of biological activities, including antitumoral properties. Inthis study we isolated isololiolide from Cystoseira tamariscifolia for the first time. To examine the therapeutical potential of isololiolide against tumor cell lines. The structure of the compound was established and confirmed by 1D and 2D NMR as well as HRMS spectral analysis. The in vitro cytotoxicity was analyzed by colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay in tumoral as well as in non-tumoral cell lines. Cell cycle arrest and induction of apoptosis were assessed by flow cytometry. Alteration of expression levels in proteins important in the apoptotic cascade was analyzed by western blotting. Isololiolidewas isolated for the first time from the brown macroalga C.tamariscifolia. Isololiolide exhibited significant cytotoxic activity against three human tumoral cell lines, namely hepatocarcinoma HepG2 cells, whereas no cytotoxicity was found in non-malignant MRC-5 and HFF-1 human fibroblasts. Isololiolide completely disrupted the HepG2 normal cell cycle and induced significant apoptosis. Moreover, western blot analysis showed that isololiolide altered the expression of proteins that are important in the apoptotic cascade, increasing PARP cleavage and p53 expression while decreasing procaspase-3 and Bcl-2 levels. Isololiolide isolated from C. tamariscifolia is able to exert a selective cytotoxic activity on hepatocarcinoma HepG2 cells as well as induce apoptosis through the modulation of apoptosis-related proteins. Copyright © 2016 Elsevier GmbH. All rights reserved.

  5. A novel microculture kinetic assay (MiCK assay) for malignant cell growth and chemosensitivity.

    Science.gov (United States)

    Kravtsov, V D

    1994-01-01

    The THERMOmax microplate reader was adapted for monitoring the growth kinetics of human leukaemic OCI/AML-2 and mouse tumour J-774.1 cell lines in continuous culture. Fluid evaporation from wells, CO2 escape and contamination were prevented by hermetic sealing of the microcultures in wells of a 96-well microplate, thus enabling the cells to grow exponentially for 72 h under the conditions of the incubated microplate reader. For both OCI/AML-2 cells, which grow in suspension, and adherent J-774.1 cells, a linear correlation was demonstrated between the number of unstained cells seeded in a given microplate well and the optical density (OD) of that well. Therefore, the OD/time curve of the culture could be deemed to be its growth curve. By the use of the linear fit equation, the actual number of the cells in the wells was computable at any time point of the assay. In the chemosensitivity test, an inhibitory effect of ARA-C on the growth of the cells could be estimated by viewing of the growth curves plotted on the screen. The maximum kinetic rates (Vmax) of the curves in the control and the ARA-C-treated wells were compared, yielding a growth inhibition index (GII). Comparison of results of the kinetic chemosensitivity assay with those of a [3H]thymidine incorporation assay revealed that the novel assay is suitable for precise quantitation of the cell chemosensitivity, is more informative and has the added technical advantage of performance without recourse to radioactive or chemically hazardous substances.

  6. Melatonin increases the effect of 5-fluorouracil-based chemotherapy in human colorectal adenocarcinoma cells in vitro.

    Science.gov (United States)

    Pariente, Roberto; Bejarano, Ignacio; Rodríguez, Ana Beatriz; Pariente, José Antonio; Espino, Javier

    2018-03-01

    Melatonin has antitumor activity via several mechanisms including its anti-proliferative and pro-apoptotic effects. Moreover, it has been proven that melatonin in combination with chemotherapeutic agents enhances chemotherapy-triggered apoptosis in several types of cancer. Therefore, this study was intended to evaluate whether melatonin is able to strengthen the anti-cancer potential of different chemotherapeutic drugs in human colorectal adenocarcinoma HT-29 cells. We found that treatment with 20 µM cisplatin (CIS) or 1 mM 5-fluorouracil (5-FU) for 72 h induced a decrease in HT-29 cell viability. Furthermore, 1 mM melatonin significantly (P < 0.05) increased the cytotoxic effects of 5-FU. Likewise, simultaneous stimulation with 1 mM melatonin and 1 mM 5-FU significantly (P < 0.05) enhanced the ratio of cells with an overproduction of intracellular reactive oxygen species and substantially augmented the population of apoptotic cells compared to the treatment with 5-FU alone. Nonetheless, melatonin only displayed moderate chemosensitizing effects in CIS-treated HT-29 cells, as suggested by a slight increment in the fraction of early apoptotic cells that was observed only after 48 h. Consistently, co-stimulation of HT-29 cells with 20 µM CIS or 1 mM 5-FU in the presence of 1 mM melatonin further increased caspase-3 activation. Apart from this, the cytostatic activity displayed by CIS due to S phase arrest was not affected by concomitant stimulation with melatonin. Overall, our results indicate that melatonin increases the sensitivity of HT-29 cells to 5-FU treatment and, consequently, the indolamine could be potentially applied to colorectal adenocarcinoma treatment as a potent chemosensitizing agent.

  7. MTHFR deficiency or reduced intake of folate or choline in pregnant mice results in impaired short-term memory and increased apoptosis in the hippocampus of wild-type offspring.

    Science.gov (United States)

    Jadavji, N M; Deng, L; Malysheva, O; Caudill, M A; Rozen, R

    2015-08-06

    Genetic or nutritional disturbances in one-carbon metabolism, with associated hyperhomocysteinemia, can result in complex disorders including pregnancy complications and neuropsychiatric diseases. In earlier work, we showed that mice with a complete deficiency of methylenetetrahydrofolate reductase (MTHFR), a critical enzyme in folate and homocysteine metabolism, had cognitive impairment with disturbances in choline metabolism. Maternal demands for folate and choline are increased during pregnancy and deficiencies of these nutrients result in several negative outcomes including increased resorption and delayed development. The goal of this study was to investigate the behavioral and neurobiological impact of a maternal genetic deficiency in MTHFR or maternal nutritional deficiency of folate or choline during pregnancy on 3-week-old Mthfr(+/+) offspring. Mthfr(+/+) and Mthfr(+/-) females were placed on control diets (CD); and Mthfr(+/+) females were placed on folate-deficient diets (FD) or choline-deficient diets (ChDD) throughout pregnancy and lactation until their offspring were 3weeks of age. Short-term memory was assessed in offspring, and hippocampal tissue was evaluated for morphological changes, apoptosis, proliferation and choline metabolism. Maternal MTHFR deficiency resulted in short-term memory impairment in offspring. These dams had elevated levels of plasma homocysteine when compared with wild-type dams. There were no differences in plasma homocysteine in offspring. Increased apoptosis and proliferation was observed in the hippocampus of offspring from Mthfr(+/-) mothers. In the maternal FD and ChDD study, offspring also showed short-term memory impairment with increased apoptosis in the hippocampus; increased neurogenesis was observed in ChDD offspring. Choline acetyltransferase protein was increased in the offspring hippocampus of both dietary groups and betaine was decreased in the hippocampus of FD offspring. Our results reveal short-term memory

  8. Myoblasts isolated from hypertrophy-responsive callipyge muscles show altered growth rates and increased resistance to serum deprivation-induced apoptosis.

    Science.gov (United States)

    Lavulo, Lopeti T; Uaesoontrachoon, Kitipong; Mirams, Michiko; White, Jason D; Cockett, Noelle E; Mackie, Eleanor J; Pagel, Charles N

    2008-01-01

    Back and hind limb muscles of sheep paternally heterozygous for the callipyge single nucleotide polymorphism undergo extensive hypertrophy shortly after birth. We have established cell cultures from foetal semitendinosus and longissimus dorsi muscles of normal and callipyge animals. Cultures were assessed for rates of proliferation, cell death, myogenicity and DLK1 expression. Myoblasts from callipyge semitendinosus, but not longissimus dorsi muscles, proliferated faster than myoblasts isolated from normal semitendinosus muscle, and cells isolated from either callipyge muscle were more resistant to serum deprivation-induced apoptosis than equivalent cells isolated from normal individuals. These observations indicate that there are intrinsic differences in the behaviour of isolated myoblasts, which are associated with their muscle and genotype of origin. As myoblasts are the cells responsible for hypertrophy of muscle fibres, the observed differences in cell growth may play a role in the hypertrophy of certain muscles in callipyge animals. Copyright 2007 S. Karger AG, Basel.

  9. Oridonin induces apoptosis and senescence in colorectal cancer cells by increasing histone hyperacetylation and regulation of p16, p21, p27 and c-myc

    International Nuclear Information System (INIS)

    Gao, Feng-Hou; Liu, Feng; Zhao, Ying-Zheng; Fang, Yong; Chen, Fang-Yuan; Wu, Ying-Li; Hu, Xiao-Hui; Li, Wei; Liu, Hua; Zhang, Yan-Jie; Guo, Zhu-Ying; Xu, Mang-Hua; Wang, Shi-Ting; Jiang, Bin

    2010-01-01

    Oridonin, a tetracycline diterpenoid compound, has the potential antitumor activities. Here, we evaluate the antitumor activity and action mechanisms of oridonin in colorectal cancer. Effects of oridonin on cell proliferation were determined by using a CCK-8 Kit. Cell cycle distribution was determined by flow cytometry. Apoptosis was examined by analyzing subdiploid population and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. Senescent cells were determined by senescence-associated β-galactosidase activity analysis. Semi-quantitative RT-PCR was used to examine the changes of mRNA of p16, p21, p27 and c-myc. The concomitant changes of protein expression were analyzed with Western blot. Expression of AcH3 and AcH4 were examined by immunofluorescence staining and Western blots. Effects of oridonin on colony formation of SW1116 were examined by Soft Agar assay. The in vivo efficacy of oridonin was detected using a xenograft colorectal cancer model in nude mice. Oridonin induced potent growth inhibition, cell cycle arrest, apoptosis, senescence and colony-forming inhibition in three colorectal cancer cell lines in a dose-dependent manner in vitro. Daily i.p. injection of oridonin (6.25, 12.5 or 25 mg/kg) for 28 days significantly inhibited the growth of SW1116 s.c. xenografts in BABL/C nude mice. With western blot and reverse transcription-PCR, we further showed that the antitumor activities of oridonin correlated with induction of histone (H3 and H4) hyperacetylation, activation of p21, p27 and p16, and suppression of c-myc expression. Oridonin possesses potent in vitro and in vivo anti-colorectal cancer activities that correlated with induction of histone hyperacetylation and regulation of pathways critical for maintaining growth inhibition and cell cycle arrest. Therefore, oridonin may represent a novel therapeutic option in colorectal cancer treatment

  10. Oridonin induces apoptosis and senescence in colorectal cancer cells by increasing histone hyperacetylation and regulation of p16, p21, p27 and c-myc

    Directory of Open Access Journals (Sweden)

    Zhao Ying-Zheng

    2010-11-01

    Full Text Available Abstract Background Oridonin, a tetracycline diterpenoid compound, has the potential antitumor activities. Here, we evaluate the antitumor activity and action mechanisms of oridonin in colorectal cancer. Methods Effects of oridonin on cell proliferation were determined by using a CCK-8 Kit. Cell cycle distribution was determined by flow cytometry. Apoptosis was examined by analyzing subdiploid population and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. Senescent cells were determined by senescence-associated β-galactosidase activity analysis. Semi-quantitative RT-PCR was used to examine the changes of mRNA of p16, p21, p27 and c-myc. The concomitant changes of protein expression were analyzed with Western blot. Expression of AcH3 and AcH4 were examined by immunofluorescence staining and Western blots. Effects of oridonin on colony formation of SW1116 were examined by Soft Agar assay. The in vivo efficacy of oridonin was detected using a xenograft colorectal cancer model in nude mice. Results Oridonin induced potent growth inhibition, cell cycle arrest, apoptosis, senescence and colony-forming inhibition in three colorectal cancer cell lines in a dose-dependent manner in vitro. Daily i.p. injection of oridonin (6.25, 12.5 or 25 mg/kg for 28 days significantly inhibited the growth of SW1116 s.c. xenografts in BABL/C nude mice. With western blot and reverse transcription-PCR, we further showed that the antitumor activities of oridonin correlated with induction of histone (H3 and H4 hyperacetylation, activation of p21, p27 and p16, and suppression of c-myc expression. Conclusion Oridonin possesses potent in vitro and in vivo anti-colorectal cancer activities that correlated with induction of histone hyperacetylation and regulation of pathways critical for maintaining growth inhibition and cell cycle arrest. Therefore, oridonin may represent a novel therapeutic option in colorectal cancer treatment.

  11. Tumor-derived hepatocyte growth factor is associated with poor prognosis of patients with glioma and influences the chemosensitivity of glioma cell line to cisplatin in vitro

    Directory of Open Access Journals (Sweden)

    Guo You-feng

    2012-06-01

    Full Text Available Abstract Background We examined the association of tumor-derived hepatocyte growth factor (HGF with the clinicopathological features of gliomas and investigated the effect of HGF inhibition on the biological behavior of tumor cells in vitro in order to determine whether HGF is a valuable prognostic predictor for glioma patients. Methods Seventy-six cases of glioma were collected. The tumor-derived HGF expression, cell proliferation index (PI and intratumoral microvessels were evaluated by immunohistochemistry. Correlation between immunostaining and clinicopathological parameters, as well as the follow-up data of patients, was analyzed statistically. U87MG glioma cells were transfected with short interference (si-RNA for HGF, and the cell viability, migratory ability and chemosensitivity to cisplatin were evaluated in vitro. Results Both high HGF expression in tumor cells (59.2%, 45/76 and high PI were significantly associated with high-grade glioma and increased microvessels in tumors (P P = 0.004 and high-expression of HGF (P = 0.008 emerged as independent prognostic factors for the overall survival of glioma patients. The tumor-derived HGF mRNA and protein expressions were significantly decreased in vitro after transfection of HGF siRNA. HGF siRNA inhibited the cell growth and reduced cell migratory ability. Moreover, HGF siRNA transfection enhanced the chemosensitivity of U87MG glioma cells to cisplatin. Conclusion This study indicated that there was significant correlation among tumor cell-derived HGF, cell proliferation and microvessel proliferation in gliomas. HGF might influence tumor progression by modulating the cell growth, migration and chemoresistance to drugs. Increased expression of HGF may be a valuable predictor for prognostic evaluation of glioma patients.

  12. Expression of TP53 isoforms p53β or p53γ enhances chemosensitivity in TP53(null cell lines.

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    Elisabeth Silden

    Full Text Available The carboxy-terminal truncated p53 alternative spliced isoforms, p53β and p53γ, are expressed at disparate levels in cancer and are suggested to influence treatment response and therapy outcome. However, their functional role in cancer remains to be elucidated. We investigated their individual functionality in the p53(null background of cell lines H1299 and SAOS-2 by stable retroviral transduction or transient transfection. Expression status of p53β and p53γ protein was found to correlate with increased response to camptothecin and doxorubicin chemotherapy. Decreased DNA synthesis and clonogenicity in p53β and p53γ congenic H1299 was accompanied by increased p21((CIP1/WAF1, Bax and Mdm2 proteins. Chemotherapy induced p53 isoform degradation, most prominent for p53γ. The proteasome inhibitor bortezomib substantially increased basal p53γ protein level, while the level of p53β protein was unaffected. Treatment with dicoumarol, a putative blocker of the proteasome-related NAD(PH quinone oxidoreductase NQO1, effectively attenuated basal p53γ protein level in spite of bortezomib treatment. Although in vitro proliferation and clonogenicity assays indicated a weak suppressive effect by p53β and p53γ expression, studies of in vivo subcutaneous H1299 tumor growth demonstrated a significantly increased growth by expression of either p53 isoforms. This study suggests that p53β and p53γ share functionality in chemosensitizing and tumor growth enhancement but comprise distinct regulation at the protein level.

  13. Microculture-based chemosensitivity testing: a feasibility study comparing freshly explanted human melanoma cells with human melanoma cell lines.

    Science.gov (United States)

    Marshall, E S; Finlay, G J; Matthews, J H; Shaw, J H; Nixon, J; Baguley, B C

    1992-03-04

    The culture of cancer cells has many applications in chemosensitivity testing and new drug development. Our goal was to adapt simple semiautomated microculture methods for testing the chemosensitivity of melanoma cells freshly recovered from patients' tumors. Cells were cultured on a substrate of agarose and exposed continuously to cytotoxic drugs, the effects of which were measured by determining the uptake of [3H]thymidine 4-7 days later. Immunocytochemical staining of cells cultured with 5-bromo-2'-deoxyuridine demonstrated that tumor cells were responsible for the measured thymidine incorporation. The effects of cytotoxic drugs were calculated as logarithmic 50% inhibitory concentrations and expressed as divergences from the mean in a log-mean graph. The inhibitory effects of amsacrine, etoposide, doxorubicin, cisplatin, mitomycin C, and fluorouracil were tested. Tumors differed widely in their sensitivity to these drugs, although sensitivity to the three topoisomerase-II-directed agents was highly correlated. Cells from two non-neoplastic hematopoietic progenitor cell lines (FT and 32D) showed chemosensitivity patterns distinct from those in the melanoma cells, indicating tissue selectivity. Two established melanoma cell lines, MM-96 and FME, were tested under the same conditions and showed sensitivity typical of at least some fresh specimens. These results support the validity of melanoma cell lines as models of freshly resected melanoma cells. If successfully applied to other tumor types, such semiautomated approaches could find wide application in routine hospital laboratories for the chemosensitivity testing of patients' tumor cells.

  14. A Simple Non-destructive Test of Cellular Activity (NTCA) for in vitro Assessment of Cancer Cell Chemosensitivity/resistance

    Czech Academy of Sciences Publication Activity Database

    Netíková, I.; Buršíková, Eva; Veselý, Pavel; Prchalová, M.; Kleibl, Z.; Matoušková, Eva

    2007-01-01

    Roč. 27, 4B (2007), s. 2331-2337 ISSN 0250-7005 R&D Projects: GA MZd NR8145 Institutional research plan: CEZ:AV0Z50520514 Keywords : in vitro chemosensitivity * primary cells * non-destructive test Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.414, year: 2007

  15. Natural killer cell cytokine response to M. bovis BCG Is associated with inhibited proliferation, increased apoptosis and ultimate depletion of NKp44(+CD56(bright cells.

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    Damien Portevin

    Full Text Available Mycobacterium bovis BCG, a live attenuated strain of M. bovis initially developed as a vaccine against tuberculosis, is also used as an adjuvant for immunotherapy of cancers and for treatment of parasitic infections. The underlying mechanisms are thought to rely on its immunomodulatory properties including the recruitment of natural killer (NK cells. In that context, we aimed to study the impact of M. bovis BCG on NK cell functions. We looked at cytotoxicity, cytokine production, proliferation and cell survival of purified human NK cells following exposure to single live particles of mycobacteria. We found that M. bovis BCG mediates apoptosis of NK cells only in the context of IL-2 stimulation during which CD56(bright NK cells are releasing IFN-γ in response to mycobacteria. We found that the presence of mycobacteria prevented the IL-2 induced proliferation and surface expression of NKp44 receptor by the CD56(bright population. In summary, we observed that M. bovis BCG is modulating the functions of CD56(bright NK cells to drive this subset to produce IFN-γ before subsequent programmed cell death. Therefore, IFN-γ production by CD56(bright cells constitutes the main effector mechanism of NK cells that would contribute to the benefits observed for M. bovis BCG as an immunotherapeutic agent.

  16. Protective effect of Tanshinone IIA against infarct size and increased HMGB1, NFκB, GFAP and apoptosis consequent to transient middle cerebral artery occlusion.

    Science.gov (United States)

    Wang, Jian-Gang; Bondy, Stephen C; Zhou, Li; Yang, Feng-Zhen; Ding, Zhi-Gang; Hu, Yu; Tian, Yun; Wen, Pu-Yuan; Luo, Hao; Wang, Fang; Li, Wen-Wen; Zhou, Jun

    2014-02-01

    Acute inflammation plays an important role in brain damage following cerebral ischemia and reperfusion (I/R) injury. The present study employed a rat model of middle cerebral artery occlusion to explore the neuroprotective effects of tanshinone IIA (TSN), which is widely used in China for treating cerebrovascular and cardiovascular diseases. Rats were divided into a sham-operated group and I/R transiently occluded then reperfused groups. Some of the I/R animals were treated daily for 7 or 15 days with two different doses of TSN. After 15 days, triphenyl tetrazolium chloride staining revealed less unstained area indicating fewer lesions in the TSN-treated I/R group relative to the untreated corresponding I/R group. TSN treatment dramatically reduced infarct sizes and reduced content of high mobility group box 1 protein following I/R. Nuclear translocation of NFκB was also attenuated in I/R animals subsequently receiving TSN. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining revealed more apoptosis in the I/R model group and this was reduced in the I/R animals treated with TSN for 15 days. Thus, TSN mitigates the severity of damage effected by I/R.

  17. Natural killer cell cytokine response to M. bovis BCG Is associated with inhibited proliferation, increased apoptosis and ultimate depletion of NKp44(+)CD56(bright) cells.

    Science.gov (United States)

    Portevin, Damien; Young, Douglas

    2013-01-01

    Mycobacterium bovis BCG, a live attenuated strain of M. bovis initially developed as a vaccine against tuberculosis, is also used as an adjuvant for immunotherapy of cancers and for treatment of parasitic infections. The underlying mechanisms are thought to rely on its immunomodulatory properties including the recruitment of natural killer (NK) cells. In that context, we aimed to study the impact of M. bovis BCG on NK cell functions. We looked at cytotoxicity, cytokine production, proliferation and cell survival of purified human NK cells following exposure to single live particles of mycobacteria. We found that M. bovis BCG mediates apoptosis of NK cells only in the context of IL-2 stimulation during which CD56(bright) NK cells are releasing IFN-γ in response to mycobacteria. We found that the presence of mycobacteria prevented the IL-2 induced proliferation and surface expression of NKp44 receptor by the CD56(bright) population. In summary, we observed that M. bovis BCG is modulating the functions of CD56(bright) NK cells to drive this subset to produce IFN-γ before subsequent programmed cell death. Therefore, IFN-γ production by CD56(bright) cells constitutes the main effector mechanism of NK cells that would contribute to the benefits observed for M. bovis BCG as an immunotherapeutic agent.

  18. Probenecid is a chemosensitizer in cancer cell lines.

    Science.gov (United States)

    Campos-Arroyo, Denise; Martínez-Lazcano, Juan Carlos; Melendez-Zajgla, Jorge

    2012-02-01

    Resistance and toxicity are the major barriers to successful cancer chemotherapies. Developing molecules that reduce drug resistance and improve antineoplastic effects is of great interest for cancer research; ideally, these substances should not affect the pharmacodynamics of the chemotherapeutic agent while providing a synergistic antineoplastic effect. In this study, we tested in vitro co-administration of the antineoplastic agents cisplatin or paclitaxel with probenecid, an anion channel inhibitor, in a panel of cancer cell lines to determine the cytotoxicity and synergistic effects of these drug combinations. In addition, we measured the clonogenicity and apoptotic index in these cells. We observed a synergistic interaction between probenecid and the chemotherapeutic agents, and increasing doses of probenecid resulted in a significant decrease in the effective doses of the chemotherapeutic agents. For the antineoplastic agent and probenecid combinations, we found increased cell death, reduced colony formation, and a higher number of apoptotic cells, compared with treatment of cisplatin or paclitaxel alone. Further research is necessary to elucidate the molecular mechanisms by which the synergistic effect occurs. If these synergistic effects can be reproduced in vivo, the co-administration of probenecid with different chemotherapeutic agents may provide a valid treatment in patients with chemotherapy resistance.

  19. Placental apoptosis in recurrent miscarriage

    Directory of Open Access Journals (Sweden)

    Tarek A. Atia

    2017-09-01

    Full Text Available Apoptosis is an interactive and dynamic biological process involved in all phases of embryogenesis. We aimed to study the effect of placental apoptosis on recurrent miscarriage (RM. Placental tissue samples were collected from 40 women with RM (study group and 30 women with sporadic spontaneous abortion (control group. Samples were prepared and stained immunohistochemically with markers for both the apoptotic protein (p53 and anti-apoptotic Bcl-2 antibodies. Our results showed that expression of the apoptotic (p53 protein was significantly increased in the placental tissues of the RM group (p = 0.003. By contrast, the expression of anti-apoptotic (Bcl-2 antibodies was significantly increased in the placental tissues of the control group (p = 0.025. We concluded that placental apoptosis plays a crucial role in pregnancy continuation. However, increased p53 expression in placental tissue in early pregnancy could negatively affect pregnancy continuation.

  20. Increased apoptosis and DNA double-strand breaks in the embryonic mouse brain in response to very low-dose X-rays but not 50 Hz magnetic fields.

    Science.gov (United States)

    Saha, Shreya; Woodbine, Lisa; Haines, Jackie; Coster, Margaret; Ricket, Nicole; Barazzuol, Lara; Ainsbury, Elizabeth; Sienkiewicz, Zenon; Jeggo, Penny

    2014-11-06

    The use of X-rays for medical diagnosis is enhancing exposure to low radiation doses. Exposure to extremely low-frequency electromagnetic or magnetic fields is also increasing. Epidemiological studies show consistent associations of childhood leukaemia with exposure to magnetic fields but any causal relationship is unclear. A limitation in assessing the consequence of such exposure is the availability of sensitive assays. The embryonic neuronal stem and progenitor cell compartments are radiosensitive tissues. Using sensitive assays, we report a statistically significant increase in DNA double-strand break (DSB) formation and apoptosis in the embryonic neuronal stem cell compartment following in utero exposure to 10-200 mGy X-rays. Both endpoints show a linear response. We also show that DSB repair is delayed following exposure to doses below 50 mGy compared with 100 mGy. Thus, we demonstrate in vivo consequences of low-dose radiation. In contrast to these impacts, we did not observe any significant induction of DSBs or apoptosis following exposure to 50 Hz magnetic fields (100 or 300 µT). We conclude that any DSB induction by treatment with magnetic fields is lower than following exposure to 10 mGy X-rays. For comparison, certain procedures involving computed tomography scanning are equivalent to 1-5 mGy X-rays.

  1. Nutrient Sensing: Another Chemosensitivity of the Olfactory System

    Directory of Open Access Journals (Sweden)

    A-Karyn Julliard

    2017-07-01

    Full Text Available Olfaction is a major sensory modality involved in real time perception of the chemical composition of the external environment. Olfaction favors anticipation and rapid adaptation of behavioral responses necessary for animal survival. Furthermore, recent studies have demonstrated that there is a direct action of metabolic peptides on the olfactory network. Orexigenic peptides such as ghrelin and orexin increase olfactory sensitivity, which in turn, is decreased by anorexigenic hormones such as insulin and leptin. In addition to peptides, nutrients can play a key role on neuronal activity. Very little is known about nutrient sensing in olfactory areas. Nutrients, such as carbohydrates, amino acids, and lipids, could play a key role in modulating olfactory sensitivity to adjust feeding behavior according to metabolic need. Here we summarize recent findings on nutrient-sensing neurons in olfactory areas and delineate the limits of our knowledge on this topic. The present review opens new lines of investigations on the relationship between olfaction and food intake, which could contribute to determining the etiology of metabolic disorders.

  2. Experimental pneumococcal meningitis: impaired clearance of bacteria from the blood due to increased apoptosis in the spleen in Bcl-2-deficient mice.

    Science.gov (United States)

    Wellmer, Andreas; von Mering, Matthias; Spreer, Annette; Diem, Ricarda; Eiffert, Helmut; Noeske, Christiane; Bunkowski, Stefanie; Gold, Ralf; Nau, Roland

    2004-06-01

    Necrotic and apoptotic neuronal cell death can be found in pneumococcal meningitis. We investigated the role of Bcl-2 as an antiapoptotic gene product in pneumococcal meningitis using Bcl-2 knockout (Bcl-2(-/-)) mice. By using a model of pneumococcal meningitis induced by intracerebral infection, Bcl-2-deficient mice and control littermates were assessed by clinical score and a tight rope test at 0, 12, 24, 32, and 36 h after infection. Then mice were sacrificed, the bacterial titers in blood, spleen, and cerebellar homogenates were determined, and the brain and spleen were evaluated histologically. The Bcl-2-deficient mice developed more severe clinical illness, and there were significant differences in the clinical score at 24, 32, and 36 h and in the tight rope test at 12 and 32 h. The bacterial titers in the blood were greater in Bcl-2-deficient mice than in the controls (7.46 +/- 1.93 log CFU/ml versus 5.16 +/- 0.96 log CFU/ml [mean +/- standard deviation]; P < 0.01). Neuronal damage was most prominent in the hippocampal formation, but there were no significant differences between groups. In situ tailing revealed only a few apoptotic neurons in the brain. In the spleen, however, there were significantly more apoptotic leukocytes in Bcl-2-deficient mice than in controls (5,148 +/- 3,406 leukocytes/mm2 versus 1,070 +/- 395 leukocytes/mm2; P < 0.005). Bcl-2 appears to counteract sepsis-induced apoptosis of splenic lymphocytes, thereby enhancing clearance of bacteria from the blood.

  3. Mutation of a Nopp140 gene dao-5 alters rDNA transcription and increases germ cell apoptosis in C. elegans.

    Science.gov (United States)

    Lee, C-C; Tsai, Y-T; Kao, C-W; Lee, L-W; Lai, H-J; Ma, T-H; Chang, Y-S; Yeh, N-H; Lo, S J

    2014-04-10

    Human diseases of impaired ribosome biogenesis resulting from disruption of rRNA biosynthesis or loss of ribosomal components are collectively described as 'ribosomopathies'. Treacher Collins syndrome (TCS), a representative human ribosomopathy with craniofacial abnormalities, is attributed to mutations in the tcof1 gene that has a homologous gene called nopp140. Previous studies demonstrated that the dao-5 (dauer and aged animal overexpression gene 5) of Caenorhabditis elegans is a member of nopp140 gene family and plays a role in nucleogenesis in the early embryo. Here, we established a C. elegans model for studying Nopp140-associated ribosomopathy. A null dao-5 mutant ok542 with a semi-infertile phenotype showed a delay in gonadogenesis, as well as a higher incidence of germline apoptosis. These phenotypes in dao-5(ok542) are likely resulted from inefficient rDNA transcription that was observed by run-on analyses and chromatin immunoprecipitation (ChIP) assays measuring the RNA Pol I occupancy on the rDNA promoter. ChIP assays further showed that the modifications of acetylated histone 4 (H4Ac) and dimethylation at the lysine 9 of histone 3 (H3K9me2) around the rDNA promoter were altered in dao-5 mutants compared with the N2 wild type. In addition, activated CEP-1 (a C. elegans p53 homolog) activity was also linked to the loss of DAO-5 in terms of the transcriptional upregulation of two CEP-1 downstream effectors, EGL-1 and CED-13. We propose that the dao-5 mutant of C. elegans can be a valuable model for studying human Nopp140-associated ribosomopathy at the cellular and molecular levels.

  4. Fe3O4 nanoparticle loaded paclitaxel induce multiple myeloma apoptosis by cell cycle arrest and increase cleavage of caspases in vitro

    Science.gov (United States)

    Yang, Cuiping; He, Xiangfeng; Chen, Junsong; Chen, Dengyu; Liu, Yunjing; Xiong, Fei; Shi, Fangfang; Dou, Jun; Gu, Ning

    2013-08-01

    Multiple myeloma (MM) still remains an incurable disease in spite of extending the patient survival by new therapies. The hypothesis of cancer stem cells (CSCs) states that although chemotherapy kills most tumor cells, it is believed to leave a reservoir of CSCs that allows the tumor cell propagation. The objective of this research was to evaluate the therapeutic effect of new paclitaxel-Fe3O4 nanoparticles (PTX-NPs) with an average size range of 7.17 ± 1.31 nm on MM CSCs in vitro. The characteristics of CD138-CD34- cells, isolated from human MM RPMI 8226 and NCI-H929 cell lines by the magnetic associated cell sorting method, were identified by the assays of colony formation, cell proliferation, drug resistance, cell migration, and tumorigenicity in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, respectively. Inhibitory effects of PTX-NPs on CD138-CD34- cells were evaluated by a variety of assays in vitro. The results showed that the CD138-CD34- cells were capable of forming colonies, exhibited high proliferative and migratory ability, possessed a strong drug resistance, and had powerful tumorigenicity in NOD/SCID mice compared to non-CD138-CD34- cells. PTX-NPs significantly inhibited CD138- CD34- cell viability and invasive ability, and resulted in G0/G1 cell cycle arrest and apoptosis compared with PTX alone. We concluded that the CD138-CD34- phenotype cells might be CSCs in RPMI 8226 and NCI-H929 cell lines. PTX-NPs had an obvious inhibitory effect on MM CD138-CD34- CSCs. The findings may provide a guideline for PTX-NPs' treatment of MM CSCs in preclinical investigation.

  5. Fe{sub 3}O{sub 4} nanoparticle loaded paclitaxel induce multiple myeloma apoptosis by cell cycle arrest and increase cleavage of caspases in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Cuiping [Medical School, Southeast University, Department of Pathogenic Biology and Immunology (China); He, Xiangfeng [Affiliated Tumor Hospital of Nantong University, Department of Medical Oncology (China); Chen, Junsong; Chen, Dengyu; Liu, Yunjing [Medical School, Southeast University, Department of Pathogenic Biology and Immunology (China); Xiong, Fei [Southeast University, School of Biological Science and Medical Engineering (China); Shi, Fangfang [Zhongda Hospital, Southeast University, Department of Oncology (China); Dou, Jun, E-mail: njdoujun@yahoo.com.cn [Medical School, Southeast University, Department of Pathogenic Biology and Immunology (China); Gu, Ning, E-mail: guning@seu.edu.cn [Southeast University, School of Biological Science and Medical Engineering (China)

    2013-08-15

    Multiple myeloma (MM) still remains an incurable disease in spite of extending the patient survival by new therapies. The hypothesis of cancer stem cells (CSCs) states that although chemotherapy kills most tumor cells, it is believed to leave a reservoir of CSCs that allows the tumor cell propagation. The objective of this research was to evaluate the therapeutic effect of new paclitaxel-Fe{sub 3}O{sub 4} nanoparticles (PTX-NPs) with an average size range of 7.17 {+-} 1.31 nm on MM CSCs in vitro. The characteristics of CD138{sup -}CD34{sup -} cells, isolated from human MM RPMI 8226 and NCI-H929 cell lines by the magnetic associated cell sorting method, were identified by the assays of colony formation, cell proliferation, drug resistance, cell migration, and tumorigenicity in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, respectively. Inhibitory effects of PTX-NPs on CD138{sup -}CD34{sup -} cells were evaluated by a variety of assays in vitro. The results showed that the CD138{sup -}CD34{sup -} cells were capable of forming colonies, exhibited high proliferative and migratory ability, possessed a strong drug resistance, and had powerful tumorigenicity in NOD/SCID mice compared to non-CD138{sup -}CD34{sup -} cells. PTX-NPs significantly inhibited CD138{sup -} CD34{sup -} cell viability and invasive ability, and resulted in G0/G1 cell cycle arrest and apoptosis compared with PTX alone. We concluded that the CD138{sup -}CD34{sup -} phenotype cells might be CSCs in RPMI 8226 and NCI-H929 cell lines. PTX-NPs had an obvious inhibitory effect on MM CD138{sup -}CD34{sup -} CSCs. The findings may provide a guideline for PTX-NPs' treatment of MM CSCs in preclinical investigation.

  6. Fe3O4 nanoparticle loaded paclitaxel induce multiple myeloma apoptosis by cell cycle arrest and increase cleavage of caspases in vitro

    International Nuclear Information System (INIS)

    Yang, Cuiping; He, Xiangfeng; Chen, Junsong; Chen, Dengyu; Liu, Yunjing; Xiong, Fei; Shi, Fangfang; Dou, Jun; Gu, Ning

    2013-01-01

    Multiple myeloma (MM) still remains an incurable disease in spite of extending the patient survival by new therapies. The hypothesis of cancer stem cells (CSCs) states that although chemotherapy kills most tumor cells, it is believed to leave a reservoir of CSCs that allows the tumor cell propagation. The objective of this research was to evaluate the therapeutic effect of new paclitaxel-Fe 3 O 4 nanoparticles (PTX-NPs) with an average size range of 7.17 ± 1.31 nm on MM CSCs in vitro. The characteristics of CD138 − CD34 − cells, isolated from human MM RPMI 8226 and NCI-H929 cell lines by the magnetic associated cell sorting method, were identified by the assays of colony formation, cell proliferation, drug resistance, cell migration, and tumorigenicity in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, respectively. Inhibitory effects of PTX-NPs on CD138 − CD34 − cells were evaluated by a variety of assays in vitro. The results showed that the CD138 − CD34 − cells were capable of forming colonies, exhibited high proliferative and migratory ability, possessed a strong drug resistance, and had powerful tumorigenicity in NOD/SCID mice compared to non-CD138 − CD34 − cells. PTX-NPs significantly inhibited CD138 − CD34 − cell viability and invasive ability, and resulted in G0/G1 cell cycle arrest and apoptosis compared with PTX alone. We concluded that the CD138 − CD34 − phenotype cells might be CSCs in RPMI 8226 and NCI-H929 cell lines. PTX-NPs had an obvious inhibitory effect on MM CD138 − CD34 − CSCs. The findings may provide a guideline for PTX-NPs’ treatment of MM CSCs in preclinical investigation

  7. Rapid in vivo Taxotere quantitative chemosensitivity response by 4.23 Tesla sodium MRI and histo-immunostaining features in N-Methyl-N-Nitrosourea induced breast tumors in rats

    Directory of Open Access Journals (Sweden)

    Wu Ed X

    2005-08-01

    Full Text Available Abstract Background Sodium weighted images can indicate sodium signal intensities from different features in the tumor before and 24 hours following administration of Taxotere. Aim To evaluate the association of in vivo intracellular sodium magnetic resonance image intensities with immuno-biomarkers and histopathological features to monitor the early tumor response to Taxotere chemotherapy in Methyl-Nitroso-Urea induced rat xenograft breast tumors. Methods and Materials Methyl-Nitroso-Urea (MNU induced rat xenograft breast tumors were imaged for sodium MRI and compared with tumor histology, immunostaining after 24 hours chemotherapy. Results Sodium MRI signal intensities represented sodium concentrations. Excised tumor histological sections showed different in vitro histological end points i.e. single strand DNA content of cell nuclei during cell cycle (G1/S-G2/M, distinct S or M histograms (Feulgen labeling to nuclear DNA content by CAS 200, mitotic figures and apoptosis at different locations of breast tumors. Necrosis and cystic fluid appeared gray on intracellular (IC sodium images while apoptosis rich regions appeared brighter on IC sodium images. After 24 hours Taxotere-treated tumors showed lower 'IC/EC ratio' of viable cells (65–76% with higher mitotic index; apoptotic tumor cells at high risk due to cytotoxicity (>70% with high apoptotic index; reduced proliferation index (270 vs 120 per high power field associated with enhanced IC sodium in vivo MR image intensities and decreased tumor size (3%; p in vivo associated with apoptosis and different pre-malignant features within 24 hours of exposure of cancer cells to anti-neoplastic Taxotere drug. Conclusion Sodium MRI imaging may be used as in vivo rapid drug monitoring method to evaluate Taxotere chemosensitivity response associated with neoplasia, apoptosis and tumor histology features.

  8. Maternal treatment with agonistic autoantibodies against type-1 angiotensin II receptor in late pregnancy increases apoptosis of myocardial cells and myocardial susceptibility to ischemia-reperfusion injury in offspring rats.

    Science.gov (United States)

    Jin, Zhu; Zhang, Wenhui; Yang, Hailiang; Wang, Xiaofang; Zheng, Yanqian; Zhang, Qiaoyan; Zhi, Jianming

    2013-01-01

    Epidemiological studies have demonstrated that offspring born to mothers preeclampsia (PE) are at increased risk for developing cardiovascular diseases after birth, but the underlying mechanism is unknown. Angiotensin II receptor type 1 autoantibody (AT1-AA), an agonist acting via activation of the AT1 receptor, is believed to be involved in the pathogenesis of both PE and fetal growth restriction. The aim of the present study was to confirm the hypothesis that prenatal AT1-AA exposure increases the heart susceptibility to ischemia/reperfusion injury (IRI) in the offspring in an AT1-AA-induced animal model of PE, and determine whether or not the increase of maternal AT1-AA level is a factor contributing to sustained abnormalities of the heart structure during infancy. The hearts of 45-day-old offspring rats were studied using Langendorff preparation to determine the susceptibility of the heart to IRI. The results showed that the body weight of the maternal rats was not significantly different between the study and control groups, but the body weight of their offspring in AT1-AA group was decreased slightly at day 21 of gestational age, and at day 3 after birth. Although the heart weight index was not significantly affected at all ages examined, AT1-AA significantly increased the size of myocardial cells of the left ventricle (LV) at the age of 45 days. AT1-AA gained access to fetal circulation via the placenta and induced apoptosis of fetal myocardial cells. AT1-AA also significantly delayed recovery from IRI and affected the LV function of 45-day-old offspring. This was associated with a significant increase in IRI-induced LV myocardial infarct size. These results suggest that AT1-AA induced abnormal apoptosis of fetal myocardial cells during the fetal period and increased the cardiac susceptibility to IRI in adult offspring.

  9. Targeting the oxidative stress response system of fungi with safe, redox-potent chemosensitizing agents

    Directory of Open Access Journals (Sweden)

    Jong H. eKim

    2012-03-01

    Full Text Available The cellular antioxidation system is a target in the antifungal action of amphotericin B (AMB and itraconazole (ITZ, in filamentous fungi. The sakAΔ mutant of Aspergillus fumigatus, a mitogen-activated protein kinase (MAPK gene deletion mutant in the antioxidation system, was found to be more sensitive to AMB or ITZ than other A. fumigatus strains, a wild type and a mpkCΔ mutant (MAPK gene deletion mutant in polyalcohol sugar utilization system. The sakAΔ mutant showed no growth at 0.5 μg mL-1 of ITZ or reduced growth at 1.0 to 2.0 μg mL-1 of AMB, while the other strains exhibited robust growth. Complete fungal kill (≥ 99.9% by ITZ or AMB was achieved by much lower dosages for the sakAΔ mutant than for the other strains. SakA and MpkC appear to have overlapping roles in marshalling the oxidative stress response under treatment by an organic peroxide, tert-butyl hydroperoxide (t-BuOOH, or hydrogen peroxide (H2O2. The SakA signalling pathway was found to be responsible for fungal tolerance to AMB or ITZ toxicity. It appears msnA, an Aspergillus ortholog to Saccharomyces cerevisiae MSN2 (encoding a stress-responsive C2H2-type zinc-finger regulator and sakA and/or mpkC (upstream MAPKs are in the same stress response network under t-BuOOH-, H2O2- or AMB-triggered toxicity. Of note is that ITZ-sensitive yeast pathogens (Candida krusei and Cryptococcus neoformans were also sensitive to t-BuOOH, showing a connection between ITZ toxicity and oxidative stress response. This was shown by enhanced antifungal activity of AMB or ITZ when co-applied with redox-potent natural compounds, 2,3-dihydroxybenzaldehyde, thymol or salicylaldehyde, as chemosensitizing agents. Hence, redox compounds, which target the antioxidation system in fungi, possess a potent chemosensitizing capacity to enhance efficacy of conventional drugs inducing oxidative stress. Such chemosensitization can reduce costs and alleviate negative side effects associated with current

  10. Melatonin sensitizes human cervical cancer HeLa cells to cisplatin-induced cytotoxicity and apoptosis: effects on oxidative stress and DNA fragmentation.

    Science.gov (United States)

    Pariente, Roberto; Pariente, José A; Rodríguez, Ana B; Espino, Javier

    2016-01-01

    Melatonin has antitumor activity via several mechanisms including its antiproliferative and pro-apoptotic effects as well as its potent antioxidant actions, although recent evidence has indicated that melatonin may perform pro-oxidant actions in tumor cells. Therefore, melatonin may be useful in the treatment of tumors in association with chemotherapy drugs. This study was intended to evaluate the in vitro effect of melatonin on the cytotoxic and pro-apoptotic actions of various chemotherapeutic agents in cervical cancer HeLa cells. Herein, we found that both melatonin and three of the chemotherapeutic drugs tested, namely cisplatin (CIS), 5-fluorouracil (5-FU), and doxorubicin, induced a decrease in HeLa cell viability. Furthermore, melatonin significantly increased the cytotoxic effect of such chemotherapeutic agents. Consistently, costimulation of HeLa cells with any chemotherapeutic agent in the presence of melatonin further increased caspase-3 activation, particularly in CIS- and 5-FU-challenged cells. Likewise, concomitant treatments with melatonin and CIS significantly enhanced the ratio of cells entering mitochondrial apoptosis due to reactive oxygen species (ROS) overproduction, substantially augmented the population of apoptotic cells, and markedly enlarged DNA fragmentation compared to the treatments with CIS alone. Nonetheless, melatonin only displayed moderate chemosensitizing effects in 5-FU-stimulated HeLa cells, as suggested by slight increments in the percentage of cells stimulated for ROS production and in the proportion of early apoptotic cells compared to the treatments with 5-FU alone. In summary, our findings provided evidence that in vitro melatonin strongly enhances CIS-induced cytotoxicity and apoptosis in HeLa cells and, hence, the indoleamine could be potentially applied to cervical cancer treatment as a powerful synergistic agent. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. Role of Smac in apoptosis of lung cancer cells A549 induced by Taxol.

    Science.gov (United States)

    Zhang, Ying; Hao, Yingtao; Sun, Qifeng; Peng, Chuanliang

    2015-01-01

    A series of structurally unique second mitochondria-derived activators of caspase (Smac) that act as antagonists of inhibitor of apoptosis proteins (IAPs) directly have been discovered and have been shown to promote chemotherapy-induced apoptosis. In this study, we investigate the role of Smac in Taxol-induced apoptosis of lung cancer cell in vitro. PcDNA3.1/Smac recombinants were transfected into the non-small cell lung cancer cell line A549. Smac expression was detected by RT-PCR and Western blot. The invasive ability of cells was examined. Flow cytometry was used to analyze apoptosis of cells induced by Taxol with Annexin V/PI double staining technique. Smac expression was significantly higher in the PcDNA3.1/Smac recombinant group than in the untransfected group at mRNA and protein level (p Smac group. There were significant differences compared to the empty vector group and control group. The apoptosis rate was significantly higher in PcDNA3.1/Smac + Taxol group than in other groups (p Smac can enhance the chemosensitivity of the non-small cell lung cancer cell line A549 to Taxol.

  12. Chemosensitivity of irradiated resistant cells of multicellular spheroids in A549 lung adenocarcinoma

    International Nuclear Information System (INIS)

    Shi Degang; Shi Genming; Huang Gang

    2006-01-01

    Objective: To investigate the chemosensitivity of irradiated resistant cells of multicellular spheroids in A549 lung adenocarcinoma. Methods: The A549 irradiated resistant cells were the 10th regrowth generations after irradiated with 2.5 Gy of 6 MV X-ray, the control groups were A549 parent cells and MCFY/VCR resistant cells. The 6 kinds of chemotherapeutic drugs were DDP, VDS, 5-FU, HCP, MMC and ADM respectively, with verapamil (VPL) as reverse agent. The treatment effect was compared with MTT assay, and the multidrug resistant gene expressions of mdrl and MRP were measured with RT-PCR method. Results: A549 cells and irradiated resistant cells were resistant to DDP, but sensitivity to VDS,5-FU, HCP, MMC and ADM. The inhibitory rates of VPL to the above two cells were 98% and 25% respectively(P 2 -MG and MRP/β 2 -MG of all A549 cells were about 0 and 0.7 respectively, and those of MCFT/VCR cells were 35 and 4.36. Conclusion: The chemosensitivity of A549 irradiated resistant cells had not changed markedly, the decreased sensitivity to VPL could not be explained by the gene expression of mdrl and MRP. It is conferred that some kinds of changes in the cell membrane and decreased regrowth ability to result in resistance. Unlike multidrug resistance induced by chemotherapy, VPL may be not an ideal reverser to irradiated resistant cells. The new kinds of biological preparation should be sought to combine chemotherapy to treat recurring tumor with irradiated resistance. (authors)

  13. 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

  14. miRNAs associated with chemo-sensitivity in cell lines and in advanced bladder cancer

    DEFF Research Database (Denmark)

    Nordentoft, Iver; Birkenkamp-Demtroder, Karin; Agerbæk, Mads

    2012-01-01

    Background MicroRNA is a naturally occurring class of non-coding RNA molecules that mediate posttranscriptional gene regulation and are strongly implicated in cellular processes such as cell proliferation, carcinogenesis, cell survival and apoptosis. Consequently there is increasing focus on mi...... guidance. Methods We profiled the expression of 671 miRNAs in formalin fixed paraffin embedded tumors from patients with advanced bladder cancer treated with cisplatin based chemotherapy. We delineated differentially expressed miRNAs in tumors from patients with complete response vs. patients...... identified 15 miRNAs that correlated with response to chemotherapy and 5 miRNAs that correlated with survival time. Three miRNAs were associated with both response and survival (886-3p, 923, 944). By changing the cellular level of the response-identified miRNAs in eight bladder cell lines with different...

  15. Apoptosis in mammalian oocytes: a review.

    Science.gov (United States)

    Tiwari, Meenakshi; Prasad, Shilpa; Tripathi, Anima; Pandey, Ashutosh N; Ali, Irfan; Singh, Arvind K; Shrivastav, Tulsidas G; Chaube, Shail K

    2015-08-01

    Apoptosis causes elimination of more than 99% of germ cells from cohort of ovary through follicular atresia. Less than 1% of germ cells, which are culminated in oocytes further undergo apoptosis during last phases of oogenesis and depletes ovarian reserve in most of the mammalian species including human. There are several players that induce apoptosis directly or indirectly in oocytes at various stages of meiotic cell cycle. Premature removal of encircling granulosa cells from immature oocytes, reduced levels of adenosine 3',5'-cyclic monophosphate and guanosine 3',5'-cyclic monophosphate, increased levels of calcium (Ca(2+)) and oxidants, sustained reduced level of maturation promoting factor, depletion of survival factors, nutrients and cell cycle proteins, reduced meiotic competency, increased levels of proapoptotic as well as apoptotic factors lead to oocyte apoptosis. The BH3-only proteins also act as key regulators of apoptosis in oocyte within the ovary. Both intrinsic (mitochondria-mediated) as well as extrinsic (cell surface death receptor-mediated) pathways are involved in oocyte apoptosis. BID, a BH3-only protein act as a bridge between both apoptotic pathways and its cleavage activates cell death machinery of both the pathways inside the follicular microenvironment. Oocyte apoptosis leads to the depletion of ovarian reserve that directly affects reproductive outcome of various mammals including human. In this review article, we highlight some of the important players and describe the pathways involved during oocyte apoptosis in mammals.

  16. MicroRNA-137 inhibits tumor growth and sensitizes chemosensitivity to paclitaxel and cisplatin in lung cancer

    Science.gov (United States)

    Ge, Xin; Jiang, Cheng-Fei; Shi, Zhu-Mei; Li, Dong-Mei; Liu, Wei-Tao; Yu, Xiaobo; Shu, Yong-Qian

    2016-01-01

    Chemotherapy resistance frequently drives tumour progression. However, the underlying molecular mechanisms are poorly characterized. In this study, we explored miR-137's role in the chemosensitivity of lung cancer. We found that the expression level of miR-137 is down-regulated in the human lung cancer tissues and the resistant cells strains: A549/paclitaxel(A549/PTX) and A549/cisplatin (A549/CDDP) when compared with lung cancer A549 cells. Moreover, we found that overe-expression of miR-137 inhibited cell proliferation, migration, cell survival and arrest the cell cycle in G1 phase in A549/PTX and A549/CDDP. Furthermore, Repression of miR-137 significantly promoted cell growth, migration, cell survival and cell cycle G1/S transition in A549 cells. We further demonstrated that the tumor suppressive role of miR-137 was mediated by negatively regulating Nuclear casein kinase and cyclin-dependent kinase substrate1(NUCKS1) protein expression. Importantly, miR-137 inhibits A549/PTX, A549/CDDP growth and angiogenesis in vivo. Our study is the first to identify the tumor suppressive role of over-expressed miR-137 in chemosensitivity. Identification of a novel miRNA-mediated pathway that regulates chemosensitivity in lung cancer will facilitate the development of novel therapeutic strategies in the future. PMID:26989074

  17. Response prediction to neoadjuvant chemotherapy: comparison between pre-therapeutic gene expression profiles and in vitro chemosensitivity assay.

    Directory of Open Access Journals (Sweden)

    Christian F Singer

    Full Text Available Although the use of (neo-adjuvant chemotherapy in breast cancer patients has resulted in improved outcome, not all patients benefit equally. We have evaluated the utility of an in vitro chemosensitivity assay in predicting response to neoadjuvant chemotherapy. Pre-therapeutic biopsies were obtained from 30 breast cancer patients assigned to neoadjuvant epirubicin 75 mg/m2 and docetaxel 75 mg/m2 (Epi/Doc in a prospectively randomized clinical trial. Biopsies were subjected to a standardized ATP-based Epi/Doc chemosensitivity assay, and to gene expression profiling. Patients then received 3 cycles of chemotherapy, and response was evaluated by changes in tumor diameter and Ki67 expression. The efficacy of Epi/Doc in vitro was correlated with differential changes in tumor cell proliferation in response to Epi/Doc in vivo (p = 0.0011; r = 0.73670, Spearmańs rho, but did not predict for changes in tumor size. While a pre-therapeutic gene expression signature identified tumors with a clinical response to Epi/Doc, no such signature could be found for tumors that responded to Epi/Doc in vitro, or tumors in which Epi/Doc exerted an antiproliferative effect in vivo. This is the first prospective clinical trial to demonstrate the utility of a standardized in vitro chemosensitivity assay in predicting the individual biological response to chemotherapy in breast cancer.

  18. [Effect of MTRR gene on apoptosis and autophagy pathways in multiresistant epithelial ovarian cancer].

    Science.gov (United States)

    Chen, J; Wang, Q; Zhang, W; Li, L

    2016-04-25

    SKOV3/DDP cells[(14.8±2.4)%,(14.2±2.4)%; all Pcisplatin. Immunofluorescence tests revealed that the aggregates of LC3B in SKOV3/DDP-MTRRi cells were more than that of SKOV3/DDP-NC cells and SKOV3/DDP cells. The expression of LC3B of SKOV3/DDP-MTRRi cells was lower than those of SKOV3/DDP-NC cells and SKOV3/DDP cells(Pcisplatin group was higher than those of other 3 group cells(1.72±0.08, 1.43±0.04, 1.37±0.11, and 1.11±0.09; Pcisplatin group was significant decreased(0.58 ± 0.10, 0.94 ± 0.12, 1.21 ± 0.11, and 1.57 ± 0.10; Pcisplatin group was higher than that of cisplatin group[(0.78±0.03)% vs(0.62±0.03)%; P=0.018], the apoptosis rate was significant decreased in rapamycin+ cisplatin group[(59.0 ± 3.9)% vs(40.4 ± 3.0)%, P=0.019].(3)The 3 groups cells(SKOV3/DDP, SKOV3/DDP-NC, and SKOV3/DDP-MTRRi)induced by a certain concentration of cisplatin(4 μg/ml)after 48 hours, the expression of Bax in 3 groups cell were not evidently changed(P= 0.661). The expression of Bcl-2 was significantly decreased in SKOV3/DDP-MTRRi cells(P=0.030). The expression of caspase-3, caspase-7, caspase-9, and poly(ADP-ribose)polymerase(PARP)were not evidently changed(P>0.05), but cleaved caspase-3, cleaved caspase-7, cleaved caspase-9, and cleaved PARP were significantly increased in SKOV3/DDP-MTRRi cells(Pcisplatin-induced apoptosis and reduce the autophagy induced by cisplatin in SKOV3/DDP cells. Down-regulation of MTRR enhanced the chemosensitivity of cisplatinresistant ovarian cancer cells may be by activating caspase and Bcl-2 apoptosis family and inhibiting the PI3K/Akt autophagy pathway.

  19. Docosahexaenoic Acid Modulates a HER2-Associated Lipogenic Phenotype, Induces Apoptosis, and Increases Trastuzumab Action in HER2-Overexpressing Breast Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    Graziela Rosa Ravacci

    2015-01-01

    Full Text Available In breast cancer, lipid metabolic alterations have been recognized as potential oncogenic stimuli that may promote malignancy. To investigate whether the oncogenic nature of lipogenesis closely depends on the overexpression of HER2 protooncogene, the normal breast cell line, HB4a, was transfected with HER2 cDNA to obtain HER2-overexpressing HB4aC5.2 cells. Both cell lines were treated with trastuzumab and docosahexaenoic acid. HER2 overexpression was accompanied by an increase in the expression of lipogenic genes involved in uptake (CD36, transport (FABP4, and storage (DGAT of exogenous fatty acids (FA, as well as increased activation of “de novo” FA synthesis (FASN. We further investigate whether this lipogenesis reprogramming might be regulated by mTOR/PPARγ pathway. Inhibition of the mTORC1 pathway markers, p70S6 K1, SREBP1, and LIPIN1, as well as an increase in DEPTOR expression (the main inhibitor of the mTOR was detected in HB4aC5.2. Based on these results, a PPARγ selective antagonist, GW9662, was used to treat both cells lines, and the lipogenic genes remained overexpressed in the HB4aC5.2 but not HB4a cells. DHA treatment inhibited all lipogenic genes (except for FABP4 in both cell lines yet only induced death in the HB4aC5.2 cells, mainly when associated with trastuzumab. Neither trastuzumab nor GW9662 alone was able to induce cell death. In conclusion, oncogenic transformation of breast cells by HER2 overexpression may require a reprogramming of lipogenic genetic that is independent of mTORC1 pathway and PPARγ activity. This reprogramming was inhibited by DHA.

  20. Distinct functional domains of PNMA5 mediate protein-protein interaction, nuclear localization, and apoptosis signaling in human cancer cells.

    Science.gov (United States)

    Lee, Yong Hoi; Pang, Siew Wai; Poh, Chit Laa; Tan, Kuan Onn

    2016-09-01

    Members of paraneoplastic Ma (PNMA) family have been identified as onconeuronal antigens, which aberrant expressions in cancer cells of patients with paraneoplastic disorder (PND) are closely linked to manifestation of auto-immunity, neuro-degeneration, and cancer. The purpose of present study was to determine the role of PNMA5 and its functional relationship to MOAP-1 (PNMA4) in human cancer cells. PNMA5 mutants were generated through deletion or site-directed mutagenesis and transiently expressed in human cancer cell lines to investigate their role in apoptosis, subcellular localization, and potential interaction with MOAP-1 through apoptosis assays, fluorescence microscopy, and co-immunoprecipitation studies, respectively. Over-expressed human PNMA5 exhibited nuclear localization pattern in both MCF-7 and HeLa cells. Deletion mapping and mutagenesis studies showed that C-terminus of PNMA5 is responsible for nuclear localization, while the amino acid residues (391KRRR) within the C-terminus of PNMA5 are required for nuclear targeting. Deletion mapping and co-immunoprecipitation studies showed that PNMA5 interacts with MOAP-1 and N-terminal domain of PNMA5 is required for interaction with MOAP-1. Furthermore, co-expression of PNMA5 and MOAP-1 in MCF-7 cells significantly enhanced chemo-sensitivity of MCF-7 to Etoposide treatment, indicating that PNMA5 and MOAP-1 interact synergistically to promote apoptotic signaling in MCF-7 cells. Our results show that PNMA5 promotes apoptosis signaling in HeLa and MCF-7 cells and interacts synergistically with MOAP-1 through its N-terminal domain to promote apoptosis and chemo-sensitivity in human cancer cells. The C-terminal domain of PNMA5 is required for nuclear localization; however, both N-and C-terminal domains of PNMA5 appear to be required for pro-apoptotic function.

  1. Pandemic Influenza A (H1N1) Virus Infection Increases Apoptosis and HIV-1 Replication in HIV-1 Infected Jurkat Cells.

    Science.gov (United States)

    Wang, Xue; Tan, Jiying; Biswas, Santanu; Zhao, Jiangqin; Devadas, Krishnakumar; Ye, Zhiping; Hewlett, Indira

    2016-02-02

    Influenza virus infection has a significant impact on public health, since it is a major cause of morbidity and mortality. It is not well-known whether influenza virus infection affects cell death and human immunodeficiency virus (HIV)-1 replication in HIV-1-infected patients. Using a lymphoma cell line, Jurkat, we examined the in vitro effects of pandemic influenza A (H1N1) virus (pH1N1) infection on cell death and HIV-1 RNA production in infected cells. We found that pH1N1 infection increased apoptotic cell death through Fas and Bax-mediated pathways in HIV-1-infected Jurkat cells. Infection with pH1N1 virus could promote HIV-1 RNA production by activating host transcription factors including nuclear factor kappa-light-chain-enhancer of activated B cells (NF-ĸB), nuclear factor of activated T-cells (NFAT) and activator protein 1 (AP-1) through mitogen-activated protein kinases (MAPK) pathways and T-cell antigen receptor (TCR)-related pathways. The replication of HIV-1 latent infection could be reactivated by pH1N1 infection through TCR and apoptotic pathways. These data indicate that HIV-1 replication can be activated by pH1N1 virus in HIV-1-infected cells resulting in induction of cell death through apoptotic pathways.

  2. Apoptosis of neutrophils

    NARCIS (Netherlands)

    Maianski, N. A.; Maianski, A. N.; Kuijpers, T. W.; Roos, D.

    2004-01-01

    Regulation of the neutrophil life span by apoptosis provides a fine balance between their function as effector cells of host defense and a safe turnover of these potentially harmful cells. Alterations of neutrophil apoptosis are associated with a number of diseases. As do other cell types,

  3. Hyperthermia-induced apoptosis

    NARCIS (Netherlands)

    Nijhuis, E.H.A.

    2008-01-01

    This thesis describes a number of studies that investigated several aspects of heat-induced apoptosis in human lymphoid malignancies. Cells harbour both pro- and anti-apoptotic proteins and the balance between these proteins determines whether a cell is susceptible to undergo apoptosis. In this

  4. Neonatal maturation of the hypercapnic ventilatory response and central neural CO2 chemosensitivity.

    Science.gov (United States)

    Putnam, Robert W; Conrad, Susan C; Gdovin, M J; Erlichman, Joseph S; Leiter, J C

    2005-11-15

    The ventilatory response to CO2 changes as a function of neonatal development. In rats, a ventilatory response to CO2 is present in the first 5 days of life, but this ventilatory response to CO2 wanes and reaches its lowest point around postnatal day 8. Subsequently, the ventilatory response to CO2 rises towards adult levels. Similar patterns in the ventilatory response to CO2 are seen in some other species, although some animals do not exhibit all of these phases. Different developmental patterns of the ventilatory response to CO2 may be related to the state of development of the animal at birth. The triphasic pattern of responsiveness (early decline, a nadir, and subsequent achievement of adult levels of responsiveness) may arise from the development of several processes, including central neural mechanisms, gas exchange, the neuromuscular junction, respiratory muscles and respiratory mechanics. We only discuss central neural mechanisms here, including altered CO2 sensitivity of neurons among the various sites of central CO2 chemosensitivity, changes in astrocytic function during development, the maturation of electrical and chemical synaptic mechanisms (both inhibitory and excitatory mechanisms) or changes in the integration of chemosensory information originating from peripheral and multiple central CO2 chemosensory sites. Among these central processes, the maturation of synaptic mechanisms seems most important and the relative maturation of synaptic processes may also determine how plastic the response to CO2 is at any particular age.

  5. In vitro effects of imatinib mesylate on radiosensitivity and chemosensitivity of breast cancer cells

    International Nuclear Information System (INIS)

    Weigel, Marion T; Maass, Nicolai; Mundhenke, Christoph; Dahmke, Linda; Schem, Christian; Bauerschlag, Dirk O; Weber, Katrin; Niehoff, Peter; Bauer, Maret; Strauss, Alexander; Jonat, Walter

    2010-01-01

    Breast cancer treatment is based on a combination of adjuvant chemotherapy followed by radiotherapy effecting intracellular signal transduction. With the tyrosine kinase inhibitors new targeted drugs are available. Imatinib mesylate is a selective inhibitor of bcr-abl, PRGFR alpha, beta and c-kit. The purpose of this study was to determine whether Imatinib has an influence on the effectiveness of radiotherapy in breast cancer cell lines and if a combination of imatinib with standard chemotherapy could lead to increased cytoreduction. Colony-forming tests of MCF 7 and MDA MB 231 were used to study differences in cell proliferation under incubation with imatinib and radiation. Changes in expression and phosphorylation of target receptors were detected using western blot. Cell proliferation, migration and apoptosis assays were performed combining imatinib with doxorubicin. The combination of imatinib and radiotherapy showed a significantly stronger inhibition of cell proliferation compared to single radiotherapy. Differences in PDGFR expression could not be detected, but receptor phosphorylation was significantly inhibited when treated with imatinib. Combination of imatinib with standard chemotherapy lead to an additive effect on cell growth inhibition compared to single treatment. Imatinib treatment combined with radiotherapy leads in breast cancer cell lines to a significant benefit which might be influenced through inhibition of PDGFR phosphorylation. Combining imatinib with chemotherapy enhances cytoreductive effects. Further in vivo studies are needed to evaluate the benefit of Imatinib in combination with radiotherapy and chemotherapy on the treatment of breast cancer

  6. JS-K, a GST-activated nitric oxide donor prodrug, enhances chemo-sensitivity in renal carcinoma cells and prevents cardiac myocytes toxicity induced by Doxorubicin.

    Science.gov (United States)

    Qiu, Mingning; Ke, Longzhi; Zhang, Sai; Zeng, Xin; Fang, Zesong; Liu, Jianjun

    2017-08-01

    Doxorubicin, a highly effective and widely used anthracycline antibiotic in multiple chemotherapy regimens, has been limited by its cardiotoxicity. The aim of this study is to investigate the effect of nitric oxide donor prodrug JS-K on proliferation and apoptosis in renal carcinoma cells and cardiac myocytes toxicity induced by Doxorubicin and to explore possible p53-related mechanism in renal carcinoma cells. The effect of JS-K on anti-cancer activity of Doxorubicin was investigated in renal carcinoma cells via detecting cell proliferation, cytotoxicity, cell death and apoptosis and expressions of apoptotic-related proteins. Effect of p53 on the combination of JS-K and Doxorubicin was determined using p53 inhibitor Pifithrin-α and p53 activator III. Furthermore, the effect of JS-K on cardiac myocytes toxicity of Doxorubicin was investigated in H9c2 (2-1) cardiac myocytes via measuring cell growth, cell death and apoptosis, expressions of proteins involved in apoptosis and intracellular reactive oxygen species. We demonstrated that JS-K could increase Doxorubicin-induced renal carcinoma cell growth suppression and apoptosis and could increase expressions of proteins that are involved in apoptosis. Additionally, Pifithrin-α reversed the promoting effect of JS-K on Doxorubicin-induced renal carcinoma cell apoptosis; conversely, the p53 activator III exacerbated the promoting effect of JS-K on Doxorubicin-induced renal carcinoma cell apoptosis. Furthermore, JS-K protected H9c2 (2-1) cardiac myocytes against Doxorubicin-induced toxicity and decreased Doxorubicin-induced reactive oxygen species production. JS-K enhances the anti-cancer activity of Doxorubicin in renal carcinoma cells by upregulating p53 expression and prevents cardiac myocytes toxicity of Doxorubicin by decreasing oxidative stress.

  7. Caspases: An apoptosis mediator

    Directory of Open Access Journals (Sweden)

    Tapan Kumar Palai

    2015-03-01

    Full Text Available The process of programmed cell death, or apoptosis, is generally characterized by distinct morphological characteristics and energy - dependent biochemical mechanisms. Apoptosis is a widely conserved phenomenon helping many processes, including normal cell turnover, proper development and functioning of the immune system, hormone dependent atrophy etc. Inappropriate apoptosis (either low level or high level leads to many developmental abnormalities like, neurodegenerative diseases, ischemic damage, autoimmune disorders and many types of cancer. To use cells for therapeutic purposes through generating cell lines, it is critical to study the cell cycle machinery and signalling pathways that controls cell death and apoptosis. Apoptotic pathways provide a fundamental protective mechanism that decreases cellular sensitivity to damaging events and allow proper developmental process in multi-cellular organisms. Major mediator of apoptosis is a family of proteins known as caspases. There are mainly fourteen types of caspases but out of them only ten caspasese have got essential role in controlling the process of apoptosis. These ten caspases have been categorized into either initiator caspases (caspase 2, 8, 9, 10 or executioner caspases (caspase 3, 6, 7. Although various types of caspases have been identified so far, the exact mechanisms of action of these groups of proteins is still to be fully understood. The aim of this review is to provide a detail overview of role of different caspases in regulating the process of apoptosis.

  8. [Astragalus polysaccharide may increase sensitivity of cervical cancer HeLa cells to cisplatin by regulating cell autophagy].

    Science.gov (United States)

    Zhai, Qiu-Li; Hu, Xiang-Dan; Xiao, Jing; Yu, Dong-Qing

    2018-02-01

    This study aimed to investigate the possible sensitivity of Astragalus polysaccharides, in order to improve the chemosensitivity of cervical cancer HeLa cells to cisplatin by regulating the cell autophagy, and explore its possible mechanism. In this study, HeLa cells were divided into control group, cisplatin group, Astragalus polysaccharide group, and Astragalus polysaccharide combined with cisplatin group. MTT assay was used to detect the proliferation of cervical cancer HeLa cells. Flow cytometry was used to detect the apoptosis and cycle of HeLa cells in each experimental group. RT-PCR was used to detect the mRNA expression of autophagy-related proteins beclin1, LC3Ⅱ and p62. The expression levels of autophagy-related proteins beclin1, LC3Ⅱ, LC3Ⅰ and p62 were detected by WB method. MTT results showed that compared with the control group, the proliferation of HeLa cells was significantly inhibited in each administration group( P HeLa cells was significantly increased( P HeLa cells to cisplatin by regulating the cell autophagy. Its possible mechanism of action is correlated with the up-regulation of autophagy-related proteins beclin1, the promote the conversion from LC3Ⅰ to LC3Ⅱ, the down-regulation of labeled protein p62, and the enhancement of HeLa cell autophagic activity, thereby increasing the sensitivity of HeLa cells to cisplatin chemotherapy. Copyright© by the Chinese Pharmaceutical Association.

  9. Chemo-sensitivity of Two-dimensional Monolayer and Three-dimensional Spheroid of Breast Cancer MCF-7 Cells to Daunorubicin, Docetaxel, and Arsenic Disulfide.

    Science.gov (United States)

    Uematsu, Nami; Zhao, Yuxue; Kiyomi, Anna; Yuan, B O; Onda, Kenji; Tanaka, Sachiko; Sugiyama, Kentaro; Sugiura, Munetoshi; Takagi, Norio; Hayakawa, Akemi; Hirano, Toshihiko

    2018-04-01

    Chemo-sensitivity of two-dimensional (2D) monolayers and three-dimensional (3D) spheroids of human breast cancer MCF-7 cells were investigated. MCF-7 cells were cultured in monolayers or spheroids established using a thermo-reversible gelatin polymer, in the presence of daunorubicin, docetaxel, or As 2 S 2 Cell proliferation was examined by a Cell Counting Kit-8 assay. Daunorubicin, docetaxel, and As 2 S 2 dose-dependently decreased the MCF-7 cell proliferation in both 2D- and 3D-culture systems. The 3D spheroids were less sensitive to these agents than the 2D cultured cells. Verapamil, an inhibitor of P-glycoprotein, partially enhanced the antiproliferative effects of the agents. DL-buthionine-(S, R)-sulfoximine significantly increased (p<0.05), while N-acetyl-L-cysteine significantly inhibited the antiproliferative effects of As 2 S 2 (p<0.003). The 3D spheroids showed less sensitivity to the antiprolliferative efficacies of anticancer agents than the 2D cultured cells. P-Glycoprotein is suggested to be partially implicated in drug resistance. Reduction of cellular glutathione level enhanced the As 2 S 2 cytotoxicity. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  10. Suppression of STAT3 NH2-terminal domain chemosensitizes medulloblastoma cells by activation of protein inhibitor of activated STAT3 via de-repression by microRNA-21.

    Science.gov (United States)

    Ray, Sutapa; Coulter, Don W; Gray, Shawn D; Sughroue, Jason A; Roychoudhury, Shrabasti; McIntyre, Erin M; Chaturvedi, Nagendra K; Bhakat, Kishor K; Joshi, Shantaram S; McGuire, Timothy R; Sharp, John G

    2018-04-01

    Medulloblastoma (MB) is a malignant pediatric brain tumor with poor prognosis. Signal transducers and activators of transcription-3 (STAT3) is constitutively activated in MB where it functions as an oncoprotein, mediating cancer progression and metastasis. Here, we have delineated the functional role of activated STAT3 in MB, by using a cell permeable STAT3-NH 2 terminal domain inhibitor (S3-NTDi) that specifically perturbs the structure/function of STAT3. We have implemented several biochemical experiments using human MB tumor microarray (TMA) and pediatric MB cell lines, derived from high-risk SHH-TP53-mutated and MYC-amplified Non-WNT/SHH tumors. Treatment of MB cells with S3-NTDi leads to growth inhibition, cell cycle arrest, and apoptosis. S3-NTDi downregulated expression of STAT3 target genes, delayed migration of MB cells, attenuated epithelial-mesenchymal transition (EMT) marker expressions and reduced cancer stem-cell associated protein expressions in MB-spheres. To elucidate mechanisms, we showed that S3-NTDi induce expression of pro-apoptotic gene, C/EBP-homologous protein (CHOP), and decrease association of STAT3 to the proximal promoter of CCND1 and BCL2. Of note, S3-NTDi downregulated microRNA-21, which in turn, de-repressed Protein Inhibitor of Activated STAT3 (PIAS3), a negative regulator of STAT3 signaling pathway. Furthermore, combination therapy with S3-NTDi and cisplatin significantly decreased highly aggressive MYC-amplified MB cell growth and induced apoptosis by downregulating STAT3 regulated proliferation and anti-apoptotic gene expression. Together, our results revealed an important role of STAT3 in regulating MB pathogenesis. Disruption of this pathway with S3-NTDi, therefore, may serves as a promising candidate for targeted MB therapy by enhancing chemosensitivity of MB cells and potentially improving outcomes in high-risk patients. © 2017 Wiley Periodicals, Inc.

  11. Apoptosis in Pneumovirus Infection

    Directory of Open Access Journals (Sweden)

    Reinout A. Bem

    2013-01-01

    Full Text Available Pneumovirus infections cause a wide spectrum of respiratory disease in humans and animals. The airway epithelium is the major site of pneumovirus replication. Apoptosis or regulated cell death, may contribute to the host anti-viral response by limiting viral replication. However, apoptosis of lung epithelial cells may also exacerbate lung injury, depending on the extent, the timing and specific location in the lungs. Differential apoptotic responses of epithelial cells versus innate immune cells (e.g., neutrophils, macrophages during pneumovirus infection can further contribute to the complex and delicate balance between host defense and disease pathogenesis. The purpose of this manuscript is to give an overview of the role of apoptosis in pneumovirus infection. We will examine clinical and experimental data concerning the various pro-apoptotic stimuli and the roles of apoptotic epithelial and innate immune cells during pneumovirus disease. Finally, we will discuss potential therapeutic interventions targeting apoptosis in the lungs.

  12. Physician Education: Apoptosis.

    Science.gov (United States)

    Kataoka; Tsuruo

    1996-01-01

    We have come to understand apoptosis as not merely a single form of cell death, but as a fundamental theme in cell biology that has far-reaching implications in the fields of physiology and pathology. At the present time, however, the mechanism of apoptosis is not clearly understood, as research into apoptosis is still at the initial stages. Nevertheless, the links between apoptosis and a variety of pathological conditions are gradually becoming clearer. In this article, we will provide a simple explanation of apoptosis and its mechanism as a novel concept of cell death and discuss the way in which apoptosis has been linked to a variety of pathological conditions. WHAT IS APOPTOSIS?: In normal tissue, cells that are no longer needed are rapidly eliminated without affecting the overall function of the tissue. In this process cells undergo an active and spontaneous suicide called programmed cell death. In fact, the majority of physiological cell deaths take the form of apoptosis. The word apoptosis is used, in contrast to necrosis, to describe the situation in which a cell actively pursues a course toward death upon receiving certain stimuli [1]. The morphological changes of apoptosis found in most cell types first involve contraction in cell volume and condensation of the nucleus. When this happens the intracellular organelles such as the mitochondria retain their normal morphology. As apoptosis proceeds, blebbing of the plasma membrane occurs, and the nucleus becomes fragmented. Finally, the cell itself fragments to form apoptotic bodies that are engulfed by nearby phagocytes. With respect to biochemical changes, it is known that the chromosomes become fragmented into nucleosome units, and DNA forms characteristic ladder patterns when subjected to agarose gel electrophoresis. MECHANISM OF APOPTOSIS: It has been reported that apoptosis is induced in various cells by many kinds of irritations, but the precise mechanism is still unclear. Cell injuries that induce

  13. Reaper-Induced Apoptosis

    National Research Council Canada - National Science Library

    Perry, Jennifer

    2005-01-01

    Reaper is a central regulator of apoptosis in the fly, Drosophila melanogaster. At the start of this proposal our laboratory identified what was believed to be a pro-apoptotic human homolog of Reaper...

  14. Viruses and apoptosis.

    Science.gov (United States)

    Roulston, A; Marcellus, R C; Branton, P E

    1999-01-01

    Successful viral replication requires not only the efficient production and spread of progeny, but also evasion of host defense mechanisms that limit replication by killing infected cells. In addition to inducing immune and inflammatory responses, infection by most viruses triggers apoptosis or programmed cell death of the infected cell. This cell response often results as a compulsory or unavoidable by-product of the action of critical viral replicative functions. In addition, some viruses seem to use apoptosis as a mechanism of cell killing and virus spread. In both cases, successful replication relies on the ability of certain viral products to block or delay apoptosis until sufficient progeny have been produced. Such proteins target a variety of strategic points in the apoptotic pathway. In this review we summarize the great amount of recent information on viruses and apoptosis and offer insights into how this knowledge may be used for future research and novel therapies.

  15. The elevation of intraocular pressure is associated with apoptosis and increased immunoreactivity for nitric oxide synthase in rat retina whereas the effectiveness of retina derived relaxing factor is unaffected.

    Science.gov (United States)

    Takır, Selçuk; Gürel-Gürevin, Ebru; Toprak, Ayça; Demirci-Tansel, Cihan; Uydeş-Doğan, B Sönmez

    2016-04-01

    rat retina particularly in the ganglion and inner layers that is associated with marked apoptosis and increased immunoreactivity for NOS, while the functional effectiveness of retina derived relaxing factor, i.e., RRF is unaffected. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Iron dysregulation combined with aging prevents sepsis-induced apoptosis.

    Science.gov (United States)

    Javadi, Pardis; Buchman, Timothy G; Stromberg, Paul E; Turnbull, Isaiah R; Vyas, Dinesh; Hotchkiss, Richard S; Karl, Irene E; Coopersmith, Craig M

    2005-09-01

    Sepsis, iron loading, and aging cause independent increases in gut epithelial and splenic apoptosis. It is unknown how their combination will affect apoptosis and systemic cytokine levels. Hfe-/- mice (a murine homologue of hemochromatosis) abnormally accumulate iron in their tissues. Aged (24-26 months) or mature (16-18 months) Hfe-/- mice and wild type (WT) littermates were subjected to cecal ligation and puncture (CLP) or sham laparotomy. Intestine, spleen, and blood were harvested 24 h later and assessed for apoptosis and cytokine levels. Gut epithelial and splenic apoptosis were low in both aged septic and sham Hfe-/- mice, regardless of the amount of iron in their diet. Mature septic WT mice had increased apoptosis compared to age-matched sham WT mice. Mature septic Hfe-/- mice had similar levels of intestinal cell death to age-matched septic WT mice but higher levels of splenic apoptosis. Apoptosis was significantly lower in septic aged Hfe-/- mice than septic mature Hfe-/- animals. Interleukin-6 was elevated in septic aged Hfe-/- mice compared to sham mice. Although sepsis, chronic iron dysregulation, and aging each increase gut and splenic apoptosis, their combination yields cell death levels similar to sham animals despite the fact that aged Hfe-/- mice are able to mount an inflammatory response following CLP and mature Hfe-/- mice have elevated sepsis-induced apoptosis. Combining sepsis with two risk factors that ordinarily increase cell death and increase mortality in CLP yields an apoptotic response that could not have been predicted based upon each element in isolation.

  17. [Depression and treatment. Apoptosis, neuroplasticity and antidepressants].

    Science.gov (United States)

    Arantes-Gonçalves, Filipe; Coelho, Rui

    2006-01-01

    Depression's neurobiology begins to be better understood. The last decade data considers neuroplasticity and stress as implicated factors on the pathophisiology of depression. Because antidepressants have a lag-time on their action it is possible that inhibition of neurotransmitters recaptation is not sufficient to explain long term changes. For that purpose, neurogenesis increase, nervous fibers sprouting, new synapses and stabilization of the old ones can be responsible for those changes. AMPc-MAPcinases-CREB-BDNF cellular cascade can play a significant role in the mechanisms of dendritic restructuration, hippocampal neurogenesis increase and nervous cells survival. The aim of this article is to discuss if apoptosis could play a key role as an ethiopathogenic factor on the patogenesis of depression. It was done a medline search for references with apoptosis, stress, neuroplasticity, depression and antidepressants key-words. It were found 101 original or review references about these subjects. Stress plays a key role in the etiopathogeny of depression. Its deletery effects on apoptosis and neuroplasticity can be changed by antidepressants. Neurogenesis' increase is necessary for their action. This increase is reached with chronic antidepressant treatment and not with other psychotropic drugs which means some pharmacological specificity of antidepressants. AMPc, CREB, BDNF and Bcl-2 can be considered as target genes in antidepressant synthesis. At the level of this neurotrophic factors apoptosis might be included in the neuroplastic model of depression and play a prominent role in etiopathogeny of depression. To confirm that, we need more research on the field to know which are the mechanisms that trigger apoptosis and its biological significance. In relation to the last one, we can say that is possible to be physiological apoptosis in deteriorated neurons death which cannot make strong connections and pathological apoptosis because of stress via, namely, HPA axis.

  18. Chemotherapy-Induced Apoptosis in a Transgenic Model of Neuroblastoma Proceeds Through p53 Induction

    Directory of Open Access Journals (Sweden)

    Louis Chesler

    2008-11-01

    Full Text Available Chemoresistance in neuroblastoma is a significant issue complicating treatment of this common pediatric solid tumor. MYCN-amplified neuroblastomas are infrequently mutated at p53 and are chemosensitive at diagnosis but acquire p53 mutations and chemoresistance with relapse. Paradoxically, Myc-driven transformation is thought to require apoptotic blockade. We used the TH-MYCN transgenic murine model to examine the role of p53-driven apoptosis on neuroblastoma tumorigenesis and the response to chemotherapy. Tumors formed with high penetrance and low latency in p53-haploinsufficient TH-MYCN mice. Cyclophosphamide (CPM induced a complete remission in p53 wild type TH-MYCN tumors, mirroring the sensitivity of childhood neuroblastoma to this agent. Treated tumors showed a prominent proliferation block, induction of p53 protein, and massive apoptosis proceeding through induction of the Bcl-2 homology domain-3-only proteins PUMA and Bim, leading to the activation of Bax and cleavage of caspase-3 and -9. Apoptosis induced by CPM was reduced in p53-haploinsufficient tumors. Treatment of MYCN-expressing human neuroblastoma cell lines with CPM induced apoptosis that was suppressible by siRNA to p53. Taken together, the results indicate that the p53 pathway plays a significant role in opposing MYCN-driven oncogenesis in a mouse model of neuroblastoma and that basal inactivation of the pathway is achieved in progressing tumors. This, in part, explains the striking sensitivity of such tumors to chemotoxic agents that induce p53-dependent apoptosis and is consistent with clinical observations that therapy-associated mutations in p53 are a likely contributor to the biology of tumors at relapse and secondarily mediate resistance to therapy.

  19. EZH2 and histone deacetylase inhibitors induce apoptosis in triple negative breast cancer cells by differentially increasing H3 Lys27 acetylation in the BIM gene promoter and enhancers.

    Science.gov (United States)

    Huang, Julia P; Ling, Kun

    2017-11-01

    Enhancer of zeste homolog 2 (EZH2), a subunit of polycomb repressive complex 2, is a histone methyl-transferase and is considered to work cooperatively with histone deacetylases (HDACs) in the same protein complex to mediate gene transcription repression by increasing histone H3 Lys 27 trimethylation (H3K27me3), in particular in the nucleosome (s). EZH2 is overexpressed in numerous types of cancer, including triple negative breast cancer (TNBC), a subtype of breast cancer, which there are no effective treatment options for. Thus, inhibition of EZH2 may be harnessed for targeted therapy of this disease. The present study demonstrated that co-treatment with an EZH2 inhibitor and a HDAC inhibitor additively induced apoptosis in two TNBC cell lines, namely MDA-MB-231 and MDA-MB-436. The increased rate of cell death was associated with an elevation of B cell lymphoma-2 like 11 (BIM) expression level, a pro-apoptotic protein at the protein and mRNA expression levels in these two cell lines. The expression of forkhead box O1 (FOXO1), a known upstream transcriptional activator of BIM , was upregulated in both cell lines by the HDAC inhibitor, and the effect was more pronounced in MDA-MB-436 cells with higher phosphorylation levels of protein kinase B, a negative regulator of FOXO1, compared with MDA-MB-231 cells. Conversely, FOXO1 expression was inhibited following treatment with the EZH2 inhibitor, suggesting that EZH2 and HDAC inhibitors induced BIM expression via a FOXO1-independent mechanism. The present study further revealed that the EZH2 inhibitor, but not the HDAC inhibitor, induced high levels of H3K27 acetylation (H3K27ac) in the BIM promoter. By contrast, compared with the effect of the EZH2 inhibitor, HDAC inhibitor treatment resulted in an increase in H3K27ac at two BIM enhancers. Collectively, the results of the present study indicated that EZH2 and HDACs act differentially on H3K27ac levels in the nucleosome at the promoter and enhancer regions of the BIM

  20. Apoptosis – is it good or bad?

    Directory of Open Access Journals (Sweden)

    Bakir Mehić

    2012-08-01

    induced by TNF-α. The over expression of antiapoptotic proteins may allow injured cells to survive, and autophagy may assist by providing critical metabolites. Apoptotic cells induce anergy or an immunosuppressive phenotype, whereas necrotic cells augment inflammation, in part by binding the receptor C-type lectin domain family 9 on dendritic cells.Clinical implications of apoptosis are consist in more than 50% of neoplasm’s that have defects in the apoptotic machinery mutations in the tumor-suppressor gene TP53, that is called the “guardian of the genome,” initiates apoptosis in response to DNA damage induced by radiation, chemical agents, oxidative stress, and other agents.[3] Abnormalities in apoptosis can increase susceptibility to autoimmune diseases.[4] There is growing evidence that neuronal apoptosis plays a key role in neonatal brain disorders.[5] Hepatocytes are particularly prone to apoptosis in response to various types of stress, including infections.[6] Necrosis predominates in ischemic injury, but often there are apoptotic cells in the hypoxic penumbra in myocardial infarction and stroke and in globally hypoxic zones after reperfusion injury. Sepsis is perhaps the most remarkable clinical setting in which apoptosis occurs. Massive apoptosis of immune effectors’ cells and gastrointestinal epithelial cells develops in patients with sepsis.[7] The profound loss of immune effectors’ cells in sepsis inhibits the ability of the immune system to eradicate the primary infection and renders the patient susceptible to nosocomial infections. Finally, why apoptosis is good? Because without apoptosis, 2 tons of bone marrow and lymph nodes and a 16-km intestine would probably accumulate in a human by the age of 8o.[8

  1. Apoptosis signaling and radiation protection

    International Nuclear Information System (INIS)

    Morita, Akinori; Suzuki, Norio; Hosoi, Yoshio

    2005-01-01

    Radiation protection by apoptosis control is the suppression of cell death in highly radiosensitive tissues. This paper describes the outline of radiation-induced apoptosis framework, apoptosis-concerned target molecules possibly related to apoptosis by radiation and their inhibitors. Although there are intrinsic (via mitochondria) and extrinsic (via death receptor) pathways in apoptosis, this review mainly mentions the former which is more important in radiation-induced apoptosis. Those molecules known at present in the apoptosis are caspase, Bcl-2 family and p53. Caspase, a group of cystein proteases, initiates apoptosis but its inhibition is known not always to result in apoptosis suppression, suggesting the existence of caspase-independent pathways. Bcl-2 family involves apoptosis-suppressing (possessing BH domains) and -promoting (lacking BH domains or possessing BH3 domain alone/BH3-only protein) groups. Two p53-transcription-dependent and one -independent pathways in p53-induced apoptosis are known and p53 can be a most possible target molecule since it positions at the start of apoptosis. Authors have found a vanadate inactivates p53. Inhibitors affecting upstream molecules of apoptosis will be the most useful candidate for apoptosis suppression/radiation protection. (S.I.) 106 refs

  2. Array of Chemosensitive Resistors with Composites of Gas Chromatography (GC) Materials and Carbon Black for Detection and Recognition of VOCs: A Basic Study

    Science.gov (United States)

    Wyszynski, Bartosz; Yatabe, Rui; Nakao, Atsuo; Nakatani, Masaya; Oki, Akio; Oka, Hiroaki; Toko, Kiyoshi

    2017-01-01

    Mimicking the biological olfaction, large odor-sensor arrays can be used to acquire a broad range of chemical information, with a potentially high degree of redundancy, to allow for enhanced control over the sensitivity and selectivity of artificial olfaction systems. The arrays should consist of the largest possible number of individual sensing elements while being miniaturized. Chemosensitive resistors are one of the sensing platforms that have a potential to satisfy these two conditions. In this work we test viability of fabricating a 16-element chemosensitive resistor array for detection and recognition of volatile organic compounds (VOCs). The sensors were fabricated using blends of carbon black and gas chromatography (GC) stationary-phase materials preselected based on their sorption properties. Blends of the selected GC materials with carbon black particles were subsequently coated over chemosensitive resistor devices and the resulting sensors/arrays evaluated in exposure experiments against vapors of pyrrole, benzenal, nonanal, and 2-phenethylamine at 150, 300, 450, and 900 ppb. Responses of the fabricated 16-element array were stable and differed for each individual odorant sample, proving the blends of GC materials with carbon black particles can be effectively used for fabrication of large odor-sensing arrays based on chemosensitive resistors. The obtained results suggest that the proposed sensing devices could be effective in discriminating odor/vapor samples at the sub-ppm level. PMID:28696353

  3. Liposomal curcumin alters chemosensitivity of breast cancer cells to Adriamycin via regulating microRNA expression.

    Science.gov (United States)

    Zhou, Siying; Li, Jian; Xu, Hanzi; Zhang, Sijie; Chen, Xiu; Chen, Wei; Yang, Sujin; Zhong, Shanliang; Zhao, Jianhua; Tang, Jinhai

    2017-07-30

    Emerging evidence suggests that curcumin can overcome drug resistance to classical chemotherapies, but poor bioavailability and low absorption have limited its clinical use and the mechanisms remain unclear. Also, Adriamycin (Adr) is one of the most active cytotoxic agents in breast cancer; however, the high resistant rate of Adr leads to a poor prognosis. We utilized encapsulation in liposomes as a strategy to improve the bioavailability of curcumin and demonstrated that liposomal curcumin altered chemosensitivity of Adr-resistant MCF-7 human breast cancer (MCF-7/Adr) by MTT assay. The miRNA and mRNA expression profiles of MCF-7/S, MCF-7/Adr and curcumin-treated MCF-7/Adr cells were analyzed by microarray and further confirmed by real-time PCR. We focused on differentially expressed miR-29b-1-5p to explore the involvement of miR-29b-1-5p in the resistance of Adr. Candidate genes of dysregulated miRNAs were identified by prediction algorithms based on gene expression profiles. Networks of KEGG pathways were organized by the selected dysregulated miRNAs. Moreover, protein-protein interaction (PPI) was utilized to map protein interaction networks of curcumin regulated proteins. We first demonstrated liposomal curcumin could rescue part of Adriamycin resistance in breast cancer and further identified 67 differentially expressed microRNAs among MCF-7/S, MCF-7/Adr and curcumin-treated MCF-7/Adr. The results showed that lower expressed miR-29b-1-5p decreased the IC50 of MCF-7/Adr cells and higher expressed miR-29b-1-5p, weaken the effects of liposomal curcumin to Adr-resistance. Besides, we found that 20 target genes (mRNAs) of each dysregulated miRNA were not only predicted by prediction algorithms, but also differentially expressed in the microarray. The results showed that MAPK, mTOR, PI3K-Akt, AMPK, TNF, Ras signaling pathways and several target genes such as PPARG, RRM2, SRSF1and EPAS1, may associate with drug resistance of breast cancer cells to Adr. We determined

  4. Organ-Tumor-on-a-Chip for Chemosensitivity Assay: A Critical Review

    Directory of Open Access Journals (Sweden)

    Navid Kashaninejad

    2016-07-01

    Full Text Available With a mortality rate over 580,000 per year, cancer is still one of the leading causes of death worldwide. However, the emerging field of microfluidics can potentially shed light on this puzzling disease. Unique characteristics of microfluidic chips (also known as micro-total analysis system make them excellent candidates for biological applications. The ex vivo approach of tumor-on-a-chip is becoming an indispensable part of personalized medicine and can replace in vivo animal testing as well as conventional in vitro methods. In tumor-on-a-chip, the complex three-dimensional (3D nature of malignant tumor is co-cultured on a microfluidic chip and high throughput screening tools to evaluate the efficacy of anticancer drugs are integrated on the same chip. In this article, we critically review the cutting edge advances in this field and mainly categorize each tumor-on-a-chip work based on its primary organ. Specifically, design, fabrication and characterization of tumor microenvironment; cell culture technique; transferring mechanism of cultured cells into the microchip; concentration gradient generators for drug delivery; in vitro screening assays of drug efficacy; and pros and cons of each microfluidic platform used in the recent literature will be discussed separately for the tumor of following organs: (1 Lung; (2 Bone marrow; (3 Brain; (4 Breast; (5 Urinary system (kidney, bladder and prostate; (6 Intestine; and (7 Liver. By comparing these microchips, we intend to demonstrate the unique design considerations of each tumor-on-a-chip based on primary organ, e.g., how microfluidic platform of lung-tumor-on-a-chip may differ from liver-tumor-on-a-chip. In addition, the importance of heart–liver–intestine co-culture with microvasculature in tumor-on-a-chip devices for in vitro chemosensitivity assay will be discussed. Such system would be able to completely evaluate the absorption, distribution, metabolism, excretion and toxicity (ADMET of

  5. Apigenin promotes TRAIL-mediated apoptosis regardless of ROS generation.

    Science.gov (United States)

    Kang, Chang-Hee; Molagoda, Ilandarage Menu Neelaka; Choi, Yung Hyun; Park, Cheol; Moon, Dong-Oh; Kim, Gi-Young

    2018-01-01

    Apigenin is a bioactive flavone in several herbs including parsley, thyme, and peppermint. Apigenin possesses anti-cancer and anti-inflammatory properties; however, whether apigenin enhances TRAIL-mediated apoptosis in cancer cells is unknown. In the current study, we found that apigenin enhanced TRAIL-induced apoptosis by promoting caspase activation and death receptor 5 (DR5) expression and a chimeric antibody against DR5 completely blocked the apoptosis. Apigenin also upregulated reactive oxygen species (ROS) generation; however, intriguingly, ROS inhibitors, glutathione (GSH) or N-acetyl-l-cysteine (NAC), moderately increased apigenin/TRAIL-induced apoptosis. Additional results showed that an autophagy inducer, rapamycin, enhanced apigenin/TRAIL-mediated apoptosis by a slight increase of ROS generation. Accordingly, NAC and GSH rather decreased apigenin-induced autophagy formation, suggesting that apigenin-induced ROS generation increased autophagy formation. However, autophagy inhibitors, bafilomycin (BAF) and 3-methyladenine (3-MA), showed different result in apigenin/TRAIL-mediated apoptosis without ROS generation. 3-MA upregulated the apoptosis but remained ROS levels; however, no changes on apoptosis and ROS generation were observed by BAF treatment. Taken together, these findings reveal that apigenin enhances TRAIL-induced apoptosis by activating apoptotic caspases by upregulating DR5 expression regardless of ROS generation, which may be a promising strategy for an adjuvant of TRAIL. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Spaceflight Associated Apoptosis

    Science.gov (United States)

    Ichiki, Albert T.; Gibson, Linda A.; Allebban, Zuhair

    1996-01-01

    Lymphoid tissues have been shown to atrophy in rats flown on Russian spaceflights. Histological examination indicated evidence for cell degradation. Lymphoid tissues from rats flown on Spacelab Life Sciences-2 mission were analyzed for apoptosis by evidence of fragmented lymphocytes, which could be engulfed by macrophages, or DNA strand breaks using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. Apoptosis was not detected in the thymus and spleen collected inflight or from the synchronous ground rats but was detected in the thymus, spleen and inguinal lymph node of the flight animals on recovery. These results indicate that the apoptosis observed in the lymphatic tissues of the rats on recovery could have been induced by the gravitational stress of reentry, corroborating the findings from the early space-flight observations.

  7. Expression of human organic cation transporter 3 in kidney carcinoma cell lines increases chemosensitivity to melphalan, irinotecan, and vincristine.

    Science.gov (United States)

    Shnitsar, Volodymyr; Eckardt, Ronny; Gupta, Shivangi; Grottker, Julia; Müller, Gerhard A; Koepsell, Hermann; Burckhardt, Gerhard; Hagos, Yohannes

    2009-02-15

    Renal cell carcinoma (RCC) is usually chemoresistant. This chemoresistance could be overcome if specific cytostatics are applied for which the RCC expresses an uptake transporter. In the present study, we investigated the expression of solute carrier (SLC) transporters in different RCC lines and their ability to interact with chemotherapeutics. We tested five RCC lines for the expression of different SLCs by reverse transcription-PCR and TaqMan real-time PCR. In two of five RCC lines, A498 and 7860, we observed a highly significant expression of SLC22A3 (hOCT3). Uptake of the organic cation [(3)H]MPP (4-methyl-pyridinium iodide) into these cells and also into hOCT3 stably transfected Chinese hamster ovary (CHO) cells was inhibited by irinotecan, vincristine, and melphalan. The K(i) values [determined from Dixon plots] for irinotecan, vincristine, and melphalan were 1.72 +/- 0.45 micromol/L, 17 +/- 4.81 micromol/L, and 366 +/- 51 micromol/L, respectively. Cytotoxic activities of the selected drugs were tested by [(3)H]thymidine incorporation and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays on CHO-hOCT3, A498 (high expression of hOCT3), and ACHN cell lines (low expression of hOCT3). The growth of CHO-hOCT3 was inhibited by 20% more with irinotecan and by 50% more with vincristine compared with nontransfected CHO cells. Melphalan produced 20% to 30% more inhibition in hOCT3-expressing cells compared with nonexpressing control cells. Similar results were obtained for A498 and ACHN cells. Thus, our data support the hypothesis that the sensitivity of tumor cells to chemotherapeutic treatment depends on the expression of transporter proteins mediating specific drug accumulation into target cells.

  8. Aspartame-induced apoptosis in PC12 cells.

    Science.gov (United States)

    Horio, Yukari; Sun, Yongkun; Liu, Chuang; Saito, Takeshi; Kurasaki, Masaaki

    2014-01-01

    Aspartame is an artificial sweetner added to many low-calorie foods. The safety of aspartame remains controversial even though there are many studies on its risks. In this study, to understand the physiological effects of trace amounts of artificial sweetners on cells, the effects of aspartame on apoptosis were investigated using a PC12 cell system. In addition, the mechanism of apoptosis induced by aspartame in PC12 cells and effects on apoptotic factors such as cytochrome c, apoptosis-inducing factor, and caspase family proteins were studied by Western blotting and RT-PCR. Aspartame-induced apoptosis in PC12 cells in a dose-dependent manner. In addition, aspartame exposure increased the expressions of caspases 8 and 9, and cytochrome c. These results indicate that aspartame induces apoptosis mainly via mitochondrial pathway involved in apoptosis due to oxigen toxicity. Copyright © 2013 Elsevier B.V. All rights reserved.

  9. Cl- channels in apoptosis

    DEFF Research Database (Denmark)

    Wanitchakool, Podchanart; Ousingsawat, Jiraporn; Sirianant, Lalida

    2016-01-01

    A remarkable feature of apoptosis is the initial massive cell shrinkage, which requires opening of ion channels to allow release of K(+), Cl(-), and organic osmolytes to drive osmotic water movement and cell shrinkage. This article focuses on the role of the Cl(-) channels LRRC8, TMEM16/anoctamin......, and cystic fibrosis transmembrane conductance regulator (CFTR) in cellular apoptosis. LRRC8A-E has been identified as a volume-regulated anion channel expressed in many cell types. It was shown to be required for regulatory and apoptotic volume decrease (RVD, AVD) in cultured cell lines. Its presence also...

  10. Escin Chemosensitizes Human Pancreatic Cancer Cells and Inhibits the Nuclear Factor-kappaB Signaling Pathway.

    Science.gov (United States)

    Rimmon, A; Vexler, A; Berkovich, L; Earon, G; Ron, I; Lev-Ari, S

    2013-01-01

    Background. There is an urgent need to develop new treatment strategies and drugs for pancreatic cancer that is highly resistant to radio-chemotherapy. Aesculus hippocastanum (the horse chestnut) known in Chinese medicine as a plant with anti-inflammatory, antiedema, antianalgesic, and antipyretic activities. The main active compound of this plant is Escin (C54H84O23). Objective. To evaluate the effect of Escin alone and combined with chemotherapy on pancreatic cancer cell survival and to unravel mechanism(s) of Escin anticancer activity. Methods. Cell survival was measured by XTT colorimetric assay. Synergistic effect of combined therapy was determined by CalcuSyn software. Cell cycle and induction of apoptosis were evaluated by FACS analysis. Expression of NF- κ B-related proteins (p65, I κ Bα, and p-I κ Bα) and cyclin D was evaluated by western blot analysis. Results. Escin decreased the survival of pancreatic cancer cells with IC50 = 10-20 M. Escin combined with gemcitabine showed only additive effect, while its combination with cisplatin resulted in a significant synergistic cytotoxic effect in Panc-1 cells. High concentrations of Escin induced apoptosis and decreased NF- κ B-related proteins and cyclin D expression. Conclusions. Escin decreased pancreatic cancer cell survival, induced apoptosis, and downregulated NF- κ B signaling pathway. Moreover, Escin sensitized pancreatic cancer cells to chemotherapy. Further translational research is required.

  11. Escin Chemosensitizes Human Pancreatic Cancer Cells and Inhibits the Nuclear Factor-kappaB Signaling Pathway

    Directory of Open Access Journals (Sweden)

    A. Rimmon

    2013-01-01

    Full Text Available Background. There is an urgent need to develop new treatment strategies and drugs for pancreatic cancer that is highly resistant to radio-chemotherapy. Aesculus hippocastanum (the horse chestnut known in Chinese medicine as a plant with anti-inflammatory, antiedema, antianalgesic, and antipyretic activities. The main active compound of this plant is Escin (C54H84O23. Objective. To evaluate the effect of Escin alone and combined with chemotherapy on pancreatic cancer cell survival and to unravel mechanism(s of Escin anticancer activity. Methods. Cell survival was measured by XTT colorimetric assay. Synergistic effect of combined therapy was determined by CalcuSyn software. Cell cycle and induction of apoptosis were evaluated by FACS analysis. Expression of NF-κB-related proteins (p65, IκBα, and p-IκBα and cyclin D was evaluated by western blot analysis. Results. Escin decreased the survival of pancreatic cancer cells with IC50 = 10–20 M. Escin combined with gemcitabine showed only additive effect, while its combination with cisplatin resulted in a significant synergistic cytotoxic effect in Panc-1 cells. High concentrations of Escin induced apoptosis and decreased NF-κB-related proteins and cyclin D expression. Conclusions. Escin decreased pancreatic cancer cell survival, induced apoptosis, and downregulated NF-κB signaling pathway. Moreover, Escin sensitized pancreatic cancer cells to chemotherapy. Further translational research is required.

  12. Role of PUMA in methamphetamine-induced neuronal apoptosis.

    Science.gov (United States)

    Chen, Chuanxiang; Qincao, Litao; Xu, Jingtao; Du, Sihao; Huang, Enping; Liu, Chao; Lin, Zhoumeng; Xie, Wei-Bing; Wang, Huijun

    2016-01-05

    Exposure to methamphetamine (METH), a widely used illicit drug, has been shown to cause neuron apoptosis. p53 upregulated modulator of apoptosis (PUMA) is a key mediator in neuronal apoptosis. This study aimed to examine the effects of PUMA in METH-induced neuronal apoptosis. We determined PUMA protein expression in PC12 cells and SH-SY5Y cells after METH exposure using western blot. We also observed the effect of METH on neuronal apoptosis after silencing PUMA expression with siRNA using TUNEL staining and flow cytometry. Additionally, to investigate possible mechanisms of METH-induced PUMA-mediated neuronal apoptosis, we measured the protein expression of apoptotic markers, including cleaved caspase-3, cleaved PARP, Bax, B-cell leukemia/lymphoma-2 (Bcl-2) and cytochrome c (cyto c), after METH treatment with or without PUMA knockdown. Results showed that METH exposure induced cell apoptosis, increased PUMA protein levels, activated caspase-3 and PARP, elevated Bax and reduced Bcl-2 expression, as well as increased the release of cyto c from mitochondria to the cytoplasm in both PC12 and SH-SY5Y cells. All these effects were attenuated or reversed after silencing PUMA. A schematic depicting the role of PUMA in METH-induced mitochondrial apoptotic pathway was proposed. Our results suggest that PUMA plays an important role in METH-triggered apoptosis and it may be a potential target for ameliorating neuronal injury and apoptosis caused by METH. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  13. Cyclin-dependent kinases regulate apoptosis of intestinal epithelial cells

    Science.gov (United States)

    Bhattacharya, Sujoy; Ray, Ramesh M.; Johnson, Leonard R.

    2014-01-01

    Homeostasis of the gastrointestinal epithelium is dependent upon a balance between cell proliferation and apoptosis. Cyclin-dependent kinases (Cdks) are well known for their role in cell proliferation. Previous studies from our group have shown that polyamine-depletion of intestinal epithelial cells (IEC-6) decreases cyclin-dependent kinase 2 (Cdk2) activity, increases p53 and p21Cip1 protein levels, induces G1 arrest, and protects cells from camptothecin (CPT)-induced apoptosis. Although emerging evidence suggests that members of the Cdk family are involved in the regulation of apoptosis, their roles directing apoptosis of IEC-6 cells are not known. In this study, we report that inhibition of Cdk1, 2, and 9 (with the broad range Cdk inhibitor, AZD5438) in proliferating IEC-6 cells triggered DNA damage, activated p53 signaling, inhibited proliferation, and induced apoptosis. By contrast, inhibition of Cdk2 (with NU6140) increased p53 protein and activity, inhibited proliferation, but had no effect on apoptosis. Notably, AZD5438 sensitized, whereas, NU6140 rescued proliferating IEC-6 cells from CPT-induced apoptosis. However, in colon carcinoma (Caco2) cells with mutant p53, treatment with either AZD5438 or NU6140 blocked proliferation, albeit more robustly with AZD5438. Both Cdk inhibitors induced apoptosis in Caco2 cells in a p53-independent manner. In serum starved quiescent IEC-6 cells, both AZD5438 and NU6140 decreased TNF- /CPT-induced activation of p53 and, consequently, rescued cells from apoptosis, indicating that sustained Cdk activity is required for apoptosis of quiescent cells. Furthermore, AZD5438 partially reversed the protective effect of polyamine depletion whereas NU6140 had no effect. Together, these results demonstrate that Cdks possess opposing roles in the control of apoptosis in quiescent and proliferating cells. In addition, Cdk inhibitors uncouple proliferation from apoptosis in a p53-dependent manner. PMID:24242917

  14. Intrinsic and chemo-sensitizing activity of SMAC-mimetics on high-risk childhood acute lymphoblastic leukemia.

    Science.gov (United States)

    Schirmer, M; Trentin, L; Queudeville, M; Seyfried, F; Demir, S; Tausch, E; Stilgenbauer, S; Eckhoff, S M; Meyer, L H; Debatin, K-M

    2016-01-14

    SMAC-mimetics represent a targeted therapy approach to overcome apoptosis resistance in many tumors. Here, we investigated the efficacy of the SMAC-mimetic BV6 in B-cell precursor acute lymphoblastic leukemia (BCP-ALL). In ALL cell lines, intrinsic apoptosis sensitivity was associated with rapid cIAP degradation, NF-κB activation, TNF-α secretion and induction of an autocrine TNF-α-dependent cell death loop. This pattern of responsiveness was also observed upon ex vivo analysis of 40 primograft BCP-ALL samples. Treatment with BV6 induced cell death in the majority of ALL primografts including leukemias with high-risk and poor-prognosis features. Inhibition of cell death by the TNF receptor fusion protein etanercept demonstrated that BV6 activity is dependent on TNF-α. In a preclinical NOD/SCID/huALL model of high-risk ALL, marked anti-leukemia effectivity and significantly prolonged survival were observed upon BV6 treatment. Interestingly, also in vivo, intrinsic SMAC-mimetic activity was mediated by TNF-α. Importantly, BV6 increased the effectivity of conventional induction therapy including vincristine, dexamethasone and asparaginase leading to prolonged remission induction. These data suggest SMAC-mimetics as an important addendum to efficient therapy of pediatric BCP-ALL.

  15. Scutellarin Increases Cisplatin-Induced Apoptosis and Autophagy to Overcome Cisplatin Resistance in Non-small Cell Lung Cancer via ERK/p53 and c-met/AKT Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Chao-Yue Sun

    2018-02-01

    Full Text Available Cisplatin, as the first-line anti-tumor agent, is widely used for treatment of a variety of malignancies including non-small cell lung cancer (NSCLC. However, the acquired resistance has been a major obstacle for the clinical application. Scutellarin is a active flavone extracted from Erigeron breviscapus Hand-Mazz that has been shown to exhibit anticancer activities on various types of tumors. Here, we reported that scutellarin was capable of sensitizing A549/DDP cells to cisplatin by enhancing apoptosis and autophagy. Mechanistic analyses indicated that cisplatin-induced caspase-3-dependent apoptosis was elevated in the presence of scutellarin through activating extracellular signal-regulated kinases (ERK-mediated p53 pathway. Furthermore, scutellarin also promoted cisplatin-induced cytotoxic autophagy, downregulated expression of p-AKT and c-met. Deficiency of c-met reduced p-AKT level, and inhibition of p-AKT or c-met improved autophagy in A549/DDP cells. Interestingly, loss of autophagy attenuated the synergism of this combination. In vivo, the co-treatment of cisplatin and scutellarin notably reduced the tumor size when compared with cisplatin treatment alone. Notably, scutellarin significantly reduced the toxicity generated by cisplatin in tumor-bearing mice. This study identifies the unique role of scutellarin in reversing cisplatin resistance through apoptosis and autophagy, and suggests that combined cisplatin and scutellarin might be a novel therapeutic strategy for patients with NSCLC.

  16. Mitochondria in neutrophil apoptosis

    NARCIS (Netherlands)

    van Raam, B. J.; Verhoeven, A. J.; Kuijpers, T. W.

    2006-01-01

    Central in the regulation of the short life span of neutrophils are their mitochondria. These organelles hardly contribute to the energy status of neutrophils but play a vital role in the apoptotic process. Not only do the mitochondria contain cytotoxic proteins that are released during apoptosis

  17. Apoptosis and inflammation

    Directory of Open Access Journals (Sweden)

    C. Haanen

    1995-01-01

    Full Text Available During the last few decades it has been recognized that cell death is not the consequence of accidental injury, but is the expression of a cell suicide programme. Kerr et al. (1972 introduced the term apoptosis. This form of cell death is under the influence of hormones, growth factors and cytokines, which depending upon the receptors present on the target cells, may activate a genetically controlled cell elimination process. During apoptosis the cell membrane remains intact and the cell breaks into apoptotic bodies, which are phagocytosed. Apoptosis, in contrast to necrosis, is not harmful to the host and does not induce any inflammatory reaction. The principal event that leads to inflammatory disease is cell damage, induced by chemical/physical injury, anoxia or starvation. Cell damage means leakage of cell contents into the adjacent tissues, resulting in the capillary transmigration of granulocytes to the injured tissue. The accumulation of neutrophils and release of enzymes and oxygen radicals enhances the inflammatory reaction. Until now there has been little research into the factors controlling the accumulation and the tissue load of granulocytes and their histotoxic products in inflammatory processes. Neutrophil apoptosis may represent an important event in the control of intlamtnation. It has been assumed that granulocytes disintegrate to apoptotic bodies before their fragments are removed by local macrophages. Removal of neutrophils from the inflammatory site without release of granule contents is of paramount importance for cessation of inflammation. In conclusion, apoptotic cell death plays an important role in inflammatory processes and in the resolution of inflammatory reactions. The facts known at present should stimulate further research into the role of neutrophil, eosinophil and macrophage apoptosis in inflammatory diseases.

  18. Carbamate Pesticide-Induced Apoptosis in Human T Lymphocytes

    Directory of Open Access Journals (Sweden)

    Qing Li

    2015-04-01

    Full Text Available We previously found that carbamate pesticides induced significant apoptosis in human natural killer cells. To investigate whether carbamate pesticides also induce apoptosis in human T lymphocytes, in the present study Jurkat human T cells were treated in vitro with thiram, maneb, carbaryl or ziram. Apoptosis was determined by FITC-Annexin-V/PI staining. To explore the mechanism of apoptosis, intracellular levels of active caspase 3 and mitochondrial cytochrome-c release were determined by flow cytometry. We found that thiram, ziram, maneb and carbaryl also induced apoptosis in a time- and dose-dependent manner in the human T cells. However, the strength of the apoptosis-inducing effect differed among the pesticides, with the: thiram > ziram > maneb > carbaryl. Moreover, thiram significantly increased the intracellular level of active caspase 3 and caspase inhibitors significantly inhibited apoptosis. Thiram also significantly caused mitochondrial cytochrome-c release. These findings indicate that carbamate pesticides can induce apoptosis in human T cells, and the apoptosis is mediated by the activation of caspases and the release of mitochondrial cytochrome-c.

  19. Effects of euphylline on breathing pattern and chemosensitivity of the respiratory system after activation of GABAb-receptors.

    Science.gov (United States)

    Lebedeva, M A; Sanotskaya, N V; Matsievsky, D D

    2010-10-01

    We studied changes in breathing pattern in nembutal-anaesthetized mongrel rats after administration of euphylline against the background of preliminary treatment with lithium hydroxybutyrate. Two types of external respiration responses to euphylline were observed; they depended on the initial blood pressure in systemic circulation and on its drop after euphylline administration. Thus, the reaction of the respiratory system to adenosine receptors blockade against the background of hydroxybutyrate pretreatment was associated with not only the effect of euphylline, but also the state of brain hemodynamics. The effects of euphylline on chemosensitive contour of the respiratory system regulation were also investigated. It was found that euphylline did not abolish desensitization of respiratory system to hypercapnia, but smoothed the response to hypoxia under conditions of GABA(b)-receptor activation.

  20. Elucidation of chemosensitization effect of acridones in cancer cell lines: Combined pharmacophore modeling, 3D QSAR, and molecular dynamics studies.

    Science.gov (United States)

    Gade, Deepak Reddy; Makkapati, Amareswararao; Yarlagadda, Rajesh Babu; Peters, Godefridus J; Sastry, B S; Rajendra Prasad, V V S

    2018-02-24

    Overexpression of P-glycoprotein (P-gp) leads to the emergence of multidrug resistance (MDR) in cancer treatment. Acridones have the potential to reverse MDR and sensitize cells. In the present study, we aimed to elucidate the chemosensitization potential of acridones by employing various molecular modelling techniques. Pharmacophore modeling was performed for the dataset of chemosensitizing acridones earlier proved for cytotoxic activity against MCF7 breast cancer cell line. Gaussian-based QSAR studies also performed to predict the favored and disfavored region of the acridone molecules. Molecular dynamics simulations were performed for compound 10 and human P-glycoprotein (obtained from Homology modeling). An efficient pharmacophore containing 2 hydrogen bond acceptors and 3 aromatic rings (AARRR.14) was identified. NCI 2012 chemical database was screened against AARRR.14 CPH and identified 25 best-fit molecules. Potential regions of the compound were identified through Field (Gaussian) based QSAR. Regression analysis of atom-based QSAR resulted in r 2 of 0.95 and q 2 of 0.72, whereas, regression analysis of field-based QSAR resulted in r 2 of 0.92 and q 2 of 0.87 along with r 2 cv as 0.71. The fate of the acridone molecule (compound 10) in the P-glycoprotein environment is analyzed through analyzing the conformational changes occurring during the molecular dynamics simulations. Combined data of different in silico techniques provided basis for deeper understanding of structural and mechanistic insights of interaction phenomenon of acridones with P-glycoprotein and also as strategic basis for designing more potent molecules for anti-cancer and multidrug resistance reversal activities. Copyright © 2018 Elsevier Ltd. All rights reserved.

  1. Serum midkine as a biomarker for malignancy, prognosis, and chemosensitivity in head and neck squamous cell carcinoma

    International Nuclear Information System (INIS)

    Yamashita, Taku; Shimada, Hideaki; Tanaka, Shingo; Araki, Koji; Tomifuji, Masayuki; Mizokami, Daisuke; Tanaka, Nobuaki; Kamide, Daisuke; Miyagawa, Yoshihiro; Suzuki, Hiroshi; Tanaka, Yuya; Shiotani, Akihiro

    2016-01-01

    Improved therapies for individuals with head and neck squamous cell carcinoma (HNSCC) may be developed by identification of appropriate biomarkers. The aim of this study was to evaluate the usefulness of serum midkine measurement as a biomarker for HNSCC. Pretreatment serum midkine concentrations were measured in 103 patients with HNSCC and 116 control individuals by enzyme-linked immunosorbent assay. Midkine expression in tumor tissues from 33 patients with HNSCC who underwent definitive surgical resection without preoperative treatment was examined by immunohistochemistry. The cut-off serum midkine concentrations for predicting the presence of head and neck malignancy and chemosensitivity to induction chemotherapy, as determined using receiver operating characteristic curves, were 482 and 626 pg/mL, respectively. Spearman bivariate correlations showed positive correlations between serum midkine levels and immunohistochemistry staining score (r = 0.612, P < 0.001). The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of serum midkine concentration for detection of HNSCC were 57.3, 85.3, 77.6, 69.2, and 72.1%, respectively. However, for predicting the response to induction chemotherapy, the values were 84.6, 60.9, 71.0, 77.8, and 73.5%, respectively. Serum midkine concentration was identified as an independent prognostic factor by multivariate analysis, using Cox's proportional hazards model (P = 0.027). Overexpression of serum midkine yielded a relative risk of death of 3.77, with 95% confidence limits ranging from 1.15 to 17.0. Serum midkine levels in patients with HNSCC were associated with malignancy, chemosensitivity, and prognosis. Serum midkine may be a useful, minimally invasive biomarker for early detection, therapeutic decision-making, and predicting prognosis

  2. Bioinformatic identification of IGF1 as a hub gene in hepatocellular carcinoma (HCC) and in-vitro analysis of the chemosensitizing effect of miR-379 via suppressing the IGF1/IGF1R signaling pathway.

    Science.gov (United States)

    Huang, D-J; Huang, J-Z; Yang, J; Li, Y-H; Luo, Y-C; He, H-Y; Huang, H-J

    2016-12-01

    We investigated the interactions among the dysregulated genes in hepatocellular carcinoma (HCC) and identified the hub genes in the protein-protein interaction (PPI) network. Also, we also investigated the regulative effect of miR-379 on the IGF1/IGF1R signaling pathway and chemoresistance in HCC. Raw data of a microarray that compared transcriptional gene profile between 3-paired HCC tissue samples and adjacent normal tissues were downloaded from Expression Atlas (E-GEOD-33006). The raw data was reanalyzed to identify the significantly dysregulated genes, which were further used for PPI network and KEGG pathway analysis. The regulative effect of miR-379 on IGF1R expression was studied by dual luciferase assay and Western blotting. The functional role of miR-379 in chemosensitivity of HCC cells was studied by drug sensitivity and flow cytometric assay. IGF1 is a hub gene that is mostly upregulated in HCC and it is an important player in the p53 signaling pathway. Knockdown of IGF1R significantly decreased IC50 of 5-FU, paclitaxel (PTX) and Doxorubicin (DOX) in Huh7 and HepG2 cells. MiR-379 could directly bind to the 3'UTR of IGF1R and suppress IGF1R expression. MiR-379 overexpression sensitized Huh7 and HepG2 cells to 5-FU, PTX and DOX and also enhanced DOX-induced cell apoptosis. IGF1 is a hub gene in HCC and is also one of the most upregulated genes in HCC tissues compared to normal tissues. It is involved in the p53 signaling pathway regulation. MiR-379 can sensitize HCC cells to chemotherapeutic reagents via targeting IGF1R and suppressing its expression.

  3. SB225002 promotes mitotic catastrophe in chemo-sensitive and -resistant ovarian cancer cells independent of p53 status in vitro.

    Directory of Open Access Journals (Sweden)

    Meirong Du

    Full Text Available Recent evidence indicates that CXCR2 signaling is crucial for cancer progression, and its antagonist SB225002 induces apoptosis in Wilms' tumor cells. Here, we investigated the effect of SB225002 on cell cycle progression and apoptosis induction in vitro, using CDDP-sensitive and -resistant OVCA cell lines with different p53 status (wild type, mutant or null. Adenovirus infection of wild-type p53 or transfection of p53 siRNA was used to over-express or knock-down p53. Cell cycle and apoptosis were determined by flow cytometry or Hoechst staining and observation of nuclear morphology. Our data demonstrated that SB225002 induced apoptosis in both wild-type and p53-deficient ovarian cancer (OVCA cells through alternative mechanisms. SB225002 promoted mitotic catastrophe, as evidenced by the accumulation of mitotic cells with spindle abnormalities, chromosome mis-segregation, multi-polar cell division, multiple nuclei, aneuploidy/polyploidy and subsequent extensive apoptosis. SB225002-induced mitotic catastrophe appeared to be mediated by down-regulation of checkpoint kinase Chk1 and Cdk1-cyclin B activation. In cells expressing wild-type p53 (OV2008 and C13*, SB225002 increased total and phospho-Ser p53 levels, and p53 knock-down decreased SB225002-induced apoptosis, without affecting premature mitosis. These results suggest that SB225002 induces p53-dependent apoptosis, and provokes mitotic catastrophe in p53-independent manner in p53 wild-type cells. Reconstitution with wild-type P53 in P53-null SKOV3 cell attenuated SB225002-induced mitotic catastrophe, suggesting p53 prevented mitotic catastrophe induced by SB225002 in p53-deficient OVCA cells. Finally, the effect of SB225002 could not be prevented by pretreatment with CXCR2 ligand or its neutralizing antibody. The present studies demonstrate for the first time that SB225002 has dual actions in OVCA cells, inducing classic apoptosis through p53 activation and provoking mitotic catastrophe in

  4. Spermine inhibits Endoplasmic Reticulum Stress - induced Apoptosis: a New Strategy to Prevent Cardiomyocyte Apoptosis

    Directory of Open Access Journals (Sweden)

    Can Wei

    2016-02-01

    Full Text Available Background/Aims: Endoplasmic reticulum stress (ERS plays an important role in the progression of acute myocardial infarction (AMI, in part by mediating apoptosis. Polyamines, including putrescine, spermidine, and spermine, are polycations with anti-oxidative, anti-aging, and cell growth-promoting activities. This study aimed to determine the mechanisms by which spermine protects against ERS-induced apoptosis in rats following AMI. Methods and Results: AMI was established by ligation of the left anterior descending coronary artery (LAD in rats, and exogenous spermine was administered by intraperitoneal injection (2.5 mg/ml daily for 7 days pre-AMI. Spermine treatment limited infarct size, attenuated cardiac troponin I and creatinine kinase-MB release, improved cardiac function, and decreased ERS and apoptosis related protein expression. Isolated cardiomyocytes subjected to hypoxia showed significant increase in reactive oxygen species (ROS and the expression of apoptosis and ERS related proteins; these effects occurred through PERK and eIF2α phosphorylation. The addition of spermine attenuated cardiomyocyte apoptosis, suppressed the production of ROS, and inhibited ERS related pathways. Conclusions: Spermine was an effective pre-treatment strategy to attenuate cardiac ERS injury in rats, and the cardioprotective mechanism occurring through inhibition of ROS production and down regulation of the PERK-eIF2α pathway. These findings provide a novel target for the prevention of apoptosis in the setting of AMI.

  5. Fullerene and apoptosis

    Directory of Open Access Journals (Sweden)

    M. A. Orlova

    2013-01-01

    Full Text Available Fullerene derivatives superfamily attracts a serious attention as antiviral and anticancer agents and drug delivery carriers as well. A large number of such fullerene С60 derivatives obtained to date. However, there is an obvious deficit of information about causes and mechanisms of immediately and long-term consequences of their effects in vivo which is a true obstacle on the way leading to practical medical use of them. First, this concerns their impact on the proliferation, apoptosis and necrosis regulation. Fullerene nanoparticle functionalization type, their sizes and surface nanopathology are of great importance to further promoting of either cytoprotective or cytotoxic effects. This lecture provides modern concept analysis regarding fullerenes effects on apoptosis pathway in normal and tumor cells.

  6. Khz-cp (crude polysaccharide extract obtained from the fusion of Ganoderma lucidum and Polyporus umbellatus mycelia) induces apoptosis by increasing intracellular calcium levels and activating P38 and NADPH oxidase-dependent generation of reactive oxygen species in SNU-1 cells.

    Science.gov (United States)

    Kim, Tae Hwan; Kim, Ju Sung; Kim, Zoo Haye; Huang, Ren Bin; Chae, Young Lye; Wang, Ren Sheng

    2014-07-10

    Khz-cp is a crude polysaccharide extract that is obtained after nuclear fusion in Ganoderma lucidum and Polyporus umbellatus mycelia (Khz). It inhibits the growth of cancer cells. Khz-cp was extracted by solvent extraction. The anti-proliferative activity of Khz-cp was confirmed by using Annexin-V/PI-flow cytometry analysis. Intracellular calcium increase and measurement of intracellular reactive oxygen species (ROS) were performed by using flow cytometry and inverted microscope. SNU-1 cells were treated with p38, Bcl-2 and Nox family siRNA. siRNA transfected cells was employed to investigate the expression of apoptotic, growth and survival genes in SNU-1 cells. Western blot analysis was performed to confirm the expression of the genes. In the present study, Khz-cp induced apoptosis preferentially in transformed cells and had only minimal effects on non-transformed cells. Furthermore, Khz-cp was found to induce apoptosis by increasing the intracellular Ca2+ concentration ([Ca2+]i) and activating P38 to generate reactive oxygen species (ROS) via NADPH oxidase and the mitochondria. Khz-cp-induced apoptosis was caspase dependent and occurred via a mitochondrial pathway. ROS generation by NADPH oxidase was critical for Khz-cp-induced apoptosis, and although mitochondrial ROS production was also required, it appeared to occur secondary to ROS generation by NADPH oxidase. Activation of NADPH oxidase was shown by the translocation of the regulatory subunits p47phox and p67phox to the cell membrane and was necessary for ROS generation by Khz-cp. Khz-cp triggered a rapid and sustained increase in [Ca2+]i that activated P38. P38 was considered to play a key role in the activation of NADPH oxidase because inhibition of its expression or activity abrogated membrane translocation of the p47phox and p67phox subunits and ROS generation. In summary, these data indicate that Khz-cp preferentially induces apoptosis in cancer cells and that the signaling mechanisms involve an

  7. Effects of hypoxia on expression of a panel of stem cell and chemosensitivity markers in glioblastoma cell line-derived spheroids

    DEFF Research Database (Denmark)

    Kolenda, Jesper; Jensen, Stine Skov; Aaberg-Jessen, Charlotte

    Glioblastomas are the most frequent and malignant primary brain tumor. Tumor stem cells in these tumors have recently been suggested to possess innate resistance mechanisms against radiation and chemotherapy possibly explaining their high level of therapeutic resistance. Moreover tumor hypoxia...... of a panel of stem cell and chemosensitivity markers was therefore investigated using glioma spheroids derived from the conventional glioblastoma cell line U87. The glioma spheroids were derived at normoxic (21% O2) and hypoxic (1% O2) culturing conditions in serum-free medium with EGF and bFGF. The entire...... immunohistochemical panel included hypoxia (HIF-1α, HIF-2α), proliferation (Ki-67) and stem cell (CD133, nestin, podoplanin, Bmi-1, Sox-2) markers as well as markers related to chemosensitivity (MGMT, MDR-1, TIMP-1, Lamp-1). Since spheroids derived in hypoxia were smaller than in normoxia, a set of experiments...

  8. Apoptosis transcriptional mechanism of feline infectious peritonitis virus infected cells.

    Science.gov (United States)

    Shuid, Ahmad Naqib; Safi, Nikoo; Haghani, Amin; Mehrbod, Parvaneh; Haron, Mohd Syamsul Reza; Tan, Sheau Wei; Omar, Abdul Rahman

    2015-11-01

    Apoptosis has been postulated to play an important role during feline infectious peritonitis virus (FIPV) infection; however, its mechanism is not well characterized. This study is focused on apoptosis and transcriptional profiling of FIPV-infected cells following in vitro infection of CRFK cells with FIPV 79-1146 WSU. Flow cytometry was used to determine mode of cell death in first 42 h post infection (hpi). FIPV infected cells underwent early apoptosis at 9 hpi (p apoptosis at 12 hpi (p apoptosis cluster (80 down-regulated and 51 up-regulated) along with increase of apoptosis, p53, p38 MAPK, VEGF and chemokines/cytokines signaling pathways were probably involved in apoptosis process. Six of the de-regulated genes expression (RASSF1, BATF2, MAGEB16, PDCD5, TNFα and TRAF2) and TNFα protein concentration were analyzed by RT-qPCR and ELISA, respectively, at different time-points. Up-regulations of both pro-apoptotic (i.e. PDCD5) and anti-apoptotic (i.e. TRAF2) were detected from first hpi and continuing to deregulate during apoptosis process in the infected cells.

  9. Apoptosis signal-regulating kinase 1 mediates denbinobin-induced apoptosis in human lung adenocarcinoma cells

    Directory of Open Access Journals (Sweden)

    Pan Shiow-Lin

    2009-05-01

    Full Text Available Abstract In the present study, we explore the role of apoptosis signal-regulating kinase 1 (ASK1 in denbinobin-induced apoptosis in human lung adenocarcinoma (A549 cells. Denbinobin-induced cell apoptosis was attenuated by an ASK1 dominant-negative mutant (ASK1DN, two antioxidants (N-acetyl-L-cysteine (NAC and glutathione (GSH, a c-Jun N-terminal kinase (JNK inhibitor (SP600125, and an activator protein-1 (AP-1 inhibitor (curcumin. Treatment of A549 cells with denbinobin caused increases in ASK1 activity and reactive oxygen species (ROS production, and these effects were inhibited by NAC and GSH. Stimulation of A549 cells with denbinobin caused JNK activation; this effect was markedly inhibited by NAC, GSH, and ASK1DN. Denbinobin induced c-Jun phosphorylation, the formation of an AP-1-specific DNA-protein complex, and Bim expression. Bim knockdown using a bim short interfering RNA strategy also reduced denbinobin-induced A549 cell apoptosis. The denbinobin-mediated increases in c-Jun phosphorylation and Bim expression were inhibited by NAC, GSH, SP600125, ASK1DN, JNK1DN, and JNK2DN. These results suggest that denbinobin might activate ASK1 through ROS production to cause JNK/AP-1 activation, which in turn induces Bim expression, and ultimately results in A549 cell apoptosis.

  10. Apoptosis and DNA Methylation

    International Nuclear Information System (INIS)

    Meng, Huan X.; Hackett, James A.; Nestor, Colm; Dunican, Donncha S.; Madej, Monika; Reddington, James P.; Pennings, Sari; Harrison, David J.; Meehan, Richard R.

    2011-01-01

    Epigenetic mechanisms assist in maintaining gene expression patterns and cellular properties in developing and adult tissues. The molecular pathology of disease states frequently includes perturbation of DNA and histone methylation patterns, which can activate apoptotic pathways associated with maintenance of genome integrity. This perspective focuses on the pathways linking DNA methyltransferases and methyl-CpG binding proteins to apoptosis, and includes new bioinformatic analyses to characterize the evolutionary origin of two G/T mismatch-specific thymine DNA glycosylases, MBD4 and TDG

  11. Reassessing apoptosis in plants.

    Science.gov (United States)

    Dickman, Martin; Williams, Brett; Li, Yurong; de Figueiredo, Paul; Wolpert, Thomas

    2017-10-01

    Cell death can be driven by a genetically programmed signalling pathway known as programmed cell death (PCD). In plants, PCD occurs during development as well as in response to environmental and biotic stimuli. Our understanding of PCD regulation in plants has advanced significantly over the past two decades; however, the molecular machinery responsible for driving the system remains elusive. Thus, whether conserved PCD regulatory mechanisms include plant apoptosis remains enigmatic. Animal apoptotic regulators, including Bcl-2 family members, have not been identified in plants but expression of such regulators can trigger or suppress plant PCD. Moreover, plants exhibit nearly all of the biochemical and morphological features of apoptosis. One difference between plant and animal PCD is the absence of phagocytosis in plants. Evidence is emerging that the vacuole may be key to removal of unwanted plant cells, and may carry out functions that are analogous to animal phagocytosis. Here, we provide context for the argument that apoptotic-like cell death occurs in plants.

  12. A novel method for detection of apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Zagariya, Alexander M., E-mail: zagariya@uic.edu

    2012-04-15

    There are two different Angiotensin II (ANG II) peptides in nature: Human type (ANG II) and Bovine type (ANG II*). These eight amino acid peptides differ only at position 5 where Valine is replaced by Isoleucine in the Bovine type. They are present in all species studied so far. These amino acids are different by only one atom of carbon. This difference is so small, that it will allow any of ANG II, Bovine or Human antibodies to interact with all species and create a universal method for apoptosis detection. ANG II concentrations are found at substantially higher levels in apoptotic, compared to non-apoptotic, tissues. ANG II accumulation can lead to DNA damage, mutations, carcinogenesis and cell death. We demonstrate that Bovine antiserum can be used for universal detection of apoptosis. In 2010, the worldwide market for apoptosis detection reached the $20 billion mark and significantly increases each year. Most commercially available methods are related to Annexin V and TUNNEL. Our new method based on ANG II is more widely known to physicians and scientists compared to previously used methods. Our approach offers a novel alternative for assessing apoptosis activity with enhanced sensitivity, at a lower cost and ease of use.

  13. A novel method for detection of apoptosis

    International Nuclear Information System (INIS)

    Zagariya, Alexander M.

    2012-01-01

    There are two different Angiotensin II (ANG II) peptides in nature: Human type (ANG II) and Bovine type (ANG II*). These eight amino acid peptides differ only at position 5 where Valine is replaced by Isoleucine in the Bovine type. They are present in all species studied so far. These amino acids are different by only one atom of carbon. This difference is so small, that it will allow any of ANG II, Bovine or Human antibodies to interact with all species and create a universal method for apoptosis detection. ANG II concentrations are found at substantially higher levels in apoptotic, compared to non-apoptotic, tissues. ANG II accumulation can lead to DNA damage, mutations, carcinogenesis and cell death. We demonstrate that Bovine antiserum can be used for universal detection of apoptosis. In 2010, the worldwide market for apoptosis detection reached the $20 billion mark and significantly increases each year. Most commercially available methods are related to Annexin V and TUNNEL. Our new method based on ANG II is more widely known to physicians and scientists compared to previously used methods. Our approach offers a novel alternative for assessing apoptosis activity with enhanced sensitivity, at a lower cost and ease of use.

  14. Long-term results of 2 adjuvant trials reveal differences in chemosensitivity and the pattern of metastases between colon cancer and rectal cancer.

    Science.gov (United States)

    Kornmann, Marko; Staib, Ludger; Wiegel, Thomas; Kron, Martina; Henne-Bruns, Doris; Link, Karl-Heinrich; Formentini, Andrea

    2013-03-01

    Two identical randomized controlled trials designed to optimize adjuvant treatment of colon cancer (CC) (n =855) and rectal cancer (RC) (n = 796) were performed. Long-term evaluation confirmed that the addition of folinic acid (FA) to 5-fluorouracil (5-FU) improved 7-year overall survival (OS) in CC but not in RC and revealed different patterns of recurrence in patients with CC and those with RC. Our aim was to compare long-term results of adjuvant treatment of colon cancer (CC) and rectal cancer (RC). Adjuvant chemotherapy of CC improved overall survival (OS), whereas that of RC remained at the level achieved by 5-fluorouracil (5-FU). We separately conducted 2 identically designed adjuvant trials in CC and RC. Patients were assigned to adjuvant chemotherapy with 5-FU alone, 5-FU + folinic acid (FA), or 5-FU + interferon-alfa. The first study enrolled patients with stage IIb/III CC, and the second study enrolled patients with stage II/III RC. All patients with RC received postoperative irradiation. Median follow-up for all patients with CC (n = 855) and RC (n = 796) was 4.9 years. The pattern and frequency of recurrence differed significantly, especially lung metastases, which occurred more frequently in RC (12.7%) than in CC (7.3%; P < .001). Seven-year OS rates for 5-FU, 5-FU + FA, and 5-FU + IFN-alfa were 54.1% (95% confidence interval [CI], 46.5-61.0), 66.8% (95% CI, 59.4-73.1), and 56.7% (95% CI, 49.3-63.4) in CC and 50.6% (95% CI, 43.0-57.7), 56.3% (95% CI, 49.4-62.7), and 54.8% (95% CI, 46.7-62.2) in RC, respectively. A subgroup analysis pointed to a reduced local recurrence (LR) rate and an increased OS by the addition of FA in stage II RC (n = 271) but not in stage III RC (n = 525). FA increased 7-year OS by 12.7 percentage points in CC but was not effective in RC. Based on these results and the pattern of metastases, our results suggest that the chemosensitivity of CC and RC may be different. Strategies different from those used in CC may be successful to

  15. Induction of Apoptosis and expression of Apoptosis-related gene products in response to radiation in murine tumors

    International Nuclear Information System (INIS)

    Seong, J. S.

    1997-01-01

    To analyze the involvement of apoptosis regulatory genes p53, p21 waf1/cip1 , bax and bcl-2 in induction of apoptosis by radiation in murine tumors. The radiation-sensitive ovarian carcinoma OCa-I and the radiation-resistant hepatocarcinoma HCa-I were used. Tumors, 8mm in diameter, were irradiated with 25Gy and at various times after irradiation, ranging from 1 to 48 h, were analyzed histologically for apoptosis and by western blot for alterations in the expression of these genes. The p53 status of the tumors were determined by the polymerase chain reaction-single strand conformation polymorphism assay. Both tumors were positive for wild-type p53. Radiation induced apoptosis in OCa-I but not in HCa-I. Apoptosis developed rapidly, peaked at 2 h after irradiation and returned to almost the background level at 48 h. In OCa-I radiation upregulated the expression of p53, p21 waf1/cip1 , and the bcl-2/bax ratio was decreased. In HCa-I radiation increased the expression of both p53 and p21 waf1/cip1 , although the increase of the latter was small. The bcl-2/bax ratio was greatly increased. In general the observed changes occurred within a few hours after irradiation, and either preceded or coincided with development of apoptosis. The development of apoptosis required upregulation of both p53 and p21 waf1/cip1 as well as a decrease in bcl-2/bax ratio. In contrast, an increase in bcl-2/bax ratio prevented apoptosis in the presence of upregulated p53 and p21 waf1/cip1 . These findings identified the involvement of multiple oncogenes in apoptosis regulation in vivo and demonstrate the complexity that may be associated with the use of a single oncogene assessment for predicting the outcome of cancer therapy with cytotoxic agents. (author)

  16. Induction of Apoptosis and expression of Apoptosis-related gene products in response to radiation in murine tumors

    Energy Technology Data Exchange (ETDEWEB)

    Seong, J. S. [Yonsei Univ., Seoul (Korea, Republic of). Coll. of Medicine; Hunter, N. R.; Milas, L. [Texas Univ., Houston, TX (United States)

    1997-09-01

    To analyze the involvement of apoptosis regulatory genes p53, p21{sup waf1/cip1}, bax and bcl-2 in induction of apoptosis by radiation in murine tumors. The radiation-sensitive ovarian carcinoma OCa-I and the radiation-resistant hepatocarcinoma HCa-I were used. Tumors, 8mm in diameter, were irradiated with 25Gy and at various times after irradiation, ranging from 1 to 48 h, were analyzed histologically for apoptosis and by western blot for alterations in the expression of these genes. The p53 status of the tumors were determined by the polymerase chain reaction-single strand conformation polymorphism assay. Both tumors were positive for wild-type p53. Radiation induced apoptosis in OCa-I but not in HCa-I. Apoptosis developed rapidly, peaked at 2 h after irradiation and returned to almost the background level at 48 h. In OCa-I radiation upregulated the expression of p53, p21{sup waf1/cip1}, and the bcl-2/bax ratio was decreased. In HCa-I radiation increased the expression of both p53 and p21{sup waf1/cip1}, although the increase of the latter was small. The bcl-2/bax ratio was greatly increased. In general the observed changes occurred within a few hours after irradiation, and either preceded or coincided with development of apoptosis. The development of apoptosis required upregulation of both p53 and p21{sup waf1/cip1} as well as a decrease in bcl-2/bax ratio. In contrast, an increase in bcl-2/bax ratio prevented apoptosis in the presence of upregulated p53 and p21{sup waf1/cip1}. These findings identified the involvement of multiple oncogenes in apoptosis regulation in vivo and demonstrate the complexity that may be associated with the use of a single oncogene assessment for predicting the outcome of cancer therapy with cytotoxic agents. (author).

  17. Apoptosis-inducing factor (Aif1) mediates anacardic acid-induced apoptosis in Saccharomyces cerevisiae.

    Science.gov (United States)

    Muzaffar, Suhail; Chattoo, Bharat B

    2017-03-01

    Anacardic acid is a medicinal phytochemical that inhibits proliferation of fungal as well as several types of cancer cells. It induces apoptotic cell death in various cell types, but very little is known about the mechanism involved in the process. Here, we used budding yeast Saccharomyces cerevisiae as a model to study the involvement of some key elements of apoptosis in the anacardic acid-induced cell death. Plasma membrane constriction, chromatin condensation, DNA degradation, and externalization of phosphatidylserine (PS) indicated that anacardic acid induces apoptotic cell death in S. cerevisiae. However, the exogenous addition of broad-spectrum caspase inhibitor Z-VAD-FMK or deletion of the yeast caspase Yca1 showed that the anacardic acid-induced cell death is caspase independent. Apoptosis-inducing factor (AIF1) deletion mutant was resistant to the anacardic acid-induced cell death, suggesting a key role of Aif1. Overexpression of Aif1 made cells highly susceptible to anacardic acid, further confirming that Aif1 mediates anacardic acid-induced apoptosis. Interestingly, instead of the increase in the intracellular reactive oxygen species (ROS) normally observed during apoptosis, anacardic acid caused a decrease in the intracellular ROS levels. Quantitative real-time PCR analysis showed downregulation of the BIR1 survivin mRNA expression during the anacardic acid-induced apoptosis.

  18. β-Elemene: Mechanistic Studies on Cancer Cell Interaction and Its Chemosensitization Effect

    Science.gov (United States)

    Jiang, Ziyu; Jacob, Joe A.; Loganathachetti, Dinesh S.; Nainangu, Prasannabalaji; Chen, Baoan

    2017-01-01

    Over the past decade, screening and identifying novel compounds for their biomedical applications has become an upcoming area of research. Identifying the molecular mechanisms of these compounds has become an integral part of anticancer research. β-elemene, a sesquiterpene, is renowned for its anticancer activity against a variety of cell lines. Recent studies on β-elemene have elucidated that it possesses anti-proliferative effect on cancer cells by creating an apoptotic trigger. Interestingly, it also induces protective autophagy in some cancerous cell lines and is less cytotoxic compared to other widely accepted chemotherapeutic agents. This provides an edge with the perception of limited toxicity to normal cells. This mini-review precisely focuses on the studies performed to identify the mechanism of anticancer activity of β-elemene against cancer cells of multiple origin. In accordance to the evaluation made by the studies mentioned, apoptosis has been identified to be most possible reason behind anticancer activity exerted by β-elemene against a variety of cancer cell lines. Cell cycle arrest and necrosis have been credited to be possible alternate mechanisms for the anticancer effect of β-elemene. PMID:28337141

  19. Chemosensitizing and cytotoxic effects of 2-deoxy-D-glucose on breast cancer cells

    Directory of Open Access Journals (Sweden)

    Zhang Fanjie

    2009-09-01

    Full Text Available Background: Accelerated glucose uptake for anerobic glycolysis is one of the major metabolic changes found in malignant cells. This property has been exploited for imaging malignancies and as a possible anticancer therapy. The nonmetabolizable glucose analog 2-deoxyglucose (2 DG interferes with glucose metabolism leading to breast cancer cell death. Aims: To determine whether 2DG can synergize with chemotherapeutic agents commonly used in breast cancer treatment and identify cellular characteristics associated with sensitivity to 2DG. Materials and Methods: SkBr3 breast cancer cells were incubated with varying concentrations of 5-fluorouracil (5FU, doxorubicin, cisplatin, cyclophosphamide, or herceptin with or without 2DG. Cell viability was measured using the MTT (3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide assay. Results: Combining 2DG with doxorubicin, 5 FU, cyclophosphamide, and herceptin resulted in enhanced cell death compared with each agent alone, while in combination with cisplatin, the amount of cell death was additive. Mouse embryo fibroblasts (MEF mutated for p53 (-/- were 30% more sensitive to the cytotoxic effects of 2DG than the parental cell lines. Cells mutated for Bax/Bac, genes involved in protection from apoptosis, are slightly more sensitive than the parental cell lines. Conclusions: These results indicate that 2DG acts synergistically with specific chemotherapeutic agents in causing cell death and the class of chemicals most sensitive appear to be those which cause DNA damage.

  20. CIAPIN1 gene silencing enhances chemosensitivity in a drug-resistant animal model in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Wang, X.M.; Gao, S.J.; Guo, X.F.; Sun, W.J. [Department of Oncology, The Second Affiliated Hospital, Harbin Medical University, Harbin (China); Yan, Z.Q. [Department of Breast Surgery, The Second Affiliated Hospital, Harbin Medical University, Harbin (China); Wang, W.X.; Xu, Y.Q.; Lu, D. [Department of Oncology, The Second Affiliated Hospital, Harbin Medical University, Harbin (China)

    2014-03-21

    Overexpression of cytokine-induced apoptosis inhibitor 1 (CIAPIN1) contributes to multidrug resistance (MDR) in breast cancer. This study aimed to evaluate the potential of CIAPIN1 gene silencing by RNA interference (RNAi) as a treatment for drug-resistant breast cancer and to investigate the effect of CIAPIN1 on the drug resistance of breast cancer in vivo. We used lentivirus-vector-based RNAi to knock down CIAPIN1 in nude mice bearing MDR breast cancer tumors and found that lentivirus-vector-mediated silencing of CIAPIN1 could efficiently and significantly inhibit tumor growth when combined with chemotherapy in vivo. Furthermore, Western blot analysis showed that both CIAPIN1 and P-glycoprotein expression were efficiently downregulated, and P53 was upregulated, after RNAi. Therefore, we concluded that lentivirus-vector-mediated RNAi targeting of CIAPIN1 is a potential approach to reverse MDR of breast cancer. In addition, CIAPIN1 may participate in MDR of breast cancer by regulating P-glycoprotein and P53 expression.

  1. Bactofection of sequences encoding a Bax protein peptide chemosensitizes prostate cancer tumor cells.

    Science.gov (United States)

    Hernández-Luna, Marco Antonio; Díaz de León-Ortega, Ricardo; Hernández-Cueto, Daniel Dimitri; Gaxiola-Centeno, Ricardo; Castro-Luna, Raúl; Martínez-Cristóbal, Leonel; Huerta-Yépez, Sara; Luria-Pérez, Rosendo

    Tumor cell resistance to chemotherapy agents is one of the main problems in the eradication of different neoplasias. One of the mechanisms of this process is the overexpression of anti-apoptotic proteins such as Bcl-2 and Bcl- XL ; blocking the activity of these proteins may contribute to the sensitization of tumor cells and allow the adequate effects of chemotherapeutic drugs. This study adressed the transfection of prostate cancer cells (PC3) with a plasmid encoding a recombinant protein with an antagonist peptide from the BH3 region of the Bax protein fused to the GFP reporter protein (BaxGFP). This protein induced apoptosis of these tumor cells; further, selective transport of this plasmid to the tumor cell with Salmonella enterica serovar Typhimurium (strain SL3261), a live-attenuated bacterial vector, can induce sensitization of the tumor cell to the action of drugs such as cisplatin, through a process known as bactofection. These results suggest that Salmonella enterica can be used as a carrier vector of nucleotide sequences encoding heterologous molecules used in antitumor therapy. Copyright © 2016 Hospital Infantil de México Federico Gómez. Publicado por Masson Doyma México S.A. All rights reserved.

  2. Cardiovascular molecular imaging of apoptosis

    International Nuclear Information System (INIS)

    Wolters, S.L.; Reutelingsperger, C.P.M.; Corsten, M.F.; Hofstra, L.; Narula, J.

    2007-01-01

    Molecular imaging strives to visualise processes at the molecular and cellular level in vivo. Understanding these processes supports diagnosis and evaluation of therapeutic efficacy on an individual basis and thereby makes personalised medicine possible. Apoptosis is a well-organised mode of cell suicide that plays a role in cardiovascular diseases (CVD). Apoptosis is associated with loss of cardiomyocytes following myocardial infarction, atherosclerotic plaque instability, congestive heart failure and allograft rejection of the transplanted heart. Thus, apoptosis constitutes an attractive target for molecular imaging of CVD. Our current knowledge about the molecular players and mechanisms underlying apoptosis offers a rich palette of potential molecular targets for molecular imaging. However, only a few have been successfully developed so far. This review highlights aspects of the molecular machinery and biochemistry of apoptosis relevant to the development of molecular imaging probes. It surveys the role of apoptosis in four major areas of CVD and portrays the importance and future perspectives of apoptosis imaging. The annexin A5 imaging protocol is emphasised since it is the most advanced protocol to measure apoptosis in both preclinical and clinical studies. (orig.)

  3. Cardiovascular molecular imaging of apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Wolters, S.L.; Reutelingsperger, C.P.M. [Maastricht University, Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht (Netherlands); Corsten, M.F.; Hofstra, L. [Maastricht University, Department of Cardiology, Cardiovascular Research Institute Maastricht, P.O. Box 616, Maastricht (Netherlands); Narula, J. [University of California Irvine, Department of Cardiology, Irvine (United States)

    2007-06-15

    Molecular imaging strives to visualise processes at the molecular and cellular level in vivo. Understanding these processes supports diagnosis and evaluation of therapeutic efficacy on an individual basis and thereby makes personalised medicine possible. Apoptosis is a well-organised mode of cell suicide that plays a role in cardiovascular diseases (CVD). Apoptosis is associated with loss of cardiomyocytes following myocardial infarction, atherosclerotic plaque instability, congestive heart failure and allograft rejection of the transplanted heart. Thus, apoptosis constitutes an attractive target for molecular imaging of CVD. Our current knowledge about the molecular players and mechanisms underlying apoptosis offers a rich palette of potential molecular targets for molecular imaging. However, only a few have been successfully developed so far. This review highlights aspects of the molecular machinery and biochemistry of apoptosis relevant to the development of molecular imaging probes. It surveys the role of apoptosis in four major areas of CVD and portrays the importance and future perspectives of apoptosis imaging. The annexin A5 imaging protocol is emphasised since it is the most advanced protocol to measure apoptosis in both preclinical and clinical studies. (orig.)

  4. Studies on hematopoietic cell apoptosis and the relative gene expression in irradiated mouse bone marrow

    International Nuclear Information System (INIS)

    Peng Ruiyun; Wang Dewen; Xiong Chengqi; Gao Yabing; Yang Hong; Cui Yufang; Wang Baozhen

    2001-01-01

    Objective: To study apoptosis and expressions bcl-2 and p53 in irradiated mouse bone marrow. Methods: LACA mice were irradiated with 60 Co γ-rays. By means of in situ terminal labelling, in situ hybridization and image analysis, the authors studied radiation-induced apoptosis of hematopoietic cells and the expressions of bcl-2 and p53. Results: The characteristics of apoptosis appeared in hematopoietic cells at 6 hrs after irradiation. The expression of bcl-2 was obviously decreased when apoptosis of hematopoietic cells occurred, whereas it increased in the early recovery phase; p53 protein increased during both apoptosis of hematopoietic cells and the recovery phase, and mutant type p53 DNA was positive only in the recovery phase. Conclusion: Radiation may induced apoptosis of hematopoietic cells in a dose-dependent manner; Both bcl-2 and p53 genes play an important role in apoptosis and recovery phase

  5. Spironolactone induces apoptosis in human mononuclear cells. Association between apoptosis and cytokine suppression

    DEFF Research Database (Denmark)

    Mikkelsen, Martin; Sønder, S U; Nersting, J

    2006-01-01

    Spironolactone (SPIR) has been described to suppress accumulation of pro-inflammatory cytokines. Here, the suppression of TNF-alpha in lipopolysaccharide (LPS)-stimulated mononuclear cell cultures was confirmed. However, SPIR was also found to induce apoptosis, prompting the investigations...... of a possible association between the two effects: The apoptosis-inducing and the cytokine-suppressive effects of SPIR correlated with regard to the effective concentration range. Also, pre-incubation experiments demonstrated a temporal separation of the two effects of ... preceding apoptosis. An association between the two effects was also seen when testing several SPIR analogues. Contrary to TNF-alpha, the levels of IL-1beta increased in SPIR-treated cultures. However, the amount of IL-1beta in the supernatants depended upon the order of SPIR and LPS addition, as IL-1beta...

  6. Clinical predictors and hemodynamic consequences of elevated peripheral chemosensitivity in optimally treated men with chronic systolic heart failure.

    Science.gov (United States)

    Niewinski, Piotr; Engelman, Zoar J; Fudim, Marat; Tubek, Stanislaw; Paleczny, Bartlomiej; Jankowska, Ewa A; Banasiak, Waldemar; Sobotka, Paul A; Ponikowski, Piotr

    2013-06-01

    Augmented peripheral chemoreflex response is an important mechanism in the pathophysiology of chronic heart failure (CHF). This study characterizes prevalence and clinical predictors of this phenomenon in optimally managed male CHF patients, and seeks to describe the hemodynamic consequences of chemoreceptor hypersensitivity. Thirty-four optimally managed CHF patients and 16 control subjects were prospectively studied. Hypoxic ventilatory response (HVR)-a measure of peripheral chemosensitivity-was calculated with the use of short nitrogen gas administrations. Systolic blood pressure (SBP) and heart rate (HR) following transient hypoxic challenges were recorded with a Nexfin monitor. Hemodynamic responses to hypoxia were expressed by the linear slopes between oxygen saturation (%) and SBP (mm Hg) or HR (beats/min). Elevated HVR was present in 15 (44%) of the CHF patients. Patients with elevated HVR exhibited higher levels of N-terminal pro-B-type natriuretic peptide, lower left ventricular ejection fraction, and higher prevalence of atrial fibrillation. CHF patients with elevated HVR had significantly greater SBP and HR responses to hypoxia than CHF patients with normal HVR. Despite comprehensive pharmacotherapy, elevated HVR is prevalent in CHF patients, related to severity of the disease and associated with augmented hemodynamic responses to hypoxia. CHF patients with elevated HVR may be prone to unfavorable hemodynamic changes. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. Prenylated Chalcone 2 Acts as an Antimitotic Agent and Enhances the Chemosensitivity of Tumor Cells to Paclitaxel

    Directory of Open Access Journals (Sweden)

    Joana Fonseca

    2016-07-01

    Full Text Available We previously reported that prenylated chalcone 2 (PC2, the O-prenyl derivative (2 of 2′-hydroxy-3,4,4′,5,6′-pentamethoxychalcone (1, induced cytotoxicity of tumor cells via disruption of p53-MDM2 interaction. However, the cellular changes through which PC2 exerts its cytotoxic activity and its antitumor potential, remain to be addressed. In the present work, we aimed to (i characterize the effect of PC2 on mitotic progression and the underlying mechanism; and to (ii explore this information to evaluate its ability to sensitize tumor cells to paclitaxel in a combination regimen. PC2 was able to arrest breast adenocarcinoma MCF-7 and non-small cell lung cancer NCI-H460 cells in mitosis. All mitosis-arrested cells showed collapsed mitotic spindles with randomly distributed chromosomes, and activated spindle assembly checkpoint. Live-cell imaging revealed that the compound induced a prolonged delay (up to 14 h in mitosis, culminating in massive cell death by blebbing. Importantly, PC2 in combination with paclitaxel enhanced the effect on cell growth inhibition as determined by cell viability and proliferation assays. Our findings demonstrate that the cytotoxicity induced by PC2 is mediated through antimitotic activity as a result of mitotic spindle damage. The enhancement effects of PC2 on chemosensitivity of cancer cells to paclitaxel encourage further validation of the clinical potential of this combination.

  8. Establishment, characterization, chemosensitivity, and radiosensitivity of two different cell lines derived from a human breast cancer biopsy

    International Nuclear Information System (INIS)

    Gioanni, J.; Courdi, A.; Lalanne, C.M.; Fischel, J.L.; Zanghellini, E.; Lambert, J.C.; Ettore, F.; Namer, M.

    1985-01-01

    In vitro culture of a human breast cancer biopsy fragment gave rise to two permanent cell lines, CAL 18 A and CAL 18 B, which were differentiated by both morphological and ultrastructural analysis. The karyotypic and growth properties of these two cell lines also differed, providing further evidence of cell heterogeneity within a given tumor. Both cell lines lost their hormone receptors in vitro. CAL 18 A cells grew in agar and were tumorigenic after inoculation into nude mice; neither of these properties was observed in CAL 18 B cells. The chemosensitivity of 12 antineoplastic drugs was assessed by a short-term assay, using inhibition of tritiated thymidine incorporation by the cells after contact with the drugs as the end point. Only a few drugs were active at moderate concentrations. The overall responses of both cell lines were similar. The cell survival curves, established by the colony method following a single dose of radiation, were also very similar, despite the greater heterogeneity of CAL 18 B cells. The two cell lines appear to be interrelated, since CAL 18 B cells were occasionally observed to emerge from CAL 18 A clones, suggesting that malignant cell redifferentiation may occur spontaneously in vitro

  9. A hybrid mathematical modeling approach of the metabolic fate of a fluorescent sphingolipid analogue to predict cancer chemosensitivity.

    Science.gov (United States)

    Molina-Mora, J A; Kop-Montero, M; Quirós-Fernández, I; Quiros, S; Crespo-Mariño, J L; Mora-Rodríguez, R A

    2018-04-13

    Sphingolipid (SL) metabolism is a complex biological system that produces and transforms ceramides and other molecules able to modulate other cellular processes, including survival or death pathways key to cell fate decisions. This signaling pathway integrates several types of stress signals, including chemotherapy, into changes in the activity of its metabolic enzymes, altering thereby the cellular composition of bioactive SLs. Therefore, the SL pathway is a promising sensor of chemosensitivity in cancer and a target hub to overcome resistance. However, there is still a gap in our understanding of how chemotherapeutic drugs can disturb the SL pathway in order to control cellular fate. We propose to bridge this gap by a systems biology approach to integrate i) a dynamic model of SL analogue (BODIPY-FL fluorescent-sphingomyelin analogue, SM-BOD) metabolism, ii) a Gaussian mixture model (GMM) of the fluorescence features to identify how the SL pathway senses the effect of chemotherapy and iii) a fuzzy logic model (FLM) to associate SL composition with cell viability by semi-quantitative rules. Altogether, this hybrid model approach was able to predict the cell viability of double experimental perturbations with chemotherapy, indicating that the SL pathway is a promising sensor to design strategies to overcome drug resistance in cancer. Copyright © 2018 Elsevier Ltd. All rights reserved.

  10. Activity of mevalonate pathway inhibitors against breast and ovarian cancers in the ATP-based tumour chemosensitivity assay

    International Nuclear Information System (INIS)

    Knight, Louise A; Kurbacher, Christian M; Glaysher, Sharon; Fernando, Augusta; Reichelt, Ralf; Dexel, Susanne; Reinhold, Uwe; Cree, Ian A

    2009-01-01

    Previous data suggest that lipophilic statins such as fluvastatin and N-bisphosphonates such as zoledronic acid, both inhibitors of the mevalonate metabolic pathway, have anti-cancer effects in vitro and in patients. We have examined the effect of fluvastatin alone and in combination with zoledronic acid in the ATP-based tumour chemosensitivity assay (ATP-TCA) for effects on breast and ovarian cancer tumour-derived cells. Both zoledronic acid and fluvastatin showed activity in the ATP-TCA against breast and ovarian cancer, though fluvastatin alone was less active, particularly against breast cancer. The combination of zoledronic acid and fluvastatin was more active than either single agent in the ATP-TCA with some synergy against breast and ovarian cancer tumour-derived cells. Sequential drug experiments showed that pre-treatment of ovarian tumour cells with fluvastatin resulted in decreased sensitivity to zoledronic acid. Addition of mevalonate pathway components with zoledronic acid with or without fluvastatin showed little effect, while mevalonate did reduced inhibition due to fluvastatin. These data suggest that the combination of zoledronic acid and fluvastatin may have activity against breast and ovarian cancer based on direct anti-cancer cell effects. A clinical trial to test this is in preparation

  11. Arctigenin enhances chemosensitivity of cancer cells to cisplatin through inhibition of the STAT3 signaling pathway.

    Science.gov (United States)

    Yao, Xiangyang; Zhu, Fenfen; Zhao, Zhihui; Liu, Chang; Luo, Lan; Yin, Zhimin

    2011-10-01

    Arctigenin is a dibenzylbutyrolactone lignan isolated from Bardanae fructus, Arctium lappa L, Saussureamedusa, Torreya nucifera, and Ipomea cairica. It has been reported to exhibit anti-inflammatory activities, which is mainly mediated through its inhibitory effect on nuclear transcription factor-kappaB (NF-κB). But the role of arctigenin in JAK-STAT3 signaling pathways is still unclear. In present study, we investigated the effect of arctigenin on signal transducer and activator of transcription 3 (STAT3) pathway and evaluated whether suppression of STAT3 activity by arctigenin could sensitize cancer cells to a chemotherapeutic drug cisplatin. Our results show that arctigenin significantly suppressed both constitutively activated and IL-6-induced STAT3 phosphorylation and subsequent nuclear translocation in cancer cells. Inhibition of STAT3 tyrosine phosphorylation was found to be achieved through suppression of Src, JAK1, and JAK2, while suppression of STAT3 serine phosphorylation was mediated by inhibition of ERK activation. Pervanadate reversed the arctigenin-induced downregulation of STAT3 activation, suggesting the involvement of a protein tyrosine phosphatase. Indeed, arctigenin can obviously induce the expression of the PTP SHP-2. Furthermore, the constitutive activation level of STAT3 was found to be correlated to the resistance of cancer cells to cisplatin-induced apoptosis. Arctigenin dramatically promoted cisplatin-induced cell death in cancer cells, indicating that arctigenin enhanced the sensitivity of cancer cells to cisplatin mainly via STAT3 suppression. These observations suggest a novel anticancer function of arctigenin and a potential therapeutic strategy of using arctigenin in combination with chemotherapeutic agents for cancer treatment. Copyright © 2011 Wiley-Liss, Inc.

  12. A systems analysis of the chemosensitivity of breast cancer cells to the polyamine analogue PG-11047

    Energy Technology Data Exchange (ETDEWEB)

    Kuo, Wen-Lin; Das, Debopriya; Ziyad, Safiyyah; Bhattacharya, Sanchita; Gibb, William J.; Heiser, Laura M.; Sadanandam, Anguraj; Fontenay, Gerald V.; Hu, Zhi; Wang, Nicholas J.; Bayani, Nora; Feiler, Heidi S.; Neve, Richard M.; Wyrobek, Andrew J.; Spellman, Paul T.; Marton, Laurence J.; Gray, Joe W.

    2009-11-14

    Polyamines regulate important cellular functions and polyamine dysregulation frequently occurs in cancer. The objective of this study was to use a systems approach to study the relative effects of PG-11047, a polyamine analogue, across breast cancer cells derived from different patients and to identify genetic markers associated with differential cytotoxicity. A panel of 48 breast cell lines that mirror many transcriptional and genomic features present in primary human breast tumours were used to study the antiproliferative activity of PG-11047. Sensitive cell lines were further examined for cell cycle distribution and apoptotic response. Cell line responses, quantified by the GI50 (dose required for 50% relative growth inhibition) were correlated with the omic profiles of the cell lines to identify markers that predict response and cellular functions associated with drug sensitivity. The concentrations of PG-11047 needed to inhibit growth of members of the panel of breast cell lines varied over a wide range, with basal-like cell lines being inhibited at lower concentrations than the luminal cell lines. Sensitive cell lines showed a significant decrease in S phase fraction at doses that produced little apoptosis. Correlation of the GI50 values with the omic profiles of the cell lines identified genomic, transcriptional and proteomic variables associated with response. A 13-gene transcriptional marker set was developed as a predictor of response to PG-11047 that warrants clinical evaluation. Analyses of the pathways, networks and genes associated with response to PG-11047 suggest that response may be influenced by interferon signaling and differential inhibition of aspects of motility and epithelial to mesenchymal transition.

  13. Roles of Apoptosis and Cellular Senescence in Cancer and Aging.

    Science.gov (United States)

    Cerella, Claudia; Grandjenette, Cindy; Dicato, Mario; Diederich, Marc

    2016-01-01

    Cancer and aging are two similar processes representing the final outcome of timedependent accumulation of various irreversible dysfunctions, mainly caused by stress-induced DNA and cellular damages. Apoptosis and senescence are two types of cellular response to damages that are altered in both cancer and aging, albeit through different mechanisms. Carcinogenesis is associated with a progressive reduction in the ability of the cells to trigger apoptosis and senescence. In contrast, in aging tissues, there is an increased accumulation of senescent cells, and the nature of apoptosis deregulation varies depending on the tissue. Thus, the prevailing model suggests that apoptosis and cellular senescence function as two essential tumor-suppressor mechanisms, ensuring the health of the individual during early and reproductive stages of life, but become detrimental and promote aging later in life. The recent discovery that various anticancer agents, including canonical inducers of apoptosis, act also as inducers of cellular senescence indicates that pro-senescence strategies may have applications in cancer prevention therapy. Therefore, dissection of the mechanisms mediating the delicate balance between apoptosis and cellular senescence will be beneficial in the therapeutic exploitation of both processes in the development of future anticancer and anti-aging strategies, including minimizing the side effects of such strategies. Here, we provide an overview of the roles of apoptosis and cellular senescence in cancer and aging.

  14. Evaluation of preventive and therapeutic activity of novel non-steroidal anti-inflammatory drug, CG100649, in colon cancer: Increased expression of TNF-related apoptosis-inducing ligand receptors enhance the apoptotic response to combination treatment with TRAIL.

    Science.gov (United States)

    Woo, Jong Kyu; Kang, Ju-Hee; Jang, Yeong-Su; Ro, Seonggu; Cho, Joong Myung; Kim, Hwan-Mook; Lee, Sang-Jin; Oh, Seung Hyun

    2015-04-01

    Non-steroidal anti-inflammatory drugs (NSAIDs) have been suggested as the potential new class of preventive or therapeutic antitumor agents. The aim of the present study was to evaluate the antitumor activity of the novel NSAID, CG100649. CG100649 is a novel NSAID dual inhibitor for COX-2 and carbonic anhydrase (CA)-I/-II. In the present study, we investigated the alternative mechanism by which CG100649 mediated suppression of the colon cancer growth and development. The anchorage‑dependent and -independent clonogenic assay showed that CG100649 inhibited the clonogenicity of human colon cancer cells. The flow cytometric analysis showed that CG100649 induced the G2/M cell cycle arrest in colon cancer cells. Animal studies showed that CG100649 inhibited the tumor growth in colon cancer xenograft in nude mice. Furthermore, quantitative PCR and FACS analysis demonstrated that CG100649 upregulated the expression of TNF-related apoptosis-inducing ligand (TRAIL) receptors (DR4 and DR5) but decreased the expression of decoy receptors (DcR1 and DcR2) in colon cancer cells. The results showed that CG100649 treatment sensitized TRAIL‑mediated growth suppression and apoptotic cell death. The combination treatment resulted in significant repression of the intestinal polyp formation in APCmin/+ mice. Our data clearly demonstrated that CG100649 contains preventive and therapeutic activity for colon cancer. The present study may be useful for identification of the potential benefit of the NSAID CG100649, for the achievement of a better treatment response in colon cancer.

  15. Proliferation and apoptosis in infection with infectious bursal disease virus: a flow cytometric study.

    Science.gov (United States)

    Ojeda, F; Skardova, I; Guarda, M I; Ulloa, J; Folch, H

    1997-01-01

    Programmed cell death, or apoptosis, is involved in the normal physiology of many immunocompetent organs, including lymphocytes of the bursa of Fabricius in chickens. Involvement of apoptosis has also been described in some viral diseases such as AIDS. The purpose of this work was to study the potential role of apoptosis in the pathogenesis of Gumboro disease in the bursa of Fabricius. Our results show that 1-3 days after infection of young chickens with infectious bursal disease virus, the number of apoptotic cells increases and cellularity and proliferation decrease. Because of the dynamic nature of bursal lymphocyte populations and the involvement of apoptosis in lymphocyte cell physiology, the increased level of cells undergoing apoptosis may be due to an impairment in the withdrawal of apoptotic cells. A concomitant increase in macrophages in infected bursae and a dramatic decrease in cellularity suggest that an increase in apoptosis may be an important cause of cell depletion.

  16. Bisphenol A induces spermatocyte apoptosis in rare minnow Gobiocypris rarus

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Yingying; Cheng, Mengqian; Wu, Lang; Zhang, Guo; Wang, Zaizhao, E-mail: zzwang@nwsuaf.edu.cn

    2016-10-15

    Highlights: • Adult male G. rarus were exposed to 225 μg/L BPA for 7, 21 and 63 days. • BPA could induce spermatocyte apoptosis in rare minnow testis. • The mitochondrial apoptotic pathway participated in the germ cell apoptosis. • The spermatocyte apoptosis was likely initiated by BPA induced meiosis arrest. - Abstract: Bisphenol A (BPA) is an endocrine disruptor, and could induce germ cells apoptosis in the testis of mammals. But whether it could affect fish in the same mechanism has not’ been studied till now. In the present study, to investigate the influence of BPA on testis germ cells in fish, adult male rare minnow Gobiocypris rarus were exposed to 225 μg L{sup −1} (0.99 μM) BPA for 1, 3 and 9 weeks. Through TdT-mediated dUTP nick end labeling (TUNEL) and transmission electron microscope (TEM) analysis, we found that the amount of apoptotic spermatocytes significantly increased in a time dependent manner following BPA exposure. Western Blot results showed that the ratio of Bcl2/Bax, the important apoptosis regulators in intrinsic mitochondrial apoptotic pathway, was significantly decreased. qPCR showed that mRNA expression of several genes in mitochondrial apoptotic pathway including bcl2, bax, casp9, cytc and mcl1b were significantly changed following BPA exposure. In addition, mRNA expression of meiosis regulation genes (kpna7 and wee2), and genes involved in both apoptosis and meiosis (birc5, ccna1, and gsa1a) were also affected by BPA. Taken together, the present study demonstrated that BPA could induce spermatocytes apoptosis in rare minnow testis, and the apoptosis was probably under regulation of intrinsic mitochondrial apoptotic pathway. Moreover, the spermatocyte apoptosis was likely initiated by BPA induced meiosis arrest.

  17. Apoptosis in HEp-2 cells infected with Ureaplasma diversum

    Directory of Open Access Journals (Sweden)

    Aline Teixeira Amorim

    2014-01-01

    Full Text Available BACKGROUND: Bacterial pathogens have many strategies for infecting and persisting in host cells. Adhesion, invasion and intracellular life are important features in the biology of mollicutes. The intracellular location ofUreaplasma diversum may trigger disturbances in the host cell. This includes activation or inhibition of pro and anti-apoptotic factors, which facilitate the development of host damage. The aim of the present study was to associate U. diversum infection in HEp-2 cells and apoptosis induction. Cells were infected for 72hs with four U. diversum clinical isolates and an ATCC strain. The U. diversuminvasion was analyzed by Confocal Laser Scanning Microscopy and gentamicin invasion assay. The apoptosis was evaluated using pro-apoptotic and anti-apoptotic gene expression, and FITC Annexin V/Dead Cell Apoptosis Kit. RESULTS: The number of internalized ureaplasma in HEp-2 cells increased significantly throughout the infection. The flow cytometry analysis with fluorochromes to detect membrane depolarization and gene expression for caspase 2, 3 and 9 increased in infected cells after 24 hours. However, after 72 hours a considerable decrease of apoptotic cells was observed. CONCLUSIONS: The data suggests that apoptosis may be initially induced by some isolates in association with HEp-2 cells, but over time, there was no evidence of apoptosis in the presence of ureaplasma and HEp-2 cells. The initial increase and then decrease in apoptosis could be related to bacterial pathogen-associated molecular pattern (PAMPS. Moreover, the isolates of U. diversum presented differences in the studied parameters for apoptosis. It was also observed that the amount of microorganisms was not proportional to the induction of apoptosis in HEp-2 cells.

  18. Apoptosis in HEp-2 cells infected with Ureaplasma diversum.

    Science.gov (United States)

    Amorim, Aline Teixeira; Marques, Lucas Miranda; Santos, Angelita Maria Oliveira Gusmão; Martins, Hellen Braga; Barbosa, Maysa Santos; Rezende, Izadora Souza; Andrade, Ewerton Ferraz; Campos, Guilherme Barreto; Lobão, Tássia Neves; Cortez, Beatriz Araujo; Monezi, Telma Alvez; Machado-Santelli, Glaucia Maria; Timenetsky, Jorge

    2014-09-04

    Bacterial pathogens have many strategies for infecting and persisting in host cells. Adhesion, invasion and intracellular life are important features in the biology of mollicutes. The intracellular location of Ureaplasma diversum may trigger disturbances in the host cell. This includes activation or inhibition of pro and anti-apoptotic factors, which facilitate the development of host damage. The aim of the present study was to associate U. diversum infection in HEp-2 cells and apoptosis induction. Cells were infected for 72hs with four U. diversum clinical isolates and an ATCC strain. The U. diversum invasion was analyzed by Confocal Laser Scanning Microscopy and gentamicin invasion assay. The apoptosis was evaluated using pro-apoptotic and anti-apoptotic gene expression, and FITC Annexin V/Dead Cell Apoptosis Kit. The number of internalized ureaplasma in HEp-2 cells increased significantly throughout the infection. The flow cytometry analysis with fluorochromes to detect membrane depolarization and gene expression for caspase 2, 3 and 9 increased in infected cells after 24 hours. However, after 72 hours a considerable decrease of apoptotic cells was observed. The data suggests that apoptosis may be initially induced by some isolates in association with HEp-2 cells, but over time, there was no evidence of apoptosis in the presence of ureaplasma and HEp-2 cells. The initial increase and then decrease in apoptosis could be related to bacterial pathogen-associated molecular pattern (PAMPS). Moreover, the isolates of U. diversum presented differences in the studied parameters for apoptosis. It was also observed that the amount of microorganisms was not proportional to the induction of apoptosis in HEp-2 cells.

  19. X-linked inhibitor of apoptosis regulates T cell effector function

    DEFF Research Database (Denmark)

    Zehntner, Simone P; Bourbonnière, Lyne; Moore, Craig S

    2007-01-01

    To understand how the balance between pro- and anti-apoptotic signals influences effector function in the immune system, we studied the X-linked inhibitor of apoptosis (XIAP), an endogenous regulator of cellular apoptosis. Real-time PCR showed increased XIAP expression in blood of mice...... and oligodendrocytes were not affected; neither did apoptosis increase in liver, where XIAP knockdown also occurred. ASO-XIAP increased susceptibility of T cells to activation-induced apoptosis in vitro. Our results identify XIAP as a critical controller of apoptotic susceptibility of effector T cell function...

  20. Methylselenium and Prostate Cancer Apoptosis

    National Research Council Canada - National Science Library

    Lu, Junxuan

    2007-01-01

    The purpose of this research is to gain a better understanding of the biochemical pathways and molecular targets for the selective induction of apoptosis signaling and execution of prostate cancer (PCa...

  1. Methylselenium and Prostate Cancer Apoptosis

    National Research Council Canada - National Science Library

    Lu, Junxuan

    2005-01-01

    The purpose of this research is to gain a better understanding of the biochemical pathways and molecular targets for the selective induction of apoptosis signaling and execution of prostate cancer (PCa...

  2. Methylselenium and Prostate Cancer Apoptosis

    National Research Council Canada - National Science Library

    Lu, Junxuan

    2006-01-01

    The purpose of this research is to gain a better understanding of the biochemical pathways and molecular targets for the selective induction of apoptosis signaling and execution of prostate cancer (PCa...

  3. Methylselenium and Prostate Cancer Apoptosis

    National Research Council Canada - National Science Library

    Lu, Junxuan

    2004-01-01

    The purpose of this research is to gain a better understanding of the biochemical pathways and molecular targets for the selective induction of apoptosis signaling and execution of PCa cells by methyl selenium (Se)/selenol...

  4. Methylselenium and Prostate Cancer Apoptosis

    National Research Council Canada - National Science Library

    Lu, Junxuan

    2008-01-01

    The purpose of this research is to gain a better understanding of the biochemical pathways and molecular targets for the selective induction of apoptosis signaling and execution of prostate cancer (PCa...

  5. Methylselenium and Prostate Cancer Apoptosis

    National Research Council Canada - National Science Library

    Lu, Junxuan

    2003-01-01

    The purpose of this research is to gain a better understanding of the biochemical pathways and molecular targets for the selective induction of apoptosis signaling and execution of PCa cells by methyl selenium (Se...

  6. [Taurine induces apoptosis in pulmonary artery smooth muscle cells].

    Science.gov (United States)

    Zhang, Xiaodan; Sheng, Jiejing; Zhang, Caixiaz; Zhao, Fenghua

    2012-03-01

    To study the effect of taurine on apoptosis in PASMCs, and whether the death-receptor pathway act in the mechanism. Culture the PASMCs, and divided the cells into control, SD. Acridine orange(AO) assay and western-blot analysis on the expression of Bax, Bcl-2, Procaspase-3 and Fas were used to study the mechanism. A major finding of this study is that the Tau effects many apoptosis index, such as increasing the expression of Bax and Fas, decreasing the expression of Procaspase-3, and Bcl-2, accrescencing the mitochondrial depolarization, causing the nuclear shrinkage, all these datas demonstrated that Tau induced the apoptosis in pulmonary artery smooth muscle cells through mitochondrial-dependent pathway. Tau induces the apoptosis in pulmonary artery smooth muscle cells through death-receptor.

  7. Accelerated apoptosis of neutrophils in familial Mediterranean fever

    DEFF Research Database (Denmark)

    Manukyan, Gayane; Aminov, Rustam; Hakobyan, Gagik

    2015-01-01

    not revealed any conventional mechanisms contributing to the enhanced apoptotic rate of neutrophils in FMF. Although the exact molecular mechanisms of accelerated neutrophil apoptosis in FMF remain unknown, it may provide a protection against excessive inflammation and tissue damage due to a massive...... of systemic neutrophil apoptosis as well as the levels of proteins involved in apoptosis were investigated ex vivo in patients with FMF using flow cytometry and RT-qPCR. The freshly collected neutrophils from the patients in FMF remission displayed a significantly larger number of cells spontaneously entering...... apoptosis compared to control (6.27 ± 2.14 vs. 1.69 ± 0.18%). This elevated ratio was retained after 24 h incubation of neutrophils in the growth medium (32.4 ± 7.41 vs. 7.65 ± 1.32%). Correspondingly, the mRNA level for caspase-3 was also significantly increased under these conditions. In response...

  8. Environmentally induced return to juvenile-like chemosensitivity in the respiratory control system of adult bullfrog, Lithobates catesbeianus.

    Science.gov (United States)

    Santin, Joseph M; Hartzler, Lynn K

    2016-11-01

    The degree to which developmental programmes or environmental signals determine physiological phenotypes remains a major question in physiology. Vertebrates change environments during development, confounding interpretation of the degree to which development (i.e. permanent processes) or phenotypic plasticity (i.e. reversible processes) produces phenotypes. Tadpoles mainly breathe water for gas exchange and frogs may breathe water or air depending on their environment and are, therefore, exemplary models to differentiate the degree to which life-stage vs. environmental context drives developmental phenotypes associated with neural control of lung breathing. Using isolated brainstem preparations and patch clamp electrophysiology, we demonstrate that adult bullfrogs acclimatized to water-breathing conditions do not exhibit CO 2 and O 2 chemosensitivity of lung breathing, similar to water-breathing tadpoles. Our results establish that phenotypes associated with developmental stage may arise from plasticity per se and suggest that a developmental trajectory coinciding with environmental change obscures origins of stage-dependent physiological phenotypes by masking plasticity. An unanswered question in developmental physiology is to what extent does the environment vs. a genetic programme produce phenotypes? Developing animals inhabit different environments and switch from one to another. Thus a developmental time course overlapping with environmental change confounds interpretations as to whether development (i.e. permanent processes) or phenotypic plasticity (i.e. reversible processes) generates phenotypes. Tadpoles of the American bullfrog, Lithobates catesbeianus, breathe water at early life-stages and minimally use lungs for gas exchange. As adults, bullfrogs rely on lungs for gas exchange, but spend months per year in ice-covered ponds without lung breathing. Aquatic submergence, therefore, removes environmental pressures requiring lung breathing and enables

  9. Chemosensitivity of human small cell carcinoma of the lung detected by flow cytometric DNA analysis of drug-induced cell cycle perturbations in vitro

    DEFF Research Database (Denmark)

    Engelholm, S A; Spang-Thomsen, M; Vindeløv, L L

    1986-01-01

    A method based on detection of drug-induced cell cycle perturbation by flow cytometric DNA analysis has previously been described in Ehrlich ascites tumors as a way to estimate chemosensitivity. The method is extended to test human small-cell carcinoma of the lung. Three tumors with different...... sensitivities to melphalan in nude mice were used. Tumors were disaggregated by a combined mechanical and enzymatic method and thereafter have incubated with different doses of melphalan. After incubation the cells were plated in vitro on agar, and drug induced cell cycle changes were monitored by flow...

  10. MPO density in primary cancer biopsies of ovarian carcinoma enhances the indicative value of IL-17 for chemosensitivity

    International Nuclear Information System (INIS)

    Droeser, Raoul A.; Mechera, Robert; Däster, Silvio; Weixler, Benjamin; Kraljević, Marko; Delko, Tarik; Güth, Uwe; Stadlmann, Sylvia; Terracciano, Luigi; Singer, Gad

    2016-01-01

    Cancer of the ovary is mostly discovered at a late stage and cannot be removed by surgery alone. Therefore surgery is usually followed by adjuvant chemotherapy. However, few reliable biomarkers exist to predict response to chemotherapy of ovarian cancer. Previously, we could demonstrate that IL-17 density is indicative for chemosensitivity. This study focuses on the predictive value of myeloperoxidase (MPO) concerning response to chemotherapy of ovarian cancer. Biopsies of mostly high-grade primary serous ovarian carcinomas and their matched recurrences were stained with MPO after fixation in formalin and embedding in paraffin. For this staining the technique of tissue-microarray was used. Recurrence within 6 months of the completion of platinum-based chemotherapy was defined as chemoresistance as previously publised. Data for MPO could be analyzed in 92 biopsies. MPO and IL-17 positive immune cells correlated significantly in biopsies of primary and recurrent carcinomas (r s = 0.41; p = 0.004 and r s = 0.40; p = 0.007, respectively). MPO expression alone did not predict response to chemotherapy, but in multivariate cox regression analysis including age, residual disease, number of chemotherapy cycles, FIGO classification and combined categorized MPO and IL-17 cell densities of primary cancer biopsies, the combination of both immune markers was an independent prognostic factor for recurrence-free survival (p = 0.013, HR = .23, 95CI = 0.07–0.73). There was no chemoresistant patient in the subgroup of MPO + IL-17+, neither in primary nor in recurrent cancer biopsies. High MPO positive cell density enhances the indicative value of IL-17 for response to chemotherapy in ovarian carcinoma. Although, these results have to be validated in a larger cohort

  11. Human TRPA1 is intrinsically cold- and chemosensitive with and without its N-terminal ankyrin repeat domain.

    Science.gov (United States)

    Moparthi, Lavanya; Survery, Sabeen; Kreir, Mohamed; Simonsen, Charlotte; Kjellbom, Per; Högestätt, Edward D; Johanson, Urban; Zygmunt, Peter M

    2014-11-25

    We have purified and reconstituted human transient receptor potential (TRP) subtype A1 (hTRPA1) into lipid bilayers and recorded single-channel currents to understand its inherent thermo- and chemosensory properties as well as the role of the ankyrin repeat domain (ARD) of the N terminus in channel behavior. We report that hTRPA1 with and without its N-terminal ARD (Δ1-688 hTRPA1) is intrinsically cold-sensitive, and thus, cold-sensing properties of hTRPA1 reside outside the N-terminal ARD. We show activation of hTRPA1 by the thiol oxidant 2-((biotinoyl)amino)ethyl methanethiosulfonate (MTSEA-biotin) and that electrophilic compounds activate hTRPA1 in the presence and absence of the N-terminal ARD. The nonelectrophilic compounds menthol and the cannabinoid Δ(9)-tetrahydrocannabiorcol (C16) directly activate hTRPA1 at different sites independent of the N-terminal ARD. The TRPA1 antagonist HC030031 inhibited cold and chemical activation of hTRPA1 and Δ1-688 hTRPA1, supporting a direct interaction with hTRPA1 outside the N-terminal ARD. These findings show that hTRPA1 is an intrinsically cold- and chemosensitive ion channel. Thus, second messengers, including Ca(2+), or accessory proteins are not needed for hTRPA1 responses to cold or chemical activators. We suggest that conformational changes outside the N-terminal ARD by cold, electrophiles, and nonelectrophiles are important in hTRPA1 channel gating and that targeting chemical interaction sites outside the N-terminal ARD provides possibilities to fine tune TRPA1-based drug therapies (e.g., for treatment of pain associated with cold hypersensitivity and cardiovascular disease).

  12. VEGF masks BNIP3-mediated apoptosis of hypoxic endothelial cells.

    Science.gov (United States)

    Jurasz, Paul; Yurkova, Natasha; Kirshenbaum, Lorrie; Stewart, Duncan J

    2011-05-01

    Hypoxia results in the apoptotic death of myocytes, neurons, and epithelial cells, through the actions of Bcl-2 and Nineteen kilodalton Interacting Protein-3 (BNIP3). On the contrary, endothelial cells are especially adept at surviving conditions of oxygen deprivation via up-regulation of vascular endothelial growth factor (VEGF) the most potent endothelial survival factor. Both VEGF and BNIP3 expression are transcriptionally regulated by hypoxia inducible factor and may antagonize each other's affects in endothelial cells (ECs). Since factors that promote and inhibit apoptosis may be expressed at the same time in endothelial cells under hypoxic conditions, we decided to investigate whether VEGF and BNIP3 have opposing actions in endothelial cells. Human microvascular endothelial cells were exposed to hypoxic conditions in a Billups-Rothenburg chamber. Under hypoxic conditions BNIP3 expression by endothelial cells increased as measured by real-time PCR and immunoblot. After 48 h of hypoxia, EC apoptosis was assessed by flow cytometry and was lower than in corresponding normoxia serum starved controls. The increase in EC survival under hypoxic conditions corresponded with an increase in the expression of VEGF. Under normoxic conditions adenoviral BNIP3 over-expression promoted apoptosis of ECs; however, recombinant VEGF (100 pg/ml) antagonized the BNIP3 apoptosis promoting affects. SiRNA knockdown of VEGF expression by hypoxic ECs resulted in increased apoptosis with a concomitant increase in BNIP3 expression. SiRNA knockdown of BNIP3 expression by hypoxic ECs reduced the increase in EC apoptosis as a result of VEGF knockdown. We conclude that under hypoxic conditions VEGF counteracts and masks the apoptosis promoting affects of BNIP3.

  13. Nicotine plus a high-fat diet triggers cardiomyocyte apoptosis.

    Science.gov (United States)

    Sinha-Hikim, Indrani; Friedman, Theodore C; Falz, Mark; Chalfant, Victor; Hasan, Mohammad Kamrul; Espinoza-Derout, Jorge; Lee, Desean L; Sims, Carl; Tran, Peter; Mahata, Sushil K; Sinha-Hikim, Amiya P

    2017-04-01

    Cigarette smoking is an important risk factor for diabetes, cardiovascular disease and non-alcoholic fatty liver disease. The health risk associated with smoking can be aggravated by obesity. Smoking might also trigger cardiomyocyte (CM) apoptosis. Given that CM apoptosis has been implicated as a potential mechanism in the development of cardiomyopathy and heart failure, we characterize the key signaling pathways in nicotine plus high-fat diet (HFD)-induced CM apoptosis. Adult C57BL6 male mice were fed a normal diet (ND) or HFD and received twice-daily intraperitoneal (IP) injections of nicotine (0.75 mg/kg body weight [BW]) or saline for 16 weeks. An additional group of nicotine-treated mice on HFD received twice-daily IP injections of mecamylamine (1 mg/kg BW), a non-selective nicotinic acetylcholine receptor antagonist, for 16 weeks. Nicotine when combined with HFD led to a massive increase in CM apoptosis that was fully prevented by mecamylamine treatment. Induction of CM apoptosis was associated with increased oxidative stress and activation of caspase-2-mediated intrinsic pathway signaling coupled with inactivation of AMP-activated protein kinase (AMPK). Furthermore, nicotine treatment significantly (P nicotine, when combined with HFD, triggers CM apoptosis through the generation of oxidative stress and inactivation of AMPK together with the activation of caspase-2-mediated intrinsic apoptotic signaling independently of FGF21 and SIRT1.

  14. Wogonin Induces Eosinophil Apoptosis and Attenuates Allergic Airway Inflammation

    Science.gov (United States)

    Dorward, David A.; Sharma, Sidharth; Rennie, Jillian; Felton, Jennifer M.; Alessandri, Ana L.; Duffin, Rodger; Schwarze, Jurgen; Haslett, Christopher; Rossi, Adriano G.

    2015-01-01

    Rationale: Eosinophils are key effector cells in allergic diseases, including allergic rhinitis, eczema, and asthma. Their tissue presence is regulated by both recruitment and increased longevity at inflamed sites. Objectives: To investigate the ability of the flavone wogonin to induce eosinophil apoptosis in vitro and attenuate eosinophil-dominant allergic inflammation in vivo in mice. Methods: Human and mouse eosinophil apoptosis in response to wogonin was investigated by cellular morphology, flow cytometry, mitochondrial membrane permeability, and pharmacological caspase inhibition. Allergic lung inflammation was modeled in mice sensitized and challenged with ovalbumin. Bronchoalveolar lavage (BAL) and lung tissue were examined for inflammation, mucus production, and inflammatory mediator production. Airway hyperresponsiveness to aerosolized methacholine was measured. Measurements and Main Results: Wogonin induced time- and concentration-dependent human and mouse eosinophil apoptosis in vitro. Wogonin-induced eosinophil apoptosis occurred with activation of caspase-3 and was inhibited by pharmacological caspase inhibition. Wogonin administration attenuated allergic airway inflammation in vivo with reductions in BAL and interstitial eosinophil numbers, increased eosinophil apoptosis, reduced airway mucus production, and attenuated airway hyperresponsiveness. This wogonin-induced reduction in allergic airway inflammation was prevented by concurrent caspase inhibition in vivo. Conclusions: Wogonin induces eosinophil apoptosis and attenuates allergic airway inflammation, suggesting that it has therapeutic potential for the treatment of allergic inflammation in humans. PMID:25629436

  15. Apoptosis in chondrogenesis of human mesenchymal stem cells: effect of serum and medium supplements.

    Science.gov (United States)

    Wang, Chien-Yuan; Chen, Ling-Lan; Kuo, Pei-Yin; Chang, Jia-Ling; Wang, Yng-Jiin; Hung, Shih-Chieh

    2010-04-01

    Apoptosis is an inevitable process during development and is evident in the formation of articular cartilage and endochondral ossification of growth plate. Mesenchymal stem cells (MSCs) can serve as alternative sources for cell therapy in focal chondral lesions or diffuse osteoarthritis. But there are few, if any, studies investigating apoptosis during chondrogenesis by MSCs. The aim of this study was to find the better condition to prevent apoptosis during chondrogenesis by MSCs. Apoptosis were evaluated in MSCs induced in different chondrogenic media by the use of Annexin V, TUNEL staining, lysosomal labeling with lysotracker and immunostaining of apoptotic markers. We found apparent apoptosis was demonstrated by Annexin V, TUNEL staining and lysosomal labeling during chondrogenesis. Meanwhile, the degree of apoptosis was related to the reagents of the defined chondrogenic medium. Adding serum in medium increased apoptosis, however, TGF-beta1 inhibited apoptosis. The apoptosis was associated with the activation of caspase-3, the increase in the Bax/Bcl-2 ratio, the loss of lysosomal integrity, and the increase of PARP-cleavage. Pro-inflammatory cytokines, IL-1alpha, IL-1beta and TNFalpha did not induce any increase in apoptosis. Interestingly, the inhibition of apoptosis by serum free medium supplemented with ITS was also associated with an increase in the expression of type II collagen, and a decrease in the expression of type X collagen, Runx2, and other osteogenic genes, while TGF-beta1 increased the expression of Sox9, type II and type X collagen and decreased the expression of osteogenic genes. These data suggest apoptosis occurs during chondrogenesis by MSCs by cell death intrinsic pathway activation and this process may be modulated by culture conditions.

  16. Apoptosis and peripheral blood lymphocyte depletion in coeliac disease

    Science.gov (United States)

    Di Sabatino, Antonio; D'Alò, Simona; Millimaggi, Danilo; Ciccocioppo, Rachele; Parroni, Raffaella; Sciarra, Giuseppe; Cifone, Maria Grazia; Corazza, Gino Roberto

    2001-01-01

    In coeliac disease (CD) immunological abnormalities are not confined to the small bowel and it has been suggested that changes in peripheral blood lymphocytes (PBL), such as lymphopenia and increased T-cell activation, may predispose to malignant or autoimmune complications of this condition. In the light of the recent findings about the Fas–Fas ligand (FasL) system in regulating lymphocyte homeostasis, the aim of the present study was to investigate peripheral lymphocyte Fas-mediated apoptosis in CD to establish whether the homeostatic role of apoptosis in peripheral T-cell selection is maintained. Moreover, because a soluble form of Fas has been described to be functionally implicated in the Fas signalling system, suggesting a relationship between some disorders and soluble Fas function, we measured levels of soluble Fas in sera of coeliac patients and analysed the relationship between these levels and the proportions of apoptotic and Fas+ PBL to further explore the function of the Fas–FasL pathway in this condition. Finally, we evaluated whether the increased prevalence of anticardiolipin antibodies, recently described in CD, could be related to PBL apoptosis in this condition. We demonstrated an increased apoptosis and higher levels of Fas and FasL expression in PBL isolated from untreated coeliac patients when compared to treated coeliac patients and controls. In addition, low levels of soluble Fas and a significant positive correlation between anticardiolipin antibodies and PBL apoptosis were found in untreated CD. Then, our results showed an increased susceptibility of PBL to undergo Fas-mediated apoptosis in active CD. This increased apoptosis could be responsible for both lymphopenia and immunogenic exposure of phospholipids with subsequent production of autoantibodies. PMID:11529933

  17. Knockdown of HIF-1α and IL-8 induced apoptosis of hepatocellular carcinoma triggers apoptosis of vascular endothelial cells.

    Science.gov (United States)

    Choi, Sung Hoon; Park, Jun Yong; Kang, Wonseok; Kim, Seung Up; Kim, Do Young; Ahn, Sang Hoon; Ro, Simon Wonsang; Han, Kwang-Hyub

    2016-01-01

    A local hypoxic microenvironment is one of the most important characteristics of solid tumors. Hypoxia inducible factor-1α (HIF-1α) and Interleukin-8 (IL-8) activate tumor survival from hypoxic-induced apoptosis in each pathway. This study aimed to evaluate whether knockdown of HIF-1α and IL-8 induced apoptosis of the hepatocellular carcinoma (HCC) and endothelial cell lines. HCC cell lines were infected with adenovirus-expressing shRNA for HIF-1α and IL-8 and maintained under hypoxic conditions (1% O2, 24 h). The expression levels of HIF-1α and both apoptotic and growth factors were examined by real-time quantitative PCR and western blot. We also investigated apoptosis by TUNEL assay (FACS and Immunofluorescence) and measured the concentration of cytochrome C. Inhibition of HIF-1α and IL-8 up-regulated the expression of apoptotic factors while downregulating anti-apoptotic factors simultaneously. Knockdown of HIF-1α and IL-8 increased the concentration of cytochrome C and enhanced DNA fragmentation in HCC cell lines. Moreover, culture supernatant collected from the knockdown of HIF-1α and IL-8 in HCC cell lines induced apoptosis in human umbilical vein endothelial cells under hypoxia, and the expression of variable apoptotic ligand increased from HCC cell lines, time-dependently. These data suggest that adenovirus-mediated knockdown of HIF-1α and IL-8 induced apoptosis in HCC cells and triggered apoptosis of vascular endothelial cells.

  18. The minus of a plus is a minus. Mass death of selected neuron populations in sporadic late-onset neurodegenerative disease may be due to a combination of subtly decreased capacity to repair oxidative DNA damage and increased propensity for damage-related apoptosis

    Directory of Open Access Journals (Sweden)

    Rumena Petkova

    2016-07-01

    Full Text Available Neurons in the adult central nervous system (CNS are subjected to high levels of oxidative damage that is usually promptly repaired. Transcribed genomic regions are repaired with priority over untranscribed regions. The prioritization of DNA repair in neurons results in modification of the input into the assessment of genomic integrity in order to delay or avoid damage-related apoptosis unless the damage interferes directly with the functioning of the neuron. CNS neurons may be replaced, albeit rarely. Over-stimulation of adult neural progenitor niche caused by accelerated neuronal loss may result in its premature depletion. The combination of the two pathologic mechanisms (increased rates of neuronal death and depletion of the progenitor niche may eventually result in irreversible loss of specific cell populations in the CNS and/or generalized neuronal loss. Here we propose that the risk of developing sporadic late-onset neurodegenerative disease (LONDD may be modulated by the individual capacity for detection and repair of DNA damage and the genetic propensity to repair moderate-degree damage or to assess it as irreparable and route the cell towards apoptosis. Thus, subtly deficient DNA damage repair coupled with a tendency to repair the damage rather than kill the damaged cell may be associated with increased risk of cancer, whereas deficient DNA repair coupled with a propensity to destroy damaged cells may increase the risk of LONDD. Extensive studies of individual repair capacity may be needed to test this hypothesis and, potentially, use the results in the assessment of the risk of common late-onset disease.

  19. Avenanthramides Prevent Osteoblast and Osteocyte Apoptosis and Induce Osteoclast Apoptosis in Vitro in an Nrf2-Independent Manner

    Science.gov (United States)

    Pellegrini, Gretel G.; Morales, Cynthya C.; Wallace, Taylor C.; Plotkin, Lilian I.; Bellido, Teresita

    2016-01-01

    Oats contain unique bioactive compounds known as avenanthramides (AVAs) with antioxidant properties. AVAs might enhance the endogenous antioxidant cellular response by activation of the transcription factor Nrf2. Accumulation of reactive oxygen species plays a critical role in many chronic and degenerative diseases, including osteoporosis. In this disease, there is an imbalance between bone formation by osteoblasts and bone resorption by osteoclasts, which is accompanied by increased osteoblast/osteocyte apoptosis and decreased osteoclast apoptosis. We investigated the ability of the synthethic AVAs 2c, 2f and 2p, to 1-regulate gene expression in bone cells, 2-affect the viability of osteoblasts, osteocytes and osteoclasts, and the generation of osteoclasts from their precursors, and 3-examine the potential involvement of the transcription factor Nrf2 in these actions. All doses of AVA 2c and 1 and 5 µM dose of 2p up-regulated collagen 1A expression. Lower doses of AVAs up-regulated OPG (osteoprotegerin) in OB-6 osteoblastic cells, whereas 100 μM dose of 2f and all concentrations of 2c down-regulated RANKL gene expression in MLO-Y4 osteocytic cells. AVAs did not affect apoptosis of OB-6 osteoblastic cells or MLO-Y4 osteocytic cells; however, they prevented apoptosis induced by the DNA topoisomerase inhibitor etoposide, the glucocorticoid dexamethasone, and hydrogen peroxide. AVAs prevented apoptosis of both wild type (WT) and Nrf2 Knockout (KO) osteoblasts, demonstrating that AVAs-induced survival does not require Nrf2 expression. Further, KO osteoclast precursors produced more mature osteoclasts than WT; and KO cultures exhibited less apoptotic osteoclasts than WT cultures. Although AVAs did not affect WT osteoclasts, AVA 2p reversed the low apoptosis of KO osteoclasts. These in vitro results demonstrate that AVAs regulate, in part, the function of osteoblasts and osteocytes and prevent osteoblast/osteocyte apoptosis and increase osteoclast apoptosis; further

  20. Avenanthramides Prevent Osteoblast and Osteocyte Apoptosis and Induce Osteoclast Apoptosis in Vitro in an Nrf2-Independent Manner

    Directory of Open Access Journals (Sweden)

    Gretel G. Pellegrini

    2016-07-01

    Full Text Available Oats contain unique bioactive compounds known as avenanthramides (AVAs with antioxidant properties. AVAs might enhance the endogenous antioxidant cellular response by activation of the transcription factor Nrf2. Accumulation of reactive oxygen species plays a critical role in many chronic and degenerative diseases, including osteoporosis. In this disease, there is an imbalance between bone formation by osteoblasts and bone resorption by osteoclasts, which is accompanied by increased osteoblast/osteocyte apoptosis and decreased osteoclast apoptosis. We investigated the ability of the synthethic AVAs 2c, 2f and 2p, to 1-regulate gene expression in bone cells, 2-affect the viability of osteoblasts, osteocytes and osteoclasts, and the generation of osteoclasts from their precursors, and 3-examine the potential involvement of the transcription factor Nrf2 in these actions. All doses of AVA 2c and 1 and 5 µM dose of 2p up-regulated collagen 1A expression. Lower doses of AVAs up-regulated OPG (osteoprotegerin in OB-6 osteoblastic cells, whereas 100 μM dose of 2f and all concentrations of 2c down-regulated RANKL gene expression in MLO-Y4 osteocytic cells. AVAs did not affect apoptosis of OB-6 osteoblastic cells or MLO-Y4 osteocytic cells; however, they prevented apoptosis induced by the DNA topoisomerase inhibitor etoposide, the glucocorticoid dexamethasone, and hydrogen peroxide. AVAs prevented apoptosis of both wild type (WT and Nrf2 Knockout (KO osteoblasts, demonstrating that AVAs-induced survival does not require Nrf2 expression. Further, KO osteoclast precursors produced more mature osteoclasts than WT; and KO cultures exhibited less apoptotic osteoclasts than WT cultures. Although AVAs did not affect WT osteoclasts, AVA 2p reversed the low apoptosis of KO osteoclasts. These in vitro results demonstrate that AVAs regulate, in part, the function of osteoblasts and osteocytes and prevent osteoblast/osteocyte apoptosis and increase osteoclast

  1. Alcohol and Apoptosis: Friends or Foes?

    Science.gov (United States)

    Rodriguez, Ana; Chawla, Karan; Umoh, Nsini A; Cousins, Valerie M; Ketegou, Assama; Reddy, Madhumati G; AlRubaiee, Mustafa; Haddad, Georges E; Burke, Mark W

    2015-11-19

    Alcohol abuse causes 79,000 deaths stemming from severe organ damage in the United States every year. Clinical manifestations of long-term alcohol abuse on the cardiac muscle include defective contractility with the development of dilated cardiomyopathy and low-output heart failure; which has poor prognosis with less than 25% survival for more than three years. In contrast, low alcohol consumption has been associated with reduced risk of cardiovascular disease, however the mechanism of this phenomenon remains elusive. The aim of this study was to determine the significance of apoptosis as a mediating factor in cardiac function following chronic high alcohol versus low alcohol exposure. Adult rats were provided 5 mM (low alcohol), 100 mM (high alcohol) or pair-fed non-alcohol controls for 4-5 months. The hearts were dissected, sectioned and stained with cresyl violet or immunohistochemically for caspase-3, a putative marker for apoptosis. Cardiomyocytes were isolated to determine the effects of alcohol exposure on cell contraction and relaxation. High alcohol animals displayed a marked thinning of the left ventricular wall combined with elevated caspase-3 activity and decreased contractility. In contrast, low alcohol was associated with increased contractility and decreased apoptosis suggesting an overall protective mechanism induced by low levels of alcohol exposure.

  2. [Ca2+]i in exterior of cells effected on apoptosis of HL-60 cells induced by irradiation

    International Nuclear Information System (INIS)

    He Ziyi; Meng Qingyong

    2005-01-01

    Objective: To investigate of the different [Ca 2+ ]i in exterior of cells promotion function on apoptosis of HL-60 cells induced by irradiation. Methods: To put ration dose 32 P and different [Ca 2+ ]i into culture of HL-60 and measure the apoptosis rate with FCM after 24 and 48 hours. Result: Apoptosis rate increased with the increase of [Ca 2+ ]i which shows an obvious function to promote apoptosis, r 24 =0.9001 (P=0.0145); r48=0.9343 (P=0.0063). Conclusion: [Ca 2+ ]i in exterior of cells has a obvious function in promoteing apoptosis induced by irradiation. (authors)

  3. Altered Chondrocyte Apoptosis Status in Developmental Hip Dysplasia in Rabbits

    Directory of Open Access Journals (Sweden)

    Yi-Shan Wei

    2016-12-01

    Full Text Available Background: Developmental dysplasia of the hip (DDH is an important factor leading to early adult osteoarthritis. Chondrocyte apoptosis has been proven to be an important factor causing osteoarthritis. Aims: The current study aims to explore whether a rabbit model of developmental dysplasia of the hip through cast immobilization in the legs results in chondrocyte apoptosis. Study Design: Animal experimentation. Methods: Thirty-two New Zealand white rabbits were divided in three groups with cast plaster-induced dislocation at 2, 4 and 6 weeks. The contralateral hip joint was utilized as a control group. Ten rabbits in each group were sacrificed, and hip specimens were obtained. Bcl-2/Bax, cleaved caspase-3 and cleaved caspase-8 expression were examined by western blot analysis. Chondrocyte apoptosis was analyzed through transmission electron microscopy (TEM and TUNEL analysis. All experiments were repeated at least three times. Results: In the experimental group, Bcl-2/Bax, cleaved caspase-3 and cleaved caspase-8 expression were significantly altered. The Bcl-2/Bax ratio decreased with time (all p<0.01, whereas levels of cleaved caspase-3 (p<0.01 and p<0.05 and cleaved caspase-8 (all p<0.05 gradually increased. Chondrocyte apoptosis was observed through transmission electron microscopy (TEM and TUNEL analysis (p<0.05 at 4 weeks and p<0.01 at 6 weeks. Conclusion: Prolonged immobilization of rabbit hip caused chondrocyte apoptosis. Reduction of the hip joint may protect chondrocytes from apoptosis, thus preventing secondary osteoarthritis.

  4. Ketamine-induced apoptosis in the mouse cerebral cortex follows similar characteristic of physiological apoptosis and can be regulated by neuronal activity.

    Science.gov (United States)

    Wang, Qi; Shen, Feng-Yan; Zou, Rong; Zheng, Jing-Jing; Yu, Xiang; Wang, Ying-Wei

    2017-06-17

    The effects of general anesthetics on inducing neuronal apoptosis during early brain development are well-documented. However, since physiological apoptosis also occurs during this developmental window, it is important to determine whether anesthesia-induced apoptosis targets the same cell population as physiological apoptosis or different cell types altogether. To provide an adequate plane of surgery, ketamine was co-administered with dexmedetomidine. The apoptotic neurons in the mouse primary somatosensory cortex (S1) were quantitated by immunohistochemistry. To explore the effect of neural activity on ketamine-induced apoptosis, the approaches of Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) and an environmental enrichment (EE) were performed. Ketamine-induced apoptosis in S1 is most prominent at postnatal days 5 and 7 (P5 - P7), and becomes insignificant by P12. Physiological and ketamine-induced apoptosis follow similar developmental patterns, mostly comprised of layer V pyramidal neurons at P5 and shifting to mostly layer II to IV GABAergic neurons by P9. Changes in neuronal activity induced by the DREADD system bidirectionally regulated the pattern of ketamine-induced apoptosis, with reduced activity inducing increased apoptosis and shifting the lamination pattern to a more immature form. Importantly, rearing mice in an EE significantly reduced the magnitude of ketamine-induced apoptosis and shifted its developmental pattern to a more mature form. Together, these results demonstrate that lamination pattern and cell-type dependent vulnerability to ketamine-induced apoptosis follow the physiological apoptosis pattern and are age- and activity-dependent. Naturally elevating neuronal activity is a possible method for reducing the adverse effects of general anesthesia.

  5. Effect of Sulfated Polysaccharides from Brown Algae on Apoptosis of Human Peripheral Blood Lymphocytes.

    Science.gov (United States)

    Gazha, A K; Zaporozhets, T S; Kuznetsova, T A; Zvyaguintseva, T N; Besednova, N N

    2015-09-01

    We studied the influence of fucoidans from brown algae Fucus evanescens, Laminaria cichorioides and Laminaria japonica on apoptosis of human peripheral blood lymphocytes. It was demonstrated that fucoidans induced lymphocyte apoptosis, increased the proportion of cells with low mitochondrial transmembrane potential, and inhibit expression of Bcl-xL gene in blood lymphocytes. These findings suggest that lymphocyte apoptosis induced by the analyzed sulfated polysaccharides is mediated by mitochondrial pathway.

  6. Advanced glycation end products (AGEs) and their receptor (RAGE) induce apoptosis of periodontal ligament fibroblasts

    OpenAIRE

    Li,D.X.; Deng,T.Z.; Lv,J.; Ke,J.

    2014-01-01

    Diabetics have an increased prevalence of periodontitis, and diabetes is one of the causative factors of severe periodontitis. Apoptosis is thought to be involved in this pathogenic relationship. The aim of this study was to investigate apoptosis in human periodontal ligament (PDL) fibroblasts induced by advanced glycation end products (AGEs) and their receptor (RAGE). We examined the roles of apoptosis, AGEs, and RAGE during periodontitis in diabetes mellitus using cultured PDL fibroblasts t...

  7. Csk regulates angiotensin II-induced podocyte apoptosis.

    Science.gov (United States)

    Zhang, Lu; Ren, Zhilong; Yang, Qian; Ding, Guohua

    2016-07-01

    Increasing data have shown that angiotensin II (Ang II) perpetuates podocyte injury and promotes progression to end-stage kidney disease. The mechanism underlying Ang II-induced podocyte apoptosis has not been established. C-terminal Src kinase (Csk) is a cytoplasmic kinase that interacts with scaffolding proteins involved in cell growth, adhesion, and polarization, and the role of Csk in regulating cellular apoptosis has gradually attracted attention. This study evaluates the role of Csk in Ang II-induced podocyte apoptosis. In vivo, Wistar rats were randomly subjected to a normal saline or Ang II infusion. In vitro, we exposed differentiated mouse podocytes to Ang II. Ang II increased Csk expression and induced podocyte apoptosis, stimulated Csk translocation and binding to Caveolin-1, and stimulated decreased Fyn pY416, increased Fyn pY529, and nephrin dephosphorylation. Csk knockdown prevented Ang II-induced podocyte apoptosis, reduced Fyn kinase inactivation, and increased the interaction between nephrin and the activated form of Fyn, accompanied by a reduced interaction between Csk and Caveolin-1. These findings indicate that Ang II induces podocyte injury via a Csk-dependent pathway.

  8. Apigenin induces apoptosis by targeting inhibitor of apoptosis proteins and Ku70–Bax interaction in prostate cancer

    Science.gov (United States)

    Shukla, Sanjeev; Fu, Pingfu; Gupta, Sanjay

    2014-01-01

    Dysfunction of the apoptotic pathway in prostate cancer cells confers apoptosis resistance towards various therapies. A novel strategy to overcome resistance is to directly target the apoptotic pathway in cancer cells. Apigenin, an anticancer agent, selectively toxic to cancer cells induces cell cycle arrest and apoptosis through mechanisms which are not fully explored. In the present study we provide novel insight into the mechanisms of apoptosis induction by apigenin. Treatment of androgen-refractory human prostate cancer PC-3 and DU145 cells with apigenin resulted in dose-dependent suppression of XIAP, c-IAP1, c-IAP2 and survivin protein levels. Apigenin treatment resulted in significant decrease in cell viability and apoptosis induction with the increase of cytochrome C in time-dependent manner. These effects of apigenin were accompanied by decrease in Bcl-xL and Bcl-2 and increase in the active form of Bax protein. The apigenin-mediated increase in Bax was due to dissociation of Bax from Ku70 which is essential for apoptotic activity of Bax. Apigenin treatment resulted in the inhibition of class I histone deacetylases and HDAC1 protein expression, thereby increasing the acetylation of Ku70 and the dissociation of Bax resulting in apoptosis of cancer cells. Furthermore, apigenin significantly reduced HDAC1 occupancy at the XIAP promoter, suggesting that histone deacetylation might be critical for XIAP downregulation. These results suggest that apigenin targets inhibitor of apoptosis proteins and Ku70–Bax interaction in the induction of apoptosis in prostate cancer cells and in athymic nude mouse xenograft model endorsing its in vivo efficacy. PMID:24563225

  9. Autophagy Regulates Colistin-Induced Apoptosis in PC-12 Cells

    Science.gov (United States)

    Zhang, Ling; Zhao, Yonghao; Ding, Wenjian; Jiang, Guozheng; Lu, Ziyin; Li, Li; Wang, Jinli

    2015-01-01

    Colistin is a cyclic cationic polypeptide antibiotic with activity against multidrug-resistant Gram-negative bacteria. Our recent study demonstrated that colistin induces apoptosis in primary chick cortex neurons and PC-12 cells. Although apoptosis and autophagy have different impacts on cell fate, there is a complex interaction between them. Autophagy plays an important role as a homeostasis regulator by removing excessive or unnecessary proteins and damaged organelles. The aim of the present study was to investigate the modulation of autophagy and apoptosis regulation in PC-12 cells in response to colistin treatment. PC-12 cells were exposed to colistin (125 to 250 μg/ml), and autophagy was detected by visualization of monodansylcadaverine (MDC)-labeled vacuoles, LC3 (microtubule-associated protein 1 light chain 3) immunofluorescence microscopic examination, and Western blotting. Apoptosis was measured by flow cytometry, Hoechst 33258 staining, and Western blotting. Autophagosomes were observed after treatment with colistin for 12 h, and the levels of LC3-II gene expression were determined; observation and protein levels both indicated that colistin induced a high level of autophagy. Colistin treatment also led to apoptosis in PC-12 cells, and the level of caspase-3 expression increased over the 24-h period. Pretreatment of cells with 3-methyladenine (3-MA) increased colistin toxicity in PC-12 cells remarkably. However, rapamycin treatment significantly increased the expression levels of LC3-II and beclin 1 and decreased the rate of apoptosis of PC-12 cells. Our results demonstrate that colistin induced autophagy and apoptosis in PC-12 cells and that the latter was affected by the regulation of autophagy. It is very likely that autophagy plays a protective role in the reduction of colistin-induced cytotoxicity in neurons. PMID:25645826

  10. Andrographolide sensitizes prostate cancer cells to TRAIL-induced apoptosis

    Directory of Open Access Journals (Sweden)

    Ruo-Jing Wei

    2018-01-01

    Full Text Available Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL is a promising agent for anticancer therapy. The identification of small molecules that can establish the sensitivity of prostate cancer (PCa cells to TRAIL-induced apoptosis is crucial for the targeted treatment of PCa. PC3, DU145, JAC-1, TsuPr1, and LNCaP cells were treated with Andrographolide (Andro and TRAIL, and the apoptosis was measured using the Annexin V/PI double staining method. Real time-polymerase chain reaction (PCR and Western blot analysis were performed to measure the expression levels of target molecules. RNA interference technique was used to down-regulate the expression of the target protein. We established a nude mouse xenograft model of PCa, which was used to measure the caspase-3 activity in the tumor cells using flow cytometry. In this research study, our results demonstrated that Andro preferentially increased the sensitivity of PCa cells to TRAIL-induced apoptosis at subtoxic concentrations, and the regulation mechanism was related to the up-regulation of DR4. In addition, it also increased the p53 expression and led to the generation of reactive oxygen species (ROS in the cells. Further research revealed that the DR4 inhibition, p53 expression, and ROS generation can significantly reduce the apoptosis induced by the combination of TRAIL and Andro in PCa cells. In conclusion, Andro increases the sensitivity of PCa cells to TRAIL-induced apoptosis through the generation of ROS and up-regulation of p53 and then promotes PCa cell apoptosis associated with the activation of DR4.

  11. Andrographolide sensitizes prostate cancer cells to TRAIL-induced apoptosis.

    Science.gov (United States)

    Wei, Ruo-Jing; Zhang, Xin-Shi; He, Da-Lin

    2018-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising agent for anticancer therapy. The identification of small molecules that can establish the sensitivity of prostate cancer (PCa) cells to TRAIL-induced apoptosis is crucial for the targeted treatment of PCa. PC3, DU145, JAC-1, TsuPr1, and LNCaP cells were treated with Andrographolide (Andro) and TRAIL, and the apoptosis was measured using the Annexin V/PI double staining method. Real time-polymerase chain reaction (PCR) and Western blot analysis were performed to measure the expression levels of target molecules. RNA interference technique was used to down-regulate the expression of the target protein. We established a nude mouse xenograft model of PCa, which was used to measure the caspase-3 activity in the tumor cells using flow cytometry. In this research study, our results demonstrated that Andro preferentially increased the sensitivity of PCa cells to TRAIL-induced apoptosis at subtoxic concentrations, and the regulation mechanism was related to the up-regulation of DR4. In addition, it also increased the p53 expression and led to the generation of reactive oxygen species (ROS) in the cells. Further research revealed that the DR4 inhibition, p53 expression, and ROS generation can significantly reduce the apoptosis induced by the combination of TRAIL and Andro in PCa cells. In conclusion, Andro increases the sensitivity of PCa cells to TRAIL-induced apoptosis through the generation of ROS and up-regulation of p53 and then promotes PCa cell apoptosis associated with the activation of DR4.

  12. Temporal characteristics of gustatory responses in rat parabrachial neurons vary by stimulus and chemosensitive neuron type.

    Science.gov (United States)

    Geran, Laura; Travers, Susan

    2013-01-01

    It has been demonstrated that temporal features of spike trains can increase the amount of information available for gustatory processing. However, the nature of these temporal characteristics and their relationship to different taste qualities and neuron types are not well-defined. The present study analyzed the time course of taste responses from parabrachial (PBN) neurons elicited by multiple applications of "sweet" (sucrose), "salty" (NaCl), "sour" (citric acid), and "bitter" (quinine and cycloheximide) stimuli in an acute preparation. Time course varied significantly by taste stimulus and best-stimulus classification. Across neurons, the ensemble code for the three electrolytes was similar initially but quinine diverged from NaCl and acid during the second 500 ms of stimulation and all four qualities became distinct just after 1s. This temporal evolution was reflected in significantly broader tuning during the initial response. Metric space analyses of quality discrimination by individual neurons showed that increases in information (H) afforded by temporal factors was usually explained by differences in rate envelope, which had a greater impact during the initial 2s (22.5% increase in H) compared to the later response (9.5%). Moreover, timing had a differential impact according to cell type, with between-quality discrimination in neurons activated maximally by NaCl or citric acid most affected. Timing was also found to dramatically improve within-quality discrimination (80% increase in H) in neurons that responded optimally to bitter stimuli (B-best). Spikes from B-best neurons were also more likely to occur in bursts. These findings suggest that among PBN taste neurons, time-dependent increases in mutual information can arise from stimulus- and neuron-specific differences in response envelope during the initial dynamic period. A stable rate code predominates in later epochs.

  13. Inhibition of apoptosis by BCL2 prevents leukemic transformation of a murine myelodysplastic syndrome

    Science.gov (United States)

    Saw, Jesslyn; Jowett, Jeremy B. M.; Aplan, Peter D.; Strasser, Andreas; Jane, Stephen M.; Curtis, David J.

    2012-01-01

    Programmed cell death or apoptosis is a prominent feature of low-risk myelodysplastic syndromes (MDS), although the underlying mechanism remains controversial. High-risk MDS have less apoptosis associated with increased expression of the prosurvival BCL2-related proteins. To address the mechanism and pathogenic role of apoptosis and BCL2 expression in MDS, we used a mouse model resembling human MDS, in which the fusion protein NUP98-HOXD13 (NHD13) of the chromosomal translocation t(2;11)(q31;p15) is expressed in hematopoietic cells. Hematopoietic stem and progenitor cells from 3-month-old mice had increased rates of apoptosis associated with increased cell cycling and DNA damage. Gene expression profiling of these MDS progenitors revealed a specific reduction in Bcl2. Restoration of Bcl2 expression by a BCL2 transgene blocked apoptosis of the MDS progenitors, which corrected the macrocytic anemia. Blocking apoptosis also restored cell-cycle quiescence and reduced DNA damage in the MDS progenitors. We expected that preventing apoptosis would accelerate malignant transformation to acute myeloid leukemia (AML). However, contrary to expectations, preventing apoptosis of premalignant cells abrogated transformation to AML. In contrast to the current dogma that overcoming apoptosis is an important step toward cancer, this work demonstrates that gaining a survival advantage of premalignant cells may delay or prevent leukemic progression. PMID:22855610

  14. Restraining reactive oxygen species in Listeria monocytogenes promotes the apoptosis of glial cells.

    Science.gov (United States)

    Li, Sen; Li, Yixuan; Chen, Guowei; Zhang, Jingchen; Xu, Fei; Wu, Man

    2017-07-01

    Listeria monocytogenes is a facultative anaerobic foodborne pathogen that can traverse the blood-brain barrier and cause brain infection. L. monocytogenes infection induces host cell apoptosis in several cell types. In this study, we investigated the apoptosis of human glioma cell line U251 invaded by L. monocytogenes and evaluated the function of bacterial reactive oxygen species (ROS) during infection. Bacterial ROS level was reduced by carrying out treatment with N-acetyl cysteine (NAC) and diphenyleneiodonium chloride (DPI). After infection, the apoptosis of U251 cells was examined by flow cytometry assay and propidium iodide staining. DPI and NAC efficiently decreased ROS level in L. monocytogenes without affecting bacterial growth. Moreover, the apoptosis of glial cells was enhanced upon invasion of DPI- and NAC-pretreated L. monocytogenes. Results indicate that the apoptosis of glial cells can be induced by L. monocytogenes, and that the inhibition of bacterial ROS increases the apoptosis of host cells.

  15. Neuroprotective effects of ganoderma lucidum polysaccharides against oxidative stress-induced neuronal apoptosis.

    Science.gov (United States)

    Sun, Xin-Zhi; Liao, Ying; Li, Wei; Guo, Li-Mei

    2017-06-01

    Ganoderma lucidum polysaccharides have protective effects against apoptosis in neurons exposed to ischemia/reperfusion injury, but the mechanisms are unclear. The goal of this study was to investigate the underlying mechanisms of the effects of ganoderma lucidum polysaccharides against oxidative stress-induced neuronal apoptosis. Hydrogen peroxide (H 2 O 2 ) was used to induce apoptosis in cultured cerebellar granule cells. In these cells, ganoderma lucidum polysaccharides remarkably suppressed H 2 O 2 -induced apoptosis, decreased expression of caspase-3, Bax and Bim and increased that of Bcl-2. These findings suggested that ganoderma lucidum polysaccharides regulate expression of apoptosis-associated proteins, inhibit oxidative stress-induced neuronal apoptosis and, therefore, have significant neuroprotective effects.

  16. Neuroprotective effects of ganoderma lucidum polysaccharides against oxidative stress-induced neuronal apoptosis

    Science.gov (United States)

    Sun, Xin-zhi; Liao, Ying; Li, Wei; Guo, Li-mei

    2017-01-01

    Ganoderma lucidum polysaccharides have protective effects against apoptosis in neurons exposed to ischemia/reperfusion injury, but the mechanisms are unclear. The goal of this study was to investigate the underlying mechanisms of the effects of ganoderma lucidum polysaccharides against oxidative stress-induced neuronal apoptosis. Hydrogen peroxide (H2O2) was used to induce apoptosis in cultured cerebellar granule cells. In these cells, ganoderma lucidum polysaccharides remarkably suppressed H2O2-induced apoptosis, decreased expression of caspase-3, Bax and Bim and increased that of Bcl-2. These findings suggested that ganoderma lucidum polysaccharides regulate expression of apoptosis-associated proteins, inhibit oxidative stress-induced neuronal apoptosis and, therefore, have significant neuroprotective effects. PMID:28761429

  17. Podocyte hypertrophy precedes apoptosis under experimental diabetic conditions.

    Science.gov (United States)

    Lee, Sun Ha; Moon, Sung Jin; Paeng, Jisun; Kang, Hye-Young; Nam, Bo Young; Kim, Seonghun; Kim, Chan Ho; Lee, Mi Jung; Oh, Hyung Jung; Park, Jung Tak; Han, Seung Hyeok; Yoo, Tae-Hyun; Kang, Shin-Wook

    2015-08-01

    Podocyte hypertrophy and apoptosis are two hallmarks of diabetic glomeruli, but the sequence in which these processes occur remains a matter of debate. Here we investigated the effects of inhibiting hypertrophy on apoptosis, and vice versa, in both podocytes and glomeruli, under diabetic conditions. Hypertrophy and apoptosis were inhibited using an epidermal growth factor receptor inhibitor (PKI 166) and a pan-caspase inhibitor (zAsp-DCB), respectively. We observed significant increases in the protein expression of p27, p21, phospho-eukaryotic elongation factor 4E-binding protein 1, and phospho-p70 S6 ribosomal protein kinase, in both cultured podocytes exposed to high-glucose (HG) medium, and streptozotocin-induced diabetes mellitus (DM) rat glomeruli. These increases were significantly inhibited by PKI 166, but not by zAsp-DCB. In addition, the amount of protein per cell, the relative cell size, and the glomerular volume were all significantly increased under diabetic conditions, and these changes were also blocked by treatment with PKI 166, but not zAsp-DCB. Increased protein expression of cleaved caspase-3 and cleaved poly (ADP-ribose) polymerase, together with increased Bax/Bcl-2 ratios, were also observed in HG-stimulated podocytes and DM glomeruli. Treatment with either zAsp-DCB or PKI 166 resulted in a significant attenuation of these effects. Both PKI 166 and zAsp-DCB also inhibited the increase in number of apoptotic cells, as assessed by Hoechst 33342 staining and TUNEL assay. Under diabetic conditions, inhibition of podocyte hypertrophy results in attenuated apoptosis, whe