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Sample records for mrna inhibits proliferation

  1. Sodium arsenite-induced inhibition of cell proliferation is related to inhibition of IL-2 mRNA expression in mouse activated T cells.

    PubMed

    Conde, Patricia; Acosta-Saavedra, Leonor C; Goytia-Acevedo, Raquel C; Calderon-Aranda, Emma S

    2007-04-01

    A proposed mechanism for the As-induced inhibition of cell proliferation is the inhibition of IL-2 secretion. However, the effects of arsenite on IL-2 mRNA expression or on the ERK pathway in activated-T cells have not yet been described. We examined the effect of arsenite on IL-2 mRNA expression, cell activation and proliferation in PHA-stimulated murine lymphocytes. Arsenite (1 and 10 microM) decreased IL-2 mRNA expression, IL-2 secretion and cell proliferation. Arsenite (10 microM) strongly inhibited ERK-phosphorylation. However, the partial inhibition (50%) of IL-2 mRNA produced by 1 microM, consistent with the effects on IL-2 secretion and cell proliferation, could not be explained by the inhibition of ERK-phosphorylation, which was not affected at this concentration. The inhibition of IL-2 mRNA expression caused by 1 microM could be associated to effects on pathways located downstream or parallel to ERK. Arsenite also decreased early activation (surface CD69+ expression) in both CD4+ and CD8+, and decreased total CD8+ count without significantly affecting CD4+, supporting that the cellular immune response mediated by cytotoxic T cells is an arsenic target. Thus, our results suggest that arsenite decreases IL-2 mRNA levels and T-cell activation and proliferation. However, further studies on the effects of arsenite on IL-2 gene transcription and IL-2 mRNA stability are needed.

  2. Rapamycin inhibits human renal epithelial cell proliferation: effect on cyclin D3 mRNA expression and stability.

    PubMed

    Pallet, Nicolas; Thervet, Eric; Le Corre, Delphine; Knebelmann, Bertrand; Nusbaum, Patrick; Tomkiewicz, Céline; Meria, Paul; Flinois, Jean-Pierre; Beaune, Philippe; Legendre, Christophe; Anglicheau, Dany

    2005-06-01

    Recent data have suggested that rapamycin use during the initial period after transplantation is associated with prolonged delayed graft function (DGF). Because of the known effects of rapamycin in other cell types, we speculated that this action may be secondary to human renal epithelial cells (HRECs) inhibition of proliferation. Primary cultures of HRECs were incubated with various concentrations of rapamycin. Cell proliferation was evaluated by cytotoxicity assays. The cell cycle was analyzed by flow cytometry. Protein expression levels were assessed by Western blot. Cyclin D3 mRNA levels were measured by quantitative real-time polymerase chain reaction (PCR). The transcriptional activity of the cyclin D3 gene was evaluated using transient transfection. Rapamycin exerted a significant concentration-dependent antiproliferative effect on growing HRECs by inhibiting the G(1) to S transition. The p70(S6) kinase pathway leading to cell cycle progression was found to be active, and low concentrations of rapamycin dramatically reduced p70(S6) kinase phosphorylation. Rapamycin completely inhibited the increase in cyclin D3 protein expression and mRNA accumulation induced by fetal calf serum, but did not affect cyclin E or cdk-inhibitor expression levels. This regulation of cyclin D3 protein expression is mainly due to a destabilization of its mRNA. Rapamycin reduced the mRNA half-life by 26% (4.8 +/- 1.3 hours vs. 6.5 +/- 1.0 hours, P < 0.001). Rapamycin inhibits the proliferative response of HRECs to mitogenic stimuli, and causes cell cycle arrest in the early G(1) phase, not only by a nonspecific process due to inhibition of the p70(S6k) pathway, but also by a direct effect on cyclin D3 mRNA stability.

  3. Runx3 Expression Inhibits Proliferation and Distinctly Alters mRNA Expression of Bax in AGS and A549 Cancer Cells.

    PubMed

    Torshabi, Maryam; Faramarzi, Mohammad Ali; Tabatabaei Yazdi, Mojtaba; Ostad, Seyyed Naser; Gharemani, Mohammad Hosein

    2011-01-01

    Runx3, a member of Runt-related transcription factor (Runx) proteins with tumor suppressor effect, is a tissue-restricted and cancer related transcription factor that regulate cell proliferation and growth, as well as differentiation. In the present study, exogenous Run3 was transiently expressed in AGS (human gastric adenocarcinoma), with undetectable Runx3 protein and in A549 (human lung carcinoma) with low levels of endogenous Runx3 protein. The GFP tagged Runx3 was transfected into AGS and A549 cells using fugene6 and PolyFect and Runx3 expression was confirmed by fluorescent microscopy and RT-PCR. The effect of Runx3 transfection on cell proliferation was determined by MTT assay and the results were confirmed by the trypan blue dye exclusion method. The effect of Runx3 expression on mRNA expression of BCL2-associated X protein (Bax) was evaluated using RT-PCR. In AGS and A549 cells, Runx3 expression inhibited cell proliferation (p < 0.01). The growth inhibition was less in A549 cells. We show that Runx3 expression increases Bax mRNA expression in AGS cells when compared with control (p < 0.05), but no significant differences in mRNA expression was observed in both examined cells. Runx3 expression has antiproliferative effect in AGS cell perhaps via increase in expression of Bax. The effect of Runx3 on A549 cells' viability which has endogenous level of Runx3 is not related to Bax. These findings implicate a complex regulation by Runx3 in inhibition of cell proliferation utilizing Bax.

  4. BTG/Tob family members Tob1 and Tob2 inhibit proliferation of mouse embryonic stem cells via Id3 mRNA degradation

    SciTech Connect

    Chen, Yuanfan; Wang, Chenchen; Wu, Jenny; Li, Lingsong

    2015-07-03

    The mammalian BTG/Tob family is a group of proteins with anti-proliferative ability, and there are six members including BTG1, BTG2/PC3/Tis21, BTG3/ANA, BTG4/PC3B, Tob1/Tob and Tob2. Among them, Tob subfamily members, specifically Tob1/Tob and Tob2, have the most extensive C-terminal regions. As previously reported, overexpression of BTG/Tob proteins is associated with the inhibition of G1 to S-phase cell cycle progression and decreased cell proliferation in a variety of cell types. Tob subfamily proteins have similar anti-proliferative effects on cell cycle progression in cultured tumor cells. An important unresolved question is whether or not they have function in rapidly proliferating cells, such as embryonic stem cells (ESCs). Tob1 and Tob2 were expressed ubiquitously in mouse ESCs (mESCs), suggesting a possible role in early embryonic development and mESCs. To address the above question and explore the possible functions of the Tob subfamily in ESCs, we established ESCs from different genotypic knockout inner cell mass (ICM). We found that Tob1{sup −/−}, Tob2{sup −/−}, and Tob1/2 double knockout (DKO, Tob1{sup −/−} & Tob2{sup −/−}) ESCs grew faster than wild type (WT) ESCs without losing pluripotency, and we provide a possible mechanistic explanation for these observations: Tob1 and Tob2 inhibit the cell cycle via degradation of Id3 mRNA, which is a set of directly targeted genes of BMP4 signaling in mESCs that play critical roles in the maintenance of ESC properties. Together, our data suggest that BTG/Tob family protein Tob1 and Tob2 regulation cell proliferation does not compromise the basic properties of mESCs. - Highlights: • We established mouse Tob1/2 double knockout embryonic stem cells. • Tob1 and Tob2 inhibit the proliferation of ESCs without effect on pluripotency. • Tob1 and Tob2 involved in the degradation of Id3 in mESCs.

  5. The thyroid hormone receptor (TR) beta-selective agonist GC-1 inhibits proliferation but induces differentiation and TR beta mRNA expression in mouse and rat osteoblast-like cells.

    PubMed

    Beber, Eduardo H; Capelo, Luciane P; Fonseca, Tatiana L; Costa, Cristiane C; Lotfi, Claudimara F; Scanlan, Thomas S; Gouveia, Cecilia H A

    2009-04-01

    Previous studies showed anabolic effects of GC-1, a triiodothyronine (T3) analogue that is selective for both binding and activation functions of thyroid hormone receptor (TR) beta1 over TRalpha1, on bone tissue in vivo. The aim of this study was to investigate the responsiveness of rat (ROS17/2.8) and mouse (MC3T3-E1) osteoblast-like cells to GC-1. As expected, T3 inhibited cellular proliferation and stimulated mRNA expression of osteocalcin or alkaline phosphatase in both cell lineages. Whereas equimolar doses of T3 and GC-1 equally affected these parameters in ROS17/2.8 cells, the effects of GC-1 were more modest compared to those of T3 in MC3T3-E1 cells. Interestingly, we showed that there is higher expression of TRalpha1 than TRbeta1 mRNA in rat (approximately 20-90%) and mouse (approximately 90-98%) cell lineages and that this difference is even higher in mouse cells, which highlights the importance of TRalpha1 to bone physiology and may partially explain the modest effects of GC-1 in comparison with T3 in MC3T3-E1 cells. Nevertheless, we showed that TRbeta1 mRNA expression increases (approximately 2.8- to 4.3-fold) as osteoblastic cells undergo maturation, suggesting a key role of TRbeta1 in mediating T3 effects in the bone forming cells, especially in mature osteoblasts. It is noteworthy that T3 and GC-1 induced TRbeta1 mRNA expression to a similar extent in both cell lineages (approximately 2- to 4-fold), indicating that both ligands may modulate the responsiveness of osteoblasts to T3. Taken together, these data show that TRbeta selective T3 analogues have the potential to directly induce the differentiation and activity of osteoblasts.

  6. The pterocarpanquinone LQB-118 inhibits tumor cell proliferation by downregulation of c-Myc and cyclins D1 and B1 mRNA and upregulation of p21 cell cycle inhibitor expression.

    PubMed

    Martino, Thiago; Magalhães, Fernanda C J; Justo, Graça A; Coelho, Marsen G P; Netto, Chaquip D; Costa, Paulo R R; Sabino, Kátia C C

    2014-06-15

    The incidence of cancer grows annually worldwide and in Brazil it is the second cause of death. The search for anti-cancer drugs has then become urgent. It depends on the studies of natural and chemical synthesis products. The antitumor action of LQB-118, a pterocarpanquinone structurally related to lapachol, has been demonstrated to induce mechanisms linked to leukemia cell apoptosis. This work investigated some mechanisms of the in vitro antitumor action of LQB-118 on prostate cancer cells. LQB-118 reduced the expression of the c-Myc transcription factor, downregulated the cyclin D1 and cyclin B1 mRNA levels and upregulated the p21 cell cycle inhibitor. These effects resulted in cell cycle arrest in the S and G2/M phases and inhibition of tumor cell proliferation. LQB-118 also induced programmed cell death of the prostate cancer cells, as evidenced by internucleosomal DNA fragmentation and annexin-V positive cells. Except the cell cycle arrest in the S phase and enhanced c-Myc expression, all the mechanisms observed here for the in vitro antitumor action of LQB-118 were also found for Paclitaxel, a traditional antineoplastic drug. These findings suggest new molecular mechanisms for the LQB-118 in vitro antitumor action. Copyright © 2014 Elsevier Ltd. All rights reserved.

  7. Inhibiting macrophage proliferation suppresses atherosclerotic plaque inflammation.

    PubMed

    Tang, Jun; Lobatto, Mark E; Hassing, Laurien; van der Staay, Susanne; van Rijs, Sarian M; Calcagno, Claudia; Braza, Mounia S; Baxter, Samantha; Fay, Francois; Sanchez-Gaytan, Brenda L; Duivenvoorden, Raphaël; Sager, Hendrik; Astudillo, Yaritzy M; Leong, Wei; Ramachandran, Sarayu; Storm, Gert; Pérez-Medina, Carlos; Reiner, Thomas; Cormode, David P; Strijkers, Gustav J; Stroes, Erik S G; Swirski, Filip K; Nahrendorf, Matthias; Fisher, Edward A; Fayad, Zahi A; Mulder, Willem J M

    2015-04-01

    Inflammation drives atherosclerotic plaque progression and rupture, and is a compelling therapeutic target. Consequently, attenuating inflammation by reducing local macrophage accumulation is an appealing approach. This can potentially be accomplished by either blocking blood monocyte recruitment to the plaque or increasing macrophage apoptosis and emigration. Because macrophage proliferation was recently shown to dominate macrophage accumulation in advanced plaques, locally inhibiting macrophage proliferation may reduce plaque inflammation and produce long-term therapeutic benefits. To test this hypothesis, we used nanoparticle-based delivery of simvastatin to inhibit plaque macrophage proliferation in apolipoprotein E deficient mice (Apoe(-/-) ) with advanced atherosclerotic plaques. This resulted in rapid reduction of plaque inflammation and favorable phenotype remodeling. We then combined this short-term nanoparticle intervention with an eight-week oral statin treatment, and this regimen rapidly reduced and continuously suppressed plaque inflammation. Our results demonstrate that pharmacologically inhibiting local macrophage proliferation can effectively treat inflammation in atherosclerosis.

  8. Inhibiting macrophage proliferation suppresses atherosclerotic plaque inflammation

    PubMed Central

    Tang, Jun; Lobatto, Mark E.; Hassing, Laurien; van der Staay, Susanne; van Rijs, Sarian M.; Calcagno, Claudia; Braza, Mounia S.; Baxter, Samantha; Fay, Francois; Sanchez-Gaytan, Brenda L.; Duivenvoorden, Raphaël; Sager, Hendrik B.; Astudillo, Yaritzy M.; Leong, Wei; Ramachandran, Sarayu; Storm, Gert; Pérez-Medina, Carlos; Reiner, Thomas; Cormode, David P.; Strijkers, Gustav J.; Stroes, Erik S. G.; Swirski, Filip K.; Nahrendorf, Matthias; Fisher, Edward A.; Fayad, Zahi A.; Mulder, Willem J. M.

    2015-01-01

    Inflammation drives atherosclerotic plaque progression and rupture, and is a compelling therapeutic target. Consequently, attenuating inflammation by reducing local macrophage accumulation is an appealing approach. This can potentially be accomplished by either blocking blood monocyte recruitment to the plaque or increasing macrophage apoptosis and emigration. Because macrophage proliferation was recently shown to dominate macrophage accumulation in advanced plaques, locally inhibiting macrophage proliferation may reduce plaque inflammation and produce long-term therapeutic benefits. To test this hypothesis, we used nanoparticle-based delivery of simvastatin to inhibit plaque macrophage proliferation in apolipoprotein E–deficient mice (Apoe−/−) with advanced atherosclerotic plaques. This resulted in the rapid reduction of plaque inflammation and favorable phenotype remodeling. We then combined this short-term nanoparticle intervention with an 8-week oral statin treatment, and this regimen rapidly reduced and continuously suppressed plaque inflammation. Our results demonstrate that pharmacologically inhibiting local macrophage proliferation can effectively treat inflammation in atherosclerosis. PMID:26295063

  9. Inhibition of fibroblast proliferation by Actinobacillus actinomycetemcomitans.

    PubMed Central

    Shenker, B J; Kushner, M E; Tsai, C C

    1982-01-01

    We have examined soluble sonic extracts of Actinobacillus actinomycetemcomitans for their ability to alter human and murine fibroblast proliferation. We found that extracts of all A. actinomycetemcomitans strains examined (both leukotoxic and nonleukotoxic) caused a dose-dependent inhibition of both murine and human fibroblast proliferation as assessed by DNA synthesis ([3H]thymidine incorporation). Addition of sonic extract simultaneously with [3H]thymidine had no effect on incorporation, indicating that suppression was not due to the presence of excessive amounts of cold thymidine. Inhibition of DNA synthesis was also paralleled by decreased RNA synthesis ([3H]uridine incorporation) and by a decrease in cell growth as assessed by direct cell counts; there was no effect on cell viability. The suppressive factor(s) is heat labile; preliminary purification and characterization studies indicate that it is a distinct and separate moiety from other A. actinomycetemcomitans mediators previously reported, including leukotoxin, immune suppressive factor, and endotoxin. Although it is not clear how A. actinomycetemcomitans acts to cause disease, we propose that one aspect of the pathogenicity of this organism rests in its ability to inhibit fibroblast growth, which in turn could contribute to the collagen loss associated with certain forms of periodontal disease, in particular juvenile periodontitis. PMID:7152684

  10. Aluminum Trichloride Inhibited Osteoblastic Proliferation and Downregulated the Wnt/β-Catenin Pathway.

    PubMed

    Huang, Wanyue; Wang, Peiyan; Shen, Tongtong; Hu, Chongwei; Han, Yanfei; Song, Miao; Bian, Yu; Li, Yanfei; Zhu, Yanzhu

    2016-11-10

    Aluminum (Al) exposure inhibits bone formation. Osteoblastic proliferation promotes bone formation. Therefore, we inferred that Al may inhibit bone formation by the inhibition of osteoblastic proliferation. However, the effects and molecular mechanisms of Al on osteoblastic proliferation are still under investigation. Osteoblastic proliferation can be regulated by Wnt/β-catenin signaling pathway. To investigate the effects of Al on osteoblastic proliferation and whether Wnt/β-catenin signaling pathway is involved in it, osteoblasts from neonatal rats were cultured and exposed to 0, 0.4 mM (1/20 IC50), 0.8 mM (1/10 IC50), and 1.6 mM (1/5 IC50) of aluminum trichloride (AlCl3) for 24 h, respectively. The osteoblastic proliferation rates; Wnt3a, lipoprotein receptor-related protein 5 (LRP-5), T cell factor 1 (TCF-1), cyclin D1, and c-Myc messenger RNA (mRNA) expressions; and p-glycogen synthase kinase 3β (GSK3β), GSK3β, and β-catenin protein expressions indicated that AlCl3 inhibited osteoblastic proliferation and downregulated Wnt/β-catenin signaling pathway. In addition, the AlCl3 concentration was negatively correlated with osteoblastic proliferation rates and the mRNA expressions of Wnt3a, c-Myc, and cyclin D1, while the osteoblastic proliferation rates were positively correlated with mRNA expressions of Wnt3a, c-Myc, and cyclin D1. Taken together, these findings indicated that AlCl3 inhibits osteoblastic proliferation may be associated with the inactivation of Wnt/β-catenin signaling pathway.

  11. Inhibitory effects of antisense oligodeoxynucleotides targeting c-myc mRNA on smooth muscle cell proliferation and migration.

    PubMed Central

    Biro, S; Fu, Y M; Yu, Z X; Epstein, S E

    1993-01-01

    Smooth muscle cell (SMC) proliferation and migration play pivotal roles in restenosis following angioplasty. c-myc is an immediate early response gene induced by various mitogens, and several lines of evidence derived from experiments using transformed or hematopoietic cell lines, or transgenic mice, suggest its protein product plays a role in numerous signaling transduction pathways, including those modulating cell division. We therefore reasoned that a strategy employing oligodeoxynucleotides (ODNs) complementary to c-myc mRNA (antisense ODNs) might be potent inhibitors of SMC proliferation and, perhaps, of SMC migration. To evaluate this concept, we tested several antisense ODNs targeted to c-myc mRNA (15- or 18-mer ODNs complementary to different c-myc mRNA sequences) by introducing them individually into the medium of cultured rat aortic SMCs. Phosphoroamidate-modified ODNs were employed to retard degradation. Antisense ODNs inhibited, in a concentration-dependent manner, SMC proliferation and SMC migration. Maximal inhibitory effect was 50% for proliferation and > 90% for migration. These effects were associated with decreased SMC expression of c-myc-encoded protein by Western immunoblotting and immunocytochemical staining. ODNs with the same nucleotides but a scrambled sequence caused no effect. These results indicate that the c-myc gene product is involved in the signal transduction pathways mediating SMC proliferation and migration in the in vitro model we employed. The results also suggest a potential role of antisense strategies designed to inhibit c-myc expression for the prevention of coronary restenosis. Images PMID:8421701

  12. Mifepristone inhibits GRβ-coupled prostate cancer cell proliferation

    PubMed Central

    Ligr, Martin; Li, Yirong; Logan, Susan K.; Taneja, Semir; Melamed, Jonathan; Lepor, Hebert; Garabedian, Michael J.; Lee, Peng

    2013-01-01

    Purpose The glucocorticoid receptor (GR) gene produces GRα and GRβ isoforms by alternative splicing of a C-terminal exon. GRα binds glucocorticoids, modulates transcription in a glucocorticoid-dependent manner, and plays a growth inhibitory role in prostate cells. Due to this role, glucocorticoids are often used to treat androgen-independent prostate cancer. By contrast, GRβ possesses intrinsic transcriptional activity and binds mifepristone (RU486), but not glucocorticoids, to control gene expression. The role of GRβ in prostate cell proliferation is unknown. Materials and Methods We determined the levels of GRβ in various prostate cancer cell lines by RT-PCR and western blotting. The effect of GRβ on the kinetics of prostate cancer cell growth was determined by cell counting and flow cytometry upon mifepristone and dexamethasone treatment. Cell proliferation was also examined following siRNA-mediated knock-down and overexpression of GRβ. Results GRβ mRNA and protein were upregulated in LNCaP cells overexpressing the androgen receptor co-factor ARA70β. Treatment of LNCaP-ARA70β with mifepristone or siRNA targeting GRβ inhibited proliferation, compared to parental LNCaP cells. An immortal but non-tumorigenic (RC165) prostate cell line, as well as a tumorigenic (DU145) prostate cell line with endogenous GRβ also showed partial growth reduction upon depletion of GRβ, albeit to a lesser extent than LNCaP-ARA70β cells. The growth-stimulatory effect of ARA70β on LNCaP cells is, in part, GRβ-dependent, as is the proliferation of RC165 and, to a lesser extent, DU145 cells. Conclusions These results suggest that patients whose primary tumors express GRβ and ARA70β may benefit from mifepristone treatment. PMID:22819113

  13. SAMHD1 is down regulated in lung cancer by methylation and inhibits tumor cell proliferation

    SciTech Connect

    Wang, Jia-lei; Lu, Fan-zhen; Shen, Xiao-Yong; Wu, Yun; Zhao, Li-ting

    2014-12-12

    Highlights: • SAMHD1 expression level is down regulated in lung adenocarcinoma. • The promoter of SAMHD1 is methylated in lung adenocarcinoma. • Over expression of SAMHD1 inhibits the proliferation of lung cancer cells. - Abstract: The function of dNTP hydrolase SAMHD1 as a viral restriction factor to inhibit the replication of several viruses in human immune cells was well established. However, its regulation and function in lung cancer have been elusive. Here, we report that SAMHD1 is down regulated both on protein and mRNA levels in lung adenocarcinoma compared to adjacent normal tissue. We also found that SAMHD1 promoter is highly methylated in lung adenocarcinoma, which may inhibit its gene expression. Furthermore, over expression of the SAMHD1 reduces dNTP level and inhibits the proliferation of lung tumor cells. These results reveal the regulation and function of SAMHD1 in lung cancer, which is important for the proliferation of lung tumor cells.

  14. CCL5 promotes proliferation of MCF-7 cells through mTOR-dependent mRNA translation

    SciTech Connect

    Murooka, Thomas T.; Rahbar, Ramtin; Fish, Eleanor N.

    2009-09-18

    The proliferative capacity of cancer cells is regulated by factors intrinsic to cancer cells and by secreted factors in the microenvironment. Here, we investigated the proto-oncogenic potential of the chemokine receptor, CCR5, in MCF-7 breast cancer cell lines. At physiological levels, CCL5, a ligand for CCR5, enhanced MCF-7.CCR5 proliferation. Treatment with the mTOR inhibitor, rapamycin, inhibited this CCL5-inducible proliferation. Because mTOR directly modulates mRNA translation, we investigated whether CCL5 activation of CCR5 leads to increased translation. CCL5 induced the formation of the eIF4F translation initiation complex through an mTOR-dependent process. Indeed, CCL5 initiated mRNA translation, shown by an increase in high-molecular-weight polysomes. Specifically, we show that CCL5 mediated a rapid up-regulation of protein expression for cyclin D1, c-Myc and Dad-1, without affecting their mRNA levels. Taken together, we describe a mechanism by which CCL5 influences translation of rapamycin-sensitive mRNAs, thereby providing CCR5-positive breast cancer cells with a proliferative advantage.

  15. SIRT1 inhibits the mouse intestinal motility and epithelial proliferation

    USDA-ARS?s Scientific Manuscript database

    SIRT1 inhibits the mouse intestinal motility and epithelial proliferation. Sirtuin 1 (SIRT1), a NAD+-dependent histone deacetylase, is involved in a wide array of cellular processes, including glucose homeostasis, energy metabolism, proliferation and apoptosis, and immune response. However, it is un...

  16. Sustained endoplasmic reticulum stress inhibits hepatocyte proliferation via downregulation of c-Met expression.

    PubMed

    He, Yihuai; Long, Jun; Zhong, Weiwei; Fu, Yu; Li, Ying; Lin, Shide

    2014-04-01

    The molecular mechanisms of impaired liver regeneration in several liver diseases remain poorly understood. Endoplasmic reticulum (ER) stress has been observed in a variety of liver diseases. The aims of this study were to explore the impacts of ER stress on hepatocyte growth factor (HGF)-induced proliferation and c-Met expression in human hepatocyte L02 cells. Human hepatocyte L02 cells were incubated with thapsigargin (TG) to induce ER stress. 4-Phenylbutyric acid (PBA) was used to rescue ER stress. Activation of glucose-regulated protein 78, phosphorylation of PKR-like ER kinase and eukaryotic translation initiation factor-2α, and the expression of c-Met were determined by western blotting. The expression of c-Met mRNA was observed by reverse transcription polymerase chain reaction. L02 cell proliferation was determined by the MTS assay. L02 cell proliferation was significantly impaired in TG-treated L02 cells from 24 to 48 h, while PBA partly restored the proliferation of L02 cells. In addition, TG treatment significantly decreased the sensitivity of L02 cells to HGF-induced proliferation. PBA partly resumed the sensitivity of L02 cells to HGF-induced proliferation. The expression of c-Met protein in L02 cells was downregulated from 6 h after TG treatment, and PBA partly restored c-Met expression inhibited by TG. The expression of c-Met mRNA was also significantly downregulated from 24 to 48 h after TG treatment. Our results strongly suggest that sustained ER stress inhibits hepatocyte proliferation via downregulation of both c-Met mRNA and protein expression in human hepatocyte L02 cells.

  17. BCR/ABL mRNA targeting small interfering RNA effects on proliferation and apoptosis in chronic myeloid leukemia.

    PubMed

    Zhu, Xi-Shan; Lin, Zi-Ying; Du, Jing; Cao, Guang-Xin; Liu, Gang

    2014-01-01

    To investigate the effects of small interference RNA (siRNA) targeting BCR/ABL mRNA on proliferation and apoptosis in the K562 human chronic myeloid leukemia (CML) cell line and to provide a theoretical rationale and experimental evidence for its potential clinical application for anti-CML treatment. The gene sequence for BCR/ABL mRNA was found from the GeneBank. The target gene site on the BCR/ABL mRNA were selected according to Max-Planck-Institute (MPI) and rational siRNA design rules, the secondary structure of the candidate targeted mRNA was predicted, the relevant thermodynamic parameters were analyzed, and the targeted gene sequences were compared with BLAST to eliminate any sequences with significant homology. Inhibition of proliferation was evaluated by MTT assay and colony-formation inhibiting test. Apoptosis was determined by flow cytometry (FCM) and the morphology of apoptotic cells was identified by Giemsa-Wright staining. Western blotting was used to analyze the expression of BCR/ABL fusion protein in K562 cells after siRNA treatment. The mRNA local secondary structure calculated by RNA structure software, and the optimal design of specific siRNA were contributed by bioinformatics rules. Five sequences of BCR/ABL siRNAs were designed and synthesized in vitro. Three sequences, siRNA1384, siRNA1276 and siRNA1786, which showed the most effective inhibition of K562 cell growth, were identified among the five candidate siRNAs, with a cell proliferative inhibitory rate nearly 50% after exposure to 12.5 nmol/L~50 nmol/L siRNA1384 for 24,48 and 72 hours. The 50% inhibitory concentrations (IC50) of siRNA1384, siRNA1276 and siRNA1786 for 24 hours were 46.6 nmol/L, 59.3 nmol/L and 62.6 nmol/L, respectively, and 65.668 nmol/L, 76.6 nmol/L, 74.4 nmol/L for 72 hours. The colony-formation inhibiting test also indicated that, compared with control, cell growth of siRNA treated group was inhibited. FCM results showed that the rate of cell apoptosis increased 24 hours

  18. Triptolide inhibits COX-2 expression by regulating mRNA stability in TNF-{alpha}-treated A549 cells

    SciTech Connect

    Sun, Lixin; Zhang, Shuang; Jiang, Zhenzhou; Huang, Xin; Wang, Tao; Huang, Xiao; Li, Han; Zhang, Luyong

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer Triptolide inhibited COX-2 expression and the half-life of COX-2 mRNA is decreased. Black-Right-Pointing-Pointer The HuR protein shuttling from nucleus to cytoplasm is inhibited by triptolide. Black-Right-Pointing-Pointer Triptolide inhibited 3 Prime -UTR fluorescence reporter gene activity. Black-Right-Pointing-Pointer COX-2 mRNA binding to HuR is decreased by triptolide in pull-down experiments. -- Abstract: Cyclooxygenase-2 (COX-2) over-expression is frequently associated with human non-small-cell lung cancer (NSCLC) and involved in tumor proliferation, invasion, angiogenesis and resistance to apoptosis. In the present study, the effects of triptolide on COX-2 expression in A549 cells were investigated and triptolide was found to inhibit TNF-{alpha}-induced COX-2 expression. In our further studies, it was found that triptolide decreased the half-life of COX-2 mRNA dramatically and that it inhibited 3 Prime -untranslated region (3 Prime -UTR) fluorescence reporter gene activity. Meanwhile, triptolide inhibited the HuR shuttling from nucleus to cytoplasm. After triptolide treatment, decreased COX-2 mRNA in pull-down experiments with anti-HuR antibodies was observed, indicating that the decreased cytoplasmic HuR is responsible for the decreased COX-2 mRNA. Taken together, our results provided evidence for the first time that triptolide inhibited COX-2 expression by COX-2 mRNA stability modulation and post-transcriptional regulation. These results provide a novel mechanism of action for triptolide which may be important in the treatment of lung cancer.

  19. Blue light inhibits proliferation of melanoma cells

    NASA Astrophysics Data System (ADS)

    Becker, Anja; Distler, Elisabeth; Klapczynski, Anna; Arpino, Fabiola; Kuch, Natalia; Simon-Keller, Katja; Sticht, Carsten; van Abeelen, Frank A.; Gretz, Norbert; Oversluizen, Gerrit

    2016-03-01

    Photobiomodulation with blue light is used for several treatment paradigms such as neonatal jaundice, psoriasis and back pain. However, little is known about possible side effects concerning melanoma cells in the skin. The aim of this study was to assess the safety of blue LED irradiation with respect to proliferation of melanoma cells. For that purpose we used the human malignant melanoma cell line SK-MEL28. Cell proliferation was decreased in blue light irradiated cells where the effect size depended on light irradiation dosage. Furthermore, with a repeated irradiation of the melanoma cells on two consecutive days the effect could be intensified. Fluorescence-activated cell sorting with Annexin V and Propidium iodide labeling did not show a higher number of dead cells after blue light irradiation compared to non-irradiated cells. Gene expression analysis revealed down-regulated genes in pathways connected to anti-inflammatory response, like B cell signaling and phagosome. Most prominent pathways with up-regulation of genes were cytochrome P450, steroid hormone biosynthesis. Furthermore, even though cells showed a decrease in proliferation, genes connected to the cell cycle were up-regulated after 24h. This result is concordant with XTT test 48h after irradiation, where irradiated cells showed the same proliferation as the no light negative control. In summary, proliferation of melanoma cells can be decreased using blue light irradiation. Nevertheless, the gene expression analysis has to be further evaluated and more studies, such as in-vivo experiments, are warranted to further assess the safety of blue light treatment.

  20. Platelet-derived growth factor activity and mRNA expression in healing vascular grafts in baboons. Association in vivo of platelet-derived growth factor mRNA and protein with cellular proliferation.

    PubMed Central

    Golden, M A; Au, Y P; Kirkman, T R; Wilcox, J N; Raines, E W; Ross, R; Clowes, A W

    1991-01-01

    In a baboon graft model of arterial intimal thickening, smooth muscle cells (SMC) have been observed to proliferate underneath an intact monolayer of endothelium and in the absence of platelet adherence. Because platelets are not present and therefore cannot be a major source of growth stimulus, we have proposed that the vascular wall cells in the graft intima express mitogens and regulate SMC proliferation. To test this hypothesis, we assayed the grafts for mitogenic activity and expression of growth factor genes. Segments of healing graft and of normal artery, when perfused ex vivo, released mitogenic activity into the perfusate. The graft released more mitogen than the normal arterial segment, and some of the activity was inhibitable with an antibody to human platelet-derived growth factor (PDGF). In addition, Northern analysis of total RNA demonstrated higher expression of PDGF-A chain mRNA in the graft intima compared to normal artery. PDGF-B chain mRNA was barely detectable in both tissues. PDGF mRNA levels within the graft interstices were not measured. In situ hybridization of 7.5- or 12-wk grafts indicated that some luminal endothelial cells and adjacent intimal SMC contained PDGF-A chain mRNA. By thymidine autoradiography, intimal SMC were observed to be proliferating in the inner third of the intima. These data demonstrate a difference in the pattern of PDGF transcript expression and luminal perfusate activity in graft as compared with control arteries. The association of intimal smooth muscle cell proliferation with intimal PDGF mRNA expression and release of PDGF-like protein supports the hypothesis that factors from cells that have grown into the graft or populated its surface rather than platelets may regulate intimal smooth muscle cell proliferation in this model. Images PMID:1825089

  1. Free fatty acid receptor (FFAR) agonists inhibit proliferation of human ovarian cancer cells.

    PubMed

    Hopkins, Mandi M; Meier, Kathryn E

    2017-07-01

    Many cellular actions of omega-3 fatty acids are mediated by two G protein-coupled receptors, FFA1 and FFA4, free fatty acid receptor (FFAR) family members that are activated by these dietary constituents. FFAR agonists inhibit proliferation of human prostate and breast cancer cells. Since omega-3 fatty acids can inhibit ovarian cancer cell growth, the current study tested the potential role of FFARs in the response. OVCAR3 and SKOV3 human ovarian cancer cell lines express mRNA for FFA1; FFA4 mRNA was detected at low levels in SKOV3 but not OVCAR3. Lysophosphatidic acid (LPA) and epidermal growth factor (EGF) stimulated proliferation of both cell lines; these responses were inhibited by eicosopentaneoic acid (EPA) and by GW9508, a synthetic FFAR agonist. The LPA antagonist Ki16425 also inhibited LPA- and EGF-induced proliferation; FFAR agonists had no further effect when added with Ki16425. The results suggest that FFARs are potential targets for ovarian cancer therapy. Copyright © 2017. Published by Elsevier Ltd.

  2. The endogenous cannabinoid anandamide inhibits human breast cancer cell proliferation

    PubMed Central

    De Petrocellis, Luciano; Melck, Dominique; Palmisano, Antonella; Bisogno, Tiziana; Laezza, Chiara; Bifulco, Maurizio; Di Marzo, Vincenzo

    1998-01-01

    Anandamide was the first brain metabolite shown to act as a ligand of “central” CB1 cannabinoid receptors. Here we report that the endogenous cannabinoid potently and selectively inhibits the proliferation of human breast cancer cells in vitro. Anandamide dose-dependently inhibited the proliferation of MCF-7 and EFM-19 cells with IC50 values between 0.5 and 1.5 μM and 83–92% maximal inhibition at 5–10 μM. The proliferation of several other nonmammary tumoral cell lines was not affected by 10 μM anandamide. The anti-proliferative effect of anandamide was not due to toxicity or to apoptosis of cells but was accompanied by a reduction of cells in the S phase of the cell cycle. A stable analogue of anandamide (R)-methanandamide, another endogenous cannabinoid, 2-arachidonoylglycerol, and the synthetic cannabinoid HU-210 also inhibited EFM-19 cell proliferation, whereas arachidonic acid was much less effective. These cannabimimetic substances displaced the binding of the selective cannabinoid agonist [3H]CP 55,940 to EFM-19 membranes with an order of potency identical to that observed for the inhibition of EFM-19 cell proliferation. Moreover, anandamide cytostatic effect was inhibited by the selective CB1 receptor antagonist SR 141716A. Cell proliferation was arrested by a prolactin mAb and enhanced by exogenous human prolactin, whose mitogenic action was reverted by very low (0.1–0.5 μM) doses of anandamide. Anandamide suppressed the levels of the long form of the prolactin receptor in both EFM-19 and MCF-7 cells, as well as a typical prolactin-induced response, i.e., the expression of the breast cancer cell susceptibility gene brca1. These data suggest that anandamide blocks human breast cancer cell proliferation through CB1-like receptor-mediated inhibition of endogenous prolactin action at the level of prolactin receptor. PMID:9653194

  3. Inhibition of FHL1 inhibits cigarette smoke extract-induced proliferation in pulmonary arterial smooth muscle cells.

    PubMed

    Li, Yuping; Pu, Guimei; Chen, Chengshui; Yang, Li

    2015-09-01

    Cigarette smoke can induce pulmonary vascular remodeling, which involves pulmonary artery smooth muscle cell (PASMC) proliferation, resulting in pulmonary hypertension in chronic obstructive pulmonary disease. FHL1 is a member of the FHL subfamily, characterized by an N‑terminal half LIM domain, followed by four complete LIM domains, and has been suggested to be critical in cell proliferation. However, the effects of FHL1 on cigarette smoke‑induced PASMC proliferation and the precise molecular mechanism remain to be elucidated. The present study demonstrated that the protein expression of FHL1 correlated with cigarette smoke extract (CSE)‑induced PASMC proliferation. Knockdown of the expression of FHL1 using siRNA significantly suppressed cell proliferation and inhibited the cell cycle transition between the G1 and S phase by regulating the cyclin‑dependent kinase pathway at the basal level and following CSE stimulation. By contrast, overexpressing FHL1 using an adenovirus increased cell proliferation and promoted the cell cycle transition between the G1 and S phase. Furthermore, CSE significantly increased the protein expression of FHL1, however, exerted no effect on the mRNA expression levels. This alteration was due to the prolonged FHL1 half‑life, leading to the antagonizing of protein degradation. Collectively, these data suggested that FHL1 may be involved in excessive cell proliferation and may represent a potential therapeutic target for pulmonary hypertension.

  4. Mitophagy inhibits proliferation by decreasing cyclooxygenase-2 (COX-2) in arsenic trioxide-treated HepG2 cells.

    PubMed

    Niu, Zhidan; Zhang, Wenya; Gu, Xueyan; Zhang, Xiaoning; Qi, Yongmei; Zhang, Yingmei

    2016-07-01

    Mitochondrial damage can trigger mitophagy and eventually suppress proliferation. However, the effect of mitophagy on proliferation remains unclear. In this study, HepG2 cells were used to assess mitophagy and proliferation arrest in response to As2O3 exposure. The stimulatory effect of As2O3 on mitophagy was investigated by assessing morphology (mitophagosome and mitolysosome) and relevant proteins (PINK1, LC3 II/I, and COX IV). Additionally, the relationship of mitophagy and proliferation was explored through the use of mitophagy inhibitors (CsA, Mdivi-1). Interestingly, the inhibition of mitophagy rescued proliferation arrest by restoring COX-2 protein level and countered the elimination of mitochondria-located COX-2 and up-regulated the COX-2 mRNA level. Taken together, our findings indicated that mitophagy can be induced and can inhibit proliferation by reducing COX-2 in HepG2 cells during As2O3 treatment.

  5. 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.

  6. Nesfatin-1 inhibits ovarian epithelial carcinoma cell proliferation in vitro

    SciTech Connect

    Xu, Yang; Pang, Xiaoyan; Dong, Mei; Wen, Fang Zhang, Yi

    2013-11-01

    Highlights: •Nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest. •Nesfatin-1 enhances HO-8910 cell apoptosis. •Nesfatin-1 inhibits HO-8910 cell proliferation via mTOR and RhoA/ROCK signaling pathway. •The first report of nesfatin-1-mediated proliferation in ovarian epithelial carcinoma. -- Abstract: Nesfatin-1, an 82-amino-acid peptide derived from a 396-amino-acid precursor protein nucleobindin 2 (NUCB2), was originally identified in hypothalamic nuclei involved in the regulation of food intake. It was recently reported that nesfatin-1 is a novel depot specific adipokine preferentially produced by subcutaneous tissue, with obesity- and food deprivation-regulated expression. Although a relation between ovarian cancer mortality and obesity has been previously established, a role of nesfatin-1 in ovarian epithelial carcinoma remains unknown. The aim of the present study is to examine the effect of nesfatin-1 on ovary carcinoma cells proliferation. We found that nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest, this inhibition could be abolished by nesfatin-1 neutralizing antibody. Nesfatin-1 enhances HO-8910 cell apoptosis, activation of mammalian target of rapamycin (mTOR) and RhoA/ROCK signaling pathway block the effects of nesfatin-1-induced apoptosis, therefore reverses the inhibition of HO-8910 cell proliferation by nesfatin-1. In conclusion, the present study demonstrated that nesfatin-1 can inhibit the proliferation in human ovarian epithelial carcinoma cell line HO-8910 cells through inducing apoptosis via mTOR and RhoA/ROCK signaling pathway. This study provides a novel regulatory signaling pathway of nesfatin-1-regulated ovarian epithelial carcinoma growth and may contribute to ovarian cancer prevention and therapy, especially in obese patients.

  7. Cell proliferation inhibition in reduced gravity

    NASA Technical Reports Server (NTRS)

    Moos, P. J.; Fattaey, H. K.; Johnson, T. C.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    Extended durations of spaceflight have been shown to be deleterious on an organismic level; however, mechanisms underlying cellular sensitivity to the gravitational environment remain to be elucidated. The majority of the gravitational studies to date indicates that cell regulatory pathways may be influenced by their gravitational environment. Still, few cell biology experiments have been performed in space flight and even fewer experiments have been repeated on subsequent flights. With flight opportunities on STS-50, 54, and 57, Sf9 cells were flown in the BioServe Fluids Processing Apparatus and cell proliferation was measured with and without exposure to a cell regulatory sialoglycopeptide (CeReS) inhibitor. Results from these flights indicate that the Sf9 cells grew comparable to ground controls, that the CeReS inhibitor bound to its specific receptor, and that its signal transduction cascade was not gravity sensitive.

  8. Cell proliferation inhibition in reduced gravity

    NASA Technical Reports Server (NTRS)

    Moos, P. J.; Fattaey, H. K.; Johnson, T. C.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    Extended durations of spaceflight have been shown to be deleterious on an organismic level; however, mechanisms underlying cellular sensitivity to the gravitational environment remain to be elucidated. The majority of the gravitational studies to date indicates that cell regulatory pathways may be influenced by their gravitational environment. Still, few cell biology experiments have been performed in space flight and even fewer experiments have been repeated on subsequent flights. With flight opportunities on STS-50, 54, and 57, Sf9 cells were flown in the BioServe Fluids Processing Apparatus and cell proliferation was measured with and without exposure to a cell regulatory sialoglycopeptide (CeReS) inhibitor. Results from these flights indicate that the Sf9 cells grew comparable to ground controls, that the CeReS inhibitor bound to its specific receptor, and that its signal transduction cascade was not gravity sensitive.

  9. MiR-520b suppresses proliferation of hepatoma cells through targeting ten-eleven translocation 1 (TET1) mRNA

    SciTech Connect

    Zhang, Weiying; Lu, Zhanping; Gao, Yuen; Ye, Lihong; Song, Tianqiang; Zhang, Xiaodong

    2015-05-08

    Accumulating evidence indicates that microRNAs are able to act as oncogenes or tumor suppressor genes in human cancer. We previously reported that miR-520b was down-regulated in hepatocellular carcinoma (HCC) and its deregulation was involved in hepatocarcinogenesis. In the present study, we report that miR-520b suppresses cell proliferation in HCC through targeting the ten-eleven translocation 1 (TET1) mRNA. Notably, we identified that miR-520b was able to target 3′-untranslated region (3′UTR) of TET1 mRNA by luciferase reporter gene assays. Then, we revealed that miR-520b was able to reduce the expression of TET1 at the levels of mRNA and protein using reverse transcription-polymerase chain reaction and Western blotting analysis. In terms of function, 5-ethynyl-2-deoxyuridine (EdU) incorporation and colony formation assays demonstrated that the forced miR-520b expression remarkably inhibited proliferation of hepatoma cells, but TET1 overexpression could rescue the inhibition of cell proliferation mediated by miR-520b. Furthermore, anti-miR-520b enhanced proliferation of hepatoma cells, whereas silencing of TET1 abolished anti-miR-520b-induced acceleration of cell proliferation. Then, we validated that the expression levels of miR-520b were negatively related to those of TET1 mRNA in clinical HCC tissues. Thus, we conclude that miR-520b depresses proliferation of liver cancer cells through targeting 3′UTR of TET1 mRNA. Our finding provides new insights into the mechanism of hepatocarcinogenesis. - Highlights: • TET1 is a novel target gene of miR-520b. • TET1 is upregulated in clinical HCC tissues. • MiR-520b is negatively correlated with TET1 in clinical HCC tissues. • MiR-520b depresses the proliferation of HCC cells through targeting TET1 mRNA.

  10. Sinomenine inhibits proliferation of SGC-7901 gastric adenocarcinoma cells via suppression of cyclooxygenase-2 expression

    PubMed Central

    LV, YIFEI; LI, CHANGSHUN; LI, SHUANG; HAO, ZHIMING

    2011-01-01

    Sinomenine (SIN) is a bioactive alkaloid extracted from the Chinese medicinal plant Sinomenium acutum. Results of studies have shown that the anti-inflammatory, immunosuppressive and anti-arthritic effects of SIN are partially attributed to the inhibition of cyclooxygenase-2 (COX-2) expression. COX-2 overexpression is associated with enhanced proliferation and angiogenesis of gastric cancer (GC). SGC-7901 cells were treated with different concentrations of SIN in order to observe its effect on the proliferation of human gastric adenocarcinoma cells and to explore the potential underlying molecular mechanism via the detection of COX-2 expression. Celecoxib was used as the positive control. Morphological alterations of the cells were observed microscopically. Cell proliferation was evaluated using MTT assay. COX-2 expression was detected using semi-quantitative RT-PCR and Western blotting. The results showed that SIN inhibited the proliferation of SGC-7901 cells in a time- and dose-dependent manner. In the presence of SIN or celecoxib, SGC-7901 cells became round and detached morphologically, indicating cell apoptosis. The expression of COX-2 was inhibited by SIN in a dose-dependent manner at both the mRNA and protein levels. Our findings indicate that the protective effects of SIN are mediated through the inhibition of COX-2 expression. These findings suggest a novel therapy to treat inflammation-mediated gastric adenocarcinomata. PMID:22848259

  11. Inhibition of brain tumor cell proliferation by alternating electric fields

    SciTech Connect

    Jeong, Hyesun; Oh, Seung-ick; Hong, Sunghoi E-mail: radioyoon@korea.ac.kr; Sung, Jiwon; Jeong, Seonghoon; Yoon, Myonggeun E-mail: radioyoon@korea.ac.kr; Koh, Eui Kwan

    2014-11-17

    This study was designed to investigate the mechanism by which electric fields affect cell function, and to determine the optimal conditions for electric field inhibition of cancer cell proliferation. Low-intensity (<2 V/cm) and intermediate-frequency (100–300 kHz) alternating electric fields were applied to glioblastoma cell lines. These electric fields inhibited cell proliferation by inducing cell cycle arrest and abnormal mitosis due to the malformation of microtubules. These effects were significantly dependent on the intensity and frequency of applied electric fields.

  12. A c-myc antisense oligonucleotide inhibits human retinal pigment epithelial cell proliferation.

    PubMed

    Capeáns, C; Piñeiro, A; Domínguez, F; Loidi, L; Buceta, M; Carneiro, C; Garcia-Caballero, T; Sanchez-Salorio, M

    1998-05-01

    The purpose of this work was to investigate if MYC-dependent intracellular mitogenic pathway is active in cultures of human retinal pigment epithelial (hRPE) cells and whether myc antisense phosphorotioate oligonucleotides (c-myc-AS-ODN) are useful tools for inhibiting the proliferation of hRPE cells. Cultures of hRPE cells were established from adult human corneal donors. These cells were positively stained for cytokeratins and vimentin. Myc mRNA expression was determined by Northern blot analysis and it was determined by means of immunofluorescence if MYC was expressed. C-myc-AS-ODN effect on cell proliferation was estimated by evaluating the incorporation of 5-bromo-2'-deoxy-uridine into cellular DNA. Cell number was estimated by using a tetrazolium bromide based colorimetric method. Human RPE cells in culture expressed MYC and myc mRNA as well as prothymosin alpha mRNA--a gene whose transcription is under MYC control--indicating that MYC-dependent intracellular mitogenic pathway is active in these cells. In accordance with this, we found that blocking the expression of myc by the addition of c-myc-AS-ODN to the culture medium inhibited hRPE cell proliferation. The effect of the c-myc-AS-ODN was found to be sequence specific (the use of a control oligonucleotide with the same sequence but in an opposite direction had no effect) and dose-dependent (4 microM was the lowest effective dose tested). By using RT-PCR we found that the c-myc-AS-ODN inhibition of cell proliferation was related to a diminution in c-myc mRNA expression, and by immunofluorescence we detected a diminution in c-MYC protein staining in RPE cells after 48 hr of treatment with c-myc-AS-ODN. Furthermore, growth inhibition remained for at least 5 days after addition of a single dose of the c-myc-AS-ODN to the culture. We conclude that hRPE cell proliferation is under MYC control. Blocking the expression of myc by c-myc-AS-ODN inhibited hRPE cell proliferation. These findings establish a rationale

  13. Lipocalin 2 deficiency inhibits cell proliferation, autophagy, and mitochondrial biogenesis in mouse embryonic cells.

    PubMed

    Jin, Daozhong; Zhang, Yuanyuan; Chen, Xiaoli

    2011-05-01

    Lipocalin 2 (LCN2) has been recently implicated as a critical player in multiple cancer tumorigeneses. However, the molecular mechanisms for its tumorigenic role are poorly understood. Herein, we investigated the effects of LCN2 on cell proliferation, autophagy, and mitochondrial biogenesis in MEF cells. We observed that LCN2 deficiency significantly inhibited cell proliferation and autophagy in MEF cells. Furthermore, mitochondrial DNA content, mRNA expression levels of mitochondrial-encoded gene cytochrome oxidase 2 and PGC-1α were all markedly reduced in LCN2⁻/⁻ MEF cells. Additionally, when compared with wild-type MEF cells, LCN2⁻/⁻ MEF cells expressed significantly higher levels of IRS-1, and displayed more potent TNFα-stimulated NF-κB activation. These findings demonstrate that LCN2 is a critical regulator of cell proliferation, autophagy, and mitochondrial biogenesis.

  14. Inhibition of Pancreatic Cancer Cell Proliferation by LRH-1 Inhibitors

    DTIC Science & Technology

    2013-09-01

    AD_________________ Award Number: W81XWH-12-1-0396 TITLE: INHIBITION OF PANCREATIC CANCER CELL...DATES COVERED 15September2012–14September2013 4. TITLE AND SUBTITLE INHIBITION OF PANCREATIC CANCER CELL PROLIFERATION BY LRH-1 INHIBITORS 5a...of pancreatic cancer is devastating, with mortality rates nearing its incidence rates. To date, there are no effective targeted anti-pancreatic

  15. Mechanism of inhibition of cell proliferation by Vinca alkaloids.

    PubMed

    Jordan, M A; Thrower, D; Wilson, L

    1991-04-15

    We have used a structure-activity approach to investigate whether the Vinca alkaloids inhibit cell proliferation primarily by means of their effects on mitotic spindle microtubules or by another mechanism or by a combination of mechanisms. Five Vinca alkaloids were used to investigate the relationship in HeLa cells between inhibition of cell proliferation and blockage of mitosis, alteration of spindle organization, and depolymerization of microtubules. Indirect immunofluorescence staining of microtubules and 4,6-diamidino-2-phenylindole staining of chromatin were used to characterize the effects of the drugs on the distributions of cells in stages of the cell cycle and on the organization of microtubules and chromosomes in metaphase spindles. The microtubule polymer was isolated from cells and quantified using a competitive enzyme-linked immunoadsorbent assay for tubulin. We observed a nearly perfect coincidence between the concentration of each Vinca derivative that inhibited cell proliferation and the concentration that caused 50% accumulation of cells at metaphase, despite the fact that the antiproliferative potencies of the drugs varied over a broad concentration range. Inhibition of cell proliferation and blockage of cells at metaphase at the lowest effective concentrations of all Vinca derivatives occurred with little or no microtubule depolymerization or spindle disorganization. With increasing drug concentrations, the organization of microtubules and chromosomes in arrested mitotic spindles deteriorated in a manner that was common to all five congeners. These results indicate that the antiproliferative activity of the Vinca alkaloids at their lowest effective concentrations in HeLa cells is due to inhibition of mitotic spindle function. The results suggest further that the Vinca alkaloids inhibit cell proliferation by altering the dynamics of tubulin addition and loss at the ends of mitotic spindle microtubules rather than by depolymerizing the microtubules

  16. SIRT1 controls cell proliferation by regulating contact inhibition.

    PubMed

    Cho, Elizabeth H; Dai, Yan

    2016-09-16

    Contact inhibition keeps cell proliferation in check and serves as a built-in protection against cancer development by arresting cell division upon cell-cell contact. Yet the complete mechanism behind this anti-cancer process remains largely unclear. Here we present SIRT1 as a novel regulator of contact inhibition. SIRT1 performs a wide variety of functions in biological processes, but its involvement in contact inhibition has not been explored to date. We used NIH3T3 cells, which are sensitive to contact inhibition, and H460 and DU145 cancer cells, which lack contact inhibition, to investigate the relationship between SIRT1 and contact inhibition. We show that SIRT1 overexpression in NIH3T3 cells overcomes contact inhibition while SIRT1 knockdown in cancer cells restores their lost contact inhibition. Moreover, we demonstrate that p27 protein expression is controlled by SIRT1 in contact inhibition. Overall, our findings underline the critical role of SIRT1 in contact inhibition and suggest SIRT1 inhibition as a potential strategy to suppress cancer cell growth by restoring contact inhibition. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Poria cocos inhibits high glucose-induced proliferation of rat mesangial cells.

    PubMed

    Yoon, Jung Joo; Lee, Yun Jung; Lee, So Min; Jin, Song Nan; Kang, Dae Gill; Lee, Ho Sub

    2013-01-01

    Mesangial cell proliferation is correlated with the progression of renal failure. The purpose of this study was to determine whether a water extract of Poria cocos Wolf (WPC), a well-known medicinal plant, regulates rat mesangial cell proliferation in the presence of high glucose (HG). HG significantly accelerated [(3)H]-thymidine incorporation, which was inhibited by WPC (1-50 μg/mL) in a dose-dependent manner. Cell migration and fibronectin mRNA expression data also supported the anti-proliferative effect of WPC. Western blot analysis revealed that pretreatment with WPC decreased the expression of cyclins and cyclin-dependent kinases (CDKs) and promoted the expression of p21(waf1/cip1) and p27(kip1). WPC also suppressed HG-induced p38 mitogen-activated protein kinase (p38 MAPK) and extracellular-signal-regulated kinase 1/2 (ERK 1/2) phosphorylation. Furthermore, WPC inhibited HG-induced production of dichlorofluorescein (DCF)-sensitive intracellular reactive oxygen species (ROS). In conclusion, HG promoted mesangial cell proliferation, and WPC inhibited this activity, at least in part, via induction of cell cycle arrest and activation of anti-oxidant properties. Taken together, these results suggest that P. cocos may be a potent regulator of HG-induced proliferation.

  18. HPV16E7-specific siRNA inhibits cell proliferation in CaSki cells.

    PubMed

    Li, Jun-guo; Li, Li; Zhang, Shui-Wen; Wei, Xiaoguang

    2015-03-01

    High-risk human papilloma virus (HPV) infection is the main cause for the genesis of cervical carcinomas. After infection, E6 and E7 genes of HPV were integrated to the genome of the cervical epithelium. Continued expression of the transforming oncoproteins E6 and E7 not only drives the neoplastic progression in cervical epithelium, but also plays an important role in maintaining the malignant phenotype of cervical cancer cells. The aim of this study is to explore the effects of liposomal transfection of HPV16E7 siRNA on the proliferation of cervical carcinoma cell line CaSki. The siRNA interfering HPV16E7 gene was synthesized and transfected into CaSki cells by liposome to observe the cell morphology changes under microscope. The cell proliferation index was detected by flow cytometry; HPV16E7 mRNA expression was determined by RT-PCR and its protein level was determined by Western blot. After transfection of the CaSki cell by siRNA, cell proliferation was inhibited significantly, and the expression of HPV16E7 mRNA and protein level of HPV16E7 decreased. HPV16E7 siRNA is able to inhibit growth of CaSki cells. HPV16E7 might become a new target for genetic therapy of cervical carcinoma.

  19. Metformin inhibits the proliferation of benign prostatic epithelial cells

    PubMed Central

    Ge, Rongbin; Li, Jijun; Johnson, Cameron W.; Rassoulian, Cyrus; Olumi, Aria F.

    2017-01-01

    Objective Benign prostatic hyperplasia (BPH) is the most common proliferative abnormality of the prostate affecting elderly men throughout the world. Epidemiologic studies have shown that diabetes significantly increases the risk of developing BPH, although whether anti-diabetic medications preventing the development of BPH remains to be defined. We have previously found that stromally expressed insulin-like growth factor 1 (IGF-1) promotes benign prostatic epithelial cell proliferation through paracrine mechanisms. Here, we seek to understand if metformin, a first line medication for the treatment of type 2 diabetes, inhibits the proliferation of benign prostatic epithelial cells through reducing the expression of IGF-1 receptor (IGF-1R) and regulating cell cycle. Methods BPE cell lines BPH-1 and P69, murine fibroblasts3T3 and primary human prostatic fibroblasts were cultured and tested in this study. Cell proliferation and the cell cycle were analyzed by MTS assay and flow cytometry, respectively. The expression of IGF-1R was determined by western-blot and immunocytochemistry. The level of IGF-1 secretion in culture medium was measured by ELISA. Results Metformin (0.5-10mM, 6-48h) significantly inhibited the proliferation of BPH-1 and P69 cells in a dose-dependent and time-dependent manner. Treatment with metformin for 24 hours lowered the G2/M cell population by 43.24% in P69 cells and 24.22% in BPH-1 cells. On the other hand, IGF-1 (100ng/mL, 24h) stimulated the cell proliferation (increased by 28.81% in P69 cells and 20.95% in BPH-1 cells) and significantly enhanced the expression of IGF-1R in benign prostatic epithelial cells. Metformin (5mM) abrogated the proliferation of benign prostatic epithelial cells induced by IGF-1. In 3T3 cells, the secretion of IGF-1 was significantly inhibited by metformin from 574.31pg/ml to 197.61pg/ml. The conditioned media of 3T3 cells and human prostatic fibroblasts promoted the proliferation of epithelial cells and the

  20. Inhibition of cyclooxygenase (COX)-2 affects endothelial progenitor cell proliferation

    SciTech Connect

    Colleselli, Daniela; Bijuklic, Klaudija; Mosheimer, Birgit A.; Kaehler, Christian M. . E-mail: C.M.Kaehler@uibk.ac.at

    2006-09-10

    Growing evidence indicates that inducible cyclooxygenase-2 (COX-2) is involved in the pathogenesis of inflammatory disorders and various types of cancer. Endothelial progenitor cells recruited from the bone marrow have been shown to be involved in the formation of new vessels in malignancies and discussed for being a key point in tumour progression and metastasis. However, until now, nothing is known about an interaction between COX and endothelial progenitor cells (EPC). Expression of COX-1 and COX-2 was detected by semiquantitative RT-PCR and Western blot. Proliferation kinetics, cell cycle distribution and rate of apoptosis were analysed by MTT test and FACS analysis. Further analyses revealed an implication of Akt phosphorylation and caspase-3 activation. Both COX-1 and COX-2 expression can be found in bone-marrow-derived endothelial progenitor cells in vitro. COX-2 inhibition leads to a significant reduction in proliferation of endothelial progenitor cells by an increase in apoptosis and cell cycle arrest. COX-2 inhibition leads further to an increased cleavage of caspase-3 protein and inversely to inhibition of Akt activation. Highly proliferating endothelial progenitor cells can be targeted by selective COX-2 inhibition in vitro. These results indicate that upcoming therapy strategies in cancer patients targeting COX-2 may be effective in inhibiting tumour vasculogenesis as well as angiogenic processes.

  1. 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.

  2. Ethylene Inhibits Cell Proliferation of the Arabidopsis Root Meristem.

    PubMed

    Street, Ian H; Aman, Sitwat; Zubo, Yan; Ramzan, Aleena; Wang, Xiaomin; Shakeel, Samina N; Kieber, Joseph J; Schaller, G Eric

    2015-09-01

    The root system of plants plays a critical role in plant growth and survival, with root growth being dependent on both cell proliferation and cell elongation. Multiple phytohormones interact to control root growth, including ethylene, which is primarily known for its role in controlling root cell elongation. We find that ethylene also negatively regulates cell proliferation at the root meristem of Arabidopsis (Arabidopsis thaliana). Genetic analysis indicates that the inhibition of cell proliferation involves two pathways operating downstream of the ethylene receptors. The major pathway is the canonical ethylene signal transduction pathway that incorporates CONSTITUTIVE TRIPLE RESPONSE1, ETHYLENE INSENSITIVE2, and the ETHYLENE INSENSITIVE3 family of transcription factors. The secondary pathway is a phosphorelay based on genetic analysis of receptor histidine kinase activity and mutants involving the type B response regulators. Analysis of ethylene-dependent gene expression and genetic analysis supports SHORT HYPOCOTYL2, a repressor of auxin signaling, as one mediator of the ethylene response and furthermore, indicates that SHORT HYPOCOTYL2 is a point of convergence for both ethylene and cytokinin in negatively regulating cell proliferation. Additional analysis indicates that ethylene signaling contributes but is not required for cytokinin to inhibit activity of the root meristem. These results identify key elements, along with points of cross talk with cytokinin and auxin, by which ethylene negatively regulates cell proliferation at the root apical meristem.

  3. MiR-30e suppresses proliferation of hepatoma cells via targeting prolyl 4-hydroxylase subunit alpha-1 (P4HA1) mRNA

    SciTech Connect

    Feng, Guoxing; Shi, Hui; Li, Jiong; Yang, Zhe; Fang, Runping; Ye, Lihong; Zhang, Weiying; Zhang, Xiaodong

    2016-04-08

    Aberrant microRNA expression has been shown to be characteristic of many cancers. It has been reported that the expression levels of miR-30e are decreased in liver cancer tissues. However, the role of miR-30e in hepatocellular carcinoma remains poorly understood. In the present study, we investigated the significance of miR-30e in hepatocarcinogenesis. Bioinformatics analysis reveals a putative target site of miR-30e in the 3′-untranslated region (3′UTR) of prolyl 4-hydroxylase subunit alpha-1 (P4HA1) mRNA. Moreover, luciferase reporter gene assays verified that miR-30e directly targeted 3′UTR of P4HA1 mRNA. Then, we demonstrated that miR-30e was able to reduce the expression of P4HA1 at the levels of mRNA and protein using reverse transcription-polymerase chain reaction and Western blot analysis. Enforced expression of miR-30e suppressed proliferation of HepG2 cells by 5-ethynyl-2-deoxyuridine (EdU) assay and reduced colony formation of these cells by colony formation analysis. Conversely, anti-miR-30e enhanced the proliferation of hepatoma cells in vitro. Interestingly, the ectopic expression of P4HA1 could efficiently rescue the inhibition of cell proliferation mediated by miR-30e in HepG2 cells. Meanwhile, silencing of P4HA1 abolished the anti-miR-30e-induced proliferation of cells. Clinically, quantitative real-time PCR showed that miR-30e was down-regulated in liver tumor tissues relative to their peritumor tissues. The expression levels of miR-30e were negatively correlated to those of P4HA1 mRNA in clinical liver tumor tissues. Thus, we conclude that miR-30e suppresses proliferation of hepatoma cells through targeting P4HA1 mRNA. Our finding provides new insights into the mechanism of hepatocarcinogenesis. - Highlights: • P4HA1 is a novel target gene of miR-30e. • P4HA1 is increased in clinical HCC tissues. • MiR-30e is negatively correlated with P4HA1 in clinical HCC tissues. • MiR-30e suppresses the proliferation of HCC cells through

  4. Metformin inhibits proliferation and migration of glioblastoma cells independently of TGF-β2.

    PubMed

    Seliger, Corinna; Meyer, Anne-Louise; Renner, Kathrin; Leidgens, Verena; Moeckel, Sylvia; Jachnik, Birgit; Dettmer, Katja; Tischler, Ulrike; Gerthofer, Valeria; Rauer, Lisa; Uhl, Martin; Proescholdt, Martin; Bogdahn, Ulrich; Riemenschneider, Markus J; Oefner, Peter J; Kreutz, Marina; Vollmann-Zwerenz, Arabel; Hau, Peter

    2016-07-02

    To this day, glioblastoma (GBM) remains an incurable brain tumor. Previous research has shown that metformin, an oral anti-diabetic drug, may decrease GBM cell proliferation and migration especially in brain tumor initiating cells (BTICs). As transforming growth factor β 2 (TGF-β2) has been reported to promote high-grade glioma and is inhibited by metformin in other tumors, we explored whether metformin directly interferes with TGF-β2-signaling. Functional investigation of proliferation and migration of primary BTICs after treatment with metformin+/-TGF-β2 revealed that metformin doses as low as 0.01 mM metformin thrice a day were able to inhibit proliferation of susceptible cell lines, whereas migration was impacted only at higher doses. Known cellular mechanisms of metformin, such as increased lactate secretion, reduced oxygen consumption and activated AMPK-signaling, could be confirmed. However, TGF-β2 and metformin did not act as functional antagonists, but both rather inhibited proliferation and/or migration, if significant effects were present. We did not observe a relevant influence of metformin on TGF-β2 mRNA expression (qRT-PCR), TGF-β2 protein expression (ELISA) or SMAD-signaling (Western blot). Therefore, it seems that metformin does not exert its inhibitory effects on GBM BTIC proliferation and migration by altering TGF-β2-signaling. Nonetheless, as low doses of metformin are able to reduce proliferation of certain GBM cells, further exploration of predictors of BTICs' susceptibility to metformin appears justified.

  5. MicroRNA-29a inhibits mesenchymal stem cell viability and proliferation by targeting Roundabout 1.

    PubMed

    Zhang, Yudong; Zhou, Shenghua

    2015-10-01

    Secreted Slit glycoproteins and their Roundabout (Robo) receptors have been identified as important axon guidance molecules. The pivotal role of Slit‑Robo signaling is in regulating cell proliferation. MicroRNAs (miRNAs), a class of small non‑coding RNAs, function as critical regulators of gene expression by binding to the 3'‑untranslated region of mRNAs and causing mRNA degradation or translational repression. The present study demonstrated that downregulation of Robo1 using small interfering RNA inhibited mesenchymal stem cell (MSC) proliferation. Additionally, four miRNAs (miR), including miR‑218, miR‑29a, miR‑146 and miR‑148, inhibited the protein expression of Robo1 in the MSCs, with miR‑29 having the most marked effect. A luciferase reporter assay identified Robo1 as a novel target of miR‑29a. Overexpression of miR‑29a suppressed the protein expression levels of Robo1 and Slit2 and inhibited the viability and proliferation of the MSCs. By contrast, overexpression of Robo1 partly rescued these inhibitory effects of miR‑29a on the MSCs confirming that miR‑29a inhibited MSC viability and proliferation, at least partially, by directly targeting Robo1. These results indicated that the miR‑29a/Robo1 axis is crucial for the regulation of MSC viability and proliferation, suggesting that miR‑29a may serve as a potential clinical target for MSC expansion and stem cell transplantation.

  6. Long non-coding RNA GHET1 promotes gastric carcinoma cell proliferation by increasing c-Myc mRNA stability.

    PubMed

    Yang, Feng; Xue, Xuchao; Zheng, Luming; Bi, Jianwei; Zhou, Yuhong; Zhi, Kangkang; Gu, Yan; Fang, Guoen

    2014-02-01

    Long non-coding RNAs (lncRNAs), a recently characterized class of non-coding RNAs, have been shown to have important regulatory roles and are de-regulated in a variety of tumors. However, the contributions of lncRNAs to gastric carcinoma and their functional mechanisms remain largely unknown. In this study, we found that lncRNA gastric carcinoma high expressed transcript 1 (lncRNA-GHET1) was up-regulated in gastric carcinoma. The over-expression of this lncRNA correlates with tumor size, tumor invasion and poor survival. Gain-of-function and loss-of-function analyses demonstrated that GHET1 over-expression promotes the proliferation of gastric carcinoma cells in vitro and in vivo. Knockdown of GHET1 inhibits the proliferation of gastric carcinoma cells. RNA pull-down and immunoprecipitation assays confirmed that GHET1 physically associates with insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) and enhances the physical interaction between c-Myc mRNA and IGF2BP1, consequently increasing the stability of c-Myc mRNA and expression. The expression of GHET1 and c-Myc is strongly correlated in gastric carcinoma tissues. Depletion of c-Myc abolishes the effects of GHET1 on proliferation of gastric carcinoma cells. Taken together, these findings indicate that GHET1 plays a pivotal role in gastric carcinoma cell proliferation via increasing c-Myc mRNA stability and expression, which suggests potential use of GHET1 for the prognosis and treatment of gastric carcinoma. © 2013 FEBS.

  7. Ionizing irradiation inhibits keloid fibroblast cell proliferation and induces premature cellular senescence.

    PubMed

    Ji, Jiang; Tian, Ye; Zhu, Ya-qun; Zhang, Li-yuan; Ji, Sheng-jun; Huan, Jian; Zhou, Xiao-zhong; Cao, Jian-ping

    2015-01-01

    Keloids are one of the common refractory conditions in dermatology and aesthetic plastic surgery. The most effective treatment is superficial radiotherapy followed by surgical removal. The rate of recurrence is strongly associated with the total dose of ionizing irradiation, and the underlying mechanism remains unclear. In this study, we used primary keloid fibroblasts (KFb) isolated from patient samples to investigate the effects of X-ray radiation on cell proliferation, cell toxicity and cell cycle, as detected by CCK-8 assay kit and flow cytometer. In addition, we examined senescence-associated β-galactosidase activity and the associated gene expression using real-time polymerase chain reaction and western blot in KFb exposed to X-ray radiation. X-ray radiation inhibited cell proliferation and induced cell senescence in KFb in a dose-dependent manner. Inhibition of cell proliferation and induction of cellular senescence were mediated by interruption of the cell cycle with an extended G0/G1 phase. Furthermore, the expressions of senescence-associated genes p21, p16 and p27 were upregulated both at mRNA and protein levels in KFb exposed to X-ray radiation. Taken together, our data indicate that X-ray radiation may prevent the recurrence of keloids by controlling fibroblast proliferation, arresting the cell cycle and inducing premature cellular senescence. © 2014 Japanese Dermatological Association.

  8. MiR-661 inhibits glioma cell proliferation, migration and invasion by targeting hTERT

    SciTech Connect

    Li, Zhen; Liu, Yun-hui; Diao, Hong-yu; Ma, Jun; Yao, Yi-long

    2015-12-25

    In this study, we analyzed the functional role of miR-661 in glioma cell proliferation, migration and invasion. We found that overexpression of miR-661 obviously suppressed the proliferation, migration and invasion of glioma cells. MiRNA target prediction algorithms implied that hTERT is a candidate target gene for miR-661. A fluorescent reporter assay confirmed that miR-661 could lead to hTERT gene silencing by recognizing and specifically binding to the predicted site of the hTERT mRNA 3′ untranslated region (3′UTR) specifically. Furthermore, hTERT knockdown significantly decreased the growth and viability of glioma cells. These results indicate that miR-661 can inhibit glioma cell proliferation, migration and invasion by targeting hTERT. - Highlights: • MiR-661 was downregulated in glioma tissues and functional as a tumor suppressor. • MiR-661 modulates cell proliferation, invasion and migration of glioma cells. • MiR-661 directly target hTERT in glioma cells. • MiR-661 inhibits glioma cell tumorgenesis by targeting hTERT.

  9. Oxymatrine inhibits the proliferation of CaSki cells via downregulating HPV16E7 expression.

    PubMed

    Pei, Zhijun; Zeng, Jing; Gao, Yan; Li, Fuyan; Li, Wei; Zhou, Hong; Yang, Yi; Wu, Ruimin; Chen, Yijia; Liu, Jie

    2016-07-01

    Treatment of recurrent and metastatic cervical cancer remains a challenge, especially in developing countries, which lack efficient screening programs. HPV16E7 has been reported to play an important role in the development of cervical cancer. In recent years, oxymatrine, which was traditionally used as anti-malarial agent, has been shown to inhibit tumor growth with low toxicity to normal cells. In the present study, we investigated the mechanisms underlying the antitumor effect of oxymatrine in cervical cancer. The CCK-8 assay was used to compare the proliferation of untreated and oxymatrine-treated cervical cancer CaSki cells. Flow cytometry was applied to observe the effect of oxymatrine on apoptosis and the cell cycle distribution of CaSki cells. We used qRT-PCR and western blot analysis to determine the mRNA level and protein level of HPV16E7. The HPV16E7 siRNA inhibition was also performed to confirm the effect of downregulating HPV16E7 on the proliferation in CaSki cells. Our results revealed that oxymatrine-treated cells showed time-dependent and dose-dependent inhibition of proliferation and a significant increase in apoptosis. Oxymatrine arrested CaSki cells in G0/G1 phase and S phase while decreased the cells in G2/M phase. The expression of HPV16E7 was significantly downregulated in oxymatrine-treated cells compared with control cells. Knock-down of HPV16E7 effectively inhibited the proliferation of CaSki cells. In conclusion, our data suggest that oxymatrine inhibits cervical cancer growth via downregulation of HPV16E7. Oxymatrine can be considered to be a potential preventive and therapeutic target for cervical cancer.

  10. Myostatin inhibits proliferation of human urethral rhabdosphincter satellite cells.

    PubMed

    Akita, Yasuyuki; Sumino, Yasuhiro; Mori, Ken-ichi; Nomura, Takeo; Sato, Fuminori; Mimata, Hiromitsu

    2013-05-01

    Myostatin, a member of the transforming growth factor-β superfamily, is a negative regulator of myogenesis in skeletal muscle. We examined the effect of myostatin and myostatin inhibition by an antagonistic agent, follistatin, on growth of human urethral rhabdosphincter satellite cells (muscle stem cells) to develop a new strategy for treatment of stress urinary incontinence. Rhabdosphincter satellite cells were cultured and selected by magnetic affinity cell sorting using an anti-neural cell adhesion molecule antibody. The cells were transfected with simian virus-40 antigen to extend their lifespan. A cell proliferation assay, a cell cycle analysis and an investigation of signal transduction were carried out. The autocrine action of endogenous myostatin by western blotting, real-time reverse transcription polymerase chain reaction and immunoneutralization using an anti-myostatin antibody was also evaluated. Selectively cultured cells expressed markers of striated muscles and successfully differentiated into myotubes. Myostatin inhibited proliferation of these cells through Smad2 phosphorylation and cell cycle arrest. Inhibitory effects of myostatin were reversed by addition of follistatin. However, rhabdosphincter satellite cells did not appear to use autocrine secretion of myostatin to regulate their proliferation. Inhibition of myostatin function might be a useful pathway in the development of novel strategies for stimulating rhabdosphincter cells regeneration to treat stress urinary incontinence. © 2012 The Japanese Urological Association.

  11. Dexamethasone inhibits induction of liver tumor necrosis factor-alpha mRNA and liver growth induced by lead nitrate and ethylene dibromide.

    PubMed Central

    Ledda-Columbano, G. M.; Columbano, A.; Cannas, A.; Simbula, G.; Okita, K.; Kayano, K.; Kubo, Y.; Katyal, S. L.; Shinozuka, H.

    1994-01-01

    We have recently demonstrated that a single injection of the mitogen lead nitrate to rats induced a rapid increase of tumor necrosis factor-alpha (TNF-alpha) mRNA in the liver and suggested that this cytokine may be involved in triggering hepatocyte proliferation in this model of direct hyperplasia. In this study, we examined whether a similar induction of liver TNF-alpha mRNA could be observed preceding the onset of hepatocyte proliferation induced by ethylene dibromide, another hepatocyte mitogen. In addition, we used dexamethasone, a well known inhibitor of TNF-alpha production, to determine whether its administration could suppress hepatocyte proliferation induced by lead nitrate and ethylene dibromide. A single intragastric administration of ethylene dibromide (100 mg/kg) to male Wistar rats enhanced liver TNF-alpha mRNA after 4 and 7 hours, which then returned to control levels by 24 hours. TNF-alpha mRNA was detectable only in a nonparenchymal cell fraction of the liver. Pretreatment of rats with a single dose of dexamethasone (2 mg/kg) 60 minutes before lead nitrate (100 mumol/kg) or ethylene dibromide completely abolished the increased levels of liver TNF-alpha mRNA induced by these agents. Inhibition by dexamethasone of TNF-alpha mRNA was associated with an inhibition of liver cell proliferation induced by these mitogens, as measured by [3H]thymidine incorporation into hepatic DNA, mitotic index, and DNA content. These results further support the hypothesis that TNF-alpha may be involved in triggering hepatocyte proliferation induced by primary mitogens. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:7943184

  12. Lysyl oxidase propeptide inhibits smooth muscle cell signaling and proliferation

    SciTech Connect

    Hurtado, Paola A.; Vora, Siddharth; Sume, Siddika Selva; Yang, Dan; Hilaire, Cynthia St.; Guo Ying; Palamakumbura, Amitha H.; Schreiber, Barbara M.; Ravid, Katya; Trackman, Philip C.

    2008-02-01

    Lysyl oxidase is required for the normal biosynthesis and maturation of collagen and elastin. It is expressed by vascular smooth muscle cells, and its increased expression has been previously found in atherosclerosis and in models of balloon angioplasty. The lysyl oxidase propeptide (LOX-PP) has more recently been found to have biological activity as a tumor suppressor, and it inhibits Erk1/2 Map kinase activation. We reasoned that LOX-PP may have functions in normal non-transformed cells. We, therefore, investigated its effects on smooth muscle cells, focusing on important biological processes mediated by Erk1/2-dependent signaling pathways including proliferation and matrix metalloproteinase-9 (MMP-9) expression. In addition, we investigated whether evidence for accumulation of LOX-PP could be found in vivo in a femoral artery injury model. Recombinant LOX-PP was expressed and purified, and was found to inhibit primary rat aorta smooth muscle cell proliferation and DNA synthesis by more than 50%. TNF-{alpha}-stimulated MMP-9 expression and Erk1/2 activation were both significantly inhibited by LOX-PP. Immunohistochemistry studies carried out with affinity purified anti-LOX-PP antibody showed that LOX-PP epitopes were expressed at elevated levels in vascular lesions of injured arteries. These novel data suggest that LOX-PP may provide a feedback control mechanism that serves to inhibit properties associated with the development of vascular pathology.

  13. Zedoary oil (Ezhu You) inhibits proliferation of AGS cells

    PubMed Central

    2013-01-01

    Background Zedoary (Curcumae Rhizoma, Ezhu), a Chinese medicinal herb, has been reported to show anticancer activity. This study aims to investigate the effect of zedoary oil (Ezhu You) on the proliferation of AGS cells which is one gastric cancer cell line. Methods The main ingredients of the herb were detected by GC-MS for herbal quality control. Cell viability was measured by MTT assay and cell proliferation was investigated by immunocytochemical staining for proliferating cell nuclear antigen (PCNA) protein. In addition, the cell cycle distributions were detected by flow cytometry with propidium iodine (PI) staining and the apoptosis rates were evaluated by flow cytometry with annexin V/PI double-staining. The morphological changes associated with apoptosis were observed by Hoechst 33342/PI double-staining. Protein expression was determined by western blot analysis. Results The main ingredients of the herb, including curzerene (26.45%), eucalyptol (12.04%), curcumol (9.04%), pyridine (7.97%), germacrone (7.89%), β-elemene (7.36%), τ-elemene (4.11%) and 28 other ingredients, including curdione, were consistent with the chemical profiles of zedoary. Zedoary oil significantly decreased the cell viability of AGS cells (P < 0.01) and MGC 803 cells (P < 0.01), and the inhibitory effects were attenuated by elevated concentrations of FBS. At high concentrations (≥90 μg/mL), zedoary oil killed GES-1 cells. At low concentrations (≤60 μg/mL), zedoary oil was less inhibitory toward normal gastric epithelial cells than gastric cancer cell lines. In AGS cells, zedoary oil inhibited cell proliferation in a dose- and time-dependent manner, with decreased PCNA protein expression in the zedoary oil-treated cells, and arrested the cell cycle at S, G2/M and G0/G1 stages after treatment for 6–48 h. At concentrations of 30, 60 and 90 μg/mL, which resulted in significant inhibition of proliferation and cell cycle arrest, zedoary oil induced cell apoptosis. In

  14. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) inhibits human ovarian cancer cell proliferation

    PubMed Central

    Li, Yan; Wang, Kai; Jiang, Yi-Zhou; Chang, Xin-Wen; Dai, Cai-Feng

    2017-01-01

    Purpose The aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor, mediates a broad spectrum of biological processes, including ovarian growth and ovulation. Recently, we found that an endogenous AhR ligand (ITE) can inhibit ovarian cancer proliferation and migration via the AhR. Here, we tested whether 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, an exogenous AhR ligand) may exert similar anti-ovarian cancer activities using human ovarian cancer and non-cancerous human ovarian surface epithelial cells. Methods Two human ovarian cancer cell lines (SKOV-3 and OVCAR-3) and one human ovarian surface epithelial cell line (IOSE-385) were used. Cell proliferation and migration activities were determined using crystal violet and FluoroBlok insert system assays, respectively. AhR protein expression was assessed by Western blotting. Expression of cytochrome P450, family 1, member A1 (CYP1A1) and member B1 (CYP1B1) mRNA was assessed by qPCR. Small interfering RNAs (siRNAs) were used to knock down AhR expression. Results We found that TCDD dose-dependently suppressed OVCAR-3 cell proliferation, with a maximum effect (~70 % reduction) at 100 nM. However, TCDD did not affect SKOV-3 and IOSE-385 cell proliferation and migration. The estimated IC50 of TCDD for inhibiting OVCAR-3 cell proliferation was 4.6 nM. At 10 nM, TCDD time-dependently decreased AhR protein levels, while it significantly increased CYP1A1 and CYP1B1 mRNA levels in SKOV-3, OVCAR-3 and IOSE-385 cells, indicating activation of AhR signaling. siRNA-mediated AhR knockdown readily blocked TCDD-mediated suppression of OVCAR-3 cell proliferation. Conclusion Our data indicate that TCDD can suppress human ovarian cancer cell proliferation via the AhR signaling pathway and that TCDD exhibits an anti-proliferative activity in at least a subset of human ovarian cancer cells. PMID:25404385

  15. Target mRNA inhibition by oligonucleotide drugs in man

    PubMed Central

    Lightfoot, Helen L.; Hall, Jonathan

    2012-01-01

    Oligonucleotide delivery in vivo is commonly seen as the principal hurdle to the successful development of oligonucleotide drugs. In an analysis of 26 oligonucleotide drugs recently evaluated in late-stage clinical trials we found that to date at least half have demonstrated suppression of the target mRNA and/or protein levels in the relevant cell types in man, including those present in liver, muscle, bone marrow, lung, blood and solid tumors. Overall, this strongly implies that the drugs are being delivered to the appropriate disease tissues. Strikingly we also found that the majority of the drug targets of the oligonucleotides lie outside of the drugable genome and represent new mechanisms of action not previously investigated in a clinical setting. Despite the high risk of failure of novel mechanisms of action in the clinic, a subset of the targets has been validated by the drugs. While not wishing to downplay the technical challenges of oligonucleotide delivery in vivo, here we demonstrate that target selection and validation are of equal importance for the success of this field. PMID:22989709

  16. Effects of AFP gene silencing on Survivin mRNA expression inhibition in HepG2 cells.

    PubMed

    Fang, Z L; Fang, N; Han, X N; Huang, G; Fu, X J; Xie, G S; Wang, N R; Xiong, J P

    2015-04-10

    We investigated the effects of alpha-fetoprotein (AFP) gene silencing on Survivin expression in HepG2 cells. Small interfering RNA technology was used to downregulate AFP expression in HepG2 cells. An enzyme-linked immunosorbent assay was used to measure AFP concentration in the supernatant before and after transfection. An MTT assay was used to detect cell proliferation activity before and after transfection. We performed flow cytometric analysis to detect the cell apoptosis rate, and reverse transcription-polymerase chain reaction to detect Survivin mRNA levels before and after transfection. Forty-eight hours after transfection, AFP concentration in the supernatant of the experimental group significantly decreased, hepatocellular carcinoma cell growth was inhibited by 43.1%, and the apoptosis rate increased by 24.3%. Survivin mRNA expression was reduced by 78.0% in HepG2 cells. These indicators in the control group and in the blank group did not change significantly. Silencing of AFP expression in HepG2 cells can effectively inhibit the growth of hepatoma cells and promote apoptosis, which may be useful for reducing intracellular Survivin mRNA levels.

  17. TRPM7 channel inhibition mediates midazolam-induced proliferation loss in human malignant glioma.

    PubMed

    Chen, Jingkao; Dou, Yunling; Zheng, Xiaoke; Leng, Tiandong; Lu, Xiaofang; Ouyang, Ying; Sun, Huawei; Xing, Fan; Mai, Jialuo; Gu, Jiayu; Lu, Bingzheng; Yan, Guangmei; Lin, Jun; Zhu, Wenbo

    2016-11-01

    The melastatin-like transient receptor potential 7 (TRPM7) has been implicated in proliferation or apoptosis of some cancers, indicating the potential of TRPM7 as an anti-anaplastic target. Here, we identified the characteristic TRPM7 channel currents in human malignant glioma MGR2 cells, which could be blocked by a pharmacologic inhibitor Gd(3+). We mined the clinical sample data from Oncomine Database and found that human malignant glioma tissues expressed higher TRPM7 mRNA than normal brain ones. Importantly, we identified a widely used clinical anesthetic midazolam as a TRPM7 inhibitor. Midazolam treatment for seconds suppressed the TRPM7 currents and calcium influx, and treatment for 48 h inhibited the TRPM7 expression. The inhibitory effect on TRPM7 accounts for the proliferation loss and G0/G1 phase cell cycle arrest induced by midazolam. Our data demonstrates that midazolam represses proliferation of human malignant glioma cells through inhibiting TRPM7 currents, which may be further potentiated by suppressing the expression of TRPM7. Our result indicates midazolam as a pharmacologic lead compound with brain-blood barrier permeability for targeting TRPM7 in the glioma.

  18. ESR1 inhibits hCG-induced steroidogenesis and proliferation of progenitor Leydig cells in mice.

    PubMed

    Oh, Yeong Seok; Koh, Il Kyoo; Choi, Bomi; Gye, Myung Chan

    2017-03-07

    Oestrogen is an important regulator in reproduction. To understand the role of oestrogen receptor 1 (ESR1) in Leydig cells, we investigated the expression of ESR1 in mouse Leydig cells during postnatal development and the effects of oestrogen on steroidogenesis and proliferation of progenitor Leydig cells (PLCs). In Leydig cells, the ESR1 expression was low at birth, increased until postnatal day 14 at which PLCs were predominant, and then decreased until adulthood. In foetal Leydig cells, ESR1 immunoreactivity increased from birth to postnatal day 14. These suggest that ESR1 is a potential biomarker of Leydig cell development. In PLCs, 17β-estradiol and the ESR1-selective agonist propylpyrazoletriol suppressed human chorionic gonadotropin (hCG)-induced progesterone production and steroidogenic gene expression. The ESR2-selective agonist diarylpropionitrile did not affect steroidogenesis. In PLCs from Esr1 knockout mice, hCG-stimulated steroidogenesis was not suppressed by 17β-estradiol, suggesting that oestrogen inhibits PLC steroidogenesis via ESR1. 17β-estradiol, propylpyrazoletriol, and diarylpropionitrile decreased bromodeoxyuridine uptake in PLCs in the neonatal mice. In cultured PLCs, 17β-estradiol, propylpyrazoletriol, and diarylpropionitrile reduced hCG-stimulated Ki67 and Pcna mRNA expression and the number of KI67-positive PLCs, suggesting that oestrogen inhibits PLC proliferation via both ESR1 and ESR2. In PLCs, ESR1 mediates the oestrogen-induced negative regulation of steroidogenesis and proliferation.

  19. ESR1 inhibits hCG-induced steroidogenesis and proliferation of progenitor Leydig cells in mice

    PubMed Central

    Oh, Yeong Seok; Koh, Il Kyoo; Choi, Bomi; Gye, Myung Chan

    2017-01-01

    Oestrogen is an important regulator in reproduction. To understand the role of oestrogen receptor 1 (ESR1) in Leydig cells, we investigated the expression of ESR1 in mouse Leydig cells during postnatal development and the effects of oestrogen on steroidogenesis and proliferation of progenitor Leydig cells (PLCs). In Leydig cells, the ESR1 expression was low at birth, increased until postnatal day 14 at which PLCs were predominant, and then decreased until adulthood. In foetal Leydig cells, ESR1 immunoreactivity increased from birth to postnatal day 14. These suggest that ESR1 is a potential biomarker of Leydig cell development. In PLCs, 17β-estradiol and the ESR1-selective agonist propylpyrazoletriol suppressed human chorionic gonadotropin (hCG)-induced progesterone production and steroidogenic gene expression. The ESR2-selective agonist diarylpropionitrile did not affect steroidogenesis. In PLCs from Esr1 knockout mice, hCG-stimulated steroidogenesis was not suppressed by 17β-estradiol, suggesting that oestrogen inhibits PLC steroidogenesis via ESR1. 17β-estradiol, propylpyrazoletriol, and diarylpropionitrile decreased bromodeoxyuridine uptake in PLCs in the neonatal mice. In cultured PLCs, 17β-estradiol, propylpyrazoletriol, and diarylpropionitrile reduced hCG-stimulated Ki67 and Pcna mRNA expression and the number of KI67-positive PLCs, suggesting that oestrogen inhibits PLC proliferation via both ESR1 and ESR2. In PLCs, ESR1 mediates the oestrogen-induced negative regulation of steroidogenesis and proliferation. PMID:28266530

  20. LRG1 promotes proliferation and inhibits apoptosis in colorectal cancer cells via RUNX1 activation.

    PubMed

    Zhou, Ying; Zhang, Xintian; Zhang, Jingjing; Fang, Jingyuan; Ge, Zhizheng; Li, Xiaobo

    2017-01-01

    Leucine-rich-alpha-2-glycoprotein 1 (LRG1) has been shown to be involved in various human malignancies. Whether it plays a role in colorectal cancer (CRC) development remains unclear. Here, we investigated whether and through what mechanism LRG1 functions in human CRC cells. The plasma level of LRG1 was significantly increased in CRC patients, but it was remarkably decreased in patients with resected colorectal cancers. Meanwhile, both mRNA and protein levels of LRG1 were remarkable overexpressed in CRC tissues than normal tissues. The knockdown of LRG1 significantly inhibited cell proliferation, induced cell cycle arrest at the G0/G1 phase, and promoted apoptosis in SW480 and HCT116 cells in vitro. In addition, LRG1 silencing led to the downregulation of the levels of key cell cycle factors, such as cyclin D1, B, and E and anti-apoptotic B-cell lymphoma-2(Bcl-2). However, it up-regulated the expression of pro-apoptotic Bax and cleaved caspase-3. Furthermore, RUNX1 could be induced by LRG1 in a concentration-dependent manner, while the knockdown of RUNX1 blocked the promotion of the proliferation and inhibition of apoptosis induced by LRG1. Collectively, these findings indicate that LRG1 plays a crucial role in the proliferation and apoptosis of CRC by regulating RUNX1 expression. Thus, LRG1 may be a potential detection biomarker as well as a marker for monitoring recurrence and therapeutic target for CRC.

  1. Skeletal unloading inhibits the in vitro proliferation and differentiation of rat osteoprogenitor cells

    NASA Technical Reports Server (NTRS)

    Kostenuik, P. J.; Halloran, B. P.; Morey-Holton, E. R.; Bikle, D. D.

    1997-01-01

    Loss of weight bearing in the growing rat decreases bone formation, osteoblast numbers, and bone maturation in unloaded bones. These responses suggest an impairment of osteoblast proliferation and differentiation. To test this assumption, we assessed the effects of skeletal unloading using an in vitro model of osteoprogenitor cell differentiation. Rats were hindlimb elevated for 0 (control), 2, or 5 days, after which their tibial bone marrow stromal cells (BMSCs) were harvested and cultured. Five days of hindlimb elevation led to significant decreases in proliferation, alkaline phosphatase (AP) enzyme activity, and mineralization of BMSC cultures. Differentiation of BMSCs was analyzed by quantitative competitive polymerase chain reaction of cDNA after 10, 15, 20, and 28 days of culture. cDNA pools were analyzed for the expression of c-fos (an index of proliferation), AP (an index of early osteoblast differentiation), and osteocalcin (a marker of late differentiation). BMSCs from 5-day unloaded rats expressed 50% less c-fos, 61% more AP, and 35% less osteocalcin mRNA compared with controls. These data demonstrate that cultured osteoprogenitor cells retain a memory of their in vivo loading history and indicate that skeletal unloading inhibits proliferation and differentiation of osteoprogenitor cells in vitro.

  2. Skeletal unloading inhibits the in vitro proliferation and differentiation of rat osteoprogenitor cells

    NASA Technical Reports Server (NTRS)

    Kostenuik, P. J.; Halloran, B. P.; Morey-Holton, E. R.; Bikle, D. D.

    1997-01-01

    Loss of weight bearing in the growing rat decreases bone formation, osteoblast numbers, and bone maturation in unloaded bones. These responses suggest an impairment of osteoblast proliferation and differentiation. To test this assumption, we assessed the effects of skeletal unloading using an in vitro model of osteoprogenitor cell differentiation. Rats were hindlimb elevated for 0 (control), 2, or 5 days, after which their tibial bone marrow stromal cells (BMSCs) were harvested and cultured. Five days of hindlimb elevation led to significant decreases in proliferation, alkaline phosphatase (AP) enzyme activity, and mineralization of BMSC cultures. Differentiation of BMSCs was analyzed by quantitative competitive polymerase chain reaction of cDNA after 10, 15, 20, and 28 days of culture. cDNA pools were analyzed for the expression of c-fos (an index of proliferation), AP (an index of early osteoblast differentiation), and osteocalcin (a marker of late differentiation). BMSCs from 5-day unloaded rats expressed 50% less c-fos, 61% more AP, and 35% less osteocalcin mRNA compared with controls. These data demonstrate that cultured osteoprogenitor cells retain a memory of their in vivo loading history and indicate that skeletal unloading inhibits proliferation and differentiation of osteoprogenitor cells in vitro.

  3. Oleanolic acid inhibits proliferation and induces apoptosis in NB4 cells by targeting PML/RARα

    PubMed

    Li, Hongmei; He, Ning; Li, Xueyan; Zhou, Li; Zhao, Mingyu; Jiang, Hairui; Zhang, Xiaojie

    2013-10-01

    Oleanolic acid (OA), a naturally occurring pentacyclic triterpenoid contained in a variety of plant species, exhibits broad biological properties, including anticancer effects. Acute promyelocytic leukemia (APL) is a distinct subtype of acute myeloid leukemia. APL has a unique and specific chromosomal aberration, t(15;17), which results in the formation of a fusion gene and protein PML/RARα, which is not only necessary for the diagnosis of APL, but is also critical for APL pathogenesis. In the present study, the cytotoxic effect of OA on NB4 cells was investigated. Cell viability was assessed via the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The expression levels of bax and bcl-2 mRNA were determined by quantitative PCR. Apoptosis was analyzed using DNA fragment analysis and cell cycle distributions were analyzed by flow cytometry. The activity of caspase-3 and caspase-9 was determined by colorimetric assays. The expression of the PML/RARα fusion protein was analyzed by western blotting. The MTT assay showed that OA inhibited the proliferation of the NB4 cells. The expression levels of pro-apoptotic bax mRNA were increased and the levels of anti-apoptotic bcl-2 mRNA were decreased following the treatment of the NB4 cells with OA at 80 μmol/l. Treatment of the NB4 cells with OA at 80 μmol/l induced apoptosis and G1 phase arrest, while caspase-9 and caspase-3 activity was significantly increased. Furthermore, the expression of the PML/RARα fusion protein was decreased. Together, these data suggest that OA exerts a cytotoxic effect that inhibits proliferation and induces apoptosis in NB4 cells by targeting PML/RARα, making it a potent therapeutic agent against leukemia.

  4. Momordin Ic, a new natural SENP1 inhibitor, inhibits prostate cancer cell proliferation

    PubMed Central

    Tang, Caixia; Wang, Weiwei; Xu, Hanzhang; Xiao, Weilie; Gu, Wenli; Wu, Yingli

    2016-01-01

    SUMO-specific protease 1 (SENP1), a member of the de-SUMOylation protease family, is elevated in prostate cancer (PCa) cells and is involved in PCa pathogenesis. Momordin Ιc (Mc), a natural pentacyclic triterpenoid, inhibited SENP1 in vitro, as reflected by reduced SENP1C-induced cleavage of SUMO2-ΔRanGAP1. Mc also altered the thermal stability of SENP1 in a newly developed cellular thermal shift assay, indicating that Mc directly interacts with SENP1 in PCa cells. Consistent with SENP1 inhibition, Mc increased SUMOylated protein levels, which was further confirmed by the accumulation of two known SUMOylated proteins, hypoxia inducible factor-1a and nucleus accumbens associated protein 1 in PC3 cells. Compared to LNCaP and normal prostate epithelial RWPE-1 cells, PC3 cells had higher levels of SENP1 mRNA and were more sensitive to Mc-induced growth inhibition. Mc also reduced SENP1 mRNA levels in PCa cells. Overexpression of SENP1 rescued PC3 cells from Mc-induced apoptosis. Finally, Mc suppressed cell proliferation and induced cell death in vivo in a xenograft PC3 tumor mouse model. These findings demonstrate that Mc is a novel SENP1 inhibitor with potential therapeutic value for PCa. Investigation of other pentacyclic triterpenoids may aid in the development of novel SENP1 inhibitor drugs. PMID:27449295

  5. Momordin Ic, a new natural SENP1 inhibitor, inhibits prostate cancer cell proliferation.

    PubMed

    Wu, Jingjing; Lei, Hu; Zhang, Jinfu; Chen, Xiangyun; Tang, Caixia; Wang, Weiwei; Xu, Hanzhang; Xiao, Weilie; Gu, Wenli; Wu, Yingli

    2016-09-13

    SUMO-specific protease 1 (SENP1), a member of the de-SUMOylation protease family, is elevated in prostate cancer (PCa) cells and is involved in PCa pathogenesis. Momordin Ιc (Mc), a natural pentacyclic triterpenoid, inhibited SENP1 in vitro, as reflected by reduced SENP1C-induced cleavage of SUMO2-ΔRanGAP1. Mc also altered the thermal stability of SENP1 in a newly developed cellular thermal shift assay, indicating that Mc directly interacts with SENP1 in PCa cells. Consistent with SENP1 inhibition, Mc increased SUMOylated protein levels, which was further confirmed by the accumulation of two known SUMOylated proteins, hypoxia inducible factor-1a and nucleus accumbens associated protein 1 in PC3 cells. Compared to LNCaP and normal prostate epithelial RWPE-1 cells, PC3 cells had higher levels of SENP1 mRNA and were more sensitive to Mc-induced growth inhibition. Mc also reduced SENP1 mRNA levels in PCa cells. Overexpression of SENP1 rescued PC3 cells from Mc-induced apoptosis. Finally, Mc suppressed cell proliferation and induced cell death in vivo in a xenograft PC3 tumor mouse model. These findings demonstrate that Mc is a novel SENP1 inhibitor with potential therapeutic value for PCa. Investigation of other pentacyclic triterpenoids may aid in the development of novel SENP1 inhibitor drugs.

  6. Axl receptor tyrosine kinase is a novel target of apigenin for the inhibition of cell proliferation.

    PubMed

    Kim, Kyung-Chan; Choi, Eun-Ha; Lee, Chuhee

    2014-08-01

    The Axl receptor tyrosine kinase (RTK), along with Tyro 3 and Mer, belongs to the TAM subfamily that promotes survival, stimulates proliferation and/or inhibits apoptosis. In various types of human cancer, including breast, lung and prostate cancer, Axl expression is increased and correlates with an advanced clinical stage. In this study, we examined whether apigenin has an effect on Axl expression, which in turn can affect cell proliferation. The treatment of the non‑small cell lung cancer (NSCLC) cells, A549 and H460, with apigenin decreased Axl mRNA and protein expression in a dose‑dependent manner. Axl promoter activity was also inhibited by apigenin, indicating that apigenin suppressed Axl expression at the transcriptional level. Upon treatment with apigenin, the viability of both the A549 and H460 cells was gradually decreased and the anti-proliferative effects were further confirmed by the dose‑dependent decrease in the clonogenic ability of the apigenin‑treated cells. Subsequently, we found that the viability and clonogenic ability of the cells treated with apigenin was less or more affected by transfection of the cells with a Axl-expressing plasmid or Axl targeting siRNA, compared to transfection with the empty vector or control siRNA, respectively. In addition, apigenin increased the expression of p21, a cyclin-dependent kinase inhibitor, but reduced the expression of X-linked inhibitor of apoptosis protein (XIAP). These cell cycle arrest and pro-apoptotic effects of apigenin were also attenuated or augmented by the up- or downregulation of Axl expression, respectively, which suggests that Axl is a novel target of apigenin through which it exerts its inhibitory effects on cell proliferation. Taken together, our data indicate that apigenin downregulates Axl expression, which subsequently results in the inhibition of NSCLC cell proliferation through the increase and decrease of p21 and XIAP expression, respectively.

  7. Miconia sp. Increases mRNA Levels of PPAR Gamma and Inhibits Alpha Amylase and Alpha Glucosidase.

    PubMed

    Ortíz-Martinez, David Mizael; Rivas-Morales, Catalina; de la Garza-Ramos, Myriam Angelica; Verde-Star, Maria Julia; Nuñez-Gonzalez, Maria Adriana; Leos-Rivas, Catalina

    2016-01-01

    Diabetes mellitus is a public health problem worldwide. For this reason, ethanolic extract of Miconia sp. from Oaxaca, Mexico, was selected in search of an alternative against this disease. The effect of Miconia sp. on mRNA expression of PPARγ on cell line 3T3-L1, its effect on alpha amylase and alpha glucosidase, lipid accumulation during adipogenesis, and cell viability on VERO cells were evaluated. The mRNA levels of PPARγ increased on 1.393 ± 0.008 folds, lipid accumulation was increased by 29.55% with Miconia sp. extract and 34.57% with rosiglitazone, and α-amylase and α-glycosidase were inhibited with IC50 values from 28.23 ± 2.15 μg/mL and 1.95 ± 0.15 μg/mL, respectively; the IC50 on antiproliferative activity on VERO cells was 314.54 ± 45.40 μg/mL. In case of α-amylase and α-glycosidase assays, IC50 (inhibitory concentration 50) refers to necessary extract amounts to inhibit 50% of enzymatic activity. On the other hand, on antiproliferative activity, IC50 (inhibitory concentration 50) refers to necessary extract amounts to inhibit 50% of cell proliferation. It was concluded that the compounds present in Miconia sp. ethanolic extract increase mRNA expression of PPARγ, inhibit α-amylase and α-glucosidase, and increase lipid accumulation. It constitutes an alternative as adjuvant in diabetes mellitus treatment; therefore, we recommend continuing identifying the compounds responsible for its promising in vivo antidiabetic activity.

  8. Miconia sp. Increases mRNA Levels of PPAR Gamma and Inhibits Alpha Amylase and Alpha Glucosidase

    PubMed Central

    Ortíz-Martinez, David Mizael; Rivas-Morales, Catalina; de la Garza-Ramos, Myriam Angelica; Verde-Star, Maria Julia; Nuñez-Gonzalez, Maria Adriana

    2016-01-01

    Diabetes mellitus is a public health problem worldwide. For this reason, ethanolic extract of Miconia sp. from Oaxaca, Mexico, was selected in search of an alternative against this disease. The effect of Miconia sp. on mRNA expression of PPARγ on cell line 3T3-L1, its effect on alpha amylase and alpha glucosidase, lipid accumulation during adipogenesis, and cell viability on VERO cells were evaluated. The mRNA levels of PPARγ increased on 1.393 ± 0.008 folds, lipid accumulation was increased by 29.55% with Miconia sp. extract and 34.57% with rosiglitazone, and α-amylase and α-glycosidase were inhibited with IC50 values from 28.23 ± 2.15 μg/mL and 1.95 ± 0.15 μg/mL, respectively; the IC50 on antiproliferative activity on VERO cells was 314.54 ± 45.40 μg/mL. In case of α-amylase and α-glycosidase assays, IC50 (inhibitory concentration 50) refers to necessary extract amounts to inhibit 50% of enzymatic activity. On the other hand, on antiproliferative activity, IC50 (inhibitory concentration 50) refers to necessary extract amounts to inhibit 50% of cell proliferation. It was concluded that the compounds present in Miconia sp. ethanolic extract increase mRNA expression of PPARγ, inhibit α-amylase and α-glucosidase, and increase lipid accumulation. It constitutes an alternative as adjuvant in diabetes mellitus treatment; therefore, we recommend continuing identifying the compounds responsible for its promising in vivo antidiabetic activity. PMID:27478477

  9. MiR-34a inhibits colon cancer proliferation and metastasis by inhibiting platelet-derived growth factor receptor α.

    PubMed

    Li, Chunyan; Wang, Yulin; Lu, Shuming; Zhang, Zhuqing; Meng, Hua; Liang, Lina; Zhang, Yan; Song, Bo

    2015-11-01

    The microRNA (miRNA), miR‑34a is significant in colon cancer progression. In the present study, the role of miR‑34a in colon cancer cell proliferation and metastasis was investigated. It was found that the expression of miR‑34a in colon cancer tissues and cell lines was lower when compared with that of normal tissues and cells. Further research demonstrated that miR‑34a inhibited cell proliferation, induced G1 phase arrest, and suppressed metastasis and epithelial mesenchymal transition in colon cancer cells. Bioinformatic prediction indicated that platelet‑derived growth factor receptor α (PDGFRA) was a potential target gene of miR‑34a and a luciferase assay identified that PDGFRA was a novel direct target gene of miR‑34a. In addition, assays of western blot analyses and quantitative reverse‑transcription polymerase chain reaction confirmed that miR‑34a decreased PDGFRA mRNA expression and protein levels in colon cancer cells. Assessment of cellular function indicated that miR‑34a inhibited colon cancer progression via PDGFRA. These findings demonstrate that miR‑34a may act as a negative regulator in colon cancer by targeting PDGFRA.

  10. Vitamin E inhibits retinal pigment epithelium cell proliferation in vitro.

    PubMed

    Mojon, D; Boscoboinik, D; Haas, A; Bohnke, M; Azzi, A

    1994-01-01

    Retinal pigment epithelium (RPE) cells migrating through the damaged retina play an important role in the pathogenesis of proliferative vitreoretinopathy (PVR). We found that alpha-tocopherol (vitamin E) inhibits proliferation of human RPE in culture without exerting cytotoxic effects. Maximal inhibition was achieved with 100 microM alpha-tocopherol. Our result could explain the observation that vitamin E supplements have an adverse effect on light-damaged retina and on the course of retinitis pigmentosa. Since it has been shown that supplemental oral administrations of vitamin E can raise the RPE concentration of alpha-tocopherol well above 100 microM and supplementation is not associated with any clinical relevant adverse effect, we believe that vitamin E could be beneficial in the treatment of PVR.

  11. MicroRNA-29a inhibited epididymal epithelial cell proliferation by targeting nuclear autoantigenic sperm protein (NASP).

    PubMed

    Ma, Wubin; Xie, Shengsong; Ni, Minjie; Huang, Xingxu; Hu, Shuanggang; Liu, Qiang; Liu, Aihua; Zhang, Jinsong; Zhang, Yonglian

    2012-03-23

    Cell proliferation often decreases gradually during postnatal development of some organs. However, the underlying molecular mechanisms remain unclear. Epididymis, playing important roles in sperm maturation, is a typical organ of this type, which displays a decreased proliferation during postnatal development and even ceased at the adult stage. Here, epididymis was employed as a model to explore the underlying mechanisms. We profiled the microRNA and mRNA expression of newborn (1 day) and adult (90 day) rat epididymis by microarray analysis, and found that the level of miR-29a was dramatically up-regulated during postnatal development of rat epididymis. Subsequent investigations demonstrated that overexpression of miR-29a inhibited the proliferation of epididymal epithelial cells in vitro. The nuclear autoantigenic sperm protein (NASP), a novel target of miR-29a, was significantly down-regulated during postnatal development of rat epididymis. Further analysis showed that silence of NASP mimicked the anti-proliferation effect of miR-29a, whereas overexpression of this protein attenuated the effect of miR-29a. As in rat epididymis, miR-29a was up-regulated and Nasp was down-regulated during postnatal development of mouse epididymis, heart, liver, and lung. Moreover, miR-29a can also inhibit the proliferation of cancer cells by targeting Nasp. Thus, an increase of miR-29a, and hence decrease of Nasp, may contribute to inhibit cell proliferation during postnatal organ development.

  12. Identification of Novel Tumor Suppressor Genes in Human Breast Cancer Using Nonsense-Mediated mRNA Decay Inhibition (NMDI)-Microarray Analysis

    DTIC Science & Technology

    2007-08-01

    polymorphism array, deletion, allelic imbalance, p53, peroxisome proliferator activated receptor gamma, caffeine , Actinomycin D, emetine. 16... effects on the overall fitness of the organism. One such mechanism is nonsense-mediated mRNA decay (NMD), a pathway conserved from yeasts to humans...using caffeine (10 mM), rather than employing a global inhibition of translation with emetine (11). As a result of these protocol modifications

  13. The Hippo pathway mediates inhibition of vascular smooth muscle cell proliferation by cAMP

    PubMed Central

    Kimura, Tomomi E.; Duggirala, Aparna; Smith, Madeleine C.; White, Stephen; Sala-Newby, Graciela B.; Newby, Andrew C.; Bond, Mark

    2016-01-01

    Aims Inhibition of vascular smooth muscle cell (VSMC) proliferation by intracellular cAMP prevents excessive neointima formation and hence angioplasty restenosis and vein-graft failure. These protective effects are mediated via actin-cytoskeleton remodelling and subsequent regulation of gene expression by mechanisms that are incompletely understood. Here we investigated the role of components of the growth-regulatory Hippo pathway, specifically the transcription factor TEAD and its co-factors YAP and TAZ in VSMC. Methods and results Elevation of cAMP using forskolin, dibutyryl-cAMP or the physiological agonists, Cicaprost or adenosine, significantly increased phosphorylation and nuclear export YAP and TAZ and inhibited TEAD-luciferase report gene activity. Similar effects were obtained by inhibiting RhoA activity with C3-transferase, its downstream kinase, ROCK, with Y27632, or actin-polymerisation with Latrunculin-B. Conversely, expression of constitutively-active RhoA reversed the inhibitory effects of forskolin on TEAD-luciferase. Forskolin significantly inhibited the mRNA expression of the pro-mitogenic genes, CCN1, CTGF, c-MYC and TGFB2 and this was reversed by expression of constitutively-active YAP or TAZ phospho-mutants. Inhibition of YAP and TAZ function with RNAi or Verteporfin significantly reduced VSMC proliferation. Furthermore, the anti-mitogenic effects of forskolin were reversed by overexpression of constitutively-active YAP or TAZ. Conclusion Taken together, these data demonstrate that cAMP-induced actin-cytoskeleton remodelling inhibits YAP/TAZ–TEAD dependent expression of pro-mitogenic genes in VSMC. This mechanism contributes novel insight into the anti-mitogenic effects of cAMP in VSMC and suggests a new target for intervention. PMID:26625714

  14. The Hippo pathway mediates inhibition of vascular smooth muscle cell proliferation by cAMP.

    PubMed

    Kimura, Tomomi E; Duggirala, Aparna; Smith, Madeleine C; White, Stephen; Sala-Newby, Graciela B; Newby, Andrew C; Bond, Mark

    2016-01-01

    Inhibition of vascular smooth muscle cell (VSMC) proliferation by intracellular cAMP prevents excessive neointima formation and hence angioplasty restenosis and vein-graft failure. These protective effects are mediated via actin-cytoskeleton remodelling and subsequent regulation of gene expression by mechanisms that are incompletely understood. Here we investigated the role of components of the growth-regulatory Hippo pathway, specifically the transcription factor TEAD and its co-factors YAP and TAZ in VSMC. Elevation of cAMP using forskolin, dibutyryl-cAMP or the physiological agonists, Cicaprost or adenosine, significantly increased phosphorylation and nuclear export YAP and TAZ and inhibited TEAD-luciferase report gene activity. Similar effects were obtained by inhibiting RhoA activity with C3-transferase, its downstream kinase, ROCK, with Y27632, or actin-polymerisation with Latrunculin-B. Conversely, expression of constitutively-active RhoA reversed the inhibitory effects of forskolin on TEAD-luciferase. Forskolin significantly inhibited the mRNA expression of the pro-mitogenic genes, CCN1, CTGF, c-MYC and TGFB2 and this was reversed by expression of constitutively-active YAP or TAZ phospho-mutants. Inhibition of YAP and TAZ function with RNAi or Verteporfin significantly reduced VSMC proliferation. Furthermore, the anti-mitogenic effects of forskolin were reversed by overexpression of constitutively-active YAP or TAZ. Taken together, these data demonstrate that cAMP-induced actin-cytoskeleton remodelling inhibits YAP/TAZ-TEAD dependent expression of pro-mitogenic genes in VSMC. This mechanism contributes novel insight into the anti-mitogenic effects of cAMP in VSMC and suggests a new target for intervention. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  15. Astragalus extract inhibits proliferation but enhances apoptosis in gastric cancer.

    PubMed

    Wang, Zhengguang; Dong, Liuyi; Zhen, Yinan; Wang, Yanan; Qi, Dongjiang; Xu, Aman; Meng, Xiangling; Li, Weiping

    2016-09-01

    We and others have shown that Astragalus extract (AE) regulates various cellular processes including inflammation and apoptosis. It remains elusive whether and how AE modulates apoptosis in gastric cancer cells in vitro and in vivo. The objective of this study is to determine the effects and mechanisms of AE on the proliferation and apoptosis of human gastric cancer SGC-7901 cells and on tumor growth in orthotopic transplantation gastric tumor model in nude mice. Human gastric adenocarcinoma SGC-7901 cells and nude mice implanted with gastric cancer cells were treated with different concentration of AE and 5-fluorouracil as control. Cellular proliferation, apoptosis and tumor growth as well as interleukin (IL)-6/signal transducer and activator of transcription (Stat) 3 signals pathway were determined. We found that AE inhibited proliferation but caused apoptosis in human gastric cancer cells. Furthermore, the tumor growth and volume were reduced by AE administration in nude mice implanted with gastric cancer cells. In addition, treatments with AE decreased the expression of Bcl-2 proteins, whereas the expression of Bax was increased after AE treatment in tumor tissues of nude mice transplanted with human gastric cancer cells. This was associated with AE-mediated reduction of IL-6, phosphorylated Stat3, survivin and vascular endothelial growth factor. Overall, AE enhances apoptosis in gastric cancer cells in vitro and in vivo, which is associated with decreased activation of IL-6/Stat3 signals.

  16. Ganciclovir inhibits lymphocyte proliferation by impairing DNA synthesis.

    PubMed

    Battiwalla, Minoo; Wu, Yiyuan; Bajwa, Rajinder P S; Radovic, Marija; Almyroudis, Nikolaos G; Segal, Brahm H; Wallace, Paul K; Nakamura, Ryotaro; Padmanabhan, Swaminathan; Hahn, Theresa; McCarthy, Philip L

    2007-07-01

    Cytomegalovirus (CMV) disease-related mortality in allogeneic hematopoietic stem cell transplant (HSCT) recipients has dramatically declined because of ganciclovir prophylaxis and preemptive therapeutic strategies. However, ganciclovir has not improved overall survival in randomized studies despite effectively preventing overt CMV disease. Moreover, recurrent posttransplant CMV antigenemia, associated with prolonged ganciclovir exposure, is a predictor of increased relapse of malignancy. We examined the hypothesis that ganciclovir itself may have a negative impact on immune reconstitution by testing the effect of ganciclovir on normal human lymphocytes in vitro. T-lymphocyte activation and proliferation, as measured by PHA-induced (3)H-thymidine uptake, was greatly reduced at therapeutic concentrations of ganciclovir (10 microg/mL) but not for foscarnet (300 microM/L). Moreover, ganciclovir impaired bromodeoxyuridine incorporation in proliferating lymphocytes, but did not impair lymphocyte survival or induce lymphocyte apoptosis. Collectively, these results show that ganciclovir suppresses T-lymphocyte proliferation in vitro by inhibiting DNA synthesis; with implications for T-lymphocyte function following allogeneic BMT.

  17. RNA interference targeting raptor inhibits proliferation of gastric cancer cells

    SciTech Connect

    Wu, William Ka Kei; Lee, Chung Wa; Cho, Chi Hin; Chan, Francis Ka Leung; Yu, Jun; Sung, Joseph Jao Yiu

    2011-06-10

    Mammalian target of rapamycin complex 1 (mTORC1) is dysregulated in gastric cancer. The biologic function of mTORC1 in gastric carcinogenesis is unclear. Here, we demonstrate that disruption of mTORC1 function by RNA interference-mediated downregulation of raptor substantially inhibited gastric cancer cell proliferation through induction of G{sub 0}/G{sub 1}-phase cell cycle arrest. The anti-proliferative effect was accompanied by concomitant downregulation of activator protein-1 and upregulation of Smad2/3 transcriptional activities. In addition, the expression of cyclin D{sub 3} and p21{sup Waf1}, which stabilizes cyclin D/cdk4 complex for G{sub 1}-S transition, was reduced by raptor knockdown. In conclusion, disruption of mTORC1 inhibits gastric cancer cell proliferation through multiple pathways. This discovery may have an implication in the application of mTORC1-directed therapy for the treatment of gastric cancer.

  18. Inhibition of cell proliferation by the Mad1 transcriptional repressor.

    PubMed Central

    Roussel, M F; Ashmun, R A; Sherr, C J; Eisenman, R N; Ayer, D E

    1996-01-01

    Mad1 is a basic helix-loop-helix-leucine zipper protein that is induced upon differentiation of a number of distinct cell types. Mad1 dimerizes with Max and recognizes the same DNA sequences as do Myc:Max dimers. However, Mad1 and Myc appear to have opposing functions. Myc:Max heterodimers activate transcription while Mad:Max heterodimers repress transcription from the same promoter. In addition Mad1 has been shown to block the oncogenic activity of Myc. Here we show that ectopic expression of Mad1 inhibits the proliferative response of 3T3 cells to signaling through the colony-stimulating factor-1 (CSF-1) receptor. The ability of over-expressed Myc and cyclin D1 to complement the mutant CSF-1 receptor Y809F (containing a Y-to-F mutation at position 809) is also inhibited by Mad1. Cell cycle analysis of proliferating 3T3 cells transfected with Mad1 demonstrates a significant decrease in the fraction of cells in the S and G2/M phases and a concomitant increase in the fraction of G1 phase cells, indicating that Mad1 negatively influences cell cycle progression from the G1 to the S phase. Mutations in Mad1 which inhibit its activity as a transcription repressor also result in loss of Mad1 cell cycle inhibitory activity. Thus, the ability of Mad1 to inhibit cell cycle progression is tightly coupled to its function as a transcriptional repressor. PMID:8649388

  19. Stromelysin generates a fibronectin fragment that inhibits Schwann cell proliferation

    PubMed Central

    1992-01-01

    Our previous report (Muir, D., S. Varon, and M. Manthorpe. 1990. J. Cell Biol. 109:2663-2672) described the isolation and partial characterization of a 55-kD antiproliferative protein found in Schwann cell (SC) and schwannoma cell line-conditioned media and we concluded that SC proliferation is under negative autocrine control. In the present study the 55-kD protein was found to possess metalloprotease activity and stromelysin immunoreactivity. The SC-derived metalloprotease shares many properties with stromelysin isolated from other sources including the ability to cleave fibronectin (FN). Furthermore, limited proteolysis of FN by the SC-derived protease generated a FN fragment which itself expresses a potent antiproliferative activity for SCs. The active FN fragment corresponds to the 29-kD amino-terminal region of the FN molecule which was also identified as an active component in SC CM. Additional evidence that a proteolytic fragment of FN can possess antiproliferative activity for SCs was provided by the finding that plasmin can generate an amino- terminal FN fragment which mimicked the activity of the SC metalloprotease-generated antiproliferative FN fragment. Both the 55-kD SC metalloprotease and the 29-kD FN fragment could completely and reversibly inhibit proliferation of SCs treated with various mitogens and both were largely ineffective at inhibiting proliferation by immortalized or transformed SC lines. Normal and transformed SC types do secrete the proform of stromelysin, however, transformed cultures do not produce activated stromelysin and thus cannot generate the antiproliferative fragment of FN. These results suggest that, once activated, a SC-derived protease similar to stromelysin cleaves FN and generates an antiproliferative activity which can maintain normal SC quiescence in vitro. PMID:1730742

  20. Bruceantin inhibits multiple myeloma cancer stem cell proliferation.

    PubMed

    Issa, Mark E; Berndt, Sarah; Carpentier, Gilles; Pezzuto, John M; Cuendet, Muriel

    2016-09-01

    Multiple myeloma (MM) continues to claim the lives of a majority of patients. MM cancer stem cells (CSCs) have been demonstrated to sustain tumor growth. Due to their ability to self-renew and to express detoxifying enzymes and efflux transporters, MM-CSCs are rendered highly resistant to conventional therapies. Therefore, managing MM-CSCs characteristics could have profound clinical implications. Bruceantin (BCT) is a natural product previously demonstrated to inhibit the growth of MM in RPMI 8226 cells-inoculated mouse xenograft models, and to cause regression in already established tumors. The objectives of the present study were to test the inhibitory effects of BCT on MM-CSCs growth derived from a human primary tumor, and to explore a mechanism of action underlying these effects. BCT exhibited potent antiproliferative activity in MM-CSCs starting at 25 nM. BCT induced cell cycle arrest, cell death and apoptosis in MM-CSCs as well as inhibited cell migration and angiogenesis in vitro. Using a qPCR screen, it was found that the gene expression of a number of Notch pathway members was altered. Pretreatment of MM-CSCs with the γ-secretase inhibitor RO4929097, a Notch pathway inhibitor, reversed BCT-induced effects on MM-CSCs proliferation. In this study, BCT was shown to be an effective agent in controlling the proliferation, viability and migration of MM-CSCs as well as angiogenesis in vitro. The effect on MM-CSCs proliferation may be mediated by the Notch pathway. These results warrant further investigation of BCT in a broader set of human-derived MM-CSCs and with in vivo models representative of MM.

  1. Hammerhead ribozyme targeting connective tissue growth factor mRNA blocks transforming growth factor-beta mediated cell proliferation.

    PubMed

    Blalock, Timothy D; Yuan, Rong; Lewin, Alfred S; Schultz, Gregory S

    2004-06-01

    Excessive scarring following trauma or surgery of cornea, conjunctiva or retina can greatly impair visual outcome. At present, no agents are clinically available that selectively reduce activity of genes that regulate fibrosis. Connective tissue growth factor (CTGF) has been linked to fibrosis in several tissues, including cornea and conjunctiva. In this study, hammerhead ribozymes targeting CTGF mRNA were synthesized, kinetic parameters were measured, and the effect on TGF-beta-mediated cell proliferation was measured in cultured human fibroblasts. The mRNA sequence of human CTGF was scanned for potential hammerhead ribozyme cleavage sites, and predicted secondary folding structures around the sites were calculated. Synthetic 12mer ribozymes and 33mer oligonucleotide mRNA targets corresponding to two sites were synthesized, and kinetic constants calculated from Hanes-Wolff plots of in vitro cleavage reactions. The ribozyme with higher percentage cleavage and kinetic rate was cloned into an expression plasmid (pTR-UF21) and stably transfected into cultured human fibroblasts. An inactive ribozyme plasmid served as a negative control. The effects of the ribozyme on expression of TGF-beta-induced CTGF mRNA and protein levels were measured using ELISA and real-time TaqMan quantitative RT-PCR. Finally, the effect of the CTGF ribozyme on TGF-beta-mediated proliferation of fibroblasts was measured using a non-radioactive cell proliferation microtiter assay. Of the eight potential hammerhead ribozyme cleavage sites in human CTGF mRNA, two sites (CHR 745, and CHR 859) were identified with optimal secondary folding. CHR 859 cleaved 94% of the target mRNA, compared to 46% cleavage for CHR 745 after 16 hr of reaction. CHR 859 had a K(m) of 1.56 microM and a K(cat) of 2.97 min(-1), while CHR 745 had a K(m) of 7.80 microM and a K(cat) of 5.7 min(-1). The turnover numbers (K(cat)/K(m)) of CHR 859 and CHR 745 were 1.9 x 10(6) M min(-1) and 7.4 x 10(5) M min(-1), respectively

  2. Inhibition of TRPM7 by carvacrol suppresses glioblastoma cell proliferation, migration and invasion.

    PubMed

    Chen, Wen-Liang; Barszczyk, Andrew; Turlova, Ekaterina; Deurloo, Marielle; Liu, Baosong; Yang, Burton B; Rutka, James T; Feng, Zhong-Ping; Sun, Hong-Shuo

    2015-06-30

    Glioblastomas are progressive brain tumors with devastating proliferative and invasive characteristics. Ion channels are the second largest target class for drug development. In this study, we investigated the effects of the TRPM7 inhibitor carvacrol on the viability, resistance to apoptosis, migration, and invasiveness of the human U87 glioblastoma cell line.The expression levels of TRPM7 mRNA and protein in U87 cells were detected by RT-PCR, western blotting and immunofluorescence. TRPM7 currents were recorded using whole-cell patch-clamp techniques. An MTT assay was used to assess cell viability and proliferation. Wound healing and transwell experiments were used to evaluate cell migration and invasion. Protein levels of p-Akt/t-Akt, p-ERK1/2/t-ERK1/2, cleaved caspase-3, MMP-2 and phosphorylated cofilin were also detected.TRPM7 mRNA and protein expression in U87 cells is higher than in normal human astrocytes. Whole-cell patch-clamp recording showed that carvacrol blocks recombinant TRPM7 current in HEK293 cells and endogenous TRPM7-like current in U87 cells. Carvacrol treatment reduced the viability, migration and invasion of U87 cells. Carvacrol also decreased MMP-2 protein expression and promoted the phosphorylation of cofilin. Furthermore, carvacrol inhibited the Ras/MEK/MAPK and PI3K/Akt signaling pathways.Therefore, carvacrol may have therapeutic potential for the treatment of glioblastomas through its inhibition of TRPM7 channels.

  3. Inhibition of TRPM7 by carvacrol suppresses glioblastoma cell proliferation, migration and invasion

    PubMed Central

    Chen, Wen-Liang; Barszczyk, Andrew; Turlova, Ekaterina; Deurloo, Marielle; Liu, Baosong; Yang, Burton B.; Rutka, James T.; Feng, Zhong-Ping; Sun, Hong-Shuo

    2015-01-01

    Glioblastomas are progressive brain tumors with devastating proliferative and invasive characteristics. Ion channels are the second largest target class for drug development. In this study, we investigated the effects of the TRPM7 inhibitor carvacrol on the viability, resistance to apoptosis, migration, and invasiveness of the human U87 glioblastoma cell line. The expression levels of TRPM7 mRNA and protein in U87 cells were detected by RT-PCR, western blotting and immunofluorescence. TRPM7 currents were recorded using whole-cell patch-clamp techniques. An MTT assay was used to assess cell viability and proliferation. Wound healing and transwell experiments were used to evaluate cell migration and invasion. Protein levels of p-Akt/t-Akt, p-ERK1/2/t-ERK1/2, cleaved caspase-3, MMP-2 and phosphorylated cofilin were also detected. TRPM7 mRNA and protein expression in U87 cells is higher than in normal human astrocytes. Whole-cell patch-clamp recording showed that carvacrol blocks recombinant TRPM7 current in HEK293 cells and endogenous TRPM7-like current in U87 cells. Carvacrol treatment reduced the viability, migration and invasion of U87 cells. Carvacrol also decreased MMP-2 protein expression and promoted the phosphorylation of cofilin. Furthermore, carvacrol inhibited the Ras/MEK/MAPK and PI3K/Akt signaling pathways. Therefore, carvacrol may have therapeutic potential for the treatment of glioblastomas through its inhibition of TRPM7 channels. PMID:25965832

  4. Local anesthetics inhibit tissue factor expression in activated monocytes via inhibition of tissue factor mRNA synthesis.

    PubMed

    Kim, Ji-Eun; Kim, Ki Jun; Ahn, Wonsik; Han, Kyou-Sup; Kim, Hyun Kyung

    2011-01-01

    Local anesthetics have been reported to have anticoagulant properties, but the mechanisms responsible for this action are poorly understood. Here, we evaluated the in vitro effects of 3 local anesthetics--lidocaine, ropivacaine, and bupivacaine--on the tissue factor expression by monocytes. Monocytes from peripheral blood were stimulated with lipopolysaccharide (LPS) in the presence or absence of local anesthetics. All 3 local anesthetics inhibited the expression of tissue factor antigen and tissue factor activity in LPS-stimulated monocytes in a dose- and time-dependent manner and reduced tissue factor messenger RNA (mRNA) expression in endothelial cells and a monocytic cell line. None of the 3 drugs induced apoptosis or affected the viability of monocytes. Our findings that local anesthetics inhibited the tissue factor induction in activated monocytes by inhibiting tissue factor mRNA level may demonstrate the feasibility of using local anesthetics in hypercoagulable and inflammatory conditions.

  5. Shikonin Suppresses Skin Carcinogenesis via Inhibiting Cell Proliferation.

    PubMed

    Li, Wenjuan; Zhang, Chunjing; Ren, Amy; Li, Teena; Jin, Rong; Li, Guohong; Gu, Xin; Shi, Runhua; Zhao, Yunfeng

    2015-01-01

    The M2 isoform of pyruvate kinase M2 (PKM2) has been shown to be up-regulated in human skin cancers. To test whether PKM2 may be a target for chemoprevention, shikonin, a natural product from the root of Lithospermum erythrorhizon and a specific inhibitor of PKM2, was used in a chemically-induced mouse skin carcinogenesis study. The results revealed that shikonin treatment suppressed skin tumor formation. Morphological examinations and immunohistochemical staining of the skin epidermal tissues suggested that shikonin inhibited cell proliferation without inducing apoptosis. Although shikonin alone suppressed PKM2 activity, it did not suppress tumor promoter-induced PKM2 activation in the skin epidermal tissues at the end of the skin carcinogenesis study. To reveal the potential chemopreventive mechanism of shikonin, an antibody microarray analysis was performed, and the results showed that the transcription factor ATF2 and its downstream target Cdk4 were up-regulated by chemical carcinogen treatment; whereas these up-regulations were suppressed by shikonin. In a promotable skin cell model, the nuclear levels of ATF2 were increased during tumor promotion, whereas this increase was inhibited by shikonin. Furthermore, knockdown of ATF2 decreased the expression levels of Cdk4 and Fra-1 (a key subunit of the activator protein 1. In summary, these results suggest that shikonin, rather than inhibiting PKM2 in vivo, suppresses the ATF2 pathway in skin carcinogenesis.

  6. Betulin inhibits lung carcinoma proliferation through activation of AMPK signaling.

    PubMed

    Li, Xian-Dong; Zhang, Yi-Jie; Han, Ji-Chang

    2014-11-01

    Betulin (lup-20(29)-ene-3β, 28-diol) is an abundant, naturally occurring triterpene. It is commonly isolated from the bark of birch trees and forms up to 30% of the dry weight of the extractive. In the present study, we revealed its antiproliferative effects and mechanisms using two lung carcinoma cells (A549 and NCI-292). By 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and bromodeoxyuridine (BrdU) incorporation assays, we found that betulin could efficiently inhibit cell growth and proliferation. Besides, several key genes of cell-cycle regulators were also affected by betulin treatment. At the molecular level, our results demonstrated that treatment with betulin was also associated with activation of AMP kinase and inhibition of mTOR/p70S6K/pS6 signaling in these cells. In agreement, inhibition of AMPK signaling largely reversed the antiproliferative roles of betulin. Taken together, these data provide evidence for a mechanism that may contribute to the antineoplastic effects of betulin and justify further work to explore its potential roles in lung cancer prevention and treatment.

  7. Kuwanon V Inhibits Proliferation, Promotes Cell Survival and Increases Neurogenesis of Neural Stem Cells

    PubMed Central

    Kong, Sun-Young; Park, Min-Hye; Lee, Mina; Kim, Jae-Ouk; Lee, Ha-Rim; Han, Byung Woo; Svendsen, Clive N.; Sung, Sang Hyun; Kim, Hyun-Jung

    2015-01-01

    Neural stem cells (NSCs) have the ability to proliferate and differentiate into neurons and glia. Regulation of NSC fate by small molecules is important for the generation of a certain type of cell. The identification of small molecules that can induce new neurons from NSCs could facilitate regenerative medicine and drug development for neurodegenerative diseases. In this study, we screened natural compounds to identify molecules that are effective on NSC cell fate determination. We found that Kuwanon V (KWV), which was isolated from the mulberry tree (Morus bombycis) root, increased neurogenesis in rat NSCs. In addition, during NSC differentiation, KWV increased cell survival and inhibited cell proliferation as shown by 5-bromo-2-deoxyuridine pulse experiments, Ki67 immunostaining and neurosphere forming assays. Interestingly, KWV enhanced neuronal differentiation and decreased NSC proliferation even in the presence of mitogens such as epidermal growth factor and fibroblast growth factor 2. KWV treatment of NSCs reduced the phosphorylation of extracellular signal-regulated kinase 1/2, increased mRNA expression levels of the cyclin-dependent kinase inhibitor p21, down-regulated Notch/Hairy expression levels and up-regulated microRNA miR-9, miR-29a and miR-181a. Taken together, our data suggest that KWV modulates NSC fate to induce neurogenesis, and it may be considered as a new drug candidate that can regenerate or protect neurons in neurodegenerative diseases. PMID:25706719

  8. MiR-21 inhibits c-Ski signaling to promote the proliferation of rat vascular smooth muscle cells.

    PubMed

    Li, Jun; Zhao, Li; He, Xie; Yang, Ting; Yang, Kang

    2014-04-01

    Previously, we reported that the decrease of endogenous c-Ski expression is implicated in the progression of vascular smooth muscle cell (VSMC) proliferation after arterial injury. However, the molecular mechanism of the down-regulation of c-Ski is not clear. In this study, a potential miR-21 recognition element was identified in the 3'-untranslated region (UTR) of rat c-Ski mRNA. A reporter assay revealed that miR-21 could recognize the miR-21 recognition element of c-Ski mRNA. In A10 rat aortic smooth muscle cells, overexpression of miR-21 significantly inhibited the expression of c-Ski protein and promoted cell proliferation, which could be blocked by inhibition of miR-21 or overexpression of c-Ski. Further investigation demonstrated that the effect of miR-21 on VSMC proliferation resulted from negative regulation of c-Ski to suppress p38-p21/p27 signaling, the downstream pathway of c-Ski in VSMCs. These results indicate that c-Ski is a target gene of miR-21. miR-21 specifically binds to the 3'-untranslated region of c-Ski and negatively regulates c-Ski expression to diminish the protective effects of c-Ski and stimulate VSMC proliferation in the progression of arterial injury.

  9. A novel multi-target RNAi adenovirus inhibits hepatoma cell proliferation, migration, and induction of angiogenesis

    PubMed Central

    Pan, Tingting; Cheng, Ya; Ren, Weihua; Jia, Weidong; Ma, Jinliang; Xu, Geliang

    2016-01-01

    The pathogenesis of hepatocellular carcinoma (HCC) is a multi-step process involving many genes. Consequently, single gene targeting therapy has limited efficacy, making combination therapy targeting multiple genes a necessity. Based on our previous findings, we constructed a single vector mediating simultaneous expression of multiple short hairpin RNAs (shRNAs) against human vascular endothelial growth factor receptor 2 (VEGFR2), chemokine C-C motif receptor 1 (CCR1), and epithelial cell adhesion molecule (EpCAM), three genes closely related to HCC progression that act through separate pathways. The shRNA vector efficiently downregulated the mRNA and protein of all three molecules in Huh7 hepatoma cells. The vector also inhibited cell proliferation and migration and reduced angiogenesis. Furthermore, this shRNA vector can be recombined into adenovirus, a gene therapy vector, for better in vivo application. It thus offers a potentially effective future gene therapy approach to treating human liver cancer. PMID:27221035

  10. CXCL5 knockdown expression inhibits human bladder cancer T24 cells proliferation and migration

    SciTech Connect

    Zheng, Jiajia; Zhu, Xi; Zhang, Jie

    2014-03-28

    Highlights: • We first demonstrated CXCL5 is highly expressed in human bladder tumor tissues and cells. • CXCL5 knockdown inhibits proliferation, migration and promotes apoptosis in T24 cells. • CXCL5 knockdown inhibits Snail, PI3K-AKT and ERK1/2 signaling pathways in T24 cells. • CXCL5 is critical for bladder tumor growth and progression. - Abstract: CXCL5 (epithelial neutrophil activating peptide-78) which acts as a potent chemoattractant and activator of neutrophil function was reported to play a multifaceted role in tumorigenesis. To investigate the role of CXCL5 in bladder cancer progression, we examined the CXCL5 expression in bladder cancer tissues by real-time PCR and Western blot, additionally, we used shRNA-mediated silencing to generate stable CXCL5 silenced bladder cancer T24 cells and defined its biological functions. Our results demonstrated that mRNA and protein of CXCL5 is increased in human bladder tumor tissues and cell lines, down-regulation of CXCL5 in T24 cells resulted in significantly decreased cell proliferation, migration and increased cell apoptosis in vitro through Snail, PI3K-AKT and ERK1/2 signaling pathways. These data suggest that CXCL5 is critical for bladder tumor growth and progression, it may represent a potential application in cancer diagnosis and therapy.

  11. Andrographolide Inhibits Proliferation and Metastasis of SGC7901 Gastric Cancer Cells

    PubMed Central

    Dai, Lei; Wang, Gang

    2017-01-01

    To explore the mechanisms by which andrographolide inhibits gastric cancer cell proliferation and metastasis, we employed the gastric cell line SGC7901 to investigate the anticancer effects of andrographolide. The cell survival ratio, cell migration and invasion, cell cycle, apoptosis, and matrix metalloproteinase activity were assessed. Moreover, western blotting and real-time PCR were used to examine the protein expression levels and the mRNA expression levels, respectively. The survival ratio of cells decreased with an increasing concentration of andrographolide in a dose-dependent manner. Consistent results were also obtained using an apoptosis assay, as detected by flow cytometry. The cell cycle was blocked at the G2/M2 phase by andrographolide treatment, and the proportion of cells arrested at G1/M was enhanced as the dose increased. Similarly, wound healing and Transwell assays showed reduced migration and invasion of the gastric cancer cells at various concentrations of andrographolide. Andrographolide can inhibit cell proliferation, invasion, and migration, block the cell cycle, and promote apoptosis in SGC7901 cells. The mechanisms may include upregulated expression of Timp-1/2, cyclin B1, p-Cdc2, Bax, and Bik and downregulated expression of MMP-2/9 and antiapoptosis protein Bcl-2. PMID:28194420

  12. Overexpression of miR-21 promotes the proliferation and migration of cervical cancer cells via the inhibition of PTEN.

    PubMed

    Xu, Jingjie; Zhang, Wei; Lv, Qiongying; Zhu, Dingjun

    2015-06-01

    The oncogenic miR-21 has been widely recognized to promote the development and progression of various types of malignant tumors, but not cervical cancers. The aim of this study was to examine the expression of miR-21 and PTEN in cervical cancer specimens using quantitative PCR. The miR-21 level was then manipulated in the cervical cancer lines and the regulation of miR-21 on the proliferation, migration and invasion of cervical cancer cells was determined. Additionally, we determined the role of PTEN in the miR-21-regulated proliferation, migration and invasion of cervical cancer cells. miR-21 was upregulated in the cervical cancer specimens, negatively correlating with the PTEN mRNA level. Transfection of the miR-21 mimics was markedly promoted, whereas the miR-21 inhibitor suppressed the proliferation, migration and invasion of cervical cancer cells, with a significant inhibition of PTEN expression. In addition, the overexpression of PTEN markedly inhibited the proliferation and migration of the cervical cancer cells. The present study showed the upregulation of miR-21 in invasive cervical cancers, and confirmed the promotion of miR-21 with regard to the proliferation, migration and invasion in cervical cancer cells via inhibiting the PTEN expression. To the best of our knowledge, this is the first study to confirm that the miR-21/PTEN pathway promotes cervical cancer.

  13. Blockade of MUC1 expression by glycerol guaiacolate inhibits proliferation of human breast cancer cells.

    PubMed

    Smith, J S; Colon, J; Madero-Visbal, R; Isley, B; Konduri, S D; Baker, C H

    2010-10-01

    We sought to determine whether administration of glycerol guaiacolate at an optimal biological dose inhibits human breast cancer cell growth. Human breast cancer MCF-7 and ZR-75-1 cells were treated with glycerol guaiacolate and the therapeutic efficacy and biological activity of this drug was investigated on breast cancer cell growth. MCF-7 cells were injected into the mammary fat pad of overectamized female athymic nude mice. Ten days later, animals were treated with daily intraperitoneal injections of glycerol guaiacolate for six weeks. Tumor size and volume was monitored and immunohistochemistry analysis on MUC1, p21 and ki-67 was performed. Glycerol guaiacolate decreased breast cancer cell growth in a dose-dependent manner, decreased cell migration, and caused G1 cell cycle arrest. Our results demonstrate that glycerol guaiacolate inhibits MUC1 protein and mRNA expression levels and significantly increased p21 expression in human breast cancer cells as well as induced PARP cleavage. Similarly, glycerol guaiacolate inhibited breast tumor growth in vivo as well as enhanced p21 expression and decreased breast tumor cell proliferation (ki-67 expression). Collectively, our results demonstrate that glycerol guaiacolate decreased MUC1 expression and enhanced cell growth inhibition by inducing p21 expression in breast cancer cells. These findings suggest that glycerol guaiacolate may provide a novel and effective approach for the treatment of human breast cancer.

  14. Secreted ovarian stromal substance inhibits ovarian epithelial cell proliferation.

    PubMed

    Karlan, B Y; Baldwin, R L; Cirisano, F D; Mamula, P W; Jones, J; Lagasse, L D

    1995-10-01

    Determine the effects of factors secreted by normal human ovarian stroma on the proliferation of benign and malignant ovarian epithelia, in vitro. Primary cultures of normal human ovarian surface epithelium (HOSE), human ovarian stromal tissue (HOST), and epithelial ovarian carcinomas (CSOC) were established from surgical specimens and characterized immunohistochemically using anti-cytokeratin, vimentin, and Factor VIII antibodies. Stroma-conditioned media (SCM) were collected over 3 days from confluent HOST cultures. The SCM were dialyzed, lyophilized, resuspended, and added to HOSE, CSOC, SKOV-3, and Caov-3 ovarian cancer cell cultures and growth inhibitory effects were assayed by MTS and [3H]thymidine uptake. SCM inhibited the growth and DNA synthesis of normal HOSE cells and cancer cells by 79-99% in > 10-cell lines studied to date. The inhibitory effect was rapid in onset with 31-82% reduction in DNA synthesis at 1 hr and approximately 50% return of activity by 23 hr following a 1-hr SCM pulse treatment. The SCM inhibitory activity was not abolished by boiling or by absorption with heparin-agarose. Size exclusion filtration places the molecular weight of the inhibitory substance between 1 and 3 kDa. Neither trypsin nor proteinase K treatments altered the inhibitory activity of SCM, while a Bligh-Dyer organic extraction placed the activity in the aqueous phase. A heat-stable, non-heparin-binding, low-molecular-weight, water-soluble substance secreted by normal ovarian stroma significantly inhibits HOSE and ovarian cancer cell proliferation. Derangements in normal ovarian stroma-epithelial interactions may contribute to growth dysregulation of the surface epithelia and result in ovarian carcinogenesis.

  15. eIF4GI links nutrient sensing by mTOR to cell proliferation and inhibition of autophagy.

    PubMed

    Ramírez-Valle, Francisco; Braunstein, Steve; Zavadil, Jiri; Formenti, Silvia C; Schneider, Robert J

    2008-04-21

    Translation initiation factors have complex functions in cells that are not yet understood. We show that depletion of initiation factor eIF4GI only modestly reduces overall protein synthesis in cells, but phenocopies nutrient starvation or inhibition of protein kinase mTOR, a key nutrient sensor. eIF4GI depletion impairs cell proliferation, bioenergetics, and mitochondrial activity, thereby promoting autophagy. Translation of mRNAs involved in cell growth, proliferation, and bioenergetics were selectively inhibited by reduction of eIF4GI, as was the mRNA encoding Skp2 that inhibits p27, whereas catabolic pathway factors were increased. Depletion or overexpression of other eIF4G family members did not recapitulate these results. The majority of mRNAs that were translationally impaired with eIF4GI depletion were excluded from polyribosomes due to the presence of multiple upstream open reading frames and low mRNA abundance. These results suggest that the high levels of eIF4GI observed in many breast cancers might act to specifically increase proliferation, prevent autophagy, and release tumor cells from control by nutrient sensing.

  16. Oxidative stress induces proliferation of colorectal cancer cells by inhibiting RUNX3 and activating the Akt signaling pathway.

    PubMed

    Kang, Kyoung Ah; Kim, Ki Cheon; Bae, Suk Chul; Hyun, Jin Won

    2013-11-01

    We recently reported that the tumor suppressor Runt-related transcription factor 3 (RUNX3) is silenced in colorectal cancer cells via oxidative stress-induced hypermethylation of its promoter. The resulting downregulation of RUNX3 expression influences cell proliferation. Activation of the Akt signaling pathway is also associated with cell survival and proliferation; however, the effects of oxidative stress on the relationship between RUNX3 and Akt signaling are largely unknown. Therefore, this study investigated the mechanisms involved in cell proliferation caused by oxidative stress-induced silencing of RUNX3. The levels of RUNX3 mRNA and protein were downregulated in response to treatment of the human colorectal cancer cell line SNU-407 with H2O2. Treatment of the cells with H2O2 also upregulated Akt mRNA and protein expression, and inhibited the binding of RUNX3 to the Akt promoter. The inverse correlation between the expression levels of RUNX3 and Akt in H2O2-treated cells was also associated with nuclear translocation of β-catenin and upregulation of cyclin D1 expression, which induced cell proliferation. H2O2 treatment also increased the binding of β-catenin to the cyclin D1 promoter. The results presented here demonstrate that reactive oxygen species silence the tumor suppressor RUNX3, enhance the Akt-mediated signaling pathway, and promote the proliferation of colorectal cancer cells.

  17. Proliferation of macrophages due to the inhibition of inducible nitric oxide synthesis by oxidized low-density lipoproteins

    PubMed Central

    Brunner, Monika; Gruber, Miriam; Schmid, Diethart; Baran, Halina; Moeslinger, Thomas

    2015-01-01

    Oxidized low-density lipoprotein (ox-LDL) is assumed to be a major causal agent in hypercholesteraemia-induced atherosclerosis. Because the proliferation of lipid-loaden macrophages within atherosclerotic lesions has been described, we investigated the dependence of macrophage proliferation on the inhibition of inducible nitric oxide synthase (iNOS) by hypochlorite oxidized LDL. Ox-LDL induces a dose dependent inhibition of inducible nitric oxide synthesis in lipopolysaccharide-interferon stimulated mouse macrophages (J774.A1) with concomitant macrophage proliferation as assayed by cell counting, tritiated-thymidine incorporation and measurement of cell protein. Native LDL did not influence macrophage proliferation and inducible nitric oxide synthesis. iNOS protein and mRNA was reduced by HOCl-oxidized LDL (0-40 µg/ml) as revealed by immunoblotting and competitive semiquantitative PCR. Macrophage proliferation was increased by the addition of the iNOS inhibitor L-NAME. The addition of ox-LDL to L-NAME containing incubations induced no further statistically significant increase in cell number. Nitric oxide donors decreased ox-LDL induced macrophage proliferation and nitric oxide scavengers restored macrophage proliferation to the initial values achieved by ox-LDL. The decrease of cytosolic DNA fragments in stimulated macrophages incubated with ox-LDL demonstrates that the proliferative actions of ox-LDL are associated with a decrease of NO-induced apoptosis. Our data show that inhibition of iNOS dependent nitric oxide production caused by hypochlorite oxidized LDL enhances macrophage proliferation. This might be a key event in the pathogenesis of atherosclerotic lesions. PMID:26600745

  18. Progesterone inhibits proliferation and modulates expression of proliferation-Related genes in classical progesterone receptor-negative human BxPC3 pancreatic adenocarcinoma cells.

    PubMed

    Goncharov, Alexey I; Maslakova, Aitsana A; Polikarpova, Anna V; Bulanova, Elena A; Guseva, Alexandra A; Morozov, Ivan A; Rubtsov, Petr M; Smirnova, Olga V; Shchelkunova, Tatiana A

    2017-01-01

    Recent studies suggest that progesterone may possess anti-tumorigenic properties. However, a growth-modulatory role of progestins in human cancer cells remains obscure. With the discovery of a new class of membrane progesterone receptors (mPRs) belonging to the progestin and adipoQ receptor gene family, it becomes important to study the effect of this hormone on proliferation of tumor cells that do not express classical nuclear progesterone receptors (nPRs). To identify a cell line expressing high levels of mPRs and lacking nPRs, we examined mRNA levels of nPRs and three forms of mPRs in sixteen human tumor cell lines of different origin. High expression of mPR mRNA has been found in pancreatic adenocarcinoma BxPC3 cells, while nPR mRNA has not been detected in these cells. Western blot analysis confirmed these findings at the protein level. We revealed specific binding of labeled progesterone in these cells with affinity constant similar to that of human mPR expressed in yeast cells. Progesterone at high concentration of 20 μM significantly reduced the mRNA levels of proliferation markers Ki67 and PCNA, as well as of cyclin D1, and increased the mRNA levels of cyclin dependent kinase inhibitors p21 and p27. Progesterone (1 μM and 20 μM) significantly inhibited proliferative activity of BxPC3 cells. These results point to anti-proliferative effects of the progesterone high concentrations on BxPC3 cells and suggest that activation of mPRs may mediate this action. Our data are a starting point for further investigations regarding the application of progesterone in pancreatic cancer. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Viscum album-Mediated COX-2 Inhibition Implicates Destabilization of COX-2 mRNA

    PubMed Central

    Saha, Chaitrali; Hegde, Pushpa; Friboulet, Alain; Bayry, Jagadeesh; Kaveri, Srinivas V.

    2015-01-01

    Extensive use of Viscum album (VA) preparations in the complementary therapy of cancer and in several other human pathologies has led to an increasing number of cellular and molecular approaches to explore the mechanisms of action of VA. We have recently demonstrated that, VA preparations exert a potent anti-inflammatory effect by selectively down-regulating the COX-2-mediated cytokine-induced secretion of prostaglandin E2 (PGE2), one of the important molecular signatures of inflammatory reactions. In this study, we observed a significant down-regulation of COX-2 protein expression in VA-treated A549 cells however COX-2 mRNA levels were unaltered. Therefore, we hypothesized that VA induces destabilisation of COX-2 mRNA, thereby depleting the available functional COX-2 mRNA for the protein synthesis and for the subsequent secretion of PGE2. To address this question, we analyzed the molecular degradation of COX-2 protein and its corresponding mRNA in A549 cell line. Using cyclohexamide pulse chase experiment, we demonstrate that, COX-2 protein degradation is not affected by the treatment with VA whereas experiments on transcriptional blockade with actinomycin D, revealed a marked reduction in the half life of COX-2 mRNA due to its rapid degradation in the cells treated with VA compared to that in IL-1β-stimulated cells. These results thus demonstrate that VA-mediated inhibition of PGE2 implicates destabilization of COX-2 mRNA. PMID:25664986

  20. Conditioned Fear Inhibits c-fos mRNA Expression in the Central Extended Amygdala

    PubMed Central

    Day, Heidi E.W.; Kryskow, Elisa M.; Nyhuis, Tara J.; Herlihy, Lauren; Campeau, Serge

    2008-01-01

    We have shown previously that unconditioned stressors inhibit neurons of the lateral/capsular division of the central nucleus of the amygdala (CEAl/c) and oval division of the bed nucleus of the stria terminalis (BSTov), which form part of the central extended amygdala. The current study investigated whether conditioned fear inhibits c-fos mRNA expression in these regions. Male rats were trained either to associate a visual stimulus (light) with footshock or were exposed to the light alone. After training, animals were replaced in the apparatus, and 2 hours later injected remotely, via a catheter, with amphetamine (2 mg/kg i.p.), to induce c-fos mRNA and allow inhibition of expression to be measured. The rats were then presented with 15 visual stimuli over a 30 minute period. As expected, fear conditioned animals that were not injected with amphetamine, had extremely low levels of c-fos mRNA in the central extended amygdala. In contrast, animals that were trained with the light alone (no fear conditioning) and were injected with amphetamine had high levels of c-fos mRNA in the CEAl/c and BSTov. Animals that underwent fear-conditioning, and were re-exposed to the conditioned stimulus after amphetamine injection had significantly reduced levels of c-fos mRNA in both the BSTov and CEAl/c, compared to the non-conditioned animals. These data suggest that conditioned fear can inhibit neurons of the central extended amygdala. Because these neurons are GABAergic, and project to the medial CEA (an amygdaloid output region), this may be a novel mechanism whereby conditioned fear potentiates amygdaloid output. PMID:18634767

  1. Ginger phytochemicals exhibit synergy to inhibit prostate cancer cell proliferation

    PubMed Central

    Brahmbhatt, Meera; Gundala, Sushma R.; Asif, Ghazia; Shamsi, Shahab A; Aneja, Ritu

    2014-01-01

    Dietary phytochemicals offer non-toxic therapeutic management as well as chemopreventive intervention for slow-growing prostate cancers. However, the limited success of several single-agent clinical trials suggest a paradigm shift that the health benefits of fruits and vegetables are not ascribable due to individual phytochemicals rather may be ascribed to but to synergistic interactions among them. We recently reported growth-inhibiting and apoptosis-inducing properties of ginger extract (GE) in in vitro and in vivo prostate cancer models. Nevertheless, the nature of interactions among the constituent ginger biophenolics, viz. 6-gingerol, 8-gingerol, 10-gingerol, and 6-shogoal, remains elusive. Here we show antiproliferative efficacy of the most-active GE biophenolics as single-agents and in binary combinations, and investigate the nature of their interactions using the Chou-Talalay combination-index (CI) method. Our data demonstrate that binary combinations of ginger phytochemicals synergistically inhibit proliferation of PC-3 cells with CI values ranging from 0.03-0.88. To appreciate synergy among phytochemicals present in GE, the natural abundance of ginger biophenolics was quantitated using LC-UV/MS. Interestingly, combining GE with its constituents (in particular, 6-gingerol) resulted in significant augmentation of GE’s antiproliferative activity. These data generate compelling grounds for further preclinical evaluation of GE alone and in combination with individual ginger biophenols for prostate cancer management. PMID:23441614

  2. A Herpesvirus Protein Selectively Inhibits Cellular mRNA Nuclear Export.

    PubMed

    Gong, Danyang; Kim, Yong Hoon; Xiao, Yuchen; Du, Yushen; Xie, Yafang; Lee, Kevin K; Feng, Jun; Farhat, Nisar; Zhao, Dawei; Shu, Sara; Dai, Xinghong; Chanda, Sumit K; Rana, Tariq M; Krogan, Nevan J; Sun, Ren; Wu, Ting-Ting

    2016-11-09

    Nuclear mRNA export is highly regulated to ensure accurate cellular gene expression. Viral inhibition of cellular mRNA export can enhance viral access to the cellular translation machinery and prevent anti-viral protein production but is generally thought to be nonselective. We report that ORF10 of Kaposi's sarcoma-associated herpesvirus (KSHV), a nuclear DNA virus, inhibits mRNA export in a transcript-selective manner to control cellular gene expression. Nuclear export inhibition by ORF10 requires an interaction with an RNA export factor, Rae1. Genome-wide analysis reveals a subset of cellular mRNAs whose nuclear export is blocked by ORF10 with the 3' UTRs of ORF10-targeted transcripts conferring sensitivity to export inhibition. The ORF10-Rae1 interaction is important for the virus to express viral genes and produce infectious virions. These results suggest that a nuclear DNA virus can selectively interfere with RNA export to restrict host gene expression for optimal replication.

  3. Metformin inhibits the proliferation of human prostate cancer PC-3 cells via the downregulation of insulin-like growth factor 1 receptor

    SciTech Connect

    Kato, Haruo Sekine, Yoshitaka; Furuya, Yosuke; Miyazawa, Yoshiyuki; Koike, Hidekazu; Suzuki, Kazuhiro

    2015-05-22

    Metformin is a biguanide drug that is widely used for the treatment of type 2 diabetes. Recent studies have shown that metformin inhibits cancer cell proliferation and tumor growth both in vitro and in vivo. The anti-tumor mechanisms of metformin include activation of the AMP-activated protein kinase/mTOR pathway and direct inhibition of insulin/insulin-like growth factor (IGF)-mediated cellular proliferation. However, the anti-tumor mechanism in prostate cancer remains unclear. Because activation of the IGF-1 receptor (IGF-1R) is required for prostate cell proliferation, IGF-1R inhibitors may be of therapeutic value. Accordingly, we examined the effects of metformin on IGF-1R signaling in prostate cancer cells. Metformin significantly inhibited PC-3 cell proliferation, migration, and invasion. IGF-1R mRNA expression decreased significantly after 48 h of treatment, and IGF-1R protein expression decreased in a similar manner. IGF-1R knockdown by siRNA transfection led to inhibited proliferation, migration and invasion of PC-3 cells. IGF-1 activated both ERK1/2 and Akt, but these effects were attenuated by metformin treatment. In addition, intraperitoneal treatment with metformin significantly reduced tumor growth and IGF-1R mRNA expression in PC-3 xenografts. Our results suggest that metformin is a potent inhibitor of the IGF-1/IGF-1R system and may be beneficial in prostate cancer treatment. - Highlights: • Metformin inhibited PC-3 cell proliferation, migration, and invasion. • Metformin decreased IGF-1R mRNA and protein expressions in PC-3 cells. • Metformin inhibited IGF-1 induced ERK and Akt phosphorylations in PC-3 cells. • Metformin treatment inhibited PC-3 cell growth and IGF-1R expression in vivo. • Metformin may be a potent inhibitor of the IGF-1/IGF-1R signaling.

  4. Nitric oxide inhibits irreversibly P815 cell proliferation: involvement of potassium channels.

    PubMed

    Costa, R S A; Assreuy, J

    2002-12-01

    Nitric oxide (NO) has been shown to inhibit both normal and cancer cell proliferation. Potassium channels are involved in cell proliferation and, as NO activates these channels, we investigated the effect of NO on the proliferation of murine mastocytoma cell lines and the putative involvement of potassium channels. NO (in the form of NO donors) caused dose-dependent inhibition of cell proliferation in the P815 cell line inducing growth arrest in the mitosis phase. Incubation with NO donor for 4 or 24 h had a similar inhibitory effect on cell proliferation, indicating that this effect is irreversible. The inhibitory effect of NO was completely prevented by the blockade of voltage- and calcium-dependent potassium channels, but not by blockade of ATP-dependent channels. NO inhibition of cell proliferation was unaffected by guanylate cyclase and by cytoskeleton disruptors. Therefore, NO inhibits cell proliferation irreversibly via a potassium channel-dependent but guanylate cyclase-independent pathway in murine mastocytoma cells.

  5. Sulforaphane inhibits restenosis by suppressing inflammation and the proliferation of vascular smooth muscle cells.

    PubMed

    Kwon, Jin-Sook; Joung, Hosouk; Kim, Yong Sook; Shim, Young-Sun; Ahn, Youngkeun; Jeong, Myung Ho; Kee, Hae Jin

    2012-11-01

    Sulforaphane, a naturally occurring organosulfur compound in broccoli, has chemopreventive properties in cancer. However, the effects of sulforaphane in vascular diseases have not been examined. We therefore aimed to investigate the effects of sulforaphane on vascular smooth muscle cell (VSMC) proliferation and neointimal formation and the related mechanisms. The expression of vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1) was examined in VSMCs. The nuclear translocation of nuclear factor-κB (NF-κB) and GATA6 expression was examined in VSMCs and in a carotid artery injury model by Western blot and immunohistochemistry. We also investigated whether local delivery of sulforaphane affected neointimal formation. Sulforaphane inhibited the mRNA and protein expression of VCAM-1 induced by tumor necrosis factor (TNF)-α in VSMCs. Treatment of VSMCs with sulforaphane blocked TNF-α-induced IκBα degradation and NF-κB p65 and GATA6 expression. Furthermore, NF-κB p65 and GATA6 expression were reduced in sulforaphane-treated carotid injury sections. Notably, binding of GATA6 to the VCAM-1 promoter was dramatically reduced by sulforaphane. The MTT, BrdU incorporation, and in vitro scratch assays revealed that the proliferation and migration of VSMCs were reduced by sulforaphane. Furthermore, local administration of sulforaphane significantly reduced neointima formation 14 days after vascular injury in rats. Our results indicate that sulforaphane inhibits neointima formation via targeting of adhesion molecules through the suppression of NF-κB/GATA6. Furthermore, sulforaphane regulates migration and proliferation in VSMCs. Sulforaphane may be a potential therapeutic agent for preventing restenosis after vascular injury. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  6. Amicoumacin a inhibits translation by stabilizing mRNA interaction with the ribosome.

    PubMed

    Polikanov, Yury S; Osterman, Ilya A; Szal, Teresa; Tashlitsky, Vadim N; Serebryakova, Marina V; Kusochek, Pavel; Bulkley, David; Malanicheva, Irina A; Efimenko, Tatyana A; Efremenkova, Olga V; Konevega, Andrey L; Shaw, Karen J; Bogdanov, Alexey A; Rodnina, Marina V; Dontsova, Olga A; Mankin, Alexander S; Steitz, Thomas A; Sergiev, Petr V

    2014-11-20

    We demonstrate that the antibiotic amicoumacin A (AMI) is a potent inhibitor of protein synthesis. Resistance mutations in helix 24 of the 16S rRNA mapped the AMI binding site to the small ribosomal subunit. The crystal structure of bacterial ribosome in complex with AMI solved at 2.4 Å resolution revealed that the antibiotic makes contacts with universally conserved nucleotides of 16S rRNA in the E site and the mRNA backbone. Simultaneous interactions of AMI with 16S rRNA and mRNA and the in vivo experimental evidence suggest that it may inhibit the progression of the ribosome along mRNA. Consistent with this proposal, binding of AMI interferes with translocation in vitro. The inhibitory action of AMI can be partly compensated by mutations in the translation elongation factor G. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. Specific inhibition of mRNA translation by complementary oligonucleotides covalently linked to intercalating agents.

    PubMed Central

    Toulmé, J J; Krisch, H M; Loreau, N; Thuong, N T; Hélène, C

    1986-01-01

    Synthetic oligodeoxynucleotides that are covalently linked at their 3' end to an acridine derivative and are complementary to the repeated sequence UUAAAUUAAAUUAAA adjacent to the ribosome binding site of the gene 32-encoded mRNA from phage T4 have been used to regulate the synthesis of gene 32-encoded protein in vitro. These modified, synthetic oligonucleotides specifically block the translation of gene 32-encoded mRNA with a higher efficiency than the homologous unsubstituted oligonucleotides. The inhibition produced by these short "anti-messengers" is due to the formation of specific mRNA . oligodeoxynucleotide hybrids that are stabilized by the intercalation of the acridine ring in the RNA . DNA duplex. Images PMID:3513172

  8. Human peroxisome proliferator-activated receptor mRNA and protein expression during development

    EPA Science Inventory

    The peroxisome proliferator-activated receptors (PPAR) are nuclear hormone receptors that regulate lipid and glucose homeostasis and are important in reproduction and development. PPARs are targets ofpharmaceuticals and are also activated by environmental contaminants, including ...

  9. Human peroxisome proliferator-activated receptor mRNA and protein expression during development

    EPA Science Inventory

    The peroxisome proliferator-activated receptors (PPAR) are nuclear hormone receptors that regulate lipid and glucose homeostasis and are important in reproduction and development. PPARs are targets ofpharmaceuticals and are also activated by environmental contaminants, including ...

  10. Adiponectin-induced antitumor activity on prostatic cancers through inhibiting proliferation.

    PubMed

    Gao, Qiruo; Zheng, Junhua

    2014-09-01

    Adiponectin, an adipose tissue-derived hormone, has been studied intensively for the past decade because of its anti-inflammatory, anti-atherogenic, and anti-diabetic properties. Recent advances suggest that adiponectin also plays an important role in the development and progression of various cancers. Accumulating evidence suggests that adiponectin may have an important protective role in carcinogenesis. Adiponectin circulates at high concentrations in human plasma. Plasma levels of adiponectin are approximately 50 % lower in obese than in lean subjects. An association between low plasma levels of adiponectin and higher risk of developing prostate and other cancers was recently reported. Obesity and overweight have also been associated with increased mortality from cancer. To test the hypothesis that adiponectin exerts direct antiproliferative and/or pro-apoptotic effects on cancer cells, we used the PC-3 human prostate adenocarcinoma cell line. The proliferation rate of the PC-3 cells was measured using the MTT method, and apoptosis was examined by quantifying the DNA fragmentation using an ELISA assay. In addition, adiponectin receptor 1 (AdipoR1) and AdipoR2 mRNA expression was detected using RT-PCR. Adiponectin diminished the proliferation rate of PC-3 cells; this effect was significant after 48-96 h of treatment. The presence of receptor expression suggested that the effect of adiponectin on cell proliferation was most likely specific and adiponectin receptor-mediated. Adiponectin induced no apoptosis of PC-3 cells over 48 h. We conclude that adiponectin inhibits proliferation but causes no apoptosis of PC-3 prostate cancer cells.

  11. MicroRNA-195 inhibits proliferation of cervical cancer cells by targeting cyclin D1a.

    PubMed

    Wang, Ning; Wei, Heng; Yin, Duo; Lu, Yanming; Zhang, Yao; Zhang, Qiao; Ma, Xiaoxin; Zhang, Shulan

    2016-04-01

    Cervical cancer is one of the most frequent gynecological malignancies in women worldwide. MicroRNA-195 (miR-195) was recently found highly expressed in cervical cancer. However, the role of miR-195 in the pathology of cervical cancer remains poorly understood. In this study, we first confirmed the downregulation of miR-195 in primary cervical cancer tissues. For the functional study, we introduced the sequences of miR-195 or miR-195 inhibitor into Hela and SiHa cervical cancer cell lines. Overexpression of miR-195 inhibited the proliferation of both Hela and SiHa cells. In contrast, reducing the endogenous miR-195 level by miR-195 inhibitor promoted the proliferation of cervical cancer cells. Flow cytometric assay showed that overexpression of miR-195 induced G1 phase arrest, whereas miR-195 inhibitor shortened G1 phase of cervical cancer cells. In addition, the suppressive role of miR-195 in cell cycle was also demonstrated by the western blot results of various cell cycle indicators, such as phosphorylated retinoblastoma (p-Rb) and proliferating cell nuclear antigen (PCNA), in the gain and loss of function experiments. Furthermore, Dual-Luciferase Reporter Assay revealed that miR-195 targeted the 3'-untranslated region of cyclin D1a transcript, such as to regulate cyclin D1 expression. In summary, our results suggest that miR-195 acts as a suppressor in the proliferation and cell cycle of cervical cancer cells by directly targeting cyclin D1a mRNA.

  12. Inhibition of cyclooxygenase-2 and activation of peroxisome proliferator-activated receptor-gamma synergistically induces apoptosis and inhibits growth of human breast cancer cells.

    PubMed

    Michael, Michael S; Badr, Mostafa Z; Badawi, Alaa F

    2003-06-01

    Cyclooxygenase-2 (COX-2) expression and peroxisome proliferator-activated receptor-gamma (PPARgamma) inactivation are linked to increased risk of human breast cancer. This study examines the effect of simultaneous targeting of COX-2 and PPARgamma on the proliferation of human breast cancer cells and on the expression of Bcl-2, BAX, and caspases-3 and -9, modulators of apoptotic cell death. Treatment of MDA-MB-231 breast cancer cells with NS-398 (a COX-2 inhibitor) or ciglitazone (CGZ, a PPARgamma-ligand) significantly inhibited cell proliferation and markedly increased apoptotic rates. These effects were accompanied by upregulation of BAX and caspases-3 and -9 mRNA expression and downregulation of Bcl-2. Compared to the influence of separate treatments, simultaneous treatment with NS-398 and CGZ synergistically inhibited cell proliferation and induced apoptotic cell death. In conclusion, combinational targeting of COX-2 and PPARgamma can inhibit the growth of human breast cancer cells and induce apoptosis to an extent more suprior to that produced by targeting each molecule alone. COX-2 and PPARgamma can be promising molecular targets for combinational chemoprevention or treatment of breast cancer.

  13. Xanthohumol inhibits proliferation of laryngeal squamous cell carcinoma.

    PubMed

    Li, Yan; Wang, Kai; Yin, Shankai; Zheng, Hongliang; Min, Daliu

    2016-12-01

    Xanthohumol is a flavonoid compound that exhibits antioxidant and anticancer effects, and is used to treat atherosclerosis. The aim of the present study was to investigate the effect of xanthohumol on the cell proliferation of laryngeal squamous cell carcinoma and to understand the mechanism of its action. The effects of xanthohumol on the cell viability and apoptosis rate of laryngeal squamous cell carcinoma SCC4 cells were assessed by Annexin V-fluorescein isothiocyanate/propidium iodide staining. In addition, the expression levels of pro-apoptotic proteins, caspase-3, caspase-8, caspase-9, poly ADP ribose polymerase (PARP) p53 and apoptosis-inducing factor (AIF), as well as anti-apoptotic markers, B-cell lymphoma 2 (Bcl-2) and myeloid cell leukemia 1 (Mcl-1), were analyzed by western blotting. The results revealed that treatment with 40 µM xanthohumol significantly inhibited the proliferation of SCC4 cells. Furthermore, xanthohumol treatment (40 µM) induced SCC4 cell apoptosis, as indicated by the significant increase in activity and expression of caspase-3, caspase-8, caspase-9, PARP, p53 and AIF. By contrast, the protein expression of Bcl-2 and Mcl-1 was significantly decreased following treatment with 40 µM xanthohumol. Taken together, the results of the present study indicated that xanthohumol mediates growth suppression and apoptosis induction, which was mediated via the suppression of Bcl-2 and Mcl-1 and activation of PARP, p53 and AIF signaling pathways. Therefore, future studies that investigate xanthohumol as a potential therapeutic agent for laryngeal squamous cell carcinoma are required.

  14. Xanthohumol inhibits proliferation of laryngeal squamous cell carcinoma

    PubMed Central

    Li, Yan; Wang, Kai; Yin, Shankai; Zheng, Hongliang; Min, Daliu

    2016-01-01

    Xanthohumol is a flavonoid compound that exhibits antioxidant and anticancer effects, and is used to treat atherosclerosis. The aim of the present study was to investigate the effect of xanthohumol on the cell proliferation of laryngeal squamous cell carcinoma and to understand the mechanism of its action. The effects of xanthohumol on the cell viability and apoptosis rate of laryngeal squamous cell carcinoma SCC4 cells were assessed by Annexin V-fluorescein isothiocyanate/propidium iodide staining. In addition, the expression levels of pro-apoptotic proteins, caspase-3, caspase-8, caspase-9, poly ADP ribose polymerase (PARP) p53 and apoptosis-inducing factor (AIF), as well as anti-apoptotic markers, B-cell lymphoma 2 (Bcl-2) and myeloid cell leukemia 1 (Mcl-1), were analyzed by western blotting. The results revealed that treatment with 40 µM xanthohumol significantly inhibited the proliferation of SCC4 cells. Furthermore, xanthohumol treatment (40 µM) induced SCC4 cell apoptosis, as indicated by the significant increase in activity and expression of caspase-3, caspase-8, caspase-9, PARP, p53 and AIF. By contrast, the protein expression of Bcl-2 and Mcl-1 was significantly decreased following treatment with 40 µM xanthohumol. Taken together, the results of the present study indicated that xanthohumol mediates growth suppression and apoptosis induction, which was mediated via the suppression of Bcl-2 and Mcl-1 and activation of PARP, p53 and AIF signaling pathways. Therefore, future studies that investigate xanthohumol as a potential therapeutic agent for laryngeal squamous cell carcinoma are required. PMID:28105237

  15. The downregulation of OPN inhibits proliferation and migration and regulate activation of Erk1/2 in ECA-109 cells

    PubMed Central

    Xu, Song-Tao; Zou, Fa-Zhang; Cai, Li-Na; Xu, Wan-Ling

    2015-01-01

    Osteopontin (OPN) involves in tumor formation, and strongly correlated with the tumor progression. It was overexpressed in human esophageal squamous cell carcinoma (ESCC). To study the molecular mechanisms of OPN in ESCC, we examined its roles in inhibiting proliferation and invasion of ECA-109 (esophageal squamous cell carcinoma) cells. The expression of OPN gene was knockdown by RNA interference (RNAi) in the Eca-109 cell. The transcription level of OPN was to detect by reverse transcription-quantitative PCR (RT-qPCR). Western blot assay was performed to detect the expression of OPN, Caspase-3,Caspase-8, Caspase-9, ERK1/2, phospho-ERK1/2 and MMP2 after RNAi. The cell proliferation and apoptosis were detected by MTT and Hoechst33342 assay. Transwell inserts was used for detecting ECA-109 cell’s migration ability. The results shown that the level of OPN mRNA and protein was significantly reduced after RNAi. Proliferation and migration of cell line (ECA-109) was significantly inhibited in vitro. The protein phosphorylation and activation of ERK1/2 in the OPN RNAi group reduced significantly than the negative control groups. In Conclusion, the proliferation and migration of human ESCC can be inhibited by RNAi-targeting OPN. OPN can promote the expression of MMP2 through the ERK signaling pathways. OPN could serve as a potential therapeutic target for human ESCC. PMID:26131112

  16. Inositol hexaphosphate (IP6) inhibits cellular proliferation in melanoma.

    PubMed

    Rizvi, Irfan; Riggs, Dale R; Jackson, Barbara J; Ng, Alex; Cunningham, Cynthia; McFadden, David W

    2006-06-01

    Inositol Hexaphosphate (IP6) is a naturally occurring polyphosphorylated carbohydrate found in food sources high in fiber content. We have previously reported IP6 to have significant inhibitory effects against pancreatic cancer in vitro. We hypothesized that the IP6 would significantly inhibit cell growth of cutaneous melanoma in vitro. The melanoma line HTB68 was cultured using standard techniques and treated with IP6 at doses ranging from 0.2 to 1.0 mM/well. Cell viability was measured by MTT at 72 h. VEGF production was measured in the cell supernatants by ELISA. Apoptosis was evaluated by Annexin V-FITC and results calculated using FACS analysis. Statistical analysis was performed by ANOVA. Significant reductions (P < 0.001) in cellular proliferation were observed with IP6. Overall, IP6 exhibited a mean inhibition of cell growth of 52.1 +/- 11.5% (range, 1.6-83.0%) at 72 h of incubation. VEGF production was significantly reduced (P < 0.001) by the addition of IP6 (7.5 pg/ml) compared to control (40.9 pg/ml). IP6 significantly increased (P = 0.029) late apoptosis from 5.3 to 7.0% gated events. No changes in necrosis or early apoptosis were observed. Adjuvant treatment of melanoma continues to challenge clinicians and patients. Our findings that IP6 significantly decreased cellular growth, VEGF production and increased late apoptosis in melanoma suggest its potential therapeutic value. Further in vivo studies are planned to evaluate safety and clinical utility of this agent.

  17. Dehydroepiandrosterone inhibits cell proliferation and improves viability by regulating S phase and mitochondrial permeability in primary rat Leydig cells.

    PubMed

    Liu, Lin; Wang, Dian; Li, Longlong; Ding, Xiao; Ma, Haitian

    2016-07-01

    Dehydroepiandrosterone (DHEA) is widely used as a nutritional supplement and exhibits putative anti‑aging properties. However, the molecular basis of the actions of DHEA, particularly on the biological characteristics of target cells, remain unclear. The aim of the current study was to investigate the effects of DHEA on cell viability, cell proliferation, cell cycle and mitochondrial function in primary rat Leydig cells. Adult Leydig cells were purified by Percoll gradient centrifugation, and cell proliferation was detected using a Click-iT® EdU Assay kit and cell cycle assessment performed using flow cytometry. Mitochondrial membrane potential was detected using JC-1 staining assay. The results of the current study demonstrate that DHEA decreased cell proliferation in a dose‑dependent manner, whereas it improved cell viability in a time‑dependent and dose‑dependent manner. Flow cytometry analysis demonstrated that DHEA treatment increased the S phase cell population and decreased the G2/M cell population. Cyclin A and CDK2 mRNA levels were decreased in primary rat Leydig cells following DHEA treatment. DHEA treatment decreased the transmembrane electrical gradient in primary Leydig cells, whereas treatment significantly increased succinate dehydrogenase activity. These results indicated that DHEA inhibits primary rat Leydig cell proliferation by decreasing cyclin mRNA level, whereas it improves cells viability by modulating the permeability of the mitochondrial membrane and succinate dehydrogenase activity. These findings may demonstrate an important molecular mechanism by which DHEA activity is mediated.

  18. Regulation of PMP22 mRNA by G3BP1 affects cell proliferation in breast cancer cells

    PubMed Central

    2013-01-01

    Background Regulation of mRNAs is one way to control protein levels and thereby important cellular processes such as growth, invasion and apoptosis. G3BPs constitute a family of mRNA-binding proteins, shown to be overexpressed in several cancer types, including breast, colon and pancreas cancer. G3BP has been reported to both stabilize and induce degradation of specific mRNAs. Results Here, we show that G3BP1, but not G3BP2, supports proliferation of several breast cancer cell lines. Global gene expression analyses of G3BP1- and G3BP2-depleted cells indicate that primarily G3BP1, and much less G3BP2, influences mRNA expression levels. Peripheral myelin protein 22 (PMP22) was one gene that was significantly influenced by G3BP1 depletion which led to a 2–3 fold increased expression. Depletion of PMP22 resulted in increased proliferation and the G3BP1-mediated effect on proliferation was not seen upon PMP22-depletion. Conclusions This indicates a novel role for G3BP1 in the regulation of cell proliferation in breast cancer cells, perhaps via a regulatory effect on PMP22 expression. PMID:24321297

  19. Deviating the level of proliferating cell nuclear antigen in Trypanosoma brucei elicits distinct mechanisms for inhibiting proliferation and cell cycle progression.

    PubMed

    Valenciano, Ana L; Ramsey, Aaron C; Mackey, Zachary B

    2015-01-01

    The DNA replication machinery is spatially and temporally coordinated in all cells to reproduce a single exact copy of the genome per division, but its regulation in the protozoan parasite Trypanosoma brucei is not well characterized. We characterized the effects of altering the levels of proliferating cell nuclear antigen, a key component of the DNA replication machinery, in bloodstream form T. brucei. This study demonstrated that tight regulation of TbPCNA levels was critical for normal proliferation and DNA replication in the parasite. Depleting TbPCNA mRNA reduced proliferation, severely diminished DNA replication, arrested the synthesis of new DNA and caused the parasites to accumulated in G2/M. Attenuating the parasite by downregulating TbPCNA caused it to become hypersensitive to hydroxyurea. Overexpressing TbPCNA in T. brucei arrested proliferation, inhibited DNA replication and prevented the parasite from exiting G2/M. These results indicate that distinct mechanisms of cell cycle arrest are associated with upregulating or downregulating TbPCNA. The findings of this study validate deregulating intra-parasite levels of TbPCNA as a potential strategy for therapeutically exploiting this target in bloodstream form T. brucei.

  20. Overexpression of TOR (target of rapamycin) inhibits cell proliferation in Dictyostelium discoideum.

    PubMed

    Swer, Pynskhem Bok; Mishra, Himanshu; Lohia, Rakhee; Saran, Shweta

    2016-05-01

    TOR (target of rapamycin) protein kinase acts as a central controller of cell growth and development of an organism. Present study was undertaken to find the expression pattern and role of TOR during growth and development of Dictyostelium discoideum. Failures to generate either knockout and/or knockdown mutants indicate that interference with its levels led to cellular defects. Thus, the effects of TOR (DDB_G0281569) overexpression specifically, cells expressing Dd(Δ211-TOR)-Eyfp mutant was analyzed. Elevated expression of (Δ211-TOR)-Eyfp reduced both cell size and cell proliferation. DdTOR was found to be closer to fungus. mRNA level of TOR was found maximally in the freshly starved/aggregate cells that gradually declined. This was also strengthened by the expression patterns observed by in situ and the analysis of β-galactosidase reporter driven by the putative TOR promoter. The TOR protein was found to be highest at the aggregate stage. The fusion protein, (Δ211-TOR)-Eyfp was localized to the cell membrane, cytosol, and the nucleus. We suggest, DdTOR to be an essential protein and high TOR expression inhibits cell proliferation. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Trk inhibition reduces cell proliferation and potentiates the effects of chemotherapeutic agents in Ewing sarcoma

    PubMed Central

    Heinen, Tiago Elias; dos Santos, Rafael Pereira; da Rocha, Amanda; dos Santos, Michel Pinheiro; da Costa Lopez, Patrícia Luciana; Filho, Marco Aurélio Silva; Souza, Bárbara Kunzler; da Rosa Rivero, Luís Fernando; Becker, Ricardo Gehrke; Gregianin, Lauro José; Brunetto, Algemir Lunardi; Brunetto, André Tesainer; de Farias, Caroline Brunetto; Roesler, Rafael

    2016-01-01

    Ewing sarcoma (ES) is a highly aggressive pediatric cancer that may arise from neuronal precursors. Neurotrophins stimulate neuronal devlopment and plasticity. Here, we found that neurotrophins nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF), as well as their receptors (TrkA and TrkB, respectively) are expressed in ES tumors. Treatment with TrkA (GW-441756) or TrkB (Ana-12) selective inhibitors decreased ES cell proliferation, and the effect was increased when the two inhibitors were combined. ES cells treated with a pan-Trk inhibitor, K252a, showed changes in morphology, reduced levels of β-III tubulin, and decreased mRNA expression of NGF, BDNF, TrkA and TrkB. Furthermore, combining K252a with subeffective doses of cytotoxic chemotherapeutic drugs resulted in a decrease in ES cell proliferation and colony formation, even in chemoresistant cells. These results indicate that Trk inhibition may be an emerging approach for the treatment of ES. PMID:27145455

  2. NLS-RARα promotes proliferation and inhibits differentiation in HL-60 cells.

    PubMed

    Hu, Xiu-Xiu; Zhong, Liang; Zhang, Xi; Gao, Yuan-Mei; Liu, Bei-Zhong

    2014-01-01

    A unique mRNA produced in leukemic cells from a t(15;17) acute promyelocytic leukemia (APL) patient encodes a fusion protein between the retinoic acid receptor α (RARα) and a myeloid gene product called PML. Studies have reported that neutrophil elastase (NE) cleaves bcr-1-derived PML-RARα in early myeloid cells, leaving only the nuclear localization signal (NLS) of PML attached to RARα. The resultant NLS-RARα fusion protein mainly localizes to, and functions within, the cell nucleus. It is speculated that NLS-RARα may act in different ways from the wild-type RARα, but its biological characteristics have not been reported. This study takes two approaches. Firstly, the NLS-RARα was silenced with pNLS-RARα-shRNA. The mRNA and protein expression of NLS-RARα were detected by RT-PCR and Western blot respectively. Cell proliferation in vitro was assessed by MTT assay. Flow cytometry (FCM) was used to detect the differentiation of cells. Secondly, the NLS-RARα was over-expressed by preparation of recombinant adenovirus HL-60/pAd-NLS-RARα. The assays of mRNA and protein expression of NLS-RARα, and cell proliferation, were as above. By contrast, cell differentiation was stimulated by all trans retinoic acid (ATRA) (2.5µmol/L) at 24h after virus infection of pAd-NLS-RARα, and then detected by CD11b labeling two days later. The transcription and translation of C-MYC was detected in HL-60/pAd-NLS-RARα cells which treated by ATRA. Our results showed that compared to the control groups, the expression of NLS-RARα was significantly reduced in the HL-60/pNLS-RARα-shRNA cells, and increased dramatically in the HL-60/pAd-NLS-RARα cells. The proliferation was remarkably inhibited in the HL-60/pNLS-RARα-shRNA cells in a time-dependent manner, but markedly promoted in the HL-60/pAd-NLS-RARα cells. FCM outcome revealed the differentiation increased in HL-60/pNLS-RARα-shRNA cells, and decreased in the HL-60/pAd-NLS-RARα cells treated with 2.5µmol/L ATRA. The

  3. Silencing homeobox C6 inhibits colorectal cancer cell proliferation

    PubMed Central

    Tang, Wentao; Lao, Xinyuan; Zhu, Dexiang; Lin, Qi; Xu, Pingping; Wei, Ye; Xu, Jianmin

    2016-01-01

    Homeobox C6 (HOXC6), a member of the homeobox family that encodes highly conserved transcription factors, plays a vital role in various carcinomas. In this study, we used a tissue microarray (TMA) consisting of 462 CRC samples to demonstrate that HOXC6 is more abundantly expressed in colorectal cancer (CRC) tissues than adjacent normal mucosa. Clinicopathological data indicated that higher HOXC6 expression correlated with poor overall survival and was associated with primary tumor location in the right colon, primary tumor (pT) stage 3/4 and primary node (pN) stage 1/2. Multivariate analysis showed that high HOXC6 expression was an independent risk factor for poor CRC patient prognosis. HOXC6 downregulation via lentivirus-mediated expression of HOXC6-targeting shRNA reduced HCT116 cell viability and colony formation in vitro, and reduced growth of subcutaneous xenografts in nude mouse. HOXC6 thus appears to promote CRC cell proliferation and tumorigenesis through autophagy inhibition and mTOR pathway activation. PMID:27081081

  4. Patrinia scabiosaefolia inhibits the proliferation of colorectal cancer in vitro and in vivo via G1/S cell cycle arrest.

    PubMed

    Zhang, Mingyue; Sun, Guodong; Shen, Aling; Liu, Liya; Ding, Jingzhen; Peng, Jun

    2015-02-01

    Patrinia scabiosaefolia (PS) has long been used as an important component in traditional Chinese medicine formulas to treat gastrointestinal malignancies including colorectal cancer (CRC). We recently reported that PS can inhibit CRC growth through induction of apoptosis and inhibition of tumor angiogenesis. To further elucidate the mode of action of PS, in the present study, we used a CRC mouse xenograft model and a human CRC cell line HT-29 to evaluate the effect of the ethanol extract of PS (EEPS) on cancer cell proliferation and investigated the underlying molecular mechanisms. We found that EEPS inhibited CRC growth both in vivo and in vitro, which was associated with the inhibitory effects of EEPS on cancer cell proliferation. In addition, EEPS treatment significantly blocked G1 to S phase cell cycle progression in HT-29 cells. Moreover, EEPS treatment decreased the expression of pro-proliferative CyclinD1 and CDK4, at both the mRNA and protein levels. Thus, inhibition of cell proliferation via G1/S cell cycle arrest might be a potential mechanism whereby PS effectively treats cancers.

  5. Inhibition of mRNA export in vertebrate cells by nuclear export signal conjugates.

    PubMed

    Pasquinelli, A E; Powers, M A; Lund, E; Forbes, D; Dahlberg, J E

    1997-12-23

    Leucine-rich nuclear export signals (NESs) are recognized by the NES receptor exportin 1 and are central to the export of multiple shuttling proteins and RNAs. The export of messenger RNA in vertebrates was, however, thought to occur by a different pathway, because inhibition by injection of a synthetic Rev NES conjugate could not be demonstrated. Here we find that peptide conjugates composed of the NES of either protein kinase A inhibitor protein (PKI) or the HIV-1 Rev protein, when coupled to human serum albumin, are potent inhibitors of mRNA and small nuclear RNA export. These results provide direct evidence that mRNA export in vertebrates depends on interactions between an NES and its cognate NES receptors. PKI NES conjugates are significantly more efficient at inhibiting RNA export than are REV NES conjugates, indicating that different NESs may have different abilities to promote protein and RNA export. Surprisingly, an expected control conjugate containing the mutant Rev NES sequence M10 strongly inhibited the export of intronless dihydrofolate reductase mRNA. Nuclear injection of NES peptide conjugates led to mislocalization to the nucleus of 10-20% of the cytoplasmic Ran GTPase-binding protein (RanBP1) indicating that RanBP1 shuttles between the nucleus and the cytoplasm via an NES pathway. These results demonstrate that in vertebrates the export of mRNA, like that of small nuclear RNA, 5S rRNA, and transport factors such as RanBP1, employs NES-mediated molecular machinery.

  6. Antisense oligonucleotides adsorbed to polyalkylcyanoacrylate nanoparticles specifically inhibit mutated Ha-ras-mediated cell proliferation and tumorigenicity in nude mice.

    PubMed Central

    Schwab, G; Chavany, C; Duroux, I; Goubin, G; Lebeau, J; Hélène, C; Saison-Behmoaras, T

    1994-01-01

    Ras oncogenes owe their transforming properties to single point mutations in the sequence coding for the active site of the p21 protein. These mutations lead to changes in cellular proliferation and induce tumorigenic properties. Point mutations represent a well-defined target for antisense oligonucleotides that can specifically suppress the translation of the targeted mutant mRNA. We show that the stability and cellular disponibility of antisense oligonucleotides can be markedly improved by adsorption to polyalkylcyanoacrylate nanoparticles. Nanoparticle-adsorbed antisense oligonucleotides directed to a point mutation (G-->U) in codon 12 of the Ha-ras mRNA selectively inhibited the proliferation of cells expressing the point-mutated Ha-ras gene at a concentration 100 times lower than free oligonucleotides. In addition they markedly inhibited Ha-ras-dependent tumor growth in nude mice after subcutaneous injection. These experiments show that inhibition of ras oncogenes by antisense oligonucleotides can block tumor development even though ras oncogenic activation might be an early event in tumor progression. Images PMID:7937975

  7. SPARC expression in CML is associated to imatinib treatment and to inhibition of leukemia cell proliferation

    PubMed Central

    2013-01-01

    Background SPARC is a matricellular glycoprotein with growth-inhibitory and antiangiogenic activity in some cell types. The study of this protein in hematopoietic malignancies led to conflicting reports about its role as a tumor suppressor or promoter, depending on its different functions in the tumor microenvironment. In this study we investigated the variations in SPARC production by peripheral blood cells from chronic myeloid leukemia (CML) patients at diagnosis and after treatment and we identified the subpopulation of cells that are the prevalent source of SPARC. Methods We evaluated SPARC expression using real-time PCR and western blotting. SPARC serum levels were detected by ELISA assay. Finally we analyzed the interaction between exogenous SPARC and imatinib (IM), in vitro, using ATP-lite and cell cycle analysis. Results Our study shows that the CML cells of patients at diagnosis have a low mRNA and protein expression of SPARC. Low serum levels of this protein are also recorded in CML patients at diagnosis. However, after IM treatment we observed an increase of SPARC mRNA, protein, and serum level in the peripheral blood of these patients that had already started at 3 months and was maintained for at least the 18 months of observation. This SPARC increase was predominantly due to monocyte production. In addition, exogenous SPARC protein reduced the growth of K562 cell line and synergized in vitro with IM by inhibiting cell cycle progression from G1 to S phase. Conclusion Our results suggest that low endogenous SPARC expression is a constant feature of BCR/ABL positive cells and that IM treatment induces SPARC overproduction by normal cells. This exogenous SPARC may inhibit CML cell proliferation and may synergize with IM activity against CML. PMID:23383963

  8. Overexpression of Tau Downregulated the mRNA Levels of Kv Channels and Improved Proliferation in N2A Cells

    PubMed Central

    Li, Xiantao; Hu, Ximu; Li, Xiaoqing; Hao, Xuran

    2015-01-01

    Microtubule binding protein tau has a crucial function in promoting the assembly and stabilization of microtubule. Besides tuning the action potentials, voltage-gated K+ channels (Kv) are important for cell proliferation and appear to play a role in the development of cancer. However, little is known about the possible interaction of tau with Kv channels in various tissues. In the present study, tau plasmids were transiently transfected into mouse neuroblastoma N2A cells to explore the possible linkages between tau and Kv channels. This treatment led to a downregulation of mRNA levels of several Kv channels, including Kv2.1, Kv3.1, Kv4.1, Kv9.2, and KCNH4, but no significant alteration was observed for Kv5.1 and KCNQ4. Furthermore, the macroscopic currents through Kv channels were reduced by 36.5% at +60 mV in tau-tranfected N2A cells. The proliferation rates of N2A cells were also improved by the induction of tau expression and the incubation of TEA (tetraethylammonium) for 48 h by 120.9% and 149.3%, respectively. Following the cotransfection with tau in HEK293 cells, the mRNA levels and corresponding currents of Kv2.1 were significantly declined compared with single Kv2.1 transfection. Our data indicated that overexpression of tau declined the mRNA levels of Kv channels and related currents. The effects of tau overexpression on Kv channels provided an alternative explanation for low sensitivity to anti-cancer chemicals in some specific cancer tissues. PMID:25590133

  9. Overexpression of tau downregulated the mRNA levels of Kv channels and improved proliferation in N2A cells.

    PubMed

    Li, Xiantao; Hu, Ximu; Li, Xiaoqing; Hao, Xuran

    2015-01-01

    Microtubule binding protein tau has a crucial function in promoting the assembly and stabilization of microtubule. Besides tuning the action potentials, voltage-gated K+ channels (Kv) are important for cell proliferation and appear to play a role in the development of cancer. However, little is known about the possible interaction of tau with Kv channels in various tissues. In the present study, tau plasmids were transiently transfected into mouse neuroblastoma N2A cells to explore the possible linkages between tau and Kv channels. This treatment led to a downregulation of mRNA levels of several Kv channels, including Kv2.1, Kv3.1, Kv4.1, Kv9.2, and KCNH4, but no significant alteration was observed for Kv5.1 and KCNQ4. Furthermore, the macroscopic currents through Kv channels were reduced by 36.5% at +60 mV in tau-transfected N2A cells. The proliferation rates of N2A cells were also improved by the induction of tau expression and the incubation of TEA (tetraethylammonium) for 48 h by 120.9% and 149.3%, respectively. Following the cotransfection with tau in HEK293 cells, the mRNA levels and corresponding currents of Kv2.1 were significantly declined compared with single Kv2.1 transfection. Our data indicated that overexpression of tau declined the mRNA levels of Kv channels and related currents. The effects of tau overexpression on Kv channels provided an alternative explanation for low sensitivity to anti-cancer chemicals in some specific cancer tissues.

  10. Naltrindole Inhibits Human Multiple Myeloma Cell Proliferation In Vitro and in a Murine Xenograft Model In Vivo

    PubMed Central

    Mundra, Jyoti Joshi; Terskiy, Alexandra

    2012-01-01

    It has been demonstrated previously that immune cell activation and proliferation were sensitive to the effects of naltrindole, a nonpeptidic δ-opioid receptor-selective antagonist; therefore, we hypothesized that human multiple myeloma (MM) would be a valuable model for studying potential antineoplastic properties of naltrindole. [3H]naltrindole exhibited saturable, low-affinity binding to intact human MM cells; however, the pharmacological profile of the binding site differed considerably from the properties of δ-, κ-, and μ-opioid receptors, and opioid receptor mRNA was not detected in MM cells by reverse transcriptase-polymerase chain reaction. Naltrindole inhibited the proliferation of cultured human U266 MM cells in a time- and dose-dependent manner with an EC50 of 16 μM. The naltrindole-induced inhibition of U266 cell proliferation was not blocked by a 10-fold molar excess of naltrexone, a nonselective opioid antagonist. Additive inhibition of MM cell proliferation was observed when using a combination of naltrindole with the histone deacetylase inhibitor sodium valproate, the proteasome inhibitor bortezomib, the glucocorticoid receptor agonist dexamethasone, and the HMG CoA reductase inhibitor simvastatin. Treatment of U266 cells with naltrindole significantly decreased the level of the active, phosphorylated form of the kinases, extracellular signal-regulated kinase and Akt, which may be related to its antiproliferative activity. The antiproliferative activity of naltrindole toward MM cells was maintained in cocultures of MM and bone marrow-derived stromal cells, mimicking the bone marrow microenvironment. In vivo, naltrindole significantly decreased tumor cell volumes in human MM cell xenografts in severe combined immunodeficient mice. We hypothesize that naltrindole inhibits the proliferation of MM cells through a nonopioid receptor-dependent mechanism. PMID:22537770

  11. miRNA-145 inhibits VSMC proliferation by targeting CD40

    PubMed Central

    Guo, Xin; Li, Dai; Chen, Min; Chen, Lei; Zhang, Bikui; Wu, Tian; Guo, Ren

    2016-01-01

    Recent studies have demonstrated functions of miR-145 in vascular smooth muscle cells (VSMCs) phenotypes and vascular diseases. In this study, we aim to determine whether CD40 is involved in miR-145 mediated switch of VSMC phenotypes. In cultured VSMCs, the effects of miR-145 and CD40 on TNF-α, TGF-β, and Homocysteine (Hcy) induced cell proliferation were evaluated by over-expression of miR-145 or by siRNA-mediated knockdown of CD40. We also used ultrasound imaging to explore the effect of miR-145 on carotid artery intima-media thickness (CIMT) in atherosclerotic cerebral infarction (ACI) patients. The results showed 50 ng/mL TNF-α, 5 ng/mL TGF-β, and 500 μmol/L Hcy significantly increased the expression of CD40, both at mRNA and protein levels, and also induced the proliferation of VSMCs. We found that over-expression of miR-145 significantly inhibited the expression of CD40 and the differentiation of VSMCs, and over-expression of miR-145 decreased IL-6 levels in VSMC supernatants. In ACI patients, the lower expression of miR-145 was associated with thicker CIMT and higher levels of plasma IL-6. Our results suggest that the miR-145/CD40 pathway is involved in regulating VSMC phenotypes in TNF-α, TGF-β, and Hcy induced VSMCs proliferation model. Targeting miR-145/CD40 might be a useful strategy for treating atherosclerosis. PMID:27731400

  12. Pharmacological WNT Inhibition Reduces Proliferation, Survival and Clonogenicity of Glioblastoma Cells

    PubMed Central

    Kahlert, Ulf D.; Suwala, Abigail K.; Koch, Katharina; Natsumeda, Manabu; Orr, Brent A.; Hayashi, Masanori; Maciaczyk, Jarek; Eberhart, Charles G.

    2015-01-01

    Wingless (WNT) signaling has been shown to be an important pathway in gliomagenesis and in the growth of stem-like glioma cells. Using immunohistochemistry to assess translocation of β-catenin protein, we identified intranuclear staining, which suggest WNT pathway activation, in 8 of 43 (19%) adult and 9 of 30 (30%) pediatric glioblastoma patient surgical samples. WNT activity, evidenced by nuclear β-catenin in our cohort and high expression of its target AXIN2 in published glioma datasets, was associated with shorter patient survival, although this was not statistically significant. We determined the effects of the porcupine inhibitor LGK974 in 3 glioblastoma cell lines with elevated AXIN2 and found that it reduced WNT pathway activity by 50% or more as assessed by T cell factor-luciferase reporters. WNT inhibition led to suppression of growth and proliferation in the cultures and a modest induction of cell death. LGK974 reduced NANOG mRNA levels and the fraction of cells expressing the stem cell marker CD133 in neurosphere cultures, induced glial differentiation, and suppressed clonogenicity. These data indicate that LGK974 represents a promising new agent that can inhibit the canonical WNT pathway in vitro, slow tumor growth and deplete stem-like clonogenic cells, thereby providing further support for targeting WNT in patients with glioblastoma. PMID:26222502

  13. Hispidulin inhibits proliferation and enhances chemosensitivity of gallbladder cancer cells by targeting HIF-1α

    SciTech Connect

    Gao, Hui; Xie, Jing; Peng, Jianjun; Han, Yantao; Jiang, Qixiao; Han, Mei; Wang, Chunbo

    2015-03-15

    Gallbladder cancer (GBC) is an aggressive malignancy of the bile duct, which is associated with a low (5-year) survival and poor prognosis. The transcription factor HIF-1α is implicated in the angiogenesis, cell survival, epithelial mesenchymal transition (EMT) and invasiveness of GBC. In this study, we have investigated the role of HIF-1α in the pathobilogy of GBC and effect of hispidulin on the molecular events controlled by this transcription factor. We observed that hispidulin caused induction of apoptosis, blockade of growth and cell cycle progression in GBC cells. Our results have demonstrated for the first time that hispidulin-exerted anti-tumor effect involved the suppression of HIF-1α signaling. Hispidulin was found to repress the expression of HIF-1α protein dose-dependently without affecting the HIF-1α mRNA expression. In addition, the inhibition of HIF-1α protein synthesis was revealed to be mediated through the activation of AMPK signaling. Hispidulin also sensitized the tumor cells to Gemcitabine and 5-Fluoroucil by down-regulating HIF-1α/P-gp signaling. Given the low cost and exceedingly safe profile, hispidulin appears to be a promising and novel chemosensitizer for GBC treatment. - Highlights: • Hispidulin inhibits proliferation of gallbladder cancer cells by targeting HIF-1α. • Hispidulin regulates HIF-1α via activating AMPK signaling. • Hispidulin sensitized the GBC cells to chemotherapeutics by down-regulating P-gp.

  14. Cytochalasin releases mRNA from the cytoskeletal framework and inhibits protein synthesis.

    PubMed Central

    Ornelles, D A; Fey, E G; Penman, S

    1986-01-01

    Cytochalasin D was shown to be a reversible inhibitor of protein synthesis in HeLa cells. The inhibition was detectable at drug levels typically used to perturb cell structure and increased in a dose-dependent manner. The drug also released mRNA from the cytoskeletal framework in direct proportion to the inhibition of protein synthesis. The released mRNA was unaltered in its translatability as measured in vitro but was no longer translated in the cytochalasin-treated HeLa cells. The residual protein synthesis occurred on polyribosomes that were reduced in amount but displayed a normal sedimentation distribution. The results support the hypothesis that mRNA binding to the cytoskeletal framework is necessary although not sufficient for translation. Analysis of the cytoskeletal framework, which binds the polyribosomes, revealed no alterations in composition or amount of protein as a result of treatment with cytochalasin D. Electron microscopy with embedment-free sections shows the framework in great detail. The micrographs revealed the profound reorganization effected by the drug but did not indicate substantial disaggregation of the cytoskeletal elements. Images PMID:3785175

  15. Apigenin prevents UVB-induced cyclooxygenase 2 expression: coupled mRNA stabilization and translational inhibition.

    PubMed

    Tong, Xin; Van Dross, Rukiyah T; Abu-Yousif, Adnan; Morrison, Aubrey R; Pelling, Jill C

    2007-01-01

    Cyclooxygenase 2 (COX-2) is a key enzyme in the conversion of arachidonic acid to prostaglandins, and COX-2 overexpression plays an important role in carcinogenesis. Exposure to UVB strongly increased COX-2 protein expression in mouse 308 keratinocytes, and this induction was inhibited by apigenin, a nonmutagenic bioflavonoid that has been shown to prevent mouse skin carcinogenesis induced by both chemical carcinogens and UV exposure. Our previous study suggested that one pathway by which apigenin inhibits UV-induced and basal COX-2 expression is through modulation of USF transcriptional activity in the 5' upstream region of the COX-2 gene. Here, we found that apigenin treatment also increased COX-2 mRNA stability, and the inhibitory effect of apigenin on UVB-induced luciferase reporter gene activity was dependent on the AU-rich element of the COX-2 3'-untranslated region. Furthermore, we identified two RNA-binding proteins, HuR and the T-cell-restricted intracellular antigen 1-related protein (TIAR), which were associated with endogenous COX-2 mRNA in 308 keratinocytes, and apigenin treatment increased their localization to cell cytoplasm. More importantly, reduction of HuR levels by small interfering RNA inhibited apigenin-mediated stabilization of COX-2 mRNA. Cells expressing reduced TIAR showed marked resistance to apigenin's ability to inhibit UVB-induced COX-2 expression. Taken together, these results indicate that in addition to transcriptional regulation, another mechanism by which apigenin prevents COX-2 expression is through mediating TIAR suppression of translation.

  16. PML(NLS(-)) inhibits cell apoptosis and promotes proliferation in HL-60 cells.

    PubMed

    Gao, Yuan-Mei; Zhong, Liang; Zhang, Xi; Hu, Xiu-Xiu; Liu, Bei-Zhong

    2013-01-01

    Promyelocytic leukemia (PML) is a cell-growth suppressor, and PML-retinoic acid receptor α (PML-RARα) is known as a fusion gene of acute promyelocytic leukemia (APL). Studies have reported that neutrophil elastase(NE) cleaved bcr-1-derived PML-RARα in early myeloid cells leading to the removal of nuclear localization signal (NLS) from PML. The resultant PML without NLS named PML(NLS(-)). PML(NLS(-)) mainly locates and functions in the cytoplasm. PML(NLS(-)) may act in different ways from PML, but its biological characteristics have not been reported. In this study, the PML (NLS(-)) was silenced with shRNA [HL-60/pPML(NLS(-))-shRNA] and over-expressed by preparation of recombinant adenovirus [HL-60/pAd-PML(NLS(-))]. The mRNA and protein expression of PML(NLS(-)) were detected by RT-PCR and Western blot respectively. Cell proliferation in vitro was assessed by MTT assay. Flow cytometry (FCM) was used to detect apoptotic cells. The transcription of BCL-2, BAX and C-MYC was detected in HL-60/pAd-PML(NLS(-)) cells. Our results showed that compared to the control group, the expression of PML(NLS(-)) was significantly reduced in the HL-60/pPML(NLS(-))-shRNA cells, and increased significantly in the HL-60/pAd-PML(NLS(-)) cells. The proliferation was significantly inhibited in the HL-60/pPML(NLS(-))-shRNA cells in a time-dependent manner, but markedly promoted in the HL-60/pAd-PML(NLS(-)) cells treated with 60 μmol/L emodin. FCM revealed the apoptosis increased in HL-60/pPML(NLS(-))-shRNA cells, and decreased in the HL-60/pAd-PML(NLS(-)) cells. The expression of BAX decreased significantly, while that of BCL-2 and C-MYC increased significantly in HL-60/ pAd-PML(NLS(-)) cells. Down-regulation of PML(NLS(-)) expression inhibits the proliferation and induces the apoptosis of HL-60 cells. On the contrary, over-expression of PML(NLS(-)) promotes the proliferation and reduce the emodin-induced apoptosis of HL-60 cells.

  17. Podocalyxin promotes glioblastoma multiforme cell invasion and proliferation by inhibiting angiotensin-(1-7)/Mas signaling.

    PubMed

    Liu, Bo; Liu, Yu; Jiang, Yugang

    2015-05-01

    Podocalyxin (PODX) reportedly enhances invasion in many human cancers including glioblastoma multiforme (GBM). Recent studies have shown that the local renin-angiotensin system (RAS) in tumor environment contributes significantly to tumor progression. As a counter-regulatory axis in RAS, angiotensin (Ang)-(1-7)/Mas signaling has been shown to inhibit the growth and invasiveness of several human cancers including GBM. In the present study, we examined the crosstalk between PODX and Ang-(1-7)/Mas signaling in GBM cells, and assessed its impact on GBM cell invasion and proliferation. A strong negative correlation between the expression of PODX and Mas in GBM tumor tissues from 10 consecutive patients (r=-0.768, p<0.01) was observed. The stable overexpression of PODX in LN-229 and U-118 MG human GBM cells decreased the expression of Mas at the mRNA and protein levels, which led to decreased density of Ang-(1-7)-binding Mas on the cell membrane. This effect was completely abolished by selective phosphatidylinositol 3-kinase (PI3K) inhibitor BKM120. By contrast, the stable knockdown of PODX in LN-229 and U-118 MG cells increased the expression of Mas and the density of Ang-(1-7)-binding Mas on the cell membrane. Overexpression and knockdown of PODX respectively reversed and enhanced the inhibitory effects of Ang-(1-7) on the expression/activity of matrix metalloproteinase-9 and cell invasion and proliferation in GBM cells. Although the overexpression of Mas showed no significant effect on the promoting effect of PODX on GBM cell invasion and proliferation in the absence of Ang-(1-7), it completely eliminated the effect of PODX in the presence of Ang-(1-7). In conclusion, to the best of our knowledge, the present study provided the first evidence that PODX inhibits Ang-(1-7)/Mas signaling by downregulating the expression of Mas through a PI3K-dependent mechanism in GBM cells. This effect led to enhanced GBM cell invasion and proliferation. The results of this study add

  18. Compound Astragalus and Salvia miltiorrhiza extract suppresses hepatocellular carcinoma progression by inhibiting fibrosis and PAI-1 mRNA transcription.

    PubMed

    Rui, Wenjuan; Xie, Lei; Liu, Xin; He, Shufang; Wu, Chao; Zhang, Xiaoxiang; Zhang, Linjie; Yang, Yan

    2014-01-01

    Astragalus membranaceus and Salvia miltiorrhiza have been used for centuries in China to treat liver diseases. Previous studies have shown that these herbs and their extracts inhibit the development of liver fibrosis and the proliferation and invasion of human hepatoma HepG2 cells. Further study of their pharmacological effects on hepatocellular carcinoma (HCC) is needed. To investigate the effects of Compound Astragalus and Salvia miltiorrhiza Extract (CASE) on diethylinitrosamine (DEN)-induced hepatocarcinogenesis in rats. Male rats were divided into five groups, with the first group serving as normal control, the second group receiving 0.2% DEN solution five times a week for 14 weeks, and the third to fifth group receiving the same DEN as in the second group together with CASE at the doses of 60, 120, and 240 mg/kg per day for 16 weeks, respectively. Hepatoma incidence, serum enzymes levels, degree of fibrosis and hydroxyproline content were evaluated and compared across the five groups to determine CASE's suppression of fibrosis and HCC progression. In addition, an in vitro experiment using HepG2 cells was conduct to verify CASE's effect on the transcription of plasminogen activator inhibitor-1 (PAI-1) mRNA. CASE treatment significantly reduced the incidence and multiplicity of DEN-induced HCC development in a dose-dependent manner. It significantly suppressed the elevation of alanine transaminase, aspartate aminotransferase, gamma-glutamyl transferase, alkaline phosphatase, hyaluronic acid, direct bilirubin and total bilirubin, and significantly lessened the depression of serum total protein in DEN-induced HCC rats. CASE treatment also significantly suppressed the elevated expression of GST-P and α-SMA. The in vitro experiment confirmed that CASE inhibits the transcription of PAI-1 mRNA in HepG2 cells induced by TGF-β1 in a dose-dependent manner. CASE suppresses DEN-induced hepatocarcinogenesis by inhibiting fibrosis and PAI-1 mRNA transcription, suggesting

  19. Catechins inhibit angiotensin II-induced vascular smooth muscle cell proliferation via mitogen-activated protein kinase pathway.

    PubMed

    Won, Sun-Mi; Park, Youn-Hee; Kim, Hee-Jung; Park, Kwon-Moo; Lee, Won-Jung

    2006-10-31

    Catechins, components of green tea, reduce the incidence of cardiovascular diseases such as atherosclerosis. Angiotensin II (Ang II) is highly implicated in the proliferation of vascular smooth muscle cells (VSMC), resulting in atherosclerosis. The acting mechanisms of the catechins remain to be defined in the proliferation of VSMC induced by Ang II. Here we report that catechin, epicatechin (EC), epicatechingallate (ECG) or epigallocatechingallate (EGCG) significantly inhibits the Ang II-induced [3H]thymidine incorporation into the primary cultured rat aortic VSMC. Ang II increases the phosphorylation of the extracellular signal-regulated protein kinase 1/2 (ERK 1/2), c-jun-N-terminal kinase 1/2 (JNK 1/2), or p38 mitogen-activated protein kinases (MAPKs) and mRNA expression of c-jun and c-fos. The EGCG pretreatment inhibits the Ang II-induced phosphorylation of ERK 1/2, JNK 1/2, or p38 MAPK, and the expression of c-jun or c-fos mRNA. U0126, a MEK inhibitor, SP600125, a JNK inhibitor, or SB203580, a p38 inhibitor, attenuates the Ang II-induced [3H]thymidine incorporation into the VSMC. In conclusion, catechins inhibit the Ang II-stimulated VSMC proliferation via the inhibition of the Ang II-stimulated activation of MAPK and activator protein-1 signaling pathways. The antiproliferative effect of catechins may be associated with the reduced risk of cardiovascular diseases by the intake of green tea. Catechins may be useful in the development of prevention and therapeutics of vascular diseases.

  20. Gga-miR-21 inhibits chicken pre-adipocyte proliferation in part by down-regulating Kruppel-like factor 5.

    PubMed

    Wang, Weishi; Cheng, Min; Qiao, Shupei; Wang, Yuxiang; Li, Hui; Wang, Ning

    2017-01-01

    Gga-miR-21 is abundantly expressed in chicken pre-adipocytes, but its role is unclear. The present study investigated the role of gga-miR-21 in chicken pre-adipocyte proliferation. Cell proliferation assay and gene expression analysis of proliferating cell nuclear antigen (PCNA) showed that the gga-miR-21 mimic inhibited pre-adipocyte proliferation. In contrast, the gga-miR-21 inhibitor enhanced pre-adipocyte proliferation. The subsequent investigation identified Kruppel-like factor 5 (KLF5) mRNA as a target of gga-miR-21. The gga-miR-21 mimic inhibited KLF5 3(')UTR reporter activity and decreased endogenous KLF5 expression in primary pre-adipocytes. KLF5 knockdown using RNAi had a similar effect to that of the gga-miR-21 mimic on cell proliferation. The promoting effect of the gga-miR-21 inhibitor on pre-adipocyte proliferation was partially attenuated by KLF5 knockdown. Taken together, our results demonstrated that miR-21 inhibits chicken pre-adipocyte proliferation, at least in part, by targeting KLF5.

  1. JunD Is Required for Proliferation of Prostate Cancer Cells and Plays a Role in Transforming Growth Factor-β (TGF-β)-induced Inhibition of Cell Proliferation.

    PubMed

    Millena, Ana Cecilia; Vo, BaoHan T; Khan, Shafiq A

    2016-08-19

    TGF-β inhibits proliferation of prostate epithelial cells. However, prostate cancer cells in advanced stages become resistant to inhibitory effects of TGF-β. The intracellular signaling mechanisms involved in differential effects of TGF-β during different stages are largely unknown. Using cell line models, we have shown that TGF-β inhibits proliferation in normal (RWPE-1) and prostate cancer (DU145) cells but does not have any effect on proliferation of prostate cancer (PC3) cells. We have investigated the role of Jun family proteins (c-Jun, JunB, and JunD) in TGF-β effects on cell proliferation. Jun family members were expressed at different levels and responded differentially to TGF-β treatment. TGF-β effects on JunD protein levels, but not mRNA levels, correlated with its effects on cell proliferation. TGF-β induced significant reduction in JunD protein in RWPE-1 and DU145 cells but not in PC3 cells. Selective knockdown of JunD expression using siRNA in DU145 and PC3 cells resulted in significant reduction in cell proliferation, and forced overexpression of JunD increased the proliferation rate. On the other hand, knockdown of c-Jun or JunB had little, if any, effect on cell proliferation; overexpression of c-Jun and JunB decreased the proliferation rate in DU145 cells. Further studies showed that down-regulation of JunD in response to TGF-β treatment is mediated via the proteasomal degradation pathway. In conclusion, we show that specific Jun family members exert differential effects on proliferation in prostate cancer cells in response to TGF-β, and inhibition of cell proliferation by TGF-β requires degradation of JunD protein.

  2. Polypyrimidine tract-binding protein induces p19(Ink4d) expression and inhibits the proliferation of H1299 cells.

    PubMed

    Lin, Shankung; Wang, Ming Jen; Tseng, Kuo-Yun

    2013-01-01

    The expression of polypyrimidine tract-binding protein (PTB) is up-regulated in many types of cancer. Here, we studied the role of PTB in the growth of non small cell lung cancer cells. Data showed that PTB overexpression inhibited the growth of H1299 cells at least by inhibiting DNA synthesis. Quantitative real-time PCR and Western blot analyses showed that PTB overexpression in H1299 cells specifically induced the expression of p19(Ink4d), an inhibitor of cyclin-dependent kinase 4. Repression of p19(Ink4d) expression partially rescued PTB-caused proliferation inhibition. PTB overexpression also inhibited the growth and induced the expression of p19(Ink4d) mRNA in A549 cells. However, Western blot analyses failed to detect the presence of p19(Ink4d) protein in A549 cells. To address how PTB induced p19(Ink4d) in H1299 cells, we showed that PTB might up-regulate the activity of p19(Ink4d) gene (CDKN2D) promoter. Besides, PTB lacking the RNA recognition motif 3 (RRM3) was less effective in growth inhibition and p19(Ink4d) induction, suggesting that RNA-binding activity of PTB plays an important role in p19(Ink4d) induction. However, immunoprecipitation of ribonuclearprotein complexes plus quantitative real-time PCR analyses showed that PTB might not bind p19(Ink4d) mRNA, suggesting that PTB overexpression might trigger the other RNA-binding protein(s) to bind p19(Ink4d) mRNA. Subsequently, RNA electrophoretic mobility-shift assays revealed a 300-base segment (designated as B2) within the 3'UTR of p19(Ink4d) mRNA, with which the cytoplasmic lysates of PTB-overexpressing cells formed more prominent complexes than did control cell lysates. Insertion of B2 into a reporter construct increased the expression of the chimeric luciferase transcripts in transfected PTB-overexpressing cells but not in control cells; conversely, overexpression of B2-containing reporter construct in PTB-overexpressing cells abolished the induction of p19(Ink4d) mRNA. In sum, we have shown that

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

    SciTech Connect

    Yoon, Jeongyeon; Ryoo, Sungwoo

    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) 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.

  4. Cigarette smoke extract inhibits the proliferation of alveolar epithelial cells and augments the expression of P21WAF1.

    PubMed

    Jiao, Zongxian; Ao, Qilin; Ge, Xiaona; Xiong, Mi

    2008-02-01

    Cigarette smoking is intimately related with the development of chronic obstructive pulmonary diseases, and alveolar epithelium is a major target for the exposure of cigarette smoke extract. In order to investigate the effect of cigarette smoke extract on the proliferation of alveolar epithelial cell type II and its relationship with P21WAF1, the alveolar epithelial type II cell line (A549) cells were chosen as surrogate cells to represent alveolar epithelial type II cells. MTT assay was used to detect cell viability after interfered with different concentrations of cigarette smoke extract. It was observed cigarette smoke extract inhibited the growth of A549 cells in a dose-and time-dependent manner. The morphological changes, involving the condensation and margination of nuclear chromatin, even karyorrhexis, were observed by both Hoechst staining and electronic microscopy. Flow cytometry analysis demonstrated the increased cell percentages in G1 and subG1 phases after the cells were incubated with cigarette smoke extract. The expression of p21WAF1 protein and mRNA was also significantly increased as detected by the methods of Western blot or reverse transcription-polymerase chain reaction respectively. In conclusion, cigarette smoke extract inhibits the proliferation of alveolar epithelial cell type II and blocks them in G1/S phase. The intracellular accumulation of P21WAF1 may be one of the mechanisms which contribute to cigarette smoke extract-induced inhibition of cell proliferation.

  5. Spica Prunellae extract inhibits the proliferation of human colon carcinoma cells via the regulation of the cell cycle.

    PubMed

    Lin, Wei; Zheng, Liangpu; Zhuang, Qunchuan; Shen, Aling; Liu, Liya; Chen, Youqin; Sferra, Thomas J; Peng, Jun

    2013-10-01

    Spica Prunellae has long been used as a significant component in numerous traditional Chinese medicine (TCM) formulas to clinically treat cancers. Previously, Spica Prunellae was shown to promote cancer cell apoptosis and inhibit angiogenesis in vivo and in vitro. To further elucidate the precise mechanism of its tumoricidal activity, the effect of the ethanol extract of Spica Prunellae (EESP) on the proliferation of human colon carcinoma HT-29 cells was elucidated and the underlying molecular mechanisms were investigated. The proliferation of HT-29 cells was evaluated using 3-(4, 5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and colony formation analyses. The cell cycle was determined using fluorescence-activated cell sorting (FACS) with propidium iodide (PI) staining. The mRNA and protein expression of cyclin-dependent kinase 4 (CDK4) and cyclin D1 was examined using RT-PCR and western blotting, respectively. EESP was observed to inhibit HT-29 viability and survival in a dose- and time-dependent manner. Furthermore, EESP treatment blocked G1/S cell cycle progression and reduced the expression of pro-proliferative cyclin D1 and CDK4 at the transcriptional and translational levels. Altogether, these data suggest that the inhibition of cell proliferation via G1/S cell cycle arrest may be one of the mechanisms through which Spica Prunellae treats cancer.

  6. Spica Prunellae extract inhibits the proliferation of human colon carcinoma cells via the regulation of the cell cycle

    PubMed Central

    LIN, WEI; ZHENG, LIANGPU; ZHUANG, QUNCHUAN; SHEN, ALING; LIU, LIYA; CHEN, YOUQIN; SFERRA, THOMAS J.; PENG, JUN

    2013-01-01

    Spica Prunellae has long been used as a significant component in numerous traditional Chinese medicine (TCM) formulas to clinically treat cancers. Previously, Spica Prunellae was shown to promote cancer cell apoptosis and inhibit angiogenesis in vivo and in vitro. To further elucidate the precise mechanism of its tumoricidal activity, the effect of the ethanol extract of Spica Prunellae (EESP) on the proliferation of human colon carcinoma HT-29 cells was elucidated and the underlying molecular mechanisms were investigated. The proliferation of HT-29 cells was evaluated using 3-(4, 5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and colony formation analyses. The cell cycle was determined using fluorescence-activated cell sorting (FACS) with propidium iodide (PI) staining. The mRNA and protein expression of cyclin-dependent kinase 4 (CDK4) and cyclin D1 was examined using RT-PCR and western blotting, respectively. EESP was observed to inhibit HT-29 viability and survival in a dose- and time-dependent manner. Furthermore, EESP treatment blocked G1/S cell cycle progression and reduced the expression of pro-proliferative cyclin D1 and CDK4 at the transcriptional and translational levels. Altogether, these data suggest that the inhibition of cell proliferation via G1/S cell cycle arrest may be one of the mechanisms through which Spica Prunellae treats cancer. PMID:24137475

  7. Chemically modified tetracyclines selectively inhibit IL-6 expression in osteoblasts by decreasing mRNA stability.

    PubMed

    Kirkwood, Keith; Martin, Thomas; Andreadis, Stelios T; Kim, Young Joon

    2003-11-01

    In bone biology, interleukin (IL)-6 is an autocrine/paracrine cytokine which can induce osteoclasts formation and activation to help mediate inflammatory bone destruction. Previous studies have shown that tetracycline and its derivatives have potentially beneficial therapeutic effects in the prevention and treatment of metabolic bone diseases by modulating osteoblast and osteoclast activities. Our previous studies indicated that non-antimicrobial chemically modified tetracyclines (CMTs) can dose-dependently inhibit IL-1 beta-induced IL-6 secretion in osteoblastic cells. In the present study, we explored the molecular mechanisms underlying the ability of doxycycline analogs CMT-8 and its non-chelating pyrazole derivative, CMT-5 to affect IL-6 gene expression in murine osteoblasts. Steady-state IL-6 mRNA was decreased with CMT-8 (ca. 50%) but not by CMT-5 when stimulated by IL-1 beta. CMT-8 regulation of IL-1 beta-induced IL-6 gene expression was further explored. CMT-8 did not affect IL-6 promoter activity in reporter gene assays. However, the IL-6 mRNA stability was decreased in the presence of CMT-8. These effects require de novo protein synthesis as they were inhibited by cycloheximide. Western blot analysis indicated that CMT-8 did not affect p38 mitogen-activated protein kinase, c-jun NH(2)-terminal kinases, or extracellular signal-regulated kinases (1 and 2) phosphorylation in response to IL-1 beta. These data suggest that CMT-8 can modulate inhibit IL-1 beta-induced IL-6 expression in MC3T3-E1 cells at the post-transcriptional level affecting IL-6 mRNA stability. These observations may offer a novel molecular basis for this treatment of metabolic bone diseases that are mediated by IL-6.

  8. Inhibition of pyrimidine synthesis reverses viral virulence factor-mediated block of mRNA nuclear export

    PubMed Central

    Zhang, Liang; Das, Priyabrata; Schmolke, Mirco; Manicassamy, Balaji; Wang, Yaming; Deng, Xiaoyi; Cai, Ling; Tu, Benjamin P.; Forst, Christian V.; Roth, Michael G.; Levy, David E.; García-Sastre, Adolfo; de Brabander, Jef; Phillips, Margaret A.

    2012-01-01

    The NS1 protein of influenza virus is a major virulence factor essential for virus replication, as it redirects the host cell to promote viral protein expression. NS1 inhibits cellular messenger ribonucleic acid (mRNA) processing and export, down-regulating host gene expression and enhancing viral gene expression. We report in this paper the identification of a nontoxic quinoline carboxylic acid that reverts the inhibition of mRNA nuclear export by NS1, in the absence or presence of the virus. This quinoline carboxylic acid directly inhibited dihydroorotate dehydrogenase (DHODH), a host enzyme required for de novo pyrimidine biosynthesis, and partially reduced pyrimidine levels. This effect induced NXF1 expression, which promoted mRNA nuclear export in the presence of NS1. The release of NS1-mediated mRNA export block by DHODH inhibition also occurred in the presence of vesicular stomatitis virus M (matrix) protein, another viral inhibitor of mRNA export. This reversal of mRNA export block allowed expression of antiviral factors. Thus, pyrimidines play a necessary role in the inhibition of mRNA nuclear export by virulence factors. PMID:22312003

  9. BC047440 antisense eukaryotic expression vectors inhibited HepG2 cell proliferation and suppressed xenograft tumorigenicity.

    PubMed

    Zheng, Lu; Liang, Ping; Zhou, JianBo; Huang, XiaoBing; Wen, Yu; Wang, Zheng; Li, Jing

    2012-02-01

    The biological functions of the BC047440 gene highly expressed by hepatocellular carcinoma (HCC) are unknown. The objective of this study was to reconstruct antisense eukaryotic expression vectors of the gene for inhibiting HepG(2) cell proliferation and suppressing their xenograft tumorigenicity. The full-length BC047440 cDNA was cloned from human primary HCC by RT-PCR. BC047440 gene fragments were ligated with pMD18-T simple vectors and subsequent pcDNA3.1(+) plasmids to construct the recombinant antisense eukaryotic vector pcDNA3.1(+)BC047440AS. The endogenous BC047440 mRNA abundance in target gene-transfected, vector-transfected and naive HepG(2) cells was semiquantitatively analyzed by RT-PCR and cell proliferation was measured by the MTT assay. Cell cycle distribution and apoptosis were profiled by flow cytometry. The in vivo xenograft experiment was performed on nude mice to examine the effects of antisense vector on tumorigenicity. BC047440 cDNA fragments were reversely inserted into pcDNA3.1(+) plasmids. The antisense vector significantly reduced the endogenous BC047440 mRNA abundance by 41% in HepG(2) cells and inhibited their proliferation in vitro (P < 0.01). More cells were arrested by the antisense vector at the G(1) phase in an apoptosis-independent manner (P = 0.014). Additionally, transfection with pcDNA3.1(+)BC047440AS significantly reduced the xenograft tumorigenicity in nude mice. As a novel cell cycle regulator associated with HCC, the BC047440 gene was involved in cell proliferation in vitro and xenograft tumorigenicity in vivo through apoptosis-independent mechanisms.

  10. CBP knockdown inhibits angiotensin II-induced vascular smooth muscle cells proliferation through downregulating NF-kB transcriptional activity.

    PubMed

    Yang, Jian; Jiang, Hong; Chen, Si-si; Chen, Jing; Xu, Sheng-kai; Li, Wan-qiang; Wang, Ji-chun

    2010-07-01

    CREB binding protein (CBP), a powerful transcriptional co-activator for various transcriptional factors, regulates cell behavior in many cell types. Angiotensin II (Ang II) contributes to vascular lesion by promoting vascular smooth muscle cells (VSMCs) proliferation and migration. Therefore, we examined whether CBP knockdown could suppress Ang II-induced VSMCs proliferation, and elucidated its underlying molecular mechanism. We constructed lentiviral vector expressing CBP-specific short hairpin RNAs (shRNAs) that efficiently silenced CBP. VSMCs proliferation was evaluated by bromodeoxyuridine (BrdU) incorporation assay. Protein and mRNA expression of CBP and relevant cytokines were examined by Western blot, ELISA, and real-time PCR, respectively. We also used luciferase reporter gene and electrophoretic mobility shift assay (EMSA) to detect Nuclear factor kappaB (NF-kB) transcriptional activity and DNA binding. Meanwhile, NF-kB p65 subunit nuclear translocation was confirmed by immunoblotting. Lentiviral-mediated CBP-shRNAs at different multiplicities of infection (MOI = 100, 150) both significantly suppressed Ang II-induced CBP expression. Knockdown of CBP markedly inhibited Ang II-stimulated VSMCs proliferation and cytokines (TNF-alpha and IL-6) production. However, this inhibitory effect was not enhanced at MOI of 150 compared with MOI of 100 (P > 0.05). CBP siRNA showed the potent inhibition on Ang II-induced NF-kB transcriptional activity. Similarly, no significant difference was found between CBP siRNA lentivirus treatment groups. Furthermore, CBP gene silencing had no effect on NF-kB nuclear translocation and DNA binding. These findings suggest that CBP knockdown inhibits Ang II-induced VSMCs proliferation and the mechanism is involved with downregulation of NF-kB transcriptional activity, not through reduction in NF-kB nuclear translocation or DNA binding. Maintaining proper CBP level may be a potential therapeutic target for Ang II-induced cardiovascular

  11. SPARC is expressed by mesangial cells in experimental mesangial proliferative nephritis and inhibits platelet-derived-growth-factor-medicated mesangial cell proliferation in vitro.

    PubMed Central

    Pichler, R. H.; Bassuk, J. A.; Hugo, C.; Reed, M. J.; Eng, E.; Gordon, K. L.; Pippin, J.; Alpers, C. E.; Couser, W. G.; Sage, E. H.; Johnson, R. J.

    1996-01-01

    Mesangial cell proliferation is a characteristic feature of many glomerular diseases and often precedes extracellular matrix expansion and glomerulosclerosis. This study provides the first evidence that SPARC (secreted protein acidic and rich in cysteine) could be an endogenous factor mediating resolution of experimental mesangial proliferative nephritis in the rat. SPARC is a platelet-derived-growth-factor-binding glycoprotein that inhibits proliferation of endothelial cells and fibroblasts. We now show that SPARC is synthesized by mesangial cells in culture and that SPARC mRNA levels are increased by platelet-derived growth factor and basic fibroblast growth factor. Recombinant SPARC or the synthetic SPARC peptide 2.1 inhibited platelet-derived-growth-factor-induced mesangial cell DNA synthesis in vitro. In a model of experimental mesangioproliferative glomerulonephritis, SPARC mRNA was increased 5-fold by day 7 and was identified in the mesangium by in situ hybridization. Similarly, SPARC was increased in glomerular mesangial cells and visceral epithelial cells by day 5 and reached maximal expression levels by day 7. Mesangial cell proliferation increased by 36-fold on day 5 and decreased abruptly on day 7. Maximal expression of SPARC was correlated with the resolution of mesangial cell proliferation. We propose that SPARC functions in part as an endogenous inhibitor of platelet-derived-growth-factor-mediated mesangial cell proliferation in glomerulonephritis and that it could account for the resolution of cellular proliferation in this disease. Images Figure 1 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:8644857

  12. p53-dependent NDRG1 expression induces inhibition of intestinal epithelial cell proliferation but not apoptosis after polyamine depletion.

    PubMed

    Zhang, Ai-Hong; Rao, Jaladanki N; Zou, Tongtong; Liu, Lan; Marasa, Bernard S; Xiao, Lan; Chen, Jie; Turner, Douglas J; Wang, Jian-Ying

    2007-07-01

    Normal intestinal mucosal growth requires polyamines that regulate expression of various genes involved in cell proliferation, growth arrest, and apoptosis. Our previous studies have shown that polyamine depletion stabilizes p53, resulting in inhibition of intestinal epithelial cell (IEC) proliferation, but the exact downstream targets of induced p53 are still unclear. The NDRG1 (N-myc downregulated gene-1) gene encodes a growth-related protein, and its transcription can be induced in response to stress. The current study tests the hypothesis that induced p53 inhibits IEC proliferation by upregulating NDRG1 expression following polyamine depletion. Depletion of cellular polyamines by inhibiting ornithine decarboxylase (ODC) with alpha-difluoromethylornithine not only induced p53 but also increased NDRG1 transcription as indicated by induction of the NDRG1 promoter activity and increased levels of NDRG1 mRNA and protein, all of which were prevented by using specific p53 siRNA and in cells with a targeted deletion of p53. In contrast, increased levels of cellular polyamines by ectopic expression of the ODC gene decreased p53 and repressed expression of NDRG1. Consistently, polyamine depletion-induced activation of the NDRG1-promoter was decreased when p53-binding sites within the NDRG1 proximal promoter region were deleted. Ectopic expression of the wild-type NDRG1 gene inhibited DNA synthesis and decreased final cell numbers regardless of the presence or absence of endogenous p53, whereas silencing NDRG1 promoted cell growth. However, overexpression of NDRG1 failed to directly induce cell death and to alter susceptibility to apoptosis induced by tumor necrosis factor-alpha/cycloheximide. These results indicate that NDRG1 is one of the direct mediators of induced p53 following polyamine depletion and that p53-dependent NDRG1 expression plays a critical role in the negative control of IEC proliferation.

  13. Equine mesenchymal stem cells inhibit T cell proliferation through different mechanisms depending on tissue source.

    PubMed

    Carrade Holt, Danielle D; Wood, Joshua A; Granick, Jennifer L; Walker, Naomi J; Clark, Kaitlin C; Borjesson, Dori L

    2014-06-01

    Mesenchymal stem cells (MSCs) are used in both human clinical trials and veterinary medicine for the treatment of inflammatory and immune-mediated diseases. MSCs modulate inflammation by decreasing the cells and products of the inflammatory response. Stimulated equine MSCs from bone marrow (BM), adipose tissue (AT), cord blood (CB), and umbilical cord tissue (CT) inhibit lymphocyte proliferation and decrease inflammatory cytokine production. We hypothesized that equine MSCs inhibit T cell proliferation through secreted mediators and that MSCs from different tissue sources decrease T cell proliferation through different mechanisms. To test our hypotheses, we inhibited interleukin-6 (IL-6), nitric oxide (NO), and prostaglandin E2 (PGE2) to determine their impact on stimulated T cell proliferation. We also determined how equine MSCs modulate lymphocyte proliferation either via cell cycle arrest or apoptosis. Inhibition of IL-6 or NO did not reverse the immunomodulatory effect of MSCs on activated T cells. In contrast, inhibition of PGE2 restored T cell proliferation, restored the secretion of tumor necrosis factor-α and interferon-γ, and increased IL-10 levels. MSCs from solid-tissue-derived sources, AT and CT, inhibited T cell proliferation through induction of lymphocyte apoptosis while blood-derived MSCs, BM and CB, induced lymphocyte cell cycle arrest. Equine MSCs from different tissue sources modulated immune cell function by both overlapping and unique mechanisms. MSC tissue source may determine immunomodulatory properties of MSCs and may have very practical implications for MSC selection in the application of MSC therapy.

  14. Inhibition of proliferation of human lung cancer cells by green tea catechins is mediated by upregulation of let-7.

    PubMed

    Zhong, Zhiwei; Dong, Zhuo; Yang, Lihua; Chen, Xiaoqiang; Gong, Zhaohui

    2012-08-01

    Green tea catechins are known to function as anticancer agents via inhibition of carcinogenesis during the initiation, promotion and progression stages. Many potential mechanisms have been proposed, yet the precise mechanism of lung cancer prevention by green tea catechins remains unclear. microRNAs (miRs) are a class of 21-24 nucleotide small non-coding RNAs and play critical roles throughout cellular development and regulation. Emerging evidence demonstrates that tea catechins influence the expression of miRs in human cancer cells to inhibit tumorigenesis. Both let-7a-1 and let-7g were detected in the human lung cancer cells treated with tea catechins. The cell viability and cell cycle were analyzed after tea catechins treatment. In the present study, we found that tea catechins upregulated the tumor-suppressor miRs, let-7a-1 and let-7g, in lung cancer cell lines. The upregulation of let-7a/7g repressed the expression of their targets, C-MYC and the regulatory protein of LIN-28, at the mRNA and protein levels. Moreover, the cell growth assay indicated that tea catechins significantly inhibited cell proliferation, and the flow cytometric analysis revealed an increase in the number of cells in the G2/M phase and a decrease in the number of cells in the S phase after treatment with tea catechins. These observations suggest that green tea catechins mediate the inhibition of proliferation of lung cancer cells through the let-7 signaling pathway.

  15. Dietary feeding of Opuntia humifusa inhibits UVB radiation-induced carcinogenesis by reducing inflammation and proliferation in hairless mouse model.

    PubMed

    Lee, Jin-A; Jung, Bock-Gie; Kim, Tae-Hoon; Lee, Su-Gil; Park, Young-Seok; Lee, Bong-Joo

    2013-01-01

    It has been validated that ultraviolet B (UVB) irradiation induced both squamous and basal cell carcinomas, as a tumor initiator and promoter. Opuntia humifusa is a member of the Cactaceae family which has been demonstrated in our previous study to have a chemopreventive effect in 7, 12-dimethylbenz[a]anthracene and 12-O-tetradecanoylphorbol-13-acetate induced skin carcinogenesis models. Therefore, this study was designed to determine the protective effects of O. humifusa against photocarcinogenesis. O. humifusa was administrated to mice as a dietary feeding, following exposure to UVB radiation (180 mJ/cm(2)) twice a week of 30 weeks for skin tumor development in hairless mice. Dietary O. humifusa inhibited UVB-induced epidermal hyperplasia, infiltration of leukocytes, level of myeloperoxidase and the levels of proinflammatory cytokines, tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6), in UVB exposed skin. Also, O. humifusa significantly inhibited both protein and mRNA expression level of cyclooxygenase-2 (COX-2), nitric oxide synthase (iNOS), proliferating cell nuclear antigen (PCNA) and cyclin D1 compared to the non-O. humifusa treated group. Collectively, these results suggest that O. humifusa could inhibit photocarcinogenesis in mouse skin and that protective effect is associated with the inhibition of not only UVB-induced inflammatory responses involving COX-2, iNOS and proinflammatory cytokines, but also the down-regulation of UVB-induced cellular proliferation. © 2013 The American Society of Photobiology.

  16. CCL22 and IL-37 inhibit the proliferation and epithelial-mesenchymal transition process of NSCLC A549 cells.

    PubMed

    Chen, Yu-Hua; Zhou, Bi-Yun; Wu, Xian-Jing; Xu, Jun-Fa; Zhang, Jun-Ai; Chen, Yong-Hua; Liang, Si-Si

    2016-10-01

    In the present study, we aimed to investigate the effects of CC chemokine ligand 22 (CCL22) and interleukin-37 (IL-37) on the proliferation and epithelial-mesenchymal transition (EMT) of non-small cell lung cancer (NSCLC) A549 cells. pDsRed-CCL22 and pEGFP-IL-37 plasmids were constructed. A549 cells were divided into six groups: the control, the pDsRed-N1 blank plasmid, the pEGFP-C1 blank plasmid, the pDsRed-CCL22 plasmid, the pEGFP‑IL-37 plasmid and the pDsRed-CCL22 + pEGFP-IL-37 plasmid group. Expression levels and localization of CCL22 and IL-37 in cells were detected by confocal microscopy. Phase-contrast microscopy was applied for observing cellular morphology. Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) was used for detecting the mRNA levels of vimentin, N-cadherin and E-cadherin, and their protein expression levels were tested using western blotting. Constructed plasmids expressed CCL22 and IL-37, both of which had a co-localization in the cell membrane. MTT assay and cell observation results revealed that CCL22 and IL-37 inhibited the proliferation and EMT process of the A549 cells. The results of RT-qPCR and western blotting revealed that decreased vimentin and N-cadherin mRNA and protein expression levels, and increased E-cadherin mRNA and protein expression levels were found in the pDsRed-CCL22 plasmid, pEGFP-IL-37 plasmid and pDsRed‑CCL22 + pEGFP‑IL-37 plasmid groups when compared with the control, the pDsRed-N1 blank plasmid and the pEGFP-C1 blank plasmid groups (all P<0.05), and decreased vimentin and N-cadherin mRNA and protein expression levels and increased E-cadherin mRNA and protein expression levels were found in the pDsRed‑CCL22 + pEGFP‑IL-37 plasmid group when compared with the pDsRed-CCL22 plasmid and the pEGFP‑IL-37 plasmid groups (all P<0.05). CCL22 and IL-37 with a co-localization in the A549 cells inhibited the proliferation and EMT process in A549 cells. The antitumor effects of CCL22 and IL

  17. Silencing E1A mRNA by RNA interference inhibits adenovirus replication.

    PubMed

    Chung, Y-S; Kim, M-K; Lee, W-J; Kang, C

    2007-01-01

    The adenovirus family contains 51 human serotypes, and most human adenoviruses cause widespread respiratory tract infections. Adenovirus infections can result in severe complications in some cases, such as in adenovirus type 11 infection in immunocompromised patients. However, effective treatment methods for adenovirus infections are currently unavailable. This prompted the search for antiviral agents effective against adenovirus infections. In the present study, adenovirus E1A was targeted by RNA interference (RNAi) using synthetic small interfering RNAs (siRNAs) in an attempt to inhibit viral replication, since adenovirus E1A proteins are known to be involved in the transcriptional activation of the viral and cellular genes necessary for controlling the cell cycle and viral replication. The results indicated that the siRNAs effectively reduced the amount of adenovirus E1A mRNA and the levels of replicative intermediates. Additionally, siRNA-mediated gene silencing inhibited adenovirus replication by suppressing the E1A mRNA. These results suggest that the RNAi-mediated targeting of adenovirus E1A may have a potentially therapeutic effect in controlling adenovirus infections.

  18. Imatinib mesylate inhibits platelet derived growth factor stimulated proliferation of rheumatoid synovial fibroblasts

    SciTech Connect

    Sandler, Charlotta; Joutsiniemi, Saima; Lindstedt, Ken A.; Juutilainen, Timo; Kovanen, Petri T.; Eklund, Kari K. . E-mail: kari.eklund@hus.fi

    2006-08-18

    Synovial fibroblast is the key cell type in the growth of the pathological synovial tissue in arthritis. Here, we show that platelet-derived growth factor (PDGF) is a potent mitogen for synovial fibroblasts isolated from patients with rheumatoid arthritis. Inhibition of PDGF-receptor signalling by imatinib mesylate (1 {mu}M) completely abrogated the PDGF-stimulated proliferation and inhibited approximately 70% of serum-stimulated proliferation of synovial fibroblasts. Similar extent of inhibition was observed when PDGF was neutralized with anti-PDGF antibodies, suggesting that imatinib mesylate does not inhibit pathways other than those mediated by PDGF-receptors. No signs of apoptosis were detected in synovial fibroblasts cultured in the presence of imatinib. These results suggest that imatinib mesylate specifically inhibits PDGF-stimulated proliferation of synovial fibroblasts, and that inhibition of PDGF-receptors could represent a feasible target for novel antirheumatic therapies.

  19. Short hairpin RNA targeting NP mRNA inhibiting Newcastle disease virus production and other viral structural mRNA transcription.

    PubMed

    Yue, Hua; Deng, Shu; Yang, Fa-Long; Li, Ding-Fei; Fu, An-Jing; Yang, Fan; Tang, Cheng

    2009-02-01

    Newcastle disease virus (NDV), formally recognized as avian paramyxovirus 1 (APMV-1), is the etiological agent of Newcastle disease (ND), an affliction which can cause severe losses in the poultry industry. Better understanding of the molecular basis of viral structural genes involved with production should contribute significantly toward the development of improved prophylactic and therapeutic reagents to control the infection. Here we show that a short hairpin RNA (shRNA) eukaryotic expression vector targeting nucleocapsid (NP) gene of NDV can potently inhibit NDV production in both primary cells and embryonated chicken eggs. Moreover, shRNA specific for NP abolished the accumulation of not only the corresponding mRNA but also P, HN, F, M gene mRNA. The findings reveal that newly synthesized NP mRNA is essential for NDV transcription and replication, and provide a basis for the development of shRNAs as a prophylaxis and therapy for NDV infection in poultry.

  20. The transcription factor FOXN3 inhibits cell proliferation by downregulating E2F5 expression in hepatocellular carcinoma cells

    PubMed Central

    Huo, Qi; Cui, Meiling; Ge, Chao; Zhao, Fangyu; Tian, Hua; Chen, Taoyang; Yao, Ming; Li, Jinjun

    2016-01-01

    Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death worldwide, and the mechanisms underlying the development of HCC remain to be elucidated. Forkhead box N3 (FOXN3) is an important member of the FOX family of transcription factors that plays an essential role in several cancers but has not been investigated in HCC. In this study, we demonstrate that FOXN3 is downregulated in human primary HCC tissues compared with their matched adjacent liver tissues. Functional tests of FOXN3 demonstrated that FOXN3 inhibits the proliferation of HCC cells in vitro and in vivo. Additionally, FOXN3 repressed the mRNA and protein expression of E2F5, a reported potential oncogene, by inhibiting the promoter activity of E2F5. Collectively, our findings indicate that FOXN3 functions as a tumor suppressor in HCC by downregulating the expression of E2F5. PMID:27259277

  1. D1-like receptors inhibit insulin-induced vascular smooth muscle cell proliferation via down-regulation of insulin receptor expression

    PubMed Central

    Zeng, Chunyu; Han, Yu; Huang, Hefei; Yu, Changqing; Ren, Hongmei; Shi, Weibin; He, Duofen; Huang, Lan; Yang, Chengming; Wang, Xukai; Zhou, Lin; Jose, Pedro A.

    2013-01-01

    Objective Vascular smooth muscle cell (VSMC) proliferation is central to the development of vascular diseases, including hypertension, which is regulated by numerous hormones and humoral factors. Our previous study showed that the stimulatory effect of norepinephrine on VSMC proliferation is inhibited by D1-like receptors and the D3 dopamine receptor, a member of the D2-like receptor family. Insulin is a proliferative hormone but it is not known if there is any interaction between insulin and D1-like receptors. We hypothesized that Dl-like receptors may have an inhibitory effect on the insulin-induced VSMC proliferation; aberrant insulin and Dl-like receptor functions could be involved in the pathogenesis of essential hypertension. Methods VSMC proliferation was determined by [3H]-thymidine incorporation; insulin receptor mRNA and protein expressions were determined by RT-PCR, immunoblotting, and immunohistochemistry. Results Insulin increased VSMC proliferation in immortalized aortic A10 cells, determined by [3H]-thymidine incorporation. Although the D1-like receptor, by itself, had no effect on VSMC proliferation, stimulation with fenoldopam, a D1-like receptor agonist, inhibited the stimulatory effect of insulin. The inhibitory effect of fenoldopam on insulin-mediated VSMC proliferation was receptor specific, because its effect could be blocked by SCH23390, a D1-like receptor antagonist. Fenoldopam also inhibited insulin receptor mRNA and protein expression, which was time dependent and concentration dependent. A PKC or MAP kinase inhibitor blocked the inhibitory effect of fenoldopam on insulin receptor expression, indicating that PKC and MAP kinase were involved in the signaling pathway. Conclusion The inhibitory effect of D1-like receptors on insulin-mediated VSMC proliferation may play an important role in the regulation of blood pressure. PMID:19293728

  2. Amantadine inhibits cellular proliferation and induces the apoptosis of hepatocellular cancer cells in vitro.

    PubMed

    Lan, Zengmei; Chong, Zhaoyang; Liu, Cong; Feng, Danyang; Fang, Dihai; Zang, Weijin; Zhou, Jun

    2015-09-01

    Hepatocellular carcinoma (HCC) is one of the most aggressive malignancies worldwide, and its incidence associated with viral infection has increased in recent years. Amantadine is a tricyclic symmetric amine that can effectively protect against the hepatitis C virus. However, its antitumor properties remain unclear. In the present study, the effects of amantadine on tumor cell viability, cell cycle regulation and apoptosis were investigated. The growth of HepG2 and SMMC‑7721 cells (HCC cell lines) was detected by an MTT assay. Flow cytometry was used to investigate cell cycle regulation and apoptosis. Reverse transcription‑quantitative polymerase chain reaction and western blot analysis were also performed to examine the expression of cell cycle‑ and apoptosis‑related genes and proteins, including cyclin E, cyclin D1, cyclin‑dependent kinase 2 (CDK2), B‑cell lymphoma 2 (Bcl‑2) and Bax. Our results demonstrated that amantadine markedly inhibited the proliferation of HepG2 and SMMC‑7721 cells in a dose‑ and time‑dependent manner and arrested the cell cycle at the G0/G1 phase. The levels of the cell cycle‑related genes and proteins (cyclin D1, cyclin E and CDK2) were reduced by amantadine, and apoptosis was significantly induced. Amantadine treatment also reduced Bcl‑2 and increased the Bax protein and mRNA levels. Additionally, Bcl‑2/Bax ratios were lower in the two HCC cell lines following amantadine treatment. Collectively, these results emphasize the role of amantadine in suppressing proliferation and inducing apoptosis in HCC cells, advocating its use as a novel tumor-suppressive therapeutic candidate.

  3. Protective effect of silymarin against rapamycin-induced apoptosis and proliferation inhibition in endothelial progenitor cells.

    PubMed

    Zhang, Peng; Han, Guohua; Gao, Pei; Qiao, Kun; Ren, Yusheng; Liang, Chun; Leng, Bing; Wu, Zonggui

    2015-02-01

    For this study, peripheral blood samples were collected from human volunteers. Mononuclear cells (MNC) were separated by density centrifugation and were induced to differentiate into endothelial progenitor cells (EPCs) in vitro. Different concentrations of rapamycin and silymarin were introduced to the EPCs over 24 hours and then EPCs were analyzed for proliferation, migration, apoptosis and angiogenesis. Compared with the control group, rapamycin (1, 10, 100 ng/mL) inhibited the proliferation and migration of EPCs in a concentration dependent manner (P<0.05). Silymarin (50, 100 μg/mL) enhanced the proliferation and migration of EPCs and inhibited apoptosis in a concentration dependent manner (P<0.05). By adding rapamycin (1 ng/mL) and silymarin (25, 50, 100 μg/mL) over 24 hours, silymarin inhibited the pro-apoptotic effect of rapamycin on EPCs, and reversed the inhibition of proliferation, migration and angiogenesis of EPCs by rapamycin (P<0.05).

  4. Cyclosporin A Inhibits Smooth Muscle Proliferation in the Vascular Response to Injury

    NASA Astrophysics Data System (ADS)

    Jonasson, Lena; Holm, Jan; Hansson, Goran K.

    1988-04-01

    The arterial response to injury is dominated by proliferation of smooth muscle cells and infiltration of blood-borne cells in the vascular intima. Arterial smooth muscle cell proliferation is under growth factor control, but how this regulation operates in vivo is unclear. We studied the effect on arterial response to mechanical injury of cyclosporin A, a drug that inhibits T-lymphocyte activation. Cyclosporin A treatment at surgery caused a persistent inhibition of the intimal proliferative lesion. Cyclosporin A also inhibited expression of Ia antigens on smooth muscle cells in situ but had no direct effects on smooth muscle cell proliferation in culture. Therefore, the inhibition of intimal cell proliferation appears to be mediated via the immune system.

  5. Knockdown of REGγ inhibits proliferation by inducing apoptosis and cell cycle arrest in prostate cancer

    PubMed Central

    Chen, Shaojun; Wang, Longsheng; Xu, Chen; Chen, Hui; Peng, Bo; Xu, Yunfei; Yao, Xudong; Li, Lei; Zheng, Junhua

    2017-01-01

    Prostate cancer (PCa) is the most common malignant tumor and the second leading cause of cancer related death among men in western countries. REGγ, a proteasome activator, is reported to play important roles in various human cancers. However, the expression patterns and potential roles of REGγ in prostate cancer have never been reported. In this study, we found for the first time that REGγ is overexpressed in prostate cancer tissues and cell lines at both protein and mRNA levels. In addition, we demonstrated that knockdown of REGγ significantly inhibited cell proliferation and induced apoptosis and cell cycle arrest in PCa cell lines PC-3 and DU145. Moreover, we observed that the expressions of P21 were increased while the levels of cycinD1 and bcl-2 were decreased after knockdown of REGγ in PCa cells. And the stabilization of P21 was enhanced after REGγ knockdown in PC-3 cells. In summary, our findings suggest that REGγ may play important roles in prostate cancer and may serve as a novel therapeutic target in the treatment of PCa patients. PMID:28861169

  6. Thyroid hormone inhibits the proliferation of piglet Sertoli cell via PI3K signaling pathway.

    PubMed

    Sun, Yan; Yang, WeiRong; Luo, HongLin; Wang, XianZhong; Chen, ZhongQiong; Zhang, JiaoJiao; Wang, Yi; Li, XiaoMin

    2015-01-01

    Accumulating researches show that thyroid hormone (TH) inhibits Sertoli cells (SCs) proliferation and stimulates their functional maturation in prepubertal rat testis, confirming that TH plays a key role in testicular development. However, the mechanism under the T3 regulation of piglet SC proliferation remains unclear. In the present study, in order to investigate the possible mechanism of T3 on the suppression of SC proliferation, the expression pattern of TRα1 and cell cycle-related molecules, effect of T3 on SC proliferation, and the role of phosphoinositide 3-kinase (PI3K)/Akt signaling pathway on the T3-mediated SC proliferation in piglet testis were explored. Our results demonstrated that TRα1 was expressed in all tested stages of SCs and decreased along with the ages. T3 inhibited the proliferation of SCs in a time- and dose-dependent manner, and T3 treatment downregulated the expressions of cell cycling molecules, such as cyclinA2, cyclinD1, cyclinE1, PCNA, and Skp2, but upregulated the p27 expression in SCs. Most importantly, the suppressive effects of T3 on SC proliferation seemed dependent on the inhibition of PI3K/Akt signaling pathway, and pre-stimulation of PI3K could enhance such suppressive effects. Together, our findings demonstrate that TH inhibits the proliferation of piglet SCs via the suppression of PI3K/Akt signaling pathway.

  7. Suberoylanilide hydroxamic acid (SAHA) inhibits EGF-induced cell transformation via reduction of cyclin D1 mRNA stability

    SciTech Connect

    Zhang, Jingjie; Ouyang, Weiming; Li, Jingxia; Zhang, Dongyun; Yu, Yonghui; Wang, York; Li, Xuejun; Huang, Chuanshu

    2012-09-01

    Suberoylanilide hydroxamic acid (SAHA) inhibiting cancer cell growth has been associated with its downregulation of cyclin D1 protein expression at transcription level or translation level. Here, we have demonstrated that SAHA inhibited EGF-induced Cl41 cell transformation via the decrease of cyclin D1 mRNA stability and induction of G0/G1 growth arrest. We found that SAHA treatment resulted in the dramatic inhibition of EGF-induced cell transformation, cyclin D1 protein expression and induction of G0/G1 growth arrest. Further studies showed that SAHA downregulation of cyclin D1 was only observed with endogenous cyclin D1, but not with reconstitutionally expressed cyclin D1 in the same cells, excluding the possibility of SAHA regulating cyclin D1 at level of protein degradation. Moreover, SAHA inhibited EGF-induced cyclin d1 mRNA level, whereas it did not show any inhibitory effect on cyclin D1 promoter-driven luciferase reporter activity under the same experimental conditions, suggesting that SAHA may decrease cyclin D1 mRNA stability. This notion was supported by the results that treatment of cells with SAHA decreased the half-life of cyclin D1 mRNA from 6.95 h to 2.57 h. Consistent with downregulation of cyclin D1 mRNA stability, SAHA treatment also attenuated HuR expression, which has been well-characterized as a positive regulator of cyclin D1 mRNA stability. Thus, our study identifies a novel mechanism responsible for SAHA inhibiting cell transformation via decreasing cyclin D1 mRNA stability and induction of G0/G1 growth arrest in Cl41 cells. -- Highlights: ► SAHA inhibits cell transformation in Cl41 cells. ► SAHA suppresses Cyclin D1 protein expression. ► SAHA decreases cyclin D1 mRNA stability.

  8. Evidence That Base-pairing Interaction between Intron and mRNA Leader Sequences Inhibits Initiation of HAC1 mRNA Translation in Yeast*

    PubMed Central

    Sathe, Leena; Bolinger, Cheryl; Mannan, M. Amin-ul; Dever, Thomas E.; Dey, Madhusudan

    2015-01-01

    The Hac1 transcription factor in yeast up-regulates a collection of genes that control protein homeostasis. Base-pairing interactions between sequences in the intron and the 5′-untranslated region (5′ UTR) of the HAC1 mRNA represses Hac1 protein production under basal conditions, whereas cytoplasmic splicing of the intron by the Ire1 kinase-endonuclease, activated under endoplasmic reticulum stress conditions, relieves the inhibition and enables Hac1 synthesis. Using a random mutational screen as well as site-directed mutagenesis, we identify point mutations within the 5′ UTR-intron interaction site that derepress translation of the unspliced HAC1 mRNA. We also show that insertion of an in-frame AUG start codon upstream of the interaction site releases the translational block, demonstrating that an elongating ribosome can disrupt the interaction. Moreover, overexpression of translation initiation factor eIF4A, a helicase, enhances production of Hac1 from an mRNA containing an upstream AUG start codon at the beginning of the base-paired region. These results suggest that the major block of translation occurs at the initiation stage. Supporting this interpretation, the point mutations that enhanced Hac1 production resulted in an increased percentage of the HAC1 mRNA associating with polysomes versus free ribosomal subunits. Thus, our results provide evidence that the 5′ UTR-intron interaction represses translation initiation on the unspliced HAC1 mRNA. PMID:26175153

  9. miR-33a inhibits cell proliferation and invasion by targeting CAND1 in lung cancer.

    PubMed

    Kang, M; Li, Y; Zhao, Y; He, S; Shi, J

    2017-09-04

    Lung cancer continues to be one of the top five causes of cancer-related mortality. This study aims to identify down- and upregulated miRNAs and mRNA which can be used as potential biomarkers and/or therapeutic targets for lung cancer. Integrated analysis of differential expression profiles of miRNA and mRNA in lung cancer was performed by searching Gene Expression Omnibus datasets. Based on miRNA expression profiles, direct mRNA targets of miRNAs with experimental support were identified through miRTarBase. The levels of representative miRNAs and mRNAs were confirmed through qualitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR). The miR-33a was decreased in non-small cell lung cancer (NSCLC) tissues compared with the para-carcinoma tissues, whereas its target mRNA of cullin-associated NEDD8-dissociated protein 1 (CAND1) was increased in NSCLC tissues. Further research has shown that miR-33a can inhibit lung cancer cell proliferation, cell cycle progression, and migration by targeting CAND1. Moreover, the CAND1 knockout lung cancer cells showed similar results as cells transfected with miR-33a mimic. These results suggested that the data mining based on online databases was an effective method in finding novel target in cancer research, and the miR-33a and CAND1 played an important role in lung cancer proliferation and cell migration.

  10. Predators inhibit brain cell proliferation in natural populations of electric fish, Brachyhypopomus occidentalis.

    PubMed

    Dunlap, Kent D; Tran, Alex; Ragazzi, Michael A; Krahe, Rüdiger; Salazar, Vielka L

    2016-02-10

    Compared with laboratory environments, complex natural environments promote brain cell proliferation and neurogenesis. Predators are one important feature of many natural environments, but, in the laboratory, predatory stimuli tend to inhibit brain cell proliferation. Often, laboratory predatory stimuli also elevate plasma glucocorticoids, which can then reduce brain cell proliferation. However, it is unknown how natural predators affect cell proliferation or whether glucocorticoids mediate the neurogenic response to natural predators. We examined brain cell proliferation in six populations of the electric fish, Brachyhypopomus occidentalis, exposed to three forms of predator stimuli: (i) natural variation in the density of predatory catfish; (ii) tail injury, presumably from predation attempts; and (iii) the acute stress of capture. Populations with higher predation pressure had lower density of proliferating (PCNA+) cells, and fish with injured tails had lower proliferating cell density than those with intact tails. However, plasma cortisol did not vary at the population level according to predation pressure or at the individual level according to tail injury. Capture stress significantly increased cortisol, but only marginally decreased cell proliferation. Thus, it appears that the presence of natural predators inhibits brain cell proliferation, but not via mechanisms that depend on changes in basal cortisol levels. This study is the first demonstration of predator-induced alteration of brain cell proliferation in a free-living vertebrate.

  11. Interleukin 37 Expression Inhibits STAT3 to Suppress the Proliferation and Invasion of Human Cervical Cancer Cells

    PubMed Central

    Wang, Sen; An, Weifang; Yao, Yunhong; Chen, Renhuai; Zheng, Xiaoxuan; Yang, Wanyong; Zhao, Yi; Hu, Xinrong; Jiang, Enping; Bie, Yanhong; Chen, Zhangquan; Ouyang, Ping; Zhang, He; Xiong, Hui

    2015-01-01

    Objectives: The most recently discovered cytokine interleukin 37 (IL-37) received growing attention. Its function on tumor is largely unknown. Here, we investigated the biological function of IL-37 on cervical cancer (CC). Materials and methods: HPV+ Hela cells and HPV- C33A cells were used. RT-qPCR was performed to detect the transcription of IL-37, STAT3, TNF-αand IL-1β. Western blotting was used for protein detection. CCK-8 assay and transwell assay were employed for cell proliferation and invasion detection, respectively. Results: Successful gene transfection of IL-37 suppressed the proliferation and invasion of CC. Interestingly, IL-37 showed higher anticancer ability in HPV+ Hela cells than that in HPV- C33A cells. Then, the molecular mechanism of IL-37 anticancer was explored. Firstly, we found that IL-37 inhibited STAT3 expression at both mRNA and protein levels. IL-37 also down regulated the phosphorylation of STAT3. Secondly, blockage of STAT3 using siRNAs reduced significantly the ability of IL-37 to suppress cell proliferation and invasion. Thirdly, STAT3 knockdown reduced markedly the inhibition of IL-37 on the transcription of tumor-derived TNF-α and IL-1β, indicating the contribution of STAT3 for the cancer associated antiinflammation of IL-37. Finally, STAT3 up regulation restored the ability of cell proliferation, cell invasion and the expression of inflammatory cytokines, TNF-α and IL-1β. Conclusions: IL-37 suppressed cell proliferation and invasion of CC and STAT3 is involved in this process. Thus, IL-37 emerges as a new anticancer cytokine for CC. This study demonstrated a new biological function of IL-37 and offered a potential molecule for CC treatment. PMID:26316892

  12. Mifepristone acts as progesterone antagonist of non-genomic responses but inhibits phytohemagglutinin-induced proliferation in human T cells.

    PubMed

    Chien, C H; Lai, J N; Liao, C F; Wang, O Y; Lu, L M; Huang, M I; Lee, W F; Shie, M C; Chien, E J

    2009-08-01

    Progesterone is an endogenous immunomodulator that suppresses T cell activation during pregnancy. The stimulation of membrane progesterone receptors (mPRs) would seem to be the cause of rapid non-genomic responses in human peripheral T cells, such as an elevation of intracellular calcium ([Ca(2+)](i)) and decreased intracellular pH (pH(i)). Mifepristone (RU486) produces mixed agonist/antagonist effects on immune cells compared with progesterone. We explored whether RU486 is an antagonist to mPRs and can block rapid non-genomic responses and the induction by phytohemagglutinin (PHA) of cell proliferation. Human male peripheral T cell responses in terms of pH(i) and [Ca(2+)](i) changes were measured using the fluorescent dyes, 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF) and fura-2, respectively. Expression of mPR mRNA was determined by RT-PCR analysis. Cell proliferation and cell toxicity were determined by [(3)H]-thymidine incorporation and MTT assay, respectively. The mRNAs of mPRalpha, mPRbeta and mPRgamma were expressed in T cells. RU486 blocked progesterone-mediated rapid responses including, the [Ca(2+)](i) increase and pH(i) decrease, in a dose related manner. RU486 did not block, but enhanced, the inhibitory effect of progesterone on PHA induced cell proliferation. RU486 alone inhibited proliferation induced by PHA and at >25 microM seems to be cytotoxic against resting T cells (P < 0.01). RU486 is antagonistic to the rapid mPR-mediated non-genomic responses, but is synergistic with progesterone with respect to the inhibition of PHA-induced cell proliferation. Our findings shine new light on RU486's clinical application and how this relates to the non-genomic rapid physiological responses caused by progesterone.

  13. Inhibition of proliferation and induction of apoptosis in soft tissue sarcoma cells by interferon-α and retinoids

    PubMed Central

    Brodowicz, T; Wiltschke, C; Kandioler-Eckersberger, D; Grunt, T W; Rudas, M; Schneider, S M; Hejna, M; Budinsky, A; Zielinski, C C

    1999-01-01

    Uncontrolled proliferation and a defect of apoptosis constitute crucial elements in the development and progression of tumours. Among many other biological response modifiers known to influence these mechanisms, the efficacy of retinoids and interferons in the treatment of various malignant entities is currently matter of discussion. In the present study, we have investigated the effects of 9-cis-retinoic acid (9cRA), 13-cis-retinoic acid (13cRA), all-trans-retinoic acid (tRA) and interferon-α on proliferation and apoptosis of human soft tissue sarcoma (STS) cell lines HTB-82 (rhabdomyosarcoma), HTB-91 (fibrosarcoma), HTB-92 (liposarcoma), HTB-93 (synovial sarcoma) and HTB-94 (chondrosarcoma) in relation to p53 genotype as well as p53 expression. HTB-91, HTB-92 and HTB-94 STS cells exhibited mutant p53, whereas wild-type p53 was found in HTB-93 STS cells, and a normal p53 status in HTB-82 STS cells, carrying a silent point mutation only. Interferon-α, irrespective of p53 status, inhibited the proliferation of all five cell lines dose- and time-dependently. Similarly, 9cRA, 13cRA and tRA decreased the proliferation of HTB-82 and HTB-93 STS cells, whereas the proliferation of p53-mutated HTB-91, HTB-92 and HTB-94 STS cells remained unchanged. Furthermore, only 9cRA and tRA were capable of inducing apoptosis in HTB-82 and HTB-93 STS cells, whereas HTB-91, HTB-92 and HTB-94 STS cells did not undergo apoptosis under the influence of 9cRA or tRA. Retinoic acid receptor (RAR)-α and RAR-β mRNA were not detectable by Northern blot analysis in the five STS cell lines, whereas mRNA for the universal retinoic acid receptor, RAR-γ, was expressed in all STS cell lines indicating that retinoid resistance was not associated with a lack of RAR expression. Apoptosis was not induced by interferon-α or 13cRA in any of the five STS cell lines tested. Our results indicate that within the panel of tested STS cell lines, inhibition of proliferation and induction of apoptosis result

  14. Inhibition of cell proliferation by alpha-tocopherol. Role of protein kinase C.

    PubMed

    Boscoboinik, D; Szewczyk, A; Hensey, C; Azzi, A

    1991-04-05

    The effect of alpha-tocopherol (vitamin E) on the proliferation of vascular smooth muscle cells (A7r5), human osteosarcoma cells (Saos-2), fibroblasts (Balb/3T3), and neuroblastoma cells (NB2A) has been studied. The proliferation of vascular smooth muscle cells was inhibited by physiologically relevant concentrations of alpha-tocopherol, neuroblastoma cells were only sensitive to higher alpha-tocopherol concentrations, and proliferation of the other cell lines was not inhibited. The inhibition of smooth muscle cell proliferation was specific for alpha-tocopherol. Trolox, phytol, and alpha-tocopherol esters had no effect. Proliferation of smooth muscle cells stimulated by platelet-derived growth factor or endothelin was completely sensitive to alpha-tocopherol. If smooth muscle cells were stimulated by fetal calf serum, proliferation was 50% inhibited by alpha-tocopherol. No effect of alpha-tocopherol was observed when proliferation of smooth muscle cells was stimulated by bombesin and lysophosphatidic acid. The possibility of an involvement of protein kinase C in the cell response to alpha-tocopherol was suggested by experiments with the isolated enzyme and supported by the 2- to 3-fold stimulation of phorbol ester binding induced by alpha-tocopherol in sensitive cells. Moreover, alpha-tocopherol also caused inhibition of protein kinase C translocation induced by phorbol esters and inhibition of the phosphorylation of its 80-kDa protein substrate in smooth muscle cells. A model is discussed by which alpha-tocopherol inhibits cell proliferation by interacting with the cytosolic protein kinase C, thus preventing its membrane translocation and activation.

  15. Oral pirfenidone protects against fibrosis by inhibiting fibroblast proliferation and TGF-β signaling in a murine colitis model.

    PubMed

    Li, Guanwei; Ren, Jianan; Hu, Qiongyuan; Deng, Youming; Chen, Guopu; Guo, Kun; Li, Ranran; Li, Yuan; Wu, Lei; Wang, Gefei; Gu, Guosheng; Li, Jieshou

    2016-10-01

    Inflammatory bowel disease (IBD), particularly Crohn's disease, frequently causes intestinal fibrosis that ultimately leads to formation of strictures requiring bowel resection. Currently there is no effective antifibrotic therapy available for this disease. Pirfenidone is a small compound that has a broad spectrum of antifibrogenic effect and has been used for the treatment of fibrotic diseases in various organs. The present study aimed to investigate the antifibrogenic effect of pirfenidone in a dextran sulfate sodium (DSS)-induced murine colitis model. C57BL/6 mice were used and animals were randomly divided into groups receiving pirfenidone or vehicle by oral or transanal routes. Inflammation- and fibrosis-related indexes including body weight, colon length, disease activity, histological change, mRNA expression of pro-inflammatory and pro-fibrogenic cytokines were assessed. Furthermore, we performed in vitro analysis using CCD18-Co fibroblasts to evaluate cell proliferation, transdifferentiation, and viability after the cells were cultured with pirfenidone. It was found that oral administration of pirfenidone reduced deposition of collagen in colitis-associated fibrosis, and significantly suppressed the mRNA expression of col1a2, col3a1, and TGF-β. Moreover, pirfenidone inhibited the activation of TGF-β-related smad and MAPK pathways both in vitro and in vivo. Clinical and histological evaluation demonstrated that pirfenidone had no anti-inflammatory effect. The antifibrogenic effect was reduced when pirfenidone was administered in a delayed manner and was unobserved if given locally. Pirfenidone suppressed fibroblast proliferation and transdifferentiation without observed toxicity. Altogether, our results suggested that oral pirfenidone protects against fibrosis of DSS-induced colitis through inhibiting the proliferation of colonic fibroblasts and TGF-β signaling pathways.

  16. Inhibition of Hedgehog signaling pathway impedes cancer cell proliferation by promotion of autophagy.

    PubMed

    Tang, Xiaoli; Deng, Libin; Chen, Qi; Wang, Yao; Xu, Rong; Shi, Chao; Shao, Jia; Hu, Guohui; Gao, Meng; Rao, Hai; Luo, Shiwen; Lu, Quqin

    2015-05-01

    Multiple lines of evidence implicate that aberrant activation of Hedgehog (Hh) signaling is involved in a variety of human cancers. However, the molecular mechanisms underlying how cancer cells respond to Hh inhibition remain to be elucidated. In this study, we found that blockade of Hh signaling suppresses cell proliferation in human cancer cells. Microarray analysis revealed that differentially expressed genes (DEGs) in human cancer cells are enriched in autophagy pathway in response to the inhibition of Hh signaling. Interestingly, inhibition of Hh signaling induced autophagy, whereas activation of Hh signaling by ligand treatments prevented the induction of autophagy. In addition, inhibition of autophagy by 3-methyladenine (3-MA) partially suppressed cytotoxicity induced by inhibition of Hh signaling. Finally, in autophagy deficient cells, cytotoxic effect triggered by inhibition of Hh signaling was partially reversed, indicating the modulation of autophagy by Hh signaling is autophagy-specific. These results suggest that inhibition of Hh signaling impedes cancer cell proliferation in part through induction of autophagy.

  17. Activation and proliferation signals in primary human T lymphocytes inhibited by ergosterol peroxide isolated from Cordyceps cicadae

    PubMed Central

    Kuo, Y C; Weng, S C; Chou, C J; Chang, T T; Tsai, W J

    2003-01-01

    Effects of ergosterol peroxide (C28H44O3; Cpd 6A) from Cordyceps cicadae on phytohemagglutinin (PHA)-stimulated cell proliferation were studied in primary human T cells. The results showed that Cpd 6A suppressed T-cell proliferation for about 24 h after stimulation with PHA. Cell cycle analysis indicated that Cpd 6A arrested the cell cycle progression of activated T cells from the G1 transition to the S phase. To localize the point in the cell cycle where arrest occurred, a set of key regulatory events leading to the G1/S boundary, including the expression of cyclins D2, E, A1, and B1, interleukin (IL)-2, IL-4, interferon-γ (IFN-γ), and activating protein-1 (AP-1), was examined. Cpd 6A suppressed, in activated T lymphocytes, the production and mRNA expression of cyclin E, IL-2, IL-4, IL-10, and IFN-γ in a dose-dependent manner. Expression of AP-1 proteins, consisting of c-Fos and c-Jun, in activated T lymphocytes was decreased by Cpd 6A. The kinetic study indicated that the inhibitory effects of Cpd 6A on IL-2 mRNA expressed in T cells might be related to blocking c-Fos protein synthesis. T-cell proliferation after Cpd 6A treatment was partially restored by addition of IL-2, IL-4, and IFN-γ. These suppressant effects of Cpd 6A on T-cell proliferation, activated by PHA, appeared to be mediated, at least in part, through the inhibition of early gene transcripts, especially those of cyclin E, IFN-γ, IL-2, and IL-4, and by arresting cell cycle progression in the cells. PMID:14504132

  18. MiR-181b targets Six2 and inhibits the proliferation of metanephric mesenchymal cells in vitro.

    PubMed

    lyu, Zhongshi; Mao, Zhaomin; Wang, Honglian; Fang, Yin; Chen, Tielin; Wan, Qianya; Wang, Ming; Wang, Nian; Xiao, Jiangming; Wei, Hongyuan; Li, Xun; Liu, Yi; Zhou, Qin

    2013-11-01

    MicroRNAs (miRNAs) are small non-coding RNAs that down-regulate gene expression by binding to target mRNA for cleavage or translational repression, and play important regulatory roles in renal development. Despite increasing genes have been predicted to be miRNA targets by bioinformatic analysis during kidney development, few of them have been verified by experiment. The objective of our study is to identify the miRNAs targeting Six2, a critical transcription factor that maintains the mesenchymal progenitor pool via self-renewal (proliferation) during renal development. We initially analyzed the 3'UTR of Six2 and found 37 binding sites targeted by 50 putative miRNAs in the 3'UTR of Six2. Among the 50 miRNAs, miR-181b is the miRNAs predicted by the three used websites. In our study, the results of luciferase reporter assay, realtime-PCR and Western blot demonstrated that miR-181b directly targeted on the 3'UTR of Six2 and down-regulate the expression of Six2 at mRNA and protein levels. Furthermore, EdU proliferation assay along with the Six2 rescue strategy showed that miR-181b suppresses the proliferation of metanephric mesenchymal by targeting Six2 in part. In our research, we concluded that by targeting the transcription factor gene Six2, miR-181b inhibits the proliferation of metanephric mesenchymal cells in vitro and might play an important role in the formation of nephrons.

  19. Sirt1 AS lncRNA interacts with its mRNA to inhibit muscle formation by attenuating function of miR-34a

    PubMed Central

    Wang, Guo-qiang; Wang, Yu; Xiong, Yan; Chen, Xiao-Chang; Ma, Mei-ling; Cai, Rui; Gao, Yun; Sun, Yun-mei; Yang, Gong-She; Pang, Wei-Jun

    2016-01-01

    Recent studies demonstrate the functions of long non-coding RNAs (lncRNAs) in mediating gene expression at the transcriptional or translational level. Our previous study identified a Sirt1 antisense (AS) lncRNA transcribed from the Sirt1 AS strand. However, its role and regulatory mechanism is still unknown in myogenesis. Here, functional analyses showed that Sirt1 AS lncRNA overexpression promoted myoblast proliferation, but inhibited differentiation. Mechanistically, Sirt1 AS lncRNA was found to activate its sense gene, Sirt1. The luciferase assay provided evidences that Sirt1 AS lncRNA interacted with Sirt1 3′ UTR and rescued Sirt1 transcriptional suppression by competing with miR-34a. In addition, RNA stability assay showed that Sirt1 AS lncRNA prolonged Sirt1 mRNA half-life from 2 to 10 h. Ribonuclease protection assay further indicated that it fully bound to Sirt1 mRNA in the myoblast cytoplasm. Moreover, Sirt1 AS overexpression led to less mouse weight than the control because of less lean mass and greater levels of Sirt1, whereas the fat mass and levels of miR-34a were not altered. Based on the findings, a novel regulatory mechanism was found that Sirt1 AS lncRNA preferably interacted with Sirt1 mRNA forming RNA duplex to promote Sirt1 translation by competing with miR-34a, inhibiting muscle formation. PMID:26902620

  20. [Inhibition of proliferation of H5N1 subtype AIV in CEF by chemosynthetic siRNA].

    PubMed

    Li, Ru-Shu; Yu, Dan; Luo, Bao-Zheng; Bo, Qing-Ru; Xu, Hai-Nie; Sha, Cai-Hua; Liao, Xiu-Yun

    2013-06-01

    In order to study the proliferation inhibition effect of H5N1 subtype avian influenza virus (AIV) with small interfere RNA (siRNA), a total of 4 siRNAs were designed in accordance with the NP and PA genes of H5N1 subtype AIV, the siRNAs were then transfected to chicken embryo fibroblast(CEF), CEF was infected with H5N1 subtype AIV after 6 hrs. Virus titer of cell supernatant was tested at 16-56hrs post infection, and pathological changes of the cells was observed; mRNA levels of NP, PA, HA and p13-actin gene were tested at 36hrs post infection. The results showed that these 4 siRNAs could inhibit the prolif-eration of H5N1 subtype AIV in CEF in varying degrees, and one siRNA targeting PA was best per-formed. The experimental results also showed that the inhibition effect was decreased with the time prolonged. This research provides a basis for further studying RNAi on AIV prevention and control.

  1. Ethanol inhibits human bone cell proliferation and function in vitro

    SciTech Connect

    Friday, K.E.; Howard, G.A. )

    1991-06-01

    The direct effects of ethanol on human bone cell proliferation and function were studied in vitro. Normal human osteoblasts from trabecular bone chips were prepared by collagenase digestion. Exposure of these osteoblasts to ethanol in concentrations of 0.05% to 1% for 22 hours induced a dose-dependent reduction in bone cell DNA synthesis as assessed by incorporation of 3H-thymidine. After 72 hours of ethanol exposure in concentrations of 0.01% to 1%, protein synthesis as measured by 3H-proline incorporation into trichbroacetic acid (TCA)-precipitable material was reduced in a dose-dependent manner. Human bone cell protein concentrations and alkaline phosphatase total activity were significantly reduced after exposure to 1% ethanol for 72 hours, but not with lower concentrations of ethanol. This reduction in osteoblast proliferation and activity may partially explain the development of osteopenia in humans consuming excessive amounts of ethanol.

  2. HIV-1 Vif binds to APOBEC3G mRNA and inhibits its translation.

    PubMed

    Mercenne, Gaëlle; Bernacchi, Serena; Richer, Delphine; Bec, Guillaume; Henriet, Simon; Paillart, Jean-Christophe; Marquet, Roland

    2010-01-01

    The HIV-1 viral infectivity factor (Vif) allows productive infection of non-permissive cells (including most natural HIV-1 targets) by counteracting the cellular cytosine deaminases APOBEC-3G (hA3G) and hA3F. The Vif-induced degradation of these restriction factors by the proteasome has been extensively studied, but little is known about the translational repression of hA3G and hA3F by Vif, which has also been proposed to participate in Vif function. Here, we studied Vif binding to hA3G mRNA and its role in translational repression. Filter binding assays and fluorescence titration curves revealed that Vif tightly binds to hA3G mRNA. Vif overall binding affinity was higher for the 3'UTR than for the 5'UTR, even though this region contained at least one high affinity Vif binding site (apparent K(d) = 27 +/- 6 nM). Several Vif binding sites were identified in 5' and 3'UTRs using RNase footprinting. In vitro translation evidenced that Vif inhibited hA3G translation by two mechanisms: a main time-independent process requiring the 5'UTR and an additional time-dependent, UTR-independent process. Results using a Vif protein mutated in the multimerization domain suggested that the molecular mechanism of translational control is more complicated than a simple physical blockage of scanning ribosomes.

  3. HIV-1 Vif binds to APOBEC3G mRNA and inhibits its translation

    PubMed Central

    Mercenne, Gaëlle; Bernacchi, Serena; Richer, Delphine; Bec, Guillaume; Henriet, Simon; Paillart, Jean-Christophe; Marquet, Roland

    2010-01-01

    The HIV-1 viral infectivity factor (Vif) allows productive infection of non-permissive cells (including most natural HIV-1 targets) by counteracting the cellular cytosine deaminases APOBEC-3G (hA3G) and hA3F. The Vif-induced degradation of these restriction factors by the proteasome has been extensively studied, but little is known about the translational repression of hA3G and hA3F by Vif, which has also been proposed to participate in Vif function. Here, we studied Vif binding to hA3G mRNA and its role in translational repression. Filter binding assays and fluorescence titration curves revealed that Vif tightly binds to hA3G mRNA. Vif overall binding affinity was higher for the 3′UTR than for the 5′UTR, even though this region contained at least one high affinity Vif binding site (apparent Kd = 27 ± 6 nM). Several Vif binding sites were identified in 5′ and 3′UTRs using RNase footprinting. In vitro translation evidenced that Vif inhibited hA3G translation by two mechanisms: a main time-independent process requiring the 5′UTR and an additional time-dependent, UTR-independent process. Results using a Vif protein mutated in the multimerization domain suggested that the molecular mechanism of translational control is more complicated than a simple physical blockage of scanning ribosomes. PMID:19910370

  4. Anticancer agent xanthohumol inhibits IL-2 induced signaling pathways involved in T cell proliferation

    PubMed Central

    Liu, Yongbo; Gao, Xiaohua; Deeb, Dorrah; Arbab, Ali S.; Dulchavsky, Scott A.; Gautam, Subhash C.

    2013-01-01

    Xanthohumol (XN), a prenylated chalcone present in hops exhibits anti-inflammatory, antioxidant and anticancer activity. In the present study we show that XN inhibits the proliferation of mouse lymphoma cells and IL-2 induced proliferation and cell cycle progression in mouse splenic T cells. The suppression of T cell proliferation by XN was due to the inhibition of IL-2 induced Janus kinase/signal transducers and activators of transcription (Jak/STAT) and extracellular signal-regulated kinase 1 and 2 (Erk1/2) signaling pathways. XN also inhibited proliferation-related cellular proteins such as c-Myc, c-Fos and NF-κB and cyclin D1. Thus, understanding of IL-2 induced cell signaling pathways in normal T cells, which are constitutively turned on in T cell lymphomas may facilitate development of XN for the treatment of hematologic cancers. PMID:22946339

  5. Inhibition of proliferation and induction of differentiation of glioma cells with Datura stramonium agglutinin.

    PubMed

    Sasaki, T; Yamazaki, K; Yamori, T; Endo, T

    2002-10-07

    We found that a lectin, Datura stramonium agglutinin, induced irreversible differentiation in C6 glioma cells. The differentiated cells had long processes, a low rate of proliferation and a high content of glial fibrillary acidic protein. When the medium was replaced with Datura stramonium agglutinin-free medium after 1 h, cell proliferation continued to be inhibited. Experiments with several other lectins indicated that both recognition of linear N-acetyllactosamine repeats and recognition of multiantennary units of cell-surface glycans were required for the inhibition of C6 proliferation. Proliferation of four human glial tumour cells was also inhibited by Datura stramonium agglutinin. Further, these differentiated human glial tumour cells had long processes and a high content of glial fibrillary acidic protein similar to differentiated C6 glioma cells. Taken together, these observations suggest that Datura stramonium agglutinin may be useful as a new therapy for treating glioma without side effects.

  6. Oxymatrine downregulates HPV16E7 expression and inhibits cell proliferation in laryngeal squamous cell carcinoma Hep-2 cells in vitro.

    PubMed

    Ying, Xin-Jiang; Jin, Bin; Chen, Xin-Wei; Xie, Jin; Xu, Hong-Ming; Dong, Pin

    2015-01-01

    To investigate the possible mechanisms of oxymatrine's role in anti laryngeal squamous cell carcinoma. We examined the effects of oxymatrine on the proliferation, cell cycle phase distribution, apoptosis, and the protein and mRNA expression levels of HPV16E7 gene in laryngeal carcinoma Hep-2 cells in vitro. The HPV16E7 siRNA inhibition was also done to confirm the effect of downregulating HPV16E7 on the proliferation in Hep-2 cells. Oxymatrine significantly inhibited the growth and proliferation of Hep-2 cells in a dose-dependence and time-dependence manner. Oxymatrine blocked Hep-2 cells in G0/G1 phase, resulting in an obvious accumulation of G0/G1 phase cells while decreasing S phase cells. Oxymatrine induced apoptosis of Hep-2 cells, whose apoptotic rate amounted to about 42% after treatment with 7 mg/mL oxymatrine for 72 h. Oxymatrine also downregulated the expression of HPV16E7 gene, as determined by the western blotting and reverse transcription-polymerase chain reaction analysis. Knockdown of HPV16E7 effectively inhibited the proliferation of Hep-2 cells. Oxymatrine inhibits the proliferation and induces apoptosis of laryngeal carcinoma Hep-2 cells, which might be mediated by a significant cell cycle arrest in G0/G1 phase and downregulation of HPV16E7 gene. Oxymatrine is considered to be a likely preventive and curative candidate for laryngeal cancer.

  7. NS-398, a selective COX-2 inhibitor, inhibits proliferation of IL-1{beta}-stimulated vascular smooth muscle cells by induction of {eta}{omicron}-1

    SciTech Connect

    Choi, Hyoung Chul; Kim, Hee Sun; Lee, Kwang Youn; Chang, Ki Churl Kang, Young Jin

    2008-11-28

    We investigated whether NS-398, a selective inhibitor of COX-2, induces HO-1 in IL-1{beta}-stimulated vascular smooth muscle cells (VSMC). NS-398 reduced the production of PGE{sub 2} without modulation of expression of COX-2 in IL-1{beta}-stimulated VSMC. NS-398 increased HO-1 mRNA and protein in a dose-dependent manner, but inhibited proliferation of IL-1{beta}-stimulated VSMC. Furthermore, SnPPIX, a HO-1 inhibitor, reversed the effects of NS-398 on PGE{sub 2} production, suggesting that COX-2 activity can be affected by HO-1. Hemin, a HO-1 inducer, also reduced the production of PGE{sub 2} and proliferation of IL-1{beta}-stimulated VSMC. CORM-2, a CO-releasing molecule, but not bilirubin inhibited proliferation of IL-1{beta}-stimulated VSMC. NS-398 inhibited proliferation of IL-1{beta}-stimulated VSMC in a HbO{sub 2}-sensitive manner. In conclusion, NS-398 inhibits proliferation of IL-1{beta}-stimulated VSMC by HO-1-derived CO. Thus, NS-398 may facilitate the healing process of vessels in vascular inflammatory disorders such as atherosclerosis.

  8. Inositol hexaphosphate hydrolysate competitively binds to AKT to inhibit the proliferation of colon carcinoma.

    PubMed

    Chen, Chen; Yang, Fuguo; Liu, Cuiping; Cui, Lianhua; Fu, Min; Song, Yang

    2017-09-07

    Phytate, myto-inositol 1,2,3,4,5,6 hexaphosphate (IP6), is recognized as an anti-nutrition phytochemical for decades. Recently, numerous studies have indicated that IP6 and its hydrolysates could suppress colon oncogenesis. However, very little is known concerning the mechanism of IP6 hydrolysates in regulating colon oncogenesis. The aim of the present study was to identify the underlying relationship between IP6 hydrolysates and colon cancer. Three types of human colorectal cancer cells were utilized in the present study. The proliferation inhibition and migration assays were employed to reveal that IP6 hydrolysates inhibited the proliferation of SW620 cells. Real-time PCR, cell-based ELISA and the AKT inhibitor assay were utilized to reveal that 20 and 30% degree of hydrolysis hydrolysates of IP6 inhibited SW620 cell growth by inhibiting the activation of AKT protein. The docking simulation study revealed that IP4 and IP5 could inhibit the activation of AKT by binding to PIP3 receptor. Collectively, our results indicated that the IP6 hydrolysates inhibit SW620 cell proliferation; IP4 and IP5, the probable primary constituents of the 20-30% degree of hydrolysis hydrolysates of IP6, inhibited the proliferation of SW620 cells by competitively inhibiting the AKT protein.

  9. Acetylsalicylic acid inhibits cell proliferation by involving transforming growth factor-beta.

    PubMed

    Redondo, Santiago; Santos-Gallego, Carlos G; Ganado, Patricia; García, Marta; Rico, Laura; Del Rio, Marcela; Tejerina, Teresa

    2003-02-04

    Acetylsalicylic acid (ASA) inhibits cell proliferation. This may be mediated by transforming growth factor-beta (TGF-beta). TGF-beta directly stops cell proliferation, restrains cells in G(0), and inhibits the uptake of platelet-derived growth factor and insulin-like growth factor. These effects are identical to those observed with ASA treatment. We cultured rat thoracic aorta vascular smooth muscle cells and measured cytotoxicity, cell proliferation, cell cycle, transcription of TGF-beta1, and concentration of TGF-beta1 in supernatant medium. ASA dose-dependently restrained cells in G(0) phase with no cytotoxic effect and inhibited cell proliferation by 30.86%. Anti-TGF-beta1 reversed this inhibition by 30.21%. However, ASA treatment decreased TGF-beta1 transcription and had no significant effect on TGF-beta1 concentration. TGF-beta seems to play an important role in ASA-mediated inhibition of cell proliferation. Therefore, treatment with ASA prevents coronary disease not only by means of its antiplatelet properties but also by an important inhibition of plaque growth. This relationship between ASA and TGF-beta explains many other effects, such as cancer chemoprevention, immunomodulation, and wound healing. The aim of this study was to demonstrate this link.

  10. The exodus subfamily of CC chemokines inhibits the proliferation of chronic myelogenous leukemia progenitors.

    PubMed

    Hromas, R; Cripe, L; Hangoc, G; Cooper, S; Broxmeyer, H E

    2000-02-15

    Chemokines are a family of related proteins that regulate leukocyte infiltration into inflamed tissue and play important roles in disease processes. Among the biologic activities of chemokines is inhibition of proliferation of normal hematopoietic progenitors. However, chemokines that inhibit normal progenitors rarely inhibit proliferation of hematopoietic progenitors from patients with chronic myelogenous leukemia (CML). We and others recently cloned a subfamily of CC chemokines that share similar amino-terminal peptide sequences and a remarkable ability to chemoattract T cells. These chemokines, Exodus-1/LARC/MIP-3alpha, Exodus-2/SLC/6Ckine/TCA4, and Exodus-3/CKbeta11/MIP-3beta, were found to inhibit proliferation of normal human marrow progenitors. The study described here found that these chemokines also inhibited the proliferation of progenitors in every sample of marrow from patients with CML that was tested. This demonstration of consistent inhibition of CML progenitor proliferation makes the 3 Exodus chemokines unique among chemokines. (Blood. 2000;95:1506-1508)

  11. ETOH inhibits embryonic neural stem/precursor cell proliferation via PLD signaling

    SciTech Connect

    Fujita, Yuko; Hiroyama, Masami; Sanbe, Atsushi Yamauchi, Junji; Murase, Shoko; Tanoue, Akito

    2008-05-23

    While a mother's excessive alcohol consumption during pregnancy is known to have adverse effects on fetal neural development, little is known about the underlying mechanism of these effects. In order to investigate these mechanisms, we investigated the toxic effect of ethanol (ETOH) on neural stem/precursor cell (NSC) proliferation. In cultures of NSCs, phospholipase D (PLD) is activated following stimulation with epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2). Exposure of NSCs to ETOH suppresses cell proliferation, while it has no effect on cell death. Phosphatidic acid (PA), which is a signaling messenger produced by PLD, reverses ETOH inhibition of NSC proliferation. Blocking the PLD signal by 1-butanol suppresses the proliferation. ETOH-induced suppression of NSC proliferation and the protective effect of PA for ETOH-induced suppression are mediated through extracellular signal-regulated kinase signaling. These results indicate that exposure to ETOH impairs NSC proliferation by altering the PLD signaling pathway.

  12. ETOH inhibits embryonic neural stem/precursor cell proliferation via PLD signaling.

    PubMed

    Fujita, Yuko; Hiroyama, Masami; Sanbe, Atsushi; Yamauchi, Junji; Murase, Shoko; Tanoue, Akito

    2008-05-23

    While a mother's excessive alcohol consumption during pregnancy is known to have adverse effects on fetal neural development, little is known about the underlying mechanism of these effects. In order to investigate these mechanisms, we investigated the toxic effect of ethanol (ETOH) on neural stem/precursor cell (NSC) proliferation. In cultures of NSCs, phospholipase D (PLD) is activated following stimulation with epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2). Exposure of NSCs to ETOH suppresses cell proliferation, while it has no effect on cell death. Phosphatidic acid (PA), which is a signaling messenger produced by PLD, reverses ETOH inhibition of NSC proliferation. Blocking the PLD signal by 1-butanol suppresses the proliferation. ETOH-induced suppression of NSC proliferation and the protective effect of PA for ETOH-induced suppression are mediated through extracellular signal-regulated kinase signaling. These results indicate that exposure to ETOH impairs NSC proliferation by altering the PLD signaling pathway.

  13. miR-455 inhibits breast cancer cell proliferation through targeting CDK14.

    PubMed

    Wang, Bing; Zou, Aimei; Ma, Liqiang; Chen, Xiong; Wang, Lie; Zeng, Ximing; Tan, Ting

    2017-03-11

    Breast cancer is the most frequently occurring cancer in women worldwide, microRNAs (miRNAs) play critical role in the initiation and progression of breast cancer. Here, we studied the effect of miR-455 on cell proliferation of breast cancer, and found that miR-455 was downregulated in breast cancer tissues and cells. Its overexpression inhibited cell proliferation, whereas its knockdown promoted cell proliferation of breast cancer. We found a Cdc2-related protein kinase CDK14 was the target of miR-455, when the 3(')UTR of CDK14 was cloned into luciferase reporter vector and transfected into cells, miR-455 mimic could inhibit the luciferase activity in a dose-dependent manner, miR-455 inhibitor increased the luciferase activity, but the mutant miR-455 mimic couldn't change the luciferase activity, suggesting miR-455 directly bound to the 3(')UTR of CDK14. Meanwhile, we also found miR-455 inhibited Cyclin D1 expression and promoted p21 expression, confirming miR-455 inhibited cell proliferation. Double knockdown of miR-455 and CDK14 inhibited the proliferation of breast cancer cell, confirming miR-455 inhibiting cell proliferation by targeting CDK14. Moreover, miR-455 levels were negatively correlated with CDK14 levels in breast cancer tissues. Our finding revealed miR-455 inhibits breast cancer cell proliferation through targeting CDK14, it might be a target for breast cancer therapy.

  14. Involvement of the mRNA binding protein CRD-BP in the regulation of metastatic melanoma cell proliferation and invasion by hypoxia.

    PubMed

    Craig, Evisabel A; Weber, Jonathan D; Spiegelman, Vladimir S

    2012-12-15

    We have previously shown that the mRNA binding protein CRD-BP is overexpressed in human melanomas, where it promotes cell survival and resistance to chemotherapy. The present study investigates the role of hypoxia, a common characteristic of the tumor microenvironment, in the regulation of CRD-BP expression and melanoma cell responses. We found that hypoxia increases CRD-BP levels in metastatic melanoma cell lines but not in melanocytes or primary melanoma cells. Hypoxic stimulation transcriptionally regulates CRD-BP by facilitating the acetylation of histones within the CRD-BP gene and by modulating the extent of HIF1α binding to the CRD-BP promoter. Hypoxia significantly enhances the proliferative and invasive potential of metastatic melanoma cells but not that of normal or primary melanoma cells. Furthermore, inhibition of CRD-BP impairs the ability of metastatic cells to proliferate and invade in response to hypoxia. These findings identify CRD-BP as a novel effector of hypoxic responses that is relevant for the selection of metastatic cells. This work also describes a previously unknown role for CRD-BP in the regulation of melanoma cell invasion and highlights the importance of the hypoxic microenvironment in determining cell fate.

  15. [Effect of C23 down-regulation on proliferation inhibition and apoptosis induced by cisplatin in human osteosarcoma SaOS-2 cells].

    PubMed

    Wu, Bei; Wang, Wanchun; Li, Zhihong

    2012-01-01

    To investigate the effect of C23 down-regulation on proliferation inhibition and apoptosis induced by cisplatin in human osteosarcoma SaOS-2 cells. SaOS-2 cells were randomly divided in to 5 groups: a control group, a DDP group, an S+DDP group, an AS+DDP group, and an R+DDP group. Cell growth suppression of each group was analyzed by MTT assay. RT-PCR and Western blot were used to detect the expression of C23, Bcl-2 and Bax mRNA and protein in each group. Morphologic character of apoptosis was evaluated by DAPI staining. The percentage of apoptotic cells was analyzed by flowcytometer assay. C23 antisense oligonucleotides remarkably inhibited the expression of C23 mRNA and protein in SaOS-2 cells (P<0.01). The proliferation of SaOS-2 cells was remarkably inhibited in the AS+DDP group compared with the DDP group (P<0.01). The expression of C23, Bcl-2 mRNA and protein was weaker and that of Bax mRNA and protein was stronger in the AS+DDP group compared with the DDP group (P<0.01). Cisplatin-mediated apoptosis neclei in SaOS-2 cells and the rate of apoptotic cells were higher in the AS+DDP group than those of the DDP group (P<0.01). Down-regulation of C23 can contribute to cisplatin-mediated proliferation inhibition and apoptosis in SaOS-2 cells.

  16. Regulation of mRNA abundance in activated T lymphocytes: identification of mRNA species affected by the inhibition of protein synthesis.

    PubMed Central

    Coleclough, C; Kuhn, L; Lefkovits, I

    1990-01-01

    Inhibition of protein synthesis has often been observed to increase the concentration of mRNAs that encode proteins associated with the regulation of cell division. As two-dimensional gel electrophoresis permits the simultaneous monitoring of individual elements in large populations of gene products, we have used this technique to assess the effect of cycloheximide treatment on the mRNA complement of activated mouse T cells in an objective fashion. Two-dimensional gels of proteins generated by cell-free translation of mRNA from T-cell blasts display about 400 spots; only 5 of these are reproducibly enhanced by cycloheximide treatment and about 4 are diminished. The cDNA cloning vector lambda jac allows analysis of large arrays of molecular clones by cell-free expression, and we have used it in a sibling selection scheme to isolate a clone of one of the prominently induced mRNA species, which we refer to as chx1. chx1 mRNA concentration is increased by cycloheximide treatment of activated B cells, as well as T cells, and it is rapidly and transiently induced, in a cycloheximide-enhanced manner, upon serum stimulation of resting 3T3 fibroblastoid cells. The chx1 protein is hydrophilic, is slightly basic, and has patches of homology with the Jun-D gene product. The chx1 gene is remarkable in its lack of detectable introns and of strong bias against CpG dinucleotides. Images PMID:2308934

  17. C/EBPalpha arrests cell proliferation through direct inhibition of Cdk2 and Cdk4.

    PubMed

    Wang, H; Iakova, P; Wilde, M; Welm, A; Goode, T; Roesler, W J; Timchenko, N A

    2001-10-01

    The transcription factor CCAAT/enhancer binding protein alpha (C/EBPalpha) is a strong inhibitor of cell proliferation. We found that C/EBPalpha directly interacts with cdk2 and cdk4 and arrests cell proliferation by inhibiting these kinases. We mapped a short growth inhibitory region of C/EBPalpha between amino acids 175 and 187. This portion of C/EBPalpha is responsible for direct inhibition of cyclin-dependent kinases and causes growth arrest in cultured cells. C/EBPalpha inhibits cdk2 activity by blocking the association of cdk2 with cyclins. Importantly, the activities of cdk4 and cdk2 are increased in C/EBPalpha knockout livers, leading to increased proliferation. Our data demonstrate that the liver-specific transcription factor C/EBPalpha brings about growth arrest through direct inhibition of cdk2 and cdk4.

  18. Vitamin E inhibits proliferation of human Tenon's capsule fibroblasts in vitro.

    PubMed

    Haas, A L; Boscoboinik, D; Mojon, D S; Böhnke, M; Azzi, A

    1996-01-01

    Failure of glaucoma surgery is mostly due to fibrocellular scar formation, derived from Tenon's capsule fibroblasts. In high-risk cases, postoperative Tenon's capsule fibroblast proliferation is inhibited by mitomycin C or 5-fluorouracil. Toxicity to other ocular cell types and the risk of ocular hypotony limits the use of these agents. We have found that d-alpha-tocopherol (vitamin E) was able to inhibit proliferation of in vitro human Tenon's capsule fibroblasts obtained from seven different donors. At 48 h, inhibition of cell proliferation was 30-78% (mean 60%) for 50 microM d-alpha-tocopherol and 46-97% (mean 77%) for 100 microM d-alpha-tocopherol. This inhibition was statistically significant. No cytotoxic effects were observed.

  19. Inhibition of FOXO1 by small interfering RNA enhances proliferation and inhibits apoptosis of papillary thyroid carcinoma cells via Akt/FOXO1/Bim pathway

    PubMed Central

    Song, Hong-ming; Song, Jia-lu; Li, Deng-feng; Hua, Kai-yao; Zhao, Bing-kun; Fang, Lin

    2015-01-01

    Forkhead box protein O1 (FOXO1) is a multifunctional transcription factor of the forkhead family. It may function as a tumor suppressor through its ability to regulate cellular events, including cell proliferation, apoptosis, and cell cycle control. As reported, FOXO1 is downregulated in papillary thyroid carcinoma (PTC). However, the function of FOXO1 in human PTC remains unclear. In this study, we investigated the function and underlying regulatory mechanisms of FOXO1 in PTC cells. PTC cell lines K1 and TPC1 were transiently transfected with FOXO1 small interfering RNA (siRNA) and negative control RNA. Successful transfection was confirmed by RT-qPCR and Western blot analysis. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide cell proliferation assays, colony formation assays, apoptosis, and cell cycle assays were used to explore the potential function of FOXO1 in the PTC cell lines. We found that downregulation of FOXO1 promoted cellular proliferation, enhanced clonogenesis, and inhibited cellular apoptosis. However, the cell cycle was not markedly affected by FOXO1 siRNA. Furthermore, Bim, a downstream target of the Akt/FOXO1 signaling pathway, was downregulated at both mRNA and protein levels in cells transfected with FOXO1 siRNA. Collectively, these results indicate that FOXO1 may play an important role in inhibiting PTC development by regulating cellular proliferation, growth, and apoptosis. FOXO1 expression is a potentially useful biomarker for human PTC. Moreover, tumorigenesis of PTC may be associated with repression of the Akt/FOXO1/Bim signaling pathway. PMID:26664140

  20. Let-7b Regulates Myoblast Proliferation by Inhibiting IGF2BP3 Expression in Dwarf and Normal Chicken

    PubMed Central

    Lin, Shumao; Luo, Wen; Ye, Yaqiong; Bekele, Endashaw J.; Nie, Qinghua; Li, Yugu; Zhang, Xiquan

    2017-01-01

    The sex-linked dwarf chicken is caused by the mutation of growth hormone receptor (GHR) gene and characterized by shorter shanks, lower body weight, smaller muscle fiber diameter and fewer muscle fiber number. However, the precise regulatory pathways that lead to the inhibition of skeletal muscle growth in dwarf chickens still remain unclear. Here we found a let-7b mediated pathway might play important role in the regulation of dwarf chicken skeletal muscle growth. Let-7b has higher expression in the skeletal muscle of dwarf chicken than in normal chicken, and the expression of insulin-like growth factor 2 mRNA binding protein 3 (IGF2BP3), which is a translational activator of IGF2, showed opposite expression trend to let-7b. In vitro cellular assays validated that let-7b directly inhibits IGF2BP3 expression through binding to its 3′UTR region, and the protein level but not mRNA level of IGF2 would be reduced in let-7b overexpressed chicken myoblast. Let-7b can inhibit cell proliferation and induce cell cycle arrest in chicken myoblast through let-7b-IGF2BP3-IGF2 signaling pathway. Additionally, let-7b can also regulate skeletal muscle growth through let-7b-GHR-GHR downstream genes pathway, but this pathway is non-existent in dwarf chicken because of the deletion mutation of GHR 3′UTR. Notably, as the loss binding site of GHR for let-7b, let-7b has enhanced its binding and inhibition on IGF2BP3 in dwarf myoblast, suggesting that the miRNA can balance its inhibiting effect through dynamic regulate its binding to target genes. Collectively, these results not only indicate that let-7b can inhibit skeletal muscle growth through let-7b-IGF2BP3-IGF2 signaling pathway, but also show that let-7b regulates myoblast proliferation by inhibiting IGF2BP3 expression in dwarf and normal chickens. PMID:28736533

  1. Mesenchymal stem cells inhibit lymphocyte proliferation by mitogens and alloantigens by different mechanisms

    SciTech Connect

    Rasmusson, Ida . E-mail: Ida.Rasmusson@labmed.ki.se; Ringden, Olle; Sundberg, Berit; Le Blanc, Katarina

    2005-04-15

    Human mesenchymal stem cells (MSCs) have immuno-modulatory properties. They inhibit T-cell proliferation to mitogens and alloantigens in vitro and prolong skin graft survival in vivo. We found that MSCs inhibited the proliferation of peripheral blood lymphocytes (PBLs) to phorbol myristate acetate (PMA), suggesting that MSCs exert an inhibitory effect downstream of the receptor level. We analyzed cytokine profiles of PBLs co-cultured with MSCs. MSCs increased interleukin (IL)-2 and soluble IL-2 receptor in mixed lymphocyte cultures (MLCs), while IL-2 and IL-2R decreased in phytohemagglutinin (PHA)-stimulated PBL cultures. MSCs inhibited IL-2 induced proliferation, without absorbing IL-2. IL-10 levels increased in MLCs co-cultured with 10% MSCs, while the levels were not affected in PHA cultures. In MLCs inhibited by MSCs, antibodies against IL-10 further suppressed proliferation but had no effect in PHA cultures. Addition of indomethacin, an inhibitor of prostaglandin-synthesis, restored part of the inhibition by MSCs in PHA cultures. However, indomethacin did not affect MSC-induced inhibition in MLCs. To conclude, our data indicate that MSC-induced suppression is a complex mechanism affecting IL-2 and IL-10 signaling and may function differently, depending on T-cell stimuli. Prostaglandins are important in the inhibition by MSCs when the T cells were activated by PHA, but not alloantigens.

  2. A knock-in mouse model reveals roles for nuclear Apc in cell proliferation, Wnt signal inhibition and tumor suppression.

    PubMed

    Zeineldin, M; Cunningham, J; McGuinness, W; Alltizer, P; Cowley, B; Blanchat, B; Xu, W; Pinson, D; Neufeld, K L

    2012-05-10

    Mutation of the tumor suppressor adenomatous polyposis coli (APC) is considered an initiating step in the genesis of the vast majority of colorectal cancers. APC inhibits the Wnt-signaling pathway by targeting the proto-oncogene β-catenin for destruction by cytoplasmic proteasomes. In the presence of a Wnt signal, or in the absence of functional APC, β-catenin can serve as a transcription cofactor for genes required for cell proliferation such as cyclin-D1 and c-Myc. In cultured cells, APC shuttles between the nucleus and the cytoplasm, with nuclear APC implicated in the inhibition of Wnt target gene expression. Adopting a genetic approach to evaluate the functions of nuclear APC in the context of a whole organism, we generated a mouse model with mutations that inactivate the nuclear localization signals (NLSs) of Apc (Apc(mNLS)). Apc(mNLS/mNLS) mice are viable and fractionation of mouse embryonic fibroblasts (MEFs) isolated from these mice revealed a significant reduction in nuclear Apc as compared with Apc(+/+) MEFs. The levels of Apc and β-catenin protein were not significantly altered in small intestinal epithelia from Apc(mNLS/mNLS) mice. Compared with Apc(+/+) mice, Apc(mNLS/mNLS) mice showed increased proliferation in epithelial cells from the jejunum, ileum and colon. These same tissues from Apc(mNLS/mNLS) mice showed more mRNA from three genes upregulated in response to canonical Wnt signal, c-Myc, axin-2 and cyclin-D1, and less mRNA from Hath-1, which is downregulated in response to Wnt. These observations suggest a role for nuclear Apc in the inhibition of canonical Wnt signaling and the control of epithelial proliferation in intestinal tissue. Furthermore, we found Apc(Min/+) mice, which harbor a mutation that truncates Apc, to have an increased polyp size and multiplicity if they also carry the Apc(mNLS) allele. Taken together, this analysis of the novel Apc(mNLS) mouse model supports a role for nuclear Apc in the control of Wnt target genes

  3. A knock-in mouse model reveals roles for nuclear Apc in cell proliferation, Wnt signal inhibition and tumor suppression

    PubMed Central

    Zeineldin, Maged; Cunningham, Jamie; McGuinness, William; Alltizer, Preston; Cowley, Brett; Blanchat, Bryan; Xu, Wenhao; Pinson, David; Neufeld, Kristi L.

    2011-01-01

    Mutation of the tumor suppressor adenomatous polyposis coli (APC) is considered an initiating step in the genesis of the vast majority of colorectal cancers. APC inhibits the Wnt signaling pathway by targeting proto-oncogene β-catenin for destruction by cytoplasmic proteasomes. In the presence of a Wnt signal, or in the absence of functional APC, β-catenin can serve as a transcription co-factor for genes required for cell proliferation such as cyclin D1 and c-Myc. In cultured cells, APC shuttles between the nucleus and cytoplasm, with nuclear APC implicated in inhibition of Wnt target gene expression. Taking a genetic approach to evaluate the functions of nuclear APC in the context of a whole organism, we generated a mouse model with mutations that inactivate the nuclear localization signals of Apc (ApcmNLS). ApcmNLS/mNLS mice are viable and fractionation of embryonic fibroblasts (MEFs) isolated from these mice revealed a significant reduction in nuclear Apc compared to Apc+/+ MEFs. The levels of Apc and β-catenin protein were not significantly altered in small intestinal epithelia from ApcmNLS/mNLS mice. Compared to Apc+/+ mice, ApcmNLS/mNLS mice displayed increased proliferation in epithelial cells from the jejunum, ileum, and colon. These same tissues from ApcmNLS/mNLS mice displayed more mRNA from three genes up-regulated in response to canonical Wnt signal, c-Myc, Axin2, and Cyclin D1, and less mRNA from Hath 1 which is down-regulated in response to Wnt. These observations suggest a role for nuclear Apc in inhibition of canonical Wnt signaling and control of epithelial proliferation in intestinal tissue. Furthermore, we found ApcMin/+ mice, which harbor a mutation that truncates Apc, have increased polyp size and multiplicity if they also carry the ApcmNLS allele. Taken together, this analysis of the novel ApcmNLS mouse model supports a role for nuclear Apc in control of Wnt target genes, intestinal epithelial cell proliferation and polyp formation. PMID

  4. Apigenin inhibits TGF-β1-induced proliferation and migration of airway smooth muscle cells.

    PubMed

    Li, Li-Hua; Lu, Bin; Wu, Hong-Ke; Zhang, Hao; Yao, Fei-Fei

    2015-01-01

    It is well known that the proliferation and migration of ASM cells (ASMCs) plays an important role in the pathogenesis of airway remodeling in asthma. Previous studies reported that apigenin can inhibit airway remodeling in a mouse asthma model. However, its effects on the proliferation and migration of ASMCs in asthma remain unknown. Therefore, the aim of our present study was to investigate the effects of apigenin on ASMC proliferation and migration, and explore the possible molecular mechanism. We found that apigenin inhibited transforming growth factor-β1 (TGF-β1)-induced ASMC proliferation. The cell cycle was blocked at G1/S-interphase by apigenin. It also suppressed TGF-β1-induced ASMCs migration. Furthermore, apigenin inhibited TGF-β1-induced Smad 2 and Smad 3 phosphorylation in ASMCs. Taken together, these results suggested that apigenin inhibited the proliferation and migration of TGF-β1-stimulated ASMCs by inhibiting Smad signaling pathway. These data might provide useful information for treating asthma and show that apigenin has potential for attenuating airway remodeling.

  5. Apigenin inhibits TGF-β1-induced proliferation and migration of airway smooth muscle cells

    PubMed Central

    Li, Li-Hua; Lu, Bin; Wu, Hong-Ke; Zhang, Hao; Yao, Fei-Fei

    2015-01-01

    It is well known that the proliferation and migration of ASM cells (ASMCs) plays an important role in the pathogenesis of airway remodeling in asthma. Previous studies reported that apigenin can inhibit airway remodeling in a mouse asthma model. However, its effects on the proliferation and migration of ASMCs in asthma remain unknown. Therefore, the aim of our present study was to investigate the effects of apigenin on ASMC proliferation and migration, and explore the possible molecular mechanism. We found that apigenin inhibited transforming growth factor-β1 (TGF-β1)-induced ASMC proliferation. The cell cycle was blocked at G1/S-interphase by apigenin. It also suppressed TGF-β1-induced ASMCs migration. Furthermore, apigenin inhibited TGF-β1-induced Smad 2 and Smad 3 phosphorylation in ASMCs. Taken together, these results suggested that apigenin inhibited the proliferation and migration of TGF-β1-stimulated ASMCs by inhibiting Smad signaling pathway. These data might provide useful information for treating asthma and show that apigenin has potential for attenuating airway remodeling. PMID:26722444

  6. KISS-1 inhibits the proliferation and invasion of gastric carcinoma cells

    PubMed Central

    Li, Na; Wang, Hong-Xing; Zhang, Jie; Ye, Ya-Ping; He, Guo-Yang

    2012-01-01

    AIM: To investigate the function of the KISS-1 gene in gastric carcinoma cells and to explore its potential mechanism. METHODS: A KISS-1 eukaryotic expression vector was constructed and transfected into BGC-823 cells. Resistant clones were obtained through G418 selection. reverse transcription-polymerase chain reaction and western blotting were used to detect KISS-1 and matrix metalloproteinase-9 (MMP-9) expression in transfected cells. The growth of transfected cells was investigated by 3-(4, 5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) proliferation assays, and the cells’ invasive potential was analyzed by basement membrane (Matrigel) invasion assays. The anti-tumor effects of KISS-1 were tested in vivo using allografts in nude mice. RESULTS: The expression level of KISS-1 mRNA and protein in BGC-823/KISS-1 transfected cells were significantly higher than in BGC-823/pcDNA3.1 transfected cells (P < 0.05) or the parental BGC-823 cell line (P < 0.05). The expression level of MMP-9 mRNA and protein in BGC-823/KISS-1 were significantly less than in BGC-823/pcDNA3.1 (P < 0.05) or BGC-823 cells (P < 0.05). MTT growth assays show the proliferation of BGC-823/KISS-1 cells at 48 h (0.642 ± 0.130) and 72 h (0.530 ± 0.164) were significantly reduced compared to BGC-823/pcDNA3.1 (0.750 ± 0.163, 0.645 ± 0.140) (P < 0.05) and BGC-823 cells (0.782 ± 0.137, 0.685 ± 0.111) (P < 0.05). Invasion assays indicate the invasive potential of BGC-823/KISS-1 cells (16.50 ± 14.88) is significantly reduced compared to BGC-823/pcDNA3.1 (20.22 ± 14.87) (P < 0.05) and BGC-823 cells after 24 h (22.12 ± 16.12) (P < 0.05). In vivo studies demonstrate the rate of pcDNA3.1-KISS-1 tumor growth is significantly slower than pcDNA3.1 and control cell tumor growth in nude mice. Furthermore, tumor volume of pcDNA3.1-KISS-1 tumors (939.38 ± 82.08 mm3) was significantly less than pcDNA3.1 (1250.46 ± 44.36 mm3) and control tumors (1284.36 ± 55.26 mm3) (P < 0.05). Moreover, the

  7. Stiehopus japonieus acidic mucopolysaccharide inhibits the proliferation of pancreatic cancer SW1990 cells through Hippo-YAP pathway.

    PubMed

    Li, Xiaoyu; Liu, Yi; Zhang, Cuiping; Niu, Qinghui; Wang, Hui; Che, Cong; Xie, Man; Zhou, Bin; Xu, Yonghong; Zhang, Qi; Wu, Jun; Tian, Zibin

    2017-03-07

    Previous studies have indicated that stiehopus japonieus acidic mucopolysaccharide (SJAMP) could inhibit the proliferation of pancreatic cancer cell SW1990. However, the mechanism remains unclear. In our study, YAP expression was identified by immunohistochemistry and quantitative Real-time PCR from 45 pairs of human pancreatic ductal adenocarcinoma (PDAC) tissues and their adjacent non-tumor samples. We found that the YAP expression was associated with the histological differentiation degree, and negatively correlated with pancreatic cancer patients' survival. More YAP localization in nuclear and enhanced expression of YAP mRNA in pancreatic cancer tissue was found in comparison with in the normal tissue. These results identify YAP acts as an amazing regulator in the pathogenesis of pancreatic cancer. After affected by SJAMP, YAP and TEAD1 were down regulated, while MST1 and pYAP were upregulated gradually with the prolong of effect time. SJAMP also improved YAP phosphorylation, nuclear-to-cytoplasmic translocation and inactivation. After successfully knocked-down by YAP siRNA, the inhibition of proliferation of SJAMP to cancer cells was attenuated. Interestingly, we indicated a down-regulation of that TEAD with SJAMP 4 mg/ml, 8 mg/ml for 24 h and with 8 mg/ml SJAMP for 24 h, 48 h even after YAP silencing. That might mean that the SJAMP has other targets, not only YAP, to downregulate TEAD. We proposed a hypothesis that Hippo-YAP pathway involved in carcinogenesis of pancreatic cancer and in the inhibition effect of SJAMP to the proliferation of pancreatic cancer cell, although maybe not the sole signaling pathway.

  8. Involvement of basic fibroblast growth factor in suramin-induced inhibition of V79/AP4 fibroblast cell proliferation.

    PubMed Central

    Bernardini, N.; Giannessi, F.; Bianchi, F.; Dolfi, A.; Lupetti, M.; Citti, L.; Danesi, R.; Del Tacca, M.

    1993-01-01

    The V79/AP4 Chinese hamster fibroblasts were densely stained with the anti-basic fibroblast growth factor (bFGF) antibody demonstrating an endogenous production of the peptide. The in vitro proliferation of these cells was stimulated by exogenous bFGF and the maximum growth (259% increase in 3H-thymidine incorporation into DNA) was reached with bFGF 10 ng ml-1. Inhibition of bFGF-mediated mitogenic pathway was obtained with a 15-mer antisense oligodeoxynucleotide targeted against bFGF mRNA and with suramin, a drug which blocks the biological activity of heparin-binding growth factors. bFGF antisense oligomer reduced the synthesis of DNA by 79.5 and 89.5% at 20 and 60 microM, respectively; this effect was reversed by the addition of exogenous bFGF to the culture medium. A short-term exposure to suramin 300 micrograms ml-1 produced a modest reduction in 3H-thymidine incorporation but suppressed the mitogenic effect of bFGF on V79/AP4 cells. In cells treated with suramin 300 micrograms ml-1 the drug concentration increased linearly over 3 days, reaching 13.15 micrograms mg-1 of protein; cell proliferation was inhibited in a dose-related manner as evaluated by the colony formation assay (IC50: 344.22 micrograms ml-1) and by the number of mitoses observed in culture. Furthermore, the drug induced ultrastructural alterations, consisting of perinuclear cisternae swelling, chromatin condensation, nucleolar segregation and cytoplasmic vacuolations. These findings demonstrated that the endogenous production of bFGF plays an important role in V79/AP4 fibroblasts proliferation, and the inhibition of bFGF-mediated mitogenic signalling with bFGF antisense oligomer or suramin is an effective mean of reducing cell growth. Images Figure 1 Figure 5 Figure 6 PMID:7685616

  9. The effect of Rho kinase inhibition on long-term keratinocyte proliferation is rapid and conditional

    PubMed Central

    2014-01-01

    Introduction We previously demonstrated that the lifespan of primary human keratinocytes could be extended indefinitely by culture in the presence of the Rho kinase (ROCK) inhibitor Y-27632. This technique has proven to be very useful in diverse areas of basic and clinical research. Methods In this follow-up study we determine whether the continual presence of Y-27632 is required for sustained proliferation. We also test whether different ROCK inhibitors can be used for this technique and whether it can also promote indefinite proliferation of animal keratinocytes. We measure keratinocyte gene expression, proliferation, behaviour and lifespan in the presence and absence of Y-27632. Results We demonstrate that the extension of lifespan observed by culture of keratinocytes in the presence of fibroblast feeders and a ROCK inhibitor is reversible and that cells senesce gradually when the inhibitor is removed from the medium. Conversely, keratinocytes that are close to the end of their replicative life span can be revived by ROCK inhibition. We demonstrate that different inhibitors of ROCK can also efficiently extend the lifespan of human keratinocytes and that ROCK inhibition extends the lifespan of animal keratinocytes derived from mouse and bovine epithelia. Gene expression analysis of human epidermal keratinocytes cells grown in the presence of Y-27632 demonstrates that ROCK inhibition primarily inhibits keratinocyte differentiation. Live-imaging of keratinocytes cultured with ROCK inhibitors show that the effect of ROCK inhibition on cellular proliferation is immediate and ROCK inhibited cells proliferate rapidly without differentiation or stratification. Conclusions ROCK inhibition rapidly and conditionally induces indefinite proliferation of keratinocytes. This method has far-reaching applications for basic research, as well as for regenerative and personalized medicine. PMID:24774536

  10. The effect of Rho kinase inhibition on long-term keratinocyte proliferation is rapid and conditional.

    PubMed

    Chapman, Sandra; McDermott, David H; Shen, Kui; Jang, Moon Kyoo; McBride, Alison A

    2014-04-28

    We previously demonstrated that the lifespan of primary human keratinocytes could be extended indefinitely by culture in the presence of the Rho kinase (ROCK) inhibitor Y-27632. This technique has proven to be very useful in diverse areas of basic and clinical research. In this follow-up study we determine whether the continual presence of Y-27632 is required for sustained proliferation. We also test whether different ROCK inhibitors can be used for this technique and whether it can also promote indefinite proliferation of animal keratinocytes. We measure keratinocyte gene expression, proliferation, behaviour and lifespan in the presence and absence of Y-27632. We demonstrate that the extension of lifespan observed by culture of keratinocytes in the presence of fibroblast feeders and a ROCK inhibitor is reversible and that cells senesce gradually when the inhibitor is removed from the medium. Conversely, keratinocytes that are close to the end of their replicative life span can be revived by ROCK inhibition. We demonstrate that different inhibitors of ROCK can also efficiently extend the lifespan of human keratinocytes and that ROCK inhibition extends the lifespan of animal keratinocytes derived from mouse and bovine epithelia. Gene expression analysis of human epidermal keratinocytes cells grown in the presence of Y-27632 demonstrates that ROCK inhibition primarily inhibits keratinocyte differentiation. Live-imaging of keratinocytes cultured with ROCK inhibitors show that the effect of ROCK inhibition on cellular proliferation is immediate and ROCK inhibited cells proliferate rapidly without differentiation or stratification. ROCK inhibition rapidly and conditionally induces indefinite proliferation of keratinocytes. This method has far-reaching applications for basic research, as well as for regenerative and personalized medicine.

  11. PKCη promotes a proliferation to differentiation switch in keratinocytes via upregulation of p27Kip1 mRNA through suppression of JNK/c-Jun signaling under stress conditions

    PubMed Central

    Hara, T; Miyazaki, M; Hakuno, F; Takahashi, S; Chida, K

    2011-01-01

    To maintain epidermal homeostasis, the balance between keratinocyte proliferation and differentiation is tightly controlled. However, the molecular mechanisms underlying this balance remain unclear. In 3D organotypic coculture with mouse keratinocytes and fibroblasts, the thickness of stratified cell layers was prolonged, and growth arrest and terminal differentiation were delayed when PKCη-null keratinocytes were used. Re-expression of PKCη in PKCη-null keratinocytes restored stratified cell layer thickness, growth arrest and terminal differentiation. We show that in 3D cocultured PKCη-null keratinocytes, p27Kip1 mRNA was downregulated, whereas JNK/c-Jun signaling was enhanced. Furthermore, inhibition of JNK/c-Jun signaling in PKCη-null keratinocytes led to upregulation of p27Kip1 mRNA, and to thinner stratified cell layers. Collectively, our findings indicate that PKCη upregulates p27Kip1 mRNA through suppression of JNK/c-Jun signaling. This results in promoting a proliferation to differentiation switch in keratinocytes. PMID:21593789

  12. Allyl Isothiocyanate Inhibits the Proliferation of Renal Carcinoma Cell Line GRC-1 by Inducing an Imbalance Between Bcl2 and Bax

    PubMed Central

    Jiang, Zhongyong; Liu, Xi; Chang, Kai; Liu, Xia; Xiong, Jie

    2016-01-01

    Background Because of the insensitivity of renal cell carcinoma (RCC) to both chemotherapy and radiotherapy, surgery remains the primary approach for anticancer treatment. However, patients who do not receive timely diagnoses may not be suitable for surgery, especially in the late phase of tumor development. Thus, the discovery of novel effective treatment is of great importance. Allyl isothiocyanate (AITC) can inhibit the proliferation and induce apoptosis in many cancer cells. In this paper, we report on an in vitro study to determine the effect of AITC on proliferation and apoptosis of RCC line GRC-1. Material/Methods CCK8 assay was used to detect cell proliferation under gradient concentrations of AITC. Flow cytometry was employed to evaluate cell apoptosis. Real-time fluorescent polymerase chain reaction quantified mRNA levels of Bax and Bcl-2 genes. Western blotting was further employed for protein expression assay. Results AITC inhibited GRC-1 cell proliferation and induced cell apoptosis in a dose-dependent manner; it also elevated Bax while suppressing Bcl-2 gene expression at both mRNA and protein levels. In general, increasing concentration of AITC decreased Bcl-2/Bax ratio. Conclusions The inhibitory effect of AITC on GRC-1 cells is exerted via cell apoptosis, in which the imbalance of Bcl-2/Bax plays a significant role. PMID:27834342

  13. Pirfenidone inhibits proliferation, arrests the cell cycle, and downregulates heat shock protein-47 and collagen type I in rat hepatic stellate cells in vitro.

    PubMed

    Xiang, Xian-Hong; Jiang, Tian-Peng; Zhang, Shuai; Song, Jie; Li, Xing; Yang, Jian-Yong; Zhou, Shi

    2015-07-01

    Pirfenidone (esbiret) is an established anti-fibrotic and anti-inflammatory drug used to treat idiopathic pulmonary fibrosis. In the present study, the dose-dependent effects of pirfenidone on the cell cycle, proliferation and expression of heat shock protein (HSP)-47 and collagen type I in a cultured rat hepatic stellate cell line (HSC-T6) were investigated. Following pirfenidone treatment, cell proliferation was determined using the cell counting kit-8 assay and the cell cycle was measured using flow cytometry. HSP-47 expression was estimated using western blot analysis and collagen type I mRNA was assessed using reverse transcription quantitative polymerase chain reaction. Pirfenidone induced significant dose-dependent inhibition of proliferation in HSC-T6 cells. Cell viability was unaffected by treatment with pirfenidone (0, 10 or 100 µM) for 24 and 72 h. However, after 24 h, HSC-T6 cells exhibited dose-dependent decreases in HSP-47 protein and collagen I mRNA levels. In conclusion, pirfenidone inhibited HSC-T6 cell proliferation, arrested the cell cycle and reduced the expression of HSP-47 and collagen type I, indicating that pirfenidone may be a promising drug in the treatment of liver fibrosis.

  14. 3-Methylcholanthrene, which binds to the arylhydrocarbon receptor, inhibits proliferation and differentiation of osteoblasts in vitro and ossification in vivo.

    PubMed

    Naruse, Masae; Ishihara, Yoko; Miyagawa-Tomita, Sachiko; Koyama, Atsushi; Hagiwara, Hiromi

    2002-09-01

    3-Methylcholanthrene (3MC) is a ligand for arylhydrocarbon receptor (AhR), which binds dioxin. We examined the effects of 3MC on the proliferation and differentiation of osteoblasts using cultures of rat calvarial osteoblast-like cells (ROB cells) and mouse calvarial clonal preosteoblastic cells (MC3T3-E1 cells). Analysis by RT-PCR revealed that the mRNAs for AhR and AhR nuclear translocators were expressed in both ROB and MC3T3-E1 cells. Cell proliferation and the synthesis of DNA by ROB cells and MC3T3-E1 cells were markedly inhibited on exposure of cells to 3MC. Furthermore, 3MC reduced the activity of alkaline phosphatase and the rate of deposition of calcium by cells. The level of expression of mRNA for osteocalcin, which is a marker of osteoblastic differentiation, was also depressed by 3MC. Moreover, when 3MC (1 mg/kg body weight) was administered sc to pregnant mice at 10.5, 12.5, and 14.5 d post coitus, fetuses examined subsequently at 15.5 or 17.5 d post coitus revealed evidence of inhibition of appropriate calcification of bones. The treated metacarpals showed no subperiosteal bone matrix histologically. Our findings indicate that 3MC might have critical effects on the formation of bone both in vivo and in vitro.

  15. miR-22 inhibits the proliferation, motility, and invasion of human glioblastoma cells by directly targeting SIRT1.

    PubMed

    Chen, Hanchun; Lu, Qiong; Fei, Xifeng; Shen, Likui; Jiang, Dongyi; Dai, Dongwei

    2016-05-01

    Recently, microRNAs (miRNAs), a kind of small and non-coding RNA, can target the downstream molecules. Increasing evidence demonstrates that miRNAs meditate the onset and progression of a variety of tumors. In the present study, we carried out gene transfection, western blot, and reverse transcription PCR (RT-PCR) to explore the role of miR-22 in glioblastoma tissues and cell lines. Here, we verified that the expression of miR-22 was downregulated in glioblastoma tissues and cells rather than matched non-tumor tissues and normal human astrocyte (NHA) cells (p < 0.001). By contrast, SIRT1 messenger RNA (mRNA) and protein were upregulated in glioblastoma tissues and cells (p < 0.001). In vitro miR-22 mimics interfered with cell proliferation, migration, and invasion of U87 and U251 cells. Mechanically, the 3'-untranslated regions (3'-UTRs) of SIRT1 were a direct target of miR-22, leading to the decreased expression of SIRT1 protein in U87 and U251 cells. Meanwhile, miR-22 mimics also inhibited the expression of epidermal growth factor receptor (EGFR) and matrix metallopeptidase 9 (MMP9). In conclusion, miR-22 inhibited cell proliferation, migration, and invasion via targeting the 3'-UTR of SIRT1 in the progression of glioblastoma and miR-22-SIRT1 pathway can be recommended as a potential target for treatment of glioblastoma.

  16. Evodiamine inhibits PDGF-BB-induced proliferation of rat vascular smooth muscle cells through the suppression of cell cycle progression and oxidative stress

    PubMed Central

    Ge, Xie; Chen, Si-Yu; Liu, Mei; Liang, Ting-Ming; Liu, Chang

    2016-01-01

    Vascular smooth muscle cell (VSMC) proliferation is a key event in the development of in-stent restenosis. Evodiamine is an indole alkaloid extracted from the Chinese medicine, evodia, and has been shown to inhibit tumor cell proliferation and protect the cardiovascular system. However, whether evodiamine affects VSMC proliferation remains to be elucidated. Therefore, the present study examined the effects and the mechanisms of action of evodiamine on the proliferation of rat VSMCs. The cells were treated with evodiamine alone or in combination with platelet-derived growth factor-BB (PDGF-BB) stimulation. It was found that evodiamine inhibited PDGF-BB-induced VSMC proliferation in a dose-dependent manner, without inducing cell death. Evodiamine also retarded cell cycle progression, evidenced by the suppression of the expression of cell cycle-promoting cyclin proteins and cyclin-dependent kinases. In addition, evodiamine attenuated the PDGF-BB-induced phosphorylation of mitogen-activated protein kinases p38 and extracellular signal-regulated kinases 1/2, however, it had no effect on the phosphorylation of Akt. Evodiamine also inhibited the increase of reactive oxygen species generation and upregulated the mRNA expression levels of genes encoding antioxidant enzymes. These findings provide important insights into the mechanisms underlying the vasoprotective actions of evodiamine and suggest that it may be a useful therapeutic agent for the treatment of vascular occlusive disease. PMID:27748810

  17. Nifedipine inhibits advanced glycation end products (AGEs) and their receptor (RAGE) interaction-mediated proximal tubular cell injury via peroxisome proliferator-activated receptor-gamma activation

    SciTech Connect

    Matsui, Takanori; Yamagishi, Sho-ichi; Takeuchi, Masayoshi; Ueda, Seiji; Fukami, Kei; Okuda, Seiya

    2010-07-23

    Research highlights: {yields} Nifedipine inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by GW9662, an inhibitor of peroxisome proliferator-activated receptor-{gamma}. {yields} GW9662 treatment alone increased RAGE mRNA levels in tubular cells. {yields} Nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-{kappa}B activation and increases in intercellular adhesion molecule-1 and transforming growth factor-{beta} gene expression in tubular cells, all of which were blocked by GW9662. -- Abstract: There is a growing body of evidence that advanced glycation end products (AGEs) and their receptor (RAGE) interaction evokes oxidative stress generation and subsequently elicits inflammatory and fibrogenic reactions, thereby contributing to the development and progression of diabetic nephropathy. We have previously found that nifedipine, a calcium-channel blocker (CCB), inhibits the AGE-induced mesangial cell damage in vitro. However, effects of nifedipine on proximal tubular cell injury remain unknown. We examined here whether and how nifedipine blocked the AGE-induced tubular cell damage. Nifedipine, but not amlodipine, a control CCB, inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by the simultaneous treatment of GW9662, an inhibitor of peroxisome proliferator-activated receptor-{gamma} (PPAR{gamma}). GW9662 treatment alone was found to increase RAGE mRNA levels in tubular cells. Further, nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-{kappa}B activation and increases in intercellular adhesion molecule-1 and transforming growth factor-beta gene expression in tubular cells, all of which were blocked by GW9662. Our present study provides a unique beneficial aspect of nifedipine on diabetic nephropathy; it could work as an anti-oxidative and anti-inflammatory agent against AGEs in tubular cells by suppressing RAGE expression

  18. Nicotine inhibits the proliferation by upregulation of nitric oxide and increased HDAC1 in mouse neural stem cells.

    PubMed

    Lee, Hanbyeol; Park, Jeong-Ran; Yang, Jungwon; Kim, Eunjeong; Hong, Seok-Ho; Woo, Heung-Myong; Ryu, Se-Min; Cho, Sung-Joon; Park, Sung-Min; Yang, Se-Ran

    2014-09-01

    Cigarette smoking (CS) is considered one of the major risk factors to cause neurodegenerative disorders. Nicotine is the main chemical in CS which is responsible for dysfunction of the brain as a neuroteratogen. Also, nicotine dependency is a real mental illness and disease. Recently, chronic nicotine exposure has been shown to cause oxidative/nitrosative stress leading to a deleterious condition to cellular death in different brain regions. However, little is known about the effects of nicotine on mouse neural stem cells (mNSCs). The aim of this study is to investigate the effects of nicotine on mNSCs and elucidate underlying mechanisms involved in expression of a diversity of genes regulated by nicotine. When mNSCs were isolated from the whole brain of embryonic day 16 mice treated with nicotine at vehicle, 100, 400, and 800 μM for 5 d, nicotine significantly decreased the number and size of neurospheres. In immunocytochemistry, nicotine-exposed mNSCs expressing nestin showed the shortened filaments and condensed nuclei. In RT-PCR, messenger RNA (mRNA) levels of proliferating cell nuclear antigen (PCNA) and sirtuin1 (SIRT1) were significantly decreased, while the production of nitric oxide and mRNA levels of cyclooxygenase2 (COX-2), tumor necrosis factor-alpha TNF-α, and histone deacetylase 1 (HDAC1) were increased in a dose-dependent manner. In addition, sodium butyrate and valproic acid, HDAC inhibitors, partially rescue proliferation of mNSCs via inhibition of HDAC1 expression and NO production. Taken together, these data demonstrate that prolonged exposure of nicotine decreased proliferation of mNSCs by increased NO and inflammatory cytokine through increased HDAC1. Furthermore, this study could help in the development of a therapy for nicotine-induced neurodegenerative disorder and drug abuse.

  19. Purinergic signaling inhibits human acute myeloblastic leukemia cell proliferation, migration, and engraftment in immunodeficient mice.

    PubMed

    Salvestrini, Valentina; Zini, Roberta; Rossi, Lara; Gulinelli, Sara; Manfredini, Rossella; Bianchi, Elisa; Piacibello, Wanda; Caione, Luisa; Migliardi, Giorgia; Ricciardi, Maria Rosaria; Tafuri, Agostino; Romano, Marco; Salati, Simona; Di Virgilio, Francesco; Ferrari, Sergio; Baccarani, Michele; Ferrari, Davide; Lemoli, Roberto M

    2012-01-05

    Extracellular ATP and UTP nucleotides increase the proliferation and engraftment potential of normal human hematopoietic stem cells via the engagement of purinergic receptors (P2Rs). In the present study, we show that ATP and UTP have strikingly opposite effects on human acute myeloblastic leukemia (AML) cells. Leukemic cells express P2Rs. ATP-stimulated leukemic cells, but not normal CD34+ cells, undergo down-regulation of genes involved in cell proliferation and migration, whereas cell-cycle inhibitors are up-regulated. Functionally, ATP induced the inhibition of proliferation and accumulation of AML cells, but not of normal cells, in the G0 phase of the cell cycle. Exposure to ATP or UTP inhibited AML-cell migration in vitro. In vivo, xenotransplantation experiments demonstrated that the homing and engraftment capacity of AML blasts and CD34+CD38- cells to immunodeficient mice BM was significantly inhibited by pretreatment with nucleotides. P2R-expression analysis and pharmacologic profiling suggested that the inhibition of proliferation by ATP was mediated by the down-regulation of the P2X7R, which is up-regulated on untreated blasts, whereas the inhibition of chemotaxis was mainly mediated via P2Y2R and P2Y4R subtypes. We conclude that, unlike normal cells, P2R signaling inhibits leukemic cells and therefore its pharmacologic modulation may represent a novel therapeutic strategy.

  20. Advanced Glycation End Products Inhibit the Proliferation of Human Umbilical Vein Endothelial Cells by Inhibiting Cathepsin D

    PubMed Central

    Li, Yuan; Chang, Ye; Ye, Ning; Dai, Dongxue; Chen, Yintao; Zhang, Naijin; Sun, Guozhe; Sun, Yingxian

    2017-01-01

    We aimed to investigate the effect of advanced glycation end products (AGEs) on the proliferation and migration ability of human umbilical vein endothelial cells (HUVECs). Cell proliferation was detected by methyl thiazolyl tetrazolium (MTT) assay, real-time cell analyzer and 5-Ethynyl-2′-deoxyuridine (EdU) staining. Cell migration was detected by wound-healing and transwell assay. AGEs significantly inhibited the proliferation and migration of HUVECs in a time-and dose-dependent way. Western blotting revealed that AGEs dramatically increased the expression of microtubule-associated protein 1 light chain 3 (LC3) II/I and p62. Immunofluorescence of p62 and acridine orange staining revealed that AGEs significantly increased the expression of p62 and the accumulation of autophagic vacuoles, respectively. Chloroquine (CQ) could further promote the expression of LC3 II/I and p62, increase the accumulation of autophagic vacuoles and promote cell injury induced by AGEs. In addition, AGEs reduced cathepsin D (CTSD) expression in a time-dependent way. Overexpression of wild-type CTSD significantly decreased the ratio of LC 3 II/I as well as p62 accumulation induced by AGEs, but overexpression of catalytically inactive mutant CTSD had no such effects. Only overexpression of wild-type CTSD could restore the proliferation of HUVECs inhibited by AGEs. However, overexpression of both wild-type CTSD and catalytically inactive mutant CTSD could promote the migration of HUVECs inhibited by AGEs. Collectively, our study found that AGEs inhibited the proliferation and migration in HUVECs and promoted autophagic flux, which in turn played a protective role against AGEs-induced cell injury. CTSD, in need of its catalytic activity, may promote proliferation in AGEs-treated HUVECs independent of the autophagy-lysosome pathway. Meanwhile, CTSD could improve the migration of AGEs-treated HUVECs regardless of its enzymatic activity. PMID:28218663

  1. Advanced Glycation End Products Inhibit the Proliferation of Human Umbilical Vein Endothelial Cells by Inhibiting Cathepsin D.

    PubMed

    Li, Yuan; Chang, Ye; Ye, Ning; Dai, Dongxue; Chen, Yintao; Zhang, Naijin; Sun, Guozhe; Sun, Yingxian

    2017-02-17

    We aimed to investigate the effect of advanced glycation end products (AGEs) on the proliferation and migration ability of human umbilical vein endothelial cells (HUVECs). Cell proliferation was detected by methyl thiazolyl tetrazolium (MTT) assay, real-time cell analyzer and 5-Ethynyl-2'-deoxyuridine (EdU) staining. Cell migration was detected by wound-healing and transwell assay. AGEs significantly inhibited the proliferation and migration of HUVECs in a time-and dose-dependent way. Western blotting revealed that AGEs dramatically increased the expression of microtubule-associated protein 1 light chain 3 (LC3) II/I and p62. Immunofluorescence of p62 and acridine orange staining revealed that AGEs significantly increased the expression of p62 and the accumulation of autophagic vacuoles, respectively. Chloroquine (CQ) could further promote the expression of LC3 II/I and p62, increase the accumulation of autophagic vacuoles and promote cell injury induced by AGEs. In addition, AGEs reduced cathepsin D (CTSD) expression in a time-dependent way. Overexpression of wild-type CTSD significantly decreased the ratio of LC 3 II/I as well as p62 accumulation induced by AGEs, but overexpression of catalytically inactive mutant CTSD had no such effects. Only overexpression of wild-type CTSD could restore the proliferation of HUVECs inhibited by AGEs. However, overexpression of both wild-type CTSD and catalytically inactive mutant CTSD could promote the migration of HUVECs inhibited by AGEs. Collectively, our study found that AGEs inhibited the proliferation and migration in HUVECs and promoted autophagic flux, which in turn played a protective role against AGEs-induced cell injury. CTSD, in need of its catalytic activity, may promote proliferation in AGEs-treated HUVECs independent of the autophagy-lysosome pathway. Meanwhile, CTSD could improve the migration of AGEs-treated HUVECs regardless of its enzymatic activity.

  2. MicroRNA-506 inhibits esophageal cancer cell proliferation via targeting CREB1

    PubMed Central

    Yao, Wen-Jian; Wang, Yong-Lian; Lu, Jian-Guo; Guo, Ling; Qi, Bo; Chen, Zhi-Jun

    2015-01-01

    MicroRNAs (miRNAs) act as key regulators of multiple cancers. MicroRNA-506 (miR-506) functions as a tumor suppressor in various types of cancers. However, its role in esophageal cancer remains unclear. In our study, we found that miR-506 was significantly down-regulated in esophageal cancer tissues and cell lines. In vitro assay, our results showed that ectopic over-expression of miR-506 inhibited esophageal cancer cells proliferation, meanwhile, cells proliferation was promoted by miR-506 inhibition. In exploring mechanisms underlying the inhibitive role, we found that miR-506 significantly decreased the expression and transcription activity of cAMP responsive element binding protein 1 (CREB1). CREB1, tumor oncogene, exhibited significantly promote effect on esophageal cancer cell proliferation. Taken together, our data identify a new role of miR-506 in esophageal cancer involving CREB1 suppression. PMID:26617801

  3. Stochastic model of contact inhibition and the proliferation of melanoma in situ.

    PubMed

    Morais, Mauro César Cafundó; Stuhl, Izabella; Sabino, Alan U; Lautenschlager, Willian W; Queiroga, Alexandre S; Tortelli, Tharcisio Citrangulo; Chammas, Roger; Suhov, Yuri; Ramos, Alexandre F

    2017-08-14

    Contact inhibition is a central feature orchestrating cell proliferation in culture experiments; its loss is associated with malignant transformation and tumorigenesis. We performed a co-culture experiment with human metastatic melanoma cell line (SKMEL- 147) and immortalized keratinocyte cells (HaCaT). After 8 days a spatial pattern was detected, characterized by the formation of clusters of melanoma cells surrounded by keratinocytes constraining their proliferation. In addition, we observed that the proportion of melanoma cells within the total population has increased. To explain our results we propose a spatial stochastic model (following a philosophy of the Widom-Rowlinson model from Statistical Physics and Molecular Chemistry) which considers cell proliferation, death, migration, and cell-to-cell interaction through contact inhibition. Our numerical simulations demonstrate that loss of contact inhibition is a sufficient mechanism, appropriate for an explanation of the increase in the proportion of tumor cells and generation of spatial patterns established in the conducted experiments.

  4. Astaxanthin Inhibits Proliferation and Induces Apoptosis and Cell Cycle Arrest of Mice H22 Hepatoma Cells

    PubMed Central

    Shao, Yiye; Ni, Yanbo; Yang, Jing; Lin, Xutao; Li, Jun; Zhang, Lixia

    2016-01-01

    Background It is widely recognized that astaxanthin (ASX), a member of the carotenoid family, has strong biological activities including antioxidant, anti-inflammation, and immune-modulation activities. Previous studies have confirmed that ASX can effectively inhibit hepatoma cells in vitro. Material/Methods MTT was used to assay proliferation of mice H22 cells, and flow cytometry was used to determine apoptosis and cell cycle arrest of H22 cells in vitro and in vivo. Moreover, anti-tumor activity of ASX was observed in mice. Results ASX inhibited the proliferation of H22 cells, promoted cell necrosis, and induced cell cycle arrest in G2 phase in vitro and in vivo. Conclusions This study indicated that ASX can inhibit proliferation and induce apoptosis and cell cycle arrest in mice H22 hepatoma cells in vitro and in vivo. PMID:27333866

  5. Apoptosis and inhibition of proliferation of cancer cells induced by cordycepin.

    PubMed

    Tian, Xuewen; Li, Yujian; Shen, Yinyu; Li, Qiaoqiao; Wang, Qinglu; Feng, Lianshi

    2015-08-01

    Cordycepin, a 3-deoxyadenosine, is the predominant functional component of the fungus Cordyceps militaris, a traditional Chinese medicine. Previous studies investigating the inhibition of cancer cells by cordycepin identified that it not only promotes cell apoptosis, but also controls cell proliferation. Furthermore, studies have elucidated the molecular mechanisms of inhibiting cell proliferation by cordycepin binding the A3 adenosine receptor, activating G protein, inhibiting cAMP formation, decreasing glycogen synthase kinase-3β/β-catenin activation and suppressing cyclin D1 and c-myc expression. The most significant signaling pathway in which cell apoptosis is induced by cordycepin is the caspase pathway. Cordycepin induces cell apoptosis via binding the DR3 receptor and consequently activating caspase-8/-3. Taken together, these studies demonstrate that cordycepin may be used as a natural medicine, as it can not only control tumor cell proliferation, but also induce cancer cell apoptosis.

  6. RKIP suppresses the proliferation and invasion ofchoriocarcinoma cells through inhibiting the MAPK signaling pathway.

    PubMed

    Zhang, Meng; Zhang, Di; Fan, Qingxia

    2015-01-01

    Raf kinase inhibitor protein (RKIP), an inhibitor of Raf-mediated activation of mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK), has been identified as a metastasis suppressor gene. However, the role of RKIP in human choriocarcinoma remains undetermined. Therefore, in the present study, we investigated the expression of RKIP in human choriocarcinoma cells and evaluated the effects of RKIP on choriocarcinoma cell proliferation and invasion. Our results indicated that RKIP was lowly expressed in human choriocarcinoma cells. Overexpression of RKIP inhibits choriocarcinoma cell proliferation, migration and invasion, promotes the apoptosis of choriocarcinoma cells. Furthermore, overexpression of RKIP significantly inhibited the expression of MEK and ERK phosphorylation in choriocarcinoma cells. Taken together, these data suggest that RKIP suppresses cell proliferation and invasion in choriocarcinoma cells through inhibiting the MAPK signaling pathway, implying that RKIP may serve as a potential molecular target for the treatment of human choriocarcinoma.

  7. Cell proliferation depends on mitochondrial Ca2+ uptake: inhibition by salicylate

    PubMed Central

    Núñez, Lucía; Valero, Ruth A; Senovilla, Laura; Sanz-Blasco, Sara; García-Sancho, Javier; Villalobos, Carlos

    2006-01-01

    Store-operated Ca2+ entry (SOCE) is a ubiquitous Ca2+ influx pathway involved in control of multiple cellular and physiological processes including cell proliferation. Recent evidence has shown that SOCE depends critically on mitochondrial sinking of entering Ca2+ to avoid Ca2+-dependent inactivation. Thus, a role of mitochondria in control of cell proliferation could be anticipated. We show here that activation of SOCE induces cytosolic high [Ca2+] domains that are large enough to be sensed and avidly taken up by a pool of nearby mitochondria. Prevention of mitochondrial clearance of the entering Ca2+ inhibited both SOCE and cell proliferation in several cell types including Jurkat and human colon cancer cells. In addition, we find that therapeutic concentrations of salicylate, the major metabolite of aspirin, depolarize partially mitochondria and inhibit mitochondrial Ca2+ uptake, as revealed by mitochondrial Ca2+ measurements with targeted aequorins. This salicylate-induced inhibition of mitochondrial Ca2+ sinking prevented SOCE and impaired cell growth of Jurkat and human colon cancer cells. Finally, direct blockade of SOCE by the pyrazole derivative BTP-2 was sufficient to arrest cell growth. Taken together, our results reveal that cell proliferation depends critically on mitochondrial Ca2+ uptake and suggest that inhibition of tumour cell proliferation by salicylate may be due to interference with mitochondrial Ca2+ uptake, which is essential for sustaining SOCE. This novel mechanism may contribute to explaining the reported anti-proliferative and anti-tumoral actions of aspirin and dietary salicylates. PMID:16339178

  8. miR-613 inhibits proliferation and invasion of breast cancer cell via VEGFA

    SciTech Connect

    Wu, Junzhao; Yuan, Peng; Mao, Qixin; Lu, Peng; Xie, Tian; Yang, Hanzhao; Wang, Chengzheng

    2016-09-09

    MicroRNAs (miRNAs) play important roles in the pathogenesis of many types of cancers by negatively regulating gene expression at posttranscriptional level. However, the role of microRNAs in breast cancer, has remained elusive. Here, we identified that miR-613 inhibits breast cancer cell proliferation by negatively regulates its target gene VEGFA. In breast cancer cell lines, CCK-8 proliferation assay indicated that the cell proliferation was inhibited by miR-613, while miR-613 inhibitor significantly promoted the cell proliferation. Transwell assay showed that miR-613 mimics significantly inhibited the migration and invasion of breast cancer cells, whereas miR-613 inhibitors significantly increased cell migration and invasion. Luciferase assays confirmed that miR-613 directly bound to the 3′ untranslated region of VEGFA, and western blotting showed that miR-613 suppressed the expression of VEGFA at the protein levels. This study indicated that miR-613 negatively regulates VEGFA and inhibits proliferation and invasion of breast cancer cell lines. Thus, miR-613 may represent a potential therapeutic molecule for breast cancer intervention.

  9. Cyclooxygenase-2 Inhibition Enhances Proliferation of NKT Cells Derived from Patients with Laryngeal Cancer.

    PubMed

    Klatka, Janusz; Grywalska, Ewelina; Hymos, Anna; Guz, Małgorzata; Polberg, Krzysztof; Roliński, Jacek; Stepulak, Andrzej

    2017-08-01

    The aim of this study was to analyze whether inhibition of cyclooxygenase-2 by celecoxib and the subsequent enhancement in the proliferation of natural killer T (NKT) cells could play a role in dendritic cell (DC)-based laryngeal cancer (LC) immunotherapy. Peripheral blood mononuclear cells were obtained from 48 male patients diagnosed with LC and 30 control patients without cancer disease. Neoplastic cell lysate preparations were made from cancer tissues obtained after surgery and used for in vitro DCs generation. NKT cells proliferation assay was performed based on (3)H-thymidine incorporation assay. An increased proliferation of NKT cells was obtained from control patients compared to NKT cells obtained from LC patients regardless of the type of stimulation or treatment. In the patient group diagnosed with LC, COX-2 inhibition resulted in a significantly enhanced proliferation of NKT cells when stimulated with autologous DCs than NKT cells stimulated with DCs without COX-2 inhibition. These correlations were not present in the control group. Higher proliferation rate of NKT cells was also observed in non-metastatic and highly differentiated LC, which was independent of the type of stimulation or treatment. COX-2 inhibition could be regarded as immunotherapy-enhancing tool in patients with LC. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  10. An adenosine nucleoside analogue NITD008 inhibits EV71 proliferation.

    PubMed

    Shang, Luqing; Wang, Yaxin; Qing, Jie; Shu, Bo; Cao, Lin; Lou, Zhiyong; Gong, Peng; Sun, Yuna; Yin, Zheng

    2014-12-01

    Enterovirus 71 (EV71), one of the major causative agents of Hand-Foot-Mouth Disease (HFMD), causes severe pandemics and hundreds of deaths in the Asia-Pacific region annually and is an enormous public health threat. However, effective therapeutic antiviral drugs against EV71 are rare. Nucleoside analogues have been successfully used in the clinic for the treatment of various viral infections. We evaluated a total of 27 nucleoside analogues and discovered that an adenosine nucleoside analogue NITD008, which has been reported to be an antiviral reagent that specifically inhibits flaviviruses, effectively suppressed the propagation of different strains of EV71 in RD, 293T and Vero cells with a relatively high selectivity index. Triphosphorylated NITD008 (ppp-NITD008) functions as a chain terminator to directly inhibit the RNA-dependent RNA polymerase activity of EV71, and it does not affect the EV71 VPg uridylylation process. A significant synergistic anti-EV71 effect of NITD008 with rupintrivir (AG7088) (a protease inhibitor) was documented, supporting the potential combination therapy of NITD008 with other inhibitors for the treatment of EV71 infections.

  11. RNA interference-mediated silencing of eukaryotic translation initiation factor 3, subunit B (EIF3B) gene expression inhibits proliferation of colon cancer cells.

    PubMed

    Wang, Zheng; Chen, Jinxian; Sun, Jianhua; Cui, Zhe; Wu, Hui

    2012-06-26

    A key factor underlying the control of the cellular growth, size and proliferation involves the regulation of the total protein synthesis. Most often, the initial stages of mRNA translation are rate limiting, which involves a group of eukaryotic translation initiation factors (EIFs). Research advances focused on the inhibition of their expression and activity hold the key to the initiation and progression of tumor and tumor prognosis. We performed RNA interference (RNAi) with the lentivirus vector system to silence the EIF3B gene using the colon cancer cell strain SW1116. The negative control included the normal target cells infected with the negative control virus whereas the knockdown cells included the normal target cells transfected with the RNAi target virus. We tested the inhibition resulting from the decreased expression of EIF3B gene on the proliferation rate of SW1116 cells, including the cell cycle, apoptosis and clonability. Compared with the negative control, the impact of EIF3B gene expression in SW1116 cells on the levels of mRNA and protein in the knockdown group, was significantly inhibited (P < 0.01). Furthermore, the cell proliferation rate and clonability were also significantly inhibited (P < 0.01). The apoptosis rate increased significantly (P < 0.05). A significant decrease in the number of cells in the G1 phase (P < 0.01) and significant increases in S (P < 0.01) and G2 phases (P < 0.05) were observed. The silencing of EIF3B gene expression inhibits the proliferation of colon cancer cells.

  12. MOXIBUSTION ALLEVIATES GASTRIC PRECANCEROUS LESIONS IN RATS BY PROMOTING CELL APOPTOSIS AND INHIBITING PROLIFERATION-RELATED ONCOGENES

    PubMed Central

    Peng, Li; Xie, Yu-feng; Wang, Chen-guang; Wu, Huan-gan; Liu, Mi; Wang, Ya-dong; Ma, Fu-qiang; Chang, Xiao-rong; Yang, Zong-bao

    2017-01-01

    Background: It is well known that gastric mucosa dysplasia and intestinal metaplasia are gastric precancerous lesions (GPL). Moxibustion treatment of Liangmen (ST21) and Zusanli (ST36) alleviated the inflammatory response and dysplasia of gastric mucosa in our previous study. The purpose of this study was to further examine the underlying mechanism of moxibustion treatment of ST21 and ST36 on GPL. Materials and Methods: Sixty SD rats were divided into five groups and rats with GPL were treated with either moxibustion (ST), moxibustion (Sham), or vitacoenzyme. B-cell lymphoma 2 (bcl-2), tumor protein p53 (P53) and cellular Myc (C-MYC), which are related to cell apoptosis, proliferating cell nuclear antigen (PCNA), vascular endothelial growth factor (VEGF), argyrophilic nucleolar organizer region proteins (Ag-NORs), which are associated with cell proliferation, and cell signaling proteins, nuclear factor kappa B (NF-κB), epidermal growth factor receptor (EGFR) and phosphorylated extracellular signal regulated kinase (p-ERK), were measured after moxibustion treatment. Results: Compared with Control group, gastric mucosa in GPL group showed abnormal mucosal proliferation and pathological mitotic figure, the mRNA expression of bcl-2, P53 and C-MYC increased significantly (P < 0.01), the protein expression of PCNA, VEGF, Ag-NORs and the activity of NF-κβ as well as EGFR/ERK signaling proteins also increased significantly (P < 0.01). Moxibustion treatment decreased gastric mucosal proliferation and pathological mitotic figure, down-regulated the mRNA expression of bcl-2, P53, C-MYC (P < 0.01), decreased the protein expression of PCNA, VEGF, Ag-NORs and the activity of NF-κβ as well as EGFR/ERK signaling proteins significantly (P < 0.01). But moxibustion treatment of Sham didn’t show the same effect on GPL. Conclusion: Moxibustion treatment inhibited cell apoptosis and reduced gastric mucosa dysplasia by inhibiting the expression of bcl-2, P53, C-MYC and decreased

  13. Gliotoxin Inhibits Proliferation and Induces Apoptosis in Colorectal Cancer Cells

    PubMed Central

    Chen, Junxiong; Wang, Chenliang; Lan, Wenjian; Huang, Chunying; Lin, Mengmeng; Wang, Zhongyang; Liang, Wanling; Iwamoto, Aikichi; Yang, Xiangling; Liu, Huanliang

    2015-01-01

    The discovery of new bioactive compounds from marine natural sources is very important in pharmacological research. Here we developed a Wnt responsive luciferase reporter assay to screen small molecule inhibitors of cancer associated constitutive Wnt signaling pathway. We identified that gliotoxin (GTX) and some of its analogues, the secondary metabolites from marine fungus Neosartorya pseufofischeri, acted as inhibitors of the Wnt signaling pathway. In addition, we found that GTX downregulated the β-catenin levels in colorectal cancer cells with inactivating mutations of adenomatous polyposis coli (APC) or activating mutations of β-catenin. Furthermore, we demonstrated that GTX induced growth inhibition and apoptosis in multiple colorectal cancer cell lines with mutations of the Wnt signaling pathway. Together, we illustrated a practical approach to identify small-molecule inhibitors of the Wnt signaling pathway and our study indicated that GTX has therapeutic potential for the prevention or treatment of Wnt dependent cancers and other Wnt related diseases. PMID:26445050

  14. Fermented red ginseng extract inhibits cancer cell proliferation and viability.

    PubMed

    Oh, Jisun; Jeon, Seong Bin; Lee, Yuri; Lee, Hyeji; Kim, Ju; Kwon, Bo Ra; Yu, Kang-Yeol; Cha, Jeong-Dan; Hwang, Seung-Mi; Choi, Kyung-Min; Jeong, Yong-Seob

    2015-04-01

    Red ginseng (Panax ginseng C.A. Meyer) is the most widely recognized medicinal herb due to its remedial effects in various disorders, such as cancers, diabetes, and heart problems. In this study, we investigated the anticancer effect of fermented red ginseng extract (f-RGE; provided by Jeonju Biomaterials Institute, Jeonju, South Korea) in a parallel comparison with the effect of nonfermented red ginseng extract (nf-RGE; control) on several cancer cell lines--MCF-7 breast cancer cells, HepG2 hepatocellular carcinoma cells, and reprogrammed MCF-7 cells (mimicking cancer stem cells). Cells were cultured at various concentrations of RGE (from 0.5 up to 5 mg/mL) and their viabilities and proliferative properties were examined. Our data demonstrate the following: (1) nf-RGE inhibited cell viability at ≥1 mg/mL for MCF-7 cells and ≥2 mg/mL for HepG2 cells, (2) in the presence of a carcinogenic agent, 12-O-tetradecanoylphorbol-13-acetate (TPA), nf-RGE treatment in combination with paclitaxel synergistically decreased MCF-7 as well as HepG2 cell viability, (3) f-RGE (which contained a greater level of Rg3 content) more effectively decreased the viability of MCF-7 and HepG2 cells compared to nf-RGE, and (4) f-RGE appeared more potent for inhibiting cancerous differentiation of reprogrammed MCF-7 cells in a synergistic fashion with paclitaxel, especially in the presence of TPA, compared to nf-RGE. These findings suggest that f-RGE treatment may be more effective for decreasing cancer cell survival by inducing apoptotic cell death and also presumably for preventing cancer stem cell differentiation compared to nf-RGE.

  15. Equol inhibits proliferation of human gastric carcinoma cells via modulating Akt pathway

    PubMed Central

    Yang, Zhi-Ping; Zhao, Yan; Huang, Fang; Chen, Jie; Yao, Ya-Hong; Li, Jun; Wu, Xiao-Nan

    2015-01-01

    AIM: To investigate the anti-tumor effects of equol in gastric cancer cells and the underlying molecular mechanisms. METHODS: MGC-803 cells were employed for in vitro experiments in this study. Cells were treated with control (vehicle, 0.1% DMSO) or equol under specified dose titration or time courses. Cell viability was examined by MTS assay, and the levels of Ki67 were determined by qPCR and immunofluorescent assay. Changes in cell cycle distribution and apoptosis rate were detected by flow cytometry. The mRNA expression of cyclin E1 and P21WAF1 was determined by qPCR. The protein levels of cell cycle regulators, PARP and Caspase-3 cleavage, and the phosphorylation of Akt were examined by Western blot. In addition, to characterize the role of elevated Akt activation in the anti-tumor effect exerted by equol, Ly294002, a PI3K/AKT pathway inhibitor, was used to pretreat MGC-803 cells. RESULTS: Equol (5, 10, 20, 40, or 80 μmol/L) inhibited viability of MGC-803 cells in a dose- and time-dependent manner after treatment for 24, 36, or 48 h (P < 0.05 for all). Equol also decreased the mRNA (P < 0.05 for 12 and 24 h treatment) and protein levels of Ki67. Equol treatment significantly induced G0/G1 cell cycle arrest (P < 0.05), with the percentages of G0/G1 cells of 32.23% ± 3.62%, 36.31% ± 0.24%, 45.58% ± 2.29%, and 65.10% ± 2.04% for equol (0, 10, 20, or 30 μmol/L) treatment, respectively, accompanied by a significant decrease of CDK2/4 (P < 0.05 for 24 and 48 h treatment) and Cyclin D1/Cyclin E1 (P < 0.05), and an increased level of P21WAF1 (P < 0.05). A marked increase of apoptosis was observed, with the percentages of apoptotic cells of 5.01% ± 0.91%, 14.57% ± 0.99%, 37.40% ± 0.58%, and 38.46% ± 2.01% for equol (0, 5, 10, or 20 μmol/L) treatment, respectively, accompanied by increased levels of cleaved PARP and caspase-3. In addition, we found that equol treatment increased P-Akt (Ser473 and Thr308) at 12 and 24 h compared to vehicle-treated control

  16. Heparin fragments inhibit human vascular smooth muscle cell proliferation in vitro

    SciTech Connect

    Selden, S.C.; Johnson, W.V.; Maciag, T.

    1986-03-01

    The authors have examined the effect of heparin on human abdominal aortic smooth muscle cell growth. Cell proliferation was inhibited by more than 90% at a concentration of 20 ..mu..g/ml in a 12 day growth assay using heparin from Sigma, Upjohn or Calbiochem. Additionally, 200 ..mu..g/ml Upjohn heparin inhibits /sup 3/H-thymidine incorporation by 50% in short term assays using serum or purified platelet-derived growth factor (25-100ng/ml) to initiate the cell cycle. Homogeneous size classes of heparin fragments were prepared by nitrous acid cleavage and BioGel P-10 filtration chromatography. Deca-, octa-, hexa-, tetra-, and di-saccharides inhibited proliferation by 90% at concentrations of 280, 320, 260, 180 and 100 ..mu..g/ml, respectively, in a 12 day growth assay. These data confirm the work of Castellot et.al. and extend the range of inhibitory fragments down to the tetra- and di-saccharide size. These data suggest, therefore, that di-saccharide subunit of heparin is sufficient to inhibit vascular smooth muscle cell proliferation. The authors are now examining the role of the anhydromannose moiety on the reducing end of the nitrous acid generated fragments as a possible mediator of heparin-induced inhibition of vascular smooth muscle cell proliferation.

  17. BCAS2 promotes prostate cancer cells proliferation by enhancing AR mRNA transcription and protein stability

    PubMed Central

    Kuo, P-C; Huang, C-W; Lee, C-I; Chang, H-W; Hsieh, S-W; Chung, Y-P; Lee, M-S; Huang, C-S; Tsao, L-P; Tsao, Y-P; Chen, S-L

    2015-01-01

    Background: We showed previously that breast carcinoma amplified sequence 2 (BCAS2) functions as a negative regulator of p53. We also found that BCAS2 is a potential AR-associated protein. AR is essential for the growth and survival of prostate carcinoma. Therefore we characterised the correlation between BCAS2 and AR. Methods: Protein interactions were examined by GST pull-down assay and co-immunoprecipitation. Clinical prostate cancer (PCa) specimens were evaluated by immunohistochemical assay. AR transcriptional activity and LNCaP cell growth were assessed by luciferase assay and MTT assay, respectively. Results: BCAS2 expression was significantly increased in PCa. BCAS2 stabilised AR protein through both hormone-dependent and -independent manners. There are at least two mechanisms for BCAS2-mediated AR protein upregulation: One is p53-dependent. The p53 is suppressed by BCAS2 that results in increasing AR mRNA and protein expression. The other is via p53-independent inhibition of proteasome degradation. As BCAS2 can form a complex with AR and HSP90, it may function with HSP90 to stabilise AR protein from being degraded by proteasome. Conclusions: In this study, we show that BCAS2 is a novel AR-interacting protein and characterise the correlation between BCAS2 and PCa. Thus we propose that BCAS2 could be a diagnostic marker and therapeutic target for PCa. PMID:25461807

  18. Activation of PPAR-γ inhibits PDGF-induced proliferation of mouse renal fibroblasts.

    PubMed

    Lu, Jiamei; Shi, Jianhua; Gui, Baosong; Yao, Ganglian; Wang, Li; Ou, Yan; Zhu, Dan; Ma, Liqun; Ge, Heng; Fu, Rongguo

    2016-10-15

    Recent studies have shown that activation of peroxisome proliferators activated receptor-γ (PPAR-γ) ameliorates renal interstitial fibrosis (RIF) in animal model. Yet, the underlying molecular mechanisms remain still largely unknown. Here, we investigated the hypothesis that activation of PPAR-γ regulates renal remodeling by modulating proliferation of primary cultured renal fibroblasts. In our present study, platelet-derived growth factor-AA (PDGF-AA), a key isoform of PDGF superfamily as mitogen in RIF, was applied to stimulate renal fibroblasts, the selective inhibitor or sequence specific siRNA of PI3K, skp2 or PPAR-γ was used to investigate the involvement of above molecular mediators in PDGF-AA-induced cell proliferation. Our results demonstrate that PDGF-AA induced proliferation of renal fibroblasts by activating PI3K/AKT signaling and resultant skp2 production. Pre-stimulation of cells with rosiglitazone or adenovirus carrying PPAR-γ cDNA (AdPPAR-γ) blocked PDGF-AA-stimulated cell proliferation, this effect was particularly coupled to PPAR-γ inhibition of AKT phosphorylation and skp2 expression. Inhibition of PPAR-γ by GW9662 restored the suppression of activated PPAR-γ on phosphorylation of AKT and subsequent skp2 production. Our results indicate that activation of PI3K/AKT signaling and resultant skp2 generation mediated PDGF-induced proliferation of renal fibroblasts. Activation of PPAR-γ inhibited cell proliferation by inhibition of AKT phosphorylation and its down-streams.

  19. MiR-181b targets Six2 and inhibits the proliferation of metanephric mesenchymal cells in vitro

    SciTech Connect

    Lyu, Zhongshi; Mao, Zhaomin; Wang, Honglian; Fang, Yin; Chen, Tielin; Wan, Qianya; Wang, Ming; Wang, Nian; Xiao, Jiangming; Wei, Hongyuan; Li, Xun; Liu, Yi; Zhou, Qin

    2013-11-01

    Highlights: •We do bio-informatics websites to analysis of Six2 3′UTR. •MiR181b is a putative miRNA which can targets Six2 3′UTR. •MiR-181b binding site in the 3′UTR of Six2 is functional. •MiR-181b suppresses MK3 cells cell proliferation by targeting Six2. -- Abstract: MicroRNAs (miRNAs) are small non-coding RNAs that down-regulate gene expression by binding to target mRNA for cleavage or translational repression, and play important regulatory roles in renal development. Despite increasing genes have been predicted to be miRNA targets by bioinformatic analysis during kidney development, few of them have been verified by experiment. The objective of our study is to identify the miRNAs targeting Six2, a critical transcription factor that maintains the mesenchymal progenitor pool via self-renewal (proliferation) during renal development. We initially analyzed the 3′UTR of Six2 and found 37 binding sites targeted by 50 putative miRNAs in the 3′UTR of Six2. Among the 50 miRNAs, miR-181b is the miRNAs predicted by the three used websites. In our study, the results of luciferase reporter assay, realtime-PCR and Western blot demonstrated that miR-181b directly targeted on the 3′UTR of Six2 and down-regulate the expression of Six2 at mRNA and protein levels. Furthermore, EdU proliferation assay along with the Six2 rescue strategy showed that miR-181b suppresses the proliferation of metanephric mesenchymal by targeting Six2 in part. In our research, we concluded that by targeting the transcription factor gene Six2, miR-181b inhibits the proliferation of metanephric mesenchymal cells in vitro and might play an important role in the formation of nephrons.

  20. Isoliquiritigenin Inhibits Proliferation and Induces Apoptosis via Alleviating Hypoxia and Reducing Glycolysis in Mouse Melanoma B16F10 Cells.

    PubMed

    Wang, Yanming; Ma, Jun; Yan, Xinyan; Chen, Xiaoyu; Si, Lingling; Liu, Ying; Han, Jichun; Hao, Wenjin; Zheng, Qiusheng

    2016-01-01

    Isoliquiritigenin (ISL) is a licorice chalcone. According to CN104758274, CN101658513 and US009089546, it is claimed that ISL has anti-inflammatory, anti-oxidative, and anti-tumoral effects. This study aimed to investigate the potential therapeutic effect of ISL in mouse melanoma B16F10 cells. Sulforhodamine B (SRB) colorimetric assay was used to test the effects of ISL on proliferation. Commercial assay kits were applied to assess glucose uptake, lactate production and ATP levels. Measurement of apoptosis was involved with Hoechst 33258, JC-1 and annexin V-FITC/PI staining. H2DCFDA probe was employed to detect ROS generation. Quantitative RT-PCR and western blot were utilized to measure the mRNA and protein levels. ISL abated hypoxia-inducible factor 1α (HIF-1α) stability and reduced a series of glycolysis-relevant enzymes expression, including glucose transporters 1/4 (GLUT 1/4), hexokinase 2 (HK2), pyruvate kinase M2 (PKM2) and lactate dehydrogenase A (LDHA). Exposure to ISL induced the mitochondrial membrane potential depolarization and increased intracellular reactive oxygen species (ROS) level. ISL could effectively inhibit proliferation and alleviate hypoxia in mouse melanoma B16F10 cells via inducing apoptosis and reducing the expression of significant enzymes in the glycolysis. ISL significantly inhibited B16F10 cell proliferation via inducing apoptosis, and alleviated hypoxia by recovering mitochondrial function and reversing high glycolysis. Our findings propose that ISL can be a promising therapeutic agent for the melanoma via reliving hypoxia of microenvironment and targeting energy metabolism system of cancer cells. Consistent with WO2015079213 and WO2014084494, targeting glycolysis can be an effective means to anti-cancer.

  1. Statins induce immunosuppressive effect on heterotopic limb allografts in rat through inhibiting T cell activation and proliferation.

    PubMed

    Nie, Chunlei; Yang, Daping; Liu, Guofeng; Dong, Deli; Ma, Zhiqiang; Fu, Hailiang; Zhao, Zhengyu; Sun, Zhiyong

    2009-01-05

    Long-term use of immunosuppressive agents could bring many side effects. Recently, 3-Hydroxy-3-methyl-gutaryl coenzyme A reductase inhibitors (statins) have been reported to be immunomodulatory besides lowering serum cholesterol level. The aim of this study was to investigate the effects of statins on composite tissue allografts and T lymphocyte in vivo and in vitro. Rats were divided into 5 groups: syngeneic transplantation group (Lewis-Lewis); allogeneic control group (Brown Norway-Lewis, no treatment); low-dose statins group (15 mg /kg); high-dose statins group (30 mg /kg) and cyclosporin A group. In vivo, treatment of statins significantly prolonged allografts survival as compared to control group. Histological findings further supported these clinical results and demonstrated less extent of rejection. Immunohistochemical analysis showed that there was a remarkably reduced T cells infiltration in statins groups. Moreover, the serum levels of IL-2 and IFN-gamma were decreased after statins therapy, while these in control group increased significantly. Meanwhile, transcriptional activities of IL-2 and IFN-gamma were also dramatically down-regulated after statins treatment. In vitro, mixed lymphocyte reaction assay was performed and the results revealed lymphocyte proliferation was inhibited by statins in a dose-dependent manner. Furthermore, administration of statins exhibited inhibitory effects on CD3/CD28 mediated T cell activation and proliferation. Besides, the results demonstrated that statins significantly down-regulated mRNA expression and suppress cytokine production of IL-2 and IFN-gamma in vitro. In conclusion, our data demonstrated that application of statins could induce immunosuppressive effect and prolong allografts survival through inhibiting activation and proliferation of T cell and reducing production of IL-2 and IFN-gamma.

  2. Cyclin D1 represses peroxisome proliferator-activated receptor alpha and inhibits fatty acid oxidation

    PubMed Central

    Hanse, Eric A.; Mashek, Douglas G.; Mashek, Mara T.; Hendrickson, Anna M.; Mullany, Lisa K.; Albrecht, Jeffrey H.

    2016-01-01

    Cyclin D1 is a cell cycle protein that promotes proliferation by mediating progression through key checkpoints in G1 phase. It is also a proto-oncogene that is commonly overexpressed in human cancers. In addition to its canonical role in controlling cell cycle progression, cyclin D1 affects other aspects of cell physiology, in part through transcriptional regulation. In this study, we find that cyclin D1 inhibits the activity of a key metabolic transcription factor, peroxisome proliferator-activated receptor α (PPARα), a member of nuclear receptor family that induces fatty acid oxidation and may play an anti-neoplastic role. In primary hepatocytes, cyclin D1 inhibits PPARα transcriptional activity and target gene expression in a cdk4-independent manner. In liver and breast cancer cells, knockdown of cyclin D1 leads to increased PPARα transcriptional activity, expression of PPARα target genes, and fatty acid oxidation. Similarly, cyclin D1 depletion enhances binding of PPARα to target sequences by chromatin immunoprecipitation. In proliferating hepatocytes and regenerating liver in vivo, induction of endogenous cyclin D1 is associated with diminished PPARα activity. Cyclin D1 expression is both necessary and sufficient for growth factor-mediated repression of fatty acid oxidation in proliferating hepatocytes. These studies indicate that in addition to playing a pivotal role in cell cycle progression, cyclin D1 represses PPARα activity and inhibits fatty acid oxidation. Our findings establish a new link between cyclin D1 and metabolism in both tumor cells and physiologic hepatocyte proliferation. PMID:27351284

  3. Cyclin D1 represses peroxisome proliferator-activated receptor alpha and inhibits fatty acid oxidation.

    PubMed

    Kamarajugadda, Sushama; Becker, Jennifer R; Hanse, Eric A; Mashek, Douglas G; Mashek, Mara T; Hendrickson, Anna M; Mullany, Lisa K; Albrecht, Jeffrey H

    2016-07-26

    Cyclin D1 is a cell cycle protein that promotes proliferation by mediating progression through key checkpoints in G1 phase. It is also a proto-oncogene that is commonly overexpressed in human cancers. In addition to its canonical role in controlling cell cycle progression, cyclin D1 affects other aspects of cell physiology, in part through transcriptional regulation. In this study, we find that cyclin D1 inhibits the activity of a key metabolic transcription factor, peroxisome proliferator-activated receptor α (PPARα), a member of nuclear receptor family that induces fatty acid oxidation and may play an anti-neoplastic role. In primary hepatocytes, cyclin D1 inhibits PPARα transcriptional activity and target gene expression in a cdk4-independent manner. In liver and breast cancer cells, knockdown of cyclin D1 leads to increased PPARα transcriptional activity, expression of PPARα target genes, and fatty acid oxidation. Similarly, cyclin D1 depletion enhances binding of PPARα to target sequences by chromatin immunoprecipitation. In proliferating hepatocytes and regenerating liver in vivo, induction of endogenous cyclin D1 is associated with diminished PPARα activity. Cyclin D1 expression is both necessary and sufficient for growth factor-mediated repression of fatty acid oxidation in proliferating hepatocytes. These studies indicate that in addition to playing a pivotal role in cell cycle progression, cyclin D1 represses PPARα activity and inhibits fatty acid oxidation. Our findings establish a new link between cyclin D1 and metabolism in both tumor cells and physiologic hepatocyte proliferation.

  4. Inhibition of cancer cell proliferation by midazolam by targeting transient receptor potential melastatin 7.

    PubMed

    Dou, Yunling; Li, Yuan; Chen, Jingkao; Wu, Sihan; Xiao, Xiao; Xie, Shanshan; Tang, Lipeng; Yan, Min; Wang, Youqiong; Lin, Jun; Zhu, Wenbo; Yan, Guangmei

    2013-03-01

    Transient receptor potential melastatin 7 (TRPM7), a Ca(2+)-permeable channel, has been demonstrated to be present in cancer cells and involved in their growth and proliferation. The present study used midazolam, a benzodiazepine class anesthesic, to pharmacologically intervene in the expression of TRPM7 and to inhibit cancer cell proliferation. Midazolam significantly inhibited the growth and proliferation of FaDu human hypopharyngeal squamous cell carcinoma cells, concurring with the induction of G(0)/G(1) cell cycle arrest and blockage of Rb activation. Central-type and peripheral-type benzodiazepine receptor antagonists did not abrogate proliferation inhibition by midazolam, while the specific TRPM7 agonist bradykinin reversed this effect. In addition, other benzodiazepines, diazepam and clonazepam also exhibited anti-proliferative activities. The inhibitory activity on cancer cell growth and proliferation, combined with the TRPM-dependent mechanism, reveals the anticancer potential of midazolam as a TRPM7 inhibitor and supports the suggestion that TRPM7 is a valuable target for pharmaceutical intervention.

  5. Inhibition of cancer cell proliferation by midazolam by targeting transient receptor potential melastatin 7

    PubMed Central

    DOU, YUNLING; LI, YUAN; CHEN, JINGKAO; WU, SIHAN; XIAO, XIAO; XIE, SHANSHAN; TANG, LIPENG; YAN, MIN; WANG, YOUQIONG; LIN, JUN; ZHU, WENBO; YAN, GUANGMEI

    2013-01-01

    Transient receptor potential melastatin 7 (TRPM7), a Ca2+-permeable channel, has been demonstrated to be present in cancer cells and involved in their growth and proliferation. The present study used midazolam, a benzodiazepine class anesthesic, to pharmacologically intervene in the expression of TRPM7 and to inhibit cancer cell proliferation. Midazolam significantly inhibited the growth and proliferation of FaDu human hypopharyngeal squamous cell carcinoma cells, concurring with the induction of G0/G1 cell cycle arrest and blockage of Rb activation. Central-type and peripheral-type benzodiazepine receptor antagonists did not abrogate proliferation inhibition by midazolam, while the specific TRPM7 agonist bradykinin reversed this effect. In addition, other benzodiazepines, diazepam and clonazepam also exhibited anti-proliferative activities. The inhibitory activity on cancer cell growth and proliferation, combined with the TRPM-dependent mechanism, reveals the anticancer potential of midazolam as a TRPM7 inhibitor and supports the suggestion that TRPM7 is a valuable target for pharmaceutical intervention. PMID:23426784

  6. Mefloquine inhibits chondrocytic proliferation by arresting cell cycle in G2/M phase.

    PubMed

    Li, Qiong; Chen, Zeng-Gan; Xia, Qing; Lin, Jian-Ping; Yan, Zuo-Qin; Yao, Zheng-Jun; Dong, Jian

    2015-01-01

    Mefloquine (MQ), an analog of chloroquine, exhibits a promising cytotoxic activity against carcinoma cell lines and for the treatment of glioblastoma patients. The present study demonstrates the effect of mefloquine on proliferation and cell cycle in chondrocytes. MTT assay and propidium iodide staining were used for the analysis of proliferation and cell cycle distribution, respectively. Western blot analysis was used to examine the expression levels of cyclin B1/cdc2, cdc25c, p21WAF1/CIP1 and p53. The results revealed that mefloquine inhibited the proliferation of chondrocytes and caused cell cycle arrests in the G2/M phase. The proliferation of chondrocytes was reduced to 27% at 40 μM concentration of mefloquine after 48 h. The population of chondrocytes in G2/M phase was found to be 15.7 and 48.4%, respectively at 10 and 40 μM concentration of mefloquine at 48 h following treatment. The expression of the cell cycle regulatory proteins including, cyclin B1/cdc2 and cdc25c was inhibited. On the other hand, mefloquine treatment promoted the expression of p21WAF1/CIP1 and p53 at 40 μM concentration after 48 h. Therefore, mefloquine inhibits proliferation and induces cell cycle arrest in chondrocytes.

  7. Inhibition of proliferation of retinal vascular endothelial cells more effectively than choroidal vascular endothelial cell proliferation by bevacizumab

    PubMed Central

    Mynampati, Bharani Krishna; Sambhav, Kumar; Grover, Sandeep; Chalam, Kakarla V.

    2017-01-01

    AIM To evaluate the differential inhibitory effects of bevacizumab on cell proliferation of vascular endothelial growth factor (VEGF)-stimulated choroidal vascular endothelial cells (CVECs) and retinal vascular endothelial cells (RVECs) in vitro. METHODS VEGF (400 ng/mL) enriched CVECs and RVECs were treated with escalating doses of bevacizumab (0.1, 0.5, 1, 1.5 and 2 mg/mL). Cell proliferation changes were analyzed with WST-1 assay and trypan blue exclusion assay at 48, 72h and 1wk. Morphological changes were recorded with bright field microscopy. RESULTS VEGF enriched RVECs showed significantly more decline of cell viability than CVECs after bevacizumab treatment. One week after treatment, RVEC cell proliferation decreased by 29.7%, 37.5%, 52.8%, 35.9% and 45.6% at 0.1, 0.5, 1.0, 1.5 and 2 mg/mL bevacizumab respectively compared to CVEC proliferation decrease of 4.1%, 7.7%, 2.4%, 4.1% and 17.7% (P<0.05) by WST-1 assay. Trypan blue exclusion assay also revealed similar decrease in RVEC proliferation of 20%, 60%, 73.3%, 80% and 93.3% compared to CVEC proliferation decrease of 4%, 12%, 22.9%, 16.7% and 22.2% respectively (P<0.05). The maximum differential effect between the two cell types was observed at bevacizumab doses of 1.0 and 1.5 mg/mL at all time points. RVECs were 22 fold more sensitive (P<0.01) compared to CVECs (52.8% vs 2.4%) at concentration of 1.0 mg/mL, and 8.7 fold more at 1.5 mg/mL (35.9% vs 4.1%) 1wk after treatment (P<0.05 respectively). CONCLUSION VEGF-enriched RVECs are more susceptible to bevacizumab inhibition than CVECs at clinically used dosage of 1.25 mg and this differential sensitivity between two cell types should be taken into consideration in dosage selection. PMID:28149771

  8. Statins inhibited erythropoietin-induced proliferation of rat vascular smooth muscle cells.

    PubMed

    Kaneda, Tae; Tsuruoka, Shuichi; Fujimura, Akio

    2010-12-15

    Erythropoietin (EPO) directly stimulates the proliferation of vascular smooth muscle cells, and this is believed to be one of the mechanisms of vascular access failure of hemodialysis patients. However, precise mechanisms of the EPO-induced proliferation of vascular smooth muscle cells are not certain. HMG-CoA reductase inhibitors (statins) are primarily used to reduce cholesterol levels, but also exert other effects, including reno-protective effects. We evaluated the effect of several statins with various hydrophilicities on the EPO-induced proliferation of primary cultured rat vascular smooth muscle cells (VSMCs) in vitro. EPO significantly and concentration-dependently increased DNA synthesis as assessed by [³H]thymidine incorporation, cell proliferation as assessed by WST-1 assay, and activation of the p44/42MAPK pathway. Therapeutic doses of statins (pravastatin, simvastatin, atorvastatin and fluvastatin) in patients with hypercholesterolemia almost completely suppressed all of the EPO-induced effects in a concentration-dependent manner. Co-addition of mevalonic acid almost completely reversed the effects of statins. Statin alone did not affect the basal proliferation capacity of the cells. The effects were almost similar among the statins. We concluded that statins inhibited EPO-induced proliferation in rat VSMCs at least partly through their inhibition of HMG-CoA reductase activity. In the future, statins might prove useful for the treatment of EPO-induced hyperplasia of vascular access. Because the statins all showed comparable effects irrespective of their hydrophilicities, these effects might be a class effect.

  9. MicroRNA let-7a inhibits the proliferation and invasion of nonsmall cell lung cancer cell line 95D by regulating K-Ras and HMGA2 gene expression.

    PubMed

    Wang, Yan-Ying; Ren, Tao; Cai, Ying-Yun; He, Xiao-Ye

    2013-03-01

    MicroRNAs are closely linked to tumor metastasis and let-7a may play a role in inhibiting the proliferation, invasion, and metastasis of lung cancer. In vitro, we aim to observe the impact of let-7a on the proliferation and invasion of the nonsmall cell lung cancer cell line 95D by constructing a lentiviral vector that expresses let-7a. Cell proliferation assays and Transwell experiments were used to compare the proliferation and invasion of the 95D cell group with let-7a overexpressed or inhibited. Real-time polymerase chain reaction and immunoblotting analysis were used to compare the expression of K-RAS and HMGA2 at mRNA and the protein level in the above groups. The results showed the cells in the let-7a overexpressed group were significantly less proliferative and invasive than those in the let-7a inhibited group (p < 0.05). K-RAS and HMGA2 mRNA levels were significantly higher in the let-7a overexpressed group than those in the let-7a inhibited group (p < 0.05). However, the protein levels of K-RAS and HMGA2 were significantly lower in the let-7a overexpressed group than those in the let-7a inhibited group (p < 0.05). We suppose that let-7a inhibits the proliferation and invasion of the cell line 95D by regulating the translation of K-RAS and HMGA2 mRNA, not the transcription of the mRNA itself.

  10. Progranulin Inhibits Human T Lymphocyte Proliferation by Inducing the Formation of Regulatory T Lymphocytes

    PubMed Central

    Kwack, Kyu Hwan

    2017-01-01

    We have examined the effect of progranulin (PGRN) on human T cell proliferation and its underlying mechanism. We show that PGRN inhibits the PHA-induced multiplication of T lymphocytes. It increases the number of iTregs when T lymphocytes are activated by PHA but does not do so in the absence of PHA. PGRN-mediated inhibition of T lymphocyte proliferation, as well as the induction of iTregs, was completely reversed by a TGF-β inhibitor or a Treg inhibitor. PGRN induced TGF-β secretion in the presence of PHA whereas it did not in the absence of PHA. Our findings indicate that PGRN suppresses T lymphocyte proliferation by enhancing the formation of iTregs from activated T lymphocytes in response to TGF-β. PMID:28194047

  11. Corosolic acid inhibits the proliferation of glomerular mesangial cells and protects against diabetic renal damage

    PubMed Central

    Li, Xiao-Qiang; Tian, Wen; Liu, Xiao-Xiao; Zhang, Kai; Huo, Jun-Cheng; Liu, Wen-Juan; Li, Ping; Xiao, Xiong; Zhao, Ming-Gao; Cao, Wei

    2016-01-01

    Diabetic nephropathy (DN) is one of the major complications of diabetes mellitus (DM). This study aimed to explore the effects of corosolic acid (CA) on the renal damage of DM and the mechanisms behind these effects. The renoprotective effect of CA was investigated in type 1 diabetic rats and db/db mice. The kidneys and glomerular mesangial cells (GMCs) were used to study the proliferation of GMCs by immunostaining and MTT assay. Further immunoblotting, siRNA, qPCR analysis, and detecting of NADPH oxidase activity and reactive oxygen species (ROS) generation were performed to explore relevant molecular mechanisms. In CA-treated diabetic animals, diabetes-induced albuminuria, increased serum creatinine and blood urea nitrogen were significantly attenuated, and glomerular hypertrophy, mesangial expansion and fibrosis were ameliorated. Furthermore, CA significantly inhibited proliferation of GMCs and phosphorylation of ERK1/2 and p38 MAPK in both diabetic animals and high glucose (HG)-induced GMCs. CA also normalized Δψm and inhibited HG-induced NADPH oxidase activity, ROS generation and NOX4, NOX2, p22phox and p47phox expression. More importantly, CA inhibited GMC proliferation mediated by NADPH/ERK1/2 and p38 MAPK signaling pathways. These findings suggest that CA exert the protective effect on DN by anti-proliferation resulted from inhibition of p38 MAPK- and NADPH-mediated inactivation of ERK1/2. PMID:27229751

  12. Silibinin inhibits HIV-1 infection by reducing cellular activation and proliferation.

    PubMed

    McClure, Janela; Lovelace, Erica S; Elahi, Shokrollah; Maurice, Nicholas J; Wagoner, Jessica; Dragavon, Joan; Mittler, John E; Kraft, Zane; Stamatatos, Leonidas; Stamatatos, Leonidis; Horton, Helen; De Rosa, Stephen C; Coombs, Robert W; Polyak, Stephen J

    2012-01-01

    Purified silymarin-derived natural products from the milk thistle plant (Silybum marianum) block hepatitis C virus (HCV) infection and inhibit T cell proliferation in vitro. An intravenous formulation of silibinin (SIL), a major component of silymarin, displays anti-HCV effects in humans and also inhibits T-cell proliferation in vitro. We show that SIL inhibited replication of HIV-1 in TZM-bl cells, PBMCs, and CEM cells in vitro. SIL suppression of HIV-1 coincided with dose-dependent reductions in actively proliferating CD19+, CD4+, and CD8+ cells, resulting in fewer CD4+ T cells expressing the HIV-1 co-receptors CXCR4 and CCR5. SIL inhibition of T-cell growth was not due to cytotoxicity measured by cell cycle arrest, apoptosis, or necrosis. SIL also blocked induction of the activation markers CD38, HLA-DR, Ki67, and CCR5 on CD4+ T cells. The data suggest that SIL attenuated cellular functions involved in T-cell activation, proliferation, and HIV-1 infection. Silymarin-derived compounds provide cytoprotection by suppressing virus infection, immune activation, and inflammation, and as such may be relevant for both HIV mono-infected and HIV/HCV co-infected subjects.

  13. Lipocalin-2 inhibits osteoclast formation by suppressing the proliferation and differentiation of osteoclast lineage cells

    SciTech Connect

    Kim, Hyun-Ju; Yoon, Hye-Jin; Yoon, Kyung-Ae; Gwon, Mi-Ri; Jin Seong, Sook; Suk, Kyoungho; Kim, Shin-Yoon; Yoon, Young-Ran

    2015-06-10

    Lipocalin-2 (LCN2) is a member of the lipocalin superfamily and plays a critical role in the regulation of various physiological processes, such as inflammation and obesity. In this study, we report that LCN2 negatively modulates the proliferation and differentiation of osteoclast precursors, resulting in impaired osteoclast formation. The overexpression of LCN2 in bone marrow-derived macrophages or the addition of recombinant LCN2 protein inhibits the formation of multinuclear osteoclasts. LCN2 suppresses macrophage colony-stimulating factor (M-CSF)-induced proliferation of osteoclast precursor cells without affecting their apoptotic cell death. Interestingly, LCN2 decreases the expression of the M-CSF receptor, c-Fms, and subsequently blocks its downstream signaling cascades. In addition, LCN2 inhibits RANKL-induced osteoclast differentiation and attenuates the expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which are important modulators in osteoclastogenesis. Mechanistically, LCN2 inhibits NF-κB signaling pathways, as demonstrated by the suppression of IκBα phosphorylation, nuclear translocation of p65, and NF-κB transcriptional activity. Thus, LCN2 is an anti-osteoclastogenic molecule that exerts its effects by retarding the proliferation and differentiation of osteoclast lineage cells. - Highlights: • LCN2 expression is regulated during osteoclast development. • LCN2 suppresses M-CSF-mediated osteoclast precursor proliferation. • LCN2 inhibits RANKL-induced osteoclast differentiation.

  14. Inhibition of colonic cancer cell proliferation and COX2 by oats avenanthramides (Avns)

    USDA-ARS?s Scientific Manuscript database

    High intake of whole grain foods is associated with reduced risk of colon cancer, but the mechanism underlying this protection has yet to be elucidated. Avns are polyphenols unique to oats. We have reported that Avns inhibited pro-inflammatory cytokines and vascular smooth muscle cell proliferation....

  15. White button mushroom phytochemicals inhibit aromatase activity and breast cancer cell proliferation.

    PubMed

    Grube, B J; Eng, E T; Kao, Y C; Kwon, A; Chen, S

    2001-12-01

    Estrogen is a major factor in the development of breast cancer. In situ estrogen production by aromatase/estrogen synthetase in breast cancer plays a dominant role in tumor proliferation. Because natural compounds such as flavones and isoflavones have been shown to be inhibitors of aromatase, it is thought that vegetables that contain these phytochemicals can inhibit aromatase activity and suppress breast cancer cell proliferation. Heat-stable extracts were prepared from vegetables and screened for their ability to inhibit aromatase activity in a human placental microsome assay. The white button mushroom (species Agaricus bisporus) suppressed aromatase activity dose dependently. Enzyme kinetics demonstrated mixed inhibition, suggesting the presence of multiple inhibitors or more than one inhibitory mechanism. "In cell" aromatase activity and cell proliferation were measured using MCF-7aro, an aromatase-transfected breast cancer cell line. Phytochemicals in the mushroom aqueous extract inhibited aromatase activity and proliferation of MCF-7aro cells. These results suggest that diets high in mushrooms may modulate the aromatase activity and function in chemoprevention in postmenopausal women by reducing the in situ production of estrogen.

  16. DICHLOROACETIC ACID (DCA) INHIBITS PROLIFERATION AND APOPTOSIS IN NORMAL HEPATOCYTES OF MALE F344 RATS

    EPA Science Inventory

    Dichloroacetic acid (DCA} inhibits proliferation and apoptosis in nonnal hepatocytes of
    male F344 rats.

    Large segments of the population are chronically exposed to dichloroacetic acid (DCA}: DCA is a by product of the chlorine disinfection of drinking water, a metab...

  17. [IL-37 inhibits the proliferation, invasion and migration of SMMC-7721 cells in vitro].

    PubMed

    Zhang, Guoqing; Tang, Chengyoug; Tan, Jingyu; Liu, Rui; Zhou, Mi; Wu, Zhongjun

    2015-10-01

    To analyze the impact of interleukin-37 (IL-37) on proliferation, migration and invasion of SMMC-7721 hepatocellular carcinoma cells and the related molecular mechanism. SMMC-7721 cells cultured in vitro were divided into the (50, 100, 200, 500) ng/mL recombinant human IL-37 (rhIL-37) treated groups and control group. Cell proliferation ability was detected by CCK-8 assay. Wound healing assay was used to observe cell migration ability. Transwell(TM) assay was performed to test the cell invasion ability. The levels of signal transducers and activators of transcription 3 (STAT3), phospho-STAT3 (p-STAT3) and matrix metalloproteinase-2 (MMP-2) were determined using Western blotting. CCK-8 assay showed that rhIL-37 inhibited the proliferation of SMMC-7721 cells. Wound healing assay revealed that rhIL-37 inhibited the migration of SMMC-7721 cells. Transwell(TM) indicated that rhIL-37 inhibited the invasion of SMMC-7721 cells. Western blotting demonstrated that rhIL-37 down-regulated the expressions of p-STAT3 and MMP-2. IL-37 can inhibit the proliferation, migration and invasion of SMMC-7721 cells. It may be related with the down-regulation of p-STAT3 and MMP-2.

  18. MicroRNA‑320a inhibition decreases insulin‑induced KGN cell proliferation and apoptosis by targeting PCGF1.

    PubMed

    Yang, Lei; Li, Yan; Wang, Xiaojing; Liu, Yuling; Yang, Lingzhu

    2017-10-01

    MicroRNAs (miRNAs) are widely involved in regulation of cellular processes of polycystic ovary syndrome (PCOS). However, the function of miR‑320a in PCOS remains unclear. The present study aimed to explore the effect of miR‑320a on PCOS cell proliferation and apoptosis following treatment with insulin, and to clarify the underlying mechanism. PCOS tissues and corresponding normal tissues were collected from 16 female patients with PCOS. KGN cells were pre‑treated with insulin, and KGN cells were transfected with ASO‑miR‑320a, miR‑320a mimics and polycomb group ring finger 1 (PCGF1) overexpression plasmids. Expressions of miR‑320a and PCGF1 were detected using the reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR). Dual‑Luciferase reporter assays were performed to investigate the target gene of miR‑320a. MTS, colony formation and flow cytometry assays were performed to determine cell viability, colony formation, and apoptosis, respectively. Furthermore, mRNA and protein expression levels of B‑cell lymphoma 2 apoptosis regulator (Bcl‑2) and Bcl‑2 associated protein X apoptosis regulator (Bax) were examined using RT‑qPCR and western blotting. The results demonstrated that miR‑320a expression was significantly increased in PCOS tissues compared with normal tissues. Moreover, miR‑320a was upregulated in insulin‑induced cells in a dose‑dependent manner. Inhibition of miR‑320a suppressed insulin‑induced cell viability and colony formation, and promoted apoptosis. Luciferase reporter assays demonstrated that PCGF1 was a target of miR‑320a. Additionally, PCGF1 overexpression inhibited cell viability and colony formation and promoted apoptosis. Additionally, the mRNA and protein levels of Bcl‑2 were inhibited by miR‑320a suppression and PCGF1 overexpression, while Bax expression was promoted by them in insulin‑induced cells. The results of the present study demonstrated that miR‑320a inhibition decreased

  19. 5-aminosalicylic acid in combination with nimesulide inhibits proliferation of colon carcinoma cells in vitro

    PubMed Central

    Fang, Hai-Ming; Mei, Qiao; Xu, Jian-Ming; Ma, Wei-Juan

    2007-01-01

    AIM: To investigate the effects of 5-aminosalicylic acid (5-ASA) in combination with nimesulide on the proliferation of HT-29 colon carcinoma cells and its potential mechanisms. METHODS: Inhibitory effects of drugs (5-ASA, nimesulide and their combination) on HT-29 colon carcinoma cells were investigated by thiazolyl blue tetrazolium bromide (MTT) assay. Cellular apoptosis and proliferation were detected by TUNEL assay and immunocytochemical staining, respectively. RESULTS: Pretreatment with 5-ASA or nimesulide at the concentration of 10-1000 μmol/L inhibited proliferation of HT-29 colon carcinoma cells in a dose-dependent manner in vitro (t = 5.122, P < 0.05; t = 3.086, P < 0.05, respectively). The inhibition rate of HT-29 colon carcinoma cell proliferation was also increased when pretreated with 5-ASA (100 μmol/L) or nimesulide (100 μmol/L) for 12-96 h, which showed an obvious time-effect relationship (t = 6.149, P < 0.05; t = 4.159, P < 0.05, respectively). At the concentration of 10-500 μmol/L, the apoptotic rate of HT-29 colon carcinoma cells significantly increased (t = 18.156, P < 0.001; t = 19.983, P < 0.001, respectively), while expression of proliferating cell nuclear antigen (PCNA) was remarkably decreased (t = 6.828, P < 0.05; t = 14.024, P < 0.05, respectively). 5-ASA in combination with nimesulide suppressed the proliferation of HT-29 colon carcinoma cells more than either of these agents in a dose-dependent and time-dependent manner (t = 5.448, P < 0.05; t = 4.428, P < 0.05, respectively). CONCLUSION: 5-ASA and nimesulide may inhibit the proliferation of HT-29 colon carcinoma cells and coadministration of these agents may have additional chemopreventive potential. PMID:17569127

  20. Protocatechuic aldehyde inhibits migration and proliferation of vascular smooth muscle cells and intravascular thrombosis

    SciTech Connect

    Moon, Chang Yoon; Ku, Cheol Ryong; Cho, Yoon Hee; Lee, Eun Jig

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer Protocatechuic aldehyde (PCA) inhibits ROS production in VSMCs. Black-Right-Pointing-Pointer PCA inhibits proliferation and migration in PDGF-induced VSMCs. Black-Right-Pointing-Pointer PCA has anti-platelet effects in ex vivo rat whole blood. Black-Right-Pointing-Pointer We report the potential therapeutic role of PCA in atherosclerosis. -- Abstract: The migration and proliferation of vascular smooth muscle cells (VSMCs) and formation of intravascular thrombosis play crucial roles in the development of atherosclerotic lesions. This study examined the effects of protocatechuic aldehyde (PCA), a compound isolated from the aqueous extract of the root of Salvia miltiorrhiza, an herb used in traditional Chinese medicine to treat a variety of vascular diseases, on the migration and proliferation of VSMCs and platelets due to platelet-derived growth factor (PDGF). DNA 5-bromo-2 Prime -deoxy-uridine (BrdU) incorporation and wound-healing assays indicated that PCA significantly attenuated PDGF-induced proliferation and migration of VSMCs at a pharmacologically relevant concentration (100 {mu}M). On a molecular level, we observed down-regulation of the phosphatidylinositol 3-kinase (PI3K)/Akt and the mitogen-activated protein kinase (MAPK) pathways, both of which regulate key enzymes associated with migration and proliferation. We also found that PCA induced S-phase arrest of the VSMC cell cycle and suppressed cyclin D2 expression. In addition, PCA inhibited PDGF-BB-stimulated reactive oxygen species production in VSMCs, indicating that PCA's antioxidant properties may contribute to its suppression of PDGF-induced migration and proliferation in VSMCs. Finally, PCA exhibited an anti-thrombotic effect related to its inhibition of platelet aggregation, confirmed with an aggregometer. Together, these findings suggest a potential therapeutic role of PCA in the treatment of atherosclerosis and angioplasty-induced vascular restenosis.

  1. The matricellular protein CCN1 suppresses hepatocarcinogenesis by inhibiting compensatory proliferation.

    PubMed

    Chen, C-C; Kim, K-H; Lau, L F

    2016-03-10

    Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related deaths worldwide, and is on the rise in the United States. Previous studies showed that the matricellular protein CCN1 (CYR61) is induced during hepatic injuries and functions to restrict and resolve liver fibrosis. Here, we show that CCN1 suppresses hepatocarcinogenesis by inhibiting carcinogen-induced compensatory hepatocyte proliferation, thus limiting the expansion of damaged and potentially oncogenic hepatocytes. Consistent with tumor suppression, CCN1 expression is downregulated in human HCC. Ccn1(ΔHep) mice with hepatocyte-specific deletion of Ccn1 suffer increased HCC tumor multiplicity induced by the hepatocarcinogen diethylnitrosamine (DEN). Knockin mice (Ccn1(dm/dm)) that express an integrin α6β1-binding defective CCN1 phenocopied Ccn1(ΔHep) mice, indicating that CCN1 acts through its α6β1 binding sites in this context. CCN1 effectively inhibits epidermal growth factor receptor (EGFR)-dependent hepatocyte proliferation through integrin α6-mediated accumulation of reactive oxygen species (ROS), thereby triggering p53 activation and cell cycle block. Consequently, Ccn1(dm/dm) mice exhibit diminished p53 activation and elevated compensatory hepatocyte proliferation, resulting in increased HCC. Furthermore, we show that a single dose of the EGFR inhibitor erlotinib delivered prior to DEN-induced injury was sufficient to block compensatory proliferation and annihilate development of HCC nodules observed 8 months later, suggesting potential chemoprevention by targeting CCN1-inhibitable EGFR-dependent hepatocyte proliferation. Together, these results show that CCN1 is an injury response protein that functions not only to restrict fibrosis in the liver, but also to suppress hepatocarcinogenesis by inhibiting EGFR-dependent hepatocyte compensatory proliferation.

  2. Peroxisome proliferator-activated receptor-alpha and -gamma mRNA levels are reduced in chronic hepatitis C with steatosis and genotype 3 infection.

    PubMed

    de Gottardi, A; Pazienza, V; Pugnale, P; Bruttin, F; Rubbia-Brandt, L; Juge-Aubry, C E; Meier, C A; Hadengue, A; Negro, F

    2006-01-01

    Steatosis in chronic hepatitis C is associated with inflammation and accelerated fibrogenesis. To assess the contribution of peroxisome proliferator-activated receptor-alpha and -gamma to the pathogenesis of hepatitis C virus associated steatosis is unknown. We measured peroxisome proliferator-activated receptor (PPAR)-alpha and -gamma mRNA by quantitative polymerase chain reaction in liver biopsies of 35 genotype 1 and 22 genotype 3 infected patients and in Huh7 cells expressing hepatitis C virus 1b or 3a core protein. PPAR-alpha mRNA was significantly reduced in livers of patients with genotype 3 compared with genotype 1. Steatosis was associated to a decreased expression of PPAR-alpha in genotype 1, but not in genotype 3. PPAR-gamma expression was significantly lower in genotype 3 compared with genotype 1 and steatosis was associated to decreased levels of PPAR-gamma, but only in genotype 1. There was no significant relationship between PPARs mRNA levels and liver activity or fibrosis. Expression of the hepatitis C virus 3a core protein was associated with an increase in triglyceride accumulation and with a significant reduction of PPAR-gamma mRNA compared with hepatitis C virus 1b. The presence of steatosis and hepatitis C virus genotype 3 are both associated with a significant down-regulation of PPARs. These receptors, and also additional factors, seem to play a role in the pathogenesis of hepatitis C virus-associated steatosis.

  3. Small heterodimer partner-interacting leucine zipper protein inhibits adipogenesis by regulating peroxisome proliferator-activated receptor γ activity.

    PubMed

    Jang, Hoon; Kim, Hyoung-Joo; Kim, Dong-Hwan; Park, Jae-Kyung; Sun, Wu-Sheng; Hwang, Seongsoo; Oh, Keon-Bong; Jang, Won-Gu; Lee, Jeong-Woong

    2015-07-01

    Adipocytes play a critical role in energy balance. Growth of fat tissue is achieved via an increase in adipocyte mass and the formation of newly differentiated adipocytes from precursor cells. Understanding the cellular and molecular mechanisms of adipocyte differentiation is crucial for the study of obesity- and fat-related diseases. The present study was designed to study whether small heterodimer partner-interacting leucine zipper protein (SMILE), a novel co-repressor, could regulate differentiation of adipocyte in 3T3-L1 cells. Treatment of endoplasmic stress inducers, thapsigargin and tunicamycin, inhibited adipocyte differentiation, stimulated Smile mRNA expression, and repressed the expression of adiponectin (Adipoq) in 3T3-L1 pre-adipocyte. Overexpression of SMILE in 3T3-L1 cells decreased the expression of the mRNA encoding Adipoq, a major marker of adipocytes, significantly. Furthermore, knockdown of SMILE recovered the thapsigargin-mediated repression of Adipoq transcription. Co-immunoprecipitation experiments revealed that SMILE interacted physically with PPARγ in 3T3-L1 cells. In addition, chromatin immunoprecipitation experiments revealed that SMILE suppressed the binding affinity of PPARγ for the Adipoq promoter. We demonstrate that SMILE controls adipocyte differentiation by regulating the transactivity of peroxisome proliferator-activated receptor γ (PPARγ). These findings demonstrate that SMILE represses adipocyte differentiation by regulating PPARγ transactivity; hence, SMILE is a potential regulator of PPARγ-related diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Sulforaphane inhibits IL-1β-induced proliferation of rheumatoid arthritis synovial fibroblasts and the production of MMPs, COX-2, and PGE2.

    PubMed

    Choi, Yun Jung; Lee, Won-Seok; Lee, Eun-Gyeong; Sung, Myung-Soon; Yoo, Wan-Hee

    2014-10-01

    This study was performed to define the effects of sulforaphane on interleukin-1β (IL-1β)-induced proliferation of rheumatoid arthritis synovial fibroblasts (RASFs), the expression of matrix metalloproteinases (MMPs) and cyclooxygenase (COX), and the production of prostaglandin E2 (PGE2) by RASFs. The proliferation of RASFs was evaluated with CCK-8 reagent in the presence of IL-1β with/without sulforaphane. The expression of MMPs, tissue inhibitor of metalloproteinase-1, COXs, intracellular mitogen-activated protein kinase signalings, including p-ERK, p-p38, p-JNK, and nuclear factor-kappaB (NF-kB), and the production of PGE2 were examined by Western blotting or semi-quantitative RT-PCR and ELISA. Sulforaphane inhibits unstimulated and IL-1β-induced proliferation of RASFs; the expression of MMP-1, MMP-3, and COX-2 mRNA and protein; and the PGE2 production induced by IL-1β. Sulforaphane also inhibits the phosphorylation of ERK-1/2, p-38, and JNK and activation of NF-kB by IL-1β. These results indicate that sulforaphane inhibits the proliferation of synovial fibroblasts, the expression of MMPs and COX-2, and the production of PGE2, which are involved in synovitis and destruction of RA, and suggest that sulforaphane might be a new therapeutic agent for RA.

  5. mRNA levels of peroxisome proliferator-activated receptors and their coactivators are affected by glucose deprivation and oleate in human hepatoma hepG2 cells.

    PubMed

    Bogdanova, Katerina; Uherkova, Lenka; Poczatkova, Hana; Rypka, Miroslav; Vesely, Jaroslav

    2007-12-01

    Very modest changes in mRNA stability can affect critical points in cellular energy pathways. The aim of this study was to investigate the impact of energy abundant substrates on peroxisome proliferator-activated receptors (PPARs) and PPAR-gamma coactivators (PGCs) mRNA's steady-state levels. Quantitative RT-PCR study was performed to assess the effect of zero or normal (5 mmol/l) glucose and/or oleic acid (0.3 mmol/l) on mRNA levels of (PPARs) (PGCs) in HepG2 cells. PGC-1alpha mRNA was significantly upregulated in glucose deprived cells (123 % of the control level; p < 0.05), while PGC-1beta mRNA was significantly enhanced in oleate-fed cells (134 % and 160 % of control levels for zero glucose plus oleate and normal glucose plus oleate, respectively; p < 0.05) during the 0.5 h incubation. Upon the 4 h incubation, PPAR-gamma1 and PGC-1alpha mRNAs were significantly elevated in cells lacking glucose (142 % and 163 % of control levels, respectively; p < 0.05). Oleate significantly suppressed PPAR-alpha and PGC-1beta mRNA levels in glucose-deprived cells (58 % and 49 % of control levels, respectively; p < 0.05). PPAR-gamma1 and -gamma2 mRNAs were significantly superinduced when the cells were treated with cycloheximide, whereas PPAR-alpha and PGC-1alpha and-1beta mRNAs were destabilized. Upon actinomycin D treatment, glucose shortage significantly stabilized PPAR-alpha mRNA, while PGC-1alpha mRNA was destabilized by oleate in glucose-deprived cells. Our findings provide evidence that transcriptional processes that are under the control of energetic substrates are interconnected with concurrent translational processes that can change stability of mRNAs.

  6. Proliferation of smooth muscle cells stimulated by Porphyromonas gingivalis is inhibited by apple polyphenol.

    PubMed

    Inaba, Hiroaki; Tagashira, Motoyuki; Kanda, Tomomasa; Amano, Atsuo

    2011-11-01

    Porphyromonas gingivalis (Pg) is thought to be involved in the progression of occlusive arterial lesions, whereas vascular smooth muscle cell (SMC) proliferation is considered to be involved in occlusive arterial disease. We previously showed that bacteremia caused by Pg infection induced proliferation of mouse aortic SMCs. Furthermore, human SMCs stimulated with human plasma incubated with Pg showed a marked transformation from the contractile to proliferative phenotype. In the present study, we examine the involvement of Pg gingipains and fimbriae in induction of the SMC transformation and proliferation, and effective inhibitors. Pg strains including gingipain- and fimbria-null mutants were incubated in human plasma, after which the bacteria were removed and the supernatants were added to cultured SMCs. To evaluate the effects of inhibitors, Pg organisms were incubated in plasma in the presence of apple polyphenol (AP), epigallocatechin gallate, KYT-1 (Arg-gingipain inhibitor), and KYT-36 (Lys-gingipain inhibitor). Plasma supernatants from wild-type and fimbria-mutant cultures markedly stimulated cellular proliferation, whereas those containing gingipain-null mutants showed negligible effects. SMC proliferation was also induced by plasma treated with trypsin. Furthermore, plasma supernatants cultured in the presence of KYT-1/KYT-36 and AP showed significant inhibitory effects on SMC proliferation, whereas cultures with epigallocatechin gallate did not. Our results suggest that Pg gingipains are involved in the induction of SMC transformation and proliferation, whereas this was inhibited by AP.

  7. Baicalin inhibits PDGF-induced proliferation and migration of airway smooth muscle cells.

    PubMed

    Yang, Guang; Li, Jian-Qiang; Bo, Jian-Ping; Wang, Bei; Tian, Xin-Rui; Liu, Tan-Zhen; Liu, Zhuo-La

    2015-01-01

    Airway smooth muscle (ASM) cell proliferation and migration play important roles in airway remodeling in asthma. In vitro platelet-derived growth factor (PDGF) induced ASM cell proliferation and migration. Baicalin is one of flavonoid extracts from Scutellaria baicalensis, which has an anti-asthma effect. However, little is known about its role in PDGF-induced proliferation and migration in rat ASM (RASM) cells. In this study, we aimed to investigate the effects of baicalin on PDGF-induced RASM cell proliferation and migration. We also identified the signaling pathway by which baicalin influences RASM cell proliferation and migration. In the current study, we demonstrated that baicalin suppressed PDGF-induced RASM cell proliferation, arrested PDGF-induced cell-cycle progression. It also suppressed PDGF-induced RASM cell migration. Furthermore, baicalin suppressed PDGF-induced expression of phosphorylated p38, ERK1/2 and JNK in RASM cells. In summary, our study is the first to show that baicalin pretreatment can significantly inhibit PDGF-induced RASM cell proliferation and migration by suppressing the MAPK signaling pathway, and baicalin may be a useful chemotherapeutic agent for asthma.

  8. Curcumin ameliorates renal fibrosis by inhibiting local fibroblast proliferation and extracellular matrix deposition.

    PubMed

    Zhou, Xiangjun; Zhang, Jie; Xu, Changgeng; Wang, Wei

    2014-01-01

    Renal fibrosis is mainly characterized by activation and proliferation of interstitial fibroblasts and by excessive synthesis and accumulation of extracellular matrix (ECM) components, including fibronectin (FN) and collagen. This study investigated the effects of curcumin on proliferation of renal interstitial fibroblasts and their underlying mechanisms in vivo and in vitro. ECM components were visualized by Sirius red and immunohistochemistry staining and quantified by western blot analysis in mice with unilateral ureteral obstruction (UUO). Duplex staining for proliferating cell nuclear antigen and α-smooth muscle actin (α-SMA), as well as MTT and flow cytometry assays, were performed to measure fibroblast proliferation. Protein expression of phosphorylated Smad2/3 (p-Smad2/3) and peroxisome proliferator-activated receptor-γ (PPAR-γ) were assessed by western blotting. Curcumin treatment decreased the accumulation of type I collagen and FN in the kidney of animals with UUO. Activation of rat renal interstitial fibroblasts (NRK-49F) was induced by TGF-β1. Curcumin treatment inhibited fibroblast proliferation and the cell cycle was arrested in the G1 phase. Curcumin treatment upregulated the expression of PPAR-γ and downregulated the expression of p-Smad2/3. These results suggest that curcumin treatment ameliorates renal fibrosis by reducing fibroblast proliferation and ECM accumulation mediated by PPAR-γ and Smad-dependent TGF-β1 signaling.

  9. Far-infrared radiation inhibits proliferation, migration, and angiogenesis of human umbilical vein endothelial cells by suppressing secretory clusterin levels.

    PubMed

    Hwang, Soojin; Lee, Dong-Hoon; Lee, In-Kyu; Park, Young Mi; Jo, Inho

    2014-04-28

    Far-infrared (FIR) radiation is known to lessen the risk of angiogenesis-related diseases including cancer. Because deficiency of secretory clusterin (sCLU) has been reported to inhibit angiogenesis of endothelial cells (EC), we investigated using human umbilical vein EC (HUVEC) whether sCLU mediates the inhibitory effects of FIR radiation. Although FIR radiation ranging 3-25μm wavelength at room temperature for 60min did not alter EC viability, further incubation in the culture incubator (at 37°C under 5% CO2) after radiation significantly inhibited EC proliferation, in vitro migration, and tube formation in a time-dependent manner. Under these conditions, we found decreased sCLU mRNA and protein expression in HUVEC and decreased sCLU protein secreted in culture medium. Expectedly, the replacement of control culture medium with the FIR-irradiated conditioned medium significantly decreased wound closure and tube formation of HUVEC, and vice versa. Furthermore, neutralization of sCLU with anti-sCLU antibody also mimicked all observed inhibitory effects of FIR radiation. Moreover, treatment with recombinant human sCLU protein completely reversed the inhibitory effects of FIR radiation on EC migration and angiogenesis. Lastly, vascular endothelial growth factor also increased sCLU secretion in the culture medium, and wound closure and tube formation of HUVEC, which were significantly reduced by FIR radiation. Our results demonstrate a novel mechanism by which FIR radiation inhibits the proliferation, migration, and angiogenesis of HUVEC, via decreasing sCLU.

  10. Inhibition of GLI1 Expression by Targeting the CRD-BP-GLI1 mRNA Interaction Using a Specific Oligonucleotide.

    PubMed

    Mehmood, Kashif; Akhtar, Daud; Mackedenski, Sebastian; Wang, Chuyi; Lee, Chow H

    2016-06-01

    The stabilization of glioma-associated oncogene 1 (GLI1) mRNA by coding region determinant binding protein (CRD-BP) through the Wnt/β-catenin signaling pathway is implicated in the proliferation of colorectal cancer and basal cell carcinoma. Here, we set out to characterize the physical interaction between CRD-BP and GLI1 mRNA so as to find inhibitors for such interaction. Studies using CRD-BP variants with a point mutation in the GXXG motif at each KH domain showed that KH1 and KH2 domain are critical for the binding of GLI1 RNA. The smallest region of GLI1 RNA binding to CRD-BP was mapped to nucleotides (nts) 320-380. A 37-nt S1 RNA sense oligonucleotide, containing two distinct stem-loops present in nts 320-380 of GLI1 RNA, was found to be effective in blocking CRD-BP-GLI1 RNA interaction. Studies using various competitor RNAs with modifications to S1 RNA oligonucleotide further displayed that both the sequences and the structure of the two stem-loops are important for CRD-BP-GLI1 RNA binding. The role of the two-stem-loop motif in influencing CRD-BP-RNA interaction was further investigated in cells. The 2'-O-methyl derivative of the S1 RNA oligonucleotide significantly decreased GLI1, c-myc, and CD44 mRNA levels, in a panel of colon and breast cancer cells. The results from this study demonstrate the potential importance of the two-stem-loop motif as a target region for the inhibition of the CRD-BP-GLI1 RNA interaction and Hedgehog signaling pathway. Such results pave the way for the development of novel inhibitors that act by destabilizing the CRD-BP-GLI1 mRNA interaction.

  11. hnRNP-Q1 represses nascent axon growth in cortical neurons by inhibiting Gap-43 mRNA translation.

    PubMed

    Williams, Kathryn R; McAninch, Damian S; Stefanovic, Snezana; Xing, Lei; Allen, Megan; Li, Wenqi; Feng, Yue; Mihailescu, Mihaela Rita; Bassell, Gary J

    2016-02-01

    Posttranscriptional regulation of gene expression by mRNA-binding proteins is critical for neuronal development and function. hnRNP-Q1 is an mRNA-binding protein that regulates mRNA processing events, including translational repression. hnRNP-Q1 is highly expressed in brain tissue, suggesting a function in regulating genes critical for neuronal development. In this study, we have identified Growth-associated protein 43 (Gap-43) mRNA as a novel target of hnRNP-Q1 and have demonstrated that hnRNP-Q1 represses Gap-43 mRNA translation and consequently GAP-43 function. GAP-43 is a neuronal protein that regulates actin dynamics in growth cones and facilitates axonal growth. Previous studies have identified factors that regulate Gap-43 mRNA stability and localization, but it remains unclear whether Gap-43 mRNA translation is also regulated. Our results reveal that hnRNP-Q1 knockdown increased nascent axon length, total neurite length, and neurite number in mouse embryonic cortical neurons and enhanced Neuro2a cell process extension; these phenotypes were rescued by GAP-43 knockdown. Additionally, we have identified a G-quadruplex structure in the 5' untranslated region of Gap-43 mRNA that directly interacts with hnRNP-Q1 as a means to inhibit Gap-43 mRNA translation. Therefore hnRNP-Q1-mediated repression of Gap-43 mRNA translation provides an additional mechanism for regulating GAP-43 expression and function and may be critical for neuronal development.

  12. hnRNP-Q1 represses nascent axon growth in cortical neurons by inhibiting Gap-43 mRNA translation

    PubMed Central

    Williams, Kathryn R.; McAninch, Damian S.; Stefanovic, Snezana; Xing, Lei; Allen, Megan; Li, Wenqi; Feng, Yue; Mihailescu, Mihaela Rita; Bassell, Gary J.

    2016-01-01

    Posttranscriptional regulation of gene expression by mRNA-binding proteins is critical for neuronal development and function. hnRNP-Q1 is an mRNA-binding protein that regulates mRNA processing events, including translational repression. hnRNP-Q1 is highly expressed in brain tissue, suggesting a function in regulating genes critical for neuronal development. In this study, we have identified Growth-associated protein 43 (Gap-43) mRNA as a novel target of hnRNP-Q1 and have demonstrated that hnRNP-Q1 represses Gap-43 mRNA translation and consequently GAP-43 function. GAP-43 is a neuronal protein that regulates actin dynamics in growth cones and facilitates axonal growth. Previous studies have identified factors that regulate Gap-43 mRNA stability and localization, but it remains unclear whether Gap-43 mRNA translation is also regulated. Our results reveal that hnRNP-Q1 knockdown increased nascent axon length, total neurite length, and neurite number in mouse embryonic cortical neurons and enhanced Neuro2a cell process extension; these phenotypes were rescued by GAP-43 knockdown. Additionally, we have identified a G-quadruplex structure in the 5′ untranslated region of Gap-43 mRNA that directly interacts with hnRNP-Q1 as a means to inhibit Gap-43 mRNA translation. Therefore hnRNP-Q1–mediated repression of Gap-43 mRNA translation provides an additional mechanism for regulating GAP-43 expression and function and may be critical for neuronal development. PMID:26658614

  13. miR-130b targets NKD2 and regulates the Wnt signaling to promote proliferation and inhibit apoptosis in osteosarcoma cells

    SciTech Connect

    Li, Zhi; Li, Youjun; Wang, Nan; Yang, Lifeng; Zhao, Wei; Zeng, Xiandong

    2016-03-18

    miR-130b was significantly up-regulated in osteosarcoma (OS) cells. Naked cuticle homolog 2 (NKD2) inhibited tumor growth and metastasis in OS by suppressing Wnt signaling. We used three miRNA target analysis tools to identify potential targets of miR-130b, and found that NKD2 is a potential target of miR-130b. Based on these findings, we hypothesize that miR-130b might target NKD2 and regulate the Wnt signaling to promote OS growth. We detected the expression of miR-130b and NKD2 mRNA and protein by quantitative Real-Time PCR (qRT-PCR) and western blot assays, respectively, and found up-regulation of miR-130b and down-regulation of NKD2 mRNA and protein exist in OS cell lines. MTT and flow cytometry assays showed that miR-130b inhibitors inhibit proliferation and promote apoptosis in OS cells. Furthermore, we showed that NKD2 is a direct target of miR-130b, and miR-130b regulated proliferation and apoptosis of OS cells by targeting NKD2. We further investigated whether miR-130b and NKD2 regulate OS cell proliferation and apoptosis by inhibiting Wnt signaling, and the results confirmed our speculation that miR-130b targets NKD2 and regulates the Wnt signaling to promote proliferation and inhibit apoptosis of OS cells. These findings will offer new clues for OS development and progression, and novel potential therapeutic targets for OS. - Highlights: • miR-130b is up-regulated and NKD2 is down-regulated in osteosarcoma cell lines. • Down-regulation of miR-130b inhibits proliferation of osteosarcoma cells. • Down-regulation of miR-130b promotes apoptosis of osteosarcoma cells. • miR-130b directly targets NKD2. • NKD2 regulates OS cell proliferation and apoptosis by inhibiting the Wnt signaling.

  14. Overexpression of FABP3 inhibits human bone marrow derived mesenchymal stem cell proliferation but enhances their survival in hypoxia

    SciTech Connect

    Wang, Suna Zhou, Yifu; Andreyev, Oleg; Hoyt, Robert F.; Singh, Avneesh; Hunt, Timothy; Horvath, Keith A.

    2014-04-15

    Studying the proliferative ability of human bone marrow derived mesenchymal stem cells in hypoxic conditions can help us achieve the effective regeneration of ischemic injured myocardium. Cardiac-type fatty acid binding protein (FABP3) is a specific biomarker of muscle and heart tissue injury. This protein is purported to be involved in early myocardial development, adult myocardial tissue repair and responsible for the modulation of cell growth and proliferation. We have investigated the role of FABP3 in human bone marrow derived mesenchymal stem cells under ischemic conditions. MSCs from 12 donors were cultured either in standard normoxic or modified hypoxic conditions, and the differential expression of FABP3 was tested by quantitative {sup RT}PCR and western blot. We also established stable FABP3 expression in MSCs and searched for variation in cellular proliferation and differentiation bioprocesses affected by hypoxic conditions. We identified: (1) the FABP3 differential expression pattern in the MSCs under hypoxic conditions; (2) over-expression of FABP3 inhibited the growth and proliferation of the MSCs; however, improved their survival in low oxygen environments; (3) the cell growth factors and positive cell cycle regulation genes, such as PCNA, APC, CCNB1, CCNB2 and CDC6 were all down-regulated; while the key negative cell cycle regulation genes TP53, BRCA1, CASP3 and CDKN1A were significantly up-regulated in the cells with FABP3 overexpression. Our data suggested that FABP3 was up-regulated under hypoxia; also negatively regulated the cell metabolic process and the mitotic cell cycle. Overexpression of FABP3 inhibited cell growth and proliferation via negative regulation of the cell cycle and down-regulation of cell growth factors, but enhances cell survival in hypoxic or ischemic conditions. - Highlights: • FABP3 expression pattern was studied in 12 human hypoxic-MSCs. • FABP3 mRNA and proteins are upregulated in the MSCs under hypoxic conditions.

  15. Inhibition of miR-29c promotes proliferation, and inhibits apoptosis and differentiation in P19 embryonic carcinoma cells.

    PubMed

    Chen, Bin; Song, Guixian; Liu, Ming; Qian, Lingmei; Wang, Lihua; Gu, Haitao; Shen, Yahui

    2016-03-01

    In our previous study, the upregulation of microRNA (miR)-29c was identified in the mother of a fetus with a congenital heart defect. However, the functional and regulatory mechanisms of miR‑29c in the development of the heart remain to be elucidated. In the present study, the role and mechanism of miR‑29c inhibition in heart development were investigated in an embryonic carcinoma cell model. Inhibition of miR‑29c promoted proliferation, and suppressed the apoptosis and differentiation of P19 cells. It was also demonstrated that Wingless‑related MMTV integration site 4 (Wnt4) was a target of miR‑29c, determined using bioinformatic analysis combined with luciferase assays. The inhibition of miR‑29c stimulated the WNT4/β‑catenin pathway, promoting proliferation of the P19 cells, but suppressing their differentiation into cardiomyocytes. Furthermore, the inhibition of miR‑29c promoted the expression of B cell lymphoma‑2 and inhibited cell apoptosis. These results demonstrate the significance of miR‑29c in the process of cardiac development and suggest that miR-29c dysregulation may be associated with the occurrence of CHD. Thus, miR-29c may have therapeutic potential in the future.

  16. Proliferation and mRNA expression of absorptive villous cell markers and mineral transporters in prolactin-exposed IEC-6 intestinal crypt cells.

    PubMed

    Teerapornpuntakit, Jarinthorn; Wongdee, Kannikar; Thongbunchoo, Jirawan; Krishnamra, Nateetip; Charoenphandhu, Narattaphol

    2012-06-01

    During pregnancy and lactation, prolactin (PRL) enhances intestinal absorption of calcium and other minerals for fetal development and milk production. Although an enhanced absorptive efficiency is believed to mainly result from the upregulation of mineral transporters in the absorptive villous cells, some other possibilities, such as PRL-enhanced crypt cell proliferation and differentiation to increase the absorptive area, have never been ruled out. Here, we investigated cell proliferation and mRNA expression of mineral absorption-related genes in the PRL-exposed IEC-6 crypt cells. As expected, the cell proliferation was not altered by PRL. Inasmuch as the mRNA expressions of villous cell markers, including dipeptidylpeptidase-4, lactase and glucose transporter-5, were not increased, PRL was not likely to enhance crypt cell differentiation into the absorptive villous cells. In contrast to the previous findings in villous cells, PRL was found to downregulate the expression of calbindin-D(9k), claudin-3 and occludin in IEC-6 crypt cells, while having no effect on transient receptor potential vanilloid family channels-5/6, plasma membrane Ca(2+)-ATPase (PMCA)-1b and Na(+)/Ca(2+) exchanger-1 expression. In conclusion, IEC-6 crypt cells did not respond to PRL by increasing proliferation or differentiation into villous cells. The present results thus supported the previous hypothesis that PRL enhanced mineral absorption predominantly by increasing transporter expression and activity in the absorptive villous cells. Copyright © 2012 John Wiley & Sons, Ltd.

  17. MicroRNA-125b-5p inhibits proliferation and promotes adipogenic differentiation in 3T3-L1 preadipocytes.

    PubMed

    Ouyang, Dan; Ye, Yaqiong; Guo, Dongguang; Yu, Xiaofang; Chen, Jian; Qi, Junjie; Tan, Xiaotong; Zhang, Yuan; Ma, Yongjiang; Li, Yugu

    2015-05-01

    Previous evidence has indicated that the microRNA-125b (miR-125b) family plays important roles in the regulation of cancer cell growth, development, differentiation, and apoptosis. However, whether they contribute to the process of adipocyte differentiation remains unclear. In the present study, we revealed that the expression level of miR-125b-5p, a member of miR-125b family, was dramatically up-regulated during differentiation of 3T3-L1 preadipocyte into mature adipocyte. Supplement of miR-125b-5p into 3T3-L1 cells promoted adipogenic differentiation as evidenced by increased lipid droplets and mRNA levels of adipocyte-specific molecular markers, including peroxisome proliferators-activated receptor γ, CCAAT/enhancer-binding protein α, fatty acid-binding protein 4, and lipoprotein lipase, and by triglyceride accumulation. CCK-8 assay showed that miR-125b-5p supplementation significantly inhibited cell proliferation. Flow cytometry analysis showed that miR-125b-5p impaired G1/S phase transition as well as the mRNA and protein expression of G1/S-related genes, such as Cyclin D2, Cyclin D3, and CDK4. Nevertheless, it had no effect on apoptosis. Additionally, by target gene prediction, we demonstrated that smad4 may be a potential target of miR-125b-5p in mouse 3T3-L1 preadipocytes, accounting for some of miR-125b-5p's functions. Taken together, these data indicated that miR-125b-5p may serve as an important positive regulator in adipocyte differentiation, at least partially through down-regulating smad4.

  18. MicroRNA-490-5p inhibits proliferation of bladder cancer by targeting c-Fos

    SciTech Connect

    Li, Shiqi; Xu, Xianglai; Xu, Xin; Hu, Zhenghui; Wu, Jian; Zhu, Yi; Chen, Hong; Mao, Yeqing; Lin, Yiwei; Luo, Jindan; Zheng, Xiangyi; Xie, Liping

    2013-11-29

    Highlights: •We examined the level of miR-490-5p in bladder cancer tissues and three cancer cell lines. •We are the first to show the function of miR-490-5p in bladder cancer. •We demonstrate c-Fos may be a target of miR-490-5p. -- Abstract: MicroRNAs (miRNAs) are non-protein-coding sequences that play a crucial role in tumorigenesis by negatively regulating gene expression. Here, we found that miR-490-5p is down-regulated in human bladder cancer tissue and cell lines compared to normal adjacent tissue and a non-malignant cell line. To better characterize the function of miR-490-5p in bladder cancer, we over-expressed miR-490-5p in bladder cancer cell lines with chemically synthesized mimics. Enforced expression of miR-490-5p in bladder cancer cells significantly inhibited the cell proliferation via G1-phase arrest. Further studies found the decreased c-Fos expression at both mRNA and protein levels and Luciferase reporter assays demonstrated that c-Fos is a direct target of miR-490-5p in bladder cancer. These findings indicate miR-490-5p to be a novel tumor suppressor of bladder cancer cell proliferation through targeting c-Fos.

  19. Inhibition of human copper trafficking by a small molecule significantly attenuates cancer cell proliferation

    NASA Astrophysics Data System (ADS)

    Wang, Jing; Luo, Cheng; Shan, Changliang; You, Qiancheng; Lu, Junyan; Elf, Shannon; Zhou, Yu; Wen, Yi; Vinkenborg, Jan L.; Fan, Jun; Kang, Heebum; Lin, Ruiting; Han, Dali; Xie, Yuxin; Karpus, Jason; Chen, Shijie; Ouyang, Shisheng; Luan, Chihao; Zhang, Naixia; Ding, Hong; Merkx, Maarten; Liu, Hong; Chen, Jing; Jiang, Hualiang; He, Chuan

    2015-12-01

    Copper is a transition metal that plays critical roles in many life processes. Controlling the cellular concentration and trafficking of copper offers a route to disrupt these processes. Here we report small molecules that inhibit the human copper-trafficking proteins Atox1 and CCS, and so provide a selective approach to disrupt cellular copper transport. The knockdown of Atox1 and CCS or their inhibition leads to a significantly reduced proliferation of cancer cells, but not of normal cells, as well as to attenuated tumour growth in mouse models. We show that blocking copper trafficking induces cellular oxidative stress and reduces levels of cellular ATP. The reduced level of ATP results in activation of the AMP-activated protein kinase that leads to reduced lipogenesis. Both effects contribute to the inhibition of cancer cell proliferation. Our results establish copper chaperones as new targets for future developments in anticancer therapies.

  20. Inhibition of human copper trafficking by a small molecule significantly attenuates cancer cell proliferation.

    PubMed

    Wang, Jing; Luo, Cheng; Shan, Changliang; You, Qiancheng; Lu, Junyan; Elf, Shannon; Zhou, Yu; Wen, Yi; Vinkenborg, Jan L; Fan, Jun; Kang, Heebum; Lin, Ruiting; Han, Dali; Xie, Yuxin; Karpus, Jason; Chen, Shijie; Ouyang, Shisheng; Luan, Chihao; Zhang, Naixia; Ding, Hong; Merkx, Maarten; Liu, Hong; Chen, Jing; Jiang, Hualiang; He, Chuan

    2015-12-01

    Copper is a transition metal that plays critical roles in many life processes. Controlling the cellular concentration and trafficking of copper offers a route to disrupt these processes. Here we report small molecules that inhibit the human copper-trafficking proteins Atox1 and CCS, and so provide a selective approach to disrupt cellular copper transport. The knockdown of Atox1 and CCS or their inhibition leads to a significantly reduced proliferation of cancer cells, but not of normal cells, as well as to attenuated tumour growth in mouse models. We show that blocking copper trafficking induces cellular oxidative stress and reduces levels of cellular ATP. The reduced level of ATP results in activation of the AMP-activated protein kinase that leads to reduced lipogenesis. Both effects contribute to the inhibition of cancer cell proliferation. Our results establish copper chaperones as new targets for future developments in anticancer therapies.

  1. Inhibition of human copper trafficking by a small molecule significantly attenuates cancer cell proliferation

    PubMed Central

    Wang, Jing; Luo, Cheng; Shan, Changliang; You, Qiancheng; Lu, Junyan; Elf, Shannon; Zhou, Yu; Wen, Yi; Vinkenborg, Jan L.; Fan, Jun; Kang, Heebum; Lin, Ruiting; Han, Dali; Xie, Yuxin; Karpus, Jason; Chen, Shijie; Ouyang, Shisheng; Luan, Chihao; Zhang, Naixia; Ding, Hong; Merkx, Maarten; Liu, Hong; Chen, Jing; Jiang, Hualiang; He, Chuan

    2016-01-01

    Copper is a transition metal that plays critical roles in many life processes. Controlling the cellular concentration and trafficking of copper offers a route to disrupt these processes. Here we report small molecules that inhibit the human copper-trafficking proteins Atox1 and CCS, and so provide a selective approach to disrupt cellular copper transport. The knockdown of Atox1 and CCS or their inhibition leads to a significantly reduced proliferation of cancer cells, but not of normal cells, as well as to attenuated tumour growth in mouse models. We show that blocking copper trafficking induces cellular oxidative stress and reduces levels of cellular ATP. The reduced level of ATP results in activation of the AMP-activated protein kinase that leads to reduced lipogenesis. Both effects contribute to the inhibition of cancer cell proliferation. Our results establish copper chaperones as new targets for future developments in anticancer therapies. PMID:26587712

  2. Inhibition of smooth muscle cell proliferation and protein kinase C activity by tocopherols and tocotrienols.

    PubMed

    Chatelain, E; Boscoboinik, D O; Bartoli, G M; Kagan, V E; Gey, F K; Packer, L; Azzi, A

    1993-03-10

    alpha-Tocopherol, the most active form of vitamin E, causes a dose-dependent inhibition of serum-induced proliferation of smooth muscle cells (A7r5) in culture. Some tocopherol-related compounds exhibiting various degrees of antioxidant potency have also been tested on cellular proliferation. No direct correlation between the antioxidant activity of these compounds and their effect on smooth muscle cell growth could be observed. While most of the derivatives employed were not effective in inhibiting protein kinase C, in the case of alpha-tocopherol the antiproliferative effect was found to be parallel to the inhibition of protein kinase C activity, as measured in streptolysin-O permeabilized cells.

  3. Purification and Properties of a Rat Liver Protein That Specifically Inhibits the Proliferation of Nonmalignant Epithelial Cells from Rat Liver

    NASA Astrophysics Data System (ADS)

    McMahon, James B.; Farrelly, James G.; Iype, P. Thomas

    1982-01-01

    An inhibitor of cell proliferation was purified from rat liver by alcohol precipitation, ultrafiltration, and DEAE-cellulose chromatography. The hepatic proliferation inhibitor was shown to be pure by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, analytical isoelectric focusing, and high-performance liquid chromatography. The hepatic proliferation inhibitor was found to have a molecular weight of 26,000 and an isoelectric point of 4.65. This protein inhibited the proliferation of nonmalignant rat liver cells in culture, and removal of the protein reversed the inhibition produced by low doses. It exerted no effect on the proliferation of malignant rat liver cells.

  4. The Ba fragment of complement factor B inhibits human B lymphocyte proliferation.

    PubMed

    Ambrus, J L; Peters, M G; Fauci, A S; Brown, E J

    1990-03-01

    Normal human B lymphocyte function is finely regulated by both positive and negative signals at each stage of activation, proliferation, and differentiation. Activation signals include antigen and surface Ig cross-linking agents such as anti-mu or anti-delta. Signals inducing proliferation include IL-2, high m.w.-B cell growth factor (BCGF), and low m.w.-BCGF. IL-2 as well as IL-6 and other partially characterized B cell differentiation factors can induce terminal differentiation of proliferating B cells into Ig-secreting plasma cells. Various C components have been described to regulate B cell function including Bb that enhances proliferation, C5a that enhances Ig production, and C3a that inhibits Ig production. In our study, we examined the ability of the factor B cleavage fragment Ba to influence human B cell function. Ba did not affect the activation of resting B cells but inhibited the proliferation of activated B cells stimulated with either high m.w.-BCGF or low m.w.-BCGF. The inhibition occurred with doses of Ba as low as 1 microgram/ml (29 nM). Ba was found to bind to activated human B lymphocytes in a saturable manner with an apparent K of approximately 25 nM and an apparent Bmax of 56,000 sites/cell. A peptide made of the carboxy terminal 10 amino acids of Ba (GHGPGEQQKR), was also found to inhibit growth factor induced proliferation of activated B cells but at an ID50 of approximately 5 microM. Finally, Ba was found to inhibit the terminal differentiation of Staphylococcus aweus Cowan-activated B cells stimulated with B cell differentiation factors but not Ig secretion by the partially differentiated EBV-transformed cell line SKW.6. Thus, concentrations of Ba achievable in vivo at sites of active inflammation were found to act on human B lymphocytes by inhibiting their proliferation. This may act to limit the immune response to a specific antigenic challenge.

  5. MicroRNA-126 inhibits tumor proliferation and angiogenesis of hepatocellular carcinoma by down-regulating EGFL7 expression

    PubMed Central

    Hu, Ming-Hua; Ma, Chen-Yang; Wang, Xiao-Ming; Ye, Chen-Dong; Zhang, Guang-Xian; Chen, Lin; Wang, Jin-Guo

    2016-01-01

    This study aims to explore the effects of microRNA-126 (miR-126) on tumor proliferation and angiogenesis of hepatocellular carcinoma (HCC) by targeting EGFL7. HCC tissues and adjacent normal tissues were obtained from 71 HCC patients. Immunohistochemistry (IHC) was conducted to detect expressions of EGFL7 and VEGF and the micro-vessel density (MVD). HCC cell lines were collected and assigned into the blank, miR-126 mimics, miR-126 inhibitors, miR-126 mimics negative control (NC), miR-126 inhibitors NC, si-EGFL7, and miR-126 inhibitors + si-EGFL7 groups. Expressions of miR-126 and EGFL7 mRNA were detected by qRT-PCR assay. The protein expressions of EGFL7 and VEGF were measured by Western blotting. MTT assay was used to measure the proliferation of HCC cells. Tumor xenograft model in nude mice was utilized to evaluate the influence of miR-126 on tumor growth. HCC tissues had higher miR-126 expression and lower EGFL7 mRNA expression than adjacent normal tissues. Compared with the blank, miR-126 mimic NC, miR-126 inhibitor NC and miR-126 inhibitors + si-EGFL7 groups, the protein expressions of EGFL7 and VEGF and cell proliferation were reduced in the miR-126 mimics and si-EGFL7 groups, while the opposite trend was found in the miR-126 inhibitors group. Compared with the blank and miR-126 inhibitors + siRNA-EGFL7 groups, tumor size, tumor weight, and MVD of transplanted tumors in nude mice were significantly reduced in the miR-126 mimics and siRNA-EGFL7 groups, while the opposite trend was found in the miR-126 inhibitors group. In conclusion, miR-126 could inhibit tumor proliferation and angiogenesis of HCC by down-regulating EGFL7 expression. PMID:27611944

  6. Inhibition of Excessive Cell Proliferation by Calcilytics in Idiopathic Pulmonary Arterial Hypertension

    PubMed Central

    Yamamura, Aya; Ohara, Naoki; Tsukamoto, Kikuo

    2015-01-01

    Idiopathic pulmonary arterial hypertension (IPAH) is a rare and progressive disease of unknown pathogenesis. Vascular remodeling due to excessive proliferation of pulmonary arterial smooth muscle cells (PASMCs) is a critical pathogenic event that leads to early morbidity and mortality. The excessive cell proliferation is closely linked to the augmented Ca2+ signaling in PASMCs. More recently, we have shown by an siRNA knockdown method that the Ca2+-sensing receptor (CaSR) is upregulated in PASMCs from IPAH patients, involved in the enhanced Ca2+ response and subsequent excessive cell proliferation. In this study, we examined whether pharmacological blockade of CaSR attenuated the excessive proliferation of PASMCs from IPAH patients by MTT assay. The proliferation rate of PASMCs from IPAH patients was much higher (~1.5-fold) than that of PASMCs from normal subjects and patients with chronic thromboembolic pulmonary hypertension (CTEPH). Treatment with NPS2143, an antagonist of CaSR or calcilytic, clearly suppressed the cell proliferation in a concentration-dependent manner (IC50 = 2.64 μM) in IPAH-PASMCs, but not in normal and CTEPH PASMCs. Another calcilytic, Calhex 231, which is structurally unrelated to NPS2143, also concentration-dependently inhibited the excessive proliferation of IPAH-PASMCs (IC50 = 1.89 μM). In contrast, R568, an activator of CaSR or calcimimetic, significantly facilitated the proliferation of IPAH-PASMCs (EC50 = 0.33 μM). Similar results were obtained by BrdU incorporation assay. These results reveal that the excessive PASMC proliferation was modulated by pharmacological tools of CaSR, showing us that calcilytics are useful for a novel therapeutic approach for pulmonary arterial hypertension. PMID:26375676

  7. Let-7a Inhibits T-Cell Proliferation and IFN-γ Secretion by Down-Regulating STAT3 Expression in Patients with Psoriasis.

    PubMed

    Hu, Xiao-Ping; Xie, Qian; Chen, Chao-Feng; Zhang, Wei; Yu, Bo

    2017-01-01

    This study aimed to explore the effects of STAT3 targeting by let-7a on T-cell proliferation and IFN-γ secretion in psoriasis. From January 2013 to January 2015, 40 patients with psoriasis (psoriasis group) and 38 volunteers undergoing plastic surgery (control group) were enrolled in this study. Pearson correlation analysis was performed to evaluate the correlation between let-7a and STAT3 expression. T-cells were isolated and subjected to different transfection methods. A dual luciferase reporter assay was carried out to confirm STAT3 as a target gene of let-7a. Let-7a, STAT3 and IFN-γ mRNA expression was detected by quantitative real-time fluorescent polymerase chain reaction (qRT-PCR), and pSTAT3 protein levels were determined by Western blot. T-cell proliferation was evaluated with a cell counting kit-8 (CCK-8) assay. The level of STAT3 mRNA and pSTAT3 was higher, but let-7a expression was lower in the psoriasis group than the control group. Pearson correlation analysis indicated that STAT3 expression was negatively correlated with let-7a expression. T-cells transfected with inhibitors exhibited greater IFN-γ mRNA expression and T-cell proliferation than transfected T-cells and T-cells transfected with a non-sense sequence, while T-cells transfected with let-7a mimics exhibited lower IFN-γ mRNA expression and T-cell proliferation than transfected T-cells and T-cells transfected with a non-sense sequence. This suggested that siRNA-STAT3 could reverse the increase in IFN-y mRNA expression and T-cell proliferation induced by let-7a inhibitors. Our results demonstrated that let-7a inhibits T-cell proliferation and IFN-γ secretion by down-regulating STAT3 in psoriasis. © 2017 The Author(s). Published by S. Karger AG, Basel.

  8. CKIP-1 regulates macrophage proliferation by inhibiting TRAF6-mediated Akt activation

    PubMed Central

    Zhang, Luo; Wang, Yiwu; Xiao, Fengjun; Wang, Shaoxia; Xing, Guichun; Li, Yang; Yin, Xiushan; Lu, Kefeng; Wei, Rongfei; Fan, Jiao; Chen, Yuhan; Li, Tao; Xie, Ping; Yuan, Lin; Song, Lei; Ma, Lanzhi; Ding, Lujing; He, Fuchu; Zhang, Lingqiang

    2014-01-01

    Macrophages play pivotal roles in development, homeostasis, tissue repair and immunity. Macrophage proliferation is promoted by macrophage colony-stimulating factor (M-CSF)-induced Akt signaling; yet, how this process is terminated remains unclear. Here, we identify casein kinase 2-interacting protein-1 (CKIP-1) as a novel inhibitor of macrophage proliferation. In resting macrophages, CKIP-1 was phosphorylated at Serine 342 by constitutively active GSK3β, the downstream target of Akt. This phosphorylation triggers the polyubiquitination and proteasomal degradation of CKIP-1. Upon M-CSF stimulation, Akt is activated by CSF-1R-PI3K and then inactivates GSK3β, leading to the stabilization of CKIP-1 and β-catenin proteins. β-catenin promotes the expression of proliferation genes including cyclin D and c-Myc. CKIP-1 interacts with TRAF6, a ubiquitin ligase required for K63-linked ubiquitination and plasma membrane recruitment of Akt, and terminates TRAF6-mediated Akt activation. By this means, CKIP-1 inhibits macrophage proliferation specifically at the late stage after M-CSF stimulation. Furthermore, CKIP-1 deficiency results in increased proliferation and decreased apoptosis of macrophages in vitro and CKIP-1−/− mice spontaneously develop a macrophage-dominated splenomegaly and myeloproliferation. Together, these data demonstrate that CKIP-1 plays a critical role in the regulation of macrophage homeostasis by inhibiting TRAF6-mediated Akt activation. PMID:24777252

  9. Radish (Raphanus sativus L. leaf) ethanol extract inhibits protein and mRNA expression of ErbB(2) and ErbB(3) in MDA-MB-231 human breast cancer cells.

    PubMed

    Kim, Woo Kyoung; Kim, Ji Hae; Jeong, Da Hee; Chun, Young Hee; Kim, Sun Hee; Cho, Kang Jin; Chang, Moon-Jeong

    2011-08-01

    In this study, we investigated the effects of the ethanol extract of aerial parts of Raphanus sativus L. (ERL) on breast cancer cell proliferation and gene expression associated with cell proliferation and apoptosis in MDA-MB-231 human breast cancer cells. The MDA-MB-231 cells were cultured in the presence or absence of various concentrations (100, 200, or 300 µg/mL) of ERL. ERL significantly decreased cell proliferation after 48 h of incubation (P < 0.05). The protein and mRNA expression of ErbB(2) were decreased significantly in a dose-dependent manner (P < 0.05). The protein expression of ErbB(3) was decreased significantly at an ERL concentration of 300 µg/mL (P < 0.05), and mRNA expression of ErbB(3) was decreased significantly in a dose-dependent manner (P < 0.05). The protein expression of Akt was decreased significantly at the ERL concentration of 200 µg/mL (P < 0.05), and the protein expression of pAkt was decreased significantly in a dose-dependent manner (P < 0.05). The mRNA expression of Akt was decreased significantly at the ERL concentration of 200 µg/mL ERL (P < 0.05). The protein and mRNA expression of Bax were increased significantly at ERL concentrations of 200 µg/mL or higher (P < 0.05). The protein expression of Bcl(2) was increased significantly at ERL concentrations of 100 µg/mL or higher (P < 0.05), and mRNA expression of Bcl(2) was increased significantly at an ERL concentration of 300 µg/mL (P < 0.05). In conclusion, we suggest that Raphanus sativus, L. inhibits cell proliferation via the ErbB-Akt pathway in MDA-MB-231 cells.

  10. Effects of Thalidomide Combined with Interferon on Inhibiting Kasumi-1 Cell Proliferation.

    PubMed

    Xu, Hao; Mi, Ruihua; Fan, Ruihua; Yin, Qingsong; Wei, Xudong

    2016-01-01

    Our previous clinical observations proved that the combination of thalidomide and interferon (IFN) had certain effects in relapsed or refractory AML. The aim of this study was to investigate the effects and its mechanism of thalidomide and IFN on inhibiting the proliferation of Kasumi-1 cells. Thalidomide, IFN and a combination of both drugs were used to treat Kasumi-1 cells. The inhibition of cell proliferation and the apoptosis rate were measured. Vascular endothelial growth factor levels and the expression of apoptosis-related proteins were detected by ELISA and Western blotting, respectively. Thalidomide and IFN could both inhibit Kasumi-1 cell proliferation in a dose-dependent manner. When Kasumi-1 cells were treated with thalidomide 350 μg/mL or IFN1400 U/mL for 48 h, the proliferation inhibition rates were (48.8 ± 4.64)% and (50.19 ± 2.59)% and the rates of apoptosis were (14.68 ± 2.61)% and (21.71 ± 0.71)%, respectively; when treated with a combination, the cell proliferation inhibition rate and apoptotic rate were statistically significantly higher than both the control group and the groups treated with a single drug. The ELISA assay revealed that both 350 μg/mL of thalidomide and 1400 U/mL of IFN could reduce the VEGF levels in cell culture supernatants; the two-drug combination group had a further decreased VEGF concentration. Forty-eighthour treatment of thalidomide 350 μg/mL and IFN 1400 U/mL could significantly decrease Bcl-2 expression and increase the expression levels of phosphor-P38, BAX, cytochrome c, and cleaved caspase-3, -8, and -9 as compared to the control group. The combination group exhibited significantly greater extents of reduction in Bcl-2 protein and increases in p-P38, BAX, and cytochrome c, and cleaved caspase-3, -8, and -9 protein expression as compared to the single drug groups. Thalidomide and IFN can synergistically inhibit Kasumi-1 cell proliferation, which is possibly achieved through the mitochondrial and death

  11. Targeted inhibition of p57 and p15 blocks transforming growth factor β-inhibited proliferation of primary cultured human limbal epithelial cells

    PubMed Central

    Chen, Zhuo; Li, De-quan; Tong, Louis; Stewart, Paul; Chu, Claire; Pflugfelder, Stephen C.

    2010-01-01

    Purpose To evaluate the role of cyclin-dependent kinase inhibitors p57 and p15 in transforming growth factor (TGF)-β1 or TGF-β2 inhibited proliferation of primary cultured human limbal epithelial cells using short interfering RNA (siRNA). Methods Primary cultured human limbal epithelial cells were treated with TGF-β1 or TGF-β2 for 6 and 24 h, and total RNA extracted for RT-PCR and real-time PCR using primers for p21, p27, and p57 (CipP/Kip family) and p15 and p19 (INK4 family). Proteins were extracted for western blot analysis of p57 and p15. For RNA interference, primary cultured human limbal epithelial cells were transfected with annealed double-stranded siRNA (67 nM) specific for p57, p15, or siRNA-Fluorescein (siRNA-F; as a negative control) followed by treatment with TGF-β1 or TGF-β2 at 1 ng/ml. P57 and p15 were quantitatively detected by real-time PCR and western blot; and immunolocalized by immunofluorescent staining. The effects of TGF-β1 or TGF-β2 on cell proliferation were evaluated by BrdU incorporation and MTT assay. Results TGF-β1 or TGF-β2 significantly inhibited primary cultured human limbal epithelial cell proliferation measured by BrdU incorporation and MTT assay. TGF-β1 or TGF-β2 upregulated the expression of p57 and p15 mRNA and protein, but did not effect the expression of p19, p21, or p27. The siRNA transfection efficiency of these cells was 75% and no cellular toxicity was observed by 24 h. The TGF-β1 or TGF-β2 stimulated expression of p57 and p15 mRNA were markedly blocked by siRNA-p57 or siRNA-p15, respectively, but not by siRNA-F. The TGF-β1 or TGF-β2 suppression of epithelial proliferation measured by BrdU incorporation and MTT generation was increased to near normal levels by siRNA-p57 or siRNA-p15. Western blot and immunofluorescent staining showed that levels of p57 and p15 proteins were equally reduced in the cytoplasm and nucleus. Conclusions These findings demonstrate that TGF-β1 and/or TGF-β2 inhibit proliferation

  12. Knockdown of Golgi phosphoprotein 2 inhibits hepatocellular carcinoma cell proliferation and motility

    PubMed Central

    Liu, Yiming; Zhang, Xiaodi; Sun, Ting; Jiang, Junchang; Li, Ying; Chen, Mingliang; Wei, Zhen; Jiang, Weiqin; Zhou, Linfu

    2016-01-01

    Golgi phosphoprotein 2 (GP73) is highly expressed in hepatocellular carcinoma (HCC) cells, where it serves as a biomarker and indicator of disease progression. We used MTS assays, anchorage-independent cell colony formation assays and a xenograft tumor model to show that GP73-specific siRNAs inhibit HCC proliferation in HepG2, SMMC-7721, and Huh7 cell lines and in vivo. Following GP73 silencing, levels of p-Rb, a factor related to metastasis, were reduced, but cell cycle progression was unaffected. Our results suggest that GP73 silencing may not directly suppress proliferation, but may instead inhibit cell motility. Results from proliferation assays suggest GP73 reduces expression of epithelial mesenchymal transition (EMT)-related factors and promotes cell motility, while transwell migration and invasion assays indicated a possible role in metastasis. Immunofluorescence co-localization microscopy and immunoblotting showed that GP73 decreases expression of N-cadherin and E-cadherin, two key factors in EMT, which may in turn decrease intracellular adhesive forces and promote cell motility. This study confirmed that GP73 expression leads to increased expression of EMT-related proteins and that GP73 silencing reduces HCC cell migration in vitro. These findings suggest that GP73 silencing through siRNA delivery may provide a novel low-toxicity therapy for the inhibition of tumor proliferation and metastasis. PMID:26870893

  13. WNT3 Inhibits Cerebellar Granule Neuron Progenitor Proliferation and Medulloblastoma Formation via MAPK Activation

    PubMed Central

    Ayrault, Olivier; Kim, Jee Hae; Zhu, Xiaodong; Murphy, David A.; Van Aelst, Linda; Roussel, Martine F.; Hatten, Mary E.

    2013-01-01

    During normal cerebellar development, the remarkable expansion of granule cell progenitors (GCPs) generates a population of granule neurons that outnumbers the total neuronal population of the cerebral cortex, and provides a model for identifying signaling pathways that may be defective in medulloblastoma. While many studies focus on identifying pathways that promote growth of GCPs, a critical unanswered question concerns the identification of signaling pathways that block mitogenic stimulation and induce early steps in differentiation. Here we identify WNT3 as a novel suppressor of GCP proliferation during cerebellar development and an inhibitor of medulloblastoma growth in mice. WNT3, produced in early postnatal cerebellum, inhibits GCP proliferation by down-regulating pro-proliferative target genes of the mitogen Sonic Hedgehog (SHH) and the bHLH transcription factor Atoh1. WNT3 suppresses GCP growth through a non-canonical Wnt signaling pathway, activating prototypic mitogen-activated protein kinases (MAPKs), the Ras-dependent extracellular-signal-regulated kinases 1/2 (ERK1/2) and ERK5, instead of the classical β-catenin pathway. Inhibition of MAPK activity using a MAPK kinase (MEK) inhibitor reversed the inhibitory effect of WNT3 on GCP proliferation. Importantly, WNT3 inhibits proliferation of medulloblastoma tumor growth in mouse models by a similar mechanism. Thus, the present study suggests a novel role for WNT3 as a regulator of neurogenesis and repressor of neural tumors. PMID:24303070

  14. Inhibition of fibroblast proliferation in cardiac myocyte cultures by surface microtopography.

    PubMed

    Boateng, Samuel Y; Hartman, Thomas J; Ahluwalia, Neil; Vidula, Himabindu; Desai, Tejal A; Russell, Brenda

    2003-07-01

    Cardiac myocyte cultures usually require pharmacological intervention to prevent overproliferation of contaminating nonmyocytes. Our aim is to prevent excessive fibroblast cell proliferation without the use of cytostatins. We have produced a silicone surface with 10-microm vertical projections that we term "pegs," to which over 80% of rat neonatal cardiac fibroblasts attach within 48 h after plating. There was a 50% decrease in cell proliferation by 5 days of culture compared with flat membranes (P < 0.001) and a concomitant 60% decrease (P < 0.01) in cyclin D1 protein levels, suggesting a G1/S1 cell cycle arrest due to microtopography. Inhibition of Rho kinase with 5 or 20 microM Y-27632 reduced attachment of fibroblasts to the pegs by over 50% (P < 0.001), suggesting that this signaling pathway plays an important role in the process. Using mobile and immobile 10-microm polystyrene spheres, we show that reactive forces are important for inhibiting fibroblast cell proliferation, because mobile spheres failed to reduce cell proliferation. In primary myocyte cultures, pegs also inhibit fibroblast proliferation in the absence of cytostatins. The ratio of aminopropeptide of collagen protein from fibroblasts to myosin from myocytes was significantly reduced in cultures from pegged surfaces (P < 0.01), suggesting an increase in the proportion of myocytes on the pegged surfaces. Connexin43 protein expression was also increased, suggesting improved myocyte-myocyte interaction in the presence of pegs. We conclude that this microtextured culture system is useful for preventing proliferation of fibroblasts in myocyte cultures and may ultimately be useful for tissue engineering applications in vivo.

  15. Quercetin inhibits the migration and proliferation of astrocytes in wound healing.

    PubMed

    Yuan, Zhaohu; Yao, Fang; Hu, Ziyou; Sun, Shumei; Wu, Bingyi

    2015-05-06

    A previous study showed that quercetin inhibits astrogliosis in a scratch-wound model, but did not identify the underlying mechanisms. Here, we show that quercetin exerts no effect on apoptosis or the viability of astrocytes, but significantly inhibits their proliferation, arresting them in the G1 phase and decreasing the percentage of cells in the S and G2 phase. In addition, we found that quercetin significantly decreased the phosphorylation of ERK1/2 and FAK, a downstream ERK signaling protein. Inhibition of this pathway with U0126, an inhibitor of MAP kinase, retarded wound closure, whereas sustained p-ERK1/2 activation, induced by vanadate, restored astrocyte migration. Our findings thus indicate that quercetin inhibits healing in the scratch-wound model of primary astrocytes in two ways: blockade of the G1 to S phase cell cycle transition and inhibition of the ERK/FAK signaling pathway, which may contribute toward decreasing astroglial scar formation in vivo.

  16. Vasoactive Intestinal Peptide Inhibits Human Small-Cell Lung Cancer Proliferation in vitro and in vivo

    NASA Astrophysics Data System (ADS)

    Maruno, Kaname; Absood, Afaf; Said, Sami I.

    1998-11-01

    Small-cell lung carcinoma (SCLC) is an aggressive, rapidly growing and metastasizing, and highly fatal neoplasm. We report that vasoactive intestinal peptide inhibits the proliferation of SCLC cells in culture and dramatically suppresses the growth of SCLC tumor-cell implants in athymic nude mice. In both cases, the inhibition was mediated apparently by a cAMP-dependent mechanism, because the inhibition was enhanced by the adenylate cyclase activator forskolin and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine in proportion to increases in intracellular cAMP levels, and the inhibition was abolished by selective inhibition of cAMP-dependent protein kinase. If confirmed in clinical trials, this antiproliferative action of vasoactive intestinal peptide may offer a new and promising means of suppressing SCLC in human subjects, without the toxic side effects of chemotherapeutic agents.

  17. Mammalian Tead proteins regulate cell proliferation and contact inhibition as transcriptional mediators of Hippo signaling.

    PubMed

    Ota, Mitsunori; Sasaki, Hiroshi

    2008-12-01

    Regulation of organ size is important for development and tissue homeostasis. In Drosophila, Hippo signaling controls organ size by regulating the activity of a TEAD transcription factor, Scalloped, through modulation of its co-activator protein Yki. Here, we show that mouse Tead proteins regulate cell proliferation by mediating Hippo signaling. In NIH3T3 cells, cell density and Hippo signaling regulated the activity of endogenous Tead proteins by modulating nuclear localization of a Yki homolog, Yap1, and the resulting change in Tead activity altered cell proliferation. Tead2-VP16 mimicked Yap1 overexpression, including increased cell proliferation, reduced cell death, promotion of EMT, lack of cell contact inhibition and promotion of tumor formation. Growth-promoting activities of various Yap1 mutants correlated with their Tead-co-activator activities. Tead2-VP16 and Yap1 regulated largely overlapping sets of genes. However, only a few of the Tead/Yap1-regulated genes in NIH3T3 cells were affected in Tead1(-/-);Tead2(-/-) or Yap1(-/-) embryos. Most of the previously identified Yap1-regulated genes were not affected in NIH3T3 cells or mutant mice. In embryos, levels of nuclear Yap1 and Tead1 varied depending on cell type. Strong nuclear accumulation of Yap1 and Tead1 were seen in myocardium, correlating with requirements of Tead1 for proliferation. However, their distribution did not always correlate with proliferation. Taken together, mammalian Tead proteins regulate cell proliferation and contact inhibition as a transcriptional mediator of Hippo signaling, but the mechanisms by which Tead/Yap1 regulate cell proliferation differ depending on the cell type, and Tead, Yap1 and Hippo signaling may play multiple roles in mouse embryos.

  18. Iptakalim inhibits PDGF-BB-induced human airway smooth muscle cells proliferation and migration.

    PubMed

    Liu, Wenrui; Kong, Hui; Zeng, Xiaoning; Wang, Jingjing; Wang, Zailiang; Yan, Xiaopei; Wang, Yanli; Xie, Weiping; Wang, Hong

    2015-08-15

    Chronic airway diseases are characterized by airway remodeling which is attributed partly to the proliferation and migration of airway smooth muscle cells (ASMCs). ATP-sensitive potassium (KATP) channels have been identified in ASMCs. Mount evidence has suggested that KATP channel openers can reduce airway hyperresponsiveness and alleviate airway remodeling. Opening K(+) channels triggers K(+) efflux, which leading to membrane hyperpolarization, preventing Ca(2+)entry through closing voltage-operated Ca(2+) channels. Intracellular Ca(2+) is the most important regulator of muscle contraction, cell proliferation and migration. K(+) efflux decreases Ca(2+) influx, which consequently influences ASMCs proliferation and migration. As a KATP channel opener, iptakalim (Ipt) has been reported to restrain the proliferation of pulmonary arterial smooth muscle cells (PASMCs) involved in vascular remodeling, while little is known about its impact on ASMCs. The present study was designed to investigate the effects of Ipt on human ASMCs and the mechanisms underlying. Results obtained from cell counting kit-8 (CCK-8), flow cytometry and 5-ethynyl-2'-deoxyuridine (EdU) incorporation showed that Ipt significantly inhibited platelet-derived growth factor (PDGF)-BB-induced ASMCs proliferation. ASMCs migration induced by PDGF-BB was also suppressed by Ipt in transwell migration and scratch assay. Besides, the phosphorylation of Ca(2+)/calmodulin-dependent kinase II (CaMKII), extracellular regulated protein kinases 1/2 (ERK1/2), protein kinase B (Akt), and cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) were as well alleviated by Ipt administration. Furthermore, we found that the inhibition of Ipt on the PDGF-BB-induced proliferation and migration in human ASMCs was blocked by glibenclamide (Gli), a selective KATP channel antagonist. These findings provide a strong evidence to support that Ipt antagonize the proliferating and migrating effects of PDGF-BB on

  19. Cdh1 inhibits reactive astrocyte proliferation after oxygen-glucose deprivation and reperfusion.

    PubMed

    Qiu, Jin; Zhang, Chuanhan; Lv, Youyou; Zhang, Yue; Zhu, Chang; Wang, Xueren; Yao, Wenlong

    2013-08-01

    Anaphase-promoting complex (APC) and its co-activator Cdh1 are required for cell cycle regulation in proliferating cells. Recent studies have defined diverse functions of APC-Cdh1 in nervous system development and injury. Our previous studies have demonstrated the activity of APC-Cdh1 is down-regulated in hippocampus after global cerebral ischemia. But the detailed mechanisms of APC-Cdh1 in ischemic nervous injury are unclear. It is known that astrocyte proliferation is an important pathophysiological process following cerebral ischemia. However, the role of APC-Cdh1 in reactive astrocyte proliferation is not determined yet. In the present study, we cultured primary cerebral astrocytes and set up in vitro oxygen-glucose deprivation and reperfusion model. Our results showed that the expression of Cdh1 was decreased while Skp2 (the downstream substrate of APC-Cdh1) was increased in astrocytes after 1h oxygen-glucose deprivation and reperfusion. The down-regulation of APC-Cdh1 was coupled with reactive astrocyte proliferation. By constructing Cdh1 expressing lentivirus system, we also found exogenous Cdh1 can down-regulate Skp2 and inhibit reactive astrocyte proliferation induced by oxygen-glucose deprivation and reperfusion. Moreover, Western blot showed that other downstream proteins of APC-Cdh1, PFK-1 and SnoN, were decreased in the inhibition of reactive astrocyte proliferation with Cdh1 expressing lentivirus treatment. These results suggest that Cdh1 plays an important role in the regulation of reactive astrocyte proliferation induced by oxygen-glucose deprivation and reperfusion.

  20. TGF-{beta}2 inhibits AKT activation and FGF-2-induced corneal endothelial cell proliferation

    SciTech Connect

    Lu Jiawei; Lu Zhenyu; Reinach, Peter

    2006-11-01

    The corneal endothelial cells form a boundary layer between anterior chamber and cornea. This single cell layer is important to maintain cornea transparency by eliciting net fluid transport into the anterior chamber. Injuries of the corneal endothelial layer in humans lead to corneal swelling and translucence. This hindrance is thought to be due to limited proliferative capacity of the endothelial layer. Fibroblast growth factor 2 (FGF-2) and transforming growth factor-beta 2 (TGF-{beta}2) are both found in aqueous humor, and these two cytokines promote and inhibit cell growth, respectively. The intracellular signaling mechanisms by which TGF-{beta}2 suppresses the mitogenic response to FGF-2, however, remain unclear. We have addressed this question by investigating potential crosstalk between FGF-2-induced and TGF-{beta}2-regulated intracellular signaling events in cultured bovine corneal endothelial (BCE) cells. We found that TGF-{beta}2 and FGF-2 oppositely affect BCE cell proliferation and TGF-{beta}2 can override the stimulating effects of FGF-2 by increasing COX-2 expression in these cells. Consistent with these findings, overexpression of COX-2 significantly reduced FGF-2-induced cell proliferation whereas a COX-2 specific inhibitor NS398 reversed the effect of TGF-{beta}2 on FGF-2-induced cell proliferation. The COX-2 product prostaglandin E2 (PGE-2) blocks FGF-2-induced cell proliferation. Whereas FGF-2 stimulates cell proliferation by activating the AKT pathway, TGF-{beta}2 and PGE-2 both inhibit this pathway. In accordance with the effect of PGE-2, cAMP also inhibits FGF-2-induced AKT activation. These findings suggest that the mitogenic response to FGF-2 in vivo in the corneal endothelial layer may be inhibited by TGF-{beta}2-induced suppression of the PI3-kinase/AKT signaling pathway.

  1. Arbutin inhibits TCCSUP human bladder cancer cell proliferation via up-regulation of p21.

    PubMed

    Li, Hailan; Jeong, Yun-Mi; Kim, Su Yeon; Kim, Myo-Kyoung; Kim, Dong-Seok

    2011-04-01

    Arbutin is a glycosylated hydroquinone extracted from the bearberry plant (Arctostaphylos species). In the present study, we determined the effects of arbutin on TCCSUP human bladder carcinoma cell proliferation. Arbutin did not exhibit any cytotoxic effects in TCCSUP cells at concentrations of < 500 microg/ml. To determine the effects of arbutin on cell proliferation, TCCSUP cells were treated with arbutin at various concentrations, and the cell proliferation was measured using the MTT assay. Arbutin significantly decreased TCCSUP cell proliferation in a concentration- and time-dependent manner. Furthermore, cell cycle analysis revealed that arbutin strongly disrupted the cell cycle in a time-dependent manner. Western blot analysis demonstrated that arbutin led to the inactivation of extracellular signal-regulated kinase (ERK), which is known to critically regulate cell proliferation. In addition, arbutin markedly increased the expression of p21WAF1/CIP1 (p21), which is known to be highly involved in cell cycle regulation. Therefore, this study suggests that arbutin inhibits TCCSUP cell proliferation via ERK inactivation and p21 up-regulation.

  2. Passive leg movement enhances interstitial VEGF protein, endothelial cell proliferation, and eNOS mRNA content in human skeletal muscle.

    PubMed

    Hellsten, Ylva; Rufener, Nora; Nielsen, Jens J; Høier, Birgitte; Krustrup, Peter; Bangsbo, Jens

    2008-03-01

    The present study used passive limb movement as an experimental model to study the effect of increased blood flow and passive stretch, without enhanced metabolic demand, in young healthy male subjects. The model used was 90 min of passive movement of the leg leading to a 2.8-fold increase (P < 0.05) in blood flow without a significant enhancement in oxygen uptake. Muscle interstitial fluid was sampled with microdialysis technique and analyzed for vascular endothelial growth factor (VEGF) protein and for the effect on endothelial cell proliferation. Biopsies obtained from the musculus vastus lateralis were analyzed for mRNA content of VEGF, endothelial nitric oxide synthase (eNOS), and matrix metalloproteinase-2 (MMP-2). The passive leg movement caused an increase (P < 0.05) in interstitial VEGF protein concentration above rest (73 +/- 21 vs. 344 +/- 83 pg/ml). Addition of muscle dialysate to cultured endothelial cells revealed that dialysate obtained during leg movement induced a 3.2-fold higher proliferation rate (P < 0.05) than dialysate obtained at rest. Passive movement also enhanced (P < 0.05) the eNOS mRNA level fourfold above resting levels. VEGF mRNA and MMP-2 mRNA levels were unaffected. The results show that a session of passive leg movement, elevating blood flow and causing passive stretch, augments the interstitial concentrations of VEGF, the proliferative effect of interstitial fluid, and eNOS mRNA content in muscle tissue. We propose that enhanced blood flow and passive stretch are positive physiological stimulators of factors associated with capillary growth in human muscle.

  3. Rapamycin Inhibits Proliferation of Hemangioma Endothelial Cells by Reducing HIF-1-Dependent Expression of VEGF

    PubMed Central

    Medici, Damian; Olsen, Bjorn R.

    2012-01-01

    Hemangiomas are tumors formed by hyper-proliferation of vascular endothelial cells. This is caused by elevated vascular endothelial growth factor (VEGF) signaling through VEGF receptor 2 (VEGFR2). Here we show that elevated VEGF levels produced by hemangioma endothelial cells are reduced by the mTOR inhibitor rapamycin. mTOR activates p70S6K, which controls translation of mRNA to generate proteins such as hypoxia inducible factor-1 (HIF-1). VEGF is a known HIF-1 target gene, and our data show that VEGF levels in hemangioma endothelial cells are reduced by HIF-1α siRNA. Over-expression of HIF-1α increases VEGF levels and endothelial cell proliferation. Furthermore, both rapamycin and HIF-1α siRNA reduce proliferation of hemangioma endothelial cells. These data suggest that mTOR and HIF-1 contribute to hemangioma endothelial cell proliferation by stimulating an autocrine loop of VEGF signaling. Furthermore, mTOR and HIF-1 may be therapeutic targets for the treatment of hemangiomas. PMID:22900063

  4. Rebamipide-induced downregulation of phospholipase D inhibits inflammation and proliferation in gastric cancer cells

    PubMed Central

    Kang, Dong Woo; Min, Gyesik; Park, Do Yoon; Hong, Ki Whan

    2010-01-01

    Rebamipide a gastroprotective drug, is clinically used for the treatment of gastric ulcers and gastritis, but its actions on gastric cancer are not clearly understood. Phospholipase D (PLD) is overexpressed in various types of cancer tissues and has been implicated as a critical factor in inflammation and carcinogenesis. However, whether rebamipide is involved in the regulation of PLD in gastric cancer cells is not known. In this study, we showed that rebamipide significantly suppressed the expression of both PLD1 and PLD2 at a transcriptional level in AGS and MKN-1 gastric cancer cells. Downregulation of PLD expression by rebamipide inhibited its enzymatic activity. In addition, rebamipide inhibited the transactivation of nuclear factor kappa B (NFκB), which increased PLD1 expression. Rebamipide or PLD knockdown significantly suppressed the expression of genes involved in inflammation and proliferation and inhibited the proliferation of gastric cancer cells. In conclusion, rebamipide-induced downregulation of PLD may contribute to the inhibition of inflammation and proliferation in gastric cancer. PMID:20625243

  5. Implication of unfolded protein response in resveratrol-induced inhibition of K562 cell proliferation

    SciTech Connect

    Liu, Bao-Qin; Gao, Yan-Yan; Niu, Xiao-Fang; Xie, Ji-Sheng; Meng, Xin; Guan, Yifu; Wang, Hua-Qin

    2010-01-01

    Resveratrol (RES), a natural plant polyphenol, is an effective inducer of cell cycle arrest and apoptosis in a variety of carcinoma cell types. In addition, RES has been reported to inhibit tumorigenesis in several animal models suggesting that it functions as a chemopreventive and anti-tumor agent in vivo. The chemopreventive and chemotherapeutic properties associated with resveratrol offer promise for the design of new chemotherapeutic agents. However, the mechanisms by which RES mediates its effects are not yet fully understood. In this study, we showed that RES caused cell cycle arrest and proliferation inhibition via induction of unfolded protein response (UPR) in human leukemia K562 cell line. Treatment of K562 cells with RES induced a number of signature UPR markers, including transcriptional induction of GRP78 and CHOP, phosphorylation of eukaryotic initiation factor 2{alpha} (eIF2{alpha}), ER stress-specific XBP-1 splicing, suggesting the induction of UPR by RES. RES inhibited proliferation of K562 in a concentration-dependent manner. Flow cytometric analyses revealed that K562 cells were arrested in G1 phase upon RES treatment. Salubrinal, an eIF2{alpha} inhibitor, or overexpression of dominant negative mutants of PERK or eIF2{alpha}, effectively restored RES-induced cell cycle arrest, underscoring the important role of PERK/eIF2{alpha} branch of UPR in RES-induced inhibition of cell proliferation.

  6. Rapamycin inhibits smooth muscle cell proliferation and obstructive arteriopathy attributable to elastin deficiency.

    PubMed

    Li, Wei; Li, Qingle; Qin, Lingfeng; Ali, Rahmat; Qyang, Yibing; Tassabehji, May; Pober, Barbara R; Sessa, William C; Giordano, Frank J; Tellides, George

    2013-05-01

    Patients with elastin deficiency attributable to gene mutation (supravalvular aortic stenosis) or chromosomal microdeletion (Williams syndrome) are characterized by obstructive arteriopathy resulting from excessive smooth muscle cell (SMC) proliferation, mural expansion, and inadequate vessel size. We investigated whether rapamycin, an inhibitor of the cell growth regulator mammalian target of rapamycin (mTOR) and effective against other SMC proliferative disorders, is of therapeutic benefit in experimental models of elastin deficiency. As previously reported, Eln(-/-) mice demonstrated SMC hyperplasia and severe stenosis of the aorta, whereas Eln(+/-) mice exhibited a smaller diameter aorta with more numerous but thinner elastic lamellae. Increased mTOR signaling was detected in elastin-deficient aortas of newborn pups that was inhibited by maternal administration of rapamycin. mTOR inhibition reduced SMC proliferation and aortic obstruction in Eln(-/-) pups and prevented medial hyperlamellation in Eln(+/-) weanlings without compromising aortic size. However, rapamycin did not prolong the survival of Eln(-/-) pups, and it retarded the somatic growth of juvenile Eln(+/-) and Eln(+/+) mice. In cell cultures, rapamycin inhibited prolonged mTOR activation and enhanced proliferation of SMC derived from patients with supravalvular aortic stenosis and with Williams syndrome. mTOR inhibition may represent a pharmacological strategy to treat diffuse arteriopathy resulting from elastin deficiency.

  7. Induction of sister chromatid exchanges and inhibition of cellular proliferation in vitro. I. Caffeine

    SciTech Connect

    Guglielmi, G.E.; Vogt, T.F.; Tice, R.R.

    1982-01-01

    While many agents have been examined for their ability to induce SCE's, complete dose-response information has often been lacking. We have reexamined the ability of one such compound - caffeine - to induce SCEs and also to inhibit cellular proliferation in human peripheral lymphocytes in vitro. An acute exposure to caffeine prior to the DNA synthetic period did not affect either SCE frequency or the rate of cellular proliferation. Chronic exposure to caffeine throughout the culture period lead to both a dose-dependent increase in SCEs (SCE/sub d/ or doubling dose = 2.4 mM; SCE/sub 10/ or the dose capable of inducing 10 SCE = 1.4 mM) and a dose-dependent inhibition of cellular proliferation (IC/sub 50/ or the 50% inhibition concentration = 2.6 mM). The relative proportion of first generation metaphase cells, an assessment of proliferative inhibiton, increased linearly with increasing caffeine concentrations. However, SCE frequency increased nonlinearly over the same range of caffeine concentrations. Examination of the ratio of nonsymmetrical to symmetrical SCEs in third generation metaphase cells indicated that caffeine induced SCEs in equal frequency in each of three successive generations. The dependency of SCE induction and cellular proliferative inhibition on caffeine's presence during the DNA synthetic period suggests that caffeine may act as an antimetabolite in normal human cells.

  8. Lovastatin inhibits human B lymphoma cell proliferation by reducing intracellular ROS and TRPC6 expression

    PubMed Central

    Song, Xiang; Liu, Bing-Chen; Lu, Xiao-Yu; Yang, Li-Li; Zhai, Yu-Jia; Eaton, Amity F.; Thai, Tiffany L.; Eaton, Douglas C.; Ma, He-Ping; Shen, Bao-Zhong

    2014-01-01

    Clinical evidence suggests that statins reduce cancer incidence and mortality. However, there is lack of in vitro data to show the mechanism by which statins can reduce the malignancies of cancer cells. We used a human B lymphoma Daudi cells as a model and found that lovastatin inhibited, whereas exogenous cholesterol (Cho) stimulated, proliferation cell cycle progression in control Daudi cells, but not in the cells when transient receptor potential canonical 6 (TRPC6) channel was knocked down. Lovastatin decreased, whereas Cho increased, the levels of intracellular reactive oxygen species (ROS) respectively by decreasing or increasing the expression of p47-phox and gp91-phox (NOX2). Reducing intracellular ROS with either a mimetic superoxide dismutase (TEMPOL) or a NADPH oxidase inhibitor (apocynin) inhibited cell proliferation, particularly in Cho-treated cells. The effects of TEMPOL or apocynin were mimicked by inhibition of TRPC6 with SKF-96365. Lovastatin decreased TRPC6 expression and activity via a Cho-dependent mechanism, whereas Cho increased TRPC6 expression and activity via an ROS-dependent mechanism. Consistent with the fact that TRPC6 is a Ca2+-permeable channel, lovastatin decreased, but Cho increased, intracellular Ca2+ also via ROS. These data suggest that lovastatin inhibits malignant B cell proliferation by reducing membrane Cho, intracellular ROS, TRPC6 expression and activity, and intracellular Ca2+. PMID:24518247

  9. Lovastatin inhibits human B lymphoma cell proliferation by reducing intracellular ROS and TRPC6 expression.

    PubMed

    Song, Xiang; Liu, Bing-Chen; Lu, Xiao-Yu; Yang, Li-Li; Zhai, Yu-Jia; Eaton, Amity F; Thai, Tiffany L; Eaton, Douglas C; Ma, He-Ping; Shen, Bao-Zhong

    2014-05-01

    Clinical evidence suggests that statins reduce cancer incidence and mortality. However, there is lack of in vitro data to show the mechanism by which statins can reduce the malignancies of cancer cells. We used a human B lymphoma Daudi cells as a model and found that lovastatin inhibited, whereas exogenous cholesterol (Cho) stimulated, proliferation cell cycle progression in control Daudi cells, but not in the cells when transient receptor potential canonical 6 (TRPC6) channel was knocked down. Lovastatin decreased, whereas Cho increased, the levels of intracellular reactive oxygen species (ROS) respectively by decreasing or increasing the expression of p47-phox and gp91-phox (NOX2). Reducing intracellular ROS with either a mimetic superoxide dismutase (TEMPOL) or an NADPH oxidase inhibitor (apocynin) inhibited cell proliferation, particularly in Cho-treated cells. The effects of TEMPOL or apocynin were mimicked by inhibition of TRPC6 with SKF-96365. Lovastatin decreased TRPC6 expression and activity via a Cho-dependent mechanism, whereas Cho increased TRPC6 expression and activity via an ROS-dependent mechanism. Consistent with the fact that TRPC6 is a Ca(2+)-permeable channel, lovastatin decreased, but Cho increased, intracellular Ca(2+) also via ROS. These data suggest that lovastatin inhibits malignant B cell proliferation by reducing membrane Cho, intracellular ROS, TRPC6 expression and activity, and intracellular Ca(2+). Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Loureirin B inhibits fibroblast proliferation and extracellular matrix deposition in hypertrophic scar via TGF-β/Smad pathway.

    PubMed

    Bai, Xiaozhi; He, Ting; Liu, Jiaqi; Wang, Yunchuan; Fan, Lei; Tao, Ke; Shi, Jihong; Tang, Chaowu; Su, Linlin; Hu, Dahai

    2015-05-01

    The ethanolic extract of Resina Draconis (RDEE) has been reported beneficial to normal wound healing yielding more regularly arranged collagen fibres. Loureirin B, a major component in RDEE, has been supposed to be effective on the prevention and treatment of pathological scars. To investigate the therapeutic effects of loureirin B on hypertrophic scar (HS), fibroblasts from human HS and normal skin (NS) were isolated. Results showed that loureirin B dose-dependently downregulated both mRNA and protein levels of type I collagen (ColI), type III collagen (ColIII) and α-smooth muscle actin (α-SMA) in HS fibroblasts. Loureirin B also suppressed fibroblast proliferative activity and redistributed cell cycle, but did not affect cell apoptosis. In vivo rabbit ear scar model, loureirin B significantly improved the arrangement and deposition of collagen fibres, decreased protein levels of ColI, ColIII and α-SMA and suppressed myofibroblast differentiation and scar proliferative activity. In NS fibroblasts, loureirin B effectively inhibited TGF-β1-induced upregulation of ColI, ColIII and α-SMA levels, myofibroblast differentiation and the activation of Smad2 and Smad3. Loureirin B also affected mRNA levels of major MMPs and TIMPs in TGF-β1-stimulated fibroblasts. Taken together, this study demonstrates that loureirin B could downregulate the expression of fibrosis-related molecules by regulating MMPs and TIMPs levels, inhibit scar fibroblast proliferation and suppress TGF-β1-induced fibrosis, during which TGF-β1/Smad2/3 pathway is likely involved. These findings suggest that loureirin B is a potential therapeutic compound for HS treatment.

  11. Gambogic acid potentiates the chemosensitivity of colorectal cancer cells to 5-fluorouracil by inhibiting proliferation and inducing apoptosis

    PubMed Central

    Wei, Jianchang; Yang, Ping; Li, Wanglin; He, Feng; Zeng, Shanqi; Zhang, Tong; Zhong, Junbin; Huang, Di; Chen, Zhuanpeng; Wang, Chengxing; Chen, Huacui; Hu, He; Cao, Jie

    2017-01-01

    Chemotherapy using 5-fluorouracil (5-FU) for colorectal cancer (CRC) has low specificity and response rates, leading to severe side effects. Gambogic acid (GA), a traditional Chinese medicine, has multi-targeted anticancer effects, including growth inhibition and apoptosis induction. However, it is unclear whether a combination of 5-FU and GA has synergistic anticancer effects in CRC cells. In this study, SW480 and HCT116 human CRC cells and human intestinal epithelial cells (IECs) were treated with different concentrations of 5-FU, GA or 5-FU+GA. A Cell Counting kit-8 assay was conducted to quantify cell proliferation. The combination index (CI) was calculated and the median-effect principle was applied to analyze the interaction between 5-FU and GA. Flow cytometry was used to determine the percentage of cells undergoing apoptosis. Reverse transcription-quantitative polymerase chain reaction and western blotting were applied to measure P53, survivin and thymidylate synthase (TS) mRNA and protein levels. It was found that 5-FU+GA more pronouncedly inhibited cell growth and induced apoptosis, compared with either monotherapy. CI values <1 indicated the synergistic effects of the drugs. 5-FU+GA further decreased P53, survivin and TS mRNA and protein levels in the two CRC cell lines compared with single drugs, whereas increased P53 protein levels were observed in HCT116 cells. Moreover, 5-FU+GA did not increase cytotoxicity to IECs. These results demonstrate that GA enhances the anticancer effects of 5-FU on CRC cells. Combined treatment with 5-FU and GA is effective and safe for CRC cells, and may become a promising chemotherapy treatment. PMID:28352348

  12. PKI-587 and sorafenib alone and in combination on inhibition of liver cancer stem cell proliferation

    PubMed Central

    Gedaly, Roberto; Galuppo, Roberto; Musgrave, Yolanda; Angulo, Paul; Hundley, Jonathan; Shah, Malay; Daily, Michael F.; Chen, Changguo; Cohen, Donald A.; Spear, Brett T.; Evers, B. Mark

    2015-01-01

    Background Deregulated Ras/Raf/mitogen-activated protein kinase and PI3 K/AKT/mTOR signaling pathways are significant in hepatocellular carcinoma proliferation (HCC). In this study we evaluated differences in the antiproliferative effect of dual PI3 K/Akt/mTOR and Ras/Raf/mitogen-activated protein kinase inhibition of non liver cancer stem cell lines (PLC and HuH7) and liver cancer stem cell (LCSC) lines (CD133, CD44, CD24, and aldehyde dehydrogenase 1-positive cells). Materials and methods Flow cytometry was performed on the resulting tumors to identify the LCSC markers CD133, CD44, CD24, and aldehyde dehydrogenase 1. Methylthiazol tetrazolium assay was used to assess cellular proliferation. Finally, a Western blot assay was used to evaluate for inhibition of specific enzymes in these two signaling pathways. Results Using flow cytometry, we found that LCSC contain 64.4% CD133 + cells, 83.2% CD44 + cells, and 96.4% CD24 + cells. PKI-587 and sorafenib caused inhibiton of LCSC and HCC cell proliferation. PLC cells were more sensitive to PKI-587 than LCSC or Huh7 (P < 0.001). Interestingly, HuH7 cells were more sensitive to sorafenib than LCSC or PLC cells. Additionally, combination therapy with PKI-587 and sorafenib caused significantly more inhibition than monotherapy in HuH7, PLC, and LCSC. Using the methylthiazol tetrazolium assay, we found that the LCSC proliferation was inhibited with sorafenib monotherapy 39% at 5 μM (P < 0.001; n = 12) and 67% by PKI-587 at 0.1 μM (P = 0.002, n = 12) compared with control. The combination of PKI-587 and sorafenib, however, synergistically inhibited LCSC proliferation by 86% (P = 0.002; n = 12). Conclusions LCSC (CD133+, CD44+, CD24+) were able to develop very aggressive tumors with low cell concentrations at 4 to 6 wk. Cells CD133+, CD44+, CD24+ demonstrated at least moderate resistance to therapy in vitro. The combination of PKI-587 and sorafenib was better than either drug alone at inhibiting of LCSC and on HCC cell

  13. Statins inhibit proliferation and cytotoxicity of a human leukemic natural killer cell line

    PubMed Central

    2013-01-01

    Background Natural killer cells comprise the body’s first line of defense against virus-infected cells. As is true of all lymphocytes, natural killer cell malignancies can develop, however natural killer cell leukemias can be very difficult to treat due to their intrinsic resistance to chemotherapeutic agents. With the recent understanding that statin drugs may have anti-cancer properties, our investigations have focused on the ability of statins to inhibit the growth and cytotoxicity of the YT-INDY natural killer cell leukemia cell line. Results Our findings indicate that several statin compounds can inhibit YT-INDY proliferation disrupt cell cycle progression and abrogate natural killer cell cytotoxicity. Since natural killer cell leukemia cytotoxicity may play a role in the pulmonary damage seen in these patients, this is an important finding. Cytotoxicity, proliferation and cell cycle progression could be restored by the addition of mevalonate, signifying that the statin effects are brought about through HMG CoA reductase inhibition. The mevalonate pathway intermediate geranylgeranyl pyrophosphate, but not other intermediates in the mevalonate pathway, partially reversed statin-induced inhibition of YT-INDY proliferation and cytotoxicity. These results suggest that blockage of products made in the latter part of the mevalonate pathway may account for the observed inhibitory effects on YT-INDY proliferation and cytotoxicity. However, geranylgeranyl pyrophosphate could not reverse the statin-induced inhibition of the cell cycle. Conclusions These results suggest that the statin drugs should be investigated as a potential therapeutic strategy for human natural killer cell leukemias possibly in combination with chemotherapeutic agents. PMID:24359683

  14. PinX1 suppresses bladder urothelial carcinoma cell proliferation via the inhibition of telomerase activity and p16/cyclin D1 pathway.

    PubMed

    Liu, Jian-Ye; Qian, Dong; He, Li-Ru; Li, Yong-Hong; Liao, Yi-Ji; Mai, Shi-Juan; Tian, Xiao-Peng; Liu, Yan-Hui; Zhang, Jia-Xing; Kung, Hsiang-Fu; Zeng, Yi-Xin; Zhou, Fang-Jian; Xie, Dan

    2013-11-23

    PIN2/TRF1-interacting telomerase inhibitor1 (PinX1) was recently suggested as a putative tumor suppressor in several types of human cancer, based on its binding to and inhibition of telomerase. Moreover, loss of PinX1 has been detected in many human malignancies. However, the possible involvement of PinX1 and its clinical/prognostic significance in urothelial carcinoma of the bladder (UCB) are unclear. The PinX1 expression profile was examined by quantitative real-time polymerase chain reaction (qRT-PCR), western blotting, and immunohistochemistry (IHC) in UCB tissues and adjacent normal urothelial bladder epithelial tissues. PinX1 was overexpressed and silenced in UCB cell lines to determine its role in tumorigenesis, development of UCB, and the possible mechanism. PinX1 expression in UCB was significantly down-regulated at both mRNA and protein level as compared with that in normal urothelial bladder epithelial tissues. PinX1 levels were inversely correlated with tumor multiplicity, advanced N classification, high proliferation index (Ki-67), and poor survival (P < 0.05). Moreover, overexpression of PinX1 in UCB cells significantly inhibited cell proliferation in vitro and in vivo, whereas silencing PinX1 dramatically enhanced cell proliferation. Overexpression of PinX1 resulted in G1/S phase arrest and cell growth/proliferation inhibition, while silencing PinX1 led to acceleration of G1/S transition, and cell growth/proliferation promotion by inhibiting/enhancing telomerase activity and via the p16/cyclin D1 pathway. These findings suggest that down-regulation of PinX1 play an important role in the tumorigenesis and development of UCB and that the expression of PinX1 as detected by IHC is an independent molecular marker in patients with UCB.

  15. Roles of p38 and JNK protein kinase pathways activated by compound cantharidin capsules containing serum on proliferation inhibition and apoptosis of human gastric cancer cell line

    PubMed Central

    Sun, Yonghao; Zhang, Dejuan; Mao, Mao; Lu, Yangping; Jiao, Ning

    2017-01-01

    The aim of the present study was to investigate the inhibitory effect of compound cantharides capsules (CCCs) on the viability and apoptosis of human gastric cancer cell lines, BGC-823 and SGC-7901, and to detect its regulation of gene expression levels, as well as its inhibition mechanisms. Each cell line was grouped into a control group, CCC serum group, 5-fluorouracil (5-FU) group, combination therapy group (CCC serum + 5-FU) and serum control group. Growth curves were measured and flow cytometry was used to detect cell apoptosis and cell viability. The mRNA expression level of proliferation-related C-MYC and p53 genes were assayed by reverse transcription-quantitative polymerase chain reaction. Protein phosphorylation levels of proliferating cell nuclear antigen, p38 mitogen-activated protein kinase, extracellular signal-related kinase 1/2, c-Jun N-terminal kinase (JNK) and IκB were assayed by western blotting. The combined CCC serum and 5-FU group exhibited a higher inhibition rate in both cell lines and CCC serum therapy demonstrated a similar effect to 5-FU treatment, as demonstrated in the MTT and cell growth assay. Combined therapy significantly decreased the C-MYC mRNA expression levels and increased p53 mRNA expression levels (P<0.05). Combined therapy of 5-FU and CCC was more significant compared with CCC serum or 5-FU only (P<0.05). P38 and JNK-related protein phosphorylation are involved in apoptosis initiated by CCC combined 5-FU therapy. Combined therapy was able to significantly inhibit human gastric cancer cell growth (P<0.05), and advance cell apoptosis compared with CCC serum only. CCC serum resulted in downregulation of the c-Myc gene and upregulation of the p53 gene. p38 and JNK-related protein phosphorylation is involved in the inhibition of cell viability and apoptosis of human gastric cancer cell lines. PMID:28810654

  16. Roles of p38 and JNK protein kinase pathways activated by compound cantharidin capsules containing serum on proliferation inhibition and apoptosis of human gastric cancer cell line.

    PubMed

    Sun, Yonghao; Zhang, Dejuan; Mao, Mao; Lu, Yangping; Jiao, Ning

    2017-08-01

    The aim of the present study was to investigate the inhibitory effect of compound cantharides capsules (CCCs) on the viability and apoptosis of human gastric cancer cell lines, BGC-823 and SGC-7901, and to detect its regulation of gene expression levels, as well as its inhibition mechanisms. Each cell line was grouped into a control group, CCC serum group, 5-fluorouracil (5-FU) group, combination therapy group (CCC serum + 5-FU) and serum control group. Growth curves were measured and flow cytometry was used to detect cell apoptosis and cell viability. The mRNA expression level of proliferation-related C-MYC and p53 genes were assayed by reverse transcription-quantitative polymerase chain reaction. Protein phosphorylation levels of proliferating cell nuclear antigen, p38 mitogen-activated protein kinase, extracellular signal-related kinase 1/2, c-Jun N-terminal kinase (JNK) and IκB were assayed by western blotting. The combined CCC serum and 5-FU group exhibited a higher inhibition rate in both cell lines and CCC serum therapy demonstrated a similar effect to 5-FU treatment, as demonstrated in the MTT and cell growth assay. Combined therapy significantly decreased the C-MYC mRNA expression levels and increased p53 mRNA expression levels (P<0.05). Combined therapy of 5-FU and CCC was more significant compared with CCC serum or 5-FU only (P<0.05). P38 and JNK-related protein phosphorylation are involved in apoptosis initiated by CCC combined 5-FU therapy. Combined therapy was able to significantly inhibit human gastric cancer cell growth (P<0.05), and advance cell apoptosis compared with CCC serum only. CCC serum resulted in downregulation of the c-Myc gene and upregulation of the p53 gene. p38 and JNK-related protein phosphorylation is involved in the inhibition of cell viability and apoptosis of human gastric cancer cell lines.

  17. Inhibition of DYRK1A Stimulates Human β-Cell Proliferation.

    PubMed

    Dirice, Ercument; Walpita, Deepika; Vetere, Amedeo; Meier, Bennett C; Kahraman, Sevim; Hu, Jiang; Dančík, Vlado; Burns, Sean M; Gilbert, Tamara J; Olson, David E; Clemons, Paul A; Kulkarni, Rohit N; Wagner, Bridget K

    2016-06-01

    Restoring functional β-cell mass is an important therapeutic goal for both type 1 and type 2 diabetes (1). While proliferation of existing β-cells is the primary means of β-cell replacement in rodents (2), it is unclear whether a similar principle applies to humans, as human β-cells are remarkably resistant to stimulation of division (3,4). Here, we show that 5-iodotubercidin (5-IT), an annotated adenosine kinase inhibitor previously reported to increase proliferation in rodent and porcine islets (5), strongly and selectively increases human β-cell proliferation in vitro and in vivo. Remarkably, 5-IT also increased glucose-dependent insulin secretion after prolonged treatment. Kinome profiling revealed 5-IT to be a potent and selective inhibitor of the dual-specificity tyrosine phosphorylation-regulated kinase (DYRK) and cell division cycle-like kinase families. Induction of β-cell proliferation by either 5-IT or harmine, another natural product DYRK1A inhibitor, was suppressed by coincubation with the calcineurin inhibitor FK506, suggesting involvement of DYRK1A and nuclear factor of activated T cells signaling. Gene expression profiling in whole islets treated with 5-IT revealed induction of proliferation- and cell cycle-related genes, suggesting that true proliferation is induced by 5-IT. Furthermore, 5-IT promotes β-cell proliferation in human islets grafted under the kidney capsule of NOD-scid IL2Rg(null) mice. These results point to inhibition of DYRK1A as a therapeutic strategy to increase human β-cell proliferation.

  18. 6-Shogaol inhibits breast and colon cancer cell proliferation through activation of peroxisomal proliferator activated receptor γ (PPARγ).

    PubMed

    Tan, Boon Shing; Kang, Owen; Mai, Chun Wai; Tiong, Kai Hung; Khoo, Alan Soo-Beng; Pichika, Mallikarjuna Rao; Bradshaw, Tracey D; Leong, Chee-Onn

    2013-08-09

    6-Shogaol has been shown to possess many antitumor properties including inhibition of cancer cell growth, inhibition of cancer metastasis, induction of apoptosis in cancer cells and induction of cancer cell differentiation. Despite its prominent antitumor effects, the direct molecular target of 6-shogaol has remained elusive. To identify the direct targets of 6-shogaol, a comprehensive antitumor profile of 6-shogaol (NSC752389) was tested in the NCI-60 cell line in an in vitro screen. The results show that 6-shogaol is COMPARE negative suggesting that it functions via a mechanism of action distinct from existing classes of therapeutic agents. Further analysis using microarray gene profiling and Connectivity Map analysis showed that MCF-7 cells treated with 6-shogaol display gene expression signatures characteristic of peroxisome proliferator activated receptor γ (PPARγ) agonists, suggesting that 6-shogaol may activate the PPARγ signaling pathway for its antitumor effects. Indeed, treatment of MCF-7 and HT29 cells with 6-shogaol induced PPARγ transcriptional activity, suppressed NFκB activity, and induced apoptosis in breast and colon cancer cells in a PPARγ-dependent manner. Furthermore, 6-shogaol is capable of binding to PPARγ with a binding affinity comparable to 15-delta prostaglandin J2, a natural ligand for PPARγ. Together, our findings suggest that the antitumor effects of 6-shogaol are mediated through activation of PPARγ and imply that activation of PPARγ might be beneficial for breast and colon cancer treatment. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  19. CCAAT/enhancer-binding protein alpha (C/EBP alpha) inhibits cell proliferation through the p21 (WAF-1/CIP-1/SDI-1) protein.

    PubMed

    Timchenko, N A; Wilde, M; Nakanishi, M; Smith, J R; Darlington, G J

    1996-04-01

    C/EBPalpha has a role in growth arrest and differentiation of mouse preadipocytes. To study the mechanism of C/EBPalpha-induced growth arrest, we developed a cell line, HT1, that contained the human C/EBPalpha gene under Lac repressor control. IPTG-induced C/EBPalpha caused inhibition of cell proliferation and DNA synthesis as measured by colony growth assays, cell counting, and BrdU uptake. A number of proteins that are known to be involved in the regulation of the cell cycle, such as cyclin-dependent kinase (CDK)2 and CDK4, proliferating cell nuclear antigen (PCNA), p53, c-fos, and the CDK inhibitor p16 and p27 were investigated by Western analysis. No change in their expression was observed. However, the p21 (WAF-1/CIP-1/SDI-1) protein was significantly elevated in growth-arrested HT1 cells. Elevation of p21/SDI-1 mRNA (threefold) and activation of the p21/SDI-1 promoter by C/EBPalpha did not account for the 12- to 20-fold increase in p21/SDI-1 protein. Protein synthesis inhibition by cycloheximide (CHX) treatment indicated that the half-life of p21/SDI-1 in dividing HT1 cells was approximately 30 min. However, in C/EBPalpha growth-arrested cells, the level of the p21/SDI-1 did not change for > 80 min after CHX addition. Our studies demonstrate that C/EBPalpha activates p21/SDI-1 by increasing p21/SDI-1 gene expression and by post-translational stabilization of p21/SDI-1 protein. Furthermore, induction of p21/SDI-1 is responsible for the ability of C/EBPalpha to inhibit proliferation because transcription of antisense p21/SDI-1 mRNA eliminated growth inhibition by C/EBPalpha.

  20. Aloe-emodin inhibits proliferation of adult human keratinocytes in vitro.

    PubMed

    Popadic, Dusan; Savic, Emina; Ramic, Zorica; Djordjevic, Vladimir; Trajkovic, Vladimir; Medenica, Ljiljana; Popadic, Svetlana

    2012-01-01

    Aloe-emodin (AE) is a plant-derived hydroxyanthraquinone with several biological activities. It is present in a variety of skin-conditioning agents containing aloe extracts, but its influence on keratinocyte growth was not examined so far. We investigated the influence of AE on human keratinocyte proliferation and apoptosis in vitro. AE significantly inhibited proliferation of cultivated human keratinocytes at 5 μM concentration, as revealed by incorporation of radioactive thymidine. The antiproliferative effect of AE was accompanied with induction of apoptosis, but not necrosis, as demonstrated by flow cytometric analysis and lactate dehydrogenase release assay. Based on the half maximal inhibitory concentration values, we demonstrated that AE may impair proliferation of keratinocytes at concentrations far below the industry standards for commercial products containing aloe extracts. Therefore, further research of AE effects on the human skin and proper labeling of products are necessary for maximizing benefits from aloe extracts and to avoid undesired responses.

  1. Cordyceps bassiana inhibits smooth muscle cell proliferation via the ERK1/2 MAPK signaling pathway.

    PubMed

    Jin, Enze; Han, Seongho; Son, Mina; Kim, Sung-Whan

    2016-01-01

    Cordyceps belongs to a genus of acormycete fungi and is known to exhibit various pharmacological effects. The aim of this study was to investigate the effect of Cordyceps species on the proliferation of vascular smooth muscle cells (VSMC) and their underlying molecular mechanism. A cell proliferation assay showed that Cordyceps bassiana ethanol extract (CBEE) significantly inhibited VSMC proliferation. In addition, neointimal formation was significantly reduced by treatment with CBEE in the carotid artery of balloon-injured rats. We also investigated the effects of CBEE on the extracellular signal-regulated kinase (ERK) signal pathway. Western blot analysis revealed increased ERK 1/2 phosphorylation in VSMCs treated with CBEE. Pretreatment with U0126 completely abrogated CBEE-induced ERK 1/2 phosphorylation. In conclusion, CBEE exhibited anti-proliferative properties that affected VSMCs through the ERK1/2 MAPK signaling pathway. Our data may elucidate the inhibitory mechanism of this natural product.

  2. Somatostatin analogue (octreotide) inhibits bile duct epithelial cell proliferation and fibrosis after extrahepatic biliary obstruction.

    PubMed Central

    Tracy, T. F.; Tector, A. J.; Goerke, M. E.; Kitchen, S.; Lagunoff, D.

    1993-01-01

    Extrahepatic biliary obstruction leads to bile duct epithelial cell proliferation. Somatostatin and its analogue, octreotide, have been shown to inhibit DNA synthesis and proliferation in hepatocytes. We investigated the effect of octreotide on the biliary epithelial cell proliferative responses to biliary obstruction. Male Sprague-Dawley rats underwent common bile duct ligation and subcutaneous injection of either saline or octreotide (6 micrograms/kg) twice daily for 7 days. Morphometric analysis of hepatocytes, bile duct epithelial cells, and periportal connective tissue was performed by computerized point counting. Hepatocyte volume was preserved with octreotide treatment, which also significantly decreased bile duct proliferation and periportal extracellular matrix deposition in response to biliary obstruction compared with saline treated, duct-ligated animals. These results indicate that octreotide prevents the morphological changes that accompany extrahepatic biliary obstruction. Images Figure 1 PMID:8256850

  3. Griffipavixanthone, a dimeric xanthone extracted from edible plants, inhibits tumor metastasis and proliferation via downregulation of the RAF pathway in esophageal cancer.

    PubMed

    Ding, Zhijie; Lao, Yuanzhi; Zhang, Hong; Fu, Wenwei; Zhu, Lunlun; Tan, Hongsheng; Xu, Hongxi

    2016-01-12

    Metastasis causes a large number of deaths among esophageal cancer patients. The activation of RAF family proteins elevates tumor metastasis and proliferation. In screen targeting the RAF protein, we identified that Griffipavixanthone (GPX), a dimeric xanthone isolated from Garcinia esculenta, is a B-RAF and C-RAF inhibitor against esophageal cancer cells. Using wound healing, transwell migration and matrigel invasion assays, we confirmed that GPX significantly inhibited cell migration and invasion. Furthermore, exposure to GPX rendered cell proliferation and induced G2/M cell cycle arrest. Our mechanistic study showed that GPX suppressed cancer metastasis and proliferation through downregulation of RAF-MEK-ERK cascades proteins as well as RAF mRNA levels. In a pulmonary metastasis model, the intraperitoneal injection of GPX significantly suppressed esophageal tumor metastasis and ERK protein level in vivo. In conclusion, our present study suggested that GPX could inhibit tumor migration, invasion and proliferation in vitro and in vivo, which indicated the potential of GPX for preventing and treating esophageal cancer.

  4. Processed Panax ginseng, sun ginseng, inhibits the differentiation and proliferation of 3T3-L1 preadipocytes and fat accumulation in Caenorhabditis elegans.

    PubMed

    Lee, Hyejin; Kim, Jinhee; Park, Jun Yeon; Kang, Ki Sung; Park, Joeng Hill; Hwang, Gwi Seo

    2017-07-01

    Heat-processed ginseng, sun ginseng (SG), has been reported to have improved therapeutic properties compared with raw forms, such as increased antidiabetic, anti-inflammatory, and antihyperglycemic effects. The aim of this study was to investigate the antiobesity effects of SG through the suppression of cell differentiation and proliferation of mouse 3T3-L1 preadipocyte cells and the lipid accumulation in Caenorhabditis elegans. To investigate the effect of SG on adipocyte differentiation, levels of stained intracellular lipid droplets were quantified by measuring the oil red O signal in the lipid extracts of cells on differentiation Day 7. To study the effect of SG on fat accumulation in C. elegans, L4 stage worms were cultured on an Escherichia coli OP50 diet supplemented with 10 μg/mL of SG, followed by Nile red staining. To determine the effect of SG on gene expression of lipid and glucose metabolism-regulation molecules, messenger RNA (mRNA) levels of genes were analyzed by real-time reverse transcription-polymerase chain reaction analysis. In addition, the phosphorylation of Akt was examined by Western blotting. SG suppressed the differentiation of 3T3-L1 cells stimulated by a mixture of 3-isobutyl-1-methylxanthine, dexamethasone, and insulin (MDI), and inhibited the proliferation of adipocytes during differentiation. Treatment of C. elegans with SG showed reductions in lipid accumulation by Nile red staining, thus directly demonstrating an antiobesity effect for SG. Furthermore, SG treatment downregulated mRNA and protein expression levels of peroxisome proliferator-activated receptor subtype γ (PPARγ) and CCAAT/enhancer-binding protein-alpha (C/EBPα) and decreased the mRNA level of sterol regulatory element-binding protein 1c in MDI-treated adipocytes in a dose-dependent manner. In differentiated 3T3-L1 cells, mRNA expression levels of lipid metabolism-regulating factors, such as amplifying mouse fatty acid-binding protein 2, leptin, lipoprotein

  5. Calcitriol inhibits keratinocyte proliferation by upregulating leukocyte elastase inhibitor (serpin B1).

    PubMed

    Hui, Li; Yongxia, Zheng; Zhili, Guo; Jun, Gu

    2014-05-01

    Calcitriol had been proved to be effective for treatment of psoriasis vulgaris. However, the molecular events leading to the normalization of keratinocyte differentiation had not been fully explored. The aim of the study was to evaluate the role of calcitriol and serpin B1 in human keratinocyte cell line (HaCaT) proliferation. Proteins extracted from calcitriol-treated and untreated HaCaT were separated by 2-D differential gel electrophoresis (2DE). Then, the 2DE profiles were analyzed to screen for differentially expressed proteins, which were identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry. The upregulation of serpin B1 was confirmed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot analysis. The effect of serpin B1 on HaCaT proliferation was analyzed by RNA interference experiments, methylthiazoletetrazolium assay and flow cytometry. Reproducible 2-DE profiles of HaCaT were established. The result that serpin B1 was upregulated in the calcitriol-treated group was confirmed by qRT-PCR and western blot analysis. Calcitriol could inhibit proliferation of HaCaT at the concentration of 10(-9) -10(-6 ) mol/L. HaCaT proliferation was promoted when serpin B1 was interfered. The inhibition effect of calcitriol was stopped after serpin B1 was interfered. Serpin B1 was overexpressed in calcitriol-treated HaCaT cells and may play an important role in inhibiting HaCaT proliferation by calcitriol. © 2014 Japanese Dermatological Association.

  6. Fractone-heparan sulfates mediate BMP-7 inhibition of cell proliferation in the adult subventricular zone.

    PubMed

    Douet, Vanessa; Arikawa-Hirasawa, Eri; Mercier, Frederic

    2012-10-24

    Bone morphogenetic protein-7 (BMP-7) is a heparin-binding growth factor that inhibits cell proliferation in the subventricular zone (SVZ) of the lateral ventricle, the primary neurogenic niche in the adult brain. However, the physiological mechanisms regulating the activity of BMP-7 in the SVZ are unknown. Here, we report the inhibitory effect of BMP-7 on cell proliferation through the anterior SVZ after intracerebroventricular injection in the adult mouse. To determine whether the inhibition of cell proliferation induced by BMP-7 is dependant on heparin-binding, heparitinase-1 was intracerebroventricularly injected to N-desulfate heparan sulfate proteoglycans before BMP-7 was injected. Heparatinase-1 drastically reduced the inhibitory effect of BMP-7 on cell proliferation in the SVZ. To determine where BMP-7 binds within the niche, we visualized biotinylated-BMP-7 after intracerebroventricular injection, using streptavidin Texas red on frozen brain sections. BMP-7 binding was seen as puncta in the SVZ at the location of fractones, the particulate specialized extracellular matrix of the SVZ, which have been identified primarily by N-sulfated heparan sulfate immunoreactivity (NS-HS+). BMP binding was also seen in NS-HS+ blood vessels of the SVZ. Injection of heparitinase-1 prior to biotinylated BMP-7 resulted in the absence of signal for biotinylated-BMP-7 in the fractones and blood vessels, indicating that the binding is heparan sulfate dependant. These results indicate that BMP-7 requires heparan sulfates to bind and inhibit cell proliferation in the SVZ neurogenic niche. Heparan sulfates concentrated in fractones and SVZ blood vessels emerge as a functional stem cell niche component involved in growth factor activity.

  7. CaMKK2 Suppresses Muscle Regeneration through the Inhibition of Myoblast Proliferation and Differentiation

    PubMed Central

    Ye, Cheng; Zhang, Duo; Zhao, Lei; Li, Yan; Yao, Xiaohan; Wang, Hui; Zhang, Shengjie; Liu, Wei; Cao, Hongchao; Yu, Shuxian; Wang, Yucheng; Jiang, Jingjing; Wang, Hui; Li, Xihua; Ying, Hao

    2016-01-01

    Skeletal muscle has a major role in locomotion and muscle disorders are associated with poor regenerative efficiency. Therefore, a deeper understanding of muscle regeneration is needed to provide a new insight for new therapies. CaMKK2 plays a role in the calcium/calmodulin-dependent kinase cascade; however, its role in skeletal muscle remains unknown. Here, we found that CaMKK2 expression levels were altered under physiological and pathological conditions including postnatal myogensis, freeze or cardiotoxin-induced muscle regeneration, and Duchenne muscular dystrophy. Overexpression of CaMKK2 suppressed C2C12 myoblast proliferation and differentiation, while inhibition of CaMKK2 had opposite effect. We also found that CaMKK2 is able to activate AMPK in C2C12 myocytes. Inhibition of AMPK could attenuate the effect of CaMKK2 overexpression, while AMPK agonist could abrogate the effect of CaMKK2 knockdown on C2C12 cell differentiation and proliferation. These results suggest that CaMKK2 functions as an AMPK kinase in muscle cells and AMPK mediates the effect of CaMKK2 on myoblast proliferation and differentiation. Our data also indicate that CaMKK2 might inhibit myoblast proliferation through AMPK-mediated cell cycle arrest by inducing cdc2-Tyr15 phosphorylation and repress differentiation through affecting PGC1α transcription. Lastly, we show that overexpressing CaMKK2 in the muscle of mice via electroporation impaired the muscle regeneration during freeze-induced injury, indicating that CaMKK2 could serve as a potential target to treat patients with muscle injury or myopathies. Together, our study reveals a new role for CaMKK2 as a negative regulator of myoblast differentiation and proliferation and sheds new light on the molecular regulation of muscle regeneration. PMID:27783047

  8. Curcumin inhibits proliferation of human lens epithelial cells: a proteomic analysis*

    PubMed Central

    Hu, Yan-hong; Huang, Xiu-rong; Qi, Ming-xin; Hou, Bu-yuan

    2012-01-01

    Objective: The incidence of after-cataracts [also known as posterior capsular opacification (PCO)] is between 30% and 50% three years following cataract surgery. Suppressing the proliferation of lens epithelial cells (LECs) is a primary goal in preventing PCO. Here, we investigated the proteomic regulation of the inhibitory effects of curcumin (Cur) on the proliferation of human lens epithelial B3 (HLE-B3) cells. Methods: Recombinant human basic fibroblast growth factor (rhbFGF) was used to induce proliferation of HLE-B3 cells, which were incubated with 20 mg/L Cur in a CO2 incubator for 24 h. Results: We found that the absorbance (A) value of rhbFGF group was significantly higher than the A value of the control group. Furthermore, the A value of the Cur group was significantly lower compared to the rhbFGF group, with an inhibition of 53.7%. Five different protein spots were obtained from proliferative HLE-B3 cells induced by rhbFGF. Eight different protein spots were obtained in HLE-B3 cells incubated with Cur. There were the common variational protein spots at mass/charge (m/z) ratios of 8 093 and 13 767 between rhbFGF group and control group as well as between the Cur group and rhbFGF group. Conclusions: These results show that Cur effectively inhibited HLE-B3 cell proliferation induced by rhbFGF. The protein spots at m/z of 8 093 and 13 767 may be the targets of Cur-induced inhibition of HLE-B3 cell proliferation. Cur may be a reliable and effective drug for prevention and treatment of polymerase chain reaction (PCR). PMID:22556179

  9. Nickel Ions Selectively Inhibit Lipopolysaccharide-Induced Interleukin-6 Production by Decreasing Its mRNA Stability

    PubMed Central

    Asakawa, Sanki; Kishimoto, Yu; Takano, Takayuki; Okita, Kiyuki; Takakuwa, Shiho; Sato, Taiki; Hiratsuka, Masahiro; Takeuchi, Osamu; Hirasawa, Noriyasu

    2015-01-01

    Nickel (Ni) ions easily elute from many alloys and elicit inflammation and allergies. Previous studies have shown that infections due to the implantation of medical devices cause inflammation and enhance the elution of Ni ions (Ni2+). However, cross-talk between infection- and Ni2+-induced signaling pathways has not yet been elucidated in detail. In the present study, we investigated the effects of Ni2+ on the lipopolysaccharide (LPS)-induced production of cytokines in a LPS-induced air pouch-type inflammation model in BALB/c mice and the murine macrophage cell line RAW264. We demonstrated that Ni2+ inhibited the LPS-induced production of interleukin (IL)-6, but not that of tumor necrosis factor (TNF)-α both in vivo and in vitro. This inhibitory effect was also observed with cobalt ion (Co2+), but not with chloride ion (Cl-), zinc ion (Zn2+), or palladium ion (Pd2+), and was highly selective to the production of IL-6. Ni2+ did not inhibit the activation of ERK1/2, p38 MAPK, or JNK. Although Ni2+ decreased IL-6 mRNA levels, it failed to inhibit the LPS-induced activation of the IL-6 promoter. An experiment using actinomycin D, a transcription inhibitor, revealed that Ni2+ decreased the stability of IL-6 mRNA. Moreover, Ni2+ inhibited the LPS-induced expression of Arid5a, but not regnase-1. These results demonstrated that Ni2+ may have selectively inhibited the LPS-induced production of IL-6 by decreasing the Arid5a-dependent stabilization of IL-6 mRNA. PMID:25742007

  10. FoxP3 inhibits proliferation and induces apoptosis of gastric cancer cells by activating the apoptotic signaling pathway

    SciTech Connect

    Ma, Gui-Fen; Chen, Shi-Yao; Sun, Zhi-Rong; Miao, Qing; Liu, Yi-Mei; Zeng, Xiao-Qing; Luo, Tian-Cheng; Ma, Li-Li; Lian, Jing-Jing; Song, Dong-Li

    2013-01-11

    Highlights: Black-Right-Pointing-Pointer The article revealed FoxP3 gene function in gastric cancer firstly. Black-Right-Pointing-Pointer Present the novel roles of FoxP3 in inhibiting proliferation and promoting apoptosis in gastric cancer cells. Black-Right-Pointing-Pointer Overexpression of FoxP3 increased proapoptotic molecules and repressed antiapoptotic molecules. Black-Right-Pointing-Pointer Silencing of FoxP3 reduced the expression of proapoptotic genes, such as PARP, caspase-3 and caspase-9. Black-Right-Pointing-Pointer FoxP3 is sufficient for activating the apoptotic signaling pathway. -- Abstract: Forkhead Box Protein 3 (FoxP3) was identified as a key transcription factor to the occurring and function of the regulatory T cells (Tregs). However, limited evidence indicated its function in tumor cells. To elucidate the precise roles and underlying molecular mechanism of FoxP3 in gastric cancer (GC), we examined the expression of FoxP3 and the consequences of interfering with FoxP3 gene in human GC cell lines, AGS and MKN45, by multiple cellular and molecular approaches, such as immunofluorescence, gene transfection, CCK-8 assay, clone formation assay, TUNEL assay, Flow cytometry, immunoassay and quantities polymerase chain reaction (PCR). As a result, FoxP3 was expressed both in nucleus and cytoplasm of GC cells. Up-regulation of FoxP3 inhibited cell proliferation and promoted cell apoptosis. Overexpression of FoxP3 increased the protein and mRNA levels of proapoptotic molecules, such as poly ADP-ribose polymerase1 (PARP), caspase-3 and caspase-9, and repressed the expression of antiapoptotic molecules, such as cellular inhibitor of apoptosis-1 (c-IAP1) and the long isoform of B cell leukemia/lymphoma-2 (Bcl-2). Furthermore, silencing of FoxP3 by siRNA in GC cells reduced the expression of proapoptotic genes, such as PARP, caspase-3 and caspase-9. Collectively, our findings identify the novel roles of FoxP3 in inhibiting proliferation and inducing apoptosis

  11. Silencing of carboxypeptidase E inhibits cell proliferation, tumorigenicity, and metastasis of osteosarcoma cells

    PubMed Central

    Fan, Shuli; Li, Xu; Li, Leiming; Wang, Liguo; Du, Zhangzhen; Yang, Yan; Zhao, Jiansong; Li, Yan

    2016-01-01

    Carboxypeptidase E (CPE), a prohormone processing enzyme, has been implicated in the progression of multiple malignancies. However, the biological role and molecular mechanisms of CPE in osteosarcoma remain elusive. In this study, we assessed the effects of CPE on cell proliferation, tumorigenicity, migration, and invasion in osteosarcoma. Our results showed that silencing of CPE significantly inhibited cell proliferation, caused cell cycle arrest at G0/G1 phase, decreased the expression levels of cell cycle protein, cyclin D1, and inhibited tumorigenicity in vivo. Additionally, CPE downregulation repressed the migratory and invasive capacities of osteosarcoma cells in vitro. Furthermore, overexpression of CPE-ΔN (a splice variant of CPE) enhanced the cell growth, migration, and invasion of osteosarcoma cells. It is possible that both CPE forms are involved in the tumorigenesis and development of osteosarcoma, and therefore CPE may provide a promising biological target for osteosarcoma therapy. PMID:27274275

  12. Inhibition of Dopamine Receptor D4 Impedes Autophagic Flux, Proliferation, and Survival of Glioblastoma Stem Cells.

    PubMed

    Dolma, Sonam; Selvadurai, Hayden J; Lan, Xiaoyang; Lee, Lilian; Kushida, Michelle; Voisin, Veronique; Whetstone, Heather; So, Milly; Aviv, Tzvi; Park, Nicole; Zhu, Xueming; Xu, ChangJiang; Head, Renee; Rowland, Katherine J; Bernstein, Mark; Clarke, Ian D; Bader, Gary; Harrington, Lea; Brumell, John H; Tyers, Mike; Dirks, Peter B

    2016-06-13

    Glioblastomas (GBM) grow in a rich neurochemical milieu, but the impact of neurochemicals on GBM growth is largely unexplored. We interrogated 680 neurochemical compounds in patient-derived GBM neural stem cells (GNS) to determine the effects on proliferation and survival. Compounds that modulate dopaminergic, serotonergic, and cholinergic signaling pathways selectively affected GNS growth. In particular, dopamine receptor D4 (DRD4) antagonists selectively inhibited GNS growth and promoted differentiation of normal neural stem cells. DRD4 antagonists inhibited the downstream effectors PDGFRβ, ERK1/2, and mTOR and disrupted the autophagy-lysosomal pathway, leading to accumulation of autophagic vacuoles followed by G0/G1 arrest and apoptosis. These results demonstrate a role for neurochemical pathways in governing GBM stem cell proliferation and suggest therapeutic approaches for GBM. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. RASSF4 Overexpression Inhibits the Proliferation, Invasion, EMT, and Wnt Signaling Pathway in Osteosarcoma Cells.

    PubMed

    Zhang, Minglei; Wang, Dapeng; Zhu, Tongtong; Yin, Ruofeng

    2017-01-02

    RASSF4, a member of the RASSF family, is broadly expressed in normal tissues but often inactivated in human cancers. Despite various studies on RASSF4, its role in osteosarcoma remains unclear. Therefore, in this study, we investigated the effects of RASSF4 expression on osteosarcoma cells and explored the underlying mechanism. The results of our study showed that RASSF4 was lowly expressed in osteosarcoma tissues and cells. RASSF4 overexpression significantly inhibited proliferation, migration, and invasion as well as the EMT process in osteosarcoma cells. Meanwhile, we found that RASSF4 overexpression markedly decreased the protein expression of β-catenin, cyclin D1, and c-Myc in osteosarcoma cells. In conclusion, our findings showed that RASSF4 overexpression inhibits proliferation, invasion, EMT, and Wnt signaling pathway in osteosarcoma cells. Thus, RASSF4 may be considered a novel target for osteosarcoma treatment.

  14. Carnosine inhibits KRAS-mediated HCT116 proliferation by affecting ATP and ROS production.

    PubMed

    Iovine, Barbara; Iannella, Maria Luigia; Nocella, Francesca; Pricolo, Maria Rosaria; Bevilacqua, Maria Assunta

    2012-02-28

    Carnosine is a natural dipeptide that has generated particular interest for its antioxidant, anti-aging and especially for its antiproliferative properties. In this study, we demonstrate that carnosine inhibits the proliferation of human HCT116 colon cancer cells. In this cell line, the activating KRAS mutation induces mitochondrial ROS, the signaling molecules for cell proliferation. We observed that 50-100 mM carnosine decreases ATP and ROS concentration and induces cell cycle arrest in G1 phase. In HCT116 cells these effects are related to decreased ERK1/2 phosphorylation and increased p21waf1 protein. Our findings support the concept that carnosine could inhibit HCT116 cell growth via its antioxidant activity and its ability to affect glycolysis.

  15. Peroxisome Proliferator Activated Receptors Alpha, Beta, and Gamma mRNA and protein expression in human fetal tissues

    EPA Science Inventory

    Peroxisome proliferator-activated receptors (PPARs) regulate lipid and glucose homeostasis, are targets of pharmaceuticals, and are also activated by environmental contaminants. Almost nothing is known about expression of PPARs during human fetal development. This study examine...

  16. Peroxisome Proliferator Activated Receptors Alpha, Beta, and Gamma mRNA and protein expression in human fetal tissues

    EPA Science Inventory

    Peroxisome proliferator-activated receptors (PPARs) regulate lipid and glucose homeostasis, are targets of pharmaceuticals, and are also activated by environmental contaminants. Almost nothing is known about expression of PPARs during human fetal development. This study examine...

  17. CCRK depletion inhibits glioblastoma cell proliferation in a cilium-dependent manner

    PubMed Central

    Yang, Ying; Roine, Niina; Mäkelä, Tomi P

    2013-01-01

    Loss of primary cilia is frequently observed in tumour cells, including glioblastoma cells, and proposed to benefit tumour growth, but a causal link has not been established. Here, we show that CCRK (cell cycle-related kinase) and its substrate ICK (intestinal cell kinase) inhibit ciliogenesis. Depletion of CCRK leads to accumulation of ICK at ciliary tips, altered ciliary transport and inhibition of cell cycle re-entry in NIH3T3 fibroblasts. In glioblastoma cells with deregulated high levels of CCRK, its depletion restores cilia through ICK and an ICK-related kinase MAK, thereby inhibiting glioblastoma cell proliferation. These results indicate that inhibition of ciliogenesis might be a mechanism used by cancer cells to provide a growth advantage. PMID:23743448

  18. Molecular basis of alpha-tocopherol inhibition of smooth muscle cell proliferation in vitro.

    PubMed

    Boscoboinik, D; Chatelain, E; Bartoli, G M; Azzi, A

    1992-01-01

    The molecular events responsible for the inhibition of cell proliferation by alpha-tocopherol have been investigated. Smooth muscle cells in vitro have been shown to be specifically inhibited by alpha-tocopherol with a concomitant inhibition of protein kinase C activity. beta-Tocopherol was inactive, despite its similar radical scavenging activity. The point of inhibition of alpha-tocopherol relative to the cell cycle was localized in the late G1 phase. A second effect of alpha-tocopherol observed with smooth muscle cells was the stimulation of protein kinase C biosynthesis in both the S and G2 phases of the cell cycle. The implications of these findings for the onset of arteriosclerosis are discussed.

  19. TFF1 inhibits proliferation and induces apoptosis of gastric cancer cells in vitro.

    PubMed

    Ge, Yanli; Zhang, Junjie; Cao, Jianchun; Wu, Qiong; Sun, Longe; Guo, Likun; Wang, Zhirong

    2012-05-01

    Trefoil Factor Family (TFF) plays an essential role in the intestinal epithelial restitution, but the relationship between TFF1 and gastric cancer (GC) is still unclear. The present study aimed to determine the role of TFF1 in repairing gastric mucosa and in the pathogenesis of GC. The TFF1 expression in different gastric mucosas was measured with immunohistochemistry. Then, siRNA targeting TFF1 or plasmids expressing TFF1 gene were transfected into BGC823 cells, SGC7901 cells and GES-1 cells. The cell proliferation was detected with MTT assay and apoptosis and cell cycle measured by flow cytometry. From normal gastric mucosa to mucosa with dysplasia and to gastric cancer, the TFF1 expression had a decreasing trend. Down-regulation of TFF1 expression significantly reduced the apoptosis of three cell lines and markedly facilitated their proliferation but had no significant effect on cell cycle. Over-expression of TFF1 could promote apoptosis of three cell lines and inhibit proliferation but had no pronounced effect on cell cycle. TFF1 can inhibit proliferation and induce apoptosis of GC cells in vitro.

  20. Cdk4 deficiency inhibits skin tumor development but does not affect normal keratinocyte proliferation.

    PubMed

    Rodriguez-Puebla, Marcelo L; Miliani de Marval, Paula L; LaCava, Margaret; Moons, David S; Kiyokawa, Hiroaki; Conti, Claudio J

    2002-08-01

    Most human tumors have mutations that result in deregulation of the cdk4/cyclin-Ink4-Rb pathway. Overexpression of D-type cyclins or cdk4 and inactivation of Ink4 inhibitors are common in human tumors. Conversely, lack of cyclin D1 expression results in significant reduction in mouse skin and mammary tumor development. However, complete elimination of tumor development was not observed in these models, suggesting that other cyclin/cdk complexes play an important role in tumorigenesis. Here we described the effects of cdk4 deficiency on mouse skin proliferation and tumor development. Cdk4 deficiency resulted in a 98% reduction in the number of tumors generated through the two-stage carcinogenesis model. The absence of cdk4 did not affect normal keratinocyte proliferation and both wild-type and cdk4 knockout epidermis are equally affected after topical treatment with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA), resulting in epidermal hyperplasia. In similar fashion, cdk4 knockout keratinocytes proliferated well in an in vivo model of wound-induced proliferation. Biochemical studies in mouse epidermis showed that cdk6 activity increased twofold in cdk4-deficient mice compared to wild-type siblings. These results suggest that therapeutic approaches to inhibit cdk4 activity could provide a target to inhibit tumor development with minimal or no effect in normal tissue.

  1. Pyruvate fuels mitochondrial respiration and proliferation of breast cancer cells: effect of monocarboxylate transporter inhibition.

    PubMed

    Diers, Anne R; Broniowska, Katarzyna A; Chang, Ching-Fang; Hogg, Neil

    2012-06-15

    Recent studies have highlighted the fact that cancer cells have an altered metabolic phenotype, and this metabolic reprogramming is required to drive the biosynthesis pathways necessary for rapid replication and proliferation. Specifically, the importance of citric acid cycle-generated intermediates in the regulation of cancer cell proliferation has been recently appreciated. One function of MCTs (monocarboxylate transporters) is to transport the citric acid cycle substrate pyruvate across the plasma membrane and into mitochondria, and inhibition of MCTs has been proposed as a therapeutic strategy to target metabolic pathways in cancer. In the present paper, we examined the effect of different metabolic substrates (glucose and pyruvate) on mitochondrial function and proliferation in breast cancer cells. We demonstrated that cancer cells proliferate more rapidly in the presence of exogenous pyruvate when compared with lactate. Pyruvate supplementation fuelled mitochondrial oxygen consumption and the reserve respiratory capacity, and this increase in mitochondrial function correlated with proliferative potential. In addition, inhibition of cellular pyruvate uptake using the MCT inhibitor α-cyano-4-hydroxycinnamic acid impaired mitochondrial respiration and decreased cell growth. These data demonstrate the importance of mitochondrial metabolism in proliferative responses and highlight a novel mechanism of action for MCT inhibitors through suppression of pyruvate-fuelled mitochondrial respiration.

  2. Parecoxib inhibits glioblastoma cell proliferation, migration and invasion by upregulating miRNA-29c

    PubMed Central

    Li, Lin-Yong; Xiao, Jie; Liu, Qiang

    2017-01-01

    ABSTRACT Glioblastoma (GBM) is one of the most lethal brain cancers worldwide, and there is an urgent need for development of novel therapeutic approaches. Parecoxib is a well-known cyclooxygenase-2 (COX-2) inhibitor, and had already been developed for postoperative analgesia with high efficacy and low adverse reaction. A recent study has suggested that parecoxib potently enhances immunotherapeutic efficacy of GBM, but its effects on GBM growth, migration and invasion have not previously been studied. In the present study, MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] and BrdU (5-bromo-2-deoxyuridine) incorporation assays were used to evaluate the cell proliferation of GBM cells. Wound-healing and transwell assays were preformed to analyze GBM cell migration and invasion, respectively. The results suggested that parecoxib inhibits cell proliferation, migration and invasion of GBM cells in a dose-dependent manner. RT-qPCR (real-time quantitative PCR) analysis demonstrated that miRNA-29c can be significantly induced by parecoxib. Furthermore, our data suggests that a miRNA-29c inhibitor can significantly attenuate parecoxib's effect on proliferation, migration and invasion of GBM. In conclusion, the present study suggests that parecoxib inhibits GBM cell proliferation, migration and invasion by upregulating miRNA-29c. PMID:27895048

  3. miR-125b inhibits osteoblastic differentiation by down-regulation of cell proliferation

    SciTech Connect

    Mizuno, Yosuke; Yagi, Ken; Tokuzawa, Yoshimi; Kanesaki-Yatsuka, Yukiko; Suda, Tatsuo; Katagiri, Takenobu; Fukuda, Toru; Maruyama, Masayoshi; Okuda, Akihiko; Amemiya, Tomoyuki; Kondoh, Yasumitsu; Tashiro, Hideo; Okazaki, Yasushi

    2008-04-04

    Although various microRNAs regulate cell differentiation and proliferation, no miRNA has been reported so far to play an important role in the regulation of osteoblast differentiation. Here we describe the role of miR-125b in osteoblastic differentiation in mouse mesenchymal stem cells, ST2, by regulating cell proliferation. The expression of miR-125b was time-dependently increased in ST2 cells, and the increase in miR-125b expression was attenuated in osteoblastic-differentiated ST2 cells induced by BMP-4. The transfection of exogenous miR-125b inhibited proliferation of ST2 cells and caused inhibition of osteoblastic differentiation. In contrast, when the endogenous miR-125b was blocked by transfection of its antisense RNA molecule, alkaline phosphatase activity after BMP-4 treatment was elevated. These results strongly suggest that miR-125b is involved in osteoblastic differentiation through the regulation of cell proliferation.

  4. Pyruvate fuels mitochondrial respiration and proliferation of breast cancer cells: effect of monocarboxylate transporter inhibition

    PubMed Central

    Diers, Anne R.; Broniowska, Katarzyna A.; Chang, Ching-Fang; Hogg, Neil

    2016-01-01

    Recent studies have highlighted the fact that cancer cells have an altered metabolic phenotype, and this metabolic reprogramming is required to drive biosynthesis pathways necessary for rapid replication and proliferation. Specifically, the importance of tricarboxylic acid (TCA) cycle-generated intermediates in the regulation of cancer cells proliferation has been recently appreciated. One function of monocarboxylate transporters (MCTs) is to transport the TCA cycle substrate pyruvate across the plasma membrane and into mitochondria, and inhibition of MCTs has been proposed as a therapeutic strategy to target metabolic pathways in cancer. Here, we examined the effect of different metabolic substrates (glucose and pyruvate) on mitochondrial function and proliferation in breast cancer cells. We demonstrated that cancer cells proliferate more rapidly in the presence of exogenous pyruvate when compared to lactate. Pyruvate supplementation fueled mitochondrial oxygen consumption and the reserve respiratory capacity, and this increase in mitochondrial function correlated with proliferative potential. In addition, inhibition of cellular pyruvate uptake using the MCT inhibitor α-cyano-4-hydroxycinnamic acid impaired mitochondrial respiration and decreased cell growth. These data demonstrate the importance of mitochondrial metabolism in proliferative responses and highlight a novel mechanism of action for MCT inhibitors through suppression of pyruvate-fueled mitochondrial respiration. PMID:22458763

  5. Dehydroleucodine inhibits vascular smooth muscle cell proliferation in G2 phase.

    PubMed

    Cruzado, M; Castro, C; Fernandez, D; Gomez, L; Roque, M; Giordano, O E; Lopez, L A

    2005-11-08

    Vascular smooth muscle cell (VSMC) proliferation plays an important role in the development of atherosclerosis and in the vascular changes seen in hypertension. Dehydroleucodine (DhL) is a sesquiterpene lactone that inhibits cell proliferation in plant cells. In this paper, we study the effect of DhL in the proliferation of VSMCs stimulated with 10% fetal bovine serum (FBS). Very low concentrations of DhL (2-6 microM) inhibited VSMC proliferation and induced cell accumulation in G2. DhL did not affect the dynamics of 3H-thymidine incorporation, and did not modify either the activity of DNA polymerase or the incorporation of deoxyribonucleotides in an in vitro assay. Moreover, DhL did not induce apoptosis in VSMCs. These results indicate that DhL, in very low concentration, induces a transient arrest of VSMCs in G2. Our data show that VSMCs are especially sensitive to DhL effect, suggesting that DhL could be potentially useful to prevent the vascular pathological changes seen in hypertension and other vascular diseases.

  6. Tramadol inhibits proliferation, migration and invasion via α2-adrenoceptor signaling in breast cancer cells.

    PubMed

    Xia, M; Tong, J-H; Zhou, Z-Q; Duan, M-L; Xu, J-G; Zeng, H-J; Wang, S-H

    2016-01-01

    The aim of this study was to examine the function of tramadol on cell proliferation, migration and invasion in breast cancer cells in vitro, and to evaluate the effect of tramadol in vivo. Further, we explore the mechanism accounting for the role of tramadol on breast cancer cells. Cell proliferation was detected by the methyl thiazolyl tetrazolium (MTT) assay. Wound healing assay and transwell assay was applied to quantify the migration and invasion ability of MDA-MB-231 cells. The expression of endogenous α2-adrenoceptor and ERK was measured by Western blotting. Tramadol at a clinical dose of up to 2 μM significantly inhibited the proliferation, migration and invasion in a time-dependent manner from day 0 to 28 in vitro. Moreover, tramadol suppressed the growth of xenotransplant tumor in vivo markedly. Furthermore, the protein levels of α2-adrenoceptor and phosphorylated ERK were decreased by tramadol, whereas the expression of total ERK remained unchanged. In addition, downregulation of α2-adrenoceptor by yohimbine could mimic the effect of tramadol treatment. Collectively, we demonstrated that tramadol could inhibit proliferation, migration and invasion of breast cancers via inactivating α2-adrenoceptor signaling pathway. Our data provide the experimental fundamental for further investigation of the anti-cancer effect of tramadol in breast cancer cells.

  7. Nitric oxide mediates low magnesium inhibition of osteoblast-like cell proliferation.

    PubMed

    Leidi, Marzia; Dellera, Federica; Mariotti, Massimo; Banfi, Giuseppe; Crapanzano, Calogero; Albisetti, Walter; Maier, Jeanette A M

    2012-10-01

    An adequate intake of magnesium (Mg) is important for bone cell activity and contributes to the prevention of osteoporosis. Because (a) Mg is mitogenic for osteoblasts and (b) reduction of osteoblast proliferation is detected in osteoporosis, we investigated the influence of different concentrations of extracellular Mg on osteoblast-like SaOS-2 cell behavior. We found that low Mg inhibited SaOS-2 cell proliferation by increasing the release of nitric oxide through the up-regulation of inducible nitric oxide synthase (iNOS). Indeed, both pharmacological inhibition with the iNOS inhibitor l-N(6)-(iminoethyl)-lysine-HCl and genetic silencing of iNOS by small interfering RNA restored the normal proliferation rate of the cells. Because a moderate induction of nitric oxide is sufficient to potentiate bone resorption and a relative deficiency in osteoblast proliferation can result in their inadequate activity, we conclude that maintaining Mg homeostasis is relevant to ensure osteoblast function and, therefore, to prevent osteoporosis.

  8. Molecular effects of leptin on peroxisome proliferator activated receptor gamma (PPAR-γ) mRNA expression in rat's adipose and liver tissue.

    PubMed

    Abbasi, A; Moghadam, A A; Kahrarian, Z; Abbsavaran, R; Yari, K; Alizadeh, E

    2017-08-15

    Leptin is a 16-kDa peptide hormone secreted by adipose tissue that participates in the regulation of energy homeostasis. The aim of this study was to determine the effect of leptin injection on mRNA expression of peroxisome proliferator-activated receptor gamma (PPAR-γ) and comparison of PPAR-γ mRNA expression in rat's adipose and liver tissue. Twenty adult male rats were divided into the following groups: Group 1asa control (n=10) that did not receive any treatment. Group 2as a treatment (n=10) that received leptin (30 µg ⁄ kg BW) intraperitoneally (ip) for two successive days. Blood samples were taken before and one day after second leptin injection for triglyceride (TG), Free Fatty Acid (FFA), HLD-cholesterol, and LDL-cholesterol measurement. Total RNA was extractedfrom the adipose tissue and liver tissues of rats.  Adipose and liver tissue cells' cDNA was synthesized to characterize the expression of PPAR-γ. Gene expression of PPAR-γ mRNA was tested by RT- PCR technique. Results show leptin decreases expression of PPAR-γ on rat. Low levels of PPAR-γ mRNA were detected in adipose and liver tissues of treatment rats in comparison to control group. In treatment group, the level of PPAR-γ mRNA in liver tissue was very lower than the adipose tissue. The levels of HDL and FFA in treatment rats were increased whereas serum levels TG, VLDL and LDL were not changed. It is concluded that leptin signal with suppressing of PPAR-γ mRNA expression in rat's adipose and liver tissues can result in lipolysis instead of lipogenesis.

  9. β-catenin knockdown inhibits the proliferation of human glioma cells in vitro and in vivo

    PubMed Central

    WANG, ZHONG; CHEN, QIANXUE

    2016-01-01

    β-catenin is a crucial oncogene that is capable of regulating cancer progression. The aim of the present study was to clarify whether β-catenin was associated with the proliferation and progress of glioma. In order to knockdown the expression of β-catenin in human U251 glioma cells, three pairs of small interfering (si)RNA were designed and synthesized and the most effective siRNA was selected and used for silencing the endogenous β-catenin, which was detected by western blot analysis and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Proliferation was subsequently detected using a methylthiazolyl-tetrazolium bromide assay and the results demonstrated that knockdown of β-catenin significantly inhibited the proliferation of U251 cells in a time- and dose-dependent manner (P<0.01). Cell apoptosis rate was analyzed using flow cytometry and Annexin V-fluorescein isothiocyanate/propidium iodide staining demonstrated that β-catenin siRNA significantly increased the apoptosis of U251 cells (P<0.01). Furthermore, the results of an in vitro scratch assay demonstrated that β-catenin silencing suppressed the proliferation of U251 cells, as compared with the control group (P<0.01). In vivo, β-catenin expression levels in U251 cells were significantly inhibited (P<0.01) following β-catenin short hairpin (sh)RNA lentiviral-vector transfection, as detected by western blot analysis and RT-qPCR. Tumorigenicity experiments demonstrated that β-catenin inhibition significantly increased the survival rate of nude mice. The results of the present study demonstrated that knockdown of β-catenin expression significantly inhibited the progression of human glioma cancer cells, in vitro and in vivo; thus suggesting that β-catenin silencing may be a novel therapy for the treatment of human glioma. PMID:26998037

  10. Inhibition of antiviral drug cidofovir on proliferation of human papillomavirus-infected cervical cancer cells.

    PubMed

    Yang, Jing; Dai, Lv-Xia; Chen, Ming; Li, Bei; Ding, Nana; Li, Gang; Liu, Yan-Qing; Li, Ming-Yuan; Wang, Bao-Ning; Shi, Xin-Li; Tan, Hua-Bing

    2016-11-01

    In order to evaluate the potential application value of cidofovir (CDV) in the prevention of human papillomavirus (HPV) infection and treatment of cervical cancer, the inhibitory effect of CDV on the proliferation of HPV 18-positive HeLa cells in cervical cancer was preliminarily investigated, using cisplatin (DDP) as a positive control. An MTT assay was used to analyze the effects of CDV and DDP on HeLa cell proliferation. In addition, clone formation assay and Giemsa staining were used to examine the extent of HeLa cell apoptosis caused by CDV and DDP. Flow cytometry was also used to detect the shape and size of apoptotic cells following propidium iodide staining, while western blot analysis identified the expression levels of of E6 and p53 proteins in HeLa cells. A cell climbing immunofluorescence technique was used to locate the subcellular position of p53 in HeLa cells. The results demonstrated that CDV and DDP inhibited the proliferation of HeLa cells in a concentration- and time-dependent manner. Flow cytometry showed that CDV and DDP treatments resulted in cell arrest in the S-phase, and triggered programmed cell death. Furthermore, western blot analysis revealed that CDV and DDP inhibited E6 protein expression and activated p53 expression in HeLa cells. Finally, the immunofluorescence results indicated that CDV and DDP inhibited the nuclear export of p53 by E6 protein, which is required for degradation of endogenous p53 by MDM2 and human papilloma virus E6. In conclusion, CDV and DDP inhibited HeLa cell proliferation in a concentration- and time-dependent manner, reduced the expression of E6 protein, and reinstated p53 protein activity. Thus, CDV regulates cell cycle arrest and apoptosis, and may be a potential cervical cancer therapeutic strategy.

  11. Inhibition of human lung cancer cell proliferation and survival by wine.

    PubMed

    Barron, Carly C; Moore, Jessy; Tsakiridis, Theodoros; Pickering, Gary; Tsiani, Evangelia

    2014-01-23

    Compounds of plant origin and food components have attracted scientific attention for use as agents for cancer prevention and treatment. Wine contains polyphenols that were shown to have anti-cancer and other health benefits. The survival pathways of Akt and extracellular signal-regulated kinase (Erk), and the tumor suppressor p53 are key modulators of cancer cell growth and survival. In this study, we examined the effects of wine on proliferation and survival of human Non-small cell lung cancer (NSCLC) cells and its effects on signaling events. Human NSCLC adenocarcinoma A549 and H1299 cells were used. Cell proliferation was assessed by thymidine incorporation. Clonogenic assays were used to assess cell survival. Immunoblotting was used to examine total and phosphorylated levels of Akt, Erk and p53. In A549 cells red wine inhibited cell proliferation and reduced clonogenic survival at doses as low as 0.02%. Red wine significantly reduced basal and EGF-stimulated Akt and Erk phosphorylation while it increased the levels of total and phosphorylated p53 (Ser15). Control experiments indicated that the anti-proliferative effects of wine were not mediated by the associated contents of ethanol or the polyphenol resveratrol and were independent of glucose transport into cancer cells. White wine also inhibited clonogenic survival, albeit at a higher doses (0.5-2%), and reduced Akt phosphorylation. The effects of both red and white wine on Akt phosphorylation were also verified in H1299 cells. Red wine inhibits proliferation of lung cancer cells and blocks clonogenic survival at low concentrations. This is associated with inhibition of basal and EGF-stimulated Akt and Erk signals and enhancement of total and phosphorylated levels of p53. White wine mediates similar effects albeit at higher concentrations. Our data suggest that wine may have considerable anti-tumour and chemoprevention properties in lung cancer and deserves further systematic investigation in animal models

  12. Inhibition of antiviral drug cidofovir on proliferation of human papillomavirus-infected cervical cancer cells

    PubMed Central

    Yang, Jing; Dai, Lv-Xia; Chen, Ming; Li, Bei; Ding, Nana; Li, Gang; Liu, Yan-Qing; Li, Ming-Yuan; Wang, Bao-Ning; Shi, Xin-Li; Tan, Hua-Bing

    2016-01-01

    In order to evaluate the potential application value of cidofovir (CDV) in the prevention of human papillomavirus (HPV) infection and treatment of cervical cancer, the inhibitory effect of CDV on the proliferation of HPV 18-positive HeLa cells in cervical cancer was preliminarily investigated, using cisplatin (DDP) as a positive control. An MTT assay was used to analyze the effects of CDV and DDP on HeLa cell proliferation. In addition, clone formation assay and Giemsa staining were used to examine the extent of HeLa cell apoptosis caused by CDV and DDP. Flow cytometry was also used to detect the shape and size of apoptotic cells following propidium iodide staining, while western blot analysis identified the expression levels of of E6 and p53 proteins in HeLa cells. A cell climbing immunofluorescence technique was used to locate the subcellular position of p53 in HeLa cells. The results demonstrated that CDV and DDP inhibited the proliferation of HeLa cells in a concentration- and time-dependent manner. Flow cytometry showed that CDV and DDP treatments resulted in cell arrest in the S-phase, and triggered programmed cell death. Furthermore, western blot analysis revealed that CDV and DDP inhibited E6 protein expression and activated p53 expression in HeLa cells. Finally, the immunofluorescence results indicated that CDV and DDP inhibited the nuclear export of p53 by E6 protein, which is required for degradation of endogenous p53 by MDM2 and human papilloma virus E6. In conclusion, CDV and DDP inhibited HeLa cell proliferation in a concentration- and time-dependent manner, reduced the expression of E6 protein, and reinstated p53 protein activity. Thus, CDV regulates cell cycle arrest and apoptosis, and may be a potential cervical cancer therapeutic strategy. PMID:27882102

  13. Silencing Nrf2 impairs glioma cell proliferation via AMPK-activated mTOR inhibition

    SciTech Connect

    Jia, Yue; Wang, Handong; Wang, Qiang; Ding, Hui; Wu, Heming; Pan, Hao

    2016-01-15

    Gliomas are the leading cause of death among adults with primary brain malignancies. Treatment for malignant gliomas remains limited, and targeted therapies have been incompletely explored. Nuclear factor erythroid 2-related factor 2 (Nrf2), a key transcription regulator for antioxidant and detoxification enzymes, is abundantly expressed in cancer cells. In this study, the role and mechanism of Nrf2 in cancer cell proliferation was investigated in multiple glioma cell lines. We first evaluated the expression patterns of Nrf2 in four glioma cell lines and found all four cell lines expressed Nrf2, but the highest level was observed in U251 cells. We further evaluated the biological functions of Nrf2 in U251 glioma cell proliferation by specific inhibition of Nrf2 using short hairpin RNA (shRNA). We found that Nrf2 depletion inhibited glioma cell proliferation. Nrf2 depletion also decreased colony formation in U251 cells stably expressing Nrf2 shRNA compared to scrambled control shRNA. Moreover, suppression of Nrf2 expression could lead to ATP depletion (with concomitant rise in AMP/ATP ratio) and consequently to AMPK-activated mTOR inhibition. Finally, activation of adenosine monophosphate–activated protein kinase (AMPK) by treated with phenformin, an AMPK agonist, can mimic the inhibitory effect of Nrf2 knockdown in U251 cells. In conclusion, our findings will shed light to the role and mechanism of Nrf2 in regulating glioma proliferation via ATP-depletion-induced AMPK activation and consequent mTOR inhibition, a novel insight into our understanding the role and mechanism of Nrf2 in glioma pathoetiology. To our knowledge, this is also the first report to provide a rationale for the implication of cross-linking between Nrf2 and mTOR signaling.

  14. Inhibition of human lung cancer cell proliferation and survival by wine

    PubMed Central

    2014-01-01

    Background Compounds of plant origin and food components have attracted scientific attention for use as agents for cancer prevention and treatment. Wine contains polyphenols that were shown to have anti-cancer and other health benefits. The survival pathways of Akt and extracellular signal-regulated kinase (Erk), and the tumor suppressor p53 are key modulators of cancer cell growth and survival. In this study, we examined the effects of wine on proliferation and survival of human Non-small cell lung cancer (NSCLC) cells and its effects on signaling events. Methods Human NSCLC adenocarcinoma A549 and H1299 cells were used. Cell proliferation was assessed by thymidine incorporation. Clonogenic assays were used to assess cell survival. Immunoblotting was used to examine total and phosphorylated levels of Akt, Erk and p53. Results In A549 cells red wine inhibited cell proliferation and reduced clonogenic survival at doses as low as 0.02%. Red wine significantly reduced basal and EGF-stimulated Akt and Erk phosphorylation while it increased the levels of total and phosphorylated p53 (Ser15). Control experiments indicated that the anti-proliferative effects of wine were not mediated by the associated contents of ethanol or the polyphenol resveratrol and were independent of glucose transport into cancer cells. White wine also inhibited clonogenic survival, albeit at a higher doses (0.5-2%), and reduced Akt phosphorylation. The effects of both red and white wine on Akt phosphorylation were also verified in H1299 cells. Conclusions Red wine inhibits proliferation of lung cancer cells and blocks clonogenic survival at low concentrations. This is associated with inhibition of basal and EGF-stimulated Akt and Erk signals and enhancement of total and phosphorylated levels of p53. White wine mediates similar effects albeit at higher concentrations. Our data suggest that wine may have considerable anti-tumour and chemoprevention properties in lung cancer and deserves further

  15. Slow and sustained nitric oxide releasing compounds inhibit multipotent vascular stem cell proliferation and differentiation without causing cell death

    SciTech Connect

    Curtis, Brandon M.; Leix, Kyle Alexander; Ji, Yajing; Glaves, Richard Samuel Elliot; Ash, David E.; Mohanty, Dillip K.

    2014-07-18

    Highlights: • Multipotent vascular stem cells (MVSCs) proliferate and differentiate. • Nitric oxide inhibits proliferation of MVSCs. • Nitric oxide inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs). • Smooth muscle cells (SMCs) neither de-differentiate nor proliferate. - Abstract: Atherosclerosis is the leading cause of cerebral and myocardial infarction. It is believed that neointimal growth common in the later stages of atherosclerosis is a result of vascular smooth muscle cell (SMC) de-differentiation in response to endothelial injury. However, the claims of the SMC de-differentiation theory have not been substantiated by monitoring the fate of mature SMCs in response to such injuries. A recent study suggests that atherosclerosis is a consequence of multipotent vascular stem cell (MVSC) differentiation. Nitric oxide (NO) is a well-known mediator against atherosclerosis, in part because of its inhibitory effect on SMC proliferation. Using three different NO-donors, we have investigated the effects of NO on MVSC proliferation. Results indicate that NO inhibits MVSC proliferation in a concentration dependent manner. A slow and sustained delivery of NO proved to inhibit proliferation without causing cell death. On the other hand, larger, single-burst NO concentrations, inhibits proliferation, with concurrent significant cell death. Furthermore, our results indicate that endogenously produced NO inhibits MVSC differentiation to mesenchymal-like stem cells (MSCs) and subsequently to SMC as well.

  16. Suppression of Akt1 phosphorylation by adenoviral transfer of the PTEN gene inhibits hypoxia-induced proliferation of rat pulmonary arterial smooth muscle cells

    SciTech Connect

    Luo, Chunxia; Yi, Bin; Bai, Li; Xia, Yongzhi; Wang, Guansong; Qian, Guisheng; Feng, Hua

    2010-07-02

    Recent findings identify the role of proliferation of pulmonary artery smooth muscle cells (PASMCs) in pulmonary vascular remodeling. Phosphoinositide 3 kinase (PI3K) and serine/threonine kinase (Akt) proteins are expressed in vascular smooth muscle cells. In addition, phosphatase and tensin homolog deleted on chromosome 10 (PTEN) has been identified as a negative regulator of cytokine signaling that inhibits the PI3K-Akt pathway. However, little is known about the role of PTEN/Akt signaling in hypoxia-associated vascular remodeling. In this study, we found that hypoxia-induced the expression of Akt1 mRNA and phosphorylated protein by at least twofold in rat PASMCs. Phospho-PTEN significantly decreased in the nuclei of PASMCs after hypoxic stimulation. After forcing over-expression of PTEN by adenovirus-mediated PTEN (Ad-PTEN) transfection, the expression of phospho-Akt1 was significantly suppressed in PASMCs at all time-points measured. Additionally, we showed here that hypoxia increased proliferation of PASMCs by nearly twofold and over-expression of PTEN significantly inhibited hypoxia-induced PASMCs proliferation. These findings suggest that phospho-PTEN loss in the nuclei of PASMCs under hypoxic conditions may be the major cause of aberrant activation of Akt1 and may, therefore, play an important role in hypoxia-associated pulmonary arterial remodeling. Finally, the fact that transfection with Ad-PTEN inhibits the phosphorylation of Akt1 in PASMCs suggests a potential therapeutic effect on hypoxia-associated pulmonary arterial remodeling.

  17. A Mechanism for the Inhibition of Neural Progenitor Cell Proliferation by Cocaine

    PubMed Central

    Lee, Chun-Ting; Chen, Jia; Hayashi, Teruo; Tsai, Shang-Yi; Sanchez, Joseph F; Errico, Stacie L; Amable, Rose; Su, Tsung-Ping; Lowe, Ross H; Huestis, Marilyn A; Shen, James; Becker, Kevin G; Geller, Herbert M; Freed, William J

    2008-01-01

    Background Prenatal exposure of the developing brain to cocaine causes morphological and behavioral abnormalities. Recent studies indicate that cocaine-induced proliferation inhibition and/or apoptosis in neural progenitor cells may play a pivotal role in causing these abnormalities. To understand the molecular mechanism through which cocaine inhibits cell proliferation in neural progenitors, we sought to identify the molecules that are responsible for mediating the effect of cocaine on cell cycle regulation. Methods and Findings Microarray analysis followed by quantitative real-time reverse transcription PCR was used to screen cocaine-responsive and cell cycle-related genes in a neural progenitor cell line where cocaine exposure caused a robust anti-proliferative effect by interfering with the G1-to-S transition. Cyclin A2, among genes related to the G1-to-S cell cycle transition, was most strongly down-regulated by cocaine. Down-regulation of cyclin A was also found in cocaine-treated human primary neural and A2B5+ progenitor cells, as well as in rat fetal brains exposed to cocaine in utero. Reversing cyclin A down-regulation by gene transfer counteracted the proliferation inhibition caused by cocaine. Further, we found that cocaine-induced accumulation of reactive oxygen species, which involves N-oxidation of cocaine via cytochrome P450, promotes cyclin A down-regulation by causing an endoplasmic reticulum (ER) stress response, as indicated by increased phosphorylation of eIF2α and expression of ATF4. In the developing rat brain, the P450 inhibitor cimetidine counteracted cocaine-induced inhibition of neural progenitor cell proliferation as well as down-regulation of cyclin A. Conclusions Our results demonstrate that down-regulation of cyclin A underlies cocaine-induced proliferation inhibition in neural progenitors. The down-regulation of cyclin A is initiated by N-oxidative metabolism of cocaine and consequent ER stress. Inhibition of cocaine N

  18. Kaempferol inhibits cell proliferation and glycolysis in esophagus squamous cell carcinoma via targeting EGFR signaling pathway.

    PubMed

    Yao, Shihua; Wang, Xiaowei; Li, Chunguang; Zhao, Tiejun; Jin, Hai; Fang, Wentao

    2016-08-01

    Antitumor activity of kaempferol has been studied in various tumor types, but its potency in esophagus squamous cell carcinoma is rarely known. Here, we reported the activity of kaempferol against esophagus squamous cell carcinoma as well as its antitumor mechanisms. Results of cell proliferation and colony formation assay showed that kaempferol substantially inhibited tumor cell proliferation and clone formation in vitro. Flow cytometric analysis demonstrated that tumor cells were induced G0/G1 phase arrest after kaempferol treatment, and the expression of protein involved in cell cycle regulation was dramatically changed. Except the potency on cell proliferation, we also discovered that kaempferol had a significant inhibitory effect against tumor glycolysis. With the downregulation of hexokinase-2, glucose uptake and lactate production in tumor cells were dramatically declined. Mechanism studies revealed kaempferol had a direct effect on epidermal growth factor receptor (EGFR) activity, and along with the inhibition of EGFR, its downstream signaling pathways were also markedly suppressed. Further investigations found that exogenous overexpression of EGFR in tumor cells substantially attenuated glycolysis suppression induced by kaempferol, which implied that EGFR also played an important role in kaempferol-mediated glycolysis inhibition. Finally, the antitumor activity of kaempferol was validated in xenograft model and kaempferol prominently restrained tumor growth in vivo. Meanwhile, dramatic decrease of EGFR activity and hexokinase-2 expression were observed in kaempferol-treated tumor tissue, which confirmed these findings in vitro. Briefly, these studies suggested that kaempferol, or its analogues, may serve as effective candidates for esophagus squamous cell carcinoma management.

  19. The recombinant beta subunit of C-phycocyanin inhibits cell proliferation and induces apoptosis.

    PubMed

    Wang, Haizhen; Liu, Yongding; Gao, Xueliang; Carter, Christie L; Liu, Zhi-Ren

    2007-03-08

    C-Phycocyanin (C-PC) from blue-green algae has been reported to have various pharmacological characteristics, including anti-inflammatory and anti-tumor activities. In this study, we expressed the beta-subunit of C-PC (ref to as C-PC/beta) in Escherichia coli. We found that the recombinant C-PC/beta has anti-cancer properties. Under the treatment of 5 microM of the recombinant C-PC/beta, four different cancer cell lines accrued high proliferation inhibition and apoptotic induction. Substantially, a lower response occurred in non-cancer cells. We investigated the mechanism by which C-PC/beta inhibits cancer cell proliferation and induces apoptosis. We found that the C-PC/beta interacts with membrane-associated beta-tubulin and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Under the treatment of the C-PC/beta, depolymerization of microtubules and actin-filaments were observed. The cells underwent apoptosis with an increase in caspase-3, and caspase-8 activities. The cell cycle was arrested at the G0/G1 phase under the treatment of C-PC/beta. In addition, the nuclear level of GAPDH decreased significantly. Decrease in the nuclear level of GAPDH prevents the cell cycle from entering into the S phase. Inhibition of cancer cell proliferation and induction of apoptosis may potentate the C-PC/beta as a promising cancer prevention or therapy agent.

  20. Bisphenol A Inhibits Cell Proliferation and Reduces the Motile Potential of Murine LM8 Osteosarcoma Cells.

    PubMed

    Kidani, Teruki; Yasuda, Rie; Miyawaki, Joji; Oshima, Yusuke; Miura, Hiromasa; Masuno, Hiroshi

    2017-04-01

    The aim of this study was to examine the effect of bisphenol A (BPA) on the proliferation and motility potential of murine LM8 osteosarcoma cells. LM8 cells were treated for 3 days with or without 80 μM BPA. The effect of BPA on cell proliferation was determined by DNA measurement in the cultures and 5-bromo-2'-deoxyuridine (BrdU) incorporation study. Ethanol-fixed cells were stained with hematoxylin-eosin (H&E) to visualize cell morphology. Cell motility was assayed using inserts with uncoated membranes in invasion chambers. Expression of cell division cycle 42 (CDC42) was determined by immunofluorescence staining and western blotting. BPA reduced the DNA content of cultures and the number of BrdU-positive cells. BPA induced a change in morphology from cuboidal with multiple filopodia on the cell surface to spindle-shaped with a smooth cell surface. BPA-treated cells expressed less CDC42 and were less motile than untreated cells. BPA inhibited DNA replication and cell proliferation. BPA inhibited filopodia formation and motile potential by inhibiting CDC42 expression in LM8 cells. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  1. Inhibition of lipid peroxidation, cyclooxygenase enzyme and human tumor cell proliferation by compounds in herbal water.

    PubMed

    Liu, Yunbao; Mulabagal, Vanisree; Bowen-Forbes, Camille S; Aviayan, Rejanish; Nair, Muraleedharan G

    2009-09-01

    A powdered mixture of dried herbs, "Panamrutham", is sold in India for the preparation of "herbal drinking water". The hot water extract of this herbal mixture gave lipid peroxidation (LPO), cyclo-oxygenase (COX-1 and -2) enzyme and human tumor cell proliferation inhibitory activities between 25 and 250 microg/mL. The bioassay-guided purification of the water extract afforded a novel compound (1), along with phenolics (2, 4, 6, and 7) and sesquiterpenoids (3 and 5). The isolates were evaluated for LPO, COX-1 and -2 enzyme and human tumor cell proliferation inhibitory activities. At 25 microg/mL, compounds 1-7 inhibited LPO by 22-73% and COX-1 and -2 enzymes by 3-14% and 14-74%, respectively. Compounds 5 and 6 at 25 microg/mL showed growth inhibition of colon, gastric, lung, breast and central nervous system human tumor cell lines by 60 and 67, 43 and 60, 24 and 64, 34 and 65, 6 and 27%, respectively. Compounds 2, 4 and 7 displayed weak or moderate growth inhibition of colon, gastric and breast human tumor cell lines. This is the first report on the LPO inhibitory activities of compounds 1 and 3-7 and the COX and tumor cell proliferation inhibitory activities of compounds 1, 3-5 and 7.

  2. Picotamide, an antithromboxane agent, inhibits the migration and proliferation of arterial myocytes.

    PubMed

    Ratti, S; Quarato, P; Casagrande, C; Fumagalli, R; Corsini, A

    1998-08-14

    Picotamide is an antiplatelet drug with a peculiar dual mechanism of action: it inhibits thromboxane A2 synthase and antagonizes the pharmacological responses mediated by thromboxane A2 receptor. We investigated the in vitro effect of picotamide on smooth muscle cell migration and proliferation. Picotamide (1-500 microM) decreased human and rat smooth muscle cell proliferation, evaluated as cell number, in a concentration-dependent and reversible manner. Picotamide inhibited DNA synthesis induced by fetal calf serum (10%), platelet-derived growth factor (PDGF-BB (20 ng/ml)), epidermal growth factor (EGF (1 nM)) and (15S)-hydroxy-11,9-(epoxymethano)prosta-5Z,13E-dienoic acid (U46619 (10 microM, thromboxane A2 receptor agonist)). Co-incubation of U46619 together with EGF or PDGF-BB resulted in a marked amplification of [3H]thymidine incorporation that was completely reversed by picotamide. The drug also inhibited smooth muscle cell migration induced by fibrinogen (600 microg/ml) or PDGF-BB (20 ng/ml) in a concentration-dependent manner. The ability of picotamide to interfere with myocyte migration and proliferation confers, at least in vitro, a pharmacological interest on the compound in atherogenesis.

  3. Lidocaine inhibits the proliferation of lung cancer by regulating the expression of GOLT1A.

    PubMed

    Zhang, Lei; Hu, Rong; Cheng, Yanyong; Wu, Xiaoyang; Xi, Siwei; Sun, Yu; Jiang, Hong

    2017-10-01

    Lidocaine is the most commonly used local anaesthetic in clinical and can inhibit proliferation, suppress invasion and migration and induce apoptosis in human lung adenocarcinoma (LAD) cells. However, its specific downstream molecular mechanism is unclear. LAD cell lines, A549 and H1299 cells, were treated with lidocaine. The proliferation was evaluated by the methylthiazolyldiphenyl-tetrazolium bromide (MTT) and bromodeoxyuridine (BrdU) assay. The expression level of related proteins was detected by real-time quantitative PCR (qPCR) and Western blot assay. The results indicated that lidocaine dose-dependently suppressed the proliferation of A549 and H1299 cells. In the LAD patients' samples, GOLT1A was upregulated and involved in the poor prognosis and higher grade malignancy. Additionally, GOLT1A mediates the function of lidocaine on repressing proliferation by regulating the cell cycle in A549 cells. Our findings suggest that lidocaine downregulates the GOLT1A expression to repress the proliferation of lung cancer cells. © 2017 John Wiley & Sons Ltd.

  4. Stimulation of Neural Stem Cell Proliferation by Inhibition of Phosphodiesterase 5

    PubMed Central

    Santos, Ana I.; Carreira, Bruno P.; Nobre, Rui J.; Carvalho, Caetana M.; Araújo, Inês M.

    2014-01-01

    The involvement of nitric oxide (NO) and cyclic GMP (cGMP) in neurogenesis has been progressively unmasked over the last decade. Phosphodiesterase 5 (PDE5) specifically degrades cGMP and is highly abundant in the mammalian brain. Inhibition of cGMP hydrolysis by blocking PDE5 is a possible strategy to enhance the first step of neurogenesis, proliferation of neural stem cells (NSC). In this work, we have studied the effect on cell proliferation of 3 inhibitors with different selectivity and potency for PDE5, T0156, sildenafil, and zaprinast, using subventricular zone-(SVZ-) derived NSC cultures. We observed that a short- (6 h) or a long-term (24 h) treatment with PDE5 inhibitors increased SVZ-derived NSC proliferation. Cell proliferation induced by PDE5 inhibitors was dependent on the activation of the mitogen-activated protein kinase (MAPK) and was abolished by inhibitors of MAPK signaling, soluble guanylyl cyclase, and protein kinase G. Moreover, sildenafil neither activated ERK1/2 nor altered p27Kip1 levels, suggesting the involvement of pathways different from those activated by T0156 or zaprinast. In agreement with the present results, PDE5 inhibitors may be an interesting therapeutic approach for enhancing the proliferation stage of adult neurogenesis. PMID:24550991

  5. Rosiglitazone inhibits migration, proliferation, and phenotypic differentiation in cultured human lung fibroblasts.

    PubMed

    Lin, Qing; Fang, Li-Ping; Zhou, Wei-Wei; Liu, Xin-Min

    2010-03-01

    Recent studies have indicated that peroxisome proliferator-activated receptor gamma (PPARgamma) is capable of modulating inflammation, which prompted us to investigate the potential of PPARgamma ligands as lung protective agents in pulmonary fibrosis. The present study was undertaken to investigate the effects of rosiglitazone (RSG), a highly potent ligand of PPARgamma, on migration, proliferation, and phenotypic differentiation of human lung fibroblasts (MRC-5) and to explore its potential for therapy of pulmonary fibrosis. The cell migration potential was observed in a scratch wound model. Cell proliferation was determined by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) method, immunocytochemical staining, and flow cytometry, and protein expression by Western blot analysis. RSG slowed cell migration distance induced by fetal bovine serum (FBS), decreased cell proliferation initiated by FBS or platelet-derived growth factor-BB (PDGF-BB), and decreased alpha-smooth muscle actin (alpha-SMA) protein expression induced by transforming growth factor-beta1 (TGF-beta1). In addition, RSG incubation reduced the ratio of phospho-extracellular signal-regulated kinases 1/2 (p-ERK1/2) to ERK1/2 expression stimulated by FBS, PDGF-BB, and TGF-beta1. These findings show that RSG treatment inhibits lung fibroblast migration and proliferation and myofibroblast transdifferentiation stimulated by FBS and growth factors in vitro, which suggests that PPARgamma agonists could antagonize pulmonary fibrosis and have potential for therapeutic application in pulmonary fibrosis.

  6. Compound Radix Sophorae Flavescentis exerts antitumor effects by inhibiting the proliferation and inducing the apoptosis of esophageal carcinoma TE-8 cells.

    PubMed

    Yang, Xiaoyu; Cai, Weimei; Yang, Qinghui; Lu, Zhihong; Li, Jinsong; Yu, Jian

    2015-10-01

    The aim of this study was to examine the effects of compound Radix Sophorae Flavescentis on the proliferation of esophageal carcinoma TE-8 cells and to elucidate the mechanisms involved. For this purpose, we incubated TE-8 cells in medium containing various concentrations (0, 0.0125, 0.025, 0.05, 0.1, 0.2, 0.4 and 0.8 mg/ml) of the compound Radix Sophorae Flavescentis injection and its effects on the proliferation of TE-8 cells were examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. In addition, we observed the morphological changes and measured the expression levels of apoptosis-related genes (caspase-3, Bcl-2 and Bax) in the cells treated with different doses of the compound (low-dose group, 0.05 mg/ml; medium-dose group, 0.2 mg/ml; and high-dose group, 0.8 ng/ml) by reverse transcription-quantitative PCR (RT-qPCR). The apoptotic index of the cancer cells treated with different doses of the compound was determined by TUNEL assay. Our results revealed that compared with the control group (untreated cells), the proliferation of the cancer cells treated with the compound was significantly inhibited (P≤0.05); the inhibition of the proliferation of the cancer cells occured in a dose-dependent manner. Compared with the control group, the apoptotic rate of the cells in the low-dose, medium-dose and high-dose groups increased significantly (P<0.05) in a dose-dependent manner. In addition, compared with the control group, the mRNA expression of caspase-3 and Bax increased significantly in the cells treated with the compound. However, the mRNA expression of Bcl-2 markedly decreased (P<0.05). With the gradual increase in the drug concentration, the mRNA expression levels of caspase-3, Bcl-2 and Bax in the cancer cells were altered in a dose-dependent manner. In conclusion, our data demonstrate that compound Radix Sophorae Flavescentis injection significantly enhances the expression of pro-apoptotic genes in esophageal carcinoma TE-8

  7. Inhibition and Avoidance of mRNA Degradation by RNA Viruses

    PubMed Central

    Moon, Stephanie L.; Barnhart, Michael D.; Wilusz, Jeffrey

    2012-01-01

    The cellular mRNA decay machinery plays a major role in regulating the quality and quantity of gene expression in cells. This machinery involves multiple enzymes and pathways that converge to promote the exonucleolytic decay of mRNAs. The transcripts made by RNA viruses are susceptible to degradation by this machinery and, in fact, can be actively targeted. Thus, to maintain gene expression and replication, RNA viruses have evolved a number of strategies to avoid and/or inactivate aspects of the cellular mRNA decay machinery. Recent work uncovering the mechanisms used by RNA viruses to maintain the stability of their transcripts is described below. PMID:22626865

  8. Peroxisome proliferator-activated receptor gamma (PPARγ) in yellow catfish Pelteobagrus fulvidraco: molecular characterization, mRNA expression and transcriptional regulation by insulin in vivo and in vitro.

    PubMed

    Zheng, Jia-Lang; Zhuo, Mei-Qin; Luo, Zhi; Pan, Ya-Xiong; Song, Yu-Feng; Huang, Chao; Zhu, Qing-Ling; Hu, Wei; Chen, Qi-Liang

    2015-02-01

    Peroxisome proliferator-activated receptor gamma (PPARγ) is ligand-inducible transcription factor and has important roles in lipid metabolism, cell proliferation and inflammation. In the present study, yellow catfish Pelteobagrus fulvidraco PPARγ cDNA was isolated from liver by RT-PCR and RACE, and its molecular characterization and transcriptional regulation by insulin in vivo and in vitro were determined. The generation of PPARγ1 and PPARγ2 was due to alternative promoter of PPARγ gene. PPARγ1 and PPARγ2 mRNA covered 2426 bp and 2537 bp, respectively, with an open reading frame (ORF) of 1584 bp encoding 527 amino acid residues. Yellow catfish PPARγ gene was organized in a manner similar to that of their mammalian homologs, implying a modular organization of the protein's domains. A comparison between the yellow catfish PPARγ amino acid sequence and the correspondent sequences of several other species revealed the identity of 55-76.2%. Two PPARγ transcripts (PPARγ1 and PPARγ2) mRNAs were expressed in a wide range of tissues, but the abundance of each PPARγ mRNA showed the tissue- and developmental stage-dependent expression patterns. Intraperitoneal injection of insulin in vivo significantly stimulated the mRNA expression of total PPARγ and PPARγ1, but not PPARγ2 in the liver of yellow catfish. In contrast, incubation of hepatocytes with insulin in vitro increased the mRNA levels of PPARγ1, PPARγ2 and total PPARγ. To our knowledge, for the first time, the present study provides evidence that PPARγ1 and PPARγ2 are differentially expressed with and among tissues during different developmental stages and also regulated by insulin both in vivo and in vitro, which serves to increase our understanding on PPARγ physiological function in fish.

  9. The sGC activator inhibits the proliferation and migration, promotes the apoptosis of human pulmonary arterial smooth muscle cells via the up regulation of plasminogen activator inhibitor-2

    SciTech Connect

    Zhang, Shuai; Zou, Lihui; Yang, Ting; Yang, Yuanhua; Zhai, Zhenguo; Xiao, Fei; Wang, Chen

    2015-03-15

    Background: Different types of pulmonary hypertension (PH) share the same process of pulmonary vascular remodeling, the molecular mechanism of which is not entirely clarified by far. The abnormal biological behaviors of pulmonary arterial smooth muscle cells (PASMCs) play an important role in this process. Objectives: We investigated the regulation of plasminogen activator inhibitor-2 (PAI-2) by the sGC activator, and explored the effect of PAI-2 on PASMCs proliferation, apoptosis and migration. Methods: After the transfection with PAI-2 overexpression vector and specific siRNAs or treatment with BAY 41-2272 (an activator of sGC), the mRNA and protein levels of PAI-2 in cultured human PASMCs were detected, and the proliferation, apoptosis and migration of PASMCs were investigated. Results: BAY 41-2272 up regulated the endogenous PAI-2 in PASMCs, on the mRNA and protein level. In PAI-2 overexpression group, the proliferation and migration of PASMCs were inhibited significantly, and the apoptosis of PASMCs was increased. In contrast, PAI-2 knockdown with siRNA increased PASMCs proliferation and migration, inhibited the apoptosis. Conclusions: PAI-2 overexpression inhibits the proliferation and migration and promotes the apoptosis of human PASMCs. Therefore, sGC activator might alleviate or reverse vascular remodeling in PH through the up-regulation of PAI-2. - Highlights: • sGC activator BAY41-2272 up regulated PAI-2 in PASMCs, on the mRNA and protein level. • PAI-2 overexpression inhibits the proliferation and migration of human PASMCs. • PAI-2 overexpression promotes the apoptosis of human PASMCs. • sGC activator might alleviate the vascular remodeling in pulmonary hypertension.

  10. D-Glucosamine inhibits proliferation of human cancer cells through inhibition of p70S6K

    SciTech Connect

    Oh, Hyun-Ji; Lee, Jason S.; Song, Dae-Kyu; Shin, Dong-Hoon; Jang, Byeong-Churl; Suh, Seong-Il; Park, Jong-Wook; Suh, Min-Ho; Baek, Won-Ki . E-mail: wonki@dsmc.or.kr

    2007-09-07

    Although D-glucosamine has been reported as an inhibitor of tumor growth both in vivo and in vitro, the mechanism for the anticancer effect of D-glucosamine is still unclear. Since there are several reports suggesting D-glucosamine inhibits protein synthesis, we examined whether D-glucosamine affects p70S6 K activity, an important signaling molecule involved in protein translation. In the present study, we found D-glucosamine inhibited the activity of p70S6K and the proliferation of DU145 prostate cancer cells and MDA-MB-231 breast cancer cells. D-Glucosamine decreased phosphorylation of p70S6K, and its downstream substrates RPS6, and eIF-4B, but not mTOR and 4EBP1 in DU145 cells, suggesting that D-glucosamine induced inhibition of p70S6K is not through the inhibition of mTOR. In addition, D-glucosamine enhanced the growth inhibitory effects of rapamycin, a specific inhibitor of mTOR. These findings suggest that D-glucosamine can inhibit growth of cancer cells through dephosphorylation of p70S6K.

  11. Neural cell adhesion molecule (N-CAM) domains and intracellular signaling pathways involved in the inhibition of astrocyte proliferation.

    PubMed

    Krushel, L A; Tai, M H; Cunningham, B A; Edelman, G M; Crossin, K L

    1998-03-03

    The neural cell adhesion molecule (N-CAM) inhibits astrocyte proliferation in vitro and in vivo, and this effect is partially reversed by the glucocorticoid antagonist RU-486. The present studies have tested the hypothesis that N-CAM-mediated inhibition of astrocyte proliferation is caused by homophilic binding and involves the activation of glucocorticoid receptors. It was observed that all N-CAM Ig domains inhibited astrocyte proliferation in parallel with their ability to influence N-CAM binding. The proliferation of other N-CAM-expressing cells also was inhibited by the addition of N-CAM. In contrast, the proliferation of astrocytes from knockout mice lacking N-CAM was not inhibited by added N-CAM. These findings support the hypothesis that it is binding of soluble N-CAM to N-CAM on the astrocyte surface that leads to decreased proliferation. Signaling pathways stimulated by growth factors include activation of mitogen-activated protein (MAP) kinase. Addition of N-CAM inhibited MAP kinase activity induced by basic fibroblast growth factor in astrocytes. In accord with previous findings that RU-486 could partially prevent the proliferative effects of N-CAM, inhibition of MAP kinase activity by N-CAM was reversed by RU-486. The ability of N-CAM to inhibit astrocyte proliferation was unaffected, however, by agents that block the ability of N-CAM to promote neurite outgrowth. Together, these findings indicate that homophilic N-CAM binding leads to inhibition of astrocyte proliferation via a pathway involving the glucocorticoid receptor and that the ability of N-CAM to influence astrocyte proliferation and neurite outgrowth involves different signal pathways.

  12. Synergistic Inhibition of Endothelial Cell Proliferation, Tube Formation, and Sprouting by Cyclosporin A and Itraconazole

    PubMed Central

    Nacev, Benjamin A.; Liu, Jun O.

    2011-01-01

    Pathological angiogenesis contributes to a number of diseases including cancer and macular degeneration. Although angiogenesis inhibitors are available in the clinic, their efficacy against most cancers is modest due in part to the existence of alternative and compensatory signaling pathways. Given that angiogenesis is dependent on multiple growth factors and a broad signaling network in vivo, we sought to explore the potential of multidrug cocktails for angiogenesis inhibition. We have screened 741 clinical drug combinations for the synergistic inhibition of endothelial cell proliferation. We focused specifically on existing clinical drugs since the re-purposing of clinical drugs allows for a more rapid and cost effective transition to clinical studies when compared to new drug entities. Our screen identified cyclosporin A (CsA), an immunosuppressant, and itraconazole, an antifungal drug, as a synergistic pair of inhibitors of endothelial cell proliferation. In combination, the IC50 dose of each drug is reduced by 3 to 9 fold. We also tested the ability of the combination to inhibit endothelial cell tube formation and sprouting, which are dependent on two essential processes in angiogenesis, endothelial cell migration and differentiation. We found that CsA and itraconazole synergistically inhibit tube network size and sprout formation. Lastly, we tested the combination on human foreskin fibroblast viability as well as Jurkat T cell and HeLa cell proliferation, and found that endothelial cells are selectively targeted. Thus, it is possible to combine existing clinical drugs to synergistically inhibit in vitro models of angiogenesis. This strategy may be useful in pursuing the next generation of antiangiogenesis therapy. PMID:21969860

  13. MicroRNA-217 suppresses homocysteine-induced proliferation and migration of vascular smooth muscle cells via N-methyl-D-aspartic acid receptor inhibition.

    PubMed

    Duan, Hongyan; Li, Yongqiang; Yan, Lijie; Yang, Haitao; Wu, Jintao; Qian, Peng; Li, Bing; Wang, Shanling

    2016-10-01

    Hyperhomocysteine has become a critical risk for atherosclerosis and can stimulate proliferation and migration of vascular smooth muscle cells (VSMCs). N-methyl-D-aspartic acid receptor (NMDAR) is a receptor of homocysteine and mediates the effects of homocysteine on VSMCs. Bioinformatics analysis has shown NMDAR is a potential target of microRNA-217 (miR-217), which exerts multiple functions in cancer tumorigenesis and carotid plaque progression. In this study, we sought to investigate the role of miR-217 in VSMCs phenotype transition under homocysteine exposure and elucidate its effect on atherosclerotic plaque formation. After treating with several doses of homocysteine (0-8 × 10(-4)  mol/L) for 24 hours, the expression of miR-217 in HA-VSMCs and rat aortic VSMCs was not altered. Intriguingly, the expression of NMDAR mRNA and protein was reduced by homocysteine in a dose-dependent manner. Transfection of miR-217 mimic significantly inhibited the proliferation and migration of VSMCs with homocysteine treatment, while transfection of miR-217 inhibitor promoted VSMCs migration. Moreover, miR-217 mimic down-regulated while miR-217 inhibitor up-regulated NMDAR protein expression but not NMDAR mRNA expression. Through luciferase reporter assay, we showed that miR-217 could directly bind to the 3'-UTR of NMDAR. MiR-217 mimic transfection also released the inhibition of cAMP-response element-binding protein (CREB)-PGC-1α signalling induced by homocysteine. Additionally, restoration of PGC-1α expression via AdPGC-1α infection markedly suppressed VSMCs proliferation through the degradation of NADPH oxidase (NOX1) and reduction of reactive oxygen species (ROS). Collectively, our study identified the role of miR-217 in regulating VSMCs proliferation and migration, which might serve as a target for atherosclerosis therapy.

  14. MicroRNA-7 inhibits cell proliferation, migration and invasion in human non-small cell lung cancer cells by targeting FAK through ERK/MAPK signaling pathway

    PubMed Central

    Shi, Yu-Jia; Chen, Yi; Sun, Yun; Zhang, Qian; Song, Lei; Peng, Li-Ping

    2016-01-01

    Objective To investigate the effects of microRNA-7 (miR-7) on the proliferation, migration and invasion of non-small cell lung cancer NSCLC) cells by targeting FAK through ERK/MAPK signaling pathway. Methods NSCLC tissues and adjacent normal tissues were obtained from 160 NSCLC patients after operation. NSCLC cell lines (A549, H1299 and H1355) and a normal human fetal lung fibroblast cell line (MRC-5) were obtained. NSCLC cells were assigned into miR-7 inhibitors, miR-7 mimics, blank, miR-7 mimics control, miR-7 inhibitors control, FAK siRNA and miR-7 inhibitors + FAK siRNA groups. The expressions of miR-7 and FAK mRNA in tissues and cell lines were detected by qRT-PCR and Western-Blotting. Cell proliferation, migration and invasion were detected by MTT assay, wound scratch assay and Transwell assay. Results Compared with adjacent normal tissues, miR-7 expression was down-regulated, but the mRNA and protein expressions of FAK, ERK and MAPK were up-regulated. Compared with the blank and mimics control groups, miR-7 significantly increased but FAK, ERK and MAPK expressions decreased in miR-7 mimics and FAK siRNA groups. Cell proliferation, migration and invasion were inhibited in the miR-7 mimics and FAK siRNA groups, while opposite regarding miR-7 inhibitors group. Conclusion The miR-7 can inhibit the activation of ERK/MAPK signaling pathway by down-regulating FAK expression, thereby suppressing the proliferation, migration and invasion of NSCLC cells. The miR-7 and its target gene FAK may be novel targets for the diagnosis and treatment of NSCLC. PMID:27764812

  15. Iptakalim inhibits PDGF-BB-induced human airway smooth muscle cells proliferation and migration

    SciTech Connect

    Liu, Wenrui; Kong, Hui; Zeng, Xiaoning; Wang, Jingjing; Wang, Zailiang; Yan, Xiaopei; Wang, Yanli; Xie, Weiping Wang, Hong

    2015-08-15

    Chronic airway diseases are characterized by airway remodeling which is attributed partly to the proliferation and migration of airway smooth muscle cells (ASMCs). ATP-sensitive potassium (K{sub ATP}) channels have been identified in ASMCs. Mount evidence has suggested that K{sub ATP} channel openers can reduce airway hyperresponsiveness and alleviate airway remodeling. Opening K{sup +} channels triggers K{sup +} efflux, which leading to membrane hyperpolarization, preventing Ca{sup 2+}entry through closing voltage-operated Ca{sup 2+} channels. Intracellular Ca{sup 2+} is the most important regulator of muscle contraction, cell proliferation and migration. K{sup +} efflux decreases Ca{sup 2+} influx, which consequently influences ASMCs proliferation and migration. As a K{sub ATP} channel opener, iptakalim (Ipt) has been reported to restrain the proliferation of pulmonary arterial smooth muscle cells (PASMCs) involved in vascular remodeling, while little is known about its impact on ASMCs. The present study was designed to investigate the effects of Ipt on human ASMCs and the mechanisms underlying. Results obtained from cell counting kit-8 (CCK-8), flow cytometry and 5-ethynyl-2′-deoxyuridine (EdU) incorporation showed that Ipt significantly inhibited platelet-derived growth factor (PDGF)-BB-induced ASMCs proliferation. ASMCs migration induced by PDGF-BB was also suppressed by Ipt in transwell migration and scratch assay. Besides, the phosphorylation of Ca{sup 2+}/calmodulin-dependent kinase II (CaMKII), extracellular regulated protein kinases 1/2 (ERK1/2), protein kinase B (Akt), and cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) were as well alleviated by Ipt administration. Furthermore, we found that the inhibition of Ipt on the PDGF-BB-induced proliferation and migration in human ASMCs was blocked by glibenclamide (Gli), a selective K{sub ATP} channel antagonist. These findings provide a strong evidence to support that Ipt

  16. Inhibition of CYP3A mRNA and protein expression, and enzymatic activity, by enrofloxacin in chickens.

    PubMed

    Hu, H-F; Gao, Z-X; Cheng, Y-Y

    2010-12-01

    This study was to investigate the effect of enrofloxacin (EF) on CYP3A in chicken by using quantitative reverse transcription-polymerase chain reaction and immunodetected. The treated chickens were given 5, 25 and 125 mg/kg of EF while the control chickens were treated with the same volume saline. There was no significant difference between the low dose group and controls in the concentration of hepatic microsome protein and total CYP content, while the middle and high dose EF caused the down regulation. Depression of the CYP3A activity, mRNA and protein were observed in treated chickens, and the inhibition degree was different from each group. It was concluded that EF caused the inhibition of CYP3A both in genetic transcription and protein levels. But the inhibition metabolism still needs further researches. © 2010 Blackwell Publishing Ltd.

  17. Endometrial cancer cells exhibit high expression of p110β and its selective inhibition induces variable responses on PI3K signaling, cell survival and proliferation

    PubMed Central

    Karlsson, Thomas; Krakstad, Camilla; Tangen, Ingvild Løberg; Hoivik, Erling A.; Pollock, Pamela M.; Lewis, Aurélia E.

    2017-01-01

    PTEN loss and constitutive activation of the class I phosphoinositide 3-kinase (PI3K) pathway are key drivers of endometrial tumorigenesis. In some cancer types, PTEN-deficient tumors are reliant on class I PI3K p110β (encoded by PIK3CB) activity but little is known about this contribution in endometrial tumorigenesis. In this study, we find that p110β is overexpressed in a panel of 7 endometrial cancer cell lines compared to non-transformed cells. Furthermore, in 234 clinically annotated patient samples, PIK3CB mRNA levels increase significantly in the early phase of tumorigenesis from precursors to low grade primary malignant lesions whereas PIK3CA levels are higher in non-endometrioid compared to endometrioid primary tumors. While high levels of either PIK3CA or PIK3CB associate with poor prognosis, only elevated PIK3CB mRNA levels correlate with a high cell cycle signature score in clinical samples. In cancer cell lines, p110α inhibition reduces cell viability by inducing cell death in PIK3CA mutant cells while p110β inhibition delayed proliferation in PTEN-deficient cells, but not in WT cells. Taken together, our findings suggest that PIK3CB/p110β contributes to some of the pleiotropic functions of PI3K in endometrial cancer, particularly in the early steps by contributing to cell proliferation. PMID:28002804

  18. THOC5, a member of the mRNA export complex: a novel link between mRNA export machinery and signal transduction pathways in cell proliferation and differentiation.

    PubMed

    Tran, Doan D H; Koch, Alexandra; Tamura, Teruko

    2014-01-10

    Cell growth, differentiation, and commitment to a restricted lineage are guided by a timely expressed set of growth factor/cytokine receptors and their down-stream transcription factor genes. Transcriptional control mechanisms of gene expression during differentiation have been mainly studied by focusing on the cis- and trans-elements in promoters however, the role of mRNA export machinery during differentiation has not been adequately examined. THO (Suppressors of the transcriptional defects of hpr1 delta by overexpression) complex 5 (THOC5) is a member of THO complex which is a subcomplex of the transcription/export complex (TREX). THOC5 is evolutionarily conserved in higher eukaryotes, however the exact roles of THOC5 in transcription and mRNA export are still unclear. In this review, we focus on recently uncovered aspects of the role of THOC5 in signal transduction induced by extracellular stimuli. THOC5 is phosphorylated by several protein kinases at multiple residues upon extracellular stimuli. These include stimulation with growth factors/cytokines/chemokines, or DNA damage reagents. Furthermore, THOC5 is a substrate for several oncogenic tyrosine kinases, suggesting that THOC5 may be involved in cancer development. Recent THOC5 knockout mouse data reveal that THOC5 is an essential element in the maintenance of stem cells and growth factor/cytokine-mediated differentiation/proliferation. Furthermore, depletion of THOC5 influences less than 1% of total mRNA export in the steady state, however it influences more than 90% of growth factor/cytokine induced genes. THOC5, thereby contributes to the 3' processing and/or export of immediate-early genes induced by extracellular stimuli. These studies bring new insight into the link between the mRNA export complex and immediate-early gene response. The data from these studies also suggest that THOC5 may be a useful tool for studying stem cell biology, for modifying the differentiation processes and for cancer therapy.

  19. [Aliskiren inhibits proliferation of cardiac fibroblasts in AGT-REN double transgenic hypertensive mice in vitro].

    PubMed

    Wang, Li-Ping; Fan, Su-Jing; Li, Shu-Min; Wang, Xiao-Jun; Sun, Na

    2016-10-25

    The purpose of the present study is to explore the effect of aliskiren on the proliferation of cardiac fibroblasts (CFs) in AGT-REN double transgenic hypertensive (dTH) mice. The cultured CFs from AGT-REN dTH mice were divided into AGT-REN group (dTH) and aliskiren group (ALIS). Cultured CFs from C57B6 mice were served as control (WT). The effect of different concentration of aliskiren (1 × 10(-6), 1 × 10(-7), 1 × 10(-8), 1 × 10(-9) mol/L) on CFs proliferation was determined by MTT assay. After treatment with 1 × 10(-7) mol/L aliskiren for 24 h, α-SMA, collagen I, III and NADPH oxidase (NOX) protein expression in CFs of AGT-REN dTH mice were detected by Western blot. The collagen synthesis in CFs was assessed by hydroxyproline kit. The expression of ROS was determined by DHE. Results showed that the blood pressure and plasma Ang II levels were significantly increased and CFs proliferation was significantly increased as well in AGT-REN dTH mice compared with WT group. However, aliskiren intervention decreased CFs proliferation, myofibroblast transformation, as well as the collagen I and III synthesis in CFs of AGT-REN dTH mice. Meanwhile, aliskiren inhibited ROS content and NOX2/NOX4 protein expression in CFs of AGT-REN dTH mice. These results suggest that aliskiren decreases the cell proliferation, myofibroblast transformation and collagen production in CFs of AGT-REN dTH mice, which might be through inhibition of oxidative stress response.

  20. Functional inhibition of aquaporin-3 with a gold-based compound induces blockage of cell proliferation.

    PubMed

    Serna, Ana; Galán-Cobo, Ana; Rodrigues, Claudia; Sánchez-Gomar, Ismael; Toledo-Aral, Juan José; Moura, Teresa F; Casini, Angela; Soveral, Graça; Echevarría, Miriam

    2014-11-01

    AQP3 has been correlated with higher transport of glycerol, increment of ATP content, and larger proliferation capacity. Recently, we described the gold(III) complex Auphen as a very selective and potent inhibitor of AQP3's glycerol permeability (Pgly ). Here we evaluated Auphen effect on the proliferation of various mammalian cell lines differing in AQP3 expression level: no expression (PC12), moderate (NIH/3T3) or high (A431) endogenous expression, cells stably expressing AQP3 (PC12-AQP3), and human HEK293T cells transiently transfected (HEK-AQP3) for AQP3 expression. Proliferation was evaluated in the absence or presence of Auphen (5 μM) by counting number of viable cells and analyzing 5-bromo-2'-deoxyuridine (BrdU) incorporation. Auphen reduced ≈50% the proliferation in A431 and PC12-AQP3, ≈15% in HEK-AQP3 and had no effect in PC12-wt and NIH/3T3. Strong arrest in the S-G2/M phases of the cell cycle, supported by analysis of cyclins (A, B1, D1, E) levels, was observed in AQP3-expressing cells treated with Auphen. Flow-cytometry of propidium iodide incorporation and measurements of mitochondrial dehydrogenases activity confirmed absence of cytotoxic effect of the drug. Functional studies evidenced ≈50% inhibition of A431 Pgly by Auphen, showing that the compound's antiproliferative effect correlates with its ability to inhibit AQP3 Pgly . Role of Cys-40 on AQP3 permeability blockage by Auphen was confirmed by analyzing the mutated protein (AQP3-Ser-40). Accordingly, cells transfected with mutated AQP3 gained resistance to the antiproliferative effect of Auphen. These results highlight an Auphen inhibitory effect on proliferation of cells expressing AQP3 and suggest a targeted therapeutic effect on carcinomas with large AQP3 expression.

  1. Carvedilol inhibits proliferation of cultured pulmonary artery smooth muscle cells of patients with idiopathic pulmonary arterial hypertension.

    PubMed

    Fujio, Hideki; Nakamura, Kazufumi; Matsubara, Hiromi; Kusano, Kengo Fukushima; Miyaji, Katsumasa; Nagase, Satoshi; Ikeda, Tetsuya; Ogawa, Aiko; Ohta-Ogo, Keiko; Miura, Daiji; Miura, Aya; Miyazaki, Masahiro; Date, Hiroshi; Ohe, Tohru

    2006-02-01

    Idiopathic pulmonary arterial hypertension (IPAH) is associated with proliferation of smooth muscle cells (SMCs) in small pulmonary arteries. Inhibition of proliferation of pulmonary artery smooth muscle cells (PASMCs) may be an effective treatment of patients with idiopathic pulmonary arterial hypertension. Recent studies have shown that carvedilol, an alpha- and beta-blocker with antioxidant and calcium channel blocking properties, inhibits the proliferation of cultured normal human pulmonary artery smooth muscle cells. In this study, we tested the hypothesis that carvedilol has antiproliferative effects on pulmonary artery smooth muscle cells of patients with idiopathic pulmonary arterial hypertension. Pulmonary artery smooth muscle cells from six idiopathic pulmonary arterial hypertension patients who had undergone lung transplantation were cultured. To determine cell proliferation, H-thymidine incorporation was measured. Platelet-derived growth factor-induced proliferation of IPAH-PASMCs was significantly greater than that of normal control pulmonary artery smooth muscle cells. Carvedilol (0.1 microM to 10 microM) inhibited the proliferation of idiopathic pulmonary arterial hypertension-pulmonary artery smooth muscle cells in a concentration-dependent manner. Prazosin (an alpha-blocker) and N-acetyl L cysteine (an antioxidant agent) (0.1 microM to 10 microM) did not inhibit their proliferation, but the high concentration of propranolol (a beta-blocker) and nifedipine (a calcium channel blocker) (10 microM) inhibited the proliferation. The combination of propranolol and nifedipine inhibited the proliferation but only at a high concentration (10 microM) combination. Cell cycle analysis revealed that carvedilol (10 microM) significantly decreased the number of cells in S and G2/M phases. These results indicate that carvedilol inhibits the exaggerated proliferation of pulmonary artery smooth muscle cells of patients with idiopathic pulmonary arterial hypertension

  2. Metformin: Direct Inhibition of Rat Ovarian Theca-Interstitial Cell Proliferation

    PubMed Central

    Will, Matthew A.; Palaniappan, Murugesan; Peegel, Helle; Kayampilly, Pradeep; Menon, K.M.J.

    2012-01-01

    Study Objective To determine if metformin has direct effects on ovarian theca-interstitial cell proliferation through activation of AMP-activated protein kinase (AMPK). Design In vitro experimental study. Setting Academic medical center laboratory. Animal(s) Immature Sprague-Dawley female rats Interventions Ovarian theca-interstitial (T-I) cells were isolated, purified and cultured in the absence (control) or presence of insulin (1mcg/mL) with or without metformin or other activators/inhibitors of AMPK (AICAR, Compound C). Main outcome measure(s) Proliferation was assessed by determination of expression levels of proteins involved in cell cycle progression, cyclin D3 and cyclin-dependent kinase 4 (CDK4) with Western blot analysis, and determination of DNA synthesis with bromodeoxyuridine (BrdU) incorporation assay. Activation of AMPK, Erk1/2 and S6K1 was determined by Western blot analysis with the use of antibodies specific for the phosphorylated (activated) forms. Results Metformin inhibited insulin-induced ovarian T-I cell proliferation and upregulation of cell cycle regulatory proteins, cyclin D3 and CDK4. Metformin independently activated AMPK in a dose-dependent manner. Treatment with metformin inhibited insulin-induced activation of Erk1/2 and S6K1. This effect was reversed with the addition of compound C, a known AMPK inhibitor. Conclusions Metformin directly inhibits proliferation of ovarian theca-interstitial cells via an AMPK-dependent mechanism. Present findings further validate potential benefits of metformin in the treatment of conditions associated with hyperinsulinemia and excessive growth of ovarian T-I cells (such as PCOS). PMID:22608319

  3. Phytochemical-induced nucleolar stress results in the inhibition of breast cancer cell proliferation.

    PubMed

    Lewinska, Anna; Bednarz, Diana; Adamczyk-Grochala, Jagoda; Wnuk, Maciej

    2017-03-14

    The nucleolus is a stress sensor and compromised nucleolar activity may be considered as an attractive anticancer strategy. In the present study, the effects of three plant-derived natural compounds, i.e., sulforaphane (SFN), ursolic acid (UA) and betulinic acid (BA) on nucleolar state were investigated in breast cancer cell lines of different receptor status, namely MCF-7, MDA-MB-231 and SK-BR-3 cells. Cytostatic action of phytochemicals against breast cancer cells was observed at low micromolar concentration window (5-20µM) and mediated by elevated p21 levels, and cell proliferation of SFN-, UA- and BA-treated normal human mammary epithelial cells (HMEC) was unaffected. Phytochemical-mediated inhibition of cell proliferation was accompanied by increased levels of superoxide and protein carbonylation that lead to disorganization of A- and B-type lamin networks and alterations in the nuclear architecture. Phytochemicals promoted nucleolar stress as judged by the nucleoplasmic translocation of RNA polymerase I-specific transcription initiation factor RRN3/TIF-IA, inhibition of new rRNA synthesis and decrease in number of nucleoli. Phytochemicals also decreased the levels of NOP2, proliferation-associated nucleolar protein p120, and WDR12 required for maturation of 28S and 5.8S ribosomal RNAs and formation of the 60S ribosome, and phosphorylation of S6 ribosomal protein that may result in diminished translation and inhibition of cell proliferation. In summary, three novel ribotoxic stress stimuli were revealed with a potential to be used in nucleolus-focused anticancer therapy.

  4. Antibiotic drug tigecycline inhibited cell proliferation and induced autophagy in gastric cancer cells

    SciTech Connect

    Tang, Chunling; Yang, Liqun; Jiang, Xiaolan; Xu, Chuan; Wang, Mei; Wang, Qinrui; Zhou, Zhansong; Xiang, Zhonghuai; Cui, Hongjuan

    2014-03-28

    Highlights: • Tigecycline inhibited cell growth and proliferation in human gastric cancer cells. • Tigecycline induced autophagy not apoptosis in human gastric cancer cells. • AMPK/mTOR/p70S6K pathway was activated after tigecycline treatment. • Tigecycline inhibited tumor growth in xenograft model of human gastric cancer cells. - Abstract: Tigecycline acts as a glycylcycline class bacteriostatic agent, and actively resists a series of bacteria, specifically drug fast bacteria. However, accumulating evidence showed that tetracycline and their derivatives such as doxycycline and minocycline have anti-cancer properties, which are out of their broader antimicrobial activity. We found that tigecycline dramatically inhibited gastric cancer cell proliferation and provided an evidence that tigecycline induced autophagy but not apoptosis in human gastric cancer cells. Further experiments demonstrated that AMPK pathway was activated accompanied with the suppression of its downstream targets including mTOR and p70S6K, and ultimately induced cell autophagy and inhibited cell growth. So our data suggested that tigecycline might act as a candidate agent for pre-clinical evaluation in treatment of patients suffering from gastric cancer.

  5. Myostatin inhibits cell proliferation and protein synthesis in C2C12 muscle cells.

    PubMed

    Taylor, W E; Bhasin, S; Artaza, J; Byhower, F; Azam, M; Willard, D H; Kull, F C; Gonzalez-Cadavid, N

    2001-02-01

    Myostatin mutations in mice and cattle are associated with increased muscularity, suggesting that myostatin is a negative regulator of skeletal muscle mass. To test the hypothesis that myostatin inhibits muscle cell growth, we examined the effects of recombinant myostatin in mouse skeletal muscle C2C12 cells. After verification of the expression of cDNA constructs in a cell-free system and in transfected Chinese hamster ovary cells, the human recombinant protein was expressed as the full-length (375-amino acid) myostatin in Drosophila cells (Mst375D), or the 110-amino acid carboxy-terminal protein in Escherichia coli (Mst110EC). These proteins were identified by immunoblotting and were purified. Both Mst375D and Mst110EC dose dependently inhibited cell proliferation (cell count and Formazan assay), DNA synthesis ([3H]thymidine incorporation), and protein synthesis ([1-14C]leucine incorporation) in C2C12 cells. The inhibitory effects of both proteins were greater in myotubes than in myoblasts. Neither protein had any significant effects on protein degradation or apoptosis. In conclusion, recombinant myostatin proteins inhibit cell proliferation, DNA synthesis, and protein synthesis in C2C12 muscle cells, suggesting that myostatin may control muscle mass by inhibiting muscle growth or regeneration.

  6. MiRNA-125a-5p inhibits glioblastoma cell proliferation and promotes cell differentiation by targeting TAZ

    SciTech Connect

    Yuan, Jian; Xiao, Gelei; Peng, Gang; Liu, Dingyang; Wang, Zeyou; Liao, Yiwei; Liu, Qing; Wu, Minghua; Yuan, Xianrui

    2015-02-06

    Highlights: • Expression of miR-125a-5p is inversely correlated with that of TAZ in glioma cells. • MiR-125a-5p represses TAZ expression in glioma cells. • MiR-125a-5p directly targets the 3′ UTR of TAZ mRNA and promotes its degradation. • MiR-125a-5p represses CTGF and survivin via TAZ, and inhibits glioma cell growth. • MiR-125a-5p inhibits the stem cell features of HFU-251 MG cells. - Abstract: Glioblastoma (GBM) is the most lethal brain tumor due to the resistance to conventional therapies, such as radiotherapy and chemotherapy. TAZ, an important mediator of the Hippo pathway, was found to be up-regulated in diverse cancers, including in GBM, and plays important roles in tumor initiation and progression. However, little is known about the regulation of TAZ expression in tumors. In this study, we found that miR-125a-5p is an important regulator of TAZ in glioma cells by directly targeting the TAZ 3′ UTR. MiR-125a-5p levels are inversely correlated with that of TAZ in normal astrocytes and a panel of glioma cell lines. MiR-125a-5p represses the expression of TAZ target genes, including CTGF and survivin, and inhibits cell proliferation and induces the differentiation of GBM cells; whereas over-expression of TAZ rescues the effects of miR-125a-5p. This study revealed a mechanism for TAZ deregulation in glioma cells, and also demonstrated a tumor suppressor role of miR-125a-5p in glioblastoma cells.

  7. The tumor suppressor miR-124 inhibits cell proliferation and invasion by targeting B7-H3 in osteosarcoma.

    PubMed

    Wang, Ling; Kang, Fu-Biao; Sun, Nan; Wang, Juan; Chen, Wei; Li, Dong; Shan, Bao-En

    2016-11-01

    Our previous studies have shown that the expression level of B7 homolog 3 (B7-H3) was correlated with clinical staging and prognosis of osteosarcoma (OS) patients, and its silencing inhibited the proliferation and invasion of OS cells in vitro. However, its overexpression mechanism behind was far from elucidated. On the basis of bioinformatics and the preliminary screening data, we hypothesized that miR-124 might play an important role in OS development and as a lead candidate for modulating B7-H3 expression. In this study, we found that miR-124 was downregulated significantly in OS tumor tissue, compared to normal adjacent tissues (NATs). Lower miR-124 expression levels were associated with advanced Ennecking stage, lower tumor differentiation, and common pulmonary metastasis. The 5-year overall survival rate in the miR-124 upregulated group was 61.5 %, while with low miR-124 expression, only 11.8 % survived. Further studies in vitro showed that B7-H3 was a direct target of miR-124. Overexpression of miR-124 decreased B7-H3 mRNA and protein level and inhibited B7-H3 3'-UTR reporter activity. Treatment of OS cells with miR-124 mimics induced the inhibition of cell growth and invasion in vitro, which could be abrogated by transfected by B7-H3 expression vector. Our findings highlight the potential application of miR-124 as a novel onco-miRNA in OS, and its oncogenic effects are mediated chiefly through downregulation of B7-H3, thus suggesting a model for identifying miR-124 that can be exploited to improve the therapeutic potential efficacy of mAb targeting to B7-H3.

  8. Kaempferol inhibits IL-1β-induced proliferation of rheumatoid arthritis synovial fibroblasts and the production of COX-2, PGE2 and MMPs.

    PubMed

    Yoon, Ha-Yong; Lee, Eun-Gyeong; Lee, Hyun; Cho, In Jin; Choi, Yun Jung; Sung, Myung-Soon; Yoo, Han-Gyul; Yoo, Wan-Hee

    2013-10-01

    Inflammatory cytokines, matrix metalloproteinases (MMPs) and cyclooxygenase (COX)-2 released from rheumatoid arthritis synovial fibroblasts (RASFs) are involved in the destruction of both articular bone and cartilage. Kaempferol has been reported to act as an antioxidant and anti-inflammatory agent by inhibiting nitric oxide synthase and COX enzymes. The aim of the present study was to determine the effects of kaempferol on the interleukin-1β (IL-1β)-induced proliferation of RASFs and the production of MMPs, COX and prostaglandin E2 (PGE2) by RASFs. The proliferation of the RASFs stimulated with IL-1β and treated with/without kaempferol was evaluated by CCK-8 assay. The expression of MMPs, TIMP metallopeptidase inhibitor-1 (TIMP-1), COXs, PGE2 and that of intracellular MAPK signaling molecules, including p-ERK, p-p38, p-JNK and nuclear factor-κB (NF-κB) was examined by immunoblotting or semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and ELISA under the conditions described above. Kaempferol inhibited the proliferation of both unstimulated and IL-1β‑stimulated RASFs, as well as the mRNA and protein expression of MMP-1, MMP-3, COX-2 and PGE2 induced by IL-1β. Kaempferol also inhibited the phosphorylation of ERK-1/2, p38 and JNK, as well as the activation of NF-κB induced by IL-1β. These results indicate that kaempferol inhibits synovial fibroblast proliferation, as well as the production of and MMPs, COX‑2 and PGE2, which is involved in articular inflammation and destruction in rheumatoid arthritis (RA). Our data suggest that kaempferol may be a novel therapeutic agent for the treatment of RA.

  9. Triptolide inhibits proliferation of Epstein–Barr virus-positive B lymphocytes by down-regulating expression of a viral protein LMP1

    SciTech Connect

    Zhou, Heng; Guo, Wei; Long, Cong; Wang, Huan; Wang, Jingchao; Sun, Xiaoping

    2015-01-16

    Highlights: • Triptolide inhibits proliferation of EBV-positive lymphoma cells in vitro and in vivo. • Triptolide reduces expression of LMP1 by decreasing its transcription level. • Triptolide inhibits ED-L1 promoter activity. - Abstract: Epstein–Barr virus (EBV) infects various types of cells and mainly establishes latent infection in B lymphocytes. The viral latent membrane protein 1 (LMP1) plays important roles in transformation and proliferation of B lymphocytes infected with EBV. Triptolide is a compound of Tripterygium extracts, showing anti-inflammatory, immunosuppressive, and anti-cancer activities. In this study, it is determined whether triptolide inhibits proliferation of Epstein–Barr virus-positive B lymphocytes. The CCK-8 assays were performed to examine cell viabilities of EBV-positive B95-8 and P3HR-1 cells treated by triptolide. The mRNA and protein levels of LMP1 were examined by real time-PCR and Western blotting, respectively. The activities of two LMP1 promoters (ED-L1 and TR-L1) were determined by Dual luciferase reportor assay. The results showed that triptolide inhibited the cell viability of EBV-positive B lymphocytes, and the over-expression of LMP1 attenuated this inhibitory effect. Triptolide decreased the LMP1 expression and transcriptional levels in EBV-positive B cells. The activity of LMP1 promoter ED-L1 in type III latent infection was strongly suppressed by triptolide treatment. In addition, triptolide strongly reduced growth of B95-8 induced B lymphoma in BALB/c nude mice. These results suggest that triptolide decreases proliferation of EBV-induced B lymphocytes possibly by a mechanism related to down-regulation of the LMP1 expression.

  10. Antisense oligodeoxynucleotide against human telomerase reverse transcriptase inhibits the proliferation of Eca-109 esophageal carcinoma cells

    PubMed Central

    FAN, XIANG-KUI; YAN, RUI-HUA; LI, BAO-JIANG; CHEN, XIANG-MING; WEI, LIN; WANG, ZHOU

    2014-01-01

    Previous studies have demonstrated that the growth of tumor cells may be inhibited by antisense oligonucleotides (ASODNs) targeted against human telomerase (hTR) or human telomerase reverse transcriptase (hTERT), resulting in antitumor activity in a wide variety of tumors. However, few studies have investigated the effect of hTERT gene-targeted ASODNs on telomerase activity and cell proliferation in human esophageal cancer. In the present study, an MTT assay was used to determine the growth inhibition rate of Eca-109 cells treated with a hTERT-targeted phosphorothioate-ASODN (PS-ASODN). An inverted microscope was used to observe the morphologic changes of the cells following treatment with 5 μM PS-ASODN for 10 days. Telomerase activity was detected using the silver staining semi-quantitative telomeric repeat amplification protocol (TRAP) assay. Following treatment with the PS-ASODN (1–5 μmol/l), the proliferation of the Eca-109 cells was inhibited. The differences in inhibition rate between the PS-ASODN and blank control groups were statistically significant (P<0.05) when the concentration of the PS-ASODN was ≥2 μmol/l, whereas no statistically significant difference was identified between the non-specific-ASODN and blank control groups. The inhibition rate increased gradually as the concentration of the PS-ASODN increased and with time, suggesting that the PS-ASODN inhibited the growth of Eca-109 cells in a concentration-dependent, time-dependent and sequence-specific manner. The growth rate of the cells incubated with the PS-ASODN was reduced compared with that of the control cells. Cells treated with the PS-ASODN became round, suspended and reduced in size. The PS-ASODN was also found to inhibit telomerase activity. The ability of the PS-ASODN to inhibit the telomerase activity and cell proliferation of the Eca-109 cell line suggests that ASODNs have the potential to be novel therapeutic agents for the treatment of esophageal cancer. PMID:25187833

  11. Dehydroepiandrosterone effects on the mRNA levels of peroxisome proliferator-activated receptors and their coactivators in human hepatoma HepG2 cells.

    PubMed

    Poczatková, H; Bogdanová, K; Uherková, L; Cervenková, K; Riegrová, D; Rypka, M; Veselý, J

    2007-12-01

    To investigate the effect of dehydroepiandrosterone (DHEA) on intracellular mRNA levels of peroxisome proliferator-activated receptors (PPARs) and PPAR-gamma coactivators (PGCs), we conducted a quantitative real-time RT-PCR study using HepG2 cells. Treatment with 100 micromol/l DHEA for 2-20 h caused a time-dependent elevation of mRNA levels in the cells. Upon 20 h, PPAR-alpha, -gamma1, and -gamma2 mRNAs and PGC-1alpha and -1beta mRNAs increased to 157, 161, 155, 656, and 475% of control levels, respectively (p < 0.05 each). Treatment with actinomycin D for 2.5-8 h revealed a significant stabilization effect of DHEA on PPAR-gamma1 and PGC-1alpha mRNAs at both 2.5 and 8 h incubation periods and a mild but significant stabilization effect on PGC-1beta mRNA at the 8 h incubation period suggesting that DHEA can modulate turnover of these mRNA transcripts. Basal mRNA levels of PPAR-alpha and PGC-1alpha were significantly suppressed upon 20 h treatment with cycloheximide, while those of PPAR-gamma1, -gamma2, and PGC-1beta were elevated. Cycloheximide also significantly reduced DHEA-induced accumulation of PPAR-alpha, -gamma1, -gamma2, and PGC-1alpha mRNAs, demonstrating the dependence of the DHEA action on de novo protein synthesis. The findings demonstrate that a supraphysiological concentration of DHEA can substantially influence gene expression of the PPAR signalling machinery at both transcriptional and posttranscriptional levels.

  12. Telmisartan inhibits the proinflammatory effects of homocysteine on human endothelial cells through activation of the peroxisome proliferator-activated receptor-δ pathway.

    PubMed

    Xu, Shanghua; Song, Huanhuan; Huang, Maozhi; Wang, Kefeng; Xu, Changsheng; Xie, Liangdi

    2014-09-01

    The aim of this study was to investigate the inhibition capacity of telmisartan to endothelial inflammation induced by homocysteine (Hcy) and discuss the proposed mechanism in vitro. Human umbilical vein endothelial cells (HUVECs) were prepared by collagenase digestion and cultured in vitro. An increase in monocyte chemoattractant protein-1 (MCP-1) and vascular cell adhesion molecule-1 (VCAM-1) was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) as markers of Hcy-induced endothelial inflammation. HL-60 cell adhesion to HUVECs was measured by rose bengal staining. Nuclear, cytosolic and total nuclear transcription factor-κB (NF-κB) p65 levels were analyzed by western blotting. Peroxisome proliferator-activated receptor-δ (PPARδ) expression by HUVECs exposed to Hcy with or without telmisartan pretreatment was analyzed by RT-PCR and western blotting. Hcy significantly increased the levels of MCP-1 mRNA, VCAM-1 mRNA and monocyte binding to HUVECs. These effects were significantly attenuated by pretreatment with telmisartan and PPARδ agonists. The effect of telmisartan was inhibited by PPARδ antagonists. The Hcy-mediated downregulation of PPARδ mRNA and protein of HUVECs was inhibited by telmisartan. Hcy-mediated upregulation of NF-κB p65 protein levels in nuclear extracts was inhibited by telmisartan and PPARδ agonists. In conclusion, telmisartan exerts potent anti-inflammatory effects in endothelial cells, probably via a binary mechanism involving PPARδ activation and inhibition of the nuclear translocation of NF-κB.

  13. Testosterone inhibits transforming growth factor-β signaling during myogenic differentiation and proliferation of mouse satellite cells: Potential role of follistatin in mediating testosterone action

    PubMed Central

    Braga, Melissa; Bhasin, Shalender; Jasuja, Ravi; Pervin, Shehla; Singh, Rajan

    2012-01-01

    Testosterone (T) administration is associated with increased satellite cell number and skeletal muscle hypertrophy, although there is considerable heterogeneity in the response of different skeletal muscle groups to T in vivo. We investigated the effects of T on the growth and differentiation of satellite cells isolated from levator ani (LA) and gastrocnemius (gastroc) muscles. T up regulated follistatin (Fst) expression, but down regulated the mRNA and protein expression of a number of genes in the transforming growth factor-beta (TGF-β)-signaling pathway. Inhibition of Fst expression by small interfering RNA (siRNA) inhibited myogenic differentiation and blocked the pro-myogenic effects of T. Treatment of satellite cells with T or Fst up regulated the expression of Pax7 and PCNA, and increased their proliferation. T and Fst blocked TGF-β induced inhibition of growth and myogenic differentiation and down regulated TGF-β-dependent transcriptome in both LA and gastroc cells. We conclude that T stimulation of satellite cell proliferation and myogenic differentiation are associated with up regulation of Fst and inhibition of TGF-β-signaling. PMID:22138414

  14. Possible involvement of calcineurin in retinoic acid-induced inhibition of leukemic HL-60 cell proliferation.

    PubMed

    Kihira, H; Hiasa, A; Yamamoto, M; Katayama, N; Kuno, T; Ohtsuka, K; Shiku, H; Nishikawa, M

    1998-03-01

    Differentiation of leukemic HL-60 cells by all transretinoic acid (ATRA) resulted in a reduced rate of growth. Cyclosporin A and FK506, at concentrations that inhibited calcineurin activity, abrogated the ATRA-induced inhibition of HL-60 cell growth but these immunosuppressants had no effect on the ATRA-induced granulocytic differentiation. Treatment with 1 microM ATRA led to a progressive increase in calcineurin phosphatase activity of HL-60 cells; the increase in this activity appeared to parallel the functional change of HL-60 cells during granulocytic differentiation. Increase in calcineurin activity was concordant with the increased expressions of calcineurin A and calcineurin B subunit proteins. The FKBP12 expression increased during ATRA-induced differentiation and expression of cyclophilin A remained unchanged. We propose that the increased expression of calcineurin is involved in the ATRA-induced inhibition of HL-60 cell proliferation, as in the case with 1,25alpha-dihydroxy-vitamin D3.

  15. Fangchinoline inhibits the proliferation of SPC-A-1 lung cancer cells by blocking cell cycle progression.

    PubMed

    Luo, Xue; Peng, Jian-Ming; Su, Lan-DI; Wang, Dong-Yan; Yu, You-Jiang

    2016-02-01

    Fangchinoline (Fan) is a bioactive compound isolated from the Chinese herb Stephania tetrandra S. Moore (Fen Fang Ji). The aim of the present study was to investigate the effect of Fan on the proliferation of SPC-A-1 lung cancer cells, and to define the associated molecular mechanisms. Following treatment with Fan, Cell Counting Kit-8, phase contrast imaging and Giemsa staining assays were used to detect cell viability; flow cytometry was performed to analyze the cell cycle distribution; and reverse transcription-quantitative polymerase chain reaction and western blot assays were used to investigate changes in the expression levels of cell cycle-associated genes and proteins. In the present study, treatment with Fan markedly inhibited the proliferation of SPC-A-1 lung cancer cells and significantly increased the percentage of cells in the G0/G1 phase of the cell cycle in a dose-dependent manner (P<0.05 for 2.5-5 µm; P<0.01 for 10 µm), whereas the percentage of cells in the S and G2/M phases were significantly reduced following treatment (P<0.05 for 5 µm; P<0.01 for 10 µm). Mechanistically, Fan significantly reduced the mRNA expression levels of cyclin D1, cyclin-dependent kinase 4 (CDK4) and CDK6 (P<0.05 for 2.5-5 µm; P<0.01 for 10 µm), which are key genes in the regulation of the G0/G1 phase of the cell cycle. Furthermore, treatment with Fan also decreased the expression of phosphorylated retinoblastoma (Rb) and E2F transcription factor-1 (E2F-1) proteins (P<0.05 for 5 µm; P<0.01 for 10 µm). In summary, the present study demonstrated that Fan inhibited the proliferation of SPC-A-1 lung cancer cells and induced cell cycle arrest at the G0/G1 phase. These effects may be mediated by the downregulation of cellular CDK4, CDK6 and cyclin D1 levels, thus leading to hypophosphorylation of Rb and subsequent suppression of E2F-1 activity. Therefore, the present results suggest that Fan may be a potential drug candidate for the prevention of lung cancer.

  16. Piperlongumine inhibits the proliferation and survival of B-cell acute lymphoblastic leukemia cell lines irrespective of glucocorticoid resistance

    SciTech Connect

    Han, Seong-Su; Han, Sangwoo; Kamberos, Natalie L.

    2014-09-26

    Highlights: • PL inhibits the proliferation of B-ALL cell lines irrespective of GC-resistance. • PL selectively kills B-ALL cells by increasing ROS, but not normal counterpart. • PL does not sensitize majority of B-ALL cells to DEX. • PL represses the network of constitutively activated TFs and modulates their target genes. • PL may serve as a new therapeutic molecule for GC-resistant B-ALL. - Abstract: Piperlongumine (PL), a pepper plant alkaloid from Piper longum, has anti-inflammatory and anti-cancer properties. PL selectively kills both solid and hematologic cancer cells, but not normal counterparts. Here we evaluated the effect of PL on the proliferation and survival of B-cell acute lymphoblastic leukemia (B-ALL), including glucocorticoid (GC)-resistant B-ALL. Regardless of GC-resistance, PL inhibited the proliferation of all B-ALL cell lines, but not normal B cells, in a dose- and time-dependent manner and induced apoptosis via elevation of ROS. Interestingly, PL did not sensitize most of B-ALL cell lines to dexamethasone (DEX). Only UoC-B1 exhibited a weak synergistic effect between PL and DEX. All B-ALL cell lines tested exhibited constitutive activation of multiple transcription factors (TFs), including AP-1, MYC, NF-κB, SP1, STAT1, STAT3, STAT6 and YY1. Treatment of the B-ALL cells with PL significantly downregulated these TFs and modulated their target genes. While activation of AURKB, BIRC5, E2F1, and MYB mRNA levels were significantly downregulated by PL, but SOX4 and XBP levels were increased by PL. Intriguingly, PL also increased the expression of p21 in B-ALL cells through a p53-independent mechanism. Given that these TFs and their target genes play critical roles in a variety of hematological malignancies, our findings provide a strong preclinical rationale for considering PL as a new therapeutic agent for the treatment of B-cell malignancies, including B-ALL and GC-resistant B-ALL.

  17. LKB1 inhibits the proliferation of gastric cancer cells by suppressing the nuclear translocation of Yap and β-catenin.

    PubMed

    Ma, Lian-Gang; Bian, Shi-Bo; Cui, Jian-Xin; Xi, Hong-Qing; Zhang, Ke-Cheng; Qin, Hong-Zhen; Zhu, Xiao-Ming; Chen, Lin

    2016-04-01

    Liver kinase B1 (LKB1) is known to suppress the proliferation, energy metabolism and mesenchymal transition of various cancer cells, and is involved in the regulation of Hippo-Yes-associated protein (Yap) and the Wnt/β-catenin signaling pathways. However, the role of LKB1 in gastric cancer (GC) was not fully understood. Thus, in the present study, we studied LKB1 and found that protein expression (0.37±0.061 vs. 0.59±0.108, P=0.006) and the protein ratio of p-Yap/Yap (0.179±0.085 vs. 0.8±0.126, P=0.001) were reduced in 54 gastric adenocarcinoma (GAC) tissues compared with the matched adjacent non-cancerous tissues, using western blotting and RT-qPCR assays. LKB1 expression was also observed decreased in 109 GAC tissues compared with 54 adjacent non-cancerous tissues (χ2=4.678, P=0.0306), and negatively correlated with the nuclear expression of Yap (r=-0.6997) and β-catenin (r=-0.3510), using immunohistochemical analysis. In GC patients, LKB1 expression was negatively associated with tumor size, tumor infiltration, lymph node metastasis and the TNM stage. LKB1 expression was determined to be positively correlated with longer overall survival of GC patients using Kaplan-Meier analysis (P=0.001). Subsequently, LKB1 expression in human GAC AGS cells was enhanced with a full‑length LKB1 transfection. In vitro and in vivo proliferation was inhibited in LKB1-overexpressing GC cells compared with the control cells. Yap and β-catenin expression were assessed by western blotting and RT-qPCR, and were found to be increased in the cytoplasm but decreased in the nucleus in LKB1-overexpressing GC cells compared with the control cells. The increase in cytoplasmic β-catenin was reversed by the silencing of LKB1 or Yap with shRNAs in LKB1-overexpressing GC cells. Moreover, Yap and β-catenin mRNA were barely altered by LKB1 overexpression. Thus, we concluded that LKB1 expression was reduced in GAC tissues but that it correlated positively with better prognosis for GC

  18. Transient inhibition of cell proliferation does not compromise self-renewal of mouse embryonic stem cells

    SciTech Connect

    Wang, Ruoxing; Guo, Yan-Lin

    2012-10-01

    Embryonic stem cells (ESCs) have unlimited capacity for self-renewal and can differentiate into various cell types when induced. They also have an unusual cell cycle control mechanism driven by constitutively active cyclin dependent kinases (Cdks). In mouse ESCs (mESCs). It is proposed that the rapid cell proliferation could be a necessary part of mechanisms that maintain mESC self-renewal and pluripotency, but this hypothesis is not in line with the finding in human ESCs (hESCs) that the length of the cell cycle is similar to differentiated cells. Therefore, whether rapid cell proliferation is essential for the maintenance of mESC state remains unclear. We provide insight into this uncertainty through chemical intervention of mESC cell cycle. We report here that inhibition of Cdks with olomoucine II can dramatically slow down cell proliferation of mESCs with concurrent down-regulation of cyclin A, B and E, and the activation of the Rb pathway. However, mESCs display can recover upon the removal of olomoucine II and are able to resume normal cell proliferation without losing self-renewal and pluripotency, as demonstrated by the expression of ESC markers, colony formation, embryoid body formation, and induced differentiation. We provide a mechanistic explanation for these observations by demonstrating that Oct4 and Nanog, two major transcription factors that play critical roles in the maintenance of ESC properties, are up-regulated via de novo protein synthesis when the cells are exposed to olomoucine II. Together, our data suggest that short-term inhibition of cell proliferation does not compromise the basic properties of mESCs. -- Highlights: Black-Right-Pointing-Pointer Inhibition of Cdks slows down mESCs proliferation. Black-Right-Pointing-Pointer mESCs display remarkable recovery capacity from short-term cell cycle interruption. Black-Right-Pointing-Pointer Short-term cell cycle interruption does not compromise mESC self-renewal. Black

  19. miR-1181 inhibits invasion and proliferation via STAT3 in pancreatic cancer

    PubMed Central

    Wang, Jie; Guo, Xing-Jun; Ding, You-Ming; Jiang, Jian-Xin

    2017-01-01

    AIM To examine the role of microRNA 1181 (miR-1181) in invasion and proliferation in pancreatic cancer. METHODS We analyzed the expression of miR-1181 in several pancreatic cancer cell lines and generated stable MIA-PaCa-2 and PANC-1 cell lines with up-regulated miR-1181 expression using an adenovirus delivery system. We then investigated miR-1181's effect on invasion and proliferation of pancreatic cancer cells by transwell assay, wound healing assay, cell counting kit-8 assay and colony-forming assay, and explored any underlying mechanisms by western bolt. Beyond that, we observed the change of the PANC-1 cell's cytoskeleton by immunofluorescence staining. RESULTS Our data showed that miR-1181 was relatively down-regulated in pancreatic cancer cell lines compared with normal pancreatic ductal epithelial cells. And miR-1181 inhibited the migration, invasion and proliferation activities of MIA-PaCa-2 and PANC-1 cells. Notably, after over-expressing of miR-1181 in PANC-1 cells, F-actin depolymerized. Immunofluorescence staining shows decreased F-actin and β-tubulin expression in PANC-1 cells over-expressing miR-1181 compared with the control cells. Furthermore, we found that over-expressing miR-1181 inhibited the expression of signal transducer and activator of transcription 3 (STAT3) while knocking-down miR-1181 up-regulated the expression of STAT3. Knocking-down miR-1181 promoted the invasion and proliferation of pancreatic cancer cells. And inhibition of STAT3 blocked the promotion effects of knocking-down miR-1181 on proliferation and invasion in pancreatic cancer. CONCLUSION Together our findings suggest that miR-1181 may be involved in pancreatic cancer cell invasion and proliferation by targeting STAT3 and indicate that miR-1181 may be a potential therapeutic agent for pancreatic cancer. PMID:28321160

  20. Curcumin inhibits angiotensin II-induced inflammation and proliferation of rat vascular smooth muscle cells by elevating PPAR-γ activity and reducing oxidative stress.

    PubMed

    Li, Hai-Yu; Yang, Mei; Li, Ze; Meng, Zhe

    2017-05-01

    Angiotensin II (AngII)-induced production of inflammatory factors and proliferation in vascular smooth muscle cells (VSMCs) play an important role in the progression of atherosclerotic plaques. Growing evidence has demonstrated that activation of peroxisome proliferator-activated receptor-γ (PPAR-γ) effectively attenuates AngII-induced inflammation and intercellular reactive oxygen species (iROS) production. Curcumin (Cur) inhibits inflammatory responses by enhancing PPAR-γ activity and reducing oxidative stress in various tissues. The aim of the present study was to ascertain whether Cur inhibits AngII-induced inflammation and proliferation, and its underlying molecular mechanism, in VSMCs. Enzyme-linked immunosorbent assay (ELISA) and real-time PCR were used to measure the protein and mRNA expression of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). Nitric oxide (NO) production was measured by Griess reaction. Western blot analysis and a DNA-binding assay were used to measure PPAR-γ activity. iROS production was measured using the DCFH-DA method. In rat VSMCs, Cur attenuated AngII‑induced expression of IL-6 and TNF-α mRNA and protein in a concentration-dependent manner, inhibited NO production by suppressing inducible NO synthase (iNOS) activity, and suppressed proliferation of VSMCs. This was accompanied by increased PPAR-γ expression and activation in Cur-pretreated VSMCs. GW9662, a PPAR-γ antagonist, reversed the anti-inflammatory effect of Cur. Moreover, Cur attenuated AngII-induced oxidative stress by downregulating the expression of p47phox, which is a key subunit of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. In conclusion, Cur inhibited the expression of IL-6 and TNF-α, decreased the production of NO, and suppressed the proliferation of VSMCs, by elevating PPAR-γ activity and suppressing oxidative stress, leading to attenuated AngII-induced inflammatory responses in VSMCs.

  1. Cordycepin Induces Apoptosis and Inhibits Proliferation of Human Lung Cancer Cell Line H1975 via Inhibiting the Phosphorylation of EGFR.

    PubMed

    Wang, Zheng; Wu, Xue; Liang, Yan-Ni; Wang, Li; Song, Zhong-Xing; Liu, Jian-Li; Tang, Zhi-Shu

    2016-09-27

    Cordycepin is an active component of the traditional Chinese medicine Cordyceps sinensis and Cordyceps militaris with notable anticancer activity. Though the prominent inhibitory activity was reported in different kinds of cancer cell lines, the concrete mechanisms remain elusive. It was reported that cordycepin could be converted into tri-phosphates in vivo to confuse a number of enzymes and interfere the normal cell function. For the inhibitory mechanism of EGFR inhibitors and the structure similarity of ATP and tri-phosphated cordycepin, human lung cancer cell line H1975 was employed to investigate the inhibitory effect of cordycepin. The results showed that cordycepin could inhibit cell proliferation and induce apoptosis in a dose-dependent manner. Cell cycle analysis revealed that H1975 cells could be arrested at the G₀/G₁ phase after cordycepin treatment. The expression levels of apoptosis-related protein Caspase-3 and Bcl-2 and phosphorylated expression levels of EGFR, AKT and ERK1/2 were all decreased compared with the control group stimulated with EGF. However, the protein expression levels of proapoptotic protein Bax and cleaved caspase-3 were increased. These results implied that cordycepin could inhibit cell proliferation and induce apoptosis via the EGFR signaling pathway. Our results indicated that there was potential to seek a novel EGFR inhibitor from cordycepin and its chemical derivatives.

  2. Recombinant disintegrin domain of ADAM15 inhibits the proliferation and migration of Bel-7402 cells

    SciTech Connect

    Hou, Y.; Chu, M.; Du, F.F.; Lei, J.Y.; Chen, Y.; Zhu, R.Y.; Gong, X.H.; Ma, X.; Jin, J.

    2013-06-14

    Highlights: •rhddADAM15 inhibited the proliferation and migration of Bel-7402 cells. •rhddADAM15 inhibited growth and metastasis of Bel-7402 cells in zebrafish xenograft. •rhddADAM15 induced apoptosis in Bel-7402 cells and somatic cells of zebrafish. •Cell-cycle in Bel-7402 cells showed a partial G{sub 2}/S arrest. •Activity of caspases 8, 9 and 3 was increased in rhddADAM15-treated Bel-7402 cells. -- Abstract: ADAM15 (A Disintegrin And Metalloproteinase 15), a transmembrane protein containing seven domains, interacts with some integrins via its disintegrin domain and overexpresses in many solid tumors. In this study, the effect of the recombinant human disintegrin domain (rhddADAM15) on the proliferation and migration of Bel-7402 cells was evaluated in vitro and in vivo in zebrafish xenografts. rhddADAM15 (4 μM) severely inhibited the proliferation and migration of Bel-7402 cells, inducing a partial G{sub 2}/S arrest and morphological nucleus changes of apoptosis. Moreover, the activity of caspases 8, 9 and 3 in Bel-7402 cells was increased. In addition, the zebrafish was used as a model for apoptosis-induction and tumor-xenograft. rhddADAM15 (1 pM) inhibited the growth and metastasis of Bel-7402 cell xenografts in zebrafish and a lower concentration (0.1 pM) induced severe apoptosis in the somatic cells of zebrafish. In conclusion, our data identified rhddADAM15 as a potent inhibitor of tumor growth and metastasis, making it a promising tool for use in anticancer treatment.

  3. Inhibition of murine splenic T lymphocyte proliferation by 2-deoxy-D-glucose-induced metabolic stress

    NASA Technical Reports Server (NTRS)

    Miller, E. S.; Klinger, J. C.; Akin, C.; Koebel, D. A.; Sonnenfeld, G.

    1994-01-01

    Female Swiss-Webster mice were injected with the glucose analogue 2-deoxy-D-glucose (2-DG), which when administered to rodents induces acute periods of metabolic stress. A single or multiple injections of 2-DG invoked a stress response, as evidenced by increases in serum corticosterone levels. The influence of this metabolic stressor on the blastogenic potential of splenic T lymphocytes was then examined. It was found that one, two, or three injections of 2-DG resulted in depressed T cell proliferative responses, with an attenuation of the effect occurring by the fifth injection. The 2-DG-induced inhibition of T cell proliferation was not attributable to 2-DG-induced cytolysis, as in vitro incubation of naive T cells with varying concentrations of 2-DG did not result in a reduction in cell number or viability, and flow cytometric analysis demonstrated that percentages of CD3, CD4, and CD8 splenic T cells were not altered as a result of 2-DG-induced stress. Incubating naive T cells in varying concentrations of 2-DG resulted in a dose-dependent inhibition of T cell blastogenic potential. Following in vivo exposure to 2-DG, T cell proliferation did not return to normal levels until 3 days after the cessation of 2-DG injections. Administering the beta-adrenergic receptor antagonist propranolol did not reverse the inhibited lymphoproliferation in 2-DG-treated mice. The inhibition in T cell proliferation was not observed, however, in mice that had been adrenalectomized or hypophysectomized and injected with 2-DG.(ABSTRACT TRUNCATED AT 250 WORDS).

  4. Inhibition of murine splenic T lymphocyte proliferation by 2-deoxy-D-glucose-induced metabolic stress

    NASA Technical Reports Server (NTRS)

    Miller, E. S.; Klinger, J. C.; Akin, C.; Koebel, D. A.; Sonnenfeld, G.

    1994-01-01

    Female Swiss-Webster mice were injected with the glucose analogue 2-deoxy-D-glucose (2-DG), which when administered to rodents induces acute periods of metabolic stress. A single or multiple injections of 2-DG invoked a stress response, as evidenced by increases in serum corticosterone levels. The influence of this metabolic stressor on the blastogenic potential of splenic T lymphocytes was then examined. It was found that one, two, or three injections of 2-DG resulted in depressed T cell proliferative responses, with an attenuation of the effect occurring by the fifth injection. The 2-DG-induced inhibition of T cell proliferation was not attributable to 2-DG-induced cytolysis, as in vitro incubation of naive T cells with varying concentrations of 2-DG did not result in a reduction in cell number or viability, and flow cytometric analysis demonstrated that percentages of CD3, CD4, and CD8 splenic T cells were not altered as a result of 2-DG-induced stress. Incubating naive T cells in varying concentrations of 2-DG resulted in a dose-dependent inhibition of T cell blastogenic potential. Following in vivo exposure to 2-DG, T cell proliferation did not return to normal levels until 3 days after the cessation of 2-DG injections. Administering the beta-adrenergic receptor antagonist propranolol did not reverse the inhibited lymphoproliferation in 2-DG-treated mice. The inhibition in T cell proliferation was not observed, however, in mice that had been adrenalectomized or hypophysectomized and injected with 2-DG.(ABSTRACT TRUNCATED AT 250 WORDS).

  5. Surgical Marking Pen Dye Inhibits Saphenous Vein Cell Proliferation and Migration in Saphenous Vein Graft Tissue

    PubMed Central

    Kikuchi, Shinsuke; Kenagy, Richard D; Gao, Lu; Wight, Thomas N; Azuma, Nobuyoshi; Sobel, Michael; Clowes, Alexander W

    2014-01-01

    and not different in blue-stained and unstained vein wall segments. Blue outer layer explants had 65.9±8.0 ng dye/explant compared to 2.1±1.3 for non-blue outer layer explants. Dye applied in vitro to either outer or inner layer explants dose-dependently inhibited migration (IC50=8.5 ng/explant). The IC50s of crystal violet for outer layer cell proliferation and migration were 0.1 and 1.2 μg/ml, while the EC50 for death was between 1 and 10 μg/ml. Conclusion Crystal violet inhibits venous cell migration and proliferation indicating that alternative methods should be considered for marking vein grafts. PMID:25935273

  6. Induction of type 1 iodothyronine deiodinase expression inhibits proliferation and migration of renal cancer cells.

    PubMed

    Poplawski, Piotr; Rybicka, Beata; Boguslawska, Joanna; Rodzik, Katarzyna; Visser, Theo J; Nauman, Alicja; Piekielko-Witkowska, Agnieszka

    2017-02-15

    Type 1 iodothyronine deiodinase (DIO1) regulates peripheral metabolism of thyroid hormones that control cellular proliferation, differentiation and metabolism. The significance of DIO1 in cancer is unknown. In this study we hypothesized that diminished expression of DIO1, observed in renal cancer, contributes to the carcinogenic process in the kidney. Here, we demonstrate that ectopic expression of DIO1 in renal cancer cells changes the expression of genes controlling cell cycle, including cyclin E1 and E2F5, and results in inhibition of proliferation. The expression of genes encoding collagens (COL1A1, COL4A2, COL5A1), integrins (ITGA4, ITGA5, ITGB3) and transforming growth factor-β-induced (TGFBI) is significantly altered in renal cancer cells with induced expression of DIO1. Finally, we show that overexpression of DIO1 inhibits migration of renal cancer cells. In conclusion, we demonstrate for the first time that loss of DIO1 contributes to renal carcinogenesis and that its induced expression protects cells against cancerous proliferation and migration.

  7. SIRT1 promotes proliferation and inhibits the senescence-like phenotype in human melanoma cells

    PubMed Central

    Ohanna, Mickaël; Bonet, Caroline; Bille, Karine; Allegra, Maryline; Davidson, Irwin; Bahadoran, Philippe; Lacour, Jean-Philippe; Ballotti, Robert; Bertolotto, Corine

    2014-01-01

    SIRT1 operates as both a tumor suppressor and oncogenic factor depending on the cell context. Whether SIRT1 plays a role in melanoma biology remained poorly elucidated. Here, we demonstrate that SIRT1 is a critical regulator of melanoma cell proliferation. SIRT1 suppression by genetic or pharmacological approaches induces cell cycle arrest and a senescence-like phenotype. Gain and loss of function experiments show that M-MITF regulates SIRT1 expression, thereby revealing a melanocyte-specific control of SIRT1. SIRT1 over-expression relieves the senescence-like phenotype and the proliferation arrest caused by MITF suppression, demonstrating that SIRT1 is an effector of MITF-induced proliferation in melanoma cells. Interestingly, SIRT1 level and activity are enhanced in the PLX4032-resistant BRAFV600E-mutated melanoma cells compared with their sensitive counterpart. SIRT1 inhibition decreases melanoma cell growth and rescues the sensibility to PLX4032 of PLX4032-resistant BRAFV600E-mutated melanoma cells. In conclusion, we provide the first evidence that inhibition of SIRT1 warrants consideration as an anti-melanoma therapeutic option. PMID:24742694

  8. Inhibition of AGS Cancer Cell Proliferation following siRNA-Mediated Downregulation of VEGFR2

    PubMed Central

    Zarei Mahmudabadi, Ali; Masoomi Karimi, Masoomeh; Bahabadi, Majid; Bagheri Hoseinabadi, Zahra; JafariSani, Moslem; Ahmadi, Reza

    2016-01-01

    Objective Vascular endothelial growth factor (VEGF) and VEGF receptors (VEGFRs) play important roles in angiogenesis of different developmental mechanisms such as wound healing, embryogenesis and diseases, including different types of cancer. VEGFR2 is associated with cell proliferation, migration, and vascular permeability of endothelial cells. Blocking VEGF and its receptors is suggested as a therapeutic approach to prevent tumor growth. In this study, we aim to block VEGF signaling via small interfering RNA (siRNA) inhibition of VEGFR2. Materials and Methods In this experimental study, we used the RNA interference (RNAi) mechanism to suppress expression of the VEGFR2 gene. We conducted the 3-(4,5-di- methylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay, real-time polymerase chain reaction (PCR), Western blot, and flow cytometry analyses of VEGFR2 expression. Results Real-time PCR and Western blot results showed that VEGFR2 expression significantly downregulated. This suppression was followed by inhibition of cell prolifera- tion, reduction of viability, and induction of apoptosis in the cancer cells. Conclusion These findings suggest that VEGFR2 has a role in cell proliferation and tumor growth. Accordingly, it is suggested that VEGFR2 can be a therapeutic target for controlling tumor growth and proliferation. PMID:27602320

  9. Plant-Made Trastuzumab (Herceptin) Inhibits HER2/Neu+ Cell Proliferation and Retards Tumor Growth

    PubMed Central

    Komarova, Tatiana V.; Kosorukov, Vyacheslav S.; Frolova, Olga Y.; Petrunia, Igor V.; Skrypnik, Ksenia A.; Gleba, Yuri Y.; Dorokhov, Yuri L.

    2011-01-01

    Background Plant biotechnology provides a valuable contribution to global health, in part because it can decrease the cost of pharmaceutical products. Breast cancer can now be successfully treated by a humanized monoclonal antibody (mAb), trastuzumab (Herceptin). A course of treatment, however, is expensive and requires repeated administrations of the mAb. Here we used an Agrobacterium-mediated transient expression system to produce trastuzumab in plant cells. Methodology/Principal Findings We describe the cloning and expression of gene constructs in Nicotiana benthamiana plants using intron-optimized Tobacco mosaic virus- and Potato virus X-based vectors encoding, respectively, the heavy and light chains of trastuzumab. Full-size antibodies extracted and purified from plant tissues were tested for functionality and specificity by (i) binding to HER2/neu on the surface of a human mammary gland adenocarcinoma cell line, SK-BR-3, in fluorescence-activated cell sorting assay and (ii) testing the in vitro and in vivo inhibition of HER-2-expressing cancer cell proliferation. We show that plant-made trastuzumab (PMT) bound to the Her2/neu oncoprotein of SK-BR-3 cells and efficiently inhibited SK-BR-3 cell proliferation. Furthermore, mouse intraperitoneal PMT administration retarded the growth of xenografted tumors derived from human ovarian cancer SKOV3 Her2+ cells. Conclusions/Significance We conclude that PMT is active in suppression of cell proliferation and tumor growth. PMID:21390232

  10. Structural requirements for novel coenzyme-substrate derivatives to inhibit intracellular ornithine decarboxylase and cell proliferation.

    PubMed

    Wu, Fang; Gehring, Heinz

    2009-02-01

    Creating transition-state mimics has proven to be a powerful strategy in developing inhibitors to treat malignant diseases in several cases. In the present study, structurally diverse coenzyme-substrate derivatives mimicking this type for pyridoxal 5'-phosphate-dependent human ornithine decarboxylase (hODC), a potential anticancer target, were designed, synthesized, and tested to elucidate the structural requirements for optimal inhibition of intracellular ODC as well as of tumor cell proliferation. Of 23 conjugates, phosphopyridoxyl- and pyridoxyl-L-tryptophan methyl ester (pPTME, PTME) proved significantly more potent in suppression proliferation (IC(50) up to 25 microM) of glioma cells (LN229) than alpha-DL-difluoromethylornithine (DFMO), a medically used irreversible inhibitor of ODC. In agreement with molecular modeling predictions, the inhibitory action of pPTME and PTME toward intracellular ODC of LN229 cells exceeded that of the previous designed lead compound POB. The inhibitory active compounds feature hydrophobic side chain fragments and a kind of polyamine motif (-NH-(CH(X))(4)-NH-). In addition, they induce, as polyamine analogs often do, the activity of the polyamine catabolic enzymes polyamine oxidase and spermine/spermidine N(1)-acetyltransferase up to 250 and 780%, respectively. The dual-action mode of these compounds in LN229 cells affects the intracellular polyamine metabolism and might underlie the more favorable cell proliferation inhibition in comparison with DFMO.

  11. M2muscarinic receptors inhibit cell proliferation and migration in urothelial bladder cancer cells

    PubMed Central

    Pacini, Luca; De Falco, Elena; Di Bari, Maria; Coccia, Andrea; Siciliano, Camilla; Ponti, Donatella; Pastore, Antonio Luigi; Petrozza, Vincenzo; Carbone, Antonio; Tata, Ada Maria; Calogero, Antonella

    2014-01-01

    The role of muscarinic receptors in several diseases including cancer has recently emerged. To evaluate the hypothesis that muscarinic acetylcholine receptors may play a role in bladder cancer as well as in other tumor types, we investigated their expression in bladder tumor specimens. All examined samples expressed the M1, M2 and M3 receptor subtypes. We also found that the level of M2 transcripts, but not those of M1 or M3, significantly increased with the tumor histologic grade. In view of these results, we proceeded to investigate whether the M2 agonist Arecaidine had any effect on in vitro cell growth and migration of T24 cells, a bladder tumor cell line expressing the muscarinic receptors, including the M2 subtype. We observed that Arecaidine significantly reduced T24 and 5637 cell proliferation and migration in a concentration dependent manner. The silencing of M2 receptor by siRNA in T24 and 5637 cell lines showed the inability of Arecaidine (100 μM) to inhibit cell proliferation after 48 hours, whereas the use of M1 and M3 antagonists in T24 appeared not to counteract the Arecaidine effect, suggesting that the inhibition of cell proliferation was directly dependent on M2 receptor activation. These data suggest that M2 muscarinic receptors may play a relevant role in bladder cancer and represent a new attractive therapeutic target. PMID:25482946

  12. Downregulation of PAX6 by shRNA Inhibits Proliferation and Cell Cycle Progression of Human Non-Small Cell Lung Cancer Cell Lines

    PubMed Central

    Zhao, Xiaoting; Yue, Wentao; Zhang, Lina; Ma, Li; Jia, Wenyun; Qian, Zhe; Zhang, Chunyan; Wang, Yue

    2014-01-01

    Background The transcription factor PAX6 is primarily expressed in embryos. PAX6 is also expressed in several tumors and plays an oncogenic role. However, little is known about the role of PAX6 in lung cancer. Methods The function of PAX6 in lung cancer cells was evaluated by small interfering RNA-mediated depletion of the protein followed by analyses of cell proliferation, anchorage-independent growth, and cell cycle arrest. The changes of cyclin D1, pRB, ERK1/2, p38 expression caused by PAX6 inhibition were detected using western-blotting. The PAX6 mRNA level in 52 pairs of tumors and corresponding matched adjacent normal tissues from non-small cell lung cancer patients and lung cancer cell lines was detected by real-time PCR. Results Suppression of PAX6 expression inhibited cell growth and colony formation in A549 and H1299 cells. The percentage of cells in G1-phase increased when PAX6 expression was inhibited. The cyclin D1 protein level, as well as the pRB phosphorylation level, decreased as a result of PAX6 down-regulation. The activity of ERK1/2 and p38 was also suppressed in PAX6 knock-down cells. The PAX6 mRNA was highly expressed in lung cancer tissue and lung cancer cell lines. In most patients (about 65%), the relative ratio of PAX6 mRNA in primary NSCLC versus adjacent tissues exceeded 100. Conclusions Our data implicated that PAX6 accelerates cell cycle progression by activating MAPK signal pathway. PAX6 mRNA levels were significantly elevated in primary lung cancer tissues compared to their matched adjacent tissues. PMID:24454925

  13. miR-497-5p inhibits cell proliferation and invasion by targeting KCa3.1 in angiosarcoma

    PubMed Central

    Zhao, Xia; Chen, Dong; Wang, Xiaoyan; Yang, Qin; Wan, Jie; Zhu, Yuanli; Wang, Yu; Zhang, Shiying; Wang, Ying; Tang, Qiang; Masuzawa, Mikio; Wang, Guoping; Duan, Yaqi

    2016-01-01

    Angiosarcoma is a rare malignant mesenchymal tumor with poor prognosis. We aimed to identify malignancy-associated miRNAs and their target genes, and explore biological functions of miRNA and its target in angiosarcoma. By miRNA microarrays and reverse transcription polymerase chain reaction, we identified 1 up-regulated miRNA (miR-222-3p) and 3 down-regulated miRNAs (miR-497-5p, miR-378-3p and miR-483-5p) in human angiosarcomas compared with human capillary hemangiomas. The intermediate-conductance calcium activated potassium channel KCa3.1 was one of the putative target genes of miR-497-5p, and marked up-regulation of KCa3.1 was detected in angiosarcoma biopsy specimens by immunohistochemistry. The inverse correlation of miR-497-5p and KCa3.1 also was observed in the ISO-HAS angiosarcoma cell line at the mRNA and protein levels. The direct targeting of KCa3.1 by miR-497-5p was evidenced by reduced luciferase activity due to complementary binding of miR-497-5p to KCa3.1 mRNA 3′ untranslated region. For the functional role of miR-497-5p/KCa3.1 pair, we showed that application of TRAM-34, a specific KCa3.1 channel blocker, or transfection of ISO-HAS cells with KCa3.1 siRNA or miR-497-5p mimics inhibited cell proliferation, cell cycle progression, and invasion by down-regulating cell-cycle related proteins including cyclin D1, surviving and P53 and down-regulating matrix metallopeptidase 9. In an in vivo angiosarcoma xenograft model, TRAM-34 or miR-497-5p mimics both inhibited tumor growth. In conclusion, the tumor suppressor miR-497-5p down-regulates KCa3.1 expression and contributes to the inhibition of angiosarcoma malignancy development. The miR-497-5p or KCa3.1 might be potential new targets for angiosarcoma treatment. PMID:27531900

  14. S100P is a potential molecular target of cadmium-induced inhibition of human placental trophoblast cell proliferation.

    PubMed

    Zhou, Taimei; Wang, Haiying; Zhang, Shen; Jiang, Xinglin; Wei, Xiaolong

    2016-11-01

    Cadmium, a common and highly toxic pollutant, has been known to accumulate high concentrations in placenta with deleterious effects on placental structure and function. Cadmium inhibits cell proliferation in placenta via targeting metal binding proteins. S100P, a Ca(2+)-binding protein, plays an important role in promoting cell proliferation and our previous study found its downregulation was linked to cadmium exposure in Guiyu, a famous e-waste recycling town in China. So, the present study was aimed to define whether cadmium inhibited cell proliferation through interfering with S100P. Using human trophoblast-derived HTR-8/SVneo cells as a model in vitro, we showed that cadmium exposure led to decreases in both cell proliferation and S100P expression. Knockdown of S100P in HTR-8/SVneo cells led to an obvious decrease of cell proliferation, and upregulation of S100P resulted in a significant increase of cell proliferation. Furthermore, after 24h of exposure to cadmium (20μM), cells transfected with pcDNA3.1-S100P showed a 1.3-fold higher S100P protein level, 38% higher proliferation evaluated with MTT assay than cells with no transfection, indicating that S100P expression attenuated cadmium-induced inhibition of cell proliferation. Taken together, we demonstrate that cadmium inhibits S100P expression and cell proliferation in placenta, meanwhile, S100P expression affects cell proliferation. Thus, our study is the first to indicate that cadmium may induce inhibition of placental trophoblast cell proliferation through targeting S100P. Copyright © 2016 Elsevier GmbH. All rights reserved.

  15. Shade Inhibits Leaf Size by Controlling Cell Proliferation and Enlargement in Soybean.

    PubMed

    Wu, Yushan; Gong, Wanzhuo; Yang, Wenyu

    2017-08-23

    To gain more insight into the physiological function of shade and how shade affects leaf size, we investigated the growth, leaf anatomical structure, hormones and genes expressions in soybean. Soybean seeds were sown in plastic pots and were allowed to germinate and grow for 30 days under shade or full sunlight conditions. Shade treated plants showed significantly increase on stem length and petiole length, and decrease on stem diameters, shoot biomass and its partition to leaf also were significantly lower than that in full sunlight. Smaller and thinner on shade treated leaves than corresponding leaves on full sunlight plants. The decreased leaf size caused by shade was largely attributable to cell proliferation in young leaves and both cell proliferation and enlargement in old leaves. Shade induced the expression of a set of genes related to cell proliferation and/or enlargement, but depended on the developmental stage of leaf. Shade significantly increased the auxin and gibberellin content, and significantly decreased the cytokinin content in young, middle and old leaves. Taken together, these results indicated that shade inhibited leaf size by controlling cell proliferation and enlargement, auxin, gibberellin and cytokinin may play important roles in this process.

  16. Corticosterone Inhibits the Proliferation of C6 Glioma Cells via the Translocation of Unphosphorylated Glucocorticoid Receptor.

    PubMed

    Nakatani, Yoshihiko; Amano, Taku; Takeda, Hiroshi

    2016-01-01

    Astroglial cells have been considered to have passive brain function by helping to maintain neurons. However, recent studies have revealed that the dysfunction of such passive functions may be associated with various neuropathological diseases, such as schizophrenia, Alzheimer's disease, amyotrophic lateral sclerosis and major depression. Corticosterone (CORT), which is often referred to as the stress hormone, is a well-known regulator of peripheral immune responses and also shows anti-inflammatory properties in the brain. However, it is still obscure how CORT affects astroglial cell function. In this study, we investigated the effects of CORT on the proliferation and survival of astroglial cells using C6 glioma cells. Under treatment with CORT for 24h, the proliferation of C6 glioma cells decreased in a dose-dependent manner. Moreover, this inhibition was diminised by treatment with mifepristone, a glucocorticoid receptor (GR) antagonist, but not by spironolactone, a mineralocorticoid receptor (MR) antagonist, and was independent of GR phosphorylation and other GR-related intracellular signaling cascades. Furthermore, it was observed that the translocation of GR from the cytosol to the nucleus was promoted by the treatment with CORT. These results indicate that CORT decreases the proliferation of C6 glioma cells by modifying the transcription of a particular gene related to cell proliferation independent of GR phosphorylation.

  17. Knockdown of homeobox A5 by small hairpin RNA inhibits proliferation and enhances cytarabine chemosensitivity of acute myeloid leukemia cells.

    PubMed

    Li, Na; Jia, Xiuhong; Wang, Jianyong; Li, Youjie; Xie, Shuyang

    2015-11-01

    Homeobox genes encode transcription factors that are essential for embryonic morphogenesis and differentiation. Transcription factors containing the highly conserved homeobox motif show considerable promise as potential regulators of hematopoietic maturation events. Previous studies have suggested that the increased expression levels of homeobox (HOX)A genes was correlated with the cytogenetic findings associated with poor prognosis in acute myeloid leukemia and mixed lineage leukemia. The aim of the present study was to investigate the role of HOXA5 in leukemia. The U937 human leukemia cell line was transfected with a HOXA5‑targeted short hairpin RNA (shRNA) to determine the effects of downregulation of the HOXA5 on proliferation, apoptosis, cell cycle distribution and chemoresistance in leukemia cells. Reverse transcription‑quantitative polymerase chain reaction and western blot analyses demonstrated that the mRNA and protein expression levels of HOXA5 were markedly suppressed following transfection with an shRNA‑containing vector. Knockdown of HOXA5 significantly inhibited cell proliferation, as determined by Cell Counting kit‑8 assay. Flow cytometry revealed that reduced HOXA5 expression levels resulted in cell cycle arrest at the G1 phase, and induced apoptosis. In addition, western blot analysis demonstrated that HOXA5 knockdown increased the expression levels of caspase‑3, and reduced the expression levels of survivin in the U937 cells. Furthermore, knockdown of HOXA5 in the U937 cells enhanced their chemosensitivity to cytarabine. The results of the present study suggested that downregulation of HOXA5 by shRNA may trigger apoptosis and overcome drug resistance in leukemia cells. Therefore, HOXA5 may serve as a potential target for developing novel therapeutic strategies for leukemia.

  18. MicroRNA-490-3p inhibits proliferation of A549 lung cancer cells by targeting CCND1

    SciTech Connect

    Gu, Haihua; Yang, Tao; Fu, Shaozi; Chen, Xiaofan; Guo, Lei; Ni, Yiming

    2014-01-31

    Highlights: • We examined the level of miR-490-3p in A549 lung cancer cells compared with normal bronchial epithelial cell line. • We are the first to show the function of miR-490-3p in A549 lung cancer cells. • We demonstrate CCND1 may be one of the targets of miR-490-3p. - Abstract: MicroRNAs (miRNAs) are small non-coding RNAs that negatively regulate the translation of messenger RNAs by binding their 3′-untranslated region (3′UTR). In this study, we found that miR-490-3p is significantly down-regulated in A549 lung cancer cells compared with the normal bronchial epithelial cell line. To better characterize the role of miR-490-3p in A549 cells, we performed a gain-of-function analysis by transfecting the A549 cells with chemically synthesized miR-490-3P mimics. Overexpression of miR-490-3P evidently inhibits cell proliferation via G1-phase arrest. We also found that forced expression of miR-490-3P decreased both mRNA and protein levels of CCND1, which plays a key role in G1/S phase transition. In addition, the dual-luciferase reporter assays indicated that miR-490-3P directly targets CCND1 through binding its 3′UTR. These findings indicated miR-490-3P could be a potential suppressor of cellular proliferation.

  19. Ursolic acid inhibits proliferation and induces apoptosis of HT-29 colon cancer cells by inhibiting the EGFR/MAPK pathway*

    PubMed Central

    Shan, Jian-zhen; Xuan, Yan-yan; Zheng, Shu; Dong, Qi; Zhang, Su-zhan

    2009-01-01

    Objective: To investigate the effects of ursolic acid on the proliferation and apoptosis of human HT-29 colon cancer cells. Methods: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry assays were performed to evaluate the effects of ursolic acid on the growth and apoptosis of HT-29 cells. Western blot analysis was applied to investigate the inhibitory effects of ursolic acid on the phosphorylation of the epidermal growth factor receptor (EGFR), extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (p38 MAPK), and the activity of B cell leukemia-2 (Bcl-2), B cell leukemia-xL (Bcl-xL), caspase-3, and caspase-9. Results: Ursolic acid inhibited the growth of HT-29 cells in dose- and time-dependent manners. The median inhibition concentration (IC50) values for 24, 48, and 72 h treatment were 26, 20, and 18 μmol/L, respectively. The apoptotic rates of 10, 20, and 40 μmol/L ursolic acid treatments for 24 h were 5.74%, 14.49%, and 33.05%, and for 48 h were 9%, 21.39%, and 40.49%, respectively. Ursolic acid suppressed the phosphorylation of EGFR, ERK1/2, p38 MAPK, and JNK, which is well correlated with its growth inhibitory effect. 10, 20, and 40 μmol/L ursolic acid significantly inhibited the proliferation of EGF-stimulated HT-29 cells (P<0.05). Cell proliferation was most significantly inhibited when treated with 10 and 20 μmol/L ursolic acid combined with 200 nmol/L AG 1478 or 10 μmol/L U0126 (P<0.01). Besides, it also down-regulated the expression of Bcl-2 and Bcl-xL and activated caspase-3 and caspase-9. Conclusion: Ursolic acid induces apoptosis in HT-29 cells by suppressing the EGFR/MAPK pathway, suggesting that it may be a potent agent for the treatment of colorectal cancer. PMID:19735099

  20. HCV core protein promotes hepatocyte proliferation and chemoresistance by inhibiting NR4A1

    SciTech Connect

    Tan, Yongsheng Li, Yan

    2015-10-23

    This study investigated the effect of HCV core protein on the proliferation of hepatocytes and hepatocellular carcinoma cells (HCC), the influence of HCV core protein on HCC apoptosis induced by the chemotherapeutic agent cisplatin, and the mechanism through which HCV core protein acts as a potential oncoprotein in HCV-related HCC by measuring the levels of NR4A1 and Runt-related transcription factor 3 (RUNX3), which are associated with tumor suppression and chemotherapy resistance. In the present study, PcDNA3.1-core and RUNX3 siRNA were transfected into LO2 and HepG2 cells using Lipofectamine 2000. LO2-core, HepG2-core, LO2-RUNX3 {sup low} and control cells were treated with different concentrations of cisplatin for 72 h, and cell proliferation and apoptosis were assayed using the CellTiter 96{sup ®}Aqueous Non-Radioactive Cell Proliferation Assay Kit. Western blot and real time PCR analyses were used to detect NR4A1, RUNX3, smad7, Cyclin D1 and BAX. Confocal microscopy was used to determine the levels of NR4A1 in HepG2 and HepG2-core cells. The growth rate of HepG2-core cells was considerably greater than that of HepG2 cells. HCV core protein increased the expression of cyclin D1 and decreased the expressions of NR4A1 and RUNX3. In LO2 – RUNX3 {sup low}, the rate of cel