Silibinin Inhibits Platelet-Derived Growth Factor-Driven Cell Proliferation via Downregulation of N-Glycosylation in Human Tenon's Fibroblasts in a Proteasome-Dependent Manner

PubMed Central

Chen, Yi-Hao; Chen, Ching-Long; Lu, Da-Wen; Liang, Chang-Min; Tai, Ming-Cheng; Chen, Jiann-Torng

2016-01-01

The objective of this study was to evaluate the effects of silibinin on cell proliferation in platelet-derived growth factor (PDGF)-treated human Tenon's fibroblasts (HTFs). The effect of silibinin on cell proliferation in PDGF-treated HTFs was determined by examining the expression of proliferating cell nuclear antigen (PCNA) and performing WST-1 assays. Cell cycle progression was evaluated using flow cytometry. The related cyclins and cyclin-dependent kinases (CDKs) were also analyzed using western blot. A modified rat trabeculectomy model was established to evaluate the effect of silibinin on cell proliferation in vivo. Western blot analysis was carried out to determine the effect of silibinin on the expression of PDGF receptor and on the downstream signaling pathways regulated by PDGF receptor. PDGF elevated the expression of PCNA in HTFs, and this elevation was inhibited by silibinin. The inhibitory effect of silibinin on cell proliferation was also confirmed via WST-1 assay. PDGF-stimulated cell cycle in HTFs was delayed by silibinin, and the related cyclin D1 and CDK4 were also suppressed by silibinin. In the rat model of trabeculectomy, silibinin reduced the expression of PCNA at the site of blebs in vivo. The effects of silibinin on PDGF-stimulated HTFs were mediated via the downregulation of PDGF receptor-regulated signaling pathways, such as ERKs and STATs, which may be partially caused by the downregulation of N-glycosylation of PDGF receptor beta (PDGFRβ). The effect of silibinin on modulation of N-glycosylation of PDGFRβ was mediated in a proteasome-dependent manner. Silibinin inhibited cell proliferation and delayed cell cycle progression in PDGF-treated HTFs in vitro. PDGF also modulated the process of N-glycosylation of the PDGFRβ in a proteasome-dependent manner. Our findings suggest that silibinin has potential therapeutic applications in glaucoma filtering surgery. PMID:28030611

Morphogenesis checkpoint kinase Swe1 is the executor of lipolysis-dependent cell-cycle progression.

PubMed

Chauhan, Neha; Visram, Myriam; Cristobal-Sarramian, Alvaro; Sarkleti, Florian; Kohlwein, Sepp D

2015-03-10

Cell growth and division requires the precise duplication of cellular DNA content but also of membranes and organelles. Knowledge about the cell-cycle-dependent regulation of membrane and storage lipid homeostasis is only rudimentary. Previous work from our laboratory has shown that the breakdown of triacylglycerols (TGs) is regulated in a cell-cycle-dependent manner, by activation of the Tgl4 lipase by the major cyclin-dependent kinase Cdc28. The lipases Tgl3 and Tgl4 are required for efficient cell-cycle progression during the G1/S (Gap1/replication phase) transition, at the onset of bud formation, and their absence leads to a cell-cycle delay. We now show that defective lipolysis activates the Swe1 morphogenesis checkpoint kinase that halts cell-cycle progression by phosphorylation of Cdc28 at tyrosine residue 19. Saturated long-chain fatty acids and phytosphingosine supplementation rescue the cell-cycle delay in the Tgl3/Tgl4 lipase-deficient strain, suggesting that Swe1 activity responds to imbalanced sphingolipid metabolism, in the absence of TG degradation. We propose a model by which TG-derived sphingolipids are required to activate the protein phosphatase 2A (PP2A(Cdc55)) to attenuate Swe1 phosphorylation and its inhibitory effect on Cdc28 at the G1/S transition of the cell cycle.

Cyclin-dependent kinase inhibitor p20 controls circadian cell-cycle timing

PubMed Central

Laranjeiro, Ricardo; Tamai, T. Katherine; Peyric, Elodie; Krusche, Peter; Ott, Sascha; Whitmore, David

2013-01-01

Specific stages of the cell cycle are often restricted to particular times of day because of regulation by the circadian clock. In zebrafish, both mitosis (M phase) and DNA synthesis (S phase) are clock-controlled in cell lines and during embryo development. Despite the ubiquitousness of this phenomenon, relatively little is known about the underlying mechanism linking the clock to the cell cycle. In this study, we describe an evolutionarily conserved cell-cycle regulator, cyclin-dependent kinase inhibitor 1d (20 kDa protein, p20), which along with p21, is a strongly rhythmic gene and directly clock-controlled. Both p20 and p21 regulate the G1/S transition of the cell cycle. However, their expression patterns differ, with p20 predominant in developing brain and peak expression occurring 6 h earlier than p21. p20 expression is also p53-independent in contrast to p21 regulation. Such differences provide a unique mechanism whereby S phase is set to different times of day in a tissue-specific manner, depending on the balance of these two inhibitors. PMID:23569261

Cyclin-dependent kinase inhibitor p20 controls circadian cell-cycle timing.

PubMed

Laranjeiro, Ricardo; Tamai, T Katherine; Peyric, Elodie; Krusche, Peter; Ott, Sascha; Whitmore, David

2013-04-23

Specific stages of the cell cycle are often restricted to particular times of day because of regulation by the circadian clock. In zebrafish, both mitosis (M phase) and DNA synthesis (S phase) are clock-controlled in cell lines and during embryo development. Despite the ubiquitousness of this phenomenon, relatively little is known about the underlying mechanism linking the clock to the cell cycle. In this study, we describe an evolutionarily conserved cell-cycle regulator, cyclin-dependent kinase inhibitor 1d (20 kDa protein, p20), which along with p21, is a strongly rhythmic gene and directly clock-controlled. Both p20 and p21 regulate the G1/S transition of the cell cycle. However, their expression patterns differ, with p20 predominant in developing brain and peak expression occurring 6 h earlier than p21. p20 expression is also p53-independent in contrast to p21 regulation. Such differences provide a unique mechanism whereby S phase is set to different times of day in a tissue-specific manner, depending on the balance of these two inhibitors.

High fat diet triggers cell cycle arrest and excessive apoptosis of granulosa cells during the follicular development

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Yanqing; Zhang, Zhenghong; Liao, Xinghui

The regulatory mechanism of granulosa cells (GCs) proliferation during the follicular development is complicated and multifactorial, which is essential for the oocyte growth and normal ovarian functions. To investigate the role of high fat diet (HFD) on the proliferation of GCs, 4-week old female mice were fed with HFD or normal control diet (NC) for 15 weeks or 20 weeks and then detected the expression level of some regulatory molecules of cell cycle and apoptosis. The abnormal ovarian morphology was observed at 20 weeks. Further mechanistic studies indicated that HFD induced-obesity caused elevated apoptotic levels in GCs of the ovariesmore » in a time-dependent manner. Moreover, cell cycle progress was also impacted after HFD fed. The cell cycle inhibitors, p27{sup Kip1} and p21{sup Cip1}, were significantly induced in the ovaries from the mice in HFD group when compared with that in the ovaries from the mice in NC group. Subsequently, the expression levels of Cyclin D1, D3 and CDK4 were also significantly influenced in the ovaries from the mice fed with HFD in a time-dependent manner. The present results suggested that HFD induced-obesity may trigger cell cycle arrest and excessive apoptosis of GCs, causing the abnormal follicular development and ovarian function failure. - Highlights: • HFD induced-obesity leads to abnormal ovarian morphology. • HFD induced-obesity triggers excessive apoptosis in the ovary. • HFD induced-obesity up-regulates cell cycle inhibitors p21{sup Cip1} and p27{sup Kip1} in the ovary. • HFD induced-obesity causes cell cycle arrest in the ovary.« less

lH-Pyrazolo[3,4-b]quinolin-3-amine derivatives inhibit growth of colon cancer cells via apoptosis and sub G1 cell cycle arrest.

PubMed

Karthikeyan, Chandrabose; Amawi, Haneen; Viana, Arabela Guedes; Sanglard, Leticia; Hussein, Noor; Saddler, Maria; Ashby, Charles R; Moorthy, N S Hari Narayana; Trivedi, Piyush; Tiwari, Amit K

2018-07-15

A series of lH-pyrazolo[3,4-b]quinolin-3-amine derivatives were synthesized and evaluated for anticancer efficacy in a panel of ten cancer cell lines, including breast (MDAMB-231 and MCF-7), colon (HCT-116, HCT-15, HT-29 and LOVO), prostate (DU-145 and PC3), brain (LN-229), ovarian (A2780), and human embryonic kidney (HEK293) cells, a non-cancerous cell line. Among the eight derivatives screened, compound QTZ05 had the most potent and selective antitumor efficacy in the four colon cancer cell lines, with IC 50 values ranging from 2.3 to 10.2 µM. Furthermore, QTZ05 inhibited colony formation in HCT-116 cells in a concentration-dependent manner. Cell cycle analysis data indicated that QTZ05 caused an arrest in the sub G1 cell cycle in HCT-116 cells. QTZ05 induced apoptosis in HCT-116 cells in a concentration-dependent manner that was characterized by chromatin condensation and increase in the fluorescence of fluorochrome-conjugated Annexin V. The findings from our study suggest that QTZ05 may be a valuable prototype for the development of chemotherapeutics targeting apoptotic pathways in colorectal cancer cells. Copyright © 2018 Elsevier Ltd. All rights reserved.

Characterization of dependencies between growth and division in budding yeast

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mayhew, Michael B.; Iversen, Edwin S.; Hartemink, Alexander J.

Cell growth and division are processes vital to the proliferation and development of life. Coordination between these two processes has been recognized for decades in a variety of organisms. In the budding yeast Saccharomyces cerevisiae, this coordination or ‘size control’ appears as an inverse correlation between cell size and the rate of cell-cycle progression, routinely observed in G1 prior to cell division commitment. Beyond this point, cells are presumed to complete S/G 2/M at similar rates and in a size-independent manner. As such, studies of dependence between growth and division have focused on G1. Moreover, in unicellular organisms, coordination betweenmore » growth and division has commonly been analyzed within the cycle of a single cell without accounting for correlations in growth and division characteristics between cycles of related cells. In a comprehensive analysis of three published time-lapse microscopy datasets, we analyze both intra- and inter-cycle dependencies between growth and division, revisiting assumptions about the coordination between these two processes. Interestingly, we find evidence (1) that S/G 2/M durations are systematically longer in daughters than in mothers, (2) of dependencies between S/G2/M and size at budding that echo the classical G1 dependencies, and, (3) in contrast with recent bacterial studies, of negative dependencies between size at birth and size accumulated during the cell cycle. In addition, we develop a novel hierarchical model to uncover inter-cycle dependencies, and we find evidence for such dependencies in cells growing in sugar-poor environments. Our analysis highlights the need for experimentalists and modelers to account for new sources of cell-to-cell variation in growth and division, and our model provides a formal statistical framework for the continued study of dependencies between biological processes.« less

Characterization of dependencies between growth and division in budding yeast

DOE PAGES

Mayhew, Michael B.; Iversen, Edwin S.; Hartemink, Alexander J.

2017-02-01

Cell growth and division are processes vital to the proliferation and development of life. Coordination between these two processes has been recognized for decades in a variety of organisms. In the budding yeast Saccharomyces cerevisiae, this coordination or ‘size control’ appears as an inverse correlation between cell size and the rate of cell-cycle progression, routinely observed in G1 prior to cell division commitment. Beyond this point, cells are presumed to complete S/G 2/M at similar rates and in a size-independent manner. As such, studies of dependence between growth and division have focused on G1. Moreover, in unicellular organisms, coordination betweenmore » growth and division has commonly been analyzed within the cycle of a single cell without accounting for correlations in growth and division characteristics between cycles of related cells. In a comprehensive analysis of three published time-lapse microscopy datasets, we analyze both intra- and inter-cycle dependencies between growth and division, revisiting assumptions about the coordination between these two processes. Interestingly, we find evidence (1) that S/G 2/M durations are systematically longer in daughters than in mothers, (2) of dependencies between S/G2/M and size at budding that echo the classical G1 dependencies, and, (3) in contrast with recent bacterial studies, of negative dependencies between size at birth and size accumulated during the cell cycle. In addition, we develop a novel hierarchical model to uncover inter-cycle dependencies, and we find evidence for such dependencies in cells growing in sugar-poor environments. Our analysis highlights the need for experimentalists and modelers to account for new sources of cell-to-cell variation in growth and division, and our model provides a formal statistical framework for the continued study of dependencies between biological processes.« less

Characterization of dependencies between growth and division in budding yeast

PubMed Central

Iversen, Edwin S.; Hartemink, Alexander J.

2017-01-01

Cell growth and division are processes vital to the proliferation and development of life. Coordination between these two processes has been recognized for decades in a variety of organisms. In the budding yeast Saccharomyces cerevisiae, this coordination or ‘size control’ appears as an inverse correlation between cell size and the rate of cell-cycle progression, routinely observed in G1 prior to cell division commitment. Beyond this point, cells are presumed to complete S/G2/M at similar rates and in a size-independent manner. As such, studies of dependence between growth and division have focused on G1. Moreover, in unicellular organisms, coordination between growth and division has commonly been analysed within the cycle of a single cell without accounting for correlations in growth and division characteristics between cycles of related cells. In a comprehensive analysis of three published time-lapse microscopy datasets, we analyse both intra- and inter-cycle dependencies between growth and division, revisiting assumptions about the coordination between these two processes. Interestingly, we find evidence (i) that S/G2/M durations are systematically longer in daughters than in mothers, (ii) of dependencies between S/G2/M and size at budding that echo the classical G1 dependencies, and (iii) in contrast with recent bacterial studies, of negative dependencies between size at birth and size accumulated during the cell cycle. In addition, we develop a novel hierarchical model to uncover inter-cycle dependencies, and we find evidence for such dependencies in cells growing in sugar-poor environments. Our analysis highlights the need for experimentalists and modellers to account for new sources of cell-to-cell variation in growth and division, and our model provides a formal statistical framework for the continued study of dependencies between biological processes. PMID:28228543

Characterization of dependencies between growth and division in budding yeast.

PubMed

Mayhew, Michael B; Iversen, Edwin S; Hartemink, Alexander J

2017-02-01

Cell growth and division are processes vital to the proliferation and development of life. Coordination between these two processes has been recognized for decades in a variety of organisms. In the budding yeast Saccharomyces cerevisiae , this coordination or 'size control' appears as an inverse correlation between cell size and the rate of cell-cycle progression, routinely observed in G 1 prior to cell division commitment. Beyond this point, cells are presumed to complete S/G 2 /M at similar rates and in a size-independent manner. As such, studies of dependence between growth and division have focused on G 1 Moreover, in unicellular organisms, coordination between growth and division has commonly been analysed within the cycle of a single cell without accounting for correlations in growth and division characteristics between cycles of related cells. In a comprehensive analysis of three published time-lapse microscopy datasets, we analyse both intra- and inter-cycle dependencies between growth and division, revisiting assumptions about the coordination between these two processes. Interestingly, we find evidence (i) that S/G 2 /M durations are systematically longer in daughters than in mothers, (ii) of dependencies between S/G 2 /M and size at budding that echo the classical G 1 dependencies, and (iii) in contrast with recent bacterial studies, of negative dependencies between size at birth and size accumulated during the cell cycle. In addition, we develop a novel hierarchical model to uncover inter-cycle dependencies, and we find evidence for such dependencies in cells growing in sugar-poor environments. Our analysis highlights the need for experimentalists and modellers to account for new sources of cell-to-cell variation in growth and division, and our model provides a formal statistical framework for the continued study of dependencies between biological processes. © 2017 The Author(s).

Antiviral effects of artesunate on polyomavirus BK replication in primary human kidney cells.

PubMed

Sharma, Biswa Nath; Marschall, Manfred; Henriksen, Stian; Rinaldo, Christine Hanssen

2014-01-01

Polyomavirus BK (BKV) causes polyomavirus-associated nephropathy (PyVAN) and hemorrhagic cystitis (PyVHC) in renal and bone marrow transplant patients, respectively. Antiviral drugs with targeted activity against BKV are lacking. Since the antimalarial drug artesunate was recently demonstrated to have antiviral activity, the possible effects of artesunate on BKV replication in human primary renal proximal tubular epithelial cells (RPTECs), the host cells in PyVAN, were explored. At 2 h postinfection (hpi), RPTECs were treated with artesunate at concentrations ranging from 0.3 to 80 μM. After one viral replication cycle (approximately 72 hpi), the loads of extracellular BKV DNA, reflecting viral progeny production, were reduced in a concentration-dependent manner. Artesunate at 10 μM reduced the extracellular BKV load by 65%; early large T antigen mRNA and protein expression by 30% and 75%, respectively; DNA replication by 73%; and late VP1 mRNA and protein expression by 47% and 64%, respectively. Importantly, the proliferation of RPTECs was also inhibited in a concentration-dependent manner. At 72 hpi, artesunate at 10 μM reduced cellular DNA replication by 68% and total metabolic activity by 47%. Cell impedance and lactate dehydrogenase measurements indicated a cytostatic but not a cytotoxic mechanism. Flow cytometry and 5-ethynyl-2'-deoxyuridine incorporation revealed a decreased number of cells in S phase and suggested cell cycle arrest in G0 or G2 phase. Both the antiproliferative and antiviral effects of artesunate at 10 μM were reversible. Thus, artesunate inhibits BKV replication in RPTECs in a concentration-dependent manner by inhibiting BKV gene expression and genome replication. The antiviral mechanism appears to be closely connected to cytostatic effects on the host cell, underscoring the dependence of BKV on host cell proliferative functions.

Antiviral Effects of Artesunate on Polyomavirus BK Replication in Primary Human Kidney Cells

PubMed Central

Sharma, Biswa Nath; Marschall, Manfred; Henriksen, Stian

2014-01-01

Polyomavirus BK (BKV) causes polyomavirus-associated nephropathy (PyVAN) and hemorrhagic cystitis (PyVHC) in renal and bone marrow transplant patients, respectively. Antiviral drugs with targeted activity against BKV are lacking. Since the antimalarial drug artesunate was recently demonstrated to have antiviral activity, the possible effects of artesunate on BKV replication in human primary renal proximal tubular epithelial cells (RPTECs), the host cells in PyVAN, were explored. At 2 h postinfection (hpi), RPTECs were treated with artesunate at concentrations ranging from 0.3 to 80 μM. After one viral replication cycle (approximately 72 hpi), the loads of extracellular BKV DNA, reflecting viral progeny production, were reduced in a concentration-dependent manner. Artesunate at 10 μM reduced the extracellular BKV load by 65%; early large T antigen mRNA and protein expression by 30% and 75%, respectively; DNA replication by 73%; and late VP1 mRNA and protein expression by 47% and 64%, respectively. Importantly, the proliferation of RPTECs was also inhibited in a concentration-dependent manner. At 72 hpi, artesunate at 10 μM reduced cellular DNA replication by 68% and total metabolic activity by 47%. Cell impedance and lactate dehydrogenase measurements indicated a cytostatic but not a cytotoxic mechanism. Flow cytometry and 5-ethynyl-2′-deoxyuridine incorporation revealed a decreased number of cells in S phase and suggested cell cycle arrest in G0 or G2 phase. Both the antiproliferative and antiviral effects of artesunate at 10 μM were reversible. Thus, artesunate inhibits BKV replication in RPTECs in a concentration-dependent manner by inhibiting BKV gene expression and genome replication. The antiviral mechanism appears to be closely connected to cytostatic effects on the host cell, underscoring the dependence of BKV on host cell proliferative functions. PMID:24145549

Phenazine-1-carboxylic acid influences biofilm development and turnover of rhizobacterial biomass in a soil moisture-dependent manner

USDA-ARS?s Scientific Manuscript database

Rhizobacterial biofilm development influences terrestrial carbon and nitrogen cycles with ramifications for crop and soil health. Phenazine-1-carboxylic acid (PCA) is a redox-active metabolite produced by rhizobacteria in dryland wheat fields of Washington and Oregon, USA. PCA promotes biofilm dev...

Morphogenesis checkpoint kinase Swe1 is the executor of lipolysis-dependent cell-cycle progression

PubMed Central

Chauhan, Neha; Visram, Myriam; Cristobal-Sarramian, Alvaro; Sarkleti, Florian

2015-01-01

Cell growth and division requires the precise duplication of cellular DNA content but also of membranes and organelles. Knowledge about the cell-cycle–dependent regulation of membrane and storage lipid homeostasis is only rudimentary. Previous work from our laboratory has shown that the breakdown of triacylglycerols (TGs) is regulated in a cell-cycle–dependent manner, by activation of the Tgl4 lipase by the major cyclin-dependent kinase Cdc28. The lipases Tgl3 and Tgl4 are required for efficient cell-cycle progression during the G1/S (Gap1/replication phase) transition, at the onset of bud formation, and their absence leads to a cell-cycle delay. We now show that defective lipolysis activates the Swe1 morphogenesis checkpoint kinase that halts cell-cycle progression by phosphorylation of Cdc28 at tyrosine residue 19. Saturated long-chain fatty acids and phytosphingosine supplementation rescue the cell-cycle delay in the Tgl3/Tgl4 lipase-deficient strain, suggesting that Swe1 activity responds to imbalanced sphingolipid metabolism, in the absence of TG degradation. We propose a model by which TG-derived sphingolipids are required to activate the protein phosphatase 2A (PP2ACdc55) to attenuate Swe1 phosphorylation and its inhibitory effect on Cdc28 at the G1/S transition of the cell cycle. PMID:25713391

Cell cycle regulation of the BRCA1/acetyl-CoA-carboxylase complex.

PubMed

Ray, H; Suau, F; Vincent, A; Dalla Venezia, N

2009-01-16

Germ-line alterations in BRCA1 are associated with an increased susceptibility to breast and ovarian cancer. The BRCA1 protein has been implicated in multiple cellular functions. We have recently demonstrated that BRCA1 reduces acetyl-CoA-carboxylase alpha (ACCA) activity through its phospho-dependent binding to ACCA, and further established that the phosphorylation of the Ser1263 of ACCA is required for this interaction. Here, to gain more insight into the cellular conditions that trigger the BRCA1/ACCA interaction, we designed an anti-pSer1263 antibody and demonstrated that the Ser1263 of ACCA is phosphorylated in vivo, in a cell cycle-dependent manner. We further showed that the interaction between BRCA1 and ACCA is regulated during cell cycle progression. Taken together, our findings reveal a novel mechanism of regulation of ACCA distinct from the previously described phosphorylation of Ser79, and provide new insights into the control of lipogenesis through the cell cycle.

A Unique cis-Encoded Small Noncoding RNA Is Regulating Legionella pneumophila Hfq Expression in a Life Cycle-Dependent Manner.

PubMed

Oliva, Giulia; Sahr, Tobias; Rolando, Monica; Knoth, Maike; Buchrieser, Carmen

2017-01-10

Legionella pneumophila is an environmental bacterium that parasitizes protozoa, but it may also infect humans, thereby causing a severe pneumonia called Legionnaires' disease. To cycle between the environment and a eukaryotic host, L. pneumophila is regulating the expression of virulence factors in a life cycle-dependent manner: replicating bacteria do not express virulence factors, whereas transmissive bacteria are highly motile and infective. Here we show that Hfq is an important regulator in this network. Hfq is highly expressed in transmissive bacteria but is expressed at very low levels in replicating bacteria. A L. pneumophila hfq deletion mutant exhibits reduced abilities to infect and multiply in Acanthamoeba castellanii at environmental temperatures. The life cycle-dependent regulation of Hfq expression depends on a unique cis-encoded small RNA named Anti-hfq that is transcribed antisense of the hfq transcript and overlaps its 5' untranslated region. The Anti-hfq sRNA is highly expressed only in replicating L. pneumophila where it regulates hfq expression through binding to the complementary regions of the hfq transcripts. This results in reduced Hfq protein levels in exponentially growing cells. Both the small noncoding RNA (sRNA) and hfq mRNA are bound and stabilized by the Hfq protein, likely leading to the cleavage of the RNA duplex by the endoribonuclease RNase III. In contrast, after the switch to transmissive bacteria, the sRNA is not expressed, allowing now an efficient expression of the hfq gene and consequently Hfq. Our results place Hfq and its newly identified sRNA anti-hfq in the center of the regulatory network governing L. pneumophila differentiation from nonvirulent to virulent bacteria. The abilities of L. pneumophila to replicate intracellularly and to cause disease depend on its capacity to adapt to different extra- and intracellular environmental conditions. Therefore, a timely and fine-tuned expression of virulence factors and adaptation traits is crucial. Yet, the regulatory circuits governing the life cycle of L. pneumophila from replicating to virulent bacteria are only partly uncovered. Here we show that the life cycle-dependent regulation of the RNA chaperone Hfq relies on a small regulatory RNA encoded antisense to the hfq-encoding gene through a base pairing mechanism. Furthermore, Hfq regulates its own expression in an autoregulatory loop. The discovery of this RNA regulatory mechanism in L. pneumophila is an important step forward in the understanding of how the switch from inoffensive, replicating to highly virulent, transmissive L. pneumophila is regulated. Copyright © 2017 Oliva et al.

Proteomic analysis of the bacterial cell cycle

PubMed Central

Grünenfelder, Björn; Rummel, Gabriele; Vohradsky, Jiri; Röder, Daniel; Langen, Hanno; Jenal, Urs

2001-01-01

A global approach was used to analyze protein synthesis and stability during the cell cycle of the bacterium Caulobacter crescentus. Approximately one-fourth (979) of the estimated C. crescentus gene products were detected by two-dimensional gel electrophoresis, 144 of which showed differential cell cycle expression patterns. Eighty-one of these proteins were identified by mass spectrometry and were assigned to a wide variety of functional groups. Pattern analysis revealed that coexpression groups were functionally clustered. A total of 48 proteins were rapidly degraded in the course of one cell cycle. More than half of these unstable proteins were also found to be synthesized in a cell cycle-dependent manner, establishing a strong correlation between rapid protein turnover and the periodicity of the bacterial cell cycle. This is, to our knowledge, the first evidence for a global role of proteolysis in bacterial cell cycle control. PMID:11287652

Effects of Histone Deacetylase Inhibitor Panobinostat (LBH589) on Bone Marrow Mononuclear Cells of Relapsed or Refractory Multiple Myeloma Patients and Its Mechanisms

PubMed Central

Ma, Yanping; Liu, Wenhua; Zhang, Ling; Jia, Gu

2017-01-01

Background The aim of this study was to explore the impact of LBH589 alone or in combination with proteasome inhibitor bortezomib on multiple myeloma (MM) cell proliferation and its mechanism. Material/Methods MM cell line U266 and RRMM-BMMNC were treated with different concentrations of LBH589 alone or in combination with bortezomib. Cell proliferation was detected by MTT assay. Cell cycle and apoptosis was analyzed by flow cytometry. The protein and mRNA level of related genes was determined by Western blotting and qRT-PCR respectively. Results U266 cell and RRMM-BMMNC proliferation were inhibited by different concentrations of LBH589 (0, 10, 20, and 50 nmol/L) alone or 50 nmol/L of LBH589 in combination with bortezomib (10 and 20 nmol/L) in a dose- and time-dependent manner. LBH589 significantly induced G0/G1phase arrest and apoptosis in RRMM-BMMNC in a dose-dependent manner. The effects were significantly higher in all combined groups than in single-agent groups (all P<0.05). The mRNA level of Caspase3 and APAF1 were up-regulated gradually, while TOSO gene expression in RRMM-BMMNC was down-regulated gradually in a dose- and time-dependent manner. Moreover, LBH589 significantly induced hyperacetylation of histone H4, the protein level of PARP notably increased, and the level of Bcl-X decreased. Conclusions LBH589 can inhibit MM cell growth, block the cell cycle, and induce cell apoptosis, which has an anti-resistant effect on multidrug-resistant cells. LBH589 in combination with bortezomib has a synergistic effect on myeloma cells; its mechanism and reversal of drug resistance mechanism is involved in multiple changes in gene expression. PMID:29080899

Hypoxia induces p53 accumulation in the S-phase and accumulation of hypophosphorylated retinoblastoma protein in all cell cycle phases of human melanoma cells.

PubMed Central

Danielsen, T.; Hvidsten, M.; Stokke, T.; Solberg, K.; Rofstad, E. K.

1998-01-01

Hypoxia has been shown to induce accumulation of p53 and of hypophosphorylated retinoblastoma protein (pRb) in tumour cells. In this study, the cell cycle dependence of p53 accumulation and pRb hypophosphorylation in four human melanoma cell lines that are wild type for p53 was investigated using two-parameter flow cytometry measurements of p53 or pRb protein content and DNA content. The hypoxia-induced increase in p53 protein was higher in S-phase than in G1 and G2 phases in all cell lines. The accumulation of p53 in S-phase during hypoxia was not related to hypoxia-induced apoptosis or substantial cell cycle specific cell inactivation during the first 24 h of reoxygenation. pRb was hypophosphorylated in all cell cycle phases by hypoxia treatment. The results did not support a direct link between p53 and pRb during hypoxia because p53 was induced in a cell cycle-specific manner, whereas no cell cycle-dependent differences in pRb hypophosphorylation were detected. Only a fraction of the cell populations (0.60+/-0.10) showed hypophosphorylated pRb. Thus, pRb is probably not the only mediator of the hypoxia-induced cell cycle block seen in all cells and all cell cycle phases. Moreover, the cell cycle-dependent induction of p53 by hypoxia suggests that the primary function of p53 accumulation during hypoxia is other than to arrest the cells. Images Figure 4 Figure 7 PMID:9862563

DOE Office of Scientific and Technical Information (OSTI.GOV)

Heart, Emma; Palo, Meridith; Womack, Trayce

Pancreatic β-cells release insulin in response to elevation of glucose from basal (4–7 mM) to stimulatory (8–16 mM) levels. Metabolism of glucose by the β-cell results in the production of low levels of reactive oxygen intermediates (ROI), such as hydrogen peroxide (H{sub 2}O{sub 2}), a newly recognized coupling factor linking glucose metabolism to insulin secretion. However, high and toxic levels of H{sub 2}O{sub 2} inhibit insulin secretion. Menadione, which produces H{sub 2}O{sub 2} via redox cycling mechanism in a dose-dependent manner, was investigated for its effect on β-cell metabolism and insulin secretion in INS-1 832/13, a rat β-cell insulinoma cellmore » line, and primary rodent islets. Menadione-dependent redox cycling and resulting H{sub 2}O{sub 2} production under stimulatory glucose exceeded several-fold those reached at basal glucose. This was paralleled by a differential effect of menadione (0.1–10 μM) on insulin secretion, which was enhanced at basal, but inhibited at stimulatory glucose. Redox cycling of menadione and H{sub 2}O{sub 2} formation was dependent on glycolytically-derived NADH, as inhibition of glycolysis and application of non-glycogenic insulin secretagogues did not support redox cycling. In addition, activity of plasma membrane electron transport, a system dependent in part on glycolytically-derived NADH, was also inhibited by menadione. Menadione-dependent redox cycling was sensitive to the NQO1 inhibitor dicoumarol and the flavoprotein inhibitor diphenylene iodonium, suggesting a role for NQO1 and other oxidoreductases in this process. These data may explain the apparent dichotomy between the stimulatory and inhibitory effects of H{sub 2}O{sub 2} and menadione on insulin secretion. -- Highlights: ► Menadione stimulation or inhibition of insulin secretion is dependent upon applied glucose levels. ► Menadione-dependent H{sub 2}O{sub 2} production is proportional to applied glucose levels. ► Quinone-mediated redox cycling is dependent on glycolysis.« less

5-AED Enhances Survival of Irradiated Mice in a G-CSF-Dependent Manner, Stimulates Innate Immune Cell Function, Reduces Radiation-induced DNA Damage and Induces Genes that Modulate Cell Cycle Progression and Apoptosis

DTIC Science & Technology

2012-01-01

modulate cell cycle progression and apoptosis. INTRODUCTION Because of the increasing threat posed by nuclear weapons [1], there is a pressing need for both...were per- formed using the iCycler iQ Sequence Detection System ( Bio -Rad Laboratories, Hercules CA) on 96-well microtiter plates with optical caps...Thoss K, Petrow PK et al. Amelioration of murine antigen -induced arthritis by dehydroepiandrosterone (DHEA). Inflamm Res 2004;53:189–98. 56. Auci D

5-AED Enhances Survival of Irradiated Mice in a G-CSF-Dependent Manner, Stimulates Innate Immune Cell Function, Reduces Radiation-Induced DNA Damage and Induces Genes that Modulate Cell Cycle Progression and Apoptosis

DTIC Science & Technology

2012-07-22

modulate cell cycle progression and apoptosis. INTRODUCTION Because of the increasing threat posed by nuclear weapons [1], there is a pressing need for both...Detection System ( Bio -Rad Laboratories, Hercules CA) on 96-well microtiter plates with optical caps. Reactions were performed in a total volume of 50 µL... antigen -induced arthritis by dehydroepiandrosterone (DHEA). Inflamm Res 2004;53:189–98. 56. Auci D, Nicoletti F, Mangano K et al. Anti-inflammatory and

The level of menadione redox-cycling in pancreatic β-cells is proportional to the glucose concentration: role of NADH and consequences for insulin secretion

PubMed Central

Heart, Emma; Palo, Meridith; Womack, Trayce; Smith, Peter J. S.; Gray, Joshua P.

2011-01-01

Pancreatic β-cells release insulin in response to elevation of glucose from basal (4-7 mM) to stimulatory (8-16 mM) levels. Metabolism of glucose by the β-cell results in the production of low levels of reactive oxygen intermediates (ROI), such as hydrogen peroxide (H2O2), a newly recognized coupling factor linking glucose metabolism to insulin secretion. However, high and toxic levels of H2O2 inhibit insulin secretion. Menadione, which produces H2O2 via redox cycling mechanism in a dose-dependent manner, was investigated for its effect on β-cell metabolism and insulin secretion in INS-1 832/13, a rat β-cell insulinoma cell line, and primary rodent islets. Menadione-dependent redox cycling and resulting H2O2 production under stimulatory glucose exceeded several-fold those reached at basal glucose. This was paralleled by a differential effect of menadione (0.1-10 μM) on insulin secretion, which was enhanced at basal, but inhibited at stimulatory glucose. Redox cycling of menadione and H2O2 formation was dependent on glycolytically-derived NADH, as inhibition of glycolysis and application of non-glycogenic insulin secretagogues did not support redox cycling. In addition, activity of plasma membrane electron transport, a system dependent in part on glycolytically-derived NADH, was also inhibited by menadione. Menadione-dependent redox cycling was sensitive to the NQO1 inhibitor dicoumarol and the flavoprotein inhibitor diphenylene iodonium, suggesting a role for NQO1 and other oxidoreductases in this process. These data may explain the apparent dichotomy between the stimulatory and inhibitory effects of H2O2 and menadione on insulin secretion. PMID:22115979

The level of menadione redox-cycling in pancreatic β-cells is proportional to the glucose concentration: role of NADH and consequences for insulin secretion.

PubMed

Heart, Emma; Palo, Meridith; Womack, Trayce; Smith, Peter J S; Gray, Joshua P

2012-01-15

Pancreatic β-cells release insulin in response to elevation of glucose from basal (4-7mM) to stimulatory (8-16mM) levels. Metabolism of glucose by the β-cell results in the production of low levels of reactive oxygen intermediates (ROI), such as hydrogen peroxide (H(2)O(2)), a newly recognized coupling factor linking glucose metabolism to insulin secretion. However, high and toxic levels of H(2)O(2) inhibit insulin secretion. Menadione, which produces H(2)O(2) via redox cycling mechanism in a dose-dependent manner, was investigated for its effect on β-cell metabolism and insulin secretion in INS-1 832/13, a rat β-cell insulinoma cell line, and primary rodent islets. Menadione-dependent redox cycling and resulting H(2)O(2) production under stimulatory glucose exceeded several-fold those reached at basal glucose. This was paralleled by a differential effect of menadione (0.1-10μM) on insulin secretion, which was enhanced at basal, but inhibited at stimulatory glucose. Redox cycling of menadione and H(2)O(2) formation was dependent on glycolytically-derived NADH, as inhibition of glycolysis and application of non-glycogenic insulin secretagogues did not support redox cycling. In addition, activity of plasma membrane electron transport, a system dependent in part on glycolytically-derived NADH, was also inhibited by menadione. Menadione-dependent redox cycling was sensitive to the NQO1 inhibitor dicoumarol and the flavoprotein inhibitor diphenylene iodonium, suggesting a role for NQO1 and other oxidoreductases in this process. These data may explain the apparent dichotomy between the stimulatory and inhibitory effects of H(2)O(2) and menadione on insulin secretion. Published by Elsevier Inc.

Dengue Virus Modulates the Unfolded Protein Response in a Time-dependent Manner*

PubMed Central

Peña, José; Harris, Eva

2011-01-01

Flaviviruses, such as dengue virus (DENV), depend on the host endoplasmic reticulum for translation, replication, and packaging of their genomes. Here we report that DENV-2 infection modulates the unfolded protein response in a time-dependent manner. We show that early DENV-2 infection triggers and then suppresses PERK-mediated eIF2α phosphorylation and that in mid and late DENV-2 infection, the IRE1-XBP1 and ATF6 pathways are activated, respectively. Activation of IRE1-XBP1 correlated with induction of downstream targets GRP78, CHOP, and GADD34. Furthermore, induction of CHOP did not induce apoptotic markers, such as suppression of anti-apoptotic protein Bcl-2, activation of caspase-9 or caspase-3, and cleavage of poly(ADP-ribose) polymerase. Finally, we show that DENV-2 replication is affected in PERK−/− and IRE1−/− mouse embryo fibroblasts when compared with wild-type mouse embryo fibroblasts. These results demonstrate that time-dependent activation of the unfolded protein response by DENV-2 can override inhibition of translation, prevent apoptosis, and prolong the viral life cycle. PMID:21385877

Cell cycle G2/M arrest through an S phase-dependent mechanism by HIV-1 viral protein R.

PubMed

Li, Ge; Park, Hyeon U; Liang, Dong; Zhao, Richard Y

2010-07-07

Cell cycle G2 arrest induced by HIV-1 Vpr is thought to benefit viral proliferation by providing an optimized cellular environment for viral replication and by skipping host immune responses. Even though Vpr-induced G2 arrest has been studied extensively, how Vpr triggers G2 arrest remains elusive. To examine this initiation event, we measured the Vpr effect over a single cell cycle. We found that even though Vpr stops the cell cycle at the G2/M phase, but the initiation event actually occurs in the S phase of the cell cycle. Specifically, Vpr triggers activation of Chk1 through Ser345 phosphorylation in an S phase-dependent manner. The S phase-dependent requirement of Chk1-Ser345 phosphorylation by Vpr was confirmed by siRNA gene silencing and site-directed mutagenesis. Moreover, downregulation of DNA replication licensing factors Cdt1 by siRNA significantly reduced Vpr-induced Chk1-Ser345 phosphorylation and G2 arrest. Even though hydroxyurea (HU) and ultraviolet light (UV) also induce Chk1-Ser345 phosphorylation in S phase under the same conditions, neither HU nor UV-treated cells were able to pass through S phase, whereas vpr-expressing cells completed S phase and stopped at the G2/M boundary. Furthermore, unlike HU/UV, Vpr promotes Chk1- and proteasome-mediated protein degradations of Cdc25B/C for G2 induction; in contrast, Vpr had little or no effect on Cdc25A protein degradation normally mediated by HU/UV. These data suggest that Vpr induces cell cycle G2 arrest through a unique molecular mechanism that regulates host cell cycle regulation in an S-phase dependent fashion.

Human cytomegalovirus tegument protein pp150 acts as a cyclin A2-CDK-dependent sensor of the host cell cycle and differentiation state.

PubMed

Bogdanow, Boris; Weisbach, Henry; von Einem, Jens; Straschewski, Sarah; Voigt, Sebastian; Winkler, Michael; Hagemeier, Christian; Wiebusch, Lüder

2013-10-22

Upon cell entry, herpesviruses deliver a multitude of premade virion proteins to their hosts. The interplay between these incoming proteins and cell-specific regulatory factors dictates the outcome of infections at the cellular level. Here, we report a unique type of virion-host cell interaction that is essential for the cell cycle and differentiation state-dependent onset of human cytomegalovirus (HCMV) lytic gene expression. The major tegument 150-kDa phosphoprotein (pp150) of HCMV binds to cyclin A2 via a functional RXL/Cy motif resulting in its cyclin A2-dependent phosphorylation. Alanine substitution of the RXL/Cy motif prevents this interaction and allows the virus to fully escape the cyclin-dependent kinase (CDK)-mediated block of immediate early (IE) gene expression in S/G2 phase that normally restricts the onset of the HCMV replication cycle to G0/G1. Furthermore, the cyclin A2-CDK-pp150 axis is also involved in the establishment of HCMV quiescence in NTera2 cells, showing the importance of this molecular switch for differentiation state-dependent regulation of IE gene expression. Consistent with the known nucleocapsid-binding function of pp150, its RXL/Cy-dependent phosphorylation affects gene expression of the parental virion only, suggesting a cis-acting, virus particle-associated mechanism of control. The pp150 homologs of other primate and mammalian CMVs lack an RXL/Cy motif and accordingly even the nearest relative of HCMV, chimpanzee CMV, starts its lytic cycle in a cell cycle-independent manner. Thus, HCMV has evolved a molecular sensor for cyclin A2-CDK activity to restrict its IE gene expression program as a unique level of self-limitation and adaptation to its human host.

The BMI1 inhibitor PTC-209 is a potential compound to halt cellular growth in biliary tract cancer cells

PubMed Central

Mayr, Christian; Wagner, Andrej; Loeffelberger, Magdalena; Bruckner, Daniela; Jakab, Martin; Berr, Frieder; Di Fazio, Pietro; Ocker, Matthias; Neureiter, Daniel; Pichler, Martin; Kiesslich, Tobias

2016-01-01

BMI1 is a core component of the polycomb repressive complex 1 (PRC1) and is up-regulated in biliary tract cancer (BTC), contributing to aggressive clinical features. In this study we investigated the cytotoxic effects of PTC-209, a recently developed inhibitor of BMI1, in BTC cells. PTC-209 reduced overall viability in BTC cell lines in a dose-dependent fashion (0.04 - 20 μM). Treatment with PTC-209 led to slightly enhanced caspase activity and stop of cell proliferation. Cell cycle analysis revealed that PTC-209 caused cell cycle arrest at the G1/S checkpoint. A comprehensive investigation of expression changes of cell cycle-related genes showed that PTC-209 caused significant down-regulation of cell cycle-promoting genes as well as of genes that contribute to DNA synthesis initiation and DNA repair, respectively. This was accompanied by significantly elevated mRNA levels of cell cycle inhibitors. In addition, PTC-209 reduced sphere formation and, in a cell line-dependent manner, aldehyde dehydrogease-1 positive cells. We conclude that PTC-209 might be a promising drug for future in vitro and in vivo studies in BTC. PMID:26623561

A futile cycle, formed between two ATP-dependant gamma-glutamyl cycle enzymes, gamma-glutamyl cysteine synthetase and 5-oxoprolinase: the cause of cellular ATP depletion in nephrotic cystinosis?

PubMed

Kumar, Akhilesh; Bachhawat, Anand Kumar

2010-03-01

Cystinosis, an inherited disease caused by a defect in the lysosomal cystine transporter (CTNS), is characterized by renal proximal tubular dysfunction. Adenosine triphosphate (ATP) depletion appears to be a key event in the pathophysiology of the disease, even though the manner in which ATP depletion occurs is still a puzzle. We present a model that explains how a futile cycle that is generated between two ATP-utilizing enzymes of the gamma-glutamyl cycle leads to ATP depletion. The enzyme gamma-glutamyl cysteine synthetase (gamma-GCS), in the absence of cysteine, forms 5-oxoproline (instead of the normal substrate, gamma-glutamyl cysteine) and the 5-oxoproline is converted into glutamate by the ATP-dependant enzyme, 5-oxoprolinase. Thus, in cysteine-limiting conditions, glutamate is cycled back into glutamate via 5-oxoproline at the cost of two ATP molecules without production of glutathione and is the cause of the decreased levels of glutathione synthesis, as well as the ATP depletion observed in these cells. The model is also compatible with the differences seen in the human patients and the mouse model of cystinosis, where renal failure is not observed.

[Study on thaspine in inducing apoptosis of A549 cell].

PubMed

Zhang, Yan-min; He, Lang-chong

2007-04-01

To investigate the effect of thaspine on the cellular proliferation, apoptosis and cell cycle in A549 cell line. A549 cell was cultured with different concentrations of thaspine. Cellular proliferation was detected with MTT, apoptosis and cell cycle were checked with Flow Cytometer, and change of microstructure was observed by transmission electron microscope. Thaspine could inhibit the proliferation and induce apoptosis of A549 cell in a time-dose dependent manner. Cell cycle was significantly stopped at the S phase by thaspine with FCM technology. Under electronic microscope, the morphology of A549 cell showed nuclear karyopycnosis, chromatin agglutination and typical apoptotic body when the cell was treated with thaspine. Thaspine has the effects of anti-tumor and inducing apoptosis.

Separase is recruited to mitotic chromosomes to dissolve sister chromatid cohesion in a DNA-dependent manner.

PubMed

Sun, Yuxiao; Kucej, Martin; Fan, Heng-Yu; Yu, Hong; Sun, Qing-Yuan; Zou, Hui

2009-04-03

Sister chromatid separation is triggered by the separase-catalyzed cleavage of cohesin. This process is temporally controlled by cell-cycle-dependent factors, but its biochemical mechanism and spatial regulation remain poorly understood. We report that cohesin cleavage by human separase requires DNA in a sequence-nonspecific manner. Separase binds to DNA in vitro, but its proteolytic activity, measured by its autocleavage, is not stimulated by DNA. Instead, biochemical characterizations suggest that DNA mediates cohesin cleavage by bridging the interaction between separase and cohesin. In human cells, a fraction of separase localizes to the mitotic chromosome. The importance of the chromosomal DNA in cohesin cleavage is further demonstrated by the observation that the cleavage of the chromosome-associated cohesins is sensitive to nuclease treatment. Our observations explain why chromosome-associated cohesins are specifically cleaved by separase and the soluble cohesins are left intact in anaphase.

Evodiamine Induces Cell Growth Arrest, Apoptosis and Suppresses Tumorigenesis in Human Urothelial Cell Carcinoma Cells.

PubMed

Shi, Chung-Sheng; Li, Jhy-Ming; Chin, Chih-Chien; Kuo, Yi-Hung; Lee, Ying-Ray; Huang, Yun-Ching

2017-03-01

Evodiamine, an indole alkaloid derived from Evodia rutaecarpa, exhibits pharmacological activities including vasodilatation, analgesia, anti-cardiovascular disease, anti-Alzheimer's disease, anti-inflammation, and anti-tumor activity. This study analyzes the anti-tumor effects of evodiamine on cellular growth, tumorigenesis, cell cycle and apoptosis induction of human urothelial cell carcinoma (UCC) cells. The present study showed that evodiamine significantly inhibited the proliferation of UCC cells in a dose- and time-dependent manner. Also, evodiamine suppressed the tumorigenesis of UCC cells in vitro. Moreover, evodiamine caused G 2 /M cell-cycle arrest and induced caspase-dependent apoptosis in UCC cells. Finally, we demonstrated that evodiamine exhibits better cytotoxic than 5-fluorouracil, a clinical chemotherapeutic drug, for UCC cells. Evodiamine induces growth inhibition, tumorigenesis suppression, cell-cycle arrest, and apoptosis induction in human UCC cells. Therefore, this agent displays a therapeutic potential for treating human UCC cells and is worthy for further investigation. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

DNA Replication Is Required for Circadian Clock Function by Regulating Rhythmic Nucleosome Composition.

PubMed

Liu, Xiao; Dang, Yunkun; Matsu-Ura, Toru; He, Yubo; He, Qun; Hong, Christian I; Liu, Yi

2017-07-20

Although the coupling between circadian and cell cycles allows circadian clocks to gate cell division and DNA replication in many organisms, circadian clocks were thought to function independently of cell cycle. Here, we show that DNA replication is required for circadian clock function in Neurospora. Genetic and pharmacological inhibition of DNA replication abolished both overt and molecular rhythmicities by repressing frequency (frq) gene transcription. DNA replication is essential for the rhythmic changes of nucleosome composition at the frq promoter. The FACT complex, known to be involved in histone disassembly/reassembly, is required for clock function and is recruited to the frq promoter in a replication-dependent manner to promote replacement of histone H2A.Z by H2A. Finally, deletion of H2A.Z uncoupled the dependence of the circadian clock on DNA replication. Together, these results establish circadian clock and cell cycle as interdependent coupled oscillators and identify DNA replication as a critical process in the circadian mechanism. Published by Elsevier Inc.

Simvastatin Potently Induces Calcium-dependent Apoptosis of Human Leiomyoma Cells*

PubMed Central

Borahay, Mostafa A.; Kilic, Gokhan S.; Yallampalli, Chandrasekha; Snyder, Russell R.; Hankins, Gary D. V.; Al-Hendy, Ayman; Boehning, Darren

2014-01-01

Statins are drugs commonly used for the treatment of high plasma cholesterol levels. Beyond these well known lipid-lowering properties, they possess broad-reaching effects in vivo, including antitumor effects. Statins inhibit the growth of multiple tumors. However, the mechanisms remain incompletely understood. Here we show that simvastatin inhibits the proliferation of human leiomyoma cells. This was associated with decreased mitogen-activated protein kinase signaling and multiple changes in cell cycle progression. Simvastatin potently stimulated leiomyoma cell apoptosis in a manner mechanistically dependent upon apoptotic calcium release from voltage-gated calcium channels. Therefore, simvastatin possesses antitumor effects that are dependent upon the apoptotic calcium release machinery. PMID:25359773

Twitchin can regulate the ATPase cycle of actomyosin in a phosphorylation-dependent manner in skinned mammalian skeletal muscle fibres.

PubMed

Avrova, Stanislava V; Rysev, Nikita A; Matusovsky, Oleg S; Shelud'ko, Nikolay S; Borovikov, Yurii S

2012-05-01

The effect of twitchin, a thick filament protein of molluscan muscles, on the actin-myosin interaction at several mimicked sequential steps of the ATPase cycle was investigated using the polarized fluorescence of 1.5-IAEDANS bound to myosin heads, FITC-phalloidin attached to actin and acrylodan bound to twitchin in the glycerol-skinned skeletal muscle fibres of mammalian. The phosphorylation-dependent multi-step changes in mobility and spatial arrangement of myosin SH1 helix, actin subunit and twitchin during the ATPase cycle have been revealed. It was shown that nonphosphorylated twitchin inhibited the movements of SH1 helix of the myosin heads and actin subunits and decreased the affinity of myosin to actin by freezing the position and mobility of twitchin in the muscle fibres. The phosphorylation of twitchin reverses this effect by changing the spatial arrangement and mobility of the actin-binding portions of twitchin. In this case, enhanced movements of SH1 helix of the myosin heads and actin subunits are observed. The data imply a novel property of twitchin incorporated into organized contractile system: its ability to regulate the ATPase cycle in a phosphorylation-dependent fashion by changing the affinity and spatial arrangement of the actin-binding portions of twitchin. Copyright © 2012 Elsevier Inc. All rights reserved.

Ethanolic Neem (Azadirachta indica) Leaf Extract Prevents Growth of MCF-7 and HeLa Cells and Potentiates the Therapeutic Index of Cisplatin

PubMed Central

Sharma, Chhavi; Vas, Andrea J.; Goala, Payal; Gheewala, Taher M.; Rizvi, Tahir A.

2014-01-01

The present study was designed to gain insight into the antiproliferative activity of ethanolic neem leaves extract (ENLE) alone or in combination with cisplatin by cell viability assay on human breast (MCF-7) and cervical (HeLa) cancer cells. Nuclear morphological examination and cell cycle analysis were performed to determine the mode of cell death. Further, to identify its molecular targets, the expression of genes involved in apoptosis, cell cycle progression, and drug metabolism was analyzed by RT-PCR. Treatment of MCF-7, HeLa, and normal cells with ENLE differentially suppressed the growth of cancer cells in a dose- and time-dependent manner through apoptosis. Additionally, lower dose combinations of ENLE with cisplatin resulted in synergistic growth inhibition of these cells compared to the individual drugs (combination index <1). ENLE significantly modulated the expression of bax, cyclin D1, and cytochrome P450 monooxygenases (CYP 1A1 and CYP 1A2) in a time-dependent manner in these cells. Conclusively, these results emphasize the chemopreventive ability of neem alone or in combination with chemotherapeutic treatment to reduce the cytotoxic effects on normal cells, while potentiating their efficacy at lower doses. Thus, neem may be a prospective therapeutic agent to combat gynecological cancers. PMID:24624140

Annatto Tocotrienol Induces a Cytotoxic Effect on Human Prostate Cancer PC3 Cells via the Simultaneous Inhibition of Src and Stat3.

PubMed

Sugahara, Ryosuke; Sato, Ayami; Uchida, Asuka; Shiozawa, Shinya; Sato, Chiaki; Virgona, Nantiga; Yano, Tomohiro

2015-01-01

Prostate cancer is one of the most frequently occurring cancers and often acquires the potential of androgen-independent growth as a malignant phenotype. Androgen-independent prostate cancer has severe chemoresistance towards conventional chemotherapeutic agents, so a new treatment approach is required for curing such prostate cancer. In this context, the present study was undertaken to check if annatto tocotrienol (main component δ-tocotrienol) could suppress cell growth in human prostate cancer (PC3, androgen-independent type) cells via the inhibition of Src and Stat3. The tocotrienol showed cytotoxic effects on PC3 cells in a dose-dependent manner, and the effect depended on G1 arrest in the cell cycle and subsequent induction of apoptosis. In a cytotoxic dose, the tocotrienol suppressed cellular growth via the simultaneous inhibition of Src and Stat3. Similarly, the treatment combination of both Src and Stat3 inhibitors induced cytotoxic effects in PC3 cells in an additive manner compared to each by itself. With respect to cell cycle regulation and the induction of apoptosis, the combination treatment showed a similar effect to that of the tocotrienol treatment. These results suggest that annatto tocotrienol effectively induces cytotoxicity in androgen-independent prostate cancer cells via the suppression of Src and Stat3.

Quercetin suppresses HeLa cells by blocking PI3K/Akt pathway.

PubMed

Xiang, Tao; Fang, Yong; Wang, Shi-Xuan

2014-10-01

To explore the effect of quercetin on the proliferation and apoptosis of HeLa cells, HeLa cells were incubated with quercetin at different concentrations. Cell viability was evaluated by MTT assay, cell apoptosis was detected by Annexin-V/PI double labeled cytometry and DNA ladder assay. Cell cycle was flow cytometrically determined and the morphological changes of the cells were observed under a fluorescence microscope after Hoechst 33258 staining and the apoptosis-related proteins in the HeLa cells were assessed by Western blotting. The results showed that quercetin significantly inhibited the growth of HeLa cells and induced obvious apoptosis in vitro in a time- and dose-dependent manner. Moreover, quercetin induced apoptosis of HeLa cells in cell cycle-dependent manner because quercetin could induce arrest of HeLa cells at G0/G1 phase. Quercetin treatment down-regulated the expression of the PI3K and p-Akt. In addition, quercetin could down-regulate expression of bcl-2, up-regulate Bax, but exerted no effect on the overall expression of Akt. We are led to conclude that quercetin induces apoptosis via PI3k/Akt pathways, and quercetin has potential to be used as an anti-tumor agent against human cervix cancer.

Indole-3-carbinol induces G1 cell cycle arrest and apoptosis through aryl hydrocarbon receptor in THP-1 monocytic cell line.

PubMed

Mohammadi, Saeed; Seyedhosseini, Fakhri Sadat; Behnampour, Nasser; Yazdani, Yaghoub

2017-10-01

The role of aryl hydrocarbon receptor (AhR) in carcinogenesis has been studied recently. Indole-3-carbinol (I3C) is an AhR agonist and a potential anticancer agent. Here, we investigated the effects of I3C on cell cycle progression and apoptosis through activation of AhR on THP-1 acute myeloid leukemia (AML) cell line. MTT viability assay was used to measure the cytotoxic effects of I3C on THP-1 cells. Apoptosis and cell cycle assays were investigated using flow cytometry. Real time RT-PCR was conducted to measure the alterations in the expression of AhR gene, key genes associated with AhR activation (IL1β and CYP1A1) and major genes involved in cell cycle regulation and apoptosis including P27, P21, CDK2, P53, BCL2 and FasR. Our findings revealed that I3C inhibits the proliferation of THP-1 cells in a dose- and time-dependent manner with minimal toxicity over normal monocytes. The AhR target genes (CYP1A1, IL1β) were overexpressed upon I3C treatment (p < .05 to p < .001). The antiproliferative effects of I3C were in association with programed cell death. I3C downregulated BCL2 and upregulated FasR in THP-1 cells (p < .05 to p < .001). G1 cell cycle arrest was also observed using flow cytometry. G1-acting cell cycle genes (P21, P27 and P53) were overexpressed (p < .05 to p < .001), while CDK2 was downregulated upon I3C treatment (p < .01 to p < .001). I3C could exert its antileukemic effects through AhR activation which is associated with programed cell death and G1 cell cycle arrest in a dose- and time-dependent manner. Therefore, AhR could be targeted as a novel treatment possibility in AML.

Effects of demethoxycurcumin on the viability and apoptosis of skin cancer cells.

PubMed

Wu, Yaoqun; Zhang, Pei; Yang, Hongyun; Ge, Yong; Xin, Yong

2017-07-01

The present study investigated the effects and mechanisms of demethoxycurcumin (DMC) on a human skin squamous cell carcinoma cell line, A431, and a human keratinocyte cell line, HaCaT. A431 and HaCaT cells were cultured in vitro. The effects of DMC treatment on cell viability were analyzed using the Cell Counting kit‑8 (CCK‑8) assay; cell cycle distribution was analyzed by flow cytometry; apoptosis was assessed by flow cytometry and Hoechst 33258 staining; and the protein expression levels of cytochrome c, B‑cell lymphoma 2 (Bcl‑2), Bcl‑2‑associated X protein (BAX), caspase‑9 and caspase‑3 were evaluated by western blotting. CCK‑8 assay results demonstrated that DMC treatment significantly inhibited viability of A431 and HaCaT cells in a dose‑dependent manner. Flow cytometric analysis confirmed that DMC treatment induced apoptosis in a dose‑dependent manner, and significantly increased the proportion of cells in G2/M phase. Western blot analysis indicated that the protein expression levels of Bcl‑2 were decreased, whereas the expression levels of BAX, caspase‑9, caspase‑3 and cytochrome c were increased following DMC treatment compared with in untreated cells. In conclusion, DMC treatment significantly inhibited viability of A431 and HaCaT cells, and induced cell cycle arrest in G2/M phase. The present study indicated that DMC may induce apoptosis of skin cancer cells through a caspase‑dependent pathway.

p205, a potential tumor suppressor, inhibits cell proliferation via multiple pathways of cell cycle regulation.

PubMed

Asefa, Benyam; Dermott, Jonathan M; Kaldis, Philipp; Stefanisko, Karen; Garfinkel, David J; Keller, Jonathan R

2006-02-20

p205 is a member of the interferon-inducible p200 family of proteins that regulate cell proliferation. Over-expression of p205 inhibits cell growth, although its mechanism of action is currently unknown. Therefore, we evaluated the effect of p205 on the p53 and Rb-dependent pathways of cell cycle regulation. p205 expression results in elevated levels of p21, and activates the p21 promoter in vitro in a p53-dependent manner. In addition, p205 induces increased expression of Rb, and binds directly to Rb and p53. Interestingly, p205 also induces growth inhibition independent of p53 and Rb by delaying G2/M progression in proliferating cells, and is a substrate for Cdk2 kinase activity. Finally, we have identified other binding partners of p205 by a yeast two-hybrid screen, including the paired homeodomain protein HoxB2. Taken together, our results indicate that p205 induces growth arrest by interaction with multiple transcription factors that regulate the cell cycle, including but not entirely dependent on the Rb- and p53-mediated pathways of growth inhibition.

Tumor Suppression by BRCA-1: A Critical Role at DNA Replication Forks

DTIC Science & Technology

2006-10-01

replication defect. We wished to test the hypothesis that BRCA1/BARD1 function during DNA replication supporting DNA transactions at replication forks. We...are using cell-free extracts derived from Xenopus laevis eggs that support: 1. Semi-conservative, cell-cycle regulated DNA replication ; 2. Many facets...complex assembles to chromatin in a DNA replication -dependent manner. Finally, we show that BRCA1/BARD1 loading to chromatin does not dramatically

Osthole induces apoptosis, suppresses cell-cycle progression and proliferation of cancer cells.

PubMed

Jarząb, Agata; Grabarska, Aneta; Kiełbus, Michał; Jeleniewicz, Witold; Dmoszyńska-Graniczka, Magdalena; Skalicka-Woźniak, Krystyna; Sieniawska, Elwira; Polberg, Krzysztof; Stepulak, Andrzej

2014-11-01

The aim of the present study was to determine the effects of osthole on cell proliferation and viability, cell-cycle progression and induction of apoptosis in human laryngeal cancer RK33 and human medulloblastoma TE671 cell lines. Cell viability was measured by means of the MTT method and cell proliferation by the 5-bromo-2-deoxyuridine (BrdU) incorporation assay. Cell-cycle progression was determined by flow cytometry, and induction of apoptosis by release of oligonucleosomes to the cytosol. The gene expression was estimated by a quantitative polymerase chain reaction (qPCR) method. High-performance counter-current chromatography (HPCCC) was applied for isolation of osthole from fruits of Mutellina purpurea. Osthole decreased proliferation and cell viability of cancer cells in a dose-dependent manner. The tested compound induced apoptosis, increased the cell numbers in G1 and decreased cell number in S/G2 phases of the cell cycle, differentially regulating CDKN1A and TP53 gene expression depending on cancer cell type. Osthole could be considered as a potential compound for cancer therapy and chemoprevention. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

A phthalide derivative isolated from endophytic fungi Pestalotiopsis photiniae induces G1 cell cycle arrest and apoptosis in human HeLa cells.

PubMed

Chen, C; Yang, R L

2013-08-01

MP [4-(3',3'-dimethylallyloxy)-5-methyl-6-methoxyphthalide] was obtained from liquid culture of Pestalotiopsis photiniae isolated from the Chinese Podocarpaceae plant Podocarpus macrophyllus. MP significantly inhibited the proliferation of HeLa tumor cell lines. After treatment with MP, characteristic apoptotic features such as DNA fragmentation and chromatin condensation were observed in DAPI-stained HeLa cells. Flow cytometry showed that MP induced G1 cell cycle arrest and apoptosis in a dose-dependent manner. Western blotting and real-time reverse transcription-polymerase chain reaction were used to investigate protein and mRNA expression. MP caused significant cell cycle arrest by upregulating the cyclin-dependent kinase inhibitor p27(KIP1) protein and p21(CIP1) mRNA levels in HeLa cells. The expression of p73 protein was increased after treatment with various MP concentrations. mRNA expression of the cell cycle-related genes, p21(CIP1), p16(INK4a) and Gadd45α, was significantly upregulated and mRNA levels demonstrated significantly increased translation of p73, JunB, FKHR, and Bim. The results indicate that MP may be a potential treatment for cervical cancer.

Erufosine simultaneously induces apoptosis and autophagy by modulating the Akt-mTOR signaling pathway in oral squamous cell carcinoma.

PubMed

Kapoor, Vaishali; Zaharieva, Maya M; Das, Satya N; Berger, Martin R

2012-06-01

We investigated the anticancer activity of erufosine in oral squamous carcinoma cell lines in terms of cell proliferation, colony formation, induction of autophagy/apoptosis, cell cycle and mTOR signaling pathway. Erufosine showed dose-dependent cytotoxicity in all cell lines, it induced autophagy as well as apoptosis, G2 cell cycle arrest and modulation of cyclin D1 expression. Further erufosine downregulated the phosphorylation of major components of mTOR pathway, like p-Akt at Ser473 and Thr308 residues, p-Raptor, p-mTOR, p-PRAS40 and its downstream substrates p-p70S6K and p-4EBP1 in a dose-dependent manner. The pre-treatment of tumor cells with p-mTOR siRNA increased cytotoxic effects of erufosine comparable to cisplatin but higher than rapamycin. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Anti-proliferative actions of 2-decylamino-5,8-dimethoxy-1,4-naphthoquinone in vascular smooth muscle cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Jung-Jin; Institute of Drug Research and Development, Chungnam National University, Daejeon 305-764; Zhang, Wei-Yun

2011-07-22

Highlights: {yields} 2-Decylamino-DMNQ inhibited PDGF-BB-induced VSMC proliferation in a dose-dependent manner with no apparent cytotoxicity. {yields} 2-Decylamino-DMNQ inhibited PDGF-BB-induced phosphorylation of Erk1/2 and PLC{gamma}1. {yields} 2-Decylamino-DMNQ arrested a G{sub 0}/G{sub 1} cell cycle progression in association with pRb phosphorylation and PCNA expression. {yields} Both U0126, an Erk inhibitor, and U73122, a PLC{gamma} inhibitor, arrested a G{sub 0}/G{sub 1} phase of the cell cycle. -- Abstract: Naphthoquinone derivatives have been reported to possess various pharmacological activities, such as antiplatelet, anticancer, antifungal, and antiviral properties. In this study, we investigated the effects of a newly-synthesized naphthoquinone derivative, 2-decylamino-5,8-dimethoxy-1,4-naphthoquinone (2-decylamino-DMNQ), on VSMC proliferationmore » and examined the molecular basis of the underlying mechanism. In a dose-dependent manner, 2-decylamino-DMNQ inhibited PDGF-stimulated VSMC proliferation with no apparent cytotoxic effect. While 2-decylamino-DMNQ did not affect PDGF-R{beta} or Akt, it did inhibit the phosphorylation of Erk1/2 and PLC{gamma}1 induced by PDGF. Moreover, 2-decylamino-DMNQ suppressed DNA synthesis through the arrest of cell cycle progression at the G{sub 0}/G{sub 1} phase, including the suppression of pRb phosphorylation and a decrease in PCNA expression, which was related to the downregulation of cell cycle regulatory factors, such as cyclin D1/E and CDK 2/4. It was demonstrated that both U0126, an Erk1/2 inhibitor, and U73122, a PLC{gamma} inhibitor, increased the proportion of cells in the G{sub 0}/G{sub 1} phase of the cell cycle. Thus, these results suggest that 2-decylamino DMNQ has an inhibitory effect on PDGF-induced VSMC proliferation and the mechanism of this action is through cell cycle arrest at the G{sub 0}/G{sub 1} phase. This may be a useful tool for studying interventions for vascular restenosis in coronary revascularization procedures and stent implantation.« less

Arginase inhibition reduces interleukin-1β-stimulated vascular smooth muscle cell proliferation by increasing nitric oxide synthase-dependent nitric oxide production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yoon, Jeongyeon; Ryoo, Sungwoo, E-mail: ryoosw08@kangwon.ac.kr

2013-06-07

Highlights: •Arginase inhibition suppressed proliferation of IL-1β-stimulated VSMCs in dose-dependent manner. •NO production from IL-1β-induced iNOS expression was augmented by arginase inhibition, reducing VSMC proliferation. •Incubation with cGMP analogues abolished IL-1β-dependent proliferation of VSMCs. -- Abstract: We investigated whether arginase inhibition suppressed interleukin (IL)-1β-stimulated proliferation in vascular smooth muscle cells (VSMCs) and the possible mechanisms involved. IL-1β stimulation increased VSMC proliferation, while the arginase inhibitor BEC and transfection of the antisense (AS) oligonucleotide against arginase I decreased VSMC proliferation and was associated with increased protein content of the cell cycle regulator p21Waf1/Cip1. IL-1β incubation induced inducible nitric oxide synthase (iNOS)more » mRNA expression and protein levels in a dose-dependent manner, but did not affect arginase I and II expression. Consistent with this data, IL-1β stimulation resulted in increase in NO production that was significantly augmented by arginase inhibition. The specific iNOS inhibitor 1400W abolished IL-1β-mediated NO production and further accentuated IL-1β-stimulated cell proliferation. Incubation with NO donors GSNO and DETA/NO in the presence of IL-1β abolished VSMCs proliferation and increased p21Waf1/Cip1 protein content. Furthermore, incubation with the cGMP analogue 8-Br-cGMP prevented IL-1β-induced VSMCs proliferation. In conclusion, arginase inhibition augmented iNOS-dependent NO production that resulted in suppression of IL-1β-induced VSMCs proliferation in a cGMP-dependent manner.« less

Contact guidance is cell cycle-dependent.

PubMed

Pourfarhangi, Kamyar Esmaeili; De La Hoz, Edgar Cardenas; Cohen, Andrew R; Gligorijevic, Bojana

2018-09-01

Cancer cell migration is essential for metastasis, during which cancer cells move through the tumor and reach the blood vessels. In vivo , cancer cells are exposed to contact guidance and chemotactic cues. Depending on the strength of such cues, cells will migrate in a random or directed manner. While similar cues may also stimulate cell proliferation, it is not clear whether cell cycle progression affects migration of cancer cells and whether this effect is different in random versus directed migration. In this study, we tested the effect of cell cycle progression on contact guided migration in 2D and 3D environments, in the breast carcinoma cell line, FUCCI-MDA-MB-231. The results were quantified from live cell microscopy images using the open source lineage editing and validation image analysis tools (LEVER). In 2D, cells were placed inside 10 μ m-wide microchannels to stimulate contact guidance, with or without an additional chemotactic gradient of the soluble epidermal growth factor. In 3D, contact guidance was modeled by aligned collagen fibers. In both 2D and 3D, contact guidance was cell cycle-dependent, while the addition of the chemo-attractant gradient in 2D increased cell velocity and persistence in directionally migrating cells, regardless of their cell cycle phases. In both 2D and 3D contact guidance, cells in the G1 phase of the cell cycle outperformed cells in the S/G2 phase in terms of migration persistence and instantaneous velocity. These data suggest that in the presence of contact guidance cues in vivo , breast carcinoma cells in the G1 phase of the cell cycle may be more efficient in reaching the neighboring vasculature.

Sinorhizobium meliloti CtrA Stability Is Regulated in a CbrA-Dependent Manner That Is Influenced by CpdR1

PubMed Central

Schallies, Karla B.; Sadowski, Craig; Meng, Julia; Chien, Peter

2015-01-01

ABSTRACT CbrA is a DivJ/PleC-like histidine kinase of DivK that is required for cell cycle progression and symbiosis in the alphaproteobacterium Sinorhizobium meliloti. Loss of cbrA results in increased levels of CtrA as well as its phosphorylation. While many of the known Caulobacter crescentus regulators of CtrA phosphorylation and proteolysis are phylogenetically conserved within S. meliloti, the latter lacks the PopA regulator that is required for CtrA degradation in C. crescentus. In order to investigate whether CtrA proteolysis occurs in S. meliloti, CtrA stability was assessed. During exponential growth, CtrA is unstable and therefore likely to be degraded in a cell cycle-regulated manner. Loss of cbrA significantly increases CtrA stability, but this phenotype is restored to that of the wild type by constitutive ectopic expression of a CpdR1 variant that cannot be phosphorylated (CpdR1D53A). Addition of CpdR1D53A fully suppresses cbrA mutant cell cycle defects, consistent with regulation of CtrA stability playing a key role in mediating proper cell cycle progression in S. meliloti. Importantly, the cbrA mutant symbiosis defect is also suppressed in the presence of CpdR1D53A. Thus, regulation of CtrA stability by CbrA and CpdR1 is associated with free-living cell cycle outcomes and symbiosis. IMPORTANCE The cell cycle is a fundamental process required for bacterial growth, reproduction, and developmental differentiation. Our objective is to understand how a two-component signal transduction network directs cell cycle events during free-living growth and host colonization. The Sinorhizobium meliloti nitrogen-fixing symbiosis with plants is associated with novel cell cycle events. This study identifies a link between the regulated stability of an essential response regulator, free-living cell cycle progression, and symbiosis. PMID:25897034

APELA promotes tumour growth and cell migration in ovarian cancer in a p53-dependent manner.

PubMed

Yi, Yuyin; Tsai, Shu-Huei; Cheng, Jung-Chien; Wang, Evan Y; Anglesio, Michael S; Cochrane, Dawn R; Fuller, Megan; Gibb, Ewan A; Wei, Wei; Huntsman, David G; Karsan, Aly; Hoodless, Pamela A

2017-12-01

APELA is a small, secreted peptide that can function as a ligand for the G-protein coupled receptor, Apelin Receptor (APLNR, APJ). APELA plays an essential role in endoderm differentiation and cardiac development during embryogenesis. We investigated whether APELA exerts any functions in cancer progression. The Cancer Genome Atlas (TCGA) RNA sequencing datasets, microarray from an OCCC mouse model, and RNA isolated from fresh frozen and FFPE patient tissue were used to assess APELA expression. APELA knockout ovarian clear cell carcinoma (OCCC) cell lines were generated using CRISPR/Cas9. APELA was expressed in various ovarian cancer histotypes and was especially elevated in OCCC. Disruption of APELA expression in OCCC cell lines suppressed cell growth and migration, and altered cell-cycle progression. Moreover, addition of human recombinant APELA peptide to the OCCC cell line OVISE promoted cell growth and migration. Interestingly, OVISE cells do not express APLNR, suggesting that APELA can function through an APLNR-independent pathway. Furthermore, APELA affected cell growth and cell cycle progression in a p53-dependent manner. In addition, APELA knockdown induced p53 expression in cancer cell lines. Our findings uncover a potential oncogenic role for APELA in promoting ovarian tumour progression and provide a possible therapeutic strategy in ovarian cancer by targeting APELA. Copyright © 2017 Elsevier Inc. All rights reserved.

Tangeretin and nobiletin induce G1 cell cycle arrest but not apoptosis in human breast and colon cancer cells.

PubMed

Morley, Karen L; Ferguson, Peter J; Koropatnick, James

2007-06-18

Tangeretin and nobiletin are citrus flavonoids that are among the most effective at inhibiting cancer cell growth in vitro and in vivo. The antiproliferative activity of tangeretin and nobiletin was investigated in human breast cancer cell lines MDA-MB-435 and MCF-7 and human colon cancer line HT-29. Both flavonoids inhibited proliferation in a dose- and time-dependent manner, and blocked cell cycle progression at G1 in all three cell lines. At concentrations that resulted in significant inhibition of proliferation and cell cycle arrest, neither flavonoid induced apoptosis or cell death in any of the tumor cell lines. To test the ability of arrested cells to recover, cells that were incubated with tangeretin and nobiletin for 4 days were then cultured in flavonoid-free medium for an additional 4 days. Cells resumed proliferation similar to untreated control within a day of flavonoid removal. Cell cycle distribution was similar to that of control within 4 days of flavonoid removal. These data indicate that, in these cell lines at concentrations that inhibit proliferation up to 80% over 4 days, tangeretin and nobiletin are cytostatic and significantly suppress proliferation by cell cycle arrest without apoptosis. Such an agent could be expected to spare normal tissues from toxic side effects. Thus, tangeretin and nobiletin could be effective cytostatic anticancer agents. Inhibition of proliferation of human cancers without inducing cell death may be advantageous in treating tumors as it would restrict proliferation in a manner less likely to induce cytotoxicity and death in normal, non-tumor tissues.

Visualizing the complex functions and mechanisms of the anaphase promoting complex/cyclosome (APC/C)

PubMed Central

Alfieri, Claudio; Zhang, Suyang

2017-01-01

The anaphase promoting complex or cyclosome (APC/C) is a large multi-subunit E3 ubiquitin ligase that orchestrates cell cycle progression by mediating the degradation of important cell cycle regulators. During the two decades since its discovery, much has been learnt concerning its role in recognizing and ubiquitinating specific proteins in a cell-cycle-dependent manner, the mechanisms governing substrate specificity, the catalytic process of assembling polyubiquitin chains on its target proteins, and its regulation by phosphorylation and the spindle assembly checkpoint. The past few years have witnessed significant progress in understanding the quantitative mechanisms underlying these varied APC/C functions. This review integrates the overall functions and properties of the APC/C with mechanistic insights gained from recent cryo-electron microscopy (cryo-EM) studies of reconstituted human APC/C complexes. PMID:29167309

Cell cycle arrest and induction of apoptosis by cajanin stilbene acid from Cajanus cajan in breast cancer cells.

PubMed

Fu, Yujie; Kadioglu, Onat; Wiench, Benjamin; Wei, Zuofu; Gao, Chang; Luo, Meng; Gu, Chengbo; Zu, Yuangang; Efferth, Thomas

2015-04-15

The low abundant cajanin stilbene acid (CSA) from Pigeon Pea (Cajanus cajan) has been shown to kill estrogen receptor α positive cancer cells in vitro and in vivo. Downstream effects such as cell cycle and apoptosis-related mechanisms have not been analyzed yet. We analyzed the activity of CSA by means of flow cytometry (cell cycle distribution, mitochondrial membrane potential, MMP), confocal laser scanning microscopy (MMP), DNA fragmentation assay (apoptosis), Western blotting (Bax and Bcl-2 expression, caspase-3 activation) as well as mRNA microarray hybridization and Ingenuity pathway analysis. CSA induced G2/M arrest and apoptosis in a concentration-dependent manner from 8.88 to 14.79 µM. The MMP broke down, Bax was upregulated, Bcl-2 downregulated and caspase-3 activated. Microarray profiling revealed that CSA affected BRCA-related DNA damage response and cell cycle-regulated chromosomal replication pathways. CSA inhibited breast cancer cells by DNA damage and cell cycle-related signaling pathways leading to cell cycle arrest and apoptosis. Copyright © 2015 Elsevier GmbH. All rights reserved.

Nucleosome architecture throughout the cell cycle

PubMed Central

Deniz, Özgen; Flores, Oscar; Aldea, Martí; Soler-López, Montserrat; Orozco, Modesto

2016-01-01

Nucleosomes provide additional regulatory mechanisms to transcription and DNA replication by mediating the access of proteins to DNA. During the cell cycle chromatin undergoes several conformational changes, however the functional significance of these changes to cellular processes are largely unexplored. Here, we present the first comprehensive genome-wide study of nucleosome plasticity at single base-pair resolution along the cell cycle in Saccharomyces cerevisiae. We determined nucleosome organization with a specific focus on two regulatory regions: transcription start sites (TSSs) and replication origins (ORIs). During the cell cycle, nucleosomes around TSSs display rearrangements in a cyclic manner. In contrast to gap (G1 and G2) phases, nucleosomes have a fuzzier organization during S and M phases, Moreover, the choreography of nucleosome rearrangements correlate with changes in gene expression during the cell cycle, indicating a strong association between nucleosomes and cell cycle-dependent gene functionality. On the other hand, nucleosomes are more dynamic around ORIs along the cell cycle, albeit with tighter regulation in early firing origins, implying the functional role of nucleosomes on replication origins. Our study provides a dynamic picture of nucleosome organization throughout the cell cycle and highlights the subsequent impact on transcription and replication activity. PMID:26818620

Lack of a peroxiredoxin suppresses the lethality of cells devoid of electron donors by channelling electrons to oxidized ribonucleotide reductase.

PubMed

Boronat, Susanna; Domènech, Alba; Carmona, Mercè; García-Santamarina, Sarela; Bañó, M Carmen; Ayté, José; Hidalgo, Elena

2017-06-01

The thioredoxin and glutaredoxin pathways are responsible of recycling several enzymes which undergo intramolecular disulfide bond formation as part of their catalytic cycles such as the peroxide scavengers peroxiredoxins or the enzyme ribonucleotide reductase (RNR). RNR, the rate-limiting enzyme of deoxyribonucleotide synthesis, is an essential enzyme relying on these electron flow cascades for recycling. RNR is tightly regulated in a cell cycle-dependent manner at different levels, but little is known about the participation of electron donors in such regulation. Here, we show that cytosolic thioredoxins Trx1 and Trx3 are the primary electron donors for RNR in fission yeast. Unexpectedly, trx1 transcript and Trx1 protein levels are up-regulated in a G1-to-S phase-dependent manner, indicating that the supply of electron donors is also cell cycle-regulated. Indeed, genetic depletion of thioredoxins triggers a DNA replication checkpoint ruled by Rad3 and Cds1, with the final goal of up-regulating transcription of S phase genes and constitutive RNR synthesis. Regarding the thioredoxin and glutaredoxin cascades, one combination of gene deletions is synthetic lethal in fission yeast: cells lacking both thioredoxin reductase and cytosolic dithiol glutaredoxin. We have isolated a suppressor of this lethal phenotype: a mutation at the Tpx1-coding gene, leading to a frame shift and a loss-of-function of Tpx1, the main client of electron donors. We propose that in a mutant strain compromised in reducing equivalents, the absence of an abundant and competitive substrate such as the peroxiredoxin Tpx1 has been selected as a lethality suppressor to favor RNR function at the expense of the non-essential peroxide scavenging function, to allow DNA synthesis and cell growth.

Arctigenin anti-tumor activity in bladder cancer T24 cell line through induction of cell-cycle arrest and apoptosis.

PubMed

Yang, Shucai; Ma, Jing; Xiao, Jianbing; Lv, Xiaohong; Li, Xinlei; Yang, Huike; Liu, Ying; Feng, Sijia; Zhang, Yafang

2012-08-01

Bladder cancer is the most common neoplasm in the urinary system. This study assesses arctigenin anti-tumor activity in human bladder cancer T24 cells in vitro and the underlying molecular events. The flow cytometry analysis was used to detect cell-cycle distribution and apoptosis. Western blotting was used to detect changes in protein expression. The data showed that arctigenin treatment reduced viability of bladder cancer T24 cells in a dose- and time-dependent manner after treatment with arctigenin (10, 20, 40, 80, and 100 μmol/L) for 24 hr and 48 hr. Arctigenin treatment clearly arrested tumor cells in the G1 phase of the cell cycle. Apoptosis was detected by hoechst stain and flow cytometry after Annexin-V-FITC/PI double staining. Early and late apoptotic cells were accounted for 2.32-7.01% and 3.07-7.35%, respectively. At the molecular level, arctigenin treatment decreased cyclin D1 expression, whereas CDK4 and CDK6 expression levels were unaffected. Moreover, arctigenin selectively altered the phosphorylation of members of the MAPK superfamily, decreasing phosphorylation of ERK1/2 and activated phosphorylation of p38 significantly in a dose-dependent manner. These results suggest that arctigenin may inhibit cell viability and induce apoptosis by direct activation of the mitochondrial pathway, and the mitogen-activated protein kinase pathway may play an important role in the anti-tumor effect of arctigenin. The data from the current study demonstrate the usefulness of arctigenin in bladder cancer T24 cells, which should further be evaluated in vivo before translation into clinical trials for the chemoprevention of bladder cancer. Copyright © 2012 Wiley Periodicals, Inc.

[Effect of shikonin, a phytocompound from Lithospermum erythrorhizon, on rat vascular smooth muscle cells proliferation and apoptosis in vitro].

PubMed

Zhang, Zhuo-qi; Cao, Xi-chuan; Zhang, Ling; Zhu, Wen-ling

2005-06-08

To study the anti-proliferation, pro-apoptosis and cell cycle blocking effects of shikonin on rat vascular smooth muscle cell (VSMC) in vitro. VSMCs were primarily cultured by explant method from the thoracic aorta of male SD rats. Shikonin of different concentration, 4, 2, 1, 0.5, 0.25, and 0 micromol/L was added. The cell viability was detected by MTT method. Cell growth curve was drawn by trypan blue exclusion method. (3)H-thymidine incorporation was used to calculate the inhibition rate of DNA synthesis. Flow cytometry was used to detect the cell cycle. Cell apoptosis was observed by fluorescence microscopy. Western blotting was performed to detect the expression of different cell apoptosis and cell cycle regulatory proteins, such as cyclin D(1) and E, proliferating cell nuclear antigen (PCNA), p21(waf1/cip1), p27(kip1), and p53. Compared with control group, shikonin had no obvious cytotoxic effect on cell viability at the concentration of 0.25-1 micromol/L (P > 0.05). While it could inhibit, both time- and dose-dependently, the growth of VSMC, which was predominant of 1 micromol/L at 72 h (1.9 x 10(5)/well vs 5.8 x 10(5)/well, P < 0.05), and DNA synthesis was also significantly inhibited in a time- and dose-dependent manner with inhibition rate varied from 33 to 98% (P < 0.05 or P < 0.01). 1 micromol/L shikonin significantly blocked the cell cycle progression in proliferative VSMC, decreased S, G(2)/M phase (P < 0.05) and increased G(0)/G(1) phase (P < 0.05) to quiescent level with sub-G(1) apoptotic distribution at 48 h (10.9% +/- 0.3%). Shikohin at the concentration of 1-2 micromol/L significantly increased the percentage of apoptotic cells in a time- and dose-dependent manner compared with control group (2.8%-23.7% vs 0.2%-0.4%, P < 0.05), and typical apoptotic nuclear morphological changes were observed. 1 micromol/L shikonin significantly down-regulated cyclin D(1), E and PCNA expression, up-regulated p21(wif1/cip1) expression, and did not obviously influence the p27(kip1) and p53 expression. Shikonin inhibits the proliferation, promotes the apoptosis and blocks cell cycle progression of VSMC. These effects are associated with the expression changes of cell cycle regulatory proteins.

Molecular mechanisms of celery seed extract induced apoptosis via s phase cell cycle arrest in the BGC-823 human stomach cancer cell line.

PubMed

Gao, Lin-Lin; Feng, Lei; Yao, Shu-Tong; Jiao, Peng; Qin, Shu-Cun; Zhang, Wei; Zhang, Ya-Bin; Li, Fu-Rong

2011-01-01

Mechanisms of apoptosis in tumor cells is an important field of tumor therapy and cancer molecular biology. Loss of cell cycle control, leading to uncontrolled proliferation, is common in cancer. Therefore, the identification of potent and selective cyclin dependent kinase inhibitors is a priority for anti-cancer drug discovery. There are at least two major apoptotic pathways, initiated by caspase-8 and caspase-9, respectively, which can activate caspase cascades. Apoptosis triggered by activation of the mitochondrial-dependent caspase pathway represents the main programmed cell death mechanism. This is activated by various intracellular stresses that induce permeabilization of the mitochondrial membrane. Anti-tumor effects of celery seed extract (CSE) and related mechanisms regarding apoptosis were here investigated in human gastric cancer BGC-823 cells. CSE was produced by supercritical fluid extraction. Cell viability was analyzed by 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl-tetrazolium bromide (MTT) assay and apoptosis by flow cytometry using Annexin/PI staining and DAPI staining and a laser scanning confocal microscope (LSCM). Cell cycling was evaluated using PI staining with flow cytometry and expression of cell cycle and apoptosis-related proteins cyclin A, CDK2, bcl-2 and bax was assessed by immunohistochemical staining. CSE had an anti-proliferation effect on human gastric cancer BGC-823 cells in a dose- and time-dependent manner. After treatment, the apoptotic rate significantly increased, with morphological changes typical of apoptosis observed with LSCM by DAPI staining. Cell cycle and apoptosis related proteins, such as cyclin A, CDK2 and bcl-2 were all down-regulated, whereas bax was up-regulated. The molecular determinants of inhibition of cell proliferation as well as apoptosis of CSE may be associated with cycle arrest in the S phase.

A phthalide derivative isolated from endophytic fungi Pestalotiopsis photiniae induces G1 cell cycle arrest and apoptosis in human HeLa cells

PubMed Central

Chen, C.; Yang, R.L.

2013-01-01

MP [4-(3′,3′-dimethylallyloxy)-5-methyl-6-methoxyphthalide] was obtained from liquid culture of Pestalotiopsis photiniae isolated from the Chinese Podocarpaceae plant Podocarpus macrophyllus. MP significantly inhibited the proliferation of HeLa tumor cell lines. After treatment with MP, characteristic apoptotic features such as DNA fragmentation and chromatin condensation were observed in DAPI-stained HeLa cells. Flow cytometry showed that MP induced G1 cell cycle arrest and apoptosis in a dose-dependent manner. Western blotting and real-time reverse transcription-polymerase chain reaction were used to investigate protein and mRNA expression. MP caused significant cell cycle arrest by upregulating the cyclin-dependent kinase inhibitor p27KIP1 protein and p21CIP1 mRNA levels in HeLa cells. The expression of p73 protein was increased after treatment with various MP concentrations. mRNA expression of the cell cycle-related genes, p21CIP1, p16INK4a and Gadd45α, was significantly upregulated and mRNA levels demonstrated significantly increased translation of p73, JunB, FKHR, and Bim. The results indicate that MP may be a potential treatment for cervical cancer. PMID:23903687

Toxoplasma gondii influences aversive behaviors of female rats in an estrus cycle dependent manner.

PubMed

Golcu, Doruk; Gebre, Rahiwa Z; Sapolsky, Robert M

2014-08-01

The protozoan Toxoplasma gondii (T. gondii) manipulates the behavior of its rodent intermediate host to facilitate its passage to its feline definitive host. This is accomplished by a reduction of the aversive response that rodents show towards cat odors, which likely increases the predation risk. Females on average show similar changes as males. However, behaviors that relate to aversion and attraction are usually strongly influenced by the estrus cycle. In this study, we replicated behavioral effects of T. gondii in female rats, as well as expanded it to two novel behavioral paradigms. We also characterized the role of the estrus cycle in the behavioral effects of T. gondii on female rats. Uninfected females preferred to spend more time in proximity to rabbit rather than bobcat urine, and in a dark chamber rather than a lit chamber. Infected females lost both of these preferences, and also spent more time investigating social novelty (foreign bedding in their environment). Taken together, these data suggest that infection makes females less risk averse and more exploratory. Furthermore, this effect was influenced by the estrus cycle. Uninfected rats preferred rabbit urine to bobcat urine throughout the cycle except at estrus and metestrus. In contrast, infected rats lost this preference at every stage of the cycle except estrus. Commensurate with the possibility that this was a hormone-dependent effect, infected rats had elevated levels of circulating progesterone, a known anxiolytic. Copyright © 2014 Elsevier Inc. All rights reserved.

Toxoplasma gondii influences aversive behaviors of female rats in an estrus cycle dependent manner

PubMed Central

Golcu, Doruk; Gebre, Rahiwa Z.; Sapolsky, Robert M.

2016-01-01

The protozoan Toxoplasma gondii (T. gondii) manipulates the behavior of its rodent intermediate host to facilitate its passage to its feline definitive host. This is accomplished by a reduction of the aversive response that rodents show towards cat odors, which likely increases the predation risk. Females on average show similar changes as males. However, behaviors that relate to aversion and attraction are usually strongly influenced by the estrus cycle. In this study, we replicated behavioral effects of T. gondii in female rats, as well as expanded it to two novel behavioral paradigms. We also characterized the role of the estrus cycle in the behavioral effects of T. gondii on female rats. Uninfected females preferred to spend more time in proximity to rabbit rather than bobcat urine, and in a dark chamber rather than a lit chamber. Infected females lost both of these preferences, and also spent more time investigating social novelty (foreign bedding in their environment). Taken together, these data suggest that infection makes females less risk averse and more exploratory. Furthermore, this effect was influenced by the estrus cycle. Uninfected rats preferred rabbit urine to bobcat urine throughout the cycle except at estrus and metestrus. In contrast, infected rats lost this preference at every stage of the cycle except estrus. Commensurate with the possibility that this was a hormone-dependent effect, infected rats had elevated levels of circulating progesterone, a known anxiolytic. PMID:24907696

Induction of Apoptosis and Antiproliferative Activity of Naringenin in Human Epidermoid Carcinoma Cell through ROS Generation and Cell Cycle Arrest

PubMed Central

Jafri, Asif; Ahmad, Sheeba; Afzal, Mohammad; Arshad, Md

2014-01-01

A natural predominant flavanone naringenin, especially abundant in citrus fruits, has a wide range of pharmacological activities. The search for antiproliferative agents that reduce skin carcinoma is a task of great importance. The objective of this study was to analyze the anti-proliferative and apoptotic mechanism of naringenin using MTT assay, DNA fragmentation, nuclear condensation, change in mitochondrial membrane potential, cell cycle kinetics and caspase-3 as biomarkers and to investigate the ability to induce reactive oxygen species (ROS) initiating apoptotic cascade in human epidermoid carcinoma A431 cells. Results showed that naringenin exposure significantly reduced the cell viability of A431 cells (p<0.01) with a concomitant increase in nuclear condensation and DNA fragmentation in a dose dependent manner. The intracellular ROS generation assay showed statistically significant (p<0.001) dose-related increment in ROS production for naringenin. It also caused naringenin-mediated epidermoid carcinoma apoptosis by inducing mitochondrial depolarization. Cell cycle study showed that naringenin induced cell cycle arrest in G0/G1 phase of cell cycle and caspase-3 analysis revealed a dose dependent increment in caspase-3 activity which led to cell apoptosis. This study confirms the efficacy of naringenin that lead to cell death in epidermoid carcinoma cells via inducing ROS generation, mitochondrial depolarization, nuclear condensation, DNA fragmentation, cell cycle arrest in G0/G1 phase and caspase-3 activation. PMID:25330158

Induction of S phase cell arrest and caspase activation by polysaccharide peptide isolated from Coriolus versicolor enhanced the cell cycle dependent activity and apoptotic cell death of doxorubicin and etoposide, but not cytarabine in HL-60 cells.

PubMed

Hui, Kenrie Pui-Yan; Sit, Wai-Hung; Wan, Jennifer Man-Fan

2005-07-01

Activation of the cell death program (apoptosis) is a strategy for the treatment of human cancer, and unfortunately a large number of drugs identified as cell cycle-specific agents for killing cancer cells are also toxic to normal cells. The present study demonstrates that the polysaccharide peptide (PSP) extracted from the Chinese medicinal mushroom, Coriolus versicolor, used in combination therapy in China, has the ability to lower the cytotoxicity of certain anti-leukemic drugs via their interaction with cell cycle-dependent and apoptotic pathways. Flow cytometry analysis demonstrated that pre-treatment of PSP (25-100 microg/ml) dose-dependently enhanced the cell cycle perturbation and apoptotic activity of doxorubicin (Doxo) and etoposide (VP-16), but not cytarabine (Ara-C) in human promyelocytic leukemia HL-60 cells. The antagonistic result from combined treatment with Ara-C and PSP may be caused by the removal of HL-60 cells in the G1-S boundary by PSP before exposure to Ara-C. A negative correlation between the increase in apoptotic cell population (pre-G1 peak) with the S-phase cell population expression (R2=0.998), the expression of cyclin E expression (R2=0.872) and caspase 3 activity (R2=0.997) suggests that PSP enhanced the apoptotic machinery of Doxo and VP-16 in a cell cycle-dependent manner and is mediated, at least in part, by the PSP-mediated modulation of the regulatory checkpoint cyclin E and caspase 3. This study is the first to describe the cell cycle mechanistic action of PSP and its interaction with other anticancer agents. Our data support the potential development of PSP as an adjuvant for leukemia treatment, but also imply the importance of understanding its interaction with individual anticancer agents.

A pH-Regulated Quality Control Cycle for Surveillance of Secretory Protein Assembly

PubMed Central

Vavassori, Stefano; Cortini, Margherita; Masui, Shoji; Sannino, Sara; Anelli, Tiziana; Caserta, Imma R.; Fagioli, Claudio; Mossuto, Maria F.; Fornili, Arianna; van Anken, Eelco; Degano, Massimo; Inaba, Kenji; Sitia, Roberto

2013-01-01

Summary To warrant the quality of the secretory proteome, stringent control systems operate at the endoplasmic reticulum (ER)-Golgi interface, preventing the release of nonnative products. Incompletely assembled oligomeric proteins that are deemed correctly folded must rely on additional quality control mechanisms dedicated to proper assembly. Here we unveil how ERp44 cycles between cisGolgi and ER in a pH-regulated manner, patrolling assembly of disulfide-linked oligomers such as IgM and adiponectin. At neutral, ER-equivalent pH, the ERp44 carboxy-terminal tail occludes the substrate-binding site. At the lower pH of the cisGolgi, conformational rearrangements of this peptide, likely involving protonation of ERp44’s active cysteine, simultaneously unmask the substrate binding site and −RDEL motif, allowing capture of orphan secretory protein subunits and ER retrieval via KDEL receptors. The ERp44 assembly control cycle couples secretion fidelity and efficiency downstream of the calnexin/calreticulin and BiP-dependent quality control cycles. PMID:23685074

DNA damage-induced nuclear translocation of Apaf-1 is mediated by nucleoporin Nup107

PubMed Central

Jagot-Lacoussiere, Léonard; Faye, Audrey; Bruzzoni-Giovanelli, Heriberto; Villoutreix, Bruno O; Rain, Jean-Christophe; Poyet, Jean-Luc

2015-01-01

Beside its central role in the mitochondria-dependent cell death pathway, the apoptotic protease activating factor 1 (Apaf-1) is involved in the DNA damage response through cell-cycle arrest induced by genotoxic stress. This non-apoptotic function requires a nuclear translocation of Apaf-1 during the G1-to-S transition. However, the mechanisms that trigger the nuclear accumulation of Apaf-1 upon DNA damage remain to be investigated. Here we show that the main 4 isoforms of Apaf-1 can undergo nuclear translocation and restore Apaf-1 deficient MEFs cell cycle arrest in the S phase following genotoxic stress through activation of Chk-1. Interestingly, DNA damage-dependent nuclear accumulation of Apaf-1 occurs independently of p53 and the retinoblastoma (pRb) pathway. We demonstrated that Apaf-1 associates with the nucleoporin Nup107 and this association is necessary for Apaf-1 nuclear import. The CED-4 domain of Apaf-1 directly binds to the central domain of Nup107 in an ATR-regulated, phosphorylation-dependent manner. Interestingly, expression of the Apaf-1-interacting domain of Nup107 interfered with Apaf-1 nuclear translocation upon genotoxic stress, resulting in a marked reduction of Chk-1 activation and cell cycle arrest. Thus, our results confirm the crucial role of Apaf-1 nuclear relocalization in mediating cell-cycle arrest induced by genotoxic stress and implicate Nup107 as a critical regulator of the DNA damage-induced intra-S phase checkpoint response. PMID:25695197

A balance of FGF and BMP signals regulates cell cycle exit and Equarin expression in lens cells

PubMed Central

Jarrin, Miguel; Pandit, Tanushree; Gunhaga, Lena

2012-01-01

In embryonic and adult lenses, a balance of cell proliferation, cell cycle exit, and differentiation is necessary to maintain physical function. The molecular mechanisms regulating the transition of proliferating lens epithelial cells to differentiated primary lens fiber cells are poorly characterized. To investigate this question, we used gain- and loss-of-function analyses to modulate fibroblast growth factor (FGF) and/or bone morphogenetic protein (BMP) signals in chick lens/retina explants. Here we show that FGF activity plays a key role for proliferation independent of BMP signals. Moreover, a balance of FGF and BMP signals regulates cell cycle exit and the expression of Ccdc80 (also called Equarin), which is expressed at sites where differentiation of lens fiber cells occurs. BMP activity promotes cell cycle exit and induces Equarin expression in an FGF-dependent manner. In contrast, FGF activity is required but not sufficient to induce cell cycle exit or Equarin expression. Furthermore, our results show that in the absence of BMP activity, lens cells have increased cell cycle length or are arrested in the cell cycle, which leads to decreased cell cycle exit. Taken together, these findings suggest that proliferation, cell cycle exit, and early differentiation of primary lens fiber cells are regulated by counterbalancing BMP and FGF signals. PMID:22718906

Glucose and lactate are equally effective in energizing activity-dependent synaptic vesicle turnover in purified cortical neurons.

PubMed

Morgenthaler, F D; Kraftsik, R; Catsicas, S; Magistretti, P J; Chatton, J-Y

2006-08-11

This study examines the role of glucose and lactate as energy substrates to sustain synaptic vesicle cycling. Synaptic vesicle turnover was assessed in a quantitative manner by fluorescence microscopy in primary cultures of mouse cortical neurons. An electrode-equipped perfusion chamber was used to stimulate cells both by electrical field and potassium depolarization during image acquisition. An image analysis procedure was elaborated to select in an unbiased manner synaptic boutons loaded with the fluorescent dye N-(3-triethylammoniumpropyl)-4-(4-(dibutylamino)styryl)pyridinium dibromide (FM1-43). Whereas a minority of the sites fully released their dye content following electrical stimulation, others needed subsequent K(+) depolarization to achieve full release. This functional heterogeneity was not significantly altered by the nature of metabolic substrates. Repetitive stimulation sequences of FM1-43 uptake and release were then performed in the absence of any metabolic substrate and showed that the number of active sites dramatically decreased after the first cycle of loading/unloading. The presence of 1 mM glucose or lactate was sufficient to sustain synaptic vesicle cycling under these conditions. Moreover, both substrates were equivalent for recovery of function after a phase of decreased metabolic substrate availability. Thus, lactate appears to be equivalent to glucose for sustaining synaptic vesicle turnover in cultured cortical neurons during activity.

Seleno-short-chain chitosan induces apoptosis in human non-small-cell lung cancer A549 cells through ROS-mediated mitochondrial pathway.

PubMed

Zhao, Yana; Zhang, Shaojing; Wang, Pengfei; Fu, Shengnan; Wu, Di; Liu, Anjun

2017-12-01

Seleno-short-chain chitosan (SSCC) is a synthesized chitosan derivative. In this study, antitumor activity and underlying mechanism of SSCC on human non-small-cell lung cancer A549 cells were investigated in vitro. The MTT assay showed that SSCC could inhibit cell viability in a dose- and time-dependent manner, and 200 μg/ml SSCC exhibited significantly toxic effects on A549 cells. The cell cycle assay showed that SSCC triggered S phase cell cycle arrest in a dose- and time-dependent manner, which was related to a downregulation of S phase associated cyclin A. The DAPI staining and Annexin V-FITC/PI double staining identified that the SSCC could induce A549 cells apoptosis. Further studies found that SSCC led to the generation of reactive oxygen species (ROS) and the disruption of mitochondrial membrane potential (MMP) by DCFH-DA and Rhodamin 123 staining, respectively. Meanwhile, free radical scavengers N-acetyl-L-cysteine (NAC) pretreatment confirmed that SSCC-induced A549 cells apoptosis was associated with ROS generation. Furthermore, real-time PCR and western blot assay showed that SSCC up-regulated Bax and down-regulated Bcl-2, subsequently incited the release of cytochrome c from mitochondria to cytoplasm, activated the increase of cleaved-caspase 3 and finally induced A549 cells apoptosis in vitro. In general, the present study demonstrated that SSCC induced A549 cells apoptosis via ROS-mediated mitochondrial apoptosis pathway.

Dimethylaminoethanol affects the viability of human cultured fibroblasts.

PubMed

Gragnani, Alfredo; Giannoccaro, Fabiana Bocci; Sobral, Christiane S; Moraes, A A F; França, Jeronimo P; Ferreira, A T; Ferreira, Lydia Masako

2007-01-01

In clinical practice, dimethylaminoethanol (DMAE) has been used in the fight against wrinkles and flaccidity in the cervicofacial region. The firming action of DMAE is explained by the fact that its molecule, considered to be a precursor of acetylcholine, alters muscle contraction. However, no experimental studies have confirmed this theory. Because the actual mechanism of DMAE action was not defined and there were no references in the literature regarding its direct action on fibroblasts, this study was performed to evaluate the direct action of DMAE on cultured human fibroblasts. Human fibroblasts obtained from discarded fragments of total skin from patients undergoing plastic or reconstructive surgical procedures performed within the Plastic Surgery Division at the Federal University of São Paulo were used for this study. The explant technique was used. The culture medium was supplemented with different concentrations of DMAE on the fourth cell passage, and the cell proliferation rate, cytosolic calcium levels, and cell cycle were evaluated. Statistical analysis was performed using analysis of variance (ANOVA) followed by a Newman-Keuls test for multiple comparisons. A decrease in fibroblast proliferation was associated with an increase in DMAE concentration. A longer treatment time with trypsin was required for the groups treated with DMAE in a dose-dependent manner. In the presence of DMAE, cytosolic calcium increased in a dose-dependent manner. Apoptosis also increased in groups treated with DMAE. Dimethylaminoethanol reduced the proliferation of fibroblasts, increased cytosolic calcium, and changed the cell cycle, causing an increase in apoptosis in cultured human fibroblasts.

Anti-leukemic effect of a synthetic compound, (±) trans-dihydronarciclasine (HYU-01) via cell-cycle arrest and apoptosis in acute myeloid leukemia.

PubMed

Kim, Seo Ju; Park, Hyun Ki; Kim, Ju Young; Yoon, Jin Sun; Kim, Eun Shil; Cho, Cheon-Gyu; Kim, Byoung Kook; Park, Byeong Bae; Lee, Young Yiul

2012-10-01

(±) trans-Dihydronarciclasine, isolated from Chinese medicinal plant Zephyranthes candida, has been shown to possess quite potent anti-tumoral effect against selected human cancer cell lines. However, little is known about the anti-tumoral effect of (±) trans-dihydronarciclasine in acute myeloid leukemia (AML). This study was performed to investigate the effect of a novel synthetic (±) trans-dihydronarciclasine (code name; HYU-01) in AML. The HYU-01 inhibited the proliferation of various AML cell lines including HL-60 as well as primary leukemic blasts in a dose-dependent manner. To investigate the mechanism of the anti-proliferative effect of HYU-01, cell-cycle analysis was attempted in HL-60 cells, resulting in G1 arrest. The expression levels of CDK2, CDK4, CDK6, cyclin E, and cyclin A were decreased in a time-dependent manner. In addition, HYU-01 up-regulated the expression of the p27, and markedly enhanced the binding of p27 with CDK2, 4, and 6, ultimately resulting in the decrease of their kinase activities. Furthermore, HYU-01 induced the apoptosis through the induction of proapoptotic molecules and reduction of antiapoptotic molecules in association with the activation of caspase-3, -8, and -9. These results suggest that HYU-01 may inhibit the proliferation of HL-60 cells, via apoptosis, as well as G1 block in association with the induction of p27. © 2012 The Authors APMIS © 2012 APMIS.

Loss of Proliferation and Antigen Presentation Activity following Internalization of Polydispersed Carbon Nanotubes by Primary Lung Epithelial Cells

PubMed Central

Kumari, Mandavi; Sachar, Sumedha; Saxena, Rajiv K.

2012-01-01

Interactions between poly-dispersed acid functionalized single walled carbon nanotubes (AF-SWCNTs) and primary lung epithelial (PLE) cells were studied. Peritoneal macrophages (PMs, known phagocytic cells) were used as positive controls in this study. Recovery of live cells from cultures of PLE cells and PMs was significantly reduced in the presence of AF-SWCNTs, in a time and dose dependent manner. Both PLE cells as well as PMs could take up fluorescence tagged AF-SWCNTs in a time dependent manner and this uptake was significantly blocked by cytochalasin D, an agent that blocks the activity of acto-myosin fibers and therefore the phagocytic activity of cells. Confocal microscopic studies confirmed that AF-SWCNTs were internalized by both PLE cells and PMs. Intra-trachially instilled AF-SWCNTs could also be taken up by lung epithelial cells as well as alveolar macrophages. Freshly isolated PLE cells had significant cell division activity and cell cycling studies indicated that treatment with AF-SWCNTs resulted in a marked reduction in S-phase of the cell cycle. In a previously standardized system to study BCG antigen presentation by PLE cells and PMs to sensitized T helper cells, AF-SWCNTs could significantly lower the antigen presentation ability of both cell types. These results show that mouse primary lung epithelial cells can efficiently internalize AF-SWCNTs and the uptake of nanotubes interfered with biological functions of PLE cells including their ability to present BCG antigens to sensitized T helper cells. PMID:22384094

Context-dependent effects of rimonabant on ethanol-induced conditioned place preference in female mice.

PubMed

Silva, Aline A F; Barbosa-Souza, Evelyn; Confessor-Carvalho, Cassio; Silva, Raiany R R; De Brito, Ana Carolina L; Cata-Preta, Elisangela G; Silva Oliveira, Thaynara; Berro, Lais F; Oliveira-Lima, Alexandre J; Marinho, Eduardo A V

2017-10-01

The CB1 receptor antagonist rimonabant has been previously found to prevent behavioral effects of drugs of abuse in a context-dependent manner, suggesting an important role of endocannabinoid signaling in drug-induced environmental conditioning. The aim of the present study was to evaluate the effects of rimonabant on ethanol-induced conditioned place preference (CPP) in female mice. Animals were conditioned with saline or ethanol (1.8g/kg) during 8 sessions, and subsequently treated with either saline or rimonabant (1 or 10mg/kg) in the CPP environment previously associated with saline (unpaired) or ethanol (paired) for 6 consecutive days. Animals were then challenged with ethanol (1.8g/kg) in the ethanol-paired environment and ethanol-induced CPP was quantified on the following day. While treatment with 1mg/kg rimonabant in the saline-associated environment had no effects on the subsequent expression of ethanol-induced CPP, it blocked the expression of CPP to ethanol when paired to the ethanol-associated environment. When given in the ethanol-paired environment, 10mg/kg rimonabant induced aversion to the ethanol-associated environment. The same aversion effect was observed for 10mg/kg rimonabant when given in the saline-associated environment, thereby potentiating the expression of ethanol-induced CPP. Importantly, rimonabant did not induce CPP or conditioned place aversion on its own. Controlling for the estrous cycle phase showed no influences of hormonal cycle on the development and expression of ethanol-induced CPP. Our data suggest that rimonabant reduces the rewarding properties of ethanol by abolishing drug-environment conditioning in the CPP paradigm in a context-dependent manner. Copyright © 2017 Elsevier B.V. All rights reserved.

In vitro assessment of anti-proliferative effect induced by α-mangostin from Cratoxylum arborescens on HeLa cells

PubMed Central

El habbash, Aisha I.; Ibrahim, Mohamed Yousif; Yahayu, Maizatulakmal; Omer, Fatima Abd Elmutaal; Abd Rahman, Mashitoh; Nordin, Noraziah; Lian, Gwendoline Ee Cheng

2017-01-01

Natural medicinal products possess diverse chemical structures and have been an essential source for drug discovery. Therefore, in this study, α-mangostin (AM) is a plant-derived compound was investigated for the apoptotic effect on human cervical cancer cells (HeLa). The cytotoxic effects of AM on the viability of HeLa and human normal ovarian cell line (SV40) were evaluated by using MTT assay. Results showed that AM inhibited HeLa cells viability at concentration- and time-dependent manner with IC50 value of 24.53 ± 1.48 µM at 24 h. The apoptogenic effects of AM on HeLa were assessed using fluorescence microscopy analysis. The effect of AM on cell proliferation was also studied through clonogenic assay. ROS production evaluation, flow cytometry (cell cycle) analysis, caspases 3/7, 8, and 9 assessment and multiple cytotoxicity assays were conducted to determine the mechanism of cell apoptosis. This was associated with G2/M phase cell cycle arrest and elevation in ROS production. AM induced mitochondrial apoptosis which was confirmed based on the significant increase in the levels of caspases 3/7 and 9 in a dose-dependent manner. Furthermore, the MMP disruption and increased cell permeability, concurrent with cytochrome c release from the mitochondria to the cytosol provided evidence that AM can induce apoptosis via mitochondrial-dependent pathway. AM exerted a remarkable antitumor effect and induced characteristic apoptogenic morphological changes on HeLa cells, which indicates the occurrence of cell death. This study reveals that AM could be a potential antitumor compound on cervical cancer in vitro and can be considered for further cervical cancer preclinical and in vivo testing. PMID:28740747

Small-scale geochemical cycles and the distribution of uranium in central and north Florida organic deposits

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bond, P.A.

1993-03-01

The global geochemical cycle for an element tracks its path from its various sources to its sinks via processes of weathering and transportation. The cycle may then be quantified in a necessarily approximate manner. The geochemical cycle (thus quantified) reveals constraints (known and unknown) on an element's behavior imposed by the various processes which act on it. In the context of a global geochemical cycle, a continent becomes essentially a source term. If, however, an element's behavior is examined in a local or regional context, sources and their related sinks may be identified. This suggests that small-scale geochemical cycles maymore » be superimposed on global geochemical cycles. Definition of such sub-cycles may clarify the distribution of an element in the earth's near-surface environment. In Florida, phosphate minerals of the Hawthorn Group act as a widely distributed source of uranium. Uranium is transported by surface- and ground-waters. Florida is the site of extensive wetlands and peatlands. The organic matter associated with these deposits adsorbs uranium and may act as a local sink depending on its hydrogeologic setting. This work examines the role of organic matter in the distribution of uranium in the surface and shallow subsurface environments of central and north Florida.« less

Genistein induced anticancer effects on pancreatic cancer cell lines involves mitochondrial apoptosis, G0/G1cell cycle arrest and regulation of STAT3 signalling pathway.

PubMed

Bi, Yi-Liang; Min, Min; Shen, Wei; Liu, Yan

2018-01-15

Genistein is a natural flavonoid that has been reported to exhibit anticancer effects against different types of cancers which include, but are not limited to, breast and oral squamous cell carcinoma. The present study was designed to evaluate the anticancer effects of the natural flavonoid genistein against pancreatic cancer cell lines and to explore the underlying mechanism. Antiproliferative activity was investigated by MTT assay. Apoptosis was detected by DAPI and annexin V/PI staining. DNA damage was assessed by comet assay. Reactive oxygen species (ROS) and reduction of mitochondrial membrane potential (MMP) were determined by flow cytometry. Cell migration was examined by wound healing assay. Protien expressions were determined by western blotting. Antiproliferative assay revealed that genistein reduced the cell viability of pancreatic cancer cells in a dose dependent manner with an IC 50 of 20 and 25 µM against Mia-PaCa2 and PANC-1 cancer cell lines respectively. However, its antiproliferative effects were less pronounced against non-cancerous pancreatic ductal epithelial cell line (H6C7) as evident from the IC 50 of 120 µM. Genistein induced significant morphological changes in pancreatic cancer cells and triggered cell cycle arrest in G 0 /G 1 phase. DAPI staining and flow cytometric analysis revealed that genistein induced apoptosis in a dose dependent manner through generation of substantial amounts of ROS and reduction of MMP. However, treatment of the pancreatic cancer with genistein and ascorbic acid could abrogate the effects of genistein on cell viability. Protien expression analysis revealed that genistein upregulated cytosolic cytochrome c, Bax, cleaved Caspase-3 and cleaved caspase-9 expressions with concomitant downregulation of Bcl-2 expression. Moreover, genistein inhibited the phosphorylation of signal transducer and activator of transcription STAT3 proteins and downregulated the expression of survivin, cyclin D1 and ALDH1A1 in Mia-PaCa2 cells in a dose dependent manner. Interestingly, genistein could inhibit the cell migration potential of the Mia-PaCa2 cells which was further associated with the downregulation of metalloproteinases (MPP-2 and MPP-9). Taken together, we propose that genistein exerts anticancer activity in pancreatic cancer cells through induction of ROS mediated mitochondrial apoptosis, cell cycle arrest and regulation of STAT3 and may therefore prove beneficial in the management of pancreatic cancers cancer. Copyright © 2017 Elsevier GmbH. All rights reserved.

Abnormal mitosis triggers p53-dependent cell cycle arrest in human tetraploid cells.

PubMed

Kuffer, Christian; Kuznetsova, Anastasia Yurievna; Storchová, Zuzana

2013-08-01

Erroneously arising tetraploid mammalian cells are chromosomally instable and may facilitate cell transformation. An increasing body of evidence shows that the propagation of mammalian tetraploid cells is limited by a p53-dependent arrest. The trigger of this arrest has not been identified so far. Here we show by live cell imaging of tetraploid cells generated by an induced cytokinesis failure that most tetraploids arrest and die in a p53-dependent manner after the first tetraploid mitosis. Furthermore, we found that the main trigger is a mitotic defect, in particular, chromosome missegregation during bipolar mitosis or spindle multipolarity. Both a transient multipolar spindle followed by efficient clustering in anaphase as well as a multipolar spindle followed by multipolar mitosis inhibited subsequent proliferation to a similar degree. We found that the tetraploid cells did not accumulate double-strand breaks that could cause the cell cycle arrest after tetraploid mitosis. In contrast, tetraploid cells showed increased levels of oxidative DNA damage coinciding with the p53 activation. To further elucidate the pathways involved in the proliferation control of tetraploid cells, we knocked down specific kinases that had been previously linked to the cell cycle arrest and p53 phosphorylation. Our results suggest that the checkpoint kinase ATM phosphorylates p53 in tetraploid cells after abnormal mitosis and thus contributes to proliferation control of human aberrantly arising tetraploids.

Cell-cycle-dependent three-dimensional redistribution of nuclear proteins, P 120, pKi-67, and SC 35 splicing factor, in the presence of the topoisomerase I inhibitor camptothecin.

PubMed

Elias, Emmanuel; Lalun, Nathalie; Lorenzato, Marianne; Blache, Laurent; Chelidze, Pavel; O'Donohue, Marie-Françoise; Ploton, Dominique; Bobichon, Hélène

2003-11-15

Topoisomerase I (Topo I) is mostly known for its role in DNA relaxation, which is required for duplication and transcription. Topo I acts as a protein kinase mainly directed to the mRNA splicing factor SC35. Camptothecin is one of the specific Topo I inhibitors and is effective on the two functions of the enzyme. In this study we demonstrated that treatment of KB cells with camptothecin for only 30 min induced the 3D reorganization and redistribution of three proteins involved in the nucleus machinery, P 120, pKi-67, and SC 35, and this occurred in a cell cycle-dependent manner. Our data were obtained from confocal microscopic studies after immunolabeling, 3D reconstruction, and measurement of the nuclear components volumes. In the presence of camptothecin, P 120, which occupied the nucleolar volume, lost its reticulation and pKi-67 was redistributed within the nucleoplasm and even into the cytoplasm. Finally, for SC 35 the fusion of its dots into bigger volumes was observed specifically during the G1 phase. Variations of volumes were also observed for the nucleolus and for the nucleus. These results pointed out that, depending on the cell cycle phase, Topo I functions were selective toward the three different proteins.

PRMT5 is essential for the eIF4E-mediated 5′-cap dependent translation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lim, Ji-Hong; Lee, Yoon-Mi; Lee, Gibok

2014-10-03

Highlights: • PRMT5 participates in syntheses of HIF-1α, c-Myc and cyclin D1 proteins. • PRMT5 promotes the 5′-cap dependent translation. • PRMT5 is required for eIF4E binding to mRNA 5′-cap. • PRMT5 is essential for eIF4E-dependent cell proliferation. - Abstract: It is becoming clear that PRMT5 plays essential roles in cell cycle progression, survival, and responses to external stresses. However, the precise mechanisms underlying such roles of PRMT5 have not been clearly understood. Previously, we have demonstrated that PRMT5 participates in cellular adaptation to hypoxia by ensuring 5′-cap dependent translation of HIF-1α. Given that c-Myc and cyclin D1 expressions aremore » also tightly regulated in 5′-cap dependent manner, we here tested the possibility that PRMT5 promotes cell proliferation by increasing de novo syntheses of the oncoproteins. c-Myc and cyclin D1 were found to be noticeably downregulated by PRMT5 knock-down. A RNA immunoprecipitation analysis, which can identify RNA–protein interactions, showed that PRMT5 is required for the interaction among eIF4E and 5′-UTRs of HIF-1α, c-Myc and cyclin D1 mRNAs. In addition, PRMT5 knock-down inhibited cell proliferation by inducing cell cycle arrest at the G1 phase. More importantly, ectopic expression of eIF4E significantly rescued the cell cycle progression and cell proliferation even in PRMT5-deficeint condition. Based on these results, we propose that PRMT5 determines cell fate by regulating 5′-cap dependent translation of proteins essential for proliferation and survival.« less

Cytotoxic and apoptotic activities of black widow spiderling extract against HeLa cells

PubMed Central

Peng, Xiaozhen; Dai, Zhipan; Lei, Qian; Liang, Long; Yan, Shuai; Wang, Xianchun

2017-01-01

Black widow spiders contain toxic components not only in the venom glands but also in other parts of the spider body, including the legs and abdomen. Additionally, both the eggs and newborn spiderlings of the black widow spider contain venom. It is important to investigate their potential effects on cancer cells. In the present study, the effects of newborn black widow spiderling extract on human HeLa cells were evaluated in vitro. When applied at different concentrations, the total extract decreased HeLa cell viability in a dose-dependent manner, with an IC50 value of 158 µg/ml. Flow cytometry indicated that treatment of HeLa cells with the total extract of the spiderlings induced apoptosis in HeLa cells in a dose-dependent manner and led to cell cycle arrest in the S-phase. Additionally, application of the total extract at different concentrations increased apoptosis-related caspase 3 activity in a dose-dependent manner. HeLa cells treated with the total extract appeared to be morphologically changed, exhibiting membrane blebbing, nuclear fragmentation and condensation of chromatin. Further separation and activity screening demonstrated that the cytotoxic and apoptotic activities of the total extract were attributable mainly to its high molecular mass proteins, one of which was purified and characterized to determine its anti-tumor activities on HeLa cells. The results of the present study therefore have expanded understanding regarding the effect of spider toxins on cancer cells and suggested that components of black widow spiderlings may be developed as a promising novel agent to treat cancer. PMID:28587399

Cytotoxic and apoptotic activities of black widow spiderling extract against HeLa cells.

PubMed

Peng, Xiaozhen; Dai, Zhipan; Lei, Qian; Liang, Long; Yan, Shuai; Wang, Xianchun

2017-06-01

Black widow spiders contain toxic components not only in the venom glands but also in other parts of the spider body, including the legs and abdomen. Additionally, both the eggs and newborn spiderlings of the black widow spider contain venom. It is important to investigate their potential effects on cancer cells. In the present study, the effects of newborn black widow spiderling extract on human HeLa cells were evaluated in vitro . When applied at different concentrations, the total extract decreased HeLa cell viability in a dose-dependent manner, with an IC 50 value of 158 µg/ml. Flow cytometry indicated that treatment of HeLa cells with the total extract of the spiderlings induced apoptosis in HeLa cells in a dose-dependent manner and led to cell cycle arrest in the S-phase. Additionally, application of the total extract at different concentrations increased apoptosis-related caspase 3 activity in a dose-dependent manner. HeLa cells treated with the total extract appeared to be morphologically changed, exhibiting membrane blebbing, nuclear fragmentation and condensation of chromatin. Further separation and activity screening demonstrated that the cytotoxic and apoptotic activities of the total extract were attributable mainly to its high molecular mass proteins, one of which was purified and characterized to determine its anti-tumor activities on HeLa cells. The results of the present study therefore have expanded understanding regarding the effect of spider toxins on cancer cells and suggested that components of black widow spiderlings may be developed as a promising novel agent to treat cancer.

Citric acid induces cell-cycle arrest and apoptosis of human immortalized keratinocyte cell line (HaCaT) via caspase- and mitochondrial-dependent signaling pathways.

PubMed

Ying, Tsung-Ho; Chen, Chia-Wei; Hsiao, Yu-Ping; Hung, Sung-Jen; Chung, Jing-Gung; Yang, Jen-Hung

2013-10-01

Citric acid is an alpha-hydroxyacid (AHA) widely used in cosmetic dermatology and skincare products. However, there is concern regarding its safety for the skin. In this study, we investigated the cytotoxic effects of citric acid on the human keratinocyte cell line HaCaT. HaCaT cells were treated with citric acid at 2.5-12.5 mM for different time periods. Cell-cycle arrest and apoptosis were investigated by 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) staining, flow cytometry, western blot and confocal microscopy. Citric acid not only inhibited proliferation of HaCaT cells in a dose-dependent manner, but also induced apoptosis and cell cycle-arrest at the G2/M phase (before 24 h) and S phase (after 24 h). Citric acid increased the level of Bcl-2-associated X protein (BAX) and reduced the levels of B-cell lymphoma-2 (BCL-2), B-cell lymphoma-extra large (BCL-XL) and activated caspase-9 and caspase-3, which subsequently induced apoptosis via caspase-dependent and caspase-independent pathways. Citric acid also activated death receptors and increased the levels of caspase-8, activated BH3 interacting-domain death agonist (BID) protein, Apoptosis-inducing factor (AIF), and Endonuclease G (EndoG). Therefore, citric acid induces apoptosis through the mitochondrial pathway in the human keratinocyte cell line HaCaT. The study results suggest that citric acid is cytotoxic to HaCaT cells via induction of apoptosis and cell-cycle arrest in vitro.

The steady-state level and stability of TLS polymerase eta are cell cycle dependent in the yeast S. cerevisiae.

PubMed

Plachta, Michal; Halas, Agnieszka; McIntyre, Justyna; Sledziewska-Gojska, Ewa

2015-05-01

Polymerase eta (Pol eta) is a ubiquitous translesion DNA polymerase that is capable of bypassing UV-induced pyrimidine dimers in an error-free manner. However, this specialized polymerase is error prone when synthesizing through an undamaged DNA template. In Saccharomyces cerevisiae, both depletion and overproduction of Pol eta result in mutator phenotypes. Therefore, regulation of the cellular abundance of this enzyme is of particular interest. However, based on the investigation of variously tagged forms of Pol eta, mutually contradictory conclusions have been reached regarding the stability of this polymerase in yeast. Here, we optimized a protocol for the detection of untagged yeast Pol eta and established that the half-life of the native enzyme is 80 ± 14 min in asynchronously growing cultures. Experiments with synchronized cells indicated that the cellular abundance of this translesion polymerase changes throughout the cell cycle. Accordingly, we show that the stability of Pol eta, but not its mRNA level, is cell cycle stage dependent. The half-life of the polymerase is more than fourfold shorter in G1-arrested cells than in those at G2/M. Our results, in concert with previous data for Rev1, indicate that cell cycle regulation is a general property of Y family TLS polymerases in S. cerevisiae. Copyright © 2015 Elsevier B.V. All rights reserved.

Cibotium barometz polysaccharides stimulate chondrocyte proliferation in vitro by promoting G1/S cell cycle transition

PubMed Central

Fu, Changlong; Zheng, Chunsong; Lin, Jie; Ye, Jinxia; Mei, Yangyang; Pan, Caibin; Wu, Guangwen; Li, Xihai; Ye, Hongzhi; Liu, Xianxiang

2017-01-01

Cibotium barometz polysaccharides (CBPS) are one of the most important bioactive components extracted from the Cibotium barometz plant, which belongs to the Dicksoniaceae family. It has been widely used for the treatment of orthopedic diseases in traditional Chinese medicine. However, the molecular mechanisms behind the therapeutic effects of CBPS remain to be clarified. In the present study, the concentration of CBPS was detected by phenol-vitriol colorimetry. Furthermore, the effects stimulated by CBPS on the viability and G1/S cell cycle transition in primary chondrocytes from Sprague-Dawley rats were investigated. A cell viability assay demonstrated that chondrocyte proliferation may be enhanced by CBPS in a dose- and time-dependent manner. The mechanism underlying the promotion of chondrocyte cell cycle was suggested to involve the stimulation of G1 to S phase transition. To further confirm the results, reverse transcription-quantitative polymerase chain reaction and western blot analyses were used to detect the expression of mRNA and protein levels of cyclin D1, cyclin-dependent kinase 4 and retinoblastoma protein. The results suggested that CBPS may stimulate chondrocyte proliferation via promoting G1/S cell cycle transition. Since osteoarthritis is characterized by deficient proliferation in chondrocytes, the present study indicates that CBPS may potentially serve as a novel method for the treatment of osteoarthritis. PMID:28358416

Intrahippocampal administration of anandamide increases REM sleep.

PubMed

Rueda-Orozco, Pavel Ernesto; Soria-Gómez, Edgar; Montes-Rodríguez, Corinne Jennifer; Pérez-Morales, Marcel; Prospéro-García, Oscar

2010-04-05

A nascent literature has postulated endocannabinoids (eCBs) as strong sleep-inducing lipids, particularly rapid-eye-movement sleep (REMs), nevertheless the exact mechanisms behind this effect remain to be determined. Anandamide and 2-arachidonyl glycerol, two of the most important eCBS, are synthesized in the hippocampus. This structure also expresses a high concentration of cannabinoid receptor 1 (CB1). Recent extensive literature supports eCBs as important regulators of hippocampal activity. It has also been shown that these molecules vary their expression on the hippocampus depending on the light-dark cycle. In this context we decided to analyze the effect of intrahippocampal administration of the eCB anandamide (ANA) on the sleep-waking cycle at two points of the light-dark cycle. Our data indicate that the administration of ANA directly into the hippocampus increases REMs in a dose dependent manner during the dark but not during the light phase of the cycle. The increase of REMs was blocked by the CB1 antagonist AM251. This effect was specific for the hippocampus since ANA administrations in the surrounding cortex did not elicit any change in REMs. These results support the idea of a direct relationship between hippocampal activity and sleep mechanisms by means of eCBs. The data presented here show, for the first time that eCBs administered into the hippocampus trigger REMs and support previous studies where chemical stimulation of limbic areas triggered sleep.

Dbf4-dependent kinase and the Rtt107 scaffold promote Mus81-Mms4 resolvase activation during mitosis.

PubMed

Princz, Lissa N; Wild, Philipp; Bittmann, Julia; Aguado, F Javier; Blanco, Miguel G; Matos, Joao; Pfander, Boris

2017-03-01

DNA repair by homologous recombination is under stringent cell cycle control. This includes the last step of the reaction, disentanglement of DNA joint molecules (JMs). Previous work has established that JM resolving nucleases are activated specifically at the onset of mitosis. In case of budding yeast Mus81-Mms4, this cell cycle stage-specific activation is known to depend on phosphorylation by CDK and Cdc5 kinases. Here, we show that a third cell cycle kinase, Cdc7-Dbf4 (DDK), targets Mus81-Mms4 in conjunction with Cdc5-both kinases bind to as well as phosphorylate Mus81-Mms4 in an interdependent manner. Moreover, DDK-mediated phosphorylation of Mms4 is strictly required for Mus81 activation in mitosis, establishing DDK as a novel regulator of homologous recombination. The scaffold protein Rtt107, which binds the Mus81-Mms4 complex, interacts with Cdc7 and thereby targets DDK and Cdc5 to the complex enabling full Mus81 activation. Therefore, Mus81 activation in mitosis involves at least three cell cycle kinases, CDK, Cdc5 and DDK Furthermore, tethering of the kinases in a stable complex with Mus81 is critical for efficient JM resolution. © 2017 The Authors. Published under the terms of the CC BY NC ND 4.0 license.

Tyrosine phosphorylation switching of a G protein.

PubMed

Li, Bo; Tunc-Ozdemir, Meral; Urano, Daisuke; Jia, Haiyan; Werth, Emily G; Mowrey, David D; Hicks, Leslie M; Dokholyan, Nikolay V; Torres, Matthew P; Jones, Alan M

2018-03-30

Heterotrimeric G protein complexes are molecular switches relaying extracellular signals sensed by G protein-coupled receptors (GPCRs) to downstream targets in the cytoplasm, which effect cellular responses. In the plant heterotrimeric GTPase cycle, GTP hydrolysis, rather than nucleotide exchange, is the rate-limiting reaction and is accelerated by a receptor-like regulator of G signaling (RGS) protein. We hypothesized that posttranslational modification of the Gα subunit in the G protein complex regulates the RGS-dependent GTPase cycle. Our structural analyses identified an invariant phosphorylated tyrosine residue (Tyr 166 in the Arabidopsis Gα subunit AtGPA1) located in the intramolecular domain interface where nucleotide binding and hydrolysis occur. We also identified a receptor-like kinase that phosphorylates AtGPA1 in a Tyr 166 -dependent manner. Discrete molecular dynamics simulations predicted that phosphorylated Tyr 166 forms a salt bridge in this interface and potentially affects the RGS protein-accelerated GTPase cycle. Using a Tyr 166 phosphomimetic substitution, we found that the cognate RGS protein binds more tightly to the GDP-bound Gα substrate, consequently reducing its ability to accelerate GTPase activity. In conclusion, we propose that phosphorylation of Tyr 166 in AtGPA1 changes the binding pattern with AtRGS1 and thereby attenuates the steady-state rate of the GTPase cycle. We coin this newly identified mechanism "substrate phosphoswitching." © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Circadian rhythms synchronize mitosis in Neurospora crassa.

PubMed

Hong, Christian I; Zámborszky, Judit; Baek, Mokryun; Labiscsak, Laszlo; Ju, Kyungsu; Lee, Hyeyeong; Larrondo, Luis F; Goity, Alejandra; Chong, Hin Siong; Belden, William J; Csikász-Nagy, Attila

2014-01-28

The cell cycle and the circadian clock communicate with each other, resulting in circadian-gated cell division cycles. Alterations in this network may lead to diseases such as cancer. Therefore, it is critical to identify molecular components that connect these two oscillators. However, molecular mechanisms between the clock and the cell cycle remain largely unknown. A model filamentous fungus, Neurospora crassa, is a multinucleate system used to elucidate molecular mechanisms of circadian rhythms, but not used to investigate the molecular coupling between these two oscillators. In this report, we show that a conserved coupling between the circadian clock and the cell cycle exists via serine/threonine protein kinase-29 (STK-29), the Neurospora homolog of mammalian WEE1 kinase. Based on this finding, we established a mathematical model that predicts circadian oscillations of cell cycle components and circadian clock-dependent synchronized nuclear divisions. We experimentally demonstrate that G1 and G2 cyclins, CLN-1 and CLB-1, respectively, oscillate in a circadian manner with bioluminescence reporters. The oscillations of clb-1 and stk-29 gene expression are abolished in a circadian arrhythmic frq(ko) mutant. Additionally, we show the light-induced phase shifts of a core circadian component, frq, as well as the gene expression of the cell cycle components clb-1 and stk-29, which may alter the timing of divisions. We then used a histone hH1-GFP reporter to observe nuclear divisions over time, and show that a large number of nuclear divisions occur in the evening. Our findings demonstrate the circadian clock-dependent molecular dynamics of cell cycle components that result in synchronized nuclear divisions in Neurospora.

Degradation of Hof1 by SCFGrr1 is important for actomyosin contraction during cytokinesis in yeast

PubMed Central

Blondel, Marc; Bach, Stéphane; Bamps, Sophie; Dobbelaere, Jeroen; Wiget, Philippe; Longaretti, Céline; Barral, Yves; Meijer, Laurent; Peter, Matthias

2005-01-01

SCF-type (SCF: Skp1–Cullin–F-box protein complex) E3 ligases regulate ubiquitin-dependent degradation of many cell cycle regulators, mainly at the G1/S transition. Here, we show that SCFGrr1 functions during cytokinesis by degrading the PCH protein Hof1. While Hof1 is required early in mitosis to assemble a functional actomyosin ring, it is specifically degraded late in mitosis and remains unstable during the entire G1 phase of the cell cycle. Degradation of Hof1 depends on its PEST motif and a functional 26S proteasome. Interestingly, degradation of Hof1 is independent of APCCdh1, but instead requires the SCFGrr1 E3 ligase. Grr1 is recruited to the mother–bud neck region after activation of the mitotic-exit network, and interacts with Hof1 in a PEST motif-dependent manner. Our results also show that downregulation of Hof1 at the end of mitosis is necessary to allow efficient contraction of the actomyosin ring and cell separation during cytokinesis. SCFGrr1-mediated degradation of Hof1 may thus represent a novel mechanism to couple exit from mitosis with initiation of cytokinesis. PMID:15775961

Novel Synthetic Mono-triazole Glycosides Induce G0/G1 Cell-cycle Arrest and Apoptosis in Cholangiocarcinoma Cells.

PubMed

Obchoei, Sumalee; Saeeng, Rungnapha; Wongkham, Chaisiri; Wongkham, Sopit

2016-11-01

The treatment of cholangiocarcinoma (CCA) is still ineffective and the search for a novel treatment is needed. In this study, eight novel mono-triazole glycosides (W1-W8) were synthesized and tested for their anticancer activities in CCA cell lines. The anti-proliferation effect and the underlying mechanisms of the triazole glycosides were explored. Viable cells were determined using the MTT test. Among glycosides tested, W4 and W5 exhibited the most potent anticancer activity in a dose- and time-dependent fashion. Flow cytometry and wstern blot analysis revealed that W4 and W5 induced G 0 /G 1 phase cell-cycle arrest through down-regulation of cyclin D1, cyclin E and induction of cyclin-dependent kinase inhibitors, p27 and p21 protein expression. Annexin V/propidium iodide (PI) staining demonstrated that W4 and W5 also induced apoptotic cells in a dose-dependent manner via caspase signaling cascade. Together, these findings imply that the novel synthetic glycosides might be a promising anticancer agent for CCA. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Evidence for the load-dependent mechanical efficiency of individual myosin heads in skeletal muscle fibers activated by laser flash photolysis of caged calcium in the presence of a limited amount of ATP.

PubMed

Sugi, H; Iwamoto, H; Akimoto, T; Ushitani, H

1998-03-03

Although a contracting muscle regulates its energy output depending on the load imposed on it ("Fenn effect"), the mechanism underlying the load-dependent energy output remains obscure. To explore the possibility that the mechanical efficiency, with which chemical energy derived from ATP hydrolysis is converted into mechanical work, of individual myosin heads changes in a load-dependent manner, we examined the auxotonic shortening of glycerinated rabbit psoas muscle fibers, containing ATP molecules almost equal in number to the myosin heads, after laser-flash photolysis of caged calcium. Immediately before laser-flash activation, almost all of the myosin heads in the fiber are in the state M.ADP.Pi, and can undergo only one ATP hydrolysis cycle after activation. When the fibers were activated to shorten under various auxotonic loads, the length, force, and power output changes were found to be scaled according to the auxotonic load. Both the power and energy outputs were maximal under a moderate auxotonic load. The amount of M.ADP.Pi utilized at a time after activation was estimated from the amount of isometric force developed after interruption of fiber shortening. This amount was minimal in the isometric condition and increased nearly in proportion to the distance of fiber shortening. These results are taken as evidence that the efficiency of chemomechanical energy conversion in individual myosin heads changes in a load-dependent manner.

Load-dependent mechanical efficiency of individual myosin heads in skeletal muscle fibers activated by laser flash photolysis of caged calcium in the presence of a limited amount of ATP.

PubMed

Sugi, H; Iwamoto, H; Akimoto, T; Ushitani, H

1998-01-01

Although a contracting muscle regulates its energy output depending on the load imposed on it ("Fenn effect"), the mechanism underlying the load-dependent energy output remains obscure. To explore the possibility that the mechanical efficiency, with which chemical energy derived from ATP hydrolysis is converted into mechanical work, of individual myosin heads changes in a load-dependent manner, we examined the auxotonic shortening of glycerinated rabbit psoas muscle fibers, containing ATP molecules almost equal in number to the myosin heads, following laser flash photolysis of caged calcium. Immediately before laser flash activation, almost all of the myosin heads in the fiber are in the state, M.ADP.Pi, and can undergo only one ATP hydrolysis cycle after activation. When the fibers were activated to shorten under various auxotonic loads, the length, force and power output changes were found to be scaled according to the auxotonic load. Both the power and energy outputs were maximal under a moderate auxotonic load. The amount of M.ADP.Pi utilized at a time after activation was estimated from the amount of isometric force developed after interruption of fiber shortening. This amount was minimal in the isometric condition, and increased nearly in proportion to the distance of fiber shortening. These results are taken as evidence that the efficiency of chemo-mechanical energy conversion in individual myosin heads changes in a load-dependent manner.

RNA helicase MOV10 functions as a co-factor of HIV-1 Rev to facilitate Rev/RRE-dependent nuclear export of viral mRNAs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Feng; Zhang, Junsong; Zhang, Yijun

Human immunodeficiency virus type 1 (HIV-1) exploits multiple host factors during its replication. The REV/RRE-dependent nuclear export of unspliced/partially spliced viral transcripts needs the assistance of host proteins. Recent studies have shown that MOV10 overexpression inhibited HIV-1 replication at various steps. However, the endogenous MOV10 was required in certain step(s) of HIV-1 replication. In this report, we found that MOV10 potently enhances the nuclear export of viral mRNAs and subsequently increases the expression of Gag protein and other late products through affecting the Rev/RRE axis. The co-immunoprecipitation analysis indicated that MOV10 interacts with Rev in an RNA-independent manner. The DEAG-boxmore » of MOV10 was required for the enhancement of Rev/RRE-dependent nuclear export and the DEAG-box mutant showed a dominant-negative activity. Our data propose that HIV-1 utilizes the anti-viral factor MOV10 to function as a co-factor of Rev and demonstrate the complicated effects of MOV10 on HIV-1 life cycle. - Highlights: • MOV10 can function as a co-factor of HIV-1 Rev. • MOV10 facilitates Rev/RRE-dependent transport of viral mRNAs. • MOV10 interacts with Rev in an RNA-independent manner. • The DEAG-box of MOV10 is required for the enhancement of Rev/RRE-dependent export.« less

Molecular identification of PpHDAC1, the first histone deacetylase fron the slime mold Physarum polycephalum.

PubMed

Brandtner, Eva-Maria; Lechner, Thomas; Loidl, Peter; Lusser, Alexandra

2002-01-01

The dynamic state of post-translational acetylation of eukaryotic histones is maintained by histone acetyltransferases (HATs) and histone deacetylases (HDACs). HATs and HDACs have been shown to be components of various regulatory protein complexes in the cell. Their enzymatic activities, intracellular localization and substrate specificities are regulated in a complex, cell cycle related manner. In the myxomycete Physarum polycephalum multiple HATs and HDACs can be distinguished in biochemical terms and they exhibit dynamic activity patterns depending on the cell cycle stage. Here we report on the cloning of the first P. polycephalum HDAC (PpHDAC1) related to the S. cerevisiae Rpd3 protein. The expression pattern of PpHDAC1 mRNA was analysed at different time points of the cell cycle and found to be largely constant. Treatment of macroplasmodia with the HDAC inhibitor trichostatin A at several cell cycle stages resulted in a significant delay in entry into mitosis of treated versus untreated plasmodia. No effect of TSA treatment could be observed on PpHDAC1 expression itself.

Curing of Thick Thermoset Composite Laminates: Multiphysics Modeling and Experiments

NASA Astrophysics Data System (ADS)

Anandan, S.; Dhaliwal, G. S.; Huo, Z.; Chandrashekhara, K.; Apetre, N.; Iyyer, N.

2017-11-01

Fiber reinforced polymer composites are used in high-performance aerospace applications as they are resistant to fatigue, corrosion free and possess high specific strength. The mechanical properties of these composite components depend on the degree of cure and residual stresses developed during the curing process. While these parameters are difficult to determine experimentally in large and complex parts, they can be simulated using numerical models in a cost-effective manner. These simulations can be used to develop cure cycles and change processing parameters to obtain high-quality parts. In the current work, a numerical model was built in Comsol MultiPhysics to simulate the cure behavior of a carbon/epoxy prepreg system (IM7/Cycom 5320-1). A thermal spike was observed in thick laminates when the recommended cure cycle was used. The cure cycle was modified to reduce the thermal spike and maintain the degree of cure at the laminate center. A parametric study was performed to evaluate the effect of air flow in the oven, post cure cycles and cure temperatures on the thermal spike and the resultant degree of cure in the laminate.

Akt1/PKB upregulation leads to vascular smooth muscle cell hypertrophy and polyploidization

PubMed Central

Hixon, Mary L.; Muro-Cacho, Carlos; Wagner, Mark W.; Obejero-Paz, Carlos; Millie, Elise; Fujio, Yasushi; Kureishi, Yasuko; Hassold, Terry; Walsh, Kenneth; Gualberto, Antonio

2000-01-01

Vascular smooth muscle cells (VSMCs) at capacitance arteries of hypertensive individuals and animals undergo marked age- and blood pressure–dependent polyploidization and hypertrophy. We show here that VSMCs at capacitance arteries of rat models of hypertension display high levels of Akt1/PKB protein and activity. Gene transfer of Akt1 to VSMCs isolated from a normotensive rat strain was sufficient to abrogate the activity of the mitotic spindle cell–cycle checkpoint, promoting polyploidization and hypertrophy. Furthermore, the hypertrophic agent angiotensin II induced VSMC polyploidization in an Akt1-dependent manner. These results demonstrate that Akt1 regulates ploidy levels in VSMCs and contributes to vascular smooth muscle polyploidization and hypertrophy during hypertension. PMID:11032861

Subpopulation-proteomics in prokaryotic populations.

PubMed

Jahn, Michael; Seifert, Jana; von Bergen, Martin; Schmid, Andreas; Bühler, Bruno; Müller, Susann

2013-02-01

Clonal microbial cells do not behave in an identical manner and form subpopulations during cultivation. Besides varying micro-environmental conditions, cell inherent features like cell cycle dependent localization and concentration of regulatory proteins as well as epigenetic properties are well accepted mechanisms creating cell heterogeneity. Another suspected reason is molecular noise on the transcriptional and translational level. A promising tool to unravel reasons for cell heterogeneity is the combination of cell sorting and subpopulation proteomics. This review summarizes recent developments in prokaryotic single-cell analytics and provides a workflow for selection of single cells, low cell number mass spectrometry, and proteomics evaluation. This approach is useful for understanding the dependency of individual cell decisions on inherent protein profiles. Copyright © 2012 Elsevier Ltd. All rights reserved.

Abnormal Activation of RhoA/ROCK-I Signaling in Junctional Zone Smooth Muscle Cells of Patients With Adenomyosis.

PubMed

Wang, S; Duan, H; Zhang, Y; Sun, F Q

2016-03-01

Adenomyosis (ADS) is a common estrogen-dependent gynecological disease with unknown etiology. The RhoA/Rho-kinase (ROCK) signaling pathway is involved in various cellular functions, including migration, proliferation, and smooth muscle contraction. Here we examined the potential role of this pathway in junctional zone (JZ) contraction in women with and without ADS. We demonstrated that in the normal JZ, RhoA and ROCK-I messenger RNA (mRNA) and protein expression was significantly higher in the proliferative phase of the menstrual cycle than in the secretory phase. Expression of RhoA and ROCK-I in the JZ from women with ADS was significantly higher than in the control women and showed no significant differences across the menstrual cycle. Treatment of JZ smooth muscle cells (JZSMCs) with estrogen at 0, 1, 10, or 100 nmol/L for 24 hours resulted in increased expression of RhoA, ROCK-I, and myosin light-chain (MLC) phosphorylation (p-MLC) in a dose-dependent manner. In parallel to its effects on p-MLC, estrogen-mediated, dose-dependent contraction responses in JZSMCs. Estrogen-mediated contraction in the ADS group was significantly higher than in the controls and also showed no significant differences across the menstrual cycle. These effects were suppressed in the presence of ICI 182780 or Y27632, supporting an estrogen receptor-dependent and RhoA activation-dependent mechanism. Our results indicate that the level of RhoA and ROCK-I increases in patients with ADS and the cyclic change is lost. Estrogen may affect uterine JZ contraction of ADS by enhancing RhoA/ ROCK-I signaling. © The Author(s) 2015.

The effect of ATM kinase inhibition on the initial response of human dental pulp and periodontal ligament mesenchymal stem cells to ionizing radiation.

PubMed

Cmielova, Jana; Havelek, Radim; Kohlerova, Renata; Soukup, Tomas; Bruckova, Lenka; Suchanek, Jakub; Vavrova, Jirina; Mokry, Jaroslav; Rezacova, Martina

2013-07-01

This study evaluates early changes in human mesenchymal stem cells (MSC) isolated from dental pulp and periodontal ligament after γ-irradiation and the effect of ataxia-telangiectasia mutated (ATM) inhibition. MSC were irradiated with 2 and 20 Gy by (60)Co. For ATM inhibition, specific inhibitor KU55933 was used. DNA damage was measured by Comet assay and γH2AX detection. Cell cycle distribution and proteins responding to DNA damage were analyzed 2-72 h after the irradiation. The irradiation of MSC causes an increase in γH2AX; the phosphorylation was ATM-dependent. Irradiation activates ATM kinase, and the level of p53 protein is increased due to its phosphorylation on serine15. While this phosphorylation of p53 is ATM-dependent in MSC, the increase in p53 was not prevented by ATM inhibition. A similar trend was observed for Chk1 and Chk2. The increase in p21 is greater without ATM inhibition. ATM inhibition also does not fully abrogate the accumulation of irradiated MSC in the G2-phase of the cell-cycle. In irradiated MSC, double-strand breaks are tagged quickly by γH2AX in an ATM-dependent manner. Although phosphorylations of p53(ser15), Chk1(ser345) and Chk2(thr68) are ATM-dependent, the overall amount of these proteins increases when ATM is inhibited. In both types of MSC, ATM-independent mechanisms for cell-cycle arrest in the G2-phase are triggered.

Emerging players in the initiation of eukaryotic DNA replication

PubMed Central

2012-01-01

Faithful duplication of the genome in eukaryotes requires ordered assembly of a multi-protein complex called the pre-replicative complex (pre-RC) prior to S phase; transition to the pre-initiation complex (pre-IC) at the beginning of DNA replication; coordinated progression of the replisome during S phase; and well-controlled regulation of replication licensing to prevent re-replication. These events are achieved by the formation of distinct protein complexes that form in a cell cycle-dependent manner. Several components of the pre-RC and pre-IC are highly conserved across all examined eukaryotic species. Many of these proteins, in addition to their bona fide roles in DNA replication are also required for other cell cycle events including heterochromatin organization, chromosome segregation and centrosome biology. As the complexity of the genome increases dramatically from yeast to human, additional proteins have been identified in higher eukaryotes that dictate replication initiation, progression and licensing. In this review, we discuss the newly discovered components and their roles in cell cycle progression. PMID:23075259

An attempt to evaluate the effect of vitamin K3 using as an enhancer of anticancer agents.

PubMed

Matzno, Sumio; Yamaguchi, Yuka; Akiyoshi, Takeshi; Nakabayashi, Toshikatsu; Matsuyama, Kenji

2008-06-01

The possibility of vitamin K3 (VK3) as an anticancer agent was assessed. VK3 dose-dependently diminished the cell viability (measured as esterase activity) with IC50 of 13.7 microM and Hill coefficient of 3.1 in Hep G2 cells. It also decreased the population of S phase and arrested cell cycle in the G2/M phase in a dose-dependent manner. G2/M arrest was regulated by the increment of cyclin A/cdk1 and cyclin A/cdk2 complex, and contrasting cyclin B/cdk1 complex decrease. Finally, combined application demonstrated that VK3 significantly enhanced the cytotoxicity of etoposide, a G2 phase-dependent anticancer agent, whereas it reduced the cytotoxic activity of irinotecan, a S phase-dependent agent. These findings suggest that VK3 induces G2/M arrest by inhibition of cyclin B/cdk1 complex formation, and is thus useful as an enhancer of G2 phase-dependent drugs in hepatic cancer chemotherapy.

DNA fragmentation and cell cycle arrest: a hallmark of apoptosis induced by crocin from kashmiri saffron in a human pancreatic cancer cell line.

PubMed

Bakshi, Hamid; Sam, Smitha; Rozati, Roya; Sultan, Phalisteen; Islam, Tajamul; Rathore, Babita; Lone, Zahoor; Sharma, Manik; Triphati, Jagrati; Saxena, Ramesh Chand

2010-01-01

Apoptosis, a widely important mechanism that contributes to cell growth reduction, is reported to be induced by Crocus sativus in different cancer types. The present study was designed to elucidate apoptosis induction by crocin, a main component of Crocus sativus in a human pancreatic cancer cell line (BxPC-3). Cell viability was measured by MTT assay, Hoechest33258 staining was used to detect the chromatin condensation characteristic of apoptosis, and DNA fragmentation was assessed by gel electrophoresis and cell cycle analysis by flow cytometry. Crocin induced apoptosis and G1-phase cell cycle arrest of BxPC-3 cells, while decreasing cell viability in a dose dependent and time dependent manner. Cells treated with 10μg/L crocin exhibited apoptotic morphology (brightly blue-fluorescent condensed nuclei on Hoechst 33258 staining) and reduction of volume. DNA analysis revealed typical ladders as early as 12 hours after treatment indicative of apoptosis. Our preclinical study demonstrated a pancreatic cancer cell line to be highly sensitive to crocin-mediated growth inhibition and apoptotic cell death. Although the molecular mechanisms of crocin action are not yet clearly understood, it appears to have potential as a therapeutic agent.

Anaplasma phagocytophilum Rab10-dependent parasitism of the trans-Golgi network is critical for completion of the infection cycle

PubMed Central

Truchan, Hilary K.; VieBrock, Lauren; Cockburn, Chelsea L.; Ojogun, Nore; Griffin, Brian P.; Wijesinghe, Dayanjan S.; Chalfant, Charles E.; Carlyon, Jason A.

2016-01-01

Summary Anaplasma phagocytophilum is an emerging human pathogen and obligate intracellular bacterium. It inhabits a host cell-derived vacuole and cycles between replicative reticulate cell (RC) and infectious dense-cored (DC) morphotypes. Host–pathogen interactions that are critical for RC-to-DC conversion are undefined. We previously reported that A. phagocytophilum recruits green fluorescent protein (GFP)-tagged Rab10, a GTPase that directs exocytic traffic from the sphingolipid-rich trans-Golgi network (TGN) to its vacuole in a guanine nucleotide-independent manner. Here, we demonstrate that endogenous Rab10-positive TGN vesicles are not only routed to but also delivered into the A. phagocytophilum-occupied vacuole (ApV). Consistent with this finding, A. phagocytophilum incorporates sphingolipids while intracellular and retains them when naturally released from host cells. TGN vesicle delivery into the ApV is Rab10 dependent, up-regulates expression of the DC-specific marker, APH1235, and is critical for the production of infectious progeny. The A. phagocytophilum surface protein, uridine monophosphate kinase, was identified as a guanine nucleotide-independent, Rab10-specific ligand. These data delineate why Rab10 is important for the A. phagocytophilum infection cycle and expand the understanding of the benefits that exploiting host cell membrane traffic affords intracellular bacterial pathogens. PMID:26289115

Induction of a Mitosis Delay and Cell Lysis by High-Level Secretion of Mouse α-Amylase from Saccharomyces cerevisiae

PubMed Central

Wang, Bi-Dar; Kuo, Tsong-Teh

2001-01-01

Some foreign proteins are produced in yeast in a cell cycle-dependent manner, but the cause of the cell cycle dependency is unknown. In this study, we found that Saccharomyces cerevisiae cells secreting high levels of mouse α-amylase have elongated buds and are delayed in cell cycle completion in mitosis. The delayed cell mitosis suggests that critical events during exit from mitosis might be disturbed. We found that the activities of PP2A (protein phosphatase 2A) and MPF (maturation-promoting factor) were reduced in α-amylase-oversecreting cells and that these cells showed a reduced level of assembly checkpoint protein Cdc55, compared to the accumulation in wild-type cells. MPF inactivation is due to inhibitory phosphorylation on Cdc28, as a cdc28 mutant which lacks an inhibitory phosphorylation site on Cdc28 prevents MPF inactivation and prevents the defective bud morphology induced by overproduction of α-amylase. Our data also suggest that high levels of α-amylase may downregulate PPH22, leading to cell lysis. In conclusion, overproduction of heterologous α-amylase in S. cerevisiae results in a negative regulation of PP2A, which causes mitotic delay and leads to cell lysis. PMID:11472949

G protein-coupled estrogen receptor 1 agonist G-1 induces cell cycle arrest in the mitotic phase, leading to apoptosis in endometriosis.

PubMed

Mori, Taisuke; Ito, Fumitake; Matsushima, Hiroshi; Takaoka, Osamu; Tanaka, Yukiko; Koshiba, Akemi; Kusuki, Izumi; Kitawaki, Jo

2015-05-01

To demonstrate the effects of the selective G protein-coupled estrogen receptor 1 (GPER) agonist G-1 in human ovarian endometriotic stromal cells (ESCs). Experimental in vitro study. University hospital. A total of 33 patients with ovarian endometrioma. Endometriotic stromal cells from ovarian chocolate cysts were treated with the GPER agonist G-1. The primary outcomes were cell proliferation, measured using the WST-8 assay; cell cycle, as analyzed using flow cytometry, fluorescent immunocytochemistry, and cytotoxicity; caspase activity, as measured by fluorescent and luminescent enzyme assays; and protein expression levels, as determined by Western blot analysis. G-1 suppressed ESC proliferation in a concentration-dependent manner. The inhibitory effect was not blocked when GPER signaling pathways, including the GPER itself, were inhibited. G-1 induced cell cycle arrest and accumulation in the sub-G1 phase in ESCs. Immunofluorescence analysis demonstrated that G-1 interrupted microtubule assembly at the mitotic phase. G-1 also induced caspase-3-dependent apoptosis without significant cytotoxicity. G-1 suppressed proliferation and induced apoptosis in ESCs, suggesting the potential use of this compound as a therapeutic drug for the treatment of endometriosis. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Nicotine induces cell proliferation in association with cyclin D1 up-regulation and inhibits cell differentiation in association with p53 regulation in a murine pre-osteoblastic cell line

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sato, Tsuyoshi; Abe, Takahiro; Nakamoto, Norimichi

Recent studies have suggested that nicotine critically affects bone metabolism. Many studies have examined the effects of nicotine on proliferation and differentiation, but the underlying molecular mechanisms remain unclear. We examined cell cycle regulators involved in the proliferation and differentiation of MC3T3-E1 cells. Nicotine induced cell proliferation in association with p53 down-regulation and cyclin D1 up-regulation. In differentiated cells, nicotine reduced alkaline phosphatase activity and mineralized nodule formation in dose-dependent manners. Furthermore, p53 expression was sustained in nicotine-treated cells during differentiation. These findings indicate that nicotine promotes the cell cycle and inhibits differentiation in association with p53 regulation in pre-osteoblasticmore » cells.« less

Polyoma small T antigen triggers cell death via mitotic catastrophe

PubMed Central

Fernando, Arun T Pores; Andrabi, Shaida; Cizmecioglu, Onur; Zhu, Cailei; Livingston, David M.; Higgins, Jonathan M.G; Schaffhausen, Brian S; Roberts, Thomas M

2014-01-01

Polyoma small T antigen (PyST), an early gene product of the polyoma virus, has been shown to cause cell death in a number of mammalian cells in a protein phosphatase 2A (PP2A)-dependent manner. In the current study, using a cell line featuring regulated expression of PyST, we found that PyST arrests cells in mitosis. Live-cell and immunofluorescence studies showed that the majority of the PyST-expressing cells were arrested in prometaphase with almost no cells progressing beyond metaphase. These cells exhibited defects in chromosomal congression, sister chromatid cohesion and spindle positioning, resulting in the activation of the Spindle Assembly Checkpoint (SAC). Prolonged mitotic arrest then led to cell death via mitotic catastrophe. Cell cycle inhibitors that block cells in G1/S prevented PyST-induced death. PyST-induced cell death that occurs during M is not dependent on p53 status. These data suggested, and our results confirmed that, PP2A inhibition could be used to preferentially kill cancer cells with p53 mutations that proliferate normally in the presence of cell cycle inhibitors. PMID:24998850

Does autophagy in the midgut epithelium of centipedes depend on the day/night cycle?

PubMed

Rost-Roszkowska, M M; Chajec, Ł; Vilimova, J; Tajovský, K; Kszuk-Jendrysik, M

2015-01-01

The midgut epithelium of two centipedes, Lithobius forficatus and Scolopendra cingulata, is composed of digestive, secretory and regenerative cells. In L. forficatus, the autophagy occurred only in the cytoplasm of the digestive cells as a sporadic process, while in S. cingulata, it occurred intensively in the digestive, secretory and regenerative cells of the midgut epithelium. In both of the species that were analyzed, this process proceeded in a continuous manner and did not depend on the day/night cycle. Ultrastructural analysis showed that the autophagosomes and autolysosomes were located mainly in the apical and perinuclear cytoplasm of the digestive cells in L. forficatus. However, in S. cingulata, the entire cytoplasm was filled with autophagosomes and autolysosomes. Initially the membranes of phagophores surround organelles during autophagosome formation. Autolysosomes result from the fusion of autophagosomes and lysosomes. Residual bodies which are the last stage of autophagy were released into the midgut lumen due to necrosis. Autophagy in the midgut epithelia that were analyzed was confirmed using acid phosphatase and mono-dansyl-cadaverine stainings. Copyright © 2014 Elsevier Ltd. All rights reserved.

Apigenin inhibits renal cell carcinoma cell proliferation.

PubMed

Meng, Shuai; Zhu, Yi; Li, Jiang-Feng; Wang, Xiao; Liang, Zhen; Li, Shi-Qi; Xu, Xin; Chen, Hong; Liu, Ben; Zheng, Xiang-Yi; Xie, Li-Ping

2017-03-21

Apigenin, a natural flavonoid found in vegetables and fruits, has antitumor activity in several cancer types. The present study evaluated the effects and mechanism of action of apigenin in renal cell carcinoma (RCC) cells. We found that apigenin suppressed ACHN, 786-0, and Caki-1 RCC cell proliferation in a dose- and time-dependent manner. A comet assay suggested that apigenin caused DNA damage in ACHN cells, especially at higher doses, and induced G2/M phase cell cycle arrest through ATM signal modulation. Small interfering RNA (siRNA)-mediated p53 knockdown showed that apigenin-induced apoptosis was likely p53 dependent. Apigenin anti-proliferative effects were confirmed in an ACHN cell xenograft mouse model. Apigenin treatment reduced tumor growth and volume in vivo, and immunohistochemical staining revealed lower Ki-67 indices in tumors derived from apigenin-treated mice. These findings suggest that apigenin exposure induces DNA damage, G2/M phase cell cycle arrest, p53 accumulation and apoptosis, which collectively suppress ACHN RCC cell proliferation in vitro and in vivo. Given its antitumor effects and low in vivo toxicity, apigenin is a highly promising agent for treatment of RCC.

Enterolactone induces G1-phase cell cycle arrest in non-small cell lung cancer cells by down-regulating cyclins and cyclin-dependent kinases

PubMed Central

Chikara, Shireen; Lindsey, Kaitlin; Dhillon, Harsharan; Mamidi, Sujan; Kittilson, Jeffrey; Christofidou-Solomidou, Melpo; Reindl, Katie M.

2017-01-01

Flaxseed is a rich source of the plant lignan secoisolariciresinol diglucoside (SDG) which is metabolized into mammalian lignans enterodiol (ED) and enterolactone (EL) in the digestive tract. The anti-cancer properties of these lignans have been demonstrated for various cancer types, but have not been studied for lung cancer. In this study we investigated the anti-cancer effects of EL for several non-small cell lung cancer (NSCLC) cell lines of various genetic backgrounds. EL inhibited the growth of A549, H441, and H520 lung cancer cells in concentration- and time-dependent manners. The anti-proliferative effects of EL for lung cancer cells were not due to enhanced cell death, but rather due to G1-phase cell cycle arrest. Molecular studies revealed that EL- decreased mRNA or protein expression levels of the G1-phase promoters cyclin D1, cyclin E, cyclin-dependent kinases (CDK)-2, -4, and -6, and p-cdc25A; decreased phosphorylated retinoblastoma (p-pRb) protein levels; and simultaneously increased levels of p21WAF1/CIP1, a negative regulator of the G1-phase. The results suggest that EL inhibits the growth of NSCLC cell lines by down-regulating G1-phase cyclins and CDKs, and up-regulating p21WAF1/CIP1, which leads to G1-phase cell cycle arrest. Therefore, EL may hold promise as an adjuvant treatment for lung cancer therapy. PMID:28323486

Enterolactone Induces G1-phase Cell Cycle Arrest in Nonsmall Cell Lung Cancer Cells by Downregulating Cyclins and Cyclin-dependent Kinases.

PubMed

Chikara, Shireen; Lindsey, Kaitlin; Dhillon, Harsharan; Mamidi, Sujan; Kittilson, Jeffrey; Christofidou-Solomidou, Melpo; Reindl, Katie M

2017-01-01

Flaxseed is a rich source of the plant lignan secoisolariciresinol diglucoside (SDG), which is metabolized into mammalian lignans enterodiol (ED) and enterolactone (EL) in the digestive tract. The anticancer properties of these lignans have been demonstrated for various cancer types, but have not been studied for lung cancer. In this study, we investigated the anticancer effects of EL for several nonsmall cell lung cancer (NSCLC) cell lines of various genetic backgrounds. EL inhibited the growth of A549, H441, and H520 lung cancer cells in concentration- and time-dependent manners. The antiproliferative effects of EL for lung cancer cells were not due to enhanced cell death, but rather due to G 1 -phase cell cycle arrest. Molecular studies revealed that EL decreased mRNA or protein expression levels of the G 1 -phase promoters cyclin D1, cyclin E, cyclin-dependent kinases (CDK)-2, -4, and -6, and p-cdc25A; decreased phosphorylated retinoblastoma (p-pRb) protein levels; and simultaneously increased levels of p21 WAF1/CIP1 , a negative regulator of the G 1 phase. The results suggest that EL inhibits the growth of NSCLC cell lines by downregulating G 1 -phase cyclins and CDKs, and upregulating p21 WAF1/CIP1 , which leads to G 1 -phase cell cycle arrest. Therefore, EL may hold promise as an adjuvant treatment for lung cancer therapy.

[Inhibition effects of black rice pericarp extracts on cell proliferation of PC-3 cells].

PubMed

Jiang, Weiwei; Yu, Xudong; Ren, Guofeng

2013-05-01

To observe the inhibitive effects of black rice pericarp extracts on cell proliferation of human prostate cancer cell PC-3 and to explore its effecting mechanism. The black rice pericarp extract was used to treat the PC-3 cells. The inhibitory effect of black rice pericarp extract on cells proliferation of PC-3 was tested by MTT method. Cell apoptosis rates and cell cycle were measured by flow cytometric assay (FCM). Western blot was used to study the protein expression levels of p38, p-p38, JNK, p-JNK. A dose-dependent and time-dependent proliferation inhibition of black rice pericarp extract was demonstrated in PC-3. The most prominent experiment condition was inhibitory concentration with 300microg/ml and treated for 72 h. The experiment result of flow cytometry analysis demonstrates that the apoptosis rate of PC-3 cells increased along with the increasing of black rice pericarp extract concentration, and a G1-S cell cycle arrest was induced in a dose-dependent manner. After PC-3 cell was treated with black rice pericarp extract for 72 h, the expressions of p-p38, p-JNK protein increased. Black rice pericarp extract could inhibit proliferation, change the cell cycle distributions and induce apoptosis in human prostatic cancer cell PC-3. Its inhibitory effect may be through promoting activation of the JNK, p38 signaling pathway. These results suggest that black rice pericarp extract maybe has an inhibitory effect on prostatic cancer.

Quercetin, a Natural Flavonoid Interacts with DNA, Arrests Cell Cycle and Causes Tumor Regression by Activating Mitochondrial Pathway of Apoptosis

PubMed Central

Srivastava, Shikha; Somasagara, Ranganatha R.; Hegde, Mahesh; Nishana, Mayilaadumveettil; Tadi, Satish Kumar; Srivastava, Mrinal; Choudhary, Bibha; Raghavan, Sathees C.

2016-01-01

Naturally occurring compounds are considered as attractive candidates for cancer treatment and prevention. Quercetin and ellagic acid are naturally occurring flavonoids abundantly seen in several fruits and vegetables. In the present study, we evaluate and compare antitumor efficacies of quercetin and ellagic acid in animal models and cancer cell lines in a comprehensive manner. We found that quercetin induced cytotoxicity in leukemic cells in a dose-dependent manner, while ellagic acid showed only limited toxicity. Besides leukemic cells, quercetin also induced cytotoxicity in breast cancer cells, however, its effect on normal cells was limited or none. Further, quercetin caused S phase arrest during cell cycle progression in tested cancer cells. Quercetin induced tumor regression in mice at a concentration 3-fold lower than ellagic acid. Importantly, administration of quercetin lead to ~5 fold increase in the life span in tumor bearing mice compared to that of untreated controls. Further, we found that quercetin interacts with DNA directly, and could be one of the mechanisms for inducing apoptosis in both, cancer cell lines and tumor tissues by activating the intrinsic pathway. Thus, our data suggests that quercetin can be further explored for its potential to be used in cancer therapeutics and combination therapy. PMID:27068577

Evaluating antithrombotic activity of HY023016 on rat hypercoagulable model.

PubMed

Chen, Qiu-Fang; Li, Yun-Zhan; Wang, Xin-Hui; Su, You-Rui; Cui, Shuang; Miao, Ming-Xing; Jiang, Zhen-Zhou; Jiang, Mei-Ling; Jiang, Ai-Dou; Chen, Xiang; Xu, Yun-Gen; Gong, Guo-Qing

2016-06-15

The generation of thrombus is not considered as an isolated progression without other pathologic processes, which may also enhance procoagulant state. The purpose of this study was to assess whether HY023016, a novel dabigatran prodrug and an oral direct thrombin inhibitor, or dabigatran etexilate, another thrombin inhibitor can improve the state of whole blood hypercoagulability in vitro/vivo. By using whole blood flow cytometry we explored the effects of HY023016 and dabigatran etexilate on thrombin and ADP-induced human platelet-leukocyte aggregation generated in vitro. With the method of continuous infusion of thrombin intravenous, we successfully established a rat hypercoagulable model and evaluated the effect of HY023016 or dabigatran etexilate in vivo. HY023016 was able to inhibit thrombin- or ADP-induced platelet P-selectin or CD40L expression, leukocyte CD11b expression and formation of platelet-leukocyte aggregates in dose-dependent manner. Dabigatran etexilate was unable to affect ADP-induced platelet P-selectin or CD40L expression, leukocyte CD11b expression and formation of platelet-leukocyte aggregates. Based on rat hypercoagulable model, dabigatran etexilate could reverse thrombin-induced circulatory system hypercoagulable state in a concentration-dependent manner. Dabigatran etexilate also inhibited electrical stimulation induced formation of arterial thrombus in rat under hypercoagulable state, and extracorporal circulation-induced formation of thrombus in dose-dependent manner. Compared with dabigatran etexilate, HY023016 showed nearly equal or even better antithrombotic activity, regardless of reversing the cycle of rat hypercoagulable state or inhibiting platelet-leukocyte aggregation. In surrmary, HY023016 could effectively improve hypercoagulable state of circulatory system. Copyright © 2016. Published by Elsevier B.V.

On the fly quantum dynamics of electronic and nuclear wave packets

NASA Astrophysics Data System (ADS)

Komarova, Ksenia G.; Remacle, F.; Levine, R. D.

2018-05-01

Multielectronic states quantum dynamics on a grid is described in a manner motivated by on the fly classical trajectory computations. Non stationary electronic states are prepared by a few cycle laser pulse. The nuclei respond and begin moving. We solve the time dependent Schrödinger equation for the electronic and nuclear dynamics for excitation from the ground electronic state. A satisfactory accuracy is possible using a localized description on a discrete grid. This enables computing on the fly for both the nuclear and electronic dynamics including non-adiabatic couplings. Attosecond dynamics in LiH is used as an example.

Non-encapsidation Activities of the Capsid Proteins of Positive-strand RNA Viruses

PubMed Central

Ni, Peng; Kao, C. Cheng

2013-01-01

Viral capsid proteins (CPs) are characterized by their role in forming protective shells around viral genomes. However, CPs have additional and important roles in the virus infection cycles and in the cellular response to infection. These activities involve CP binding to RNAs in both sequence-specific and nonspecific manners as well as association with other proteins. This review focuses on CPs of both plant and animal-infecting viruses with positive-strand RNA genomes. We summarize the structural features of CPs and describe their modulatory roles in viral translation, RNA-dependent RNA synthesis, and host defense responses. PMID:24074574

Sequence‑dependent effect of sorafenib in combination with natural phenolic compounds on hepatic cancer cells and the possible mechanism of action.

PubMed

Bahman, Abdulmajeed A; Abaza, Mohamed Salah I; Khoushiash, Sarah I; Al-Attiyah, Rajaa J

2018-06-08

Sorafenib (Nexavar, BAY43‑9006 or Sora) is the first molecular targeted agent that has exhibited significant therapeutic benefits in advanced hepatocellular carcinoma (HCC). However, not all HCC patients respond well to Sora and novel therapeutic strategies to optimize the efficacy of Sora are urgently required. Plant‑based drugs have received increasing attention owing to their excellent chemotherapeutic and chemopreventive activities; they are also well tolerated, non‑toxic, easily available and inexpensive. It is well known that certain biologically active natural products act synergistically with synthetic drugs used in clinical applications. The present study aimed to investigate whether a combination therapy with natural phenolic compounds (NPCs), including curcumin (Cur), quercetin (Que), kaempherol (Kmf) and resveratrol (Rsv), would allow a dose reduction of Sora without concomitant loss of its effectiveness. Furthermore, the possible molecular mechanisms of this synergy were assessed. The hepatic cancer cell lines Hep3b and HepG2 were treated with Sora alone or in combination with NPCs in concomitant, sequential, and inverted sequential regimens. Cell proliferation, cell cycle, apoptosis and expression of proteins associated with the cell cycle and apoptosis were investigated. NPCs markedly potentiated the therapeutic efficacy of Sora in a sequence‑, type‑, NPC dose‑ and cell line‑dependent manner. Concomitant treatment with Sora and Cur [sensitization ratio (SR)=28], Kmf (SR=18) or Que (SR=8) was associated with the highest SRs in Hep3b cells. Rsv markedly potentiated the effect of Sora (SR=17) on Hep3b cells when administered in a reverse sequential manner. By contrast, Rsv and Que did not improve the efficacy of Sora against HepG2 cells, while concomitant treatment with Cur (SR=10) or Kmf (SR=4.01) potentiated the cytotoxicity of Sora. Concomitant treatment with Sora and Cur or Kmf caused S‑phase and G2/M phase arrest of liver cancer cells and markedly induced apoptosis compared with mono‑treatment with Sora, Cur or Kmf. Concomitant treatment with Sora and Cur reduced the protein levels of cyclins A, B2 and D1, phosphorylated retinoblastoma and B‑cell lymphoma (Bcl) extra‑large protein. By contrast, Sora and Cur co‑treatment increased the protein levels of Bcl‑2‑associated X protein, cleaved caspase‑3 and cleaved caspase‑9 in a dose‑dependent manner. In conclusion, concomitant treatment with Sora and Cur or Kmf appears to be a potent and promising therapeutic approach that may control hepatic cancer by triggering cell cycle arrest and apoptosis. Additional studies are required to examine the potential of combined treatment with Sora and NPCs in human hepatic cancer and other solid tumor types in vivo.

Krebs cycle intermediates regulate DNA and histone methylation: epigenetic impact on the aging process.

PubMed

Salminen, Antero; Kauppinen, Anu; Hiltunen, Mikko; Kaarniranta, Kai

2014-07-01

Many aging theories have proposed that mitochondria and energy metabolism have a major role in the aging process. There are recent studies indicating that Krebs cycle intermediates can shape the epigenetic landscape of chromatin by regulating DNA and histone methylation. A growing evidence indicates that epigenetics plays an important role in the regulation of healthspan but also is involved in the aging process. 2-Oxoglutarate (α-ketoglutarate) is a key metabolite in the Krebs cycle but it is also an obligatory substrate for 2-oxoglutarate-dependent dioxygenases (2-OGDO). The 2-OGDO enzyme family includes the major enzymes of DNA and histone demethylation, i.e. Ten-Eleven Translocation (TETs) and Jumonji C domain containing (JmjC) demethylases. In addition, 2-OGDO members can regulate collagen synthesis and hypoxic responses in a non-epigenetical manner. Interestingly, succinate and fumarate, also Krebs cycle intermediates, are potent inhibitors of 2-OGDO enzymes, i.e. the balance of Krebs cycle reactions can affect the level of DNA and histone methylation and thus control gene expression. We will review the epigenetic mechanisms through which Krebs cycle intermediates control the DNA and histone methylation. We propose that age-related disturbances in the Krebs cycle function induce stochastic epigenetic changes in chromatin structures which in turn promote the aging process. Copyright © 2014 Elsevier B.V. All rights reserved.

The neem limonoids azadirachtin and nimbolide induce cell cycle arrest and mitochondria-mediated apoptosis in human cervical cancer (HeLa) cells.

PubMed

Priyadarsini, R Vidya; Murugan, R Senthil; Sripriya, P; Karunagaran, D; Nagini, S

2010-06-01

Limonoids from the neem tree (Azadirachta indica) have attracted considerable research attention in recent years owing to their potent antioxidant and anti-proliferative effects. The present study was designed to investigate the cellular and molecular mechanisms by which azadirachtin and nimbolide exert cytotoxic effects in the human cervical cancer (HeLa) cell line. Both azadirachtin and nimbolide significantly suppressed the viability of HeLa cells in a dose-dependent manner by inducing cell cycle arrest at G0/G1 phase accompanied by p53-dependent p21 accumulation and down-regulation of the cell cycle regulatory proteins cyclin B, cyclin D1 and PCNA. Characteristic changes in nuclear morphology, presence of a subdiploid peak and annexin-V staining pointed to apoptosis as the mode of cell death. Increased generation of reactive oxygen species with decline in the mitochondrial transmembrane potential and release of cytochrome c confirmed that the neem limonoids transduced the apoptotic signal via the mitochondrial pathway. Altered expression of the Bcl-2 family of proteins, inhibition of NF-kappaB activation and over-expression of caspases and survivin provide compelling evidence that azadirachtin and nimbolide induce a shift of balance toward a pro-apoptotic phenotype. Antioxidants such as azadirachtin and nimbolide that can simultaneously arrest the cell cycle and target multiple molecules involved in mitochondrial apoptosis offer immense potential as anti-cancer therapeutic drugs.

Involvement of Retinoblastoma Protein and HBP1 in Histone H10 Gene Expression

PubMed Central

Lemercier, Claudie; Duncliffe, Kym; Boibessot, Isabelle; Zhang, Hui; Verdel, André; Angelov, Dimitar; Khochbin, Saadi

2000-01-01

The histone H10-encoding gene is expressed in vertebrates in differentiating cells during the arrest of proliferation. In the H10 promoter, a specific regulatory element, which we named the H4 box, exhibits features which implicate a role in mediating H10 gene expression in response to both differentiation and cell cycle control signals. For instance, within the linker histone gene family, the H4 box is found only in the promoters of differentiation-associated subtypes, suggesting that it is specifically involved in differentiation-dependent expression of these genes. In addition, an element nearly identical to the H4 box is conserved in the promoters of histone H4-encoding genes and is known to be involved in their cell cycle-dependent expression. The transcription factors interacting with the H10 H4 box were therefore expected to link differentiation-dependent expression of H10 to the cell cycle control machinery. The aim of this work was to identify such transcription factors and to obtain information concerning the regulatory pathway involved. Interestingly, our cloning strategy led to the isolation of a retinoblastoma protein (RB) partner known as HBP1. HBP1, a high-mobility group box transcription factor, interacted specifically with the H10 H4 box and moreover was expressed in a differentiation-dependent manner. We also showed that the HBP1-encoding gene is able to produce different forms of HBP1. Finally, we demonstrated that both HBP1 and RB were involved in the activation of H10 gene expression. We therefore propose that HBP1 mediates a link between the cell cycle control machinery and cell differentiation signals. Through modulating the expression of specific chromatin-associated proteins such as histone H10, HBP1 plays a vital role in chromatin remodeling events during the arrest of cell proliferation in differentiating cells. PMID:10958660

Ghrelin decreases firing activity of gonadotropin-releasing hormone (GnRH) neurons in an estrous cycle and endocannabinoid signaling dependent manner.

PubMed

Farkas, Imre; Vastagh, Csaba; Sárvári, Miklós; Liposits, Zsolt

2013-01-01

The orexigenic peptide, ghrelin is known to influence function of GnRH neurons, however, the direct effects of the hormone upon these neurons have not been explored, yet. The present study was undertaken to reveal expression of growth hormone secretagogue receptor (GHS-R) in GnRH neurons and elucidate the mechanisms of ghrelin actions upon them. Ca(2+)-imaging revealed a ghrelin-triggered increase of the Ca(2+)-content in GT1-7 neurons kept in a steroid-free medium, which was abolished by GHS-R-antagonist JMV2959 (10 µM) suggesting direct action of ghrelin. Estradiol (1nM) eliminated the ghrelin-evoked rise of Ca(2+)-content, indicating the estradiol dependency of the process. Expression of GHS-R mRNA was then confirmed in GnRH-GFP neurons of transgenic mice by single cell RT-PCR. Firing rate and burst frequency of GnRH-GFP neurons were lower in metestrous than proestrous mice. Ghrelin (40 nM-4 μM) administration resulted in a decreased firing rate and burst frequency of GnRH neurons in metestrous, but not in proestrous mice. Ghrelin also decreased the firing rate of GnRH neurons in males. The ghrelin-evoked alterations of the firing parameters were prevented by JMV2959, supporting the receptor-specific actions of ghrelin on GnRH neurons. In metestrous mice, ghrelin decreased the frequency of GABAergic mPSCs in GnRH neurons. Effects of ghrelin were abolished by the cannabinoid receptor type-1 (CB1) antagonist AM251 (1µM) and the intracellularly applied DAG-lipase inhibitor THL (10 µM), indicating the involvement of retrograde endocannabinoid signaling. These findings demonstrate that ghrelin exerts direct regulatory effects on GnRH neurons via GHS-R, and modulates the firing of GnRH neurons in an ovarian-cycle and endocannabinoid dependent manner.

Butylated Hydroxyanisole Stimulates Heme Oxygenase-1 Gene Expression and Inhibits Neointima Formation in Rat Arteries

PubMed Central

Liu, Xiao-ming; Azam, Mohammed A.; Peyton, Kelly J.; Ensenat, Diana; Keswani, Amit N.; Wang, Hong; Durante, William

2007-01-01

Objective Butylated hydroxyanisole (BHA) is a synthetic phenolic compound that is a potent inducer of phase II genes. Since heme oxygenase-1 (HO-1) is a vasoprotective protein that is upregulated by phase II inducers, the present study examined the effects of BHA on HO-1 gene expression and vascular smooth muscle cell proliferation. Methods The regulation of HO-1 gene expression and vascular cell growth by BHA was studied in cultured rat aortic smooth muscle cells and in balloon injured rat carotid arteries. Results Treatment of cultured smooth muscle cells with BHA stimulated the expression of HO-1 protein, mRNA and promoter activity in a time- and concentration-dependent manner. BHA-mediated HO-1 expression was dependent on the activation of NF-E2-related factor-2 by p38 mitogen-activated protein kinase. BHA also inhibited cell cycle progression and DNA synthesis in a HO-1-dependent manner. In addition, the local perivascular delivery of BHA immediately after arterial injury of rat carotid arteries induced HO-1 protein expression and markedly attenuated neointima formation. Conclusions These studies demonstrate that BHA stimulates HO-1 gene expression in vascular smooth muscle cells, and that the induction of HO-1 contributes to the antiproliferative actions of this phenolic antioxidant. BHA represents a potentially novel therapeutic agent in treating or preventing vasculoproliferative disease. PMID:17320844

DNA replication initiator Cdc6 also regulates ribosomal DNA transcription initiation.

PubMed

Huang, Shijiao; Xu, Xiaowei; Wang, Guopeng; Lu, Guoliang; Xie, Wenbing; Tao, Wei; Zhang, Hongyin; Jiang, Qing; Zhang, Chuanmao

2016-04-01

RNA-polymerase-I-dependent ribosomal DNA (rDNA) transcription is fundamental to rRNA processing, ribosome assembly and protein synthesis. However, how this process is initiated during the cell cycle is not fully understood. By performing a proteomic analysis of transcription factors that bind RNA polymerase I during rDNA transcription initiation, we identified that the DNA replication initiator Cdc6 interacts with RNA polymerase I and its co-factors, and promotes rDNA transcription in G1 phase in an ATPase-activity-dependent manner. We further showed that Cdc6 is targeted to the nucleolus during late mitosis and G1 phase in a manner that is dependent on B23 (also known as nucleophosmin, NPM1), and preferentially binds to the rDNA promoter through its ATP-binding domain. Overexpression of Cdc6 increases rDNA transcription, whereas knockdown of Cdc6 results in a decreased association of both RNA polymerase I and the RNA polymerase I transcription factor RRN3 with rDNA, and a reduction of rDNA transcription. Furthermore, depletion of Cdc6 impairs the interaction between RRN3 and RNA polymerase I. Taken together, our data demonstrate that Cdc6 also serves as a regulator of rDNA transcription initiation, and indicate a mechanism by which initiation of rDNA transcription and DNA replication can be coordinated in cells. © 2016. Published by The Company of Biologists Ltd.

Female Mice Avoid Male Odor from the Same Strain via the Vomeronasal System in an Estrogen-Dependent Manner.

PubMed

Yano, Saori; Sakamoto, Kentaro Q; Habara, Yoshiaki

2015-11-01

Inbreeding avoidance is essential to providing offspring with genetic diversity. Females' mate choice is more crucial than males' for successful reproduction because of the high cost of producing gametes and limited chances to mate. However, the mechanism of female inbreeding avoidance is still unclear. To elucidate the mechanism underlying inbreeding avoidance by females, we conducted Y-maze behavioral assays using BALB/c and C57BL/6 female mice. In both strains, the avoidance of male urine from the same strain was lower in the low estrogen phase than in the high estrogen phase. The estrous cycle-dependent avoidance was completely prevented by vomeronasal organ (VNO) removal. To assess the regulation of the vomeronasal system by estrogen, the neural excitability was evaluated by immunohistochemistry of the immediate early gene products. Although estrogen did not affect neural excitability in the VNO, estrogen enhanced the neural excitability of the mitral cell layer in the AOB induced by urine from the cognate males. These results suggest that female mice avoid odor from genetically similar males in an estrogen-dependent manner via the vomeronasal system and the excitability of the mitral cells in the AOB is presumed to be regulated by estrogen. © The Author 2015. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

FOXO3 Modulates Endothelial Gene Expression and Function by Classical and Alternative Mechanisms*

PubMed Central

Czymai, Tobias; Viemann, Dorothee; Sticht, Carsten; Molema, Grietje; Goebeler, Matthias; Schmidt, Marc

2010-01-01

FOXO transcription factors represent targets of the phosphatidylinositol 3-kinase/protein kinase B survival pathway controlling important biological processes, such as cell cycle progression, apoptosis, vascular remodeling, stress responses, and metabolism. Recent studies suggested the existence of alternative mechanisms of FOXO-dependent gene expression beyond classical binding to a FOXO-responsive DNA-binding element (FRE). Here we analyzed the relative contribution of those mechanisms to vascular function by comparing the transcriptional and cellular responses to conditional activation of FOXO3 and a corresponding FRE-binding mutant in human primary endothelial cells. We demonstrate that FOXO3 controls expression of vascular remodeling genes in an FRE-dependent manner. In contrast, FOXO3-induced cell cycle arrest and apoptosis occurs independently of FRE binding, albeit FRE-dependent gene expression augments the proapoptotic response. These findings are supported by bioinformatical analysis, which revealed a statistical overrepresentation of cell cycle regulators and apoptosis-related genes in the group of co-regulated genes. Molecular analysis of FOXO3-induced endothelial apoptosis excluded modulators of the extrinsic death receptor pathway and demonstrated important roles for the BCL-2 family members BIM and NOXA in this process. Although NOXA essentially contributed to FRE-dependent apoptosis, BIM was effectively induced in the absence of FRE-binding, and small interfering RNA-mediated BIM depletion could rescue apoptosis induced by both FOXO3 mutants. These data suggest BIM as a critical cell type-specific mediator of FOXO3-induced endothelial apoptosis, whereas NOXA functions as an amplifying factor. Our study provides the first comprehensive analysis of alternatively regulated FOXO3 targets in relevant primary cells and underscores the importance of such genes for endothelial function and integrity. PMID:20123982

Apoptosis induction in MV4-11 and K562 human leukemic cells by Pereskia sacharosa (Cactaceae) leaf crude extract.

PubMed

Asmaa, Mat Jusoh Siti; Al-Jamal, Hamid Ali Nagi; Ang, Cheng Yong; Asan, Jamaruddin Mat; Seeni, Azman; Johan, Muhammad Farid

2014-01-01

Pereskia sacharosa is a genus of cacti widely used in folk medicine for cancer-related treatment. Anti-proliferative effects have been studied in recent years against colon, breast, cervical and lung cancer cell lines, with promising results. We here extended study of anti-proliferative effects to a blood malignancy, leukemia. Two leukemic cell lines, MV4-11 (acute myeloid leukemia) and K562 (chronic myeloid leukemia), were studied. IC50 concentrations were determined and apoptosis and cell cycle regulation were studied by flow cytometric analysis. The expression of apoptosis and cell-cycle related regulatory proteins was assessed by Western blotting. P sacharosa inhibited growth of MV4-11 and K562 cells in a dose-dependent manner. The mode of cell death was via induction of intrinsic apoptotic pathways and cell cycle arrest. There was profound up-regulation of cytochrome c, caspases, p21 and p53 expression and repression of Akt and Bcl-2 expression in treated cells. These results suggest that P sacharosa induces leukemic cell death via apoptosis induction and changes in cell cycle checkpoint, thus deserves further study for anti-leukemic potential.

Cdc15 integrates Tem1 GTPase-mediated spatial signals with Polo kinase-mediated temporal cues to activate mitotic exit.

PubMed

Rock, Jeremy M; Amon, Angelika

2011-09-15

In budding yeast, a Ras-like GTPase signaling cascade known as the mitotic exit network (MEN) promotes exit from mitosis. To ensure the accurate execution of mitosis, MEN activity is coordinated with other cellular events and restricted to anaphase. The MEN GTPase Tem1 has been assumed to be the central switch in MEN regulation. We show here that during an unperturbed cell cycle, restricting MEN activity to anaphase can occur in a Tem1 GTPase-independent manner. We found that the anaphase-specific activation of the MEN in the absence of Tem1 is controlled by the Polo kinase Cdc5. We further show that both Tem1 and Cdc5 are required to recruit the MEN kinase Cdc15 to spindle pole bodies, which is both necessary and sufficient to induce MEN signaling. Thus, Cdc15 functions as a coincidence detector of two essential cell cycle oscillators: the Polo kinase Cdc5 synthesis/degradation cycle and the Tem1 G-protein cycle. The Cdc15-dependent integration of these temporal (Cdc5 and Tem1 activity) and spatial (Tem1 activity) signals ensures that exit from mitosis occurs only after proper genome partitioning.

Ellagic acid inhibits the proliferation of human pancreatic carcinoma PANC-1 cells in vitro and in vivo.

PubMed

Cheng, Hao; Lu, Chenglin; Tang, Ribo; Pan, Yiming; Bao, Shanhua; Qiu, Yudong; Xie, Min

2017-02-14

Ellagic aicd (EA), a dietary polyphenolic compound found in plants and fruits, possesses various pharmacological activities. This study investigated the effect of EA on human pancreatic carcinoma PANC-1 cells both in vitro and in vivo; and defined the associated molecular mechanisms. In vitro, the cell growth and repairing ability were assessed by CCK-8 assay and wound healing assay. The cell migration and invasion activity was evaluated by Tanswell assay. In vivo, PANC-1 cell tumor-bearing mice were treated with different concentrations of EA. We found that EA significantly inhibited cell growth, cell repairing activity, and cell migration and invasion in a dose-dependent manner. Treatment of PANC-1 xenografted mice with EA resulted in significant inhibition in tumor growth and prolong mice survival rate. Furthermore, flow cytometric analysis showed that EA increased the percentage of cells in the G1 phase of cell cycle. Western blot analysis revealed that EA inhibited the expression of COX-2 and NF-κB. In addition, EA reversed epithelial to mesenchymal transition by up-regulating E-cadherin and down-regulating Vimentin. In summary, the present study demonstrated that EA inhibited cell growth, cell repairing activity, cell migration and invasion in a dose-dependent manner. EA also effectively inhibit human pancreatic cancer growth in mice. The anti-tumor effect of EA might be related to cell cycle arrest, down-regulating the expression of COX-2 and NF-κB, reversing epithelial to mesenchymal transition by up-regulating E-cadherin and down-regulating Vimentin. Our findings suggest that the use of EA would be beneficial for the management of pancreatic cancer.

N,N'-di-(m-methylphenyi)-3,6-dimethyl-1,4-dihydro-1,2,4,5-tetrazine-1,4-dicarboamide (ZGDHu-1) suppresses the proliferation of PANC-1 pancreatic cancer cells via apoptosis and G2/M cell cycle arrest.

PubMed

Chen, Su-Feng; Xia, Jun; Lv, Ya-Ping; Liu, Jin-Lin; Li, Wan-Xiang; Yu, Xi-Ping; Hu, Wei-Xiao; Zhou, Yong-Lie

2015-04-01

Pancreatic cancer is one of the human gastrointestinal malignancies with a high mortality and poor prognosis. Approximately eighty percent of patients are diagnosed with unresectable or metastatic disease. Thus, development of novel chemicals in the treatment of pancreatic cancer is imperative. This study aimed to investigate the anticancer effects of N,N'-di-(m-methylphenyi)-3,6-dimethyl-1,4-dihydro-1,2,4,5-tetrazine-1,4-dicarboamide (ZGDHu-1), a new tetrazine derivative, on the PANC-1 pancreatic cancer cell line and clarify the underlying molecular mechanism. Using an MTT assay, we found that ZGDHu-1 significantly suppressed the proliferation of PANC-1 cells in a time- and dose-dependent manner. Moreover, according to the morphological and flow cytometric analysis, the results indicated that ZGDHu-1 induced PANC-1 cell apoptosis and G2/M cell cycle arrest in a dose-dependent manner. In the western blot analysis, expression of the pro-apoptotic Bax gene was upregulated while the anti-apoptotic Bcl-2 gene was downregulated following treatment with ZGDHu-1. ZGDHu-1 also activated pro-caspase-3 and PARP and increased the expression of NF-κB inhibitor IκB. Furthermore, the expression levels of G2/M regulatory molecules such as cyclin B1 and cdc2 were decreased while that of Chk1 was increased. These results suggested that ZGDHu-1 suppressed the proliferation of pancreatic cancer cells, rendering it a potential therapeutic drug for the treatment of pancreatic cancer.

Alpinia oxyphylla Miq. fruit extract activates IGFR-PI3K/Akt signaling to induce Schwann cell proliferation and sciatic nerve regeneration.

PubMed

Chang, Yung-Ming; Chang, Hen-Hong; Tsai, Chin-Chuan; Lin, Hung-Jen; Ho, Tsung-Jung; Ye, Chi-Xin; Chiu, Ping-Ling; Chen, Yueh-Sheng; Chen, Ray-Jade; Huang, Chih-Yang; Lin, Chien-Chung

2017-03-31

It is known that the medicinal herb Alpinia oxyphylla Miq. is widely used as a remedy for diarrhea as well as the symptoms accompanying hypertension and cerebrovascular disorders. Moreover, it has also been reported that Alpinia oxyphylla Miq. has beneficial effects on anti-senescence and neuro-protection. This study focuses on the molecular mechanisms by which the Alpinia oxyphylla Miq. fruits promote neuron regeneration. A piece of silicone rubber was guided across a 15 mm gap in the sciatic nerve of a rat. This nerve gap was then filled with various doses of Alpinia oxyphylla Miq. fruits to assess their regenerative effect on damaged nerves. Further, we investigated the role of Alpinia oxyphylla Miq. fruits in RSC96 Schwann cell proliferation. Our current results showed that treatment with the extract of Alpinia oxyphylla Miq. fruits triggers the phosphorylated insulin-like growth factor-1 receptor- phosphatidylinositol 3-kinase/serine-threonine kinase pathway, and up-regulated the proliferating cell nuclear antigen in a dose-dependent manner. Cell cycle analysis on RSC96 Schwann cells showed that, after exposure to Alpinia oxyphylla Miq. fruit extract, the transition from the first gap phase to the synthesis phase occurs in 12-18 h. The expression of the cell cycle regulatory proteins cyclin D1, cyclin E and cyclin A increased in a dose-dependent manner. Transfection with a small interfering RNA blocked the expression of phosphatidylinositol 3-kinase and induced down-regulation both on the mRNA and protein levels, which resulted in a reduction of the expression of the survival factor B-cell lymphoma 2. We provide positive results that demonstrate that Alpinia oxyphylla Miq. fruits facilitate the survival and proliferation of RSC96 cells via insulin-like growth factor-1 signaling.

Dehydroleucodine inhibits tumor growth in a preclinical melanoma model by inducing cell cycle arrest, senescence and apoptosis.

PubMed

Costantino, Valeria V; Lobos-Gonzalez, Lorena; Ibañez, Jorge; Fernandez, Dario; Cuello-Carrión, F Darío; Valenzuela, Manuel A; Barbieri, Manuel A; Semino, Silvana N; Jahn, Graciela A; Quest, Andrew F G; Lopez, Luis A

2016-03-01

Malignant melanoma represents the fastest growing public health risk of all cancer types worldwide. Several strategies and anti-cancer drugs have been used in an effort to improve treatments, but the development of resistance to anti-neoplastic drugs remains the major cause of chemotherapy failure in melanomas. Previously, we showed that the sesquiterpene lactone, dehydroleucodine (DhL), promotes the accumulation of DNA damage markers, such as H2AX and 53BP1, in human tumor cells. Also DhL was shown to trigger either cell senescence or apoptosis in a concentration-dependent manner in HeLa and MCF7 cells. Here, we evaluated the effects of DhL on B16F0 mouse melanoma cells in vitro and in a pre-clinical melanoma model. DhL inhibited the proliferation of B16F0 cells by inducing senescence or apoptosis in a concentration-dependent manner. Also, DhL reduced the expression of the cell cycle proteins cyclin D1 and B1 and the inhibitor of apoptosis protein, survivin. In melanomas generated by subcutaneous injection of B16F0 cells into C57/BL6 mice, the treatment with 20 mg DhL /Kg/day in preventive, simultaneous and therapeutic protocols reduced tumor volumes by 70%, 60% and 50%, respectively. DhL treatments reduced the number of proliferating, while increasing the number of senescent and apoptotic tumor cells. To estimate the long-term effects of DhL, a mathematical model was applied to fit experimental data. Extrapolation beyond experimental time points revealed that DhL administration following preventive and therapeutic protocols is predicted to be more effective than simultaneous treatments with DhL in restricting tumor growth. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Malva verticillata seed extracts upregulate the Wnt pathway in human dermal papilla cells.

PubMed

Lee, E Y; Choi, E-J; Kim, J A; Hwang, Y L; Kim, C-D; Lee, M H; Roh, S S; Kim, Y H; Han, I; Kang, S

2016-04-01

Mesenchymal-epithelial interactions are important in controlling hair growth and the hair cycle. The β-catenin pathway of dermal papilla cells (DPCs) plays a pivotal role in morphogenesis and normal regeneration of hair follicles. Deletion of β-catenin in the dermal papilla reduces proliferation of the hair follicle progenitor cells that generate the hair shaft and induces an early onset of the catagen phase. In this study, a modulator of the Wnt/β-catenin activity was studied in oriental herb extracts on cultured human DPCs. The effect of Malva verticillata (M. verticillata) seeds on human DPCs was investigated by a Wnt/β-catenin reporter activity assay system (β-catenin-TCF/LEF reporter gene) and cell proliferation analysis. The synthesis of the factors related to hair growth and cycling was measured at both the mRNA and the protein level by semi-quantitative PCR and Western blot analysis, respectively. An extract from M. verticillata seeds increased Wnt reporter activity in a concentration-dependent manner and also led to increased β-catenin levels in cultured human DPCs. Myristoleic acid, identified as an effective compound of M. verticillata seeds, stimulated the proliferation of DPCs in a dose-dependent manner and increased transcription levels of the downstream targets: IGF-1, KGF, VEGF and HGF. Myristoleic acid also enhanced the phosphorylation of MAPKs (Akt and p38). Overall, the data suggest that this extract of M. verticillata seeds could be a good candidate for treating hair loss by modulating the Wnt/β-catenin pathway in DPCs. © 2015 Society of Cosmetic Scientists and the Société Française de Cosmétologie.

Ultra-high power capabilities in amorphous FePO4 thin films

NASA Astrophysics Data System (ADS)

Gandrud, Knut B.; Nilsen, Ola; Fjellvåg, Helmer

2016-02-01

Record breaking electrochemical properties of FePO4 have been found through investigation of the thickness dependent electrochemical properties of amorphous thin film electrodes. Atomic layer deposition was used for production of thin films of amorphous FePO4 with highly accurate thickness and topography. Electrochemical characterization of these thin film electrodes revealed that the thinner electrodes behave in a pseudocapacitive manner even at high rates of Li+ de/intercalation, which enabled specific powers above 1 MW kg-1 FePO4 to be obtained with minimal capacity loss. In addition, a self-enhancing kinetic effect was observed during cycling enabling more than 10,000 cycles at current rates approaching that of a supercapacitor (11s charge/discharge). The current findings may open for construction of ultra-high power battery electrodes that combines the energy density of batteries with the power capabilities of supercapacitors.

Triggering of longitudinal combustion instabilities in solid rocket motors: Nonlinear combustion response

NASA Technical Reports Server (NTRS)

Wicker, J. M.; Greene, W. D.; Kim, S. I.; Yang, V.

1995-01-01

Pulsed oscillations in solid rocket motors are investigated with emphasis on nonlinear combustion response. The study employs a wave equation governing the unsteady motions in a two-phase flow, and a solution technique based on spatial- and time-averaging. A wide class of combustion response functions is studied to second-order in fluctuation amplitude to determine if, when, and how triggered instabilities arise. Conditions for triggering are derived from analysis of limit cycles, and regions of triggering are found in parametric space. Based on the behavior of model dynamical systems, introduction of linear cross-coupling and quadratic self-coupling among the acoustic modes appears to be the manner in which the nonlinear combustion response produces triggering to a stable limit cycle. Regions of initial conditions corresponding to stable pulses were found, suggesting that stability depends on initial phase angle and harmonic content, as well as the composite amplitude, of the pulse.

Repair of DNA Damage Induced by the Cytidine Analog Zebularine Requires ATR and ATM in Arabidopsis[OPEN

PubMed Central

Liu, Chun-Hsin; Finke, Andreas; Díaz, Mariana; Rozhon, Wilfried; Poppenberger, Brigitte; Baubec, Tuncay; Pecinka, Ales

2015-01-01

DNA damage repair is an essential cellular mechanism that maintains genome stability. Here, we show that the nonmethylable cytidine analog zebularine induces a DNA damage response in Arabidopsis thaliana, independent of changes in DNA methylation. In contrast to genotoxic agents that induce damage in a cell cycle stage-independent manner, zebularine induces damage specifically during strand synthesis in DNA replication. The signaling of this damage is mediated by additive activity of ATAXIA TELANGIECTASIA MUTATED AND RAD3-RELATED and ATAXIA TELANGIECTASIA MUTATED kinases, which cause postreplicative cell cycle arrest and increased endoreplication. The repair requires a functional STRUCTURAL MAINTENANCE OF CHROMOSOMES5 (SMC5)-SMC6 complex and is accomplished predominantly by synthesis-dependent strand-annealing homologous recombination. Here, we provide insight into the response mechanism for coping with the genotoxic effects of zebularine and identify several components of the zebularine-induced DNA damage repair pathway. PMID:26023162

Caffeine stabilizes Cdc25 independently of Rad3 in S chizosaccharomyces pombe contributing to checkpoint override

PubMed Central

Alao, John P; Sjölander, Johanna J; Baar, Juliane; Özbaki-Yagan, Nejla; Kakoschky, Bianca; Sunnerhagen, Per

2014-01-01

Cdc25 is required for Cdc2 dephosphorylation and is thus essential for cell cycle progression. Checkpoint activation requires dual inhibition of Cdc25 and Cdc2 in a Rad3-dependent manner. Caffeine is believed to override activation of the replication and DNA damage checkpoints by inhibiting Rad3-related proteins in both S chizosaccharomyces pombe and mammalian cells. In this study, we have investigated the impact of caffeine on Cdc25 stability, cell cycle progression and checkpoint override. Caffeine induced Cdc25 accumulation in S . pombe independently of Rad3. Caffeine delayed cell cycle progression under normal conditions but advanced mitosis in cells treated with replication inhibitors and DNA-damaging agents. In the absence of Cdc25, caffeine inhibited cell cycle progression even in the presence of hydroxyurea or phleomycin. Caffeine induces Cdc25 accumulation in S . pombe by suppressing its degradation independently of Rad3. The induction of Cdc25 accumulation was not associated with accelerated progression through mitosis, but rather with delayed progression through cytokinesis. Caffeine-induced Cdc25 accumulation appears to underlie its ability to override cell cycle checkpoints. The impact of Cdc25 accumulation on cell cycle progression is attenuated by Srk1 and Mad2. Together our findings suggest that caffeine overrides checkpoint enforcement by inducing the inappropriate nuclear localization of Cdc25. PMID:24666325

Cell Cycle-Dependent Recruitment of Polycomb Proteins to the ASNS Promoter Counteracts C/ebp-Mediated Transcriptional Activation in Bombyx mori

PubMed Central

Li, Zhiqing; Cheng, Daojun; Mon, Hiroaki; Zhu, Li; Xu, Jian; Tatsuke, Tsuneyuki; Lee, Jae Man; Xia, Qingyou; Kusakabe, Takahiro

2013-01-01

Epigenetic modifiers and transcription factors contribute to developmentally programmed gene expression. Here, we establish a functional link between epigenetic regulation by Polycomb group (PcG) proteins and transcriptional regulation by C/ebp that orchestrates the correct expression of Bombyx mori asparagine synthetase (BmASNS), a gene involved in the biosynthesis of asparagine. We show that the cis-regulatory elements of YY1-binding motifs and the CpG island present on the BmASNS promoter are required for the recruitment of PcG proteins and the subsequent deposition of the epigenetic repression mark H3K27me3. RNAi-mediated knockdown of PcG genes leads to derepression of the BmASNS gene via the recruitment of activators, including BmC/ebp, to the promoter. Intriguingly, we find that PcG proteins and BmC/ebp can dynamically modulate the transcriptional output of the BmASNS target in a cell cycle-dependent manner. It will be essential to suppress BmASNS expression by PcG proteins at the G2/M phase of the cell cycle in the presence of BmC/ebp activator. Thus, our results provide a novel insight into the molecular mechanism underlying the recruitment and regulation of the PcG system at a discrete gene locus in Bombyx mori. PMID:23382816

Murrayafoline A Induces a G0/G1-Phase Arrest in Platelet-Derived Growth Factor-Stimulated Vascular Smooth Muscle Cells

PubMed Central

Han, Joo-Hui; Kim, Yohan; Jung, Sang-Hyuk; Lee, Jung-Jin; Park, Hyun-Soo; Song, Gyu-Yong; Cuong, Nguyen Manh; Kim, Young Ho

2015-01-01

The increased potential for vascular smooth muscle cell (VSMC) growth is a key abnormality in the development of atherosclerosis and post-angioplasty restenosis. Abnormally high activity of platelet-derived growth factor (PDGF) is believed to play a central role in the etiology of these pathophysiological situations. Here, we investigated the anti-proliferative effects and possible mechanism(s) of murrayafoline A, a carbazole alkaloid isolated from Glycosmis stenocarpa Guillamin (Rutaceae), on PDGF-BB-stimulated VSMCs. Murrayafoline A inhibited the PDGF-BB-stimulated proliferation of VSMCs in a concentration-dependent manner, as measured using a non-radioactive colorimetric WST-1 assay and direct cell counting. Furthermore, murrayafoline A suppressed the PDGF-BB-stimulated progression through G0/G1 to S phase of the cell cycle, as measured by [3H]-thymidine incorporation assay and cell cycle progression analysis. This anti-proliferative action of murrayafoline A, arresting cell cycle progression at G0/G1 phase in PDGF-BB-stimulated VSMCs, was mediated via down-regulation of the expression of cyclin D1, cyclin E, cyclin-dependent kinase (CDK)2, CDK4, and proliferating cell nuclear antigen (PCNA), and the phosphorylation of retinoblastoma protein (pRb). These results indicate that murrayafoline A may be useful in preventing the progression of vascular complications such as restenosis after percutaneous transluminal coronary angioplasty and atherosclerosis. PMID:26330754

Acetylation of Mammalian ADA3 Is Required for Its Functional Roles in Histone Acetylation and Cell Proliferation.

PubMed

Mohibi, Shakur; Srivastava, Shashank; Bele, Aditya; Mirza, Sameer; Band, Hamid; Band, Vimla

2016-10-01

Alteration/deficiency in activation 3 (ADA3) is an essential component of specific histone acetyltransferase (HAT) complexes. We have previously shown that ADA3 is required for establishing global histone acetylation patterns and for normal cell cycle progression (S. Mohibi et al., J Biol Chem 287:29442-29456, 2012, http://dx.doi.org/10.1074/jbc.M112.378901). Here, we report that these functional roles of ADA3 require its acetylation. We show that ADA3 acetylation, which is dynamically regulated in a cell cycle-dependent manner, reflects a balance of coordinated actions of its associated HATs, GCN5, PCAF, and p300, and a new partner that we define, the deacetylase SIRT1. We use mass spectrometry and site-directed mutagenesis to identify major sites of ADA3 acetylated by GCN5 and p300. Acetylation-defective mutants are capable of interacting with HATs and other components of HAT complexes but are deficient in their ability to restore ADA3-dependent global or locus-specific histone acetylation marks and cell proliferation in Ada3-deleted murine embryonic fibroblasts (MEFs). Given the key importance of ADA3-containing HAT complexes in the regulation of various biological processes, including the cell cycle, our study presents a novel mechanism to regulate the function of these complexes through dynamic ADA3 acetylation. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Acetylation of Mammalian ADA3 Is Required for Its Functional Roles in Histone Acetylation and Cell Proliferation

PubMed Central

Mohibi, Shakur; Srivastava, Shashank; Bele, Aditya; Mirza, Sameer; Band, Hamid

2016-01-01

Alteration/deficiency in activation 3 (ADA3) is an essential component of specific histone acetyltransferase (HAT) complexes. We have previously shown that ADA3 is required for establishing global histone acetylation patterns and for normal cell cycle progression (S. Mohibi et al., J Biol Chem 287:29442–29456, 2012, http://dx.doi.org/10.1074/jbc.M112.378901). Here, we report that these functional roles of ADA3 require its acetylation. We show that ADA3 acetylation, which is dynamically regulated in a cell cycle-dependent manner, reflects a balance of coordinated actions of its associated HATs, GCN5, PCAF, and p300, and a new partner that we define, the deacetylase SIRT1. We use mass spectrometry and site-directed mutagenesis to identify major sites of ADA3 acetylated by GCN5 and p300. Acetylation-defective mutants are capable of interacting with HATs and other components of HAT complexes but are deficient in their ability to restore ADA3-dependent global or locus-specific histone acetylation marks and cell proliferation in Ada3-deleted murine embryonic fibroblasts (MEFs). Given the key importance of ADA3-containing HAT complexes in the regulation of various biological processes, including the cell cycle, our study presents a novel mechanism to regulate the function of these complexes through dynamic ADA3 acetylation. PMID:27402865

Lunar Phase-Dependent Expression of Cryptochrome and a Photoperiodic Mechanism for Lunar Phase-Recognition in a Reef Fish, Goldlined Spinefoot

PubMed Central

Fukushiro, Masato; Takeuchi, Takahiro; Takeuchi, Yuki; Hur, Sung-Pyo; Sugama, Nozomi; Takemura, Akihiro; Kubo, Yoko; Okano, Keiko; Okano, Toshiyuki

2011-01-01

Lunar cycle-associated physiology has been found in a wide variety of organisms. Recent study has revealed that mRNA levels of Cryptochrome (Cry), one of the circadian clock genes, were significantly higher on a full moon night than on a new moon night in coral, implying the involvement of a photoreception system in the lunar-synchronized spawning. To better establish the generalities surrounding such a mechanism and explore the underlying molecular mechanism, we focused on the relationship between lunar phase, Cry gene expression, and the spawning behavior in a lunar-synchronized spawner, the goldlined spinefoot (Siganus guttatus), and we identified two kinds of Cry genes in this animal. Their mRNA levels showed lunar cycle-dependent expression in the medial part of the brain (mesencephalon and diencephalon) peaking at the first quarter moon. Since this lunar phase coincided with the reproductive phase of the goldlined spinefoot, Cry gene expression was considered a state variable in the lunar phase recognition system. Based on the expression profiles of SgCrys together with the moonlight's pattern of timing and duration during its nightly lunar cycle, we have further speculated on a model of lunar phase recognition for reproductive control in the goldlined spinefoot, which integrates both moonlight and circadian signals in a manner similar to photoperiodic response. PMID:22163321

Lunar phase-dependent expression of cryptochrome and a photoperiodic mechanism for lunar phase-recognition in a reef fish, goldlined spinefoot.

PubMed

Fukushiro, Masato; Takeuchi, Takahiro; Takeuchi, Yuki; Hur, Sung-Pyo; Sugama, Nozomi; Takemura, Akihiro; Kubo, Yoko; Okano, Keiko; Okano, Toshiyuki

2011-01-01

Lunar cycle-associated physiology has been found in a wide variety of organisms. Recent study has revealed that mRNA levels of Cryptochrome (Cry), one of the circadian clock genes, were significantly higher on a full moon night than on a new moon night in coral, implying the involvement of a photoreception system in the lunar-synchronized spawning. To better establish the generalities surrounding such a mechanism and explore the underlying molecular mechanism, we focused on the relationship between lunar phase, Cry gene expression, and the spawning behavior in a lunar-synchronized spawner, the goldlined spinefoot (Siganus guttatus), and we identified two kinds of Cry genes in this animal. Their mRNA levels showed lunar cycle-dependent expression in the medial part of the brain (mesencephalon and diencephalon) peaking at the first quarter moon. Since this lunar phase coincided with the reproductive phase of the goldlined spinefoot, Cry gene expression was considered a state variable in the lunar phase recognition system. Based on the expression profiles of SgCrys together with the moonlight's pattern of timing and duration during its nightly lunar cycle, we have further speculated on a model of lunar phase recognition for reproductive control in the goldlined spinefoot, which integrates both moonlight and circadian signals in a manner similar to photoperiodic response.

FHL2 regulates cell cycle-dependent and doxorubicin-induced p21Cip1/Waf1 expression in breast cancer cells.

PubMed

Martin, Bernd T; Kleiber, Kai; Wixler, Viktor; Raab, Monika; Zimmer, Brigitte; Kaufmann, Manfred; Strebhardt, Klaus

2007-07-15

The transcriptional cofactor FHL2 interacts with a broad variety of transcription factors and its expression is often deregulated in various types of cancer. Here we analyzed for the first time the molecular function of FHL2 in breast cancer. FHL2 is overexpressed in almost all human mammary carcinoma samples tested but not in normal breast tissues and only low levels of FHL2 expression were present in four premalignant ductal carcinoma in situ (DCIS). Cell cycle analysis revealed an upregulation of endogenous FHL2 towards G2/M in MDA-MB 231 cells and an accelerated G2/M transition when FHL2 expression was suppressed in these cells. In search for G2/M specific target genes regulated by FHL2, we found that expression of the cell cycle inhibitor p21Cip1/Waf1 (hereafter p21) is dependent on FHL2 in MDA-MB 231 breast cancer cells. Downregulation of FHL2 by shRNA abrogated the cell cycle dependent upregulation of p21 as well as the induction of p21 in response to treatment with the DNA damaging agent doxorubicin. FHL2-dependent p21 expression occurs in a p53-independent manner and p21 expression can be downregulated by specific inhibition of mitogen-activated protein kinases (MAPKs), implicating an involvement of MAPK signaling in this regulation. Analysis of FHL2 contribution to the MAPK signaling identified FHL2 as an important downstream effector of MAPKs in breast cancer cells, capable of transactivating endogenous AP1 target genes as well as AP1 dependent reporter genes. Finally, downregulation of FHL2 reduces the ability of MDA-MB 231 cells to form colonies in soft agar, while FHL2 overexpression enhances colony formation of breast cancer cells. Thus, our findings indicate that overexpression of the transcriptional cofactor FHL2 contributes to breast cancer development by mediating transcriptional activation of MAPK target genes known to be involved in cancer progression, such as p21.

Biochemical effects of veterinary antibiotics on proliferation and cell cycle arrest of human HEK293 cells.

PubMed

Kim, Hyeon Young; Kim, Ki-Tae; Kim, Sang Don

2012-08-01

The purpose of this study was to examine the effects of veterinary antibiotics, including amoxicillin (AMX), chlortetracycline (CTC) and tylosin (TYL), on the biochemical mechanism of human embryonic kidney cells (HEK293). CTC and TYL inhibited HEK293 cell proliferation, in both time- and dose-dependent manners, and changed the cell morphology; whereas, AMX showed no cytotoxic effects. The cell cycle analysis of CTC and TYL revealed G1-arrest in HEK293 cells. Western blot analysis also showed that CTC and TYL affected the activation of DNA damage responsive proteins, as well as cell cycle regulatory proteins, such as p53, p21(Waf1/Cip1) and Rb protein, which are crucial in the G1-S transition. The activation of p21(Waf1/Cip1) was significantly up-regulated over time, but there was no change in the level of CDK2 expression. The results of this study suggest that veterinary antibiotics, even at low level concentrations on continuous exposure, can potentially risk the development of human cells.

qSR: a quantitative super-resolution analysis tool reveals the cell-cycle dependent organization of RNA Polymerase I in live human cells.

PubMed

Andrews, J O; Conway, W; Cho, W -K; Narayanan, A; Spille, J -H; Jayanth, N; Inoue, T; Mullen, S; Thaler, J; Cissé, I I

2018-05-09

We present qSR, an analytical tool for the quantitative analysis of single molecule based super-resolution data. The software is created as an open-source platform integrating multiple algorithms for rigorous spatial and temporal characterizations of protein clusters in super-resolution data of living cells. First, we illustrate qSR using a sample live cell data of RNA Polymerase II (Pol II) as an example of highly dynamic sub-diffractive clusters. Then we utilize qSR to investigate the organization and dynamics of endogenous RNA Polymerase I (Pol I) in live human cells, throughout the cell cycle. Our analysis reveals a previously uncharacterized transient clustering of Pol I. Both stable and transient populations of Pol I clusters co-exist in individual living cells, and their relative fraction vary during cell cycle, in a manner correlating with global gene expression. Thus, qSR serves to facilitate the study of protein organization and dynamics with very high spatial and temporal resolutions directly in live cell.

Structure based discovery of clomifene as a potent inhibitor of cancer-associated mutant IDH1

PubMed Central

Luan, Shanshan; Li, Dan; Chen, Renqi; Zhang, Qian; Chen, Lixia; Huang, Jiangeng; Li, Hua

2017-01-01

Isocitrate dehydrogenase (IDH) plays an indispensable role in the tricarboxylic acid cycle, and IDH mutations are present in nearly 75% of glioma and 20% of acute myeloid leukemia. One IDH1R132H inhibitor (clomifene citrate) was found by virtual screening method, which can selectively suppress mutant enzyme activities in vitro and in vivo with a dose-dependent manner. The molecular docking indicated that clomifene occupied the allosteric site of the mutant IDH1. Enzymatic kinetics also demonstrated that clomifene inhibited mutant enzyme in a non-competitive manner. Moreover, knockdown of mutant IDH1 in HT1080 cells decreased the sensitivity to clomifene. In vivo studies indicated that clomifene significantly suppressed the tumor growth of HT1080-bearing CB-17/Icr-scid mice with oral administration of 100 mg/kg and 50 mg/kg per day. In short, our findings highlight clomifene may have clinical potential in tumor therapies as a safe and effective inhibitor of mutant IDH1. PMID:28498812

Evidence for Formation of a Radical-Mediated Flavin-N5 Covalent Intermediate.

PubMed

Dai, Yumin; Valentino, Hannah R; Sobrado, Pablo

2018-05-18

The redox-neutral reaction catalyzed by 2-haloacrylate hydratase (2-HAH) leads to the conversion of 2-chloroacrylate to pyruvate. Previous mechanistic studies demonstrated formation of a flavin-iminium ion as an important intermediate in the 2-HAH catalytic cycle. Time-resolved flavin absorbance studies were performed in this study and the data showed that the enzyme is capable of stabilizing both anionic and neutral flavin semiquinone species. The presence of a radical scavenger decreases the activity in a concentration-dependent manner. These data are consistent with the flavin iminium intermediate occurring via radical recombination. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Formononetin promotes cell cycle arrest via downregulation of Akt/Cyclin D1/CDK4 in human prostate cancer cells.

PubMed

Li, Tianyu; Zhao, Xinge; Mo, Zengnan; Huang, Weihua; Yan, Haibiao; Ling, Zhian; Ye, Yu

2014-01-01

Formononetin is an O-methylated isoflavone isolated from the root of Astragalus membranaceus. It has already been reported that formononetin could inhibit cell proliferation and induce cell apoptosis in several cancers, including prostate cancer. This study aimed to further investigate whether cell cycle arrest is involved in formononetin-mediated antitumor effect in human prostate cancer cells, along with the underlying molecular mechanism. Human prostate cancer cells PC-3 and DU145 were respectively treated with various concentrations of formononetin. The inhibitory effect of formononetin on proliferation of prostate cancer cells was determined using MTT assays and flow cytometry. Next, formononetin-induced alterations in cyclin D1, CDK4 and Akt expression in PC-3 cells were detected by real-time PCR and western blot. Formononetin dose-dependently inhibited prostate cancer cell proliferation via the induction of cell cycle arrest at G0/G1 phase in vitro, which was more evident in PC-3 cells. Meanwhile, concomitant with reduced phosphorylation of Akt in PC-3 cells, formononetin remarkably downregulated expression levels of cyclin D1 and CDK4 in a dose-dependent manner. More interestingly, in the in vivo studies, formononetin showed a noticeable inhibition of tumor growth in recipient mice. Formononetin could exhibit inhibitory activity against human prostate cancer cells in vivo and in vitro, which is associated with G1 cell cycle arrest by inactivation of Akt/cyclin D1/CDK4. Therefore, formononetin may be used as a candidate agent for clinical treatment of prostate cancer in the future.

Novel 3-Substituted 7-Phenylpyrrolo[3,2-f]quinolin-9(6H)-ones as Single Entities with Multitarget Antiproliferative Activity.

PubMed

Carta, Davide; Bortolozzi, Roberta; Hamel, Ernest; Basso, Giuseppe; Moro, Stefano; Viola, Giampietro; Ferlin, Maria Grazia

2015-10-22

A series of chemically modified 7-phenylpyrrolo[3,2-f]quinolinones was synthesized and evaluated as anticancer agents. Among them, the most cytotoxic (subnanomolar GI50 values) amidic derivative 5f was shown to act as an inhibitor of tubulin polymerization (IC50, 0.99 μM) by binding to the colchicine site with high affinity. Moreover, 5f induced cell cycle arrest in the G2/M phase of the cell cycle in a concentration dependent manner, followed by caspase-dependent apoptotic cell death. Compound 5f also showed lower toxicity in nontumoral cells, suggesting selectivity toward cancer cells. Additional experiments revealed that 5f inhibited the enzymatic activity of multiple kinases, including AURKA, FLT3, GSK3A, MAP3K, MEK, RSK2, RSK4, PLK4, ULK1, and JAK1. Computational studies showed that 5f can be properly accommodated in the colchicine binding site of tubulin as well as in the ATP binding clefts of all examined kinases. Our data indicate that the excellent antiproliferative profile of 5f may be derived from its interactions with multiple cellular targets.

Apigenin inhibits renal cell carcinoma cell proliferation

PubMed Central

Meng, Shuai; Zhu, Yi; Li, Jiang-Feng; Wang, Xiao; Liang, Zhen; Li, Shi-Qi; Xu, Xin; Chen, Hong; Liu, Ben; Zheng, Xiang-Yi; Xie, Li-Ping

2017-01-01

Apigenin, a natural flavonoid found in vegetables and fruits, has antitumor activity in several cancer types. The present study evaluated the effects and mechanism of action of apigenin in renal cell carcinoma (RCC) cells. We found that apigenin suppressed ACHN, 786-0, and Caki-1 RCC cell proliferation in a dose- and time-dependent manner. A comet assay suggested that apigenin caused DNA damage in ACHN cells, especially at higher doses, and induced G2/M phase cell cycle arrest through ATM signal modulation. Small interfering RNA (siRNA)-mediated p53 knockdown showed that apigenin-induced apoptosis was likely p53 dependent. Apigenin anti-proliferative effects were confirmed in an ACHN cell xenograft mouse model. Apigenin treatment reduced tumor growth and volume in vivo, and immunohistochemical staining revealed lower Ki-67 indices in tumors derived from apigenin-treated mice. These findings suggest that apigenin exposure induces DNA damage, G2/M phase cell cycle arrest, p53 accumulation and apoptosis, which collectively suppress ACHN RCC cell proliferation in vitro and in vivo. Given its antitumor effects and low in vivo toxicity, apigenin is a highly promising agent for treatment of RCC. PMID:28423637

2',4'-dihydroxychalcone, a flavonoid isolated from Herba oxytropis, suppresses PC-3 human prostate cancer cell growth by induction of apoptosis.

PubMed

Sheng, Yuqing; Zou, Mingchang; Wang, Yan; Li, Qiheng

2015-12-01

Natural products are a promising source for the development of novel cancer therapies, due to their potential effectiveness and low toxicity profiles. As a main component of Herba oxytropis , 2',4'-dihydroxychalcone (TFC) is known to demonstrate anti-tumor activity in vitro . In the present study, TFC was found to potently inhibit proliferation and induce apoptosis in PC-3 human prostate cancer cells in a dose-dependent manner. The results demonstrated that the induction of apoptosis is associated with cell cycle arrest at the G0/G1 phase and activation of caspase-3/-7. Additional mechanistic studies of two biomarkers, phosphatase and tensin homolog (PTEN) and cyclin-dependent kinase inhibitor 1B (p27 Kip1 ), in prostate cancer revealed that TFC treatment significantly upregulated the expression of PTEN and p27 Kip1 . The findings of the present study indicate that TFC-induced apoptosis in PC-3 cells via upregulation of PTEN and p27 Kip1 , which results in cell cycle arrest in G0/G1 phase, activation of caspase-3/-7 and induction of apoptosis. Therefore, TFC may be a potential compound for human prostate cancer therapy.

The inhibitory effect of disulfiram encapsulated PLGA NPs on tumor growth: Different administration routes.

PubMed

Fasehee, Hamidreza; Zarrinrad, Ghazaleh; Tavangar, Seyed Mohammad; Ghaffari, Seyed Hamidollah; Faghihi, Shahab

2016-06-01

The strong anticancer activity of disulfiram is hindered by its rapid degradation in blood system. A novel folate-receptor-targeted poly (lactide-co-glycolide) (PLGA)-polyethylene glycol (PEG) nanoparticle (NP) is developed for encapsulation and delivery of disulfiram into breast cancer tumor using passive (EPR effect) and active (folate receptor) targeting. The anticancer activity of disulfiram and its effect on caspase-3 activity and cell cycle are studied. The administration of encapsulated PLGA NPs using intra-peritoneal, intravenous and intra-tumor routes is investigated using animal model. Disulfiram shows strong cytotoxicity against MCF7 cell line. The activity of caspase-3 inhibited with disulfiram via dose dependent manner while the drug causes cell cycle arrest in G0/G1 and S phase time-dependently. The encapsulated disulfiram shows higher activity in apoptosis induction as compared to free drug. In nontoxic dose of encapsulated disulfiram, the highest and lowest efficacy of NPs in tumor growth inhibition is observed for intravenous injection and intraperitoneal injection. It is suggested that administration of disulfiram by targeted PLGA nanoparticles using intravenous injection would present an alternative therapeutic approach for solid tumor treatment. Copyright © 2016 Elsevier B.V. All rights reserved.

Icotinib, a potent and specific EGFR tyrosine kinase inhibitor, inhibits growth of squamous cell carcinoma cell line A431 through negatively regulating AKT signaling.

PubMed

Gao, Zhenzhen; Chen, Wei; Zhang, Xiaohua; Cai, Peifen; Fang, Xianying; Xu, Qiang; Sun, Yang; Gu, Yanhong

2013-06-01

Icotinib is a potent and specific epidermal growth factor receptor tyrosine kinase inhibitor. In this study, we reported that icotinib had the antitumor activity on human squamous cell carcinoma cell line A431 in vitro. Meanwhile, adhesion to fibronectin and expression of integrin α3 and β1 were significantly reduced in a dose-dependent manner after the treatment of icotinib. Moreover, icotinib induced cell cycle arrested and affected expression of various cell cycle related proteins in squamous cancer cell line A431, whereas it did not cause apoptosis. Furthermore, icotinib remarkably down-regulated phosphorylation of protein kinase B (AKT) though blocking the interaction between 3-phosphoinositide-dependent protein kinase-1 (PDK1) and AKT in A431 cells. Taken together, it is shown that the small molecular compound, icotinib, has an anti-squamous cell carcinoma activity in vitro and its antitumor mechanism is associated with the blockage of the interaction between PDK1 and AKT. These results provide a novel strategy for anti-squamous cell carcinoma therapy. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Antioxidant and anticancer activity of Artemisia princeps var. orientalis extract in HepG2 and Hep3B hepatocellular carcinoma cells.

PubMed

Choi, Eun-Jeong; Kim, Gun-Hee

2013-10-01

The aim of the present study was to investigate antioxidant and the anticancerigen activity of a methanol extract from Artemisia princeps var. orientalis (APME), a well-known traditional herbal medicine in Asia, in hepatocellular cancer cells. To evaluate the antioxidant activity of APME, reactive oxygen species (ROS) and the antioxidant enzymes, superoxide dismutase (SOD) and catalase were investigated in HepG2 cells exposed to APME (5, 100, and 200 µg/mL) for 72 h. Then, to evaluate the anticancer activity of APME, we investigated the proliferation and apoptosis induction of HepG2 and Hep3B cells exposed to APME (1-200 µg/mL) for 24, 48, and 72 h. APME dose-dependently reduced the generation of ROS in the presence of H2O2 compared with control cells. Furthermore, it increased catalase and SOD activity. Moreover, APME inhibited cell proliferation in a dose- and time-dependent manner, but at concentrations lower than 100 µg/mL, the inhibition was less dose-dependent than time-dependent. HepG2 and Hep3B cells exposed to 5, 100, and 200 µg/mL APME for 72 h underwent cell cycle arrest and apoptosis. Exposure to APME resulted in a significant increase in the number of cells in G1 phase and a decrease in the G2/M phase cell population. In addition, APME induced P53 expression of HepG2 cells in a dose-dependent manner, and played a role in the downregulation of Bcl-2 and upregulation of Bax in both HepG2 and Hep3B cells. These results indicate the potential role of APME as an antioxidant and anticancerigen agent in hepatocarcinoma cell lines.

Shikonin induces apoptosis of HaCaT cells via the mitochondrial, Erk and Akt pathways

PubMed Central

JING, HUILING; SUN, WENYAN; FAN, JINGHUA; ZHANG, YANMIN; YANG, JIAO; JIA, JINJING; LI, JICHANG; GUO, JIAQI; LUO, SUJU; ZHENG, YAN

2016-01-01

Shikonin, which is a major ingredient of the traditional Chinese herb Lithospermum erythrorhizon, possesses various biological functions, including antimicrobial, anti-inflammatory, and antitumor activities. The present study aimed to determine the molecular mechanisms underlying the effects of shikonin on HaCaT cell apoptosis. Treatment with shikonin significantly inhibited the viability of HaCaT cells in a dose- and time-dependent manner, and promoted cell cycle arrest at G0/G1 phase and apoptosis. In addition, shikonin treatment reduced the mitochondrial membrane potential and induced reactive oxygen species generation. The results of a western blot analysis demonstrated that shikonin significantly activated caspase 3 expression, downregulated B-cell lymphoma 2 (Bcl-2) expression, and upregulated Bcl-2-associated X protein and Bcl-2 homologous antagonist killer expression in a dose-dependent manner in HaCaT cells. Furthermore, shikonin decreased extracellular signal-regulated kinase (Erk) and Akt phosphorylation. These results indicated that shikonin may exert its anti-proliferative effects by inducing apoptosis via activation of the mitochondrial signaling pathway and inactivation of the Akt and Erk pathways in HaCaT cells. Therefore, the present study suggested that shikonin may have potential as a component of therapeutic strategies for the treatment of skin diseases. PMID:26935874

Withaferin A modulates the Spindle assembly checkpoint by degradation of Mad2-Cdc20 complex in colorectal cancer cell lines.

PubMed

Das, Tania; Roy, Kumar Singha; Chakrabarti, Tulika; Mukhopadhyay, Sibabrata; Roychoudhury, Susanta

2014-09-01

Withania somnifera L. Dunal (Ashwagandha) is used over centuries in the ayurvedic medicines in India. Withaferin A, a withanolide, is the major compound present in leaf extract of the plant which shows anticancer activity against leukemia, breast cancer and colorectal cancer. It arrests the ovarian cancer cells in the G2/M phase in dose dependent manner. In the current study we show the effect of Withaferin A on cell cycle regulation of colorectal cancer cell lines HCT116 and SW480 and its effect on cell fate. Treatment of these cells with this compound leads to apoptosis in a dose dependent manner. It causes the G2/M arrest in both the cell lines. We show that Withaferin A (WA) causes mitotic delay by blocking Spindle assembly checkpoint (SAC) function. Apoptosis induced by Withaferin A is associated with proteasomal degradation of Mad2 and Cdc20, an important constituent of the Spindle Checkpoint Complex. Further overexpression of Mad2 partially rescues the deleterious effect of WA by restoring proper anaphase initiation and keeping more number of cells viable. We hypothesize that Withaferin A kills cancer cells by delaying the mitotic exit followed by inducing chromosome instability. Copyright © 2014 Elsevier Inc. All rights reserved.

Curcumin inhibits proliferation and induces apoptosis of human colorectal cancer cells by activating the mitochondria apoptotic pathway.

PubMed

Guo, Li-da; Chen, Xue-Jie; Hu, Yu-Hong; Yu, Zhi-Jun; Wang, Duo; Liu, Jing-Ze

2013-03-01

Curcumin, a natural plant extract from Curcuma longa, is known for its anti-carcinogenic and chemopreventive effects on a variety of experimental cancer models. In this study, we evaluated the effects of curcumin and elucidated its mechanism in human colorectal carcinoma cells. Cell viability assay showed that curcumin significantly inhibited the growth of LoVo cells. Curcumin treatment induced the apoptosis accompanied by ultra-structural changes and release of lactate dehydrogenase in a dose-dependent manner. Moreover, treatment with 0-30 µg/mL curcumin decreased the mitochondrial membrane potential and activated the caspase-3 and caspase-9 in a dose- and time-dependent manner. Nuclear and annexin V/PI staining showed that curcumin induced the apoptosis of LoVo cells. FACS analysis revealed that curcumin could induce the cell cycle arrest of LoVo cells at the S phase. Furthermore, western blotting analysis indicated that curcumin induced the release of cytochrome c, a significant increase of Bax and p53 and a marked reduction of Bcl-2 and survivin in LoVo cells. Taken together, our results suggested that curcumin inhibited the growth of LoVo cells by inducing apoptosis through a mitochondria-mediated pathway. Copyright © 2012 John Wiley & Sons, Ltd.

Physcion induces mitochondria-driven apoptosis in colorectal cancer cells via downregulating EMMPRIN.

PubMed

Chen, Xuehong; Gao, Hui; Han, Yantao; Ye, Junli; Xie, Jing; Wang, Chunbo

2015-10-05

Physcion, an anthraquinone derivative widely isolated and characterized from both terrestrial and marine sources, has anti-tumor effects on a variety of carcinoma cells, mainly through inhibition of cell proliferation, apoptosis induction and cell cycle arrest. However, little is known about the mechanisms underlying its role in tumor progression. In the present study, we investigated the molecular mechanisms involved in physcion-induced apoptosis in human colorectal cancer (CRC) lines HCT116. Our results showed that physcion inhibited tumor cell viability in a dose- and time-dependent manner, and induced cell apoptosis via intrinsic mitochondrial pathway. Our results also revealed that physcion treatment significantly inhibited extracelluar matrix metalloproteinase inducer (EMMPRIN) expression in HCT116 cells in a dose-dependent manner and overexpression of EMMPRIN protein markedly reduced physcion-induced cell apoptosis. Furthermore, our results strongly indicated the modulating effect of physcion on EMMPRIN is correlated with AMP-activated protein kinase (AMPK)/Hypoxia-inducible factor 1α (HIF-1α) signaling pathway. Our data provide the first experimental evidence that physcion induces mitochondrial apoptosis in CRC cells by downregulating of EMMPRIN via AMPK/HIF-1α signaling pathway and suggest a new mechanism to explain its anti-tumor effects. Copyright © 2015 Elsevier B.V. All rights reserved.

Proanthocyanidins from Uncaria rhynchophylla induced apoptosis in MDA-MB-231 breast cancer cells while enhancing cytotoxic effects of 5-fluorouracil.

PubMed

Chen, Xiao-Xin; Leung, George Pak-Heng; Zhang, Zhang-Jin; Xiao, Jian-Bo; Lao, Li-Xing; Feng, Feng; Mak, Judith Choi-Wo; Wang, Ying; Sze, Stephen Cho-Wing; Zhang, Kalin Yan-Bo

2017-09-01

Breast cancer is the most frequently diagnosed cancer and cause of cancer death in women worldwide. Current treatments often result in systematic toxicity and drug resistance. Combinational use of non-toxic phytochemicals with chemotherapeutic agents to enhance the efficacy and reduce toxicity would be one promising approach. In this study, bioactive proanthocyanidins from Uncaria rhynchophylla (UPAs) were isolated and their anti-breast cancer effects alone and in combination with 5- fluorouracil (5-FU) were investigated in MDA-MB-231 breast cancer cells. The results showed that UPAs significantly inhibited cell viability and migration ability in a dose-dependent manner. Moreover, UPAs induced apoptosis in a dose-dependent manner which was associated with increased cellular reactive oxygen species production, loss of mitochondrial membrane potential, increases of Bax/Bcl-2 ratio and levels of cleaved caspase 3. Treatments of the cells with UPAs resulted in an increase in G2/M cell cycle arrest. Cytotoxic effects of 5-FU against MDA-MB-231 cells were enhanced by UPAs. The combination treatment of UPAs and 5-FU for 48 h elicited a synergistic cytotoxic effect on MDA-MB-231 cells. Altogether, these data suggest that UPAs are potential therapeutic agents for breast cancer. Copyright © 2017 Elsevier Ltd. All rights reserved.

Superfluid thermodynamic cycle refrigerator

DOEpatents

Swift, G.W.; Kotsubo, V.Y.

1992-12-22

A cryogenic refrigerator cools a heat source by cyclically concentrating and diluting the amount of [sup 3]He in a single phase [sup 3]He-[sup 4]He solution. The [sup 3]He in superfluid [sup 4]He acts in a manner of an ideal gas in a vacuum. Thus, refrigeration is obtained using any conventional thermal cycle, but preferably a Stirling or Carnot cycle. A single phase solution of liquid [sup 3]He at an initial concentration in superfluid [sup 4]He is contained in a first variable volume connected to a second variable volume through a superleak device that enables free passage of [sup 4]He while restricting passage of [sup 3]He. The [sup 3]He is compressed (concentrated) and expanded (diluted) in a phased manner to carry out the selected thermal cycle to remove heat from the heat load for cooling below 1 K. 12 figs.

Superfluid thermodynamic cycle refrigerator

DOEpatents

Swift, Gregory W.; Kotsubo, Vincent Y.

1992-01-01

A cryogenic refrigerator cools a heat source by cyclically concentrating and diluting the amount of .sup.3 He in a single phase .sup.3 He-.sup.4 He solution. The .sup.3 He in superfluid .sup.4 He acts in a manner of an ideal gas in a vacuum. Thus, refrigeration is obtained using any conventional thermal cycle, but preferably a Stirling or Carnot cycle. A single phase solution of liquid .sup.3 He at an initial concentration in superfluid .sup.4 He is contained in a first variable volume connected to a second variable volume through a superleak device that enables free passage of .sup.4 He while restricting passage of .sup.3 He. The .sup.3 He is compressed (concentrated) and expanded (diluted) in a phased manner to carry out the selected thermal cycle to remove heat from the heat load for cooling below 1 K.

Berberine displays antitumor activity in esophageal cancer cells in vitro.

PubMed

Jiang, Shu-Xian; Qi, Bo; Yao, Wen-Jian; Gu, Cheng-Wei; Wei, Xiu-Feng; Zhao, Yi; Liu, Yu-Zhen; Zhao, Bao-Sheng

2017-04-14

To investigate the effects of berberine on esophageal cancer (EC) cells and its molecular mechanisms. Human esophageal squamous cell carcinoma cell line KYSE-70 and esophageal adenocarcinoma cell line SKGT4 were used. The effects of berberine on cell proliferation were evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. For cell cycle progression, KYSE-70 cells were stained with propidium iodide (PI) staining buffer (10 mg/mL PI and 100 mg/mL RNase A) for 30 min and cell cycle was analyzed using a BD FACSCalibur flow cytometer. For apoptosis assay, cells were stained with an Annexin V-FITC/PI apoptosis detection kit. The rate of apoptotic cells was analyzed using a dual laser flow cytometer and estimated using BD ModFit software. Levels of proteins related to cell cycle and apoptosis were examined by western blotting. Berberine treatment resulted in growth inhibition of KYSE-70 and SKGT4 cells in a dose-dependent and time-dependent manner. KYSE-70 cells were more susceptible to the inhibitory activities of berberine than SKGT4 cells were. In KYSE-70 cells treated with 50 μmol/L berberine for 48 h, the number of cells in G 2 /M phase (25.94% ± 5.01%) was significantly higher than that in the control group (9.77% ± 1.28%, P < 0.01), and berberine treatment resulted in p21 up-regulation in KYSE-70 cells. Flow cytometric analyses showed that berberine significantly augmented the KYSE-70 apoptotic population at 12 and 24 h post-treatment, when compared with control cells (0.83% vs 43.78% at 12 h, P < 0.05; 0.15% vs 81.86% at 24 h, P < 0.01), and berberine-induced apoptotic effect was stronger at 24 h compared with 12 h. Western blotting showed that berberine inhibited the phosphorylation of Akt, mammalian target of rapamycin and p70S6K, and enhanced AMP-activated protein kinase phosphorylation in a sustained manner. Berberine is an inhibitor of human EC cell growth and could be considered as a potential drug for the treatment of EC patients.

Berberine displays antitumor activity in esophageal cancer cells in vitro

PubMed Central

Jiang, Shu-Xian; Qi, Bo; Yao, Wen-Jian; Gu, Cheng-Wei; Wei, Xiu-Feng; Zhao, Yi; Liu, Yu-Zhen; Zhao, Bao-Sheng

2017-01-01

AIM To investigate the effects of berberine on esophageal cancer (EC) cells and its molecular mechanisms. METHODS Human esophageal squamous cell carcinoma cell line KYSE-70 and esophageal adenocarcinoma cell line SKGT4 were used. The effects of berberine on cell proliferation were evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. For cell cycle progression, KYSE-70 cells were stained with propidium iodide (PI) staining buffer (10 mg/mL PI and 100 mg/mL RNase A) for 30 min and cell cycle was analyzed using a BD FACSCalibur flow cytometer. For apoptosis assay, cells were stained with an Annexin V-FITC/PI apoptosis detection kit. The rate of apoptotic cells was analyzed using a dual laser flow cytometer and estimated using BD ModFit software. Levels of proteins related to cell cycle and apoptosis were examined by western blotting. RESULTS Berberine treatment resulted in growth inhibition of KYSE-70 and SKGT4 cells in a dose-dependent and time-dependent manner. KYSE-70 cells were more susceptible to the inhibitory activities of berberine than SKGT4 cells were. In KYSE-70 cells treated with 50 μmol/L berberine for 48 h, the number of cells in G2/M phase (25.94% ± 5.01%) was significantly higher than that in the control group (9.77% ± 1.28%, P < 0.01), and berberine treatment resulted in p21 up-regulation in KYSE-70 cells. Flow cytometric analyses showed that berberine significantly augmented the KYSE-70 apoptotic population at 12 and 24 h post-treatment, when compared with control cells (0.83% vs 43.78% at 12 h, P < 0.05; 0.15% vs 81.86% at 24 h, P < 0.01), and berberine-induced apoptotic effect was stronger at 24 h compared with 12 h. Western blotting showed that berberine inhibited the phosphorylation of Akt, mammalian target of rapamycin and p70S6K, and enhanced AMP-activated protein kinase phosphorylation in a sustained manner. CONCLUSION Berberine is an inhibitor of human EC cell growth and could be considered as a potential drug for the treatment of EC patients. PMID:28465635

Efficacy and safety of Vitex agnus-castus extract for treatment of premenstrual syndrome in Japanese patients: a prospective, open-label study.

PubMed

Momoeda, Mikio; Sasaki, Hidetaka; Tagashira, Eiko; Ogishima, Masayuki; Takano, Yuichi; Ochiai, Kazunori

2014-03-01

Herbal medicine containing Vitex agnus-castus (VAC) extract is widely used by women with premenstrual syndrome (PMS) in Europe, however, in Japan, clinical evidence remains to be determined. This study attempted to investigate the efficacy and safety profiles of VAC extract in Japanese patients with PMS. A multi-center, prospective, open-label, single-arm, phase 3 study was performed in Japanese women with PMS and aged 18-44 years. The patients received Prefemin® (Max Zeller Söhne AG, Romanshorn, Switzerland), containing 20 mg of VAC extract, once daily for three menstrual cycles. The efficacy profile was examined based on the intensity of ten PMS symptoms-irritability, depressed mood, anger, headache, bloating, breast fullness, skin disorder, fatigue, drowsiness, and sleeplessness-recorded by patients via a visual analog scale (VAS). In addition, the responder rate was calculated based on the total VAS score defined by the sum of the VAS scores of the first six symptoms mentioned above. Furthermore, physician's global assessment (PGA) scores were recorded. Adverse events including vital signs and laboratory test values were monitored as safety evaluation. Sixty-nine patients received Prefemin®. After the first menstrual cycle, a statistically significant decrease in total VAS score was observed (P<0.001), and the score continued to diminish for the following two cycles. Each of the ten symptom scores decreased significantly in this manner. In addition, the responder rate increased in a time-dependent manner; the rate at the third menstrual cycle was 91.0%, and almost all of the patients were without symptoms or exhibited only mild symptoms based on PGA. Eight patients exhibited non-serious adverse events, one of which was allergic dermatitis whose causal relationship with VAC was not ruled out. VAC extract improved PMS symptoms in Japanese patients, with no substantial adverse events. This is the first study to report the effect of VAC extract in Japanese patients.

Mangiferin induces cell cycle arrest at G2/M phase through ATR-Chk1 pathway in HL-60 leukemia cells.

PubMed

Peng, Z G; Yao, Y B; Yang, J; Tang, Y L; Huang, X

2015-05-12

This study aimed to determine the effect of mangiferin on the cell cycle in HL-60 leukemia cells and expression of the cell cycle-regulatory genes Wee1, Chk1 and CDC25C and to further investigate the molecular mechanisms of the antileukemic action of mangiferin. The inhibitory effect of mangiferin on HL-60 leukemia cell proliferation was determined by the MTT assay. The impact of mangiferin on the HL-60 cell cycle was evaluated by flow cytometry. After the cells were treated with different concentrations of mangiferin, the expression levels of Wee1, Chk1 and CDC25C mRNA were determined by RT-PCR, and Western blot was used to evaluate the expression levels of cdc25c, cyclin B1, and Akt proteins. The inhibition of HL-60 cell growth by mangiferin was dose- and time-dependent. After treatment for 24 h, cells in G2/M phase increased, and G2/M phase arrest appeared with increased mRNA expression of Wee1, Chk1 and CDC25C. Mangiferin inhibited Chk1 and cdc25c mRNA expression at high concentrations and induced Wee1 mRNA expression in a dose-dependent manner. It significantly inhibited ATR, Chk1, Wee1, Akt, and ERK1/2 phosphorylation but increased cdc2 and cyclin B1 phosphorylation. Furthermore, mangiferin reduced cdc25c, cyclin B1, and Akt protein levels while inducing Wee1 protein expression. It also antagonized the phosphorylation effect of vanadate on ATR, and the phosphorylation effect of EGF on Wee1. These findings indicated that mangiferin inhibits cell cycle progression through the ATR-Chk1 stress response DNA damage pathway, leading to cell cycle arrest at G2/M phase in leukemia cells.

Molecular effects of the phosphatidylinositol-3-kinase inhibitor NVP-BKM120 on T and B-cell acute lymphoblastic leukaemia.

PubMed

Pereira, João Kleber Novais; Machado-Neto, João Agostinho; Lopes, Matheus Rodrigues; Morini, Beatriz Corey; Traina, Fabiola; Costa, Fernando Ferreira; Saad, Sara Teresinha Olalla; Favaro, Patricia

2015-09-01

Constitutive activation of the PI3K pathway in T cell acute lymphoblastic leukaemia (T-ALL) has been reported and in a mouse model, PI3K activation, together with MYC, cooperates in Burkitt lymphoma (BL) pathogenesis. We investigated the effects of NVP-BKM120, a potent pan-class I PI3K inhibitor, in lymphoblastic leukaemia cell lines. Effects of NVP-BKM120 on cell viability, clonogenicity, apoptosis, cell cycle, cell signalling and autophagy were assessed in vitro on T-ALL (Jurkat and MOLT-4) and BL (Daudi and NAMALWA) cell lines. NVP-BKM120 treatment decreased cell viability and clonogenic growth in all tested cells. Moreover, the drug arrested cell cycling in association with a decrease in Cyclin B1 protein levels, and increased apoptosis. Immunoblotting analysis of cells treated with the drug revealed decreased phosphorylation, in a dose-dependent manner, of AKT, mTOR, P70S6K and 4EBP1, with stable total protein levels. Additionally, we observed a dose-dependent decrease in BAD phosphorylation, in association with augmented BAX:BCL2 ratio. Quantification of autophagy showed a dose-dependent increase in acidic vesicular organelles in all cells tested. In summary, our present study establishes that NVP-BKM120 presents an effective antitumour activity against T-ALL and BL cell lines. Copyright © 2015 Elsevier Ltd. All rights reserved.

βTrCP controls the lysosome-mediated degradation of CDK1, whose accumulation correlates with tumor malignancy.

PubMed

Herrero-Ruiz, Joaquín; Mora-Santos, Mar; Giráldez, Servando; Sáez, Carmen; Japón, Miguel A; Tortolero, Maria; Romero, Francisco

2014-09-15

In mammals, cell cycle progression is controlled by cyclin-dependent kinases, among which CDK1 plays important roles in the regulation of the G2/M transition, G1 progression and G1/S transition. CDK1 is highly regulated by its association to cyclins, phosphorylation and dephosphorylation, changes in subcellular localization, and by direct binding of CDK inhibitor proteins. CDK1 steady-state protein levels are held constant throughout the cell cycle by a coordinated regulation of protein synthesis and degradation. We show that CDK1 is ubiquitinated by the E3 ubiquitin ligase SCFβTrCP and degraded by the lysosome. Furthermore, we found that DNA damage not only triggers the stabilization of inhibitory phosphorylation sites on CDK1 and repression of CDK1 gene expression, but also regulates βTrCP-induced CDK1 degradation in a cell type-dependent manner. Specifically, treatment with the chemotherapeutic agent doxorubicin in certain cell lines provokes CDK1 degradation and induces apoptosis, whereas in others it inhibits destruction of the protein. These observations raise the possibility that different tumor types, depending on their pathogenic spectrum mutations, may display different sensitivity to βTrCP-induced CDK1 degradation after DNA damage. Finally, we found that CDK1 accumulation in patients' tumors shows a negative correlation with βTrCP and a positive correlation with the degree of tumor malignancy.

Tangeretin induces cell-cycle G1 arrest through inhibiting cyclin-dependent kinases 2 and 4 activities as well as elevating Cdk inhibitors p21 and p27 in human colorectal carcinoma cells.

PubMed

Pan, Min-Hsiung; Chen, Wei-Jen; Lin-Shiau, Shoei-Yn; Ho, Chi-Tang; Lin, Jen-Kun

2002-10-01

Tangeretin (5,6,7,8,4'-pentamethoxyflavone) is concentrated in the peel of citrus fruits. DNA flow cytometric analysis indicated that tangeretin blocked cell cycle progression at G1 phase in colorectal carcinoma COLO 205 cells. Over a 24 h exposure to tangeretin, the degree of phosphorylation of Rb was decreased after 12 h and G1 arrest developed. The protein expression of cyclins A, D1, and E reduced slightly under the same conditions. Immunocomplex kinase experiments showed that tangeretin inhibited the activities of cyclin-dependent kinases 2 (Cdk2) and 4 (Cdk4) in a dose-dependent manner in the cell-free system. As the cells were exposed to tangeretin (50 microM) over 48 h a gradual loss of both Cdk2 and 4 kinase activities occurred. Tangeretin also increased the content of the Cdk inhibitor p21 protein and this effect correlated with the elevation in p53 levels. In addition, tangeretin also increased the level of the Cdk inhibitor p27 protein within 18 h. These results suggest that tangeretin either exerts its growth-inhibitory effects through modulation of the activities of several key G1 regulatory proteins, such as Cdk2 and Cdk4, or mediates the increase of Cdk inhibitors p21 and p27.

Molecular basis of sodium butyrate-dependent proapoptotic activity in cancer cells.

PubMed

Pajak, B; Orzechowski, A; Gajkowska, B

2007-01-01

This review outlines the molecular events that accompany the antitumor action of sodium butyrate (NaBt). Butyrate, a low-molecular weight four-carbon chain volatile fatty acid (VFA) has been previously shown to withdraw cells from cell cycle or to promote cell differentiation, and finally to induce programmed cell death. Recent advances in molecular biology indicate, that this product of large bowel microbial fermentation of dietary fiber, might evoke the above-mentioned effects by indirect action on genes. NaBt was shown to inhibit histone deacetylase activity, allowing DNA binding of several transcription factors. Higher genomic activity leads to the higher expression of proapoptotic genes, higher level of their protein products and elevated sensitivity to death ligand-induced apoptosis. Cancer cells might be arrested in G1 phase of cell cycle in a p21-dependent manner. Proapoptotic activity of NaBt includes higher expression of membrane death receptors (DR4/5), higher level and activation of Smad3 protein in TGF-beta-dependent apoptotic pathway, lower level of antiapoptotic proteins (cFLIP, XIAP) and activation ofproapoptotic tBid protein. Thus, both intrinsic and extrinsic apoptotic pathways are stimulated to ampify the apoptotic signals. These effects are specific for tumor but not for regular cells. Unique properties of NaBt make this agent a promising metabolic inhibitor to retard tumorigenesis to suppress tumor growth.

Essential Dosage-Dependent Functions of the Transcription Factor Yin Yang 1 in Late Embryonic Development and Cell Cycle Progression†

PubMed Central

Affar, El Bachir; Gay, Frédérique; Shi, Yujiang; Liu, Huifei; Huarte, Maite; Wu, Su; Collins, Tucker; Li, En; Shi, Yang

2006-01-01

Constitutive ablation of the Yin Yang 1 (YY1) transcription factor in mice results in peri-implantation lethality. In this study, we used homologous recombination to generate knockout mice carrying yy1 alleles expressing various amounts of YY1. Phenotypic analysis of yy1 mutant embryos expressing ∼75%, ∼50%, and ∼25% of the normal complement of YY1 identified a dosage-dependent requirement for YY1 during late embryogenesis. Indeed, reduction of YY1 levels impairs embryonic growth and viability in a dose-dependent manner. Analysis of the corresponding mouse embryonic fibroblast cells also revealed a tight correlation between YY1 dosage and cell proliferation, with a complete ablation of YY1 inducing cytokinesis failure and cell cycle arrest. Consistently, RNA interference-mediated inhibition of YY1 in HeLa cells prevents cytokinesis, causes proliferative arrest, and increases cellular sensitivity to various apoptotic agents. Genome-wide expression profiling identified a plethora of YY1 target genes that have been implicated in cell growth, proliferation, cytokinesis, apoptosis, development, and differentiation, suggesting that YY1 coordinates multiple essential biological processes through a complex transcriptional network. These data not only shed new light on the molecular basis for YY1 developmental roles and cellular functions, but also provide insight into the general mechanisms controlling eukaryotic cell proliferation, apoptosis, and differentiation. PMID:16611997

Suppression of the DHX9 Helicase Induces Premature Senescence in Human Diploid Fibroblasts in a p53-dependent Manner*

PubMed Central

Lee, Teresa; Di Paola, Domenic; Malina, Abba; Mills, John R.; Kreps, Amina; Grosse, Frank; Tang, Hengli; Zannis-Hadjopoulos, Maria; Larsson, Ola; Pelletier, Jerry

2014-01-01

DHX9 is an ATP-dependent DEXH box helicase with a multitude of cellular functions. Its ability to unwind both DNA and RNA, as well as aberrant, noncanonical polynucleotide structures, has implicated it in transcriptional and translational regulation, DNA replication and repair, and maintenance of genome stability. We report that loss of DHX9 in primary human fibroblasts results in premature senescence, a state of irreversible growth arrest. This is accompanied by morphological defects, elevation of senescence-associated β-galactosidase levels, and changes in gene expression closely resembling those encountered during replicative (telomere-dependent) senescence. Activation of the p53 signaling pathway was found to be essential to this process. ChIP analysis and investigation of nascent DNA levels revealed that DHX9 is associated with origins of replication and that its suppression leads to a reduction of DNA replication. Our results demonstrate an essential role of DHX9 in DNA replication and normal cell cycle progression. PMID:24990949

MMSET is dynamically regulated during cell-cycle progression and promotes normal DNA replication.

PubMed

Evans, Debra L; Zhang, Haoxing; Ham, Hyoungjun; Pei, Huadong; Lee, SeungBaek; Kim, JungJin; Billadeau, Daniel D; Lou, Zhenkun

2016-01-01

The timely and precise duplication of cellular DNA is essential for maintaining genome integrity and is thus tightly-regulated. During mitosis and G1, the Origin Recognition Complex (ORC) binds to future replication origins, coordinating with multiple factors to load the minichromosome maintenance (MCM) complex onto future replication origins as part of the pre-replication complex (pre-RC). The pre-RC machinery, in turn, remains inactive until the subsequent S phase when it is required for replication fork formation, thereby initiating DNA replication. Multiple myeloma SET domain-containing protein (MMSET, a.k.a. WHSC1, NSD2) is a histone methyltransferase that is frequently overexpressed in aggressive cancers and is essential for normal human development. Several studies have suggested a role for MMSET in cell-cycle regulation; however, whether MMSET is itself regulated during cell-cycle progression has not been examined. In this study, we report that MMSET is degraded during S phase in a cullin-ring ligase 4-Cdt2 (CRL4(Cdt2)) and proteasome-dependent manner. Notably, we also report defects in DNA replication and a decreased association of pre-RC factors with chromatin in MMSET-depleted cells. Taken together, our results suggest a dynamic regulation of MMSET levels throughout the cell cycle, and further characterize the role of MMSET in DNA replication and cell-cycle progression.

Hexachlorobenzene induces cell proliferation, and aryl hydrocarbon receptor expression (AhR) in rat liver preneoplastic foci, and in the human hepatoma cell line HepG2. AhR is a mediator of ERK1/2 signaling, and cell cycle regulation in HCB-treated HepG2 cells.

PubMed

de Tomaso Portaz, Ana Clara; Caimi, Giselle Romero; Sánchez, Marcela; Chiappini, Florencia; Randi, Andrea S; Kleiman de Pisarev, Diana L; Alvarez, Laura

2015-10-02

Hexachlorobenzene (HCB) is a widespread environmental pollutant, and a liver tumor promoter in rodents. Depending on the particular cell lines studied, exposure to these compounds may lead to cell proliferation, terminal differentiation, or apoptosis. The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is involved in drug and xenobiotic metabolism. AhR can also modulate a variety of cellular and physiological processes that can affect cell proliferation and cell fate determination. The mechanisms by which AhR ligands, both exogenous and endogenous, affect these processes involve multiple interactions between AhR and other signaling pathways. In the present study, we examined the effect of HCB on cell proliferation and AhR expression, using an initiation-promotion hepatocarcinogenesis protocol in rat liver and in the human-derived hepatoma cell line, HepG2. Female Wistar rats were initiated with a single dose of 100 mg/kg of diethylnitrosamine (DEN) at the start of the experiment. Two weeks later, daily dosing of 100 mg/kg HCB was maintained for 10 weeks. Partial hepatectomy was performed 3 weeks after initiation. The number and area of glutathione S-transferase-P (GST-P)-positive foci, in the rat liver were used as biomarkers of liver precancerous lesions. Immunohistochemical staining showed an increase in proliferating cell nuclear antigen (PCNA)-positive cells, along with enhanced AhR protein expression in hepatocytes within GST-P-positive foci of (DEN HCB) group, when compared to DEN. In a similar manner, Western blot analysis demonstrated that HCB induced PCNA and AhR protein expression in HepG2 cells. Flow cytometry assay indicated that the cells were accumulated at S and G2/M phases of the cell cycle. HCB increased cyclin D1 protein levels and ERK1/2 phosphorylation in a dose-dependent manner. Treatment of cells with a selective MEK1 inhibitor, prevented HCB-stimulatory effect on PCNA and cyclinD1, indicating that these effects are mediated by ERK1/2. Pretreatment with an AhR antagonist, prevented HCB-induced PCNA protein levels, ERK1/2 phosphorylation and alterations in cell cycle distribution. These results demonstrate that HCB-induced HepG2 proliferation and cell cycle progression depend on ERK1/2 phosphorylation which is mediated by the AhR. Our results provide a clue to the molecular events involved in the mechanism of action of HCB-induced hepatocarcinogenesis. Copyright © 2015. Published by Elsevier Ireland Ltd.

Graphene oxide and reduced graphene oxide induced neural pheochromocytoma-derived PC12 cell lines apoptosis and cell cycle alterations via the ERK signaling pathways.

PubMed

Kang, Yiyuan; Liu, Jia; Wu, Junrong; Yin, Qian; Liang, Huimin; Chen, Aijie; Shao, Longquan

2017-01-01

Given the novel applications of graphene materials in biomedical and electronics industry, the health hazards of these particles have attracted extensive worldwide attention. Although many studies have been performed on graphene material-induced toxic effects, toxicological data for the effect of graphene materials on the nervous system are lacking. In this study, we focused on the biological effects of graphene oxide (GO) and reduced graphene oxide (rGO) materials on PC12 cells, a type of traditional neural cell line. We found that GO and rGO exerted significant toxic effects on PC12 cells in a dose- and time-dependent manner. Moreover, apoptosis appeared to be a response to toxicity. A potent increase in the number of PC12 cells at G0/G1 phase after GO and rGO exposure was detected by cell cycle analysis. We found that phosphorylation levels of ERK signaling molecules, which are related to cell cycle regulation and apoptosis, were significantly altered after GO and rGO exposure. In conclusion, our results show that GO has more potent toxic effects than rGO and that apoptosis and cell cycle arrest are the main toxicity responses to GO and rGO treatments, which are likely due to ERK pathway regulation.

Human rpL3 induces G₁/S arrest or apoptosis by modulating p21waf1/cip1 levels in a p53-independent manner

PubMed Central

Russo, Annapina; Esposito, Davide; Catillo, Morena; Pietropaolo, Concetta; Crescenzi, Elvira; Russo, Giulia

2013-01-01

It is now largely accepted that ribosomal proteins may be implicated in a variety of biological functions besides that of components of the translation machinery. Many evidences show that a subset of ribosomal proteins are involved in the regulation of the cell cycle and apoptosis through modulation of p53 activity. In addition, p53-independent mechanisms of cell cycle arrest in response to alterations of ribosomal proteins availability have been described. Here, we identify human rpL3 as a new regulator of cell cycle and apoptosis through positive regulation of p21 expression in a p53-independent system. We demonstrate that the rpL3-mediated p21 upregulation requires the specific interaction between rpL3 and Sp1. Furthermore, in our experimental system, p21 overexpression leads to a dual outcome, activating the G₁/S arrest of the cell cycle or the apoptotic pathway through mitochondria, depending on its intracellular levels. It is noteworthy that depletion of p21 abrogates both effects. Taken together, our findings unravel a novel extraribosomal function of rpL3 and reinforce the proapoptotic role of p21 in addition to its widely reported ability as an inhibitor of cell proliferation. PMID:23255119

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bae, Sijeong; Kwon, Hwang; Yoon, Hyemin

The sine oculis homeobox 1 (SIX1) is a member of the Six gene family. SIX1 is involved in tissue development by regulating proliferation, apoptosis, and differentiation. However, function of SIX1 in the uterus remains unknown. Here, we found that Six1 expression is regulated along the estrous cycle in mouse uterus. Six1 expression was significantly increased at estrus stage and decreased at the rest of stages. SIX1 is detected in the luminal and glandular epithelium of uterine endometrium at the estrus stage. Estrogen injection increased Six1 expression in the ovariectomized mouse uterus, whereas progesterone had no effect on its expression. Estrogenmore » receptor antagonist inhibited estrogen-induced Six1 expression. Our findings imply that SIX1 may play a role as an important regulator to orchestrate the dynamic of uterine endometrium in response to estrogen level during the estrous cycle. These results will give us a better understanding of uterine biology. - Highlights: • Six1 expression is regulated during the estrous cycle in mouse uterus. • Six1 is highly expressed at the estrus stage of estrous cycle. • SIX1 is detected in luminal/glandular epithelium of the uterus at the estrus stage. • Estrogen stimulates Six1 expression in an estrogen receptor-dependent manner.« less

Site-Specific Phosphorylation of Ikaros Induced by Low-Dose Ionizing Radiation Regulates Cell Cycle Progression of B Lymphoblast Through CK2 and AKT Activation.

PubMed

Cho, Seong-Jun; Kang, Hana; Kim, Min Young; Lee, Jung Eun; Kim, Sung Jin; Nam, Seon Young; Kim, Ji Young; Kim, Hee Sun; Pyo, Suhkneung; Yang, Kwang Hee

2016-04-01

To determine how low-dose ionizing radiation (LDIR) regulates B lympho-proliferation and its molecular mechanism related with Ikaros, transcription factor. Splenocytes and IM-9 cells were uniformly irradiated with various doses of a (137)Cs γ-source, and cell proliferation was analyzed. To determine the LDIR-specific phosphorylation of Ikaros, immunoprecipitation and Western blot analysis were performed. To investigate the physiologic function of LDIR-mediatied Ikaros phosphorylation, Ikaros mutants at phosphorylation sites were generated, and cell cycle analysis was performed. First, we found that LDIR enhances B lymphoblast proliferation in an Ikaros-dependent manner. Moreover, we found that LDIR elevates the phosphorylation level of Ikaros protein. Interestingly, we showed that CK2 and AKT are involved in LDIR-induced Ikaros phosphorylation and capable of regulating DNA binding activity of Ikaros via specific phosphorylation. Finally, we identified LDIR-specific Ikaros phosphorylation sites at S391/S393 and showed that the Ikaros phosphorylations at these sites control Ikaros's ability to regulate G1/S cell cycle progression. Low-dose ionizing radiation specifically phosphorylates Ikaros protein at Ser 391/393 residues to regulate cell cycle progression in B lymphoblast. Copyright © 2016 Elsevier Inc. All rights reserved.

C. elegans STK39/SPAK ortholog-mediated inhibition of ClC anion channel activity is regulated by WNK-independent ERK kinase signaling

PubMed Central

Falin, Rebecca A.; Miyazaki, Hiroaki

2011-01-01

Mammalian Ste20-like proline/alanine-rich kinase (SPAK) and oxidative stress-responsive 1 (OSR1) kinases phosphorylate and regulate cation-coupled Cl− cotransporter activity in response to cell volume changes. SPAK and OSR1 are activated via phosphorylation by upstream with-no-lysine (WNK) kinases. In Caenorhabditis elegans, the SPAK/OSR1 ortholog germinal center kinase (GCK)-3 binds to and regulates the activity of the cell volume- and meiotic cell cycle-dependent ClC anion channel CLH-3b. We tested the hypothesis that WNK kinases function in the GCK-3/CLH-3b signaling cascade. CLH-3b heterologously expressed in human embryonic kidney (HEK) cells was unaffected by coexpression with the single C. elegans WNK kinase, WNK-1, or kinase-dead WNK-1 dominant-negative mutants. RNA interference (RNAi) knockdown of the single Drosophila WNK kinase had no effect on the activity of CLH-3b expressed in Drosophila S2 cells. Similarly, RNAi silencing of C. elegans WNK-1 had no effect on basal or cell volume-sensitive activity of CLH-3b expressed endogenously in worm oocytes. Previous yeast 2-hybrid studies suggested that ERK kinases may function upstream of GCK-3. Pharmacological inhibition of ERK signaling disrupted CLH-3b activity in HEK cells in a GCK-3-dependent manner. RNAi silencing of the C. elegans ERK kinase MPK-1 or the ERK phosphorylating/activating kinase MEK-2 constitutively activated native CLH-3b. MEK-2 and MPK-1 play important roles in regulating the meiotic cell cycle in C. elegans oocytes. Cell cycle-dependent changes in MPK-1 correlate with the pattern of CLH-3b activation observed during oocyte meiotic maturation. We postulate that MEK-2/MPK-1 functions upstream from GCK-3 to regulate its activity during cell volume and meiotic cell cycle changes. PMID:21160027

Pseudolaric Acid B Induced Cell Cycle Arrest, Autophagy and Senescence in Murine Fibrosarcoma L929 Cell

PubMed Central

hua Yu, Jing; yu Liu, Chun; bin Zheng, Gui; Zhang, Li Ying; hui Yan, Ming; yan Zhang, Wen; ying Meng, Xian; fang Yu, Xiao

2013-01-01

Objective: PAB induced various cancer cell apoptosis, cell cycle arrest and senescence. But in cell line murine fibrosarcoma L929, PAB did not induce apoptosis, but autophagy, therefore it was thought by us as a good model to research the relationship of cell cycle arrest, autophagy and senescence bypass apoptosis. Methods: Inhibitory ratio was assessed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) analysis. Phase contrast microscopy visualized cell morphology. Hoechst 33258 staining for nuclear change, propidium iodode (PI) staining for cell cycle, monodansylcadaverine (MDC) staining for autophagy, and rodanmine 123 staining for mitochondrial membrane potential (MMP) were measured by fluorescence microscopy or flowcytometry. Apoptosis was determined by DNA ladder test. Protein kinase C (PKC) activity was detected by PKC assay kit. SA-β-galactosidase assay was used to detect senescence. Protein expression was examined by western blot. Results: PAB inhibited L929 cell growth in time-and dose-dependent manner. At 12 h, 80 μmol/L PAB induced obvious mitotic arrest; at 24 h, PAB began to induce autophagy; at 36 h, cell-treated with PAB slip into G1 cell cycle; and 3 d PAB induced senescence. In time sequence PAB induced firstly cell cycle arrest, then autophagy, then slippage into G1 phase, lastly senescence. Senescent cells had high level of autophagy, inhibiting autophagy led to apoptosis, and no senescence. PAB activated PKC activity to induce cell cycle arrest, autophagy and senescence, inhibiting PKC activity suppressed cell cycle arrest, autophagy and senescence. Conclusion: PAB induced cell cycle arrest, autophagy and senescence in murine fibrosarcoma L929 cell through PKC. PMID:23630435

A Vitex agnus-castus extract inhibits cell growth and induces apoptosis in prostate epithelial cell lines.

PubMed

Weisskopf, M; Schaffner, W; Jundt, G; Sulser, T; Wyler, S; Tullberg-Reinert, H

2005-10-01

Extracts of Vitex agnus-castus fruits (VACF) are described to have beneficial effects on disorders related to hyperprolactinemia (cycle disorders, premenstrual syndrome). A VACF extract has recently been shown to exhibit antitumor activities in different human cancer cell lines. In the present study, we explored the antiproliferative effects of a VACF extract with a particular focus on apoptosis-inducing and potential cytotoxic effects. Three different human prostate epithelial cell lines (BPH-1, LNCaP, PC-3) representing different disease stages and androgen responsiveness were chosen. The action of VACF on cell viability was assessed using the WST-8-tetrazolium assay. Cell proliferation in cells receiving VACF alone or in combination with a pan-caspase inhibitor (Z-VAD-fmk) was quantified using a Crystal Violet assay. Flow cytometric cell cycle analysis and measurement of DNA fragmentation using an ELISA method were used for studying the induction of apoptosis. Lactate dehydrogenase (LDH) activity was determined as a marker of cytotoxicity. The extract inhibited proliferation of all three cell lines in a concentration-dependent manner with IC (50) values below 10 microg/mL after treatment for 48 h. Cell cycle analysis and DNA fragmentation assays suggest that part of the cells were undergoing apoptosis. The VACF-induced decrease in cell number was partially inhibited by Z-VAD-fmk, indicating a caspase-dependent apoptotic cell death. However, the concentration-dependent LDH activity of VACF treated cells indicated cytotoxic effects as well. These data suggest that VACF contains components that inhibit proliferation and induce apoptosis in human prostate epithelial cell lines. The extract may be useful for the prevention and/or treatment not only of benign prostatic hyperplasia but also of human prostate cancer.

Inhibition of NF-kappaB-mediated transcription and induction of apoptosis in human breast cancer cells by epoxypseudoisoeugenol-2-methyl butyrate.

PubMed

Ma, Guoyi; Tabanca, Nurhayat; Husnu Can Baser, K; Kirimer, Nese; Pasco, David S; Khan, Ikhlas A; Khan, Shabana I

2009-03-01

Breast cancer is one of the most prevalent woman cancers. Genomic instability, accumulative mutations, and subsequent changes in intracellular signaling cascades play key roles in the development of human breast cancers. Activation of nuclear factor-kappaB (NF-kappaB) has been implicated in oncogenesis of breast cancers and is known to be associated with resistance to anticancer agents and apoptosis. Blocking NF-kappaB signaling may represent a therapeutic strategy in breast cancer therapy. The objective of this study is to investigate the in vitro effects of epoxypseudoisoeugenol-2-methyl butyrate (EPB), a phenylpropranoid isolated from Pimpinella corymbosa, on the activation of NF-kappaB, cell growth, cell cycle progression and apoptosis in MCF-7 (estrogen-dependent) and BT-549 (estrogen-independent) breast cancer cells. Transcriptional activity of NF-kappaB was measured by cell based reporter gene assay. Cell proliferation was determined by MTT assay. Cell cycle analysis was carried out by flow cytometry and apoptosis was observed by DAPI staining assy. EPB inhibited the NF-kappaB-mediated transcription activity induced by tumor necrosis factor-alpha (TNF-alpha) and phorbol myristate acetate (PMA) in MCF-7 cells. EPB also inhibited constitutive NF-kappaB transcriptional activity in BT-549 cells. EPB inhibited the proliferation of both MCF-7 and BT-549 cells in a concentration- and time-dependent manner. EPB induced cell cycle arrest in G(1)/G(0) phase and apoptosis in both MCF-7 and BT 549 cells. These in vitro results indicated that EPB has a potential for use against both hormone-dependent and hormone-independent breast cancers and its effects seem to be mediated by inhibiting the NF-kappaB activity.

Stage and season effects on cell cycle and apoptotic activities of germ cells and Sertoli cells during spermatogenesis in the spiny dogfish (Squalus acanthias).

PubMed

McClusky, L M

2005-01-01

To understand the processes involved in the spatial and temporal maturation of testicular cells in Squalus acanthias, we used standard morphometry, proliferating-cell nuclear antigen (PCNA) and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) immunohistochemistry. Except for immature spermatocysts (germinal zone, GZ; early-stage pre-meiotic, E-PrM), the number of cysts in all subsequent stages and the total number of cysts in the spermatogenic progression varied seasonally. The spermatogenic cycle spans about 2 years and is interrupted by germcell clone deletion via apoptosis at the mitosis-meiosis transition in April/May, manifesting as a zone of degeneration (ZD). Rate of displacement of the ZD across the testis diameter indicates that late-stage premeiotic (L-PrM) generations 12-13 require 9-10 months to reach the mature-spermatid stage. Also, the number of cysts completing spermatogenesis is approximately 4-5-fold less than the number that entered spermatogenesis proper 2 years earlier. Pronounced gonocytogenesis in the germinal ridge was coincident with ZD formation in April/May, but it was absent in the fall when mature spermatogonial and meiotic activities had resumed. Whereas strong Sertoli cell PCNA immunoreactivity dominated the GZ cyst cell-cycle activities throughout the year, except during the spring/summer months, the spermatogonial- and Sertoli-cell PCNA indices in E-PrM cysts were inversely related. PCNA immunoreactivity in spermatocytes was seasonal and dependent on the stage of meiosis. TUNEL labelling was limited to spermatogonia and increased stage-dependently in the PrM region (L-PrM = mid-stage PrM >E-PrM >GZ), correlating with ZD formation, in a season-dependent manner. Results imply that effects of normal regulatory factors in Squalus are stage- and process-specific.

5-demethyltangeretin inhibits human nonsmall cell lung cancer cell growth by inducing G2/M cell cycle arrest and apoptosis.

PubMed

Charoensinphon, Noppawat; Qiu, Peiju; Dong, Ping; Zheng, Jinkai; Ngauv, Pearline; Cao, Yong; Li, Shiming; Ho, Chi-Tang; Xiao, Hang

2013-12-01

Tangeretin (TAN) and 5-demethyltangeretin (5DT) are two closely related polymethoxyflavones found in citrus fruits. We investigated growth inhibitory effects on three human nonsmall cell lung cancer (NSCLC) cells. Cell viability assay demonstrated that 5DT inhibited NSCLC cell growth in a time- and dose-dependent manner, and IC50 s of 5DT were 79-fold, 57-fold, and 56-fold lower than those of TAN in A549, H460, and H1299 cells, respectively. Flow cytometry analysis showed that 5DT induced extensive G2/M cell cycle arrest and apoptosis in NSCLC cells, while TAN at tenfold higher concentrations did not. The apoptosis induced by 5DT was further confirmed by activation of caspase-3 and cleavage of PARP. Moreover, 5DT dose-dependently upregulated p53 and p21(Cip1/Waf1), and downregulated Cdc-2 (Cdk-1) and cyclin B1. HPLC analysis revealed that the intracellular levels of 5DT in NSCLC cells were 2.7-4.9 fold higher than those of TAN after the cells were treated with 5DT or TAN at the same concentration. Our results demonstrated that 5DT inhibited NSCLC cell growth by inducing G2/M cell cycle arrest and apoptosis. These effects were much stronger than those produced by TAN, which is partially due to the higher intracellular uptake of 5DT than TAN. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Resveratrol induces cell cycle arrest and apoptosis in human eosinophils from asthmatic individuals.

PubMed

Hu, Xin; Wang, Jing; Xia, Yu; Simayi, Mihereguli; Ikramullah, Syed; He, Yuanbing; Cui, Shihong; Li, Shuang; Wushouer, Qimanguli

2016-12-01

Eosinophils exert a number of inflammatory effects through the degranulation and release of intracellular mediators, and are considered to be key effector cells in allergic disorders, including asthma. In order to investigate the regulatory effects of the natural polyphenol, resveratrol, on eosinophils derived from asthmatic individuals, the cell counting Kit‑8 assay and flow cytometry analysis were used to determine cell proliferation and cell cycle progression in these cells, respectively. Cellular apoptosis was detected using annexin V-fluorescein isothiocyanate/propidium iodide double‑staining. The protein expression levels of p53, p21, cyclin‑dependent kinase 2 (CDK2), cyclin A, cyclin E, Bim, B‑cell lymphoma (Bcl)‑2 and Bcl‑2‑associated X protein (Bax) were measured by western blot analysis following resveratrol treatment. The results indicated that resveratrol effectively suppressed the proliferation of eosinophils from asthmatic patients in a concentration‑ and time‑dependent manner. In addition, resveratrol was observed to arrest cell cycle progression in G1/S phase by increasing the protein expression levels of p53 and p21, and concurrently reducing the protein expression levels of CDK2, cyclin A and cyclin E. Furthermore, resveratrol treatment significantly induced apoptosis in eosinophils, likely through the upregulation of Bim and Bax protein expression levels and the downregulation of Bcl‑2 protein expression. These findings suggested that resveratrol may be a potential agent for the treatment of asthma by decreasing the number of eosinophils.

General Protein Diffusion Barriers create Compartments within Bacterial Cells

PubMed Central

Schlimpert, Susan; Klein, Eric A.; Briegel, Ariane; Hughes, Velocity; Kahnt, Jörg; Bolte, Kathrin; Maier, Uwe G.; Brun, Yves V.; Jensen, Grant J.; Gitai, Zemer; Thanbichler, Martin

2013-01-01

SUMMARY In eukaryotes, the differentiation of cellular extensions such as cilia or neuronal axons depends on the partitioning of proteins to distinct plasma membrane domains by specialized diffusion barriers. However, examples of this compartmentalization strategy are still missing for prokaryotes, although complex cellular architectures are widespread among this group of organisms. This study reveals the existence of a protein-mediated membrane diffusion barrier in the stalked bacterium Caulobacter crescentus. We show that the Caulobacter cell envelope is compartmentalized by macromolecular complexes that prevent the exchange of both membrane and soluble proteins between the polar stalk extension and the cell body. The barrier structures span the cross-sectional area of the stalk and comprise at least four proteins that assemble in a cell cycle-dependent manner. Their presence is critical for cellular fitness, as they minimize the effective cell volume, allowing faster adaptation to environmental changes that require de novo synthesis of envelope proteins. PMID:23201141

Membrane tension controls adhesion positioning at the leading edge of cells

PubMed Central

Pontes, Bruno; Gole, Laurent; Kosmalska, Anita Joanna; Tam, Zhi Yang; Luo, Weiwei; Kan, Sophie; Viasnoff, Virgile; Roca-Cusachs, Pere; Tucker-Kellogg, Lisa

2017-01-01

Cell migration is dependent on adhesion dynamics and actin cytoskeleton remodeling at the leading edge. These events may be physically constrained by the plasma membrane. Here, we show that the mechanical signal produced by an increase in plasma membrane tension triggers the positioning of new rows of adhesions at the leading edge. During protrusion, as membrane tension increases, velocity slows, and the lamellipodium buckles upward in a myosin II–independent manner. The buckling occurs between the front of the lamellipodium, where nascent adhesions are positioned in rows, and the base of the lamellipodium, where a vinculin-dependent clutch couples actin to previously positioned adhesions. As membrane tension decreases, protrusion resumes and buckling disappears, until the next cycle. We propose that the mechanical signal of membrane tension exerts upstream control in mechanotransduction by periodically compressing and relaxing the lamellipodium, leading to the positioning of adhesions at the leading edge of cells. PMID:28687667

Boric acid inhibits human prostate cancer cell proliferation.

PubMed

Barranco, Wade T; Eckhert, Curtis D

2004-12-08

The role of boron in biology includes coordinated regulation of gene expression in mixed bacterial populations and the growth and proliferation of higher plants and lower animals. Here we report that boric acid, the dominant form of boron in plasma, inhibits the proliferation of prostate cancer cell lines, DU-145 and LNCaP, in a dose-dependent manner. Non-tumorigenic prostate cell lines, PWR-1E and RWPE-1, and the cancer line PC-3 were also inhibited, but required concentrations higher than observed human blood levels. Studies using DU-145 cells showed that boric acid induced a cell death-independent proliferative inhibition, with little effect on cell cycle stage distribution and mitochondrial function.

Vx-770 potentiates CFTR function by promoting decoupling between the gating cycle and ATP hydrolysis cycle.

PubMed

Jih, Kang-Yang; Hwang, Tzyh-Chang

2013-03-12

Vx-770 (Ivacaftor), a Food and Drug Administration (FDA)-approved drug for clinical application to patients with cystic fibrosis (CF), shifts the paradigm from conventional symptomatic treatments to therapeutics directly tackling the root of the disease: functional defects of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel caused by pathogenic mutations. The underlying mechanism for the action of Vx-770 remains elusive partly because this compound not only increases the activity of wild-type (WT) channels whose gating is primarily controlled by ATP binding/hydrolysis, but also improves the function of G551D-CFTR, a disease-associated mutation that abolishes CFTR's responsiveness to ATP. Here we provide a unified theory to account for this dual effect of Vx-770. We found that Vx-770 enhances spontaneous, ATP-independent activity of WT-CFTR to a similar magnitude as its effects on G551D channels, a result essentially explaining Vx-770's effect on G551D-CFTR. Furthermore, Vx-770 increases the open time of WT-CFTR in an [ATP]-dependent manner. This distinct kinetic effect is accountable with a newly proposed CFTR gating model depicting an [ATP]-dependent "reentry" mechanism that allows CFTR shuffling among different open states by undergoing multiple rounds of ATP hydrolysis. We further examined the effect of Vx-770 on R352C-CFTR, a unique mutant that allows direct observation of hydrolysis-triggered gating events. Our data corroborate that Vx-770 increases the open time of WT-CFTR by stabilizing a posthydrolytic open state and thereby fosters decoupling between the gating cycle and ATP hydrolysis cycle. The current study also suggests that this unique mechanism of drug action can be further exploited to develop strategies that enhance the function of CFTR.

Vx-770 potentiates CFTR function by promoting decoupling between the gating cycle and ATP hydrolysis cycle

PubMed Central

Jih, Kang-Yang; Hwang, Tzyh-Chang

2013-01-01

Vx-770 (Ivacaftor), a Food and Drug Administration (FDA)-approved drug for clinical application to patients with cystic fibrosis (CF), shifts the paradigm from conventional symptomatic treatments to therapeutics directly tackling the root of the disease: functional defects of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel caused by pathogenic mutations. The underlying mechanism for the action of Vx-770 remains elusive partly because this compound not only increases the activity of wild-type (WT) channels whose gating is primarily controlled by ATP binding/hydrolysis, but also improves the function of G551D-CFTR, a disease-associated mutation that abolishes CFTR’s responsiveness to ATP. Here we provide a unified theory to account for this dual effect of Vx-770. We found that Vx-770 enhances spontaneous, ATP-independent activity of WT-CFTR to a similar magnitude as its effects on G551D channels, a result essentially explaining Vx-770’s effect on G551D-CFTR. Furthermore, Vx-770 increases the open time of WT-CFTR in an [ATP]-dependent manner. This distinct kinetic effect is accountable with a newly proposed CFTR gating model depicting an [ATP]-dependent “reentry” mechanism that allows CFTR shuffling among different open states by undergoing multiple rounds of ATP hydrolysis. We further examined the effect of Vx-770 on R352C-CFTR, a unique mutant that allows direct observation of hydrolysis-triggered gating events. Our data corroborate that Vx-770 increases the open time of WT-CFTR by stabilizing a posthydrolytic open state and thereby fosters decoupling between the gating cycle and ATP hydrolysis cycle. The current study also suggests that this unique mechanism of drug action can be further exploited to develop strategies that enhance the function of CFTR. PMID:23440202

Silkworm Pupa Protein Hydrolysate Induces Mitochondria-Dependent Apoptosis and S Phase Cell Cycle Arrest in Human Gastric Cancer SGC-7901 Cells.

PubMed

Li, Xiaotong; Xie, Hongqing; Chen, Yajie; Lang, Mingzi; Chen, Yuyin; Shi, Liangen

2018-03-28

Silkworm pupae ( Bombyx mori ) are a high-protein nutrition source consumed in China since more than 2 thousand years ago. Recent studies revealed that silkworm pupae have therapeutic benefits to treat many diseases. However, the ability of the compounds of silkworm pupae to inhibit tumourigenesis remains to be elucidated. Here, we separated the protein of silkworm pupae and performed alcalase hydrolysis. Silkworm pupa protein hydrolysate (SPPH) can specifically inhibit the proliferation and provoke abnormal morphologic features of human gastric cancer cells SGC-7901 in a dose- and time-dependent manner. Moreover, flow cytometry indicated that SPPH can induce apoptosis and arrest the cell-cycle in S phase. Furthermore, SPPH was shown to provoke accumulation of reactive oxygen species (ROS) and depolarization of mitochondrial membrane potential. Western blotting analysis indicated that SPPH inhibited Bcl-2 expression and promoted Bax expression, and subsequently induced apoptosis-inducing factor and cytochrome C release, which led to the activation of initiator caspase-9 and executioner caspase-3, cleavage of poly (ADP-ribose) polymerase (PARP), eventually caused cell apoptosis. Moreover, SPPH-induced S-phase arrest was mediated by up-regulating the expression of E2F1 and down-regulating those of cyclin E, CDK2 and cyclin A2. Transcriptome sequencing and gene set enrichment analysis (GSEA) also revealed that SPPH treatment could affect gene expression and pathway regulation related to tumourigenesis, apoptosis and cell cycle. In summary, our results suggest that SPPH could specifically suppress cell growth of SGC-7901 through an intrinsic apoptotic pathway, ROS accumulation and cell cycle arrest, and silkworm pupae have a potential to become a source of anticancer agents in the future.

Jasmonate Controls Leaf Growth by Repressing Cell Proliferation and the Onset of Endoreduplication while Maintaining a Potential Stand-By Mode1[W][OA

PubMed Central

Noir, Sandra; Bömer, Moritz; Takahashi, Naoki; Ishida, Takashi; Tsui, Tjir-Li; Balbi, Virginia; Shanahan, Hugh; Sugimoto, Keiko; Devoto, Alessandra

2013-01-01

Phytohormones regulate plant growth from cell division to organ development. Jasmonates (JAs) are signaling molecules that have been implicated in stress-induced responses. However, they have also been shown to inhibit plant growth, but the mechanisms are not well understood. The effects of methyl jasmonate (MeJA) on leaf growth regulation were investigated in Arabidopsis (Arabidopsis thaliana) mutants altered in JA synthesis and perception, allene oxide synthase and coi1-16B (for coronatine insensitive1), respectively. We show that MeJA inhibits leaf growth through the JA receptor COI1 by reducing both cell number and size. Further investigations using flow cytometry analyses allowed us to evaluate ploidy levels and to monitor cell cycle progression in leaves and cotyledons of Arabidopsis and/or Nicotiana benthamiana at different stages of development. Additionally, a novel global transcription profiling analysis involving continuous treatment with MeJA was carried out to identify the molecular players whose expression is regulated during leaf development by this hormone and COI1. The results of these studies revealed that MeJA delays the switch from the mitotic cell cycle to the endoreduplication cycle, which accompanies cell expansion, in a COI1-dependent manner and inhibits the mitotic cycle itself, arresting cells in G1 phase prior to the S-phase transition. Significantly, we show that MeJA activates critical regulators of endoreduplication and affects the expression of key determinants of DNA replication. Our discoveries also suggest that MeJA may contribute to the maintenance of a cellular “stand-by mode” by keeping the expression of ribosomal genes at an elevated level. Finally, we propose a novel model for MeJA-regulated COI1-dependent leaf growth inhibition. PMID:23439917

Analysis of Septin Reorganization at Cytokinesis Using Polarized Fluorescence Microscopy

PubMed Central

McQuilken, Molly; Jentzsch, Maximilian S.; Verma, Amitabh; Mehta, Shalin B.; Oldenbourg, Rudolf; Gladfelter, Amy S.

2017-01-01

Septins are conserved filament-forming proteins that act in diverse cellular processes. They closely associate with membranes and, in some systems, components of the cytoskeleton. It is not well understood how filaments assemble into higher-order structures in vivo or how they are remodeled throughout the cell cycle. In the budding yeast S. cerevisiae, septins are found through most of the cell cycle in an hourglass organization at the mother-bud neck until cytokinesis when the collar splits into two rings that disassemble prior to the next cell cycle. Experiments using polarized fluorescence microscopy have suggested that septins are arranged in ordered, paired filaments in the hourglass and undergo a coordinated 90° reorientation during splitting at cytokinesis. This apparent reorganization could be due to two orthogonal populations of filaments disassembling and reassembling or being preferentially retained at cytokinesis. In support of this idea, we report a decrease in septin concentration at the mother-bud neck during cytokinesis consistent with other reports and the timing of the decrease depends on known septin regulators including the Gin4 kinase. We took a candidate-based approach to examine what factors control reorientation during splitting and used polarized fluorescence microscopy to screen mutant yeast strains deficient in septin interacting proteins. Using this method, we have linked known septin regulators to different aspects of the assembly, stability, and reorganization of septin assemblies. The data support that ring splitting requires Gin4 activity and an anillin-like protein Bud4, and normal accumulation of septins at the ring requires phosphorylation of Shs1. We found distinct regulatory requirements for septin organization in the hourglass compared to split rings. We propose that septin subpopulations can vary in their localization and assembly/disassembly behavior in a cell-cycle dependent manner at cytokinesis. PMID:28516085

Bilirubin Inhibits Neointima Formation and Vascular Smooth Muscle Cell Proliferation and Migration

PubMed Central

Peyton, Kelly J.; Shebib, Ahmad R.; Azam, Mohammad A.; Liu, Xiao-ming; Tulis, David A.; Durante, William

2012-01-01

Bilirubin is a heme metabolite generated by the concerted action of the enzymes heme oxygenase and biliverdin reductase. Although long considered a toxic byproduct of heme catabolism, recent preclinical, and clinical studies indicate the bilirubin exerts beneficial effects in the circulation. In the present study, we determined whether local administration of bilirubin attenuates neointima formation following injury of rat carotid arteries. In addition, the ability of bilirubin to regulate the proliferation and migration of human arterial smooth muscle cells (SMCs) was investigated. Local perivascular administration of bilirubin immediately following balloon injury of rat carotid arteries significantly attenuated neointima formation. Bilirubin-mediated inhibition of neointimal thickening was associated with a significant decrease in ERK activity and cyclin D1 and A protein expression, and an increase in p21 and p53 protein expression in injured blood vessels. Treatment of human aortic SMCs with bilirubin inhibited proliferation and migration in a concentration-dependent manner without affecting cell viability. In addition, bilirubin resulted in a concentration-dependent increase in the percentage of cells in the G0/G1 phase of the cell cycle and this was paralleled by a decrease in the fraction of cells in the S and G2M phases of the cell cycle. Finally, bilirubin had no effect on mitochondrial function and ATP content of vascular SMCs. In conclusion, these studies demonstrate that bilirubin inhibits neointima formation after arterial injury and this is associated with alterations in the expression of cell cycle regulatory proteins. Furthermore, bilirubin blocks proliferation and migration of human arterial SMCs and arrests SMCs in the G0/G1 phase of the cell cycle. Bilirubin represents an attractive therapeutic agent in treating occlusive vascular disease. PMID:22470341

Anticancer effect of cucurbitacin B on MKN-45 cells via inhibition of the JAK2/STAT3 signaling pathway

PubMed Central

Xie, You-Li; Tao, Wen-Hui; Yang, Ti-Xiong; Qiao, Jian-Guo

2016-01-01

The aim of the present study was to investigate the effect of cucurbitacin B on MKN-45 gastric carcinoma cells. Cell proliferation was determined using a cell counting kit-8 assay, and commercial cell cycle and apoptosis analysis kits were used to determine the cell cycle by flow cytometry. The mRNA expression of genes which mediate cell cycle checkpoints and apoptosis was detected using reverse transcription-quantitative polymerase chain reaction, and a terminal deoxynucleotidyl transferase dUTP nick end labeling assay was used to determine apoptosis rate. Western blot analysis was used to detect the protein expression levels of JAK2/STAT3 signaling pathway-associated proteins. The presented data show that cucurbitacin B significantly inhibited the proliferation of MKN-45 cells in a dose- and time-dependent manner. In accordance with these findings, cucurbitacin B blocked the progression of the cell cycle from G0/G1 to S phase, which was confirmed by the mRNA expression analysis. Cucurbitacin B treatment significantly suppressed the expression of cyclin D1, cyclin E, cyclin-dependent kinase 4 (CDK4) and CDK2, while increasing the expression of p27. Cucurbitacin B also promoted cell apoptosis, as was determined by TUNEL assay and evaluation of mRNA expression. Further experiments suggested that the beneficial effect of cucurbitacin B on blocking the proliferation and inducing the apoptosis of MKN-45 cells may have been associated with suppression of the JAK2/STAT3 signaling pathway. Thus, the present results indicate that cucurbitacin B suppresses proliferation and promoted apoptosis of MKN-45 cells, which may be mediated by inhibition of the JAK2/STAT3 signaling pathway. Cucurbitacin B therefore may warrant further investigation as a feasible therapy for gastric carcinoma. PMID:27698776

MAP30 promotes apoptosis of U251 and U87 cells by suppressing the LGR5 and Wnt/β-catenin signaling pathway, and enhancing Smac expression

PubMed Central

Jiang, Yilin; Miao, Junjie; Wang, Dongliang; Zhou, Jingru; Liu, Bo; Jiao, Feng; Liang, Jiangfeng; Wang, Yangshuo; Fan, Cungang; Zhang, Qingjun

2018-01-01

Significant antitumor activity of Momordica anti-human immunodeficiency virus protein of 30 kDa (MAP30) purified from Momordica charantia has been the subject of previous research. However, the effective mechanism of MAP30 on malignant glioma cells has not yet been clarified. The aim of the present study was to investigate the effects and mechanism of MAP30 on U87 and U251 cell lines. A Cell Counting Kit-8 assay, wound healing assay and Transwell assay were used to detect the effects on U87 and U251 cells treated with different concentrations of MAP30 (0.5, 1, 2, 4, 8 and 16 µM) over different periods of time. Proliferation, migration and invasion of each cell line were markedly inhibited by MAP30 in a dose- and time-dependent manner. Flow cytometry and fluorescence staining demonstrated that apoptosis increased and the cell cycle was arrested in S-phase in the two investigated cell lines following MAP30 treatment. Western blot analysis demonstrated that leucine-rich-repeat-containing G-protein-coupled receptor 5 (LGR5) expression and key proteins in the Wnt/β-catenin signaling pathway were apparently decreased, whereas second mitochondria-derived activator of caspase (Smac) protein expression significantly increased with MAP30 treatment in the same manner. These results suggest that MAP30 markedly induces apoptosis in U87 and U251 cell lines by suppressing LGR5 and the Wnt/β-catenin signaling pathway, and enhancing Smac expression in a dose- and time-dependent manner. PMID:29556310

The silencing of Pokemon attenuates the proliferation of hepatocellular carcinoma cells in vitro and in vivo by inhibiting the PI3K/Akt pathway.

PubMed

Lin, Chan-Chan; Zhou, Jing-Ping; Liu, Yun-Peng; Liu, Jing-Jing; Yang, Xiao-Ning; Jazag, Amarsanaa; Zhang, Zhi-Ping; Guleng, Bayasi; Ren, Jian-Lin

2012-01-01

Pokemon (POK erythroid myeloid ontogenic factor), which belongs to the POK protein family, is also called LRF, OCZF and FBI-1. As a transcriptional repressor, Pokemon assumes a critical function in cellular differentiation and oncogenesis. Our study identified an oncogenic role for Pokemon in human hepatocellular carcinoma (HCC). We successfully established human HepG2 and Huh-7 cell lines in which Pokemon was stably knocked down. We demonstrated that Pokemon silencing inhibited cell proliferation and migration. Pokemon knockdown inhibited the PI3K/Akt and c-Raf/MEK/ERK pathways and modulated the expression of various cell cycle regulators in HepG2 and Huh-7 cells. Therefore, Pokemon may also be involved in cell cycle progression in these cells. We confirmed that Pokemon silencing suppresses hepatocellular carcinoma growth in tumor xenograft mice. These results suggest that Pokemon promotes cell proliferation and migration in hepatocellular carcinoma and accelerates tumor development in an Akt- and ERK-signaling-dependent manner.

Arsenic trioxide-mediated growth inhibition in gallbladder carcinoma cells via down-regulation of Cyclin D1 transcription mediated by Sp1 transcription factor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ai, Zhilong; Lu, Weiqi; Ton, Saixiong

2007-08-31

Gallbladder carcinoma (GBC), an aggressive and mostly lethal malignancy, is known to be resistant to a number of drug stimuli. Here, we demonstrated that arsenic trioxide inhibited the proliferation of gallbladder carcinoma in vivo and in vitro as well as the transcription of cell cycle-related protein Cyclin D1. And, Cyclin D1 overexpression inhibited the negative role of arsenic trioxide in cell cycle progression. We further explored the mechanisms by which arsenic trioxide affected Cyclin D1 transcription and found that the Sp1 transcription factor was down-regulated by arsenic trioxide, with a corresponding decrease in Cyclin D1 promoter activity. Taken together, thesemore » results suggested that arsenic trioxide inhibited gallbladder carcinoma cell proliferation via down-regulation of Cyclin D1 transcription in a Sp1-dependent manner, which provided a new mechanism of arsenic trioxide-involved cell proliferation and may have important therapeutic implications in gallbladder carcinoma patients.« less

Proteasome-mediated degradation of cell division cycle 25C and cyclin-dependent kinase 1 in phenethyl isothiocyanate-induced G2-M-phase cell cycle arrest in PC-3 human prostate cancer cells.

PubMed

Xiao, Dong; Johnson, Candace S; Trump, Donald L; Singh, Shivendra V

2004-05-01

Phenethyl isothiocyanate (PEITC), a constituent of many cruciferous vegetables, offers significant protection against cancer in animals induced by a variety of carcinogens. The present study demonstrates that PEITC suppresses proliferation of PC-3 cells in a dose-dependent manner by causing G(2)-M-phase cell cycle arrest and apoptosis. Interestingly, phenyl isothiocyanate (PITC), which is a structural analogue of PEITC but lacks the -CH(2) spacers that link the aromatic ring to the -N=C=S group, neither inhibited PC-3 cell viability nor caused cell cycle arrest or apoptosis. These results indicated that even a subtle change in isothiocyanate (ITC) structure could have a significant impact on its biological activity. The PEITC-induced cell cycle arrest was associated with a >80% reduction in the protein levels of cyclin-dependent kinase 1 (Cdk1) and cell division cycle 25C (Cdc25C; 24 h after treatment with 10 micro M PEITC), which led to an accumulation of Tyr(15) phosphorylated (inactive) Cdk1. On the other hand, PITC treatment neither reduced protein levels of Cdk1 or Cdc25C nor affected Cdk1 phosphorylation. The PEITC-induced decline in Cdk1 and Cdc25C protein levels and cell cycle arrest were significantly blocked on pretreatment of PC-3 cells with proteasome inhibitor lactacystin. A 24 h exposure of PC-3 cells to 10 micro M PEITC, but not PITC, resulted in about 56% and 44% decrease in the levels of antiapoptotic proteins Bcl-2 and Bcl-X(L), respectively. However, ectopic expression of Bcl-2 failed to alter sensitivity of PC-3 cells to growth inhibition or apoptosis induction by PEITC. Treatment of cells with PEITC, but not PITC, also resulted in cleavage of procaspase-3, procaspase-9, and procaspase-8. Moreover, the PEITC-induced apoptosis was significantly attenuated in the presence of general caspase inhibitor and specific inhibitors of caspase-8 and caspase-9. In conclusion, our data indicate that PEITC-induced cell cycle arrest in PC-3 cells is likely due to proteasome-mediated degradation of Cdc25C and Cdk1, and ectopic expression of Bcl-2 fails to confer resistance to PEITC-induced apoptosis. Furthermore, the results of the present study point toward involvement of both caspase-8- and caspase-9-mediated pathways in apoptosis induction by PEITC.

The process of processing: exploring the validity of Neisser's perceptual cycle model with accounts from critical decision-making in the cockpit.

PubMed

Plant, Katherine L; Stanton, Neville A

2015-01-01

The perceptual cycle model (PCM) has been widely applied in ergonomics research in domains including road, rail and aviation. The PCM assumes that information processing occurs in a cyclical manner drawing on top-down and bottom-up influences to produce perceptual exploration and actions. However, the validity of the model has not been addressed. This paper explores the construct validity of the PCM in the context of aeronautical decision-making. The critical decision method was used to interview 20 helicopter pilots about critical decision-making. The data were qualitatively analysed using an established coding scheme, and composite PCMs for incident phases were constructed. It was found that the PCM provided a mutually exclusive and exhaustive classification of the information-processing cycles for dealing with critical incidents. However, a counter-cycle was also discovered which has been attributed to skill-based behaviour, characteristic of experts. The practical applications and future research questions are discussed. Practitioner Summary: This paper explores whether information processing, when dealing with critical incidents, occurs in the manner anticipated by the perceptual cycle model. In addition to the traditional processing cycle, a reciprocal counter-cycle was found. This research can be utilised by those who use the model as an accident analysis framework.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cho, Seong-Jun; Kang, Hana; Kim, Min Young

Purpose: To determine how low-dose ionizing radiation (LDIR) regulates B lympho-proliferation and its molecular mechanism related with Ikaros, transcription factor. Methods and Materials: Splenocytes and IM-9 cells were uniformly irradiated with various doses of a {sup 137}Cs γ-source, and cell proliferation was analyzed. To determine the LDIR-specific phosphorylation of Ikaros, immunoprecipitation and Western blot analysis were performed. To investigate the physiologic function of LDIR-mediatied Ikaros phosphorylation, Ikaros mutants at phosphorylation sites were generated, and cell cycle analysis was performed. Results: First, we found that LDIR enhances B lymphoblast proliferation in an Ikaros-dependent manner. Moreover, we found that LDIR elevates the phosphorylationmore » level of Ikaros protein. Interestingly, we showed that CK2 and AKT are involved in LDIR-induced Ikaros phosphorylation and capable of regulating DNA binding activity of Ikaros via specific phosphorylation. Finally, we identified LDIR-specific Ikaros phosphorylation sites at S391/S393 and showed that the Ikaros phosphorylations at these sites control Ikaros's ability to regulate G1/S cell cycle progression. Conclusion: Low-dose ionizing radiation specifically phosphorylates Ikaros protein at Ser 391/393 residues to regulate cell cycle progression in B lymphoblast.« less

Phloretin induces cell cycle arrest and apoptosis of human glioblastoma cells through the generation of reactive oxygen species.

PubMed

Liu, Yuanyuan; Fan, Chenghe; Pu, Lv; Wei, Cui; Jin, Haiqiang; Teng, Yuming; Zhao, Mingming; Yu, Albert Cheung Hoi; Jiang, Feng; Shu, Junlong; Li, Fan; Peng, Qing; Kong, Jian; Pan, Bing; Zheng, Lemin; Huang, Yining

2016-06-01

Phloretin, a flavonoid present in various plants, has been reported to exert anticarcinogenic effects. However, the mechanism of its chemo-preventive effect on human glioblastoma cells is not fully understood. This study aimed to investigate the molecular mechanism of phloretin and its associated chemo-preventive effect in human glioblastoma cells. The results indicate that phloretin inhibited cell proliferation by inducing cell cycle arrest at the G0-G1 phase and induced apoptosis of human glioblastoma cells. Phloretin-induced cell cycle arrest was associated with increased expression of p27 and decreased expression of cdk2, cdk4, cdk6, cyclinD and cyclinE. Moreover, the PI3K/AKT/mTOR signaling cascades were suppressed by phloretin in a dose-dependent manner. In addition, phloretin triggered the mitochondrial apoptosis pathway and generated reactive oxygen species (ROS). This was accompanied by the up-regulation of Bax, Bak and c-PARP and the down-regulation of Bcl-2. The antioxidant agents N-acetyl-L-cysteine and glutathione weakened the effect of phloretin on glioblastoma cells. In conclusion, these results demonstrate that phloretin exerts potent chemo-preventive activity in human glioblastoma cells through the generation of ROS.

Essential function of VCP/p97 in infection cycle of the nucleopolyhedrovirus AcMNPV in Spodoptera frugiperda Sf9 cells.

PubMed

Lyupina, Yulia V; Erokhov, Pavel A; Kravchuk, Oksana I; Finoshin, Alexander D; Abaturova, Svetlana B; Orlova, Olga V; Beljelarskaya, Svetlana N; Kostyuchenko, Margarita V; Mikhailov, Victor S

2018-06-08

The protein VCP/p97 (also named CDC48 and TER94) belongs to a type II subfamily of the AAA+ATPases and controls cellular proteostasis by acting upstream of proteasomes in the ubiquitin-proteasome protein degradation pathway. The function of VCP/p97 in the baculovirus infection cycle in insect cells remains unknown. Here, we identified VCP/p97 in the fall armyworm Spodoptera frugiperda (Sf9) cells and analyzed the replication of the Autographa californica multiple nucleopolyhedrovirus, AcMNPV, in Sf9 cells in which the VCP/p97 function was inhibited. The specific allosteric inhibitor of the VCP/p97 ATPase activity, NMS-873, did not deplete VCP/p97 in infected cells but caused a dose-dependent inhibition of viral DNA synthesis and efficiently suppressed expression of viral proteins and production of budded virions. NMS-873 caused accumulation of ubiquitinated proteins in a manner similar to the inhibitor of proteasome activity, Bortezomib. This suggests the essential function of VCP/p97 in the baculovirus infection cycle might be associated, at least in part, with the ubiquitin-proteasome system. Copyright © 2018 Elsevier B.V. All rights reserved.

Annexin V-induced rat Leydig cell proliferation involves Ect2 via RhoA/ROCK signaling pathway.

PubMed

Jing, Jun; Chen, Li; Fu, Hai-Yan; Fan, Kai; Yao, Qi; Ge, Yi-Feng; Lu, Jin-Chun; Yao, Bing

2015-03-24

This study investigated the effect of annexin V on the proliferation of primary rat Leydig cells and the potential mechanism. Our results showed that annexin V promoted rat Leydig cell proliferation and cell cycle progression in a dose- and time-dependent manner. Increased level of annexin V also enhanced Ect2 protein expression. However, siRNA knockdown of Ect2 attenuated annexin V-induced proliferation of rat Leydig cells. Taken together, these data suggest that increased level of annexin V induced rat Leydig cell proliferation and cell cycle progression via Ect2. Since RhoA activity was increased following Ect2 activation, we further investigated whether Ect2 was involved in annexin V-induced proliferation via the RhoA/ROCK pathway, and the results showed that annexin V increased RhoA activity too, and this effect was abolished by the knockdown of Ect2. Moreover, inhibition of the RhoA/ROCK pathway by a ROCK inhibitor, Y27632, also attenuated annexin V-induced proliferation and cell cycle progression. We thus conclude that Ect2 is involved in annexin V-induced rat Leydig cell proliferation through the RhoA/ROCK pathway.

Paris Saponin I Sensitizes Gastric Cancer Cell Lines to Cisplatin via Cell Cycle Arrest and Apoptosis.

PubMed

Song, Shuichuan; Du, Leiwen; Jiang, Hao; Zhu, Xinhai; Li, Jinhui; Xu, Ji

2016-10-18

BACKGROUND Dose-related toxicity is the major restriction of cisplatin and cisplatin-combination chemotherapy, and is a challenge for advanced gastric cancer treatment. We explored the possibility of using Paris saponin I as an agent to sensitize gastric cancer cells to cisplatin, and examined the underlying mechanism. MATERIAL AND METHODS Growth inhibition was detected by MTT assay. The cell cycle and apoptosis were detected using flow cytometry and Annexin V/PI staining. The P21waf1/cip1, Bcl-2, Bax, and caspase-3 protein expression were detected using Western blot analysis. RESULTS The results revealed that PSI sensitized gastric cancer cells to cisplatin, with low toxicity. The IC50 value of cisplatin in SGC-7901 cell lines was decreased when combined with PSI. PSI promoted cisplatin-induced G2/M phase arrest and apoptosis in a cisplatin concentration-dependent manner. Bcl-2 protein expression decreased, but Bax, caspase-3, and P21waf1/cip1 protein expression increased with PSI treatment. CONCLUSIONS The underlying mechanism of Paris saponin I may be related to targeting the apoptosis pathway and cell cycle blocking, which suggests that PSI is a potential therapeutic sensitizer for cisplatin in treating gastric cancer.

Aluminum oxide nanoparticles alter cell cycle progression through CCND1 and EGR1 gene expression in human mesenchymal stem cells.

PubMed

Periasamy, Vaiyapuri Subbarayan; Athinarayanan, Jegan; Alshatwi, Ali A

2016-05-01

Aluminum oxide nanoparticles (Al2 O3 -NPs) are important ceramic materials that have been used in a variety of commercial and industrial applications. However, the impact of acute and chronic exposure to Al2 O3 -NPs on the environment and on human health has not been well studied. In this investigation, we evaluated the cytotoxic effects of Al2 O3 -NPs on human mesenchymal stem cells (hMSCs) by using a cell viability assay and observing cellular morphological changes, analyzing cell cycle progression, and monitoring the expression of cell cycle response genes (PCNA, EGR1, E2F1, CCND1, CCNC, CCNG1, and CYCD3). The Al2 O3 -NPs reduced hMSC viability in a dose- and time-dependent manner. Nuclear condensation and fragmentation, chromosomal DNA fragmentation, and cytoplasmic vacuolization were observed in Al2 O3 -NP-exposed cells. The nuclear morphological changes indicated that Al2 O3 -NPs alter cell cycle progression and gene expression. The cell cycle distribution revealed that Al2 O3 -NPs cause cell cycle arrest in the sub-G0-G1 phase, and this is associated with a reduction in the cell population in the G2/M and G0/G1 phases. Moreover, Al2 O3 -NPs induced the upregulation of cell cycle response genes, including EGR1, E2F1, and CCND1. Our results suggested that exposure to Al2 O3 -NPs could cause acute cytotoxic effects in hMSCs through cell cycle regulatory genes. © 2015 International Union of Biochemistry and Molecular Biology, Inc.

A new indole-3-carbinol tetrameric derivative inhibits cyclin-dependent kinase 6 expression, and induces G1 cell cycle arrest in both estrogen-dependent and estrogen-independent breast cancer cell lines.

PubMed

Brandi, Giorgio; Paiardini, Mirko; Cervasi, Barbara; Fiorucci, Chiara; Filippone, Paolino; De Marco, Cinzia; Zaffaroni, Nadia; Magnani, Mauro

2003-07-15

Indole-3-carbinol (I3C), autolysis product of glucosinolates present in cruciferous vegetables, has been indicated as a promising agent in preventing the development and progression of breast cancer. I3C has been shown to inhibit the growth of human cancer cells in vitro and possesses anticarcinogenic activity in vivo. Because I3C is unstable and may be converted into many polymeric products in the digestive tract, it is not yet clear whether the biological activity observed can be attributed to I3C or some of its polymeric products. In this study we synthesized a stable I3C cyclic tetrameric derivative and investigated its effects on a panel of human breast cancer cell lines. The I3C tetramer suppressed the growth of both estrogen receptor (ER) -positive (MCF-7, 734B, and BT474) and ER-negative (BT20, MDA-MB-231, and BT539) human breast cancer cell lines, and it was found to induce G(1) cell cycle arrest in a dose-dependent manner without evidence of apoptosis, suggesting a growth arrest via a cytostatic mechanism. At the molecular level, the tetramer inhibited cyclin-dependent kinase (CDK) 6 expression and activity, induced an increase in the level of p27(kip1), and reduced the level of retinoblastoma protein expression. Contrarily to CDK6, the level of CDK4, the other kinase involved in the G(1) phase of the cell cycle, remains unchanged. Interestingly, the tetramer resulted about five times more active than I3C in suppressing the growth of human breast cancer cells. On the whole, our data suggest that the I3C tetrameric derivative is a novel lead inhibitor of breast cancer cell growth that may be a considered a new, promising therapeutic agent for both ER+ and ER- breast cancer.

Effects of magnolol on UVB-induced skin cancer development in mice and its possible mechanism of action.

PubMed

Chilampalli, Chandeshwari; Guillermo, Ruth; Zhang, Xiaoying; Kaushik, Radhey S; Young, Alan; Zeman, David; Hildreth, Michael B; Fahmy, Hesham; Dwivedi, Chandradhar

2011-10-20

Magnolol, a plant lignan isolated from the bark and seed cones of Magnolia officinalis, has been shown to have chemopreventive effects on chemically-induced skin cancer development. The objectives of this investigation are to study the anticarcinogenic effects of magnolol on UVB-induced skin tumor development in SKH-1 mice, a model relevant to humans, and determine the possible role of apoptosis and cell cycle arrest involved in the skin tumor development. UVB-induced skin carcinogenesis model in SKH-1 mice was used for determining the preventive effects of magnolol on skin cancer development. Western blottings and flow cytometric analysis were used to study the effects of magnolol on apoptosis and cell cycle. Magnolol pretreated groups (30, 60 μ g) before UVB treatments (30 mJ/cm2, 5 days/week) resulted in 27-55% reduction in tumor multiplicity as compared to control group in SKH-1 mice. Magnolol pretreatment increased the cleavage of caspase-8 and poly-(-ADP-ribose) polymerase (PARP), increased the expression of p21, a cell cycle inhibitor, and decreased the expression of proteins involved in the G2/M phase of cell cycle in skin samples from SKH-1 mice.Treatment of A431 cells with magnolol decreased cell viability and cell proliferation in a concentration dependent manner. Magnolol induced G2/M phase cell cycle arrest in A431 cells at 12 h with a decreased expression of cell cycle proteins such as cyclin B1, cyclin A, CDK4, Cdc2 and simultaneous increase in the expression of Cip/p21, a cyclin-dependent kinase inhibitor. Magnolol induced apoptosis in vivo and in vitro with an increased cleavage of caspase-8 and PARP. Phospho-signal transducers and activators of transcription 3 (Tyr705), B-Raf, p-MEK, and p-AKT were down-regulated, whereas phosphorylation of ERK was induced by magnolol in A431 cells. Magnolol pretreatments prevent UVB-induced skin cancer development by enhancing apoptosis, causing cell cycle arrest at G2/M phase, and affecting various signaling pathways. Magnolol could be a potentially safe and potent anticarcinogenic agent against skin cancer.

Effects of magnolol on UVB-induced skin cancer development in mice and its possible mechanism of action

PubMed Central

2011-01-01

Background Magnolol, a plant lignan isolated from the bark and seed cones of Magnolia officinalis, has been shown to have chemopreventive effects on chemically-induced skin cancer development. The objectives of this investigation are to study the anticarcinogenic effects of magnolol on UVB-induced skin tumor development in SKH-1 mice, a model relevant to humans, and determine the possible role of apoptosis and cell cycle arrest involved in the skin tumor development. Methods UVB-induced skin carcinogenesis model in SKH-1 mice was used for determining the preventive effects of magnolol on skin cancer development. Western blottings and flow cytometric analysis were used to study the effects of magnolol on apoptosis and cell cycle. Results Magnolol pretreated groups (30, 60 μ g) before UVB treatments (30 mJ/cm2, 5 days/week) resulted in 27-55% reduction in tumor multiplicity as compared to control group in SKH-1 mice. Magnolol pretreatment increased the cleavage of caspase-8 and poly-(-ADP-ribose) polymerase (PARP), increased the expression of p21, a cell cycle inhibitor, and decreased the expression of proteins involved in the G2/M phase of cell cycle in skin samples from SKH-1 mice. Treatment of A431 cells with magnolol decreased cell viability and cell proliferation in a concentration dependent manner. Magnolol induced G2/M phase cell cycle arrest in A431 cells at 12 h with a decreased expression of cell cycle proteins such as cyclin B1, cyclin A, CDK4, Cdc2 and simultaneous increase in the expression of Cip/p21, a cyclin-dependent kinase inhibitor. Magnolol induced apoptosis in vivo and in vitro with an increased cleavage of caspase-8 and PARP. Phospho-signal transducers and activators of transcription 3 (Tyr705), B-Raf, p-MEK, and p-AKT were down-regulated, whereas phosphorylation of ERK was induced by magnolol in A431 cells. Conclusions Magnolol pretreatments prevent UVB-induced skin cancer development by enhancing apoptosis, causing cell cycle arrest at G2/M phase, and affecting various signaling pathways. Magnolol could be a potentially safe and potent anticarcinogenic agent against skin cancer. PMID:22014088

The Growth of SGC-7901 Tumor Xenografts Was Suppressed by Chinese Bayberry Anthocyanin Extract through Upregulating KLF6 Gene Expression.

PubMed

Wang, Yue; Zhang, Xia-Nan; Xie, Wen-Hua; Zheng, Yi-Xiong; Cao, Jin-Ping; Cao, Pei-Rang; Chen, Qing-Jun; Li, Xian; Sun, Chong-de

2016-09-27

To investigate the antitumor effect of anthocyanins extracted from Chinese bayberry fruit ( Myrica rubra Sieb. et Zucc.), a nude mouse tumor xenograft model was established. Treatments with C3G (cyanidin-3-glucoside, an anthocyanin) significantly suppressed the growth of SGC-7901 tumor xenografts in a dose-dependent manner. Immunohistochemical staining showed a significant increase in p21 expression, indicating that the cell cycle of tumor xenografts was inhibited. qPCR screening showed that C3G treatment up-regulated the expression of the KLF6 gene, which is an important tumor suppressor gene inactivated in many human cancers. Western blot showed that C3G treatments markedly increased KLF6 and p21 protein levels, inhibited CDK4 and Cyclin D1 expression, but did not notably change the expression of p53. These results indicated that KLF6 up-regulates p21 in a p53-independent manner and significantly reduces tumor proliferation. This study provides important information for the possible mechanism of C3G-induced antitumor activity against gastric adenocarcinoma in vivo.

The Growth of SGC-7901 Tumor Xenografts Was Suppressed by Chinese Bayberry Anthocyanin Extract through Upregulating KLF6 Gene Expression

PubMed Central

Wang, Yue; Zhang, Xia-nan; Xie, Wen-hua; Zheng, Yi-xiong; Cao, Jin-ping; Cao, Pei-rang; Chen, Qing-jun; Li, Xian; Sun, Chong-de

2016-01-01

To investigate the antitumor effect of anthocyanins extracted from Chinese bayberry fruit (Myrica rubra Sieb. et Zucc.), a nude mouse tumor xenograft model was established. Treatments with C3G (cyanidin-3-glucoside, an anthocyanin) significantly suppressed the growth of SGC-7901 tumor xenografts in a dose-dependent manner. Immunohistochemical staining showed a significant increase in p21 expression, indicating that the cell cycle of tumor xenografts was inhibited. qPCR screening showed that C3G treatment up-regulated the expression of the KLF6 gene, which is an important tumor suppressor gene inactivated in many human cancers. Western blot showed that C3G treatments markedly increased KLF6 and p21 protein levels, inhibited CDK4 and Cyclin D1 expression, but did not notably change the expression of p53. These results indicated that KLF6 up-regulates p21 in a p53-independent manner and significantly reduces tumor proliferation. This study provides important information for the possible mechanism of C3G-induced antitumor activity against gastric adenocarcinoma in vivo. PMID:27690088

Novel Derivative of Benzofuran Induces Cell Death Mostly by G2/M Cell Cycle Arrest through p53-dependent Pathway but Partially by Inhibition of NF-κB*

PubMed Central

Manna, Sunil K.; Bose, Julie S.; Gangan, Vijay; Raviprakash, Nune; Navaneetha, Thota; Raghavendra, Pongali B.; Babajan, Banaganapalli; Kumar, Chitta S.; Jain, Swatantra K.

2010-01-01

The Dracaena resin is widely used in traditional medicine as an anticancer agent, and benzofuran lignan is the active component. In this report, we provide evidence that the synthetic derivative of benzofuran lignan (Benfur) showed antitumor activities. It induced apoptosis in p53-positive cells. Though it inhibited endotoxin-induced nuclear factor κB (NF-κB) activation in both p53-positive and -negative cells, the activation of caspase 3 was observed in p53-positive cells. It showed partial cell death effect in both p53-positive and -negative cells through inhibition of NF-κB. Cell cycle analysis using flow cytometry showed that treatment with this novel benozofuran lignan derivative to Jurkat T-cells, but not U-937 cells, resulted in a G2/M arrest in a dose- and time-dependent manner. It increased amounts of p21, p27, and cyclin B, but not phospho-Rb through p53 nuclear translocation in Jurkat T-cells, but not in U-937 cells. It inhibited amounts of MDM2 (murine double minute 2) by repressing the transcription factor Sp1, which was also proved in silico. It induced cell death in tumor cells, but not in primary T-cells. Overall, our data suggest that Benfur-mediated cell death is partially dependent upon NF-κB, but predominantly dependent on p53. Thus, this novel benzofuran lignan derivative can be effective chemopreventive or chemotherapeutic agent against malignant T-cells. PMID:20472557

The UbL-UBA Ubiquilin4 protein functions as a tumor suppressor in gastric cancer by p53-dependent and p53-independent regulation of p21.

PubMed

Huang, Shengkai; Li, Yan; Yuan, Xinghua; Zhao, Mei; Wang, Jia; Li, You; Li, Yuan; Lin, Hong; Zhang, Qiao; Wang, Wenjie; Li, Dongdong; Dong, Xin; Li, Lanfen; Liu, Min; Huang, Weiyan; Huang, Changzhi

2018-06-13

Ubiquilin4 (Ubqln4), a member of the UbL-UBA protein family, serves as an adaptor in the degradation of specific substrates via the proteasomal pathway. However, the biological function of Ubqln4 remains largely unknown, especially in cancer. Here, we reported that Ubqln4 was downregulated in gastric cancer tissues and functioned as a tumor suppressor by inhibiting gastric cancer cell proliferation in vivo and in vitro. Overexpression of Ubqln4-induced cellular senescence and G1-S cell cycle arrest in gastric cancer cells and activated the p53/p21 axis. Moreover, Ubqln4 regulated p21 through both p53-dependent and p53-independent manners. Ubqln4 interacted with RNF114, an E3 ubiquitin ligase of p21, and negatively regulated its expression level, which in turn stabilized p21 by attenuating proteasomal degradation of p21. These effects of Ubqln4 were partly abrogated in gastric cancer cells upon silencing of p21. Our findings not only establish the anti-tumor potential of Ubqln4 in gastric cancer but also reveal a role for Ubqln4 in regulation of the cell cycle and cellular senescence via stabilizing p21.

Quercetin inhibits the invasion and mobility of murine melanoma B16-BL6 cells through inducing apoptosis via decreasing Bcl-2 expression.

PubMed

Zhang, X; Xu, Q; Saiki, I

2000-01-01

Quercetin has been known to have anti-tumor and anti-oxidation activities. In the present study, we have investigated its in vitro anti-metastatic activity. Quercetin inhibited the invasion and mobility of murine melanoma B16-BL6 cells in a dose-dependent manner but did not affect their adhesion to either laminin, fibronectin, or type VI collagen. Moreover, quercetin significantly inhibited the proliferation of B16-BL6 cells only in the case of time incubation longer than 48 h. Quercetin dose-dependently decreased the cell rates in S and G2-M phases of cell cycle. The effect of quercetin to cause a remarkable apoptosis of B16-BL6 cells was also demonstrated by flow cytometric assay as well as DNA fragmentation with a typical 180-bp ladder band in agarose electrophoresis and a quantitative analysis. Furthermore, quercetin markedly inhibited the expression of anti-apoptotic protein Bcl-2 but hardly influenced Bcl-XL. These results suggest that the inhibition of quercetin on invasiveness and migration of B16-BL6 cells are closely associated with the arrest of cell cycle as well as the induction of apoptosis by decreasing the Bcl-2 expression.

2′,4′-dihydroxychalcone, a flavonoid isolated from Herba oxytropis, suppresses PC-3 human prostate cancer cell growth by induction of apoptosis

PubMed Central

SHENG, YUQING; ZOU, MINGCHANG; WANG, YAN; LI, QIHENG

2015-01-01

Natural products are a promising source for the development of novel cancer therapies, due to their potential effectiveness and low toxicity profiles. As a main component of Herba oxytropis, 2′,4′-dihydroxychalcone (TFC) is known to demonstrate anti-tumor activity in vitro. In the present study, TFC was found to potently inhibit proliferation and induce apoptosis in PC-3 human prostate cancer cells in a dose-dependent manner. The results demonstrated that the induction of apoptosis is associated with cell cycle arrest at the G0/G1 phase and activation of caspase-3/-7. Additional mechanistic studies of two biomarkers, phosphatase and tensin homolog (PTEN) and cyclin-dependent kinase inhibitor 1B (p27Kip1), in prostate cancer revealed that TFC treatment significantly upregulated the expression of PTEN and p27Kip1. The findings of the present study indicate that TFC-induced apoptosis in PC-3 cells via upregulation of PTEN and p27Kip1, which results in cell cycle arrest in G0/G1 phase, activation of caspase-3/-7 and induction of apoptosis. Therefore, TFC may be a potential compound for human prostate cancer therapy. PMID:26788200

A potent transrepression domain in the retinoblastoma protein induces a cell cycle arrest when bound to E2F sites.

PubMed Central

Sellers, W R; Rodgers, J W; Kaelin, W G

1995-01-01

An intact T/E1A-binding domain (the pocket) is necessary, but not sufficient, for the retinoblastoma protein (RB) to bind to DNA-protein complexes containing E2F and for RB to induce a G1/S block. Indirect evidence suggests that the binding of RB to E2F may, in addition to inhibiting E2F transactivation function, generate a complex capable of functioning as a transrepressor. Here we show that a chimera in which the E2F1 transactivation domain was replaced with the RB pocket could, in a DNA-binding and pocket-dependent manner, mimic the ability of RB to repress transcription and induce a cell cycle arrest. In contrast, a transdominant negative E2F1 mutant that is capable of blocking E2F-dependent transactivation did not. Fusion of the RB pocket to a heterologous DNA-binding domain unrelated to E2F likewise generated a transrepressor protein when scored against a suitable reporter. These results suggest that growth suppression by RB is due, at least in part, to transrepression mediated by the pocket domain bound to certain promoters via E2F. Images Fig. 4 Fig. 5 PMID:8524800

DOE Office of Scientific and Technical Information (OSTI.GOV)

Seo, Jae Sung; Kim, Ha Na; Kim, Sun-Jick

Highlights: •NuMA is modified by SUMO-1 in a cell cycle-dependent manner. •NuMA lysine 1766 is the primary target site for SUMOylation. •SUMOylation-deficient NuMA induces multiple spindle poles during mitosis. •SUMOylated NuMA induces microtubule bundling. -- Abstract: Covalent conjugation of proteins with small ubiquitin-like modifier 1 (SUMO-1) plays a critical role in a variety of cellular functions including cell cycle control, replication, and transcriptional regulation. Nuclear mitotic apparatus protein (NuMA) localizes to spindle poles during mitosis, and is an essential component in the formation and maintenance of mitotic spindle poles. Here we show that NuMA is a target for covalent conjugationmore » to SUMO-1. We find that the lysine 1766 residue is the primary NuMA acceptor site for SUMO-1 conjugation. Interestingly, SUMO modification of endogenous NuMA occurs at the entry into mitosis and this modification is reversed after exiting from mitosis. Knockdown of Ubc9 or forced expression of SENP1 results in impairment of the localization of NuMA to mitotic spindle poles during mitosis. The SUMOylation-deficient NuMA mutant is defective in microtubule bundling, and multiple spindles are induced during mitosis. The mitosis-dependent dynamic SUMO-1 modification of NuMA might contribute to NuMA-mediated formation and maintenance of mitotic spindle poles during mitosis.« less

Anti-tumor effects and apoptosis induction by Realgar bioleaching solution in Sarcoma-180 cells in vitro and transplanted tumors in mice in vivo.

PubMed

Xie, Qin-Jian; Cao, Xin-Li; Bai, Lu; Wu, Zheng-Rong; Ma, Ying-Ping; Li, Hong-Yu

2014-01-01

Realgar which contains arsenic components has been used in traditional Chinese medicine (TCM) as an anticancer drug. However, neither Realgar nor its formula are soluble in water. As a result, high dose of Realgar has to be administered to achieve an effective blood medicine concentration, and this is associated with adverse side effects. The objective of the present study was to increase the solubility of a formula using hydrometallurgy technology as well as investigating its effects on in vitro and in vivo cell proliferation and apoptosis in Sarcoma-180 cell line. Antiproliferative activity of Realgar Bioleaching Solution (RBS) was evaluated by MTT assay. Further, effects of RBS on cell proliferation and apoptosis were studied using flow cytometry and transmission electron microscopy. Kunming mice were administered RBS in vivo, where arsenic specifically targeted solid tumors. The results indicated that RBS extract potently inhibited the tumor growth of Sarcoma-180 cell line in a dose-dependent manner. Flow cytometry and transmission electron microscopy further indicated that RBS significantly induced cell apoptosis through the inhibition of cell cycle pathway in a dose-dependent manner. Further, on RBS administration to mice, arsenic was specifically targeted to solid tumors RBS could substitute for traditional Realgar or its formula to work as a potent tool in cancer treatment.

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

PubMed

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

2016-01-01

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

4β-Hydroxywithanolide E from Physalis peruviana (golden berry) inhibits growth of human lung cancer cells through DNA damage, apoptosis and G2/M arrest

PubMed Central

2010-01-01

Background The crude extract of the fruit bearing plant, Physalis peruviana (golden berry), demonstrated anti-hepatoma and anti-inflammatory activities. However, the cellular mechanism involved in this process is still unknown. Methods Herein, we isolated the main pure compound, 4β-Hydroxywithanolide (4βHWE) derived from golden berries, and investigated its antiproliferative effect on a human lung cancer cell line (H1299) using survival, cell cycle, and apoptosis analyses. An alkaline comet-nuclear extract (NE) assay was used to evaluate the DNA damage due to the drug. Results It was shown that DNA damage was significantly induced by 1, 5, and 10 μg/mL 4βHWE for 2 h in a dose-dependent manner (p < 0.005). A trypan blue exclusion assay showed that the proliferation of cells was inhibited by 4βHWE in both dose- and time-dependent manners (p < 0.05 and 0.001 for 24 and 48 h, respectively). The half maximal inhibitory concentrations (IC50) of 4βHWE in H1299 cells for 24 and 48 h were 0.6 and 0.71 μg/mL, respectively, suggesting it could be a potential therapeutic agent against lung cancer. In a flow cytometric analysis, 4βHWE produced cell cycle perturbation in the form of sub-G1 accumulation and slight arrest at the G2/M phase with 1 μg/mL for 12 and 24 h, respectively. Using flow cytometric and annexin V/propidium iodide immunofluorescence double-staining techniques, these phenomena were proven to be apoptosis and complete G2/M arrest for H1299 cells treated with 5 μg/mL for 24 h. Conclusions In this study, we demonstrated that golden berry-derived 4βHWE is a potential DNA-damaging and chemotherapeutic agent against lung cancer. PMID:20167063

4beta-Hydroxywithanolide E from Physalis peruviana (golden berry) inhibits growth of human lung cancer cells through DNA damage, apoptosis and G2/M arrest.

PubMed

Yen, Ching-Yu; Chiu, Chien-Chih; Chang, Fang-Rong; Chen, Jeff Yi-Fu; Hwang, Chi-Ching; Hseu, You-Cheng; Yang, Hsin-Ling; Lee, Alan Yueh-Luen; Tsai, Ming-Tz; Guo, Zong-Lun; Cheng, Yu-Shan; Liu, Yin-Chang; Lan, Yu-Hsuan; Chang, Yu-Ching; Ko, Ying-Chin; Chang, Hsueh-Wei; Wu, Yang-Chang

2010-02-18

The crude extract of the fruit bearing plant, Physalis peruviana (golden berry), demonstrated anti-hepatoma and anti-inflammatory activities. However, the cellular mechanism involved in this process is still unknown. Herein, we isolated the main pure compound, 4beta-Hydroxywithanolide (4betaHWE) derived from golden berries, and investigated its antiproliferative effect on a human lung cancer cell line (H1299) using survival, cell cycle, and apoptosis analyses. An alkaline comet-nuclear extract (NE) assay was used to evaluate the DNA damage due to the drug. It was shown that DNA damage was significantly induced by 1, 5, and 10 microg/mL 4betaHWE for 2 h in a dose-dependent manner (p < 0.005). A trypan blue exclusion assay showed that the proliferation of cells was inhibited by 4betaHWE in both dose- and time-dependent manners (p < 0.05 and 0.001 for 24 and 48 h, respectively). The half maximal inhibitory concentrations (IC50) of 4betaHWE in H1299 cells for 24 and 48 h were 0.6 and 0.71 microg/mL, respectively, suggesting it could be a potential therapeutic agent against lung cancer. In a flow cytometric analysis, 4betaHWE produced cell cycle perturbation in the form of sub-G1 accumulation and slight arrest at the G2/M phase with 1 microg/mL for 12 and 24 h, respectively. Using flow cytometric and annexin V/propidium iodide immunofluorescence double-staining techniques, these phenomena were proven to be apoptosis and complete G2/M arrest for H1299 cells treated with 5 microg/mL for 24 h. In this study, we demonstrated that golden berry-derived 4betaHWE is a potential DNA-damaging and chemotherapeutic agent against lung cancer.

Novel Pactamycin Analogs Induce p53 Dependent Cell-Cycle Arrest at S-Phase in Human Head and Neck Squamous Cell Carcinoma (HNSCC) Cells

PubMed Central

Guha, Gunjan; Liang, Xiaobo; Kulesz-Martin, Molly F.; Mahmud, Taifo; Indra, Arup Kumar; Ganguli-Indra, Gitali

2015-01-01

Pactamycin, although putatively touted as a potent antitumor agent, has never been used as an anticancer drug due to its high cytotoxicity. In this study, we characterized the effects of two novel biosynthetically engineered analogs of pactamycin, de-6MSA-7-demethyl-7-deoxypactamycin (TM-025) and 7-demethyl-7-deoxypactamycin (TM-026), in head and neck squamous cell carcinoma (HNSCC) cell lines SCC25 and SCC104. Both TM-025 and TM-026 exert growth inhibitory effects on HNSCC cells by inhibiting cell proliferation. Interestingly, unlike their parent compound pactamycin, the analogs do not inhibit synthesis of nascent protein in a cell-based assay. Furthermore, they do not induce apoptosis or autophagy in a dose- or a time-dependent manner, but induce mild senescence in the tested cell lines. Cell cycle analysis demonstrated that both analogs significantly induce cell cycle arrest of the HNSCC cells at S-phase resulting in reduced accumulation of G2/M-phase cells. The pactamycin analogs induce expression of cell cycle regulatory proteins including master regulator p53, its downstream target p21Cip1/WAF1, p27kip21, p19, cyclin E, total and phospho Cdc2 (Tyr15) and Cdc25C. Besides, the analogs mildly reduce cyclin D1 expression without affecting expression of cyclin B, Cdk2 and Cdk4. Specific inhibition of p53 by pifithrin-α reduces the percentage of cells accumulated in S-phase, suggesting contribution of p53 to S-phase increase. Altogether, our results demonstrate that Pactamycin analogs TM-025 and TM-026 induce senescence and inhibit proliferation of HNSCC cells via accumulation in S-phase through possible contribution of p53. The two PCT analogs can be widely used as research tools for cell cycle inhibition studies in proliferating cancer cells with specific mechanisms of action. PMID:25938491

In vitro anticancer property of a novel thalidomide analogue through inhibition of NF-kappaB activation in HL-60 cells.

PubMed

Li, Min; Sun, Wan; Yang, Ya-ping; Xu, Bo; Yi, Wen-yuan; Ma, Yan-xia; Li, Zhong-jun; Cui, Jing-rong

2009-01-01

To investigate the anticancer property and possible mechanism of action of a novel sugar-substituted thalidomide derivative (STA-35) on HL-60 cells in vitro. TNF-alpha-induced NF-kappaB activation was determined using a reporter gene assay. The MTT assay was used to measure cytotoxicity of the compound. The appearance of apoptotic Sub-G1 cells was detected by flow cytometry analysis. PARP cleavage and protein expression of NF-kappaB p65 and its inhibitor IkappaB were viewed by Western blotting. TA-35 (1-20 micromol/L) suppressed TNF-alpha-induced NF-kappaB activation in transfected cells (HEK293/pNiFty-SEAP) in a dose- (1-20 micromol/L) and time-dependent (0-48 h) manner. It was also shown that STA-35 exerted a dose-dependent inhibitory effect on HL-60 cell proliferation with an IC(50) value of 9.05 micromol/L. In addition, STA-35 induced apoptosis in HL-60 cells, as indicated by the appearance of a Sub-G1 peak in the cell cycle distribution, as well as poly ADP-ribose polymerase (PARP) cleavage. Subsequently, both NF-kappaB p65 and its inhibitor IkappaB gradually accumulated in cytoplasmic extracts in a dose- and time-dependent manner, indicating the blockage of NF-kappaB translocation induced by TNF-alpha from the cytoplasm to the nucleus. A novel sugar-substituted thalidomide derivative, STA-35, is potent toward HL-60 cells in vitro and induces apoptosis by the suppression of NF-kappaB activation.

Glucose capped silver nanoparticles induce cell cycle arrest in HeLa cells.

PubMed

Panzarini, Elisa; Mariano, Stefania; Vergallo, Cristian; Carata, Elisabetta; Fimia, Gian Maria; Mura, Francesco; Rossi, Marco; Vergaro, Viviana; Ciccarella, Giuseppe; Corazzari, Marco; Dini, Luciana

2017-06-01

This study aims to determine the interaction (uptake and biological effects on cell viability and cell cycle progression) of glucose capped silver nanoparticles (AgNPs-G) on human epithelioid cervix carcinoma (HeLa) cells, in relation to amount, 2×10 3 or 2×10 4 NPs/cell, and exposure time, up to 48h. The spherical and well dispersed AgNPs (30±5nm) were obtained by using glucose as reducing agent in a green synthesis method that ensures to stabilize AgNPs avoiding cytotoxic soluble silver ions Ag + release. HeLa cells take up abundantly and rapidly AgNPs-G resulting toxic to cells in amount and incubation time dependent manner. HeLa cells were arrested at S and G2/M phases of the cell cycle and subG1 population increased when incubated with 2×10 4 AgNPs-G/cell. Mitotic index decreased accordingly. The dissolution experiments demonstrated that the observed effects were due only to AgNPs-G since glucose capping prevents Ag + release. The AgNPs-G influence on HeLa cells viability and cell cycle progression suggest that AgNPs-G, alone or in combination with chemotherapeutics, may be exploited for the development of novel antiproliferative treatment in cancer therapy. However, the possible influence of the cell cycle on cellular uptake of AgNPs-G and the mechanism of AgNPs entry in cells need further investigation. Copyright © 2017 Elsevier B.V. All rights reserved.

βTrCP controls the lysosome-mediated degradation of CDK1, whose accumulation correlates with tumor malignancy

PubMed Central

Herrero-Ruiz, Joaquín; Mora-Santos, Mar; Giráldez, Servando; Sáez, Carmen; Japón, Miguel Á.; Tortolero, Maria; Romero, Francisco

2014-01-01

In mammals, cell cycle progression is controlled by cyclin-dependent kinases, among which CDK1 plays important roles in the regulation of the G2/M transition, G1 progression and G1/S transition. CDK1 is highly regulated by its association to cyclins, phosphorylation and dephosphorylation, changes in subcellular localization, and by direct binding of CDK inhibitor proteins. CDK1 steady-state protein levels are held constant throughout the cell cycle by a coordinated regulation of protein synthesis and degradation. We show that CDK1 is ubiquitinated by the E3 ubiquitin ligase SCFβTrCP and degraded by the lysosome. Furthermore, we found that DNA damage not only triggers the stabilization of inhibitory phosphorylation sites on CDK1 and repression of CDK1 gene expression, but also regulates βTrCP-induced CDK1 degradation in a cell type-dependent manner. Specifically, treatment with the chemotherapeutic agent doxorubicin in certain cell lines provokes CDK1 degradation and induces apoptosis, whereas in others it inhibits destruction of the protein. These observations raise the possibility that different tumor types, depending on their pathogenic spectrum mutations, may display different sensitivity to βTrCP-induced CDK1 degradation after DNA damage. Finally, we found that CDK1 accumulation in patients’ tumors shows a negative correlation with βTrCP and a positive correlation with the degree of tumor malignancy. PMID:25149538

A U-Rich Element in the 5′ Untranslated Region Is Necessary for the Translation of p27 mRNA

PubMed Central

Millard, S. Sean; Vidal, Anxo; Markus, Maurice; Koff, Andrew

2000-01-01

Increased translation of p27 mRNA correlates with withdrawal of cells from the cell cycle. This raised the possibility that antimitogenic signals might mediate their effects on p27 expression by altering complexes that formed on p27 mRNA, regulating its translation. In this report, we identify a U-rich sequence in the 5′ untranslated region (5′UTR) of p27 mRNA that is necessary for efficient translation in proliferating and nonproliferating cells. We show that a number of factors bind to the 5′UTR in vitro in a manner dependent on the U-rich element, and their availability in the cytosol is controlled in a growth- and cell cycle-dependent fashion. One of these factors is HuR, a protein previously implicated in mRNA stability, transport, and translation. Another is hnRNP C1 and C2, proteins implicated in mRNA processing and the translation of a specific subset of mRNAs expressed in differentiated cells. In lovastatin-treated MDA468 cells, the mobility of the associated hnRNP C1 and C2 proteins changed, and this correlated with increased p27 expression. Together, these data suggest that the U-rich dependent RNP complex on the 5′UTR may regulate the translation of p27 mRNA and may be a target of antimitogenic signals. PMID:10913178

Loglinear Approximate Solutions to Real-Business-Cycle Models: Some Observations

ERIC Educational Resources Information Center

Lau, Sau-Him Paul; Ng, Philip Hoi-Tak

2007-01-01

Following the analytical approach suggested in Campbell, the authors consider a baseline real-business-cycle (RBC) model with endogenous labor supply. They observe that the coefficients in the loglinear approximation of the dynamic equations characterizing the equilibrium are related to the fundamental parameters in a relatively simple manner.…

Cycling with BRCA2 from DNA repair to mitosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Hyunsook, E-mail: HL212@snu.ac.kr

Genetic integrity in proliferating cells is guaranteed by the harmony of DNA replication, appropriate DNA repair, and segregation of the duplicated genome. Breast cancer susceptibility gene BRCA2 is a unique tumor suppressor that is involved in all three processes. Hence, it is critical in genome maintenance. The functions of BRCA2 in DNA repair and homology-directed recombination (HDR) have been reviewed numerous times. Here, I will briefly go through the functions of BRCA2 in HDR and focus on the emerging roles of BRCA2 in telomere homeostasis and mitosis, then discuss how BRCA2 exerts distinct functions in a cell-cycle specific manner inmore » the maintenance of genomic integrity. - Highlights: • BRCA2 is a multifaceted tumor suppressor and is crucial in genetic integrity. • BRCA2 exerts distinct functions in cell cycle-specific manner. • Mitotic kinases regulate diverse functions of BRCA2 in mitosis and cytokinesis.« less

Solar Cycle Fine Structure and Surface Rotation from Ca II K-Line Time Series Data

NASA Astrophysics Data System (ADS)

Scargle, Jeff; Keil, Steve; Worden, Pete

2011-10-01

Analysis of three and a half decades of data from the NSO/AFRL/Sac Peak K-line monitoring program yields evidence for four components to the variation: (a) the solar cycle, with considerable fine structure and a quasi-periodicity of 122.4 days; (b) a stochastic process, faster than (a) and largely independent of it, (c) a quasi-periodic signal due to rotational modulation, and of course (d) observational errors (shown to be quite small). Correlation and power spectrum analyses elucidate periodic and aperiodic variation of these chromospheric parameters. Time-frequency analysis is especially useful for extracting information about differential rotation, and in particular elucidates the connection between its behavior and fine structure of the solar cycle on approximately one-year time scales. These results further suggest that similar analyses will be useful at detecting and characterizing differential rotation in stars from stellar light-curves such as those being produced by NASA's Kepler observatory. Component (b) consists of variations over a range of timescales, in the manner of a "1/f" random process. A time-dependent Wilson-Bappu effect appears to be present in the solar cycle variations (a), but not in the stochastic process (b). The data can be found at the National Solar Observatory web site http://nsosp.nso.edu/data/cak_mon.html, or by file transfer protocol at ftp://ftp.nso.edu/idl/cak.parameters.

Photospheric activity of the Sun with VIRGO and GOLF. Comparison with standard activity proxies

NASA Astrophysics Data System (ADS)

Salabert, D.; García, R. A.; Jiménez, A.; Bertello, L.; Corsaro, E.; Pallé, P. L.

2017-12-01

We study the variability of solar activity using new photospheric proxies originally developed for the analysis of stellar magnetism with the CoRoT and Kepler photometric observations. These proxies were obtained by tracking the temporal modulations in the observations associated with the spots and magnetic features as the Sun rotates. We analyzed 21 yr of observations, spanning solar cycles 23 and 24, collected by the space-based photometric VIRGO and radial velocity GOLF instruments on board the SoHO satellite. We then calculated the photospheric activity proxy Sph is for each of the three VIRGO photometers and the associated Svel proxy from the radial velocity GOLF observations. Comparisons with several standard solar activity proxies sensitive to different layers of the Sun demonstrate that these new activity proxies, Sph and Svel, provide a new manner to monitor solar activity. We show that both the long- and short-term magnetic variabilities respectively associated with the 11-yr cycle and the quasi-biennial oscillation are well monitored, and that the magnetic field interaction between the subsurface, photosphere, and chromosphere of the Sun was modified between Cycle 24 and Cycle 23. Furthermore, the photometric proxies show a wavelength dependence of the response function of the solar photosphere among the three channels of the VIRGO photometers, providing inputs for the study of the stellar magnetism of Sun-like stars.

The effective application of a discrete transition model to explore cell-cycle regulation in yeast

PubMed Central

2013-01-01

Background Bench biologists often do not take part in the development of computational models for their systems, and therefore, they frequently employ them as “black-boxes”. Our aim was to construct and test a model that does not depend on the availability of quantitative data, and can be directly used without a need for intensive computational background. Results We present a discrete transition model. We used cell-cycle in budding yeast as a paradigm for a complex network, demonstrating phenomena such as sequential protein expression and activity, and cell-cycle oscillation. The structure of the network was validated by its response to computational perturbations such as mutations, and its response to mating-pheromone or nitrogen depletion. The model has a strong predicative capability, demonstrating how the activity of a specific transcription factor, Hcm1, is regulated, and what determines commitment of cells to enter and complete the cell-cycle. Conclusion The model presented herein is intuitive, yet is expressive enough to elucidate the intrinsic structure and qualitative behavior of large and complex regulatory networks. Moreover our model allowed us to examine multiple hypotheses in a simple and intuitive manner, giving rise to testable predictions. This methodology can be easily integrated as a useful approach for the study of networks, enriching experimental biology with computational insights. PMID:23915717

Inhibitory effect of turmeric curcuminoids on FLT3 expression and cell cycle arrest in the FLT3-overexpressing EoL-1 leukemic cell line.

PubMed

Tima, Singkome; Ichikawa, Hideki; Ampasavate, Chadarat; Okonogi, Siriporn; Anuchapreeda, Songyot

2014-04-25

Leukemia is a hematologic malignancy with a frequent incidence and high mortality rate. Previous studies have shown that the FLT3 gene is overexpressed in leukemic blast cells, especially in acute myeloid leukemia. In this study, a commercially available curcuminoid mixture (1), pure curcumin (2), pure demethoxycurcumin (3), and pure bisdemethoxycurcumin (4) were investigated for their inhibitory effects on cell growth, FLT3 expression, and cell cycle progression in an FLT3-overexpressing EoL-1 leukemic cell line using an MTT assay, Western blotting, and flow cytometry, respectively. The mixture (1) and compounds 2-4 demonstrated cytotoxic effects with IC50 values ranging from 6.5 to 22.5 μM. A significant decrease in FLT3 protein levels was found after curcuminoid treatment with IC20 doses, especially with mixture 1 and compound 2. In addition, mixture 1 and curcumin (2) showed activity on cell cycle arrest at the G0/G1 phase and decreased the FLT3 and STAT5A protein levels in a dose-dependent manner. Compound 2 demonstrated the greatest potential for inhibiting cell growth, cell cycle progression, and FLT3 expression in EoL-1 cells. This investigation has provided new findings regarding the effect of turmeric curcuminoids on FLT3 expression in leukemic cells.

3,3′-Diindolylmethane Ameliorates Staphylococcal Enterotoxin B–Induced Acute Lung Injury through Alterations in the Expression of MicroRNA that Target Apoptosis and Cell-Cycle Arrest in Activated T Cells

PubMed Central

Elliott, David M.; Nagarkatti, Mitzi

2016-01-01

3,3′-Diindolylmethane (DIM), a natural indole found in cruciferous vegetables, has significant anti-cancer and anti-inflammatory properties. In this current study, we investigated the effects of DIM on acute lung injury (ALI) induced by exposure to staphylococcal enterotoxin B (SEB). We found that pretreatment of mice with DIM led to attenuation of SEB-induced inflammation in the lungs, vascular leak, and IFN-γ secretion. Additionally, DIM could induce cell-cycle arrest and cell death in SEB-activated T cells in a concentration-dependent manner. Interestingly, microRNA (miRNA) microarray analysis uncovered an altered miRNA profile in lung-infiltrating mononuclear cells after DIM treatment of SEB-exposed mice. Moreover, computational analysis of miRNA gene targets and regulation networks indicated that DIM alters miRNA in the cell death and cell-cycle progression pathways. Specifically, DIM treatment significantly downregulated several miRNA and a correlative increase associated gene targets. Furthermore, overexpression and inhibition studies demonstrated that DIM-induced cell death, at least in part, used miR-222. Collectively, these studies demonstrate for the first time that DIM treatment attenuates SEB-induced ALI and may do so through the induction of microRNAs that promote apoptosis and cell-cycle arrest in SEB-activated T cells. PMID:26818958

The flavonoid quercetin transiently inhibits the activity of taxol and nocodazole through interference with the cell cycle

PubMed Central

Samuel, Temesgen; Fadlalla, Khalda; Turner, Timothy; Yehualaeshet, Teshome E.

2010-01-01

Quercetin is a flavonoid with anticancer properties. In this study, we examined the effects of quercetin on cell cycle, viability and proliferation of cancer cells, either singly or in combination with the microtubule-targeting drugs taxol and nocodazole. Although quercetin induced cell death in a dose dependent manner, 12.5-50μM quercetin inhibited the activity of both taxol and nocodazole to induce G2/M arrest in various cell lines. Quercetin also partially restored drug-induced loss in viability of treated cells for up to 72 hours. This antagonism of microtubule-targeting drugs was accompanied by a delay in cell cycle progression and inhibition of the buildup of cyclin-B1 at the microtubule organizing center of treated cells. However, quercetin did not inhibit the microtubule targeting of taxol or nocodazole. Despite the short-term protection of cells by quercetin, colony formation and clonogenicity of HCT116 cells were still suppressed by quercetin or quercetin-taxol combination. The status of cell adherence to growth matrix was critical in determining the sensitivity of HCT116 cells to quercetin. We conclude that while long-term exposure of cancer cells to quercetin may prevent cell proliferation and survival, the interference of quercetin with cell cycle progression diminishes the efficacy of microtubule-targeting drugs to arrest cells at G2/M. PMID:21058190

Menadione induces G2/M arrest in gastric cancer cells by down-regulation of CDC25C and proteasome mediated degradation of CDK1 and cyclin B1

PubMed Central

Lee, Min Ho; Cho, Yoonjung; Kim, Do Hyun; Woo, Hyun Jun; Yang, Ji Yeong; Kwon, Hye Jin; Yeon, Min Ji; Park, Min; Kim, Sa-Hyun; Moon, Cheol; Tharmalingam, Nagendran; Kim, Tae Ue; Kim, Jong-Bae

2016-01-01

Menadione (vitamin K3) has been reported to induce apoptotic cell death and growth inhibition in various types of cancer cells. However, involvement of menadione in cell cycle control has not been considered in gastric cancer cells yet. In the current study, we have investigated whether menadione is involved in the cell cycle regulation and suppression of growth in gastric cancer cells. In the cell cycle analysis, we found that menadione induced G2/M cell cycle arrest in AGS cells. To elucidate the underlying mechanism, we investigated the cell cycle regulatory molecules involved in the G2/M cell cycle transition. After 24 h of menadione treatment, the protein level of CDK1, CDC25C and cyclin B1 in AGS cells was decreased in a menadione dose-dependent manner. In the time course experiment, the protein level of CDC25C decreased in 6 h, and CDK1and cyclin B1 protein levels began to decrease after 18 h of menadione treatment. We found that mRNA level of CDC25C decreased by menadione treatment in 6 h. Menadione did not have an influence on mRNA level of CDK1 and cyclin B1 though the protein levels were decreased. However, the decreased protein levels of CDK1 and cyclin B1 were recovered by inhibition of proteasome. Collectively, these results suggest that menadione inhibits growth of gastric cancer cells by reducing expression of CDC25C and promoting proteasome mediated degradation of CDK1 and cyclin B1 thereby blocking transition of the cell cycle from G2 phase to M phase. PMID:28077999

Animal Models for Studying the In Vivo Functions of Cell Cycle CDKs.

PubMed

Risal, Sanjiv; Adhikari, Deepak; Liu, Kui

2016-01-01

Multiple Cdks (Cdk4, Cdk6, and Cdk2) and a mitotic Cdk (Cdk1) are involved in cell cycle progression in mammals. Cyclins, Cdk inhibitors, and phosphorylations (both activating and inhibitory) at different cellular levels tightly modulate the activities of these kinases. Based on the results of biochemical studies, it was long believed that different Cdks functioned at specific stages during cell cycle progression. However, deletion of all three interphase Cdks in mice affected cell cycle entry and progression only in certain specialized cells such as hematopoietic cells, beta cells of the pancreas, pituitary lactotrophs, and cardiomyocytes. These genetic experiments challenged the prevailing biochemical model and established that Cdks function in a cell-specific, but not a stage-specific, manner during cell cycle entry and the progression of mitosis. Recent in vivo studies have further established that Cdk1 is the only Cdk that is both essential and sufficient for driving the resumption of meiosis during mouse oocyte maturation. These genetic studies suggest a minimal-essential cell cycle model in which Cdk1 is the central regulator of cell cycle progression. Cdk1 can compensate for the loss of the interphase Cdks by forming active complexes with A-, B-, E-, and D-type Cyclins in a stepwise manner. Thus, Cdk1 plays an essential role in both mitosis and meiosis in mammals, whereas interphase Cdks are dispensable.

TOFA suppresses ovarian cancer cell growth in vitro and in vivo.

PubMed

Li, Shu; Qiu, Lihua; Wu, Buchu; Shen, Haoran; Zhu, Jing; Zhou, Liang; Gu, Liying; Di, Wen

2013-08-01

A characteristic feature of cancer cells is the activation of de novo fatty acid synthesis. Acetyl‑CoA carboxylase (ACC) is a key enzyme in fatty acid synthesis, accelerating the reaction that carboxylates cytosolic acetyl‑CoA to form malonyl‑CoA. ACC is highly expressed in several types of human cancer and is important in breast and prostate cancer cell growth. The aim of the present study was to investigate the effects of 5‑tetradecyloxy‑2‑furoic acid (TOFA), an allosteric inhibitor of ACC, on the proliferation and cell cycle progression of the ovarian cancer cell lines COC1 and COC1/DDP. TOFA was found to be cytotoxic to COC1 and COC1/DDP cells with a 50% inhibitory concentration (IC50) of ~26.1 and 11.6 µg/ml, respectively. TOFA inhibited the proliferation of the cancer cells examined in a time‑ and dose‑dependent manner, arrested the cells in the G0/G1 cell cycle phase and induced apoptosis. The expression of the cell cycle regulating proteins cyclin D1 and cyclin-dependent kinase (CDK) 4, as well as the expression of the apoptosis‑related proteins caspase‑3 and Bcl‑2, were detected by western blot analysis. Cyclin D1, CDK4 and Bcl‑2 protein expression was inhibited by TOFA, while caspase‑3 was cleaved and activated. To the best of our knowledge, the present study demonstrated for the first time that TOFA inhibits COC1/DDP cell growth in ovarian tumor mouse xenografts. By inhibiting ACC, TOFA may be a promising small molecule agent for ovarian cancer therapy.

Apoptosis induction by 7-chloroquinoline-1,2,3-triazoyl carboxamides in triple negative breast cancer cells.

PubMed

Begnini, Karine Rech; Duarte, Wladimir R; da Silva, Liziane Pereira; Buss, Julieti H; Goldani, Bruna S; Fronza, Mariana; Segatto, Natália Vieira; Alves, Diego; Savegnago, Lucielli; Seixas, Fabiana Kömmling; Collares, Tiago

2017-07-01

Breast cancer is a major public health burden in both developed and developing countries and there is still a need to screen new molecules with different modes of actions. The aims of this study were to evaluate the selectivity profile, apoptotic cell death and cell cycle arrest induced by 7-chloroquinoline-1,2,3-triazoyl carboxamides derivatives in hormonal-dependent and hormonal-independent breast cancer cells. Results showed significantly decreased MCF-7 and MDA-MB-231 cells viability in vitro in a dose dependent manner after treatment with 7-chloroquinoline derivatives QTCA-1, QTCA-2 and QTCA-3. QTCA-1 displayed the highest cytotoxic activity from all the tested compounds in MDA-MB-231 with IC50 values of 20.60, 20.42 and 19.91μM in 24, 48 and 72h of treatment respectively. Apoptosis induction was also significantly higher in the hormonal-independent breast cancer cells, with 80.4% of dead cells in MDA-MB-231 and only 16.8% of dead in MCF-7 cells. As a result, G0/G1 cycle arrest was observed in MCF-7 cells and no cell cycle arrest at all was observed in MDA-MB-231 cells. Molecular docking showed a high affinity of QTCA-1 to PARP-1, Scr and PI3K/mTOR targets. These results suggest a strong activity of the 7-chloroquinoline derivative QTCA-1 in independent-hormonal cells and suggest selectivity for triple negative cells. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Erythrocyte signal transduction pathways, their oxygenation dependence and functional significance.

PubMed

Barvitenko, Nadezhda N; Adragna, Norma C; Weber, Roy E

2005-01-01

Erythrocytes play a key role in human and vertebrate metabolism. Tissue O2 supply is regulated by both hemoglobin (Hb)-O2 affinity and erythrocyte rheology, a key determinant of tissue perfusion. Oxygenation-deoxygenation transitions of Hb may lead to re-organization of the cytoskeleton and signalling pathways activation/deactivation in an O2-dependent manner. Deoxygenated Hb binds to the cytoplasmic domain of the anion exchanger band 3, which is anchored to the cytoskeleton, and is considered a major mechanism underlying the oxygenation-dependence of several erythrocyte functions. This work discusses the multiple modes of Hb-cytoskeleton interactions. In addition, it reviews the effects of Mg2+, 2,3-diphosphoglycerate, NO, shear stress and Ca2+, all factors accompanying the oxygenation-deoxygenation cycle in circulating red cells. Due to the extensive literature on the subject, the data discussed here, pertain mainly to human erythrocytes whose O2 affinity is modulated by 2,3-diphosphoglycerate, ectothermic vertebrate erythrocytes that use ATP, and to bird erythrocytes that use inositol pentaphosphate. Copyright 2005 S. Karger AG, Basel.

In vitro and in vivo anticancer effects of Lithospermum erythrorhizon extract on B16F10 murine melanoma.

PubMed

Rajasekar, Seetharaman; Park, Da Jung; Park, Cheol; Park, Sejin; Park, Young Hoon; Kim, Sun Tae; Choi, Yung Hyun; Choi, Young Whan

2012-11-21

Lithospermum erythrorhizon has long been used in traditional Asian medicine for the treatment of diseases including skin cancer. In this study, hexane extract from the roots of Lithospermum erythrorhizon (LEH) was chemically characterized and its anticancer activity was tested against the most aggressive form of skin cancer. The in vitro anticancer studies viz. cell growth, cell cycle and apoptosis, and the expression of tumor regulating proteins were analyzed against B16F10 melanoma cells. In addition, C57BL/6 mice models were used to evaluate the in vivo anticancer potential of LEH. Mice were intraperitoneally injected with LEH at doses of 0.1 and 10mg/kg every 3 days. The tumor inhibition ratio was determined after 21 days of treatment and the histopathological analyses of the tumor tissues were compared. Further, LEH was purified and its active compounds were structurally elucidated and identified by NMR spectra and quantified by HPLC analyses. LEH effectively inhibits the growth of melanoma cells with an IC(50) of 2.73μg/ml. Cell cycle analysis revealed that LEH increased the percentage of cells in sub-G1 phase by dose dependent manner. LEH exhibited down regulation of anti-apoptotic Bcl-2 family proteins and up regulation of apoptotic Bax protein expression. Importantly, LEH induced cleavage of poly (ADP-ribose) polymerase (PARP) and activated the caspase cascade (caspase 3) with this cleavage mediating the apoptosis of B16F10 cells. LEH treatment at a dose of 10mg/kg for 21 days in experimental mice implanted with tumors resulted in significant reduction of the tumor growth (43%) and weight (36%). Histopathology analysis of LEH treated tumor tissues showed evidence of increased necrotic cells in a concentration dependent manner. Meanwhile, five naphthoquinone compounds [Shikonin (1); Deoxyshikonin (2); β-Hydroxyisovalerylshikonin (3); Acetylshikonin (4) and Isobutyrylshikonin (5)] were purified from LEH and responsible for its anticancer activity. LEH induced apoptosis in B16F10 cells by activation of caspase 3 and inducing sub-G1 cell cycle arrest. LEH exhibited both in vitro and in vivo anticancer activity. Shikonin derivatives in the LEH are responsible for the anticancer activity. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Antioxidant and anticancer activity of Artemisia princeps var. orientalis extract in HepG2 and Hep3B hepatocellular carcinoma cells

PubMed Central

Choi, Eun-Jeong

2013-01-01

Objective The aim of the present study was to investigate antioxidant and the anticancerigen activity of a methanol extract from Artemisia princeps var. orientalis (APME), a well-known traditional herbal medicine in Asia, in hepatocellular cancer cells. Methods To evaluate the antioxidant activity of APME, reactive oxygen species (ROS) and the antioxidant enzymes, superoxide dismutase (SOD) and catalase were investigated in HepG2 cells exposed to APME (5, 100, and 200 µg/mL) for 72 h. Then, to evaluate the anticancer activity of APME, we investigated the proliferation and apoptosis induction of HepG2 and Hep3B cells exposed to APME (1-200 µg/mL) for 24, 48, and 72 h. Results APME dose-dependently reduced the generation of ROS in the presence of H2O2 compared with control cells. Furthermore, it increased catalase and SOD activity. Moreover, APME inhibited cell proliferation in a dose- and time-dependent manner, but at concentrations lower than 100 µg/mL, the inhibition was less dose-dependent than time-dependent. HepG2 and Hep3B cells exposed to 5, 100, and 200 µg/mL APME for 72 h underwent cell cycle arrest and apoptosis. Exposure to APME resulted in a significant increase in the number of cells in G1 phase and a decrease in the G2/M phase cell population. In addition, APME induced P53 expression of HepG2 cells in a dose-dependent manner, and played a role in the downregulation of Bcl-2 and upregulation of Bax in both HepG2 and Hep3B cells. Conclusions These results indicate the potential role of APME as an antioxidant and anticancerigen agent in hepatocarcinoma cell lines. PMID:24255577

Phosphopeptide binding by Sld3 links Dbf4-dependent kinase to MCM replicative helicase activation.

PubMed

Deegan, Tom D; Yeeles, Joseph Tp; Diffley, John Fx

2016-05-02

The initiation of eukaryotic DNA replication requires the assembly of active CMG (Cdc45-MCM-GINS) helicases at replication origins by a set of conserved and essential firing factors. This process is controlled during the cell cycle by cyclin-dependent kinase (CDK) and Dbf4-dependent kinase (DDK), and in response to DNA damage by the checkpoint kinase Rad53/Chk1. Here we show that Sld3, previously shown to be an essential CDK and Rad53 substrate, is recruited to the inactive MCM double hexamer in a DDK-dependent manner. Sld3 binds specifically to DDK-phosphorylated peptides from two MCM subunits (Mcm4, 6) and then recruits Cdc45. MCM mutants that cannot bind Sld3 or Sld3 mutants that cannot bind phospho-MCM or Cdc45 do not support replication. Moreover, phosphomimicking mutants in Mcm4 and Mcm6 bind Sld3 without DDK and facilitate DDK-independent replication. Thus, Sld3 is an essential "reader" of DDK phosphorylation, integrating signals from three distinct protein kinase pathways to coordinate DNA replication during S phase. © 2016 The Authors. Published under the terms of the CC BY 4.0 license.

Green Net Value Added as a Sustainability Metric Based on Life Cycle Assessment: An Application to Bounty® Paper Towel

EPA Science Inventory

Sustainability measurement in economics involves evaluation of environmental and economic impact in an integrated manner. In this study, system level economic data are combined with environmental impact from a life cycle assessment (LCA) of a common product. We are exploring a co...

Impairment of the Plasmodium falciparum erythrocytic cycle induced by angiotensin peptides.

PubMed

Saraiva, Victor Barbosa; de Souza Silva, Leandro; Ferreira-DaSilva, Claudio Teixeira; da Silva-Filho, João Luiz; Teixeira-Ferreira, André; Perales, Jonas; Souza, Mariana Conceição; Henriques, Maria das Graças; Caruso-Neves, Celso; de Sá Pinheiro, Ana Acacia

2011-02-18

Plasmodium falciparum causes the most serious complications of malaria and is a public health problem worldwide with over 2 million deaths each year. The erythrocyte invasion mechanisms by Plasmodium sp. have been well described, however the physiological aspects involving host components in this process are still poorly understood. Here, we provide evidence for the role of renin-angiotensin system (RAS) components in reducing erythrocyte invasion by P. falciparum. Angiotensin II (Ang II) reduced erythrocyte invasion in an enriched schizont culture of P. falciparum in a dose-dependent manner. Using mass spectroscopy, we showed that Ang II was metabolized by erythrocytes to Ang IV and Ang-(1-7). Parasite infection decreased Ang-(1-7) and completely abolished Ang IV formation. Similar to Ang II, Ang-(1-7) decreased the level of infection in an A779 (specific antagonist of Ang-(1-7) receptor, MAS)-sensitive manner. 10(-7) M PD123319, an AT(2) receptor antagonist, partially reversed the effects of Ang-(1-7) and Ang II. However, 10(-6) M losartan, an antagonist of the AT(1) receptor, had no effect. Gs protein is a crucial player in the Plasmodium falciparum blood cycle and angiotensin peptides can modulate protein kinase A (PKA) activity; 10(-8) M Ang II or 10(-8) M Ang-(1-7) inhibited this activity in erythrocytes by 60% and this effect was reversed by 10(-7) M A779. 10(-6) M dibutyryl-cAMP increased the level of infection and 10(-7) M PKA inhibitor decreased the level of infection by 30%. These results indicate that the effect of Ang-(1-7) on P. falciparum blood stage involves a MAS-mediated PKA inhibition. Our results indicate a crucial role for Ang II conversion into Ang-(1-7) in controlling the erythrocytic cycle of the malaria parasite, adding new functions to peptides initially described to be involved in the regulation of vascular tonus.

Entrainment of the Mammalian Cell Cycle by the Circadian Clock: Modeling Two Coupled Cellular Rhythms

PubMed Central

Gérard, Claude; Goldbeter, Albert

2012-01-01

The cell division cycle and the circadian clock represent two major cellular rhythms. These two periodic processes are coupled in multiple ways, given that several molecular components of the cell cycle network are controlled in a circadian manner. For example, in the network of cyclin-dependent kinases (Cdks) that governs progression along the successive phases of the cell cycle, the synthesis of the kinase Wee1, which inhibits the G2/M transition, is enhanced by the complex CLOCK-BMAL1 that plays a central role in the circadian clock network. Another component of the latter network, REV-ERBα, inhibits the synthesis of the Cdk inhibitor p21. Moreover, the synthesis of the oncogene c-Myc, which promotes G1 cyclin synthesis, is repressed by CLOCK-BMAL1. Using detailed computational models for the two networks we investigate the conditions in which the mammalian cell cycle can be entrained by the circadian clock. We show that the cell cycle can be brought to oscillate at a period of 24 h or 48 h when its autonomous period prior to coupling is in an appropriate range. The model indicates that the combination of multiple modes of coupling does not necessarily facilitate entrainment of the cell cycle by the circadian clock. Entrainment can also occur as a result of circadian variations in the level of a growth factor controlling entry into G1. Outside the range of entrainment, the coupling to the circadian clock may lead to disconnected oscillations in the cell cycle and the circadian system, or to complex oscillatory dynamics of the cell cycle in the form of endoreplication, complex periodic oscillations or chaos. The model predicts that the transition from entrainment to 24 h or 48 h might occur when the strength of coupling to the circadian clock or the level of growth factor decrease below critical values. PMID:22693436

Mertensene, a Halogenated Monoterpene, Induces G2/M Cell Cycle Arrest and Caspase Dependent Apoptosis of Human Colon Adenocarcinoma HT29 Cell Line through the Modulation of ERK-1/-2, AKT and NF-κB Signaling.

PubMed

Tarhouni-Jabberi, Safa; Zakraoui, Ons; Ioannou, Efstathia; Riahi-Chebbi, Ichrak; Haoues, Meriam; Roussis, Vassilios; Kharrat, Riadh; Essafi-Benkhadir, Khadija

2017-07-20

Conventional treatment of advanced colorectal cancer is associated with tumor resistance and toxicity towards normal tissues. Therefore, development of effective anticancer therapeutic alternatives is still urgently required. Nowadays, marine secondary metabolites have been extensively investigated due to the fact that they frequently exhibit anti-tumor properties. However, little attention has been given to terpenoids isolated from seaweeds. In this study, we isolated the halogenated monoterpene mertensene from the red alga Pterocladiella capillacea (S.G. Gmelin) Santelices and Hommersand and we highlight its inhibitory effect on the viability of two human colorectal adenocarcinoma cell lines HT29 and LS174. Interestingly, exposure of HT29 cells to different concentrations of mertensene correlated with the activation of MAPK ERK-1/-2, Akt and NF-κB pathways. Moreover, mertensene-induced G2/M cell cycle arrest was associated with a decrease in the phosphorylated forms of the anti-tumor transcription factor p53 , retinoblastoma protein (Rb), cdc2 and chkp2. Indeed, a reduction of the cellular level of cyclin-dependent kinases CDK2 and CDK4 was observed in mertensene-treated cells. We also demonstrated that mertensene triggers a caspase-dependent apoptosis in HT29 cancer cells characterized by the activation of caspase-3 and the cleavage of poly (ADP-ribose) polymerase (PARP). Besides, the level of death receptor-associated protein TRADD increased significantly in a concentration-dependent manner. Taken together, these results demonstrate the potential of mertensene as a drug candidate for the treatment of colon cancer.

Indigo naturalis and its component tryptanthrin exert anti-angiogenic effect by arresting cell cycle and inhibiting Akt and FAK signaling in human vascular endothelial cells.

PubMed

Chang, Hsin-Ning; Huang, Sheng-Teng; Yeh, Yuan-Chieh; Wang, Hsin-Shih; Wang, Tzu-Hao; Wu, Yi-Hong; Pang, Jong-Hwei S

2015-11-04

Indigo naturalis has been used to treat inflammatory diseases and dermatosis, including psoriasis, since thousands of years in China. It has been proven effective in our previous clinical studies on treating psoriasis, but the active component and the mechanism of how indigo naturalis working still needs to be clarified. Since the dysregulated angiogenesis is known to play an important role in the pathogenesis of psoriasis, the anti-angiogenic effect of indigo naturalis and tryptanthrin, a pure component of indigo naturalis, was investigated. The in vivo angiogenesis was studied by chick chorioallantoic membrane assay. The in vitro studies were performed using human vascular endothelial cells. Cell viability was determined by MTT assay. Cell cycle distribution was revealed by flow cytometry. The cellular messenger (m)RNA or protein expression level was analyzed by real-time RT-PCR or Western blot, respectively. Transwell filter migration assay and matrix gel-induced tube formation method were applied to examine the angiogenic potential. Indigo naturalis significantly inhibited the in vivo vascular endothelial growth factor (VEGF)-induced angiogenesis, as well as tryptanthrin. In vitro studies confirmed that indigo naturalis and tryptanthrin reduced the number of viable vascular endothelial cells. Tryptanthrin resulted in a cell cycle arrest and dose-dependently decreased the expressions of cyclin A, cyclin B, cyclin dependent kinase(CDK) 1 and 2, but not cyclin D and cyclin E, at both the mRNA and protein levels. The migration and tube formation of vascular endothelial cells were significantly inhibited by tryptanthrin in a dose-dependent manner. Result also showed that tryptanthrin could reduce the phosphorylated levels of both protein kinase B (PKB or Akt) and focal adhesion kinase (FAK). All together, these results demonstrated the anti-angiogenic effect of tryptanthrin, the acting component of indigo naturalis and revealed the underlying mechanism by inhibiting the cell cycle progression, cell migration and tube formation, likely mediated through blocking the Akt and FAK pathways. Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Life in the Clouds of Venus? An Experimental Synthetic Biology Approach

NASA Technical Reports Server (NTRS)

Rothschild, L. J.; Paulino-Lima, I. G.; Amatya, D.; Bajar, B.; Geilich, B.; Hu, J.; Jackson, C. J.

2015-01-01

The surface of Venus constitutes the most hellish and biologically inhospitable planetary surface in our solar system, boasting a pH of 0, blistering winds that can melt lead, and pressures of 60 atm. However, during the earlier years of the solar system, without the runaway greenhouse effect that has plagued the planet, Venus potentially housed oceans and perhaps even life. There is a possibility that microbes could have retreated into hospitable niches in the atmosphere, as suggested by Carl Sagan as early as 1967 [1]. For example, 50 km above the raging hell of the Venusian surface, exists a relatively temperate environment that might serve as reservoir for life. This astrobiology project seeks to explore life at the extremes and to theorize whether microbial communities could not only survive but also reproduce in the Venusian atmosphere. Specifically, we ask: are aerosols viable microbial environments? But before we can test for life in the clouds, we have to develop a proper reporter to visualize cell growth in situ. For this purpose, we aimed to develop cell-growth dependent reporters to serve as remote biosensors for cell growth. We developed two using the polA promoter, a DNA-replication dependent promoter, and nrd operon promoter, a cell-cycle dependent promoter. Using these cell-growth reporters, the next step is to aerosolize microbes expressing these reporters in a suspension chamber adapted from a Millikan Drop Apparatus to assay reproduction in an aerosolized environment. Better yet is to test the reproduction of microbes in a microgravity regime such as on ISS.Approach: We engineered two cell-cycle dependent genetic reporters. One was the polA promoter which codes for DNA Polymerase I, a gene active in DNA replication [2]. The other was the nrdP. The activation of ribonucleotide reductase reduces ribonucleotides into deoxyribonucleotides and is involved in the bacterial cell cycle [3]. This promoter began activation during the initiation of DNA replication and is cell-cycle dependent [4]. These promoters were fused to a GFP reporter, transformed into E. coli. The constructs were deposited in the iGEM registry as K847210: Escherichia coli DNA-replication dependent polA promoter K847211: Escherichia coli cell-division dependent nrd promoter Results: Our constructs displayed fluorescence when transformed into NEB-5alpha competent cells. While nrdP-E0840 displayed sufficient fluorescence as verified by fluorescent microscopy, the original polAP-E0840 construct (which uses mut3b GFP) exhibited low expression; while fluorescence was visible under the microscope, the signal was too weak for the camera to recognize. The polA promoter was therefore digest-ed with EcoRI and SpeI then ligated into plasmid pNCS containing a RBS, Clover, and a terminator. Clover is a highly engineered green fluorescent protein that exhibits extreme brightness [5] Fluoresence time course data demonsrated that the genes were induced in a cell cycle dependant manner [6]. Our assays via microscopy and the bulk assay shows that our promoters are functional as cell cycle reporters.Conclusions: The application of such tools are widespread and not limited to astrobiology; nrdP could be used to determine doubling times empirically and could possibly extrapolate DNA content from intensity of signals expressed by polAP. However, we are pri-marily interested in its use in astrobiology.

DELAY OF GERMINATION 1 mediates a conserved coat-dormancy mechanism for the temperature- and gibberellin-dependent control of seed germination.

PubMed

Graeber, Kai; Linkies, Ada; Steinbrecher, Tina; Mummenhoff, Klaus; Tarkowská, Danuše; Turečková, Veronika; Ignatz, Michael; Sperber, Katja; Voegele, Antje; de Jong, Hans; Urbanová, Terezie; Strnad, Miroslav; Leubner-Metzger, Gerhard

2014-08-26

Seed germination is an important life-cycle transition because it determines subsequent plant survival and reproductive success. To detect optimal spatiotemporal conditions for germination, seeds act as sophisticated environmental sensors integrating information such as ambient temperature. Here we show that the delay of germination 1 (DOG1) gene, known for providing dormancy adaptation to distinct environments, determines the optimal temperature for seed germination. By reciprocal gene-swapping experiments between Brassicaceae species we show that the DOG1-mediated dormancy mechanism is conserved. Biomechanical analyses show that this mechanism regulates the material properties of the endosperm, a seed tissue layer acting as germination barrier to control coat dormancy. We found that DOG1 inhibits the expression of gibberellin (GA)-regulated genes encoding cell-wall remodeling proteins in a temperature-dependent manner. Furthermore we demonstrate that DOG1 causes temperature-dependent alterations in the seed GA metabolism. These alterations in hormone metabolism are brought about by the temperature-dependent differential expression of genes encoding key enzymes of the GA biosynthetic pathway. These effects of DOG1 lead to a temperature-dependent control of endosperm weakening and determine the optimal temperature for germination. The conserved DOG1-mediated coat-dormancy mechanism provides a highly adaptable temperature-sensing mechanism to control the timing of germination.

Tea polyphenols induce S phase arrest and apoptosis in gallbladder cancer cells

PubMed Central

Wang, Jiaqi; Pan, Yixuan; Hu, Jiacheng; Ma, Qiang; Xu, Yi; Zhang, Yijian; Zhang, Fei; Liu, Yingbin

2018-01-01

Gallbladder cancer (GBC) is the most common malignancy in the biliary tract. Without effective treatment, its prognosis is notoriously poor. Tea polyphenols (TPs) have many pharmacological and health benefits, including antioxidant, anti-inflammatory, anti-tumor, anti-thrombotic, antibacterial, and vasodilatory properties. However, the anti-cancer effect of TPs in human gallbladder cancer has not yet been determined. Cell viability and colony formation assay were used to investigate the cell growth. Cell cycle and apoptosis were evaluated by flow cytometry analysis. Western blot assay was used to detect the expression of proteins related to cell cycle and apoptosis. Human tumor xenografts were used to examine the effect of TPs on gallbladder cancer cells in vivo. TPs significantly inhibited cell growth of gallbladder cancer cell lines in a dose- and time-dependent manner. Cell cycle progression in GBC cells was blocked at the S phase by TPs. TPs also induced mitochondrial-related apoptosis in GBC cells by upregulating Bax, cleaved caspase-3, and cleaved PARP expressions and downregulating Bcl-2, cyclin A, and Cdk2 expressions. The effects of TPs on GBC were further proven in vivo in a mouse xenograft model. Our study is the first to report that TPs inhibit GBC cell growth and these compounds may have potential as novel therapeutic agents for treating gallbladder cancer. PMID:29513793

Alternariol induce toxicity via cell death and mitochondrial damage on Caco-2 cells.

PubMed

Fernández-Blanco, Celia; Juan-García, Ana; Juan, Cristina; Font, Guillermina; Ruiz, Maria-Jose

2016-02-01

Alternariol (AOH), a mycotoxin produced by Alternaria sp, appears as food contaminant in fruit, vegetables and cereal products. Its toxicity has been demonstrated, but the mechanisms involved have not been elucidated yet. In this study, the pathways triggered by AOH and degradation products generated on Caco-2 cells were evaluated. Cells were exposed to AOH sub-cytotoxic concentrations of 15, 30 and 60 μM. Cell cycle disruption, the induction of apoptosis/necrosis and changes in mitochondrial membrane potential (Δψm) after 24 and 48 h was asses by flow cytometry. Also, AOH and its degradation products were evaluated after 24 and 48 h by high-performance liquid chromatography with tandem mass spectrometric (LC-MS/MS) to detect and quantify its levels. Cell cycle was significantly decreased at G1 phase and increased at S and G2/M phase at the time of exposure. AOH induced necrosis, apoptosis/necrosis and loss of Δψm in a dose and time-dependent manner. The concentrations of AOH quantified in the culture media exposed to AOH decreased as the exposure time was increased. In conclusion, AOH caused cytotoxic effects supported by blocking cell cycle, decreasing cell proliferation and increasing apoptosis/necrosis cells. Copyright © 2015 Elsevier Ltd. All rights reserved.

Terrestrosin D, a steroidal saponin from Tribulus terrestris L., inhibits growth and angiogenesis of human prostate cancer in vitro and in vivo.

PubMed

Wei, Shihu; Fukuhara, Hideo; Chen, Guang; Kawada, Chiaki; Kurabayashi, Atsushi; Furihata, Mutsuo; Inoue, Keiji; Shuin, Taro

2014-01-01

The aim of this study was to investigate whether terrestrosin D (TED) inhibits the progression of castration-resistant prostate cancer and consider its mechanism. Cell cycle, mitochondrial membrane potential (ΔΨm) and apoptosis were determined by flow cytometry. Caspase-3 activity and vascular endothelial growth factor secretion were detected by a caspase-3 assay and human vascular endothelial growth factor kit, respectively. A PC-3 xenograft mouse model was used to evaluate the anticancer effect of TED in vivo. In vitro, TED strongly suppressed the growth of prostate cancer cells and endothelial cells in a dose-dependent manner. TED induced cell cycle arrest and apoptosis in PC-3 cells and human umbilical vascular endothelial cells (HUVECs). TED-induced apoptosis did not involve the caspase pathway. TED also decreased ΔΨm in PC-3 cells and HUVECs. In vivo, TED significantly suppressed tumor growth in nude mice bearing PC-3 cells, without any overt toxicity. Immunohistochemical analysis showed TED induced apoptotic cell death and inhibited angiogenesis in xenograft tumor cells. Cell cycle arrest and induction of apoptosis in cancer cells and endothelial cells might be plausible mechanisms of actions for the observed antitumor and antiangiogenic activities of TED. © 2014 S. Karger AG, Basel.

Gossypol inhibition of mitosis, cyclin D1 and Rb protein in human mammary cancer cells and cyclin-D1 transfected human fibrosarcoma cells.

PubMed Central

Ligueros, M.; Jeoung, D.; Tang, B.; Hochhauser, D.; Reidenberg, M. M.; Sonenberg, M.

1997-01-01

The antiproliferative effects of gossypol on human MCF-7 mammary cancer cells and cyclin D1-transfected HT-1060 human fibrosarcoma cells were investigated by cell cycle analysis and effects on the cell cycle regulatory proteins Rb and cyclin D1. Flow cytometry of MCF-7 cells at 24 h indicated that 10 microM gossypol inhibited DNA synthesis by producing a G1/S block. Western blot analysis using anti-human Rb antibodies and anti-human cyclin D1 antibodies in MCF-7 cells and high- and low-expression cyclin D1-transfected fibrosarcoma cells indicated that, after 6 h exposure, gossypol decreased the expression levels of these proteins in a dose-dependent manner. Gossypol also decreased the ratio of phosphorylated to unphosphorylated Rb protein in human mammary cancer and fibrosarcoma cell lines. Gossypol (10 microM) treated also decreased cyclin D1-associated kinase activity on histone H1 used as a substrate in MCF-7 cells. These results suggest that gossypol might suppress growth by modulating the expression of cell cycle regulatory proteins Rb and cyclin D1 and the phosphorylation of Rb protein. Images Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 PMID:9218727

Targeting TRPM2 Channels Impairs Radiation-Induced Cell Cycle Arrest and Fosters Cell Death of T Cell Leukemia Cells in a Bcl-2-Dependent Manner

PubMed Central

Klumpp, Dominik; Misovic, Milan; Szteyn, Kalina; Shumilina, Ekaterina; Rudner, Justine; Huber, Stephan M.

2016-01-01

Messenger RNA data of lymphohematopoietic cancer lines suggest a correlation between expression of the cation channel TRPM2 and the antiapoptotic protein Bcl-2. The latter is overexpressed in various tumor entities and mediates therapy resistance. Here, we analyzed the crosstalk between Bcl-2 and TRPM2 channels in T cell leukemia cells during oxidative stress as conferred by ionizing radiation (IR). To this end, the effects of TRPM2 inhibition or knock-down on plasma membrane currents, Ca2+ signaling, mitochondrial superoxide anion formation, and cell cycle progression were compared between irradiated (0–10 Gy) Bcl-2-overexpressing and empty vector-transfected Jurkat cells. As a result, IR stimulated a TRPM2-mediated Ca2+-entry, which was higher in Bcl-2-overexpressing than in control cells and which contributed to IR-induced G2/M cell cycle arrest. TRPM2 inhibition induced a release from G2/M arrest resulting in cell death. Collectively, this data suggests a pivotal function of TRPM2 in the DNA damage response of T cell leukemia cells. Apoptosis-resistant Bcl-2-overexpressing cells even can afford higher TRPM2 activity without risking a hazardous Ca2+-overload-induced mitochondrial superoxide anion formation. PMID:26839633

Nucleolar TRF2 attenuated nucleolus stress-induced HCC cell-cycle arrest by altering rRNA synthesis.

PubMed

Yuan, Fuwen; Xu, Chenzhong; Li, Guodong; Tong, Tanjun

2018-05-03

The nucleolus is an important organelle that is responsible for the biogenesis of ribosome RNA (rRNA) and ribosomal subunits assembly. It is also deemed to be the center of metabolic control, considering the critical role of ribosomes in protein translation. Perturbations of rRNA synthesis are closely related to cell proliferation and tumor progression. Telomeric repeat-binding factor 2 (TRF2) is a member of shelterin complex that is responsible for telomere DNA protection. Interestingly, it was recently reported to localize in the nucleolus of human cells in a cell-cycle-dependent manner, while the underlying mechanism and its role on the nucleolus remained unclear. In this study, we found that nucleolar and coiled-body phosphoprotein 1 (NOLC1), a nucleolar protein that is responsible for the nucleolus construction and rRNA synthesis, interacted with TRF2 and mediated the shuttle of TRF2 between the nucleolus and nucleus. Abating the expression of NOLC1 decreased the nucleolar-resident TRF2. Besides, the nucleolar TRF2 could bind rDNA and promoted rRNA transcription. Furthermore, in hepatocellular carcinoma (HCC) cell lines HepG2 and SMMC7721, TRF2 overexpression participated in the nucleolus stress-induced rRNA inhibition and cell-cycle arrest.

Double strand breaks and cell-cycle arrest induced by the cyanobacterial toxin cylindrospermopsin in HepG2 cells.

PubMed

Alja, Štraser; Filipič, Metka; Novak, Matjaž; Žegura, Bojana

2013-08-21

The newly emerging cyanobacterial cytotoxin cylindrospermopsin (CYN) is increasingly found in surface freshwaters, worldwide. It poses a potential threat to humans after chronic exposure as it was shown to be genotoxic in a range of test systems and is potentially carcinogenic. However, the mechanisms of CYN toxicity and genotoxicity are not well understood. In the present study CYN induced formation of DNA double strand breaks (DSBs), after prolonged exposure (72 h), in human hepatoma cells, HepG2. CYN (0.1-0.5 µg/mL, 24-96 h) induced morphological changes and reduced cell viability in a dose and time dependent manner. No significant increase in lactate dehydrogenase (LDH) leakage could be observed after CYN exposure, indicating that the reduction in cell number was due to decreased cell proliferation and not due to cytotoxicity. This was confirmed by imunocytochemical analysis of the cell-proliferation marker Ki67. Analysis of the cell-cycle using flow-cytometry showed that CYN has an impact on the cell cycle, indicating G0/G1 arrest after 24 h and S-phase arrest after longer exposure (72 and 96 h). Our results provide new evidence that CYN is a direct acting genotoxin, causing DSBs, and these facts need to be considered in the human health risk assessment.

The crucial role of cyclic GMP in the eclosion hormone mediated signal transduction in the silkworm metamorphoses.

PubMed

Shibanaka, Y; Hayashi, H; Okada, N; Fujita, N

1991-10-31

The signal transduction of the peptide, eclosion hormone, in the silkworm Bombyx mori appears to be mediated via the second messenger cyclic GMP throughout their life cycle. Injection of 8-bromo-cGMP induced the ecdysis behavior in pharate adults with similar latency to eclosion hormone-induced ecdysis; the moulting occurred 50-70 min after the injection. The potency of 8Br-cGMP was 10(2) fold higher than that of cGMP and the efficacy was increased by the co-injection of the phosphodiesterase inhibitor IBMX. On the other hand, in the silkworm pupal ecdysis the eclosion hormone and also 8Br-cGMP induced the moulting behavior in a dose-dependent manner. The adult development of the ability to respond to 8Br-cGMP took place concomitantly with the response to the eclosion hormone. Both the developmental time courses were shifted by a shift of light and dark cycles. Accordingly, the sensitivities to the peptide and cyclic nucleotide developed correspondently under the light and dark circadian rhythm. Thus throughout the silkworm life cycle, eclosion hormone is effective to trigger the ecdysis behavior and cGMP plays a crucial role as the second messenger in the eclosion hormone-mediated signal transduction.

New Heart Failure Treatment Capability for Remote Environments

DTIC Science & Technology

2013-06-01

valve in a cycle manner. Control of the absolute pressure, rate of actuation and resistance in the 19 drive lines were the manner in which...Hegde SS, Lowe JE. Sensing Lead Insulation Fractures Following Implantable Cardioverter-Defibrillator Placement. ASAIO Journal 1993;39:M711-M714...14:45-46,1985. Anstadt MP, Galbraith TA, Murray KD, Howanitz EP, Myerowitz PD: Bridge to Cardiac Transplantation Using Prosthetic Biventricular

Regulation of cell function by methionine oxidation and reduction

PubMed Central

Hoshi, Toshinori; Heinemann, Stefan H

2001-01-01

Reactive oxygen species (ROS) are generated during normal cellular activity and may exist in excess in some pathophysiological conditions, such as inflammation or reperfusion injury. These molecules oxidize a variety of cellular constituents, but sulfur-containing amino acid residues are especially susceptible. While reversible cysteine oxidation and reduction is part of well-established signalling systems, the oxidation and the enzymatically catalysed reduction of methionine is just emerging as a novel molecular mechanism for cellular regulation. Here we discuss how the oxidation of methionine to methionine sulfoxide in signalling proteins such as ion channels affects the function of these target proteins. Methionine sulfoxide reductase, which reduces methionine sulfoxide to methionine in a thioredoxin-dependent manner, is therefore not only an enzyme important for the repair of age- or degenerative disease-related protein modifications. It is also a potential missing link in the post-translational modification cycle involved in the specific oxidation and reduction of methionine residues in cellular signalling proteins, which may give rise to activity-dependent plastic changes in cellular excitability. PMID:11179387

Critical ligand binding reagent preparation/selection: when specificity depends on reagents.

PubMed

Rup, Bonita; O'Hara, Denise

2007-05-11

Throughout the life cycle of biopharmaceutical products, bioanalytical support is provided using ligand binding assays to measure the drug product for pharmacokinetic, pharmacodynamic, and immunogenicity studies. The specificity and selectivity of these ligand binding assays are highly dependent on the ligand binding reagents. Thus the selection, characterization, and management processes for ligand binding reagents are crucial to successful assay development and application. This report describes process considerations for selection and characterization of ligand binding reagents that are integral parts of the different phases of assay development. Changes in expression, purification, modification, and storage of the ligand binding reagents may have a profound effect on the ligand binding assay performance. Thus long-term management of the critical ligand binding assay reagents is addressed including suggested characterization criteria that allow ligand binding reagents to be used in as consistent a manner as possible. Examples of challenges related to the selection, modification, and characterization of ligand binding reagents are included.

Functional link between DNA damage responses and transcriptional regulation by ATM in response to a histone deacetylase inhibitor TSA.

PubMed

Lee, Jong-Soo

2007-09-01

Mutations in the ATM (ataxia-telangiectasia mutated) gene, which encodes a 370 kd protein with a kinase catalytic domain, predisposes people to cancers, and these mutations are also linked to ataxia-telangiectasia (A-T). The histone acetylaion/deacetylation- dependent chromatin remodeling can activate the ATM kinase-mediated DNA damage signal pathway (in an accompanying work, Lee, 2007). This has led us to study whether this modification can impinge on the ATM-mediated DNA damage response via transcriptional modulation in order to understand the function of ATM in the regulation of gene transcription. To identify the genes whose expression is regulated by ATM in response to histone deaceylase (HDAC) inhibition, we performed an analysis of oligonucleotide microarrays with using the appropriate cell lines, isogenic A-T (ATM(-)) and control (ATM(+)) cells, following treatment with a HDAC inhibitor TSA. Treatment with TSA reprograms the differential gene expression profile in response to HDAC inhibition in ATM(-) cells and ATM(+) cells. We analyzed the genes that are regulated by TSA in the ATM-dependent manner, and we classified these genes into different functional categories, including those involved in cell cycle/DNA replication, DNA repair, apoptosis, growth/differentiation, cell- cell adhesion, signal transduction, metabolism and transcription. We found that while some genes are regulated by TSA without regard to ATM, the patterns of gene regulation are differentially regulated in an ATM-dependent manner. Taken together, these finding indicate that ATM can regulate the transcription of genes that play critical roles in the molecular response to DNA damage, and this response is modulated through an altered HDAC inhibition-mediated gene expression.

Osthole inhibits gastric cancer cell proliferation through regulation of PI3K/AKT

PubMed Central

Zhang, Yan

2018-01-01

Osthole is an active compound isolated from Chinese herb Cnidium monnieri (L.) Cusson, and had been reported to possess antitumor effect. However, the effect of osthole on the gastric cancer cells has not been investigated. In this study, the effects of osthole on the proliferation of human gastric cancer cells were tested. The data showed that osthole treatment significantly inhibited the proliferation of gastric cancer cells and resulted in the cell cycle arrest at G2/M phase in a dose-dependent manner. Western-blot study showed that the expression of cyclin B1 and cdc2 was markedly reduced by osthole. Moreover, expression of PI3K and pAKT was also significantly suppressed, and the results indicated that the inhibition of pAKT, cyclin B1, and cdc2 levels by osthole was notably enhanced by a PI3K inhibitor. These results demonstrate that osthole could inhibit gastric cancer cells proliferation via induction of cell cycle arrest at G2/M phase by the reduction of PI3K/AKT. PMID:29590128

Osthole inhibits gastric cancer cell proliferation through regulation of PI3K/AKT.

PubMed

Xu, Xiaojun; Liu, Xiaoyuan; Zhang, Yan

2018-01-01

Osthole is an active compound isolated from Chinese herb Cnidium monnieri (L.) Cusson, and had been reported to possess antitumor effect. However, the effect of osthole on the gastric cancer cells has not been investigated. In this study, the effects of osthole on the proliferation of human gastric cancer cells were tested. The data showed that osthole treatment significantly inhibited the proliferation of gastric cancer cells and resulted in the cell cycle arrest at G2/M phase in a dose-dependent manner. Western-blot study showed that the expression of cyclin B1 and cdc2 was markedly reduced by osthole. Moreover, expression of PI3K and pAKT was also significantly suppressed, and the results indicated that the inhibition of pAKT, cyclin B1, and cdc2 levels by osthole was notably enhanced by a PI3K inhibitor. These results demonstrate that osthole could inhibit gastric cancer cells proliferation via induction of cell cycle arrest at G2/M phase by the reduction of PI3K/AKT.

The Silencing of Pokemon Attenuates the Proliferation of Hepatocellular Carcinoma Cells In Vitro and In Vivo by Inhibiting the PI3K/Akt Pathway

PubMed Central

Liu, Yun-Peng; Liu, Jing-Jing; Yang, Xiao-Ning; Jazag, Amarsanaa; Zhang, Zhi-Ping; Guleng, Bayasi; Ren, Jian-Lin

2012-01-01

Pokemon (POK erythroid myeloid ontogenic factor), which belongs to the POK protein family, is also called LRF, OCZF and FBI-1. As a transcriptional repressor, Pokemon assumes a critical function in cellular differentiation and oncogenesis. Our study identified an oncogenic role for Pokemon in human hepatocellular carcinoma (HCC). We successfully established human HepG2 and Huh-7 cell lines in which Pokemon was stably knocked down. We demonstrated that Pokemon silencing inhibited cell proliferation and migration. Pokemon knockdown inhibited the PI3K/Akt and c-Raf/MEK/ERK pathways and modulated the expression of various cell cycle regulators in HepG2 and Huh-7 cells. Therefore, Pokemon may also be involved in cell cycle progression in these cells. We confirmed that Pokemon silencing suppresses hepatocellular carcinoma growth in tumor xenograft mice. These results suggest that Pokemon promotes cell proliferation and migration in hepatocellular carcinoma and accelerates tumor development in an Akt- and ERK-signaling-dependent manner. PMID:23300578

Ulk4 Regulates Neural Stem Cell Pool.

PubMed

Liu, Min; Guan, Zhenlong; Shen, Qin; Flinter, Frances; Domínguez, Laura; Ahn, Joo Wook; Collier, David A; O'Brien, Timothy; Shen, Sanbing

2016-09-01

The size of neural stem cell (NSC) pool at birth determines the starting point of adult neurogenesis. Aberrant neurogenesis is associated with major mental illness, in which ULK4 is proposed as a rare risk factor. Little is known about factors regulating the NSC pool, or function of the ULK4. Here, we showed that Ulk4(tm1a/tm1a) mice displayed a dramatically reduced NSC pool at birth. Ulk4 was expressed in a cell cycle-dependent manner and peaked in G2/M phases. Targeted disruption of the Ulk4 perturbed mid-neurogenesis and significantly reduced cerebral cortex in postnatal mice. Pathway analyses of dysregulated genes in Ulk4(tm1a/tm1a) mice revealed Ulk4 as a key regulator of cell cycle and NSC proliferation, partially through regulation of the Wnt signaling. In addition, we identified hemizygous deletion of ULK4 gene in 1.2/1,000 patients with pleiotropic symptoms including severe language delay and learning difficulties. ULK4, therefore, may significantly contribute to neurodevelopmental, neuropsychiatric, and neurodegenerative disorders. Stem Cells 2016;34:2318-2331. © 2016 AlphaMed Press.

Coordination of size-control, reproduction and generational memory in freshwater planarians

NASA Astrophysics Data System (ADS)

Yang, Xingbo; Kaj, Kelson; Schwab, David; Collins, Eva-Maria

Uncovering the mechanisms that control size, growth, and division rates of systems reproducing through binary division means understanding basic principles of their life cycle. Recent work has focused on how division rates are regulated in bacteria and yeast, but this question has not yet been addressed in more complex, multicellular organisms. We have acquired a unique large-scale data set on the growth and asexual reproduction of two freshwater planarian species, Dugesia japonica and Dugesia tigrina, which reproduce by transverse fission and succeeding regeneration of head and tail pieces into new worms. We developed a new additive theoretical model that mixes multiple size control strategies based on worm size, growth, and waiting time. Our model quantifies the proportions of each strategy in the mixed dynamics, revealing the ability of the two planarian species to utilize different strategies in a coordinated manner for size control. Additionally, we found that head and tail offspring of both species employ different mechanisms to monitor and trigger their reproduction cycles. Finally, we show that generation-dependent memory effects in planarians need to be taken into account to accurately capture the experimental data.

Intercellular propagation of extracellular signal-regulated kinase activation revealed by in vivo imaging of mouse skin

PubMed Central

Hiratsuka, Toru; Fujita, Yoshihisa; Naoki, Honda; Aoki, Kazuhiro; Kamioka, Yuji; Matsuda, Michiyuki

2015-01-01

Extracellular signal-regulated kinase (ERK) is a key effector of many growth signalling pathways. In this study, we visualise epidermal ERK activity in living mice using an ERK FRET biosensor. Under steady-state conditions, the epidermis occasionally revealed bursts of ERK activation patterns where ERK activity radially propagated from cell to cell. The frequency of this spatial propagation of radial ERK activity distribution (SPREAD) correlated with the rate of epidermal cell division. SPREADs and proliferation were stimulated by 12-O-tetradecanoylphorbol 13-acetate (TPA) in a manner dependent on EGF receptors and their cognate ligands. At the wounded skin, ERK activation propagated as trigger wave in parallel to the wound edge, suggesting that ERK activation propagation can be superimposed. Furthermore, by visualising the cell cycle, we found that SPREADs were associated with G2/M cell cycle progression. Our results provide new insights into how cell proliferation and transient ERK activity are synchronised in a living tissue. DOI: http://dx.doi.org/10.7554/eLife.05178.001 PMID:25668746

Host cell subversion by Toxoplasma GRA16, an exported dense granule protein that targets the host cell nucleus and alters gene expression.

PubMed

Bougdour, Alexandre; Durandau, Eric; Brenier-Pinchart, Marie-Pierre; Ortet, Philippe; Barakat, Mohamed; Kieffer, Sylvie; Curt-Varesano, Aurélie; Curt-Bertini, Rose-Laurence; Bastien, Olivier; Coute, Yohann; Pelloux, Hervé; Hakimi, Mohamed-Ali

2013-04-17

After invading host cells, Toxoplasma gondii multiplies within a parasitophorous vacuole (PV) that is maintained by parasite proteins secreted from organelles called dense granules. Most dense granule proteins remain within the PV, and few are known to access the host cell cytosol. We identify GRA16 as a dense granule protein that is exported through the PV membrane and reaches the host cell nucleus, where it positively modulates genes involved in cell-cycle progression and the p53 tumor suppressor pathway. GRA16 binds two host enzymes, the deubiquitinase HAUSP and PP2A phosphatase, which exert several functions, including regulation of p53 and the cell cycle. GRA16 alters p53 levels in a HAUSP-dependent manner and induces nuclear translocation of the PP2A holoenzyme. Additionally, certain GRA16-deficient strains exhibit attenuated virulence, indicating the importance of these host alterations in pathogenesis. Therefore, GRA16 represents a potentially emerging subfamily of exported dense granule proteins that modulate host function. Copyright © 2013 Elsevier Inc. All rights reserved.

Inhibitory effects of β,β-dimethylacrylshikonin on hepatocellular carcinoma in vitro and in vivo.

PubMed

Wu, Yi-ying; Wan, Li-hong; Zheng, Xiao-wei; Shao, Zhen-jun; Chen, Jian; Chen, Xia-jing; Liu, Li-tao; Kuang, Wen-juan; Tan, Xian-shu; Zhou, Li-ming

2012-05-01

β,β-Dimethylacrylshikonin is one of the most abundant naphthoquinones in the root extracts of Lithospermum erythrorhizon Sieb. et Zucc. (Boraginaceae), which have been reported to have antitumor effects. This study evaluated the antiproliferative activity of β,β-dimethylacrylshikonin on human hepatocellular carcinoma (HCC) cells both in vitro and in vivo. In vitro, the MTT assay showed that β,β-dimethylacrylshikonin inhibited the proliferation of SMMC-7721 cells in both dose- and time-dependent manners with its 50% inhibitory concentration (IC(50) ) at 48 h being 15.01 ± 0.76 µg/mL. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) and Hoechst staining detected the characteristics of cell apoptosis in β,β-dimethylacrylshikonin-treated cells and the apoptotic rates of treated groups were increased in a dose-dependent manner. Flow cytometric analysis revealed that β,β-dimethylacrylshikonin could block the cell cycle arrest at G2 phase. Furthermore, β,β-dimethylacrylshikonin down-regulated the mRNA and protein expression of Bcl-2 but up-regulated that of Bax. The cleaved caspase-3 protein was also detected in treated cells. The experiment in vivo showed that β,β-dimethylacrylshikonin significantly suppressed the growth of H(22) transplantable hepatoma, and induced the activation of caspase-3 determined by immunohistochemistry. The results indicate that β,β-dimethylacrylshikonin has significant antitumor effects on hepatocellular carcinoma both in vitro and in vivo. Copyright © 2011 John Wiley & Sons, Ltd.

Menstrual Cycle Effects on Anaerobic Power, Muscular Strength, and Muscular Endurance in Trained and Untrained Females.

ERIC Educational Resources Information Center

Rosenburg, Beth S.; And Others

A study determined if anaerobic power, isometric strength, and isometric endurance are affected by the menstrual cycle and if endurance trained females and untrained females are affected in the same manner on these performance parameters. Subjects were healthy, normally menstruating females, ages 18-34 years who were classified as either trained…

Teaching Complex Activities to Students with Moderate Handicaps through the Forward Chaining of Shorter Total Cycle Response Sequences.

ERIC Educational Resources Information Center

McWilliams, Renee; And Others

1990-01-01

Three adolescents with moderate mental handicaps were taught bedmaking skills through a technique which involved dividing the entire chain into sections, teaching the first section in a total cycle fashion, and adding instruction on subsequent sections in a forward chaining manner. The technique promoted skill acquisition and substantial…

Scorpion (Androctonus bicolor) venom exhibits cytotoxicity and induces cell cycle arrest and apoptosis in breast and colorectal cancer cell lines

PubMed Central

Al-Asmari, Abdulrahman K.; Riyasdeen, Anvarbatcha; Abbasmanthiri, Rajamohamed; Arshaduddin, Mohammed; Al-Harthi, Fahad Ali

2016-01-01

Objectives: The defective apoptosis is believed to play a major role in the survival and proliferation of neoplastic cells. Hence, the induction of apoptosis in cancer cells is one of the targets for cancer treatment. Researchers are considering scorpion venom as a potent natural source for cancer treatment because it contains many bioactive compounds. The main objective of the current study is to evaluate the anticancer property of Androctonus bicolor scorpion venom on cancer cells. Materials and Methods: Scorpions were milked by electrical stimulation of telsons and lyophilized. The breast (MDA-MB-231) and colorectal (HCT-8) cancer cells were maintained in appropriate condition. The venom cytotoxicity was assessed by 3-(4,5-di-methylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay, and the cellular and nuclear changes were studied with propidium iodide and 4’,6-diamidino-2-phenylindole stain, respectively. The cell cycle arrest was examined using muse cell analyzer. Results: The A. bicolor venom exerted cytotoxic effects on MDA-MB-231 and HCT-8 cells in a dose- and duration-dependent manner and induced apoptotic cell death. The treatment with this venom arrests the cancer cells in G0/G1 phase of cell cycle. Conclusions: The venom selectively induces the rate of apoptosis in MDA-MB-231 and HCT-8 cells as reflected by morphological and cell cycle studies. To the best of our knowledge, this is the first scientific evidence demonstrating the induction of apoptosis and cell cycle arrest by A. bicolor scorpion venom. PMID:27721540

Scorpion (Androctonus bicolor) venom exhibits cytotoxicity and induces cell cycle arrest and apoptosis in breast and colorectal cancer cell lines.

PubMed

Al-Asmari, Abdulrahman K; Riyasdeen, Anvarbatcha; Abbasmanthiri, Rajamohamed; Arshaduddin, Mohammed; Al-Harthi, Fahad Ali

2016-01-01

The defective apoptosis is believed to play a major role in the survival and proliferation of neoplastic cells. Hence, the induction of apoptosis in cancer cells is one of the targets for cancer treatment. Researchers are considering scorpion venom as a potent natural source for cancer treatment because it contains many bioactive compounds. The main objective of the current study is to evaluate the anticancer property of Androctonus bicolor scorpion venom on cancer cells. Scorpions were milked by electrical stimulation of telsons and lyophilized. The breast (MDA-MB-231) and colorectal (HCT-8) cancer cells were maintained in appropriate condition. The venom cytotoxicity was assessed by 3-(4,5-di-methylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay, and the cellular and nuclear changes were studied with propidium iodide and 4',6-diamidino-2-phenylindole stain, respectively. The cell cycle arrest was examined using muse cell analyzer. The A. bicolor venom exerted cytotoxic effects on MDA-MB-231 and HCT-8 cells in a dose- and duration-dependent manner and induced apoptotic cell death. The treatment with this venom arrests the cancer cells in G0/G1 phase of cell cycle. The venom selectively induces the rate of apoptosis in MDA-MB-231 and HCT-8 cells as reflected by morphological and cell cycle studies. To the best of our knowledge, this is the first scientific evidence demonstrating the induction of apoptosis and cell cycle arrest by A. bicolor scorpion venom.

Identification of Novel Saccharomyces cerevisiae Proteins with Nuclear Export Activity: Cell Cycle-Regulated Transcription Factor Ace2p Shows Cell Cycle-Independent Nucleocytoplasmic Shuttling

PubMed Central

Jensen, Torben Heick; Neville, Megan; Rain, Jean Christophe; McCarthy, Terri; Legrain, Pierre; Rosbash, Michael

2000-01-01

Nuclear export of proteins containing leucine-rich nuclear export signals (NESs) is mediated by the NES receptor CRM1/Crm1p. We have carried out a yeast two-hybrid screen with Crm1p as a bait. The Crm1p-interacting clones were subscreened for nuclear export activity in a visual assay utilizing the Crm1p-inhibitor leptomycin B (LMB). This approach identified three Saccharomyces cerevisiae proteins not previously known to have nuclear export activity. These proteins are the 5′ RNA triphosphatase Ctl1p, the cell cycle-regulated transcription factor Ace2p, and a protein encoded by the previously uncharacterized open reading frame YDR499W. Mutagenesis analysis show that YDR499Wp contains an NES that conforms to the consensus sequence for leucine-rich NESs. Mutagenesis of Ctl1p and Ace2p were unable to identify specific NES residues. However, a 29-amino-acid region of Ace2p, rich in hydrophobic residues, contains nuclear export activity. Ace2p accumulates in the nucleus at the end of mitosis and activates early-G1-specific genes. We now provide evidence that Ace2p is nuclear not only in late M-early G1 but also during other stages of the cell cycle. This feature of Ace2p localization explains its ability to activate genes such as CUP1, which are not expressed in a cell cycle-dependent manner. PMID:11027275

An APC/C-Cdh1 Biosensor Reveals the Dynamics of Cdh1 Inactivation at the G1/S Transition.

PubMed

Ondracka, Andrej; Robbins, Jonathan A; Cross, Frederick R

2016-01-01

B-type cyclin-dependent kinase activity must be turned off for mitotic exit and G1 stabilization. B-type cyclin degradation is mediated by the anaphase-promoting complex/cyclosome (APC/C); during and after mitotic exit, APC/C is dependent on Cdh1. Cdh1 is in turn phosphorylated and inactivated by cyclin-CDK at the Start transition of the new cell cycle. We developed a biosensor to assess the cell cycle dynamics of APC/C-Cdh1. Nuclear exit of the G1 transcriptional repressor Whi5 is a known marker of Start; APC/C-Cdh1 is inactivated 12 min after Whi5 nuclear exit with little measurable cell-to-cell timing variability. Multiple phosphorylation sites on Cdh1 act in a redundant manner to repress its activity. Reducing the number of phosphorylation sites on Cdh1 can to some extent be tolerated for cell viability, but it increases variability in timing of APC/C-Cdh1 inactivation. Mutants with minimal subsets of phosphorylation sites required for viability exhibit striking stochasticity in multiple responses including budding, nuclear division, and APC/C-Cdh1 activity itself. Multiple cyclin-CDK complexes, as well as the stoichiometric inhibitor Acm1, contribute to APC/C-Cdh1 inactivation; this redundant control is likely to promote rapid and reliable APC/C-Cdh1 inactivation immediately following the Start transition.

Regulation of Cdk7 activity through a phosphatidylinositol (3)-kinase/PKC-ι-mediated signaling cascade in glioblastoma

PubMed Central

Desai, Shraddha R.; Pillai, Prajit P.; Patel, Rekha S.; McCray, Andrea N.; Win-Piazza, Hla Y.; Acevedo-Duncan, Mildred E.

2012-01-01

The objective of this research was to study the potential function of protein kinase C (PKC)-ι in cell cycle progression and proliferation in glioblastoma. PKC-ι is highly overexpressed in human glioma and benign and malignant meningioma; however, little is understood about its role in regulating cell proliferation of glioblastoma. Several upstream molecular aberrations and/or loss of PTEN have been implicated to constitutively activate the phosphatidylinositol (PI) (3)-kinase pathway. PKC-ι is a targeted mediator in the PI (3)-kinase signal transduction repertoire. Results showed that PKC-ι was highly activated and overexpressed in glioma cells. PKC-ι directly associated and phosphorylated Cdk7 at T170 in a cell cycle-dependent manner, phosphorylating its downstream target, cdk2 at T160. Cdk2 has a major role in inducing G1–S phase progression of cells. Purified PKC-ι phosphorylated both endogenous and exogenous Cdk7. PKC-ι downregulation reduced Cdk7 and cdk2 phosphorylation following PI (3)-kinase inhibition, phosphotidylinositol-dependent kinase 1 knockdown as well as PKC-ι silencing (by siRNA treatment). It also diminished cdk2 activity. PKC-ι knockdown inhibited overall proliferation rates and induced apoptosis in glioma cells. These findings suggest that glioma cells may be proliferating through a novel PI (3)-kinase-/PKC-ι/Cdk7/cdk2-mediated pathway. PMID:22021906

Regulation of Cdk7 activity through a phosphatidylinositol (3)-kinase/PKC-ι-mediated signaling cascade in glioblastoma.

PubMed

Desai, Shraddha R; Pillai, Prajit P; Patel, Rekha S; McCray, Andrea N; Win-Piazza, Hla Y; Acevedo-Duncan, Mildred E

2012-01-01

The objective of this research was to study the potential function of protein kinase C (PKC)-ι in cell cycle progression and proliferation in glioblastoma. PKC-ι is highly overexpressed in human glioma and benign and malignant meningioma; however, little is understood about its role in regulating cell proliferation of glioblastoma. Several upstream molecular aberrations and/or loss of PTEN have been implicated to constitutively activate the phosphatidylinositol (PI) (3)-kinase pathway. PKC-ι is a targeted mediator in the PI (3)-kinase signal transduction repertoire. Results showed that PKC-ι was highly activated and overexpressed in glioma cells. PKC-ι directly associated and phosphorylated Cdk7 at T170 in a cell cycle-dependent manner, phosphorylating its downstream target, cdk2 at T160. Cdk2 has a major role in inducing G(1)-S phase progression of cells. Purified PKC-ι phosphorylated both endogenous and exogenous Cdk7. PKC-ι downregulation reduced Cdk7 and cdk2 phosphorylation following PI (3)-kinase inhibition, phosphotidylinositol-dependent kinase 1 knockdown as well as PKC-ι silencing (by siRNA treatment). It also diminished cdk2 activity. PKC-ι knockdown inhibited overall proliferation rates and induced apoptosis in glioma cells. These findings suggest that glioma cells may be proliferating through a novel PI (3)-kinase-/PKC-ι/Cdk7/cdk2-mediated pathway.

The p53 inhibitor, pifithrin-{alpha}, suppresses self-renewal of embryonic stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Abdelalim, Essam Mohamed, E-mail: essam_abdelalim@yahoo.com; Department of Cytology and Histology, Faculty of Veterinary Medicine, Suez Canal University, Ismailia 41522; Tooyama, Ikuo

2012-04-13

Highlights: Black-Right-Pointing-Pointer We determine the role of p53 in ES cells under unstressful conditions. Black-Right-Pointing-Pointer PFT-{alpha} suppresses ES cell proliferation. Black-Right-Pointing-Pointer PFT-{alpha} induces ES cell cycle arrest. Black-Right-Pointing-Pointer PFT-{alpha} downregulates Nanog and cyclin D1. -- Abstract: Recent studies have reported the role of p53 in suppressing the pluripotency of embryonic stem (ES) cells after DNA damage and blocking the reprogramming of somatic cells into induced pluripotent stem (iPS) cells. However, to date no evidence has been presented to support the function of p53 in unstressed ES cells. In this study, we investigated the effect of pifithrin (PFT)-{alpha}, an inhibitor ofmore » p53-dependent transcriptional activation, on self-renewal of ES cells. Our results revealed that treatment of ES cells with PFT-{alpha} resulted in the inhibition of ES cell propagation in a dose-dependent manner, as indicated by a marked reduction in the cell number and colony size. Also, PFT-{alpha} caused a cell cycle arrest and significant reduction in DNA synthesis. In addition, inhibition of p53 activity reduced the expression levels of cyclin D1 and Nanog. These findings indicate that p53 pathway in ES cells rather than acting as an inactive gene, is required for ES cell proliferation and self-renewal under unstressful conditions.« less

5-AED enhances survival of irradiated mice in a G-CSF-dependent manner, stimulates innate immune cell function, reduces radiation-induced DNA damage and induces genes that modulate cell cycle progression and apoptosis

PubMed Central

Grace, Marcy B.; Singh, Vijay K.; Rhee, Juong G.; Jackson, William E.; Kao, Tzu-Cheg; Whitnall, Mark H.

2012-01-01

The steroid androst-5-ene-3ß,17ß-diol (5-androstenediol, 5-AED) elevates circulating granulocytes and platelets in animals and humans, and enhances survival during the acute radiation syndrome (ARS) in mice and non-human primates. 5-AED promotes survival of irradiated human hematopoietic progenitors in vitro through induction of Nuclear Factor-κB (NFκB)-dependent Granulocyte Colony-Stimulating Factor (G-CSF) expression, and causes elevations of circulating G-CSF and interleukin-6 (IL-6). However, the in vivo cellular and molecular effects of 5-AED are not well understood. The aim of this study was to investigate the mechanisms of action of 5-AED administered subcutaneously (s.c.) to mice 24 h before total body γ- or X-irradiation (TBI). We used neutralizing antibodies, flow cytometric functional assays of circulating innate immune cells, analysis of expression of genes related to cell cycle progression, DNA repair and apoptosis, and assessment of DNA strand breaks with halo-comet assays. Neutralization experiments indicated endogenous G-CSF but not IL-6 was involved in survival enhancement by 5-AED. In keeping with known effects of G-CSF on the innate immune system, s.c. 5-AED stimulated phagocytosis in circulating granulocytes and oxidative burst in monocytes. 5-AED induced expression of both bax and bcl-2 in irradiated animals. Cdkn1a and ddb1, but not gadd45a expression, were upregulated by 5-AED in irradiated mice. S.c. 5-AED administration caused decreased DNA strand breaks in splenocytes from irradiated mice. Our results suggest 5-AED survival enhancement is G-CSF-dependent, and that it stimulates innate immune cell function and reduces radiation-induced DNA damage via induction of genes that modulate cell cycle progression and apoptosis. PMID:22843381

5-AED enhances survival of irradiated mice in a G-CSF-dependent manner, stimulates innate immune cell function, reduces radiation-induced DNA damage and induces genes that modulate cell cycle progression and apoptosis.

PubMed

Grace, Marcy B; Singh, Vijay K; Rhee, Juong G; Jackson, William E; Kao, Tzu-Cheg; Whitnall, Mark H

2012-11-01

The steroid androst-5-ene-3ß,17ß-diol (5-androstenediol, 5-AED) elevates circulating granulocytes and platelets in animals and humans, and enhances survival during the acute radiation syndrome (ARS) in mice and non-human primates. 5-AED promotes survival of irradiated human hematopoietic progenitors in vitro through induction of Nuclear Factor-κB (NFκB)-dependent Granulocyte Colony-Stimulating Factor (G-CSF) expression, and causes elevations of circulating G-CSF and interleukin-6 (IL-6). However, the in vivo cellular and molecular effects of 5-AED are not well understood. The aim of this study was to investigate the mechanisms of action of 5-AED administered subcutaneously (s.c.) to mice 24 h before total body γ- or X-irradiation (TBI). We used neutralizing antibodies, flow cytometric functional assays of circulating innate immune cells, analysis of expression of genes related to cell cycle progression, DNA repair and apoptosis, and assessment of DNA strand breaks with halo-comet assays. Neutralization experiments indicated endogenous G-CSF but not IL-6 was involved in survival enhancement by 5-AED. In keeping with known effects of G-CSF on the innate immune system, s.c. 5-AED stimulated phagocytosis in circulating granulocytes and oxidative burst in monocytes. 5-AED induced expression of both bax and bcl-2 in irradiated animals. Cdkn1a and ddb1, but not gadd45a expression, were upregulated by 5-AED in irradiated mice. S.c. 5-AED administration caused decreased DNA strand breaks in splenocytes from irradiated mice. Our results suggest 5-AED survival enhancement is G-CSF-dependent, and that it stimulates innate immune cell function and reduces radiation-induced DNA damage via induction of genes that modulate cell cycle progression and apoptosis.

Nucleotide-dependent assembly of the peroxisomal receptor export complex

PubMed Central

Grimm, Immanuel; Saffian, Delia; Girzalsky, Wolfgang; Erdmann, Ralf

2016-01-01

Pex1p and Pex6p are two AAA-ATPases required for biogenesis of peroxisomes. Both proteins form a hetero-hexameric complex in an ATP-dependent manner, which has a dual localization in the cytosol and at the peroxisomal membrane. At the peroxisomal membrane, the complex is responsible for the release of the import receptor Pex5p at the end of the matrix protein import cycle. In this study, we analyzed the recruitment of the AAA-complex to its anchor protein Pex15p at the peroxisomal membrane. We show that the AAA-complex is properly assembled even under ADP-conditions and is able to bind efficiently to Pex15p in vivo. We reconstituted binding of the Pex1/6p-complex to Pex15p in vitro and show that Pex6p mediates binding to the cytosolic part of Pex15p via a direct interaction. Analysis of the isolated complex revealed a stoichiometry of Pex1p/Pex6p/Pex15p of 3:3:3, indicating that each Pex6p molecule of the AAA-complex binds Pex15p. Binding of the AAA-complex to Pex15p in particular and to the import machinery in general is stabilized when ATP is bound to the second AAA-domain of Pex6p and its hydrolysis is prevented. The data indicate that receptor release in peroxisomal protein import is associated with a nucleotide-depending Pex1/6p-cycle of Pex15p-binding and release. PMID:26842748

Preferential invasion of mitotic cells by Salmonella reveals that cell surface cholesterol is maximal during metaphase.

PubMed

Santos, António J M; Meinecke, Michael; Fessler, Michael B; Holden, David W; Boucrot, Emmanuel

2013-07-15

Cell surface-exposed cholesterol is crucial for cell attachment and invasion of many viruses and bacteria, including the bacterium Salmonella, which causes typhoid fever and gastroenteritis. Using flow cytometry and 3D confocal fluorescence microscopy, we found that mitotic cells, although representing only 1-4% of an exponentially growing population, were much more efficiently targeted for invasion by Salmonella. This targeting was not dependent on the spherical shape of mitotic cells, but was instead SipB and cholesterol dependent. Thus, we measured the levels of plasma membrane and cell surface cholesterol throughout the cell cycle using, respectively, brief staining with filipin and a fluorescent ester of polyethylene glycol-cholesterol that cannot flip through the plasma membrane, and found that both were maximal during mitosis. This increase was due not only to the rise in global cell cholesterol levels along the cell cycle but also to a transient loss in cholesterol asymmetry at the plasma membrane during mitosis. We measured that cholesterol, but not phosphatidylserine, changed from a ∼2080 outerinner leaflet repartition during interphase to ∼5050 during metaphase, suggesting this was specific to cholesterol and not due to a broad change of lipid asymmetry during metaphase. This explains the increase in outer surface levels that make dividing cells more susceptible to Salmonella invasion and perhaps to other viruses and bacteria entering cells in a cholesterol-dependent manner. The change in cholesterol partitioning also favoured the recruitment of activated ERM (Ezrin, Radixin, Moesin) proteins at the plasma membrane and thus supported mitotic cell rounding.

Method for converting heat energy to mechanical energy with 1,2-dichloro-1,1-difluoroethane

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, C.C.; Stiel, L.I.

1980-09-30

1,2-dichloro-1,1-difluoroethane is useful as a power fluid with particular suitability for moderate scale Rankine cycle applications based on systems with moderate temperature heat sources. The fluid is utilized in a Rankine cycle application by vaporizing the fluid by passing the same in heat exchange relationship with a heat source and utilizing the kinetic energy of the resulting expanding vapors to perform work. In this manner heat energy is converted to mechanical energy. The fluid is particularly advantageous in a dual cycle system consisting of a Rankine power cycle combined with a vapor compression cooling or heating cycle.

Functional role of diverse changes in sympathetic nerve activity in regulating arterial pressure during REM sleep.

PubMed

Yoshimoto, Misa; Yoshida, Ikue; Miki, Kenju

2011-08-01

This study aimed to investigate whether REM sleep evoked diverse changes in sympathetic outflows and, if so, to elucidate why REM sleep evokes diverse changes in sympathetic outflows. Male Wistar rats were chronically implanted with electrodes to measure renal (RSNA) and lumbar sympathetic nerve activity (LSNA), electroencephalogram, electromyogram, and electrocardiogram, and catheters to measure systemic arterial and central venous pressure; these parameters were measured simultaneously and continuously during the sleep-awake cycle in the same rat. REM sleep resulted in a step reduction in RNSA by 36.1% ± 2.7% (P < 0.05), while LSNA increased in a step manner by 15.3% ± 2% (P < 0.05) relative to the NREM level. Systemic arterial pressure increased gradually (P < 0.05), while heart rate decreased in a step manner (P < 0.05) during REM sleep. In contrast to REM sleep, RSNA, LSNA, systemic arterial pressure, and heart rate increased in a unidirectional manner associated with increases in physical activity levels in the order from NREM sleep, quiet awake, moving, and grooming state. Thus, the relationship between RSNA vs. LSNA and systemic arterial pressure vs. heart rate observed during REM sleep was dissociated compared with that obtained during the other behavioral states. It is suggested that the diverse changes in sympathetic outflows during REM sleep may be needed to increase systemic arterial pressure by balancing vascular resistance between muscles and vegetative organs without depending on the heart.

Emerging links between the biological clock and the DNA damage response.

PubMed

Collis, Spencer J; Boulton, Simon J

2007-08-01

For life forms to survive, they must adapt to their environmental conditions. One such factor that impacts on both prokaryotic and eukaryotic organisms is the light-dark cycle, a consequence of planetary rotation in relation to our sun. In mammals, the daily light cycle has affected the regulation of many cellular processes such as sleep-wake and calorific intake activities, hormone secretion, blood pressure and immune system responses. Such rhythmic behaviour is the consequence of circadian rhythm/biological clock (BC) systems which are controlled in a light stimulus-dependent manner by a master clock called the suprachiasmatic nucleus (SCN) situated within the anterior hypothalamus. Peripheral clocks located in other organs such as the liver and kidneys relay signals from the SCN, which ultimately leads to tightly controlled expression of several protein families that in turn act on a broad range of cellular functions. Work in lower organisms has demonstrated a link between aging processes and BC factors, and studies in both animal models and clinical trials have postulated a role for certain BC-associated proteins in tumourigenesis and cancer progression. Recent exciting data reported within the last year or so have now established a molecular link between specific BC proteins and factors that control the mammalian cell cycle and DNA damage checkpoints. This mini review will focus on these discoveries and emphasise how such BC proteins may be involved, through their interplay with cell cycle/DNA damage response pathways, in the development of human disease such as cancer.

Cypermethrin Induces Macrophages Death through Cell Cycle Arrest and Oxidative Stress-Mediated JNK/ERK Signaling Regulated Apoptosis

PubMed Central

Huang, Fang; Liu, Qiaoyun; Xie, Shujun; Xu, Jian; Huang, Bo; Wu, Yihua; Xia, Dajing

2016-01-01

Cypermethrin is one of the most highly effective synthetic pyrethroid insecticides. The toxicity of cypermethrin to the reproductive and nervous systems has been well studied. However, little is known about the toxic effect of cypermethrin on immune cells such as macrophages. Here, we investigated the cytotoxicity of cypermethrin on macrophages and the underlying molecular mechanisms. We found that cypermethrin reduced cell viability and induced apoptosis in RAW 264.7 cells. Cypermethrin also increased reactive oxygen species (ROS) production and DNA damage in a dose-dependent manner. Moreover, cypermethrin-induced G1 cell cycle arrest was associated with an enhanced expression of p21, wild-type p53, and down-regulation of cyclin D1, cyclin E and CDK4. In addition, cypermethrin treatment activated MAPK signal pathways by inducing c-Jun N-terminal kinase (JNK) and extracellular regulated protein kinases 1/2 ERK1/2 phosphorylation, and increased the cleaved poly ADP-ribose polymerase (PARP). Further, pretreatment with antioxidant N-acetylcysteine (NAC) effectively abrogated cypermethrin-induced cell cytotoxicity, G1 cell cycle arrest, DNA damage, PARP activity, and JNK and ERK1/2 activation. The specific JNK inhibitor (SP600125) and ERK1/2 inhibitor (PD98059) effectively reversed the phosphorylation level of JNK and ERK1/2, and attenuated the apoptosis. Taken together, these data suggested that cypermethrin caused immune cell death via inducing cell cycle arrest and apoptosis regulated by ROS-mediated JNK/ERK pathway. PMID:27322250

Arctigenin induces cell cycle arrest by blocking the phosphorylation of Rb via the modulation of cell cycle regulatory proteins in human gastric cancer cells.

PubMed

Jeong, Jin Boo; Hong, Se Chul; Jeong, Hyung Jin; Koo, Jin Suk

2011-10-01

Gastric cancer is a leading cause of cancer-related deaths, worldwide being second only to lung cancer as a cause of death. Arctigenin, a representative dibenzylbutyrolactone lignan, occurs in a variety of plants. However, the molecular mechanisms of arctigenin for anti-tumor effect on gastric cancer have not been examined. This study examined the biological effects of arctigenin on the human gastric cancer cell line SNU-1 and AGS. Cell proliferation was determined by MTT assay. In MTT assay, the proliferation of SNU-1 and AGS cells was significantly inhibited by arctigenin in a time and dose dependent manner, as compared with SNU-1 and AGS cells cultured in the absence of arctigenin. Inhibition of cell proliferation by arctigenin was in part associated with apoptotic cell death, as shown by changes in the expression ratio of Bcl-2 to Bax by arctigenin. Also, arctigenin blocked cell cycle arrest from G(1) to S phase by regulating the expression of cell cycle regulatory proteins such as Rb, cyclin D1, cyclin E, CDK4, CDK2, p21Waf1/Cip1 and p15 INK4b. The antiproliferative effect of arctigenin on SNU-1 and AGS gastric cancer cells revealed in this study suggests that arctigenin has intriguing potential as a chemopreventive or chemotherapeutic agent. Crown Copyright © 2011. Published by Elsevier B.V. All rights reserved.

Microscopy and microRaman study of periodically poled domains in deeply thinned lithium niobate wafers

NASA Astrophysics Data System (ADS)

Bullen, P. S.; Huang, H.-C.; Yang, H.; Dadap, J. I.; Kymissis, I.; Osgood, R. M.

2016-07-01

The domain structure of poled deeply thinned lithium niobate is investigated as a function of sample thickness. Free-standing samples of thickness from 25 to 500 μm are prepared by a multiple-cycle polish and annealing procedure and then periodically poled. Using these samples and employing micro-Raman scattering and scanning electron, atomic force, and optical microscopy together, the domain broadening and poling voltage are found to vary in a regular and significant manner. The poled domains show a reduction in width spreading of 38% as the sample thickness is reduced from 500 to 25 μm. Micro-Raman probe measurements verify the quality and the uniformity of the poled domains and provide insight into their thickness-dependent poling contrast.

Antineoplastic Effects of α-Santalol on Estrogen Receptor-Positive and Estrogen Receptor-Negative Breast Cancer Cells through Cell Cycle Arrest at G2/M Phase and Induction of Apoptosis

PubMed Central

Santha, Sreevidya; Bommareddy, Ajay; Rule, Brittny; Guillermo, Ruth; Kaushik, Radhey S.; Young, Alan; Dwivedi, Chandradhar

2013-01-01

Anticancer efficacy and the mechanism of action of α-santalol, a terpenoid isolated from sandalwood oil, were investigated in human breast cancer cells by using p53 wild-type MCF-7 cells as a model for estrogen receptor(ER)-positive and p53 mutated MDA-MB-231 cells as a model for ER-negative breast cancer. α-Santalol inhibited cell viability and proliferation in a concentration and time-dependent manner in both cells regardless of their ER and/or p53 status. However, α-santalol produced relatively less toxic effect on normal breast epithelial cell line, MCF-10A. It induced G2/M cell cycle arrest and apoptosis in both MCF-7 and MDA-MB-231 cells. Cell cycle arrest induced by α-santalol was associated with changes in the protein levels of BRCA1, Chk1, G2/M regulatory cyclins, Cyclin dependent kinases (CDKs), Cell division cycle 25B (Cdc25B), Cdc25C and Ser-216 phosphorylation of Cdc25C. An up-regulated expression of CDK inhibitor p21 along with suppressed expression of mutated p53 was observed in MDA-MB-231 cells treated with α-santalol. On the contrary, α-santalol did not increase the expression of wild-type p53 and p21 in MCF-7 cells. In addition, α-santalol induced extrinsic and intrinsic pathways of apoptosis in both cells with activation of caspase-8 and caspase-9. It led to the activation of the executioner caspase-6 and caspase-7 in α-santalol-treated MCF-7 cells and caspase-3 and caspase-6 in MDA-MB-231 cells along with strong cleavage of poly(ADP-ribose) polymerase (PARP) in both cells. Taken together, this study for the first time identified strong anti-neoplastic effects of α-santalol against both ER-positive and ER-negative breast cancer cells. PMID:23451128

[1-9-NαC]-crourorb A1 isolated from Croton urucurana latex induces G2/M cell cycle arrest and apoptosis in human hepatocarcinoma cells.

PubMed

de Matos Cândido-Bacani, Priscila; Ezan, Frédéric; de Oliveira Figueiredo, Patrícia; Matos, Maria de Fátima Cepa; Rodrigues Garcez, Fernanda; Silva Garcez, Walmir; Baffet, Georges

2017-05-05

[1-9-NαC]-crourorb A1 is a cyclic peptide isolated from Croton urucurana Baillon latex, found in midwestern Brazil, that has been shown to exert cytotoxic effects against a panel of cancer cell lines. However, the underlying mechanisms responsible for the crourorb A1-induced cytotoxicity in cancer cells remain unknown. In this study, the effects of crourorb A1 on the viability, apoptosis, cell cycle and migration of Huh-7 (human hepatocarcinoma) cells were investigated. We evaluated the viability of Huh-7 cells treated with crourorb A1 in 2D and 3D collagen cultures and found that cells in 3D culture exhibited increased resistance to crourorb A1 compared to cells in 2D culture (IC 50 : 62μg/ml versus 35.75μg/ml). Crourorb A1 treatment decreases the viability of Huh-7 cells in a dose- and time-dependent manner and is associated with the induction of apoptosis, in the absence of necrotic cells, through the activation of caspase-3/7 and increased expression of the pro-apoptotic proteins Bak, Bid, Bax, Puma, Bim, and Bad. The effects of crourorb A1 are also associated with G2/M phase cell cycle arrest and increases in cyclin-dependent kinase (CDK1) and cyclin B1 expression. A significant reduction in Huh-7 cell migration induced by crourorb A1 was also observed in the presence of mitomycin C. Finally, we showed that the JNK/MAP pathway, but not ERK signaling, is involved in crourorb A1-induced hepatocarcinoma cell mortality. Copyright © 2017 Elsevier B.V. All rights reserved.

Sulforaphane inhibits PDGF-induced proliferation of rat aortic vascular smooth muscle cell by up-regulation of p53 leading to G1/S cell cycle arrest.

PubMed

Yoo, Su-Hyang; Lim, Yong; Kim, Seung-Jung; Yoo, Kyu-Dong; Yoo, Hwan-Soo; Hong, Jin-Tae; Lee, Mi-Yea; Yun, Yeo-Pyo

2013-01-01

Vascular diseases such as atherosclerosis and restenosis artery angioplasty are associated with vascular smooth muscle cell (VSMC) proliferation and intimal thickening arterial walls. In the present study, we investigated the inhibitory effects of sulforaphane, an isothiocyanate produced in cruciferous vegetables, on VSMC proliferation and neointimal formation in a rat carotid artery injury model. Sulforaphane at the concentrations of 0.5, 1.0, and 2.0 μM significantly inhibited platelet-derived growth factor (PDGF)-BB-induced VSMC proliferation in a concentration-dependent manner, determined by cell count. The IC50 value of sulforaphane-inhibited VSMC proliferation was 0.8 μM. Sulforaphane increased the cyclin-dependent kinase inhibitor p21 and p53 levels, while it decreased CDK2 and cyclin E expression. The effects of sulforaphane on vascular thickening were determined 14 days after the injury to the rat carotid artery. The angiographic mean luminary diameters of the group treated with 2 and 4 μM sulforaphane were 0.25±0.1 and 0.09±0.1 mm², respectively, while the value of the control groups was 0.40±0.1 mm², indicating that sulforaphane may inhibit neointimal formation. The expression of PCNA, maker for cell cycle arrest, was decreased, while that of p53 and p21 was increased, which showed the same pattern as one in in-vitro study. These results suggest that sulforaphane-inhibited VSMC proliferation may occur through the G1/S cell cycle arrest by up-regulation of p53 signaling pathway, and then lead to the decreased neointimal hyperplasia thickening. Thus, sulforaphane may be a promising candidate for the therapy of atherosclerosis and post-angiography restenosis. © 2013.

Small-molecule histone methyltransferase inhibitors display rapid antimalarial activity against all blood stage forms in Plasmodium falciparum

PubMed Central

Malmquist, Nicholas A.; Moss, Thomas A.; Mecheri, Salah; Scherf, Artur; Fuchter, Matthew J.

2012-01-01

Epigenetic factors such as histone methylation control the developmental progression of malaria parasites during the complex life cycle in the human host. We investigated Plasmodium falciparum histone lysine methyltransferases as a potential target class for the development of novel antimalarials. We synthesized a compound library based upon a known specific inhibitor (BIX-01294) of the human G9a histone methyltransferase. Two compounds, BIX-01294 and its derivative TM2-115, inhibited P. falciparum 3D7 parasites in culture with IC50 values of ∼100 nM, values at least 22-fold more potent than their apparent IC50 toward two human cell lines and one mouse cell line. These compounds irreversibly arrested parasite growth at all stages of the intraerythrocytic life cycle. Decrease in parasite viability (>40%) was seen after a 3-h incubation with 1 µM BIX-01294 and resulted in complete parasite killing after a 12-h incubation. Additionally, mice with patent Plasmodium berghei ANKA strain infection treated with a single dose (40 mg/kg) of TM2-115 had 18-fold reduced parasitemia the following day. Importantly, treatment of P. falciparum parasites in culture with BIX-01294 or TM2-115 resulted in significant reductions in histone H3K4me3 levels in a concentration-dependent and exposure time-dependent manner. Together, these results suggest that BIX-01294 and TM2-115 inhibit malaria parasite histone methyltransferases, resulting in rapid and irreversible parasite death. Our data position histone lysine methyltransferases as a previously unrecognized target class, and BIX-01294 as a promising lead compound, in a presently unexploited avenue for antimalarial drug discovery targeting multiple life-cycle stages. PMID:23011794

Activation of pyruvate dehydrogenase by dichloroacetate has the potential to induce epigenetic remodeling in the heart.

PubMed

Matsuhashi, Tomohiro; Hishiki, Takako; Zhou, Heping; Ono, Tomohiko; Kaneda, Ruri; Iso, Tatsuya; Yamaguchi, Aiko; Endo, Jin; Katsumata, Yoshinori; Atsushi, Anzai; Yamamoto, Tsunehisa; Shirakawa, Kohsuke; Yan, Xiaoxiang; Shinmura, Ken; Suematsu, Makoto; Fukuda, Keiichi; Sano, Motoaki

2015-05-01

Dichloroacetate (DCA) promotes pyruvate entry into the Krebs cycle by inhibiting pyruvate dehydrogenase (PDH) kinase and thereby maintaining PDH in the active dephosphorylated state. DCA has recently gained attention as a potential metabolic-targeting therapy for heart failure but the molecular basis of the therapeutic effect of DCA in the heart remains a mystery. Once-daily oral administration of DCA alleviates pressure overload-induced left ventricular remodeling. We examined changes in the metabolic fate of pyruvate carbon (derived from glucose) entering the Krebs cycle by metabolic interventions of DCA. (13)C6-glucose pathway tracing analysis revealed that instead of being completely oxidized in the mitochondria for ATP production, DCA-mediated PDH dephosphorylation results in an increased acetyl-CoA pool both in control and pressure-overloaded hearts. DCA induces hyperacetylation of histone H3K9 and H4 in a dose-dependent manner in parallel to the dephosphorylation of PDH in cultured cardiomyocytes. DCA administration increases histone H3K9 acetylation in in vivo mouse heart. Interestingly, DCA-dependent histone acetylation was associated with an up-regulation of 2.3% of genes (545 out of 23,474 examined). Gene ontology analysis revealed that these genes are highly enriched in transcription-related categories. This evidence suggests that sustained activation of PDH by DCA results in an overproduction of acetyl-CoA, which exceeds oxidation in the Krebs cycle and results in histone acetylation. We propose that DCA-mediated PDH activation has the potential to induce epigenetic remodeling in the heart, which, at least in part, forms the molecular basis for the therapeutic effect of DCA in the heart. Copyright © 2015 Elsevier Ltd. All rights reserved.

Identification of a cast iron alloy containing nonstrategic elements

NASA Technical Reports Server (NTRS)

Cooper, C. V.; Anton, D. L.; Lemkey, F. D.; Nowotny, H.; Bailey, R. S.; Favrow, L. H.; Smeggil, J. G.; Snow, D. B.

1989-01-01

A program was performed to address the mechanical and environmental needs of Stirling engine heater head and regenerator housing components, while reducing the dependence on strategic materials. An alloy was developed which contained no strategic elemental additions per se. The base is iron with additions of manganese, molybdenum, carbon, silicon, niobium, and ferro-chromium. Such an alloy should be producible on a large scale at very low cost. The resulting alloy, designated as NASAUT 4G-Al, contained 15 Mn, 15 Cr, 2 Mo, 1.5 C, 1.0 Si, 1.0 Nb (in weight percent) with a balance of Fe. This alloy was optimized for chemistry, based upon tensile strength, creep-rupture strength, fracture behavior, and fatigue resistance up to 800 C. Alloys were also tested for environmental compatibility. The microstructure and mechanic properties (including hardness) were assessed in the as-cast condition and following several heat treatments, including one designed to simulate a required braze cycle. The alloy was fabricated and characterized in the form of both equiaxed and columnar-grained castings. The columnar grains were produced by directional solidification, and the properties were characterized in both the longitudinal and transverse orientations. The NASAUT 4G-Al alloy was found to be good in cyclic-oxidation resistance and excellent in both hydrogen and hot-corrosion resistance, especially in comparison to the baseline XF-818 alloy. The mechanical properties of yield strength, stress-rupture life, high-cycle-fatigue resistance, and low-cycle-fatigue resistance were good to excellent in comparison to the current alloy for this application, HS-31 (X-40), with precise results depending in a complex manner on grain orientation and temperature. If required, the ductility could be improved by lowering the carbon content.

In vitro and in vivo anti-tumour activities of echinoside A and ds-echinoside A from Pearsonothuria graeffei.

PubMed

Zhao, Qin; Xue, Yong; Wang, Jing-feng; Li, Hui; Long, Teng-teng; Li, Zhaojie; Wang, Yu-ming; Dong, Ping; Xue, Chang-hu

2012-03-15

Echinoside A (EA) and ds-echinoside A (DSEA) are triterpene glycosides isolated from the sea cucumber Pearsonothuria graeffei. DSEA, the desulfurisation product of EA, has the following structure: β-D-xylopyranosyl-holost-8(9),11(12)-diene-3β,17α-diol. In the present study, we examined the anti-tumour activities-in particular, the structure-activity relationships-of EA and DSEA in vitro and in vivo. Both EA and DSEA exhibited an inhibitory effect on cell proliferation, along with apoptosis-inducing activity, in HepG2 cells. Moreover, they significantly arrested the cell cycle in the G₀/G₁ phase. A reverse transcriptase-polymerase chain reaction assay revealed that EA and DSEA significantly increased the expression of the cell-cycle-related genes, namely, p16, p21 and c-myc, and decreased that of cyclin D₁. Western blotting analysis demonstrated that they down-regulated the expression of Bcl-2, and enhanced mitochondria cytochrome c release, caspase-3 activation, and poly(adenosine diphosphate ribose) polymerase, cleavage. Nuclear factor kappa B (NF-κB) expression was significantly decreased by DSEA, but was unaffected by EA. EA and DSEA (2.5 mg kg⁻¹) treatment of mice bearing H22 hepatocarcinoma tumours reduced the tumour weight by 49.8% and 55.0%, respectively. EA and DSEA exhibit marked anti-cancer activity in HepG2 cells, by blocking cell-cycle progression and inducing apoptosis through the mitochondrial pathway. DSEA-induced apoptosis was more potent than EA-induced apoptosis. Furthermore, the two triterpene glycosides derived from P. graeffei may induce apoptosis of HepG2 cells in an NF-κB-dependent or NF-κB-independent manner, depending on their structure. Copyright © 2011 Society of Chemical Industry.

Polyalthia longifolia Methanolic Leaf Extracts (PLME) induce apoptosis, cell cycle arrest and mitochondrial potential depolarization by possibly modulating the redox status in hela cells.

PubMed

Vijayarathna, Soundararajan; Oon, Chern Ein; Chen, Yeng; Kanwar, Jagat R; Sasidharan, Sreenivasan

2017-05-01

Medicinal plants have been accepted as a gold mine, with respect to the diversity of their phytochemicals. Many medicinal plants extracts are potential anticancer agents. Polyalthia longifolia var. angustifolia Thw. (Annonaceae) is one of the most significant native medicinal plants and is found throughout Malaysia. Hence, the present study was intended to assess the anticancer properties of P. longifolia leaf methanolic extract (PLME) and its underlying mechanisms. The Annexin V/PI flow cytometry analysis showed that PLME induces apoptosis in HeLa cells in dose-dependent manner whereas the PI flow cytometric analysis for cell cycle demonstrated the accumulation of cells at sub G0/G1, G0/G1 and G2/M phases. Investigation with JC-1 flow cytometry analysis indicated increase in mitochondria membrane potential depolarisation corresponding to increase in PLME concentrations. PLME was also shown to influence intracellular reactive oxygen species (ROS) by exerting anti-oxidant (half IC 50 ) and pro-oxidant (IC 50 and double IC 50 ) affect against HeLa cells. PLME treatment also displayed DNA damage in HeLa cells in concentration depended fashion. The proteomic profiling array exposed the expression of pro-apoptotic and anti-apoptotic proteins upon PLME treatment at IC 50 concentration in HeLa cells. Pro-apoptotic proteins; BAX, BAD, cytochrome c, caspase-3, p21, p27 and p53 were found to be significantly up-regulated while anti-apoptotic proteins; BCL-2 and BCL-w were found to be significantly down-regulated. This investigation postulated the role of p53 into mediating apoptosis, cell cycle arrest and mitochondrial potential depolarisation by modulating the redox status of HeLa cells. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Dimethylfumarate induces cell cycle arrest and apoptosis via regulating intracellular redox systems in HeLa cells.

PubMed

Han, Guocan; Zhou, Qiang

2016-12-01

Dimethylfumarate (DMF) is cytotoxic to several kinds of cells and serves as an anti-tumor drug. This study was designed to investigate the effects and underlying mechanism of DMF on cervical cancer cells. HeLa cells were cultured and treated with 0, 50, 100, 150, and 200 μM DMF, respectively. After 24 h, cell growth was evaluated using Cell Counting Kit-8 (CCK-8) assay and the cell cycle was examined using flow cytometry. In addition, cell apoptosis was detected by Annexin V/propidium iodide (PI) staining and the expressions of caspase-3 and poly-ADP-ribose polymerase (PARP) were detected using western blotting. The redox-related factors were then assessed. Furthermore, all of the indicators were detected in HeLa cells after combined treatment of DMF and N-acetyl-L-cysteine (NAC, an oxygen-free radical scavenger). The cell number and cell growth of HeLa were obviously inhibited by DMF in a dose-dependent manner, as the cell cycle was arrested at G0/G1 phase (P < 0.05). The apoptotic HeLa cells were markedly increased, and the expression levels of caspase-3 and PARP were significantly increased in a DMF concentration-dependent way (P < 0.05). Meanwhile, loss of △Ψm, increase in reactive oxygen species and O 2 ·- , and the decrease in catalase activity and glutathione (GSH) level were found after DMF treatment (P < 0.05). All these changes were significantly attenuated and even completely disappeared by adding NAC (P < 0.05). In conclusion, the cytotoxicity of DMF on cell proliferation and apoptosis of HeLa cells was mainly related to the intracellular redox systems by depletion of intracellular GSH.

DIFFERENT CONCENTRATIONS OF SIJUNZI DECOCTION INHIBIT PROLIFERATION AND INDUCE APOPTOSIS OF HUMAN GASTRIC CANCER SGC-7901 SIDE POPULATION.

PubMed

Qian, Jun; Li, Jing; Jia, Jianguang; Jin, Xin; Yu, Dajun; Guo, Chenxu; Xie, Bo; Qian, Liyu

2016-01-01

Sijunzi Decoction (SD) is a traditional Chinese medicine which is composed of Ginseng, Atractylodes, Poria and Licorice. It is one of the commonly used Chinese traditional medicines that showed anti-gastric cancer activity in clinical studies. Previous evidence demonstrated SD parties (Ginseng, Atractylodes, Poria, Licorice) can inhibit proliferation and induced apoptosis for gastric cancer cell. In order to further investigate the anticancer effect of SD in gastric cancer, we observed the effects of different concentrations of SD on proliferation and apoptosis of Side Population Cells (SP) of human gastric cancer SGC-7901. SGC-7901 SP and Non- Side Population Cells (NSP) were sorted through flow cytometry; to detect the changes of proliferation of SP and NSP before and after the intervention of serum containing different concentrations of SD using cck-8 method; to detect the changes of cell cycle and apoptosis of SP and NSP before and after the intervention of serum containing different concentrations of SD through flow cytometry; to detect the effects of serum containing different concentrations of SD on apoptosis-related proteins Bax and Bcl-2 of SP and NSP before and after the intervention by western-blot. It was found that different concentrations of SD serum treatments inhibited cell proliferation in a time-dependent and concentration-dependent manner. Compared with the control group (normal saline serum treatment), there were increase in G1/G0 phase population of SP and NSP, and decrease in G2/M and S phase population ( P <0.05). Meanwhile, we found G1/G0 arrest induced by different concentrations of SD serum which was followed by apoptosis in a time-dependent and concentration-dependent manner. The apoptosis rate of SD serum treatment group was higher than the control group ( P <0.05), the apoptosis rate of 48 h treatment was higher than 24 h treatment ( P <0.05), and as the SD serum concentration increases, apoptosis rate is higher and higher ( P <0.05). The expression of Bax protein of SP and NSP was higher than the control group in a time-dependent and concentration dependent manner. The expression of Bcl-2 protein of SP and NSP was lower than the control group in a time-dependent and concentration- dependent manner. With the increase of SD serum concentrations, SD can gradually inhibits the proliferation of SP of SGC-7901 cell lines through G1/G0 phase arrest and followed by apoptosis which involves the up-regulation of Bax and the down-regulation of Bcl-2. List of Abbreviations: (SD) Sijunzi Decoction, (SP) side population, (NSP) non-side population, (Control) normal saline serum group, (L) low concentration SD serum group, (N) normal concentration SD serum group, (H) high concentration SD serum group, (ABCG-2) Adenosine triphosphate Binding Cassette super family G member-2 of transport protein, (Bcl-2) B-cell lymphoma 2, (BAX) Bcl-2 Associated X Protein, (FBS) Fetal bovine serum, (PBS) Phosphate buffer solution, (CCK-8) Cell Counting Kit-8 reagent, (AV) Annexin V-FITC, (PI) Propidium iodide, (EDTA) Ethylene Diamine Tetraacetic Acid, (PMSF) Phenylmethanesulfonyl fluoride, (RIPA) Radio Immunoprecipitation Assay, (PVDF) Poly (vinylidene fluoride), (TBST) Tris-buffered saline containing Tween-20.

Interactive Effects of Dopamine Baseline Levels and Cycle Phase on Executive Functions: The Role of Progesterone.

PubMed

Hidalgo-Lopez, Esmeralda; Pletzer, Belinda

2017-01-01

Estradiol and progesterone levels vary along the menstrual cycle and have multiple neuroactive effects, including on the dopaminergic system. Dopamine relates to executive functions in an "inverted U-shaped" manner and its levels are increased by estradiol. Accordingly, dopamine dependent changes in executive functions along the menstrual cycle have been previously studied in the pre-ovulatory phase, when estradiol levels peak. Specifically it has been demonstrated that working memory is enhanced during the pre-ovulatory phase in women with low dopamine baseline levels, but impaired in women with high dopamine baseline levels. However, the role of progesterone, which peaks in the luteal cycle phase, has not been taken into account previously. Therefore, the main goals of the present study were to extend these findings (i) to the luteal cycle phase and (ii) to other executive functions. Furthermore, the usefulness of the eye blink rate (EBR) as an indicator of dopamine baseline levels in menstrual cycle research was explored. 36 naturally cycling women were tested during three cycle phases (menses-low sex hormones; pre-ovulatory-high estradiol; luteal-high progesterone and estradiol). During each session, women performed a verbal N-back task, as measure of working memory, and a single trial version of the Stroop task, as measure of response inhibition and cognitive flexibility. Hormone levels were assessed from saliva samples and spontaneous eye blink rate was recorded during menses. In the N-back task, women were faster during the luteal phase the higher their progesterone levels, irrespective of their dopamine baseline levels. In the Stroop task, we found a dopamine-cycle interaction, which was also driven by the luteal phase and progesterone levels. For women with higher EBR performance decreased during the luteal phase, whereas for women with lower EBR performance improved during the luteal phase. These findings suggest an important role of progesterone in modulating dopamine-cycle interactions. Additionally, we identified the eye blink rate as a non-invasive indicator of baseline dopamine function in menstrual cycle research.

Conceptual design of closed Brayton cycle for coal-fired power generation

NASA Technical Reports Server (NTRS)

Shah, R. P.; Corman, J. C.

1977-01-01

The objectives to be realized in developing a closed cycle gas turbine are (1) to exploit high temperature gas turbine technology while maintaining a working fluid which is free from combustion gas contamination, (2) to achieve compact turbo-equipment designs through pressurization of the working fluid, and (3) to obtain relatively simple cycle configurations. The technical/economic performance of a specific closed cycle gas turbine system was evaluated through the development of a conceptual plant and system design. This energy conversion system is designed for electric utility service and to utilize coal directly in an environmentally acceptable manner.

Apoptosis induction by silica nanoparticles mediated through reactive oxygen species in human liver cell line HepG2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ahmad, Javed; Ahamed, Maqusood, E-mail: maqusood@gmail.com; Akhtar, Mohd Javed

Silica nanoparticles are increasingly utilized in various applications including agriculture and medicine. In vivo studies have shown that liver is one of the primary target organ of silica nanoparticles. However, possible mechanisms of hepatotoxicity caused by silica nanoparticles still remain unclear. In this study, we explored the reactive oxygen species (ROS) mediated apoptosis induced by well-characterized 14 nm silica nanoparticles in human liver cell line HepG2. Silica nanoparticles (25–200 μg/ml) induced a dose-dependent cytotoxicity in HepG2 cells. Silica nanoparticles were also found to induce oxidative stress in dose-dependent manner indicated by induction of ROS and lipid peroxidation and depletion ofmore » glutathione (GSH). Quantitative real-time PCR and immunoblotting results showed that both the mRNA and protein expressions of cell cycle checkpoint gene p53 and apoptotic genes (bax and caspase-3) were up-regulated while the anti-apoptotic gene bcl-2 was down-regulated in silica nanoparticles treated cells. Moreover, co-treatment of ROS scavenger vitamin C significantly attenuated the modulation of apoptotic markers along with the preservation of cell viability caused by silica nanoparticles. Our data demonstrated that silica nanoparticles induced apoptosis in human liver cells, which is ROS mediated and regulated through p53, bax/bcl-2 and caspase pathways. This study suggests that toxicity mechanisms of silica nanoparticles should be further investigated at in vivo level. -- Highlights: ► We explored the mechanisms of toxicity caused by silica NPs in human liver HepG2 cells. ► Silica NPs induced a dose-dependent cytotoxicity in HepG2 cells. ► Silica NPs induced ROS generation and oxidative stress in a dose-dependent manner. ► Silica NPs were also modulated apoptosis markers both at mRNA and protein levels. ► ROS mediated apoptosis induced by silica NPs was preserved by vitamin C.« less

Helicobacter pylori Induced Phosphatidylinositol-3-OH Kinase/mTOR Activation Increases Hypoxia Inducible Factor-1α to Promote Loss of Cyclin D1 and G0/G1 Cell Cycle Arrest in Human Gastric Cells.

PubMed

Canales, Jimena; Valenzuela, Manuel; Bravo, Jimena; Cerda-Opazo, Paulina; Jorquera, Carla; Toledo, Héctor; Bravo, Denisse; Quest, Andrew F G

2017-01-01

Helicobacter pylori ( H. pylori ) is a human gastric pathogen that has been linked to the development of several gastric pathologies, such as gastritis, peptic ulcer, and gastric cancer. In the gastric epithelium, the bacterium modifies many signaling pathways, resulting in contradictory responses that favor both proliferation and apoptosis. Consistent with such observations, H. pylori activates routes associated with cell cycle progression and cell cycle arrest. H. pylori infection also induces the hypoxia-induced factor HIF-1α, a transcription factor known to promote expression of genes that permit metabolic adaptation to the hypoxic environment in tumors and angiogenesis. Recently, however, also roles for HIF-1α in the repair of damaged DNA and inhibition of gene expression were described. Here, we investigated signaling pathways induced by H. pylori in gastric cells that favor HIF-1α expression and the consequences thereof in infected cells. Our results revealed that H. pylori promoted PI3K/mTOR-dependent HIF-1α induction, HIF-1α translocation to the nucleus, and activity as a transcription factor as evidenced using a reporter assay. Surprisingly, however, transcription of known HIF-1α effector genes evaluated by qPCR analysis, revealed either no change (LDHA and GAPDH), statistically insignificant increases SLC2A1 (GLUT-1) or greatly enhance transcription (VEGFA), but in an HIF-1α-independent manner, as quantified by PCR analysis in cells with shRNA-mediated silencing of HIF-1α. Instead, HIF-1α knockdown facilitated G1/S progression and increased Cyclin D1 protein half-life, via a post-translational pathway. Taken together, these findings link H. pylori -induced PI3K-mTOR activation to HIF-1α induced G0/G1 cell cycle arrest by a Cyclin D1-dependent mechanism. Thus, HIF-1α is identified here as a mediator between survival and cell cycle arrest signaling activated by H. pylori infection.

Bax Translocation Mediated Mitochondrial Apoptosis and Caspase Dependent Photosensitizing Effect of Ficus religiosa on Cancer Cells

PubMed Central

Thankayyan R, Santhosh Kumar; Sithul, Hima; Sreeharshan, Sreeja

2012-01-01

The main aim of the present work was to investigate the potential effect of acetone extract of Ficus religosa leaf (FAE) in multiple apoptosis signalling in human breast cancer cells. FAE treatment significantly induced dose and time dependent, irreversible inhibition of breast cancer cell growth with moderate toxicity to normal breast epithelial cells. This observation was validated using Sulforhodamine B assay. Cell cycle analysis by Flow cytometry showed cell cycle arrest in G1 phase and induction of sub-G0 peak. FAE induced chromatin condensation and displayed an increase in apoptotic population in Annexin V-FITC/PI (Fluorescein isothiocyanate/Propidium iodide) double staining. FAE stimulated the loss of mitochondrial membrane potential in multiple breast cancer cell lines when compared to normal diploid cells. To understand the role of Bax in FAE induced apoptosis, we employed a sensitive cell based platform of MCF-7 cells expressing Bax-EGFP. Bax translocation to mitochondria was accompanied by the disruption of mitochondrial membrane potential and marked elevation in LEHDase activity (Caspase 9). Consistent with this data, FAE induced Caspase activation as evidenced by ratio change in FRET Caspase sensor expressing MCF-7 cell line and cleavage of prominent Caspases and PARP. Interestingly, FAE accelerated cell death in a mitochondrial dependent manner in continuous live cell imaging mode indicating its possible photosensitizing effect. Intracellular generation of reactive oxygen species (ROS) by FAE played a critical role in mediating apoptotic cell death and photosensitizing activity. FAE induced dose and time dependent inhibition of cancer cell growth which was associated with Bax translocation and mitochondria mediated apoptosis with the activation of Caspase 9 dependent Caspase cascade. FAE also possessed strong photosensitizing effect on cancer cell line that was mediated through rapid mitochondrial transmembrane potential loss and partial Caspase activation involving generation of intracellular ROS. PMID:22792212

Effect of Different Stressors on Voluntary Ethanol Intake in Ethanol-Dependent and Nondependent C57BL/6J Mice

PubMed Central

Lopez, Marcelo F.; Anderson, Rachel I.; Becker, Howard C.

2016-01-01

Several animal models have evaluated the effect of stress on voluntary ethanol intake with mixed results. The experiments reported here examined the effects of different stressors on voluntary ethanol consumption in dependent and nondependent adult male C57BL/6J mice. In Experiment 1, restraint, forced swim, and social defeat stress procedures all tended to reduce ethanol intake in nondependent mice regardless of whether the stress experience occurred 1 h or 4 h prior to ethanol access. The reduction in ethanol consumption was most robust following restraint stress. Experiment 2 examined the effects of forced swim stress and social defeat stress on drinking in a dependence model that involved repeated cycles of chronic intermittent ethanol (CIE) exposure. Repeated exposure to forced swim stress prior to intervening test drinking periods that followed repeated cycles of CIE exposure further increased ethanol consumption in CIE-exposed mice while not altering intake in nondependent mice. In contrast, repeated exposure to the social defeat stressor in a similar manner reduced ethanol consumption in CIE-exposed mice while not altering drinking in nondependent mice. Results from Experiment 3 confirmed this selective effect of forced swim stress increasing ethanol consumption in mice with a history of CIE exposure, and also demonstrated that enhanced drinking is only observed when the forced swim stressor is administered during each test drinking week, but not if it is applied only during the final test week. Collectively, these studies point to a unique interaction between repeated stress experience and CIE exposure, and also suggest that such an effect depends on the nature of the stressor. Future studies will need to further explore the generalizability of these results, as well as mechanisms underlying the ability of forced swim stress to selectively further enhance ethanol consumption in dependent (CIE-exposed) mice but not alter intake in nondependent animals. PMID:26992696

Effect of different stressors on voluntary ethanol intake in ethanol-dependent and nondependent C57BL/6J mice.

PubMed

Lopez, Marcelo F; Anderson, Rachel I; Becker, Howard C

2016-03-01

Several animal models have evaluated the effect of stress on voluntary ethanol intake with mixed results. The experiments reported here examined the effects of different stressors on voluntary ethanol consumption in dependent and nondependent adult male C57BL/6J mice. In Experiment 1, restraint, forced swim, and social defeat stress procedures all tended to reduce ethanol intake in nondependent mice regardless of whether the stress experience occurred 1 h or 4 h prior to ethanol access. The reduction in ethanol consumption was most robust following restraint stress. Experiment 2 examined the effects of forced swim stress and social defeat stress on drinking in a dependence model that involved repeated cycles of chronic intermittent ethanol (CIE) exposure. Repeated exposure to forced swim stress prior to intervening test drinking periods that followed repeated cycles of CIE exposure further increased ethanol consumption in CIE-exposed mice while not altering intake in nondependent mice. In contrast, repeated exposure to the social defeat stressor in a similar manner reduced ethanol consumption in CIE-exposed mice while not altering drinking in nondependent mice. Results from Experiment 3 confirmed this selective effect of forced swim stress increasing ethanol consumption in mice with a history of CIE exposure, and also demonstrated that enhanced drinking is only observed when the forced swim stressor is administered during each test drinking week, but not if it is applied only during the final test week. Collectively, these studies point to a unique interaction between repeated stress experience and CIE exposure, and also suggest that such an effect depends on the nature of the stressor. Future studies will need to further explore the generalizability of these results, as well as mechanisms underlying the ability of forced swim stress to selectively further enhance ethanol consumption in dependent (CIE-exposed) mice but not alter intake in nondependent animals. Copyright © 2016 Elsevier Inc. All rights reserved.

Sulforaphane inhibits TGF-β-induced epithelial-mesenchymal transition of hepatocellular carcinoma cells via the reactive oxygen species-dependent pathway.

PubMed

Wu, Jinsheng; Han, Jingli; Hou, Benxin; Deng, Chengwei; Wu, Huanliang; Shen, Liangfang

2016-05-01

Sulforaphane is recognized as a safe antitumor agent derived from various cruciferous vegetables, including broccoli. It has been demonstrated that sulforaphase is a potent antitumor agent in diverse cancers. However, its effect on hepatocellular carcinoma remains largely unknown. Here, we show that sulforaphane inhibits TGF-β-induced epithelial-mesenchymal transition of hepatocellular carcinoma cell via the reactive oxygen species-dependent pathway. We found sulforaphane inhibited hepatocellular carcinoma cell proliferation in a dose- and time-dependent manner. Sulforaphane induced G0/G1 phase cell cycle arrest and promoted cell apoptosis. A set of experiments showed that sulforaphase inhibited hepatocellular carcinoma cell migration and invasion, inhibited the formation of fibroblast like mesenchymal cells and the expression of Vimentin, but increased the expression of E-cadherin, suggesting sulforaphane suppresses epithelial-mesenchymal transition (EMT) process. Cotreatment with N-acetyl-L-cysteine inhibited sulforaphane-inhibited invasion and upregulation of E-cadherin and almost completely abolished the sulforaphane-induced expression of Vimentin. The effect of sulforaphane on the growth of hepatocellular carcinoma cells was confirmed by a xenograft tumor growth model. All our finding indicated that sulforaphane is a promising and safe strategy for treating hepatocellular carcinoma.

Rescue of p53 function by small-molecule RITA in cervical carcinoma by blocking E6-mediated degradation.

PubMed

Zhao, Carolyn Ying; Szekely, Laszlo; Bao, Wenjie; Selivanova, Galina

2010-04-15

Proteasomal degradation of p53 by human papilloma virus (HPV) E6 oncoprotein plays a pivotal role in the survival of cervical carcinoma cells. Abrogation of HPV-E6-dependent p53 destruction can therefore be a good strategy to combat cervical carcinomas. Here, we show that a small-molecule reactivation of p53 and induction of tumor cell apoptosis (RITA) is able to induce the accumulation of p53 and rescue its tumor suppressor function in cells containing high-risk HPV16 and HPV18 by inhibiting HPV-E6-mediated proteasomal degradation. RITA blocks p53 ubiquitination by preventing p53 interaction with E6-associated protein, required for HPV-E6-mediated degradation. RITA activates the transcription of proapoptotic p53 targets Noxa, PUMA, and BAX, and repressed the expression of pro-proliferative factors CyclinB1, CDC2, and CDC25C, resulting in p53-dependent apoptosis and cell cycle arrest. Importantly, RITA showed substantial suppression of cervical carcinoma xenografts in vivo. These results provide a proof of principle for the treatment of cervical cancer in a p53-dependent manner by using small molecules that target p53. (c)2010 AACR.

RNA helicase MOV10 functions as a co-factor of HIV-1 Rev to facilitate Rev/RRE-dependent nuclear export of viral mRNAs.

PubMed

Huang, Feng; Zhang, Junsong; Zhang, Yijun; Geng, Guannan; Liang, Juanran; Li, Yingniang; Chen, Jingliang; Liu, Chao; Zhang, Hui

2015-12-01

Human immunodeficiency virus type 1 (HIV-1) exploits multiple host factors during its replication. The REV/RRE-dependent nuclear export of unspliced/partially spliced viral transcripts needs the assistance of host proteins. Recent studies have shown that MOV10 overexpression inhibited HIV-1 replication at various steps. However, the endogenous MOV10 was required in certain step(s) of HIV-1 replication. In this report, we found that MOV10 potently enhances the nuclear export of viral mRNAs and subsequently increases the expression of Gag protein and other late products through affecting the Rev/RRE axis. The co-immunoprecipitation analysis indicated that MOV10 interacts with Rev in an RNA-independent manner. The DEAG-box of MOV10 was required for the enhancement of Rev/RRE-dependent nuclear export and the DEAG-box mutant showed a dominant-negative activity. Our data propose that HIV-1 utilizes the anti-viral factor MOV10 to function as a co-factor of Rev and demonstrate the complicated effects of MOV10 on HIV-1 life cycle. Copyright © 2015 Elsevier Inc. All rights reserved.

A common microstructure in behavioral hearing thresholds and stimulus-frequency otoacoustic emissions.

PubMed

Dewey, James B; Dhar, Sumitrajit

2017-11-01

Behavioral hearing thresholds and otoacoustic emission (OAE) spectra often exhibit quasiperiodic fluctuations with frequency. For behavioral and OAE responses to single tones-the latter referred to as stimulus-frequency otoacoustic emissions (SFOAEs)-this microstructure has been attributed to intracochlear reflections of SFOAE energy between its region of generation and the middle ear boundary. However, the relationship between behavioral and SFOAE microstructures, as well as their presumed dependence on the properties of the SFOAE-generation mechanism, have yet to be adequately examined. To address this, behavioral thresholds and SFOAEs evoked by near-threshold tones were compared in 12 normal-hearing female subjects. The microstructures observed in thresholds and both SFOAE amplitudes and delays were found to be strikingly similar. SFOAE phase accumulated an integer number of cycles between the frequencies of microstructure maxima, consistent with a dependence of microstructure periodicity on SFOAE propagation delays. Additionally, microstructure depth was correlated with SFOAE magnitude in a manner resembling that predicted by the intracochlear reflection framework, after assuming reasonable values of parameters related to middle ear transmission. Further exploration of this framework may yield more precise estimates of such parameters and provide insight into their frequency dependence.

Simulation Propulsion System and Trajectory Optimization

NASA Technical Reports Server (NTRS)

Hendricks, Eric S.; Falck, Robert D.; Gray, Justin S.

2017-01-01

A number of new aircraft concepts have recently been proposed which tightly couple the propulsion system design and operation with the overall vehicle design and performance characteristics. These concepts include propulsion technology such as boundary layer ingestion, hybrid electric propulsion systems, distributed propulsion systems and variable cycle engines. Initial studies examining these concepts have typically used a traditional decoupled approach to aircraft design where the aerodynamics and propulsion designs are done a-priori and tabular data is used to provide inexpensive look ups to the trajectory ana-ysis. However the cost of generating the tabular data begins to grow exponentially when newer aircraft concepts require consideration of additional operational parameters such as multiple throttle settings, angle-of-attack effects on the propulsion system, or propulsion throttle setting effects on aerodynamics. This paper proposes a new modeling approach that eliminated the need to generate tabular data, instead allowing an expensive propulsion or aerodynamic analysis to be directly integrated into the trajectory analysis model and the entire design problem optimized in a fully coupled manner. The new method is demonstrated by implementing a canonical optimal control problem, the F-4 minimum time-to-climb trajectory optimization using three relatively new analysis tools: Open M-DAO, PyCycle and Pointer. Pycycle and Pointer both provide analytic derivatives and Open MDAO enables the two tools to be combined into a coupled model that can be run in an efficient parallel manner that helps to cost the increased cost of the more expensive propulsion analysis. Results generated with this model serve as a validation of the tightly coupled design method and guide future studies to examine aircraft concepts with more complex operational dependencies for the aerodynamic and propulsion models.

Effects of nitrogen-doped multi-walled carbon nanotubes compared to pristine multi-walled carbon nanotubes on human small airway epithelial cells.

PubMed

Mihalchik, Amy L; Ding, Weiqiang; Porter, Dale W; McLoughlin, Colleen; Schwegler-Berry, Diane; Sisler, Jennifer D; Stefaniak, Aleksandr B; Snyder-Talkington, Brandi N; Cruz-Silva, Rodolfo; Terrones, Mauricio; Tsuruoka, Shuji; Endo, Morinobu; Castranova, Vincent; Qian, Yong

2015-07-03

Nitrogen-doped multi-walled carbon nanotubes (ND-MWCNTs) are modified multi-walled carbon nanotubes (MWCNTs) with enhanced electrical properties that are used in a variety of applications, including fuel cells and sensors; however, the mode of toxic action of ND-MWCNT has yet to be fully elucidated. In the present study, we compared the interaction of ND-MWCNT or pristine MWCNT-7 with human small airway epithelial cells (SAEC) and evaluated their subsequent bioactive effects. Transmission electron microscopy, X-ray photoelectron spectroscopy, Raman spectroscopy, and X-ray diffraction suggested the presence of N-containing defects in the lattice of the nanotube. The ND-MWCNTs were determined to be 93.3% carbon, 3.8% oxygen, and 2.9% nitrogen. A dose-response cell proliferation assay showed that low doses of ND-MWCNT (1.2μg/ml) or MWCNT-7 (0.12μg/ml) increased cellular proliferation, while the highest dose of 120μg/ml of either material decreased proliferation. ND-MWCNT and MWCNT-7 appeared to interact with SAEC at 6h and were internalized by 24h. ROS were elevated at 6 and 24h in ND-MWCNT exposed cells, but only at 6h in MWCNT-7 exposed cells. Significant alterations to the cell cycle were observed in SAEC exposed to either 1.2μg/ml of ND-MWCNT or MWCNT-7 in a time and material-dependent manner, possibly suggesting potential damage or alterations to cell cycle machinery. Our results indicate that ND-MWCNT induce effects in SAEC over a time and dose-related manner which differ from MWCNT-7. Therefore, the physicochemical characteristics of the materials appear to alter their biological effects. Published by Elsevier Ireland Ltd.

Development of real-world driving cycles and estimation of emission factors for in-use light-duty gasoline vehicles in urban areas.

PubMed

Hwa, Mei-Yin; Yu, Tai-Yi

2014-07-01

This investigation adopts vehicle tracking manner to establish real-world driving patterns and estimates emission factors with dynamometers with 23 traffic-driving variables for 384 in-use light-duty passenger vehicles during non-rush hour. Adequate numbers of driving variables were decided with factor analysis and cluster analysis. The dynamometer tests were performed on FTP75 cycle and five local driving cycles derived from real-world speed profiles. Results presented that local driving cycles and FTP75 cycle were completely different in driving characteristic parameters of typical driving cycles and emission factors. The highest values of emission factor ratios of local driving cycle and FTP75 cycle for CO, NMHC, NO x , CH4, and CO2 were 1.38, 1.65, 1.58, 1.39, and 1.14, respectively.

41 CFR 109-1.5302 - Policies.

Code of Federal Regulations, 2011 CFR

2011-01-01

... efficient manner. High risk personal property will be managed throughout its life cycle so as to protect... transferred or disposed unless it receives a high risk assessment and is handled accordingly. ...

Ascorbyl stearate and ionizing radiation potentiate apoptosis through intracellular thiols and oxidative stress in murine T lymphoma cells.

PubMed

Mane, Shirish D; Kamatham, Akhilender Naidu

2018-02-01

Ascorbyl stearate (Asc-s) is a derivative of ascorbic acid with better anti-tumour efficacy compared to its parent compound ascorbic acid. In this study, we have examined radio-sensitizing effect of Asc-s in murine T cell lymphoma (EL4) cells at 4 Gy. Asc-s and radiation treatment reduced cell proliferation, induced apoptosis in a dose dependent manner by arresting the cells at S/G2-M phase of cell cycle. It also decreased the frequency of cancer stem cells per se, with significantly higher decrease in combination with radiation treatment./Further, Asc-s and radiation treatment increased the level of reactive oxygen species (ROS), drop in mitochondrial membrane potential (MMP) and increased caspase-3 activity resulting in apoptosis of EL4 cells. Further it also significantly decreased GSH/GSSG ratio due to binding of Asc-s with thiols. The increase in oxidative stress induced by Asc-s and radiation treatment was abrogated by thiol antioxidants in EL4 cells. Interestingly, this redox modulation triggered significant increase in protein glutathionylation in a time dependent manner. Asc-s treatment resulted in glutathionylation of IKK, p50-NF-kB and mutated p53, thereby inhibiting cancer progression during oxidative stress. Asc-s quenches GSH ensuing Asc-s + GSH adduct thereby further modulating GSH/GSSG ratio as evident from HPLC and docking studies. The anti-tumour effect of Asc-s along with radiation was studied by injecting EL4 cells in synegenicC57/BL6 male mice. Intraperitoneal injection of Asc-s followed by radiation exposure at 4 Gy to the tumour bearing mice resulted in radio-sensitization which is evident from significant regression of tumour as evident from tumour burden index. The survival study supports the data that Asc-s pre-treatment enhances radio-sensitization in murine lymphoma. Our data, suggest that Asc-s and ionizing radiation induced cell cycle arrest and apoptosis by perturbing redox balance through irreversible complexes of thiols with Asc-s, disturbed mitochondrial membrane permeability and activation of caspase-3 in EL4 cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Mechanisms and rates of strength recovery in laboratory fault zones

NASA Astrophysics Data System (ADS)

Muhuri, Sankar Kumar

2001-07-01

The life cycle of a typical fault zone consists of repeated catastrophic seismic events during which much of the slip is accommodated interspersed with creep during the inter-seismic cycle. Fault strength is regenerated during this period as a result of several time-dependent, fluid assisted deformation mechanisms that are favored by high stresses along active fault zones. The strengthening is thought to be a function of the sum total of the rates of recovery due to these multiple creep processes as well as the rate of tectonic loading. Mechanisms and rates of strength recovery in laboratory fault zones were investigated in this research with the aid of several experimental designs. It was observed that wet faults recover strength in a time-dependent manner after slip due to operative creep processes. Subsequent loading results in unstable failure of a cohesive gouge zone with large associated stress drops. The failure process is similar to that observed for intact rocks. Dry laboratory faults in contrast do not recover strength and slip along them is always stable with no observable drop in stress. Strengthening in laboratory faults proceeds in a manner that is a logarithmic function of time. The recovery is attributable to fluid mediated mechanisms such as pressure solution, crack sealing and Ostwald ripening that collectively cause a reduction in porosity and enhance lithification of an unconsolidated gouge. Rates for the individual deformation mechanisms investigated in separate experimental setups were also observed to be a non-linear function of time. Pressure solution and Ostwald ripening are especially enhanced due to the significant volume fraction of fine particles within the gouge created due to cataclasis during slip. The results of this investigation may be applied to explain observations of rapid strengthening along large, active crustal fault zones such as parts of the San Andreas Fault system in California and the Nojima fault in Japan. Presence of fault seals in clean hydrocarbon reservoirs with minor clay content as in several North Sea fields may also be a manifestation of similar deformation processes.

Bipolar mood cycles and lunar tidal cycles.

PubMed

Wehr, T A

2018-04-01

In 17 patients with rapid cycling bipolar disorder, time-series analyses detected synchronies between mood cycles and three lunar cycles that modulate the amplitude of the moon's semi-diurnal gravimetric tides: the 14.8-day spring-neap cycle, the 13.7-day declination cycle and the 206-day cycle of perigee-syzygies ('supermoons'). The analyses also revealed shifts among 1:2, 1:3, 2:3 and other modes of coupling of mood cycles to the two bi-weekly lunar cycles. These shifts appear to be responses to the conflicting demands of the mood cycles' being entrained simultaneously to two different bi-weekly lunar cycles with slightly different periods. Measurements of circadian rhythms in body temperature suggest a biological mechanism through which transits of one of the moon's semi-diurnal gravimetric tides might have driven the patients' bipolar cycles, by periodically entraining the circadian pacemaker to its 24.84-h rhythm and altering the pacemaker's phase-relationship to sleep in a manner that is known to cause switches from depression to mania.

Aging and insulin signaling differentially control normal and tumorous germline stem cells.

PubMed

Kao, Shih-Han; Tseng, Chen-Yuan; Wan, Chih-Ling; Su, Yu-Han; Hsieh, Chang-Che; Pi, Haiwei; Hsu, Hwei-Jan

2015-02-01

Aging influences stem cells, but the processes involved remain unclear. Insulin signaling, which controls cellular nutrient sensing and organismal aging, regulates the G2 phase of Drosophila female germ line stem cell (GSC) division cycle in response to diet; furthermore, this signaling pathway is attenuated with age. The role of insulin signaling in GSCs as organisms age, however, is also unclear. Here, we report that aging results in the accumulation of tumorous GSCs, accompanied by a decline in GSC number and proliferation rate. Intriguingly, GSC loss with age is hastened by either accelerating (through eliminating expression of Myt1, a cell cycle inhibitory regulator) or delaying (through mutation of insulin receptor (dinR) GSC division, implying that disrupted cell cycle progression and insulin signaling contribute to age-dependent GSC loss. As flies age, DNA damage accumulates in GSCs, and the S phase of the GSC cell cycle is prolonged. In addition, GSC tumors (which escape the normal stem cell regulatory microenvironment, known as the niche) still respond to aging in a similar manner to normal GSCs, suggesting that niche signals are not required for GSCs to sense or respond to aging. Finally, we show that GSCs from mated and unmated females behave similarly, indicating that female GSC-male communication does not affect GSCs with age. Our results indicate the differential effects of aging and diet mediated by insulin signaling on the stem cell division cycle, highlight the complexity of the regulation of stem cell aging, and describe a link between ovarian cancer and aging. © 2014 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

Changes of Constituents and Activity to Apoptosis and Cell Cycle During Fermentation of Tea

PubMed Central

Zhao, Hang; Zhang, Min; Zhao, Lu; Ge, Ya-kun; Sheng, Jun; Shi, Wei

2011-01-01

Tea is believed to be beneficial for health, and the effects of the fermentation process on its contributions to apoptosis and cell cycle arrest of gastric cancer cells have not been completely investigated. In this study, the chemical components in green tea, black tea and pu-erh tea aqueous extracts were analyzed and compared. The polysaccharide and caffeine levels were substantially higher in the fermented black tea and pu-erh tea, while the polyphenol level was higher in the unfermented green tea. Hence, a treatment of tea aqueous extract and the components, which are emerging as promising anticancer agents, were pursued to determine whether this treatment could lead to enhance apoptosis and cell cycle arrest. In the human gastric cancer cell line SGC-7901, the cell viability and flow cytometry analysis for apoptotic cells indicated effects in a dose-dependent inhibition manner for the three tea treatment groups. The apoptosis rates were found to be elevated after 48 h of treatment with 31.2, 125, and 500 μg/mL of green tea extract, the higher catechins content may be involved in the mechanism. Cell cycle was arrested in S phase in the fermented black tea and pu-erh tea, and the populations were significantly decreased in G2/M phases, possibly due to the oxidation of tea polyphenols, which causes an increase of theabrownins. CCC-HEL-1 normal cells were not sensitive to tea extract. These findings suggest that the fermentation process causes changes of the compounds which might be involved in the changes of cell proliferation inhibition, apoptosis induction and cell cycle arrest. PMID:21673927

Effect of sodium butyrate on cell proliferation and cell cycle in porcine intestinal epithelial (IPEC-J2) cells.

PubMed

Qiu, Yueqin; Ma, Xianyong; Yang, Xuefen; Wang, Li; Jiang, Zongyong

2017-04-01

Conflicting results have been reported that butyrate in normal piglets leads either to an increase or to a decrease of jejunal villus length, implying a possible effect on the proliferation of enterocytes. No definitive study was found for the biological effects of butyrate in porcine jejunal epithelial cells. The present study used IPEC-J2 cells, a non-transformed jejunal epithelial line to evaluate the direct effects of sodium butyrate on cell proliferation, cell cycle regulation, and apoptosis. Low concentrations (0.5 and 1 mM) of butyrate had no effect on cell proliferation. However, at 5 and 10 mM, sodium butyrate significantly decreased cell viability, accompanied by reduced levels of p-mTOR and PCNA protein. Sodium butyrate, in a dose-dependent manner, induced cell cycle arrest in G0/G1 phase and reduced the numbers of cells in S phase. In addition, relative expression of p21, p27, and pro-apoptosis bak genes, and protein levels of p21Waf1/Cip1, p27Kip1, cyclinD3, CDK4, and Cleave-caspase3 were increased by higher concentrations of sodium butyrate (1, 5, 10 mM), and the levels of cyclinD1 and CDK6 were reduced by 5 and 10 mM butyrate. Butyrate increased the phosphorylated form of the signaling molecule p38 and phosphorylated JNK. In conclusion, the present in vitro study indicated that sodium butyrate inhibited the proliferation of IPEC-J2 cells by inducing cell cycle arrest in the G0/G1 phase of cell cycles and by increasing apoptosis at high concentrations.

Rare sugar D-allose induces specific up-regulation of TXNIP and subsequent G1 cell cycle arrest in hepatocellular carcinoma cells by stabilization of p27kip1.

PubMed

Yamaguchi, Fuminori; Takata, Maki; Kamitori, Kazuyo; Nonaka, Machiko; Dong, Youyi; Sui, Li; Tokuda, Masaaki

2008-02-01

'Rare sugars' are defined as monosaccharides that exist in nature but are only present in limited quantities. The development of mass production method of rare sugars revealed some interesting physiological effects of these on animal cells, but the mechanisms have not been well studied. We examined the effect of D-allose on the proliferation of cancer cells and the underlying molecular mechanism of the action. The HuH-7 hepatocellular carcinoma cells were treated with various monosaccharides for 48 h and D-allose was shown to inhibit cell growth by 40% in a dose-dependent manner. D-allose induced G1 cell cycle arrest but not apoptosis. The microarray analysis revealed that D-allose significantly up-regulated thioredoxin interacting protein (TXNIP) gene expression, which is often suppressed in tumor cells and western blot analysis confirmed its increase at protein level. The overexpression of TXNIP also induced G1 cell cycle arrest. Analysis of cell cycle regulatory genes showed p27kip1, a key regulator of G1/S cell cycle transition, to be increased at the protein but not the transcriptional level. Protein interaction between TXNIP and jab1, and p27kip1 and jab1, was observed, suggesting stabilization of p27kip1 protein by the competitive inhibition of jab1-mediated nuclear export of p27kip1 by TXNIP. In addition, increased interaction and nuclear localization of TXNIP and p27kip1 were apparent after D-allose treatment. Our findings surprisingly suggest that D-allose, a simple monosaccharide, may act as a novel anticancer agent via unique TXNIP induction and p27kip1 protein stabilization.

pRb phosphorylation regulates the proliferation of supporting cells in gentamicin-damaged neonatal avian utricle.

PubMed

Wu, Jingfang; Sun, Shan; Li, Wenyan; Chen, Yan; Li, Huawei

2014-10-01

The ability of nonmammalian vertebrates to regenerate hair cells (HCs) after damage-induced HC loss has stimulated and inspired research in the field of HC regeneration. The protein pRb encoded by retinoblastoma gene Rb1 forces sensory progenitor cells to exit cell cycle and maintain differentiated HCs and supporting cells (SCs) in a quiescent state. pRb function is regulated by phosphorylation through the MEK/ERK or the pRb/Raf-1 signaling pathway. In our previous study, we have shown that pRb phosphorylation is crucial for progenitor cell proliferation and survival during the early embryonic stage of avian otocyst sensory epithelium development. However, in damaged avian utricle, the role of pRb in regulating the cell cycling of SCs or HCs regeneration still remains unclear. To further elucidate the function of pRb phosphorylation on SCs re-entering the cell cycle triggered by gentamycin-induced HCs damage, we isolated neonatal chicken utricles and treated them with the MEK inhibitor U0126 or the pRb/Raf-1 inhibitor RRD-251, respectively in vitro. We found that after gentamycin-induced HCs damage, pRb phosphorylation is important for the quiescent SCs re-entering the cell cycle in the neonatal chicken utricle. In addition, the proliferation of SCs decreased in a dose-dependent manner in response to both U0126 and RRD-251, which indicates that both the MEK/ERK and the pRb/Raf-1 signaling pathway play important roles in pRb phosphorylation in damaged neonatal chicken utricle. Together, these findings on the function of pRb in damaged neonatal chicken utricle improve our understanding of the regulation of the cell cycle of SCs after HCs loss and may shed light on the mammalian HC regeneration from SCs in damaged organs.

Changes of constituents and activity to apoptosis and cell cycle during fermentation of tea.

PubMed

Zhao, Hang; Zhang, Min; Zhao, Lu; Ge, Ya-Kun; Sheng, Jun; Shi, Wei

2011-01-01

Tea is believed to be beneficial for health, and the effects of the fermentation process on its contributions to apoptosis and cell cycle arrest of gastric cancer cells have not been completely investigated. In this study, the chemical components in green tea, black tea and pu-erh tea aqueous extracts were analyzed and compared. The polysaccharide and caffeine levels were substantially higher in the fermented black tea and pu-erh tea, while the polyphenol level was higher in the unfermented green tea. Hence, a treatment of tea aqueous extract and the components, which are emerging as promising anticancer agents, were pursued to determine whether this treatment could lead to enhance apoptosis and cell cycle arrest. In the human gastric cancer cell line SGC-7901, the cell viability and flow cytometry analysis for apoptotic cells indicated effects in a dose-dependent inhibition manner for the three tea treatment groups. The apoptosis rates were found to be elevated after 48 h of treatment with 31.2, 125, and 500 μg/mL of green tea extract, the higher catechins content may be involved in the mechanism. Cell cycle was arrested in S phase in the fermented black tea and pu-erh tea, and the populations were significantly decreased in G2/M phases, possibly due to the oxidation of tea polyphenols, which causes an increase of theabrownins. CCC-HEL-1 normal cells were not sensitive to tea extract. These findings suggest that the fermentation process causes changes of the compounds which might be involved in the changes of cell proliferation inhibition, apoptosis induction and cell cycle arrest.

DELAY OF GERMINATION 1 mediates a conserved coat-dormancy mechanism for the temperature- and gibberellin-dependent control of seed germination

PubMed Central

Graeber, Kai; Linkies, Ada; Steinbrecher, Tina; Mummenhoff, Klaus; Tarkowská, Danuše; Turečková, Veronika; Ignatz, Michael; Sperber, Katja; Voegele, Antje; de Jong, Hans; Urbanová, Terezie; Strnad, Miroslav; Leubner-Metzger, Gerhard

2014-01-01

Seed germination is an important life-cycle transition because it determines subsequent plant survival and reproductive success. To detect optimal spatiotemporal conditions for germination, seeds act as sophisticated environmental sensors integrating information such as ambient temperature. Here we show that the DELAY OF GERMINATION 1 (DOG1) gene, known for providing dormancy adaptation to distinct environments, determines the optimal temperature for seed germination. By reciprocal gene-swapping experiments between Brassicaceae species we show that the DOG1-mediated dormancy mechanism is conserved. Biomechanical analyses show that this mechanism regulates the material properties of the endosperm, a seed tissue layer acting as germination barrier to control coat dormancy. We found that DOG1 inhibits the expression of gibberellin (GA)-regulated genes encoding cell-wall remodeling proteins in a temperature-dependent manner. Furthermore we demonstrate that DOG1 causes temperature-dependent alterations in the seed GA metabolism. These alterations in hormone metabolism are brought about by the temperature-dependent differential expression of genes encoding key enzymes of the GA biosynthetic pathway. These effects of DOG1 lead to a temperature-dependent control of endosperm weakening and determine the optimal temperature for germination. The conserved DOG1-mediated coat-dormancy mechanism provides a highly adaptable temperature-sensing mechanism to control the timing of germination. PMID:25114251

Osthole Induces Cell Cycle Arrest and Inhibits Migration and Invasion via PTEN/Akt Pathways in Osteosarcoma.

PubMed

Wang, Lu; Yang, Lei; Lu, Ying; Chen, Yingzhun; Liu, Tianhua; Peng, Yanli; Zhou, Yuhong; Cao, Yang; Bi, Zhenggang; Liu, Tianyi; Liu, Zhenhong; Shan, Hongli

2016-01-01

Osteosarcoma is the second highest cause of cancer-related death in children and adolescents. Majority of osteosarcoma patients (90%) show metastasis. Previous reports revealed that osthole showed antitumor activities via induction of apoptosis and inhibition of proliferation. However, the potential effects and detailed molecular mechanisms involved remained unclear. Cell viability was analyzed by MTT assay in osteosarcoma cell lines MG-63 and SAOS-2. Cell cycle was detected by flow cytometry. The effects of migration and invasion were evaluated by wound healing assay and transwell assays. Moreover, the level of proteins expression was determined by Western blot. The cell viability of MG63 and SAOS-2 were markedly inhibited by osthole in a dose- and time-dependent manner. Cell cycle was arrested and the ability of migration and invasion was obviously reduced when cells were exposed to osthole. Moreover, enzymes involved in PTEN/Akt pathway were regulated such as PTEN and p-Akt proteins. Furthermore, osthole inhibited the tumor growth in vivo. Our study unraveled, for the first time, the ability of osthole to suppress osteosarcoma and elucidated the regulation of PTEN/Akt pathway as a signaling mechanism for the anti-tumor action of osthole. These findings indicate that osthole may represent a novel therapeutic strategy in the treatment of osteosarcoma. © 2016 The Author(s) Published by S. Karger AG, Basel.

Light and the circadian clock mediate time-specific changes in sensitivity to UV-B stress under light/dark cycles

PubMed Central

Takeuchi, Tomomi; Newton, Linsey; Burkhardt, Alyssa; Mason, Saundra; Farré, Eva M.

2014-01-01

In Arabidopsis, the circadian clock regulates UV-B-mediated changes in gene expression. Here it is shown that circadian clock components are able to inhibit UV-B-induced gene expression in a gene-by-gene-specific manner and act downstream of the initial UV-B sensing by COP1 (CONSTITUTIVE PHOTOMORPHOGENIC 1) and UVR8 (UV RESISTANCE LOCUS 8). For example, the UV-B induction of ELIP1 (EARLY LIGHT INDUCIBLE PROTEIN 1) and PRR9 (PSEUDO-RESPONSE REGULATOR 9) is directly regulated by LUX (LUX ARRYTHMO), ELF4 (EARLY FLOWERING 4), and ELF3. Moreover, time-dependent changes in plant sensitivity to UV-B damage were observed. Wild-type Arabidopsis plants, but not circadian clock mutants, were more sensitive to UV-B treatment during the night periods than during the light periods under diel cycles. Experiments performed under short cycles of 6h light and 6h darkness showed that the increased stress sensitivity of plants to UV-B in the dark only occurred during the subjective night and not during the subjective day in wild-type seedlings. In contrast, the stress sensitivity of Arabidopsis mutants with a compromised circadian clock was still influenced by the light condition during the subjective day. Taken together, the results show that the clock and light modulate plant sensitivity to UV-B stress at different times of the day. PMID:25147271

Next-generation sequencing reveals low-dose effects of cationic dendrimers in primary human bronchial epithelial cells.

PubMed

Feliu, Neus; Kohonen, Pekka; Ji, Jie; Zhang, Yuning; Karlsson, Hanna L; Palmberg, Lena; Nyström, Andreas; Fadeel, Bengt

2015-01-27

Gene expression profiling has developed rapidly in recent years with the advent of deep sequencing technologies such as RNA sequencing (RNA Seq) and could be harnessed to predict and define mechanisms of toxicity of chemicals and nanomaterials. However, the full potential of these technologies in (nano)toxicology is yet to be realized. Here, we show that systems biology approaches can uncover mechanisms underlying cellular responses to nanomaterials. Using RNA Seq and computational approaches, we found that cationic poly(amidoamine) dendrimers (PAMAM-NH2) are capable of triggering down-regulation of cell-cycle-related genes in primary human bronchial epithelial cells at doses that do not elicit acute cytotoxicity, as demonstrated using conventional cell viability assays, while gene transcription was not affected by neutral PAMAM-OH dendrimers. The PAMAMs were internalized in an active manner by lung cells and localized mainly in lysosomes; amine-terminated dendrimers were internalized more efficiently when compared to the hydroxyl-terminated dendrimers. Upstream regulator analysis implicated NF-κB as a putative transcriptional regulator, and subsequent cell-based assays confirmed that PAMAM-NH2 caused NF-κB-dependent cell cycle arrest. However, PAMAM-NH2 did not affect cell cycle progression in the human A549 adenocarcinoma cell line. These results demonstrate the feasibility of applying systems biology approaches to predict cellular responses to nanomaterials and highlight the importance of using relevant (primary) cell models.

Esculetin Inhibits the Survival of Human Prostate Cancer Cells by Inducing Apoptosis and Arresting the Cell Cycle.

PubMed

Turkekul, Kader; Colpan, R Dilsu; Baykul, Talha; Ozdemir, Mehmet D; Erdogan, Suat

2018-03-01

Prostate cancer (PCa) is one of the most important causes of death in men and thus new therapeutic approaches are needed. In this study, antiproliferative and anti-migration properties of a coumarin derivative esculetin were evaluated. Human PCa cell lines PC3, DU145, and LNCaP were treated with various concentrations of esculetin for 24 to 72 hours, and cell viability was determined by the MTT test. Cell cycle and apoptosis were analyzed by using cell-based cytometer. Gene expression levels were assessed by reverse transcription and quantitative real-time PCR, cell migration was determined by the wound healing assay. The protein expression was measured by Western blotting. Esculetin inhibited cell proliferation in a dose- and time-dependent manner. Cell migration was inhibited by esculetin treatment. Administration of esculetin significantly reduced the cells survival, induced apoptosis and caused the G1 phase cell cycle arrest shown by image-based cytometer. The induced expression of cytochrome c , p53, p21 and p27, and down-regulated CDK2 and CDK4 may be the underlying molecular mechanisms of esculetin effect. Esculetin suppressed phosphorylation of Akt and enhanced protein expression of tumor-suppressor phosphatase and tensin homologue. Our findings showed that the coumarin derivative esculetin could be used in the management of PCa. However, further in vivo research is needed.

Esculetin Inhibits the Survival of Human Prostate Cancer Cells by Inducing Apoptosis and Arresting the Cell Cycle

PubMed Central

Turkekul, Kader; Colpan, R. Dilsu; Baykul, Talha; Ozdemir, Mehmet D.

2018-01-01

Background Prostate cancer (PCa) is one of the most important causes of death in men and thus new therapeutic approaches are needed. In this study, antiproliferative and anti-migration properties of a coumarin derivative esculetin were evaluated. Methods Human PCa cell lines PC3, DU145, and LNCaP were treated with various concentrations of esculetin for 24 to 72 hours, and cell viability was determined by the MTT test. Cell cycle and apoptosis were analyzed by using cell-based cytometer. Gene expression levels were assessed by reverse transcription and quantitative real-time PCR, cell migration was determined by the wound healing assay. The protein expression was measured by Western blotting. Results Esculetin inhibited cell proliferation in a dose- and time-dependent manner. Cell migration was inhibited by esculetin treatment. Administration of esculetin significantly reduced the cells survival, induced apoptosis and caused the G1 phase cell cycle arrest shown by image-based cytometer. The induced expression of cytochrome c, p53, p21 and p27, and down-regulated CDK2 and CDK4 may be the underlying molecular mechanisms of esculetin effect. Esculetin suppressed phosphorylation of Akt and enhanced protein expression of tumor-suppressor phosphatase and tensin homologue. Conclusions Our findings showed that the coumarin derivative esculetin could be used in the management of PCa. However, further in vivo research is needed. PMID:29629344

Identification by Random Mutagenesis of Functional Domains in KREPB5 That Differentially Affect RNA Editing between Life Cycle Stages of Trypanosoma brucei

PubMed Central

McDermott, Suzanne M.; Carnes, Jason

2015-01-01

KREPB5 is an essential component of ∼20S editosomes in Trypanosoma brucei which contains a degenerate, noncatalytic RNase III domain. To explore the function of this protein, we used a novel approach to make and screen numerous conditional null T. brucei bloodstream form cell lines that express randomly mutagenized KREPB5 alleles. We identified nine single amino acid substitutions that could not complement the conditional loss of wild-type KREPB5. Seven of these were within the RNase III domain, and two were in the C-terminal region that has no homology to known motifs. Exclusive expression of these mutated KREPB5 alleles in the absence of wild-type allele expression resulted in growth inhibition, the loss of ∼20S editosomes, and inhibition of RNA editing in BF cells. Eight of these mutations were lethal in bloodstream form parasites but not in procyclic-form parasites, showing that multiple domains function in a life cycle-dependent manner. Amino acid changes at a substantial number of positions, including up to 7 per allele, allowed complementation and thus did not block KREPB5 function. Hence, the degenerate RNase III domain and a newly identified domain are critical for KREPB5 function and have differential effects between the life cycle stages of T. brucei that differentially edit mRNAs. PMID:26370513

FOXC2 regulates the G2/M transition of stem cell-rich breast cancer cells and sensitizes them to PLK1 inhibition

PubMed Central

Pietilä, Mika; Vijay, Geraldine V.; Soundararajan, Rama; Yu, Xian; Symmans, William F.; Sphyris, Nathalie; Mani, Sendurai A.

2016-01-01

Cancer cells with stem cell properties (CSCs) underpin the chemotherapy resistance and high therapeutic failure of triple-negative breast cancers (TNBCs). Even though CSCs are known to proliferate more slowly, they are sensitive to inhibitors of G2/M kinases such as polo-like kinase 1 (PLK1). Understanding the cell cycle regulatory mechanisms of CSCs will help target these cells more efficiently. Herein, we identify a novel role for the transcription factor FOXC2, which is mostly expressed in CSCs, in the regulation of cell cycle of CSC-enriched breast cancer cells. We demonstrate that FOXC2 expression is regulated in a cell cycle-dependent manner, with FOXC2 protein levels accumulating in G2, and rapidly decreasing during mitosis. Knockdown of FOXC2 in CSC-enriched TNBC cells delays mitotic entry without significantly affecting the overall proliferation rate of these cells. Moreover, PLK1 activity is important for FOXC2 protein stability, since PLK1 inhibition reduces FOXC2 protein levels. Indeed, FOXC2 expressing CSC-enriched TNBC cells are sensitive to PLK1 inhibition. Collectively, our findings demonstrate a novel role for FOXC2 as a regulator of the G2/M transition and elucidate the reason for the observed sensitivity of CSC-enriched breast cancer cells to PLK1 inhibitor. PMID:27064522

Dynamics of Wolbachia pipientis Gene Expression Across the Drosophila melanogaster Life Cycle

PubMed Central

Gutzwiller, Florence; Carmo, Catarina R.; Miller, Danny E.; Rice, Danny W.; Newton, Irene L. G.; Hawley, R. Scott; Teixeira, Luis; Bergman, Casey M.

2015-01-01

Symbiotic interactions between microbes and their multicellular hosts have manifold biological consequences. To better understand how bacteria maintain symbiotic associations with animal hosts, we analyzed genome-wide gene expression for the endosymbiotic α-proteobacteria Wolbachia pipientis across the entire life cycle of Drosophila melanogaster. We found that the majority of Wolbachia genes are expressed stably across the D. melanogaster life cycle, but that 7.8% of Wolbachia genes exhibit robust stage- or sex-specific expression differences when studied in the whole-organism context. Differentially-expressed Wolbachia genes are typically up-regulated after Drosophila embryogenesis and include many bacterial membrane, secretion system, and ankyrin repeat-containing proteins. Sex-biased genes are often organized as small operons of uncharacterized genes and are mainly up-regulated in adult Drosophila males in an age-dependent manner. We also systematically investigated expression levels of previously-reported candidate genes thought to be involved in host-microbe interaction, including those in the WO-A and WO-B prophages and in the Octomom region, which has been implicated in regulating bacterial titer and pathogenicity. Our work provides comprehensive insight into the developmental dynamics of gene expression for a widespread endosymbiont in its natural host context, and shows that public gene expression data harbor rich resources to probe the functional basis of the Wolbachia-Drosophila symbiosis and annotate the transcriptional outputs of the Wolbachia genome. PMID:26497146

Juglans mandshurica Maxim extracts exhibit antitumor activity on HeLa cells in vitro.

PubMed

Xin, Nian; Hasan, Murtaza; Li, Wei; Li, Yan

2014-04-01

The present study examined the potential application of Juglans mandshurica Maxim extracts (HT) for cancer therapy by assessing their anti‑proliferative activity, reduction of telomerase activity, induction of apoptosis and cell cycle arrest in S phase in HeLa cells. From the perspective of using HT as a herbal medicine, photomicroscopy and florescent microscopy techniques were utilized to characterize the effect of the extracts on telomerase activity and cell morphology. Flow cytometry was employed to study apoptosis and cell cycle of HeLa cells, and DNA laddering was performed. The results showed that HT inhibited cell proliferation and telomerase activity, induced apoptosis and caused S phase arrest of HeLa cells in vitro. HT inhibited HeLa cell proliferation significantly, and the highest inhibition rate was 83.7%. A trap‑silver staining assay showed that HT was capable of markedly decreasing telomerase activity of HeLa cells and this inhibition was enhanced in a time‑ and dose‑dependent manner. Results of a Hoechst 33258 staining assay showed that HeLa cells treated by HT induced cell death. Through DNA agarose gel electrophoresis, DNA ladders of HeLa cells treated with HT were observed, indicating apoptosis. In conclusion, the present study demonstrated that HT exhibited anti‑tumor effects comprising the inhibition of growth and telomerase activity as well as apoptosis and cell cycle arrest in HeLa cells.

Phosphorylated Akt Protein at Ser473 Enables HeLa Cells to Tolerate Nutrient-Deprived Conditions

PubMed

Fathy, Moustafa; Awale, Suresh; Nikaido, Toshio

2017-12-29

Background: Despite angiogenesis, many tumours remain hypovascular and starved of nutrients while continuing to grow rapidly. The specific biochemical mechanisms associated with starvation resistance, austerity, may be new biological characters of cancer that are critical for cancer progression. Objective: This study aim was to investigate the effect of nutrient starvation on HeLa cells and the possible mechanism by which the cells are able to tolerate nutrient-deprived conditions. Methods: Nutrient starvation was achieved by culturing HeLa cells in nutrient-deprived medium (NDM) and cell survival was estimated by using cell counting kit-8. The effect of starvation on cell cycle distribution and the quantitative analysis of apoptotic cells were investigated by flow cytometry using propidium iodide staining. Western blotting was used to detect the expression levels of Akt and phosphorylated Akt at Ser473 (Ser473p-Akt) proteins. Results: HeLa cells displayed extremely long survival when cultured in NDM. The percentage of apoptotic HeLa cells was significantly increased by starvation in a time-dependent manner. A significant increase in the expression of Ser473p-Akt protein after starvation was also observed. Furthermore, it was found that Akt inhibitor III molecule inhibited the cells proliferation in a concentration- and time-dependent manner. Conclusion: Results of the present study provide evidence that Akt activation may be implicated in the tolerance of HeLa cells for nutrient starvation and may help to suggest new therapeutic strategies designed to prevent austerity of cervical cancer cells through inhibition of Akt activation. Creative Commons Attribution License

Sodium fluoride induces apoptosis in mouse embryonic stem cells through ROS-dependent and caspase- and JNK-mediated pathways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nguyen Ngoc, Tam Dan; Son, Young-Ok; Lim, Shin-Saeng

2012-03-15

Sodium fluoride (NaF) is used as a source of fluoride ions in diverse applications. Fluoride salt is an effective prophylactic for dental caries and is an essential element required for bone health. However, fluoride is known to cause cytotoxicity in a concentration-dependent manner. Further, no information is available on the effects of NaF on mouse embryonic stem cells (mESCs). We investigated the mode of cell death induced by NaF and the mechanisms involved. NaF treatment greater than 1 mM reduced viability and DNA synthesis in mESCs and induced cell cycle arrest in the G{sub 2}/M phase. The addition of NaFmore » induced cell death mainly by apoptosis rather than necrosis. Catalase (CAT) treatment significantly inhibited the NaF-mediated cell death and also suppressed the NaF-mediated increase in phospho-c-Jun N-terminal kinase (p-JNK) levels. Pre-treatment with SP600125 or z-VAD-fmk significantly attenuated the NaF-mediated reduction in cell viability. In contrast, intracellular free calcium chelator, but not of sodium or calcium ion channel blockers, facilitated NaF-induced toxicity in the cells. A JNK specific inhibitor (SP600125) prevented the NaF-induced increase in growth arrest and the DNA damage-inducible protein 45α. Further, NaF-mediated loss of mitochondrial membrane potential was apparently inhibited by pifithrin-α or CAT inhibitor. These findings suggest that NaF affects viability of mESCs in a concentration-dependent manner, where more than 1 mM NaF causes apoptosis through hydroxyl radical-dependent and caspase- and JNK-mediated pathways. -- Highlights: ► The mode of NaF-induced cell death and the mechanisms involved were examined. ► NaF induced mainly apoptotic death of mouse embryonic stem cells (mESCs). ► NaF induced mitochondrial-mediated and caspase-dependent apoptosis. ► JNK- and p53-mediated pathways are involved in NaF-mediated apoptosis in the cells. ► ROS are the up-stream effector in NaF-mediated activation of JNK and p53 in mESCs.« less

17-DMAG induces heat shock protein 90 functional impairment in human bladder cancer cells: knocking down the hallmark traits of malignancy.

PubMed

Karkoulis, Panagiotis K; Stravopodis, Dimitrios J; Voutsinas, Gerassimos E

2016-05-01

Heat shock protein 90 (Hsp90) is a molecular chaperone that maintains the structural and functional integrity of various protein clients involved in multiple oncogenic signaling pathways. Hsp90 holds a prominent role in tumorigenesis, as numerous members of its broad clientele are involved in the generation of the hallmark traits of cancer. 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) specifically targets Hsp90 and interferes with its function as a molecular chaperone, impairing its intrinsic ATPase activity and undermining proper folding of multiple protein clients. In this study, we have examined the effects of 17-DMAG on the regulation of Hsp90-dependent tumorigenic signaling pathways directly implicated in cell cycle progression, survival, and motility of human urinary bladder cancer cell lines. We have used MTT-based assays, FACS analysis, Western blotting, semiquantitative PCR (sqPCR), immunofluorescence, and scratch-wound assays in RT4 (p53(wt)), RT112 (p53(wt)), T24 (p53(mt)), and TCCSUP (p53(mt)) human urinary bladder cancer cell lines. We have demonstrated that, upon exposure to 17-DMAG, bladder cancer cells display prominent cell cycle arrest and commitment to apoptotic and autophagic cell death, in a dose-dependent manner. Furthermore, 17-DMAG administration induced pronounced downregulation of multiple Hsp90 protein clients and other downstream oncogenic effectors, therefore causing inhibition of cell proliferation and decline of cell motility due to the molecular "freezing" of critical cytoskeletal components. In toto, we have clearly demonstrated the dose-dependent and cell type-specific effects of 17-DMAG on the hallmark traits of cancer, appointing Hsp90 as a key molecular component in bladder cancer targeted therapy.

Phosphorylation of Nucleotide Excision Repair Factor Xeroderma Pigmentosum Group A by Ataxia Telangiectasia Mutated and Rad3-Related-Dependent Checkpoint Pathway Promotes Cell Survival in Response to UV Irradiation

PubMed Central

Wu, Xiaoming; Shell, Steven M.; Yang, Zhengguan; Zou, Yue

2006-01-01

DNA damage triggers complex cellular responses in eukaryotic cells, including initiation of DNA repair and activation of cell cycle checkpoints. In addition to inducing cell cycle arrest, checkpoint also has been suggested to modulate a variety of other cellular processes in response to DNA damage. In this study, we present evidence showing that the cellular function of xeroderma pigmentosum group A (XPA), a major nucleotide excision repair (NER) factor, could be modulated by checkpoint kinase ataxia-telangiectasia mutated and Rad3-related (ATR) in response to UV irradiation. We observed the apparent interaction and colocalization of XPA with ATR in response to UV irradiation. We showed that XPA was a substrate for in vitro phosphorylation by phosphatidylinositol-3-kinase-related kinase family kinases whereas in cells XPA was phosphorylated in an ATR-dependent manner and stimulated by UV irradiation. The Ser196 of XPA was identified as a biologically significant residue to be phosphorylated in vivo. The XPA-deficient cells complemented with XPA-S196A mutant, in which Ser196 was substituted with an alanine, displayed significantly higher UV sensitivity compared with the XPA cells complemented with wild-type XPA. Moreover, substitution of Ser196 with aspartic acid for mimicking the phosphorylation of XPA increased the cell survival to UV irradiation. Taken together, our results revealed a potential physical and functional link between NER and the ATR-dependent checkpoint pathway in human cells and suggested that the ATR checkpoint pathway could modulate the cellular activity of NER through phosphorylation of XPA at Ser196 on UV irradiation. PMID:16540648

Effects of Ionizing Radiation on Human Adipose Derived Mesenchymal Stem Cells and their Differentiation towards the Osteoblastic Lineage

NASA Astrophysics Data System (ADS)

Konda, Bikash; Baumstark-Khan, Christa; Hellweg, Christine; Reitz, Guenther; Lau, Patrick

Radiation exposure and musculoskeletal disuse are among the major challenges during space missions. Astronauts face the problem to lose bone calcium due to uncoupling of bone formation and resorption. Bone forming osteoblasts can be derived from the undifferentiated mesenchymal stem cell compartment (MSC). In this study, the ability of human adipose tissue derived stem cells (ATSC) to differentiate into the osteoblastic lineage was examined after radiation exposure in presence of medium supplementation with osteogenic additives (ß-glycerophosphate, ascorbic acid and dexamethasone). The SAOS-2 cell line (human osteosarcoma cell line) was used as control for osteoblastic differentiation. Changes in cellular morphology, cell cycle progression, as well as cellular radiation sensitivity were characterized after ionizing radiation exposure with X-rays and heavy ions (Ti). Rapidly proliferating SAOS-2 cells are less radiation-sensitive than slowly proliferating ATSC cells after X-ray (CFA: dose effect curves show D0 values of 1 Gy and 0.75 Gy for SAOS-2 and ATSC, respectively) exposure. Heavy ion (Ti) exposure resulted in a greater extent of cells accumulating in the G2/M phase of the cell cycle in a dose-dependent manner when compared to X-ray exposure. Differentiation of cells towards the osteoblastic lineage was quantified by hydroxyapatite (HA) deposition using Lonza OsteoImageTM mineralization assay. The deposition of HA after X- and Ti-irradiation for highly proliferating SAOS-2 cells showed a dose-dependent time delay while slowly proliferating ATSC showed no effect from radiation exposure. More detailed investigation is required to reveal the radiation dependent mechanism of bone loss in astronauts.

Galangin Induces Apoptosis in MCF-7 Human Breast Cancer Cells Through Mitochondrial Pathway and Phosphatidylinositol 3-Kinase/Akt Inhibition.

PubMed

Liu, Dan; You, Pengtao; Luo, Yan; Yang, Min; Liu, Yanwen

2018-06-07

The study aimed to investigate the molecular mechanism of inhibition of proliferation and apoptosis induction by galangin against MCF-7 human breast cancer cells. Cell Counting Kit-8 assay was used to assess cell viability and flow cytometry was used to detect cell apoptosis. The expression level of apoptosis-related proteins (cleaved-caspase-9, cleaved-caspase-8, cleaved-caspase-3, Bad, cleaved-Bid, Bcl-2, Bax, p-phosphatidylinositol 3-kinase [PI3K], and p-Akt) and cell cycle-related proteins (cyclin D3, cyclin B1, cyclin-dependent kinases CDK1, CDK2, CDK4, p21, p27, p53) were evaluated by Western blotting. Galangin increased the expression of Bax and decreased the expression of Bcl-2 in a concentration-dependent manner, inhibited cell viability, and induced apoptosis. Meanwhile, the expression of cleavage of caspase-9, caspase-8, caspase-3, Bid, and Bad increased significantly while the expression of p-PI3K and p-Akt proteins decreased. In addition, the protein levels of cyclin D3, cyclin B1, CDK1, CDK2, and CDK4 were downregulated while the expression levels of p21, p27, and p53 were upregulated significantly. Galangin could suppress the viability of MCF-7 cells and induce cell apoptosis via the mitochondrial pathway and PI3K/Akt inhibition as well as cell cycle arrest. © 2018 S. Karger AG, Basel.

Sporoderm-Broken Spores of Ganoderma lucidum Inhibit the Growth of Lung Cancer: Involvement of the Akt/mTOR Signaling Pathway.

PubMed

Chen, Yali; Lv, Jing; Li, Kun; Xu, Jing; Li, Mingyan; Zhang, Wen; Pang, Xiufeng

2016-10-01

The sporoderm-broken spores of Ganoderma lucidum (SBGS) and their extracts exhibited a wide range of biological activities. In the present study, we prepare ethanol/ethanol extract (E/E-SBGS) and ethanol/aqueous extract (E/A-SBGS) from SBGS and examine their antitumor activities against human lung cancer. Our results showed that E/E-SBGS, not E/A-SBGS, inhibited the survival and migration of lung cancer cells in a dose-dependent manner. E/E-SBGS arrested cell cycle at G2/M phase and triggered apoptosis by decreasing the expression and activity of cell cycle regulators, cyclin B1 and cdc2, as well as anti-apoptotic proteins, Bcl-2 and Bcl-xl. Consequently, colony formation of lung cancer cells was markedly blocked by E/E-SBGS at subtoxic concentrations. Oral administration of both E/E-SBGS and SBGS significantly suppressed tumor volume and tumor weight without gross toxicity in mice. Mechanism study showed that E/E-SBGS dose-dependently suppressed the activation of Akt, the mammalian target of rapamycin (mTOR) and their downstream molecules S6 kinase and 4E-BP1 in treated tumor cells. Taken together, these results indicate that the ethanol extract of sporoderm-broken spores of G. lucidum suppresses the growth of human lung cancer, at least in part, through inhibition of the Akt/mTOR signaling pathway, suggesting its potential role in cancer treatments.

Retinoblastoma function is essential for establishing lung epithelial quiescence after injury.

PubMed

Mason-Richie, Nicole A; Mistry, Meenakshi J; Gettler, Caitlin A; Elayyadi, Asmaa; Wikenheiser-Brokamp, Kathryn A

2008-06-01

The retinoblastoma gene product (RB) regulates cell cycle, quiescence, and survival in a cell type-dependent and environment-dependent manner. RB function is critical in the pulmonary epithelium, as evidenced by nearly universal RB inactivation in lung cancer and increased lung cancer risk in persons with germline RB gene mutations. Lung carcinomas occur in the context of epithelial remodeling induced by cytotoxic damage. Whereas the role of RB in development and normal organ homeostasis has been extensively studied, RB function in the context of cellular injury and repair has remained largely unexplored. In the current studies, the RB gene was selectively deleted in the respiratory epithelium of the mouse. Although RB was not required for establishing or maintaining quiescence during lung homeostasis, RB was essential for establishing quiescence during epithelial repair after injury. Notably, aberrant cell cycle progression was sustained for 9 months after injury in RB-deficient lungs. Prenatal and postnatal RB ablation had similar effects, providing evidence that timing of RB loss was not critical to the outcome and that the injury-induced phenotype was not secondary to compensatory alterations occurring during development. These data show that RB is essential for repair of the respiratory epithelium after cytotoxic damage and support a critical unique role for RB in the context of epithelial remodeling after injury. Because human cancers are associated with chronic cellular damage, these findings have important new implications for RB-mediated tumor suppression.

Sildenafil (Viagra) prolongs cardiac repolarization by blocking the rapid component of the delayed rectifier potassium current.

PubMed

Geelen, P; Drolet, B; Rail, J; Bérubé, J; Daleau, P; Rousseau, G; Cardinal, R; O'Hara, G E; Turgeon, J

2000-07-18

BACKGROUND-Several cases of unexpected death have been reported with sildenafil in patients predisposed to ischemic cardiac events. Although acute episodes of ischemia could account for some of these deaths, we hypothesized that sildenafil may have unsuspected electrophysiological effects predisposing some patients to proarrhythmia. METHODS AND RESULTS-Studies were undertaken in 10 isolated guinea pig hearts that demonstrated prolongation of cardiac repolarization in a reverse use-dependent manner by sildenafil 30 mcmol/L. Action potential duration increased 15% from baseline 117+/-3 to 134+/-2 ms with sildenafil during pacing at 250 ms cycle length, whereas a 6% increase from 99+/-2 to 105+/-2 ms was seen with pacing at 150 ms cycle length. Experiments in human ether-a-go-go-related gene (HERG)-transfected HEK293 cells (n=30) demonstrated concentration-dependent block of the rapid component (I(Kr)) of the delayed rectifier potassium current: activating current was 50% decreased at 100 mcmol/L. This effect was confirmed using HERG-transfected Chinese hamster ovary (CHO) cells, which exhibit no endogenous I(K)-like current. CONCLUSIONS-Sildenafil possesses direct cardiac electrophysiological effects similar to class III antiarrhythmic drugs. These effects are observed at concentrations that may be found in conditions of impaired drug elimination such as renal or hepatic insufficiency, during coadministration of another CYP3A substrate/inhibitor, or after drug overdose and offer a new potential explanation for sudden death during sildenafil treatment.

Chloroquine inhibits hepatocellular carcinoma cell growth in vitro and in vivo

PubMed Central

HU, TAO; LI, PEI; LUO, ZHONGGUANG; CHEN, XIAOYU; ZHANG, JINGYANG; WANG, CHUNYAO; CHEN, PING; DONG, ZIMING

2016-01-01

Recently, chloroquine (CQ) has been widely used to improve the efficacy of different chemotherapy drugs to treat tumors. However, the effects of single treatment of CQ on liver cancer have not been investigated. In the present study, we examined the effects of CQ on the growth and viability of liver cancer cells in vitro and in vivo, and revealed that CQ treatment triggered G0/G1 cell cycle arrest, induced DNA damage and apoptosis in a dose- and time-dependent manner in liver cancer cells. Moreover, administration of CQ to tumor-bearing mice suppressed the tumor growth in an orthotopic xenograft model of liver cancer. These findings extend our understanding and suggest that CQ could be repositioned as a treatment option for liver cancer as a single treatment or in combination. PMID:26530158

Inhibition of angiogenic attributes by decursin in endothelial cells and ex vivo rat aortic ring angiogenesis model.

PubMed

Bhat, Tariq A; Moon, Jung S; Lee, Sookyeon; Yim, Dongsool; Singh, Rana P

2011-11-01

The present study was undertaken to observe the inhibition of angiogenesis by decursin. It was the first time to show that decursin offered strong anti-angiogenic activities under the biologically relevant growth (with serum) conditions. Decursin significantly inhibited human umbilical vein endothelial cell (HUVEC) proliferation concomitant with G1 phase cell cycle arrest. Decursin also inhibited HUVEC-capillary tube formation and invasion/migration in a dose-dependant manner which was associated with the suppression of matrix metalloproteinase (MMP) -2 and -9 activities. Decursin suppressed angiogenesis in ex vivo rat aortic ring angiogenesis model where it significantly inhibited blood capillary-network sprouting from rat aortic sections. Taken together, these findings suggested anti-angiogenic activity of decursin in biologically relevant condition, and warrants further pre-clinical studies for its potential clinical usefulness.

Antichlamydial Dimeric Indole Derivatives from Marine Actinomycete Rubrobacter radiotolerans.

PubMed

Li, Jian Lin; Chen, Dandan; Huang, Lei; Ni, Min; Zhao, Yu; Fan, Huizhou; Bao, Xiaofeng

2017-06-01

Chlamydiae are widely distributed pathogens of human populations, which can lead to serious reproductive and other health problems. In our search for novel antichlamydial metabolites from marine derived-microorganisms, one new ( 1 ) and two known ( 2, 3 ) dimeric indole derivatives were isolated from the sponge-derived actinomycete Rubrobacter radiotolerans . The chemical structures of these metabolites were elucidated by NMR spectroscopic data as well as CD calculations. All three metabolites suppressed chlamydial growth in a concentration-dependent manner. Among them, compound 1 exhibited the most effective antichlamydial activity with IC 50 values of 46.6 ~ 96.4 µM in the production of infectious progeny. Compounds appeared to target the mid-stage of the chlamydial developmental cycle by interfering with reticular body replication, but not directly inactivating the infectious elementary body. Georg Thieme Verlag KG Stuttgart · New York.

Enhancement of DMNQ-induced hepatocyte toxicity by cytochrome P450 inhibition.

PubMed

Ishihara, Yasuhiro; Shiba, Dai; Shimamoto, Norio

2006-07-15

Two mechanisms have been proposed to explain quinone cytotoxicity: oxidative stress via the redox cycle and the arylation of intracellular nucleophiles. As the redox cycle is catalyzed by NADPH cytochrome P450 reductase, cytochrome P450 systems are expected to be related to the cytotoxicity induced by redox-cycling quinones. Thus, we investigated the relationship between cytochrome P450 systems and quinone toxicity for rat primary hepatocytes using an arylator, 1,4-benzoquinone (BQ), and a redox cycler, 2,3-dimethoxy-1,4-naphthoquinone (DMNQ). The hepatocyte toxicity of both BQ and DMNQ increased in a time- and dose-dependent manner. Pretreatment with cytochrome P450 inhibitors, such as SKF-525A (SKF), ketoconazole and 2-methy-1,2-di-3-pyridyl-1-propanone, enhanced the hepatocyte toxicity induced by DMNQ but did not affect BQ-induced hepatocyte toxicity. The production of superoxide anion and the levels of glutathione disulfide and thiobarbituric-acid-reactive substances were increased by treatment with DMNQ, and SKF pretreatment further enhanced their increases. In addition, NADPH oxidation in microsomes was increased by treatment with DMNQ and further augmented by pretreatment with SKF, and a NADPH cytochrome P450 reductase inhibitor, diphenyleneiodonium chloride completely suppressed NADPH oxidations increased by treatment with either DMNQ- or DMNQ + SKF. Pretreatment with antioxidants, such as alpha-tocopherol, reduced glutathione, N-acetyl cysteine or an iron ion chelator deferoxamine, totally suppressed DMNQ- and DMNQ + SKF-induced hepatocyte toxicity. These results indicate that the hepatocyte toxicity of redox-cycling quinones is enhanced under cytochrome P450 inhibition, and that this enhancement is caused by the potentiation of oxidative stress.

The influence of high glucose on the Cip/Kip family expression profiles in HRECs.

PubMed

Tian, Jingyi; Ma, Hongjie; Luo, Yan; Hu, Andina; Lin, Shaofen; Li, Tao; Guo, Kai; Li, Jing; Cai, Meng; Tang, Shibo

2013-12-01

Neovascularization is the main characteristic of the proliferative stage of diabetic retinopathy. It has been proven that cell cycle regulation is involved in angiogenesis. The cell cycle regulators, Cip/Kip protein family, belong to the cyclin-dependent kinase inhibitors, are versatile proteins, and except for their function in cell cycle regulation, they also participate in transcription, apoptosis and migration. The expression profiles of the Cip/Kip family in human retina microvascular endothelial cells (HRECs) under normal or high glucose conditions has not been described before. This study was undertaken to determine the expression profiles of the Cip/Kip family proteins, e.g., proteins which are influenced by high glucose and in what manner. Western blot and immunofluorescence analyses were used to investigate the protein expression profiles. Only p21(cip1) and p27(kip1) were detected in HRECs, and they were located in the nucleus. P21(cip1) protein abundance was higher than p27(kip1) in HRECs. Incubation of HRECs in medium containing 30 mM D-glucose for 48 h resulted in downregulation of p21(cip1) protein expression, but had no influence on p27(kip1) protein levels or p21(cip1) mRNA abundance. These results were accompanied by cell cycle G1 phase exit and a lower cell survival rate. Our data show for the first time that high glucose changes the Cip/Kip family expression profiles in HRECs, which may be the foundation for the investigation of the role of the Cip/Kip family in the pathogenesis of diabetic retinopathy.

Protective role of Delphinium denudatum (Jadwar) against morphine induced tolerance and dependence in mice.

PubMed

Zafar, S; Ahmad, M A; Siddiqui, T A

2001-11-01

Chronic treatment with Delphinium denudatum (Dd) (Jadwar) (family: Ranunculaceae, 200-1600 mg/kg) suppressed morphine withdrawal jumps in a dose-dependent manner, a sign of the development of dependence to opiate as assessed by naloxone (2 mg/kg) precipitation withdrawal on day 10 of testing in mice. Repeated administration of Dd (200-1600 mg/kg) for 9 days attenuated the development of tolerance to the analgesic effect of morphine (10 mg/kg), also produces significant change in tail-flick latency from the saline pretreated group in a dose-dependent manner.

Aeginetia indica Decoction Inhibits Hepatitis C Virus Life Cycle

PubMed Central

Lin, Cheng-Wei; Lo, Chieh-Wen; Tsai, Chia-Ni; Pan, Ting-Chun; Chen, Pin-Yin

2018-01-01

Chronic hepatitis C virus (HCV) infection is still a global epidemic despite the introduction of several highly effective direct-acting antivirals that are tagged with sky-high prices. The present study aimed to identify an herbal decoction that ameliorates HCV infection. Among six herbal decoctions tested, the Aeginetia indica decoction had the most profound effect on the HCV reporter activity in infected Huh7.5.1 liver cells in a dose- and time-dependent manner. The Aeginetia indica decoction exerted multiple inhibitory effects on the HCV life cycle. Pretreatment of the cells with the Aeginetia indica decoction prior to HCV infection reduced the HCV RNA and non-structural protein 3 (NS3) protein levels in the infected cells. The Aeginetia indica decoction reduced HCV internal ribosome entry site-mediated protein translation activity. It also reduced the HCV RNA level in the infected cells in association with reduced NS5A phosphorylation at serine 235, a predominant phosphorylation event indispensable to HCV replication. Thus, the Aeginetia indica decoction inhibits HCV infection, translation, and replication. Mechanistically, the Aeginetia indica decoction probably reduced HCV replication via reducing NS5A phosphorylation at serine 235. PMID:29315273

Cytotoxic effect of the serotonergic drug 1-(1-Naphthyl)piperazine against melanoma cells.

PubMed

Menezes, Ana Catarina; Carvalheiro, Manuela; Ferreira de Oliveira, José Miguel P; Ascenso, Andreia; Oliveira, Helena

2018-03-01

1-(1-Naphthyl)piperazine (1-NPZ) is a serotonergic derivative of quipazine acting both as antagonist and agonist of different serotonin receptors, with promising results for the management of skin cancer. In this work, we studied the effect of 1-NPZ on human MNT-1 melanoma cells by evaluating its effects on cell viability, ability to form colonies, cell cycle dynamics, reactive oxygen species (ROS) production and apoptosis. Treatment of MNT-1 cells with 1-NPZ for 24h decreased cell viability and induced apoptosis in a dose-dependent manner. Activity against melanoma was confirmed with a different melanoma cell line, SK-MEL-28. Simultaneously, 1-NPZ affected cell cycle progression by mediating a S-phase delay. Higher levels of ROS were also detected in MNT-1 cells after treatment with 1-NPZ. Furthermore, 1-NPZ significantly increased the expression of cyclooxygenase-2 in MNT-1 cells. These findings suggest that 1-NPZ pretreatment is able to induce oxidative stress, and consequently apoptotic cell death in melanoma cells. In conclusion, this study demonstrates the cytotoxic and genotoxic potential of 1-NPZ against melanoma cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

EWSR1 regulates mitosis by dynamically influencing microtubule acetylation.

PubMed

Wang, Yi-Long; Chen, Hui; Zhan, Yi-Qun; Yin, Rong-Hua; Li, Chang-Yan; Ge, Chang-Hui; Yu, Miao; Yang, Xiao-Ming

2016-08-17

EWSR1, participating in transcription and splicing, has been identified as a translocation partner for various transcription factors, resulting in translocation, which in turn plays crucial roles in tumorigenesis. Recent studies have investigated the role of EWSR1 in mitosis. However, the effect of EWSR1 on mitosis is poorly understood. Here, we observed that depletion of EWSR1 resulted in cell cycle arrest in the mitotic phase, mainly due to an increase in the time from nuclear envelope breakdown to metaphase, resulting in a high percentage of unaligned chromosomes and multipolar spindles. We also demonstrated that EWSR1 is a spindle-associated protein that interacts with α-tubulin during mitosis. EWSR1 depletion increased the cold-sensitivity of spindle microtubules, and decreased the rate of spindle assembly. EWSR1 regulated the level of microtubule acetylation in the mitotic spindle; microtubule acetylation was rescued in EWSR1-depleted mitotic cells following suppression of HDAC6 activity by its specific inhibitor or siRNA treatment. In summary, these results suggest that EWSR1 regulates the acetylation of microtubules in a cell cycle-dependent manner through its dynamic location on spindle MTs, and may be a novel regulator for mitosis progress independent of its translocation.

On the sources of drift in a turbine-based spirometer.

PubMed

Ha, Jonathan K; Perlow, Daniel B; Yi, Byong Yong; Yu, Cedric X

2008-08-21

A systematic study on the sources of drift in a turbine-based spirometer (VMM-400) is presented. The study utilized an air-tight cylinder to pump air through the spirometer in a precise and programmable manner. Factors contributing to the drift were isolated and quantified. The drift due to imbalance in the electronics and the mechanical blade increased from 1% per breathing cycle to as much as 10% when the flow rate decreased from 0.24 to 0.08 l s(-1). A temperature difference of 16 degrees between the ambient and the air in the cylinder contributed about 3.5%. Most significantly, a difference in the breathing between inhalation and exhalation could produce a drift of 40% per breathing cycle, or even higher, depending on the extent of the breathing asymmetry. The origin of this drift was found to be rooted in the differential response of the spirometer to the different flow rate. Some ideas and suggestions for a correction strategy are provided for future work. The present work provides an important first step for eventual utilization of a spirometer as a stand-alone breathing surrogate for gating or tracking radiation therapy.

Non-gassing nickel-cadmium battery electrodes and cells

NASA Technical Reports Server (NTRS)

Luksha, E.; Gordy, D. J.

1972-01-01

The concept of a negative limited nongassing nickel-cadmium battery was demonstrated by constructing and testing practical size experimental cells of approximately 25 Ah capacity. These batteries operated in a gas-free manner and had measured energy densities of 10-11 Wh/lb. Thirty cells were constructed for extensive testing. Some small cells were tested for over 200 cycles at 100% depth. For example, a small cell with an electrodeposited cadmium active mass on a silver screen still had 55% of its theoretical capacity (initial efficiency was 85%). There was no evidence of deterioration of gassing properties with cycling of the nickel electrodes. The charge temperature was observed to be the most critical variable governing nickel electrode gassing. This variable was shown to be age dependent. Four types of cadmium electrodes were tested: an electrodeposited cadmium active mass on a cadmium or silver substrate, a porous sintered silver substrate based electrode, and a Teflon bonded pressed cadmium electrode. The electrodeposited cadmium mass on a silver screen was found to be the best all-around electrode from a performance point of view and from the point of view of manufacturing them in a size required for a 25 Ah size battery.

Dynamic DNA methylation reconfiguration during seed development and germination.

PubMed

Kawakatsu, Taiji; Nery, Joseph R; Castanon, Rosa; Ecker, Joseph R

2017-09-15

Unlike animals, plants can pause their life cycle as dormant seeds. In both plants and animals, DNA methylation is involved in the regulation of gene expression and genome integrity. In animals, reprogramming erases and re-establishes DNA methylation during development. However, knowledge of reprogramming or reconfiguration in plants has been limited to pollen and the central cell. To better understand epigenetic reconfiguration in the embryo, which forms the plant body, we compared time-series methylomes of dry and germinating seeds to publicly available seed development methylomes. Time-series whole genome bisulfite sequencing reveals extensive gain of CHH methylation during seed development and drastic loss of CHH methylation during germination. These dynamic changes in methylation mainly occur within transposable elements. Active DNA methylation during seed development depends on both RNA-directed DNA methylation and heterochromatin formation pathways, whereas global demethylation during germination occurs in a passive manner. However, an active DNA demethylation pathway is initiated during late seed development. This study provides new insights into dynamic DNA methylation reprogramming events during seed development and germination and suggests possible mechanisms of regulation. The observed sequential methylation/demethylation cycle suggests an important role of DNA methylation in seed dormancy.

Cytotoxic effect of artocarpin on T47D cells.

PubMed

Arung, Enos Tangke; Wicaksono, Britanto Dani; Handoko, Yohana Ayupriyanti; Kusuma, Irawan Wijaya; Shimizu, Kuniyoshi; Yulia, Dina; Sandra, Ferry

2010-10-01

In our screening projects for anticancer agents from natural resources, artocarpin [6-(3-methyl-1-butenyl)-5,2',4'-trihydroxy-3-isoprenyl-7-methoxyflavone] isolated from wood of jack fruit (Artocarpus heterophyllus) showed potent cytotoxic activity on human T47D breast cancer cells. The mode of action of artocarpin was evaluated by its effect on cell viability, nuclear morphology, cell cycle progression, expression of protein markers for apoptosis, and mitochondrial membrane potential (Delta psi m). These results showed that artocarpin caused a reduction of cell viability in a concentration-dependent manner and an alteration of cell and nuclear morphology. Moreover, the percentage of the sub-G1 phase formation was elevated dose-dependently. Artocarpin induced activation of caspase 8 and 10 as indicated by stronger signal intensity of cleaved-caspase 8 and weaker signal intensity of caspase 10 markers detected after artocarpin treatment. In addition, we also noticed the activation of caspase 3 by artocarpin. There were negligible changes in mitochondrial membrane potential (Delta psi m) due to artocarpin treatment. All together, these data indicated that artocarpin induced apoptosis in T47D cells possibly via an extrinsic pathway.

Glutamine promotes ovarian cancer cell proliferation through the mTOR/S6 pathway

PubMed Central

Yuan, Lingqin; Sheng, Xiugui; Willson, Adam K; Roque, Dario R; Stine, Jessica E; Guo, Hui; Jones, Hannah M; Zhou, Chunxiao; Bae-Jump, Victoria L

2015-01-01

Glutamine is one of the main nutrients used by tumor cells for biosynthesis. Therefore, targeted inhibition of glutamine metabolism may have anti-tumorigenic implications. In the present study, we aimed to evaluate the effects of glutamine on ovarian cancer cell growth. Three ovarian cancer cell lines, HEY, SKOV3, and IGROV-1, were assayed for glutamine dependence by analyzing cytotoxicity, cell cycle progression, apoptosis, cell stress, and glucose/glutamine metabolism. Our results revealed that administration of glutamine increased cell proliferation in all three ovarian cancer cell lines in a dose dependent manner. Depletion of glutamine induced reactive oxygen species and expression of endoplasmic reticulum stress proteins. In addition, glutamine increased the activity of glutaminase (GLS) and glutamate dehydrogenase (GDH) by modulating the mTOR/S6 and MAPK pathways. Inhibition of mTOR activity by rapamycin or blocking S6 expression by siRNA inhibited GDH and GLS activity, leading to a decrease in glutamine-induced cell proliferation. These studies suggest that targeting glutamine metabolism may be a promising therapeutic strategy in the treatment of ovarian cancer. PMID:26045471

Cisplatin radiosensitizes radioresistant human mesenchymal stem cells.

PubMed

Rühle, Alexander; Perez, Ramon Lopez; Glowa, Christin; Weber, Klaus-Josef; Ho, Anthony D; Debus, Jürgen; Saffrich, Rainer; Huber, Peter E; Nicolay, Nils H

2017-10-20

Cisplatin-based chemo-radiotherapy is widely used to treat cancers with often severe therapy-associated late toxicities. While mesenchymal stem cells (MSCs) were shown to aid regeneration of cisplatin- or radiation-induced tissue lesions, the effect of the combined treatment on the stem cells remains unknown. Here we demonstrate that cisplatin treatment radiosensitized human bone marrow-derived MSCs in a dose-dependent manner and increased levels of radiation-induced apoptosis. However, the defining stem cell properties of MSCs remained largely intact after cisplatin-based chemo-radiation, and stem cell motility, adhesion, surface marker expression and the characteristic differentiation potential were not significantly influenced. The increased cisplatin-mediated radiosensitivity was associated with a cell cycle shift of MSCs towards the radiosensitive G2/M phase and increased residual DNA double-strand breaks. These data demonstrate for the first time a dose-dependent radiosensitization effect of MSCs by cisplatin. Clinically, the observed increase in radiation sensitivity and subsequent loss of regenerative MSCs may contribute to the often severe late toxicities observed after cisplatin-based chemo-radiotherapy in cancer patients.

Glabridin induces apoptosis and cell cycle arrest in oral cancer cells through the JNK1/2 signaling pathway.

PubMed

Chen, Chang-Tai; Chen, Yi-Tzu; Hsieh, Yi-Hsien; Weng, Chia-Jui; Yeh, Jung-Chun; Yang, Shun-Fa; Lin, Chiao-Wen; Yang, Jia-Sin

2018-06-01

Glabridin, a flavonoid extracted from licorice (Glycyrrhiza glabra), possesses various biological properties, including anticancer activities. However, the effect of glabridin on oral cancer cell apoptosis and the underlying molecular mechanisms has not been elucidated. In this study, we demonstrated that glabridin treatment significantly inhibits cell proliferation in human oral cancer SCC-9 and SAS cell lines. Flow cytometric assays demonstrated that glabridin induced several features of apoptosis, such as sub-G1 phase cell increase and phosphatidylserine externalization. Furthermore, glabridin induced apoptosis dose-dependently in SCC-9 cells through caspase-3, -8, and -9 activation and poly (ADP-ribose) polymerase cleavage. Moreover, glabridin increased the phosphorylation of the extracellular signal-regulated kinase, p38, and c-Jun N-terminal kinase (JNK) pathways in a dose-dependent manner. Moreover, the inhibition of the JNK1/2 inhibitor significantly reversed the glabridin-induced activation of the caspase pathway. In conclusion, our findings suggest that glabridin induces oral cancer cell apoptosis through the JNK1/2 pathway and is a potential therapeutic agent for oral cancer. © 2018 Wiley Periodicals, Inc.

A comprehensive analysis of coregulator recruitment, androgen receptor function and gene expression in prostate cancer

PubMed Central

Hu, Qiang; Senapati, Dhirodatta; Venkadakrishnan, Varadha Balaji; Wang, Dan; DePriest, Adam D; Schlanger, Simon E; Ben-Salem, Salma; Valenzuela, Malyn May; Willard, Belinda; Mudambi, Shaila; Swetzig, Wendy M; Das, Gokul M; Shourideh, Mojgan; Koochekpour, Shahriah; Falzarano, Sara Moscovita; Magi-Galluzzi, Cristina; Yadav, Neelu; Chen, Xiwei; Lao, Changshi; Wang, Jianmin; Billaud, Jean-Noel

2017-01-01

Standard treatment for metastatic prostate cancer (CaP) prevents ligand-activation of androgen receptor (AR). Despite initial remission, CaP progresses while relying on AR. AR transcriptional output controls CaP behavior and is an alternative therapeutic target, but its molecular regulation is poorly understood. Here, we show that action of activated AR partitions into fractions that are controlled preferentially by different coregulators. In a 452-AR-target gene panel, each of 18 clinically relevant coregulators mediates androgen-responsiveness of 0–57% genes and acts as a coactivator or corepressor in a gene-specific manner. Selectivity in coregulator-dependent AR action is reflected in differential AR binding site composition and involvement with CaP biology and progression. Isolation of a novel transcriptional mechanism in which WDR77 unites the actions of AR and p53, the major genomic drivers of lethal CaP, to control cell cycle progression provides proof-of-principle for treatment via selective interference with AR action by exploiting AR dependence on coregulators. PMID:28826481

Depletion of ribosomal protein L37 occurs in response to DNA damage and activates p53 through the L11/MDM2 pathway.

PubMed

Llanos, Susana; Serrano, Manuel

2010-10-01

Perturbation of ribosomal biogenesis has recently emerged as a relevant p53-activating pathway. This pathway can be initiated by depletion of certain ribosomal proteins, which is followed by the binding and inhibition of MDM2 by a different subset of ribosomal proteins that includes L11. Here, we report that depletion of L37 leads to cell cycle arrest in a L11- and p53-dependent manner. DNA damage can initiate ribosomal stress, although little is known about the mechanisms involved. We have found that some genotoxic insults, namely, UV light and cisplatin, lead to proteasomal degradation of L37 in the nucleoplasm and to the ensuing L11-dependent stabilization of p53. Moreover, ectopic L37 overexpression can attenuate the DNA damage response mediated by p53. These results support the concept that DNA damage-induced proteasomal degradation of L37 constitutes a mechanistic link between DNA damage and the ribosomal stress pathway, and is a relevant contributing signaling pathway for the activation of p53 in response to DNA damage.

Gallic acid induced apoptotic events in HCT-15 colon cancer cells.

PubMed

Subramanian, Aruna Priyadharshni; Jaganathan, Saravana Kumar; Mandal, Mahitosh; Supriyanto, Eko; Muhamad, Ida Idayu

2016-04-21

To investigate the inhibitory action of diet-derived phenolic compound gallic acid (GA) against HCT-15 colon cancer cells. The antiproliferative effect of GA against colon cancer cells was determined by performing thiazolyl blue tetrazolium bromide (MTT) assay. The colony forming ability of GA treated colon cancer cells was evaluated using the colony forming assay. The cell cycle changes induced by GA in HCT-15 cells were analyzed by propidium iodide staining. Levels of reactive oxygen species (ROS) and mitochondrial membrane potential of HCT-15 exposed to GA was assessed using 2',7'-dichlorfluorescein-diacetate and rhodamine-123 respectively, with the help of flow cytometry. Morphological changes caused by GA treatment in the colon cancer cells were identified by scanning electron microscope and photomicrograph examination. Apoptosis was confirmed using flow cytometric analysis of GA treated HCT-15 cells after staining with Yo-Pro-1. MTT assay results illustrated that GA has an inhibitory effect on HCT-15 cells with IC50 value of 740 μmol/L. A time-dependent inhibition of colony formation was evident with GA treatment. Cell cycle arrest was evident from the accumulation of GA treated HCT-15 cells at sub-G1 phase (0.98 ± 1.03 vs 58.01 ± 2.05) with increasing exposure time. Flow cytometric analysis of GA treated HCT-15 cells depicted early events associated with apoptosis like lipid layer breakage and fall in mitochondrial membrane potential apart from an increase in the generation of ROS which were in a time dependent manner. SEM and photomicrograph images of the GA-treated cells displayed membrane blebbing and cell shrinking characteristics of apoptosis. Further apoptosis confirmation by Yo-Pro-1 staining also showed the time-dependent increase of apoptotic cells after treatment. These results show that GA induced ROS dependent apoptosis and inhibited the growth of colon cancer cells.

Gallic acid induced apoptotic events in HCT-15 colon cancer cells

PubMed Central

Subramanian, Aruna Priyadharshni; Jaganathan, Saravana Kumar; Mandal, Mahitosh; Supriyanto, Eko; Muhamad, Ida Idayu

2016-01-01

AIM: To investigate the inhibitory action of diet-derived phenolic compound gallic acid (GA) against HCT-15 colon cancer cells. METHODS: The antiproliferative effect of GA against colon cancer cells was determined by performing thiazolyl blue tetrazolium bromide (MTT) assay. The colony forming ability of GA treated colon cancer cells was evaluated using the colony forming assay. The cell cycle changes induced by GA in HCT-15 cells were analyzed by propidium iodide staining. Levels of reactive oxygen species (ROS) and mitochondrial membrane potential of HCT-15 exposed to GA was assessed using 2’,7’-dichlorfluorescein-diacetate and rhodamine-123 respectively, with the help of flow cytometry. Morphological changes caused by GA treatment in the colon cancer cells were identified by scanning electron microscope and photomicrograph examination. Apoptosis was confirmed using flow cytometric analysis of GA treated HCT-15 cells after staining with Yo-Pro-1. RESULTS: MTT assay results illustrated that GA has an inhibitory effect on HCT-15 cells with IC50 value of 740 μmol/L. A time-dependent inhibition of colony formation was evident with GA treatment. Cell cycle arrest was evident from the accumulation of GA treated HCT-15 cells at sub-G1 phase (0.98 ± 1.03 vs 58.01 ± 2.05) with increasing exposure time. Flow cytometric analysis of GA treated HCT-15 cells depicted early events associated with apoptosis like lipid layer breakage and fall in mitochondrial membrane potential apart from an increase in the generation of ROS which were in a time dependent manner. SEM and photomicrograph images of the GA-treated cells displayed membrane blebbing and cell shrinking characteristics of apoptosis. Further apoptosis confirmation by Yo-Pro-1 staining also showed the time-dependent increase of apoptotic cells after treatment. CONCLUSION: These results show that GA induced ROS dependent apoptosis and inhibited the growth of colon cancer cells. PMID:27099438

YC-1 induces G0/G1 phase arrest and mitochondria-dependent apoptosis in cisplatin-resistant human oral cancer CAR cells.

PubMed

Lee, Miau-Rong; Lin, Chingju; Lu, Chi-Cheng; Kuo, Sheng-Chu; Tsao, Je-Wei; Juan, Yu-Ning; Chiu, Hong-Yi; Lee, Fang-Yu; Yang, Jai-Sing; Tsai, Fuu-Jen

2017-06-01

Oral cancer is a serious and fatal disease. Cisplatin is the first line of chemotherapeutic agent for oral cancer therapy. However, the development of drug resistance and severe side effects cause tremendous problems clinically. In this study, we investigated the pharmacologic mechanisms of YC-1 on cisplatin-resistant human oral cancer cell line, CAR. Our results indicated that YC-1 induced a concentration-dependent and time-dependent decrease in viability of CAR cells analyzed by MTT assay. Real-time image analysis of CAR cells by IncuCyte™ Kinetic Live Cell Imaging System demonstrated that YC-1 inhibited cell proliferation and reduced cell confluence in a time-dependent manner. Results from flow cytometric analysis revealed that YC-1 promoted G 0 /G 1 phase arrest and provoked apoptosis in CAR cells. The effects of cell cycle arrest by YC-1 were further supported by up-regulation of p21 and down-regulation of cyclin A, D, E and CDK2 protein levels. TUNEL staining showed that YC-1 caused DNA fragmentation, a late stage feature of apoptosis. In addition, YC-1 increased the activities of caspase-9 and caspase-3, disrupted the mitochondrial membrane potential (AYm) and stimulated ROS production in CAR cells. The protein levels of cytochrome c, Bax and Bak were elevated while Bcl-2 protein expression was attenuated in YC-1-treated CAR cells. In summary, YC-1 suppressed the viability of cisplatin-resistant CAR cells through inhibiting cell proliferation, arresting cell cycle at G 0 /G 1 phase and triggering mitochondria-mediated apoptosis. Our results provide evidences to support the potentially therapeutic application of YC-1 on fighting against drug resistant oral cancer in the future. © Author(s) 2017. This article is published with open access by China Medical University.

Bipolar mood cycles and lunar tidal cycles

PubMed Central

Wehr, T A

2018-01-01

In 17 patients with rapid cycling bipolar disorder, time-series analyses detected synchronies between mood cycles and three lunar cycles that modulate the amplitude of the moon’s semi-diurnal gravimetric tides: the 14.8-day spring–neap cycle, the 13.7-day declination cycle and the 206-day cycle of perigee-syzygies (‘supermoons’). The analyses also revealed shifts among 1:2, 1:3, 2:3 and other modes of coupling of mood cycles to the two bi-weekly lunar cycles. These shifts appear to be responses to the conflicting demands of the mood cycles’ being entrained simultaneously to two different bi-weekly lunar cycles with slightly different periods. Measurements of circadian rhythms in body temperature suggest a biological mechanism through which transits of one of the moon’s semi-diurnal gravimetric tides might have driven the patients’ bipolar cycles, by periodically entraining the circadian pacemaker to its 24.84-h rhythm and altering the pacemaker’s phase-relationship to sleep in a manner that is known to cause switches from depression to mania. PMID:28115741

Estrous cycle modulation of extracellular serotonin in mediobasal hypothalamus: role of the serotonin transporter and terminal autoreceptors.

PubMed

Maswood, S; Truitt, W; Hotema, M; Caldarola-Pastuszka, M; Uphouse, L

1999-06-12

In vivo microdialysis was used to examine extracellular serotonin (5-HT) in the mediobasal hypothalamus (MBH) of male and female Fischer (CDF-344) rats. Females from the stages of diestrus, proestrus, and estrus were used. Additionally, ovariectomized rats, primed subcutaneously (s.c.) with estradiol benzoate or estradiol benzoate plus progesterone were examined. Extracellular 5-HT in the MBH varied with stage of the estrous cycle and with the light/dark cycle. Proestrous females had the highest microdialysate concentrations of 5-HT during the light portion of the light/dark cycle and lowest concentrations during the dark portion of the cycle. Diestrous females had the highest levels during the dark portion of the cycle, while males and estrous females showed little change between light and dark portions of the cycle. In ovariectomized rats, there was no effect of 2.5 microg or 25 microg estradiol benzoate (s.c.) on extracellular 5-HT; but the addition of 500 microg progesterone, 48 h after estrogen priming, reduced microdialysate 5-HT near the threshold for detection. In intact females and in males, reverse perfusion with 3 microM fluoxetine, a selective serotonin reuptake inhibitor (SSRI), or 2 microM methiothepin, a 5-HT receptor antagonist, increased microdialysate concentrations of 5-HT. Estrous females and males showed nearly a 4-fold increase in microdialysate 5-HT in response to fluoxetine while smaller responses were seen in diestrous and proestrous rats. In contrast, proestrous rats showed the largest response to methiothepin. Estrous females showed a delayed response to methiothepin, but there was no methiothepin-induced increase in extracellular 5-HT in males. These findings are discussed in reference to the suggestion that extracellular 5-HT in the MBH is regulated in a manner that is gender and estrous cycle dependent. The 5-HT terminal autoreceptor may exert a greater role in proestrous females; the serotonin transporter appears to play a more active role in the regulation of extracellular 5-HT in estrous females and in males. Copyright 1999 Elsevier Science B.V.

Cycling the hot CNO: a teaching methodology

NASA Astrophysics Data System (ADS)

Frost-Schenk, J. W.; Diget, C. Aa; Bentley, M. A.; Tuff, A.

2018-03-01

An interactive activity to teach the hot Carbon, Nitrogen and Oxygen (HCNO) cycle is proposed. Justification for why the HCNO cycle is important is included via an example of x-ray bursts. The activity allows teaching and demonstration of half-life, nuclear isotopes, nuclear reactions, protons and α-particles, and catalytic processes. Whilst the process example is specific to astrophysics it may be used to teach more broadly about catalytic processes. This practical is designed for use with 10-20 participants, with the intention that the exercise will convey nuclear physics principles in a fun and interactive manner.

A global, myosin light chain kinase-dependent increase in myosin II contractility accompanies the metaphase-anaphase transition in sea urchin eggs.

PubMed

Lucero, Amy; Stack, Christianna; Bresnick, Anne R; Shuster, Charles B

2006-09-01

Myosin II is the force-generating motor for cytokinesis, and although it is accepted that myosin contractility is greatest at the cell equator, the temporal and spatial cues that direct equatorial contractility are not known. Dividing sea urchin eggs were placed under compression to study myosin II-based contractile dynamics, and cells manipulated in this manner underwent an abrupt, global increase in cortical contractility concomitant with the metaphase-anaphase transition, followed by a brief relaxation and the onset of furrowing. Prefurrow cortical contractility both preceded and was independent of astral microtubule elongation, suggesting that the initial activation of myosin II preceded cleavage plane specification. The initial rise in contractility required myosin light chain kinase but not Rho-kinase, but both signaling pathways were required for successful cytokinesis. Last, mobilization of intracellular calcium during metaphase induced a contractile response, suggesting that calcium transients may be partially responsible for the timing of this initial contractile event. Together, these findings suggest that myosin II-based contractility is initiated at the metaphase-anaphase transition by Ca2+-dependent myosin light chain kinase (MLCK) activity and is maintained through cytokinesis by both MLCK- and Rho-dependent signaling. Moreover, the signals that initiate myosin II contractility respond to specific cell cycle transitions independently of the microtubule-dependent cleavage stimulus.

A Global, Myosin Light Chain Kinase-dependent Increase in Myosin II Contractility Accompanies the Metaphase–Anaphase Transition in Sea Urchin Eggs

PubMed Central

Lucero, Amy; Stack, Christianna; Bresnick, Anne R.

2006-01-01

Myosin II is the force-generating motor for cytokinesis, and although it is accepted that myosin contractility is greatest at the cell equator, the temporal and spatial cues that direct equatorial contractility are not known. Dividing sea urchin eggs were placed under compression to study myosin II-based contractile dynamics, and cells manipulated in this manner underwent an abrupt, global increase in cortical contractility concomitant with the metaphase–anaphase transition, followed by a brief relaxation and the onset of furrowing. Prefurrow cortical contractility both preceded and was independent of astral microtubule elongation, suggesting that the initial activation of myosin II preceded cleavage plane specification. The initial rise in contractility required myosin light chain kinase but not Rho-kinase, but both signaling pathways were required for successful cytokinesis. Last, mobilization of intracellular calcium during metaphase induced a contractile response, suggesting that calcium transients may be partially responsible for the timing of this initial contractile event. Together, these findings suggest that myosin II-based contractility is initiated at the metaphase–anaphase transition by Ca2+-dependent myosin light chain kinase (MLCK) activity and is maintained through cytokinesis by both MLCK- and Rho-dependent signaling. Moreover, the signals that initiate myosin II contractility respond to specific cell cycle transitions independently of the microtubule-dependent cleavage stimulus. PMID:16837551

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mohapatra, Purusottam; Satapathy, Shakti Ranjan; Das, Dipon

Cigarette smoking is a key factor for the development and progression of different cancers including mammary tumor in women. Resveratrol (Res) is a promising natural chemotherapeutic agent that regulates many cellular targets including p21, a cip/kip family of cyclin kinase inhibitors involved in DNA damage-induced cell cycle arrest and blocking of DNA replication and repair. We have recently shown that cigarette smoke condensate (CSC) prepared from commercially available Indian cigarette can cause neoplastic transformation of normal breast epithelial MCF-10A cell. Here we studied the mechanism of Res mediated apoptosis in CSC transformed (MCF-10A-Tr) cells in vitro and in vivo. Resmore » mediated apoptosis in MCF-10A-Tr cells was a p21 dependent event. It increased the p21 protein expression in MCF-10A-Tr cells and MCF-10A-Tr cells-mediated tumors in xenograft mice. Res treatment reduced the tumor size(s) and expression of anti-apoptotic proteins (e.g. PI3K, AKT, NFκB) in solid tumor. The expressions of cell cycle regulatory (Cyclins, CDC-2, CDC-6, etc.), BER associated (Pol-β, Pol-δ, Pol-ε, Pol-η, RPA, Fen-1, DNA-Ligase-I, etc.) proteins and LP-BER activity decreased in MCF-10A-Tr cells but remain significantly unaltered in isogenic p21 null MCF-10A-Tr cells after Res treatment. Interestingly, no significant changes were noted in SP-BER activity in both the cell lines after Res exposure. Finally, it was observed that increased p21 blocks the LP-BER in MCF-10A-Tr cells by increasing its interaction with PCNA via competing with Fen-1 after Res treatment. Thus, Res caused apoptosis in CSC-induced cancer cells by reduction of LP-BER activity and this phenomenon largely depends on p21. - Highlights: • Resveratrol (Res) caused reduction of MCF-10A-Tr cell growth by inducing apoptosis. • Res caused cell cycle arrest and DNA damage in p21 dependent manner. • Res mediated LP-BER reduction in MCF-10A-Tr cells was a p21 dependent phenomenon. • Res inhibits BER and PI3K, AKT, and NFκB protein expressions in tumor and xenografts. • Res-induced-p21 inhibited DNA repair by modulating Fen-1 binding to PCNA complex.« less

PKI 166 induced redox signalling and apoptosis through activation of p53, MAP kinase and caspase pathway in epidermoid carcinoma.

PubMed

Das, Subhasis; Dey, Kaushik Kumar; Bharti, Rashmi; MaitiChoudhury, Sujata; Maiti, Sukumar; Mandal, Mahitosh

2012-01-01

Cellular redox changes have emerged as a pivotal and proximal event in cancer. PKI 166 is used to determine the effects of redox sensitive inhibition of EGFR, metastasis and apoptosis in epidermoid carcinoma. Cytotoxicity study of PKI 166 (IC50 1.0 microM) treated A431 cells were performed by MTT assay for 48 and 72 hrs. Morphological analysis of PKI 166 treated A431 cells for 48 hrs. revealed the cell shrinkage, loss of filopodia and lamellipodia by phase contrast and SEM images in dose dependent manner. It has cytotoxic effects through inhibiting cellular proliferation, leads to the induction of apoptosis, as increased fraction of sub-G1 phase of the cell cycle, chromatin condensation and DNA ladder. It inhibited cyclin-D1 and cyclin-E expression and induced p53, p21 expression in dose dependent manner. Consequently, an imbalance of Bax/Bcl-2 ratio triggered caspase cascade and subsequent cleavage of PARP, thereby shifting the balance in favour of apoptosis. PKI 166 treatment actively stimulated reactive oxygen species (ROS) and mitochondrial membrane depolarization. It inhibited some metastatic properties of A431 cells supressing colony formation by soft agar assay and inhibition of MMP 9 activity by gelatin zymography and western blot analysis. PKI 166 inhibited growth factor induced phosphorylation of EGFR, Akt, MAPK, JNK and colony formation in A431 cells. Thus the inhibition of proliferation was associated with redox regulation of the caspase cascade, EGFR, Akt/PI3K, MAPK/ ERK and JNK pathway. On the other hand, increased antioxidant activity leads to decreased ROS generation inhibit the anti-proliferative and apoptotic properties of PKI 166 in A431 cells. These observations indicated PKI 166 induced redox signalling dependent inhibition of cell proliferation, metastatic properties and induction of apoptotic potential in epidermoid carcinoma.

Ablation of an Ovarian Tumor Family Deubiquitinase Exposes the Underlying Regulation Governing the Plasticity of Cell Cycle Progression in Toxoplasma gondii.

PubMed

Dhara, Animesh; de Paula Baptista, Rodrigo; Kissinger, Jessica C; Snow, E Charles; Sinai, Anthony P

2017-11-21

The Toxoplasma genome encodes the capacity for distinct architectures underlying cell cycle progression in a life cycle stage-dependent manner. Replication in intermediate hosts occurs by endodyogeny, whereas a hybrid of schizogony and endopolygeny occurs in the gut of the definitive feline host. Here, we characterize the consequence of the loss of a cell cycle-regulated o varian tu mor (OTU family) deubiquitinase, OTUD3A of Toxoplasma gondii (TgOTUD3A; TGGT1_258780), in T. gondii tachyzoites. Rather than the mutation being detrimental, mutant parasites exhibited a fitness advantage, outcompeting the wild type. This phenotype was due to roughly one-third of TgOTUD3A-knockout (TgOTUD3A-KO) tachyzoites exhibiting deviations from endodyogeny by employing replication strategies that produced 3, 4, or 5 viable progeny within a gravid mother instead of the usual 2. We established the mechanistic basis underlying these altered replication strategies to be a dysregulation of centrosome duplication, causing a transient loss of stoichiometry between the inner and outer cores that resulted in a failure to terminate S phase at the attainment of 2N ploidy and/or the decoupling of mitosis and cytokinesis. The resulting dysregulation manifested as deviations in the normal transitions from S phase to mitosis (S/M) (endopolygeny-like) or M phase to cytokinesis (M/C) (schizogony-like). Notably, these imbalances are corrected prior to cytokinesis, resulting in the generation of normal progeny. Our findings suggest that decisions regarding the utilization of specific cell cycle architectures are controlled by a ubiquitin-mediated mechanism that is dependent on the absolute threshold levels of an as-yet-unknown target(s). Analysis of the TgOTUD3A-KO mutant provides new insights into mechanisms underlying the plasticity of apicomplexan cell cycle architecture. IMPORTANCE Replication by Toxoplasma gondii can occur by 3 distinct cell cycle architectures. Endodyogeny is used by asexual stages, while a hybrid of schizogony and endopolygeny is used by merozoites in the definitive feline host. Here, we establish that the disruption of an o varian- tu mor (OTU) family deubiquitinase, TgOTUD3A, in tachyzoites results in dysregulation of the mechanism controlling the selection of replication strategy in a subset of parasites. The mechanistic basis for these altered cell cycles lies in the unique biology of the bipartite centrosome that is associated with the transient loss of stoichiometry between the inner and outer centrosome cores in the TgOTUD3A-KO mutant. This highlights the importance of ubiquitin-mediated regulation in the transition from the nuclear to the budding phases of the cell cycle and provides new mechanistic insights into the regulation of the organization of the apicomplexan cell cycle. Copyright © 2017 Dhara et al.

Effects of allicin on both telomerase activity and apoptosis in gastric cancer SGC-7901 cells.

PubMed

Sun, Li; Wang, Xu

2003-09-01

To investigate the effects of allicin on both telomerase activity and apoptosis in gastric cancer SGC-7901 cells. The gastric cancer SGC-7901 adenocarcinoma cells were treated with allicin and the cell cycle, inhibitory rate, apoptosis, telomerase activity and morphologic changes were studied by MTT assay, flow cytometry (FCM), TRAP-PCR-ELISA assay, light microscope, electron microscope respectively. Results were compared with that of AZT (3'-Azido-3'-deoxythymidine). SGC-7901 cells were suppressed after exposure to allicin of 0.016 mg/ml, 0.05 mg/ml, and 0.1 mg/ml for 48 h. Compared with the control, the difference was significant (P<0.05). Allicin could induce apoptosis of the cells in a dose-dependent and non-linear manner and increase the proportion of cells in the G(2)/M phase. Compared with the control, the difference was significant in terms of the percentage of cells in the G2/M phase (P<0.05). Allicin could inhibit telomerase activity in a time-dependent and dose-dependent pattern. After exposure to allicin at 0.016 mg/ml for 24 hours, SGC-7901 cells showed typical morphologic change. Allicin can inhibit telomerase activity and induce apoptosis of gastric cancer SGC-7901 cells. Allicin may be more effective than AZT.

Loss of Cyclin-dependent Kinase 2 in the Pancreas Links Primary β-Cell Dysfunction to Progressive Depletion of β-Cell Mass and Diabetes*

PubMed Central

Kim, So Yoon; Lee, Ji-Hyeon; Merrins, Matthew J.; Gavrilova, Oksana; Bisteau, Xavier; Kaldis, Philipp; Satin, Leslie S.; Rane, Sushil G.

2017-01-01

The failure of pancreatic islet β-cells is a major contributor to the etiology of type 2 diabetes. β-Cell dysfunction and declining β-cell mass are two mechanisms that contribute to this failure, although it is unclear whether they are molecularly linked. Here, we show that the cell cycle regulator, cyclin-dependent kinase 2 (CDK2), couples primary β-cell dysfunction to the progressive deterioration of β-cell mass in diabetes. Mice with pancreas-specific deletion of Cdk2 are glucose-intolerant, primarily due to defects in glucose-stimulated insulin secretion. Accompanying this loss of secretion are defects in β-cell metabolism and perturbed mitochondrial structure. Persistent insulin secretion defects culminate in progressive deficits in β-cell proliferation, reduced β-cell mass, and diabetes. These outcomes may be mediated directly by the loss of CDK2, which binds to and phosphorylates the transcription factor FOXO1 in a glucose-dependent manner. Further, we identified a requirement for CDK2 in the compensatory increases in β-cell mass that occur in response to age- and diet-induced stress. Thus, CDK2 serves as an important nexus linking primary β-cell dysfunction to progressive β-cell mass deterioration in diabetes. PMID:28100774

A Comparison of Simple Methods to Incorporate Material Temperature Dependency in the Green's Function Method for Estimating Transient Thermal Stresses in Thick-Walled Power Plant Components.

PubMed

Rouse, James; Hyde, Christopher

2016-01-06

The threat of thermal fatigue is an increasing concern for thermal power plant operators due to the increasing tendency to adopt "two-shifting" operating procedures. Thermal plants are likely to remain part of the energy portfolio for the foreseeable future and are under societal pressures to generate in a highly flexible and efficient manner. The Green's function method offers a flexible approach to determine reference elastic solutions for transient thermal stress problems. In order to simplify integration, it is often assumed that Green's functions (derived from finite element unit temperature step solutions) are temperature independent (this is not the case due to the temperature dependency of material parameters). The present work offers a simple method to approximate a material's temperature dependency using multiple reference unit solutions and an interpolation procedure. Thermal stress histories are predicted and compared for realistic temperature cycles using distinct techniques. The proposed interpolation method generally performs as well as (if not better) than the optimum single Green's function or the previously-suggested weighting function technique (particularly for large temperature increments). Coefficients of determination are typically above 0 . 96 , and peak stress differences between true and predicted datasets are always less than 10 MPa.

Endocytosis of ABCG2 drug transporter caused by binding of 5D3 antibody: trafficking mechanisms and intracellular fate.

PubMed

Studzian, Maciej; Bartosz, Grzegorz; Pulaski, Lukasz

2015-08-01

ABCG2, a metabolite and xenobiotic transporter located at the plasma membrane (predominantly in barrier tissues and progenitor cells), undergoes a direct progressive endocytosis process from plasma membrane to intracellular compartments upon binding of 5D3 monoclonal antibody. This antibody is specific to an external epitope on the protein molecule and locks it in a discrete conformation within its activity cycle, presumably providing a structural trigger for the observed internalization phenomenon. Using routine and novel assays, we show that ABCG2 is endocytosed by a mixed mechanism: partially via a rapid, clathrin-dependent pathway and partially in a cholesterol-dependent, caveolin-independent manner. While the internalization process is entirely dynamin-dependent and converges initially at the early endosome, subsequent intracellular fate of ABCG2 is again twofold: endocytosis leads to only partial lysosomal degradation, while a significant fraction of the protein is retained in a post-endosomal compartment with the possibility of at least partial recycling back to the cell surface. This externally triggered, conformation-related trafficking pathway may serve as a general regulatory paradigm for membrane transporters, and its discovery was made possible thanks to consistent application of quantitative methods. Copyright © 2015 Elsevier B.V. All rights reserved.

Estradiol induces endothelial cell migration and proliferation through estrogen receptor-enhanced RhoA/ROCK pathway.

PubMed

Oviedo, Pilar J; Sobrino, Agua; Laguna-Fernandez, Andrés; Novella, Susana; Tarín, Juan J; García-Pérez, Miguel-Angel; Sanchís, Juan; Cano, Antonio; Hermenegildo, Carlos

2011-03-30

Migration and proliferation of endothelial cells are involved in re-endothelialization and angiogenesis, two important cardiovascular processes that are increased in response to estrogens. RhoA, a small GTPase which controls multiple cellular processes, is involved in the control of cell migration and proliferation. Our aim was to study the role of RhoA on estradiol-induced migration and proliferation and its dependence on estrogen receptors activity. Human umbilical vein endothelial cells were stimulated with estradiol, in the presence or absence of ICI 182780 (estrogen receptors antagonist) and Y-27632 (Rho kinase inhibitor). Estradiol increased Rho GEF-1 gene expression and RhoA (gene and protein expression and activity) in an estrogen receptor-dependent manner. Cell migration, stress fiber formation and cell proliferation were increased in response to estradiol and were also dependent on the estrogen receptors and RhoA activation. Estradiol decreased p27 levels, and significantly raised the expression of cyclins and CDK. These effects were counteracted by the use of either ICI 182780 or Y-27632. In conclusion, estradiol enhances the RhoA/ROCK pathway and increases cell cycle-related protein expression by acting through estrogen receptors. This results in an enhanced migration and proliferation of endothelial cells. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

The molecular basis of cytotoxicity of α-spinasterol from Ganoderma resinaceum: Induction of apoptosis and overexpression of p53 in breast and ovarian cancer cell lines.

PubMed

Sedky, Nada K; El Gammal, Zaynab H; Wahba, Amir E; Mosad, Eman; Waly, Zahraa Y; El-Fallal, Amira Ali; Arafa, Reem K; El-Badri, Nagwa

2018-05-01

Despite advances in therapy of breast and ovarian cancers, they still remain among the most imperative causes of cancer death in women. The first can be considered one of the most widespread diseases among females, while the latter is more lethal and needs prompt treatment. Thus, the research field can still benefit from discovery of new compounds that can be of potential use in management of these grave illnesses. We hereby aimed to assess the antitumor activity of the phytosterol α-spinasterol isolated from Ganoderma resinaceum mushroom on human breast cancer cell lines (MCF-7, MDA-MB-231), as well as, on human ovarian cancer cell line (SKOV-3). The anti-tumor activity of α-spinasterol, isolated from the mycelial extract of the Egyptian G. resinaceum, on human breast and ovarian cancer cell lines was evaluated by MTT cell viability assay and AnnexinV/propidium iodide apoptosis assay. The molecular mechanism underlying this effect was assessed by the relative expression of the following markers; tumor suppressor (p53, BRCA1, BRCA2), apoptotic marker (Bax) and cell cycle progression markers (cyclin dependent kinases cdk4/6) using real-time PCR. Cell cycle analysis was performed for the three investigated cancer cell lines to explore the effect on cell cycle progression. Our findings showed that α-spinasterol exhibited a higher antitumor activity on MCF-7 cells relative to SKOV-3 cells, while its lowest antitumor activity was against MDA-MB-231 cells. A significant increase in the expression of p53 and Bax was observed in cells treated with α-spinasterol, while cdk4/6 were significantly down-regulated upon exposure to α-spinasterol. Cell cycle analysis of α-spinasterol treated cells showed a G 0 -G 1 arrest. In conclusion, α-spinasterol isolated from G. resinaceum mushroom exerts a potent inhibitory activity on breast and ovarian cancer cell lines in a time- and dose-dependent manner. This can be reasonified in lights of the compound's ability to increase p53 and Bax expressions, and to lower the expression of cdk4/6. © 2017 Wiley Periodicals, Inc.

Interaction of plant chimeric calcium/calmodulin-dependent protein kinase with a homolog of eukaryotic elongation factor-1alpha

NASA Technical Reports Server (NTRS)

Wang, W.; Poovaiah, B. W.

1999-01-01

A chimeric Ca2+/calmodulin-dependent protein kinase (CCaMK) was previously cloned and characterized in this laboratory. To investigate the biological functions of CCaMK, the yeast two-hybrid system was used to isolate genes encoding proteins that interact with CCaMK. One of the cDNA clones obtained from the screening (LlEF-1alpha1) has high similarity with the eukaryotic elongation factor-1alpha (EF-1alpha). CCaMK phosphorylated LlEF-1alpha1 in a Ca2+/calmodulin-dependent manner. The phosphorylation site for CCaMK (Thr-257) was identified by site-directed mutagenesis. Interestingly, Thr-257 is located in the putative tRNA-binding region of LlEF-1alpha1. An isoform of Ca2+-dependent protein kinase (CDPK) phosphorylated multiple sites of LlEF-1alpha1 in a Ca2+-dependent but calmodulin-independent manner. Unlike CDPK, CCaMK phosphorylated only one site, and this site is different from CDPK phosphorylation sites. This suggests that the phosphorylation of EF-1alpha by these two kinases may have different functional significance. Although the phosphorylation of LlEF-1alpha1 by CCaMK is Ca2+/calmodulin-dependent, in vitro binding assays revealed that CCaMK binds to LlEF-1alpha1 in a Ca2+-independent manner. This was further substantiated by coimmunoprecipitation of CCaMK and EF-1alpha using the protein extract from lily anthers. Dissociation of CCaMK from EF-1alpha by Ca2+ and phosphorylation of EF-1alpha by CCaMK in a Ca2+/calmodulin-dependent manner suggests that these interactions may play a role in regulating the biological functions of EF-1alpha.

Treadmill training with partial body weight support after stroke.

PubMed

Hesse, Stefan; Werner, Cordula; von Frankenberg, Sophie; Bardeleben, Anita

2003-02-01

Treadmill therapy with partial BWS is a promising new approach to improve gait ability after stroke. This task-specific approach enables nonambulatory patients the repetitive practice of complex gait cycles instead of single-limb gait-preparatory maneuvers. Patients walk more symmetrically with less spasticity and better cardiovascular efficiency on the treadmill than with floor walking. Several controlled, clinical studies have shown the potential of treadmill training as a therapeutic intervention for nonambulatory patients with chronic stroke-related hemiplegia. Furthermore, controlled trials in acute stroke survivors have shown that treadmill training is as effective as other physiotherapy approaches that stress the repetitive practice of gait. Controlled multicenter trials comparing locomotor training with conventional therapy will be forthcoming. An electromechanical gait trainer that relieves the strenuous effort of the therapists and provides control of the trunk in a phase-dependent manner is a new technical alternative for gait training in severely impaired stroke patients.

Skin cancer chemoprevention by α-santalol.

PubMed

Zhang, Xiaoying; Dwivedi, Chandradhar

2011-01-01

Alpha-santalol, a naturally occurring terpenoid, has been shown to have chemopreventive effects on both 7, 12-dimethylbenz(a)anthracene (DMBA)-initiated and 12-O- tetradecanoylphorbol-13-acetate (TPA)-promoted skin cancer development in CD-1 and SENCAR mice, and UVB-induced skin cancer developments in SKH-1 hairless mice in a concentration-dependent manner. Studies have demonstrated that α-santalol could be effective against skin carcinogenesis through both induction of apoptosis via caspase activation together with dissipation of mitochondria membrane potential and cytochrome c release in A431 cells, and inhibition of cell growth via induction of G2/M phase arrest in both A431 cells and melanoma UACC-62 cells by altering multiple cell cycle regulatory proteins and complexes. This review summarizes the chemopreventive effects and molecular mechanisms of α-santalol on skin cancer development in both animal models and skin cancer cell lines.

Cyclophilin A Associates with Enterovirus-71 Virus Capsid and Plays an Essential Role in Viral Infection as an Uncoating Regulator

PubMed Central

Huang, Jiaoyan; Yan, Wenzhong; Wang, Jinglan; Su, Dan; Ni, Cheng; Li, Jian; Rao, Zihe; Liu, Lei; Lou, Zhiyong

2014-01-01

Viruses utilize host factors for their efficient proliferation. By evaluating the inhibitory effects of compounds in our library, we identified inhibitors of cyclophilin A (CypA), a known immunosuppressor with peptidyl-prolyl cis-trans isomerase activity, can significantly attenuate EV71 proliferation. We demonstrated that CypA played an essential role in EV71 entry and that the RNA interference-mediated reduction of endogenous CypA expression led to decreased EV71 multiplication. We further revealed that CypA directly interacted with and modified the conformation of H-I loop of the VP1 protein in EV71 capsid, and thus regulated the uncoating process of EV71 entry step in a pH-dependent manner. Our results aid in the understanding of how host factors influence EV71 life cycle and provide new potential targets for developing antiviral agents against EV71 infection. PMID:25275585

Salt-mediated multicell formation in Deinococcus radiodurans.

PubMed Central

Chou, F I; Tan, S T

1991-01-01

The highly radiation-resistant tetracoccal bacterium Deinococcus radiodurans exhibited a reversible multi-cell-form transition which depended on the NaCl concentration in the medium. In response to 0.8% NaCl addition into the medium, the pair/tetrad (designated 2/4) cells in a young culture grew and divided but did not separate and became 8-, 16-, and 32-cell units successively. In exponential growth phase, the cells divided in a 16/32 pattern. Potassium ions were equally effective as Na+ in mediating this multicell-formation effect; Mg2+, Li+, and Ca2+ also worked but produced less multiplicity. This effect appears to be species specific. This-section micrographs revealed that in a 16/32-cell unit, eight 2/4 cells were encased in an orderly manner within a large peripheral wall, showing five cycles of septation. Our results suggest the presence of a salt-sensitive mechanism for controlling cell separation in D. radiodurans. Images PMID:2022617

NLRP3 activation and mitosis are mutually exclusive events coordinated by NEK7, a new inflammasome component

PubMed Central

Shi, Hexin; Wang, Ying; Li, Xiaohong; Zhan, Xiaoming; Tan, Miao; Fina, Maggy; Su, Lijing; Pratt, David; Bu, Chun Hui; Hildebrand, Sara; Lyon, Stephen; Scott, Lindsay; Quan, Jiexia; Sun, Qihua; Russell, Jamie; Arnett, Stephanie; Jurek, Peter; Chen, Ding; Kravchenko, Vladimir V.; Mathison, John C.; Moresco, Eva Marie Y.; Monson, Nancy L.; Ulevitch, Richard J.; Beutler, Bruce

2015-01-01

The NLRP3 inflammasome responds to microbes and danger signals by processing and activating proinflammatory cytokines including IL-1β and IL-18. We show that NLRP3 inflammasome activation is restricted to interphase of the cell cycle by NEK7, a serine/threonine kinase previously implicated in mitosis. NLRP3 inflammasome activation requires NEK7, which binds to the NLRP3 leucine-rich repeat domain in a kinase-independent manner downstream from the induction of mitochondrial ROS. This interaction is necessary for NLRP3-ASC complex formation, ASC oligomerization, and caspase-1 activation. NEK7 promotes the NLRP3-dependent cellular inflammatory response to intraperitoneal monosodium urate challenge, and the development of experimental autoimmune encephalitis in mice. Our findings suggest NEK7 serves as a cellular switch that enforces mutual exclusivity between the inflammasome response and cell division. PMID:26642356

Redifferentiation of human hepatoma cells (SMMC-7721) induced by two new highly oxygenated bisabolane-type sesquiterpenes.

PubMed

Miao, Ruidong; Wei, Juan; Zhang, Qi; Sajja, Venkateswara; Yang, Jinbo; Wang, Qin

2008-12-01

Bisabolane-type sesquiterpenes are a class of biologically active compounds that has antitumour,antifungal, antibacterial,antioxidant and antivenom properties.We investigated the effect of two new highly oxygenated bisabolane-type sesquiterpenes (HOBS)isolated from Cremanthodium discoideum (C.discoideum) on tumour cells. Our results showed that HOBS induced morphological differentiation and reduced microvilli formation on the cell surface in SMMC-7721 cells.Flow cytometry analysis demonstrated that HOBS could induce cell-cycle arrest in the G1 phase. Moreover,HOBS was able to increase tyrosine-alpha ketoglutarate transaminase activity,decrease alpha- foetoprotein level and gamma-glutamyl transferase activity. In addition,we found that HOBS inhibited the anchorage- independent growth of SMMC-7721 cells in a dose-dependent manner.Taken together,all the above observations indicate that HOBS might be able to normalize malignant SMMC-7721 cells by inhibiting cell proliferation and inducing redifferentiation.

Design and synthesis of formononetin-dithiocarbamate hybrids that inhibit growth and migration of PC-3 cells via MAPK/Wnt signaling pathways

PubMed Central

Fu, Dong-Jun; Zhang, Li; Song, Jian; Mao, Ruo-Wang; Zhao, Ruo-Han; Liu, Ying-Chao; Hou, Yu-Hui; Li, Jia-Huan; Yang, Jia-Jia; Jin, Cheng-Yun; Li, Ping; Zi, Xiao-Lin; Liu, Hong-Min; Zhang, Sai-Yang; Zhang, Yan-Bing

2017-01-01

A series of novel formononetin-dithiocarbamate derivatives were designed, synthesized and evaluated for antiproliferative activity against three selected cancer cell line (MGC-803, EC-109, PC-3). The first structure-activity relationship (SAR) for this formononetin-dithiocarbamate scaffold is explored in this report with evaluation of 14 variants of the structural class. Among these analogues, tert-butyl 4-(((3-((3-(4-methoxyphenyl)-4-oxo-4H–chromen-7-yl)oxy)propyl)thio)carbonothioyl)piperazine-1-carboxylate (8i) showed the best inhibitory activity against PC-3 cells (IC50 = 1. 97 µM). Cellular mechanism studies elucidated 8i arrests cell cycle at G1 phase and regulates the expression of G1 checkpoint-related proteins in concentration-dependent manners. Furthermore, 8i could inhibit cell growth via MAPK signaling pathway and inhibit migration via Wnt pathway in PC-3 cells. PMID:28038329

Mechanisms of Dihydroartemisinin and Dihydroartemisinin/Holotransferrin Cytotoxicity in T-Cell Lymphoma Cells

PubMed Central

Zhao, Xindong; Zhao, Chunting; Zhao, Hongguo; Huo, Lanfen

2015-01-01

The validated therapeutic effects of dihydroartemisinin (DHA) in solid tumors have encouraged us to explore its potential in treating T-cell lymphoma. We found that Jurkat cells (a T-cell lyphoma cell line) were sensitive to DHA treatment with a IC50 of dihydroartemisinin. The cytotoxic effect of DHA in Jurkat cells showed a dose- and time- dependent manner. Interestingly, the cytotoxic effect of DHA was further enhanced by holotransferrin (HTF) due to the high expression of transferrin receptors in T-cell lymphoma. Mechanistically, DHA significantly increased the production of intracellular reactive oxygen species, which led to cell cycle arrest and apoptosis. The DHA treatment also inhibited the expression of protumorgenic factors including VEGF and telomerase catalytic subunit. Our results have proved the therapeutic effect of DHA in T-cell lymphoma. Especially in combination with HTF, DHA may provide a novel efficient approach in combating the deadly disease. PMID:26502166

Proliferation marker pKi-67 affects the cell cycle in a self-regulated manner.

PubMed

Schmidt, Mirko H H; Broll, Rainer; Bruch, Hans-Peter; Duchrow, Michael

2002-01-01

The proliferation marker pKi-67 is commonly used in research and pathology to detect proliferating cells. In a previous work, we found the protein to be associated with regulators of the cell cycle, controlling S-phase progression, as well as entry into and exit from mitosis. Here we investigate whether pKi-67 has a regulative effect on the cell cycle itself. For that purpose we cloned four fragments of pKi-67, together representing nearly the whole protein, and an N-terminal pKi-67 antisense oligonucleotide into a tetracycline inducible gene expression system. The sense fragments were C-terminally modified by addition of either a nuclear localization sequence (NLS) or a STOP codon to address the impact of their intracellular distribution. FACS based cell cycle analysis revealed that expression of nearly all pKi-67 domains and the antisense oligonucleotide led to a decreased amount of cells in S-phase and an increased number of cells in G(2)/M- and G(1)-phase. Subsequent analysis of the endogenous pKi-67 mRNA and protein levels revealed that the constructs with the most significant impact on the cell cycle were able to silence pKi-67 transcription as well. We conclude from the data that pKi-67 influences progression of S-phase and mitosis in a self-regulated manner and, therefore, effects the cell cycle checkpoints within both phases. Furthermore, we found pKi-67 mediates an anti-apoptotic effect on the cell and we verified that this marker, although it is a potential ribosomal catalyst, is not expressed in differentiated tissues with a high transcriptional activity. Copyright 2002 Wiley-Liss, Inc.

Assessment of Non-traditional Isotopic Ratios by Mass Spectrometry for Analysis of Nuclear Activities. Annual Report 2011

DOE Office of Scientific and Technical Information (OSTI.GOV)

Biegalski, Steven R.; Buchholz, Bruce A.

2011-08-24

The objective of this work is to identify isotopic ratios suitable for analysis via mass spectrometry that distinguish between commercial nuclear reactor fuel cycles, fuel cycles for weapons grade plutonium, and products from nuclear weapons explosions. Methods will also be determined to distinguish the above from medical and industrial radionuclide sources. Mass spectrometry systems will be identified that are suitable for field measurement of such isotopes in an expedient manner.

Dimethyl Sulfoxide Promotes the Multiple Functions of the Tumor Suppressor HLJ1 through Activator Protein-1 Activation in NSCLC Cells

PubMed Central

Wang, Chi-Chung; Lin, Sheng-Yi; Lai, Yi-Hua; Liu, Ya-Jung; Hsu, Yuan-Lin; Chen, Jeremy J. W.

2012-01-01

Background Dimethyl sulfoxide (DMSO) is an amphipathic molecule that displays a diversity of antitumor activities. Previous studies have demonstrated that DMSO can modulate AP-1 activity and lead to cell cycle arrest at the G1 phase. HLJ1 is a newly identified tumor and invasion suppressor that inhibits tumorigenesis and cancer metastasis. Its transcriptional activity is regulated by the transcription factor AP-1. However, the effects of DMSO on HLJ1 are still unknown. In the present study, we investigate the antitumor effects of DMSO through HLJ1 induction and demonstrate the mechanisms involved. Methods and Findings Low-HLJ1-expressing highly invasive CL1–5 lung adenocarcinoma cells were treated with various concentrations of DMSO. We found that DMSO can significantly inhibit cancer cell invasion, migration, proliferation, and colony formation capabilities through upregulation of HLJ1 in a concentration-dependent manner, whereas ethanol has no effect. In addition, the HLJ1 promoter and enhancer reporter assay revealed that DMSO transcriptionally upregulates HLJ1 expression through an AP-1 site within the HLJ1 enhancer. The AP-1 subfamily members JunD and JunB were significantly upregulated by DMSO in a concentration-dependent manner. Furthermore, pretreatment with DMSO led to a significant increase in the percentage of UV-induced apoptotic cells. Conclusions Our results suggest that DMSO may be an important stimulator of the tumor suppressor protein HLJ1 through AP-1 activation in highly invasive lung adenocarcinoma cells. Targeted induction of HLJ1 represents a promising approach for cancer therapy, which also implied that DMSO may serve as a potential lead compound or coordinated ligand for the development of novel anticancer drugs. PMID:22529897

Novel microtubule-targeted agent 6-chloro-4-(methoxyphenyl) coumarin induces G2-M arrest and apoptosis in HeLa cells

PubMed Central

Ma, Yi-ming; Zhou, Yu-bo; Xie, Chuan-ming; Chen, Dong-mei; Li, Jia

2012-01-01

Aim: To identify a novel coumarin analogue with the highest anticancer activity and to further investigate its anticancer mechanisms. Methods: The viability of cancer cells was investigated using the MTT assay. The cell cycle progression was evaluated using both flow cytometric and Western blotting analysis. Microtubule depolymerization was observed with immunocytochemistry in vivo and a tubulin depolymerization assay in vitro. Apoptosis was demonstrated using Annexin V/Propidium Iodide (PI) double-staining and sub-G1 analysis. Results: Among 36 analogues of coumarin, 6-chloro-4-(methoxyphenyl) coumarin showed the best anticancer activity (IC50 value about 200 nmol/L) in HCT116 cells. The compound had a broad spectrum of anticancer activity against 9 cancer cell lines derived from colon cancer, breast cancer, liver cancer, cervical cancer, leukemia, epidermoid cancer with IC50 value of 75 nmol/L–1.57 μmol/L but with low cytotocitity against WI-38 human lung fibroblasts (IC50 value of 12.128 μmol/L). The compound (0.04–10 μmol/L) induced G2-M phase arrest in HeLa cells in a dose-dependent manner, which was reversible after the compound was removed. The compound (10–300 μmol/L) induced the depolymerization of purified porcine tubulin in vitro. Finally, the compound (0.04–2.5 μmol/L) induced apoptosis of HeLa cells in dose- and time-dependent manners. Conclusion: 6-Chloro-4-(methoxyphenyl) coumarin is a novel microtubule-targeting agent that induces G2–M arrest and apoptosis in HeLa cells. PMID:22266726

Artemisia leaf extract induces apoptosis in human endometriotic cells through regulation of the p38 and NFκB pathways.

PubMed

Kim, Ji-Hyun; Jung, Seung-Hyun; Yang, Yeong-In; Ahn, Ji-Hye; Cho, Jin-Gyeong; Lee, Kyung-Tae; Baek, Nam-In; Choi, Jung-Hye

2013-02-13

Artemisia leaves have long been used for the treatment of gynecological disorders, including infertility and dysmenorrhea, which can be commonly caused by endometriosis. In the present study, we investigated the effect of Artemisia princeps extract (APE) on the cell growth and apoptosis of human endometriotic cells. MTT assays and FACS analysis using PI and Annexin staining were performed to study cell viability, cell cycle progression, and apoptosis. We also explored the mechanism of APE-induced effects by evaluating the activation of caspases, Akt, p38, and NFκB. The expressions of XIAP, Bcl-2, and Bcl-xL were measured by real-time RT-PCR and Western blot analyses. APE significantly inhibited the cell viability of 11Z and 12Z human endometriotic epithelial cells. Interestingly, endometriotic cells were more sensitive to APE treatment than immortalized endometrial cells (HES). Treatment with APE induced apoptosis of 11Z cells in a time-dependent manner, as shown by accumulation of sub G1 and apoptotic cell populations. In addition, treatment with APE stimulated the activation of caspase -3, -8, and -9 in a dose- and time-dependent manner. Furthermore, p38 was activated by APE treatment, and the p38 inhibitor SB203580 markedly inhibited APE-induced cell death in 11Z cells. Moreover, treatment with APE suppressed the activation of NFκB and the expressions of anti-apoptotic factors such as XIAP, Bcl-2, and Bcl-xL. These results indicate that APE is a potential anti-endometriotic agent, acting to induce apoptosis of endometrial cells through the modulation of the p38 and NFκB pathways. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Stimulation of mucosal secretion by lubiprostone (SPI-0211) in guinea pig small intestine and colon

PubMed Central

Fei, Guijun; Wang, Yu-Zhong; Liu, Sumei; Hu, Hong-Zhen; Wang, Guo-Du; Qu, Mei-Hua; Wang, Xi-Yu; Xia, Yun; Sun, Xiaohong; Bohn, Laura M.; Cooke, Helen J.; Wood, Jackie D.

2009-01-01

Actions of lubiprostone, a selective type-2 chloride channel activator, on mucosal secretion were investigated in guinea pig small intestine and colon. Flat-sheet preparations were mounted in Ussing flux chambers for recording short-circuit current (Isc) as a marker for electrogenic chloride secretion. Lubiprostone, applied to the small intestinal mucosa in eight concentrations ranging from 1–3000 nM, evoked increases in Isc in a concentration-dependent manner with an EC50 of 42.5 nM. Lubiprostone applied to the mucosa of the colon in eight concentrations ranging from 1–3000 nM evoked increases in Isc in a concentration-dependent manner with an EC50 of 31.7 nM. Blockade of enteric nerves by tetrodotoxin did not influence stimulation of Isc by lubiprostone. Antagonists acting at prostaglandin (PG)E2, EP1–3, or EP4 receptors did not suppress stimulation of Isc by lubiprostone but suppressed or abolished PGE2-evoked responses. Substitution of gluconate for chloride abolished all responses to lubiprostone. The selective CFTR channel blocker, CFTR(inh)-172, did not suppress lubiprostone-evoked Isc. The broadly acting blocker, glibenclamide, suppressed (P < 0.001) lubiprostone-evoked Isc. Lubiprostone, in the presence of tetrodotoxin, enhanced carbachol-evoked Isc. The cholinergic component, but not the putative vasoactive intestinal peptide component, of neural responses to electrical field stimulation was enhanced by lubiprostone. Application of any of the prostaglandins, E2, F2, or I2, evoked depolarization of the resting membrane potential in enteric neurons. Unlike the prostaglandins, lubiprostone did not alter the electrical behavior of enteric neurons. Exposure to the histamine H2 receptor agonists increased basal Isc followed by persistent cyclical increases in Isc. Lubiprostone increased the peak amplitude of the dimaprit-evoked cycles. PMID:19179625

Stimulation of mucosal secretion by lubiprostone (SPI-0211) in guinea pig small intestine and colon.

PubMed

Fei, Guijun; Wang, Yu-Zhong; Liu, Sumei; Hu, Hong-Zhen; Wang, Guo-Du; Qu, Mei-Hua; Wang, Xi-Yu; Xia, Yun; Sun, Xiaohong; Bohn, Laura M; Cooke, Helen J; Wood, Jackie D

2009-04-01

Actions of lubiprostone, a selective type-2 chloride channel activator, on mucosal secretion were investigated in guinea pig small intestine and colon. Flat-sheet preparations were mounted in Ussing flux chambers for recording short-circuit current (Isc) as a marker for electrogenic chloride secretion. Lubiprostone, applied to the small intestinal mucosa in eight concentrations ranging from 1-3000 nM, evoked increases in Isc in a concentration-dependent manner with an EC50 of 42.5 nM. Lubiprostone applied to the mucosa of the colon in eight concentrations ranging from 1-3000 nM evoked increases in Isc in a concentration-dependent manner with an EC50 of 31.7 nM. Blockade of enteric nerves by tetrodotoxin did not influence stimulation of Isc by lubiprostone. Antagonists acting at prostaglandin (PG)E2, EP1-3, or EP4 receptors did not suppress stimulation of Isc by lubiprostone but suppressed or abolished PGE2-evoked responses. Substitution of gluconate for chloride abolished all responses to lubiprostone. The selective CFTR channel blocker, CFTR(inh)-172, did not suppress lubiprostone-evoked Isc. The broadly acting blocker, glibenclamide, suppressed (P<0.001) lubiprostone-evoked Isc. Lubiprostone, in the presence of tetrodotoxin, enhanced carbachol-evoked Isc. The cholinergic component, but not the putative vasoactive intestinal peptide component, of neural responses to electrical field stimulation was enhanced by lubiprostone. Application of any of the prostaglandins, E2, F2, or I2, evoked depolarization of the resting membrane potential in enteric neurons. Unlike the prostaglandins, lubiprostone did not alter the electrical behavior of enteric neurons. Exposure to the histamine H2 receptor agonists increased basal Isc followed by persistent cyclical increases in Isc. Lubiprostone increased the peak amplitude of the dimaprit-evoked cycles.

Homocysteine induces oxidative stress to damage trabecular meshwork cells.

PubMed

You, Zhi-Peng; Zhang, Yue-Zhi; Zhang, Yu-Lan; Shi, Lu; Shi, Ke

2018-05-01

The aim of the present study was to investigate the effect of homocysteine (Hcy) in on human trabecular meshwork cells (HTMCs). A total of 41 patients with primary open-angle glaucoma (POAG) and 53 patients with senile cataracts (control group) were recruited. Plasma and aqueous humor samples were collected and the Hcy concentrations were determined using enzymatic cycling assays. In cell experiments, normal HTMCs were passaged and randomly divided into a blank control group, a normal HTMC group and experimental groups, which were treated with different concentrations of Hcy. The HTMC activities were detected using the Cell Counting Kit-8 method and the HTMC mitochondrial membrane potential (MMP) was detected using JC-1 staining. Reactive oxygen species (ROS) released by trabecular meshwork cells was detected using flow cytometry and superoxide dismjutase-1 (SOD1) expression was detected using immunoblotting. The results revealed that the concentration of Hcy in the plasma and aqueous humor of the POAG group (14.44±0.86 and 1.60±0.27 µmol/l, respectively) was significantly higher compared with the control group (10.82±0.29 and 0.69±0.39 µmol/l). All tested concentrations (30, 100, 300 and 1,000 µmol/l) of Hcy reduced the MMP in HTMCs and inhibited HTMC proliferation in a dose-dependent manner. ROS production by HTMCs significantly increased with increased concentrations of Hcy, whereas SOD1 expression significantly decreased in a dose-dependent manner. In summary, patients with POAG were demonstrated to have increased concentrations of Hcy in the plasma and aqueous humor. High concentrations of Hcy in HTMCs induced an oxidative stress state, thereby further inhibiting HTMC proliferation. The results of the present study demonstrate that Hcy may be a potential treatment target in patients with POAG.

Persea declinata (Bl.) Kosterm Bark Crude Extract Induces Apoptosis in MCF-7 Cells via G0/G1 Cell Cycle Arrest, Bcl-2/Bax/Bcl-xl Signaling Pathways, and ROS Generation

PubMed Central

Wong, Yi Li; Wong, Won Fen; Ali Mohd, Mustafa; Hadi, A. Hamid A.

2014-01-01

Persea declinata (Bl.) Kosterm is a member of the Lauraceae family, widely distributed in Southeast Asia. It is from the same genus with avocado (Persea americana Mill), which is widely consumed as food and for medicinal purposes. In the present study, we examined the anticancer properties of Persea declinata (Bl.) Kosterm bark methanolic crude extract (PDM). PDM exhibited a potent antiproliferative effect in MCF-7 human breast cancer cells, with an IC50 value of 16.68 µg/mL after 48 h of treatment. We observed that PDM caused cell cycle arrest and subsequent apoptosis in MCF-7 cells, as exhibited by increased population at G0/G1 phase, higher lactate dehydrogenase (LDH) release, and DNA fragmentation. Mechanistic studies showed that PDM caused significant elevation in ROS production, leading to perturbation of mitochondrial membrane potential, cell permeability, and activation of caspases-3/7. On the other hand, real-time PCR and Western blot analysis showed that PDM treatment increased the expression of the proapoptotic molecule, Bax, but decreased the expression of prosurvival proteins, Bcl-2 and Bcl-xL, in a dose-dependent manner. These findings imply that PDM could inhibit proliferation in MCF-7 cells via cell cycle arrest and apoptosis induction, indicating its potential as a therapeutic agent worthy of further development. PMID:24808916

NaTi2(PO4)3 as an Aqueous Anode: Degradation Mechanisms and Mitigation Techniques

NASA Astrophysics Data System (ADS)

Mohamed, Alexander I.

With the proliferation of renewable energy sources, there has been a growing interest in battery chemistries for grid scale energy storage. Aqueous sodium ion batteries are particularly interesting for large scale energy storage because of their low cost and high safety, however, they tend to show poor long term stability. NaTi2(PO4)3 (NTP) shows promise as an anode for these systems with excellent long term stability when cycled quickly. When cycled slowly, NaTi2(PO4) 3 shows rapid capacity fade. The reasons for this rate depend capacity fade is poorly understood and is the topic of this document. It has been found that the products of the hydrogen evolution reaction, H2(g) and OH-, are the two largest contributors to capacity fade. High electrolyte pH caused by generation of OH- promotes dissolution of NTP during extend cycling, this is exacerbated when the pH increase above 11. The single greatest cause of apparent capacity fade for this material is loss of electrochemical surface area due to hydrogen gas entrapment within the porous structure of the electrode. Capacity lost in this manner can be recovered through reinfiltration of the electrode. The detrimental effects of gas entrapment within the electrode can be partially mitigated through compositing of the electrode with activated carbon and enhancing the wettability of the pores through addition of a surfactant to the electrolyte.

CFS-1686 causes cell cycle arrest at intra-S phase by interference of interaction of topoisomerase 1 with DNA.

PubMed

Lin, Ru-Wei; Yang, Chia-Ning; Ku, ShengYu; Ho, Cheng-Jung; Huang, Shih-Bo; Yang, Min-Chi; Chang, Hsin-Wen; Lin, Chun-Mao; Hwang, Jaulang; Chen, Yeh-Long; Tzeng, Cherg-Chyi; Wang, Chihuei

2014-01-01

CFS-1686 (chemical name (E)-N-(2-(diethylamino)ethyl)-4-(2-(2-(5-nitrofuran-2-yl)vinyl)quinolin-4-ylamino)benzamide) inhibits cell proliferation and triggers late apoptosis in prostate cancer cell lines. Comparing the effect of CFS-1686 on cell cycle progression with the topoisomerase 1 inhibitor camptothecin revealed that CFS-1686 and camptothecin reduced DNA synthesis in S-phase, resulting in cell cycle arrest at the intra-S phase and G1-S boundary, respectively. The DNA damage in CFS-1686 and camptothecin treated cells was evaluated by the level of ATM phosphorylation, γH2AX, and γH2AX foci, showing that camptothecin was more effective than CFS-1686. However, despite its lower DNA damage capacity, CFS-1686 demonstrated 4-fold higher inhibition of topoisomerase 1 than camptothecin in a DNA relaxation assay. Unlike camptothecin, CFS-1686 demonstrated no activity on topoisomerase 1 in a DNA cleavage assay, but nevertheless it reduced the camptothecin-induced DNA cleavage of topoisomerase 1 in a dose-dependent manner. Our results indicate that CFS-1686 might bind to topoisomerase 1 to inhibit this enzyme from interacting with DNA relaxation activity, unlike campothecin's induction of a topoisomerase 1-DNA cleavage complex. Finally, we used a computer docking strategy to localize the potential binding site of CFS-1686 to topoisomerase 1, further indicating that CFS-1686 might inhibit the binding of Top1 to DNA.

Isorhapontigenin (ISO) inhibited cell transformation by inducing G0/G1 phase arrest via increasing MKP-1 mRNA Stability.

PubMed

Gao, Guangxun; Chen, Liang; Li, Jingxia; Zhang, Dongyun; Fang, Yong; Huang, Haishan; Chen, Xiequn; Huang, Chuanshu

2014-05-15

The cancer chemopreventive property of Chinese herb new isolate isorhapontigenin (ISO) and mechanisms underlying its activity have never been explored. Here we demonstrated that ISO treatment with various concentrations for 3 weeks could dramatically inhibit TPA/EGF-induced cell transformation of Cl41 cells in Soft Agar assay, whereas co-incubation of cells with ISO at the same concentrations could elicit G0/G1 cell-cycle arrest without redundant cytotoxic effects on non-transformed cells. Further studies showed that ISO treatment resulted in cyclin D1 downregulation in dose- and time-dependent manner. Our results indicated that ISO regulated cyclin D1 at transcription level via targeting JNK/C-Jun/AP-1 activation. Moreover, we found that ISO-inhibited JNK/C-Jun/AP-1 activation was mediated by both upregulation of MKP-1 expression through increasing its mRNA stability and deactivating MKK7. Most importantly, MKP-1 knockdown could attenuate ISO-mediated suppression of JNK/C-Jun activation and cyclin D1 expression, as well as G0/G1 cell cycle arrest and cell transformation inhibition, while ectopic expression of FLAG-cyclin D1 T286A mutant also reversed ISO-induced G0/G1 cell-cycle arrest and inhibition of cell transformation. Our results demonstrated that ISO is a promising chemopreventive agent via upregulating mkp-1 mRNA stability, which is distinct from its cancer therapeutic effect with downregulation of XIAP and cyclin D1 expression.

Cryptotanshinone suppresses the proliferation and induces the apoptosis of pancreatic cancer cells via the STAT3 signaling pathway.

PubMed

Ge, Yuqing; Yang, Bo; Chen, Zhe; Cheng, Rubin

2015-11-01

Pancreatic cancer remains a challenging disease worldwide. Cryptotanshinone (CPT) is one of the active constituents of Salvia miltiorrhiza Bunge and exhibits significant antitumor activities in several human cancer cells. However, the efficacy and molecular mechanism of CPT in pancreatic cancer remains to be elucidated. In the present study, the effect of CPT on the proliferation, apoptosis and cell cycle of human pancreatic cancer cell BxPC‑3 cells was evaluated. The results demonstrated that CPT inhibited proliferation of the BxPC‑3 cells in a concentration‑dependent manner, and significantly induced cell apoptosis and cell cycle arrest. The protein levels of cleaved caspase‑3, caspase‑9 and poly ADP ribose polymerase were upregulated, while the levels of c‑myc, survivin and cyclin D1 were downregulated following treatment with CPT. In addition, CPT decreased the activities of signal transducer and activator of transcription 3 (STAT3) and several upstream regulatory signaling pathways after 24 h. However, CPT only inhibited the phosphorylation of STAT3 Tyr705 within 30 min, without marked effects on the phosphorylation of the other proteins. These results suggested that the inhibition of STAT3 activity by CPT was directly and independent of the upstream regulators in human pancreatic cancer. The present study demonstrated that CPT exerts anticancer effects by inducing apoptosis and cell cycle arrest via inhibition of the STAT3 signaling pathway in human BxPC-3 cells.

Identification by Random Mutagenesis of Functional Domains in KREPB5 That Differentially Affect RNA Editing between Life Cycle Stages of Trypanosoma brucei.

PubMed

McDermott, Suzanne M; Carnes, Jason; Stuart, Kenneth

2015-12-01

KREPB5 is an essential component of ∼ 20S editosomes in Trypanosoma brucei which contains a degenerate, noncatalytic RNase III domain. To explore the function of this protein, we used a novel approach to make and screen numerous conditional null T. brucei bloodstream form cell lines that express randomly mutagenized KREPB5 alleles. We identified nine single amino acid substitutions that could not complement the conditional loss of wild-type KREPB5. Seven of these were within the RNase III domain, and two were in the C-terminal region that has no homology to known motifs. Exclusive expression of these mutated KREPB5 alleles in the absence of wild-type allele expression resulted in growth inhibition, the loss of ∼ 20S editosomes, and inhibition of RNA editing in BF cells. Eight of these mutations were lethal in bloodstream form parasites but not in procyclic-form parasites, showing that multiple domains function in a life cycle-dependent manner. Amino acid changes at a substantial number of positions, including up to 7 per allele, allowed complementation and thus did not block KREPB5 function. Hence, the degenerate RNase III domain and a newly identified domain are critical for KREPB5 function and have differential effects between the life cycle stages of T. brucei that differentially edit mRNAs. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Effects of Wannachawee Recipe with Antipsoriatic Activity on Suppressing Inflammatory Cytokine Production in HaCaT Human Keratinocytes

PubMed Central

Na Takuathung, Mingkwan; Wongnoppavich, Ariyaphong; Pitchakarn, Pornsiri; Panthong, Ampai; Khonsung, Parirat; Soonthornchareonnon, Noppamas

2017-01-01

Psoriasis is a chronic inflammatory and immune-mediated skin disease. The pathogenesis involves T cells activation via the IL-23/Th17 axis. Conventional treatments of psoriasis have adverse events influencing patients' adherence. Wannachawee Recipe (WCR) has been effectively used as Thai folk remedy for psoriasis patients; however, preclinical evidence defining how WCR works is still lacking. This study defined mechanisms for its antiproliferation and anti-inflammatory effects in HaCaT cells. The cytotoxicity and antiproliferation results from SRB and CCK-8 assays showed that WCR inhibited the growth and viability of HaCaT cells in a concentration-dependent manner. The distribution of cell cycle phases determined by flow cytometry showed that WCR did not interrupt cell cycle progression. Interestingly, RT-qPCR revealed that WCR significantly decreased the mRNA expression of IL-1β, IL-6, IL-8, IL-17A, IL-22, IL-23, and TNF-α but induced IL-10 expression in TNF-α- and IFN-γ-induced HaCaT cells. At the protein level determined by ELISA, WCR significantly reduced the secretion of IL-17A, IL-22, and IL-23. The WCR at low concentrations was proved to possess anti-inflammatory effect without cytotoxicity and it did not interfere with cell cycle of keratinocytes. This is the first study to provide convincing evidence that WCR is a potential candidate for development of effective psoriasis therapies. PMID:28900461

Effects of Wannachawee Recipe with Antipsoriatic Activity on Suppressing Inflammatory Cytokine Production in HaCaT Human Keratinocytes.

PubMed

Na Takuathung, Mingkwan; Wongnoppavich, Ariyaphong; Pitchakarn, Pornsiri; Panthong, Ampai; Khonsung, Parirat; Chiranthanut, Natthakarn; Soonthornchareonnon, Noppamas; Sireeratawong, Seewaboon

2017-01-01

Psoriasis is a chronic inflammatory and immune-mediated skin disease. The pathogenesis involves T cells activation via the IL-23/Th17 axis. Conventional treatments of psoriasis have adverse events influencing patients' adherence. Wannachawee Recipe (WCR) has been effectively used as Thai folk remedy for psoriasis patients; however, preclinical evidence defining how WCR works is still lacking. This study defined mechanisms for its antiproliferation and anti-inflammatory effects in HaCaT cells. The cytotoxicity and antiproliferation results from SRB and CCK-8 assays showed that WCR inhibited the growth and viability of HaCaT cells in a concentration-dependent manner. The distribution of cell cycle phases determined by flow cytometry showed that WCR did not interrupt cell cycle progression. Interestingly, RT-qPCR revealed that WCR significantly decreased the mRNA expression of IL-1 β , IL-6, IL-8, IL-17A, IL-22, IL-23, and TNF- α but induced IL-10 expression in TNF- α - and IFN- γ -induced HaCaT cells. At the protein level determined by ELISA, WCR significantly reduced the secretion of IL-17A, IL-22, and IL-23. The WCR at low concentrations was proved to possess anti-inflammatory effect without cytotoxicity and it did not interfere with cell cycle of keratinocytes. This is the first study to provide convincing evidence that WCR is a potential candidate for development of effective psoriasis therapies.

Berberine sensitizes nasopharyngeal carcinoma cells to radiation through inhibition of Sp1 and EMT.

PubMed

Wang, Jun; Kang, Min; Wen, Qin; Qin, Yu-Tao; Wei, Zhu-Xin; Xiao, Jing-Jian; Wang, Ren-Sheng

2017-04-01

Nasopharyngeal carcinoma (NPC) is a tumor of epithelial origin with radiotherapy as its standard treatment. However, radioresistance remains a critical issue in the treatment of NPC. This study aimed to investigate the effect of berberine on the proliferation, cell cycle regulation, apoptosis, radioresistance of NPC cells and whether specificity protein 1 (Sp1) is a functional target of berberine. Our results showed that treatment with berberine reduced the proliferation and viability of CNE-2 cells in a dose- and time‑dependent manner. Berberine induced cell cycle arrest in the G0/G1 phase and apoptosis. In CNE-2 cells exposed to gamma‑ray irradiation, berberine reduced cell viability at various concentrations (25, 50, 75 and 100 µmol/l). Berberine significantly decreased mRNA and protein expression of Sp1 in the CNE-2 cells. Mithramycin A, a selective Sp1 inhibitor, enhanced the radiosensitivity and the rate of apoptosis in the CNE-2 cells. Berberine inhibited transforming growth factor-β (TGF-β)-induced tumor invasion and suppressed epithelial-to-mesenchymal transition (EMT) process, as evidenced by increased E-cadherin and decreased vimentin proteins. Sp1 may be required for the TGF-β1-induced invasion and EMT by berberine. In conclusion, berberine demonstrated the ability to suppress proliferation, induce cell cycle arrest and apoptosis, and enhance radiosensitivity of the CNE-2 NPC cells. Sp1 may be a target of berberine which is decreased during the radiosensitization of berberine.

Expression, regulation, and function of drug transporters in cervicovaginal tissues of a mouse model used for microbicide testing

PubMed Central

Zhou, Tian; Hu, Minlu; Pearlman, Andrew; Rohan, Lisa C.

2016-01-01

P-glycoprotein (P-gp), breast cancer resistance protein (BCRP), and multidrug resistance protein 4 (MRP4) are three efflux transporters that play key roles in the pharmacokinetics of antiretroviral drugs used in the pre-exposure prophylaxis of HIV sexual transmission. In this study, we investigated the expression, regulation, and function of these transporters in cervicovaginal tissues of a mouse model. Expression and regulation were examined using real-time RT-PCR and immunohistochemical staining, in the mouse tissues harvested at estrus and diestrus stages under natural cycling or after hormone synchronization. The three transporters were expressed at moderate to high levels compared to the liver. Transporter proteins were localized in various cell types in different tissue segments. Estrous cycle and exogenous hormone treatment affected transporter mRNA and protein expression, in a tissue- and transporter-dependent manner. Depo-Provera-synchronized mice were dosed vaginally or intraperitoneally with 3H-TFV, with or without MK571 co-administration, to delineate the function of cervicovaginal Mrp4. Co-administration of MK571 significantly increased the concentration of vaginally-administered TFV in endocervix and vagina. MK571 increased the concentration of intraperitoneally-administered TFV in the cervicovaginal lavage and vagina by several fold. Overall, P-gp, Bcrp, and Mrp4 were positively expressed in mouse cervicovaginal tissues, and their expression can be regulated by the estrous cycle or by exogenous hormones. In this model, the Mrp4 transporter impacted TFV distribution in cervicovaginal tissues. PMID:27453435

Cadmium induces histopathological injuries and ultrastructural changes in the liver of freshwater turtle (Chinemys reevesii).

PubMed

Huo, Junfeng; Dong, Aiguo; Wang, Yonghui; Lee, Shaochin; Ma, Cungen; Wang, Lan

2017-11-01

The study investigated the histopathological and ultrastructural lesions of liver of freshwater turtle Chinemys reevesii exposed to Cadmium (Cd). The animals were exposed to 0 mg kg -1 (0.85% normal saline (NS)), 7.5 mg kg -1 , 15 mg kg -1 , 30 mg kg -1 Cd chloride separately by intraperitoneal injection. Liver samples were collected for examination of lesions under light and electronic microscopes. Results showed that liver tissues from Cd -treated animals presented various degrees of histopathological lesions. Liver cells showed swollen, degeneration and necrosis with dose-dependent manner. Under electronic microscope, nucleus, mitochondria and rough endoplasmic reticulum presented various degrees of lesions with dose-dependent manner. In conclusion, Cd has significant toxicity on liver tissue of the freshwater turtle, which occurs in a dose-dependent manner. Copyright © 2017 Elsevier Ltd. All rights reserved.

Thiocoraline alters neuroendocrine phenotype and activates the Notch pathway in MTC-TT cell line

PubMed Central

Tesfazghi, Sara; Eide, Jacob; Dammalapati, Ajitha; Korlesky, Colin; Wyche, Thomas P; Bugni, Tim S; Chen, Herbert; Jaskula-Sztul, Renata

2013-01-01

Medullary thyroid cancer (MTC) is an aggressive neuroendocrine tumor (NET). Previous research has shown that activation of Notch signaling has a tumor suppressor role in NETs. The potential therapeutic effect of thiocoraline on the activation of the Notch pathway in an MTC cell line (TT) was investigated. Thiocoraline was isolated from a marine bacterium Verrucosispora sp. MTT assay (3-[4, 5-dimethylthiazole-2-yl]-2, 5-diphenyltetrazolium bromide) was used to determine the IC50 value and to measure cell proliferation. Western blot revealed the expression of Notch isoforms, NET, and cell cycle markers. Cell cycle progression was validated by flow cytometry. The mRNA expression of Notch isoforms and downstream targets were measured using real-time PCR. The IC50 value for thiocoraline treatment in TT cells was determined to be 7.6 nmol/L. Thiocoraline treatment decreased cell proliferation in a dose- and time-dependent manner. The mechanism of growth inhibition was found to be cell cycle arrest in G1 phase. Thiocoraline activated the Notch pathway as demonstrated by the dose-dependent increase in mRNA and protein expression of Notch isoforms. Furthermore, treatment with thiocoraline resulted in changes in the expression of downstream targets of the Notch pathway (HES1, HES2, HES6, HEY1, and HEY2) and reduced expression of NET markers, CgA, and ASCL1. Thiocoraline is a potent Notch pathway activator and an inhibitor of MTC-TT cell proliferation at low nanomolar concentrations. These results provide exciting evidence for the use of thiocoraline as a potential treatment for intractable MTC. Thiocoraline is a potent Notch pathway activator and an inhibitor of medullary thyroid cancer cell line (MTC-TT) cell proliferation at low nanomolar concentrations. These results provide evidence for the use of thiocoraline as a potential treatment for intractable MTC. PMID:24403239

The role of p21Waf1/CIP1 as a Cip/Kip type cell-cycle regulator in oral squamous cell carcinoma (Review).

PubMed

Pérez-Sayáns, Mario; Suárez-Peñaranda, José-Manuel; Gayoso-Diz, Pilar; Barros-Angueira, Francisco; Gándara-Rey, José-Manuel; García-García, Abel

2013-03-01

Oral Squamous Cell Carcinoma (OSCC) is biologically characterized by the accumulation of multiple genetic and molecular alterations that end up clinically characterized as a malignant neoplasm through a phenomenon known as multistep. The members of the Cip/Kip family, specifically p21Waf1/CIP1, are responsible for cell cycle control, blocking the transition from phase G1 to phase S. We made a search of articles of peer-reviewed Journals in PubMed/ Medline, crossing the keywords. The goal of this paper is to determine the relationship between p21Waf1/CIP1 expression and several clinical and pathological aspects of OSCC, their relationship with p53 and HPV, as well as genetic alterations in their expression pattern, their use as a prognosis market in the evolution of precancerous lesions and their roles in anticancer treatments. The results of p21WAF1/CIP1 expression in OSCC showed mixed results in terms of positivity/negativity throughout different studies. It seems that, although p21Waf1/CIP1 expression is controlled in a p53-dependent manner, coexpression of both in OSCC is not intrinsically related. Although the presence of HPV viral oncoproteins increases p21Waf1/CIP1 levels, the small number of studies, have forced us to disregard the hypothesis that HPV infected lesions that present better prognosis are due to a p21Waf1/CIP1-dependent control. The role of p21WAF1/CIP1 as cell-cycle regulator has been well described; however, its relationship to OSCC, the clinical and pathological variables of tumors, HPV and different treatments are not entirely clear. Thus, it would be very interesting to pursue further study of this protein, which may have a significant value for the diagnosis, prognosis and therapy of this type of tumors.

In vitro studies of the antirhinovirus activity of soluble intercellular adhesion molecule-1.

PubMed Central

Arruda, E; Crump, C E; Marlin, S D; Merluzzi, V J; Hayden, F G

1992-01-01

We studied the in vitro antirhinovirus activity of a soluble form of intercellular adhesion molecule-1 (sICAM-1). sICAM-1 inhibited the cytopathic effect of 10 representative human rhinovirus (HRV) serotypes of the major receptor group with, 50% effective concentrations (EC50s) of 0.1 to 7.9 micrograms/ml. Cell type-dependent variation in the inhibitory activity of sICAM-1 was observed for two major receptor group serotypes in HeLa cells (EC50, greater than 32 micrograms/ml), and no inhibitory effect was observed for two serotypes which use different cell receptors. Yield reduction assays showed that sICAM-1 inhibited the replication of HRV serotype 39 (HRV-39) in human adenoid explants in a concentration-dependent manner. No direct inactivation of infectivity of HRV-39 (EC50, 0.5 microgram/ml) was observed after incubation with sICAM-1 (32 micrograms/ml) for up to 24 h. Single-cycle-of-replication experiments with the addition of sICAM-1 at 10 micrograms/ml at different times showed that the inhibitory effect occurs only when sICAM-1 is added within 30 min after infection. In experiments in which absorption was carried out at 4 degrees C and then a single cycle of replication incubation was carried out at 33 degrees C, it was found that sICAM-1 at 10 micrograms/ml was inhibitory only when it was present during the absorption period. Our data show that sICAM-1 is inhibitory for representative major receptor group serotypes of HRV in two cell lines and human respiratory epithelium, that the interaction of sICAM-1 with HRV is readily reversible by dilution, and that the inhibitory effect of sICAM-1 on virus replication is present early in the infection cycle. PMID:1358025

Amplified Self-replication of DNA Origami Nanostructures through Multi-cycle Fast-annealing Process

NASA Astrophysics Data System (ADS)

Zhou, Feng; Zhuo, Rebecca; He, Xiaojin; Sha, Ruojie; Seeman, Nadrian; Chaikin, Paul

We have developed a non-biological self-replication process using templated reversible association of components and irreversible linking with annealing and UV cycles. The current method requires a long annealing time, up to several days, to achieve the specific self-assembly of DNA nanostructures. In this work, we accomplished the self-replication with a shorter time and smaller replication rate per cycle. By decreasing the ramping time, we obtained the comparable replication yield within 90 min. Systematic studies show that the temperature and annealing time play essential roles in the self-replication process. In this manner, we can amplify the self-replication process to a factor of 20 by increasing the number of cycles within the same amount of time.

Aqueous extract of Tribulus terrestris Linn induces cell growth arrest and apoptosis by down-regulating NF-κB signaling in liver cancer cells.

PubMed

Kim, Hye Jin; Kim, Jin Chul; Min, Jung Sun; Kim, Mi-Jee; Kim, Ji Ae; Kor, Myung Ho; Yoo, Hwa Seung; Ahn, Jeong Keun

2011-06-14

A medicinal herb Tribulus terrestris Linn has been used to treat various diseases including hepatocellular carcinoma. The aim of the present study was to investigate the anticancer activity of Tribulus terrestris Linn (TT) in liver cancer cells. The antitumor activity of aqueous TT extract was analyzed by testing the cytotoxicity and the effect on clonogenecity in HepG2 cells. Apoptosis and cell cycle arrest induced by TT were dissected by flow cytometry and its inhibitory effect on NF-κB activity was determined by analyzing the expression levels of NF-κB/IκB subunit proteins. The suppression of NF-κB-regulated gene expression by TT was assessed by RT-PCR. TT extract repressed clonogenecity and proliferation, induced apoptosis, and enhanced accumulation in the G0/G1 phase of liver cancer cells. It also turned out that TT extract inhibited NF-κB-dependent reporter gene expression and NF-κB subunit p50 expression, while it enhanced the cellular level of IκBα by inhibiting the phosphorylation and degradation of IκBα. In addition, IKK activity was inhibited in a dose-dependent manner. Furthermore, TT extract suppressed the transcription of genes associated with cell cycle regulation, anti-apoptosis, and invasion. These data showed that TT extract blocks proliferation and induces apoptosis in human liver cancer cells through the inhibition of NF-κB signaling. Aqueous TT extract can be used as an anticancer drug for hepatocellular carcinoma patients. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Preventive effects of 125I seeds on benign restenosis following esophageal stent implantation in a dog model

PubMed Central

GAN, ZHEN; JING, JIAN; ZHU, GUANGYU; QIN, YONGLIN; TENG, GAOJUN; GUO, JINHE

2015-01-01

The present study aimed to evaluate the effects of iodine-125 (125I) seeds on the proliferation of primary esophageal fibroblasts in dogs, and to assess the safety and preventive efficacy of 125I seed-pre-loaded esophageal stents in benign restenosis following implantation. Primary fibroblasts were cultured with various 125I seed activities, which were then evaluated using cell proliferation and apoptosis assays as well as cell cycle analysis using Annexin V/propidium iodide (PI) double staining and PI staining. Prior to sacrification, animals were submitted to esophageal radiography under digital subtraction angiography. Esophageal tissues were collected and examined for macroscopic, microscopic and pathological alterations. The results demonstrated a significant and dose-dependent inhibition of fibroblast proliferation and increased apoptosis following exposure to 125I seeds. G0/G1 fibroblast populations increased in a dose-dependent manner following treatment with 125I seeds, in contrast to cells in S phase. Four weeks following implantation, α-smooth muscle actin and proliferating cell nuclear antigen expression levels in the experimental group were significantly lower compared with those in the control group; in addition, eight weeks following implantation, esophageal inner diameters were increased in the experimental group. 125I seeds inhibited proliferation of dog esophageal fibroblasts via cell cycle arrest and apoptosis. In conclusion, 125I seed-pre-loaded esophageal stents inhibited benign hyperplasia in the upper edge of the stent to a certain extent, which relieved benign restenosis following implantation with a good safety profile. PMID:25543838

The increase of cell-membranous phosphatidylcholines containing polyunsaturated fatty acid residues induces phosphorylation of p53 through activation of ATR

PubMed Central

Zhang, Xu Hannah; Zhao, Chunying; Ma, Zhongmin Alex

2010-01-01

Summary The G1 phase of the cell cycle is marked by the rapid turnover of phospholipids. This turnover is regulated by CTP:phosphocholine-cytidylyltransferase (CCT) and group VIA Ca2+-independent-phospholipase A2 (iPLA2). We previously reported that inhibition of iPLA2 arrests cells in G1 phase of the cell cycle by activating the p53-p21 checkpoint. Here we further characterize the mechanism of p53 activation. We show that specific inhibition of iPLA2 induces a time dependent phosphorylation of Ser15 in p53 in the absence of DNA damage. This phosphorylation requires the kinase ataxia-telangiectasia and Rad-3-related (ATR) but not the ataxia-telangiectasia-mutated (ATM) kinase. Moreover, we identify in cell membranes a significant increase of phosphatidylcholines (PCs) containing chains of polyunsaturated fatty acids and a decrease of PCs containing saturated fatty acids in response to inhibition of iPLA2. The time course of phosphorylation of Ser15 in p53 correlates with increasing levels of PCs containing polyunsaturated fatty acids. We further demonstrate that the PCs with linoleic acid in their sn-2 position (18:2n6) induce phosphorylation of Ser15 in p53 in an ATR-dependent manner. Our findings establish that cells can regulate the levels of polyunsaturated fatty acids in phospholipids through iPLA2-mediated deacylation of PCs. Disruption of this regulation increases the proportions of PCs containing polyunsaturated fatty acids and activates the ATR-p53 signalling pathway. PMID:18032786

The increase of cell-membranous phosphatidylcholines containing polyunsaturated fatty acid residues induces phosphorylation of p53 through activation of ATR.

PubMed

Zhang, Xu Hannah; Zhao, Chunying; Ma, Zhongmin Alex

2007-12-01

The G1 phase of the cell cycle is marked by the rapid turnover of phospholipids. This turnover is regulated by CTP:phosphocholine-cytidylyltransferase (CCT) and group VIA Ca(2+)-independent-phospholipase A(2) (iPLA(2)). We previously reported that inhibition of iPLA(2) arrests cells in G1 phase of the cell cycle by activating the p53-p21 checkpoint. Here we further characterize the mechanism of p53 activation. We show that specific inhibition of iPLA(2) induces a time dependent phosphorylation of Ser15 in p53 in the absence of DNA damage. This phosphorylation requires the kinase ataxia-telangiectasia and Rad-3-related (ATR) but not the ataxia-telangiectasia-mutated (ATM) kinase. Moreover, we identify in cell membranes a significant increase of phosphatidylcholines (PCs) containing chains of polyunsaturated fatty acids and a decrease of PCs containing saturated fatty acids in response to inhibition of iPLA(2). The time course of phosphorylation of Ser15 in p53 correlates with increasing levels of PCs containing polyunsaturated fatty acids. We further demonstrate that the PCs with linoleic acid in their sn-2 position (18:2n6) induce phosphorylation of Ser15 in p53 in an ATR-dependent manner. Our findings establish that cells can regulate the levels of polyunsaturated fatty acids in phospholipids through iPLA(2)-mediated deacylation of PCs. Disruption of this regulation increases the proportions of PCs containing polyunsaturated fatty acids and activates the ATR-p53 signalling pathway.

Effect of lithium chloride and antineoplastic drugs on survival and cell cycle of androgen-dependent prostate cancer LNCap cells

PubMed Central

Azimian-Zavareh, Vajihe; Hossein, Ghamartaj; Janzamin, Ehsan

2012-01-01

Objective: Glycogen synthase kinase-3β (GSK-3β) has been reported to be required for androgen receptor (AR) activity. This study sought to determine the usefulness of lithium chloride (LiCl) as a highly selective inhibitor of GSK-3β to increase the sensitivity of LNCap cells to doxorubicin (Dox), etoposide (Eto), and vinblastine (Vin) drugs. Materials and Methods: Thiazolyl Blue Tetrazolium Blue (MTT) assay was used to determine the cytotoxic effect to LiCl alone or in combination with low dose and IC50 doses of drugs. Subsequently, cell cycle analysis was performed by using flow cytometry. Results: LiCl showed cytotoxic effect in a dose- and time-dependent manner (P<0.001). Both Dox (100 or 280 nM) and Vin IC50 (5 nM) doses caused G2/M-phase arrest (P<0.001) compared with control. However, low dose (10 μM) or IC50 (70 μM) Eto doses showed G2/M or S-phase arrests, respectively (P<0.001). Combination of low dose or IC50 dose of Eto with LiCl showed increased apoptosis as revealed by high percent of cells in SubG1 (P<0.05, P<0.01, respectively). Moreover, Eto (10 μM) led to decreased percent of cells in G2/M phase when combined with LiCl (P<0.05). Conclusion: This study showed that LiCl increases apoptosis of (LNCap) Lymph Node Carcinoma of the Prostate cells in the presence of Eto, which is S- and G2-phase-specific drug. PMID:23248400

An Extended Eddy-Diffusivity Mass-Flux Scheme for Unified Representation of Subgrid-Scale Turbulence and Convection

NASA Astrophysics Data System (ADS)

Tan, Zhihong; Kaul, Colleen M.; Pressel, Kyle G.; Cohen, Yair; Schneider, Tapio; Teixeira, João.

2018-03-01

Large-scale weather forecasting and climate models are beginning to reach horizontal resolutions of kilometers, at which common assumptions made in existing parameterization schemes of subgrid-scale turbulence and convection—such as that they adjust instantaneously to changes in resolved-scale dynamics—cease to be justifiable. Additionally, the common practice of representing boundary-layer turbulence, shallow convection, and deep convection by discontinuously different parameterizations schemes, each with its own set of parameters, has contributed to the proliferation of adjustable parameters in large-scale models. Here we lay the theoretical foundations for an extended eddy-diffusivity mass-flux (EDMF) scheme that has explicit time-dependence and memory of subgrid-scale variables and is designed to represent all subgrid-scale turbulence and convection, from boundary layer dynamics to deep convection, in a unified manner. Coherent up and downdrafts in the scheme are represented as prognostic plumes that interact with their environment and potentially with each other through entrainment and detrainment. The more isotropic turbulence in their environment is represented through diffusive fluxes, with diffusivities obtained from a turbulence kinetic energy budget that consistently partitions turbulence kinetic energy between plumes and environment. The cross-sectional area of up and downdrafts satisfies a prognostic continuity equation, which allows the plumes to cover variable and arbitrarily large fractions of a large-scale grid box and to have life cycles governed by their own internal dynamics. Relatively simple preliminary proposals for closure parameters are presented and are shown to lead to a successful simulation of shallow convection, including a time-dependent life cycle.

Therapeutic treatments potentially mediated by melatonin receptors: potential clinical uses in the prevention of osteoporosis, cancer and as an adjuvant therapy.

PubMed

Witt-Enderby, Paula A; Radio, Nicholas M; Doctor, John S; Davis, Vicki L

2006-11-01

Melatonin's therapeutic potential is grossly underestimated because its functional roles are diverse and its mechanism(s) of action are complex and varied. Melatonin produces cellular effects via a variety of mechanisms in a receptor independent and dependent manner. In addition, melatonin is a chronobiotic agent secreted from the pineal gland during the hours of darkness. This diurnal release of melatonin impacts the sensitivity of melatonin receptors throughout a 24-hr period. This changing sensitivity probably contributes to the narrow therapeutic window for use of melatonin in treating sleep disorders, that is, at the light-to-dark (dusk) or dark-to-light (dawn) transition states. In addition to the cyclic changes in melatonin receptors, many genes cycle over the 24-hr period, independent or dependent upon the light/dark cycle. Interestingly, many of these genes support a role for melatonin in modulating metabolic and cardiovascular physiology as well as bone metabolism and immune function and detoxification of chemical agents and cancer reduction. Melatonin also enhances the actions of a variety of drugs or hormones; however, the role of melatonin receptors in modulating these processes is not known. The goal of this review is to summarize the evidence related to the utility of melatonin as a therapeutic agent by focusing on its other potential uses besides sleep disorders. In particular, its use in cancer prevention, osteoporosis and, as an adjuvant to other therapies are discussed. Also, the role that melatonin and, particularly, its receptors play in these processes are highlighted.

Hydralazine and Organic Nitrates Restore Impaired Excitation-Contraction Coupling by Reducing Calcium Leak Associated with Nitroso-Redox Imbalance*

PubMed Central

Dulce, Raul A.; Yiginer, Omer; Gonzalez, Daniel R.; Goss, Garrett; Feng, Ning; Zheng, Meizi; Hare, Joshua M.

2013-01-01

Although the combined use of hydralazine and isosorbide dinitrate confers important clinical benefits in patients with heart failure, the underlying mechanism of action is still controversial. We used two models of nitroso-redox imbalance, neuronal NO synthase-deficient (NOS1−/−) mice and spontaneously hypertensive heart failure rats, to test the hypothesis that hydralazine (HYD) alone or in combination with nitroglycerin (NTG) or isosorbide dinitrate restores Ca2+ cycling and contractile performance and controls superoxide production in isolated cardiomyocytes. The response to increased pacing frequency was depressed in NOS1−/− compared with wild type myocytes. Both sarcomere length shortening and intracellular Ca2+ transient (Δ[Ca2+]i) responses in NOS1−/− cardiomyocytes were augmented by HYD in a dose-dependent manner. NTG alone did not affect myocyte shortening but reduced Δ[Ca2+]i across the range of pacing frequencies and increased myofilament Ca2+ sensitivity thereby enhancing contractile efficiency. Similar results were seen in failing myocytes from the heart failure rat model. HYD alone or in combination with NTG reduced sarcoplasmic reticulum (SR) leak, improved SR Ca2+ reuptake, and restored SR Ca2+ content. HYD and NTG at low concentrations (1 μm), scavenged superoxide in isolated cardiomyocytes, whereas in cardiac homogenates, NTG inhibited xanthine oxidoreductase activity and scavenged NADPH oxidase-dependent superoxide more efficiently than HYD. Together, these results revealed that by reducing SR Ca2+ leak, HYD improves Ca2+ cycling and contractility impaired by nitroso-redox imbalance, and NTG enhanced contractile efficiency, restoring cardiac excitation-contraction coupling. PMID:23319593

Piperlongumine decreases cell proliferation and the expression of cell cycle-associated proteins by inhibiting Akt pathway in human lung cancer cells.

PubMed

Seok, Jin Sil; Jeong, Chang Hee; Petriello, Michael C; Seo, Han Geuk; Yoo, Hyunjin; Hong, Kwonho; Han, Sung Gu

2018-01-01

Piperlongumine (PL) is an alkaloid of a pepper plant found in Southeast Asia. PL is known to induce selective toxicity towards a variety of cancer cell types. To explore the possible anti-lung cancer effects of PL, A549 cells were treated with PL (0-40 μM) for 24 h. Alterations in the expression of cell cycle-associated proteins (cyclin D1, cyclin-dependent kinase 4 (CDK4), CDK6 and retinoblastoma (Rb)) and intracellular signaling molecules (extracellular signal receptor-activated kinase 1/2 (ERK1/2), Akt, p38 and nuclear factor-κB (NF-κB)) were examined in cells following treatment of PL using Western blot analysis. Results showed that proliferation of cells were significantly decreased by PL in a dose-dependent manner. Flow cytometry results demonstrated increased number of cells in G1 phase in PL (40 μM)-treated group. Reactive oxygen species was significantly increased in cells treated with PL at 20-40 μM. The expression of cyclin D1, CDK4, CDK6 and p-Rb were markedly decreased in cells treated with PL at 40 μM. Treatment of cells with PL suppressed phosphorylation of Akt but increased ERK1/2 phosphorylation. Treatment of PL significantly decreased nuclear translocation of NF-κB p65 in cells. These results suggest that PL possesses antiproliferative properties in A549 cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

Regulation of p53 Target Gene Expression by Peptidylarginine Deiminase 4 ▿ †

PubMed Central

Li, Pingxin; Yao, Hongjie; Zhang, Zhiqiang; Li, Ming; Luo, Yuan; Thompson, Paul R.; Gilmour, David S.; Wang, Yanming

2008-01-01

Histone Arg methylation has been correlated with transcriptional activation of p53 target genes. However, whether this modification is reversed to repress the expression of p53 target genes is unclear. Here, we report that peptidylarginine deiminase 4, a histone citrullination enzyme, is involved in the repression of p53 target genes. Inhibition or depletion of PAD4 elevated the expression of a subset of p53 target genes, including p21/CIP1/WAF1, leading to cell cycle arrest and apoptosis. Moreover, the induction of p21, cell cycle arrest, and apoptosis by PAD4 depletion is p53 dependent. Protein-protein interaction studies showed an interaction between p53 and PAD4. Chromatin immunoprecipitation assays showed that PAD4 is recruited to the p21 promoter in a p53-dependent manner. RNA polymerase II (Pol II) activities and the association of PAD4 are dynamically regulated at the p21 promoter during UV irradiation. Paused RNA Pol II and high levels of PAD4 were detected before UV treatment. At early time points after UV treatment, an increase of histone Arg methylation and a decrease of citrullination were correlated with a transient activation of p21. At later times after UV irradiation, a loss of RNA Pol II and an increase of PAD4 were detected at the p21 promoter. The dynamics of RNA Pol II activities after UV treatment were further corroborated by permanganate footprinting. Together, these results suggest a role of PAD4 in the regulation of p53 target gene expression. PMID:18505818

CP-31398 inhibits the growth of p53-mutated liver cancer cells in vitro and in vivo.

PubMed

He, Xing-Xing; Zhang, Yu-Nan; Yan, Jun-Wei; Yan, Jing-Jun; Wu, Qian; Song, Yu-Hu

2016-01-01

The tumor suppressor p53 is one of the most frequently mutated genes in hepatocellular carcinoma (HCC). Previous studies demonstrated that CP-31398 restored the native conformation of mutant p53 and trans-activated p53 downstream genes in tumor cells. However, the research on the application of CP-31398 to liver cancer has not been reported. Here, we investigated the effects of CP-31398 on the phenotype of HCC cells carrying p53 mutation. The effects of CP-31398 on the characteristic of p53-mutated HCC cells were evaluated through analyzing cell cycle, cell apoptosis, cell proliferation, and the expression of p53 downstream genes. In tumor xenografts developed by PLC/PRF/5 cells, the inhibition of tumor growth by CP-31398 was analyzed through gross morphology, growth curve, and the expression of p53-related genes. Firstly, we demonstrated that CP-31398 inhibited the growth of p53-mutated liver cancer cells in a dose-dependent and p53-dependent manner. Then, further study showed that CP-31398 re-activated wild-type p53 function in p53-mutated HCC cells, which resulted in inhibitive response of cell proliferation and an induction of cell-cycle arrest and apoptosis. Finally, in vivo data confirmed that CP-31398 blocked the growth of xenografts tumors through transactivation of p53-responsive downstream molecules. Our results demonstrated that CP-31398 induced desired phenotypic change of p53-mutated HCC cells in vitro and in vivo, which revealed that CP-31398 would be developed as a therapeutic candidate for HCC carrying p53 mutation.

6-Gingerol Inhibits Growth of Colon Cancer Cell LoVo via Induction of G2/M Arrest

PubMed Central

Lin, Ching-Bin; Lin, Chun-Che; Tsay, Gregory J.

2012-01-01

6-Gingerol, a natural component of ginger, has been widely reported to possess antiinflammatory and antitumorigenic activities. Despite its potential efficacy against cancer, the anti-tumor mechanisms of 6-gingerol are complicated and remain sketchy. In the present study, we aimed to investigate the anti-tumor effects of 6-gingerol on colon cancer cells. Our results revealed that 6-gingerol treatment significantly reduced the cell viability of human colon cancer cell, LoVo, in a dose-dependent manner. Further flow cytometric analysis showed that 6-gingerol induced significant G2/M phase arrest and had slight influence on sub-G1 phase in LoVo cells. Therefore, levels of cyclins, cyclin-dependent kinases (CDKs), and their regulatory proteins involved in S-G2/M transition were investigated. Our findings revealed that levels of cyclin A, cyclin B1, and CDK1 were diminished; in contrast, levels of the negative cell cycle regulators p27Kip1 and p21Cip1 were increased in response to 6-gingerol treatment. In addition, 6-gingerol treatment elevated intracellular reactive oxygen species (ROS) and phosphorylation level of p53. These findings indicate that exposure of 6-gingerol may induce intracellular ROS and upregulate p53, p27Kip1, and p21Cip1 levels leading to consequent decrease of CDK1, cyclin A, and cyclin B1 as result of cell cycle arrest in LoVo cells. It would be suggested that 6-gingerol should be beneficial to treatment of colon cancer. PMID:22719783

An Extended Eddy‐Diffusivity Mass‐Flux Scheme for Unified Representation of Subgrid‐Scale Turbulence and Convection

PubMed Central

Tan, Zhihong; Kaul, Colleen M.; Pressel, Kyle G.; Cohen, Yair; Teixeira, João

2018-01-01

Abstract Large‐scale weather forecasting and climate models are beginning to reach horizontal resolutions of kilometers, at which common assumptions made in existing parameterization schemes of subgrid‐scale turbulence and convection—such as that they adjust instantaneously to changes in resolved‐scale dynamics—cease to be justifiable. Additionally, the common practice of representing boundary‐layer turbulence, shallow convection, and deep convection by discontinuously different parameterizations schemes, each with its own set of parameters, has contributed to the proliferation of adjustable parameters in large‐scale models. Here we lay the theoretical foundations for an extended eddy‐diffusivity mass‐flux (EDMF) scheme that has explicit time‐dependence and memory of subgrid‐scale variables and is designed to represent all subgrid‐scale turbulence and convection, from boundary layer dynamics to deep convection, in a unified manner. Coherent up and downdrafts in the scheme are represented as prognostic plumes that interact with their environment and potentially with each other through entrainment and detrainment. The more isotropic turbulence in their environment is represented through diffusive fluxes, with diffusivities obtained from a turbulence kinetic energy budget that consistently partitions turbulence kinetic energy between plumes and environment. The cross‐sectional area of up and downdrafts satisfies a prognostic continuity equation, which allows the plumes to cover variable and arbitrarily large fractions of a large‐scale grid box and to have life cycles governed by their own internal dynamics. Relatively simple preliminary proposals for closure parameters are presented and are shown to lead to a successful simulation of shallow convection, including a time‐dependent life cycle. PMID:29780442

Cnidium officinale Makino extract induces apoptosis through activation of caspase-3 and p53 in human liver cancer HepG2 cells

PubMed Central

Hong, Heeok; An, Jeong Cheol; de La Cruz, Joseph F.; Hwang, Seong-Gu

2017-01-01

A number of diverse studies have reported the anticancer properties of Cnidium officinale Makino (CO). However, the apoptotic effect of this traditional medicinal herb in human hepatocellular carcinoma cells (HepG2) remains to be elucidated. Therefore, the present study investigated the ability of CO to reduce cell viability through apoptotic pathways. Cell viability was determined using the 2,3-bis [2-methyloxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide assay. CO extract-induced apoptosis in HepG2 cells was assessed by Hoechst 33258 staining. The cell cycle was monitored using fluorescence-activated cell sorting analysis with propidium iodide staining. Furthermore, the present study explored whether various signaling molecules associated with HepG2 cell death were affected by CO treatment, including caspase-3, B-cell lymphoma 2 (Bcl-2), tumor protein p53 (p53), cyclin-dependent kinase 4 (CDK4) and cyclin D. The expression levels of these genes were examined by reverse-transcription polymerase chain reaction and western blotting. The expression levels of caspase-3 and p53 were upregulated with CO extract treatment, whereas those of Bcl-2, CDK4 and cyclin D were significantly downregulated. Cleaved caspase-3 expression was upregulated following treatment with CO extract in a dose-dependent manner. Collectively, the data suggest that CO extract has the potential to induce apoptosis of HepG2 cells and may act by suppressing the cell cycle, which leads to caspase-3 cleavage and p53 signaling. PMID:28966688

6-Gingerol Inhibits Growth of Colon Cancer Cell LoVo via Induction of G2/M Arrest.

PubMed

Lin, Ching-Bin; Lin, Chun-Che; Tsay, Gregory J

2012-01-01

6-Gingerol, a natural component of ginger, has been widely reported to possess antiinflammatory and antitumorigenic activities. Despite its potential efficacy against cancer, the anti-tumor mechanisms of 6-gingerol are complicated and remain sketchy. In the present study, we aimed to investigate the anti-tumor effects of 6-gingerol on colon cancer cells. Our results revealed that 6-gingerol treatment significantly reduced the cell viability of human colon cancer cell, LoVo, in a dose-dependent manner. Further flow cytometric analysis showed that 6-gingerol induced significant G2/M phase arrest and had slight influence on sub-G1 phase in LoVo cells. Therefore, levels of cyclins, cyclin-dependent kinases (CDKs), and their regulatory proteins involved in S-G2/M transition were investigated. Our findings revealed that levels of cyclin A, cyclin B1, and CDK1 were diminished; in contrast, levels of the negative cell cycle regulators p27(Kip1) and p21(Cip1) were increased in response to 6-gingerol treatment. In addition, 6-gingerol treatment elevated intracellular reactive oxygen species (ROS) and phosphorylation level of p53. These findings indicate that exposure of 6-gingerol may induce intracellular ROS and upregulate p53, p27(Kip1), and p21(Cip1) levels leading to consequent decrease of CDK1, cyclin A, and cyclin B1 as result of cell cycle arrest in LoVo cells. It would be suggested that 6-gingerol should be beneficial to treatment of colon cancer.

Apoptosis Induction by Polygonum minus is related to antioxidant capacity, alterations in expression of apoptotic-related genes, and S-phase cell cycle arrest in HepG2 cell line.

PubMed

Mohd Ghazali, Mohd Alfazari; Al-Naqeb, Ghanya; Krishnan Selvarajan, Kesavanarayanan; Hazizul Hasan, Mizaton; Adam, Aishah

2014-01-01

Polygonum minus (Polygonaceae) is a medicinal herb distributed throughout eastern Asia. The present study investigated antiproliferative effect of P. minus and its possible mechanisms. Four extracts (petroleum ether, methanol, ethyl acetate, and water) were prepared by cold maceration. Extracts were subjected to phytochemical screening, antioxidant, and antiproliferative assays; the most bioactive was fractionated using vacuum liquid chromatography into seven fractions (F1-F7). Antioxidant activity was measured via total phenolic content (TPC), 2,2-diphenyl-1-picrylhydrazyl (DPPH), and ferric reducing antioxidant power (FRAP) assays. Antiproliferative activity was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Most active fraction was tested for apoptosis induction and cell cycle arrest in HepG2 cells using flow cytometry and confocal microscopy. Apoptotic-related gene expression was studied by RT-PCR. Ethyl acetate extract was bioactive in initial assays. Its fraction, F7, exhibited highest antioxidant capacity (TPC; 113.16 ± 6.2 mg GAE/g extract, DPPH; EC50: 30.5 ± 3.2 μg/mL, FRAP; 1169 ± 20.3 μmol Fe (II)/mg extract) and selective antiproliferative effect (IC50: 25.75 ± 1.5 μg/mL). F7 induced apoptosis in concentration- and time-dependent manner and caused cell cycle arrest at S-phase. Upregulation of proapoptotic genes (Bax, p53, and caspase-3) and downregulation of antiapoptotic gene, Bcl-2, were observed. In conclusion, F7 was antiproliferative to HepG2 cells by inducing apoptosis, cell cycle arrest, and via antioxidative effects.

Apoptosis Induction by Polygonum minus Is Related to Antioxidant Capacity, Alterations in Expression of Apoptotic-Related Genes, and S-Phase Cell Cycle Arrest in HepG2 Cell Line

PubMed Central

Mohd Ghazali, Mohd Alfazari; Al-Naqeb, Ghanya; Krishnan Selvarajan, Kesavanarayanan; Hazizul Hasan, Mizaton; Adam, Aishah

2014-01-01

Polygonum minus (Polygonaceae) is a medicinal herb distributed throughout eastern Asia. The present study investigated antiproliferative effect of P. minus and its possible mechanisms. Four extracts (petroleum ether, methanol, ethyl acetate, and water) were prepared by cold maceration. Extracts were subjected to phytochemical screening, antioxidant, and antiproliferative assays; the most bioactive was fractionated using vacuum liquid chromatography into seven fractions (F1–F7). Antioxidant activity was measured via total phenolic content (TPC), 2,2-diphenyl-1-picrylhydrazyl (DPPH), and ferric reducing antioxidant power (FRAP) assays. Antiproliferative activity was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Most active fraction was tested for apoptosis induction and cell cycle arrest in HepG2 cells using flow cytometry and confocal microscopy. Apoptotic-related gene expression was studied by RT-PCR. Ethyl acetate extract was bioactive in initial assays. Its fraction, F7, exhibited highest antioxidant capacity (TPC; 113.16 ± 6.2 mg GAE/g extract, DPPH; EC50: 30.5 ± 3.2 μg/mL, FRAP; 1169 ± 20.3 μmol Fe (II)/mg extract) and selective antiproliferative effect (IC50: 25.75 ± 1.5 μg/mL). F7 induced apoptosis in concentration- and time-dependent manner and caused cell cycle arrest at S-phase. Upregulation of proapoptotic genes (Bax, p53, and caspase-3) and downregulation of antiapoptotic gene, Bcl-2, were observed. In conclusion, F7 was antiproliferative to HepG2 cells by inducing apoptosis, cell cycle arrest, and via antioxidative effects. PMID:24955361

Role of insulin-like growth factor-1 (IGF-1) in regulating cell cycle progression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ma, Qi-lin; Yang, Tian-lun; Yin, Ji-ye

2009-11-06

Aims: Insulin-like growth factor-1 (IGF-1) is a polypeptide protein hormone, similar in molecular structure to insulin, which plays an important role in cell migration, cell cycle progression, cell survival and proliferation. In this study, we investigated the possible mechanisms of IGF-1 mediated cell cycle redistribution and apoptosis of vascular endothelial cells. Method: Human umbilical vein endothelial cells (HUVECs) were pretreated with 0.1, 0.5, or 2.5 {mu}g/mL of IGF-1 for 30 min before the addition of Ang II. Cell cycle redistribution and apoptosis were examined by flow cytometry. Expression of Ang II type 1 (AT{sub 1}) mRNA and cyclin E proteinmore » were determined by RT-PCR and Western blot, respectively. Results: Ang II (1 {mu}mol/L) induced HUVECs arrested at G{sub 0}/G{sub 1}, enhanced the expression level of AT{sub 1} mRNA in a time-dependent manner, reduced the enzymatic activity of nitric oxide synthase (NOS) and nitric oxide (NO) content as well as the expression level of cyclin E protein. However, IGF-1 enhanced NOS activity, NO content, and the expression level of cyclin E protein, and reduced the expression level of AT{sub 1} mRNA. L-NAME significantly counteracted these effects of IGF-1. Conclusions: Our data suggests that IGF-1 can reverse vascular endothelial cells arrested at G{sub 0}/G{sub 1} and apoptosis induced by Ang II, which might be mediated via a NOS-NO signaling pathway and is likely associated with the expression levels of AT1 mRNA and cyclin E proteins.« less

Epidermal Growth Factor Receptor activation promotes ADA3 acetylation through the AKT-p300 pathway

PubMed Central

Srivastava, Shashank; Mohibi, Shakur; Mirza, Sameer; Band, Hamid; Band, Vimla

2017-01-01

ABSTRACT The ADA3 (Alteration/Deficiency in Activation 3) protein is an essential adaptor component of several Lysine Acetyltransferase (KAT) complexes involved in chromatin modifications. Previously, we and others have demonstrated a crucial role of ADA3 in cell cycle progression and in maintenance of genomic stability. Recently, we have shown that acetylation of ADA3 is key to its role in cell cycle progression. Here, we demonstrate that AKT activation downstream of Epidermal Growth Factor Receptor (EGFR) family proteins stimulation leads to phosphorylation of p300, which in turn promotes the acetylation of ADA3. Inhibition of upstream receptor tyrosine kinases (RTKs), HER1 (EGFR)/HER2 by lapatinib and the accompanying reduction of phospho-AKT levels led to a decrease in p300 phosphorylation and ADA3 protein levels. The p300/PCAF inhibitor garcinol also destabilized the ADA3 protein in a proteasome-dependent manner and an ADA3 mutant with K→R mutations exhibited a marked increase in half-life, consistent with opposite role of acetylation and ubiquitination of ADA3 on shared lysine residues. ADA3 knockdown led to cell cycle inhibitory effects, as well as apoptosis similar to those induced by lapatinib treatment of HER2+ breast cancer cells, as seen by accumulation of CDK inhibitor p27, reduction in mitotic marker pH3(S10), and a decrease in the S-phase marker PCNA, as well as the appearance of cleaved PARP. Taken together our results reveal a novel RTK-AKT-p300-ADA3 signaling pathway involved in growth factor-induced cell cycle progression. PMID:28759294

Mechanism of gemcitabine-induced suppression of human cholangiocellular carcinoma cell growth.

PubMed

Toyota, Yuka; Iwama, Hisakazu; Kato, Kiyohito; Tani, Joji; Katsura, Akiko; Miyata, Miwa; Fujiwara, Shintaro; Fujita, Koji; Sakamoto, Teppei; Fujimori, Takayuki; Okura, Ryoichi; Kobayashi, Kiyoyuki; Tadokoro, Tomoko; Mimura, Shima; Nomura, Takako; Miyoshi, Hisaaki; Morishita, Asahiro; Kamada, Hideki; Yoneyama, Hirohito; Okano, Keiichi; Suzuki, Yasuyuki; Masaki, Tsutomu

2015-10-01

Although gemcitabine (2',2'-difluorocytidine monohydrochloride) is a common anticancer agent of cholangiocellular carcinoma (CCC), its growth inhibitory effects and gemcitabine resistance in CCC cells are poorly understood. Our aims were to uncover the mechanism underlying the antitumor effect of gemcitabine and to analyze the mechanism regulating in vitro CCC cell gemcitabine resistance. In addition, we sought to identify miRNAs associated with the antitumor effects of gemcitabine in CCCs. Using a cell proliferation assay and flow cytometry, we examined the ability of gemcitabine to inhibit cell proliferation in three types of human CCC cell lines (HuCCT-1, Huh28, TKKK). We also employed western blotting to investigate the effects of gemcitabine on cell cycle-related molecules in CCC cells. In addition, we used array chips to assess gemcitabine-mediated changes in angiogenic molecules and activated tyrosine kinase receptors in CCC cells. We used miRNA array chips to comprehensively analyze gemcitabine-induced miRNAs and examined clusters of differentially expressed miRNAs in cells with and without gemcitabine treatment. Gemcitabine inhibited cell proliferation in a dose- and time-dependent manner in HuCCT-1 cells, whereas cell proliferation was unchanged in Huh28 and TKKK cells. Gemcitabine inhibited cell cycle progression in HuCCT-1 cells from G0/G1 to S phase, resulting in G1 cell cycle arrest due to the reduction of cyclin D1 expression. In addition, gemcitabine upregulated the angiogenic molecules IL-6, IL-8, ENA-78 and MCP-1. In TKKK cells, by contrast, gemcitabine did not arrest the cell cycle or modify angiogenic molecules. Furthermore, in gemcitabine-sensitive HuCCT-1 cells, gemcitabine markedly altered miRNA expression. The miRNAs and angiogenic molecules altered by gemcitabine contribute to the inhibition of tumor growth in vitro.

Jellyfish extract induces apoptotic cell death through the p38 pathway and cell cycle arrest in chronic myelogenous leukemia K562 cells

PubMed Central

Kwak, Choong-Hwan; Abekura, Fukushi; Park, Jun-Young; Park, Nam Gyu; Chang, Young-Chae; Lee, Young-Choon; Chung, Tae-Wook; Ha, Ki-Tae; Son, Jong-Keun

2017-01-01

Jellyfish species are widely distributed in the world’s oceans, and their population is rapidly increasing. Jellyfish extracts have several biological functions, such as cytotoxic, anti-microbial, and antioxidant activities in cells and organisms. However, the anti-cancer effect of Jellyfish extract has not yet been examined. We used chronic myelogenous leukemia K562 cells to evaluate the mechanisms of anti-cancer activity of hexane extracts from Nomura’s jellyfish in vitro. In this study, jellyfish are subjected to hexane extraction, and the extract is shown to have an anticancer effect on chronic myelogenous leukemia K562 cells. Interestingly, the present results show that jellyfish hexane extract (Jellyfish-HE) induces apoptosis in a dose- and time-dependent manner. To identify the mechanism(s) underlying Jellyfish-HE-induced apoptosis in K562 cells, we examined the effects of Jellyfish-HE on activation of caspase and mitogen-activated protein kinases (MAPKs), which are responsible for cell cycle progression. Induction of apoptosis by Jellyfish-HE occurred through the activation of caspases-3,-8 and -9 and phosphorylation of p38. Jellyfish-HE-induced apoptosis was blocked by a caspase inhibitor, Z-VAD. Moreover, during apoptosis in K562 cells, p38 MAPK was inhibited by pretreatment with SB203580, an inhibitor of p38. SB203580 blocked jellyfish-HE-induced apoptosis. Additionally, Jellyfish-HE markedly arrests the cell cycle in the G0/G1 phase. Therefore, taken together, the results imply that the anti-cancer activity of Jellyfish-HE may be mediated apoptosis by induction of caspases and activation of MAPK, especially phosphorylation of p38, and cell cycle arrest at the Go/G1 phase in K562 cells. PMID:28133573

Effects of different doses of nandrolone decanoate on estrous cycle and ovarian tissue of rats after treatment and recovery periods.

PubMed

Simão, Vinícius Augusto; Berloffa Belardin, Larissa; Araújo Leite, Gabriel Adan; de Almeida Chuffa, Luiz Gustavo; Camargo, Isabel Cristina Cherici

2015-10-01

This study tested the hypothesis that different doses of nandrolone decanoate (ND) will cause changes in the estrous cycle and ovarian tissue of adult rats; and investigated the duration of the recovery period that is sufficient to restore the damage in the animals treated with different doses. Wistar rats were treated with ND at doses of 1.87, 3.75, 7.5 and 15 mg/kg body weight, or received mineral oil (control group) for 15 days, subcutaneously. All animals were divided into three groups according to the treatment periods: (i) ND treatment for 15 days; (ii) ND treatment followed by a 30-day recovery; and (iii) ND treatment followed by a 60-day recovery. Estrous cycle was monitored daily, and at the end of each period, the animals were euthanized for histopathological analysis. During ND treatment and after 30-day recovery, all animals exhibited persistent diestrus. After a 60-day recovery, persistent diestrus was only maintained in the group that had received the highest dose. Ovarian weight was decreased significantly after the 30-day recovery, regardless of ND doses, compared with the control group. There was a reduction (P < 0.05) in the number of corpora lutea and antral and growing follicles, in contrast to an increase (P < 0.05) in atretic follicles in a dose- and time-dependent manner. Remarkable histopathological changes occurred in the ovaries of all ND-treated groups. In conclusion, the different doses of ND caused changes in the estrous cycle and ovarian tissue of rats, and recovery periods (30 and 60 days) were insufficient to completely restore the damage in the animals treated with the highest dose. © 2015 The Authors. International Journal of Experimental Pathology © 2015 International Journal of Experimental Pathology.

The Missing Link in Epstein-Barr Virus Immune Evasion: the BDLF3 Gene Induces Ubiquitination and Downregulation of Major Histocompatibility Complex Class I (MHC-I) and MHC-II

PubMed Central

Quinn, Laura L.; Williams, Luke R.; White, Claire; Forrest, Calum; Rowe, Martin

2015-01-01

ABSTRACT The ability of Epstein-Barr virus (EBV) to spread and persist in human populations relies on a balance between host immune responses and EBV immune evasion. CD8+ cells specific for EBV late lytic cycle antigens show poor recognition of target cells compared to immediate early and early antigen-specific CD8+ cells. This phenomenon is due in part to the early EBV protein BILF1, whose immunosuppressive activity increases with lytic cycle progression. However, published data suggest the existence of a hitherto unidentified immune evasion protein further enhancing protection against late EBV antigen-specific CD8+ cells. We have now identified the late lytic BDLF3 gene as the missing link accounting for efficient evasion during the late lytic cycle. Interestingly, BDLF3 also contributes to evasion of CD4+ cell responses to EBV. We report that BDLF3 downregulates expression of surface major histocompatibility complex (MHC) class I and class II molecules in the absence of any effect upon other surface molecules screened, including CD54 (ICAM-1) and CD71 (transferrin receptor). BDLF3 both enhanced internalization of surface MHC molecules and reduced the rate of their appearance at the cell surface. The reduced expression of surface MHC molecules correlated with functional protection against CD8+ and CD4+ T cell recognition. The molecular mechanism was identified as BDLF3-induced ubiquitination of MHC molecules and their subsequent downregulation in a proteasome-dependent manner. IMPORTANCE Immune evasion is a necessary feature of viruses that establish lifelong persistent infections in the face of strong immune responses. EBV is an important human pathogen whose immune evasion mechanisms are only partly understood. Of the EBV immune evasion mechanisms identified to date, none could explain why CD8+ T cell responses to late lytic cycle genes are so infrequent and, when present, recognize lytically infected target cells so poorly relative to CD8+ T cells specific for early lytic cycle antigens. The present work identifies an additional immune evasion protein, BDLF3, that is expressed late in the lytic cycle and impairs CD8+ T cell recognition by targeting cell surface MHC class I molecules for ubiquitination and proteasome-dependent downregulation. Interestingly, BDLF3 also targets MHC class II molecules to impair CD4+ T cell recognition. BDLF3 is therefore a rare example of a viral protein that impairs both the MHC class I and class II antigen-presenting pathways. PMID:26468525

The Missing Link in Epstein-Barr Virus Immune Evasion: the BDLF3 Gene Induces Ubiquitination and Downregulation of Major Histocompatibility Complex Class I (MHC-I) and MHC-II.

PubMed

Quinn, Laura L; Williams, Luke R; White, Claire; Forrest, Calum; Zuo, Jianmin; Rowe, Martin

2016-01-01

The ability of Epstein-Barr virus (EBV) to spread and persist in human populations relies on a balance between host immune responses and EBV immune evasion. CD8(+) cells specific for EBV late lytic cycle antigens show poor recognition of target cells compared to immediate early and early antigen-specific CD8(+) cells. This phenomenon is due in part to the early EBV protein BILF1, whose immunosuppressive activity increases with lytic cycle progression. However, published data suggest the existence of a hitherto unidentified immune evasion protein further enhancing protection against late EBV antigen-specific CD8(+) cells. We have now identified the late lytic BDLF3 gene as the missing link accounting for efficient evasion during the late lytic cycle. Interestingly, BDLF3 also contributes to evasion of CD4(+) cell responses to EBV. We report that BDLF3 downregulates expression of surface major histocompatibility complex (MHC) class I and class II molecules in the absence of any effect upon other surface molecules screened, including CD54 (ICAM-1) and CD71 (transferrin receptor). BDLF3 both enhanced internalization of surface MHC molecules and reduced the rate of their appearance at the cell surface. The reduced expression of surface MHC molecules correlated with functional protection against CD8(+) and CD4(+) T cell recognition. The molecular mechanism was identified as BDLF3-induced ubiquitination of MHC molecules and their subsequent downregulation in a proteasome-dependent manner. Immune evasion is a necessary feature of viruses that establish lifelong persistent infections in the face of strong immune responses. EBV is an important human pathogen whose immune evasion mechanisms are only partly understood. Of the EBV immune evasion mechanisms identified to date, none could explain why CD8(+) T cell responses to late lytic cycle genes are so infrequent and, when present, recognize lytically infected target cells so poorly relative to CD8(+) T cells specific for early lytic cycle antigens. The present work identifies an additional immune evasion protein, BDLF3, that is expressed late in the lytic cycle and impairs CD8(+) T cell recognition by targeting cell surface MHC class I molecules for ubiquitination and proteasome-dependent downregulation. Interestingly, BDLF3 also targets MHC class II molecules to impair CD4(+) T cell recognition. BDLF3 is therefore a rare example of a viral protein that impairs both the MHC class I and class II antigen-presenting pathways. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Mental model mapping as a new tool to analyse the use of information in decision-making in integrated water management

NASA Astrophysics Data System (ADS)

Kolkman, M. J.; Kok, M.; van der Veen, A.

The solution of complex, unstructured problems is faced with policy controversy and dispute, unused and misused knowledge, project delay and failure, and decline of public trust in governmental decisions. Mental model mapping (also called concept mapping) is a technique to analyse these difficulties on a fundamental cognitive level, which can reveal experiences, perceptions, assumptions, knowledge and subjective beliefs of stakeholders, experts and other actors, and can stimulate communication and learning. This article presents the theoretical framework from which the use of mental model mapping techniques to analyse this type of problems emerges as a promising technique. The framework consists of the problem solving or policy design cycle, the knowledge production or modelling cycle, and the (computer) model as interface between the cycles. Literature attributes difficulties in the decision-making process to communication gaps between decision makers, stakeholders and scientists, and to the construction of knowledge within different paradigm groups that leads to different interpretation of the problem situation. Analysis of the decision-making process literature indicates that choices, which are made in all steps of the problem solving cycle, are based on an individual decision maker’s frame of perception. This frame, in turn, depends on the mental model residing in the mind of the individual. Thus we identify three levels of awareness on which the decision process can be analysed. This research focuses on the third level. Mental models can be elicited using mapping techniques. In this way, analysing an individual’s mental model can shed light on decision-making problems. The steps of the knowledge production cycle are, in the same manner, ultimately driven by the mental models of the scientist in a specific discipline. Remnants of this mental model can be found in the resulting computer model. The characteristics of unstructured problems (complexity, uncertainty and disagreement) can be positioned in the framework, as can the communities of knowledge construction and valuation involved in the solution of these problems (core science, applied science, and professional consultancy, and “post-normal” science). Mental model maps, this research hypothesises, are suitable to analyse the above aspects of the problem. This hypothesis is tested for the case of the Zwolle storm surch barrier. Analysis can aid integration between disciplines, participation of public stakeholders, and can stimulate learning processes. Mental model mapping is recommended to visualise the use of knowledge, to analyse difficulties in problem solving process, and to aid information transfer and communication. Mental model mapping help scientists to shape their new, post-normal responsibilities in a manner that complies with integrity when dealing with unstructured problems in complex, multifunctional systems.

Systems and methods for compensating for electrical converter nonlinearities

DOEpatents

Perisic, Milun; Ransom, Ray M.; Kajouke, Lateef A.

2013-06-18

Systems and methods are provided for delivering energy from an input interface to an output interface. An electrical system includes an input interface, an output interface, an energy conversion module coupled between the input interface and the output interface, and a control module. The control module determines a duty cycle control value for operating the energy conversion module to produce a desired voltage at the output interface. The control module determines an input power error at the input interface and adjusts the duty cycle control value in a manner that is influenced by the input power error, resulting in a compensated duty cycle control value. The control module operates switching elements of the energy conversion module to deliver energy to the output interface with a duty cycle that is influenced by the compensated duty cycle control value.

DNA Damage Signaling Is Induced in the Absence of Epstein-Barr Virus (EBV) Lytic DNA Replication and in Response to Expression of ZEBRA.

PubMed

Wang'ondu, Ruth; Teal, Stuart; Park, Richard; Heston, Lee; Delecluse, Henri; Miller, George

2015-01-01

Epstein Barr virus (EBV), like other oncogenic viruses, modulates the activity of cellular DNA damage responses (DDR) during its life cycle. Our aim was to characterize the role of early lytic proteins and viral lytic DNA replication in activation of DNA damage signaling during the EBV lytic cycle. Our data challenge the prevalent hypothesis that activation of DDR pathways during the EBV lytic cycle occurs solely in response to large amounts of exogenous double stranded DNA products generated during lytic viral DNA replication. In immunofluorescence or immunoblot assays, DDR activation markers, specifically phosphorylated ATM (pATM), H2AX (γH2AX), or 53BP1 (p53BP1), were induced in the presence or absence of viral DNA amplification or replication compartments during the EBV lytic cycle. In assays with an ATM inhibitor and DNA damaging reagents in Burkitt lymphoma cell lines, γH2AX induction was necessary for optimal expression of early EBV genes, but not sufficient for lytic reactivation. Studies in lytically reactivated EBV-positive cells in which early EBV proteins, BGLF4, BGLF5, or BALF2, were not expressed showed that these proteins were not necessary for DDR activation during the EBV lytic cycle. Expression of ZEBRA, a viral protein that is necessary for EBV entry into the lytic phase, induced pATM foci and γH2AX independent of other EBV gene products. ZEBRA mutants deficient in DNA binding, Z(R183E) and Z(S186E), did not induce foci of pATM. ZEBRA co-localized with HP1β, a heterochromatin associated protein involved in DNA damage signaling. We propose a model of DDR activation during the EBV lytic cycle in which ZEBRA induces ATM kinase phosphorylation, in a DNA binding dependent manner, to modulate gene expression. ATM and H2AX phosphorylation induced prior to EBV replication may be critical for creating a microenvironment of viral and cellular gene expression that enables lytic cycle progression.

Anti-inflammatory activity of 6-hydroxy-2,7-dimethoxy-1,4-henanthraquinone from tuberous roots of yam (Dioscorea batatas) through inhibition of prostaglandin D₂ and leukotriene C₄ production in mouse bone marrow-derived mast cells.

PubMed

Jin, Meihua; Lu, Yue; Yang, Ju Hye; Jo, Tae Hyung; Park, Young In; Lee, Chong-Kil; Park, Sang-Jo; Son, Kun Ho; Chang, Hyeun Wook

2011-09-01

6-Hydroxy-2,7-dimethoxy-1,4-phenanthraquinone (PAQ) isolated from the tuberous roots of Yam (Dioscorea batatas) inhibited cyclooxygenase-2 (COX-2) and cyclooxygenase-1 (COX-1) dependent prostaglandin D(2) (PGD(2)) generation in mouse bone marrow-derived mast cells in a concentration-dependent manner with IC(50) values of 0.08 μM and 0.27 μM, respectively. In the Western blotting with specific anti-COX-2 antibodies, the decrease of the quantity of PGD(2) was accompanied by a decrease in the COX-2 protein level. But PAQ did not affect COX-1 protein level. In addition, this compound inhibited 5-lipoxygenase (5-LOX) dependent production of leukotriene C(4) in a dose-dependent manner, with an IC(50) of 0.032 μM. These results demonstrate that PAQ has a dual COX-2/5-LOX inhibitory activity. This compound also inhibited the degranulation reaction in a dose-dependent manner with an IC(50) of 2.7 μM. Thus, these results suggest that PAQ may be useful in regulating mast cell-mediated inflammatory diseases.

Resistance to cisplatin and paclitaxel does not affect the sensitivity of human ovarian cancer cells to antiprogestin-induced cytotoxicity.

PubMed

Gamarra-Luques, Carlos D; Hapon, Maria B; Goyeneche, Alicia A; Telleria, Carlos M

2014-01-01

Antiprogestin compounds have been shown to be effective in blocking the growth of ovarian cancer cells of different genetic backgrounds. Herein we studied the anti-ovarian cancer effect of a series of antiprogestins sharing the chemical backbone of the most characterized antiprogestin, mifepristone, but with unique modifications in position C-17 of the steroid ring. We assessed the effect of mifepristone-like antiprogestins on the growth of ovarian cancer cells sensitive to the standard combination therapy cisplatin-paclitaxel or made double-resistant upon six cycles of pulse-selection with the drugs used at clinically relevant concentrations and exposure times. IGROV-1 and SKOV-3 cells were pulsed with 20 μM cisplatin for 1 h followed by 100 nM paclitaxel for 3 h once a week for six weeks. The cells that did not die and repopulate the culture after the chemotherapies were termed Platinum-Taxane-EScape cells (PTES). Parental cells were compared against their PTES derivatives in their responses to further platinum-taxane treatments. Moreover, both ovarian cancer cells and their PTES siblings were exposed to escalating doses of the various antiprogestin derivatives. We assessed cell growth, viability and sub-G1 DNA content using microcapillary cytometry. Cyclin-dependent kinase inhibitors p21(cip1) and p27(kip1) and cleavage of downstream caspase-3 substrate PARP were used to assess whether cell fate, as a consequence of treatment, was limited to cytostasis or progressed to lethality. Cells subjected to six pulse-selection cycles of cisplatin-paclitaxel gave rise to sibling derivatives that displayed ~2-7 fold reduction in their sensitivities to further chemotherapy. However, regardless of the sensitivity the cells developed to the combination cisplatin-paclitaxel, they displayed similar sensitivity to the antiprogestins, which blocked their growth in a dose-related manner, with lower concentrations causing cytostasis, and higher concentrations causing lethality. Antiprogestins carrying a backbone similar to mifepristone are cytotoxic to ovarian cancer cells in a manner that does not depend on the sensitivity the cells have to the standard ovarian cancer chemotherapeutics, cisplatin and paclitaxel. Thus, antiprogestin therapy could be used to treat ovarian cancer cells showing resistance to both platinum and taxanes.

Resistance to cisplatin and paclitaxel does not affect the sensitivity of human ovarian cancer cells to antiprogestin-induced cytotoxicity

PubMed Central

2014-01-01

Background Antiprogestin compounds have been shown to be effective in blocking the growth of ovarian cancer cells of different genetic backgrounds. Herein we studied the anti-ovarian cancer effect of a series of antiprogestins sharing the chemical backbone of the most characterized antiprogestin, mifepristone, but with unique modifications in position C-17 of the steroid ring. We assessed the effect of mifepristone-like antiprogestins on the growth of ovarian cancer cells sensitive to the standard combination therapy cisplatin-paclitaxel or made double-resistant upon six cycles of pulse-selection with the drugs used at clinically relevant concentrations and exposure times. Methods IGROV-1 and SKOV-3 cells were pulsed with 20 μM cisplatin for 1 h followed by 100 nM paclitaxel for 3 h once a week for six weeks. The cells that did not die and repopulate the culture after the chemotherapies were termed Platinum-Taxane-EScape cells (PTES). Parental cells were compared against their PTES derivatives in their responses to further platinum-taxane treatments. Moreover, both ovarian cancer cells and their PTES siblings were exposed to escalating doses of the various antiprogestin derivatives. We assessed cell growth, viability and sub-G1 DNA content using microcapillary cytometry. Cyclin-dependent kinase inhibitors p21cip1 and p27kip1 and cleavage of downstream caspase-3 substrate PARP were used to assess whether cell fate, as a consequence of treatment, was limited to cytostasis or progressed to lethality. Results Cells subjected to six pulse-selection cycles of cisplatin-paclitaxel gave rise to sibling derivatives that displayed ~2-7 fold reduction in their sensitivities to further chemotherapy. However, regardless of the sensitivity the cells developed to the combination cisplatin-paclitaxel, they displayed similar sensitivity to the antiprogestins, which blocked their growth in a dose-related manner, with lower concentrations causing cytostasis, and higher concentrations causing lethality. Conclusions Antiprogestins carrying a backbone similar to mifepristone are cytotoxic to ovarian cancer cells in a manner that does not depend on the sensitivity the cells have to the standard ovarian cancer chemotherapeutics, cisplatin and paclitaxel. Thus, antiprogestin therapy could be used to treat ovarian cancer cells showing resistance to both platinum and taxanes. PMID:24795781

Oscillating in synchrony with a metronome: serial dependence, limit cycle dynamics, and modeling.

PubMed

Torre, Kjerstin; Balasubramaniam, Ramesh; Delignières, Didier

2010-07-01

We analyzed serial dependencies in periods and asynchronies collected during oscillations performed in synchrony with a metronome. Results showed that asynchronies contain 1/f fluctuations, and the series of periods contain antipersistent dependence. The analysis of the phase portrait revealed a specific asymmetry induced by synchronization. We propose a hybrid limit cycle model including a cycle-dependent stiffness parameter provided with fractal properties, and a parametric driving function based on velocity. This model accounts for most experimentally evidenced statistical features, including serial dependence and limit cycle dynamics. We discuss the results and modeling choices within the framework of event-based and emergent timing.

Tidal-cycle changes in oscillation ripples on the inner part of an estuarine sand flat

USGS Publications Warehouse

Dingler, J.R.; Clifton, H.E.

1984-01-01

Oscillation ripples form on subaqueous sand beds when wave-generated, near-bottom water motions are strong enough to move sand grains. The threshold of grain motion is the lower bound of the regime of oscillation ripples and the onset of sheet flow is the upper bound. Based on the relation between ripple spacing and orbital diameter, three types of symmetrical ripples occur within the ripple regime. In the lower part of the ripple regime (orbital ripples), spacing is proportional to orbital diameter; in the upper part (anorbital ripples) spacing is independent of orbital diameter. Between these regions occurs a transitional region (suborbital ripples). Oscillation ripples develop on a sandy tidal flat in Willapa Bay, Washington, as a result of waves traversing the area when it is submerged. Because wave energy is usually low within the bay, the ripples are primarily orbital in type. This means that their spacing should respond in a systematic way to changes in wave conditions. During the high-water parts of some tidal cycles, ripples near the beach decrease in spacing during the latter stage of the ebb tide while ripples farther offshore do not change. Observations made over several tidal cycles show that the zone of active ripples shifts on- or offshore in response to different wave conditions. Detailed bed profiles and current measurements taken during the high-water part of spring tides show the manner in which the oscillation ripples change with changes in orbital diameter. Changes in ripple spacing at the study site could be correlated with changes in orbital diameter in the manner suggested by the criterion for orbital ripples. However, there appeared to be a lag time between a decrease in orbital diameter and the corresponding decrease in ripple spacing. Absence of change during a tidal cycle could be attributed to orbital velocities below the threshold for grain motion that negated the effects of changes in orbital diameter. Because changes in sand-flat ripples depend both upon changes in orbital diameter and upon the magnitude of the orbital velocity, exposed ripples were not necessarily produced during the preceding high tide. In fact, some ripples may have been just produced, while others, farther offshore, may have been produced an unknown number of tides earlier. Therefore, when interpreting past wave conditions over tidal flats from low-tide ripples, one must remember that wave periods have to be short enough to produce velocities greater than the threshold velocity for the orbital diameters calculated from the observed ripple spacings. ?? 1984.

Tobacco components stimulate Akt-dependent proliferation and NFkappaB-dependent survival in lung cancer cells.

PubMed

Tsurutani, Junji; Castillo, S Sianna; Brognard, John; Granville, Courtney A; Zhang, Chunyu; Gills, Joell J; Sayyah, Jacqueline; Dennis, Phillip A

2005-07-01

Retrospective studies have shown that patients with tobacco-related cancers who continue to smoke after their diagnoses have lower response rates and shorter median survival compared with patients who stop smoking. To provide insight into the biologic basis for these clinical observations, we tested whether two tobacco components, nicotine or the tobacco-specific carcinogen, 4-(methylnitrosoamino)-1-(3-pyridyl)-1-butanone (NNK), could activate the Akt pathway and increase lung cancer cell proliferation and survival. Nicotine or NNK, rapidly and potently, activated Akt in non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC) cells. Nicotinic activation of Akt increased phosphorylation of multiple downstream substrates of Akt in a time-dependent manner, including GSK-3, FKHR, tuberin, mTOR and S6K1. Since nicotine or NNK bind to cell surface nicotinic acetylcholine receptors (nAchR), we used RT-PCR to assess expression of nine alpha and three beta nAchR subunits in five NSCLC cell lines and two types of primary lung epithelial cells. NSCLC cells express multiple nAchR subunits in a cell line-specific manner. Agonists of alpha3/alpha4 or alpha7 subunits activated Akt in a time-dependent manner, suggesting that tobacco components utilize these subunits to activate Akt. Cellular outcomes after nicotine or NNK administration were also assessed. Nicotine or NNK increased proliferation of NSCLC cells in an Akt-dependent manner that was closely linked with changes in cyclin D1 expression. Despite similar induction of proliferation, only nicotine decreased apoptosis caused by serum deprivation and/or chemotherapy. Protection conferred by nicotine was NFkappaB-dependent. Collectively, these results identify tobacco component-induced, Akt-dependent proliferation and NFkappaB-dependent survival as cellular processes that could underlie the detrimental effects of smoking in cancer patients.

The reproductive-cell cycle theory of aging: an update.

PubMed

Atwood, Craig S; Bowen, Richard L

2011-01-01

The Reproductive-Cell Cycle Theory posits that the hormones that regulate reproduction act in an antagonistic pleiotrophic manner to control aging via cell cycle signaling; promoting growth and development early in life in order to achieve reproduction, but later in life, in a futile attempt to maintain reproduction, become dysregulated and drive senescence. Since reproduction is the most important function of an organism from the perspective of the survival of the species, if reproductive-cell cycle signaling factors determine the rate of growth, determine the rate of development, determine the rate of reproduction, and determine the rate of senescence, then by definition they determine the rate of aging and thus lifespan. The theory is able to explain: 1) the simultaneous regulation of the rate of aging and reproduction as evidenced by the fact that environmental conditions and experimental interventions known to extend longevity are associated with decreased reproductive-cell cycle signaling factors, thereby slowing aging and preserving fertility in a hostile reproductive environment; 2) two phenomena that are closely related to species lifespan-the rate of growth and development and the ultimate size of the animal; 3). the apparent paradox that size is directly proportional to lifespan and inversely proportional to fertility between species but vice versa within a species; 4). how differing rates of reproduction between species is associated with differences in their lifespan; 5). why we develop aging-related diseases; and 6). an evolutionarily credible reason for why and how aging occurs-these hormones act in an antagonistic pleiotrophic manner via cell cycle signaling; promoting growth and development early in life in order to achieve reproduction, but later in life, in a futile attempt to maintain reproduction, become dysregulated and drive senescence (dyosis). In essence, the Reproductive-Cell Cycle Theory can explain aging in all sexually reproductive life forms. Published by Elsevier Inc.

Osthole inhibits the PI3K/AKT signaling pathway via activation of PTEN and induces cell cycle arrest and apoptosis in esophageal squamous cell carcinoma.

PubMed

Zhu, Xinbing; Li, Zhengzheng; Li, Tongtong; Long, Fei; Lv, Yuesheng; Liu, Lei; Liu, Xuefeng; Zhan, Qimin

2018-06-01

Esophageal squamous cell carcinoma (ESCC) is one of the most common lethal tumors and is known to be lack of effective therapy. Thus, novel therapeutic strategies are greatly needed for treatment of ESCC. Osthole, a natural active extract, has been documented to have anti-tumor activity. However, the effect of osthole on ESCC cells has not been elucidated. In this study, we demonstrated that osthole could inhibit the ESCC cell proliferation in dose- and time-dependent manner. Osthole treatment also induced G2/M phase arrest and apoptosis of ESCC cells. Furthermore, upon exposure to osthole, the expression of Cyclin B1, Cdc2, Bcl-2, PARP1 and Survivin was decreased, while the expression of BAX, cleaved PARP1, cleaved Caspase3 and cleaved Caspase9 was increased. In addition, osthole treatment elicited upregulation of PTEN and downregulation of PI3K and phosphorylated AKT (p-AKT). Taken together, our study demonstrates that osthole could suppress ESCC proliferation through inducing cell cycle arrest and apoptosis. Moreover, PTEN-PI3K/AKT signaling pathway can be regulated by osthole. Our results indicate that osthole may find therapeutic application in the treatment of ESCC patients. Copyright © 2018. Published by Elsevier Masson SAS.

Growth inhibitory and proapoptotic effects of l-asparaginase from Fusarium culmorum ASP-87 on human leukemia cells (Jurkat).

PubMed

Meghavarnam, Anil K; Salah, Maryam; Sreepriya, Meenakshisundaram; Janakiraman, Savitha

2017-06-01

The objective of this study was to evaluate the anticancer properties of l-asparaginase purified from fungal isolate Fusarium culmorum ASP-87 against human T-cell leukemia cell line (Jurkat). The growth inhibitory and proapoptotic effects of purified l-asparaginase on Jurkat cell lines were investigated by determining its influence on cell viability, colony formation, DNA fragmentation, and cell cycle progression. The results revealed that purified l-asparaginase showed significant decrease in cell survival with IC 50 value of 90 μg/mL (9 IU/mL). The enzyme inhibited colony formation and showed characteristic laddering pattern on agarose gel thereby confirming the induction of apoptosis. Further, cell cycle analysis revealed that the enzyme induced apoptotic cell death by arresting the growth of cells at G 2 -M phase. However, the enzyme did not elicit any toxic effects on human erythrocytes. l-asparaginase purified from F. culmorum ASP-87 showed significant and selective cytotoxic and apoptotic effects on human T-cell leukemic cells in dose-dependent manner. Results of the study give leads for the anticancer effects of fungal l-asparaginase and its potential usefulness in the chemotherapy of leukemia. © 2016 Société Française de Pharmacologie et de Thérapeutique.

Targeting c-Myc: JQ1 as a promising option for c-Myc-amplified esophageal squamous cell carcinoma.

PubMed

Wang, Jingyuan; Liu, Zhentao; Wang, Ziqi; Wang, Shubin; Chen, Zuhua; Li, Zhongwu; Zhang, Mengqi; Zou, Jianling; Dong, Bin; Gao, Jing; Shen, Lin

2018-04-10

c-Myc amplification-induced cell cycle dysregulation is a common cause for esophageal squamous cell carcinoma (ESCC), but no approved targeted drug is available so far. The bromodomain inhibitor JQ1, which targets c-Myc, exerts anti-tumor activity in multiple cancers. However, the role of JQ1 in ESCC remains unknown. In this study, we reported that JQ1 had potent anti-proliferative effects on ESCC cells in both time- and dose-dependent manners by inducing cell cycle arrest at G1 phase, cell apoptosis, and the mesenchymal-epithelial transition. Follow-up studies revealed that both c-Myc/cyclin/Rb and PI3K/AKT signaling pathways were inactivated by JQ1, as indicated by the downregulation of c-Myc, cyclin A/E, and phosphorylated Rb, AKT and S6. Tumor suppression induced by JQ1 in c-Myc amplified or highly expressed xenografts was higher than that in xenografts with low expression, suggesting its potential role in prediction. In conclusion, targeting c-Myc by JQ1 could cause significant tumor suppression in ESCC both in vitro and in vivo. Also, c-Myc amplification or high expression might serve as a potential biomarker and provide a promising therapeutic option for ESCC. Copyright © 2018 Elsevier B.V. All rights reserved.

NF-κB mediates the antiproliferative and proapoptotic effects of bergamot juice in HepG2 cells.

PubMed

Ferlazzo, Nadia; Cirmi, Santa; Russo, Marina; Trapasso, Elena; Ursino, Maria Rita; Lombardo, Giovanni Enrico; Gangemi, Sebastiano; Calapai, Gioacchino; Navarra, Michele

2016-02-01

Among cancers, hepatocellular carcinoma is one of the commonest worldwide, and its incidence is increasing around the world. A lot of evidence underlines that natural substances usually consumed in the diet can have an important role in the prevention of cancer. In this study we investigated the molecular mechanisms underlying the antiproliferative activity of Citrus bergamia (bergamot) juice (BJ) in human hepatocellular carcinoma HepG2 cells. HepG2 cells were exposed to BJ and then cell proliferation, cell cycle progression, apoptosis and NF-κB nuclear translocation were evaluated. Here we present results demonstrating that BJ reduced the growth rate of human hepatocellular carcinoma HepG2 cells in a time- and concentration-dependent manner, by a mechanism involving the activation of apoptotic machinery via both intrinsic and extrinsic pathways. Moreover, BJ increased expression of P53 and P21 proteins that may be responsible for the HepG2 cell cycle arrest in G2 phase. In addition, BJ reduced NF-κB nuclear translocation. Our data demonstrate the ability of BJ in reducing the growth of HepG2 cells, revealing its mechanism of action and suggesting a promising role as anticancer drugs. Copyright © 2016 Elsevier Inc. All rights reserved.

Influence of homeobox B2 antisense oligodeoxynucleotides on the biological characteristics of in vitro cultured primary human umbilical vein endothelial cells.

PubMed

Liu, X S; Zhang, X Q; Tian, T; Liu, L; Ming, J

2008-01-01

This study aims to explore the influence of homeobox B2 (HOXB2) antisense oligodeoxynucleotides (asodn) on the biological characteristics of in vitro cultured primary human umbilical vein endothelial cells (HUVECs). The distribution of HOXB2 asodn in the HUVECs was observed by fluorescent labelling, and the influence of different concentrations of HOXB2 asodn on the DNA synthesis of HUVECs was assessed. Flow cytometry and a reverse transcriptase-polymerase chain reaction (RT- PCR) method were employed to observe the influence of HOXB2 asodn on HOXB2 expression and the HUVEC cell cycle. After the induction of liposome, the nuclear fluorescent staining of HOXB2 asodn was weaker 15 min after transfection and the staining reached the strongest level at 4-8 h but then weakened and disappeared by 16 h after transfection. This indicated that endothelial DNA synthesis could be inhibited by HOXB2 asodn in a dose-dependent manner. Furthermore, the HUVECs could be delayed in their passage from G1 to S. Simultaneously, expression of HOXB2 mRNA had decreased significantly by 24-48 h after transfection. Clearly, HOXB2 plays important roles in the proliferation of endothelial cells and also affects the cell cycle.

The Arabidopsis CSLD 5 functions in cell plate formation in a cell cycle-dependent manner

DOE PAGES

Gu, Fangwei; Bringmann, Martin; Combs, Jonathon; ...

2016-06-27

In plants, the presence of a load-bearing cell wall presents unique challenges during cell division. Unlike other eukaryotes, which undergo contractile cytokinesis upon completion of mitosis, plants instead synthesize and assemble a new dividing cell wall to separate newly formed daughter cells. In this study, we mine transcriptome data from individual cell types in the Arabidopsis thaliana stomatal lineage and identify CSLD5, a member of the Cellulose Synthase Like-D family, as a cell wall biosynthesis enzyme uniquely enriched in rapidly dividing cell populations. We further show that CSLD5 is a direct target of SPEECHLESS, the master transcriptional regulator of thesemore » divisions during stomatal development. Using a combination of genetic analysis and in vivo localization of fluorescently tagged fusion proteins, we show that CSLD5 preferentially accumulates in dividing plant cells where it participates in the construction of newly forming cell plates. We show that CSLD5 is an unstable protein that is rapidly degraded upon completion of cell division and that the protein turnover characteristics of CSLD5 are altered in ccs52a2 mutants, indicating that CSLD5 turnover may be regulated by a cell cycle-associated E3-ubiquitin ligase, the anaphase-promoting complex.« less

Tetramethoxychalcone, a chalcone derivative, suppresses proliferation, blocks cell cycle progression, and induces apoptosis of human ovarian cancer cells.

PubMed

Qi, Zihao; Liu, Mingming; Liu, Yang; Zhang, Meiqin; Yang, Gong

2014-01-01

In the present study, we investigated the in vitro antitumor functions of a synthetic chalcone derivative 4,3',4',5'- tetramethoxychalcone (TMOC) in ovarian cancer cells. We found that TMOC inhibited the proliferation and colony formation of cisplatin sensitive cell line A2780 and resistant cell line A2780/CDDP, as well as ovarian cancer cell line SKOV3 in a time- and dose-dependent manner. Treatment of A2780 cells with TMOC resulted in G0/G1 cell cycle arrest through the down-regulation of cyclin D1 and CDK4, and the up-regulation of p16, p21 and p27 proteins. We demonstrated that TMOC might induce cell apoptosis through suppressing Bcl-2 and Bcl-xL, but enhancing the expression of Bax and the cleavage of PARP-1. Treatment of TMOC also reduced the invasion and migration of A2780 cells. Finally, we found that TMOC inhibited the constitutive activation of STAT3 signaling pathway and induced the expression of the tumor suppressor PTEN regardless of the p53 status in cell lines. These data suggest that TMOC may be developed as a potential chemotherapeutic agent to effectively treat certain cancers including ovarian cancer.

Tetramethoxychalcone, a Chalcone Derivative, Suppresses Proliferation, Blocks Cell Cycle Progression, and Induces Apoptosis of Human Ovarian Cancer Cells

PubMed Central

Liu, Yang; Zhang, Meiqin; Yang, Gong

2014-01-01

In the present study, we investigated the in vitro antitumor functions of a synthetic chalcone derivative 4,3′,4′,5′- tetramethoxychalcone (TMOC) in ovarian cancer cells. We found that TMOC inhibited the proliferation and colony formation of cisplatin sensitive cell line A2780 and resistant cell line A2780/CDDP, as well as ovarian cancer cell line SKOV3 in a time- and dose-dependent manner. Treatment of A2780 cells with TMOC resulted in G0/G1 cell cycle arrest through the down-regulation of cyclin D1 and CDK4, and the up-regulation of p16, p21 and p27 proteins. We demonstrated that TMOC might induce cell apoptosis through suppressing Bcl-2 and Bcl-xL, but enhancing the expression of Bax and the cleavage of PARP-1. Treatment of TMOC also reduced the invasion and migration of A2780 cells. Finally, we found that TMOC inhibited the constitutive activation of STAT3 signaling pathway and induced the expression of the tumor suppressor PTEN regardless of the p53 status in cell lines. These data suggest that TMOC may be developed as a potential chemotherapeutic agent to effectively treat certain cancers including ovarian cancer. PMID:25180593

Isolation and characterization of an anticancer catechol compound from Semecarpus anacardium.

PubMed

Nair, P K Raveedran; Melnick, Steven J; Wnuk, Stanislaw F; Rapp, Magdalena; Escalon, Enrique; Ramachandran, Cheppail

2009-04-21

The fruits and seeds of Semecarpus anacardium are used widely for the treatment of human cancers and other diseases in the Ayurvedic and Sidda systems of medicine in India. The principal aim of this investigation was to isolate and characterize the anticancer compound from the kernel of Semecarpus anacardium nut. The bioactivity-tailored isolation and detailed chemical characterization were used to identify the active compound. Cytotoxicity, apoptosis, cell cycle arrest as well as synergism between the identified anticancer compound and doxorubicin in human tumor cell lines were analyzed. GC/MS, IR, proton NMR, carbon NMR and collisionally induced dissociation (CID) spectra analysis showed that the isolated active compound is 3-(8'(Z),11'(Z)-pentadecadienyl) catechol (SA-3C). SA-3C is cytotoxic to tumor cell lines with IC(50) values lower than doxorubicin and even multidrug resistant tumor cell lines were equally sensitive to SA-3C. SA-3C induced apoptosis in human leukemia cell lines in a dose-dependent manner and showed synergistic cytotoxicity with doxorubicin. The cell cycle arrest induced by SA-3C at S- and G(2)/M-phases correlated with inhibition of checkpoint kinases. SA-3C isolated from the kernel of Semecarpus anacardium can be developed as an important anticancer agent for single agent and/or multiagent cancer therapy.

The spatio-temporal dynamics of PKA activity profile during mitosis and its correlation to chromosome segregation.

PubMed

Vandame, Pauline; Spriet, Corentin; Trinel, Dave; Gelaude, Armance; Caillau, Katia; Bompard, Coralie; Biondi, Emanuele; Bodart, Jean-François

2014-01-01

The cyclic adenosine monophosphate dependent kinase protein (PKA) controls a variety of cellular processes including cell cycle regulation. Here, we took advantages of genetically encoded FRET-based biosensors, using an AKAR-derived biosensor to characterize PKA activity during mitosis in living HeLa cells using a single-cell approach. We measured PKA activity changes during mitosis. HeLa cells exhibit a substantial increase during mitosis, which ends with telophase. An AKAREV T>A inactive form of the biosensor and H89 inhibitor were used to ascertain for the specificity of the PKA activity measured. On a spatial point of view, high levels of activity near to chromosomal plate during metaphase and anaphase were detected. By using the PKA inhibitor H89, we assessed the role of PKA in the maintenance of a proper division phenotype. While this treatment in our hands did not impaired cell cycle progression in a drastic manner, inhibition of PKA leads to a dramatic increase in chromososme misalignement on the spindle during metaphase that could result in aneuploidies. Our study emphasizes the insights that can be gained with genetically encoded FRET-based biosensors, which enable to overcome the shortcomings of classical methologies and unveil in vivo PKA spatiotemporal profiles in HeLa cells.

Intercellular Variation in Signaling through the TGF-β Pathway and Its Relation to Cell Density and Cell Cycle Phase*

PubMed Central

Zieba, Agata; Pardali, Katerina; Söderberg, Ola; Lindbom, Lena; Nyström, Erik; Moustakas, Aristidis; Heldin, Carl-Henrik; Landegren, Ulf

2012-01-01

Fundamental open questions in signal transduction remain concerning the sequence and distribution of molecular signaling events among individual cells. In this work, we have characterized the intercellular variability of transforming growth factor β-induced Smad interactions, providing essential information about TGF-β signaling and its dependence on the density of cell populations and the cell cycle phase. By employing the recently developed in situ proximity ligation assay, we investigated the dynamics of interactions and modifications of Smad proteins and their partners under native and physiological conditions. We analyzed the kinetics of assembly of Smad complexes and the influence of cellular environment and relation to mitosis. We report rapid kinetics of formation of Smad complexes, including native Smad2-Smad3-Smad4 trimeric complexes, in a manner influenced by the rate of proteasomal degradation of these proteins, and we found a striking cell to cell variation of signaling complexes. The single-cell analysis of TGF-β signaling in genetically unmodified cells revealed previously unknown aspects of regulation of this pathway, and it provided a basis for analysis of these signaling events to diagnose pathological perturbations in patient samples and to evaluate their susceptibility to drug treatment. PMID:22442258

Antitumor Activity of Ethanolic Extract of Dendrobium formosum in T-Cell Lymphoma: An In Vitro and In Vivo Study

PubMed Central

Prasad, Ritika; Koch, Biplob

2014-01-01

Dendrobium, a genus of orchid, was found to possess useful therapeutic activities like anticancer, hypoglycaemic, antimicrobial, immunomodulatory, hepatoprotective, antioxidant, and neuroprotective activities. The study was aimed to evaluate the anticancer property of the ethanolic extract of Dendrobium formosum on Dalton's lymphoma. In vitro cytotoxicity was determined by MTT assay, apoptosis was determined by fluorescence microscopy, and cell cycle progression was analysed using flow cytometry; in vivo antitumor activity was performed in Dalton's lymphoma bearing mice. The IC50 value of ethanolic extract was obtained at 350 μg/mL in Dalton's lymphoma cells. Fluorescence microscopy analysis showed significant increase in apoptotic cell death in dose- and time-dependent manner which was further confirmed through the resulting DNA fragmentation. Further, flow cytometry analysis showed that the ethanolic extract arrests the cells in G2/M phase of the cell cycle. The in vivo anticancer activity study illustrates significant increase in the survival time of Dalton's lymphoma bearing mice on treatment with ethanolic extract when compared to control. These results substantiate the antitumor properties of ethanolic extract of Dendrobium formosum and suggest an alternative in treatment of cancer. Further studies are required regarding the isolation and characterization of bioactive components along with the analysis of molecular mechanism involved. PMID:24959588

Molecular response to phototoxic stress of UVB-irradiated ketoprofen through arresting cell cycle in G2/M phase and inducing apoptosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu Shicheng; Mizu, Hideo; Yamauchi, Hitoshi

The phototoxicity of ketoprofen (KP), a non-steroidal anti-inflammatory drug, has recently attracted considerable attention, because it is photolabile and undergoes degradation when irradiated by sunlight to induce various skin diseases. The present study shows that combination of UVB irradiation with KP induced the cytotoxicity and suppressed DNA synthesis in HaCaT cells in a concentration-dependent manner. UVB-irradiated KP inhibited the cell growth and induced G2/M cell cycle arrest by modulating the levels of cdc2, cyclin B1, Chk1, Tyr15-phosphorylated cdc2 and p21. It also provoked a striking accumulation of cyclin B1-cdc2-p21 complexes, concomitantly with an increase in the levels of Tyr15-phosphorylated cdc2more » and p21 protein. The presence of KP accentuated the apoptotic response to UVB radiation in HaCaT cells as evidenced by DAPI staining. The apoptotic process was associated with activation of caspase-9, caspase-3 and cleavage of PARP, and this activation could be prevented by a specific caspase-3 inhibitor. Taken together, our results suggest that KP-photoinduced apoptosis may be a useful approach to reduce or prevent skin carcinogenesis.« less

RPA accumulation during class switch recombination represents 5'-3' DNA-end resection during the S-G2/M phase of the cell cycle.

PubMed

Yamane, Arito; Robbiani, Davide F; Resch, Wolfgang; Bothmer, Anne; Nakahashi, Hirotaka; Oliveira, Thiago; Rommel, Philipp C; Brown, Eric J; Nussenzweig, Andre; Nussenzweig, Michel C; Casellas, Rafael

2013-01-31

Activation-induced cytidine deaminase (AID) promotes chromosomal translocations by inducing DNA double-strand breaks (DSBs) at immunoglobulin (Ig) genes and oncogenes in the G1 phase. RPA is a single-stranded DNA (ssDNA)-binding protein that associates with resected DSBs in the S phase and facilitates the assembly of factors involved in homologous repair (HR), such as Rad51. Notably, RPA deposition also marks sites of AID-mediated damage, but its role in Ig gene recombination remains unclear. Here, we demonstrate that RPA associates asymmetrically with resected ssDNA in response to lesions created by AID, recombination-activating genes (RAG), or other nucleases. Small amounts of RPA are deposited at AID targets in G1 in an ATM-dependent manner. In contrast, recruitment in the S-G2/M phase is extensive, ATM independent, and associated with Rad51 accumulation. In the S-G2/M phase, RPA increases in nonhomologous-end-joining-deficient lymphocytes, where there is more extensive DNA-end resection. Thus, most RPA recruitment during class switch recombination represents salvage of unrepaired breaks by homology-based pathways during the S-G2/M phase of the cell cycle. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

The antibacterial peptide from Bombyx mori cecropinXJ induced growth arrest and apoptosis in human hepatocellular carcinoma cells.

PubMed

Xia, Lijie; Wu, Yanling; Ma, J I; Yang, Jianhua; Zhang, Fuchun

2016-07-01

CecropinXJ is a cationic antimicrobial peptide originally isolated from the larvae of Bombyx mori . The anticancer effect of cecropinXJ has been reported in various tumor cells, including leukemia, gastric and esophageal cancer cells. However, the activity of cecropinXJ on hepatocellular carcinoma (HCC) and its underlying mechanism have not been investigated to date. Therefore, the present study investigated the efficacy and associated mechanism of cecropinXJ in Huh-7 cells. Flow cytometric analysis was performed to determine the presence of cell cycle arrested and apoptotic cells. CecropinXJ significantly inhibited the growth of Huh-7 cells in a dose- and time-dependent manner. CecropinXJ treatment for 24 h induced S cell cycle arrest and apoptosis, in addition to loss of the mitochondrial membrane potential, in hepatoma cells. CecropinXJ induced HCC cell apoptosis by activating caspase-3 and poly(ADP-ribose) polymerase. Furthermore, cecropinXJ downregulated the expression of B-cell lymphoma 2 (Bcl-2), while upregulated the expression of Bcl-2-associated death promoter and Bcl-2-associated X protein. In conclusion, the results of the present study suggest that cecropinXJ may be an active anti-HCC agent and provide novel insights into the mechanism of cecropinXJ.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Han, Dong-Wook; Department of Medical Engineering, Yonsei University College of Medicine, 134 Shinchon-dong, Seodaemun-gu, Seoul 120-752; Lim, Hye Ryeon

The abnormal growth of vascular smooth muscle cells (VSMCs) plays an important role in vascular diseases, including atherosclerosis and restenosis after angioplasty. Although (-)-epigallocatechin-3-O-gallate (EGCG) has antiproliferative effects on various cells, relatively a little is known about precise mechanisms of the inhibitory effects of EGCG on SMCs. In this study, the inhibitory effects of EGCG on attachment, proliferation, migration, and cell cycle of rat aortic SMCs (RASMCs) with serum stimulation were investigated. Also, the involvement of nuclear factor-{kappa}B (NF-{kappa}B) during these inhibitions by EGCG was examined. EGCG treatment resulted in significant (p < 0.05) inhibition in attachment and proliferation ofmore » RASMCs induced by serum. While non-treated RASMCs migrated into denuded area in response to serum and showed essentially complete closure after 36 h, EGCG-treated cells covered only 31% of the area even after 48 h of incubation. Furthermore, EGCG treatment resulted in an appreciable cell cycle arrest at both G0/G1- and G2/M-phases. The immunoblot analysis revealed that the constitutive expression of NF-{kappa}B/p65 nuclear protein in RASMCs was lowered by EGCG in both the cytosol and the nucleus in a dose-dependent manner. These results suggest that the EGCG-caused inhibitory effects on RASMCs may be mediated through NF-{kappa}B down-modulation.« less

Cortex and hippocampus DNA epigenetic response to a long-term arsenic exposure via drinking water.

PubMed

Du, Xiaoyan; Tian, Meiping; Wang, Xiaoxue; Zhang, Jie; Huang, Qingyu; Liu, Liangpo; Shen, Heqing

2018-03-01

The neurotoxicity of arsenic is a serious health problem, especially for children. DNA epigenetic change may be an important pathogenic mechanism, but the molecular pathway remains obscure. In this study, the weaned male Sprague-Dawly (SD) rats were treated with arsenic trioxide via drinking water for 6 months, simulating real developmental exposure situation of children. Arsenic exposure impaired the cognitive abilities, and altered the expression of neuronal activity-regulated genes. Total arsenic concentrations of cortex and hippocampus tissues were significantly increased in a dose-dependent manner. The reduction in 5-methylcytosine (5 mC) and 5-hydroxymethylcytosine (5hmC) levels as well as the down-regulation of DNA methyltransferases (DNMTs) and ten-eleven translocations (TETs) expression suggested that DNA methylation/demethylation processes were significantly suppressed in brain tissues. S-adenosylmethionine (SAM) level wasn't changed, but the expression of the important indicators of oxidative/anti-oxidative balance and tricarboxylic acid (TCA) cycle was significantly deregulated. Overall, arsenic can disrupt oxidative/anti-oxidative balance, further inhibit TETs expression through TCA cycle and alpha-ketoglutarate (α-KG) pathway, and consequently cause DNA methylation/demethylation disruption. The present study implies oxidative stress but not SAM depletion may lead to DNA epigenetic alteration and arsenic neurotoxicity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Rhythm Defects Caused by Newly Engineered Null Mutations in Drosophila's cryptochrome Gene

PubMed Central

Dolezelova, Eva; Dolezel, David; Hall, Jeffrey C.

2007-01-01

Much of the knowledge about cryptochrome function in Drosophila stems from analyzing the cryb mutant. Several features of this variant's light responsiveness imply either that CRYb retains circadian-photoreceptive capacities or that additional CRY-independent light-input routes subserve these processes. Potentially to resolve these issues, we generated cry knock-out mutants (cry0's) by gene replacement. They behaved in an anomalously rhythmic manner in constant light (LL). However, cry0 flies frequently exhibited two separate circadian components in LL, not observed in most previous cryb analyses. Temperature-dependent circadian phenotypes exhibited by cry0 flies suggest that CRY is involved in core pacemaking. Further locomotor experiments combined cry0 with an externally blinding mutation (norpAP24), which caused the most severe decrements of circadian photoreception observed so far. cryb cultures were shown previously to exhibit either aperiodic or rhythmic eclosion in separate studies. We found cry0 to eclose in a solidly periodic manner in light:dark cycles or constant darkness. Furthermore, both cry0 and cryb eclosed rhythmically in LL. These findings indicate that the novel cry0 type causes more profound defects than does the cryb mutation, implying that CRYb retains residual activity. Because some norpAP24 cry0 individuals can resynchronize to novel photic regimes, an as-yet undetermined light-input route exists in Drosophila. PMID:17720919

The Novel Fission Yeast Protein Pal1p Interacts with Hip1-related Sla2p/End4p and Is Involved in Cellular Morphogenesis

PubMed Central

Ge, Wanzhong; Chew, Ting Gang; Wachtler, Volker; Naqvi, Suniti N.; Balasubramanian, Mohan K.

2005-01-01

The establishment and maintenance of characteristic cellular morphologies is a fundamental property of all cells. Here we describe Schizosaccharomyces pombe Pal1p, a protein important for maintenance of cylindrical cellular morphology. Pal1p is a novel membrane-associated protein that localizes to the growing tips of interphase cells and to the division site in cells undergoing cytokinesis in an F-actin- and microtubule-independent manner. Cells deleted for pal1 display morphological defects, characterized by the occurrence of spherical and pear-shaped cells with an abnormal cell wall. Pal1p physically interacts and displays overlapping localization with the Huntingtin-interacting-protein (Hip1)-related protein Sla2p/End4p, which is also required for establishment of cylindrical cellular morphology. Sla2p is important for efficient localization of Pal1p to the sites of polarized growth and appears to function upstream of Pal1p. Interestingly, spherical pal1Δ mutants polarize to establish a pearlike morphology before mitosis in a manner dependent on the kelch-repeat protein Tea1p and the cell cycle inhibitory kinase Wee1p. Thus, overlapping mechanisms involving Pal1p, Tea1p, and Sla2p contribute to the establishment of cylindrical cellular morphology, which is important for proper spatial regulation of cytokinesis. PMID:15975911

The novel fission yeast protein Pal1p interacts with Hip1-related Sla2p/End4p and is involved in cellular morphogenesis.

PubMed

Ge, Wanzhong; Chew, Ting Gang; Wachtler, Volker; Naqvi, Suniti N; Balasubramanian, Mohan K

2005-09-01

The establishment and maintenance of characteristic cellular morphologies is a fundamental property of all cells. Here we describe Schizosaccharomyces pombe Pal1p, a protein important for maintenance of cylindrical cellular morphology. Pal1p is a novel membrane-associated protein that localizes to the growing tips of interphase cells and to the division site in cells undergoing cytokinesis in an F-actin- and microtubule-independent manner. Cells deleted for pal1 display morphological defects, characterized by the occurrence of spherical and pear-shaped cells with an abnormal cell wall. Pal1p physically interacts and displays overlapping localization with the Huntingtin-interacting-protein (Hip1)-related protein Sla2p/End4p, which is also required for establishment of cylindrical cellular morphology. Sla2p is important for efficient localization of Pal1p to the sites of polarized growth and appears to function upstream of Pal1p. Interestingly, spherical pal1Delta mutants polarize to establish a pearlike morphology before mitosis in a manner dependent on the kelch-repeat protein Tea1p and the cell cycle inhibitory kinase Wee1p. Thus, overlapping mechanisms involving Pal1p, Tea1p, and Sla2p contribute to the establishment of cylindrical cellular morphology, which is important for proper spatial regulation of cytokinesis.

Integration of multi-scale molecular modeling approaches with experiments for the in silico guided design and discovery of novel hERG-Neutral antihypertensive oxazalone and imidazolone derivatives and analysis of their potential restrictive effects on cell proliferation.

PubMed

Durdagi, Serdar; Aksoydan, Busecan; Erol, Ismail; Kantarcioglu, Isik; Ergun, Yavuz; Bulut, Gulay; Acar, Melih; Avsar, Timucin; Liapakis, George; Karageorgos, Vlasios; Salmas, Ramin E; Sergi, Barış; Alkhatib, Sara; Turan, Gizem; Yigit, Berfu Nur; Cantasir, Kutay; Kurt, Bahar; Kilic, Turker

2018-02-10

AT1 antagonists is the most recent drug class of molecules against hypertension and they mediate their actions through blocking detrimental effects of angiotensin II (A-II) when acts on type I (AT1) A-II receptor. The effects of AT1 antagonists are not limited to cardiovascular diseases. AT1 receptor blockers may be used as potential anti-cancer agents - due to the inhibition of cell proliferation stimulated by A-II. Therefore, AT1 receptors and the A-II biosynthesis mechanisms are targets for the development of new synthetic drugs and therapeutic treatment of various cardiovascular and other diseases. In this work, multi-scale molecular modeling approaches were performed and it is found that oxazolone and imidazolone derivatives reveal similar/better interaction energy profiles compared to the FDA approved sartan molecules at the binding site of the AT1 receptor. In silico-guided designed hit molecules were then synthesized and tested for their binding affinities to human AT1 receptor in radioligand binding studies, using [ 125 I-Sar 1 -Ile 8 ] AngII. Among the compounds tested, 19d and 9j molecules bound to receptor in a dose response manner and with relatively high affinities. Next, cytotoxicity and wound healing assays were performed for these hit molecules. Since hit molecule 19d led to deceleration of cell motility in all three cell lines (NIH3T3, A549, and H358) tested in this study, this molecule is investigated in further tests. In two cell lines (HUVEC and MCF-7) tested, 19d induced G2/M cell cycle arrest in a concentration dependent manner. Adherent cells detached from the plates and underwent cell death possibly due to apoptosis at 19d concentrations that induced cell cycle arrest. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Modes of Overinitiation, dnaA Gene Expression, and Inhibition of Cell Division in a Novel Cold-Sensitive hda Mutant of Escherichia coli▿

PubMed Central

Fujimitsu, Kazuyuki; Su'etsugu, Masayuki; Yamaguchi, Yoko; Mazda, Kensaku; Fu, Nisi; Kawakami, Hironori; Katayama, Tsutomu

2008-01-01

The chromosomal replication cycle is strictly coordinated with cell cycle progression in Escherichia coli. ATP-DnaA initiates replication, leading to loading of the DNA polymerase III holoenzyme. The DNA-loaded form of the β clamp subunit of the polymerase binds the Hda protein, which promotes ATP-DnaA hydrolysis, yielding inactive ADP-DnaA. This regulation is required to repress overinitiation. In this study, we have isolated a novel cold-sensitive hda mutant, the hda-185 mutant. The hda-185 mutant caused overinitiation of chromosomal replication at 25°C, which most likely led to blockage of replication fork progress. Consistently, the inhibition of colony formation at 25°C was suppressed by disruption of the diaA gene, an initiation stimulator. Disruption of the seqA gene, an initiation inhibitor, showed synthetic lethality with hda-185 even at 42°C. The cellular ATP-DnaA level was increased in an hda-185-dependent manner. The cellular concentrations of DnaA protein and dnaA mRNA were comparable at 25°C to those in a wild-type hda strain. We also found that multiple copies of the ribonucleotide reductase genes (nrdAB or nrdEF) or dnaB gene repressed overinitiation. The cellular levels of dATP and dCTP were elevated in cells bearing multiple copies of nrdAB. The catalytic site within NrdA was required for multicopy suppression, suggesting the importance of an active form of NrdA or elevated levels of deoxyribonucleotides in inhibition of overinitiation in the hda-185 cells. Cell division in the hda-185 mutant was inhibited at 25°C in a LexA regulon-independent manner, suggesting that overinitiation in the hda-185 mutant induced a unique division inhibition pathway. PMID:18502852

Modes of overinitiation, dnaA gene expression, and inhibition of cell division in a novel cold-sensitive hda mutant of Escherichia coli.

PubMed

Fujimitsu, Kazuyuki; Su'etsugu, Masayuki; Yamaguchi, Yoko; Mazda, Kensaku; Fu, Nisi; Kawakami, Hironori; Katayama, Tsutomu

2008-08-01

The chromosomal replication cycle is strictly coordinated with cell cycle progression in Escherichia coli. ATP-DnaA initiates replication, leading to loading of the DNA polymerase III holoenzyme. The DNA-loaded form of the beta clamp subunit of the polymerase binds the Hda protein, which promotes ATP-DnaA hydrolysis, yielding inactive ADP-DnaA. This regulation is required to repress overinitiation. In this study, we have isolated a novel cold-sensitive hda mutant, the hda-185 mutant. The hda-185 mutant caused overinitiation of chromosomal replication at 25 degrees C, which most likely led to blockage of replication fork progress. Consistently, the inhibition of colony formation at 25 degrees C was suppressed by disruption of the diaA gene, an initiation stimulator. Disruption of the seqA gene, an initiation inhibitor, showed synthetic lethality with hda-185 even at 42 degrees C. The cellular ATP-DnaA level was increased in an hda-185-dependent manner. The cellular concentrations of DnaA protein and dnaA mRNA were comparable at 25 degrees C to those in a wild-type hda strain. We also found that multiple copies of the ribonucleotide reductase genes (nrdAB or nrdEF) or dnaB gene repressed overinitiation. The cellular levels of dATP and dCTP were elevated in cells bearing multiple copies of nrdAB. The catalytic site within NrdA was required for multicopy suppression, suggesting the importance of an active form of NrdA or elevated levels of deoxyribonucleotides in inhibition of overinitiation in the hda-185 cells. Cell division in the hda-185 mutant was inhibited at 25 degrees C in a LexA regulon-independent manner, suggesting that overinitiation in the hda-185 mutant induced a unique division inhibition pathway.

Iodine-131 treatment of thyroid cancer cells leads to suppression of cell proliferation followed by induction of cell apoptosis and cell cycle arrest by regulation of B-cell translocation gene 2-mediated JNK/NF-κB pathways.

PubMed

Zhao, L M; Pang, A X

2017-01-16

Iodine-131 (131I) is widely used for the treatment of thyroid-related diseases. This study aimed to investigate the expression of p53 and BTG2 genes following 131I therapy in thyroid cancer cell line SW579 and the possible underlying mechanism. SW579 human thyroid squamous carcinoma cells were cultured and treated with 131I. They were then assessed for 131I uptake, cell viability, apoptosis, cell cycle arrest, p53 expression, and BTG2 gene expression. SW579 cells were transfected with BTG2 siRNA, p53 siRNA and siNC and were then examined for the same aforementioned parameters. When treated with a JNK inhibitor of SP600125 and 131I or with a NF-κB inhibitor of BMS-345541 and 131I, non-transfected SW579 cells were assessed in JNK/NFκB pathways. It was observed that 131I significantly inhibited cell proliferation, promoted cell apoptosis and cell cycle arrest. Both BTG2 and p53 expression were enhanced in a dose-dependent manner. An increase in cell viability by up-regulation in Bcl2 gene, a decrease in apoptosis by enhanced CDK2 gene expression and a decrease in cell cycle arrest at G0/G1 phase were also observed in SW579 cell lines transfected with silenced BTG2 gene. When treated with SP600125 and 131I, the non-transfected SW579 cell lines significantly inhibited JNK pathway, NF-κB pathway and the expression of BTG2. However, when treated with BMS-345541 and 131I, only the NF-κB pathway was suppressed. 131I suppressed cell proliferation, induced cell apoptosis, and promoted cell cycle arrest of thyroid cancer cells by up-regulating B-cell translocation gene 2-mediated activation of JNK/NF-κB pathways.

Sodium ascorbate inhibits growth via the induction of cell cycle arrest and apoptosis in human malignant melanoma A375.S2 cells.

PubMed

Lin, Shuw-Yuan; Lai, Wan-Wen; Chou, Chi-Chung; Kuo, Hsiu-Maan; Li, Te-Mao; Chung, Jing-Gung; Yang, Jen-Hung

2006-12-01

Vitamin C has been reported to be useful in the treatment and prevention of cancer. Inconsistent effects from growth stimulation to induction of apoptosis of malignant tumor cells, however, have been reported. Melanoma is an increasingly common and potentially lethal malignancy. It was reported that melanoma cells were more susceptible to ascorbate toxicity than any other tumor cells. The mechanisms accounting for ascorbate-induced apoptosis in human melanoma cells, however, have remained unclear. This study was undertaken to investigate the effect of sodium ascorbate on cytotoxicity and apoptosis in human malignant melanoma A375.S2 cells. A375.S2 cells were incubated with a certain range of concentrations of sodium ascorbate for various time periods. In order to examine the effects of sodium ascorbate on cell proliferation, cell cycle, apoptosis and necrosis, we performed 4,6-diamidino-2-phenylindole dihydrochloride assays and flow cytometry analysis. Polymerase chain reaction was used to examine the mRNA levels of p53, p21, p27, cyclin A, cyclin E, CDK2 and CDK4, which are associated with cell cycle S-phase arrest and apoptosis. Flow cytometric analysis showed that sodium ascorbate significantly induced cell cycle arrest and apoptosis in the A375.S2 cell line in a dose-dependent manner. The increased expressions of p53 and p21, and the decreased expressions of cyclin A, cyclin E, CDK2 and CDK4, indicated the cell cycle arrest at G1/S phase after the cells had been treated with sodium ascorbate. Induction of apoptosis involved an increase in the levels of p53, p21 and cellular Ca, and a decrease in mitochondrial membrane potential and activation of caspase 3 before culminating in apoptosis in sodium ascorbate-treated A375.S2 cells.

Inhibition of Aurora A Kinase by Alisertib Induces Autophagy and Cell Cycle Arrest and Increases Chemosensitivity in Human Hepatocellular Carcinoma HepG2 Cells.

PubMed

Zhu, Qiaohua; Yu, Xinfa; Zhou, Zhi-Wei; Zhou, Chengyu; Chen, Xiao-Wu; Zhou, Shu-Feng

2017-01-01

Aurora A kinase represent a feasible target in cancer therapy. To evaluate the proteomic response of human liver carcinoma cells to alisertib (ALS) and identify the molecular targets of ALS, we examined the effects of ALS on the proliferation, cell cycle, autophagy, apoptosis, and chemosensitivity in HepG2 cells. The stable-isotope labeling by amino acids in cell culture (SILAC) based quantitative proteomic study was performed to evaluate the proteomic response to ALS. Cell cycle distribution and apoptosis were assessed using flow cytometry and autophagy was determined using flow cytometry and confocal microscopy. Our SILAC proteomic study showed that ALS regulated the expression of 914 proteins, with 407 molecules being up-regulated and 507 molecules being down-regulated in HepG2 cells. Ingenuity pathway analysis (IPA) and KEGG pathway analysis identified 146 and 32 signaling pathways were regulated by ALS, respectively, which were associated with cell survival, programmed cell death, and nutrition-energy metabolism. Subsequently, the verification experiments showed that ALS remarkably arrested HepG2 cells in G2/M phase and led to an accumulation of aneuploidy via regulating the expression of key cell cycle regulators. ALS induced a marked autophagy in a concentration- and time-dependent manner via the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway. Autophagy inhibition promoted the pro-apoptotic effect of ALS, indicating a cyto-protective role of ALS-induced autophagy. ALS increased the chemosensitivity of HepG2 cells to cisplatin and doxorubicin. Taken together, ALS induces autophagy and cell cycle arrest in HepG2 cells via PI3K/Akt/mTOR-mediated pathway. Autophagy inhibition may promote the anticancer effect of ALS and sensitize the chemotherapy in HepG2 cells. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Ulex europaeus agglutinin II (UEA-II) is a novel, potent inhibitor of complement activation.

PubMed

Lekowski, R; Collard, C D; Reenstra, W R; Stahl, G L

2001-02-01

Complement is an important mediator of vascular injury following oxidative stress. We recently demonstrated that complement activation following endothelial oxidative stress is mediated by mannose-binding lectin (MBL) and activation of the lectin complement pathway. Here, we investigated whether nine plant lectins which have a binding profile similar to that of MBL competitively inhibit MBL deposition and subsequent complement activation following human umbilical vein endothelial cell (HUVEC) oxidative stress. HUVEC oxidative stress (1% O(2), 24 hr) significantly increased Ulex europaeus agglutinin II (UEA-II) binding by 72 +/- 9% compared to normoxic cells. UEA-II inhibited MBL binding to HUVEC in a concentration-dependent manner following oxidative stress. Further, MBL inhibited UEA-II binding to HUVEC in a concentration-dependent manner following oxidative stress, suggesting a common ligand. UEA-II (< or = 100 micromol/L) did not attenuate the hemolytic activity, nor did it inhibit C3a des Arg formation from alternative or classical complement pathway-specific hemolytic assays. C3 deposition (measured by ELISA) following HUVEC oxidative stress was inhibited by UEA-II in a concentration-dependent manner (IC(50) = 10 pmol/L). UEA-II inhibited C3 and MBL co-localization (confocal microscopy) in a concentration-dependent manner on HUVEC following oxidative stress (IC(50) approximately 1 pmol/L). Finally, UEA-II significantly inhibited complement-dependent neutrophil chemotaxis, but failed to inhibit fMLP-mediated chemotaxis, following endothelial oxidative stress. These data demonstrate that UEA-II is a novel, potent inhibitor of human MBL deposition and complement activation following human endothelial oxidative stress.

Ulex europaeus agglutinin II (UEA-II) is a novel, potent inhibitor of complement activation

PubMed Central

Lekowski, Robert; Collard, Charles D.; Reenstra, Wende R.; Stahl, Gregory L.

2001-01-01

Complement is an important mediator of vascular injury following oxidative stress. We recently demonstrated that complement activation following endothelial oxidative stress is mediated by mannose-binding lectin (MBL) and activation of the lectin complement pathway. Here, we investigated whether nine plant lectins which have a binding profile similar to that of MBL competitively inhibit MBL deposition and subsequent complement activation following human umbilical vein endothelial cell (HUVEC) oxidative stress. HUVEC oxidative stress (1% O2, 24 hr) significantly increased Ulex europaeus agglutinin II (UEA-II) binding by 72 ± 9% compared to normoxic cells. UEA-II inhibited MBL binding to HUVEC in a concentration-dependent manner following oxidative stress. Further, MBL inhibited UEA-II binding to HUVEC in a concentration-dependent manner following oxidative stress, suggesting a common ligand. UEA-II (≤ 100 μmol/L) did not attenuate the hemolytic activity, nor did it inhibit C3a des Arg formation from alternative or classical complement pathway-specific hemolytic assays. C3 deposition (measured by ELISA) following HUVEC oxidative stress was inhibited by UEA-II in a concentration-dependent manner (IC50 = 10 pmol/L). UEA-II inhibited C3 and MBL co-localization (confocal microscopy) in a concentration-dependent manner on HUVEC following oxidative stress (IC50 ≈ 1 pmol/L). Finally, UEA-II significantly inhibited complement-dependent neutrophil chemotaxis, but failed to inhibit fMLP-mediated chemotaxis, following endothelial oxidative stress. These data demonstrate that UEA-II is a novel, potent inhibitor of human MBL deposition and complement activation following human endothelial oxidative stress. PMID:11266613

ARID3B: a Novel Regulator of the Kaposi's Sarcoma-Associated Herpesvirus Lytic Cycle

PubMed Central

Wood, Jennifer J.; Boyne, James R.; Paulus, Christina; Jackson, Brian R.; Nevels, Michael M.

2016-01-01

ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of commonly fatal malignancies of immunocompromised individuals, including primary effusion lymphoma (PEL) and Kaposi's sarcoma (KS). A hallmark of all herpesviruses is their biphasic life cycle—viral latency and the productive lytic cycle—and it is well established that reactivation of the KSHV lytic cycle is associated with KS pathogenesis. Therefore, a thorough appreciation of the mechanisms that govern reactivation is required to better understand disease progression. The viral protein replication and transcription activator (RTA) is the KSHV lytic switch protein due to its ability to drive the expression of various lytic genes, leading to reactivation of the entire lytic cycle. While the mechanisms for activating lytic gene expression have received much attention, how RTA impacts cellular function is less well understood. To address this, we developed a cell line with doxycycline-inducible RTA expression and applied stable isotope labeling of amino acids in cell culture (SILAC)-based quantitative proteomics. Using this methodology, we have identified a novel cellular protein (AT-rich interacting domain containing 3B [ARID3B]) whose expression was enhanced by RTA and that relocalized to replication compartments upon lytic reactivation. We also show that small interfering RNA (siRNA) knockdown or overexpression of ARID3B led to an enhancement or inhibition of lytic reactivation, respectively. Furthermore, DNA affinity and chromatin immunoprecipitation assays demonstrated that ARID3B specifically interacts with A/T-rich elements in the KSHV origin of lytic replication (oriLyt), and this was dependent on lytic cycle reactivation. Therefore, we have identified a novel cellular protein whose expression is enhanced by KSHV RTA with the ability to inhibit KSHV reactivation. IMPORTANCE Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of fatal malignancies of immunocompromised individuals, including Kaposi's sarcoma (KS). Herpesviruses are able to establish a latent infection, in which they escape immune detection by restricting viral gene expression. Importantly, however, reactivation of productive viral replication (the lytic cycle) is necessary for the pathogenesis of KS. Therefore, it is important that we comprehensively understand the mechanisms that govern lytic reactivation, to better understand disease progression. In this study, we have identified a novel cellular protein (AT-rich interacting domain protein 3B [ARID3B]) that we show is able to temper lytic reactivation. We showed that the master lytic switch protein, RTA, enhanced ARID3B levels, which then interacted with viral DNA in a lytic cycle-dependent manner. Therefore, we have added a new factor to the list of cellular proteins that regulate the KSHV lytic cycle, which has implications for our understanding of KSHV biology. PMID:27512077

Pan-Cancer Analysis Links PARK2 to BCL-XL-Dependent Control of Apoptosis.

PubMed

Gong, Yongxing; Schumacher, Steven E; Wu, Wei H; Tang, Fanying; Beroukhim, Rameen; Chan, Timothy A

2017-02-01

Mutation of the PARK2 gene can promote both Parkinson's Disease and cancer, yet the underlying mechanisms of how PARK2 controls cellular physiology is incompletely understood. Here, we show that the PARK2 tumor suppressor controls the apoptotic regulator BCL-XL and modulates programmed cell death. Analysis of approximately 10,000 tumor genomes uncovers a striking pattern of mutual exclusivity between PARK2 genetic loss and amplification of BCL2L1, implicating these genes in a common pathway. PARK2 directly binds to and ubiquitinates BCL-XL. Inactivation of PARK2 leads to aberrant accumulation of BCL-XL both in vitro and in vivo, and cancer-specific mutations in PARK2 abrogate the ability of the ubiquitin E3 ligase to target BCL-XL for degradation. Furthermore, PARK2 modulates mitochondrial depolarization and apoptosis in a BCL-XL-dependent manner. Thus, like genes at the nodal points of growth arrest pathways such as p53, the PARK2 tumor suppressor is able to exert its antiproliferative effects by regulating both cell cycle progression and programmed cell death. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Alpha-ketoglutarate dehydrogenase complex-dependent succinylation of proteins in neurons and neuronal cell lines

PubMed Central

Gibson, Gary E.; Xu, Hui; Chen, Huan-Lian; Chen, Wei; Denton, Travis; Zhang, Sheng

2015-01-01

Reversible post-translation modifications of proteins are common in all cells and appear to regulate many processes. Nevertheless, the enzyme(s) responsible for the alterations and the significance of the modification are largely unknown. Succinylation of proteins occurs and causes large changes in the structure of proteins; however, the source of the succinyl groups, the targets, and the consequences of these modifications on other proteins are unknown. These studies focused on succinylation of mitochondrial proteins. The results demonstrate that the α-ketoglutarate dehydrogenase complex (KGDHC) can serve as a trans-succinylase that mediates succinylation in an α-ketoglutarate-dependent manner. Inhibition of KGDHC reduced suc-cinylation of both cytosolic and mitochondrial proteins in cultured neurons and in a neuronal cell line. Purified KGDHC can succinylate multiple proteins including other enzymes of the tricarboxylic acid (TCA) cycle leading to modification of their activity. Inhibition of KGDHC also modifies acetylation by modifying the pyruvate dehydrogenase complex. The much greater effectiveness of KGDHC than succinyl CoA suggests that the catalysis due to the E2k suc-cinyltransferase is important. Succinylation appears to be a major signaling system and it can be mediated by KGDHC. PMID:25772995

Activation of Intestinal Human Pregnane X Receptor Protects against Azoxymethane/Dextran Sulfate Sodium–Induced Colon Cancer

PubMed Central

Cheng, Jie; Fang, Zhong-Ze; Nagaoka, Kenjiro; Okamoto, Minoru; Qu, Aijuan; Tanaka, Naoki; Kimura, Shioko

2014-01-01

The role of intestinal human pregnane X receptor (PXR) in colon cancer was determined through investigation of the chemopreventive role of rifaximin, a specific agonist of intestinal human PXR, toward azoxymethane (AOM)/dextran sulfate sodium (DSS)–induced colon cancer. Rifaximin treatment significantly decreased the number of colon tumors induced by AOM/DSS treatment in PXR-humanized mice, but not wild-type or Pxr-null mice. Additionally, rifaximin treatment markedly increased the survival rate of PXR-humanized mice, but not wild-type or Pxr-null mice. These data indicated a human PXR–dependent therapeutic chemoprevention of rifaximin toward AOM/DSS-induced colon cancer. Nuclear factor κ-light-chain-enhancer of activated B cells–mediated inflammatory signaling was upregulated in AOM/DSS-treated mice, and inhibited by rifaximin in PXR-humanized mice. Cell proliferation and apoptosis were also modulated by rifaximin treatment in the AOM/DSS model. In vitro cell-based assays further revealed that rifaximin regulated cell apoptosis and cell cycle in a human PXR-dependent manner. These results suggested that specific activation of intestinal human PXR exhibited a chemopreventive role toward AOM/DSS-induced colon cancer by mediating anti-inflammation, antiproliferation, and proapoptotic events. PMID:25277138

Hydra as a model organism to decipher the toxic effects of copper oxide nanorod: Eco-toxicogenomics approach.

PubMed

Murugadas, Anbazhagan; Zeeshan, Mohammed; Thamaraiselvi, Kaliannan; Ghaskadbi, Surendra; Akbarsha, Mohammad Abdulkader

2016-07-15

Nanotechnology has emerged as a powerful field of applied research. However, the potential toxicity of nano-materials is a cause of concern. A thorough toxicological investigation is required before a nanomaterial is evaluated for application of any kind. In this context, there is concerted effort to find appropriate test systems to assess the toxicity of nanomaterials. Toxicity of a nanomaterial greatly depends on its physicochemical properties and the biological system with which it interacts. The present research was carried out with a view to generate data on eco-toxicological impacts of copper oxide nanorod (CuO NR) in Hydra magnipapillata 105 at organismal, cellular and molecular levels. Exposure of hydra to CuO NR resulted in severe morphological alterations in a concentration- as well as duration-dependent manner. Impairment of feeding, population growth, and regeneration was also observed. In vivo and in vitro analyses revealed induction of oxidative stress, genotoxicity, and molecular machinery of apoptotic cell death, accompanied by disruption of cell cycle progression. Taken together, CuO nanorod is potentially toxic to the biological systems. Also, hydra offers potential to be used as a convenient model organism for aquatic ecotoxicological risk assessment of nanomaterials.

Hydra as a model organism to decipher the toxic effects of copper oxide nanorod: Eco-toxicogenomics approach

PubMed Central

Murugadas, Anbazhagan; Zeeshan, Mohammed; Thamaraiselvi, Kaliannan; Ghaskadbi, Surendra; Akbarsha, Mohammad Abdulkader

2016-01-01

Nanotechnology has emerged as a powerful field of applied research. However, the potential toxicity of nano-materials is a cause of concern. A thorough toxicological investigation is required before a nanomaterial is evaluated for application of any kind. In this context, there is concerted effort to find appropriate test systems to assess the toxicity of nanomaterials. Toxicity of a nanomaterial greatly depends on its physicochemical properties and the biological system with which it interacts. The present research was carried out with a view to generate data on eco-toxicological impacts of copper oxide nanorod (CuO NR) in Hydra magnipapillata 105 at organismal, cellular and molecular levels. Exposure of hydra to CuO NR resulted in severe morphological alterations in a concentration- as well as duration-dependent manner. Impairment of feeding, population growth, and regeneration was also observed. In vivo and in vitro analyses revealed induction of oxidative stress, genotoxicity, and molecular machinery of apoptotic cell death, accompanied by disruption of cell cycle progression. Taken together, CuO nanorod is potentially toxic to the biological systems. Also, hydra offers potential to be used as a convenient model organism for aquatic ecotoxicological risk assessment of nanomaterials. PMID:27417574

Depletion of ribosomal protein L37 occurs in response to DNA damage and activates p53 through the L11/MDM2 pathway

PubMed Central

Llanos, Susana; Serrano, Manuel

2013-01-01

Perturbation of ribosomal biogenesis has recently emerged as a relevant p53-activating pathway. This pathway can be initiated by depletion of certain ribosomal proteins, which is followed by the binding and inhibition of MDM2 by a different subset of ribosomal proteins that includes L11. Here, we report that depletion of L37 leads to cell cycle arrest in a L11- and p53-dependent manner. DNA damage can initiate ribosomal stress, although little is known about the mechanisms involved. We have found that some genotoxic insults, namely UV light and cisplatin, lead to proteasomal degradation of L37 in the nucleoplasm and to the ensuing L11-dependent stabilization of p53. Moreover, ectopic L37 overexpression can attenuate the DNA damage response mediated by p53. These results support the concept that DNA damage-induced proteasomal degradation of L37 constitutes a mechanistic link between DNA damage and the ribosomal stress pathway, and is a relevant contributing signaling pathway for the activation of p53 in response to DNA damage. PMID:20935493

A Study of the Differential Effects of Eicosapentaenoic Acid (EPA) and Docosahexaenoic Acid (DHA) on Gene Expression Profiles of Stimulated Thp-1 Macrophages.

PubMed

Allam-Ndoul, Bénédicte; Guénard, Frédéric; Barbier, Olivier; Vohl, Marie-Claude

2017-04-25

Background: An appropriate intake of omega-3 ( n -3) fatty acids (FAs) such as eicosapentaenoic and docosahexaenoic acid (EPA/DHA) from marine sources is known to have anti-inflammatory effects. However, molecular mechanisms underlying their beneficial effects on health are not fully understood. The aim of the present study was to characterize gene expression profiles of THP-1 macrophages, incubated in either EPA or DHA and stimulated with lipopolysaccharide (LPS), a pro-inflammatory agent. Methods: THP-1 macrophages were incubated into 10, 50 and 75 µM of EPA or DHA for 24 h, and 100 nM of LPS was added to the culture media for 18 h. Total mRNA was extracted and gene expression examined by microarray analysis using Illumina Human HT-12 expression beadchips (Illumina). Results: Pathway analysis revealed that EPA and DHA regulate genes involved in cell cycle regulation, apoptosis, immune response and inflammation, oxidative stress and cancer pathways in a differential and dose-dependent manner. Conclusions: EPA and DHA appear to exert differential effects on gene expression in THP-1 macrophages. Specific effects of n -3 FAs on gene expression levels are also dose-dependent.

Chronobiology of crickets: a review.

PubMed

Tomioka, Kenji

2014-10-01

Crickets provide a good model for the study of mechanisms underlying circadian rhythms and photoperiodic responses. They show clear circadian rhythms in their overt behavior and the sensitivity of the visual system. Classical neurobiological studies revealed that a pair of optic lobes is the locus of the circadian clock controlling these rhythms and that the compound eye is the major photoreceptor necessary for synchronization to environmental light cycles. The two optic lobe clocks are mutually coupled through a neural pathway and the coupling regulates an output circadian waveform and a free-running period. Recent molecular studies revealed that the cricket's clock consists of cyclic expression of so-called clock genes and that the clock mechanism is featured by both Drosophila-like and mammalian-like traits. Molecular oscillation is also observed in some extra-optic lobe tissues and depends on the optic lobe clock in a tissue dependent manner. Interestingly, the clock is also involved in adaptation to seasonally changing environment. It fits its waveform to a given photoperiod and may be an indispensable part of a photoperiodic time-measurement mechanism. With adoption of modern molecular technologies, the cricket becomes a much more important and promising model animal for the study of circadian and photoperiodic biology.

SIAH1-induced p34SEI-1 polyubiquitination/degradation mediates p53 preferential vitamin C cytotoxicity.

PubMed

Lee, Soonduck; Kim, Jinsun; Jung, Samil; Li, Chengping; Yang, Young; Kim, Keun Il; Lim, Jong-Seok; Kim, Yonghwan; Cheon, Choong-Il; Lee, Myeong-Sok

2015-03-01

Vitamin C is considered as an important anticancer therapeutic agent although this view is debatable. In this study, we introduce a physiological mechanism demonstrating how vitamin C exerts anticancer activity that induces cell cycle arrest and apoptosis. Our previous and current data reveal that p53 tumor suppressor is the prerequisite factor for stronger anticancer effects of vitamin C. In addition, vitamin C-mediated cancer cell cytotoxicity appears to be achieved at least partly through the downregulation of the p34SEI-1 oncoprotein. Our previous study showed that p34SEI-1 increases the survival of various types of cancer cells by inhibiting their apoptosis. Present data suggest that vitamin C treatment decreases the p34SEI-1 expression at the protein level and therefore alleviates its anti-apoptotic activity. Of note, SIAH1, E3 ubiquitin ligase, appears to be responsible for the p34SEI-1 polyubiquitination and its subsequent degradation, which is dependent on p53. In summary, vitamin C increases cancer cell death by inducing SIAH1-mediated polyubiquitination/degradation of the p34SEI-1 oncoprotein in a p53-dependent manner.

Greatwall is essential to prevent mitotic collapse after nuclear envelope breakdown in mammals.

PubMed

Álvarez-Fernández, Mónica; Sánchez-Martínez, Ruth; Sanz-Castillo, Belén; Gan, Pei Pei; Sanz-Flores, María; Trakala, Marianna; Ruiz-Torres, Miguel; Lorca, Thierry; Castro, Anna; Malumbres, Marcos

2013-10-22

Greatwall is a protein kinase involved in the inhibition of protein phosphatase 2 (PP2A)-B55 complexes to maintain the mitotic state. Although its biochemical activity has been deeply characterized in Xenopus, its specific relevance during the progression of mitosis is not fully understood. By using a conditional knockout of the mouse ortholog, Mastl, we show here that mammalian Greatwall is essential for mouse embryonic development and cell cycle progression. Yet, Greatwall-null cells enter into mitosis with normal kinetics. However, these cells display mitotic collapse after nuclear envelope breakdown (NEB) characterized by defective chromosome condensation and prometaphase arrest. Intriguingly, Greatwall is exported from the nucleus to the cytoplasm in a CRM1-dependent manner before NEB. This export occurs after the nuclear import of cyclin B-Cdk1 complexes, requires the kinase activity of Greatwall, and is mediated by Cdk-, but not Polo-like kinase 1-dependent phosphorylation. The mitotic collapse observed in Greatwall-deficient cells is partially rescued after concomitant depletion of B55 regulatory subunits, which are mostly cytoplasmic before NEB. These data suggest that Greatwall is an essential protein in mammals required to prevent mitotic collapse after NEB.

Identification of two novel functional p53 responsive elements in the Herpes Simplex Virus-1 genome

PubMed Central

Hsieh, Jui-Cheng; Kuta, Ryan; Armour, Courtney R.; Boehmer, Paul E.

2014-01-01

Analysis of the herpes simplex virus-1 (HSV-1) genome reveals two candidate p53 responsive elements (p53RE), located in proximity to the replication origins oriL and oriS, referred to as p53RE-L and p53RE-S, respectively. The sequences of p53RE-L and p53RE-S conform to the p53 consensus site and are present in HSV-1 strains KOS, 17, and F. p53 binds to both elements in vitro and in virus-infected cells. Both p53RE-L and p53RE-S are capable of conferring p53-dependent transcriptional activation onto a heterologous reporter gene. Importantly, expression of the essential immediate early viral transactivator ICP4 and the essential DNA replication protein ICP8, that are adjacent to p53RE-S and p53RE-L, are repressed in a p53-dependent manner. Taken together, this study identifies two novel functional p53RE in the HSV-1 genome and suggests a complex mechanism of viral gene regulation by p53 which may determine progression of the lytic viral replication cycle or the establishment of latency. PMID:25010269