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Sample records for human transitional cell

  1. Geometrical guidance and trapping transition of human sperm cells

    Science.gov (United States)

    Guidobaldi, A.; Jeyaram, Y.; Berdakin, I.; Moshchalkov, V. V.; Condat, C. A.; Marconi, V. I.; Giojalas, L.; Silhanek, A. V.

    2014-03-01

    The guidance of human sperm cells under confinement in quasi-2D microchambers is investigated using a purely physical method to control their distribution. Transport property measurements and simulations are performed with diluted sperm populations, for which effects of geometrical guidance and concentration are studied in detail. In particular, a trapping transition at convex angular wall features is identified and analyzed. We also show that highly efficient microratchets can be fabricated by using curved asymmetric obstacles to take advantage of the spermatozoa specific swimming strategy.

  2. Benzyl Isothiocyanate Inhibits Epithelial-Mesenchymal Transition in Cultured and Xenografted Human Breast Cancer Cells

    OpenAIRE

    Sehrawat, Anuradha; Singh, Shivendra V.

    2011-01-01

    We showed previously that cruciferous vegetable constituent benzyl isothiocyanate (BITC) inhibits growth of cultured and xenografted human breast cancer cells, and suppresses mammary cancer development in a transgenic mouse model. We now demonstrate, for the first time, that BITC inhibits epithelial-to-mesenchymal transition (EMT) in human breast cancer cells. Exposure of estrogen-independent MDA-MB-231 and estrogen-responsive MCF-7 human breast cancer cell lines and a pancreatic cancer cell ...

  3. Transitional B cells in early human B cell development - time to revisit the paradigm?

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    Victoria G Martin

    2016-12-01

    Full Text Available The B cell repertoire is generated in the adult bone marrow by an ordered series of gene rearrangement processes that result in massive diversity of immunoglobulin (Ig genes, and consequently an equally large number of potential specificities for antigen. As the process is essentially random, then cells exhibiting excess reactivity with self-antigens are generated and need to be removed from the repertoire before the cells are fully mature. Some of the cells are deleted, and some will undergo receptor editing to see if changing the light chain can rescue an autoreactive antibody. As a consequence, the binding properties of the B cell receptor are changed as development progresses through pre-B>>immature>>transitional>>naïve phenotypes. Using long-read, high-throughput, sequencing we have produced a unique set of sequences from these four cell types in human bone marrow and matched peripheral blood and our results describe the effects of tolerance selection on the B cell repertoire at the Ig gene level. Most strong effects of selection are seen within the heavy chain repertoire, and can be seen both in gene usage and in CDR-H3 characteristics. Age-related changes are small and only the size of the CDR-H3 shows constant and significant change in these data. The paucity of significant changes in either kappa or lambda light chain repertoires implies that either the heavy chain has more influence over autoreactivity than light chain and/or that switching between kappa and lambda light chains, as opposed to switching within the light chain loci, may effect a more successful autoreactive rescue by receptor editing. Our results show that the transitional cell population contains cells other than those that are part of the pre-B>>immature>>transitional>>naïve development pathway, since the population often shows a repertoire that is outside the trajectory of gene loss/gain between pre-B and naïve stages.

  4. Inhibition of Rho-Kinase Abrogates Migration of Human Transitional Cell Carcinoma Cells : Results of an in vitro Study

    NARCIS (Netherlands)

    vom Dorp, Frank; Sanders, Harald; Boergermann, Christof; Luemmen, Gerd; Ruebben, Herbert; Jakobs, Karl H.; Schmidt, Martina

    2011-01-01

    Introduction: Migration of cells involves a complex signaling network. The aim of the present study was to elucidate the impact of Rho-kinase (ROK) on G protein-coupled receptor-induced migration of human transitional cell carcinoma cells in an in vitro experimental setting. Materials and Methods: I

  5. Mogoltacin enhances vincristine cytotoxicity in human transitional cell carcinoma (TCC) cell line.

    Science.gov (United States)

    Behnam Rassouli, F; Matin, M M; Iranshahi, M; Bahrami, A R; Neshati, V; Mollazadeh, S; Neshati, Z

    2009-03-01

    Bladder cancer is the second common cancer of the genitourinary system throughout the world and intravesical chemotherapy is usually used to reduce tumour recurrence and progression. Human transitional cell carcinoma (TCC) is an epithelial-like adherent cell line originally established from primary bladder carcinoma. Here we report the effect of mogoltacin, a sesquiterpene coumarin from Ferula badrakema on TCC cells. Mogoltacin was isolated from the fruits of F. badrakema, using silica gel column chromatography and preparative thin layer chromatography. Mogoltacin did not have any significant cytotoxicity effect on neoplastic TCC cells at 16, 32, 64, 128, 200 and 600 microg ml(-1) concentrations. In order to analyse its combination effect, TCC cells were cultured in the presence of various combining concentrations of mogoltacin and vincristine. Cells were then observed for morphological changes (by light microscopy) and cytotoxicity using MTT assay. The effect of mogoltacin on vincristine toxicity was studied after 24, 48 and 72 h of drug administration. The results of MTT assay showed that mogoltacin can significantly enhance the cytotoxicity of vincristine and confirmed the morphological observations. Results revealed that combination of 40 microg ml(-1) vincristine with 16 microg ml(-1) mogoltacin increased the cytotoxicity of vincristine after 48 h by 32.8%.

  6. Transitional B Cells in Early Human B Cell Development – Time to Revisit the Paradigm?

    Science.gov (United States)

    Martin, Victoria G.; Wu, Yu-Chang Bryan; Townsend, Catherine L.; Lu, Grace H. C.; O’Hare, Joselli Silva; Mozeika, Alexander; Coolen, Anthonius C. C.; Kipling, David; Fraternali, Franca; Dunn-Walters, Deborah K.

    2016-01-01

    The B cell repertoire is generated in the adult bone marrow by an ordered series of gene rearrangement processes that result in massive diversity of immunoglobulin (Ig) genes and consequently an equally large number of potential specificities for antigen. As the process is essentially random, the cells exhibiting excess reactivity with self-antigens are generated and need to be removed from the repertoire before the cells are fully mature. Some of the cells are deleted, and some will undergo receptor editing to see if changing the light chain can rescue an autoreactive antibody. As a consequence, the binding properties of the B cell receptor are changed as development progresses through pre-B ≫ immature ≫ transitional ≫ naïve phenotypes. Using long-read, high-throughput, sequencing we have produced a unique set of sequences from these four cell types in human bone marrow and matched peripheral blood, and our results describe the effects of tolerance selection on the B cell repertoire at the Ig gene level. Most strong effects of selection are seen within the heavy chain repertoire and can be seen both in gene usage and in CDRH3 characteristics. Age-related changes are small, and only the size of the CDRH3 shows constant and significant change in these data. The paucity of significant changes in either kappa or lambda light chain repertoires implies that either the heavy chain has more influence over autoreactivity than light chain and/or that switching between kappa and lambda light chains, as opposed to switching within the light chain loci, may effect a more successful autoreactive rescue by receptor editing. Our results show that the transitional cell population contains cells other than those that are part of the pre-B ≫ immature ≫ transitional ≫ naïve development pathway, since the population often shows a repertoire that is outside the trajectory of gene loss/gain between pre-B and naïve stages. PMID:27994589

  7. A human breast cell model of pre-invasive to invasive transition

    Energy Technology Data Exchange (ETDEWEB)

    Bissell, Mina J; Rizki, Aylin; Weaver, Valerie M.; Lee, Sun-Young; Rozenberg, Gabriela I.; Chin, Koei; Myers, Connie A.; Bascom, Jamie L.; Mott, Joni D.; Semeiks, Jeremy R.; Grate, Leslie R.; Mian, I. Saira; Borowsky, Alexander D.; Jensen, Roy A.; Idowu, Michael O.; Chen, Fanqing; Chen, David J.; Petersen, Ole W.; Gray, Joe W.; Bissell, Mina J.

    2008-03-10

    A crucial step in human breast cancer progression is the acquisition of invasiveness. There is a distinct lack of human cell culture models to study the transition from pre-invasive to invasive phenotype as it may occur 'spontaneously' in vivo. To delineate molecular alterations important for this transition, we isolated human breast epithelial cell lines that showed partial loss of tissue polarity in three-dimensional reconstituted-basement membrane cultures. These cells remained non-invasive; however, unlike their non-malignant counterparts, they exhibited a high propensity to acquire invasiveness through basement membrane in culture. The genomic aberrations and gene expression profiles of the cells in this model showed a high degree of similarity to primary breast tumor profiles. The xenograft tumors formed by the cell lines in three different microenvironments in nude mice displayed metaplastic phenotypes, including squamous and basal characteristics, with invasive cells exhibiting features of higher grade tumors. To find functionally significant changes in transition from pre-invasive to invasive phenotype, we performed attribute profile clustering analysis on the list of genes differentially expressed between pre-invasive and invasive cells. We found integral membrane proteins, transcription factors, kinases, transport molecules, and chemokines to be highly represented. In addition, expression of matrix metalloproteinases MMP-9,-13,-15,-17 was up regulated in the invasive cells. Using siRNA based approaches, we found these MMPs to be required for the invasive phenotype. This model provides a new tool for dissection of mechanisms by which pre-invasive breast cells could acquire invasiveness in a metaplastic context.

  8. Loss of prostasin (PRSS8 in human bladder transitional cell carcinoma cell lines is associated with epithelial-mesenchymal transition (EMT

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    Chai Karl X

    2009-10-01

    Full Text Available Abstract Background The glycosylphosphatidylinositol (GPI-anchored epithelial extracellular membrane serine protease prostasin (PRSS8 is expressed abundantly in normal epithelia and essential for terminal epithelial differentiation, but down-regulated in human prostate, breast, and gastric cancers and invasive cancer cell lines. Prostasin is involved in the extracellular proteolytic modulation of the epidermal growth factor receptor (EGFR and is an invasion suppressor. The aim of this study was to evaluate prostasin expression states in the transitional cell carcinomas (TCC of the human bladder and in human TCC cell lines. Methods Normal human bladder tissues and TCC on a bladder cancer tissue microarray (TMA were evaluated for prostasin expression by means of immunohistochemistry. A panel of 16 urothelial and TCC cell lines were evaluated for prostasin and E-cadherin expression by western blot and quantitative PCR, and for prostasin gene promoter region CpG methylation by methylation-specific PCR (MSP. Results Prostasin is expressed in the normal human urothelium and in a normal human urothelial cell line, but is significantly down-regulated in high-grade TCC and lost in 9 (of 15 TCC cell lines. Loss of prostasin expression in the TCC cell lines correlated with loss of or reduced E-cadherin expression, loss of epithelial morphology, and promoter DNA hypermethylation. Prostasin expression could be reactivated by demethylation or inhibition of histone deacetylase. Re-expression of prostasin or a serine protease-inactive variant resulted in transcriptional up-regulation of E-cadherin. Conclusion Loss of prostasin expression in bladder transitional cell carcinomas is associated with epithelial-mesenchymal transition (EMT, and may have functional implications in tumor invasion and resistance to chemotherapy.

  9. Simvastatin Attenuates TGF-β1-Induced Epithelial-Mesenchymal Transition in Human Alveolar Epithelial Cells

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    Tuo Yang

    2013-06-01

    Full Text Available Background: Transforming growth factor-β1 (TGF-β1-induced epithelial-mesenchymal transition (EMT of alveolar epithelial cells (AEC may contribute to idiopathic pulmonary fibrosis (IPF. TGF-β1-induced EMT in A549 cells (a human AEC cell line resulted in the adoption of mesenchymal responses that were predominantly mediated via the TGF-β1-Smad2/3 signaling pathway. Simvastatin (Sim, a 3-hydroxy-3-methylglutaryl CoA (HMG-CoA reductase inhibitor, has been previously reported to inhibit EMT in human proximal tubular epithelial cells and porcine lens epithelial cells and to suppress Smad2/3 phosphorylation in animal models. However, whether Sim can attenuate TGF-β1-induced EMT in A549 cells and its underlying mechanisms remains unknown. Methods: Cells were incubated with TGF-β1 in the presence or absence of Sim. The epithelial marker E-cadherin (E-Cad and the mesenchymal markers, α-smooth muscle actin (α-SMA, vimentin (Vi and fibronectin (FN, were detected using western blotting analyses and immunofluorescence. Phosphorylated Smad2 and Smad3 levels and connective tissue growth factor (CTGF were analyzed using western blotting. In addition, a cell migration assay was performed. Moreover, the levels of matrix metalloproteinase (MMP-2 and -9 in the culture medium were examined using ELISA. Results: Sim significantly attenuated the TGF-β1-induced decrease in E-Cad levels and elevated the levels of α-SMA, Vi and FN via the suppression of Smad2 and Smad3 phosphorylation. Furthermore, Sim inhibited the mesenchymal-like responses in A549 cells, including cell migration, CTGF expression and secretion of MMP-2 and -9. However, Sim failed to reverse the cell morphologial changes induced by TGF-β1 in A549 cells. Conclusion: Sim attenuated TGF-β1-induced EMT in A549 cells and might be a promising therapeutic agent for treating IPF.

  10. Benzyl isothiocyanate inhibits epithelial-mesenchymal transition in cultured and xenografted human breast cancer cells.

    Science.gov (United States)

    Sehrawat, Anuradha; Singh, Shivendra V

    2011-07-01

    We showed previously that cruciferous vegetable constituent benzyl isothiocyanate (BITC) inhibits growth of cultured and xenografted human breast cancer cells and suppresses mammary cancer development in a transgenic mouse model. We now show, for the first time, that BITC inhibits epithelial-mesenchymal transition (EMT) in human breast cancer cells. Exposure of estrogen-independent MDA-MB-231 and estrogen-responsive MCF-7 human breast cancer cell lines and a pancreatic cancer cell line (PL-45) to BITC resulted in upregulation of epithelial markers (e.g., E-cadherin and/or occludin) with a concomitant decrease in protein levels of mesenchymal markers, including vimentin, fibronectin, snail, and/or c-Met. The BITC-mediated induction of E-cadherin protein was accompanied by an increase in its transcription, whereas BITC-treated MDA-MB-231 cells exhibited suppression of vimentin, snail, and slug mRNA levels. Experimental EMT induced by exposure to TGFβ and TNFα or Rb knockdown in a spontaneously immortalized nontumorigenic human mammary epithelial cell line (MCF-10A) was also partially reversed by BITC treatment. The TGFβ-/TNFα-induced migration of MCF-10A cells was inhibited in the presence of BITC, which was partially attenuated by RNA interference of E-cadherin. Inhibition of MDA-MB-231 xenograft growth in vivo in female athymic mice by BITC administration was associated with an increase in protein level of E-cadherin and suppression of vimentin and fibronectin protein expression. In conclusion, this study reports a novel anticancer effect of BITC involving inhibition of EMT, a process triggered during progression of cancer to invasive state.

  11. Triclosan potentiates epithelial-to-mesenchymal transition in anoikis-resistant human lung cancer cells.

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    Thidarat Winitthana

    Full Text Available Alteration of cancer cell toward mesenchymal phenotype has been shown to potentiate tumor aggressiveness by increasing cancer cell metastasis. Herein, we report the effect of triclosan, a widely used antibacterial agent found in many daily products, in enhancing the epithelial-to-mesenchymal transition (EMT in aggressive anoikis resistant human H460 lung cancer cells. EMT has been long known to increase abilities of the cells to increase migration, invasion, and survival in circulating system. The present study reveals that treatment of the cancer cells with triclosan at the physiologically related concentrations significantly increased the colony number of the cancer cells assessed by tumor formation assay. Also, the mesenchymal-like morphology and decrease in cell-to-cell adhesion were observed in triclosan-treated cells. Importantly, western blot analysis revealed that triclosan-treated cells exhibited decreased E-cadherin, while the levels of EMT markers, namely N-cadherin, vimentin, snail and slug were found to be significantly up-regulated. Furthermore, EMT induced by triclosan treatment was accompanied by the activation of focal adhesion kinase/ATP dependent tyrosine kinase (FAK/Akt and Ras-related C3 botulinum toxin substrate 1 (Rac1, which enhanced the ability of the cells to migrate and invade. In conclusion, we demonstrated for the first time that triclosan may potentiate cancer cells survival in detached condition and motility via the process of EMT. As mentioned capabilities are required for success in metastasis, the present study provides the novel toxicological information and encourages the awareness of triclosan use in cancer patients.

  12. Triclosan potentiates epithelial-to-mesenchymal transition in anoikis-resistant human lung cancer cells.

    Science.gov (United States)

    Winitthana, Thidarat; Lawanprasert, Somsong; Chanvorachote, Pithi

    2014-01-01

    Alteration of cancer cell toward mesenchymal phenotype has been shown to potentiate tumor aggressiveness by increasing cancer cell metastasis. Herein, we report the effect of triclosan, a widely used antibacterial agent found in many daily products, in enhancing the epithelial-to-mesenchymal transition (EMT) in aggressive anoikis resistant human H460 lung cancer cells. EMT has been long known to increase abilities of the cells to increase migration, invasion, and survival in circulating system. The present study reveals that treatment of the cancer cells with triclosan at the physiologically related concentrations significantly increased the colony number of the cancer cells assessed by tumor formation assay. Also, the mesenchymal-like morphology and decrease in cell-to-cell adhesion were observed in triclosan-treated cells. Importantly, western blot analysis revealed that triclosan-treated cells exhibited decreased E-cadherin, while the levels of EMT markers, namely N-cadherin, vimentin, snail and slug were found to be significantly up-regulated. Furthermore, EMT induced by triclosan treatment was accompanied by the activation of focal adhesion kinase/ATP dependent tyrosine kinase (FAK/Akt) and Ras-related C3 botulinum toxin substrate 1 (Rac1), which enhanced the ability of the cells to migrate and invade. In conclusion, we demonstrated for the first time that triclosan may potentiate cancer cells survival in detached condition and motility via the process of EMT. As mentioned capabilities are required for success in metastasis, the present study provides the novel toxicological information and encourages the awareness of triclosan use in cancer patients.

  13. Changing nuclear landscape and unique PML structures during early epigenetic transitions of human embryonic stem cells.

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    Butler, John T; Hall, Lisa L; Smith, Kelly P; Lawrence, Jeanne B

    2009-07-01

    The complex nuclear structure of somatic cells is important to epigenomic regulation, yet little is known about nuclear organization of human embryonic stem cells (hESC). Here we surveyed several nuclear structures in pluripotent and transitioning hESC. Observations of centromeres, telomeres, SC35 speckles, Cajal Bodies, lamin A/C and emerin, nuclear shape and size demonstrate a very different "nuclear landscape" in hESC. This landscape is remodeled during a brief transitional window, concomitant with or just prior to differentiation onset. Notably, hESC initially contain abundant signal for spliceosome assembly factor, SC35, but lack discrete SC35 domains; these form as cells begin to specialize, likely reflecting cell-type specific genomic organization. Concomitantly, nuclear size increases and shape changes as lamin A/C and emerin incorporate into the lamina. During this brief window, hESC exhibit dramatically different PML-defined structures, which in somatic cells are linked to gene regulation and cancer. Unlike the numerous, spherical somatic PML bodies, hES cells often display approximately 1-3 large PML structures of two morphological types: long linear "rods" or elaborate "rosettes", which lack substantial SUMO-1, Daxx, and Sp100. These occur primarily between Day 0-2 of differentiation and become rare thereafter. PML rods may be "taut" between other structures, such as centromeres, but clearly show some relationship with the lamina, where PML often abuts or fills a "gap" in early lamin A/C staining. Findings demonstrate that pluripotent hES cells have a markedly different overall nuclear architecture, remodeling of which is linked to early epigenomic programming and involves formation of unique PML-defined structures.

  14. Transitional features in human atherosclerosis. Intimal thickening, cholesterol clefts, and cell loss in human aortic fatty streaks.

    Science.gov (United States)

    Guyton, J. R.; Klemp, K. F.

    1993-01-01

    The possible transition from a subset of fatty streaks to fibrous plaques in human atherosclerosis has long been postulated, but transitional features in lesions have rarely been demonstrated. We examined human aortic fatty streaks to determine whether significant tendencies toward intimal thickening and toward deep extracellular lipid deposition might be found. To provide accurate ultrastructural assessment of lipid, tissues were processed by new electron microscopic cytochemical techniques. Unilateral fatty streaks exhibited a 60% increase in intimal thickness when compared to contralateral control tissue. Fat droplets in intimal cells accounted for approximately half of the increase; nonfat portions of cells and extracellular matrix accounted for the remainder. Six of 32 fatty streaks (19%) contained cholesterol clefts, which were found in the musculo-elastic (deep) layer of the intima or in the tunica media. Volume fractions occupied by cells in deep intima were reduced when cholesterol clefts were evident, suggesting loss of cells in early core regions. Light and electron microscopy showed structures consistent with lipid-rich core regions in lesions with cholesterol clefts and in a few lesions without cholesterol clefts. The findings of intimal thickening, core region formation, and disappearance of intimal cells constitute new evidence that some fatty streaks are progressive lesions and sites of eventual fibrous plaque development. The findings also suggest that the lipid-rich core region does not originate primarily from the debris of dead foam cells in the superficial intima, but instead arises from lipids accumulating gradually in the extracellular matrix of the deep intima. Images Figure 1 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 PMID:8238260

  15. Human NUMB6 Induces Epithelial-Mesenchymal Transition and Enhances Breast Cancer Cells Migration and Invasion.

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    Karaczyn, Aldona A; Adams, Tamara L; Cheng, Robert Y S; Matluk, Nicholas N; Verdi, Joseph M

    2017-02-01

    Mammalian NUMB is alternatively spliced generating four isoforms NUMB1-NUMB4 that can function as tumor suppressors. NUMB1-NUMB4 proteins, which normally determine how different cell types develop, are reduced in 21% of primary breast tumors. Our previous work has, however, indicated that two novel NUMB isoforms, NUMB5 and NUMB6 have the pro-oncogenic functions. Herein, we address a novel function of human NUMB isoform 6 (NUMB6) in promoting cancer cell migration and invasion. We found that NUMB6 induced expression of embryonic transcription factor Slug, which in turn actively repressed E-cadherin, prompting cells to undergo epithelial-mesenchymal transition (EMT). Low-metastatic breast cancer cells DB-7 stably expressing NUMB6, lost their epithelial phenotype, exhibited migratory and pro-invasive behavior, and ultimately elevated expression of mesenchymal markers. Among these markers, increased vimentin, β-catenin, and fibronectin expression elicited metalloproteinase 9 (MMP9) production. Our results revealed that NUMB6-DB-7 cells have significantly increased level of Akt1 and Akt2 phosphorylation. Therefore, antagonizing Akt signaling using a chemical inhibitor LY294002, we found that NUMB6-induced Slug expression was reduced, and ultimately accompanied with decreased cell migration and invasion. In summary, this study identified a novel molecular determinant of breast cancer progression, uncovering a potential oncogenic role for the NUMB6 protein in cancer cell migration and invasion, coupled to the maintenance of mesenchymal-like cells. J. Cell. Biochem. 118: 237-251, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  16. High glucose mediates endothelial-to-chondrocyte transition in human aortic endothelial cells

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    Tang Rining

    2012-09-01

    Full Text Available Abstract Background Vascular calcification is one of the common complications in diabetes mellitus. Many studies have shown that high glucose (HG caused cardiovascular calcification, but its underlying mechanism is not fully understood. Recently, medial calcification has been most commonly described in the vessels of patients with diabetes. Chondrocytes were involved in the medial calcification. Recent studies have shown that the conversion into mesenchymal stem cells (MSCs via the endothelial-to-mesenchymal transition (EndMT could be triggered in chondrocytes. Our previous research has indicated that HG induced EndMT in human aortic endothelial cells (HAECs. Therefore, we addressed the question of whether HG-induced EndMT could be transitioned into MSCs and differentiated into chondrocytes. Methods HAECs were divided into three groups: a normal glucose (NG group, HG group (30 mmol/L, and mannitol (5.5 mmol/L NG + 24.5 mmol/L group. Pathological changes were investigated using fluorescence microscopy and electron microscopy. Immunofluorescence staining was performed to detect the co-expression of endothelial markers, such as CD31, and fibroblast markers, such as fibroblast-specific protein 1 (FSP-1. The expression of FSP-1 was detected by real time-PCR and western blots. Endothelial-derived MSCs were grown in MSC medium for one week. The expression of the MSCs markers STRO-1, CD44, CD10 and the chondrocyte marker SOX9 was detected by immunofluorescence staining and western blots. Chondrocyte expression was detected by alcian blue staining. Calcium deposits were analyzed by alizarin red staining. Results The incubation of HAECs exposed to HG resulted in a fibroblast-like phenotype. Double staining of the HAECs indicated a co-localization of CD31 and FSP-1. The expression of FSP-1 was significantly increased in the HG group, and the cells undergoing EndMT also expressed STRO-1, CD44 and SOX9 compared with the controls (P  Conclusions Our

  17. Transitional features in human atherosclerosis. Intimal thickening, cholesterol clefts, and cell loss in human aortic fatty streaks.

    OpenAIRE

    Guyton, J. R.; Klemp, K. F.

    1993-01-01

    The possible transition from a subset of fatty streaks to fibrous plaques in human atherosclerosis has long been postulated, but transitional features in lesions have rarely been demonstrated. We examined human aortic fatty streaks to determine whether significant tendencies toward intimal thickening and toward deep extracellular lipid deposition might be found. To provide accurate ultrastructural assessment of lipid, tissues were processed by new electron microscopic cytochemical techniques....

  18. Transitional features in human atherosclerosis. Intimal thickening, cholesterol clefts, and cell loss in human aortic fatty streaks.

    OpenAIRE

    Guyton, J. R.; Klemp, K. F.

    1993-01-01

    The possible transition from a subset of fatty streaks to fibrous plaques in human atherosclerosis has long been postulated, but transitional features in lesions have rarely been demonstrated. We examined human aortic fatty streaks to determine whether significant tendencies toward intimal thickening and toward deep extracellular lipid deposition might be found. To provide accurate ultrastructural assessment of lipid, tissues were processed by new electron microscopic cytochemical techniques....

  19. Human cancer cells express Slug-based epithelial-mesenchymal transition gene expression signature obtained in vivo

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    Anastassiou Dimitris

    2011-12-01

    Full Text Available Abstract Background The biological mechanisms underlying cancer cell motility and invasiveness remain unclear, although it has been hypothesized that they involve some type of epithelial-mesenchymal transition (EMT. Methods We used xenograft models of human cancer cells in immunocompromised mice, profiling the harvested tumors separately with species-specific probes and computationally analyzing the results. Results Here we show that human cancer cells express in vivo a precise multi-cancer invasion-associated gene expression signature that prominently includes many EMT markers, among them the transcription factor Slug, fibronectin, and α-SMA. We found that human, but not mouse, cells express the signature and Slug is the only upregulated EMT-inducing transcription factor. The signature is also present in samples from many publicly available cancer gene expression datasets, suggesting that it is produced by the cancer cells themselves in multiple cancer types, including nonepithelial cancers such as neuroblastoma. Furthermore, we found that the presence of the signature in human xenografted cells was associated with a downregulation of adipocyte markers in the mouse tissue adjacent to the invasive tumor, suggesting that the signature is triggered by contextual microenvironmental interactions when the cancer cells encounter adipocytes, as previously reported. Conclusions The known, precise and consistent gene composition of this cancer mesenchymal transition signature, particularly when combined with simultaneous analysis of the adjacent microenvironment, provides unique opportunities for shedding light on the underlying mechanisms of cancer invasiveness as well as identifying potential diagnostic markers and targets for metastasis-inhibiting therapeutics.

  20. Human RAD6 Promotes G1-S Transition and Cell Proliferation through Upregulation of Cyclin D1 Expression

    Science.gov (United States)

    Biskup, Ewelina; Liu, Yan; Chen, Pei-Chao; Chang, Jian-Feng; Jiang, Wenjie; Jing, Yuanya; Chen, Youwei; Jin, Hui; Chen, Su

    2014-01-01

    Protein ubiquitinylation regulates protein stability and activity. RAD6, an E2 ubiquitin-conjugating enzyme, which that has been substantially biochemically characterized, functions in a number of biologically relevant pathways, including cell cycle progression. In this study, we show that RAD6 promotes the G1-S transition and cell proliferation by regulating the expression of cyclin D1 (CCND1) in human cells. Furthermore, our data indicate that RAD6 influences the transcription of CCND1 by increasing monoubiquitinylation of histone H2B and trimethylation of H3K4 in the CCND1 promoter region. Our study presents, for the first time, an evidence for the function of RAD6 in cell cycle progression and cell proliferation in human cells, raising the possibility that RAD6 could be a new target for molecular diagnosis and prognosis in cancer therapeutics. PMID:25409181

  1. Human RAD6 promotes G1-S transition and cell proliferation through upregulation of cyclin D1 expression.

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    Fengfeng Cai

    Full Text Available Protein ubiquitinylation regulates protein stability and activity. RAD6, an E2 ubiquitin-conjugating enzyme, which that has been substantially biochemically characterized, functions in a number of biologically relevant pathways, including cell cycle progression. In this study, we show that RAD6 promotes the G1-S transition and cell proliferation by regulating the expression of cyclin D1 (CCND1 in human cells. Furthermore, our data indicate that RAD6 influences the transcription of CCND1 by increasing monoubiquitinylation of histone H2B and trimethylation of H3K4 in the CCND1 promoter region. Our study presents, for the first time, an evidence for the function of RAD6 in cell cycle progression and cell proliferation in human cells, raising the possibility that RAD6 could be a new target for molecular diagnosis and prognosis in cancer therapeutics.

  2. Ultrastructural visualization of the Mesenchymal-to-Epithelial Transition during reprogramming of human fibroblasts to induced pluripotent stem cells

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    M.K. Høffding

    2015-01-01

    Here, we integrate a panel of morphological approaches with gene expression analyses to visualize the dynamics of episomal reprogramming of human fibroblasts to iPSCs. We provide the first ultrastructural analysis of human fibroblasts at various stages of episomal iPSC reprogramming, as well as the first real-time live cell visualization of a MET occurring during reprogramming. The results indicate that the MET manifests itself approximately 6–12 days after electroporation, in synchrony with the upregulation of early pluripotency markers, and resembles a reversal of the Epithelial-to-Mesenchymal Transition (EMT which takes place during mammalian gastrulation.

  3. Kaempferol modulates the metastasis of human non-small cell lung cancer cells by inhibiting epithelial-mesenchymal transition

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    Meng Hang

    2015-06-01

    Full Text Available The present study was done to determine whether kaempferol, a natural polyphenol of the flavonoid family, affects Epithelial-Mesenchymal Transition (EMT in non-small cell lung cancer cells. Kaempferol not only inhibited cancer cell proliferation and migration in a dose-dependent manner but also modulated the expression of EMT-related proteins E-cadherin and vimentin which are indispensible to cellular motility, invasiveness and metastasis. These results indicate that kaempferol suppresses non-small cell lung cancer migration by modulating the expression of EMT proteins. Therefore, kaempferol may be useful as a potential anticancer agent for non-small cell lung cancer.

  4. Matrine inhibits the invasive properties of human glioma cells by regulating epithelial‑to‑mesenchymal transition.

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    Wang, Zhongwei; Wu, Yi; Wang, Yali; Jin, Yingying; Ma, Xiulong; Zhang, Yang; Ren, Hongtao

    2015-05-01

    Matrine is reported to be effective in tumor therapies; however, the anti‑metastatic effect and molecular mechanism(s) of matrine on glioma remain poorly understood. Therefore, the purpose of this study was to assess the effects of matrine on glioma and the associated mechanism(s). In the study, we demonstrated that matrine inhibited the proliferation of glioma cells. We also observed that matrine inhibited the migration and invasion of glioma cells at non‑toxic concentrations. Matrine also decreased the expression of E‑cadherin and increased the expression of N‑cadherin. These results suggest that the anti‑metastatic effect of matrine may be correlated with epithelial‑to‑mesenchymal transition (EMT). Moreover, matrine could reduce the phosphorylation levels of p38 and AKT proteins. In conclusion, these results suggest matrine may be a potential alternative against invasive glioma cells via the p38 MAPK and AKT signaling‑dependent inhibition of EMT.

  5. Ursolic acid inhibits the proliferation of human ovarian cancer stem-like cells through epithelial-mesenchymal transition.

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    Zhang, Jie; Wang, Wenjing; Qian, Lin; Zhang, Qiuwan; Lai, Dongmei; Qi, Cong

    2015-11-01

    Ovarian cancer is the most frequent cause of cancer-related death among all gynecological cancers. Increasing evidence suggests that human ovarian cancer stem-like cells could be enriched under serum-free culture conditions. In the present study, SKOV3 ovarian epithelial cancer cells were cultured for sphere cells. Ursolic acid (UA) with triterpenoid compounds exist widely in food, medicinal herbs and other plants. Evidence shows that UA has anticancer activities in human ovarian cancer cells, but he role of UA in ovarian cancer stem cells (CSCs) remains unknown. The aim of the present study was to investigate the anticancer effects of UA in combination with cisplatin in ovarian CSCs (in vitro and in vivo), along with the molecular mechanism of action. Treatment with UA at various concentrations was examined in combination with cisplatin in human ovarian CSCs. MTT assay and flow cytometry were used for cell viability and apoptosis analysis, and qRT-PCR for stem cell markers and epithelial-mesenchymal transition (EMT) markers for mRNA expression. Transwell assay was employed to observe the migration and invasion of SKOV3 cells and SKOV3 sphere cells after treatment. Moreover, athymic BALB/c-nu nude mice were injected with SKOV3 sphere cells to obtain a xenograft model for in vivo studies. The results showed that CSCs possessed mesenchymal characteristics and EMT ability, and the growth of SKOV3 and sphere cells was significantly inhibited by UA. Transplanted tumors were significantly reduced after injection of UA and UA plus cisplatin. Furthermore, we found that UA could play a role in enhancing the sensitivity of CSCs to cisplatin resistance. Our findings suggested that UA is involved in EMT mechanism to affect the proliferation and apoptosis of human ovarian cancer stem-like cells and it is a potent anti-ovarian cancer agent.

  6. Loss of γ-cytoplasmic actin triggers myofibroblast transition of human epithelial cells.

    Science.gov (United States)

    Lechuga, Susana; Baranwal, Somesh; Li, Chao; Naydenov, Nayden G; Kuemmerle, John F; Dugina, Vera; Chaponnier, Christine; Ivanov, Andrei I

    2014-10-15

    Transdifferentiation of epithelial cells into mesenchymal cells and myofibroblasts plays an important role in tumor progression and tissue fibrosis. Such epithelial plasticity is accompanied by dramatic reorganizations of the actin cytoskeleton, although mechanisms underlying cytoskeletal effects on epithelial transdifferentiation remain poorly understood. In the present study, we observed that selective siRNA-mediated knockdown of γ-cytoplasmic actin (γ-CYA), but not β-cytoplasmic actin, induced epithelial-to-myofibroblast transition (EMyT) of different epithelial cells. The EMyT manifested by increased expression of α-smooth muscle actin and other contractile proteins, along with inhibition of genes responsible for cell proliferation. Induction of EMyT in γ-CYA-depleted cells depended on activation of serum response factor and its cofactors, myocardial-related transcriptional factors A and B. Loss of γ-CYA stimulated formin-mediated actin polymerization and activation of Rho GTPase, which appear to be essential for EMyT induction. Our findings demonstrate a previously unanticipated, unique role of γ-CYA in regulating epithelial phenotype and suppression of EMyT that may be essential for cell differentiation and tissue fibrosis.

  7. Paeoniflorin prevents hypoxia-induced epithelial–mesenchymal transition in human breast cancer cells

    Directory of Open Access Journals (Sweden)

    Zhou Z

    2016-04-01

    Full Text Available Zhenyu Zhou,1,* Shunchang Wang,1,* Caijuan Song,2 Zhuang Hu11Department of Thyroid and Breast, Huaihe Hospital, Henan University, Kaifeng, 2Department of Immunization Program, Zhengzhou Center for Disease Control and Prevention, Zhengzhou, People’s Republic of China*These authors contributed equally to this workAbstract: Paeoniflorin (PF is a monoterpene glycoside extracted from the root of Paeonia lactiflora Pall. Previous studies have demonstrated that PF inhibits the growth, invasion, and metastasis of tumors in vivo and in vitro. However, the effect of PF on hypoxia-induced epithelial–mesenchymal transition (EMT in breast cancer cells remains unknown. Therefore, the objective of this study was to investigate the effect of PF on hypoxia-induced EMT in breast cancer cells, as well as characterize the underlying mechanism. The results presented in this study demonstrate that PF blocks the migration and invasion of breast cancer cells by repressing EMT under hypoxic conditions. PF also significantly attenuated the hypoxia-induced increase in HIF-1α level. Furthermore, PF prevented hypoxia-induced expression of phosphorylated PI3K and Akt in MDA-MB-231 cells. In conclusion, PF prevented hypoxia-induced EMT in breast cancer cells by inhibiting HIF-1α expression via modulation of PI3K/Akt signaling pathway. This finding provides evidence that PF can serve as a therapeutic agent for the treatment of breast cancer.Keywords: paeoniflorin, breast cancer, hypoxia, epithelial–mesenchymal transition, PI3K/Akt signaling pathway

  8. Suppression of FOXQ1 in benzyl isothiocyanate-mediated inhibition of epithelial-mesenchymal transition in human breast cancer cells.

    Science.gov (United States)

    Sehrawat, Anuradha; Kim, Su-Hyeong; Vogt, Andreas; Singh, Shivendra V

    2013-04-01

    We showed previously that breast cancer chemoprevention with benzyl isothiocyanate (BITC) in MMTV-neu mice was associated with induction of E-cadherin protein in vivo. Loss of E-cadherin expression and induction of mesenchymal markers (e.g. vimentin) are biochemical hallmarks of epithelial-mesenchymal transition (EMT), a developmental process implicated in progression of cancer to aggressive state. This study offers novel insights into the mechanism by which BITC inhibits EMT. Exposure of MDA-MB-231, SUM159 and MDA-MB-468 human breast cancer cells to BITC (2.5 and 5 µM) resulted in transcriptional repression of urokinase-type plasminogen activator (uPA) as well as its receptor (uPAR). However, ectopic expression of uPAR in MDA-MB-468 cells failed to confer protection against induction of E-cadherin and inhibition of cell invasion/migration resulting from BITC treatment. The BITC-mediated induction of E-cadherin and inhibition of cell migration was sustained in MDA-MB-231 and SUM159 cells transiently transfected with an uPAR-targeted small interfering RNA. Overexpression of Forkhead Box Q1 (FOXQ1), whose protein and messenger RNA levels were decreased by BITC treatment in cells and MDA-MB-231 xenografts, conferred marked protection against BITC-mediated inhibition of EMT and cell migration. In conclusion, this study implicates FOXQ1 suppression in BITC-mediated inhibition of EMT in human breast cancer cells.

  9. Overexpression of snail induces epithelial-mesenchymal transition and a cancer stem cell-like phenotype in human colorectal cancer cells.

    Science.gov (United States)

    Fan, Fan; Samuel, Shaija; Evans, Kurt W; Lu, Jia; Xia, Ling; Zhou, Yunfei; Sceusi, Eric; Tozzi, Federico; Ye, Xiang-Cang; Mani, Sendurai A; Ellis, Lee M

    2012-08-01

    Epithelial-mesenchymal transition (EMT) is a critical process providing tumor cells with the ability to migrate and escape from the primary tumor and metastasize to distant sites. Recently, EMT was shown to be associated with the cancer stem cell (CSC) phenotype in breast cancer. Snail is a transcription factor that mediates EMT in a number of tumor types, including colorectal cancer (CRC). Our study was done to determine the role of Snail in mediating EMT and CSC function in CRC. Human CRC specimens were stained for Snail expression, and human CRC cell lines were transduced with a retroviral Snail construct or vector control. Cell proliferation and chemosensitivity to oxaliplatin of the infected cells were determined by the MTT (colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Migration and invasion were determined in vitro using modified Boyden chamber assays. EMT and putative CSC markers were analyzed using Western blotting. Intravenous injection of tumor cells was done to evaluate their metastatic potential in mice. Snail was overexpressed in human CRC surgical specimens. This overexpression induced EMT and a CSC-like phenotype in human CRC cells and enhanced cell migration and invasion (P Snail overexpression also led to an increase in metastasis formation in vivo (P Snail-overexpressing CRC cells were more chemoresistant to oxaliplatin than control cells. Increased Snail expression induces EMT and the CSC-like phenotype in CRC cells, which enhance cancer cell invasion and chemoresistance. Thus, Snail is a potential therapeutic target in metastatic CRC.

  10. AGE-RAGE interaction in the TGFβ2-mediated epithelial to mesenchymal transition of human lens epithelial cells.

    Science.gov (United States)

    Raghavan, Cibin T; Nagaraj, Ram H

    2016-08-01

    Basement membrane (BM) proteins accumulate chemical modifications with age. One such modification is glycation, which results in the formation of advanced glycation endproducts (AGEs). In a previous study, we reported that AGEs in the human lens capsule (BM) promote the TGFβ2-mediated epithelial-to-mesenchymal transition (EMT) of lens epithelial cells, which we proposed as a mechanism for posterior capsule opacification (PCO) or secondary cataract formation. In this study, we investigated the role of a receptor for AGEs (RAGE) in the TGFβ2-mediated EMT in a human lens epithelial cell line (FHL124). RAGE was present in FHL124 cells, and its levels were unaltered in cells cultured on either native or AGE-modified BM or upon treatment with TGFβ2. RAGE overexpression significantly enhanced the TGFβ2-mediated EMT responses in cells cultured on AGE-modified BM compared with the unmodified matrix. In contrast, treatment of cells with a RAGE antibody or EN-RAGE (an endogenous ligand for RAGE) resulted in a significant reduction in the TGFβ2-mediated EMT response. This was accompanied by a reduction in TGFβ2-mediated Smad signaling and ROS generation. These results imply that the interaction of matrix AGEs with RAGE plays a role in the TGFβ2-mediated EMT of lens epithelial cells and suggest that the blockade of RAGE could be a strategy to prevent PCO and other age-associated fibrosis.

  11. Cholesterol crystallization in human atherosclerosis is triggered in smooth muscle cells during the transition from fatty streak to fibroatheroma.

    Science.gov (United States)

    Ho-Tin-Noé, Benoît; Vo, Sophie; Bayles, Richard; Ferrière, Stephen; Ladjal, Hayette; Toumi, Sondes; Deschildre, Catherine; Ollivier, Véronique; Michel, Jean-Baptiste

    2017-04-01

    Recent studies have shown that in addition to being major constituents of the atheromatous core, solid cholesterol crystals (CCs) promote atherosclerotic lesion development and rupture by causing mechanical damage and exerting cytotoxic and pro-inflammatory effects. These findings suggest that targeting CCs might represent a therapeutic strategy for plaque stabilization. However, little is known about how cholesterol crystallization is initiated in human atherothrombotic disease. Here, we investigated these mechanisms. We performed a thorough immunohistological analysis of non-embedded, minimally processed human aortic tissues, combining polarized light and fluorescence microscopy. We found that CC formation was initiated during the fatty streak to fibroatheroma transition in tight association with the death of intralesional smooth muscle cells (SMCs). Cholesterol-loaded human SMCs were capable of producing CCs in vitro, a process that was enhanced by type I collagen and by inhibition of autophagy and cholesterol esterification. The fibrous transition, which was characterized by increased type I collagen expression, was associated with changes in the expression of autophagy and cholesterol flux-related genes, including a decrease in the autophagic adapter p62 and an increase in the cholesterol intracellular transporter Niemann-Pick C1. Collagen was identified as a potent inducer of these changes in SMCs. Collagen-induced changes in cholesterol metabolism and autophagy flux in smooth muscle foam cells at the fibrolipid transition likely contribute to initiate cholesterol crystallization in human atherosclerosis. Also, our data are in support of a protective role of autophagy against CC formation. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  12. HNF1α inhibition triggers epithelial-mesenchymal transition in human liver cancer cell lines

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    Vignjevic Danijela

    2011-10-01

    Full Text Available Abstract Background Hepatocyte Nuclear Factor 1α (HNF1α is an atypical homeodomain-containing transcription factor that transactivates liver-specific genes including albumin, α-1-antitrypsin and α- and β-fibrinogen. Biallelic inactivating mutations of HNF1A have been frequently identified in hepatocellular adenomas (HCA, rare benign liver tumors usually developed in women under oral contraceptives, and in rare cases of hepatocellular carcinomas developed in non-cirrhotic liver. HNF1α-mutated HCA (H-HCA are characterized by a marked steatosis and show activation of glycolysis, lipogenesis, translational machinery and mTOR pathway. We studied the consequences of HNF1α silencing in hepatic cell lines, HepG2 and Hep3B and we reproduced most of the deregulations identified in H-HCA. Methods We transfected hepatoma cell lines HepG2 and Hep3B with siRNA targeting HNF1α and obtained a strong inhibition of HNF1α expression. We then looked at the phenotypic changes by microscopy and studied changes in gene expression using qRT-PCR and Western Blot. Results Hepatocytes transfected with HNF1α siRNA underwent severe phenotypic changes with loss of cell-cell contacts and development of migration structures. In HNF1α-inhibited cells, hepatocyte and epithelial markers were diminished and mesenchymal markers were over-expressed. This epithelial-mesenchymal transition (EMT was related to the up regulation of several EMT transcription factors, in particular SNAIL and SLUG. We also found an overexpression of TGFβ1, an EMT initiator, in both cells transfected with HNF1α siRNA and H-HCA. Moreover, TGFβ1 expression is strongly correlated to HNF1α expression in cell models, suggesting regulation of TGFβ1 expression by HNF1α. Conclusion Our results suggest that HNF1α is not only important for hepatocyte differentiation, but has also a role in the maintenance of epithelial phenotype in hepatocytes.

  13. Transition of late-stage effector T cells to CD27+ CD28+ tumor-reactive effector memory T cells in humans after adoptive cell transfer therapy

    Science.gov (United States)

    Powell, Daniel J.; Dudley, Mark E.; Robbins, Paul F.; Rosenberg, Steven A.

    2007-01-01

    In humans, the pathways of memory T-cell differentiation remain poorly defined. Recently, adoptive cell transfer (ACT) of tumor-reactive T lymphocytes to metastatic melanoma patients after nonmyeloablative chemotherapy has resulted in persistence of functional, tumor-reactive lymphocytes, regression of disease, and induction of melanocyte-directed autoimmunity in some responding patients. In the current study, longitudinal phenotypic analysis was performed on melanoma antigen–specific CD8+ T cells during their transition from in vitro cultured effector cells to long-term persistent memory cells following ACT to 6 responding patients. Tumor-reactive T cells used for therapy were generally late-stage effector cells with a CD27Lo CD28Lo CD45RA− CD62 ligand− (CD62L−) CC chemokine receptor 7− (CCR7−) interleukin-7 receptor αLo (IL-7RαLo) phenotype. After transfer, rapid up-regulation and continued expression of IL-7Rα in vivo suggested an important role for IL-7R in immediate and long-term T-cell survival. Although the tumor antigen–specific T-cell population contracted between 1 and 4 weeks after transfer, stable numbers of CD27+ CD28+ tumor-reactive T cells were maintained, demonstrating their contribution to the development of long-term, melanoma-reactive memory CD8+ T cells in vivo. At 2 months after transfer, melanoma-reactive T cells persisted at high levels and displayed an effector memory phenotype, including a CD27+ CD28+ CD62L− CCR7− profile, which may explain in part their ability to mediate tumor destruction. PMID:15345595

  14. G0-G1 cell cycle phase transition as revealed by fluorescence resonance energy transfer: analysis of human fibroblast chromatin.

    Science.gov (United States)

    Bottiroli, Giovanni; Croce, A C; Bottone, M G; Vaccino, S; Pellicciari, C

    2004-01-01

    In the present study, microspectrofluorometry and digital imaging procedures were used to investigate by fluorescence Resonance Energy Transfer (FRET) analysis the changes of chromatin organization during the transition from G0 quiescent stat to G1 phase. G0 transition is a key event in cell cycle progress depending on the activation of specific genes and the concomitant silencing of others, which both entail spatial chromatin rearrangement. Normal human fibroblasts arrested in G0-phase by culture in low-serum containing medium and stimulated to re-enter G1 by serum addition were used as cell model. To investigate the occurrence and timing of these supramolecular chromatin changes, we estimated the relative FRET efficiency in single cells after double-helical DNA. Hoechst 33258 amd propidium iodide were used as a donor-acceptor dye pair since they exhibit particularly favourable spectral characteristics, that allow the calculation procedure to be semplified. The results of FRET analysis were compared to those of the immunocytochemical labelling of two nuclear proteins (i.e., Ki-67 and statin) whose expression is an established marker of potentially proliferating G1 cells or resting G0 cells, respectively. FRET efficiency was lower in G0 than G1 fibroblasts: this is likely due to higher chromatin packaging in quiescent cells which especially hinders the interaction with the donor molecules less favourable, in terms of relative distance and spatial orientation. FRET efficiency significantly increased shortly (1h) after serum stimulation of quiescent fibroblasts, thus indicating that chromatin is rearranged in parallel with activation of cycle-related gene; it is worth noting that these signs largely preceded the occurrence of immunopositivity for Ki-67, which was detectable only 24h after serum stimulation. FRET-based analyses which already proved to be suitable for studying the overall chromatin organization in differentiated cells, may now be envisaged as a

  15. Natural suppression of human immunodeficiency virus type 1 replication is mediated by transitional memory CD8+ T cells.

    Science.gov (United States)

    Killian, M Scott; Johnson, Carl; Teque, Fernando; Fujimura, Sue; Levy, Jay A

    2011-02-01

    HIV replication is suppressed in vitro by a CD8(+) cell noncytotoxic antiviral response (CNAR). This activity directly correlates with an asymptomatic clinical state. The objective of this study was to identify the phenotype of CD8(+) cell subsets having strong CNAR activity. CD8(+) cell subset frequencies and CNAR levels were measured for human immunodeficiency virus (HIV)-uninfected individuals and three groups of HIV type 1 (HIV-1)-infected individuals: asymptomatic individuals with low-level viremia (vHIV), antiretroviral-drug-treated subjects with undetectable virus levels (TxHIV), and therapy-naïve aviremic elite controllers (EC). CD8(+) cells from the vHIV individuals exhibited the highest HIV-suppressing activity and had elevated frequencies of CD45RA(-) CD27(+) and PD-1(+) (CD279(+)) cells. Functional assessments of CD8(+) cells sorted into distinct subsets established that maximal CNAR activity was mediated by CD45RA(-) CCR7(-) CD27(+) and PD-1(+) CD8(+) cells. T cell receptor (TCR) repertoire profiles of CD8(+) cell subsets having strong CNAR activity exhibited increased perturbations in comparison to those of inactive subsets. Together, these studies suggest that CNAR is driven by HIV replication and that this antiviral activity is associated with oligoclonally expanded activated CD8(+) cells expressing PD-1 and having a transitional memory cell phenotype. The findings better describe the identity of CD8(+) cells showing CNAR and should facilitate the evaluation of this important immune response in studies of HIV pathogenesis, resistance to infection, and vaccine development.

  16. Muscadine grape skin extract reverts snail-mediated epithelial mesenchymal transition via superoxide species in human prostate cancer cells.

    Science.gov (United States)

    Burton, Liza J; Barnett, Petrina; Smith, Basil; Arnold, Rebecca S; Hudson, Tamaro; Kundu, Kousik; Murthy, Niren; Odero-Marah, Valerie A

    2014-03-12

    Snail transcription factor can induce epithelial-mesenchymal transition (EMT), associated with decreased cell adhesion-associated molecules like E-cadherin, increased mesenchymal markers like vimentin, leading to increased motility, invasion and metastasis. Muscadine grape skin extract (MSKE) has been shown to inhibit prostate cancer cell growth and induce apoptosis without affecting normal prostate epithelial cells. We investigated novel molecular mechanisms by which Snail promotes EMT in prostate cancer cells via Reactive Oxygen Species (ROS) and whether it can be antagonized by MSKE. ARCaP and LNCaP cells overexpressing Snail were utilized to examine levels of reactive oxygen species (ROS), specifically, superoxide, in vitro using Dihydroethidium (DHE) or HydroCy3 dyes. Mitosox staining was performed to determine whether the source of ROS was mitochondrial in origin. We also investigated the effect of Muscadine grape skin extract (MSKE) on EMT marker expression by western blot analysis. Migration and cell viability using MTS proliferation assay was performed following MSKE treatments. Snail overexpression in ARCaP and LNCaP cells was associated with increased concentration of mitochondrial superoxide, in vitro. Interestingly, MSKE decreased superoxide levels in ARCaP and LNCaP cells. Additionally, MSKE and Superoxide Dismutase (SOD) reverted EMT as evidenced by decreased vimentin levels and re-induction of E-cadherin expression in ARCaP-Snail cells after 3 days, concomitant with reduced cell migration. MSKE also decreased Stat-3 activity in ARCaP-Snail cells. This study shows that superoxide species may play a role in Snail transcription factor-mediated EMT. Therefore, therapeutic targeting of Snail with various antioxidants such as MSKE may prove beneficial in abrogating EMT and ROS-mediated tumor progression in human prostate cancer.

  17. Muscadine grape skin extract reverts snail-mediated epithelial mesenchymal transition via superoxide species in human prostate cancer cells

    Science.gov (United States)

    2014-01-01

    Background Snail transcription factor can induce epithelial-mesenchymal transition (EMT), associated with decreased cell adhesion-associated molecules like E-cadherin, increased mesenchymal markers like vimentin, leading to increased motility, invasion and metastasis. Muscadine grape skin extract (MSKE) has been shown to inhibit prostate cancer cell growth and induce apoptosis without affecting normal prostate epithelial cells. We investigated novel molecular mechanisms by which Snail promotes EMT in prostate cancer cells via Reactive Oxygen Species (ROS) and whether it can be antagonized by MSKE. Methods ARCaP and LNCaP cells overexpressing Snail were utilized to examine levels of reactive oxygen species (ROS), specifically, superoxide, in vitro using Dihydroethidium (DHE) or HydroCy3 dyes. Mitosox staining was performed to determine whether the source of ROS was mitochondrial in origin. We also investigated the effect of Muscadine grape skin extract (MSKE) on EMT marker expression by western blot analysis. Migration and cell viability using MTS proliferation assay was performed following MSKE treatments. Results Snail overexpression in ARCaP and LNCaP cells was associated with increased concentration of mitochondrial superoxide, in vitro. Interestingly, MSKE decreased superoxide levels in ARCaP and LNCaP cells. Additionally, MSKE and Superoxide Dismutase (SOD) reverted EMT as evidenced by decreased vimentin levels and re-induction of E-cadherin expression in ARCaP-Snail cells after 3 days, concomitant with reduced cell migration. MSKE also decreased Stat-3 activity in ARCaP-Snail cells. Conclusions This study shows that superoxide species may play a role in Snail transcription factor-mediated EMT. Therefore, therapeutic targeting of Snail with various antioxidants such as MSKE may prove beneficial in abrogating EMT and ROS-mediated tumor progression in human prostate cancer. PMID:24617993

  18. Transition metal sensing by Toll-like receptor-4: next to nickel, cobalt and palladium are potent human dendritic cell stimulators

    NARCIS (Netherlands)

    Rachmawati, D.; Bontkes, H.J.; Verstege, M.I.; Muris, J.; von Blomberg, B.M.E.; Scheper, R.J.; van Hoogstraten, I.M.W.

    2013-01-01

    Background Nickel was recently identified as a potent activator of dendritic cells through ligating with human Toll-like receptor (TLR)-4. Objectives Here, we studied an extended panel of transition metals neighbouring nickel in the periodic table of elements, for their capacity to activate human mo

  19. Transition metal sensing by Toll-like receptor-4: next to nickel, cobalt and palladium are potent human dendritic cell stimulators

    NARCIS (Netherlands)

    Rachmawati, D.; Bontkes, H.J.; Verstege, M.I.; Muris, J.; von Blomberg, B.M.E.; Scheper, R.J.; van Hoogstraten, I.M.W.

    2013-01-01

    Background Nickel was recently identified as a potent activator of dendritic cells through ligating with human Toll-like receptor (TLR)-4. Objectives Here, we studied an extended panel of transition metals neighbouring nickel in the periodic table of elements, for their capacity to activate human mo

  20. Salinomycin suppresses TGF-β1-induced epithelial-to-mesenchymal transition in MCF-7 human breast cancer cells.

    Science.gov (United States)

    Zhang, Chunying; Lu, Ying; Li, Qing; Mao, Jun; Hou, Zhenhuan; Yu, Xiaotang; Fan, Shujun; Li, Jiazhi; Gao, Tong; Yan, Bing; Wang, Bo; Song, Bo; Li, Lianhong

    2016-03-25

    Epithelial-to-mesenchymal transition (EMT) is the major cause of breast cancer to initiate invasion and metastasis. Salinomycin (Sal) has been found as an effective chemical compound to kill breast cancer stem cells. However, the effect of Sal on invasion and metastasis of breast cancer is unclear. In the present study, we showed that Sal reversed transforming growth factor-β1 (TGF-β1) induced invasion and metastasis accompanied with down-regulation of MMP-2 by experiments on human breast cancer cell line MCF-7. Sal was able to inhibit TGF-β1-induced EMT phenotypic transition and the activation of key signaling molecules involved in Smad (p-Smad2/3,Snail1) and non-Smad (β-catenin, p-p38 MAPK) signals which cooperatively regulate the induction of EMT. Importantly, in a series of breast cancer specimens, we found strong correlation among E-cadherin expression, β-catenin expression, and the lymph node metastatic potential of breast cancer. Our research suggests that Sal is promised to be a chemotherapeutic drug by suppressing the metastasis of breast cancer.

  1. Interleukin-6 induces an epithelial-mesenchymal transition phenotype in human adamantinomatous craniopharyngioma cells and promotes tumor cell migration

    Science.gov (United States)

    Zhou, Jie; Zhang, Chao; Pan, Jun; Chen, Ligang; Qi, Song-Tao

    2017-01-01

    Total resection of adamantinomatous craniopharyngioma (ACP) is complex and often leads to postoperative recurrence. This is due to the tendency of the tumor to invade the surrounding brain tissue and the generation of a local inflammatory state between the tumor cells and parenchyma. While there is evidence to suggest that interleukin-6 (IL-6) induces craniopharyngioma (CP)-associated inflammation, particularly in ACP, the role of IL-6 in the progression of ACP remains unclear. The results of the present study demonstrated that CP inflammation was associated with pathological classification, extent of surgery, degree of calcification and postoperative hypothalamic status scale. Cytokine antibody arrays were conducted to measure the expression of IL-6 and other inflammatory factors in tumor tissues in response to various levels of inflammatory exposure. IL-6, IL-6 receptor (IL-6R) and glycoprotein 130 expression was detected by immunohistochemistry. In addition, an ELISA was performed to quantify the levels of soluble IL-6R (sIL-6R) in the cystic fluid and supernatants of ACP cells and tumor-associated fibroblasts. These measurements demonstrated that ACP cells produce IL-6 and its associated proteins. In addition, the results revealed that while the viability of ACP cells was not affected, the migration of ACP cells was promoted by IL-6 treatment in a concentration-dependent manner. Conversely, treatment with an IL-6-blocking monoclonal antibody significantly decreased the migration of ACP cells. In addition, IL-6 treatment increased the expression of vimentin and decreased the expression of E-cadherin in a dose-dependent manner. The findings of the present study demonstrate that IL-6 may promote migration in vitro via the classic- and trans-signaling pathways by inducing epithelial-mesenchymal transition in ACP cell cultures. PMID:28487953

  2. Phthalates stimulate the epithelial to mesenchymal transition through an HDAC6-dependent mechanism in human breast epithelial stem cells.

    Science.gov (United States)

    Hsieh, Tsung-Hua; Tsai, Cheng-Fang; Hsu, Chia-Yi; Kuo, Po-Lin; Lee, Jau-Nan; Chai, Chee-Yin; Hou, Ming-Feng; Chang, Chia-Cheng; Long, Cheng-Yu; Ko, Ying-Chin; Tsai, Eing-Mei

    2012-08-01

    Phthalates are environmental hormone-like molecules that are associated with breast cancer risk and are involved in metastasis, a process that requires the epithelial-mesenchymal transition (EMT). However, few studies have addressed the potential effects of phthalates on stem cells. Here we tested the hypothesis that phthalates such as butyl benzyl phthalate and di-n-butyl phthalate induce EMT in R2d cells, a stem cell-derived human breast epithelial cell line that is responsive to estradiol for tumor development. We observed that phthalates induced EMT as evidenced by morphological changes concomitant with increased expression of mesenchymal markers and decreased expression of epithelial markers. Molecular mechanism studies revealed that histone deacetylase 6 (HDAC6) is required for phthalate-induced cell migration and invasion during EMT in vitro and metastasis into the lungs of nude mice. We also constructed a series of mutant HDAC6 promoter fragments and found that the transcription factor AP-2a plays a novel role in regulating the HDAC6 promoter. Furthermore, phthalates stimulated estrogen receptors and triggered the downstream EGFR-PKA signaling cascade, leading to increased expression of AP-2a in the nucleus. We also observed that phthalates increased expression of the PP1/HDAC6 complex and caused Akt activation and GSK3β inactivation, leading to transcriptional activation of vimentin through the β-catenin-TCF-4/LEF1 pathway. Understanding the signaling cascades of phthalates that activate EMT through HDAC6 in breast epithelial stem cells provides the identification of novel therapeutic target for human breast cancer.

  3. Role of mitochondrial permeability transition in human hepatocellular carcinoma Hep-G2 cell death induced by rhein.

    Science.gov (United States)

    Du, Qiong; Bian, Xiao-Lan; Xu, Xiao-Le; Zhu, Bin; Yu, Bo; Zhai, Qing

    2013-12-01

    Rhein, a compound found as a glucoside in the root of rhubarb, is currently a subject of interest for its antitumor properties. The apoptosis of tumor cell lines induced by rhein was observed, and the involvement of mitochondria was established; however, the role of mitochondrial permeability transition (MPT) remains unknown. Here we report that MPT plays an important role in the apoptosis of human hepatocellular carcinoma Hep-G2 cells induced by rhein. After adding rhein to the isolated hepatic mitochondria, swelling effects and the leakage of Ca(2+) were observed. These alterations were suppressed by cyclosporin A (CsA), an MPT inhibitor. Furthermore, in Hep-G2 cells, the decrease of ATP production, the loss of mitochondrial transmembrane potential (MTP), the release of cytochrome c (Cyto c), and the activation of caspase 3 were also observed. These toxic effects of rhein can also be attenuated by CsA as well. Moreover, TUNEL assay confirmed that in the presence of CsA, rhein-induced apoptosis was largely inhibited. These results suggest that MPT plays a critical role in the pathogenesis of Hep-G2 cell injury induced by rhein, and imply that MPT may contribute to the anti-cancer activity of rhein. © 2013.

  4. G0-G1 cell cycle phase transition as revealed by fluorescence resonance energy transfer: analysis of human fibroblast chromatin

    Directory of Open Access Journals (Sweden)

    G Bottiroli

    2009-06-01

    Full Text Available In the present study, microspectrofluorometry and digital imaging procedures were used to investigate by fluorescence Resonance Energy Transfer (FRET analysis the changes of chromatin organization during the transition from G0 quiescent state to G1 phase. G0-G1 transition is a key event in cell cycle progress depending on the activation of specific genes and the concomitant silencing of others, which both entail spatial chromatin rearrangement. Normal human fibroblasts arrested in G0-phase by culture in low-serum contanining medium and stimulated to re-enter G1 by serum addition were used as cell model. To investigate the occurrence and timing of these supramolecular chromatin changes, we estimated the relative FRET efficiency in single cells after double-staining with two DNA-specific dyes which non-covalently bind to double-helical DNA. Hoechst 33258 and propidium iodide were used as a donor-acceptor dye pair since they exhibit particularly favourable spectral characteristics, that allow the calculation procedure to be semplified. The results of FRET analysis were compared to those of the immunocytochemical labelling of two nuclear proteins (i.e., Ki-67 and statin whose expression is an established marker of potentially proliferating G1 cells or resting G0 cells, respectively. FRET efficiency was lower in G0 than in G1 fibroblasts: this is likely due to a higher chromatin packaging in quiescent cells which especially hinders the intercalation of propidium iodide molecules, thus making the interaction with the donor molecules less favourable, in terms of relative distance and spatial orientation. FRET efficiency significantly increased shortly (1h after serum stimulation of quiescent fibroblasts, thus indicating that chromatin is rearranged in parallel with activation of cycle-related gene; it is worth noting that these signs largely preceded the occurrence of immunopositivity for Ki-67, which was detectable only 24h after serum stimulation

  5. α-Solanine inhibits invasion of human prostate cancer cell by suppressing epithelial-mesenchymal transition and MMPs expression.

    Science.gov (United States)

    Shen, Kun-Hung; Liao, Alex Chien-Hwa; Hung, Jui-Hsiang; Lee, Wei-Jiunn; Hu, Kai-Chieh; Lin, Pin-Tsen; Liao, Ruei-Fang; Chen, Pin-Shern

    2014-08-11

    α-Solanine, a naturally occurring steroidal glycoalkaloid found in nightshade (Solanum nigrum Linn.), was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in suppression of cancer cell metastasis by α-solanine remains unclear. This study investigates the suppression mechanism of α-solanine on motility of the human prostate cancer cell PC-3. Results show that α-solanine reduces the viability of PC-3 cells. When treated with non-toxic doses of α-solanine, cell invasion is markedly suppressed by α-solanine. α-Solanine also significantly elevates epithelial marker E-cadherin expression, while it concomitantly decreases mesenchymal marker vimentin expression, suggesting it suppresses epithelial-mesenchymal transition (EMT). α-Solanine reduces the mRNA level of matrix metalloproteinase-2 (MMP-2), MMP-9 and extracellular inducer of matrix metalloproteinase (EMMPRIN), but increases the expression of reversion-inducing cysteine-rich protein with kazal motifs (RECK), and tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2. Immunoblotting assays indicate α-solanine is effective in suppressing the phosphorylation of phosphatidylinositide-3 kinase (PI3K), Akt and ERK. Moreover, α-solanine downregulates oncogenic microRNA-21 (miR-21) and upregulates tumor suppressor miR-138 expression. Taken together, the results suggest that inhibition of PC-3 cell invasion by α-solanine may be, at least in part, through blocking EMT and MMPs expression. α-Solanine also reduces ERK and PI3K/Akt signaling pathways and regulates expression of miR-21 and miR-138. These findings suggest an attractive therapeutic potential of α-solanine for suppressing invasion of prostate cancer cell.

  6. Small interfering RNA targeting ILK inhibits metastasis in human tongue cancer cells through repression of epithelial-to-mesenchymal transition

    Energy Technology Data Exchange (ETDEWEB)

    Xing, Yu [College of Laboratory Medicine, Chongqing Medical University, Chongqing (China); Laboratory of Forensic Medicine and Biomedical Information, Chongqing Medical University, Chongqing (China); Qi, Jin [The Affiliated Hospital of Stomatology, Chongqing Medical University (China); Deng, Shixiong [College of Laboratory Medicine, Chongqing Medical University, Chongqing (China); Laboratory of Forensic Medicine and Biomedical Information, Chongqing Medical University, Chongqing (China); Wang, Cheng [Molecular Medicine and Cancer Research Center, Chongqing Medical University, Chongqing (China); Department of Cell Biology and Genetics, Chongqing Medical University, Chongqing (China); Zhang, Luyu [Molecular Medicine and Cancer Research Center, Chongqing Medical University, Chongqing (China); Chen, Junxia, E-mail: chjunxia@126.com [Molecular Medicine and Cancer Research Center, Chongqing Medical University, Chongqing (China); Department of Cell Biology and Genetics, Chongqing Medical University, Chongqing (China)

    2013-08-01

    Integrin-linked kinase (ILK) is a multifunctional serine/threonine kinase. Accumulating evidences suggest that ILK are involved in cell–matrix interactions, cell proliferation, invasion, migration, angiogenesis and Epithelial–mesenchymal transition (EMT). However, the underlying mechanisms remain largely unknown. EMT has been postulated as a prerequisite for metastasis. The reports have demonstrated that EMT was implicated in metastasis of oral squamous cell carcinomas. Therefore, here we further postulate that ILK might participate in EMT of tongue cancer. We showed that ILK siRNA inhibited EMT with low N-cadherin, Vimentin, Snail, Slug and Twist as well as high E-cadherin expression in vivo and in vitro. We found that knockdown of ILK inhibited cell proliferation, migration and invasion as well as changed cell morphology. We also demonstrated that ILK siRNA inhibited phosphorylation of downstream signaling targets Akt and GSK3β as well as reduced expression of MMP2 and MMP9. Furthermore, we found that the tongue tumor with high metastasis capability showed higher ILK, Vimentin, Snail, Slug and Twist as well as lower E-cadherin expression in clinical specimens. Finally, ILK siRNA led to the suppression for tumorigenesis and metastasis in vivo. Our findings suggest that ILK could be a novel diagnostic and therapeutic target for tongue cancer. Highlights: • ILK siRNA influences cell morphology, cell cycle, migration and invasion. • ILK siRNA affects the expression of proteins associated with EMT. • ILK expression is related to EMT in clinical human tongue tumors. • ILK siRNA inhibits metastasis of the tongue cancer cells through suppressing EMT.

  7. α-Solanine Inhibits Invasion of Human Prostate Cancer Cell by Suppressing Epithelial-Mesenchymal Transition and MMPs Expression

    Directory of Open Access Journals (Sweden)

    Kun-Hung Shen

    2014-08-01

    Full Text Available α-Solanine, a naturally occurring steroidal glycoalkaloid found in nightshade (Solanum nigrum Linn., was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in suppression of cancer cell metastasis by α-solanine remains unclear. This study investigates the suppression mechanism of α-solanine on motility of the human prostate cancer cell PC-3. Results show that α-solanine reduces the viability of PC-3 cells. When treated with non-toxic doses of α-solanine, cell invasion is markedly suppressed by α-solanine. α-Solanine also significantly elevates epithelial marker E-cadherin expression, while it concomitantly decreases mesenchymal marker vimentin expression, suggesting it suppresses epithelial-mesenchymal transition (EMT. α-Solanine reduces the mRNA level of matrix metalloproteinase-2 (MMP-2, MMP-9 and extracellular inducer of matrix metalloproteinase (EMMPRIN, but increases the expression of reversion-inducing cysteine-rich protein with kazal motifs (RECK, and tissue inhibitor of metalloproteinase-1 (TIMP-1 and TIMP-2. Immunoblotting assays indicate α-solanine is effective in suppressing the phosphorylation of phosphatidylinositide-3 kinase (PI3K, Akt and ERK. Moreover, α-solanine downregulates oncogenic microRNA-21 (miR-21 and upregulates tumor suppressor miR-138 expression. Taken together, the results suggest that inhibition of PC-3 cell invasion by α-solanine may be, at least in part, through blocking EMT and MMPs expression. α-Solanine also reduces ERK and PI3K/Akt signaling pathways and regulates expression of miR-21 and miR-138. These findings suggest an attractive therapeutic potential of α-solanine for suppressing invasion of prostate cancer cell.

  8. System-wide analysis of the transcriptional network of human myelomonocytic leukemia cells predicts attractor structure and phorbol-ester-induced differentiation and dedifferentiation transitions

    Science.gov (United States)

    Sakata, Katsumi; Ohyanagi, Hajime; Sato, Shinji; Nobori, Hiroya; Hayashi, Akiko; Ishii, Hideshi; Daub, Carsten O.; Kawai, Jun; Suzuki, Harukazu; Saito, Toshiyuki

    2015-02-01

    We present a system-wide transcriptional network structure that controls cell types in the context of expression pattern transitions that correspond to cell type transitions. Co-expression based analyses uncovered a system-wide, ladder-like transcription factor cluster structure composed of nearly 1,600 transcription factors in a human transcriptional network. Computer simulations based on a transcriptional regulatory model deduced from the system-wide, ladder-like transcription factor cluster structure reproduced expression pattern transitions when human THP-1 myelomonocytic leukaemia cells cease proliferation and differentiate under phorbol myristate acetate stimulation. The behaviour of MYC, a reprogramming Yamanaka factor that was suggested to be essential for induced pluripotent stem cells during dedifferentiation, could be interpreted based on the transcriptional regulation predicted by the system-wide, ladder-like transcription factor cluster structure. This study introduces a novel system-wide structure to transcriptional networks that provides new insights into network topology.

  9. Expression of Peroxisome Proferator-Activated Receptor γ (PPARγ) in Human Transitional Bladder Cancer and its Role in Inducing Cell Death

    OpenAIRE

    1999-01-01

    The present study examined the expression and role of the thiazolidinedione (TZD)-activated transcription factor, peroxisome proliferator-activated receptor γ (PPARγ), in human bladder cancers. In situ hybridization shows that PPARγ mRNA is highly expressed in all human transitional epithelial cell cancers (TCCa's) studied (n=11). PPARγ was also expressed in five TCCa cell lines as determined by RNase protection assays and immunoblot. Retinoid X receptor α (RXRα), a 9-cis-retinoic acid stimul...

  10. Expression of Peroxisome Proliferator-Activated Receptor γ (PPARγ) in Human Transitional Bladder Cancer and its Role in Inducing Cell Death1

    OpenAIRE

    1999-01-01

    The present study examined the expression and role of the thiazolidinedione (TZD)-activated transcription factor, peroxisome proliferator-activated receptor γ (PPARγ), in human bladder cancers. In situ hybridization shows that PPARγ mRNA is highly expressed in all human transitional epithelial cell cancers (TCCa's) studied (n=11). PPARγ was also expressed in five TCCa cell lines as determined by RNase protection assays and immunoblot. Retinoid X receptor α (RXRα), a 9-cis-retinoic acid stimul...

  11. Major transitions in human evolution

    Science.gov (United States)

    Foley, Robert A.; Martin, Lawrence; Mirazón Lahr, Marta; Stringer, Chris

    2016-01-01

    Evolutionary problems are often considered in terms of ‘origins', and research in human evolution seen as a search for human origins. However, evolution, including human evolution, is a process of transitions from one state to another, and so questions are best put in terms of understanding the nature of those transitions. This paper discusses how the contributions to the themed issue ‘Major transitions in human evolution’ throw light on the pattern of change in hominin evolution. Four questions are addressed: (1) Is there a major divide between early (australopithecine) and later (Homo) evolution? (2) Does the pattern of change fit a model of short transformations, or gradual evolution? (3) Why is the role of Africa so prominent? (4) How are different aspects of adaptation—genes, phenotypes and behaviour—integrated across the transitions? The importance of developing technologies and approaches and the enduring role of fieldwork are emphasized. This article is part of the themed issue ‘Major transitions in human evolution’. PMID:27298461

  12. Major transitions in human evolution.

    Science.gov (United States)

    Foley, Robert A; Martin, Lawrence; Mirazón Lahr, Marta; Stringer, Chris

    2016-07-05

    Evolutionary problems are often considered in terms of 'origins', and research in human evolution seen as a search for human origins. However, evolution, including human evolution, is a process of transitions from one state to another, and so questions are best put in terms of understanding the nature of those transitions. This paper discusses how the contributions to the themed issue 'Major transitions in human evolution' throw light on the pattern of change in hominin evolution. Four questions are addressed: (1) Is there a major divide between early (australopithecine) and later (Homo) evolution? (2) Does the pattern of change fit a model of short transformations, or gradual evolution? (3) Why is the role of Africa so prominent? (4) How are different aspects of adaptation-genes, phenotypes and behaviour-integrated across the transitions? The importance of developing technologies and approaches and the enduring role of fieldwork are emphasized.This article is part of the themed issue 'Major transitions in human evolution'. © 2016 The Author(s).

  13. Epithelial-mesenchymal transition stimulates human cancer cells to extend microtubule-based invasive protrusions and suppresses cell growth in collagen gel.

    Directory of Open Access Journals (Sweden)

    Jun Oyanagi

    Full Text Available Epithelial-mesenchymal transition (EMT is a crucial event in tumor invasion and metastasis. However, most of past EMT studies have been conducted in the conventional two-dimensional (2D monolayer culture. Therefore, it remains unclear what invasive phenotypes are acquired by EMT-induced cancer cells. To address this point, we attempted to characterize EMT cells in more physiological, three-dimensional (3D collagen gel culture. EMT was induced by treating three human carcinoma cell lines (A549, Panc-1 and MKN-1 with TGF-ß. The TGF-ß treatment stimulated these cells to overexpress the invasion markers laminin γ2 and MT1-MMP in 2D culture, in addition to the induction of well-known morphological change and EMT marker expression. EMT induction enhanced cell motility and adhesiveness to fibronectin and collagen in 2D culture. Although EMT cells showed comparable cell growth to control cells in 2D culture, their growth rates were extremely suppressed in soft agar and collagen gel cultures. Most characteristically, EMT-induced cancer cells commonly and markedly extended invasive protrusions in collagen gel. These protrusions were mainly supported by microtubules rather than actin cytoskeleton. Snail-introduced, stable EMT cells showed similar protrusions in 3D conditions without TGF-ß. Moreover, these protrusions were suppressed by colchicine or inhibitors of heat shock protein 90 (HSP-90 and protein phosphatase 2A. However, MMP inhibitors did not suppress the protrusion formation. These data suggest that EMT enhances tumor cell infiltration into interstitial stroma by extending microtubule-based protrusions and suppressing cell growth. The elevated cell adhesion to fibronectin and collagen and high cell motility also seem important for the tumor invasion.

  14. TGF-β1 induced epithelial to mesenchymal transition (EMT in human bronchial epithelial cells is enhanced by IL-1β but not abrogated by corticosteroids

    Directory of Open Access Journals (Sweden)

    Zuraw Bruce L

    2009-10-01

    Full Text Available Abstract Background Chronic persistent asthma is characterized by ongoing airway inflammation and airway remodeling. The processes leading to airway remodeling are poorly understood, and there is increasing evidence that even aggressive anti-inflammatory therapy does not completely prevent this process. We sought to investigate whether TGFβ1 stimulates bronchial epithelial cells to undergo transition to a mesenchymal phenotype, and whether this transition can be abrogated by corticosteroid treatment or enhanced by the pro-inflammatory cytokine IL-1β. Methods BEAS-2B and primary normal human bronchial epithelial cells were stimulated with TGFβ1 and expression of epithelial and mesenchymal markers assessed by quantitative real-time PCR, immunoblotting, immunofluorescence microscopy and zymography. In some cases the epithelial cells were also incubated with corticosteroids or IL-1β. Results were analyzed using non-parametric statistical tests. Results Treatment of BEAS-2B or primary human bronchial epithelial cells with TGFβ1 significantly reduced the expression level of the epithelial adherence junction protein E-cadherin. TGFβ1 then markedly induced mesenchymal marker proteins such as collagen I, tenascin C, fibronectin and α-smooth muscle actin mRNA in a dose dependant manner. The process of mesenchymal transition was accompanied by a morphological change towards a more spindle shaped fibroblast cell type with a more motile and invasive phenotype. Corticosteroid pre-treatment did not significantly alter the TGFβ1 induced transition but IL-1β enhanced the transition. Conclusion Our results indicate, that TGFβ1 can induce mesenchymal transition in the bronchial epithelial cell line and primary cells. Since asthma has been strongly associated with increased expression of TGFβ1 in the airway, epithelial to mesenchymal transition may contribute to the contractile and fibrotic remodeling process that accompanies chronic asthma.

  15. 6-OH-BDE-47 promotes human lung cancer cells epithelial mesenchymal transition via the AKT/Snail signal pathway.

    Science.gov (United States)

    Qu, Bao-Lin; Yu, Wei; Huang, Yu-Rong; Cai, Bo-Ning; Du, Le-Hui; Liu, Fang

    2015-01-01

    Hydroxylated polybrominated diphenyl ethers (OH-PBDEs) have been detected in the various human tissues. The OH-PBDEs are suggested to be stronger endocrine-disrupting compounds than PBDEs, therefore the toxicological effects of OH-PBDEs had received lots of attention. However, there is no study about the carcinogenic effect of OH-PBDEs and their estrogen potencies on the tumorigenesis and development of cancer. In the present study, we found that 6-hydroxy-2,2',4',4'-tetrabromodiphenyl ether (6-OH-BDE-47), the most abundant OH-PBDE congeners in human serum, promoted the in vitro migration of lung cancer A549 and H358 cells by induction of epithelial to mesenchymal transition (EMT). This was confirmed by that 6-OH-BDE-47 significantly down regulated the expression of epithelial markers E-cadherin (E-Cad) and zona occludin-1 (ZO-1) while up regulated the mesenchymal markers vimentin (Vim) and N-cadherin (N-Cad). 6-OH-BDE-47 up regulated the protein while not mRNA levels of Snail, which was the key transcription factor of EMT. Silencing of Snail by use of siRNA attenuated the 6-OH-BDE-47 induced EMT. This suggested that the stabilization of Snail was essential for 6-OH-BDE-47 induced EMT. Further, the treatment of 6-OH-BDE-47 increased the phosphorylation of AKT and ERK in A549 cells. Only PI3K/AKT inhibitor (LY294002), but not ERK inhibitor (PD98059), completely blocked the 6-OH-BDE-47 induced up regulation of Snail and down regulation of E-Cad, suggesting that PI3K/AKT pathway is important for 6-OH-BDE-47-mediated Snail stabilization and EMT in A549 cells. Generally, our results revealed for the first time that 6-OH-BDE-47 promoted the EMT of lung cancer cells via AKT/Snail signals. This suggested that more attention should be paid to the effects of OH-PBDEs on tumorigenesis and development of lung cancer. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. Cloning of Human Uroplakin Ⅱ Gene from Chinese Transitional Cell Carcinoma of Bladder and Construction of Its Eukaryotic Expression Vector

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    To clone Uroplakin Ⅱ gene from Chinese transitional cell carcinoma (TCC) of bladder and construct its eukaryotic expression vector, the molecular cloning method was used to extract total RNA from a GⅢ/ T3N0M0 tissue sample of the bladder TCC patients. The primers were designed by Primer 5.0 software. Full length cDNA of Uroplakin Ⅱ gene was amplified by reverse transcription polymerase chain reaction (RT-PCR), assayed by nucleic acid sequencing and then inserted between Xba Ⅰ and HindⅢ restrictive sites of eukaryotic expression vector pcDNA3.0. The recombinant was assayed by restricted enzyme digestion. Under the induction of Lipofectamine 2000, the recombinant was transfected into Uroplakin Ⅱ negative bladder cancer cell line EJ. Cellular expression levels of Uroplakin Ⅱ were detected by RT-PCR. The nucleic acid sequencing results indicated that Chinese Uroplakin Ⅱ cDNA (555 bp) was successfully cloned. The BLAST analysis demonstrated that the cloned sequence is 100 % homologous with sequences reported overseas. The GenBank accession number AY455312 was also registered. The results of restricted enzyme digestion indicated that eukaryotic vector pcDNA-UP Ⅱ for Uroplakin Ⅱ was successfully constructed.After being transferred with pcDNA-UPⅡ for 72 h, cellular Uroplakin Ⅱ mRNA levels were significantly improved (P<0.01). It is concluded that human Uroplakin Ⅱ gene was successfully cloned from Chinese TCC tissues, which provided a basis for further exploration of the roles of Uroplakin Ⅱ gene in TCC biological behaviors and potential strategies for targeted biological therapy of TCC.

  17. α-Solanine Inhibits Invasion of Human Prostate Cancer Cell by Suppressing Epithelial-Mesenchymal Transition and MMPs Expression

    OpenAIRE

    Kun-Hung Shen; Alex Chien-Hwa Liao; Jui-Hsiang Hung; Wei-Jiunn Lee; Kai-Chieh Hu; Pin-Tsen Lin; Ruei-Fang Liao; Pin-Shern Chen

    2014-01-01

    α-Solanine, a naturally occurring steroidal glycoalkaloid found in nightshade (Solanum nigrum Linn.), was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in suppression of cancer cell metastasis by α-solanine remains unclear. This study investigates the suppression mechanism of α-solanine on motility of the human prostate cancer cell PC-3. Results show that α-solanine reduces the viability of PC-3 cells. When treated with non-toxic doses of ...

  18. Plumbagin elicits differential proteomic responses mainly involving cell cycle, apoptosis, autophagy, and epithelial-to-mesenchymal transition pathways in human prostate cancer PC-3 and DU145 cells

    Directory of Open Access Journals (Sweden)

    Qui JX

    2015-01-01

    Full Text Available Jia-Xuan Qiu,1,2 Zhi-Wei Zhou, 3,4 Zhi-Xu He,4 Ruan Jin Zhao,5 Xueji Zhang,6 Lun Yang,7 Shu-Feng Zhou,3,4 Zong-Fu Mao11School of Public Health, Wuhan University, Wuhan, Hubei, People’s Republic of China; 2Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, People’s Republic of China; 3Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA; 4Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, Guizhou, People’s Republic of China; 5Center for Traditional Chinese Medicine, Sarasota, FL, USA; 6Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 7Bio-X Institutes, Key Laboratory for the Genetics of Development and Neuropsychiatric Disorders (Ministry of Education, Shanghai Jiao Tong University, Shanghai, People’s Republic of ChinaAbstract: Plumbagin (PLB has exhibited a potent anticancer effect in preclinical studies, but the molecular interactome remains elusive. This study aimed to compare the quantitative proteomic responses to PLB treatment in human prostate cancer PC-3 and DU145 cells using the approach of stable-isotope labeling by amino acids in cell culture (SILAC. The data were finally validated using Western blot assay. First, the bioinformatic analysis predicted that PLB could interact with 78 proteins that were involved in cell proliferation and apoptosis, immunity, and signal transduction. Our quantitative proteomic study using SILAC revealed that there were at least 1,225 and 267 proteins interacting with PLB and there were 341 and 107 signaling pathways and cellular functions potentially regulated by PLB in PC-3 and DU145 cells, respectively. These proteins and pathways played a

  19. Effect of proton and gamma irradiation on human lung carcinoma cells: Gene expression, cell cycle, cell death, epithelial–mesenchymal transition and cancer-stem cell trait as biological end points

    Energy Technology Data Exchange (ETDEWEB)

    Narang, Himanshi, E-mail: narangh@barc.gov.in [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai 400085 (India); Kumar, Amit [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai 400085 (India); Bhat, Nagesh [Radiological Physics and Advisory Division, Bhabha Atomic Research Centre, Mumbai 400085 (India); Pandey, Badri N.; Ghosh, Anu [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai 400085 (India)

    2015-10-15

    Highlights: • Biological effectiveness of proton and gamma irradiation is compared in A549 cells. • Proton irradiation is two times more cytotoxic than gamma irradiation. • It alters ten times more number of early genes, as observed by microarray study. • It does not enhance cell migration, invasion and adhesion, unlike gamma irradiation. • It was more effective in reducing the percentage of cancer stem cell like cells. - Abstract: Proton beam therapy is a cutting edge modality over conventional gamma radiotherapy because of its physical dose deposition advantage. However, not much is known about its biological effects vis-a-vis gamma irradiation. Here we investigated the effect of proton- and gamma- irradiation on cell cycle, death, epithelial-mesenchymal transition (EMT) and “stemness” in human non-small cell lung carcinoma cells (A549). Proton beam (3 MeV) was two times more cytotoxic than gamma radiation and induced higher and longer cell cycle arrest. At equivalent doses, numbers of genes responsive to proton irradiation were ten times higher than those responsive to gamma irradiation. At equitoxic doses, the proton-irradiated cells had reduced cell adhesion and migration ability as compared to the gamma-irradiated cells. It was also more effective in reducing population of Cancer Stem Cell (CSC) like cells as revealed by aldehyde dehydrogenase activity and surface phenotyping by CD44{sup +}, a CSC marker. These results can have significant implications for proton therapy in the context of suppression of molecular and cellular processes that are fundamental to tumor expansion.

  20. The bornavirus-derived human protein EBLN1 promotes efficient cell cycle transit, microtubule organisation and genome stability

    Science.gov (United States)

    Myers, Katie N.; Barone, Giancarlo; Ganesh, Anil; Staples, Christopher J.; Howard, Anna E.; Beveridge, Ryan D.; Maslen, Sarah; Skehel, J. Mark; Collis, Spencer J.

    2016-01-01

    It was recently discovered that vertebrate genomes contain multiple endogenised nucleotide sequences derived from the non-retroviral RNA bornavirus. Strikingly, some of these elements have been evolutionary maintained as open reading frames in host genomes for over 40 million years, suggesting that some endogenised bornavirus-derived elements (EBL) might encode functional proteins. EBLN1 is one such element established through endogenisation of the bornavirus N gene (BDV N). Here, we functionally characterise human EBLN1 as a novel regulator of genome stability. Cells depleted of human EBLN1 accumulate DNA damage both under non-stressed conditions and following exogenously induced DNA damage. EBLN1-depleted cells also exhibit cell cycle abnormalities and defects in microtubule organisation as well as premature centrosome splitting, which we attribute in part, to improper localisation of the nuclear envelope protein TPR. Our data therefore reveal that human EBLN1 possesses important cellular functions within human cells, and suggest that other EBLs present within vertebrate genomes may also possess important cellular functions. PMID:27739501

  1. Tacrolimus Modulates TGF-β Signaling to Induce Epithelial-Mesenchymal Transition in Human Renal Proximal Tubule Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Jason Bennett

    2016-04-01

    Full Text Available Epithelial-mesenchymal transition (EMT, a process which describes the trans-differentiation of epithelial cells into motile mesenchymal cells, is pivotal in stem cell behavior, development and wound healing, as well as contributing to disease processes including fibrosis and cancer progression. Maintenance immunosuppression with calcineurin inhibitors (CNIs has become routine management for renal transplant patient, but unfortunately the nephrotoxicity of these drugs has been well documented. HK-2 cells were exposed to Tacrolimus (FK506 and EMT markers were assessed by RT PCR and western blot. FK506 effects on TGF-β mRNA were assessed by RT PCR and TGF-β secretion was measured by ELISA. The impact of increased TGF-β secretion on Smad signaling pathways was investigated. The impact of inhibition of TGF-β signaling on EMT processes was assessed by scratch-wound assay. The results presented in this study suggest that FK506 initiates EMT processes in the HK-2 cell line, with altered expression of epithelial and myofibroblast markers evident. Additionally, the study demonstrates that FK506 activation of the TGF-β/ SMAD pathways is an essential step in the EMT process. Overall the results demonstrate that EMT is heavily involved in renal fibrosis associated with CNI nephrotoxicity.

  2. Downregulation of tight junction-associated MARVEL protein marvelD3 during epithelial-mesenchymal transition in human pancreatic cancer cells.

    Science.gov (United States)

    Kojima, Takashi; Takasawa, Akira; Kyuno, Daisuke; Ito, Tatsuya; Yamaguchi, Hiroshi; Hirata, Koichi; Tsujiwaki, Mitsuhiro; Murata, Masaki; Tanaka, Satoshi; Sawada, Norimasa

    2011-10-01

    The novel tight junction protein marvelD3 contains a conserved MARVEL (MAL and related proteins for vesicle trafficking and membrane link) domain like occludin and tricellulin. However, little is yet known about the detailed role and regulation of marvelD3 in normal epithelial cells and cancer cells, including pancreatic cancer. In the present study, we investigated marvelD3 expression in well and poorly differentiated human pancreatic cancer cell lines and normal pancreatic duct epithelial cells in which the hTERT gene was introduced into human pancreatic duct epithelial cells in primary culture, and the changes of marvelD3 during Snail-induced epithelial-mesenchymal transition (EMT) under hypoxia, TGF-β treatment and knockdown of FOXA2 in well differentiated pancreatic cancer HPAC cells. MarvelD3 was transcriptionally downregulated in poorly differentiated pancreatic cancer cells and during Snail-induced EMT of pancreatic cancer cells in which Snail was highly expressed and the fence function downregulated, whereas it was maintained in well differentiated human pancreatic cancer cells and normal pancreatic duct epithelial cells. Depletion of marvelD3 by siRNAs in HPAC cells resulted in downregulation of barrier functions indicated as a decrease in transepithelial electric resistance and an increase of permeability to fluorescent dextran tracers, whereas it did not affect fence function of tight junctions. In conclusion, marvelD3 is transcriptionally downregulated in Snail-induced EMT during the progression for the pancreatic cancer. Copyright © 2011 Elsevier Inc. All rights reserved.

  3. Blocking TGF-β expression inhibits silica particle-induced epithelial-mesenchymal transition in human lung epithelial cells.

    Science.gov (United States)

    Rong, Yi; Shen, Yan; Zhang, Zhihong; Cui, Xiuqing; Xiao, Lili; Liu, Yuewei; Luo, Xin; Chen, Weihong

    2015-11-01

    The main characteristic of silicosis is irreversible fibrosis. Certain studies have shown that epithelial-mesenchymal transition (EMT) regulated by transforming growth factor-β (TGF-β) is involved in fibrosis. Thus, we suggest that TGF-β regulated EMT may play an important role in silicosis. In this study, we determined the expression of TGF-β-Smad2/3, EMT- and ECM-related markers in lung epithelial cells treated with silica particle by RT-PCR, western-blot and ELISA. In order to explore the role of TGF-β, we used TGF-β inhibitor in the cell model. We found that the cells lost the expression of epithelial phenotypic markers and acquired increased expression of mesenchymal cells markers with ECM deposition after treatment with silica particle. Moreover, the changes of EMT-related event was restricted in response to TGF-β inhibitor. These findings suggest that EMT is essentially involved in the pathogenesis of fibrosis induced by silica particles and down-regulating the TGF-β expression can inhibit the process of EMT.

  4. Berberine reverses epithelial-to-mesenchymal transition and inhibits metastasis and tumor-induced angiogenesis in human cervical cancer cells.

    Science.gov (United States)

    Chu, Shu-Chen; Yu, Cheng-Chia; Hsu, Li-Sung; Chen, Kuo-Shuen; Su, Mei-Yu; Chen, Pei-Ni

    2014-12-01

    Metastasis is the most common cause of cancer-related death in patients, and epithelial-to-mesenchymal transition (EMT) is essential for cancer metastasis, which is a multistep complicated process that includes local invasion, intravasation, extravasation, and proliferation at distant sites. When cancer cells metastasize, angiogenesis is also required for metastatic dissemination, given that an increase in vascular density will allow easier access of tumor cells to circulation, and represents a rational target for therapeutic intervention. Berberine has several anti-inflammation and anticancer biologic effects. In this study, we provided molecular evidence that is associated with the antimetastatic effect of berberine by showing a nearly complete inhibition on invasion (P metalloproteinase-2 and urokinase-type plasminogen activator. Berberine reversed transforming growth factor-β1-induced EMT and caused upregulation of epithelial markers such as E-cadherin and inhibited mesenchymal markers such as N-cadherin and snail-1. Selective snail-1 inhibition by snail-1-specific small interfering RNA also showed increased E-cadherin expression in SiHa cells. Berberine also reduced tumor-induced angiogenesis in vitro and in vivo. Importantly, an in vivo BALB/c nude mice xenograft model and tail vein injection model showed that berberine treatment reduced tumor growth and lung metastasis by oral gavage, respectively. Taken together, these findings suggested that berberine could reduce metastasis and angiogenesis of cervical cancer cells, thereby constituting an adjuvant treatment of metastasis control.

  5. Human papillomavirus oncoproteins differentially modulate epithelial-mesenchymal transition in 5-FU-resistant cervical cancer cells.

    Science.gov (United States)

    Vishnoi, Kanchan; Mahata, Sutapa; Tyagi, Abhishek; Pandey, Arvind; Verma, Gaurav; Jadli, Mohit; Singh, Tejveer; Singh, Sukh Mahendra; Bharti, Alok C

    2016-10-01

    Etiological role of viral proteins E6 and E7 of high-risk HPV in cervical carcinogenesis is well established. However, their contribution in chemoresistance and epithelial-mesenchymal transition (EMT) that leads to advanced metastatic lesions and chemoresistance is poorly defined. In the present study, contribution of viral oncoproteins in acquisition of EMT character during onset of chemoresistance was assessed. A chemoresistant cell line (SiHaCR) was developed from an established HPV16-positive cervical cancer cell line, SiHa, by escalating selection pressure of 5-fluorouracil (5-FU). Expression of Survivin, ABCG2, Snail, Slug, Twist, and Vimentin was examined in SiHa and SiHaCR cells by reverse transcriptase-PCR (RT-PCR) and immunoblotting assays. Mesenchymal phenotype in SiHaCR cells was confirmed by assessment of migration and invasion potentials. SiHaCR cells displayed elevated level of functional and molecular markers associated with chemoresistance (Survivin, ABCG2) and EMT (Snail, Slug, Twist, Vimentin) and reduced E-cadherin. SiHaCR also showed increased levels of HPV16 E6 and E7 transcripts. Specific silencing of HPV16 E6, but not E7 using corresponding siRNA, demonstrated a differential involvement of HPV oncogenes in manifestation of EMT. HPV16 E6 silencing resulted in reduction of Slug and Twist expression. However, the expression of Snail and Vimentin was only marginally affected. In contrast, there was an increase in the expression of E-cadherin. A reduced migration and invasion capabilities were observed only in E6-silenced SiHaCR cells, which further confirmed functional contribution of HPV16 E6 in manifestation of EMT. Taken together, our study demonstrated an active involvement of HPV16 E6 in regulation of EMT, which promotes chemoresistance in cervical cancer.

  6. Plumbagin induces cell cycle arrest and autophagy and suppresses epithelial to mesenchymal transition involving PI3K/Akt/mTOR-mediated pathway in human pancreatic cancer cells

    Directory of Open Access Journals (Sweden)

    Wang F

    2015-01-01

    PLB and investigate the underlying mechanism in human pancreatic cancer PANC-1 and BxPC-3 cells. The results showed that PLB exhibited potent inducing effects on cell cycle arrest in PANC-1 and BxPC-3 cells via the modulation of cell cycle regulators including CDK1/CDC2, cyclin B1, cyclin D1, p21 Waf1/Cip1, p27 Kip1, and p53. PLB treatment concentration- and time-dependently increased the percentage of autophagic cells and significantly increased the expression level of phosphatase and tensin homolog, beclin 1, and the ratio of LC3-II over LC3-I in both PANC-1 and BxPC-3 cells. PLB induced inhibition of phosphatidylinositol 3-kinase (PI3K/protein kinase B/mammalian target of rapamycin and p38 mitogen-activated protein kinase (p38 MAPK pathways and activation of 5'-AMP-dependent kinase as indicated by their altered phosphorylation, contributing to the proautophagic activities of PLB in both cell lines. Furthermore, SB202190, a selective inhibitor of p38 MAPK, and wortmannin, a potent, irreversible, and selective PI3K inhibitor, remarkably enhanced PLB-induced autophagy in PANC-1 and BxPC-3 cells, indicating the roles of PI3K and p38 MAPK mediated signaling pathways in PLB-induced autophagic cell death in both cell lines. In addition, PLB significantly inhibited epithelial to mesenchymal transition phenotype in both cell lines with an increase in the expression level of E-cadherin and a decrease in N-cadherin. Moreover, PLB treatment significantly suppressed the expression of Sirt1 in both cell lines. These findings show that PLB promotes cell cycle arrest and autophagy but inhibits epithelial to mesenchymal transition phenotype in pancreatic cancer cells with the involvement of PI3K/protein kinase B/ mammalian target of rapamycin and p38 MAPK mediated pathways. Keywords: Plumbagin, pancreatic cancer, cell cycle, autophagy, EMT, Sirt1

  7. Fisetin inhibits human melanoma cell invasion through promotion of mesenchymal to epithelial transition and by targeting MAPK and NFκB signaling pathways.

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    Harish Chandra Pal

    Full Text Available Malignant melanoma is responsible for approximately 75% of skin cancer-related deaths. BRAF plays an important role in regulating the mitogen-activated protein kinase (MAPK signaling cascade in melanoma with activating mutations in the serine/threonine kinase BRAF occurring in 60-70% of malignant melanomas. The BRAF-MEK-ERK (MAPK pathway is a key regulator of melanoma cell invasion. In addition, activation of NFκB via the MAPK pathway is regulated through MEK-induced activation of IKK. These pathways are potential targets for prevention and treatment of melanoma. In this study, we investigated the effect of fisetin, a phytochemical present in fruits and vegetables, on melanoma cell invasion and epithelial-mesenchymal transition, and delineated the underlying molecular mechanism. Treatment of multiple human malignant melanoma cell lines with fisetin (5-20 µM resulted in inhibition of cell invasion. BRAF mutated melanoma cells were more sensitive to fisetin treatment, and this was associated with a decrease in the phosphorylation of MEK1/2 and ERK1/2. In addition, fisetin inhibited the activation of IKK leading to a reduction in the activation of the NFκB signaling pathway. Treatment of cells with an inhibitor of MEK1/2 (PD98059 or of NFκB (caffeic acid phenethyl ester also reduced melanoma cell invasion. Furthermore, treatment of fisetin promoted mesenchymal to epithelial transition in melanoma cells, which was associated with a decrease in mesenchymal markers (N-cadherin, vimentin, snail and fibronectin and an increase in epithelial markers (E-cadherin and desmoglein. Employing three dimensional skin equivalents consisting of A375 cells admixed with normal human keratinocytes embedded onto a collagen-constricted fibroblast matrix, we found that treatment of fisetin reduced the invasive potential of melanoma cells into the dermis and increased the expression of E-cadherin with a concomitant decrease in vimentin. These results indicate that

  8. Fisetin inhibits human melanoma cell invasion through promotion of mesenchymal to epithelial transition and by targeting MAPK and NFκB signaling pathways.

    Science.gov (United States)

    Pal, Harish Chandra; Sharma, Samriti; Strickland, Leah Ray; Katiyar, Santosh K; Ballestas, Mary E; Athar, Mohammad; Elmets, Craig A; Afaq, Farrukh

    2014-01-01

    Malignant melanoma is responsible for approximately 75% of skin cancer-related deaths. BRAF plays an important role in regulating the mitogen-activated protein kinase (MAPK) signaling cascade in melanoma with activating mutations in the serine/threonine kinase BRAF occurring in 60-70% of malignant melanomas. The BRAF-MEK-ERK (MAPK) pathway is a key regulator of melanoma cell invasion. In addition, activation of NFκB via the MAPK pathway is regulated through MEK-induced activation of IKK. These pathways are potential targets for prevention and treatment of melanoma. In this study, we investigated the effect of fisetin, a phytochemical present in fruits and vegetables, on melanoma cell invasion and epithelial-mesenchymal transition, and delineated the underlying molecular mechanism. Treatment of multiple human malignant melanoma cell lines with fisetin (5-20 µM) resulted in inhibition of cell invasion. BRAF mutated melanoma cells were more sensitive to fisetin treatment, and this was associated with a decrease in the phosphorylation of MEK1/2 and ERK1/2. In addition, fisetin inhibited the activation of IKK leading to a reduction in the activation of the NFκB signaling pathway. Treatment of cells with an inhibitor of MEK1/2 (PD98059) or of NFκB (caffeic acid phenethyl ester) also reduced melanoma cell invasion. Furthermore, treatment of fisetin promoted mesenchymal to epithelial transition in melanoma cells, which was associated with a decrease in mesenchymal markers (N-cadherin, vimentin, snail and fibronectin) and an increase in epithelial markers (E-cadherin and desmoglein). Employing three dimensional skin equivalents consisting of A375 cells admixed with normal human keratinocytes embedded onto a collagen-constricted fibroblast matrix, we found that treatment of fisetin reduced the invasive potential of melanoma cells into the dermis and increased the expression of E-cadherin with a concomitant decrease in vimentin. These results indicate that fisetin

  9. RNA Interference Targeting Snail Inhibits the Transforming Growth Factor β 2-Induced Epithelial-Mesenchymal Transition in Human Lens Epithelial Cells.

    Science.gov (United States)

    Li, Ping; Jing, Jiaona; Hu, Jianyan; Li, Tiejun; Sun, Yuncheng; Guan, Huaijin

    2013-01-01

    Epithelial-msenchymal transition (EMT) contributes to posterior capsule opacification (PCO) type of cataract. Transcription factors Snail is a key trigger of EMT activated by transforming growth factor β (TGF β ). This study was done to investigate the effect of Snail targeting siRNA on TGF β 2-induced EMT in human lens epithelial cells. TGF β 2 treatment of cultured human epithelial cell line (HLEB3) upregulated the expression of Snail and the EMT relevant molecules such as vimentin and α -SMA but downregulated the expression of keratin and E-cadherin. After the stimulation of TGF β 2, the HLEB3 cells became fibroblast-like in morphology, and the junctions of cell-cell disappeared. TGF β 2 treatment also enhanced migration ability of HLEB3 cells. TGF β 2-induced Snail expression and EMT were significantly inhibited by Snail siRNA. By analyzing the response characteristics of HLEB3 in TGF β 2-induced EMT model with/without Snail-specific siRNA, we concluded that Snail is an element in the EMT of HLEB3 cells induced by TGF β 2. Snail siRNA targeting can block the induced EMT and therefore has the potential to suppress the development of PCO.

  10. RNA Interference Targeting Snail Inhibits the Transforming Growth Factor β2-Induced Epithelial-Mesenchymal Transition in Human Lens Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Ping Li

    2013-01-01

    Full Text Available Epithelial-msenchymal transition (EMT contributes to posterior capsule opacification (PCO type of cataract. Transcription factors Snail is a key trigger of EMT activated by transforming growth factor β (TGFβ. This study was done to investigate the effect of Snail targeting siRNA on TGFβ2-induced EMT in human lens epithelial cells. TGFβ2 treatment of cultured human epithelial cell line (HLEB3 upregulated the expression of Snail and the EMT relevant molecules such as vimentin and α-SMA but downregulated the expression of keratin and E-cadherin. After the stimulation of TGFβ2, the HLEB3 cells became fibroblast-like in morphology, and the junctions of cell-cell disappeared. TGFβ2 treatment also enhanced migration ability of HLEB3 cells. TGFβ2-induced Snail expression and EMT were significantly inhibited by Snail siRNA. By analyzing the response characteristics of HLEB3 in TGFβ2-induced EMT model with/without Snail-specific siRNA, we concluded that Snail is an element in the EMT of HLEB3 cells induced by TGFβ2. Snail siRNA targeting can block the induced EMT and therefore has the potential to suppress the development of PCO.

  11. Expression of Peroxisome Proferator-Activated Receptor γ (PPARγ in Human Transitional Bladder Cancer and its Role in Inducing Cell Death

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    You-Fei Guan

    1999-10-01

    Full Text Available The present study examined the expression and role of the thiazolidinedione (TZD-activated transcription factor, peroxisome proliferator-activated receptor γ (PPARγ, in human bladder cancers. In situ hybridization shows that PPARγ mRNA is highly expressed in all human transitional epithelial cell cancers (TCCa's studied (n=11. PPARγ was also expressed in five TCCa cell lines as determined by RNase protection assays and immunoblot. Retinoid X receptor α (RXRα, a 9-cis-retinoic acid stimulated (9-cis-RA heterodimeric partner of PPARγ, was also co-expressed in all TCCa tissues and cell lines. Treatment of the T24 bladder cancer cells with the TZD PPARγ agonist troglitazone, dramatically inhibited 3H-thymidine incorporation and induced cell death. Addition of the RXRα ligands, 9-cis-RA or LG100268, sensitized T24 bladder cancer cells to the lethal effect of troglitazone and two other PPARγ activators, ciglitazone and 15-deoxy-Δ12,14-PGJ2 (15dPGJ2. Troglitazone treatment increased expression of two cyclin-dependent kinase inhibitors, p21wAF1/CIP1 and p16INK4, reduced cyclin D1 expression, consistent with G1 arrest. Troglitazone also induced an endogenous PPARγ target gene in T24 cells, adipocyte-type fatty acid binding protein (A-FABP, the expression of which correlates with bladder cancer differentiation. In situ hybridization shows that A-FABP expression is localized to normal uroepithelial cells as well as some TCCa's. Taken together, these results demonstrate that PPARγ is expressed in human TCCa where it may play a role in regulating TCCa differentiation and survival, thereby providing a potential target for therapy of uroepithelial cancers.

  12. Expression of peroxisome proliferator-activated receptor gamma (PPARgamma) in human transitional bladder cancer and its role in inducing cell death.

    Science.gov (United States)

    Guan, Y F; Zhang, Y H; Breyer, R M; Davis, L; Breyer, M D

    1999-10-01

    The present study examined the expression and role of the thiazolidinedione (TZD)-activated transcription factor, peroxisome proliferator-activated receptor gamma (PPARgamma), in human bladder cancers. In situ hybridization shows that PPARgamma mRNA is highly expressed in all human transitional epithelial cell cancers (TCCa's) studied (n=11). PPARgamma was also expressed in five TCCa cell lines as determined by RNase protection assays and immunoblot. Retinoid X receptor alpha (RXRalpha), a 9-cis-retinoic acid stimulated (9-cis-RA) heterodimeric partner of PPARgamma, was also co-expressed in all TCCa tissues and cell lines. Treatment of the T24 bladder cancer cells with the TZD PPARgamma agonist troglitazone, dramatically inhibited 3H-thymidine incorporation and induced cell death. Addition of the RXRalpha ligands, 9-cis-RA or LG100268, sensitized T24 bladder cancer cells to the lethal effect of troglitazone and two other PPAR- activators, ciglitazone and 15-deoxy-delta(12,14)-PGJ2 (15dPGJ(2)). Troglitazone treatment increased expression of two cyclin-dependent kinase inhibitors, p21(WAF1/CIP1) and p16(INK4), and reduced cyclin D1 expression, consistent with G1 arrest. Troglitazone also induced an endogenous PPARgamma target gene in T24 cells, adipocyte-type fatty acid binding protein (A-FABP), the expression of which correlates with bladder cancer differentiation. In situ hybridization shows that A-FABP expression is localized to normal uroepithelial cells as well as some TCCa's. Taken together, these results demonstrate that PPARgamma is expressed in human TCCa where it may play a role in regulating TCCa differentiation and survival, thereby providing a potential target for therapy of uroepithelial cancers.

  13. Inhibition of pirfenidone on TGF-beta2 induced proliferation, migration and epithlial-mesenchymal transition of human lens epithelial cells line SRA01/04.

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    Yangfan Yang

    Full Text Available BACKGROUND: Posterior capsular opacification (PCO is a common complication of cataract surgery. Transforming growth factor-β2 (TGF-β2 plays important roles in the development of PCO. The existing pharmacological treatments are not satisfactory and can have toxic side effects. METHODOLOGIES/PRINCIPAL FINDINGS: We evaluated the effect of pirfenidone on proliferation, migration and epithlial-mesenchymal transition of human lens epithelial cell line SRA01/04 (HLECs in vitro. After treatment with 0, 0.25, and 0.5 mg/ml pirfenidone, cell proliferation was measured by MTT assay. Cell viability was determined by trypan blue exclusion assay and measurement of lactate dehydrogenase (LDH activity released from the damaged cells. And cell migration was measured by scratch assay in the absence or presence of transforming growth factor-β2 (TGF-β2. The expressions of TGF-β2 and SMADs were evaluated with real-time RT-PCR, western blot, and immunofluorescence analyses. The mesenchymal phenotypic marker fibronectin (FN was demonstrated by Immunocytofluorescence analyses. The cells had high viability, which did not vary across different concentrations of pirfenidone (0 [control] 0.3, 0.5 or 1.0 mg/ml after 24 hours. Pirfenidone (0∼0.5 mg/ml had no significant cytotoxicity effect on SRA01/04 by LDH assay. Pirfenidone significantly inhibited the proliferation and TGF-β2-induced cell migration and the effects were dose-dependent, and inhibited TGF-β2-induced fibroblastic phenotypes and TGF-β2-induced expression of FN in SRA01/04 cells. The cells showed dose-dependent decreases in mRNA and protein levels of TGF-β2 and SMADs. Pirfenidone also depressed the TGF-β2-induced expression of SMADs and blocked the nuclear translocation of SMADs in cells. CONCLUSION: Pirfenidone inhibits TGF-β2-induced proliferation, migration and epithlial-mesenchymal transition of human lens epithelial cells line SRA01/04 at nontoxic concentrations. This effect may be achieved by

  14. Phase transitions in tumor growth: IV relationship between metabolic rate and fractal dimension of human tumor cells

    Science.gov (United States)

    Betancourt-Mar, J. A.; Llanos-Pérez, J. A.; Cocho, G.; Mansilla, R.; Martin, R. R.; Montero, S.; Nieto-Villar, J. M.

    2017-05-01

    By the use of thermodynamics formalism of irreversible processes, complex systems theory and systems biology, it is derived a relationship between the production of entropy per unit time, the fractal dimension and the tumor growth rate for human tumors cells. The thermodynamics framework developed demonstrates that, the dissipation function is a Landau potential and also the Lyapunov function of the dynamical behavior of tumor growth, which indicate the directional character, stability and robustness of the phenomenon. The entropy production rate may be used as a quantitative index of the metastatic potential of tumors. The current theoretical framework will hopefully provide a better understanding of cancer and contribute to improvements in cancer treatment.

  15. Paraquat induces epithelial-mesenchymal transition-like cellular response resulting in fibrogenesis and the prevention of apoptosis in human pulmonary epithelial cells.

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    Atsushi Yamada

    Full Text Available The aim of this study is to investigate the molecular mechanisms underlying delayed progressive pulmonary fibrosis, a characteristic of subacute paraquat (PQ poisoning. Epithelial-mesenchymal transition (EMT has been proposed as a cause of organ fibrosis, and transforming growth factor-β (TGF-β is suggested to be a powerful mediator of EMT. We thus examined the possibility that EMT is involved in pulmonary fibrosis during PQ poisoning using A549 human alveolar epithelial cells in vitro. The cells were treated with various concentrations of PQ (0-500 μM for 2-12 days. Short-term (2 days high-dose (>100 μM treatments with PQ induced cell death accompanied by the activation of caspase9 as well as a decrease in E-cadherin (an epithelial cell marker, suggesting apoptotic cell death with the features of anoikis (cell death due to the loss of cell-cell adhesion. In contrast, long-term (6-12 days low-dose (30 μM treatments with PQ resulted in a transformation into spindle-shaped mesenchymal-like cells with a decrease of E-cadherin as well as an increase of α-smooth muscle actin (α-SMA. The mesenchymal-like cells also secreted the extracellular matrix (ECM protein fibronectin into the culture medium. The administration of a TGF-β1 receptor antagonist, SB431542, almost completely attenuated the mesenchymal transformation as well as fibronectin secretion, suggesting a crucial role of TGF-β1 in EMT-like cellular response and subsequent fibrogenesis. It is noteworthy that despite the suppression of EMT-fibrogenesis, apoptotic death was observed in cells treated with PQ+SB431542. EMT-like cellular response and subsequent fibrogenesis were also observed in normal human bronchial epithelial (NHBE cells exposed to PQ in a TGF-β1-dependent manner. Taken together, our experimental model reflects well the etiology of PQ poisoning in human and shows the involvement of EMT-like cellular response in both fibrogenesis and resistance to cell death during

  16. SNAIL induces epithelial-to-mesenchymal transition in a human pancreatic cancer cell line (BxPC3) and promotes distant metastasis and invasiveness in vivo.

    Science.gov (United States)

    Nishioka, Ryohei; Itoh, Shunji; Gui, Ting; Gai, Zhibo; Oikawa, Kosuke; Kawai, Manabu; Tani, Masaji; Yamaue, Hiroki; Muragaki, Yasuteru

    2010-10-01

    SNAIL, a potent repressor of E-cadherin expression, plays a key role in inducing epithelial-to-mesenchymal transition (EMT) in epithelial cells. During EMT, epithelial cells lose cell polarity and adhesion, and undergo drastic morphological changes acquiring highly migratory abilities. Although there is increasing evidence that EMT is involved in the progression of some human cancers, its significance in the progression of pancreatic cancer remains elusive. In Panc-1, a well-known human pancreatic cancer cell line in which EMT is triggered by TGF-β1 treatment, SNAIL and vimentin are highly expressed, whereas E-cadherin expression is scant. In contrast, another human pancreatic cancer cell line, BxPC3, in which SNAIL expression is not detected, has high levels of E-cadherin expression and does not undergo EMT upon TGF-β1 treatment. After transfecting the SNAIL gene into BxPC3, however, the cells undergo EMT with remarkable alterations in cell morphology and molecular expression patterns without the addition of any growth factors. Furthermore, in an orthotopic transplantation model using SCID mice, SNAIL-transfected BxPC3 displayed highly metastatic and invasive activities. In the immunohistochemical analysis of the tumor derived from the SNAIL-expressing BxPC3, alterations suggestive of EMT were observed in the invasive tumor front. SNAIL enabled BxPC3 to undergo EMT, endowing it with a highly malignant potential in vivo. These results indicate that SNAIL-mediated EMT may be relevant in the progression of pancreatic cancer, and SNAIL could be a molecular target for a pancreatic cancer intervention. Copyright © 2010 Elsevier Inc. All rights reserved.

  17. High-risk human papillomavirus infections and overexpression of p53 protein as prognostic indicators in transitional cell carcinoma of the urinary bladder.

    Science.gov (United States)

    Furihata, M; Inoue, K; Ohtsuki, Y; Hashimoto, H; Terao, N; Fujita, Y

    1993-10-15

    Ninety Japanese patients with transitional cell carcinoma of the urinary bladder were investigated for tumor incorporation of DNA for high-risk human papillomavirus (HPV) types 16, 18, and 33 by in situ hybridization with biotinylated DNA probes. In addition, immunohistochemical analysis of p53 protein expression was performed with an antibody to p53 protein. Twenty-eight tumors were positive for HPV DNA, and multiple HPV infection was detected in 17 cases. Positive nuclear staining of cancer cells by the antibody to p53 protein was detected in 32 cases. DNA for HPV 16, 18, and/or 33 and the overexpression of p53 protein were simultaneously observed in 6 tumors by using a mirror section method. The overexpression of p53 protein was frequently detected in invasive and nonpapillary tumors (P infection was more common in noninvasive and papillary tumors (P infection or overexpression of p53 protein may be related to tumor behavior and may indicate a relatively poor prognosis in patients with transitional cell carcinoma.

  18. Connecticut Transit (CTTRANSIT) Fuel Cell Transit Bus: Preliminary Evaluation Results

    Energy Technology Data Exchange (ETDEWEB)

    Chandler, K.; Eudy, L.

    2008-10-01

    This report provides preliminary results from a National Renewable Energy Laboratory evaluation of a protoptye fuel cell transit bus operating at Connecticut Transit in Hartford. Included are descriptions of the planned fuel cell bus demonstration and equipment; early results and agency experience are also provided.

  19. Cigarette smoke extracts induced the colon cancer migration via regulating epithelial mesenchymal transition and metastatic genes in human colon cancer cells.

    Science.gov (United States)

    Kim, Cho-Won; Go, Ryeo-Eun; Lee, Hae-Miru; Hwang, Kyung-A; Lee, Kyuhong; Kim, Bumseok; Lee, Moo-Yeol; Choi, Kyung-Chul

    2017-02-01

    There was considerable evidence that exposure to cigarette smoke is associated with an increased risk for colon cancer. Nevertheless, the mechanism underlying the relationship between cigarette smoking and colon cancer remains unclear. Moreover, there were only a few studies on effects of complexing substance contained in cigarette smoke on colon cancer. Thus, we further investigated whether cigarette smoke extract (CSE) affects the cell cycle, apoptosis and migration of human metastatic colon cancer cells, SW-620. MTT assay revealed that SW-620 cell proliferation was significantly inhibited following treatments with all CSEs, 3R4F, and two-domestic cigarettes, for 9 days in a concentration-dependent manner. Moreover, CSE treatments decreased cyclin D1 and E1, and increased p21 and p27 proteins by Western blot analysis in SW-620 cells. Additionally, the treatment of the cells with CSE contributed to these effects expressing by apoptosis-related proteins. An increased migration or invasion ability of SW-620 cells following CSE treatment was also confirmed by a scratch or fibronectin invasion assay in vitro. In addition, the protein levels of E-cadherin as an epithelial maker were down-regulated, while the mesenchymal markers, N-cadherin, snail, and slug, were up-regulated in a time-dependent manner. A metastatic marker, cathepsin D, was also down-regulated by CSE treatment. Taken together, these results indicate that CSE exposure in colon cancer cells may deregulate the cell growth by altering the expression of cell cycle-related proteins and pro-apoptotic protein, and stimulate cell metastatic ability by altering epithelial-mesenchymal transition (EMT) markers and cathepsin D expression. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 690-704, 2017.

  20. Transitional cell carcinoma express vitamin D receptors

    DEFF Research Database (Denmark)

    Hermann, G G; Andersen, C B

    1997-01-01

    Recently, vitamin D analogues have shown antineoplastic effect in several diseases. Vitamin D analogues exert its effect by interacting with the vitamin D receptor (VDR). Studies of VDR in transitional cell carcinoma (TCC) have not been reported. The purpose of the present study was therefore...... to examine whether human bladder tumor cells express VDR. Tumor biopsies were obtained from 26 patients with TCC. Expression of VDR was examined by immunohistochemical experiments. All tumors expressed VDR. Biopsies from advanced disease contained more VDR positive cells than low stage disease (p ....05). Similarly, also tumor grade appeared to be related to the number of cells expressing the receptor. Normal urothlium also expressed VDR but only with low intensity. Our study shows that TCC cells possess the VDR receptor which may make them capable to respond to stimulation with vitamin D, but functional...

  1. Alameda-Contra Costa Transit District (AC Transit) Fuel Cell Transit Buses: Preliminary Evaluation Results

    Energy Technology Data Exchange (ETDEWEB)

    Chandler, K.; Eudy, L.

    2007-03-01

    This report provides an evaluation of three prototype fuel cell-powered transit buses operating at AC Transit in Oakland, California, and six baseline diesel buses similar in design to the fuel cell buses.

  2. Notch1 signaling regulates the proliferation and self-renewal of human dental follicle cells by modulating the G1/S phase transition and telomerase activity.

    Directory of Open Access Journals (Sweden)

    Xuepeng Chen

    Full Text Available Multipotent human dental follicle cells (HDFCs have been intensively studied in periodontal regeneration research, yet the role of Notch1 in HDFCs has not been fully understood. The aim of the current study is to explore the role of Notch1 signaling in HDFCs self-renewal and proliferation. HDFCs were obtained from the extracted wisdom teeth from adolescent patients. Regulation of Notch1 signaling in the HDFCs was achieved by overexpressing the exogenous intracellular domain of Notch1 (ICN1 or silencing Notch1 by shRNA. The regulatory effects of Notch1 on HDFC proliferation, cell cycle distribution and the expression of cell cycle regulators were investigated through various molecular technologies, including plasmid construction, retrovirus preparation and infection, qRT-PCR, western blot, RBP-Jk luciferase reporter and cell proliferation assay. Our data clearly show that constitutively activation of Notch1 stimulates the HDFCs proliferation while inhibition of the Notch1 suppresses their proliferation in vitro. In addition, the HDFCs proliferation is associated with the increased expression of cell cycle regulators, e.g. cyclin D1, cyclin D2, cyclin D3, cyclin E1, CDK2, CDK4, CDK6, and SKP2 and the decreased expression of p27 (kip1. Moreover, our data show that the G1/S phase transition (indicating proliferation and telomerase activity (indicating self-renewal can be enhanced by overexpression of ICN1 but halted by inhibition of Notch1. Together, the current study provides evidence for the first time that Notch1 signaling regulates the proliferation and self-renewal capacity of HDFCs through modulation of the G1/S phase transition and the telomerase activity.

  3. Blockade of Jagged/Notch pathway abrogates transforming growth factor β2-induced epithelial-mesenchymal transition in human retinal pigment epithelium cells.

    Science.gov (United States)

    Chen, X; Xiao, W; Liu, X; Zeng, M; Luo, L; Wu, M; Ye, S; Liu, Y

    2014-05-01

    The epithelial-mesenchymal transition (EMT) of retinal pigment epithelium (RPE) cells plays a key role in proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR), which lead to the loss of vision. The Jagged/Notch pathway has been reported to be essential in EMT during embryonic development, fibrotic diseases and cancer metastasis. However, the function of Jagged/Notch signaling in EMT of RPE cells is unknown. Thus, we hypothesized that a crosstalk between Notch and transforming growth factor β2 (TGF-β2) signaling could induce EMT in RPE cells, which subsequently contributes to PVR and PDR. Here, we demonstrate that Jagged-1/Notch pathway is involved in the TGF-β2-mediated EMT of human RPE cells. Blockade of Notch pathway with DAPT (a specific inhibitor of Notch receptor cleavage) and knockdown of Jagged-1 expression inhibited TGF-β2-induced EMT through regulating the expression of Snail, Slug and ZEB1. Besides the canonical Smad signaling pathway, the noncanonical PI3K/Akt and MAPK pathway also contributed to TGF-β2-induced up-regulation of Jagged-1 in RPE cells. Overexpression of Jagged-1 could mimic TGF-β2 induce EMT. Our data suggest that the Jagged-1/Notch signaling pathway plays a critical role in TGF-β2-induced EMT in human RPE cells, and may contribute to the development of PVR and PDR. Inhibition of the Jagged/Notch signaling pathway, therefore, may have therapeutic value in the prevention and treatment of PVR and PDR.

  4. Knockdown of Snail inhibits epithelial-mesenchymal transition of human laryngeal squamous cell carcinoma Hep-2 cells through VDR signaling pathway.

    Science.gov (United States)

    Zhao, Xue; Yu, Dan; Yang, Jingpu; Xue, Kai; Liu, Yan; Jin, Chunshun

    2017-08-14

    It has been well-documented that Snail plays a decisive role in various tumors. However, the direct effect of Snail on laryngeal squamous cell carcinoma (LSCC) has not been elaborated. In this study, we firstly detected the expression of Snail in 14 samples of patients with LSCC and found that its content was high in cancer tissues compared with adjacent tissues. Then we established LSCC Hep-2 cells with Snail silencing and validated the knockdown efficiency by western blotting and real-time PCR. Results showed that silencing of Snail significantly inhibited the ability of adhesion, migration, and invasion of Hep-2 cells. Further study revealed that knockdown of Snail suppressed the epithelial-mesenchymal transition process of Hep-2 cells, as evidenced by downregulation of matrix metallopeptidase (MMP)-2, MMP-9, integrin subunit beta 1 (ITGβ1), β-catenin, vimentin, N-cadherin, and fibronectin (FN), while upregulation of vitamin D receptor (VDR) and E-cadherin. Additionally, transfection with the small interfering RNA of VDR reversed the effect induced by Snail silencing in Hep-2 cells. Taken together, these results demonstrate that knockdown of Snail can inhibit the EMT process of LSCC cells through VDR signaling pathway in vitro.

  5. Hepatitis B virus X protein promotes renal epithelial-mesenchymal transition in human renal proximal tubule epithelial cells through the activation of NF-κB.

    Science.gov (United States)

    Li, Mei; Hu, Liping; Zhu, Fengxin; Zhou, Zhangmei; Tian, Jianwei; Ai, Jun

    2016-08-01

    Hepatitis B virus (HBV)-associated glomerulo-nephritis is the most common extra-hepatic disorder occurring with hepatitis B virus infection. In the present study, we hypothesized that HBV X protein (HBx) may play a critical role in renal interstitial fibrosis, as HBx has been shown to induce epithelial-mesenchymal transition (EMT) in renal cells. For this purpose, we successfully transfected HBx plasmid into human renal proximal tubule epithelial cells (HK-2 cells). We found that transfection with HBx plasmid significantly downregulated E-cadherin expression and upregulated α-smooth muscle actin, collagen I and fibronectin expression in a time- and concentration-dependent manner (at the lower concentrations and earlier time points). HBx also increased nuclear factor-κB (NF-κB) phosphorylation in a time- and concentration-dependent manner (again at the lower concentrations and earlier time points); however, it did not alter the phosphorylation of Smad2, Smad3, p38, phosphoinositide 3-kinase (PI3K) or extracellular signal-regulated kinase (ERK). Thus, the findings of this study demonstrate that HBx promotes EMT in renal HK-2 cells, and the potential underlying mechanisms may involve the activation of the NF-κB signaling pathway.

  6. Calpain 1 regulates TGF-β1-induced epithelial-mesenchymal transition in human lung epithelial cells via PI3K/Akt signaling pathway

    Science.gov (United States)

    Tan, Wei-Jun; Tan, Qiu-Yue; Wang, Ting; Lian, Min; Zhang, Li; Cheng, Zhen-Shun

    2017-01-01

    Cell proliferation, transformation, and epithelial-mesenchymal transition (EMT) are key processes involved in the development of idiopathic pulmonary fibrosis (IPF). This study investigated the regulatory factors and signaling pathways that mediate EMT in the human type II alveolar epithelial A549 cell line. A549 cells were cultured in RPMI-1640 medium and allocated to the following four groups: blank control group or treated with transforming growth factor-β1 (TGF-β1), TGF-β1 + PD 150606 (a calpain 1 inhibitor), or PD 150606. We examined E-cadherin (E-cad), α-smooth muscle actin (α-SMA), and calpain 1 mRNA transcript and protein expression levels in these four groups by performing RT-PCR and western blot analyses. The results indicated that TGF-β1 treatment significantly downregulated E-cad and upregulated α-SMA expression compared with that of the blank control group (Pcells. However, TGF-β1-induced ETM was not correlated with the ERK and JNK signaling pathways. These combined results indicate that calpain 1 could regulate EMT in TGF-β1-treated A549 epithelial cells via the PI3K/Akt signaling pathway.

  7. Treatment of Human Placental Choriocarcinoma Cells with Formaldehyde and Benzene Induced Growth and Epithelial Mesenchymal Transition via Induction of an Antioxidant Effect

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    Hae-Miru Lee

    2017-07-01

    Full Text Available Cigarette smoke (CS causes about 480,000 deaths each year worldwide, and it is well-known to have harmful effects on the human body, leading to heart disease, stroke, lung cancer, and cardiovascular problems. In this study, the effects of formaldehyde (FA and benzene (Bz, the main components of CS, on cell proliferation and epithelial mesenchymal transition (EMT of JEG-3 human choriocarcinoma cells were examined to confirm the relationship between CS components and placenta carcinoma. Upon MTT assay, FA (10−8 M to 10−5 M and Bz (10−11 M to 10−8 M increased JEG-3 cell proliferation. Western blot assay revealed that the protein expression of cyclin D1 and E1 increased, while the levels of p21 and p27 were reduced following treatment. In Scratch assay, FA (10−8 M and 10−5 M and Bz (10−11 M and 10−8 M increased migration of JEG-3 cells at 24 h and 48 h compared with that at 0 h. In addition, the expression of the epithelial marker, E-cadherin, was significantly decreased, while the expression of the mesenchymal marker, N-cadherin, was significantly increased by FA (10−8 M and 10−5 M and Bz (10−11 M and 10−8 M. snail and slug transcriptional factors were associated with EMT, which were also up-regulated by FA and Bz, indicating that FA and Bz lead to an increase in the EMT process in JEG-3 choriocarcinoma cells. We further evaluated reactive oxygen species (ROS and activation of antioxidant effect using dichlorofluorescin diacetate (DCFH-DA and Western blot assay. FA and Bz increased the ROS production and an antioxidant related marker, Nrf2, in JEG-3 cells. However, eIF2α levels were reduced by FA and Bz via activation of the antioxidant reaction. Taken together, these results indicated that FA and Bz induce the growth and migration of human choriocarcinoma cells via regulation of the cell cycle and EMT and activation of ROS and antioxidant related markers.

  8. Alisertib, an Aurora kinase A inhibitor, induces apoptosis and autophagy but inhibits epithelial to mesenchymal transition in human epithelial ovarian cancer cells

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    Ding YH

    2015-01-01

    Full Text Available Yong-Hui Ding,1,2 Zhi-Wei Zhou,2,3 Chun-Fang Ha,1 Xue-Yu Zhang,1 Shu-Ting Pan,4 Zhi-Xu He,3 Jeffrey L Edelman,2 Dong Wang,5 Yin-Xue Yang,6 Xueji Zhang,7 Wei Duan,8 Tianxin Yang,9 Jia-Xuan Qiu,4 Shu-Feng Zhou2,3 1Department of Gynecology, General Hospital of Ningxia Medical University, Yinchuan, People’s Republic of China; 2Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA ; 3Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, 4Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, 5Cancer Center, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, 6Department of Colorectal Surgery, General Hospital of Ningxia Medical University, Yinchuan, 7Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 8School of Medicine, Deakin University, Waurn Ponds, Australia; 9Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA Abstract: Ovarian cancer is a leading killer of women, and no cure for advanced ovarian cancer is available. Alisertib (ALS, a selective Aurora kinase A (AURKA inhibitor, has shown potent anticancer effects, and is under clinical investigation for the treatment of advanced solid tumor and hematologic malignancies. However, the role of ALS in the treatment of ovarian cancer remains unclear. This study investigated the effects of ALS on cell growth, apoptosis, autophagy, and epithelial to mesenchymal transition (EMT, and the underlying mechanisms in human epithelial ovarian cancer SKOV3 and OVCAR4 cells. Our docking study showed that ALS, MLN8054, and VX-680 preferentially bound to AURKA

  9. Serum response factor accelerates the high glucose-induced Epithelial-to-Mesenchymal Transition (EMT) via snail signaling in human peritoneal mesothelial cells.

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    He, Lijie; Lou, Weijuan; Ji, Lihua; Liang, Wei; Zhou, Meilan; Xu, Guoshang; Zhao, Lijuan; Huang, Chen; Li, Rong; Wang, Hanmin; Chen, Xiangmei; Sun, Shiren

    2014-01-01

    Epithelial-to-Mesenchymal Transition (EMT) induced by glucose in human peritoneal mesothelial cells (HPMCs) is a major cause of peritoneal membrane (PM) fibrosis and dysfunction. To investigate serum response factor (SRF) impacts on EMT-derived fibrosis in PM, we isolated HPMCs from the effluents of patients with end-stage renal disease (ESRD) to analyze alterations during peritoneal dialysis (PD) and observe the response of PM to SRF in a rat model. Our results demonstrated the activation and translocation of SRF into the nuclei of HPMCs under extensive periods of PD. Accordingly, HPMCs lost their epithelial morphology with a decrease in E-cadherin expression and an increase in α-smooth muscle actin (α-SMA) expression, implying a transition in phenotype. PD with 4.25% glucose solution significantly induced SRF up-regulation and increased peritoneal thickness. In immortal HPMCs, high glucose (HG, 60 mmol/L) stimulated SRF overexpression in transformed fibroblastic HPMCs. SRF-siRNA preserved HPMC morphology, while transfection of SRF plasmid into HPMCs caused the opposite effects. Evidence from electrophoretic mobility shift, chromatin immunoprecipitation and reporter assays further supported that SRF transcriptionally regulated Snail, a potent inducer of EMT, by directly binding to its promoter. Our data suggested that activation of SRF/Snail pathway might contribute to the progressive PM fibrosis during PD.

  10. Beta-elemene blocks epithelial-mesenchymal transition in human breast cancer cell line MCF-7 through Smad3-mediated down-regulation of nuclear transcription factors.

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    Xian Zhang

    Full Text Available Epithelial-mesenchymal transition (EMT is the first step required for breast cancer to initiate metastasis. However, the potential of drugs to block and reverse the EMT process are not well explored. In the present study, we investigated the inhibitory effect of beta-elemene (ELE, an active component of a natural plant-derived anti-neoplastic agent in an established EMT model mediated by transforming growth factor-beta1 (TGF-β1. We found that ELE (40 µg/ml blocked the TGF-β1-induced phenotypic transition in the human breast cancer cell line MCF-7. ELE was able to inhibit TGF-β1-mediated upregulation of mRNA and protein expression of nuclear transcription factors (SNAI1, SNAI2, TWIST and SIP1, potentially through decreasing the expression and phosphorylation of Smad3, a central protein mediating the TGF-β1 signalling pathway. These findings suggest a potential therapeutic benefit of ELE in treating basal-like breast cancer.

  11. Protons Sensitize Epithelial Cells to Mesenchymal Transition

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    Wang, Minli; Hada, Megumi; Saha, Janapriya; Sridharan, Deepa M.; Pluth, Janice M.; Cucinotta, Francis A.

    2012-01-01

    Proton radiotherapy has gained more favor among oncologists as a treatment option for localized and deep-seated tumors. In addition, protons are a major constituent of the space radiation astronauts receive during space flights. The potential for these exposures to lead to, or enhance cancer risk has not been well studied. Our objective is to study the biological effects of low energy protons on epithelial cells and its propensity to enhance transforming growth factor beta 1 (TGFβ1)-mediated epithelial-mesenchymal transition (EMT), a process occurring during tumor progression and critical for invasion and metastasis. Non-transformed mink lung epithelial cells (Mv1Lu) and hTERT- immortalized human esophageal epithelial cells (EPC) were used in this study. EMT was identified by alterations in cell morphology, EMT-related gene expression changes determined using real-time PCR, and EMT changes in specific cellular markers detected by immunostaining and western blotting. Although TGFβ1 treatment alone is able to induce EMT in both Mv1Lu and EPC cells, low energy protons (5 MeV) at doses as low as 0.1 Gy can enhance TGFβ1 induced EMT. Protons alone can also induce a mild induction of EMT. SD208, a potent TGFβ Receptor 1 (TGFβR1) kinase inhibitor, can efficiently block TGFβ1/Smad signaling and attenuate EMT induction. We suggest a model for EMT after proton irradiation in normal and cancerous tissue based on our results that showed that low and high doses of protons can sensitize normal human epithelial cells to mesenchymal transition, more prominently in the presence of TGFβ1, but also in the absence of TGFβ1. PMID:22844446

  12. Protons sensitize epithelial cells to mesenchymal transition.

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    Minli Wang

    Full Text Available Proton radiotherapy has gained more favor among oncologists as a treatment option for localized and deep-seated tumors. In addition, protons are a major constituent of the space radiation astronauts receive during space flights. The potential for these exposures to lead to, or enhance cancer risk has not been well studied. Our objective is to study the biological effects of low energy protons on epithelial cells and its propensity to enhance transforming growth factor beta 1 (TGFβ1-mediated epithelial-mesenchymal transition (EMT, a process occurring during tumor progression and critical for invasion and metastasis. Non-transformed mink lung epithelial cells (Mv1Lu and hTERT- immortalized human esophageal epithelial cells (EPC were used in this study. EMT was identified by alterations in cell morphology, EMT-related gene expression changes determined using real-time PCR, and EMT changes in specific cellular markers detected by immunostaining and western blotting. Although TGFβ1 treatment alone is able to induce EMT in both Mv1Lu and EPC cells, low energy protons (5 MeV at doses as low as 0.1 Gy can enhance TGFβ1 induced EMT. Protons alone can also induce a mild induction of EMT. SD208, a potent TGFβ Receptor 1 (TGFβR1 kinase inhibitor, can efficiently block TGFβ1/Smad signaling and attenuate EMT induction. We suggest a model for EMT after proton irradiation in normal and cancerous tissue based on our results that showed that low and high doses of protons can sensitize normal human epithelial cells to mesenchymal transition, more prominently in the presence of TGFβ1, but also in the absence of TGFβ1.

  13. Epithelial mesenchymal transition status is associated with anti-cancer responses towards receptor tyrosine-kinase inhibition by dovitinib in human bladder cancer cells.

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    Hänze, Jörg; Henrici, Marcus; Hegele, Axel; Hofmann, Rainer; Olbert, Peter J

    2013-12-11

    Dovitinib (TKI-258) is a receptor tyrosine kinase (RTK) inhibitor targeting fibroblast growth factor receptor (FGFR) and further related RTKs. TKI-258 is under investigation as anticancer drug for the treatment of various cancers including bladder cancer with aberrant RTK signaling. Here, we analyzed the responses of ten human bladder cancer cell lines towards TKI-258 treatment in relation to the epithelial mesenchymal transition (EMT) status of the cells. Expression of epithelial marker E-cadherin as well as mesenchymal markers N-cadherin and vimentin was determined by quantitative RT-PCR and Western-blot in RNA and protein extracts from the cultured cell lines. The cell responses were analyzed upon addition of TKI-258 by viability/proliferation (XTT assay) and colony formation assay for measurement of cell contact independent growth. The investigated bladder cancer cell lines turned out to display quite different EMT patterns as indicated by the abundance of E-cadherin or N-cadherin and vimentin. Protein and mRNA levels of the respective components strongly correlated. Based on E-cadherin and N-cadherin mRNA levels that were expressed approximately mutual exclusively, an EMT-score was calculated for each cell line. A high EMT-score indicated mesenchymal-like cells and a low EMT-score epithelial-like cells. Then, we determined the IC₅₀ values for TKI-258 by dose response curves (0-12 μM TKI-258) in XTT assays for each cell line. Also, we measured the clonogenic survival fraction after adding TKI-258 (1 μM) by colony formation assay. We observed significant correlations between EMT-score and IC₅₀ values (r = 0.637, p = 0.0474) and between EMT-score and clonogenic survival fraction (r = 0.635, p = 0.0483) as analyzed by linear regression analyses. In sum, we demonstrated that the EMT status based on E-cadherin and N-cadherin mRNA levels may be useful to predict responses towards TKI-258 treatment in bladder cancer.

  14. Upregulation of kazrin F by miR-186 suppresses apoptosis but promotes epithelial-mesenchymal transition to contribute to malignancy in human cervical cancer cells.

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    Liu, Chang; Wang, Jinghua; Hu, Yang; Xie, Hong; Liu, Min; Tang, Hua

    2017-02-01

    Previous studies have identified that kazrin is a constituent of desmosome and influences intercellular adhesion, growing development and morphology. We previously cloned another new isoform, kazrin F and found that it has anti-apoptotic effects on human glioma cell line. To further explore whether kazrin F is involved in tumorigenesis, we investigated its expression and role in cervical cancer (CC) cells. The role of kazrin F and miR-186 in CC was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, colony formation, transwell, and apoptosis assays. Using enhanced green fluorescent protein (EGFP) reporter assays, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis, we identified kazrin F post-transcriptional regulation by miR-186. We demonstrate that kazrin F is highly expressed in CC tissues compared with the adjacent noncancerous tissues and promotes cell proliferation, colony formation, migration and invasion in HeLa and C33A cells by suppressing apoptosis and facilitating epithelial-to-mesenchymal transition (EMT). Furthermore, miR-186 was confirmed as a regulator of kazrin F dysregulation. An EGFP reporter assay proved that miR-186 directly targets the 3'-untranslated region (3'UTR) of kazrin F and downregulates its expression, and miR-186 expression showed an inverse correlation with kazrin F levels in CC tissues. In addition, overexpression of miR-186 suppressed the malignant behaviors of CC cells. The ectopic expression of kazrin F rescued the inhibitory effects of miR-186. Our findings indicate that the upregulation of kazrin F due to downregulated miR-186 levels contributes to malignancy, and highlight the significance of kazrin F in CC tumorigenesis.

  15. Upregulation of kazrin F by miR-186 suppresses apoptosis but promotes epithelial-mesenchymal transition to contribute to malignancy in human cervical cancer cells

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    Liu, Chang; Wang, Jinghua; Hu, Yang; Xie, Hong; Liu, Min; Tang, Hua

    2017-01-01

    Objective Previous studies have identified that kazrin is a constituent of desmosome and influences intercellular adhesion, growing development and morphology. We previously cloned another new isoform, kazrin F and found that it has anti-apoptotic effects on human glioma cell line. To further explore whether kazrin F is involved in tumorigenesis, we investigated its expression and role in cervical cancer (CC) cells. Methods The role of kazrin F and miR-186 in CC was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, colony formation, transwell, and apoptosis assays. Using enhanced green fluorescent protein (EGFP) reporter assays, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis, we identified kazrin F post-transcriptional regulation by miR-186. Results We demonstrate that kazrin F is highly expressed in CC tissues compared with the adjacent noncancerous tissues and promotes cell proliferation, colony formation, migration and invasion in HeLa and C33A cells by suppressing apoptosis and facilitating epithelial-to-mesenchymal transition (EMT). Furthermore, miR-186 was confirmed as a regulator of kazrin F dysregulation. An EGFP reporter assay proved that miR-186 directly targets the 3’-untranslated region (3’UTR) of kazrin F and downregulates its expression, and miR-186 expression showed an inverse correlation with kazrin F levels in CC tissues. In addition, overexpression of miR-186 suppressed the malignant behaviors of CC cells. The ectopic expression of kazrin F rescued the inhibitory effects of miR-186. Conclusions Our findings indicate that the upregulation of kazrin F due to downregulated miR-186 levels contributes to malignancy, and highlight the significance of kazrin F in CC tumorigenesis. PMID:28373753

  16. The pan-inhibitor of Aurora kinases danusertib induces apoptosis and autophagy and suppresses epithelial-to-mesenchymal transition in human breast cancer cells.

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    Li, Jin-Ping; Yang, Yin-Xue; Liu, Qi-Lun; Zhou, Zhi-Wei; Pan, Shu-Ting; He, Zhi-Xu; Zhang, Xueji; Yang, Tianxin; Pan, Si-Yuan; Duan, Wei; He, Shu-Ming; Chen, Xiao-Wu; Qiu, Jia-Xuan; Zhou, Shu-Feng

    2015-01-01

    Danusertib (Danu) is a pan-inhibitor of Aurora kinases and a third-generation breakpoint cluster region-Abelson murine leukemia viral oncogene homolog 1 (Bcr-Abl) tyrosine kinase inhibitor, but its antitumor effect and underlying mechanisms in the treatment of human breast cancer remain elusive. This study aimed to investigate the effects of Danu on the growth, apoptosis, autophagy, and epithelial-to-mesenchymal transition (EMT) and the molecular mechanisms in human breast cancer MCF7 and MDA-MB-231 cells. The results demonstrated that Danu remarkably inhibited cell proliferation, induced apoptosis and autophagy, and suppressed EMT in both breast cancer cell lines. Danu arrested MCF7 and MDA-MB-231 cells in G2/M phase, accompanied by the downregulation of cyclin-dependent kinase 1 and cyclin B1 and upregulation of p21 Waf1/Cip1, p27 Kip1, and p53. Danu significantly decreased the expression of B-cell lymphoma-extra-large (Bcl-xl) and B-cell lymphoma 2 (Bcl-2), but increased the expression of Bcl-2-associated X protein (Bax) and p53-upregulated modulator of apoptosis (PUMA), and promoted the cleavage of caspases 3 and 9. Furthermore, Danu significantly increased the expression levels of the membrane-bound microtubule-associated protein 1A/1B-light chain 3 (LC3-II) and beclin 1 in breast cancer cells, two markers for autophagy. Danu induced the activation of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinases 1 and 2 (Erk1/2) and inhibited the activation of protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathways in breast cancer cells. Treatment with wortmannin (a phosphatidylinositol 3-kinase inhibitor) markedly inhibited Danu-induced activation of p38 MAPK and conversion of cytosolic LC3-I to membrane-bound LC3-II. Pharmacological inhibition and small interfering RNA-mediated knockdown of p38 MAPK suppressed Akt activation, resulting in LC3-II accumulation and enhanced autophagy. Pharmacological inhibition

  17. Advanced Glycation End Products Induce Endothelial-to-Mesenchymal Transition via Downregulating Sirt 1 and Upregulating TGF-β in Human Endothelial Cells

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    Wei He

    2015-01-01

    Full Text Available In the present study, we examined the advanced glycation end products- (AGEs- induced endothelial-to-mesenchymal transition (EndMT in human umbilical vein endothelial cells (HUVECs. Results demonstrated that AGE-BSAs significantly reduced the cluster of differentiation 31 (CD 31 expression, whereas they promoted the expression of fibroblast-specific protein-1 (FSP-1, α-smooth muscle antibody (α-SMA, and collagen I at both mRNA and protein levels in HUVECs. And the AGE-BSAs also promoted the receptors for AGEs (RAGEs and receptor I for TGF-β (TGFR I markedly with a dose dependence, whereas the Sirt 1 was significantly downregulated by the AGE-BSA at both mRNA and protein levels. Moreover, the Sirt 1 activity manipulation with its activator, resveratrol (RSV, or its inhibitor, EX527, markedly inhibited or ameliorated the AGE-mediated TGF-β upregulation. And the manipulated Sirt 1 activity positively regulated the AGE-induced CD31, whereas it negatively regulated the AGE-induced FSP-1. Thus, Sirt 1 was confirmed to regulate the AGE-induced EndMT via TGF-β. In summary, we found that AGE-BSA induced EndMT in HUVECs via upregulating TGF-β and downregulating Sirt 1, which also negatively regulated TGF-β in the cell. This study implied the EndMT probably as an important mechanism of AGE-induced cardiovascular injury.

  18. Advanced glycation end products induce endothelial-to-mesenchymal transition via downregulating Sirt 1 and upregulating TGF-β in human endothelial cells.

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    He, Wei; Zhang, Jian; Gan, Tian-yi; Xu, Guo-jun; Tang, Bao-peng

    2015-01-01

    In the present study, we examined the advanced glycation end products- (AGEs-) induced endothelial-to-mesenchymal transition (EndMT) in human umbilical vein endothelial cells (HUVECs). Results demonstrated that AGE-BSAs significantly reduced the cluster of differentiation 31 (CD 31) expression, whereas they promoted the expression of fibroblast-specific protein-1 (FSP-1), α-smooth muscle antibody (α-SMA), and collagen I at both mRNA and protein levels in HUVECs. And the AGE-BSAs also promoted the receptors for AGEs (RAGEs) and receptor I for TGF-β (TGFR I) markedly with a dose dependence, whereas the Sirt 1 was significantly downregulated by the AGE-BSA at both mRNA and protein levels. Moreover, the Sirt 1 activity manipulation with its activator, resveratrol (RSV), or its inhibitor, EX527, markedly inhibited or ameliorated the AGE-mediated TGF-β upregulation. And the manipulated Sirt 1 activity positively regulated the AGE-induced CD31, whereas it negatively regulated the AGE-induced FSP-1. Thus, Sirt 1 was confirmed to regulate the AGE-induced EndMT via TGF-β. In summary, we found that AGE-BSA induced EndMT in HUVECs via upregulating TGF-β and downregulating Sirt 1, which also negatively regulated TGF-β in the cell. This study implied the EndMT probably as an important mechanism of AGE-induced cardiovascular injury.

  19. Telmisartan counteracts TGF-β1 induced epithelial–to–mesenchymal transition via PPAR-γ in human proximal tubule epithelial cells

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    Chen, Yumin; Luo, Qiong; Xiong, Zibo; Liang, Wei; Chen, Li; Xiong, Zuying

    2012-01-01

    Chronic renal failure (CRF) mainly results from kidney fibrosis. Epithelial-to-mesenchymal transition (EMT) occurs in stressed tubular epithelial cells and contributes to renal fibrosis. Transforming growth factor-β1 (TGF-β1) has been shown to initiate and complete the whole EMT process. Peroxisome proliferators-activated receptor-γ (PPAR-γ) exerts anti-inflammatory, anti-fibrotic and vaculo-protective effects on different renal diseases. Telmisartan is a member of angiotensin II (Ang II) receptor blocker (ARB) family. Recent studies show that Telmisartan has a partial agonistic effect on PPAR-γ. Therefore, we tested the hypothesis that Telmisartan reverses the progression of induced EMT by TGF-β1 in cultured human renal proximal tubular epithelial (HK-2) cells. Cultured HK-2 cells were treated with TGF-β1 (3 ng/ml), a combination of TGF-β1 and Telmisartan (10-200umol/L) and a combination of TGF-β1, Telmisartan and GW9662, a PPAR-γ antagonist for 48 hours. EMT was determined by quantitative real-time PCR analysis of E-cadherin (E-cad), Connective Tissue Growth Factor (CTGF) and PPAR-γ transcript expression and immunocytochemical analysis of E-cad, α-Smooth Muscle Actin (α-SMA) and PPAR-γ protein expression. TGF-β1 induced phenotypic EMT in cultured HK-2 cell line via significantly reduced E-cad expression and significantly increased CTGF, α-SMA expression in association with the loss of epithelial morphology. Telmisartan reversed all EMT markers in a dose-dependent manner which was inhibited by PPAR antagonist GW9662. In the present study, it was suggested that Telmisartan attenuated TGF-β1 induced EMT by agonistic activation of PPAR-γ. PMID:22949934

  20. Epithelial-mesenchymal transition in primary human bronchial epithelial cells is Smad-dependent and enhanced by fibronectin and TNF-α

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    Câmara Joana

    2010-01-01

    Full Text Available Abstract Background Defective epithelial repair, excess fibroblasts and myofibroblasts, collagen overproduction and fibrosis occur in a number of respiratory diseases such as asthma, chronic obstructive pulmonary disease (COPD and pulmonary fibrosis. Pathological conversion of epithelial cells into fibroblasts (epithelial-mesenchymal transition, EMT has been proposed as a mechanism for the increased fibroblast numbers and has been demonstrated to occur in lung alveolar epithelial cells. Whether other airway cell types also have the capability to undergo EMT has been less explored so far. A better understanding of the full extent of EMT in airways, and the underlying mechanisms, can provide important insights into airway disease pathology and enable the development of new therapies. The main aim of this study was to test whether primary human bronchial epithelial cells are able to undergo EMT in vitro and to investigate the effect of various profibrotic factors in the process. Results Our data demonstrate that primary human bronchial epithelial cells (HBECs are able to undergo EMT in response to transforming growth factor-beta 1 (TGF-β1, as revealed by typical morphological alterations and EMT marker progression at the RNA level by real-time quantitative polymerase chain reaction and, at the protein level, by western blot. By using pharmacological inhibitors we show that this is a Smad-dependent mechanism and is independent of extracellular signal-related kinase pathway activation. Additional cytokines and growth factors such as tumour necrosis factor-alpha (TNF-α, interleukin-1 beta (IL1β and connective tissue growth factor (CTGF were also tested, alone or in combination with TGF-β1. TNF-α markedly enhances the effect of TGF-β1 on EMT, whereas IL1β shows only a very weak effect and CTGF has no significant effect. We have also found that cell-matrix contact, in particular to fibronectin, an ECM component upregulated in fibrotic lesions

  1. miR156a Mimic Represses the Epithelial-Mesenchymal Transition of Human Nasopharyngeal Cancer Cells by Targeting Junctional Adhesion Molecule A.

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    Yunhong Tian

    Full Text Available MicroRNAs (miRNAs have been documented as having an important role in the development of cancer. Broccoli is very popular in large groups of the population and has anticancer properties. Junctional adhesion molecule A (JAMA is preferentially concentrated at tight junctions and influences cell morphology and migration. Epithelial-mesenchymal transition (EMT is a developmental program associated with cancer progression and metastasis. In this study we aimed to investigate the role of miRNAs from broccoli in human nasopharyngeal cancer (NPC. We demonstrated that a total of 84 conserved miRNAs and 184 putative novel miRNAs were found in broccoli by sequencing technology. Among these, miR156a was expressed the most. In addition, synthetic miR156a mimic inhibited the EMT of NPC cells in vitro. Furthermore, it was confirmed that JAMA was the target of miR156a mimic as validated by 3' UTR luciferase reporter assays and western blotting. Knockdown of JAMA was consistent with the effects of miR156a mimic on the EMT of NPC, and the up-regulation of JAMA could partially restore EMT repressed by miR156a mimic. In conclusion, these results indicate that the miR156a mimic inhibits the EMT of NPC cells by targeting the 3' UTR of JAMA. These miRNA profiles of broccoli provide a fundamental basis for further research. Moreover, the discovery of miR156a may have clinical implications for the treatment of patients with NPC.

  2. The pan-inhibitor of Aurora kinases danusertib induces apoptosis and autophagy and suppresses epithelial-to-mesenchymal transition in human breast cancer cells

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    Li JP

    2015-02-01

    , but its antitumor effect and underlying mechanisms in the treatment of human breast cancer remain elusive. This study aimed to investigate the effects of Danu on the growth, apoptosis, autophagy, and epithelial-to-mesenchymal transition (EMT and the molecular mechanisms in human breast cancer MCF7 and MDA-MB-231 cells. The results demonstrated that Danu remarkably inhibited cell proliferation, induced apoptosis and autophagy, and suppressed EMT in both breast cancer cell lines. Danu arrested MCF7 and MDA-MB-231 cells in G2/M phase, accompanied by the downregulation of cyclin-dependent kinase 1 and cyclin B1 and upregulation of p21 Waf1/Cip1, p27 Kip1, and p53. Danu significantly decreased the expression of B-cell lymphoma-extra-large (Bcl-xl and B-cell lymphoma 2 (Bcl-2, but increased the expression of Bcl-2-associated X protein (Bax and p53-upregulated modulator of apoptosis (PUMA, and promoted the cleavage of caspases 3 and 9. Furthermore, Danu significantly increased the expression levels of the membrane-bound microtubule-associated protein 1A/1B-light chain 3 (LC3-II and beclin 1 in breast cancer cells, two markers for autophagy. Danu induced the activation of p38 mitogen-activated protein kinase (MAPK and extracellular signal-regulated kinases 1 and 2 (Erk1/2 and inhibited the activation of protein kinase B (Akt/mammalian target of rapamycin (mTOR signaling pathways in breast cancer cells. Treatment with wortmannin (a phosphatidylinositol 3-kinase inhibitor markedly inhibited Danu-induced activation of p38 MAPK and conversion of cytosolic LC3-I to membrane-bound LC3-II. Pharmacological inhibition and small interfering RNA-mediated knockdown of p38 MAPK suppressed Akt activation, resulting in LC3-II accumulation and enhanced autophagy. Pharmacological inhibition and small interfering RNA-mediated knockdown of Erk1/2 also remarkably increased the level of LC3-II in MCF7 cells. Moreover, Danu inhibited EMT in both MCF7 and MDA-MB-231 cells with upregulated E

  3. Kruppel-Like Factor 4 Overexpression Initiates a Mesenchymal-to-Epithelial Transition and Redifferentiation of Human Pancreatic Cells following Expansion in Long Term Adherent Culture.

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    Kenneth R Muir

    Full Text Available A replenishable source of insulin-producing cells has the potential to cure type 1 diabetes. Attempts to culture and expand pancreatic β-cells in vitro have resulted in their transition from insulin-producing epithelial cells to mesenchymal stromal cells (MSCs with high proliferative capacity but devoid of any hormone production. The aim of this study was to determine whether the transcription factor Krüppel-like factor 4 (KLF4, could induce a mesenchymal-to-epithelial transition (MET of the cultured cells. Islet-enriched pancreatic cells, allowed to dedifferentiate and expand in adherent cell culture, were transduced with an adenovirus containing KLF4 (Ad-Klf4. Cells were subsequently analysed for changes in cell morphology by light microscopy, and for the presence of epithelial and pancreatic markers by immunocytochemistry and quantitative RT/PCR. Infection with Ad-Klf4 resulted in morphological changes, down-regulation of mesenchymal markers, and re-expression of both epithelial and pancreatic cell markers including insulin and transcription factors specific to β-cells. This effect was further enhanced by culturing cells in suspension. However, the effects of Ad-KLf4 were transient and this was shown to be due to increased apoptosis in Klf4-expressing cells. Klf4 has been recently identified as a pioneer factor with the ability to modulate the structure of chromatin and enhance reprogramming/transdifferentiation. Our results show that Klf4 may have a role in the redifferentiation of expanded pancreatic cells in culture, but before this can be achieved the off-target effects that result in increased apoptosis would need to be overcome.

  4. CSTEA: a webserver for the Cell State Transition Expression Atlas.

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    Zhu, Guanghui; Yang, Hui; Chen, Xiao; Wu, Jun; Zhang, Yong; Zhao, Xing-Ming

    2017-05-09

    Cell state transition is one of the fundamental events in the development of multicellular organisms, and the transition trajectory path has recently attracted much attention. With the accumulation of large amounts of "-omics" data, it is becoming possible to get insights into the molecule mechanisms underlying the transitions between cell states. Here, we present CSTEA (Cell State Transition Expression Atlas), a webserver that organizes, analyzes and visualizes the time-course gene expression data during cell differentiation, cellular reprogramming and trans-differentiation in human and mouse. In particular, CSTEA defines gene signatures for uncharacterized stages during cell state transitions, thereby enabling both experimental and computational biologists to better understand the mechanisms of cell fate determination in mammals. To our best knowledge, CSTEA is the first webserver dedicated to the analysis of time-series gene expression data during cell state transitions. CSTEA is freely available at http://comp-sysbio.org/cstea/. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  5. PATTERN OF TRANSITIONAL CELL CARCINOMA OF THE ...

    African Journals Online (AJOL)

    hi-tech

    2004-03-03

    Mar 3, 2004 ... cancer of the urinary tract and transitional cell carcinoma accounts for ... Ndaguatha in a review of urinary bladder cancers in ... (ERBBI), EGFR (ERBB2), MYC and SRC. Their ..... Schairer, C., Harge, P., Hoover, R. N., et al.

  6. The mechanism of TGF-β/miR-155/c-Ski regulates endothelial-mesenchymal transition in human coronary artery endothelial cells.

    Science.gov (United States)

    Wang, Juan; He, Wen; Xu, Xiao; Guo, Liping; Zhang, Yin; Han, Suxia; Shen, Difei

    2017-08-31

    Human coronary artery endothelial cells (HCAECs) have the potential to undergo fibrogenic endothelial-mesenchymal transition (EndMT), which results in matrix-producing fibroblasts and thereby contributes to the pathogenesis of cardiac fibrosis. Recently, the profibrotic cytokine transforming growth factor-β (TGF-β) is shown to be the crucial pathogenic driver which has been verified to induce EndMT. C-Ski is an important regulator of TGF-β signaling. However, the detailed role of c-Ski and the molecular mechanisms by which c-Ski affects TGF-β-induced EndMT in HCAECs are not largely elucidated. In the present study, we treated HCAECs with TGF-β of different concentrations to induce EndMT. We found that overexpression of c-Ski in HCAECs either blocked EndMT via hindering Vimentin, Snail, Slug, and Twist expression while enhancing CD31 expression, with or without TGF-β treatment. In contrast, suppression of c-Ski further enhanced EndMT. Currently, miRNA expression disorder has been frequently reported associating with cardiac fibrosis. By using online tools, we regarded miR-155 as a candidate miRNA that could target c-Ski, which was verified using luciferase assays. C-Ski expression was negatively regulated by miR-155. TGF-β-induced EndMT was inhibited by miR-155 silence; the effect of TGF-β on Vimentin, CD31, Snail, Slug, and Twist could be partially restored by miR-155. Altogether, these findings will shed light on the role and mechanism by which miR-155 regulates TGF-β-induced HCAECs EndMT via c-Ski to affect cardiac fibrosis, and miR-155/c-Ski may represent novel biomarkers and therapeutic targets in the treatment of cardiac fibrosis. © 2017 The Author(s).

  7. Visualizing Cell State Transition Using Raman Spectroscopy

    Science.gov (United States)

    Ichimura, Taro; Chiu, Liang-da; Fujita, Katsumasa; Kawata, Satoshi; Watanabe, Tomonobu M.; Yanagida, Toshio; Fujita, Hideaki

    2014-01-01

    System level understanding of the cell requires detailed description of the cell state, which is often characterized by the expression levels of proteins. However, understanding the cell state requires comprehensive information of the cell, which is usually obtained from a large number of cells and their disruption. In this study, we used Raman spectroscopy, which can report changes in the cell state without introducing any label, as a non-invasive method with single cell capability. Significant differences in Raman spectra were observed at the levels of both the cytosol and nucleus in different cell-lines from mouse, indicating that Raman spectra reflect differences in the cell state. Difference in cell state was observed before and after the induction of differentiation in neuroblastoma and adipocytes, showing that Raman spectra can detect subtle changes in the cell state. Cell state transitions during embryonic stem cell (ESC) differentiation were visualized when Raman spectroscopy was coupled with principal component analysis (PCA), which showed gradual transition in the cell states during differentiation. Detailed analysis showed that the diversity between cells are large in undifferentiated ESC and in mesenchymal stem cells compared with terminally differentiated cells, implying that the cell state in stem cells stochastically fluctuates during the self-renewal process. The present study strongly indicates that Raman spectral morphology, in combination with PCA, can be used to establish cells' fingerprints, which can be useful for distinguishing and identifying different cellular states. PMID:24409302

  8. Epithelial-Mesenchymal Transitions and the Expression of Twist in MCF-7/ADR,Human Multidrug-Resistant Breast Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    Fei Zhang; Yurong Shi; Lin Zhang; Bin Zhang; Xiyin Wei; Yi Yang; RUi Wang; Ruifang Niu

    2007-01-01

    OBJECTIVE To study the expression levels of Twist and epithelialmesenchymal transitions in multidrug-resistant MCF-7/ADR breast cancer cells,and to study the relationship between multidrug resistance (MDR) and metastatic potential of the cells.METHODS RT-PCR,immunohislochemical and Western blotting methods were used to examine the changes of expression levels of the transcription factor Twist.E-cadherin and N-cadherin in the MCF-7 breast cancer cell line and its multidrug-resistant variant.MCF-7/ADR.RESULTS In MCF-7 cells,the expression of E-cadherin can be detected,but there is no expression of Twisl or N-cadherin.In MCF-7/ADR cells,E-cadherin expression is lost.bul the expression of two other genes was significantly positive.CONCLUSION Epithelial-mesenchymal transitions induced by Twist,may have a relationship with enhanced invasion and metastatic potential during the development of multidrug-resistant MCF-7/ADR breast cancer cells.

  9. The involvement of hematopoietic pre-B cell leukemia transcription factor-interacting protein in regulating epithelial-mesenchymal transition of human spinal glioblastoma.

    Science.gov (United States)

    Wang, Deliang; Wang, Li; Zhou, Yi; Zhao, Xinjun; Xiong, Hui

    2016-05-01

    To date, hematopoietic pre-B cell leukemia transcription factor-interacting protein (HPIP), a co-repressor for the transcription factor PBX, has been involved into the initiation and onset in a wide variety of cancers. However, the molecular mechanisms underlying HPIP-induced epithelial-mesenchymal transition (EMT) in the spinal glioblastoma have been under investigation. In the present study, spinal glioblastoma tissues, U87, and U251 cell lines were used and subjected to in vitro assays, such as RT-PCR, and Western blot. Here, in vitro assays revealed that HPIP mRNA and protein were highly expressed in five cases of spinal glioblastoma tissues, compared with non-tumor tissues. Subsequently, in vitro experiments demonstrated HPIP promoted the U87 and U251 cell growth and regulated the G1/S phase transitions in U87 and U251 cell cycle, respectively, accompanied by the increased expression of cyclin A2, cyclin B1, and cyclin D1. Furthermore, HPIP increased the expression of N-cadherin, Slug, and MMP2, and decreased the expression of E-cadherin. By contrast, knockdown of HPIP reversed HPIP-induced EMT biomarkers, migration, and invasion in U87 and U251 cells. In conclusion, our findings identified HPIP plays an important role in the progression and EMT of spinal glioblastoma, by which cell growth is improved. Thus, HPIP gene or protein could act as a useful target in the clinical practice.

  10. Kinematics of transition during human accelerated sprinting

    Directory of Open Access Journals (Sweden)

    Ryu Nagahara

    2014-07-01

    Full Text Available This study investigated kinematics of human accelerated sprinting through 50 m and examined whether there is transition and changes in acceleration strategies during the entire acceleration phase. Twelve male sprinters performed a 60-m sprint, during which step-to-step kinematics were captured using 60 infrared cameras. To detect the transition during the acceleration phase, the mean height of the whole-body centre of gravity (CG during the support phase was adopted as a measure. Detection methods found two transitions during the entire acceleration phase of maximal sprinting, and the acceleration phase could thus be divided into initial, middle, and final sections. Discriminable kinematic changes were found when the sprinters crossed the detected first transition—the foot contacting the ground in front of the CG, the knee-joint starting to flex during the support phase, terminating an increase in step frequency—and second transition—the termination of changes in body postures and the start of a slight decrease in the intensity of hip-joint movements, thus validating the employed methods. In each acceleration section, different contributions of lower-extremity segments to increase in the CG forward velocity—thigh and shank for the initial section, thigh, shank, and foot for the middle section, shank and foot for the final section—were verified, establishing different acceleration strategies during the entire acceleration phase. In conclusion, there are presumably two transitions during human maximal accelerated sprinting that divide the entire acceleration phase into three sections, and different acceleration strategies represented by the contributions of the segments for running speed are employed.

  11. N-myc downstream regulated gene 1 (NDRG1 promotes metastasis of human scirrhous gastric cancer cells through epithelial mesenchymal transition.

    Directory of Open Access Journals (Sweden)

    Hiroki Ureshino

    Full Text Available Our recent study demonstrated that higher expression of N-myc downregulated gene 1 (NDRG1 is closely correlated with poor prognosis in gastric cancer patients. In this study, we asked whether NDRG1 has pivotal roles in malignant progression including metastasis of gastric cancer cells. By gene expression microarray analysis expression of NDRG1 showed the higher increase among a total of 3691 up-regulated genes in a highly metastatic gastric cancer cell line (58As1 than their parental low metastatic counterpart (HSC-58. The highly metastatic cell lines showed decreased expression of E-cadherin, together with enhanced expression of vimentin and Snail. This decreased expression of E-cadherin was restored by Snail knockdown in highly metastatic cell lines. We next established stable NDRG1 knockdown cell lines (As1/Sic50 and As1/Sic54 from the highly metastatic cell line, and both of these cell lines showed enhanced expression of E-cadherin and decreased expression of vimentin and Snail. And also, E-cadherin promoter-driven luciferase activity was found to be increased by NDRG1 knockdown in the highly metastatic cell line. NDRG1 knockdown in gastric cancer cell showed suppressed invasion of cancer cells into surround tissues, suppressed metastasis to the peritoneum and decreased ascites accumulation in mice with significantly improved survival rates. This is the first study to demonstrate that NDRG1 plays its pivotal role in the malignant progression of gastric cancer through epithelial mesenchymal transition.

  12. [The expression of transcription factors Snail and Slug in epithelial-mesenchymal transition of human lens epithelial cells induced by transforming growth factor-β2].

    Science.gov (United States)

    Wang, Y N; Pei, C; Qin, L; Li, J M; Yi, J L; Chen, L

    2016-04-11

    To investigate the expression of transcription factors snail and slug in epithelial mesenchymal transition (EMT) of human lens epithelial cells (HLEC) induced by transforming growth factor-β2 (TGF-β2). Experimental research. HLEC were treated with different concentrations of TGF-β2 (1.0 and 10.0 μg/L) for different time. The morphological changes were observed under inverted microscope. The expression and cellular localization of snail and slug were evaluated by immunofluorescence. Expressions of snail, slug, E-Cadherin and α-SMA were further determined by Western blot analysis. Single factor analysis of variance, rank sum test and Pearson correlation were used for statistical analysis. Cultured HLEC were polygonal monolayer cells with tight intercellular adhesion closely and patchy distribution. After treatment of different doses of TGF-β2 for 24 h, HLEC became isolated, exhibited long spindle-like shape as fibroblastic phenotype. The immunofluorescence staining indicated that snail and slug were localized in the nuclei. The expressions of snail and slug appeared to be positive correlative to TGF-β2 dose (snail protein expression: 0.74±0.16, 1.13±0.03, 1.54±0.18 and slug protein expression: 1.96±0.02, 3.12±0.09, 4.07±0.12 in HLEC treated with 0.1, 1.0 and 10 μg/L TGF-β2 respectively) (χ(2)=9.62,P=0.022;F=241.10,Psnail and slug in HLEC were also increased with extending duration of TGF-β2 (1.0 μg/L). The expression levels of both proteins were modestly up-regulated at 8 hours, robustly increased at 24 h, reached peak at 48h and began to decline at 72 h (snail protein expression: 0.90±0.13, 1.43±0.14, 1.96±0.27, 1.57±0.16 and slug protein expression: 0.91±0.36, 1.24±0.16, 2.44±0.26, 1.43±0.16 in HLEC treated with 1.0 μg/L TGF-β2 for 8 h, 24 h, 48 h and 72 h respectively) (F=12.49,P=0.001;F=14.03,Psnail and slug might be time and dose-dependently involved in in-vitro TGF-β2-induced EMT of HLEC. (Chin J Ophthalmol, 2016, 52: 285-290).

  13. Cholera toxin, a typical protein kinase A activator, induces G1 phase growth arrest in human bladder transitional cell carcinoma cells via inhibiting the c-Raf/MEK/ERK signaling pathway.

    Science.gov (United States)

    Zheng, Xiaoke; Ou, Yanqiu; Shu, Minfeng; Wang, Youqiong; Zhou, Yuxi; Su, Xingwen; Zhu, Wenbo; Yin, Wei; Li, Shifeng; Qiu, Pengxin; Yan, Guangmei; Zhang, Jingxia; Hu, Jun; Xu, Dong

    2014-05-01

    The biotoxin cholera toxin has been demonstrated to have anti-tumor activity in numerous types of cancer, including glioma. However, the role of cholera toxin in the tumorigenesis of transitional cell carcinoma (TCC), the most common malignant tumor of the bladder, remains to be elucidated. To address this, in the present study, two TCC cell lines, T24 and UM-UC-3, were treated with cholera toxin [protein kinase A (PKA) activator] and KT5720 (PKA inhibitor). Cell survival and proliferation, cell cycle alterations and apoptosis were analyzed using Hoechst staining, the MTT assay, fluorescence microscopy and flow cytometry. Western blot analysis was used to detect the expression of proteins involved in cell cycle regulation. The results revealed that cholera toxin significantly induced G1 arrest and downregulated the expression of cyclin D1 and cyclin-dependent kinase 4/6 in the TCC cell lines, and this was rescued by KT5720. Furthermore, it was demonstrated that cholera toxin downregulated the activation of the c-Raf/Mek/Erk cascade, an important mediator of tumor cell proliferation, via the PKA-dependent c-Raf phosphorylation at Ser-43. Furthermore, inhibition of Mek activity with UO126 mimicked the effects of cholera toxin. In conclusion, these results confirmed that cholera toxin specifically inhibited proliferation and induced G1 phase arrest in human bladder TCC cells. This effect was due to PKA-dependent inactivation of the c-Raf/Mek/Erk pathway. This suggested that cholera toxin may be a viable therapeutic treatment against tumorigenesis and proliferation in bladder cancer.

  14. Osteopontin Promotes Invasion, Migration and Epithelial-Mesenchymal Transition of Human Endometrial Carcinoma Cell HEC-1A Through AKT and ERK1/2 Signaling

    Directory of Open Access Journals (Sweden)

    Yinghua Li

    2015-10-01

    Full Text Available Background/Aims: Osteopontin (OPN is an Extracellular Matrix (ECM molecule and is involved in many physiologic and pathologic processes, including cell adhesion, angiogenesis and tumor metastasis. OPN is a well-known multifunctional factor involved in various aspects of cancer progression, including endometrial cancer. In this study, we examined the significance of OPN in endometrial cancer. Methods: The proliferation, migration and invasion ability of HEC-1A cells were detected by Cell Counting Kit-8 (CCK-8, Wound scratch assay and transwell. Western blots were employed to detect the expression of Matrix metalloproteinase-2 (MMP-2 and epithelial-mesenchymal transition (EMT-related factors in HEC-1A cells treated with rhOPN. Results: rhOPN promotes cell proliferation, migration and invasion in HEC-1A cells. rhOPN influenced EMT-related factors and MMP-2 expression in HEC-1A cells. rhOPN promoted HEC-1A cells migration, invasion and EMT through protein kinase B (PKB/AKT and Extracellular regulated protein kinases (ERK1/2 signaling pathway. Conclusions: These results may open up a novel therapeutic strategy for endometrial cancer: namely, rhOPN have important roles in controlling growth of endometrial of cancer cells and suggest a novel target pathway for treatment of this cancer.

  15. Climate change, human health, and epidemiological transition.

    Science.gov (United States)

    Barrett, Bruce; Charles, Joel W; Temte, Jonathan L

    2015-01-01

    The health of populations depends on the availability of clean air, water, food, and sanitation, exposure to pathogens, toxins and environmental hazards, and numerous genetic, behavioral and social factors. For many thousands of years, human life expectancy was low, and population growth was slow. The development of technology-based civilizations facilitated what Abdel Omran called "epidemiological transition," with increasing life expectancy and rapid population growth. To a large extent, the spectacular growth of human populations during the past two centuries was made possible by the energy extracted from fossil fuels. We have now learned, however, that greenhouse gases from fossil fuel combustion are warming the planet's surface, causing changes in oceanic and atmospheric systems, and disrupting weather and hydrological patterns. Climate change poses unprecedented threats to human health by impacts on food and water security, heat waves and droughts, violent storms, infectious disease, and rising sea levels. Whether or not humanity can reduce greenhouse gas emissions quickly enough to slow climate change to a rate that will allow societies to successfully adapt is not yet known. This essay reviews the current state of relevant knowledge, and points in a few directions that those interested in human health may wish to consider. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. Transitional cell carcinoma of the sinonasal tract: A rare entity

    Directory of Open Access Journals (Sweden)

    Madhumita Mondal

    2015-01-01

    Full Text Available Malignant sinonasal carcinomas are a rare entity comprising less than 1% of all cancers and around 3% of all head and neck malignancies seen in humans. Among these 15-20% are transitional cell carcinoma also known as non keratinizing carcinoma of sinonasal tract. We are reporting the case of a 45 years female with history of nasal obstruction and epistaxis. A contrast enhanced computed tomography (CECT was done which showed mucosal thickening in the right nasal cavity. Endoscopy assisted biopsy was taken which revealed non keratinizing carcinoma (transitional type. Very few reported cases of this type of malignancy was found. A possible reason could be multiple synonyms like cylindrical cell carcinoma, Schneiderian carcinoma and transitional cell carcinoma.

  17. Taiwan cobra cardiotoxin III suppresses EGF/EGFR-mediated epithelial-to-mesenchymal transition and invasion of human breast cancer MDA-MB-231 cells.

    Science.gov (United States)

    Tsai, Pei-Chien; Fu, Yaw-Syan; Chang, Long-Sen; Lin, Shinne-Ren

    2016-03-01

    Breast cancer is a highly malignant carcinoma and most deaths of breast cancer are caused by metastasis. The epithelial-to-mesenchymal transition (EMT) has emerged as a pivotal event in the development of the invasive and metastatic potentials of cancer progression. Epidermal growth factor (EGF) and its receptor, EGFR, play roles in cancer metastasis. CTX III, a basic polypeptide isolated from Naja naja atra venom, has been shown to exhibit anticancer activity; however, the effect of CTX III on the EMT of cancer cells remains elusive. CTX III treatment resulted in morphological changes from elongated and spindle shape to rounded and epithelial-like shape, induced upregulation of E-cadherin and concurrent downregulation of N-cadherin and Vimentin protein levels, corresponding to observed decreases in cell migration and invasion. CTX III treatment also decreased the expression of Snail and Twist in EGF-induced MDA-MB-231 cells. Concurrently, CTX III efficiently inhibited the EGFR phosphorylation and downstream activation of phosphatidylinositol 3-kinase (PI3K)/Akt and ERK1/2. The EGFR specific inhibitor AG1478 significantly suppressed ERK1/2 and Akt phosphorylation, cell migration and invasion, as well as the expressional changes associated with EMT markers in EGF-induced MDA-MB-231 cells. CTX III inhibitory effect on EGF-evoked invasion of MDA-MB-231 cells is mediated through suppressing EGF/EGFR activation and EMT process.

  18. Phase transitions in models of human cooperation

    Science.gov (United States)

    Perc, Matjaž

    2016-08-01

    If only the fittest survive, why should one cooperate? Why should one sacrifice personal benefits for the common good? Recent research indicates that a comprehensive answer to such questions requires that we look beyond the individual and focus on the collective behavior that emerges as a result of the interactions among individuals, groups, and societies. Although undoubtedly driven also by culture and cognition, human cooperation is just as well an emergent, collective phenomenon in a complex system. Nonequilibrium statistical physics, in particular the collective behavior of interacting particles near phase transitions, has already been recognized as very valuable for understanding counterintuitive evolutionary outcomes. However, unlike pairwise interactions among particles that typically govern solid-state physics systems, interactions among humans often involve group interactions, and they also involve a larger number of possible states even for the most simplified description of reality. Here we briefly review research done in the realm of the public goods game, and we outline future research directions with an emphasis on merging the most recent advances in the social sciences with methods of nonequilibrium statistical physics. By having a firm theoretical grip on human cooperation, we can hope to engineer better social systems and develop more efficient policies for a sustainable and better future.

  19. Neuropilin-1 promotes epithelial-to-mesenchymal transition by stimulating nuclear factor-kappa B and is associated with poor prognosis in human oral squamous cell carcinoma.

    Directory of Open Access Journals (Sweden)

    Weiming Chu

    Full Text Available The epithelial-to-mesenchymal transition (EMT is a key process in carcinogenesis, invasion, and metastasis of oral squamous cell carcinoma (OSCC. In our previous studies, we found that neuropilin-1 (NRP1 is overexpressed in tongue squamous cell carcinoma and that this overexpression is associated with cell migration and invasion. Nuclear factor-kappa B (NF-κB plays an essential role both in the induction and the maintenance of EMT and tumor metastasis. Therefore, we hypothesized that NRP1 induces EMT, and that NRP1-induced migration and invasion may be an important mechanism for promoting invasion and metastasis of OSCC through NF-κB activation.The variations in gene and protein expression and the changes in the biological behavior of OSCC cell lines transfected with a vector encoding NRP1, or the corresponding vector control, were evaluated. NRP1 overexpression promoted EMT and was associated with enhanced invasive and metastatic properties. Furthermore, the induction of EMT promoted the acquisition of some cancer stem cell (CSC-like characteristics in OSCC cells. We addressed whether selective inhibition of NF-κB suppresses the NRP1-mediated EMT by treating cells with pyrrolidinedithiocarbamate ammonium (PDTC, an inhibitor of NF-κB. Immunohistochemical analysis of NRP1 in OSCC tissue samples further supported a key mediator role for NRP1 in tumor progression, lymph node metastasis, and indicated that NRP1 is a predictor for poor prognosis in OSCC patients.Our results indicate that NRP1 may regulate the EMT process in OSCC cell lines through NF-κB activation, and that higher NRP1 expression levels are associated with lymph node metastasis and poor prognosis in OSCC patients. Further investigation of the role of NRP1 in tumorigenesis may help identify novel targets for the prevention and therapy of oral cancers.

  20. Resveratrol inhibits transforming growth factor-β2-induced epithelial-to-mesenchymal transition in human retinal pigment epithelial cells by suppressing the Smad pathway

    Science.gov (United States)

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

    2017-01-01

    Proliferative vitreoretinopathy (PVR) is the main cause of failure following retinal detachment surgery. Transforming growth factor (TGF)-β2-induced epithelial-to-mesenchymal transition (EMT) plays an important role in the development of PVR, and EMT inhibition decreases collagen gel contraction and fibrotic membrane formation, resulting in prevention of PVR. Resveratrol is naturally found in red wine and has inhibitory effects on EMT. Resveratrol is widely used in cardioprotection, neuroprotection, chemotherapy, and antiaging therapy. The purpose of this study was to investigate the effects of resveratrol on TGF-β2-induced EMT in ARPE-19 cells in vitro. We found that resveratrol suppressed the decrease of zona occludens-1 (ZO-1) and caused an increase of alpha-smooth muscle actin expression in TGF-β2-treated ARPE-19 cells, assessed using Western blots; moreover, it also suppressed the decrease in ZO-1 and the increase of vimentin expression, observed using immunocytochemistry. Resveratrol attenuated TGF-β2-induced wound closure and cell migration in ARPE-19 cells in a scratch wound test and modified Boyden chamber assay, respectively. We also found that resveratrol reduced collagen gel contraction – assessed by collagen matrix contraction assay – and suppressed the phosphorylation of Smad2 and Smad3 in TGF-β2-treated ARPE-19 cells. These results suggest that resveratrol mediates anti-EMT effects, which could be used in the prevention of PVR.

  1. Resveratrol inhibits transforming growth factor-β2-induced epithelial-to-mesenchymal transition in human retinal pigment epithelial cells by suppressing the Smad pathway.

    Science.gov (United States)

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

    2017-01-01

    Proliferative vitreoretinopathy (PVR) is the main cause of failure following retinal detachment surgery. Transforming growth factor (TGF)-β2-induced epithelial-to-mesenchymal transition (EMT) plays an important role in the development of PVR, and EMT inhibition decreases collagen gel contraction and fibrotic membrane formation, resulting in prevention of PVR. Resveratrol is naturally found in red wine and has inhibitory effects on EMT. Resveratrol is widely used in cardioprotection, neuroprotection, chemotherapy, and antiaging therapy. The purpose of this study was to investigate the effects of resveratrol on TGF-β2-induced EMT in ARPE-19 cells in vitro. We found that resveratrol suppressed the decrease of zona occludens-1 (ZO-1) and caused an increase of alpha-smooth muscle actin expression in TGF-β2-treated ARPE-19 cells, assessed using Western blots; moreover, it also suppressed the decrease in ZO-1 and the increase of vimentin expression, observed using immunocytochemistry. Resveratrol attenuated TGF-β2-induced wound closure and cell migration in ARPE-19 cells in a scratch wound test and modified Boyden chamber assay, respectively. We also found that resveratrol reduced collagen gel contraction - assessed by collagen matrix contraction assay - and suppressed the phosphorylation of Smad2 and Smad3 in TGF-β2-treated ARPE-19 cells. These results suggest that resveratrol mediates anti-EMT effects, which could be used in the prevention of PVR.

  2. Regulation of Na,K-ATPase β1-subunit in TGF-β2-mediated epithelial-to-mesenchymal transition in human retinal pigmented epithelial cells.

    Science.gov (United States)

    Mony, Sridevi; Lee, Seung Joon; Harper, Jeffrey F; Barwe, Sonali P; Langhans, Sigrid A

    2013-10-01

    Proliferative vitreo retinopathy (PVR) is associated with extracellular matrix membrane (ECM) formation on the neural retina and disruption of the multilayered retinal architecture leading to distorted vision and blindness. During disease progression in PVR, retinal pigmented epithelial cells (RPE) lose cell-cell adhesion, undergo epithelial-to-mesenchymal transition (EMT), and deposit ECM leading to tissue fibrosis. The EMT process is mediated via exposure to vitreous cytokines and growth factors such as TGF-β2. Previous studies have shown that Na,K-ATPase is required for maintaining a normal polarized epithelial phenotype and that decreased Na,K-ATPase function and subunit levels are associated with TGF-β1-mediated EMT in kidney cells. In contrast to the basolateral localization of Na,K-ATPase in most epithelia, including kidney, Na,K-ATPase is found on the apical membrane in RPE cells. We now show that EMT is also associated with altered Na,K-ATPase expression in RPE cells. TGF-β2 treatment of ARPE-19 cells resulted in a time-dependent decrease in Na,K-ATPase β1 mRNA and protein levels while Na,K-ATPase α1 levels, Na,K-ATPase activity, and intracellular sodium levels remained largely unchanged. In TGF-β2-treated cells reduced Na,K-ATPase β1 mRNA inversely correlated with HIF-1α levels and analysis of the Na,K-ATPase β1 promoter revealed a putative hypoxia response element (HRE). HIF-1α bound to the Na,K-ATPase β1 promoter and inhibiting the activity of HIF-1α blocked the TGF-β2 mediated Na,K-ATPase β1 decrease suggesting that HIF-1α plays a potential role in Na,K-ATPase β1 regulation during EMT in RPE cells. Furthermore, knockdown of Na,K-ATPase β1 in ARPE-19 cells was associated with a change in cell morphology from epithelial to mesenchymal and induction of EMT markers such as α-smooth muscle actin and fibronectin, suggesting that loss of Na,K-ATPase β1 is a potential contributor to TGF-β2-mediated EMT in RPE cells.

  3. Receptor activator of NF-kB Ligand (RANKL) expression is associated with epithelial to mesenchymal transition in human prostate cancer cells

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Epithelial-mesenchymal transition (EMT) in cancer describes the phenotypic and behavioral changes of cancer cells from indolent to virulent forms with increased migratory, invasive and metastatic potential. EMT can be induced by soluble proteins like transforming growth factor β1 (TGFβ1) and transcription factors including Snail and Slug. We utilized the ARCaPE/ARCaPM prostate cancer progression model and LNCaP clones stably overexpressing Snail to identify novel markers associated with EMT. Compared to ARCaPE cells, the highly tumorigenic mesenchymai ARCaPM and ARCaPM1 variant cells displayed a higher incidence of bone metastasis after intracardiac administration in SCID mice. ARCaPM and ARCaPM1 expressed mesenchymal stromal markers of vimentin and N-cadherin in addition to elevated levels of Receptor Activator of NF-kB Ligand (RANKL). We observed that both epidermal growth factor (EGF) plus TGFβ1 treatment and Snail overexpression induced EMT in ARCaPE and LNCaP cells, and EMT was associated with increased expression of RANKL protein. Finally, we determined that the RANKL protein was functionally active, promoting osteoclastogenesis in vitro. Our results indicate that RANKL is a novel marker for EMT during prostate cancer progression. RANKL may function as a link between EMT, bone turnover, and prostate cancer skeletal metastasis.

  4. Epigenetic transitions in germ cell development and meiosis.

    Science.gov (United States)

    Kota, Satya K; Feil, Robert

    2010-11-16

    Germ cell development is controlled by unique gene expression programs and involves epigenetic reprogramming of histone modifications and DNA methylation. The central event is meiosis, during which homologous chromosomes pair and recombine, processes that involve histone alterations. At unpaired regions, chromatin is repressed by meiotic silencing. After meiosis, male germ cells undergo chromatin remodeling, including histone-to-protamine replacement. Male and female germ cells are also differentially marked by parental imprints, which contribute to sex determination in insects and mediate genomic imprinting in mammals. Here, we review epigenetic transitions during gametogenesis and discuss novel insights from animal and human studies.

  5. Alisertib induces cell cycle arrest and autophagy and suppresses epithelial-to-mesenchymal transition involving PI3K/Akt/mTOR and sirtuin 1-mediated signaling pathways in human pancreatic cancer cells

    Directory of Open Access Journals (Sweden)

    Wang F

    2015-01-01

    Full Text Available Feng Wang,1,2 Hai Li,3 Xiao-Gang Yan,4 Zhi-Wei Zhou,2 Zhi-Gang Yi,5 Zhi-Xu He,6 Shu-Ting Pan,7 Yin-Xue Yang,3 Zuo-Zheng Wang,1 Xueji Zhang,8 Tianxing Yang,9 Jia-Xuan Qiu,7 Shu-Feng Zhou21Department of Hepatobiliary Surgery, General Hospital, Ningxia Medical University, Yinchuan, People’s Republic of China; 2Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA; 3Department of Colorectal Surgery, General Hospital, Ningxia Medical University, 4Department of Oncological Surgery, The First People’s Hospital of Yinchuan, 5Department of General Surgery, Changqing Yangehu Hospital, Yinchuan, 6Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, 7Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, 8Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 9Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USAAbstract: Pancreatic cancer is the most aggressive cancer worldwide with poor response to current therapeutics. Alisertib (ALS, a potent and selective Aurora kinase A inhibitor, exhibits potent anticancer effects in preclinical and clinical studies; however, the effect and underlying mechanism of ALS in the pancreatic cancer treatment remain elusive. This study aimed to examine the effects of ALS on cell growth, autophagy, and epithelial-to-mesenchymal transition (EMT and to delineate the possible molecular mechanisms in human pancreatic cancer PANC-1 and BxPC-3 cells. The results showed that ALS exerted potent cell growth inhibitory, pro-autophagic, and EMT-suppressing effects in PANC-1 and BxPC-3 cells. ALS remarkably arrested PANC-1 and Bx

  6. Human mesenchymal stem cells

    DEFF Research Database (Denmark)

    Abdallah, Basem; Kassem, Moustapha

    2008-01-01

    Mesenchymal stem cells (MSC) are a group of clonogenic cells present among the bone marrow stroma and capable of multilineage differentiation into mesoderm-type cells such as osteoblasts, adipocytes and chondrocytes. Due to their ease of isolation and their differentiation potential, MSC are being...... introduced into clinical medicine in variety of applications and through different ways of administration. Here, we discuss approaches for isolation, characterization and directing differentiation of human mesenchymal stem cells (hMSC). An update of the current clinical use of the cells is also provided....

  7. Smad2/3-Regulated Expression of DLX2 Is Associated with Radiation-Induced Epithelial-Mesenchymal Transition and Radioresistance of A549 and MDA-MB-231 Human Cancer Cell Lines

    Science.gov (United States)

    Choi, Yeo-Jin; Baek, Ga-Young; Park, Hae-Ran; Jo, Sung-Kee; Jung, Uhee

    2016-01-01

    The control of radioresistance and metastatic potential of surviving cancer cells is important for improving cancer eradication by radiotheraphy. The distal-less homeobox2 (DLX2) gene encodes for a homeobox transcription factor involved in morphogenesis and its deregulation was found in human solid tumors and hematologic malignancies. Here we investigated the role of DLX2 in association with radiation-induced epithelial to mesenchymal transition (EMT) and stem cell-like properties and its regulation by Smad2/3 signaling in irradiated A549 and MDA-MB-231 human cancer cell lines. In irradiated A549 and MDA-MB-231 cells, EMT was induced as demonstrated by EMT marker expression, phosphorylation of Smad2/3, and migratory and invasive ability. Also, irradiated A549 and MDA-MB-231 cells showed increased cancer stem cells (CSCs) marker. Interestingly, DLX2 was overexpressed upon irradiation. Therefore, we examined the role of DLX2 in radiation-induced EMT and radioresistance. The overexpression of DLX2 alone induced EMT, migration and invasion, and CSC marker expression. The reduced colony-forming ability in irradiated cells was partially restored by DLX2 overexpression. On the other hand, the depletion of DLX2 using si-RNA abolished radiation-induced EMT, CSC marker expression, and phosphorylation of Smad2/3 in irradiated A549 and MDA-MB-231 cells. Also, depletion of DLX2 increased the radiation sensitivity in both cell lines. Moreover, knockdown of Smad2/3, a key activator of TGF-β1 pathway, abrogated the radiation-induced DLX2 expression, indicating that radiation-induced DLX2 expression is dependent on Smad2/3 signaling. These results demonstrated that DLX2 plays a crucial role in radioresistance, radiation-induced EMT and CSC marker expression, and the expression of DLX2 is regulated by Smad2/3 signaling in A549 and MDA-MB-231 cell lines. PMID:26799321

  8. Epithelial-mesenchymal transition process during human induced pluripotent stem cells differentiation%人诱导多能干细胞自然分化过程中EMT变化研究

    Institute of Scientific and Technical Information of China (English)

    林媛媛; 娄远蕾; 梁昌达; 谢淑佩; 谢安

    2013-01-01

    Objective To investigate whether the differentiation of induced pluripotent stem cells (iPS) is associated with the presence of epithelial-mesenchymal transition (EMT). Methods Human iPS cells were maintained in an undifferentiated state by culture in iPS cells medium in the presence of a fibroblast feeder layer, and were differentiated in EB cells medium. IPS cells dif-ferentiated for 0, 7, 14, 21, and 28 d were collected, and detected EMT associated genes and proteins by RT-RCR and western blot analysis. Results E-cadherin gene and protein were detected in undifferentiated human iPS cells, lower levels protein was ob-served following induction of differentiation. In contrast, N-cadherin gene and protein were absent in undifferentiated human iPS cells and subsequently detected following differentiation of the cells. Undifferentiated human iPS cells lacked transcripts encoding Slug and exhibited low levels of Snail transcripts. Upon differentiation both Snail and Slug transcripts were increased. Conclusion EMT event occurs during Human iPS cells differentiation.%目的:研究人诱导多能干细胞(induced pluripotent stem cells,iPS)自然分化过程中上皮-间质转化(epithelial-mes-enchymal transition,EMT)的变化过程。方法按常规方法iPS细胞接种于单层饲养层细胞上培养并传代,通过EB的方式令iPS细胞自然分化,收集未分化的iPS细胞以及分化7d、14d、21d和28d的细胞,采用RT-PCR和Western Blot方法检测EMT相关基因及蛋白的表达水平。结果 RT-PCR和Western Blot结果表明,未分化人iPS细胞可见E-cadherin基因和蛋白的表达,分化后E-cadherin的表达明显减少,而N-cadherin基因和蛋白在细胞未分化时均未见表达,发生分化后表达明显上调;在人iPS细胞分化过程中,Snail和Slug转录因子以及蛋白都明显发生上调。结论人iPS细胞自然分化与EMT,即E-cadherin和N-cadherin转化有关。

  9. Human Rights and Transitional Societies: Contemporary Challenges

    DEFF Research Database (Denmark)

    Hansen, Thomas Obel

    2008-01-01

    societies are said to have an obligation to apply criminal justice in dealing with such past violations. In Rwanda, the transitional government decided to prosecute the perpetrators of the 1994 genocide. As a result of widespread participation in the genocide and a devastated legal sector, difficulties...... in respecting the rights of the accused arose. A group of paralegals known as the "Corps of Judicial Defenders" was thus relied upon as to provide legal assistance for genocide suspects, but also for civil parties. This paper describes the work of these paralegals relating to the transitional trials, and, more...... generally, asserts how Judicial Defenders may have contributed to justice in other ways in post conflict Rwanda. The author argues that an efficient transitional justice policy must take sufficiently into account the context of the society in question, and aim at establishing linkages between justice...

  10. Managing Economic Transition. Dimensions of Human Resource Development in Hungary.

    Science.gov (United States)

    Benedek, Andras; Klekner, Peter

    1997-01-01

    Outlines the state of economic transition in Hungary, the status of human resource development, economic and legal reforms, and the social partnership (education, business, government) in vocational training. (SK)

  11. Response gene to complement-32 enhances metastatic phenotype by mediating transforming growth factor beta-induced epithelial-mesenchymal transition in human pancreatic cancer cell line BxPC-3

    Directory of Open Access Journals (Sweden)

    Zhu Liang

    2012-03-01

    Full Text Available Abstract Background Response gene to complement-32 (RGC-32 is comprehensively expressed in many kinds of tissues and has been reported to be expressed abnormally in different kinds of human tumors. However, the role of RGC-32 in cancer remains controversial and no reports have described the effect of RGC-32 in pancreatic cancer. The present study investigated the expression of RGC-32 in pancreatic cancer tissues and explored the role of RGC-32 in transforming growth factor-beta (TGF-β-induced epithelial-mesenchymal transition (EMT in human pancreatic cancer cell line BxPC-3. Methods Immunohistochemical staining of RGC-32 and E-cadherin was performed on specimens from 42 patients with pancreatic cancer, 12 with chronic pancreatitis and 8 with normal pancreas. To evaluate the role of RGC-32 in TGF-β-induced EMT in pancreatic cancer cells, BxPC-3 cells were treated with TGF-β1, and RGC-32 siRNA silencing and gene overexpression were performed as well. The mRNA expression and protein expression of RGC-32 and EMT markers such E-cadherin and vimentin were determined by quantitative reverse transcription-PCR (qRT-PCR and western blot respectively. Finally, migration ability of BxPC-3 cells treated with TGF-β and RGC-32 siRNA transfection was examined by transwell cell migration assay. Results We found stronger expression of RGC-32 and higher abnormal expression rate of E-cadherin in pancreatic cancer tissues than those in chronic pancreatitis tissues and normal pancreatic tissues. Immunohistochemical analysis revealed that both RGC-32 positive expression and E-cadherin abnormal expression in pancreatic cancer were correlated with lymph node metastasis and TNM staging. In addition, a significant and positive correlation was found between positive expression of RGC-32 and abnormal expression of E-cadherin. Furthermore, in vitro, we found sustained TGF-β stimuli induced EMT and up-regulated RGC-32 expression in BxPC-3 cells. By means of si

  12. Cell encoding recombinant human erythropoietin

    Energy Technology Data Exchange (ETDEWEB)

    Beck, A.K.; Withy, R.M.; Zabrecky, J.R.; Masiello, N.C.

    1990-09-04

    This patent describes a C127 cell transformed with a recombinant DNA vector. It comprises: a DNA sequence encoding human erythropoietin, the transformed cell being capable of producing N-linked and O-linked glycosylated human erythropoietin.

  13. Transitional justice as social control: political transitions, human rights norms and the reclassification of the past.

    Science.gov (United States)

    Dudai, Ron

    2017-09-12

    This article offers an interpretation of transitional justice policies - the efforts of post-conflict and post-dictatorship societies to address the legacy of past abuses - as a form of social control. While transitional justice is commonly conceptualized as responding to a core problem of impunity, this article argues that such formulation is too narrow and leads to lack of coherence in the analysis of the diverse array of transitional mechanisms, which include among others trials, truth commissions, reparations for victims and apologies. Building on the work of Stanley Cohen, the article contends that the core transitional problem is the denial of human rights violations, and consequently that the common purpose of all transitional justice mechanisms is to reclassify the past: redefining as deviant some acts and individuals which prior to the transition were considered 'normal'. The article identifies and analyses three themes in the application of a social control framework to transitional justice: (1) truth, memory and retroactive social control, pertains to the way truth-seeking transitional justice mechanisms reclassify past events by engaging in social control of and through memory; (2) censure, celebration and transitional social control refers to the reclassification of categories of individuals through expressions of both social disapproval and praise; and (3) civil society and social control from below concerns the role of social movements, organizations and groups as informal agents of social control during transitions. The concluding section recaps and briefly explores the concept of 'good moral panic' in the context of political transitions. While the concept of social control tends to have negative connotations for critical sociologists, this work suggests that efforts to categorize, punish and disapprove certain behaviours as deviant may not only be viewed as supporting a conservative status-quo, but also as promoting fledging human rights norms.

  14. Epithelial-mesenchymal transition, a novel target of sulforaphane via COX-2/MMP2, 9/Snail, ZEB1 and miR-200c/ZEB1 pathways in human bladder cancer cells.

    Science.gov (United States)

    Shan, Yujuan; Zhang, Lanwei; Bao, Yongping; Li, Baolong; He, Canxia; Gao, Mingming; Feng, Xue; Xu, Weili; Zhang, Xiaohong; Wang, Shuran

    2013-06-01

    Metastasis and recurrence of bladder cancer are the main reasons for its poor prognosis and high mortality rates. Because of its biological activity and high metabolic accumulation in urine, sulforaphane, a phytochemical exclusively occurring in cruciferous vegetables, has a powerful and specific potential for preventing bladder cancer. In this paper, sulforaphane is shown to significantly suppress a variety of biochemical pathways including the attachment, invasion, migration and chemotaxis motion in malignant transitional bladder cancer T24 cells. Transfection with cyclooxygenase-2 (COX-2) overexpression plasmid largely abolished inhibition of MMP2/9 expression as well as cell invasive capability by sulforaphane. Moreover, sulforaphane inhibited the epithelial-to-mesenchymal transition (EMT) process which underlies tumor cell invasion and migration mediated by E-cadherin induction through reducing transcriptional repressors, such as ZEB1 and Snail. Under conditions of over-expression of COX-2 and/or MMP2/9, sulforaphane was still able to induce E-cadherin or reduce Snail/ZEB1 expression, suggesting that additional pathways might be involved. Further studies indicated that miR-200c played a role in the regulation of E-cadherin via the ZEB1 repressor but not by the Snail repressor. In conclusion, the EMT and two recognized signaling pathways (COX-2/MMP2,9/ ZEB1, Snail and miR-200c/ZEB1) are all targets for sulforaphane. This study indicated that sulforaphane may possess therapeutic potential in preventing recurrence of human bladder cancer. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. Role of tumor-associated macrophages in epithelial-mesenchymal transition of human hepatocellular carcinoma cells%肿瘤相关巨噬细胞在肝癌上皮细胞间质转型中的作用

    Institute of Scientific and Technical Information of China (English)

    王皓; 李霞; 王超; 李国盛; 郭春; 朱法良; 张利宁; 石永玉

    2014-01-01

    Objective To explore the effect and mechanism of tumor-associated macrophages on human hepatocellular carcinoma ( HCC) cells.Methods The HCC cells were cocultured with macrophages from PMA-treated THP-1 cells and cell migration was detected by transwell migration test and wound-healing assay.The expressions of E-cadherin and N-cadherin were measured by RT-PCR.Results The ability of cell migration was significantly increased after being cocultured with mocrophages from PMA-treated THP-1 cells.A transition from epithelial morphology to mesenchymal morphology was observed.The expression of E-cadherin decreased and the expression of N-cadherin increased.Conclu-sion Our findings suggest that tumor-associated macrophages may promote the cell migration through epithelial-mesen-chymal transition.%目的:检测肿瘤相关巨噬细胞对肝癌细胞迁移能力的作用及机制。方法将肝癌细胞与THP-1细胞来源的巨噬细胞共培养,利用Transwell细胞迁移实验与细胞划痕实验检测肝癌细胞迁移能力的变化情况,观察肝癌细胞形态变化,并用RT-PCR检测上皮细胞间充质转化相关分子E-cadherin与N-cadherin的变化。结果与THP-1细胞来源的巨噬细胞共培养后,肝癌细胞的迁移能力明显增强,由上皮细胞形态向间质细胞形态转变,E-cadherin表达降低,而N-cadherin表达则升高。结论在肝癌中,肿瘤相关巨噬细胞可能通过EMT增强肝癌细胞的迁移能力。

  16. The ectopic expression of Snail in MDBK cells does not induce epithelial-mesenchymal transition

    OpenAIRE

    IZAWA, GENYA; Kobayashi, Wakako; Haraguchi, Misako; Sudo, Akiharu; Ozawa, Masayuki

    2015-01-01

    Epithelial-mesenchymal transition (EMT), a key process in the tumor metastatic cascade, is characterized by the loss of cell-cell junctions and cell polarity, as well as by the acquisition of migratory and invasive properties. However, the precise molecular events that initiate this complex EMT process are poorly understood. Snail expression induces EMT in Madin-Darby canine kidney (MDCK) cells and the human epidermoid carcinoma cell line, A431. Snail is a zinc finger transcription factor and...

  17. Animal model of naturally occurring bladder cancer: Characterization of four new canine transitional cell carcinoma cell lines

    OpenAIRE

    Rathore, Kusum; Cekanova, Maria

    2014-01-01

    Background Development and further characterization of animal models for human cancers is important for the improvement of cancer detection and therapy. Canine bladder cancer closely resembles human bladder cancer in many aspects. In this study, we isolated and characterized four primary transitional cell carcinoma (K9TCC) cell lines to be used for future in vitro validation of novel therapeutic agents for bladder cancer. Methods Four K9TCC cell lines were established from naturally-occurring...

  18. Assessing global transitions in human development and colorectal cancer incidence.

    Science.gov (United States)

    Fidler, Miranda M; Bray, Freddie; Vaccarella, Salvatore; Soerjomataram, Isabelle

    2017-06-15

    Colorectal cancer incidence has paralleled increases in human development across most countries. Yet, marked decreases in incidence are now observed in countries that have attained very high human development. Thus, in this study, we explored the relationship between human development and colorectal cancer incidence, and in particular assessed whether national transitions to very high human development are linked to temporal patterns in colorectal cancer incidence. For these analyses, we utilized the Human Development Index (HDI) and annual incidence data from regional and national cancer registries. Truncated (30-74 years) age-standardized incidence rates were calculated. Yearly incidence rate ratios and HDI ratios, before and after transitioning to very high human development, were also estimated. Among the 29 countries investigated, colorectal cancer incidence was observed to decrease after reaching the very high human development threshold for 12 countries; decreases were also observed in a further five countries, but the age-standardized incidence rates remained higher than that observed at the threshold. Such declines or stabilizations are likely due to colorectal cancer screening in some populations, as well as varying levels of exposure to protective factors. In summary, it appears that there is a threshold at which human development predicts a stabilization or decline in colorectal cancer incidence, though this pattern was not observed for all countries assessed. Future cancer planning must consider the increasing colorectal cancer burden expected in countries transitioning towards higher levels of human development, as well as possible declines in incidence among countries reaching the highest development level. © 2017 UICC.

  19. Variations in Humanized and Defined Culture Conditions Supporting Derivation of New Human Embryonic Stem Cell Lines

    DEFF Research Database (Denmark)

    Fletcher, Judy M; Ferrier, Patricia M; Gardner, John O

    2006-01-01

    and the potential to form cells representative of all three germinal lineages in vitro and in vivo, when transitioned off of feeders onto Laminin or Matrigel. Our study thus demonstrates the capacity to integrate derivation strategies eliminating a requirement for animal immune compliment and serum products......, with a transitional requirement for human feeder cells. This represents another sequential step in the generation of therapeutic grade stem cells with reduced risk of zoonotic pathogen transmission....

  20. Genome engineering in human cells.

    Science.gov (United States)

    Song, Minjung; Kim, Young-Hoon; Kim, Jin-Soo; Kim, Hyongbum

    2014-01-01

    Genome editing in human cells is of great value in research, medicine, and biotechnology. Programmable nucleases including zinc-finger nucleases, transcription activator-like effector nucleases, and RNA-guided engineered nucleases recognize a specific target sequence and make a double-strand break at that site, which can result in gene disruption, gene insertion, gene correction, or chromosomal rearrangements. The target sequence complexities of these programmable nucleases are higher than 3.2 mega base pairs, the size of the haploid human genome. Here, we briefly introduce the structure of the human genome and the characteristics of each programmable nuclease, and review their applications in human cells including pluripotent stem cells. In addition, we discuss various delivery methods for nucleases, programmable nickases, and enrichment of gene-edited human cells, all of which facilitate efficient and precise genome editing in human cells.

  1. BC Transit Fuel Cell Bus Project: Evaluation Results Report

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, L.; Post, M.

    2014-02-01

    This report evaluates a fuel cell electric bus demonstration led by British Columbia Transit (BC Transit) in Whistler, Canada. BC Transit is collaborating with the California Air Resources Board and the U.S. Department of Energy's National Renewable Energy Laboratory to evaluate the buses in revenue service. This evaluation report covers two years of revenue service data on the buses from April 2011 through March 2013.

  2. Human embryonic stem cells: preclinical perspectives

    Directory of Open Access Journals (Sweden)

    Sarda Kanchan

    2008-01-01

    Full Text Available Abstract Human embryonic stem cells (hESCs have been extensively discussed in public and scientific communities for their potential in treating diseases and injuries. However, not much has been achieved in turning them into safe therapeutic agents. The hurdles in transforming hESCs to therapies start right with the way these cells are derived and maintained in the laboratory, and goes up-to clinical complications related to need for patient specific cell lines, gender specific aspects, age of the cells, and several post transplantation uncertainties. The different types of cells derived through directed differentiation of hESC and used successfully in animal disease and injury models are described briefly. This review gives a brief outlook on the present and the future of hESC based therapies, and talks about the technological advances required for a safe transition from laboratory to clinic.

  3. Multi-walled carbon nanotubes directly induce epithelial-mesenchymal transition in human bronchial epithelial cells via the TGF-β-mediated Akt/GSK-3β/SNAIL-1 signalling pathway.

    Science.gov (United States)

    Polimeni, Manuela; Gulino, Giulia Rossana; Gazzano, Elena; Kopecka, Joanna; Marucco, Arianna; Fenoglio, Ivana; Cesano, Federico; Campagnolo, Luisa; Magrini, Andrea; Pietroiusti, Antonio; Ghigo, Dario; Aldieri, Elisabetta

    2016-06-01

    Multi-walled carbon nanotubes (MWCNT) are currently under intense toxicological investigation due to concern on their potential health effects. Current in vitro and in vivo data indicate that MWCNT exposure is strongly associated with lung toxicity (inflammation, fibrosis, granuloma, cancer and airway injury) and their effects might be comparable to asbestos-induced carcinogenesis. Although fibrosis is a multi-origin disease, epithelial-mesenchymal transition (EMT) is recently recognized as an important pathway in cell transformation. It is known that MWCNT exposure induces EMT through the activation of the TGF-β/Smad signalling pathway thus promoting pulmonary fibrosis, but the molecular mechanisms involved are not fully understood. In the present work we propose a new mechanism involving a TGF-β-mediated signalling pathway. Human bronchial epithelial cells were incubated with two different MWCNT samples at various concentrations for up to 96 h and several markers of EMT were investigated. Quantitative real time PCR, western blot, immunofluorescent staining and gelatin zymographies were performed to detect the marker protein alterations. ELISA was performed to evaluate TGF-β production. Experiments with neutralizing anti-TGF-β antibody, specific inhibitors of GSK-3β and Akt and siRNA were carried out in order to confirm their involvement in MWCNT-induced EMT. In vivo experiments of pharyngeal aspiration in C57BL/6 mice were also performed. Data were analyzed by a one-way ANOVA with Tukey's post-hoc test. Fully characterized MWCNT (mean length EMT in an in vitro human model (BEAS-2B cells) after long-term incubation at sub-cytotoxic concentrations. MWCNT stimulate TGF-β secretion, Akt activation and GSK-3β inhibition, which induces nuclear accumulation of SNAIL-1 and its transcriptional activity, thus contributing to switch on the EMT program. Moreover, a significant increment of nuclear β-catenin - due to E-cadherin repression and following

  4. Expression of Pol(t) in tissues and cell lines of transitional cell carcinoma

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective:To explore the expression of DNA polymerase iota in transitional cell carcinoma cells and tissues; Methods: RT-PCR was applie to detect the expression of polymerase iota in BIU87 and T24 cells, then the expression of polymerase iota was also detected in the same way in transitional cell carcinoma which was derived from clinical bladder carcinoma and renal pelvic carcinoma. Results: The expression of Polt was low in bladder normal membrana mucosa but significantly elevated in transitional cell carcinoma cells. Compared with the expression of polymerase iota in bladder normal mucous membranes, the expression of polymerase iota was significantly increased in transitional cell carcinoma tissue (P<0.01)and associated with the grade of transitional cell carcinoma. Conclusion: The significantly increased expression of polymerase iota may be associated with the generation and development of transitional cell carcinoma, even with its high heterogenicity.

  5. Regulation of B Cell to Plasma Cell Transition within the Follicular B Cell Response.

    Science.gov (United States)

    Nera, K-P; Kyläniemi, M K; Lassila, O

    2015-09-01

    Persistent humoral immunity depends on the follicular B cell response and on the generation of somatically mutated high-affinity plasma cells and memory B cells. Upon activation by an antigen, cognately activated follicular B cells and follicular T helper (TFH ) cells initiate germinal centre (GC) reaction during which high-affinity effector cells are generated. The differentiation of activated follicular B cells into plasma cells and memory B cells is guided by complex selection events, both at the cellular and molecular level. The transition of B cell into a plasma cell during the GC response involves alterations in the microenvironment and developmental state of the cell, which are guided by cell-extrinsic signals. The developmental cell fate decisions in response to these signals are coordinated by cell-intrinsic gene regulatory network functioning at epigenetic, transcriptional and post-transcriptional levels.

  6. Noise-induced transition in human reaction times

    Science.gov (United States)

    Medina, José M.; Díaz, José A.

    2016-09-01

    The human reaction/response time can be defined as the time elapsed from the onset of stimulus presentation until a response occurs in many sensory and cognitive processes. A reaction time model based on Piéron’s law is investigated. The model shows a noise-induced transition in the moments of reaction time distributions due to the presence of strong additive noise. The model also demonstrates that reaction times do not follow fluctuation scaling between the mean and the variance but follow a generalized version between the skewness and the kurtosis. The results indicate that noise-induced transitions in the moments govern fluctuations in sensory-motor transformations and open an insight into the macroscopic effects of noise in human perception and action. The conditions that lead to extreme reaction times are discussed based on the transfer of information in neurons.

  7. Spheroid cells derived from human colon cancer cells exhibit epithelial to mesenchymal transition characteristics%悬浮培养结肠癌细胞球具有上皮间质转化特征

    Institute of Scientific and Technical Information of China (English)

    冷政伟; 尹碧辉; 李勇; 谭俊; 岳中屹; 陈锦皇; 奚海林; 陶凯雄; 夏清华

    2013-01-01

    目的 无血清悬浮培养人结肠癌DLD-1细胞形成细胞球,检测其肿瘤干细胞标志表达变化,并探讨球细胞发生上皮间质转化与肿瘤干细胞及肿瘤恶性行为之间的联系.方法 在添加生长因子的无血清培养基中培养DLD-1细胞形成球细胞,运用Transwell实验检测细胞的迁移、侵袭能力变化;裸鼠皮下成瘤和软琼脂克隆形成实验分别检测体内外成瘤能力变化;流式细胞术检测细胞中CD133表达变化;实时定量聚合酶链反应(Real-time PCR),Western blot检测CD133、CD166、富含亮氨酸重复序列G-蛋白偶联受体5(Lgr5)及上皮间质转化关键基因表达变化.结果 球细胞中CD133+细胞明显增加[(0.90±0.03)%比(2.70±0.35)%,P <0.05];高表达肿瘤干细胞标志CD133、CD166、Lgr5和间质分子波形蛋白(Vimentin)及Snail而低表达上皮细胞标志ZO-1(P<0.05).球细胞的转移、侵袭、体内外成瘤能力明显增加(P<0.05).结论 虽然黏钙蛋白(E-cadherin)表达明显升高,但是结合体内外实验结果,我们认为:E-cadherin可能不是介导细胞恶性行为的独立因素,悬浮培养形成的结肠癌球细胞已发生上皮间质转化,表现出肿瘤干细胞的特征.%Objective To investigate the similarities between cancer stem cells and epithelial mesenchymal transition (EMT) cells in spheroid cells.Methods Generate cancer spheroid cells from colon cancer cell line DLD-1 in serum free medium (SFM) and to investigate the expression of colon cancer stem cells markers such as CD133,CD166,leucine-rich-repeat-containing G-protein-coupled receptor 5 (Lgr5)and the numbers of CD133 + cells.In addition,the capacity of migration,invasion and tumorigenicity in vivo and in vitro were confirmed by transwell assay,colony formation assay and mouse xenograft model respectively.Moreover,the expression of EMT-related genes was detected by real-time quantitative polymerase chain reaction (Real-time PCR) and Western

  8. Dynamic behaviour of human neuroepithelial cells in the developing forebrain

    Science.gov (United States)

    Subramanian, Lakshmi; Bershteyn, Marina; Paredes, Mercedes F.; Kriegstein, Arnold R.

    2017-01-01

    To understand how diverse progenitor cells contribute to human neocortex development, we examined forebrain progenitor behaviour using timelapse imaging. Here we find that cell cycle dynamics of human neuroepithelial (NE) cells differ from radial glial (RG) cells in both primary tissue and in stem cell-derived organoids. NE cells undergoing proliferative, symmetric divisions retract their basal processes, and both daughter cells regrow a new process following cytokinesis. The mitotic retraction of the basal process is recapitulated by NE cells in cerebral organoids generated from human-induced pluripotent stem cells. In contrast, RG cells undergoing vertical cleavage retain their basal fibres throughout mitosis, both in primary tissue and in older organoids. Our findings highlight developmentally regulated changes in mitotic behaviour that may relate to the role of RG cells to provide a stable scaffold for neuronal migration, and suggest that the transition in mitotic dynamics can be studied in organoid models. PMID:28139695

  9. In Situ Observation of Cell-to-Dendrite Transition

    Institute of Scientific and Technical Information of China (English)

    PAN Xiu-Hong; HONG Yong; JIN Wei-Qing

    2005-01-01

    @@ The cell-to-dendrite transition of succinonitrile melt suspended on a loop-shaped Pt heater is observed in real time by a differential interference microscope coupled with Schlieren technique. The transition is divided into two parts: a dendrite coalition process and a subsequent dendrite elimination process. Firstly the dendrites from the same cell are united into a single dendrite. Secondly the competitive growth of dendrites from different cells leads to the elimination of dendrites. The two processes can be understood when involving crystallographic orientation. In addition, the tip velocity and primary spacing of a cell/dendrite are also measured. It turns out that the primary spacing has a significant jump, whereas the growth velocity has no abrupt change during the cell-to-dendrite transition.

  10. Diffusion inside living human cells

    DEFF Research Database (Denmark)

    Leijnse, N.; Jeon, J. -H.; Loft, Steffen

    2012-01-01

    Naturally occurring lipid granules diffuse in the cytoplasm and can be used as tracers to map out the viscoelastic landscape inside living cells. Using optical trapping and single particle tracking we found that lipid granules exhibit anomalous diffusion inside human umbilical vein endothelial...... cells. For these cells the exact diffusional pattern of a particular granule depends on the physiological state of the cell and on the localization of the granule within the cytoplasm. Granules located close to the actin rich periphery of the cell move less than those located towards to the center...... of the cell or within the nucleus. Also, granules in cells which are stressed by intense laser illumination or which have attached to a surface for a long period of time move in a more restricted fashion than those within healthy cells. For granules diffusing in healthy cells, in regions away from the cell...

  11. Pancreatic stellate cells promote epithelial-mesenchymal transition in pancreatic cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Kikuta, Kazuhiro [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Masamune, Atsushi, E-mail: amasamune@med.tohoku.ac.jp [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Watanabe, Takashi; Ariga, Hiroyuki; Itoh, Hiromichi; Hamada, Shin; Satoh, Kennichi [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Egawa, Shinichi; Unno, Michiaki [Department of Hepatobiliary-Pancreatic Surgery, Tohoku University Graduate School of Medicine, Sendai (Japan); Shimosegawa, Tooru [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan)

    2010-12-17

    Research highlights: {yields} Recent studies have shown that pancreatic stellate cells (PSCs) promote the progression of pancreatic cancer. {yields} Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and scattered, fibroblast-like appearance. {yields} PSCs decreased the expression of epithelial markers but increased that of mesenchymal markers, along with increased migration. {yields} This study suggests epithelial-mesenchymal transition as a novel mechanism by which PSCs contribute to the aggressive behavior of pancreatic cancer cells. -- Abstract: The interaction between pancreatic cancer cells and pancreatic stellate cells (PSCs), a major profibrogenic cell type in the pancreas, is receiving increasing attention. There is accumulating evidence that PSCs promote the progression of pancreatic cancer by increasing cancer cell proliferation and invasion as well as by protecting them from radiation- and gemcitabine-induced apoptosis. Because epithelial-mesenchymal transition (EMT) plays a critical role in the progression of pancreatic cancer, we hypothesized that PSCs promote EMT in pancreatic cancer cells. Panc-1 and SUIT-2 pancreatic cancer cells were indirectly co-cultured with human PSCs isolated from patients undergoing operation for pancreatic cancer. The expression of epithelial and mesenchymal markers was examined by real-time PCR and immunofluorescent staining. The migration of pancreatic cancer cells was examined by scratch and two-chamber assays. Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and a scattered, fibroblast-like appearance. The expression of E-cadherin, cytokeratin 19, and membrane-associated {beta}-catenin was decreased, whereas vimentin and Snail (Snai-1) expression was increased more in cancer cells co-cultured with PSCs than in mono-cultured cells. The migration of pancreatic cancer cells was increased by co-culture with PSCs. The PSC-induced decrease of E-cadherin expression was not altered

  12. Fuel cell transit bus development & commercialization programs at Gerogetown University

    Energy Technology Data Exchange (ETDEWEB)

    Wimmer, R.; Larkins, J.; Romano, S. [Georgetown Univ., Washington, DC (United States)

    1996-12-31

    Fourteen years ago, Georgetown University (GU) perceived the need for a clean, efficient power systems for transportation that could operate on non-petroleum based fuels. The transit bus application was selected to begin system development. GU recognized the range and recharge constraints of a pure battery powered transit bus. A Fuel Cell power system would circumvent these limitations and, with an on board reformer, accommodate liquid fuel for rapid refueling. Feasibility studies for Fuel Cell power systems for transit buses were conducted with the Los Alamos National Laboratory in 1983. Successful results of this investigation resulted in the DOT/DOE Fuel Cell transit bus development program. The first task was to prove that small Fuel Cell power plants were possible. This was achieved with the Phase I development of two 25 kW Phosphoric Acid Fuel Cell (PAFC) brassboard systems. A liquid cooled version was selected for the Phase II activity in which three 30-foot Fuel Cell powered Test Bed Buses (TBBs) were fabricated. The first of these TBBs was delivered in the spring of 1994. All three of these development vehicles are now in Phase III of the program to conduct testing and evaluation, is conducting operational testing of the buses. The test will involve two fuel cell-operated buses; one with a proton exchange fuel cell and the other with a phosphoric acid fuel cell.

  13. Danusertib, a potent pan-Aurora kinase and ABL kinase inhibitor, induces cell cycle arrest and programmed cell death and inhibits epithelial to mesenchymal transition involving the PI3K/Akt/mTOR-mediated signaling pathway in human gastric cancer AGS and NCI-N78 cells

    Directory of Open Access Journals (Sweden)

    Yuan CX

    2015-03-01

    Full Text Available Chun-Xiu Yuan,1,2 Zhi-Wei Zhou,2,3 Yin-Xue Yang,4 Zhi-Xu He,3 Xueji Zhang,5 Dong Wang,6 Tianxing Yang,7 Si-Yuan Pan,8 Xiao-Wu Chen,9 Shu-Feng Zhou2 1Department of Oncology, General Hospital, Ningxia Medical University, Yinchuan, People’s Republic of China; 2Department of Pharmaceutical Science, College of Pharmacy, University of South Florida, Tampa, FL, USA; 3Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, 4Department of Colorectal Surgery, General Hospital, Ningxia Medical University, Yinchuan, 5Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, 6Cancer Center, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, People’s Republic of China; 7Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA; 8Department of Pharmacology, School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing, 9Department of General Surgery, The First People’s Hospital of Shunde, Southern Medical University, Shunde, People’s Republic of China Abstract: Gastric cancer is the second leading cause of cancer-related death worldwide, with a poor response to current chemotherapy. Danusertib is a pan-inhibitor of the Aurora kinases and a third-generation Bcr-Abl tyrosine kinase inhibitor with potent anticancer effects, but its antitumor effect and underlying mechanisms in the treatment of human gastric cancer are unknown. This study aimed to investigate the effects of danusertib on cell growth, apoptosis, autophagy, and epithelial to mesenchymal transition and the molecular mechanisms involved in human gastric cancer AGS and NCI-N78 cells. The results showed that danusertib had potent growth-inhibitory, apoptosis-inducing, and

  14. A theoretical model of phase transitions in the human brain.

    Science.gov (United States)

    Jirsa, V K; Friedrich, R; Haken, H; Kelso, J A

    1994-01-01

    An experiment using a multisensor SQUID (superconducting quantum interference device) array was performed by Kelso and colleagues (1992) which combined information from three different sources: perception, motor response, and brain signals. When an acoustic stimulus frequency is changed systematically, a spontaneous transition in coordination occurs at a critical frequency in both motor behavior and brain signals. Qualitatively analogous transitions are known for physical and biological systems such as changes in the coordination of human hand movements (Kelso 1981, 1984). In this paper we develop a theoretical model based on methods from the interdisciplinary field of synergetics (Haken 1983, 1987) and nonlinear oscillator theory that reproduces the main experimental features very well and suggests a formulation of a fundamental biophysical coupling.

  15. Transitional cell carcinoma express vitamin D receptors

    DEFF Research Database (Denmark)

    Hermann, G G; Andersen, C B

    1997-01-01

    .05). Similarly, also tumor grade appeared to be related to the number of cells expressing the receptor. Normal urothlium also expressed VDR but only with low intensity. Our study shows that TCC cells possess the VDR receptor which may make them capable to respond to stimulation with vitamin D, but functional...

  16. Human stromal (mesenchymal) stem cells

    DEFF Research Database (Denmark)

    Aldahmash, Abdullah; Zaher, Walid; Al-Nbaheen, May

    2012-01-01

    Human stromal (mesenchymal) stem cells (hMSC) represent a group of non-hematopoietic stem cells present in the bone marrow stroma and the stroma of other organs including subcutaneous adipose tissue, placenta, and muscles. They exhibit the characteristics of somatic stem cells of self-renewal and......Human stromal (mesenchymal) stem cells (hMSC) represent a group of non-hematopoietic stem cells present in the bone marrow stroma and the stroma of other organs including subcutaneous adipose tissue, placenta, and muscles. They exhibit the characteristics of somatic stem cells of self...... of clinical applications, e.g., non-healing bone fractures and defects and also non-skeletal degenerative diseases like heart failure. Currently, the numbers of clinical trials that employ MSC are increasing. However, several biological and biotechnological challenges need to be overcome to benefit from...

  17. BC Transit Fuel Cell Bus Project Evaluation Results: Second Report

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, L.; Post, M.

    2014-09-01

    Second report evaluating a fuel cell electric bus (FCEB) demonstration led by British Columbia Transit (BC Transit) in Whistler, Canada. BC Transit is collaborating with the California Air Resources Board and the U.S. Department of Energy's National Renewable Energy Laboratory to evaluate the buses in revenue service. NREL published its first report on the demonstration in February 2014. This report is an update to the previous report; it covers 3 full years of revenue service data on the buses from April 2011 through March 2014 and focuses on the final experiences and lessons learned.

  18. Human factors issues for resolving adverse effects of human work underload and workload transitions in complex human-machine systems

    Energy Technology Data Exchange (ETDEWEB)

    Ryan, T.G.

    1995-10-01

    A workshop was conducted whose specific purpose was to build on earlier work of the United States National Research Council, United States Federal government agencies, and the larger human factors community to: (1) clarify human factors issues pertaining to degraded performance in advanced human-machine systems (e.g., nuclear production, transportation, aerospace) due to human work underload and workload transition, and (2) develop strategies for resolving these issues. Recent history demonstrates that: (1) humans often react adversely to their diminishing roles in advanced human-machine systems, and therefore (2) new allocation models and strategies are required if humans are to be willing and able to assume diminishing and shifting roles assigned to them in these systems, and are to accept new technologies making up these systems. Problems associated with theses diminishing and shifting human roles are characterized as work underload and workload transitions. The workshop affirmed that: (1) work underload and workload transition are issues that will have to be addressed by designers of advanced human-machine systems, especially those relying on automation, if cost, performance, safety, and operator acceptability are to be optimized, (2) human machine allocation models, standards, and guidelines which go beyond simple capability approaches will be needed to preclude or seriously diminish the work underload and workload transition problems, and (3) the 16 workload definition, measurement, situational awareness, and trust issues identified during the workshop, need resolution if these models, standards, and guidelines are to be achieved.

  19. Connecticut Transit (CTTRANSIT) Fuel Cell Transit Bus: Third Evaluation Report and Appendices

    Energy Technology Data Exchange (ETDEWEB)

    Chandler, K.; Eudy, L.

    2010-01-01

    This report describes operations at Connecticut Transit (CTTRANSIT) in Hartford for one prototype fuel cell bus and three new diesel buses operating from the same location. The prototype fuel cell bus was manufactured by Van Hool and ISE Corp. and features an electric hybrid drive system with a UTC Power PureMotion 120 Fuel Cell Power System and ZEBRA batteries for energy storage. The fuel cell bus started operation in April 2007, and evaluation results through October 2009 are provided in this report.

  20. National Fuel Cell Bus Program: Accelerated Testing Evaluation Report and Appendices, Alameda-Contra Costa Transit District (AC Transit)

    Energy Technology Data Exchange (ETDEWEB)

    Chandler, K.; Eudy, L.

    2009-01-01

    This is an evaluation of hydrogen fuel cell transit buses operating at AC Transit in revenue service since March 20, 2006 compared to similar diesel buses operating from the same depot. This evaluation report includes results from November 2007 through October 2008. Evaluation results include implementation experience, fueling station operation, fuel cell bus operations at Golden Gate Transit, and evaluation results at AC Transit (bus usage, availability, fuel economy, maintenance costs, and roadcalls).

  1. Connecticut Transit (CTTRANSIT) Fuel Cell Transit Bus: Second Evaluation Report and Appendices

    Energy Technology Data Exchange (ETDEWEB)

    Chandler, K.; Eudy, L.

    2009-05-01

    This report describes operations at Connecticut Transit (CTTRANSIT) in Hartford for one prototype fuel cell bus and three new diesel buses operating from the same location. The evaluation period in this report (January 2008 through February 2009) has been chosen to coincide with a UTC Power propulsion system changeout that occurred on January 15, 2008.

  2. SunLine Transit Agency Fuel Cell Transit Bus: Fifth Evaluation Report (Report and Appendices)

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, L.; Chandler, K.

    2009-08-01

    This report describes operations at SunLine Transit Agency for a prototype fuel cell bus and five compressed natural gas (CNG) buses. This is the fifth evaluation report for this site, and it describes results and experiences from October 2008 through June 2009. These results are an addition to those provided in the previous four evaluation reports.

  3. SunLine Transit Agency Fuel Cell Transit Bus: Fourth Evaluation Report (Report and Appendices)

    Energy Technology Data Exchange (ETDEWEB)

    Chandler, K.; Eudy, L.

    2009-01-01

    This report describes operations at SunLine Transit Agency for a prototype fuel cell bus and five new compressed natural gas (CNG) buses. This is the fourth evaluation report for this site, and it describes results and experiences from April 2008 through October 2008. These results are an addition to those provided in the previous three evaluation reports.

  4. 间充质干细胞条件培养液促进肺癌细胞上皮间质转化%Human bone marrow mesenchymal stem cells promote epithelial mesenchymal transition in lung cancer cells

    Institute of Scientific and Technical Information of China (English)

    吴佳斌; 王涛; 杨伟林; 王均洁; 肖婕婓; 王儒琛; 陈振光

    2016-01-01

    BACKGROUND:The complex relationship between bone marrow mesenchymal stem cels and cancers severely limit the clinical application of mesenchymal stem cels. So it is urgent to study the role of mesenchymal stem cels in tumor growth and metastasis. OBJECTIVE:To explore the effect of human bone marrow mesenchymal stem cels on epithelial mesenchymal transition in non-smal cel lung cancer A549 and PAa cels. METHODS:The A549 and PAa cels were cultured with mesenchymal stem cel supernatant (mesenchymal stem cel conditioned medium, MSCs-CM). The celular morphology was observed under a microscope. The mRNA and protein expression of E-cadherin, N-cadherin, Vimentin, Slug, Snail, and Twist were determined by RT-PCR and western blot. Transwel and wound healing assay were used to detect the change of migration and metastatic ability. RESULTS AND CONCLUSION:Compared with the control group, the celular morphology of experimental group showed mesenchymal-like changes. In response to MSCs-CM, there was decreased E-cadherin but increased N-cadherin, Vimentin and Slug, Snail, Twist at mRNA and protein levels compared with the control group (P   目的:探讨人骨髓来源间充质干细胞对人非小细胞肺癌A549、PAa细胞上皮间质转化的影响。  方法:收集间充质干细胞上清液作为间充质干细胞条件培养液培养肺癌细胞 A549和 PAa,观察肺癌细胞形态、上皮间质转化标志物E-钙黏素、N-钙黏素、波形蛋白以及其转录因子Slug、Snail、Twist等的表达以及肺癌细胞运动迁移能力的变化。  结果与结论:①与对照组对比:加入条件培养液的肺癌细胞形态发生了间质样变化,且E-钙黏素表达降低,N-钙黏素、波形蛋白mRNA与蛋白表达水平上升,转录因子Slug、Snail、Twist mRNA表达水平也明显增加,同时细胞运动迁移能力增强。②结果表明人骨髓来源间充质干细胞可以促进非小细胞肺癌A549、PAa细胞上皮间

  5. PRIMARY TRANSITIONAL CELL CARCINOMA OF THE OVARY: A CASE REPORT

    Directory of Open Access Journals (Sweden)

    Anju

    2016-05-01

    Full Text Available A 38-year-old female presented with a history of progressively enlarging abdominal mass. Abdominal computed tomography showed a pelvic mass involving both the ovaries and omentum. CA-125 was normal. Staging surgery was performed and the histopathological diagnosis of Transitional Cell Carcinoma was made and later confirmed by immuno-histochemistry. Transitional cell carcinoma of the ovary is a rare subtype of epithelial ovarian cancer. Surgical resection is the primary therapeutic approach, and patient’s outcomes after chemotherapy are better than for other types of ovarian cancers.

  6. Metastatic transitional cell carcinoma of the tibia radiologically mimicking osteosarcoma.

    LENUS (Irish Health Repository)

    Cunningham, Laurence Patrick

    2013-01-01

    We report a case of a 73-year-old lady with transitional cell carcinoma and no evidence of metastatic disease presenting with gradual weight loss, pretibial swelling and painful weightbearing. Investigations revealed a lesion of the right tibial diaphysis. The radiological and clinical appearance was that of primary osteosarcoma. Biopsy results revealed metastatic transitional cell carcinoma of the tibia. Intramedullary nailing was performed which relieved pain on weightbearing. The patient declined radiotherapy and was started on a palliative care regimen. This case illustrates the importance of histological diagnosis in the treatment of diaphyseal lesions.

  7. An in vitro model that recapitulates the epithelial to mesenchymal transition (EMT in human breast cancer.

    Directory of Open Access Journals (Sweden)

    Elad Katz

    Full Text Available The epithelial to mesenchymal transition (EMT is a developmental program in which epithelial cells down-regulate their cell-cell junctions, acquire spindle cell morphology and exhibit cellular motility. In human breast cancer, invasion into surrounding tissue is the first step in metastatic progression. Here, we devised an in vitro model using selected cell lines, which recapitulates many features of EMT as observed in human breast cancer. By comparing the gene expression profiles of claudin-low breast cancers with the experimental model, we identified a 9-gene signature characteristic of EMT. This signature was found to distinguish a series of breast cancer cell lines that have demonstrable, classical EMT hallmarks, including loss of E-cadherin protein and acquisition of N-cadherin and vimentin expression. We subsequently developed a three-dimensional model to recapitulate the process of EMT with these cell lines. The cells maintain epithelial morphology when encapsulated in a reconstituted basement membrane, but undergo spontaneous EMT and invade into surrounding collagen in the absence of exogenous cues. Collectively, this model of EMT in vitro reveals the behaviour of breast cancer cells beyond the basement membrane breach and recapitulates the in vivo context for further investigation into EMT and drugs that may interfere with it.

  8. Human embryonic stem cells handbook

    Directory of Open Access Journals (Sweden)

    Carlo Alberto Redi

    2013-03-01

    Full Text Available After the Nobel prize in physiology or medicine was awarded jointly to Sir John Gurdon and Shinya Yamanaka for the discovery that mature cells can be reprogrammed to become pluripotent it became imperative to write down the review for a book entirely devoted to human embryonic stem cells (hES, those cells that are a urgent need for researchers, those cells that rekindle the ethical debates and finally, last but not least, those cells whose study paved the way to obtain induced pluripotent stem cells by the OSKC’s Yamanaka method (the OSKC acronim refers, for those not familiar with the topic, to the four stemness genes used to transfect somatic fibroblasts: Oct4, Sox2, Klf4 and c-Myc....

  9. Proliferation effects of sirolimus, cydosporine A and mycophenolate mofetil on human transitional cell carcinoma cells%西罗莫司、环孢素A、吗替麦考酚酯对膀胱移行细胞癌细胞增殖的影响

    Institute of Scientific and Technical Information of China (English)

    林俊; 王迎红; 张玉海; 田野

    2012-01-01

    Objective To compared the effects of three immunosuppressive agents,i.e.sirolimus (SRL),cyclosporine A (CsA) and mycophenolate mofetil (MMF),with different mechanisms of action on the in vitro growth of various tumor cell lines of human transitional cell carcinoma of bladder cell lines EJ and T24 and in vivo growth of cell line of EJ in nude mice model.Methods The effects of SRL,CsA and MMF on the proliferation of transitional cell carcinoma of bladder cell lines were examined with the method of methyl thiazolyl tetrazolium (MTT). The effects of these immunosuppressants on tumor growth and metastasis were explored in a nude mice model with human transitional cell carcinoma of bladder cell line EJ.Forty-two nude mice were divided into 7 groups to receive normal saline (control),SRL,CsA,MMF,SRL + CsA,SRL + MMF and CsA + MMF respectively ( n =6 each).Results The in vitro cell proliferation was inhibited by SRL and MMF versus the control groups. But no obvious inhibition of proliferation was observed at <1000 ng/ml in the CsA-treated group.In the in vivo nude mice mode,the tumor volume of SRL,CsA group were lower than that in control group ( (441 ± 231 ),(463 ± 110)vs (1032 ± 382 )mm3,both P <0.05).In the in vivo nude mice mode of EJ treated by SRL,CsA,SRL + CsA,SRL + MMF and CsA + MMF,tumor volume at Day 23 was the lowest in the SRL + CsA group ( (191 ±92) vs (1032 ±382)mm3,P < 0.05 ). There was an inhibition of 75.26% in SRL + CsA group versus the control groups.Conclusions SRL and MMF demonstrate dose-dependent antiproliferative effects in human transitional cell carcinoma of bladder cell both in vitro and in vivo.CsA can inhibit the growth of human transitional cell carcinoma of bladder cell lines EJ cells in vivo.%目的 探讨3种免疫抑制剂西罗莫司(SRL)、环孢素A(CsA)、吗替麦考酚酯(MMF)对膀胱移行细胞癌细胞增殖的影响.方法 体外实验,采用四甲基偶氮唑盐(MTT)比色法测定细胞吸光度(A)值,观察不

  10. Transition of mesenchymal stem/stromal cells to endothelial cells

    NARCIS (Netherlands)

    M. Crisan (Mihaela)

    2013-01-01

    textabstractMesenchymal stem/stromal cells (MSCs) are heterogeneous. A fraction of these cells constitute multipotent cells that can self-renew and mainly give rise to mesodermal lineage cells such as adipocytes, osteocytes and chondrocytes. The ability of MSCs to differentiate into endothelial cell

  11. 木材烟雾凝集物刺激人气道上皮细胞发生转分化样改变%Wood smoke condensate induced epithelial-mesenchymal transition in human airway epithelial cells

    Institute of Scientific and Technical Information of China (English)

    李雯曦; 邹威凤; 李冰; 冉丕鑫

    2014-01-01

    Objective To observe the detrimental effects of wood smoke condensate (WSC) exposure on human bronchial epithelial cells (HBEC),and to explore the expression of epithelialmesenchymal transition (EMT) markers in HBEC exposed to WSC.Methods HBEC were exposed respectively to 5,10,20,40 and 50 mg/L of WSC/CSC for 7 days,with control groups only in cell culture medium at the same time,then the total cytoactivity was detected by cell counting kit-8.After observing the cellular morphology of WSC-stimulated HBEC.Western blot and immunofluorescence method were used to evaluate the expression levels of type Ⅰ collagen,vimentin,E-cad and MMP-9 in HBEC exposed to WSC (10 mg/L) and cigarette smoke condensate (CSC) (10 mg/L) for 7 days.Statistical evaluation of the continuous data was performed by ANOVA.Independent-Samples t-test for between-group comparisons.Results After 7 days of exposure to WSC,HBEC manifested a morphological characteristic of loss of cellcell contact and elongated shape.The level of E-cad was decreased in WSC exposure groups (Western blot:0.30 ± 0.05,F =22.07,P < 0.05) compared with the groups without WSC exposure (Western blot:0.59 ± 0.08,F =22.07,P < 0.05).In contrast,an upregulation in expression of type Ⅰ collagen (Western blot:0.58 ± 0.04 vs 0.26 ± 0.02,F =119.72,P < 0.05) and MMP-9 (0.56 ± 0.08 vs 0.19 ± 0.03,F =21.79,P < 0.05) was observed in the presence of WSC,compared with the control groups.Immunofluorescence analysis showed that after a 7-day exposure to WSC in these cells,the E-cad protein was lost whereas type Ⅰ collagen,vimentin and MMP-9 were acquired.Both Western blot and immunofluorescence analysis showed no difference in expression levels of E-cad,type Ⅰ collagen,vimentin and MMP-9 between WSC and CSC exposure groups.Conclusion WSC exposure could induce EMT-like process in human airway epithelial cells.%目的 观察木材烟雾凝集物(WSC)对人支气管上皮细胞(HBEC)间充质转分

  12. Fuel Cell Buses in U.S. Transit Fleets: Current Status 2009

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, L.; Chandler, K.; Gikakis, C.

    2009-10-01

    This report documents progress in meeting the technological challenges of fuel cell propulsion for transportation based on current fuel cell transit bus demonstrations and plans for more fuel cell transit buses and hydrogen infrastructure.

  13. Cancer-associated fibroblasts in a human HEp-2 established laryngeal xenografted tumor are not derived from cancer cells through epithelial-mesenchymal transition, phenotypically activated but karyotypically normal.

    Science.gov (United States)

    Wang, Mei; Wu, Chun-Ping; Pan, Jun-Yan; Zheng, Wen-Wei; Cao, Xiao-Juan; Fan, Guo-Kang

    2015-01-01

    Cancer-associated fibroblasts (CAFs) play a crucial role in cancer progression and even initiation. However, the origins of CAFs in various cancer types remain controversial, and one of the important hypothesized origins is through epithelial-mesenchymal transition (EMT) from cancer cells. In this study, we investigated whether the HEp-2 laryngeal cancer cells are able to generate CAFs via EMT during tumor formation, which is now still unknown. The laryngeal xenografted tumor model was established by inoculating the HEp-2 laryngeal cancer cell line in nude mice. Primary cultured CAFs from the tumor nodules and matched normal fibroblasts (NFs) from the adjacent connective tissues were subcultured, purified, and verified by immunofluorescence. Migration, invasion, and proliferation potentials were compared between the CAFs and NFs. A co-culture of CAFs with HEp-2 cells and a co-injection of CAFs with HEp-2 cells in nude mice were performed to examine the cancer-promoting potential of CAFs to further verify their identity. Karyotypic analyses of the CAFs, NFs, and HEp-2 cells were conducted. A co-culture of NFs with HEp-2 cells was also performed to examine the expression of activated markers of CAFs. A pathological examination confirmed that the laryngeal xenografted tumor model was successfully established, containing abundant CAFs. Immunocytochemical staining verified the purities and identities of the CAFs and NFs. Although the CAFs manifested higher migration, invasion, proliferation, and cancer-promoting capacities compared with the NFs, an analysis of chromosomes revealed that both the CAFs and NFs showed typical normal mouse karyotypes. In addition, the NFs co-cultured with HEp-2 cells did not show induced expressions of activated markers of CAFs. Our findings reveal that the CAFs in the HEp-2 established laryngeal xenografted tumor are not of laryngeal cancer origin but of mouse origin, indicating that the HEp-2 laryngeal cancer cells cannot generate their

  14. Staghorn calculi and xanthogranulomatous pyelonephritis associated with transitional cell carcinoma

    Directory of Open Access Journals (Sweden)

    Chao-Wei Tseng

    2015-03-01

    Full Text Available Untreated staghorn calculi can cause xanthogranulomatous pyelonephritis (XGP, diminished renal function, and renal malignancy. Squamous cell carcinoma (SCC of the upper urinary tract is associated with kidney stones and chronic infection, but their association with transitional cell carcinoma (TCC has not been proven and has rarely been reported in literature. We present a rare case of staghorn calculi and XGP associated with TCC.

  15. Understanding the human dimensions of a sustainable energy transition.

    Science.gov (United States)

    Steg, Linda; Perlaviciute, Goda; van der Werff, Ellen

    2015-01-01

    Global climate change threatens the health, economic prospects, and basic food and water sources of people. A wide range of changes in household energy behavior is needed to realize a sustainable energy transition. We propose a general framework to understand and encourage sustainable energy behaviors, comprising four key issues. First, we need to identify which behaviors need to be changed. A sustainable energy transition involves changes in a wide range of energy behaviors, including the adoption of sustainable energy sources and energy-efficient technology, investments in energy efficiency measures in buildings, and changes in direct and indirect energy use behavior. Second, we need to understand which factors underlie these different types of sustainable energy behaviors. We discuss three main factors that influence sustainable energy behaviors: knowledge, motivations, and contextual factors. Third, we need to test the effects of interventions aimed to promote sustainable energy behaviors. Interventions can be aimed at changing the actual costs and benefits of behavior, or at changing people's perceptions and evaluations of different costs and benefits of behavioral options. Fourth, it is important to understand which factors affect the acceptability of energy policies and energy systems changes. We discuss important findings from psychological studies on these four topics, and propose a research agenda to further explore these topics. We emphasize the need of an integrated approach in studying the human dimensions of a sustainable energy transition that increases our understanding of which general factors affect a wide range of energy behaviors as well as the acceptability of different energy policies and energy system changes.

  16. Insulin-producing cells generated from dedifferentiated human pancreatic beta cells expanded in vitro.

    Directory of Open Access Journals (Sweden)

    Holger A Russ

    Full Text Available BACKGROUND: Expansion of beta cells from the limited number of adult human islet donors is an attractive prospect for increasing cell availability for cell therapy of diabetes. However, attempts at expanding human islet cells in tissue culture result in loss of beta-cell phenotype. Using a lineage-tracing approach we provided evidence for massive proliferation of beta-cell-derived (BCD cells within these cultures. Expansion involves dedifferentiation resembling epithelial-mesenchymal transition (EMT. Epigenetic analyses indicate that key beta-cell genes maintain open chromatin structure in expanded BCD cells, although they are not transcribed. Here we investigated whether BCD cells can be redifferentiated into beta-like cells. METHODOLOGY/PRINCIPAL FINDING: Redifferentiation conditions were screened by following activation of an insulin-DsRed2 reporter gene. Redifferentiated cells were characterized for gene expression, insulin content and secretion assays, and presence of secretory vesicles by electron microscopy. BCD cells were induced to redifferentiate by a combination of soluble factors. The redifferentiated cells expressed beta-cell genes, stored insulin in typical secretory vesicles, and released it in response to glucose. The redifferentiation process involved mesenchymal-epithelial transition, as judged by changes in gene expression. Moreover, inhibition of the EMT effector SLUG (SNAI2 using shRNA resulted in stimulation of redifferentiation. Lineage-traced cells also gave rise at a low rate to cells expressing other islet hormones, suggesting transition of BCD cells through an islet progenitor-like stage during redifferentiation. CONCLUSIONS/SIGNIFICANCE: These findings demonstrate for the first time that expanded dedifferentiated beta cells can be induced to redifferentiate in culture. The findings suggest that ex-vivo expansion of adult human islet cells is a promising approach for generation of insulin-producing cells for

  17. Phase transitions in human IgG solutions.

    Science.gov (United States)

    Wang, Ying; Lomakin, Aleksey; Latypov, Ramil F; Laubach, Jacob P; Hideshima, Teru; Richardson, Paul G; Munshi, Nikhil C; Anderson, Kenneth C; Benedek, George B

    2013-09-28

    Protein condensations, such as crystallization, liquid-liquid phase separation, aggregation, and gelation, have been observed in concentrated antibody solutions under various solution conditions. While most IgG antibodies are quite soluble, a few outliers can undergo condensation under physiological conditions. Condensation of IgGs can cause serious consequences in some human diseases and in biopharmaceutical formulations. The phase transitions underlying protein condensations in concentrated IgG solutions is also of fundamental interest for the understanding of the phase behavior of non-spherical protein molecules. Due to the high solubility of generic IgGs, the phase behavior of IgG solutions has not yet been well studied. In this work, we present an experimental approach to study IgG solutions in which the phase transitions are hidden below the freezing point of the solution. Using this method, we have investigated liquid-liquid phase separation of six human myeloma IgGs and two recombinant pharmaceutical human IgGs. We have also studied the relation between crystallization and liquid-liquid phase separation of two human cryoglobulin IgGs. Our experimental results reveal several important features of the generic phase behavior of IgG solutions: (1) the shape of the coexistence curve is similar for all IgGs but quite different from that of quasi-spherical proteins; (2) all IgGs have critical points located at roughly the same protein concentration at ~100 mg/ml while their critical temperatures vary significantly; and (3) the liquid-liquid phase separation in IgG solutions is metastable with respect to crystallization. These features of phase behavior of IgG solutions reflect the fact that all IgGs have nearly identical molecular geometry but quite diverse net inter-protein interaction energies. This work provides a foundation for further experimental and theoretical studies of the phase behavior of generic IgGs as well as outliers with large propensity to

  18. In-transit metastases from squamous cell carcinoma penis

    Directory of Open Access Journals (Sweden)

    L Padmavathy

    2012-01-01

    Full Text Available An in-transit metastasis is one that is located between the primary tumor and the closest lymph node region and results from tumor emboli getting trapped in the lymphatic channels. A 65-year-old male patient who had undergone partial amputation of the penis and bilateral inguinal lymph node resection for squamous cell carcinoma of the penis 4 months earlier developed multiple cutaneous metastatic lesions in the pubic region and scrotum. The case is reported for the uncommon presentation of in-transit metastases.

  19. Effect of anthralin on cell viability in human prostate adenocarcinoma.

    Science.gov (United States)

    Raevskaya, A A; Gorbunova, S L; Savvateeva, M V; Severin, S E; Kirpichnikov, M P

    2012-07-01

    The study revealed the key role of serine protease hepsin activity in transition of in situ prostate adenocarcinoma into the metastasizing form. Inhibition of hepsin activity suppresses the invasive growth of the tumor. Hepsin is an convenient target for pharmacological agents, so the study of its inhibitory mechanisms is a promising avenue in drug development. Assay of proteolytic activity in various tumor cell lines in vitro showed that this activity in prostate adenocarcinoma cells significantly surpasses proteolytic activity in other examined tumor cell lines. Selective cytotoxic action of anthralin, an inhibitor of hepsin activity, on human adenocarcinoma cells was demonstrated in comparison with other tumor cell lines.

  20. Transitional Cell Carcinoma of the Urinary Bladder in a Beluga Whale (Delphinapterus leucas).

    Science.gov (United States)

    Martineau, D; Lagacé, A; Massé, R; Morin, M; Béland, P

    1985-10-01

    A transitional cell carcinoma of the urinary bladder was found in a beluga whale stranded in the St. Lawrence middle estuary. Various organs of this animal were submitted to high resolution gas chromatography coupled with mass spectrometry analysis. High frequency of urinary bladder cancer in the human population of the same area and the presence of carcinogenic compounds in the marine environment of this animal are discussed.Concurrent isolation of Edwardsiella tarda from various organs of this whale is also reported.

  1. Transitional Cell Carcinoma of the Urinary Bladder in a Beluga Whale (Delphinapterus leucas)

    OpenAIRE

    Martineau, D.; Lagacé, A.; Massé, R; Morin, M.; Béland, P

    1985-01-01

    A transitional cell carcinoma of the urinary bladder was found in a beluga whale stranded in the St. Lawrence middle estuary. Various organs of this animal were submitted to high resolution gas chromatography coupled with mass spectrometry analysis. High frequency of urinary bladder cancer in the human population of the same area and the presence of carcinogenic compounds in the marine environment of this animal are discussed.

  2. Stable knockdown of Kif5b in MDCK cells leads to epithelial–mesenchymal transition

    Energy Technology Data Exchange (ETDEWEB)

    Cui, Ju, E-mail: juzi.cui@gmail.com [The Key Laboratory of Geriatrics, Beijing Hospital & Beijing Institute of Geriatrics, Ministry of Health, Beijing (China); Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); Jin, Guoxiang; Yu, Bin [Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); Wang, Zai [Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); Institute of Clinical Medical Sciences, China-Japan Friendship Hospital, Beijing (China); Lin, Raozhou [Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); Huang, Jian-Dong, E-mail: jdhuang@hku.hk [Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); The Centre for Synthetic Biology Engineering Research, Shenzhen Institutes of Advanced Technology, Shenzhen (China)

    2015-07-17

    Polarization of epithelial cells requires vectorial sorting and transport of polarity proteins to apical or basolateral domains. Kif5b is the mouse homologue of the human ubiquitous Kinesin Heavy Chain (uKHC). To investigate the function of Kif5b in epithelial cells, we examined the phenotypes of Kif5b-deficient MDCK cells. Stable knockdown of Kif5b in MDCK cells resulted in reduced cell proliferation rate, profound changes in cell morphology, loss of epithelial cell marker, and gain of mesenchymal marker, as well as increased cell migration, invasion, and tumorigenesis abilities. E-cadherin and NMMIIA could interact with Kif5b in polarized MDCK cells, and their expression levels were decreased in Kif5b-deficient MDCK cells. Overexpression of E-cadherin and NMMIIA in Kif5b depleted MDCK cells could decrease mesenchymal marker expression and cell migration ability. These results indicate that stable knockdown of Kif5b in MDCK cells can lead to epithelial–mesenchymal transition, which is mediated by defective E-cadherin and NMMIIA expression. - Highlights: • Knockdown of Kif5b in MDCK cells resulted in reduced cell proliferation rate. • Kif5b deficient MDCK cells underwent epithelial–mesenchymal transition. • E-cadherin and NMMIIA could interact with Kif5b in polarized MDCK cells. • Decreased E-cadherin and NMMIIA levels mediate EMT in Kif5b deficient MDCK cells. • Overexpression of E-cadherin and NMMIIA reverse the effects of Kif5b knockdown.

  3. Endothelial to Mesenchymal Transition (EndoMT in the Pathogenesis of Human Fibrotic Diseases

    Directory of Open Access Journals (Sweden)

    Sonsoles Piera-Velazquez

    2016-04-01

    Full Text Available Fibrotic diseases encompass a wide spectrum of clinical entities including systemic fibrotic diseases such as systemic sclerosis, sclerodermatous graft versus host disease, nephrogenic systemic fibrosis, and IgG4-associated sclerosing disease, as well as numerous organ-specific disorders including radiation-induced fibrosis, and cardiac, pulmonary, liver, and kidney fibrosis. Although their causative mechanisms are quite diverse, these diseases share the common feature of an uncontrolled and progressive accumulation of fibrous tissue macromolecules in affected organs leading to their dysfunction and ultimate failure. The pathogenesis of fibrotic diseases is complex and despite extensive investigation has remained elusive. Numerous studies have identified myofibroblasts as the cells responsible for the establishment and progression of the fibrotic process. Tissue myofibroblasts in fibrotic diseases originate from several sources including quiescent tissue fibroblasts, circulating CD34+ fibrocytes, and the phenotypic conversion of various cell types including epithelial and endothelial cells into activated myofibroblasts. However, the role of the phenotypic transition of endothelial cells into mesenchymal cells (Endothelial to Mesenchymal Transition or EndoMT in the pathogenesis of fibrotic disorders has not been fully elucidated. Here, we review the evidence supporting EndoMT’s contribution to human fibrotic disease pathogenesis.

  4. Endothelial to Mesenchymal Transition (EndoMT) in the Pathogenesis of Human Fibrotic Diseases

    Science.gov (United States)

    Piera-Velazquez, Sonsoles; Mendoza, Fabian A.; Jimenez, Sergio A.

    2016-01-01

    Fibrotic diseases encompass a wide spectrum of clinical entities including systemic fibrotic diseases such as systemic sclerosis, sclerodermatous graft versus host disease, nephrogenic systemic fibrosis, and IgG4-associated sclerosing disease, as well as numerous organ-specific disorders including radiation-induced fibrosis, and cardiac, pulmonary, liver, and kidney fibrosis. Although their causative mechanisms are quite diverse, these diseases share the common feature of an uncontrolled and progressive accumulation of fibrous tissue macromolecules in affected organs leading to their dysfunction and ultimate failure. The pathogenesis of fibrotic diseases is complex and despite extensive investigation has remained elusive. Numerous studies have identified myofibroblasts as the cells responsible for the establishment and progression of the fibrotic process. Tissue myofibroblasts in fibrotic diseases originate from several sources including quiescent tissue fibroblasts, circulating CD34+ fibrocytes, and the phenotypic conversion of various cell types including epithelial and endothelial cells into activated myofibroblasts. However, the role of the phenotypic transition of endothelial cells into mesenchymal cells (Endothelial to Mesenchymal Transition or EndoMT) in the pathogenesis of fibrotic disorders has not been fully elucidated. Here, we review the evidence supporting EndoMT’s contribution to human fibrotic disease pathogenesis. PMID:27077889

  5. Human fetal mesenchymal stem cells.

    Science.gov (United States)

    O'Donoghue, Keelin; Chan, Jerry

    2006-09-01

    Stem cells have been isolated at all stages of development from the early developing embryo to the post-reproductive adult organism. However, the fetal environment is unique as it is the only time in ontogeny that there is migration of stem cells in large numbers into different organ compartments. While fetal neural and haemopoietic stem cells (HSC) have been well characterised, only recently have mesenchymal stem cells from the human fetus been isolated and evaluated. Our group have characterised in human fetal blood, liver and bone marrow a population of non-haemopoietic, non-endothelial cells with an immunophenotype similar to adult bone marrow-derived mesenchymal stem cells (MSC). These cells, human fetal mesenchymal stem cells (hfMSC), are true multipotent stem cells with greater self-renewal and differentiation capacity than their adult counterparts. They circulate in first trimester fetal blood and have been found to traffic into the maternal circulation, engrafting in bone marrow, where they remain microchimeric for decades after pregnancy. Though fetal microchimerism has been implicated in the pathogenesis of autoimmune disease, the biological role of hfMSC microchimerism is unknown. Potential downstream applications of hfMSC include their use as a target cell for non-invasive pre-natal diagnosis from maternal blood, and for fetal cellular and gene therapy. Using hfMSC in fetal therapy offers the theoretical advantages of avoidance of immune rejection, increased engraftment, and treatment before disease pathology sets in. Aside from allogeneic hfMSC in utero transplantation, the use of autologous hfMSC has been brought a step forward with the development of early blood sampling techniques, efficient viral transduction and clonal expansion. Work is ongoing to determine hfMSC fate post-transplantation in murine models of genetic disease. In this review we will examine what is known about hfMSC biology, as well as discussing areas for future research. The

  6. A new Zero-Voltage-Transition PWM switching cell

    Energy Technology Data Exchange (ETDEWEB)

    Grigore, V. [Electronics and Telecommunications Faculty `Politebuica` University Bucharest (Romania); Kyyrae, J. [Helsinki University of Technology, Otaniemi (Finland): Institute of Intelligent Power Electronics

    1997-12-31

    In this paper a new Zero-Voltage-Transition (ZVT) PWM switching cell is presented. The proposed switching cell is composed of the normal hard-switched PWM cell (consisting of one active switch and one passive switch), plus an auxiliary circuit (consisting of one active switch and some reactive components). The auxiliary circuit is inactive during the ON and OFF intervals of the switches in the normal PWM switch. However, the transitions between the two states are controlled by the auxiliary circuit. Prior to turn-on, the voltage across the active switch in the PWM cell is forced to zero, thus the turn-on losses of the active switch are practically eliminated. At turn-off the auxiliary circuit behaves like a non-dissipative passive snubber reducing the turn-off losses to a great extent. Zero-Voltage-Transition switching technique almost eliminates switching losses. The active switch operates under ZVT conditions, the passive switch (diode) has a controlled reverse recovery, and the switch in the auxiliary circuit operates under Zero-Current-Switching (ZCS) conditions. (orig.) 6 refs.

  7. TEAD transcription factors mediate the function of TAZ in cell growth and epithelial-mesenchymal transition.

    Science.gov (United States)

    Zhang, Heng; Liu, Chen-Ying; Zha, Zheng-Yu; Zhao, Bin; Yao, Jun; Zhao, Shimin; Xiong, Yue; Lei, Qun-Ying; Guan, Kun-Liang

    2009-05-15

    The TAZ transcription co-activator has been shown to promote cell proliferation and to induce epithelial-mesenchymal transition. Recently we have demonstrated that TAZ is phosphorylated and inhibited by the Hippo tumor suppressor pathway, which is altered in human cancer. The mechanism of TAZ-mediated transcription is unclear. We demonstrate here that TEAD is a key downstream transcription factor mediating the function of TAZ. Disruption of TEAD-TAZ binding or silencing of TEAD expression blocked the function of TAZ to promote cell proliferation and to induce epithelial-mesenchymal transition, demonstrating TEAD as a key downstream effector of TAZ. We also identified CTGF, a gene that regulates cell adhesion, proliferation, and migration, as a direct target of TAZ and TEAD. Our study establishes a functional partnership between TAZ and TEAD under negative regulation by the Hippo signaling pathway.

  8. miR-335 orchestrates cell proliferation, migration and differentiation in human mesenchymal stem cells

    OpenAIRE

    Gonzalez, Manuel A.; Tomé, María; López-Romero, Pedro; Albo, Carmen; Sepúlveda, Juan Carlos; Fernández-Gutierrez, Benjamin; Dopazo, Ana; Bernad, Antonio

    2010-01-01

    Abstract In spite of the extensive potential of human mesenchymal stem cells (hMSCs) in cell therapy, little is known about the molecular mechanisms that regulate their therapeutic properties. We aimed to identify microRNAs (miRNAs) involved in controlling the transition between the resting and reparative phenotypes of hMSCs, hypothesizing that these miRNAs must be present in the undifferentiated cells and downregulated to allow initiation of distinct activation/differentiation pro...

  9. Differentiated human stem cells resemble fetal, not adult, β cells.

    Science.gov (United States)

    Hrvatin, Sinisa; O'Donnell, Charles W; Deng, Francis; Millman, Jeffrey R; Pagliuca, Felicia Walton; DiIorio, Philip; Rezania, Alireza; Gifford, David K; Melton, Douglas A

    2014-02-25

    Human pluripotent stem cells (hPSCs) have the potential to generate any human cell type, and one widely recognized goal is to make pancreatic β cells. To this end, comparisons between differentiated cell types produced in vitro and their in vivo counterparts are essential to validate hPSC-derived cells. Genome-wide transcriptional analysis of sorted insulin-expressing (INS(+)) cells derived from three independent hPSC lines, human fetal pancreata, and adult human islets points to two major conclusions: (i) Different hPSC lines produce highly similar INS(+) cells and (ii) hPSC-derived INS(+) (hPSC-INS(+)) cells more closely resemble human fetal β cells than adult β cells. This study provides a direct comparison of transcriptional programs between pure hPSC-INS(+) cells and true β cells and provides a catalog of genes whose manipulation may convert hPSC-INS(+) cells into functional β cells.

  10. Subcutaneous Transitional Cell Cancer After Percutaneous Nephrolithotomy: A Case Report

    Directory of Open Access Journals (Sweden)

    Lokman Ižrkilata

    2013-10-01

    Full Text Available Transitional cell carcinomas of the upper urinary tract are rare but, highly predisposing to tumoral seeding. Percutaneous lithotripsy (PNL recently has expanded the therapeutic choices for patients with kidney stones and gained popularity by urologic surgeons. Although unusual, renal collecting system tumours may be encountered during PNL. We present and discuss the clinical course of a 48 years old male patient who underwent PNL surgery for kidney stone in whom transitional cell carcinoma in the renal collecting system obscured by stone left undiagnosed. Three months later following PNL he admitted with a bulge on lumbar region. Excisional biopsy revealed carcinoma and therefore, he was directed to chemoradiotherapy and died 21 months later. Renal collecting system tumors undiagnosed during surgery may progress and demonstrate local invasion in a short period of time. Therefore, we recommend to take more caution during any percutaneous access and to exclude the possible existence of tumor.

  11. Isolated Meningeal Recurrence of Transitional Cell Carcinoma of the Bladder

    Directory of Open Access Journals (Sweden)

    Catherine Butchart

    2010-06-01

    Full Text Available Meningeal carcinomatosis occurs in 1–18% of patients with solid tumours, most commonly carcinomas of the breast and lung or melanomas. There are relatively few reports of meningeal carcinomatosis in transitional cell carcinoma of the bladder. Isolated meningeal recurrence is particularly uncommon, and we present an unusual case of this in a 58-year-old man. The case was further complicated by the somewhat atypical presentation with a confirmed ischaemic stroke. The patient died one month after presentation.

  12. Transitional Cell Carcinoma of Kidney- Report of a Rare Case

    Directory of Open Access Journals (Sweden)

    Priyesh Halgaonkar

    2015-01-01

    Full Text Available Hematuria is a common presentation in the surgical outpatient department. The most common causes being urinary tract infection or renal calculi that causes hematuria. Few of them are being diagnosed as Renal or Bladder mass. Transitional cell carcinoma affecting urogenital tract accounts for 5-10% of the primary renal malignancies which is relatively rare. Here we report such rare case in an elderly female who presented with painless hematuria.

  13. Induced adult stem (iAS) cells and induced transit amplifying progenitor (iTAP) cells-a possible alternative to induced pluripotent stem (iPS) cells?

    Science.gov (United States)

    Heng, Boon Chin; Richards, Mark; Ge, Zigang; Shu, Yimin

    2010-02-01

    The successful derivation of iPSC lines effectively demonstrates that it is possible to reset the 'developmental clock' of somatic cells all the way back to the initial embryonic state. Hence, it is plausible that this clock may instead be turned back half-way to a less immature developmental stage that is more directly applicable to clinical therapeutic applications or for in vitro pharmacology/toxicology screening assays. Such a suitable developmental state is postulated to be either the putative transit amplifying progenitor stage or adult stem cell stage. It is hypothetically possible to reprogram mature and terminally differentiated somatic cells back to the adult stem cell or transit amplifying progenitor stage, in a manner similar to the derivation of iPSC. It is proposed that the terminology 'Induced Adult Stem Cells' (iASC) or 'Induced Transit Amplifying Progenitor Cells' (iTAPC) be used to described such reprogrammed somatic cells. Of particular interest, is the possibility of resetting the developmental clock of mature differentiated somatic cells of the mesenchymal lineage, explanted from adipose tissue, bone marrow and cartilage. The putative adult stem cell sub-population from which these cells are derived, commonly referred to as 'mesenchymal stem cells', are highly versatile and hold much therapeutic promise in regenerative medicine, as attested to by numerous human clinical trials and animal studies. Perhaps it may be appropriate to term such reprogrammed cells as 'Induced Mesenchymal Stem Cells' (iMSC) or as 'Induced Mesenchumal Progenitor Cells' (iMPC). Given that cells from the same organ/tissue will share some commonalities in gene expression, we hypothesize that the generation of iASC or iTAPC would be more efficient as compared to iPSC generation, since a common epigenetic program must exist between the reprogrammed cells, adult stem cell or progenitor cell types and terminally differentiated cell types from the same organ/tissue.

  14. Mark the transition: chromatin modifications and cell fate decision

    Institute of Scientific and Technical Information of China (English)

    Qiang Wu; Huck-Hui Ng

    2011-01-01

    With their unique features of selfrenewal and pluripotency,human embryonic stem (hES) cells are considered to be a nearly unlimited resource for research and clinical applications [1].Accordingly,the transcriptional network specifying and governing human ES cell identity has been extensively studied.OCT4,NANOG and SOX2 form a core transcriptional network that regulates itself as well as a number of target genes [2].This transcriptional network acts together with signaling pathways to maintain ES cell identity [3].Moreover,the last decade has seen tremendous advances in understanding the epigenetic mechanisms underlying ES eell self-renewal and pluripotency.

  15. Urban transitions: on urban resilience and human-dominated ecosystems.

    Science.gov (United States)

    Ernstson, Henrik; van der Leeuw, Sander E; Redman, Charles L; Meffert, Douglas J; Davis, George; Alfsen, Christine; Elmqvist, Thomas

    2010-12-01

    Urbanization is a global multidimensional process paired with increasing uncertainty due to climate change, migration of people, and changes in the capacity to sustain ecosystem services. This article lays a foundation for discussing transitions in urban governance, which enable cities to navigate change, build capacity to withstand shocks, and use experimentation and innovation in face of uncertainty. Using the three concrete case cities--New Orleans, Cape Town, and Phoenix--the article analyzes thresholds and cross-scale interactions, and expands the scale at which urban resilience has been discussed by integrating the idea from geography that cities form part of "system of cities" (i.e., they cannot be seen as single entities). Based on this, the article argues that urban governance need to harness social networks of urban innovation to sustain ecosystem services, while nurturing discourses that situate the city as part of regional ecosystems. The article broadens the discussion on urban resilience while challenging resilience theory when addressing human-dominated ecosystems. Practical examples of harnessing urban innovation are presented, paired with an agenda for research and policy.

  16. Transition metal catalysis in the mitochondria of living cells

    Science.gov (United States)

    Tomás-Gamasa, María; Martínez-Calvo, Miguel; Couceiro, José R.; Mascareñas, José L.

    2016-09-01

    The development of transition metal catalysts capable of promoting non-natural transformations within living cells can open significant new avenues in chemical and cell biology. Unfortunately, the complexity of the cell makes it extremely difficult to translate standard organometallic chemistry to living environments. Therefore, progress in this field has been very slow, and many challenges, including the possibility of localizing active metal catalysts into specific subcellular sites or organelles, remain to be addressed. Herein, we report a designed ruthenium complex that accumulates preferentially inside the mitochondria of mammalian cells, while keeping its ability to react with exogenous substrates in a bioorthogonal way. Importantly, we show that the subcellular catalytic activity can be used for the confined release of fluorophores, and even allows selective functional alterations in the mitochondria by the localized transformation of inert precursors into uncouplers of the membrane potential.

  17. Evidencing the "robot phase transition" in experimental human-algorithmic markets

    OpenAIRE

    Cartlidge, John; Cliff, Dave

    2013-01-01

    Johnson, Zhao, Hunsader, Meng, Ravindar, Carran, and Tivnan (2012) recently suggested the existence of a phase transition in the dynamics of financial markets in which there is free interaction between human traders and algorithmic trading systems ("robots"). Above a particular time-threshold, humans and robots trade with one another; below the threshold all transactions are robot-to-robot. We refer to this abrupt system transition as the "robot phase transition". Here, we conduct controlled ...

  18. The potency of human testicular stem cells

    NARCIS (Netherlands)

    Chikhovskaya, J.V.

    2013-01-01

    In this thesis, we evaluate the stem cell state of cells present in primary human testicular cell cultures as well as their origin and relation to germ or somatic lineages within testicular tissue. We conclude that human testis-derived embryonic stem cell-like (htES-like) colonies arising in primary

  19. “人源性”乳腺微环境促进人源性乳腺癌细胞发生上皮细胞间充质转化%Human mammary microenvironment promotes epithelial mesenchymal transition in human breast cancer cells

    Institute of Scientific and Technical Information of China (English)

    张桂普; 王水; 郑明洁; 王珏

    2012-01-01

    目的 研究“人源性”乳腺微环境对人源性乳腺癌细胞生物学活性的调控作用.方法 20只重症联合免疫缺陷小鼠随机均分为“人源性”乳腺微环境小鼠模型组(实验组)和皮下注射小鼠模型组(对照组).取小鼠的乳腺癌原发灶,采用Transwell小室法检测细胞迁移和侵袭能力,免疫组织化学法检测E-钙黏蛋白(E-cadherin)和波形蛋白(vimentin)表达.结果 与对照组相比,实验组细胞迁移和细胞侵袭数量都明显增加[(29.25±5.17)个vs.(67.14±6.53)个和(63.57±6.54)个vs.(118.29±12.81)个](P<0.05).两组小鼠原发灶标本的E-cadherin均为阴性,但实验组vimentin 的表达强于对照组.结论 建立的具有“人源性”乳腺微环境的小鼠模型能很好地模拟出人源性乳腺癌细胞上皮细胞间充质转化的过程,是良好的乳腺癌转移研究的在体平台.%Objective To investigate the regulatory effect of human mammary microenvironment on biologic activity of human breast cancer cells. Methods Twenty-severe combined immunodeficiency (SCID) mice were equally randomized into two groups of A ("breast-breast" mouse model) and B (subcutaneous injection mouse model). The primary tumor of breast cancer in mice was taken,and the cell migration and invasion were detected by Transwell chamber. The expressions of E-cadherin and vimentin were measured by immunohistochemistry. Results Compared with group B, the numbers of cell migration and cell invasion were significantly increased [(29. 25 + 5. 17) pieces vs. (67. 14 + 6. 53) pieces and (63. 57 + 6. 54) pieces vs. (118. 29 + 12. 81) pieces](P<0. 05). E-cadherin was negatively expressed in both groups,but the expression of vimentin in group A was higher than that in group B. Conclusion The human mammary microenvironment can better simulate the process of epithelial mesenchymal transition in human breast cancer cells,and is a satisfactory platform for studying breast cancer metastasis in vivo.

  20. A novel aminothiazole KY-05009 with potential to inhibit Traf2- and Nck-interacting kinase (TNIK attenuates TGF-β1-mediated epithelial-to-mesenchymal transition in human lung adenocarcinoma A549 cells.

    Directory of Open Access Journals (Sweden)

    Jiyeon Kim

    Full Text Available Transforming growth factor (TGF-β triggers the epithelial-to-mesenchymal transition (EMT of cancer cells via well-orchestrated crosstalk between Smad and non-Smad signaling pathways, including Wnt/β-catenin. Since EMT-induced motility and invasion play a critical role in cancer metastasis, EMT-related molecules are emerging as novel targets of anti-cancer therapies. Traf2- and Nck-interacting kinase (TNIK has recently been considered as a first-in-class anti-cancer target molecule to regulate Wnt signaling pathway, but pharmacologic inhibition of its EMT activity has not yet been studied. Here, using 5-(4-methylbenzamido-2-(phenylaminothiazole-4-carboxamide (KY-05009 with TNIK-inhibitory activity, its efficacy to inhibit EMT in cancer cells was validated. The molecular docking/binding study revealed the binding of KY-05009 in the hinge region of TNIK, and the inhibitory activity of KY-05009 against TNIK was confirmed by an ATP competition assay (Ki, 100 nM. In A549 cells, KY-05009 significantly and strongly inhibited the TGF-β-activated EMT through the attenuation of Smad and non-Smad signaling pathways, including the Wnt, NF-κB, FAK-Src-paxillin-related focal adhesion, and MAP kinases (ERK and JNK signaling pathways. Continuing efforts to identify and validate potential therapeutic targets associated with EMT, such as TNIK, provide new and improved therapies for treating and/or preventing EMT-based disorders, such as cancer metastasis and fibrosis.

  1. Induction of the mesenchymal to epithelial transition by demethylation- activated microRNA-200c is involved in the anti-migration/invasion effects of arsenic trioxide on human breast cancer cells.

    Science.gov (United States)

    Si, Lu; Jiang, Fei; Li, Yuan; Ye, Xianqing; Mu, Juan; Wang, Xingxing; Ning, Shilong; Hu, Chunyan; Li, Zhong

    2015-09-01

    Breast cancer is a major health problem worldwide. Current standard practices for treatment of breast cancer are less than satisfactory because of high rates of metastasis. Arsenic trioxide (As(2)O(3)), which induces demethylation of DNA and causes apoptosis, has been used as an anti-tumor agent. Little is known, however, regarding its anti-metastatic effects. The microRNA-200c (miR-200c), which is frequently lowly expressed in triple negative breast cancers (TNBCs), inhibits metastasis by inducing the mesenchymal to epithelial transition (MET). Here, we report that As(2)O(3) attenuates the migratory and invasive capacities of breast cancer cells, MDA-MB-231 and BT-549. Notably, As(2)O(3) induces an MET in vitro and in vivo, as determined by the increased expression of the epithelial marker, E-cadherin and decreased expressions of mesenchymal markers, N-cadherin and vimentin. Moreover, As(2)O(3) up-regulates the expression of miR-200c through demethylation. Over-expression of miR-200c enhances the expression of E-cadherin and decreases the expressions of N-cadherin and vimentin. Further, in MDA-MB-231 cells exposed to As(2)O(3), knockdown of miR-200c blocks the As(2)O(3) -induced MET. Finally, in MDA-MB-231 and BT-549 cells exposed to As(2)O(3), knockdown of miR-200c decreases the As(2)O(3) -induced inhibition of the migratory and invasive capacities. By identifying a mechanism whereby As(2)O(3) regulates miR-200c and MET, the results establish the anti-migration/invasion potential of arsenic trioxide.

  2. A new Zero-Current-Transition PWM switching cell

    Energy Technology Data Exchange (ETDEWEB)

    Grigore, V. [Electronics and Telecommunications Faculty, `Politechnica` University Bucharest (Romania); Kyyrae, J. [Helsinki University of Technology, Otaniemi (Finland): Institute of Intelligent Power Electronics

    1997-12-31

    In this paper a new Zero-Current-Transition (ZCT) PWM switching cell is presented. The proposed switching cell is composed of the normal hard-switched PWM cell (consisting of one active switch and one passive switch), plus as auxiliary circuit. The auxiliary circuit is inactive during the ON ad OFF intervals of the switches in the normal PWM switch. The transitions between the two states are controlled by the auxiliary circuit. Prior to turn-off, the current through the active switch in the PWM cell is forced to zero, thus the turn-off losses of the active switch are practically eliminated. At turn-on the auxiliary circuit slows down the growing rate of the current through the main switch. Thus, turn-on losses are also very much reduced. The active switch operates under ZCT conditions, the passive switch (diode) has a controlled reverse recovery, while the switch in the auxiliary circuit operates under Zero-Current-Switching (ZCS) conditions. (orig.) 3 refs.

  3. Organochlorine pesticides induce epithelial to mesenchymal transition of human primary cultured hepatocytes.

    Science.gov (United States)

    Zucchini-Pascal, Nathalie; Peyre, Ludovic; de Sousa, Georges; Rahmani, Roger

    2012-11-01

    Persistent organic pollutants (POPs) are a group of organic or chemicals that adversely affect human health and are persistent in the environment. These highly toxic compounds include industrial chemicals, pesticides such as organochlorines, and unwanted wastes such as dioxins. Although studies have described the general toxicity effects of organochlorine pesticides, the mechanisms underlying its potential carcinogenic effects in the liver are not well understood. In this study, we analyzed the effect of three organochlorine pesticides (dichlorodiphenyltrichloroethane, heptachlore and endosulfan) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the epithelial to mesenchymal transition (EMT) in primary cultured human hepatocytes. We found that these compounds modified the hepatocyte phenotype, inducing cell spread, formation of lamellipodia structures and reorganization of the actin cytoskeleton in stress fibers. These morphological alterations were accompanied by disruption of cell-cell junctions, E-cadherin repression and albumin down-regulation. Interestingly, these characteristic features of dedifferentiating hepatocytes were correlated with the gain of expression of various mesenchymal genes, including vimentin, fibronectin and its receptor ITGA5. These various results show that organochlorines and TCDD accelerate cultured human hepatocyte dedifferentiation and EMT processes. These events could account, at least in part, for the carcionogenic and/or fibrogenic activities of these POPs. Copyright © 2012 Elsevier Ltd. All rights reserved.

  4. National Fuel Cell Bus Program: Accelerated Testing Evaluation Report #2, Alameda-Contra Costa Transit District (AC Transit) and Appendices

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, L.; Chandler, K.

    2010-06-01

    This is an evaluation of hydrogen fuel cell transit buses operating at AC Transit in revenue service since March 20, 2006, comparing similar diesel buses operating from the same depot. It covers November 2007 through February 2010. Results include implementation experience, fueling station operation, evaluation results at AC Transit (bus usage, availability, fuel economy, maintenance costs, and road calls), and a summary of achievements and challenges encountered during the demonstration.

  5. Stem cell differentiation and human liver disease

    Institute of Scientific and Technical Information of China (English)

    Wen-Li Zhou; Claire N Medine; Liang Zhu; David C Hay

    2012-01-01

    Human stem cells are scalable cell populations capable of cellular differentiation.This makes them a very attractive in vitro cellular resource and in theory provides unlimited amounts of primary cells.Such an approach has the potential to improve our understanding of human biology and treating disease.In the future it may be possible to deploy novel stem cell-based approaches to treat human liver diseases.In recent years,efficient hepatic differentiation from human stem cells has been achieved by several research groups including our own.In this review we provide an overview of the field and discuss the future potential and limitations of stem cell technology.

  6. The human airway epithelial basal cell transcriptome.

    Directory of Open Access Journals (Sweden)

    Neil R Hackett

    Full Text Available BACKGROUND: The human airway epithelium consists of 4 major cell types: ciliated, secretory, columnar and basal cells. During natural turnover and in response to injury, the airway basal cells function as stem/progenitor cells for the other airway cell types. The objective of this study is to better understand human airway epithelial basal cell biology by defining the gene expression signature of this cell population. METHODOLOGY/PRINCIPAL FINDINGS: Bronchial brushing was used to obtain airway epithelium from healthy nonsmokers. Microarrays were used to assess the transcriptome of basal cells purified from the airway epithelium in comparison to the transcriptome of the differentiated airway epithelium. This analysis identified the "human airway basal cell signature" as 1,161 unique genes with >5-fold higher expression level in basal cells compared to differentiated epithelium. The basal cell signature was suppressed when the basal cells differentiated into a ciliated airway epithelium in vitro. The basal cell signature displayed overlap with genes expressed in basal-like cells from other human tissues and with that of murine airway basal cells. Consistent with self-modulation as well as signaling to other airway cell types, the human airway basal cell signature was characterized by genes encoding extracellular matrix components, growth factors and growth factor receptors, including genes related to the EGF and VEGF pathways. Interestingly, while the basal cell signature overlaps that of basal-like cells of other organs, the human airway basal cell signature has features not previously associated with this cell type, including a unique pattern of genes encoding extracellular matrix components, G protein-coupled receptors, neuroactive ligands and receptors, and ion channels. CONCLUSION/SIGNIFICANCE: The human airway epithelial basal cell signature identified in the present study provides novel insights into the molecular phenotype and biology of

  7. Proteomics research on muscle-invasive bladder transitional cell carcinoma

    Directory of Open Access Journals (Sweden)

    Cao Yan

    2011-06-01

    Full Text Available Abstract Background Aimed to facilitate candidate biomarkers selection and improve network-based multi-target therapy, we perform comparative proteomics research on muscle-invasive bladder transitional cell carcinoma. Laser capture microdissection was used to harvest purified muscle-invasive bladder cancer cells and normal urothelial cells from 4 paired samples. Two-dimensional liquid chromatography tandem mass spectrometry was used to identify the proteome expression profile. The differential proteins were further analyzed using bioinformatics tools and compared with the published literature. Results A total of 885/890 proteins commonly appeared in 4 paired samples. 295/337 of the 488/493 proteins that specific expressed in tumor/normal cells own gene ontology (GO cellular component annotation. Compared with the entire list of the international protein index (IPI, there are 42/45 GO terms exhibited as enriched and 9/5 exhibited as depleted, respectively. Several pathways exhibit significantly changes between cancer and normal cells, mainly including spliceosome, endocytosis, oxidative phosphorylation, etc. Finally, descriptive statistics show that the PI Distribution of candidate biomarkers have certain regularity. Conclusions The present study identified the proteome expression profile of muscle-invasive bladder cancer cells and normal urothelial cells, providing information for subcellular pattern research of cancer and offer candidate proteins for biomarker panel and network-based multi-target therapy.

  8. Identification of muscle synergies associated with gait transition in humans

    Directory of Open Access Journals (Sweden)

    Shota eHagio

    2015-02-01

    Full Text Available There is no theoretical or empirical evidence to suggest how the central nervous system (CNS controls a variety of muscles associated with gait transition between walking and running. Here, we examined the motor control during a gait transition based on muscle synergies, which modularly organize functionally similar muscles. To this end, the subjects walked or ran on a treadmill and performed a gait transition spontaneously as the treadmill speed increased or decreased (a changing speed condition or voluntarily following an experimenter’s instruction at constant treadmill speed (a constant speed condition. Surface electromyograms (EMGs were recorded from 11 lower limb muscles bilaterally. We then extracted the muscle weightings of synergies and their activation coefficients from the EMG data using non-negative matrix factorization. As a result, the gait transition was controlled by approximately 9 muscle synergies, which were common during a walking and running, and their activation profiles were changed before and after a gait transition. Near a gait transition, the peak activation phases of the synergies, which were composed of plantar flexor muscles, were shifted to an earlier phase at the walk-to-run transition, and vice versa. The shifts were gradual in the changing speed condition, but an abrupt change was observed in the constant speed condition. These results suggest that the CNS low-dimensionally regulate the activation profiles of the specific synergies based on afferent information (spontaneous gait transition or by changing only the descending neural input to the muscle synergies (voluntary gait transition to achieve a gait transition.

  9. Expression of cyclooxygenase-2 in human transitional cell carcinoma of bladder%环氧合酶-2蛋白在人膀胱移行细胞癌中的表达

    Institute of Scientific and Technical Information of China (English)

    李国平; 杨恬

    2002-01-01

    目的研究环氧合酶-2(COX-2)在膀胱移行细胞癌(Transitional cell carcinoma,TCC)组织中的表达及其意义.方法采用SP免疫组化技术,对48例尿路上皮TCC石蜡标本切片进行免疫组化染色,对阳性表达率与肿瘤分级及侵袭性间的相关性进行统计学分析.结果 COX-2蛋白的阳性表达率与TCC分级及侵袭性呈显著相关,在低分级TCC中没有COX-2蛋白的表达(0/18),在高分级TCC中COX-2阳性表达率为44%(8/18).结论 COX-2在高分级TCC和侵袭性TCC中阳性表达率显著上升,表明COX-2可能在TCC的形成中起重要作用,有可能成为治疗人类TCC的作用靶点.

  10. Endocannabinoids and Human Sperm Cells

    Directory of Open Access Journals (Sweden)

    Giovanna Zolese

    2010-10-01

    Full Text Available N-acylethanolamides (NAEs are naturally occurring signaling lipids consisting of amides and esters of long-chain polyunsaturated fatty acids. Usually they are present in a very small amounts in many mammalian tissues and cells, including human reproductive tracts and fluids. Recently, the presence of N-arachidonoylethanolamide (anandamide, AEA, the most characterised member of endocannabinoids, and its congeners palmitoylethanolamide (PEA and oleylethanolamide (OEA in seminal plasma, oviductal fluid, and follicular fluids was demonstrated. AEA has been shown to bind not only type-1 (CB1 and type-2 (CB2 cannabinoid receptors, but also type-1 vanilloid receptor (TRPV1, while PEA and OEA are inactive with respect to classical cannabinoid CB1 and CB2 but activate TRPV1 or peroxisome proliferator activate receptors (PPARs. This review concerns the most recent experimental data on PEA and OEA, endocannabinoid-like molecules which appear to exert their action exclusively on sperm cells with altered features, such as membrane characteristics and kinematic parameters. Their beneficial effects on these cells could suggest a possible pharmacological use of PEA and OEA on patients affected by some forms of idiopathic infertility.

  11. Speeding the transition: Designing a fuel-cell hypercar

    Energy Technology Data Exchange (ETDEWEB)

    Williams, B.D.; Moore, T.C.; Lovins, A.B. [Rocky Mountain Inst., Snowmass, CO (United States). Hypercar Center

    1997-12-31

    A rapid transformation now underway in automotive technology could accelerate the transition to transportation powered by fuel cells. Ultralight, advanced-composite, low-drag, hybrid-electric hypercars--using combustion engines--could be three- to fourfold more efficient and one or two orders of magnitude cleaner than today`s cars, yet equally safe, sporty, desirable, and (probably) affordable. Further, important manufacturing advantages--including low tooling and equipment costs, greater mechanical simplicity, autobody parts consolidation, shorter product cycles, and reduced assembly effort and space--permit a free-market commercialization strategy. This paper discusses a conceptual hypercar powered by a proton-exchange-membrane fuel cell (PEMFC). It outlines the implications of platform physics and component selection for the vehicle`s mass budget and performance. The high fuel-to-traction conversion efficiency of the hypercar platform could help automakers overcome the Achilles` heel of hydrogen-powered vehicles: onboard storage. Moreover, because hypercars would require significantly less tractive power, and even less fuel-cell power, they could adopt fuel cells earlier, before fuel cells` specific cost, mass, and volume have fully matured. In the meantime, commercialization in buildings can help prepare fuel cells for hypercars. The promising performance of hydrogen-fueled PEMFC hypercars suggests important opportunities in infrastructure development for direct-hydrogen vehicles.

  12. Human regulatory B cells control the TFH cell response.

    Science.gov (United States)

    Achour, Achouak; Simon, Quentin; Mohr, Audrey; Séité, Jean-François; Youinou, Pierre; Bendaoud, Boutahar; Ghedira, Ibtissem; Pers, Jacques-Olivier; Jamin, Christophe

    2017-07-01

    Follicular helper T (TFH) cells support terminal B-cell differentiation. Human regulatory B (Breg) cells modulate cellular responses, but their control of TFH cell-dependent humoral immune responses is unknown. We sought to assess the role of Breg cells on TFH cell development and function. Human T cells were polyclonally stimulated in the presence of IL-12 and IL-21 to generate TFH cells. They were cocultured with B cells to induce their terminal differentiation. Breg cells were included in these cultures, and their effects were evaluated by using flow cytometry and ELISA. B-cell lymphoma 6, IL-21, inducible costimulator, CXCR5, and programmed cell death protein 1 (PD-1) expressions increased on stimulated human T cells, characterizing TFH cell maturation. In cocultures they differentiated B cells into CD138(+) plasma and IgD(-)CD27(+) memory cells and triggered immunoglobulin secretions. Breg cells obtained by Toll-like receptor 9 and CD40 activation of B cells prevented TFH cell development. Added to TFH cell and B-cell cocultures, they inhibited B-cell differentiation, impeded immunoglobulin secretions, and expanded Foxp3(+)CXCR5(+)PD-1(+) follicular regulatory T cells. Breg cells modulated IL-21 receptor expressions on TFH cells and B cells, and their suppressive activities involved CD40, CD80, CD86, and intercellular adhesion molecule interactions and required production of IL-10 and TGF-β. Human Breg cells control TFH cell maturation, expand follicular regulatory T cells, and inhibit the TFH cell-mediated antibody secretion. These novel observations demonstrate a role for the Breg cell in germinal center reactions and suggest that deficient activities might impair the TFH cell-dependent control of humoral immunity and might lead to the development of aberrant autoimmune responses. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  13. Human pancreatic islet progenitor cells demonstrate phenotypic plasticity in vitro

    Indian Academy of Sciences (India)

    Maithili P Dalvi; Malati R Umrani; Mugdha V Joglekar; Anandwardhan A Hardikar

    2009-10-01

    Phenotypic plasticity is a phenomenon that describes the occurrence of 2 or more distinct phenotypes under diverse conditions. This article discusses the work carried out over the past few years in understanding the potential of human pancreatic islet-derived progenitors for cell replacement therapy in diabetes. The phenotypic plasticity exhibited by pancreatic progenitors during reversible epithelial-to-mesenchymal transition (EMT) and possible role of microRNAs in regulation of this process is also presented herein.

  14. Mast cells and human hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Fabio Grizzi; Barbara Franceschini; Maurizio Chiriva-Internati; Young Liu; Paul L. Hermonat; Nicola Dioguardi

    2003-01-01

    AIM: To investigate the density of mast cells (MCs) in human hepatocellular carcinoma (HCC), and to determine whether the MCs density has any correlations with histopathological grading, staging or some baseline patient characteristics.METHODS: Tissue sections of 22 primary HCCs were histochemically stained with toluidine blue, in order to be able to quantify the MCs in and around the neoplasm using a computer-assisted image analysis system. HCC was staged and graded by two independent pathologists. To identify the sinusoidal capillarisation of each specimen 3μm thick sections were histochemically stained with sirius red, and semi-quantitatively evaluated by two independent observers. The data were statistically analysed using Spearman′s correlation and Student′s t-test when appropriate.RESULTS: MCs density did not correlate with the age or sex of the patients, the serum alanine aminotransferase (ALT) or aspartate aminotransferase (AST) levels, or the stage or grade of the HCC. No significant differences were found between the MCs density of the patients with and without hepatitis C virus infection, but they were significantly higher in the specimens showing marked sinusoidal capillarisation.CONCLUSION: The lack of any significant correlation between MCs density and the stage or grade of the neoplastic lesions suggests that there is no causal relationship between MCs recruitment and HCC. However, as capillarisation proceeds concurrently with arterial blood supply during hepatocarcinogenesis, MCs may be considered of primary importance in the transition from sinusoidal to capillary-type endothelial cells and the HCC growth.

  15. MICAL2 is a novel human cancer gene controlling mesenchymal to epithelial transition involved in cancer growth and invasion

    Science.gov (United States)

    Vindigni, Carla; Pucci, Angela; Balsamo, Michele; Libro, Rosaliana; Senchenko, Vera; Dmitriev, Alexey; Jacchetti, Emanuela; Cecchini, Marco; Roviello, Franco; Lai, Michele; Broccoli, Vania; Andreazzoli, Massimiliano; Mazzanti, Chiara M.; Angeloni, Debora

    2016-01-01

    The MICAL (Molecules Interacting with CasL) proteins catalyze actin oxidation-reduction reactions destabilizing F-actin in cytoskeletal dynamics. Here we show for the first time that MICAL2 mRNA is significantly over-expressed in aggressive, poorly differentiated/undifferentiated, primary human epithelial cancers (gastric and renal). Immunohistochemistry showed MICAL2-positive cells on the cancer invasive front and in metastasizing cancer cells inside emboli, but not at sites of metastasis, suggesting MICAL2 expression was 'on' in a subpopulation of primary cancer cells seemingly detaching from the tissue of origin, enter emboli and travel to distant sites, and was turned 'off' upon homing at metastatic sites. In vitro, MICAL2 knock-down resulted in mesenchymal to epithelial transition, reduction of viability, and loss of motility and invasion properties of human cancer cells. Moreover, expression of MICAL2 cDNA in MICAL2-depleted cells induced epithelial to mesenchymal transition. Altogether our data indicate that MICAL2 over-expression is associated with cancer progression and metastatic disease. MICAL2 might be an important regulator of epithelial to mesenchymal transition and therefore a promising target for anti-metastatic therapy. PMID:26689989

  16. MICAL2 is a novel human cancer gene controlling mesenchymal to epithelial transition involved in cancer growth and invasion.

    Science.gov (United States)

    Mariotti, Sara; Barravecchia, Ivana; Vindigni, Carla; Pucci, Angela; Balsamo, Michele; Libro, Rosaliana; Senchenko, Vera; Dmitriev, Alexey; Jacchetti, Emanuela; Cecchini, Marco; Roviello, Franco; Lai, Michele; Broccoli, Vania; Andreazzoli, Massimiliano; Mazzanti, Chiara M; Angeloni, Debora

    2016-01-12

    The MICAL (Molecules Interacting with CasL) proteins catalyze actin oxidation-reduction reactions destabilizing F-actin in cytoskeletal dynamics. Here we show for the first time that MICAL2 mRNA is significantly over-expressed in aggressive, poorly differentiated/undifferentiated, primary human epithelial cancers (gastric and renal). Immunohistochemistry showed MICAL2-positive cells on the cancer invasive front and in metastasizing cancer cells inside emboli, but not at sites of metastasis, suggesting MICAL2 expression was 'on' in a subpopulation of primary cancer cells seemingly detaching from the tissue of origin, enter emboli and travel to distant sites, and was turned 'off' upon homing at metastatic sites. In vitro, MICAL2 knock-down resulted in mesenchymal to epithelial transition, reduction of viability, and loss of motility and invasion properties of human cancer cells. Moreover, expression of MICAL2 cDNA in MICAL2-depleted cells induced epithelial to mesenchymal transition. Altogether our data indicate that MICAL2 over-expression is associated with cancer progression and metastatic disease. MICAL2 might be an important regulator of epithelial to mesenchymal transition and therefore a promising target for anti-metastatic therapy.

  17. Satellite cells in human skeletal muscle plasticity.

    Science.gov (United States)

    Snijders, Tim; Nederveen, Joshua P; McKay, Bryon R; Joanisse, Sophie; Verdijk, Lex B; van Loon, Luc J C; Parise, Gianni

    2015-01-01

    Skeletal muscle satellite cells are considered to play a crucial role in muscle fiber maintenance, repair and remodeling. Our knowledge of the role of satellite cells in muscle fiber adaptation has traditionally relied on in vitro cell and in vivo animal models. Over the past decade, a genuine effort has been made to translate these results to humans under physiological conditions. Findings from in vivo human studies suggest that satellite cells play a key role in skeletal muscle fiber repair/remodeling in response to exercise. Mounting evidence indicates that aging has a profound impact on the regulation of satellite cells in human skeletal muscle. Yet, the precise role of satellite cells in the development of muscle fiber atrophy with age remains unresolved. This review seeks to integrate recent results from in vivo human studies on satellite cell function in muscle fiber repair/remodeling in the wider context of satellite cell biology whose literature is largely based on animal and cell models.

  18. Circuits and methods for determination and control of signal transition rates in electrochemical cells

    Science.gov (United States)

    Jamison, David Kay

    2016-04-12

    A charge/discharge input is for respectively supplying charge to, or drawing charge from, an electrochemical cell. A transition modifying circuit is coupled between the charge/discharge input and a terminal of the electrochemical cell and includes at least one of an inductive constituent, a capacitive constituent and a resistive constituent selected to generate an adjusted transition rate on the terminal sufficient to reduce degradation of a charge capacity characteristic of the electrochemical cell. A method determines characteristics of the transition modifying circuit. A degradation characteristic of the electrochemical cell is analyzed relative to a transition rate of the charge/discharge input applied to the electrochemical cell. An adjusted transition rate is determined for a signal to be applied to the electrochemical cell that will reduce the degradation characteristic. At least one of an inductance, a capacitance, and a resistance is selected for the transition modifying circuit to achieve the adjusted transition rate.

  19. Expressin of cyclooxygenase-2 in human transitional cell carcinoma of bladder%膀胱移行细胞癌COX-2的表达及其意义

    Institute of Scientific and Technical Information of China (English)

    陈合群; 张向阳; 黄振国; 齐范

    2005-01-01

    目的研究环氧合酶-2(cyclooxygenase-2,COX-2)在膀胱移行细胞癌(transitional cell carcino--ma,TCC)组织中的表达及其意义.方法采用免疫组化技术,对49例膀胱TCC石蜡切片和18例正常膀胱黏膜组织进行COX-2蛋白检测,对阳性表达率与肿瘤分级及侵袭性间的相关性与TCC分级及侵袭性进行分析.结果COX-2蛋白在膀胱TCC中表达率为63.2%,正常膀胱黏膜无表达(P<0.01).低分级TCC阳性率36.4%,高分级为92.8%(P<0.05).表浅TCC阳性表达率38.9%,浸润TCC为77.4%(P<0.05).结论COX-2在高分级和侵袭性膀胱TCC中阳性表达率显著上升,可能会成为膀胱肿瘤患者恶性度和判断预后的重要生物学指标,并有可能成为治疗人类TCC的作用靶点.

  20. IMMUNORESPONSES OF HUMANIZED SCID MICE TO HUMAN LUNG CANCER CELLS

    Institute of Scientific and Technical Information of China (English)

    陈力真; 王树蕙; 张云; 王世真

    1996-01-01

    HuPBL-SCID mice were used to explore how they would response to human ttmoor cells of 801/MLC.Living 801/MLC cells appeared to be fetal to the the mice due to the production of human TNF. The huP-BL-SCID rniee did not generate any noticeable amotmt of specific human immunoglobttlin either by single immunization with living 801/MLC cells or by repeated immunization with irradiated 801/MLC cells. Our preliminary experiments with huPBL-SCID mice showed that such chimeras would he a very useful models for tumor immunological researches.

  1. Search for naive human pluripotent stem cells

    Institute of Scientific and Technical Information of China (English)

    Simone Aparecida Siqueira Fonseca; Roberta Montero Costas; Lygia Veiga Pereira

    2015-01-01

    Normal mouse pluripotent stem cells were originallyderived from the inner cell mass (ICM) of blastocystsand shown to be the in vitro equivalent of those preimplantationembryonic cells, and thus were calledembryonic stem cells (ESCs). More than a decade later,pluripotent cells were isolated from the ICM of humanblastocysts. Despite being called human ESCs, thesecells differ significantly from mouse ESCs, includingdifferent morphology and mechanisms of control ofpluripotency, suggesting distinct embryonic originsof ESCs from the two species. Subsequently, mousepluripotent stem cells were established from the ICMderivedepiblast of post-implantation embryos. Thesemouse epiblast stem cells (EpiSCs) are morphologicaland epigenetically more similar to human ESCs. Thisraised the question of whether cells from the humanICM are in a more advanced differentiation stage thantheir murine counterpart, or whether the availableculture conditions were not adequate to maintain thosehuman cells in their in vivo state, leading to a transitioninto EpiSC-like cells in vitro . More recently, novel cultureconditions allowed the conversion of human ESCs intomouse ESC-like cells called naive (or ground state)human ESCs, and the derivation of naive human ESCsfrom blastocysts. Here we will review the characteristicsof each type of pluripotent stem cells, how (andwhether) these relate to different stages of embryonicdevelopment, and discuss the potential implications ofnaive human ESCs in research and therapy.

  2. Endothelial cells derived from human embryonic stem cells

    Science.gov (United States)

    Levenberg, Shulamit; Golub, Justin S.; Amit, Michal; Itskovitz-Eldor, Joseph; Langer, Robert

    2002-04-01

    Human embryonic stem cells have the potential to differentiate into various cell types and, thus, may be useful as a source of cells for transplantation or tissue engineering. We describe here the differentiation steps of human embryonic stem cells into endothelial cells forming vascular-like structures. The human embryonic-derived endothelial cells were isolated by using platelet endothelial cell-adhesion molecule-1 (PECAM1) antibodies, their behavior was characterized in vitro and in vivo, and their potential in tissue engineering was examined. We show that the isolated embryonic PECAM1+ cells, grown in culture, display characteristics similar to vessel endothelium. The cells express endothelial cell markers in a pattern similar to human umbilical vein endothelial cells, their junctions are correctly organized, and they have high metabolism of acetylated low-density lipoprotein. In addition, the cells are able to differentiate and form tube-like structures when cultured on matrigel. In vivo, when transplanted into SCID mice, the cells appeared to form microvessels containing mouse blood cells. With further studies, these cells could provide a source of human endothelial cells that could be beneficial for potential applications such as engineering new blood vessels, endothelial cell transplantation into the heart for myocardial regeneration, and induction of angiogenesis for treatment of regional ischemia.

  3. Modeling of Cancer Stem Cell State Transitions Predicts Therapeutic Response.

    Directory of Open Access Journals (Sweden)

    Mary E Sehl

    Full Text Available Cancer stem cells (CSCs possess capacity to both self-renew and generate all cells within a tumor, and are thought to drive tumor recurrence. Targeting the stem cell niche to eradicate CSCs represents an important area of therapeutic development. The complex nature of many interacting elements of the stem cell niche, including both intracellular signals and microenvironmental growth factors and cytokines, creates a challenge in choosing which elements to target, alone or in combination. Stochastic stimulation techniques allow for the careful study of complex systems in biology and medicine and are ideal for the investigation of strategies aimed at CSC eradication. We present a mathematical model of the breast cancer stem cell (BCSC niche to predict population dynamics during carcinogenesis and in response to treatment. Using data from cell line and mouse xenograft experiments, we estimate rates of interconversion between mesenchymal and epithelial states in BCSCs and find that EMT/MET transitions occur frequently. We examine bulk tumor growth dynamics in response to alterations in the rate of symmetric self-renewal of BCSCs and find that small changes in BCSC behavior can give rise to the Gompertzian growth pattern observed in breast tumors. Finally, we examine stochastic reaction kinetic simulations in which elements of the breast cancer stem cell niche are inhibited individually and in combination. We find that slowing self-renewal and disrupting the positive feedback loop between IL-6, Stat3 activation, and NF-κB signaling by simultaneous inhibition of IL-6 and HER2 is the most effective combination to eliminate both mesenchymal and epithelial populations of BCSCs. Predictions from our model and simulations show excellent agreement with experimental data showing the efficacy of combined HER2 and Il-6 blockade in reducing BCSC populations. Our findings will be directly examined in a planned clinical trial of combined HER2 and IL-6 targeted

  4. Deep sequencing of Trichomonas vaginalis during the early infection of vaginal epithelial cells and amoeboid transition.

    Science.gov (United States)

    Gould, Sven B; Woehle, Christian; Kusdian, Gary; Landan, Giddy; Tachezy, Jan; Zimorski, Verena; Martin, William F

    2013-08-01

    The human pathogen Trichomonas vaginalis has the largest protozoan genome known, potentially encoding approximately 60,000 proteins. To what degree these genes are expressed is not well known and only a few key transcription factors and promoter domains have been identified. To shed light on the expression capacity of the parasite and transcriptional regulation during phase transitions, we deep sequenced the transcriptomes of the protozoan during two environmental stimuli of the early infection process: exposure to oxygen and contact with vaginal epithelial cells. Eleven 3' fragment libraries from different time points after exposure to oxygen only and in combination with human tissue were sequenced, generating more than 150 million reads which mapped onto 33,157 protein coding genes in total and a core set of more than 20,000 genes represented within all libraries. The data uncover gene family expression regulation in this parasite and give evidence for a concentrated response to the individual stimuli. Oxygen stress primarily reveals the parasite's strategies to deal with oxygen radicals. The exposure of oxygen-adapted parasites to human epithelial cells primarily induces cytoskeletal rearrangement and proliferation, reflecting the rapid morphological transition from spindle shaped flagellates to tissue-feeding and actively dividing amoeboids.

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

    Science.gov (United States)

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

    2007-12-21

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

  6. Layering PLGA-based electrospun membranes and cell sheets for engineering cartilage-bone transition.

    Science.gov (United States)

    Mouthuy, P-A; El-Sherbini, Y; Cui, Z; Ye, H

    2016-04-01

    It is now widely acknowledged that implants that have been designed with an effort towards reconstructing the transition between tissues might improve their functionality and integration in vivo. This paper contributes to the development of improved treatment for articular cartilage repair by exploring the potential of the combination of electrospinning technology and cell sheet engineering to create cartilage tissue. Poly(lactic-co-glycolic acid) (PLGA) was used to create the electrospun membranes. The focus being on the cartilage-bone transition, collagen type I and hydroxyapatite (HA) were also added to the scaffolds to increase the histological biocompatibility. Human mesenchymal stem cells (hMSCs) were cultured in thermoresponsive dishes to allow non-enzymatic removal of an intact cell layer after reaching confluence. The tissue constructs were created by layering electrospun membranes with sheets of hMSCs and were cultured under chondrogenic conditions for up to 21 days. High viability was found to be maintained in the multilayered construct. Under chondrogenic conditions, reverse-transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry have shown high expression levels of collagen type X, a form of collagen typically found in the calcified zone of articular cartilage, suggesting an induction of chondrocyte hypertrophy in the PLGA-based scaffolds. To conclude, this paper suggests that layering electrospun scaffolds and cell sheets is an efficient approach for the engineering of tissue transitions, and in particular the cartilage-bone transition. The use of PLGA-based scaffold might be particularly useful for the bone-cartilage reconstruction, since the differentiated tissue constructs seem to show characteristics of calcified cartilage. Copyright © 2013 John Wiley & Sons, Ltd.

  7. Cell reprogramming modelled as transitions in a hierarchy of cell cycles

    Science.gov (United States)

    Hannam, Ryan; Annibale, Alessia; Kühn, Reimer

    2017-10-01

    We construct a model of cell reprogramming (the conversion of fully differentiated cells to a state of pluripotency, known as induced pluripotent stem cells, or iPSCs) which builds on key elements of cell biology viz. cell cycles and cell lineages. Although reprogramming has been demonstrated experimentally, much of the underlying processes governing cell fate decisions remain unknown. This work aims to bridge this gap by modelling cell types as a set of hierarchically related dynamical attractors representing cell cycles. Stages of the cell cycle are characterised by the configuration of gene expression levels, and reprogramming corresponds to triggering transitions between such configurations. Two mechanisms were found for reprogramming in a two level hierarchy: cycle specific perturbations and a noise induced switching. The former corresponds to a directed perturbation that induces a transition into a cycle-state of a different cell type in the potency hierarchy (mainly a stem cell) whilst the latter is a priori undirected and could be induced, e.g. by a (stochastic) change in the cellular environment. These reprogramming protocols were found to be effective in large regimes of the parameter space and make specific predictions concerning reprogramming dynamics which are broadly in line with experimental findings.

  8. The ectopic expression of Snail in MDBK cells does not induce epithelial-mesenchymal transition.

    Science.gov (United States)

    Izawa, Genya; Kobayashi, Wakako; Haraguchi, Misako; Sudo, Akiharu; Ozawa, Masayuki

    2015-07-01

    Epithelial-mesenchymal transition (EMT), a key process in the tumor metastatic cascade, is characterized by the loss of cell-cell junctions and cell polarity, as well as by the acquisition of migratory and invasive properties. However, the precise molecular events that initiate this complex EMT process are poorly understood. Snail expression induces EMT in Madin-Darby canine kidney (MDCK) cells and the human epidermoid carcinoma cell line, A431. Snail is a zinc finger transcription factor and triggers EMT by suppressing E-cadherin expression. In the present study, to broaden our knowledge of Snail‑induced EMT, we generated stable Snail transfectants using Madin-Darby bovine kidney (MDBK) cells. Contrary to the MDCK or A431 cells examined in our previous studies, the MDBK cells transfected with the Snail construct maintained an epithelial morphology and showed no sign of reduced cell-cell adhesiveness compared to the control cells. Consistent with these observations, the downregulation of epithelial marker proteins, e.g. E-cadherin and desmoglein, and the upregulation of mesenchymal marker proteins, e.g., N-cadherin and fibronectin, were not detected. Furthermore, the E-cadherin promoter was not methylated. Therefore, in the MDBK cells, the ectopic expression of Snail failed to induce EMT. As previously demonstrated, in MDCK cells, Snail expression is accompanied by the increased expression of other EMT-inducing transcription factors, e.g., Slug and zinc finger E-box-binding homeobox 1 (ZEB1). However, the MDBK cells transfected with the Snail construct did not exhibit an increased expression of these factors. Thus, it is possible that the failure to upregulate other EMT-related transcription factors may explain the lack of Snail-mediated induction of EMT in MDBK cells.

  9. Inhibition of epithelial to mesenchymal transition in metastatic breast carcinoma cells by c-Src suppression.

    Science.gov (United States)

    Liu, Xiang; Feng, Renqing

    2010-07-01

    The aberrant activation of c-Src regulates multiple functions during tumor progression. This study was conducted to investigate the role of c-Src suppression in epithelial to mesenchymal transition (EMT) process in human breast carcinoma cells. c-Src suppression by PP2 (a Src-family kinase inhibitor) or small interfering RNA (siRNA) was carried out in MCF-7 and MDA-MB-231 cells. Cell migration was analyzed by wound-healing assay. The transcription and protein levels of EMT markers and transcription factors were evaluated by reverse transcription-PCR and Western blot analysis, respectively. The changed cell morphology was photographed by light microscope. c-Src suppression by PP2 or siRNA reversed the mesenchymal-like phenotype in MDA-MB-231 cells. E-cadherin was upregulated in MCF-7 and MDA-MB-231 cells after c-Src suppression, whereas vimentin was downregulated in MDA-MB-231 cells. Slug and SIP1 were downregulated after c-Src suppression in MCF-7 and MDA-MB-231 cells, whereas Twist was unchanged. These results suggest that c-Src suppression by PP2 or siRNA may inhibit EMT through regulation of different transcription factors in breast carcinoma cells that have different metastatic potential.

  10. Mechanisms by Which Interleukin-6 Attenuates Cell Invasion and Tumorigenesis in Human Bladder Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    Ke-Hung Tsui

    2013-01-01

    Full Text Available Interleukin-6, a multifunctional cytokine, contributes to tumor cell proliferation and differentiation. However, the biological mechanisms that are affected by the expression of interleukin-6 in bladder cancer cells remain unclear. We evaluated the effects of interleukin-6 expression in human bladder carcinoma cells in vitro and in vivo. The results of interleukin-6-knockdown experiments in T24 cells and interleukin-6-overexpression experiments in HT1376 cells revealed that interleukin-6 reduced cell proliferation, migration, and invasion in vitro. Xenograft animal studies indicated that the overexpression of interleukin-6 downregulated tumorigenesis of bladder cells and that interleukin-6 knockdown reversed this effect. The results of RT-PCR, immunoblotting, and reporter assays indicated that the overexpression of interleukin-6 upregulated the expression of the mammary serine protease inhibitor (MASPIN, N-myc downstream gene 1 (NDRG1, and KAI1 proteins in HT1376 cells and that interleukin-6 knockdown reduced the expression of these proteins in T24 cells. In addition, results of immunoblotting assays revealed that interleukin-6 modulated epithelial-mesenchymal transitions by upregulating the expression of the E-cadherin, while downregulation N-cadherin and vimentin proteins. Our results suggest that the effects of interleukin-6 on the regulation of epithelial-mesenchymal transitions and the expressions of the MASPIN, NDRG1, and KAI1 genes attribute to the modulation of tumorigenesis in human bladder carcinoma cells.

  11. Trophoblast lineage cells derived from human induced pluripotent stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ying, E-mail: ying.chen@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Wang, Kai; Chandramouli, Gadisetti V.R. [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Knott, Jason G. [Developmental Epigenetics Laboratory, Department of Animal Science, Michigan State University (United States); Leach, Richard, E-mail: Richard.leach@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Department of Obstetrics, Gynecology and Women’s Health, Spectrum Health Medical Group (United States)

    2013-07-12

    Highlights: •Epithelial-like phenotype of trophoblast lineage cells derived from human iPS cells. •Trophoblast lineage cells derived from human iPS cells exhibit trophoblast function. •Trophoblasts from iPS cells provides a proof-of-concept in regenerative medicine. -- Abstract: Background: During implantation, the blastocyst trophectoderm attaches to the endometrial epithelium and continues to differentiate into all trophoblast subtypes, which are the major components of a placenta. Aberrant trophoblast proliferation and differentiation are associated with placental diseases. However, due to ethical and practical issues, there is almost no available cell or tissue source to study the molecular mechanism of human trophoblast differentiation, which further becomes a barrier to the study of the pathogenesis of trophoblast-associated diseases of pregnancy. In this study, our goal was to generate a proof-of-concept model for deriving trophoblast lineage cells from induced pluripotency stem (iPS) cells from human fibroblasts. In future studies the generation of trophoblast lineage cells from iPS cells established from patient’s placenta will be extremely useful for studying the pathogenesis of individual trophoblast-associated diseases and for drug testing. Methods and results: Combining iPS cell technology with BMP4 induction, we derived trophoblast lineage cells from human iPS cells. The gene expression profile of these trophoblast lineage cells was distinct from fibroblasts and iPS cells. These cells expressed markers of human trophoblasts. Furthermore, when these cells were differentiated they exhibited invasive capacity and placental hormone secretive capacity, suggesting extravillous trophoblasts and syncytiotrophoblasts. Conclusion: Trophoblast lineage cells can be successfully derived from human iPS cells, which provide a proof-of-concept tool to recapitulate pathogenesis of patient placental trophoblasts in vitro.

  12. Andrographolide suppresses epithelial mesenchymal transition by inhibition of MAPK signalling pathway in lens epithelial cells

    Indian Academy of Sciences (India)

    Forum Kayastha; Kaid Johar; Devarshi Gajjar; Anshul Arora; Hardik Madhu; Darshini Ganatra; Abhay Vasavada

    2015-06-01

    Epithelial mesenchymal transition (EMT) of lens epithelial cells (LECs) may contribute to the development of posterior capsular opacification (PCO), which leads to visual impairment. Andrographolide has been shown to have therapeutic potential against various cancers. However, its effect on human LECs is still unknown. The purpose of this study is to evaluate the effect of andrographolide on EMT induced by growth factors in the fetal human lens epithelial cell line (FHL 124). Initially the LECs were treated with growth factors (TGF-2 and bFGF) to induce EMT. Subsequently these EMT-induced cells were treated with andrographolide at 100 and 500 nM concentrations for 24 h. Our results showed that FHL 124 cells treated with growth factors had a significant decrease in protein and m-RNA levels of epithelial markers pax6 and E-Cadherin. After administering andrographolide, these levels significantly increased. It was noticed that EMT markers -SMA, fibronectin and collagen IV significantly decreased after treatment with andrographolide when compared to the other group. Treatment with andrographolide significantly inhibited phosphorylation of ERK and JNK. Cell cycle analysis showed that andrographolide did not arrest cells at G0/G1 or G2/M at tested concentrations. Our findings suggest that andrographolide helps sustain epithelial characteristics by modulating EMT markers and inhibiting the mitogen-activated protein kinase (MAPK) signalling pathway in LECs. Hence it can prove to be useful in curbing EMT-mediated PCO.

  13. Human testis-derived embryonic stem cell-like cells are not pluripotent, but possess potential of mesenchymal progenitors

    NARCIS (Netherlands)

    Chikhovskaya, J.V.; Jonker, M.J.; Meissner, A.; Breit, T.M.; Repping, S.; van Pelt, A.M.M.

    2012-01-01

    BACKGROUND Spontaneous in vitro transition of undifferentiated spermatogonia into the pluripotent cell state has been achieved using neonatal and adult mouse testis tissue. In an effort to establish an analogous source of human patient-specific pluripotent stem cells, several research groups have de

  14. Effects of Combined siRNA-TR and-TERT on Telomerase Activity and Growth of Bladder Transitional Cell Cancer BIU-87 Cells

    Institute of Scientific and Technical Information of China (English)

    程文; 位志峰; 高建平; 张征宇; 葛京平; 景抗震; 徐锋; 解鹏

    2010-01-01

    The effects of combined RNA interference(RNAi) of human telomerase RNA(hTR) and human telomerase reverse transcriptase(hTERT) genes on telomerase activity in a bladder cancer cell line(BIU-87 cells) were investigated by using gene chip technology in vitro with an attempt to evaluate the role of RNAi in the gene therapy of bladder transitional cell cancer(BTCC).Three TR-specific double-stranded small interfering RNAs(siRNAs) and three TERT-specific double-stranded siRNAs were designed to target different reg...

  15. PAI-1 transcriptional regulation during the G0 --> G1 transition in human epidermal keratinocytes.

    Science.gov (United States)

    Qi, Li; Allen, Rosalie R; Lu, Qi; Higgins, Craig E; Garone, Rosemarie; Staiano-Coico, Lisa; Higgins, Paul J

    2006-10-01

    Plasminogen activator inhibitor type-1 (PAI-1) is the major negative regulator of the plasmin-dependent pericellular proteolytic cascade. PAI-1 gene expression is normally growth state regulated but frequently elevated in chronic fibroproliferative and neoplastic diseases affecting both stromal restructuring and cellular migratory activities. Kinetic modeling of cell cycle transit in synchronized human keratinocytes (HaCaT cells) indicated that PAI-1 transcription occurred early after serum stimulation of quiescent (G0) cells and prior to entry into a cycling G1 condition. PAI-1 repression (in G0) was associated with upstream stimulatory factor-1 (USF-1) occupancy of two consensus E box motifs (5'-CACGTG-3') at the PE1 and PE2 domains in the PF1 region (nucleotides -794 to -532) of the PAI-1 promoter. Chromatin immunoprecipitation (ChIP) analysis established that the PE1 and PE2 site E boxes were occupied by USF-1 in quiescent cells and by USF-2 in serum-activated, PAI-1-expressing keratinocytes. This reciprocal and growth state-dependent residence of USF family members (USF-1 vs. USF-2) at PE1/PE2 region chromatin characterized the G0 --> G1 transition period and the transcriptional status of the PAI-1 gene. A consensus E box motif was required for USF/E box interactions, as a CG --> AT substitution at the two central nucleotides inhibited formation of USF/probe complexes. The 5' flanking sites (AAT or AGAC) in the PE2 segment were not necessary for USF binding. USF recognition of the PE1/PE2 region E box sites required phosphorylation with several potential involved residues, including T153, maping to the USF-specific region (USR). A T153A substitution in USF-1 did not repress serum-induced PAI-1 expression whereas the T153D mutant was an effective suppressor. As anticipated from the ChIP results, transfection of wild-type USF-2 failed to inhibit PAI-1 induction. Collectively, these data suggest that USF family members are important regulators of PAI-1 gene

  16. Human Neural Cell-Based Biosensor

    Science.gov (United States)

    2013-05-28

    including incubation with factors such as SHH ) and proceed to Human Neural Progenitor Cells Dopaminergic Differentiation β-III Tubulin/TH...exposure in human embryonic stem cells. J Recept Signal Transduct Res. 2011 Jun;31(3):206-13. Gerwe BA, Angel PM, West FD, Hasneen K, Young A

  17. Recurrence patterns of bladder transitional cell carcinoma after radical cystectomy

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Bohyun; Choi, Hyuck Jae; Kim, Mi-hyun; Cho, Kyung-Sik [Dept. of Radiology, Asan Medical Center, Univ. of Ulsan, Seoul (Korea, Republic of); E-mail: choihj@amc.seoul.kr

    2012-10-15

    Background Multidetector computed tomography (MDCT) is widely accepted as an effective imaging modality in monitoring for bladder cancer recurrence after radical cystectomy. Elucidating the pattern of bladder cancer recurrence on CT can increase the diagnostic accuracy. Purpose To evaluate the recurrence patterns of transitional cell carcinoma of the bladder and the factors associated with cancer recurrence. Material and Methods One hundred and forty-nine consecutive patients (mean age, 66.55 years; range, 32-86 years) who underwent preoperative contrast-enhanced CT and radical cystectomy were included in this study. The presence, site, and time of tumor recurrence were recorded retrospectively by two radiologists in a consensus fashion. The association of tumor recurrence and tumor factors (T stage, lymph node metastasis, nuclear grade, and tumor diameter) were also evaluated using multiple logistic regression analysis and Kaplan-Meier statistics. Results Tumor recurrence occurred in 60 patients (40.3%) with a mean time of 14 months (range, 1-64 months). The sites of recurrence included the operation site (n = 20), lymph node (n = 20), bone (n = 11), liver (n = 6), lung (n = 5), upper urinary tract (n = 4), colon (n = 3), adrenal gland (n = 2), peritoneum (n = 1), abdominal wall (n = 1), psoas muscle (n = 1), and penile skin (n = 1). Tumor recurrence was found to be associated with advanced T stage (P = 0.002) and lymph node metastasis (P < 0.001). Conclusion Transitional cell carcinomas of the bladder recur more frequently at the operation site and lymph node, and T-stage and lymph node metastasis are closely associated with tumor recurrence.

  18. Ipsilateral synchronous renal pelvic transitional cell carcinoma, squamous cell carcinoma and adenocarcinoma

    Institute of Scientific and Technical Information of China (English)

    韩平; 魏强; 石明; 杨宇如

    2004-01-01

    @@ Reports of multiple synchronous primary renal neoplasms in the literature are rare. Although primary renal tumors of 2 distinctively dissimilar origins have been sporadically described,1-6 to our knowledge there have been no reported cases of triple primary renal neoplasms in the same kidney. Here we report a very rare case of ipsilateral synchronous renal pelvic transitional cell carcinoma, squamous cell carcinoma and adenocarcinoma with marked hydronephrosis and multiple stones in the same kidney.

  19. FGFR signaling maintains a drug persistent cell population following epithelial-mesenchymal transition.

    Science.gov (United States)

    Brown, Wells S; Akhand, Saeed Salehin; Wendt, Michael K

    2016-12-13

    An emerging characteristic of drug resistance in cancer is the induction of epithelial-mesenchymal transition (EMT). However, the mechanisms of EMT-mediated drug resistance remain poorly defined. Therefore, we conducted long-term treatments of human epidermal growth factor receptor-2 (Her2)-transformed breast cancer cells with either the EGFR/Her2 kinase inhibitor, Lapatinib or TGF-β, a known physiological inducer of EMT. Both of these treatment regimes resulted in robust EMT phenotypes, but upon withdrawal a subpopulation of TGF-β induced cells readily underwent mesenchymal-epithelial transition, where as Lapatinib-induced cells failed to reestablish an epithelial population. The mesenchymal population that remained following TGF-β stimulation and withdrawal was quickly selected for during subsequent Lapatinib treatment, manifesting in inherent drug resistance. The Nanostring cancer progression gene panel revealed a dramatic upregulation of fibroblast growth factor receptor 1 (FGFR1) and its cognate ligand FGF2 in both acquired and inherent resistance. Mechanistically, FGF:Erk1/2 signaling functions to stabilize the EMT transcription factor Twist and thus maintain the mesenchymal and drug resistant phenotype. Finally, Lapatinib resistant cells could be readily eliminated using recently characterized covalent inhibitors of FGFR. Overall our data demonstrate that next-generation targeting of FGFR can be used in combination with Her2-targeted therapies to overcome resistance in this breast cancer subtype.

  20. Colonic transit time relates to bacterial metabolism and mucosal turnover in the human gut

    DEFF Research Database (Denmark)

    Roager, Henrik Munch; Hansen, Lea Benedicte Skov; Bahl, Martin Iain

    transit time and the gut microbial composition and metabolism, we assessed the colonic transit time of 98 subjects using radiopaque markers, and profiled their gut microbiota by16S rRNA gene sequencingand their urine metabolome by ultra performance liquid chromatography mass spectrometry. Based......Little is known about how colonic transit time relates to human colonic metabolism, and its importance for host health, although stool consistency, a proxy for colonic transit time, has recently been negatively associated with gut microbial richness. To address the relationships between colonic...... on correlation analyses,we show that colonic transit time is associated with overall gutmicrobial composition, diversity and metabolism. A relatively prolonged colonic transit time associates with high microbial species richness and a shift in colonic metabolismfrom carbohydrate fermentation to protein...

  1. Colonic transit time is related to bacterial metabolism and mucosal turnover in the human gut

    DEFF Research Database (Denmark)

    Roager, Henrik Munch; Hansen, Lea Benedicte Skov; Bahl, Martin Iain

    transit time and the gut microbial composition and metabolism, we assessed the colonic transit time of 98 subjects using radiopaque markers, and profiled their gut microbiota by16S rRNA gene sequencing and their urine metabolome by ultra performance liquid chromatography mass spectrometry. Based......Little is known about how colonic transit time relates to human colonic metabolism, and its importance for host health, although stool consistency, a proxy for colonic transit time, has recently been negatively associated with gut microbial richness. To address the relationships between colonic...... on correlation analyses, we show that colonic transit time is associated with overall gut microbial composition, diversity and metabolism. A relatively prolonged colonic transit time associates with high microbial species richness and a shift in colonic metabolism from carbohydrate fermentation to protein...

  2. Cortical network from human embryonic stem cells

    OpenAIRE

    2010-01-01

    Abstract The connection of embryonic stem cell technology and developmental biology provides valuable tools to decipher the mechanisms underlying human brain development and diseases, especially among neuronal populations, that are not readily available in primary cultures. It is obviously the case of neurons forming the human cerebral cortex. In the images that are presented, the neurons were generated in vitro from human embryonic stem cells via forebrain-like progenitors. Maintained in cul...

  3. Downregulation of β-catenin decreases the tumorigenicity, but promotes epithelial-mesenchymal transition in breast cancer cells

    Directory of Open Access Journals (Sweden)

    Kai Cai

    2014-01-01

    Full Text Available Background: Wnt/β-catenin signaling pathway plays a key role in human breast cancer progression. In this study, we down regulated β-catenin expression in human breast cancer MDA-MB-231 cells and investigated the effect of β-catenin knockdown on the cell biological characteristics. Materials and Methods: The recombinant plasmids of pSUPER-enhancement green fluorescent protein 1 (EGFP1-scrabble-β-catenin-short hairpin ribonucleic acid (shRNA and pSUPER-EGFP1-β-catenin-shRNA-1 were transfected into MDA-MB-231 cells, respectively, and the stably transfected cells were isolated from G418 selected clones. The β-catenin gene silenced efficiency was measured by quantitative reverse transcriptase polymerase chain reaction (QRT-PCR and Western blot. The biological characteristics of MDA-MB-231 cells with down regulated β-catenin were evaluated by analyzing cell proliferation, clonogenicity, cell mobility and tumorigenicity. The expression of E-cadherin and Vimentin was concurrently detected by QRT-PCR. Results: The β-catenin-shRNA-1 stably transfected MDA-MB-231 cells significantly decreased β-catenin expression, cell proliferation, clonogenicity, and tumorigenicity in Balb/c nude mice compared with the MDA-MB-231 cells transfected with pSUPER-EGFP1-scrabble-β-catenin-shRNA. Interestingly, knockdown of β-catenin led to the reduction of epithelial E-cadherin expression, the increase of cell mobility and mesenchymal vimentin expression in MDA-MB-231 cells, indicating an epithelial to mesenchymal transition. Conclusion: Knockdown of β-catenin expression in human breast cancer MDA-MB-231 cells inhibits cell tumorigenicity in mice, but promotes cell epithelial-mesenchymal transition.

  4. TRPP2 Enhances Metastasis by Regulating Epithelial-Mesenchymal Transition in Laryngeal Squamous Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Kaile Wu

    2016-11-01

    Full Text Available Background/Aim: Surgery and chemotherapy treatments of human laryngeal squamous cell carcinoma (HLSCC may fail due to metastasis, in which epithelial-mesenchymal transition (EMT plays an important role. TRPP2, a nonselective cation channel, is expressed in various cell types and participates in many biological processes. Here, we show that TRPP2 enhanced metastasis by regulating EMT. Methods: We used immunohistochemistry, western blotting, Ca2+ imaging, transwell and wound healing assays to investigate TRPP2 expression levels in HLSCC tissue, and the role of TRPP2 in invasion and metastasis of a human laryngocarcinoma cell line (Hep2 cell. Results: We found that TRPP2 protein expression levels were significantly increased in HLSCC tissue; higher TRPP2 levels were associated with decreased patient survival time and degree of differentiation and advanced clinical stage. Knockdown of TRPP2 by transfection with TRPP2 siRNA markedly suppressed ATP-induced Ca2+ release, wound healing, and cell invasion in Hep2 cells. Moreover, TRPP2 siRNA significantly decreased vimentin expression but increased E-cadherin expression in Hep2 cells. In the EMT signalling pathway, TRPP2 siRNA significantly decreased Smad4, STAT3, SNAIL, SLUG and TWIST expression in Hep2 cells. Conclusion: We revealed a previously unknown function of TRPP2 in cancer development and a TRPP2-dependent mechanism underlying laryngocarcinoma cell invasion and metastasis. Our results suggest that TRPP2 may be used as a biomarker for evaluating patient prognosis and as a novel therapeutic target in HLSCC.

  5. Role of epithelial-mesenchymal transition in the enrichment of colorectal cancer stem cells

    Directory of Open Access Journals (Sweden)

    Jia-ping CHENG

    2016-10-01

    Full Text Available Objective  To explore whether the enrichment of cancer stem cells (CSCs in colorectal cancer by suspension culture method is involved with epithelial-mesenchymal transition (EMT. Methods  3D microspheres were cultured by suspension culture method to human colorectal cancer SW620 cells. The 3D microspheres and SW620 cells were used as the research objects. To clarify whether 3D microspheres were enriched with CSCs, we made tumorigenicity experiments in NOD/SCID mice, soft agar cloning experiments, and detected the expression levels of cancer stem cells markers CD44 and Ep-CAM by flow cytometry or by Western blotting. The protein expression levels of EMT markers such as E-cadherin, N-cadherin and vimentin were detected by Western blotting. Results  Compared with the parental SW620 cells, colony formation in vitro (P<0.01 and tumorigenicity in NOD/SCID mice were significantly enhanced, the percentage of CD44-positive cells and Ep-CAM protein expression levels was significantly increased (P<0.01 in 3D microspheres. The protein expression level of epithelial marker E-cadherin was obviously increased (P<0.01, while the protein expression levels of mesenchymal markers N-cadherin and vimentin were significantly decreased (P<0.01. Conclusions  Colorectal cancer stem cells can be enriched by suspension culture method, and the process may be related to EMT. DOI: 10.11855/j.issn.0577-7402.2016.09.03

  6. Human hematopoietic cell culture, transduction, and analyses

    DEFF Research Database (Denmark)

    Bonde, Jesper; Wirthlin, Louisa; Kohn, Donald B;

    2008-01-01

    This unit provides methods for introducing genes into human hematopoietic progenitor cells. The Basic Protocol describes isolation of CD34(+) cells, transduction of these cells with a retroviral vector on fibronectin-coated plates, assaying the efficiency of transduction, and establishing long...

  7. Human embryonic stem cells derived by somatic cell nuclear transfer.

    Science.gov (United States)

    Tachibana, Masahito; Amato, Paula; Sparman, Michelle; Gutierrez, Nuria Marti; Tippner-Hedges, Rebecca; Ma, Hong; Kang, Eunju; Fulati, Alimujiang; Lee, Hyo-Sang; Sritanaudomchai, Hathaitip; Masterson, Keith; Larson, Janine; Eaton, Deborah; Sadler-Fredd, Karen; Battaglia, David; Lee, David; Wu, Diana; Jensen, Jeffrey; Patton, Phillip; Gokhale, Sumita; Stouffer, Richard L; Wolf, Don; Mitalipov, Shoukhrat

    2013-06-06

    Reprogramming somatic cells into pluripotent embryonic stem cells (ESCs) by somatic cell nuclear transfer (SCNT) has been envisioned as an approach for generating patient-matched nuclear transfer (NT)-ESCs for studies of disease mechanisms and for developing specific therapies. Past attempts to produce human NT-ESCs have failed secondary to early embryonic arrest of SCNT embryos. Here, we identified premature exit from meiosis in human oocytes and suboptimal activation as key factors that are responsible for these outcomes. Optimized SCNT approaches designed to circumvent these limitations allowed derivation of human NT-ESCs. When applied to premium quality human oocytes, NT-ESC lines were derived from as few as two oocytes. NT-ESCs displayed normal diploid karyotypes and inherited their nuclear genome exclusively from parental somatic cells. Gene expression and differentiation profiles in human NT-ESCs were similar to embryo-derived ESCs, suggesting efficient reprogramming of somatic cells to a pluripotent state.

  8. MicroRNA Regulation of Human Breast Cancer Stem Cells

    Directory of Open Access Journals (Sweden)

    Yohei Shimono

    2015-12-01

    Full Text Available MicroRNAs (miRNAs are involved in virtually all biological processes, including stem cell maintenance, differentiation, and development. The dysregulation of miRNAs is associated with many human diseases including cancer. We have identified a set of miRNAs differentially expressed between human breast cancer stem cells (CSCs and non-tumorigenic cancer cells. In addition, these miRNAs are similarly upregulated or downregulated in normal mammary stem/progenitor cells. In this review, we mainly describe the miRNAs that are dysregulated in human breast CSCs directly isolated from clinical specimens. The miRNAs and their clusters, such as the miR-200 clusters, miR-183 cluster, miR-221-222 cluster, let-7, miR-142 and miR-214, target the genes and pathways important for stem cell maintenance, such as the self-renewal gene BMI1, apoptosis, Wnt signaling, Notch signaling, and epithelial-to-mesenchymal transition. In addition, the current evidence shows that metastatic breast CSCs acquire a phenotype that is different from the CSCs in a primary site. Thus, clarifying the miRNA regulation of the metastatic breast CSCs will further advance our understanding of the roles of human breast CSCs in tumor progression.

  9. Human Neuroepithelial Cells Express NMDA Receptors

    Directory of Open Access Journals (Sweden)

    Cappell B

    2003-11-01

    Full Text Available Abstract L-glutamate, an excitatory neurotransmitter, binds to both ionotropic and metabotropic glutamate receptors. In certain parts of the brain the BBB contains two normally impermeable barriers: 1 cerebral endothelial barrier and 2 cerebral epithelial barrier. Human cerebral endothelial cells express NMDA receptors; however, to date, human cerebral epithelial cells (neuroepithelial cells have not been shown to express NMDA receptor message or protein. In this study, human hypothalamic sections were examined for NMDA receptors (NMDAR expression via immunohistochemistry and murine neuroepithelial cell line (V1 were examined for NMDAR via RT-PCR and Western analysis. We found that human cerebral epithelium express protein and cultured mouse neuroepithelial cells express both mRNA and protein for the NMDA receptor. These findings may have important consequences for neuroepithelial responses during excitotoxicity and in disease.

  10. Fuel Cell Buses in U.S. Transit Fleets: Current Status 2011

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, L.; Chandler, K.; Gikakis, C.

    2011-11-01

    This status report, fifth in a series of annual status reports from the U.S. Department of Energy's National Renewable Energy Laboratory (NREL), discusses the achievements and challenges of fuel cell propulsion for transit and summarizes the introduction of fuel cell transit buses in the United States. Progress this year includes an increase in the number of fuel cell electric buses (FCEBs), from 15 to 25, operating at eight transit agencies, as well as increased diversity of the fuel cell design options for transit buses. The report also provides an analysis of the combined results from fuel cell transit bus demonstrations evaluated by NREL with a focus on the most recent data through July 2011 including fuel cell power system reliability and durability; fuel economy; roadcall; and hydrogen fueling results. These evaluations cover 22 of the 25 FCEBs currently operating.

  11. Bone formation induced in mouse thigh by cultured human cells.

    Science.gov (United States)

    Anderson, H C; Coulter, P R

    1967-04-01

    Cultured FL human amnion cells injected intramuscularly into cortisone-conditioned mice proliferate to form discrete nodules which become surrounded by fibroblasts. Within 12 days, fibroblastic zones differentiate into cartilage which calcifies to form bone. Experiments were conducted to test the hypothesis that FL cells behave as an inductor of bone formation. In the electron microscope, FL cells were readily distinguished from surrounding fibroblasts. Transitional forms between the two cell types were not recognized. Stains for acid mucopolysaccharides emphasized the sharp boundary between metachromatic fibroblastic and cartilaginous zones and nonmetachromatic FL cells. (35)S was taken up preferentially by fibroblasts and chondrocytes and then deposited extracellularly in a manner suggesting active secretion of sulfated mucopolysaccharides. FL cells showed negligible (35)S utilization and secretion. FL cells, labeled in vitro with thymidine-(3)H, were injected and followed radioautographically, during bone formation. Nuclear label of injected FL cells did not appear in adjacent fibroblasts in quantities sufficient to indicate origin of the latter from FL cells. The minimal fibroblast nuclear labeling seen may represent reutilization of label from necrotic FL cells. It is suggested that FL cells injected into the mouse thigh induced cartilage and bone formation by host fibroblasts.

  12. Proteinase-activated receptor 4 stimulation-induced epithelial-mesenchymal transition in alveolar epithelial cells

    Directory of Open Access Journals (Sweden)

    Araki Hiromasa

    2007-04-01

    Full Text Available Abstract Background Proteinase-activated receptors (PARs; PAR1–4 that can be activated by serine proteinases such as thrombin and neutrophil catepsin G are known to contribute to the pathogenesis of various pulmonary diseases including fibrosis. Among these PARs, especially PAR4, a newly identified subtype, is highly expressed in the lung. Here, we examined whether PAR4 stimulation plays a role in the formation of fibrotic response in the lung, through alveolar epithelial-mesenchymal transition (EMT which contributes to the increase in myofibroblast population. Methods EMT was assessed by measuring the changes in each specific cell markers, E-cadherin for epithelial cell, α-smooth muscle actin (α-SMA for myofibroblast, using primary cultured mouse alveolar epithelial cells and human lung carcinoma-derived alveolar epithelial cell line (A549 cells. Results Stimulation of PAR with thrombin (1 U/ml or a synthetic PAR4 agonist peptide (AYPGKF-NH2, 100 μM for 72 h induced morphological changes from cobblestone-like structure to elongated shape in primary cultured alveolar epithelial cells and A549 cells. In immunocytochemical analyses of these cells, such PAR4 stimulation decreased E-cadherin-like immunoreactivity and increased α-SMA-like immunoreactivity, as observed with a typical EMT-inducer, tumor growth factor-β (TGF-β. Western blot analyses of PAR4-stimulated A549 cells also showed similar changes in expression of these EMT-related marker proteins. Such PAR4-mediated changes were attenuated by inhibitors of epidermal growth factor receptor (EGFR kinase and Src. PAR4-mediated morphological changes in primary cultured alveolar epithelial cells were reduced in the presence of these inhibitors. PAR4 stimulation increased tyrosine phosphorylated EGFR or tyrosine phosphorylated Src level in A549 cells, and the former response being inhibited by Src inhibitor. Conclusion PAR4 stimulation of alveolar epithelial cells induced epithelial

  13. miR-506 regulates epithelial mesenchymal transition in breast cancer cell lines.

    Science.gov (United States)

    Arora, Himanshu; Qureshi, Rehana; Park, Woong-Yang

    2013-01-01

    Epithelial-mesenchymal transition (EMT) is an important parameter related to breast cancer survival. Among several microRNAs predicted to target EMT-related genes, miR-506 is a novel miRNA found to be significantly related to breast cancer patient survival in a meta-analysis. miR-506 suppressed the expression of mesenchymal genes such as Vimentin, Snai2, and CD151 in MDA-MB-231 human breast cancer cell line. Moreover, NF-κB bound to the upstream promoter region of miR-506 to suppress transcription. Overexpression of miR-506 inhibited TGFβ-induced EMT and suppressed adhesion, invasion, and migration of MDA-MB-231 cells. From these results, we concluded that miR-506 plays a key role in the process of EMT through posttranslational control of EMT-related genes.

  14. miR-506 regulates epithelial mesenchymal transition in breast cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Himanshu Arora

    Full Text Available Epithelial-mesenchymal transition (EMT is an important parameter related to breast cancer survival. Among several microRNAs predicted to target EMT-related genes, miR-506 is a novel miRNA found to be significantly related to breast cancer patient survival in a meta-analysis. miR-506 suppressed the expression of mesenchymal genes such as Vimentin, Snai2, and CD151 in MDA-MB-231 human breast cancer cell line. Moreover, NF-κB bound to the upstream promoter region of miR-506 to suppress transcription. Overexpression of miR-506 inhibited TGFβ-induced EMT and suppressed adhesion, invasion, and migration of MDA-MB-231 cells. From these results, we concluded that miR-506 plays a key role in the process of EMT through posttranslational control of EMT-related genes.

  15. Mitochondrial permeability transition increases reactive oxygen species production and induces DNA fragmentation in human spermatozoa.

    Science.gov (United States)

    Treulen, Favián; Uribe, Pamela; Boguen, Rodrigo; Villegas, Juana V

    2015-04-01

    Does mitochondrial permeability transition (MPT) induced by calcium overload cause reactive oxygen species (ROS) production and DNA fragmentation in human spermatozoa? Studies conducted in vitro suggest that in human spermatozoa, MPT occurs in response to intracellular calcium increase and is associated with mitochondrial membrane potential (ΔΨm) dissipation, increased ROS production and DNA fragmentation. Oxidative stress is a major cause of defective sperm function in male infertility. By opening calcium-dependent pores in the inner mitochondrial membrane (IMM), MPT causes, among other things, increased ROS production and ΔΨm dissipation in somatic cells. MPT as a mechanism for generating oxidative stress and DNA fragmentation in human spermatozoa has not been studied. Human sperm were exposed to ionomycin for 1.5 h (n = 8) followed by analysis of sperm IMM permeability, ΔΨm, ROS production and DNA fragmentation. To evaluate the MPT in sperm cells, the calcein-AM and cobalt chloride method was used. The ΔΨm was evaluated by JC-1 staining, intracellular ROS production was evaluated with dihydroethidium and DNA fragmentation was evaluated by a modified TUNEL assay. Measurements were performed by fluorescence microscopy, confocal laser microscopy and flow cytometry. Decreased calcein fluorescence after treatment with ionomycin (P fragmentation. ROS production occurred prior to the decrease in ΔΨm. The study was carried out in vitro using motile sperm from healthy donors; tests on sperm from infertile patients were not carried out. We propose that the MPT, due to pores opening in sperm IMM, is an important mechanism of increased ROS and DNA fragmentation. Therefore, agents that modulate the opening of these pores might contribute to the prevention of damage by oxidative stress in human spermatozoa. This study was funded by grant DI12-0102 from the Universidad de La Frontera (J.V.V.) and a doctoral scholarship from CONICYT Chile (F.T.). The authors disclose

  16. Fertility, Human Capital, and Economic Growth over the Demographic Transition.

    Science.gov (United States)

    Lee, Ronald; Mason, Andrew

    2010-05-01

    Do low fertility and population aging lead to economic decline if couples have fewer children, but invest more in each child? By addressing this question, this article extends previous work in which the authors show that population aging leads to an increased demand for wealth that can, under some conditions, lead to increased capital per worker and higher per capita consumption. This article is based on an overlapping generations (OLG) model which highlights the quantity-quality tradeoff and the links between human capital investment and economic growth. It incorporates new national level estimates of human capital investment produced by the National Transfer Accounts project. Simulation analysis is employed to show that, even in the absence of the capital dilution effect, low fertility leads to higher per capita consumption through human capital accumulation, given plausible model parameters.

  17. Cell Cycle Progression of Human Cells Cultured in Rotating Bioreactor

    Science.gov (United States)

    Parks, Kelsey

    2009-01-01

    Space flight has been shown to alter the astronauts immune systems. Because immune performance is complex and reflects the influence of multiple organ systems within the host, scientists sought to understand the potential impact of microgravity alone on the cellular mechanisms critical to immunity. Lymphocytes and their differentiated immature form, lymphoblasts, play an important and integral role in the body's defense system. T cells, one of the three major types of lymphocytes, play a central role in cell-mediated immunity. They can be distinguished from other lymphocyte types, such as B cells and natural killer cells by the presence of a special receptor on their cell surface called T cell receptors. Reported studies have shown that spaceflight can affect the expression of cell surface markers. Cell surface markers play an important role in the ability of cells to interact and to pass signals between different cells of the same phenotype and cells of different phenotypes. Recent evidence suggests that cell-cycle regulators are essential for T-cell function. To trigger an effective immune response, lymphocytes must proliferate. The objective of this project is to investigate the changes in growth of human cells cultured in rotating bioreactors and to measure the growth rate and the cell cycle distribution for different human cell types. Human lymphocytes and lymphoblasts will be cultured in a bioreactor to simulate aspects of microgravity. The bioreactor is a cylindrical culture vessel that incorporates the aspects of clinostatic rotation of a solid fluid body around a horizontal axis at a constant speed, and compensates gravity by rotation and places cells within the fluid body into a sustained free-fall. Cell cycle progression and cell proliferation of the lymphocytes will be measured for a number of days. In addition, RNA from the cells will be isolated for expression of genes related in cell cycle regulations.

  18. Cell Cycle Progression of Human Cells Cultured in Rotating Bioreactor

    Science.gov (United States)

    Parks, Kelsey

    2009-01-01

    Space flight has been shown to alter the astronauts immune systems. Because immune performance is complex and reflects the influence of multiple organ systems within the host, scientists sought to understand the potential impact of microgravity alone on the cellular mechanisms critical to immunity. Lymphocytes and their differentiated immature form, lymphoblasts, play an important and integral role in the body's defense system. T cells, one of the three major types of lymphocytes, play a central role in cell-mediated immunity. They can be distinguished from other lymphocyte types, such as B cells and natural killer cells by the presence of a special receptor on their cell surface called T cell receptors. Reported studies have shown that spaceflight can affect the expression of cell surface markers. Cell surface markers play an important role in the ability of cells to interact and to pass signals between different cells of the same phenotype and cells of different phenotypes. Recent evidence suggests that cell-cycle regulators are essential for T-cell function. To trigger an effective immune response, lymphocytes must proliferate. The objective of this project is to investigate the changes in growth of human cells cultured in rotating bioreactors and to measure the growth rate and the cell cycle distribution for different human cell types. Human lymphocytes and lymphoblasts will be cultured in a bioreactor to simulate aspects of microgravity. The bioreactor is a cylindrical culture vessel that incorporates the aspects of clinostatic rotation of a solid fluid body around a horizontal axis at a constant speed, and compensates gravity by rotation and places cells within the fluid body into a sustained free-fall. Cell cycle progression and cell proliferation of the lymphocytes will be measured for a number of days. In addition, RNA from the cells will be isolated for expression of genes related in cell cycle regulations.

  19. Lentiviral Vector Mediated Claudin1 Silencing Inhibits Epithelial to Mesenchymal Transition in Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Xianqi Zhao

    2015-06-01

    Full Text Available Breast cancer has a high incidence and mortality rate worldwide. Several viral vectors including lentiviral, adenoviral and adeno-associated viral vectors have been used in gene therapy for various forms of human cancer, and have shown promising effects in controlling tumor development. Claudin1 (CLDN1 is a member of the tetraspan transmembrane protein family that plays a major role in tight junctions and is associated with tumor metastasis. However, the role of CLDN1 in breast cancer is largely unexplored. In this study, we tested the therapeutic potential of silencing CLDN1 expression in two breast cancer (MDA-MB-231 and MCF7 cell lines using lentiviral vector mediated RNA interference. We found that a CLDN1 short hairpin (shRNA construct efficiently silenced CLDN1 expression in both breast cancer cell lines, and CLDN1 knockdown resulted in reduced cell proliferation, survival, migration and invasion. Furthermore, silencing CLDN1 inhibited epithelial to mesenchymal transition (EMT by upregulating the epithelial cell marker, E-cadherin, and downregulating mesenchymal markers, smooth muscle cell alpha-actin (SMA and Snai2. Our data demonstrated that lentiviral vector mediated CLDN1 RNA interference has great potential in breast cancer gene therapy by inhibiting EMT and controlling tumor cell growth.

  20. Discovering sparse transcription factor codes for cell states and state transitions during development

    Science.gov (United States)

    Furchtgott, Leon A; Melton, Samuel; Menon, Vilas; Ramanathan, Sharad

    2017-01-01

    Computational analysis of gene expression to determine both the sequence of lineage choices made by multipotent cells and to identify the genes influencing these decisions is challenging. Here we discover a pattern in the expression levels of a sparse subset of genes among cell types in B- and T-cell developmental lineages that correlates with developmental topologies. We develop a statistical framework using this pattern to simultaneously infer lineage transitions and the genes that determine these relationships. We use this technique to reconstruct the early hematopoietic and intestinal developmental trees. We extend this framework to analyze single-cell RNA-seq data from early human cortical development, inferring a neocortical-hindbrain split in early progenitor cells and the key genes that could control this lineage decision. Our work allows us to simultaneously infer both the identity and lineage of cell types as well as a small set of key genes whose expression patterns reflect these relationships. DOI: http://dx.doi.org/10.7554/eLife.20488.001 PMID:28296636

  1. Fuel Cell Buses in U.S. Transit Fleets: Current Status 2013

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, L.; Gikakis, C.

    2013-12-01

    This report is the seventh in an annual series of reports that summarize the progress of fuel cell electric bus (FCEB) development in the United States and discuss the achievements and challenges of introducing fuel cell propulsion in transit. The report also provides a snapshot of current FCEB performance results from August 2012 through July 2013 for five FCEB demonstrations at four transit agencies.

  2. Derivation of naive human embryonic stem cells.

    Science.gov (United States)

    Ware, Carol B; Nelson, Angelique M; Mecham, Brigham; Hesson, Jennifer; Zhou, Wenyu; Jonlin, Erica C; Jimenez-Caliani, Antonio J; Deng, Xinxian; Cavanaugh, Christopher; Cook, Savannah; Tesar, Paul J; Okada, Jeffrey; Margaretha, Lilyana; Sperber, Henrik; Choi, Michael; Blau, C Anthony; Treuting, Piper M; Hawkins, R David; Cirulli, Vincenzo; Ruohola-Baker, Hannele

    2014-03-25

    The naïve pluripotent state has been shown in mice to lead to broad and more robust developmental potential relative to primed mouse epiblast cells. The human naïve ES cell state has eluded derivation without the use of transgenes, and forced expression of OCT4, KLF4, and KLF2 allows maintenance of human cells in a naïve state [Hanna J, et al. (2010) Proc Natl Acad Sci USA 107(20):9222-9227]. We describe two routes to generate nontransgenic naïve human ES cells (hESCs). The first is by reverse toggling of preexisting primed hESC lines by preculture in the histone deacetylase inhibitors butyrate and suberoylanilide hydroxamic acid, followed by culture in MEK/ERK and GSK3 inhibitors (2i) with FGF2. The second route is by direct derivation from a human embryo in 2i with FGF2. We show that human naïve cells meet mouse criteria for the naïve state by growth characteristics, antibody labeling profile, gene expression, X-inactivation profile, mitochondrial morphology, microRNA profile and development in the context of teratomas. hESCs can exist in a naïve state without the need for transgenes. Direct derivation is an elusive, but attainable, process, leading to cells at the earliest stage of in vitro pluripotency described for humans. Reverse toggling of primed cells to naïve is efficient and reproducible.

  3. Regulatory T Cells in Human Ovarian Cancer

    Directory of Open Access Journals (Sweden)

    Dong-Jun Peng

    2012-01-01

    Full Text Available Multiple layers of suppressive components including regulatory T (TReg cells, suppressive antigen-presenting cells, and inhibitory cytokines form suppressive networks in the ovarian cancer microenvironment. It has been demonstrated that as a major suppressive element, TReg cells infiltrate tumor, interact with several types of immune cells, and mediate immune suppression through different molecular and cellular mechanisms. In this paper, we focus on human ovarian cancer and will discuss the nature of TReg cells including their subsets, trafficking, expansion, and function. We will briefly review the development of manipulation of TReg cells in preclinical and clinical settings.

  4. Generation of pancreatic islet cells from human embryonic stem cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG DongHui; JIANG Wei; SHI Yan; DENG HongKui

    2009-01-01

    Efficiently obtaining functional pancreaUc islet cells derived from human embryonic stem (hES) cells not only provides great potential to solve the shortage of islets sources for type I diabetes cell therapy,but also benefits the study of the development of the human pancreas and diabetes pathology. In 2001,hES cells were reported to have the capacity to generate insulin-producing cells by spontaneous differentiation in vitro. Since then, many strategies (such as overexpression of key transcription factors,delivery of key proteins for pancreatic development, co-transplantation of differentiated hES cells along with fetal pancreas, stepwise differentiation by mimicking in vivo pancreatic development) have been employed in order to induce the differentiation of pancreatic islet cells from hES cells. Moreover, patient-specific induced pluripotent stem (iPS) cells can be generated by reprogramming somatic cells.iPS cells have characteristics similar to those of ES cells and offer a new cell source for type I diabetes cell therapy that reduces the risk of immunologic rejection. In this review, we summarize the recent progress made in the differentiation of hES and iPS cells into functional pancreatic islet cells and discuss the challenges for their future study.

  5. Generation of pancreatic islet cells from human embryonic stem cells

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Efficiently obtaining functional pancreatic islet cells derived from human embryonic stem(hES) cells not only provides great potential to solve the shortage of islets sources for type I diabetes cell therapy,but also benefits the study of the development of the human pancreas and diabetes pathology.In 2001,hES cells were reported to have the capacity to generate insulin-producing cells by spontaneous differentiation in vitro.Since then,many strategies(such as overexpression of key transcription factors,delivery of key proteins for pancreatic development,co-transplantation of differentiated hES cells along with fetal pancreas,stepwise differentiation by mimicking in vivo pancreatic development) have been employed in order to induce the differentiation of pancreatic islet cells from hES cells.Moreover,patient-specific induced pluripotent stem(iPS) cells can be generated by reprogramming somatic cells.iPS cells have characteristics similar to those of ES cells and offer a new cell source for type I diabetes cell therapy that reduces the risk of immunologic rejection.In this review,we summarize the recent progress made in the differentiation of hES and iPS cells into functional pancreatic islet cells and discuss the challenges for their future study.

  6. Differential mRNA expression of the human DNA methyltransferases (DNMTs) 1, 3a and 3b during the G0/G1 to S phase transition in normal and tumor cells

    Science.gov (United States)

    Robertson, Keith D.; Keyomarsi, Khandan; Gonzales, Felicidad A.; Velicescu, Mihaela; Jones, Peter A.

    2000-01-01

    DNA methylation is essential for mammalian development, X-chromosome inactivation, and imprinting yet aberrant methylation patterns are one of the most common features of transformed cells. One of the proposed causes for these defects in the methylation machinery is overexpression of one or more of the three known catalytically active DNA methyltransferases (DNMTs) 1, 3a and 3b, yet there are clearly examples in which overexpression is minimal or non-existent but global methylation anomalies persist. An alternative mechanism which could give rise to global methylation errors is the improper expression of one or more of the DNMTs during the cell cycle. To begin to study the latter possibility we examined the expression of the mRNAs for DNMT1, 3a and 3b during the cell cycle of normal and transformed cells. We found that DNMT1 and 3b levels were significantly downregulated in G0/G1 while DNMT3a mRNA levels were less sensitive to cell cycle alterations and were maintained at a slightly higher level in tumor lines compared to normal cell strains. Enzymatic activity assays revealed a similar decrease in the overall methylation capacity of the cells during G0/G1 arrest and again revealed that a tumor cell line maintained a higher methylation capacity during arrest than a normal cell strain. These results reveal a new level of control exerted over the cellular DNA methylation machinery, the loss of which provides an alternative mechanism for the genesis of the aberrant methylation patterns observed in tumor cells. PMID:10773079

  7. Interrogating a cell signalling network sensitively monitors cell fate transition during early differentiation of mouse embryonic stem cells

    Institute of Scientific and Technical Information of China (English)

    LIU; Yi-Hsin; HO; Chih-ming

    2010-01-01

    The different cell types in an animal are often considered to be specified by combinations of transcription factors,and defined by marker gene expression.This paradigm is challenged,however,in stem cell research and application.Using a mouse embryonic stem cell(mESC) culture system,here we show that the expression level of many key stem cell marker genes/transcription factors such as Oct4,Sox2 and Nanog failed to monitor cell status transition during mESC differentiation.On the other hand,the response patterns of cell signalling network to external stimuli,as monitored by the dynamics of protein phosphorylation,changed dramatically.Our results also suggest that an irreversible alternation in the cell signalling network precedes the adjustment of transcription factor levels.This is consistent with the notion that signal transduction events regulate cell fate specification.We propose that interrogating a cell signalling network can assess the cell property more precisely,and provide a sensitive measurement for the early events in cell fate transition.We wish to bring attention to the potential problem of cell identification using a few marker genes,and suggest a novel methodology to address this issue.

  8. Plasma membrane proteomics of human embryonic stem cells and human embryonal carcinoma cells.

    NARCIS (Netherlands)

    Dormeyer, W.; van Hoof, D.; Braam, S.R.; Heck, A.J.R.; Mummery, C.L.; Krijgsveld, J.

    2008-01-01

    Human embryonic stem cells (hESCs) are of immense interest in regenerative medicine as they can self-renew indefinitely and can give rise to any adult cell type. Human embryonal carcinoma cells (hECCs) are the malignant counterparts of hESCs found in testis tumors. hESCs that have acquired chromosom

  9. Salinomycin alone or in combination with paclitaxel or mitomycin inhibited proliferation of human bladder transitional cell cancer cell line T24%盐霉素及其联合紫杉醇、丝裂霉素对人膀胱移行细胞癌T24细胞增殖的抑制作用

    Institute of Scientific and Technical Information of China (English)

    瞿虎; 袁浩锋; 汪中扬; 张靖; 邹自灏

    2015-01-01

    目的 观察盐霉素及其联合紫杉醇、丝裂霉素对人膀胱癌T24细胞增殖的抑制作用.方法 细胞计数试剂盒(CCK-8)方法检测0.1、0.2、0.4、0.8、1.6 g/L浓度盐霉素组24、48 h,联合紫杉醇0.5 mg/L组24、48 h及丝裂霉素2 mg/L组24、48、72 h对人膀胱癌T24细胞增殖生长抑制率及相对细胞生长抑制率;膜联蛋白V(Annexin V)/碘化丙锭(PI)双染色法流式细胞仪检测单药各组T24细胞凋亡率.结果 对照组T24细胞增殖活跃,经不同浓度盐霉素作用24h即显示出增殖抑制,随药物浓度增殖抑制作用更显著,48 h比24 h抑制更明显,除0.1 g/L外各组与对照组比较差异有统计学意义(P<0.05);流式细胞仪(FCM)检测T24细胞凋亡率在阴性对照组、0.1、0.2、0.4、0.8、1.6 g/L各组凋亡率均高于阴性对照组(P<0.05).盐霉素联合紫杉醇除紫杉醇(PAC)0.5组(阳性对照组)与PAC 0.5+SAL 10组间比较外,其余各组间相互比较差异有统计学意义(P<0.05).盐霉素联合丝裂霉素(MMC)除MMC 2.0+SAL 640组与MMC 2.0组(阳性对照组)之间比较差异有统计学意义(P<0.05),其余各组间与MMC 2.0组(阳性对照组)比较差异无统计学意义(P>0.05),联合用药24 h时,各药物处理组间差异无统计学意义,提示盐霉素对丝裂霉素协同抑制作用具有时间积累效应.结论 盐霉素单药应用、盐霉素联合紫杉醇或丝裂霉素对人膀胱癌T24细胞增殖均有时间及剂量依赖性的抑制作用及促进T24细胞凋亡作用.%Objective To investigate the inhiboty effect of salinomycin combined with paclitaxel or mitomycin on the proliferation of human bladder transitional cell cancer cell line T24.Methods The cell counting kit-8 (CCK-8) assay was adopted to investigate the growth inhibition rate and relative cell growth inhibition of T24 cellular viability after treated with different concentrations of salinomycin combined with paclitaxel or mitomycin.Annexin V/propidium iodide

  10. Transitional cell carcinoma of the ureter and struvite calculi

    Directory of Open Access Journals (Sweden)

    Danielo Garcia de Freitas

    1999-05-01

    Full Text Available CONTEXT: The association of primary carcinoma of the ureter and lithiasis is extremely rare. We report a rare case of a primary carcinoma of the ureter with corariform calculus. CASE REPORT: 60-year-old phaeodermal female, reported a history of right-side nephritic colic, hyperthermia and pyuria during the past 20 years and had received treatment for urinary infections a number of times. The first clinical presentation was related to lithiasis and the tumor had not been shown up by excretory urography, cystoscopy or ultrasonography. Two months after the calculus had been eliminated, the patient began to have serious symptoms and a grade III transitional cell carcinoma of the ureter was discovered. Total nephroureterectomy and M.V.A.C. (Metotrexate + Vinblastina + Doxo Rubicina + Cisplatina chemotherapy were tried unsuccessfully. In this report we emphasize the diagnostic difficulty caused by the concomitant presence of the two pathologies. In our opinion, the rapid evolution in this case is directly related to the high grade of the tumor.

  11. Types of HLA in the bladder transitional cell carcinoma (TCC).

    Science.gov (United States)

    Yılmaz, Erkan; Uğur Özalp, Ali; Cekmen, Arman; Eren, Bülent; Onal, Bülent; Akkuş, Emre; Erdoğan, Ergun

    2013-02-01

    HLA plays a complementary role in the interaction between tumor and body immunology. The aim of this study was to determine the existence of the association between the HLA system and transitional cell carcinoma (TCC). Using standard micro-lymphocytotoxic method of Terasaki, HLA-A, B, DR and DQ antigen types of 30 patients with TCC of the bladder were compared with the control group (30 healthy people). In the TCC patient group, HLA -DQ6(1) and HLA -DQ7(3) antigens were detected with a significantly higher frequency than in the control group (p=0.018 and p=0.038, respectively), whereas HLA-A10, B4, DR53 and DQ1 antigens were detected with significantly higher frequency in the control group (p less 0.05 in all). It suggests that patients who had the antigens detected were at higher risk of TCC, and the ones who had the antigens displaying protective features as were detected in the control group, were at lesser risk.

  12. Induction of epithelial-mesenchymal transition (EMT) in breast cancer cells is calcium signal dependent.

    Science.gov (United States)

    Davis, F M; Azimi, I; Faville, R A; Peters, A A; Jalink, K; Putney, J W; Goodhill, G J; Thompson, E W; Roberts-Thomson, S J; Monteith, G R

    2014-05-01

    Signals from the tumor microenvironment trigger cancer cells to adopt an invasive phenotype through epithelial-mesenchymal transition (EMT). Relatively little is known regarding key signal transduction pathways that serve as cytosolic bridges between cell surface receptors and nuclear transcription factors to induce EMT. A better understanding of these early EMT events may identify potential targets for the control of metastasis. One rapid intracellular signaling pathway that has not yet been explored during EMT induction is calcium. Here we show that stimuli used to induce EMT produce a transient increase in cytosolic calcium levels in human breast cancer cells. Attenuation of the calcium signal by intracellular calcium chelation significantly reduced epidermal growth factor (EGF)- and hypoxia-induced EMT. Intracellular calcium chelation also inhibited EGF-induced activation of signal transducer and activator of transcription 3 (STAT3), while preserving other signal transduction pathways such as Akt and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation. To identify calcium-permeable channels that may regulate EMT induction in breast cancer cells, we performed a targeted siRNA-based screen. We found that transient receptor potential-melastatin-like 7 (TRPM7) channel expression regulated EGF-induced STAT3 phosphorylation and expression of the EMT marker vimentin. Although intracellular calcium chelation almost completely blocked the induction of many EMT markers, including vimentin, Twist and N-cadherin, the effect of TRPM7 silencing was specific for vimentin protein expression and STAT3 phosphorylation. These results indicate that TRPM7 is a partial regulator of EMT in breast cancer cells, and that other calcium-permeable ion channels are also involved in calcium-dependent EMT induction. In summary, this work establishes an important role for the intracellular calcium signal in the induction of EMT in human breast cancer cells. Manipulation of

  13. RNA Directed Modulation of Phenotypic Plasticity in Human Cells.

    Science.gov (United States)

    Trakman, Laura; Hewson, Chris; Burdach, Jon; Morris, Kevin V

    2016-01-01

    Natural selective processes have been known to drive phenotypic plasticity, which is the emergence of different phenotypes from one genome following environmental stimulation. Long non-coding RNAs (lncRNAs) have been observed to modulate transcriptional and epigenetic states of genes in human cells. We surmised that lncRNAs are governors of phenotypic plasticity and drive natural selective processes through epigenetic modulation of gene expression. Using heat shocked human cells as a model we find several differentially expressed transcripts with the top candidates being lncRNAs derived from retro-elements. One particular retro-element derived transcripts, Retro-EIF2S2, was found to be abundantly over-expressed in heat shocked cells. Over-expression of Retro-EIF2S2 significantly enhanced cell viability and modulated a predisposition for an adherent cellular phenotype upon heat shock. Mechanistically, we find that this retro-element derived transcript interacts directly with a network of proteins including 40S ribosomal protein S30 (FAU), Eukaryotic translation initiation factor 5A (EIF5A), and Ubiquitin-60S ribosomal protein L40 (UBA52) to affect protein modulated cell adhesion pathways. We find one motif in Retro-EIF2S2 that exhibits binding to FAU and modulates phenotypic cell transitions from adherent to suspension states. The observations presented here suggest that retroviral derived transcripts actively modulate phenotypic plasticity in human cells in response to environmental selective pressures and suggest that natural selection may play out through the action of retro-elements in human cells.

  14. Derivation of Human Skin Fibroblast Lines for Feeder Cells of Human Embryonic Stem Cells.

    Science.gov (United States)

    Unger, Christian; Felldin, Ulrika; Rodin, Sergey; Nordenskjöld, Agneta; Dilber, Sirac; Hovatta, Outi

    2016-02-03

    After the first derivations of human embryonic stem cell (hESC) lines on fetal mouse feeder cell layers, the idea of using human cells instead of mouse cells as feeder cells soon arose. Mouse cells bear a risk of microbial contamination, and nonhuman immunogenic proteins are absorbed from the feeders to hESCs. Human skin fibroblasts can be effectively used as feeder cells for hESCs. The same primary cell line, which can be safely used for up to 15 passages after stock preparations, can be expanded and used for large numbers of hESC derivations and cultures. These cells are relatively easy to handle and maintain. No animal facilities or animal work is needed. Here, we describe the derivation, culture, and cryopreservation procedures for research-grade human skin fibroblast lines. We also describe how to make feeder layers for hESCs using these fibroblasts.

  15. Role of interleukin 6 in osteogenic transition and calcification of human umbilical artery smooth muscle cells in vitro and the possible cell signal transduction way%白细胞介素6对体外培养人脐动脉平滑肌细胞成骨样转化、钙化的影响

    Institute of Scientific and Technical Information of China (English)

    孙明姝; 郭永平; 顾乐怡; 戴慧莉; 严玉澄; 倪兆慧; 钱家麒

    2009-01-01

    目的 研究白细胞介素6(IL-6)对体外培养人脐动脉平滑肌细胞(HUASMC)成骨样转化、钙化的作用及可能的信号通路.方法 组织植块法原代培养HUASMC.培养基加入不同浓度重组人IL-6(rhIL-6)孵育细胞,设空白对照组.茜素红S钙沉积染色及甲氧.酚酞络合酮法榆测细胞层钙盐含量.实时定量PCR、荧光定量法以及Western印迹法分别检测骨特异性碱性磷酸酶(BAP)、骨桥蛋白(OPN)、骨形成蛋白2(BMP2)和骨保护素(OPG)基因以及蛋白表达.凝胶迁移滞后实验(EMSA)检测核心结合因子α1亚基(Cbfα1)的结合活力,以及分别应用p38丝裂原活化蛋白激酶(p38MAPK)抑制剂SB203580和蛋白激酶C二氢神经鞘氨醇(DHC)后Cbfα1的结合活力.结果 rhlL-6 50μg/L诱导12 d,细胞基质层茜素红S染色阳性.与对照组相比,细胞层钙盐含量在rhIL-6 10 μg/L组刺激9 d[(0.76+0.02)mmol/g蛋白]和12 d[(1.54±0.11)mmol/g蛋白]升高,50μg/L组刺激6 d[(1.81±0.03)mmol/g蛋白]、9 d[(2.08±0.10)mmol/g蛋白]和12 d[(3.22±0.18)mmol/g蛋白]升高,并呈时间、剂量依赖地增加.rhIL-6 10μg/L刺激12 h,BMP2 mRNA(3.04±0.07)和蛋白(8.14±0.41)及BAP mRNA(2.51±0.11)和蛋白(3.96±0.54)表达上调;刺激72 h,OPN mRNA(3.14±0.32)和蛋白(2.57±0.43)水平及OPG mRNA(4.06±0.24)和蛋白(3.46±0.34)水平上调.rhIL-6刺激6 h,Cbfα1结合活力增加;DHC能够部分抑制rhIL-6诱导的Cbfα1结合活力增加,SB203580没有明显作用.结论 IL-6体外能够诱导HUASMC发生钙化和成骨样转分化,这可能是临床观察到IL-6与血管钙化相关的机制之一.IL-6的这一作用可能与细胞内蛋白激酶C通路的活化有关.%Objective To investigate the role of recombinant human interleukin 6 (rhlL-6) in calcification and osteogenic transition of cultured human umbilical artery smooth muscle cells (HUASMC), and the possible cell signal transduction way. Methods HUASMCs were isolated by the explant method. HUASMCs were

  16. Mesenchymal stem cell like (MSCl) cells generated from human embryonic stem cells support pluripotent cell growth

    Energy Technology Data Exchange (ETDEWEB)

    Varga, Nora [Membrane Research Group of the Hungarian Academy of Sciences, Semmelweis University, Budapest (Hungary); Vereb, Zoltan; Rajnavoelgyi, Eva [Department of Immunology, Medical and Health Science Centre, University of Debrecen, Debrecen (Hungary); Nemet, Katalin; Uher, Ferenc; Sarkadi, Balazs [Membrane Research Group of the Hungarian Academy of Sciences, Semmelweis University, Budapest (Hungary); Apati, Agota, E-mail: apati@kkk.org.hu [Membrane Research Group of the Hungarian Academy of Sciences, Semmelweis University, Budapest (Hungary)

    2011-10-28

    Highlights: Black-Right-Pointing-Pointer MSC like cells were derived from hESC by a simple and reproducible method. Black-Right-Pointing-Pointer Differentiation and immunosuppressive features of MSCl cells were similar to bmMSC. Black-Right-Pointing-Pointer MSCl cells as feeder cells support the undifferentiated growth of hESC. -- Abstract: Mesenchymal stem cell like (MSCl) cells were generated from human embryonic stem cells (hESC) through embryoid body formation, and isolated by adherence to plastic surface. MSCl cell lines could be propagated without changes in morphological or functional characteristics for more than 15 passages. These cells, as well as their fluorescent protein expressing stable derivatives, efficiently supported the growth of undifferentiated human embryonic stem cells as feeder cells. The MSCl cells did not express the embryonic (Oct4, Nanog, ABCG2, PODXL, or SSEA4), or hematopoietic (CD34, CD45, CD14, CD133, HLA-DR) stem cell markers, while were positive for the characteristic cell surface markers of MSCs (CD44, CD73, CD90, CD105). MSCl cells could be differentiated toward osteogenic, chondrogenic or adipogenic directions and exhibited significant inhibition of mitogen-activated lymphocyte proliferation, and thus presented immunosuppressive features. We suggest that cultured MSCl cells can properly model human MSCs and be applied as efficient feeders in hESC cultures.

  17. Phenol-soluble modulin α induces G2/M phase transition delay in eukaryotic HeLa cells

    Science.gov (United States)

    Deplanche, Martine; Filho, Rachid Aref El-Aouar; Alekseeva, Ludmila; Ladier, Emilie; Jardin, Julien; Henry, Gwénaële; Azevedo, Vasco; Miyoshi, Anderson; Beraud, Laetitia; Laurent, Frederic; Lina, Gerard; Vandenesch, François; Steghens, Jean-Paul; Le Loir, Yves; Otto, Michael; Götz, Friedrich; Berkova, Nadia

    2015-01-01

    Staphylococcus aureus is a gram-positive bacterium responsible for a wide range of infections. Host cell cycle alteration is a sophisticated mechanism used by pathogens to hijack the defense functions of host cells. We previously demonstrated that S. aureus MW2 (USA400) bacteria induced a G2/M phase transition delay in HeLa cells. We demonstrate here that this activity is triggered by culture supernatant compounds. Using size exclusion chromatography of the MW2 supernatant, followed by mass spectroscopy analysis of corresponding peaks, we identified phenol-soluble modulin α (PSMα) peptides as the likely candidates for this effect. Indeed, synthetic PSMα1 and PSMα3 caused a G2/M phase transition delay. The implication of PSMα in cell cycle alteration was confirmed by comparison of S. aureus Los Angeles County clone (LAC) wild-type with the isogenic mutant LAC∆psmα, which lacks the psmα operon encoding PSMα1–4. PSMα-induced G2/M transition delay correlated with a decrease in the defensin genes expression suggesting a diminution of antibacterial functions of epithelial cells. By testing the supernatant of S. aureus human clinical isolates, we found that the degree of G2/M phase transition delay correlated with PSMα1 production. We show that PSMs secreted by S. aureus alter the host cell cycle, revealing a newly identified mechanism for fostering an infection.—Deplanche, M., Filho. R. A. E.–A., Alekseeva, L., Ladier, E., Jardin, J., Henry, G., Azevedo, V., Miyoshi, A., Beraud, L., Laurent, F., Lina, G., Vandenesch, F., Steghens, J.-P., Le Loir, Y., Otto, M., Götz, F., Berkova, N. Phenol-soluble modulin α induces G2/M phase transition delay in eukaryotic HeLa cells. PMID:25648996

  18. Paeonol attenuates aging MRC-5 cells and inhibits epithelial-mesenchymal transition of premalignant HaCaT cells induced by aging MRC-5 cell-conditioned medium.

    Science.gov (United States)

    Yang, Lihua; Xing, Shangping; Wang, Kun; Yi, Hua; Du, Biaoyan

    2017-08-12

    Senescence-associated secretory phenotype (SASP) factors, such as IL-6 and IL-8, are extremely critical in tissue microenvironment. Senescent human fibroblasts facilitate epithelial-mesenchymal transition (EMT) in premalignant epithelial cells mainly through the secretion of SASP factors. Meanwhile, premalignant human HaCaT Keratinocyte (HaCaT) cells as immortal epithelial cells are susceptible to malignant transformation. Paeonol, an herbal phenolic component found in peonies, exerts anti-aging and anti-tumor efficacies, while the molecular mechanisms of paeonol on EMT in premalignant HaCaT cells induced by SASP factors are unclear. In this study, we first established a senescent human fetal lung fibroblast MRC-5 cell model using hydrogen peroxide evaluated by senescence-associated β-galactosidase assay. Upon paeonol treatment, intracellular reactive oxygen species levels in aging MRC-5 cells were significantly decreased via regulation of nuclear translocation of Nrf2. Then we curiously studied whether the aging MRC-5 cell-conditioned medium could induce EMT in premalignant HaCaT cells, and the results showed that paeonol significantly reduced the clonogenic, migratory, and invasive capacities of premalignant HaCaT cells potentially induced by IL-6 and IL-8. Moreover, we found that paeonol notably altered pluripotency of EMT-associated markers via the modulation of ERK and TGF-β1/Smad pathway in premalignant HaCaT cells. These findings suggest that paeonol may be used as an adjuvant therapy for SASP factor-mediated EMT in premalignant lesion.

  19. Detecting viability transitions of umbilical cord mesenchymal stem cells by Raman micro-spectroscopy

    Science.gov (United States)

    Bai, H.; Chen, P.; Fang, H.; Lin, L.; Tang, G. Q.; Mu, G. G.; Gong, W.; Liu, Z. P.; Wu, H.; Zhao, H.; Han, Z. C.

    2011-01-01

    Recent research suggests that human umbilical cord derived mesenchymal stem cells (hUC-MSCs) can be promising candidates for cell-based therapy. Since large population and high viability are generally required, detecting viability transitions of these cells is crucial for their population expansion and quality control. Here, as a non-invasive method, Raman micro-spectroscopy is applied to examine hUC-MSCs with different viability. Using peak fitting and statistic t-test, the Raman peaks with obvious differences between the cells with high viability (> 90%) and low viability (< 20%) are extracted. It is found that the C=O out of plane bending in thymine at 744 cm-1, symmetric stretching of C-C in lipids at 877 cm-1 and CH deformation in proteins at 1342 cm-1 show the most significant changes (p < 0.001). When the cell viability decreases, the intensities of the former two peaks are both about doubled while that of the latter peak reduces by about 30%. Based on these results, we propose that the viability of hUC-MSCs can be characterized by these three peaks. And their intensity changes can be understood from the model of excessive reactive oxygen species interacting with the bio-macromolecules.

  20. Phase transition of the microvascular network architecture in human pathologies.

    Science.gov (United States)

    Bianciardi, Giorgio; Traversi, Claudio; Cattaneo, Ruggero; De Felice, Claudia; Monaco, Annalisa; Tosi, Gianmarco; Parrini, Stefano; Latini, Giuseppe

    2012-01-01

    We have investigated the microvascular pattern in acquired or genetic diseases in humans. The lower gingival and vestibular oral mucosa, as well as the optic nerve head, was chosen to characterize the vascular pattern complexity due to the simple accessibility and visibility Local fractal dimensions, fractal dimension of the minimum path and Lempel-Ziv complexity have been used as operational numerical tools to characterize the microvascular networks. In the normal healthy subjects microvascular networks show nonlinear values corresponding to the complexity of a diffusion limited aggregation (DLA) model, while in several acquired or genetic diseases they are approaching the ones of an invasion percolation model.

  1. Staphylococcus aureus-induced G2/M phase transition delay in host epithelial cells increases bacterial infective efficiency.

    Science.gov (United States)

    Alekseeva, Ludmila; Rault, Lucie; Almeida, Sintia; Legembre, Patrick; Edmond, Valérie; Azevedo, Vasco; Miyoshi, Anderson; Even, Sergine; Taieb, Frédéric; Arlot-Bonnemains, Yannick; Le Loir, Yves; Berkova, Nadia

    2013-01-01

    Staphylococcus aureus is a highly versatile, opportunistic pathogen and the etiological agent of a wide range of infections in humans and warm-blooded animals. The epithelial surface is its principal site of colonization and infection. In this work, we investigated the cytopathic effect of S. aureus strains from human and animal origins and their ability to affect the host cell cycle in human HeLa and bovine MAC-T epithelial cell lines. S. aureus invasion slowed down cell proliferation and induced a cytopathic effect, resulting in the enlargement of host cells. A dramatic decrease in the number of mitotic cells was observed in the infected cultures. Flow cytometry analysis revealed an S. aureus-induced delay in the G2/M phase transition in synchronous HeLa cells. This delay required the presence of live S. aureus since the addition of the heat-killed bacteria did not alter the cell cycle. The results of Western blot experiments showed that the G2/M transition delay was associated with the accumulation of inactive cyclin-dependent kinase Cdk1, a key inducer of mitosis entry, and with the accumulation of unphosphorylated histone H3, which was correlated with a reduction of the mitotic cell number. Analysis of S. aureus proliferation in asynchronous, G1- and G2-phase-enriched HeLa cells showed that the G2 phase was preferential for bacterial infective efficiency, suggesting that the G2 phase delay may be used by S. aureus for propagation within the host. Taken together, our results divulge the potential of S. aureus in the subversion of key cellular processes such as cell cycle progression, and shed light on the biological significance of S. aureus-induced host cell cycle alteration.

  2. Staphylococcus aureus-induced G2/M phase transition delay in host epithelial cells increases bacterial infective efficiency.

    Directory of Open Access Journals (Sweden)

    Ludmila Alekseeva

    Full Text Available Staphylococcus aureus is a highly versatile, opportunistic pathogen and the etiological agent of a wide range of infections in humans and warm-blooded animals. The epithelial surface is its principal site of colonization and infection. In this work, we investigated the cytopathic effect of S. aureus strains from human and animal origins and their ability to affect the host cell cycle in human HeLa and bovine MAC-T epithelial cell lines. S. aureus invasion slowed down cell proliferation and induced a cytopathic effect, resulting in the enlargement of host cells. A dramatic decrease in the number of mitotic cells was observed in the infected cultures. Flow cytometry analysis revealed an S. aureus-induced delay in the G2/M phase transition in synchronous HeLa cells. This delay required the presence of live S. aureus since the addition of the heat-killed bacteria did not alter the cell cycle. The results of Western blot experiments showed that the G2/M transition delay was associated with the accumulation of inactive cyclin-dependent kinase Cdk1, a key inducer of mitosis entry, and with the accumulation of unphosphorylated histone H3, which was correlated with a reduction of the mitotic cell number. Analysis of S. aureus proliferation in asynchronous, G1- and G2-phase-enriched HeLa cells showed that the G2 phase was preferential for bacterial infective efficiency, suggesting that the G2 phase delay may be used by S. aureus for propagation within the host. Taken together, our results divulge the potential of S. aureus in the subversion of key cellular processes such as cell cycle progression, and shed light on the biological significance of S. aureus-induced host cell cycle alteration.

  3. Direct Differentiation of Human Pluripotent Stem Cells into Haploid Spermatogenic Cells

    Science.gov (United States)

    Easley, Charles A.; Phillips, Bart T.; McGuire, Megan M.; Barringer, Jennifer M.; Valli, Hanna; Hermann, Brian P.; Simerly, Calvin R.; Rajkovic, Aleksander; Miki, Toshio; Orwig, Kyle E.; Schatten, Gerald P.

    2013-01-01

    Summary Human embryonic (hESCs) and induced pluripotent stem cells (hiPSCs) have been shown to differentiate into primordial germ cells (PGCs) but not into spermatogonia nor haploid spermatocytes or spermatids. Here we show that hESCs and hiPSCs differentiate directly into advanced male germ cell lineages including post-meiotic, spermatid-like cells in vitro without genetic manipulation. Furthermore, our procedure mirrors spermatogenesis in vivo by differentiating pluripotent stem cells into UTF1, PLZF and CDH1-positive spermatogonia-like cells, HIWI and HILI-positive spermatocyte-like cells, and haploid cells expressing acrosin, transition protein 1 and protamine 1, proteins found uniquely in either spermatids and/or sperm. These spermatids show uniparental genomic imprints similar to human sperm on two loci: H19 and IGF2. These results demonstrate that male pluripotent stem cells have the capability to directly differentiate into advanced germ cell lineages and may represent a novel strategy for studying spermatogenesis in vitro. PMID:22921399

  4. Stem cells in the human breast

    DEFF Research Database (Denmark)

    Petersen, Ole William; Polyak, Kornelia

    2010-01-01

    The origins of the epithelial cells participating in the development, tissue homeostasis, and cancer of the human breast are poorly understood. However, emerging evidence suggests a role for adult tissue-specific stem cells in these processes. In a hierarchical manner, these generate the two main...

  5. Human hair genealogies and stem cell latency

    Directory of Open Access Journals (Sweden)

    Tavaré Simon

    2006-02-01

    Full Text Available Abstract Background Stem cells divide to reproduce themselves and produce differentiated progeny. A fundamental problem in human biology has been the inability to measure how often stem cells divide. Although it is impossible to observe every division directly, one method for counting divisions is to count replication errors; the greater the number of divisions, the greater the numbers of errors. Stem cells with more divisions should produce progeny with more replication errors. Methods To test this approach, epigenetic errors (methylation in CpG-rich molecular clocks were measured from human hairs. Hairs exhibit growth and replacement cycles and "new" hairs physically reappear even on "old" heads. Errors may accumulate in long-lived stem cells, or in their differentiated progeny that are eventually shed. Results Average hair errors increased until two years of age, and then were constant despite decades of replacement, consistent with new hairs arising from infrequently dividing bulge stem cells. Errors were significantly more frequent in longer hairs, consistent with long-lived but eventually shed mitotic follicle cells. Conclusion Constant average hair methylation regardless of age contrasts with the age-related methylation observed in human intestine, suggesting that error accumulation and therefore stem cell latency differs among tissues. Epigenetic molecular clocks imply similar mitotic ages for hairs on young and old human heads, consistent with a restart with each new hair, and with genealogies surreptitiously written within somatic cell genomes.

  6. Transition zone cells reach G2 phase before initiating elongation in maize root apex

    Directory of Open Access Journals (Sweden)

    M. Victoria Alarcón

    2017-06-01

    Full Text Available Root elongation requires cell divisions in the meristematic zone and cell elongation in the elongation zone. The boundary between dividing and elongating cells is called the transition zone. In the meristem zone, initial cells are continuously dividing, but on the basal side of the meristem cells exit the meristem through the transition zone and enter in the elongation zone, where they stop division and rapidly elongate. Throughout this journey cells are accompanied by changes in cell cycle progression. Flow cytometry analysis showed that meristematic cells are in cycle, but exit when they enter the elongation zone. In addition, the percentage of cells in G2 phase (4C strongly increased from the meristem to the elongation zone. However, we did not observe remarkable changes in the percentage of cells in cell cycle phases along the entire elongation zone. These results suggest that meristematic cells in maize root apex stop the cell cycle in G2 phase after leaving the meristem.

  7. The activity of etoposide (VP16) in combination chemotherapy against human bladder cancer cells in vitro

    OpenAIRE

    1991-01-01

    The activity of Etoposide (VP16) in combination chemotherapy against four human transitional cell carcinoma cell lines of bladder (TCCaB) was determined by in vitro colony formation assay. Four anti-tumor agents (methotrexate: MTX, vinblastine: VBL, adriamycin: ADM, cisplatin: DDP) were used for combination chemotherapy with VP16. The ADM + VP16 combination exhibited a strong synergistic antitumor effect against the human TCCaBs compared with other combinations in this study. The combination ...

  8. Analysis of the Antitumor Activity of Clotrimazole on A375 Human Melanoma Cells

    DEFF Research Database (Denmark)

    Adinolfi, Barbara; Carpi, Sara; Romanini, Antonella

    2015-01-01

    analyses of cell viability, gene expression, cell-cycle progression, annexin V reactivity and internucleosomal DNA fragmentation. RESULTS: Clotrimazole induced cytotoxicity in A375 human melanoma cells without significant changes of human keratinocyte cell viability. Clotrimazole, at a concentration...... that approximates the inhibitory concentration 50% (IC50) value (i.e. 10 μM), reduced the expression of hexokinase type-II, induced cell-cycle arrest at G1-S phase transition, altered annexin V reactivity and induced DNA fragmentation without evidence of necrosis. CONCLUSION: The current study provides evidence...

  9. Law in Transition Biblioessay: Globalization, Human Rights, Environment, Technology

    Directory of Open Access Journals (Sweden)

    Michael Marien

    2012-04-01

    Full Text Available As globalization continues, many transformations in international and domestic laws areunderway or called for. There are too many laws and too few, too much law that is inadequateor obsolete, and too much law-breaking. This biblioessay covers some 100 recentbooks, nearly all recently published, arranged in four categories. 1 International Lawincludes six overviews/textbooks on comparative law, laws related to warfare and security,pushback against demands of globalization, and gender perspectives; 2 Human Rightsencompasses general overviews and normative visions, several books on how some statesviolate human rights, five items on how good laws can end poverty and promote prosperity,and laws regulating working conditions and health rights; 3 Environment/Resources coversgrowth of international environmental law, visions of law for a better environmental future,laws to govern genetic resources and increasingly stressed water resources, two books onprospects for climate change liability, and items on toxic hazards and problems of compliance;4 Technology, Etc. identifies eight books on global crime and the failed war on drugs,books on the response to terrorism and guarding privacy and mobility in our high-tech age,seven books on how infotech is changing law and legal processes while raising intellectualproperty questions, biomedical technologies and the law, and general views on the need forupdated laws and constitutions. In sum, this essay suggests the need for deeper and timelyanalysis of the many books on changes in law.

  10. Human neutrophils facilitate tumor cell transendothelial migration.

    LENUS (Irish Health Repository)

    Wu, Q D

    2012-02-03

    Tumor cell extravasation plays a key role in tumor metastasis. However, the precise mechanisms by which tumor cells migrate through normal vascular endothelium remain unclear. In this study, using an in vitro transendothelial migration model, we show that human polymorphonuclear neutrophils (PMN) assist the human breast tumor cell line MDA-MB-231 to cross the endothelial barrier. We found that tumor-conditioned medium (TCM) downregulated PMN cytocidal function, delayed PMN apoptosis, and concomitantly upregulated PMN adhesion molecule expression. These PMN treated with TCM attached to tumor cells and facilitated tumor cell migration through different endothelial monolayers. In contrast, MDA-MB-231 cells alone did not transmigrate. FACScan analysis revealed that these tumor cells expressed high levels of intercellular adhesion molecule-1 (ICAM-1) but did not express CD11a, CD11b, or CD18. Blockage of CD11b and CD18 on PMN and of ICAM-1 on MDA-MB-231 cells significantly attenuated TCM-treated, PMN-mediated tumor cell migration. These tumor cells still possessed the ability to proliferate after PMN-assisted transmigration. These results indicate that TCM-treated PMN may serve as a carrier to assist tumor cell transendothelial migration and suggest that tumor cells can exploit PMN and alter their function to facilitate their extravasation.

  11. Quantitative model of cell cycle arrest and cellular senescence in primary human fibroblasts.

    Directory of Open Access Journals (Sweden)

    Sascha Schäuble

    Full Text Available Primary human fibroblasts in tissue culture undergo a limited number of cell divisions before entering a non-replicative "senescent" state. At early population doublings (PD, fibroblasts are proliferation-competent displaying exponential growth. During further cell passaging, an increasing number of cells become cell cycle arrested and finally senescent. This transition from proliferating to senescent cells is driven by a number of endogenous and exogenous stress factors. Here, we have developed a new quantitative model for the stepwise transition from proliferating human fibroblasts (P via reversibly cell cycle arrested (C to irreversibly arrested senescent cells (S. In this model, the transition from P to C and to S is driven by a stress function γ and a cellular stress response function F which describes the time-delayed cellular response to experimentally induced irradiation stress. The application of this model based on senescence marker quantification at the single-cell level allowed to discriminate between the cellular states P, C, and S and delivers the transition rates between the P, C and S states for different human fibroblast cell types. Model-derived quantification unexpectedly revealed significant differences in the stress response of different fibroblast cell lines. Evaluating marker specificity, we found that SA-β-Gal is a good quantitative marker for cellular senescence in WI-38 and BJ cells, however much less so in MRC-5 cells. Furthermore we found that WI-38 cells are more sensitive to stress than BJ and MRC-5 cells. Thus, the explicit separation of stress induction from the cellular stress response, and the differentiation between three cellular states P, C and S allows for the first time to quantitatively assess the response of primary human fibroblasts towards endogenous and exogenous stress during cellular ageing.

  12. Human P-glycoprotein exhibits reduced affinity for substrates during a catalytic transition state.

    Science.gov (United States)

    Ramachandra, M; Ambudkar, S V; Chen, D; Hrycyna, C A; Dey, S; Gottesman, M M; Pastan, I

    1998-04-01

    Human P-glycoprotein (Pgp), a plasma membrane protein that confers multidrug resistance, functions as an ATP-dependent drug efflux pump. Pgp contains two ATP binding/utilization sites and exhibits ATPase activity that is stimulated in the presence of substrates and modulating agents. The mechanism of coupling of ATP hydrolysis to drug transport is not known. To understand the role of ATP hydrolysis in drug binding, it is necessary to develop methods for purifying and reconstituting Pgp that retains properties including stimulation of ATPase activity by known substrates to an extent similar to that in the native membrane. In this study, (His)6-tagged Pgp was expressed in Trichoplusia ni (High Five) cells using the recombinant baculovirus system and purified by metal affinity chromatography. Upon reconstitution into phospholipid vesicles, purified Pgp exhibited specific binding to analogues of substrates and ATP in affinity labeling experiments and displayed a high level of drug-stimulated ATPase activity (specific activity ranging from 4.5 to 6.5 micromol min-1 mg-1). The ATPase activity was inhibited by ADP in a competitive manner, and by vanadate and N-ethylmaleimide at low concentrations. Vanadate which is known to inhibit ATPase activity by trapping MgADP at the catalytic site inhibited photoaffinity labeling of Pgp with substrate analogues, [125I]iodoarylazidoprazosin and [3H]azidopine, only under ATP hydrolysis conditions. Because vanadate-trapped Pgp is known to resemble the ADP and phosphate-bound catalytic transition state, our findings indicate that ATP hydrolysis results in a conformation with reduced affinity for substrates. A catalytic transition conformation with reduced affinity would essentially result in substrate dissociation and supports a model for drug transport in which an ATP hydrolysis-induced conformational change leads to drug release toward the extracellular medium.

  13. Chronic cadmium exposure in vitro induces cancer cell characteristics in human lung cells

    Energy Technology Data Exchange (ETDEWEB)

    Person, Rachel J.; Tokar, Erik J.; Xu, Yuanyuan; Orihuela, Ruben; Ngalame, Ntube N. Olive; Waalkes, Michael P., E-mail: waalkes@niehs.nih.gov

    2013-12-01

    Cadmium is a known human lung carcinogen. Here, we attempt to develop an in vitro model of cadmium-induced human lung carcinogenesis by chronically exposing the peripheral lung epithelia cell line, HPL-1D, to a low level of cadmium. Cells were chronically exposed to 5 μM cadmium, a noncytotoxic level, and monitored for acquired cancer characteristics. By 20 weeks of continuous cadmium exposure, these chronic cadmium treated lung (CCT-LC) cells showed marked increases in secreted MMP-2 activity (3.5-fold), invasion (3.4-fold), and colony formation in soft agar (2-fold). CCT-LC cells were hyperproliferative, grew well in serum-free media, and overexpressed cyclin D1. The CCT-LC cells also showed decreased expression of the tumor suppressor genes p16 and SLC38A3 at the protein levels. Also consistent with an acquired cancer cell phenotype, CCT-LC cells showed increased expression of the oncoproteins K-RAS and N-RAS as well as the epithelial-to-mesenchymal transition marker protein Vimentin. Metallothionein (MT) expression is increased by cadmium, and is typically overexpressed in human lung cancers. The major MT isoforms, MT-1A and MT-2A were elevated in CCT-LC cells. Oxidant adaptive response genes HO-1 and HIF-1A were also activated in CCT-LC cells. Expression of the metal transport genes ZNT-1, ZNT-5, and ZIP-8 increased in CCT-LC cells culminating in reduced cadmium accumulation, suggesting adaptation to the metal. Overall, these data suggest that exposure of human lung epithelial cells to cadmium causes acquisition of cancer cell characteristics. Furthermore, transformation occurs despite the cell's ability to adapt to chronic cadmium exposure. - Highlights: • Chronic cadmium exposure induces cancer cell characteristics in human lung cells. • This provides an in vitro model of cadmium-induced human lung cell transformation. • This occurred with general and lung specific changes typical for cancer cells. • These findings add insight to the

  14. The Mu opioid receptor promotes opioid and growth factor-induced proliferation, migration and Epithelial Mesenchymal Transition (EMT in human lung cancer.

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    Frances E Lennon

    Full Text Available Recent epidemiologic studies implying differences in cancer recurrence based on anesthetic regimens raise the possibility that the mu opioid receptor (MOR can influence cancer progression. Based on our previous observations that overexpression of MOR in human non-small cell lung cancer (NSCLC cells increased tumor growth and metastasis, this study examined whether MOR regulates growth factor receptor signaling and epithelial mesenchymal transition (EMT in human NSCLC cells. We utilized specific siRNA, shRNA, chemical inhibitors and overexpression vectors in human H358 NSCLC cells that were either untreated or treated with various concentrations of DAMGO, morphine, fentanyl, EGF or IGF. Cell function assays, immunoblot and immunoprecipitation assays were then performed. Our results indicate MOR regulates opioid and growth factor-induced EGF receptor signaling (Src, Gab-1, PI3K, Akt and STAT3 activation which is crucial for consequent human NSCLC cell proliferation and migration. In addition, human NSCLC cells treated with opioids, growth factors or MOR overexpression exhibited an increase in snail, slug and vimentin and decrease ZO-1 and claudin-1 protein levels, results consistent with an EMT phenotype. Further, these effects were reversed with silencing (shRNA or chemical inhibition of MOR, Src, Gab-1, PI3K, Akt and STAT3 (p<0.05. Our data suggest a possible direct effect of MOR on opioid and growth factor-signaling and consequent proliferation, migration and EMT transition during lung cancer progression. Such an effect provides a plausible explanation for the epidemiologic findings.

  15. Metastatic transitional cell carcinoma presenting with skin metastasis.

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    Açıkgöz, Onur; Ölçücüoğlu, Erkan; Kasap, Yusuf; Yığman, Metin; Güneş, Zeki Ender; Gazel, Eymen

    2015-01-01

    Transitional cell carcinomas (TCC) of upper urinary system account for 5% of all TCCs. The incidence of such metastases ranges from 0.18% to 2%. Experimental studies reported a general unsatisfactory survival time following skin metastasis. We report in this paper a case of metastatic urinary system TCC, which had become evident with a skin lesion in the right hypogastric region. A 60-year-old female patient with a history of being operated upon due to renal pelvic TCC was admitted to our outpatient clinic with complaints of red skin lesion in the near vicinity of the operational incision scar for 3 months. Her medical history revealed nothing but nephroureterectomy operation on the upper urinary system; moreover, it was learned that she had been ignoring what was recommended to her for routine controls. Thoraco-abdominal computed tomographic (CT) examination performed on the basis of aforementioned findings depicted a mass lesion of 24*20 mm dimension with high contrast uptake detected within the subcutaneous fat tissue in the right abdominal wall. The skin lesion depicted in CT was surgically excised. The pathological examination of the excised material was reported to be compatible with TCC. The patient was referred due to abdominal lesion to medical oncology after the operation. Followed up under chemotherapy protocol, the patient died 3 months after the metastasectomy operation. Skin metastasis of upper urinary system TCCs, especially renal pelvic TCCs, are quite rare conditions. Among the likely skin sites of metastasis for genitourinary system TCCs are head, face, extremities, suprapubic region and abdomen. Taking into consideration the low survival rates, the importance of early diagnosis of recurrences and/or distant metastases should be better appreciated. These patients die soon after the skin metastasis even with the administration of aggressive therapy. Similarly, our patient died 90 days after the diagnosis of skin metastasis despite the oncologic

  16. OPIUM USE IN TRANSITIONAL CELL CARCINOMA OF THE URINARY BLADDER

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    A. Nourbakhsh

    2006-08-01

    Full Text Available Opium use is one of the most common forms of substance abuse in Iran and there are some evidence indicating it is a risk factor of transitional cell carcinoma (TCC of the urinary bladder. The majority of opium users are also cigarette smokers, so consideration of the high prevalence of smoking which is the most important risk factor of TCC of the urinary bladder among opium users is essential to assess the role of opium use as a possible risk factor of TCC. This study was done to evaluate the role of opium as a risk factor of TCC. A case-control study was performed on 255 individuals diagnosed with TCC of the urinary bladder by pathologic light microscopic examination of the tumor biopsies. Control population was chosen from individuals who had no history or presenting signs or symptoms of urinary problems. Case and control groups were matched by sex and age and also by cigarette smoking habits. Forty-one (18.1% of the cases and 12 (5% of controls were recognized to be opium users. Mantel-Haenszel analysis showed an odds ratio of 3.88, with 95% confidence interval of 1.99-7.57 and P value of < 0.001. Results indicate that opium use is a risk factor for TCC. The majority of opium users are also cigarette smokers, which is another important risk factor for TCC. Routine urine cytology and early evaluation in the patients presenting with any of the symptoms of urinary bladder malignancy by means of cystoscopy and urine cytology are highly recommended.

  17. Investigating the link between molecular subtypes of glioblastoma, epithelial-mesenchymal transition, and CD133 cell surface protein.

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    Hadi Zarkoob

    Full Text Available In this manuscript, we use genetic data to provide a three-faceted analysis on the links between molecular subclasses of glioblastoma, epithelial-to-mesenchymal transition (EMT and CD133 cell surface protein. The contribution of this paper is three-fold: First, we use a newly identified signature for epithelial-to-mesenchymal transition in human mammary epithelial cells, and demonstrate that genes in this signature have significant overlap with genes differentially expressed in all known GBM subtypes. However, the overlap between genes up regulated in the mesenchymal subtype of GBM and in the EMT signature was more significant than other GBM subtypes. Second, we provide evidence that there is a negative correlation between the genetic signature of EMT and that of CD133 cell surface protein, a putative marker for neural stem cells. Third, we study the correlation between GBM molecular subtypes and the genetic signature of CD133 cell surface protein. We demonstrate that the mesenchymal and neural subtypes of GBM have the strongest correlations with the CD133 genetic signature. While the mesenchymal subtype of GBM displays similarity with the signatures of both EMT and CD133, it also exhibits some differences with each of these signatures that are partly due to the fact that the signatures of EMT and CD133 are inversely related to each other. Taken together these data shed light on the role of the mesenchymal transition and neural stem cells, and their mutual interaction, in molecular subtypes of glioblastoma multiforme.

  18. Tungsten-induced carcinogenesis in human bronchial epithelial cells

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    Laulicht, Freda; Brocato, Jason; Cartularo, Laura; Vaughan, Joshua; Wu, Feng; Kluz, Thomas; Sun, Hong; Oksuz, Betul Akgol; Shen, Steven; Paena, Massimilano; Medici, Serenella; Zoroddu, Maria Antonietta; Costa, Max

    2015-01-01

    Metals such as arsenic, cadmium, beryllium, and nickel are known human carcinogens; however, other transition metals, such as tungsten (W), remain relatively uninvestigated with regard to their potential carcinogenic activity. Tungsten production for industrial and military applications has almost doubled over the past decade and continues to increase. Here, for the first time, we demonstrate tungsten’s ability to induce carcinogenic related endpoints including cell transformation, increased migration, xenograft growth in nude mice, and the activation of multiple cancer related pathways in transformed clones as determined by RNA seq. Human bronchial epithelial cell line (Beas-2B) exposed to tungsten developed carcinogenic properties. In a soft agar assay, tungsten-treated cells formed more colonies than controls and the tungsten-transformed clones formed tumors in nude mice. RNA-sequencing data revealed that the tungsten-transformed clones altered the expression of many cancer-associated genes when compared to control clones. Genes involved in lung cancer, leukemia, and general cancer genes were deregulated by tungsten. Taken together, our data shows the carcinogenic potential of tungsten. Further tests are needed, including in vivo and human studies, in order to validate tungsten as a carcinogen to humans. PMID:26164860

  19. Overexpression of c-myc induces epithelial mesenchymal transition in mammary epithelial cells.

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    Cho, Kyoung Bin; Cho, Min Kyong; Lee, Won Young; Kang, Keon Wook

    2010-07-28

    The c-myc gene is frequently overexpressed in human breast cancer and its target genes are involved in tumorigenesis. Epithelial mesenchymal transitions (EMT), where cells undergo a developmental switch from a polarized epithelial phenotype to a highly motile mesenchymal phenotype, are associated with invasion and motility of cancer cells. Basal E-cadherin expression was down-regulated in c-myc overexpressing MCF10A (c-myc-MCF10A) cells compared to GFP-overexpressing MCF10A (GFP-MCF10A) cells, while N-cadherin was distinctly increased in c-myc-MCF10A cells. Given that glycogen synthase kinase-3beta (GSK-3beta) and the snail axis have key roles in E-cadherin deregulation during EMT, we investigated the role of GSK-3beta/snail signaling pathways in the induction of EMT by c-myc overexpression. In contrast to GFP-MCF10A cells, both the transcriptional activity and the ubiquitination-dependent protein stability of snail were enhanced in c-myc-MCF10A cells, and this was reversed by GSK-3beta overexpression. We also found that c-myc overexpression inhibits GSK-3beta activity through activation of extracellular signal-regulated kinase (ERK). Inhibition of ERK by dominant negative mutant transfection or chemical inhibitor significantly suppressed snail gene transcription. These results suggest that c-myc overexpression during transformation of mammary epithelial cells (MEC) is involved in EMTs via ERK-dependent GSK-3beta inactivation and subsequent snail activation.

  20. Leptin promotes metastasis by inducing an epithelial-mesenchymal transition in A549 lung cancer cells.

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    Feng, Helin; Liu, Qingyi; Zhang, Ning; Zheng, Lihua; Sang, Meixiang; Feng, Jiangang; Zhang, Jinming; Wu, Xiangyun; Shan, Baoen

    2013-01-01

    Leptin, an adipocyte-derived cytokine associated with obesity, has been reported to participate in carcinogenesis. Epithelial-mesenchymal transition (EMT) is also considered as a key event in tumor metastasis. The aim of this study is to investigate the mechanism of leptin in the promotion of EMT leading to metastasis in A549 lung cancer cells. We investigated the effect of leptin on migration of A549 cells using wound healing and transwell assays. The incidence of EMT in A549 cells was examined by real-time PCR and immunofluorescence staining. The expression of TGF-β in A549 cells was detected by real-time PCR, and blocking of TGF-β in A549 cells was achieved by siRNA techniques. Additional work was performed using 100 patient samples, which included samples from 50 patients diagnosed with lung cancer and an additional 50 patients diagnosed with lung cancer with metastatic bone lesions. Leptin expression was measured using immunohistochemistry techniques. We demonstrated that leptin can effectively enhance the metastasis of human lung cancer A549 cell line using both wound healing and transwell assays. We also found the incidence of EMT in A549 cells after leptin exposure. Furthermore, we detected the expression of TGF-β in A549 cells, which had been reported to play an important role in inducing EMT. We showed that leptin can significantly upregulate TGF-β at both the mRNA and protein levels in A549 cells. Using siRNA to block the expression of TGF-β in A549 cells, we confirmed the role of TGF-β in the promotion of metastasis and induction of EMT. Furthermore, we found that in patient samples leptin was present at higher levels in samples associated with diagnosis of lung cancer bone metastases tissue than lung cancer tissue. Our results indicated that leptin promoted the metastasis of A549 human lung cancer cell lines by inducing EMT in a TGF-β-dependent manner.

  1. INTERACTIONS BETWEEN THE HUMAN GASTRIC CARCINOMA CELL AND THE HUMAN VASCULAR ENDOTHELIAL CELL

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective To definite the interactions between the human gastric carcinoma cell and the human vascular endothelial cell during the establishment and maintenance of the tumor vascular system and the tumor hematogenous metastasis.Methods We prepared the conditioned mediums of each cell so as to study the effect of the conditioned medium on itself or others by MTT colorimetry. The comprehensive effect of interactions between two cells was determined by stratified transfilter co-culture or direct contact co-culture.Results The conditioned medium of human gastric carcinoma cell can stimulate the proliferation of the human vascular endothelial cell, but the CM of HVEC can inhibit the growth of HGCC. Both kinds of cells can inhibit the growth of itself. The ultimate comprehensive effect of the interactions between two kinds of cells was increase of total cell numbers.Conclusion There exist the complicated interactions between the human gastric carcinoma cell and the human vascular endothelial cell during the tumor angiogenesis and the tumor hematogenous metastasis. The ultimate comprehensive effect of the interactions is increase of total cells numbers and tumor volume.

  2. Snail involves in the transforming growth factor β1-mediated epithelial-mesenchymal transition of retinal pigment epithelial cells.

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    Hui Li

    Full Text Available BACKGROUND: The proliferation of retinal pigment epithelium (RPE cells resulting from an epithelial-mesenchymal transition (EMT plays a key role in proliferative vitreoretinopathy (PVR, which leads to complex retinal detachment and the loss of vision. Genes of Snail family encode the zinc finger transcription factors that have been reported to be essential in EMT during embryonic development and cancer metastasis. However, the function of Snail in RPE cells undergoing EMT is largely unknown. PRINCIPAL FINDINGS: Transforming growth factor beta(TGF-β-1 resulted in EMT in human RPE cells (ARPE-19, which was characterized by the expected decrease in E-cadherin and Zona occludin-1(ZO-1 expression, and the increase in fibronectin and α-smooth muscle actin (α-SMA expression, as well as the associated increase of Snail expression at both mRNA and protein levels. Furthermore, TGF-β1 treatment caused a significant change in ARPE-19 cells morphology, with transition from a typical epithelial morphology to mesenchymal spindle-shaped. More interestingly, Snail silencing significantly attenuated TGF-β1-induced EMT in ARPE-19 cells by decreasing the mesenchymal markers fibronectin and a-SMA and increasing the epithelial marker E-cadherin and ZO-1. Snail knockdown could effectively suppress ARPE-19 cell migration. Finally, Snail was activated in epiretinal membranes from PVR patients. Taken together, Snail plays very important roles in TGF-β-1-induced EMT in human RPE cells and may contribute to the development of PVR. SIGNIFICANCE: Snail transcription factor plays a critical role in TGF-β1-induced EMT in human RPE cells, which provides deep insight into the pathogenesis of human PVR disease. The specific inhibition of Snail may provide a new approach to treat and prevent PVR.

  3. Fibronectin production by human mammary cells

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    Stampfer, M.R. (Univ. of California, Berkeley); Vlodavsky, I.; Smith, H.S.; Ford, R.; Becker, F.F.; Riggs, J.

    1981-01-01

    Human mammary cells were examined for the presence of the high-molecular-weight surface glycoprotein fibronectin. Early passage mammary epithelial cell and fibroblast cultures from both carcinomas and normal tissues were tested for the presence of cell-associated fibronectin by immunofluorescence microscopy and for the synthesis and secretion of fibronectin by specific immunoprecipitation of metabolically labeled protein. In vivo frozen sections of primary carcinomas and normal tissues were tested for the localization of fibronectin by immunofluorescence microscopy. In contrast to the extensive fibrillar networks of fibronectin found in the fibroblast cultures, the epithelial cell cultures from both tissue sources displayed a pattern of cell-associated fibronectin characterizd by powdery, punctate staining. However, the cultured epithelial cells, as well as the fibroblasts, secreted large quantities of fibronectin into the medium. Putative myoepithelial cells also displayed extensive fibrillar networks of fibronectin. The difference in cell-associated fibronectin distribution between the epithelial cells and the fibroblasts and putative myoepithelial cells provided a simple means of quantitating stromal and myoepithelial cell contamination of the mammary epithelial cells in culture. In vivo, normal tissues showed fibronectin primarily localized in the basement membrane surrounding the epithelial cells and in the stroma. Most primary carcinomas displayed powdery, punctate staining on the epithelial cells in addition to the fibronectin present in the surrounding stroma.

  4. Direct Differentiation of Human Pluripotent Stem Cells into Haploid Spermatogenic Cells

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    Charles A. Easley, IV

    2012-09-01

    Full Text Available Human embryonic stem cells (hESCs and induced pluripotent stem cells (hiPSCs have been shown to differentiate into primordial germ cells (PGCs but not into spermatogonia, haploid spermatocytes, or spermatids. Here, we show that hESCs and hiPSCs differentiate directly into advanced male germ cell lineages, including postmeiotic, spermatid-like cells, in vitro without genetic manipulation. Furthermore, our procedure mirrors spermatogenesis in vivo by differentiating PSCs into UTF1-, PLZF-, and CDH1-positive spermatogonia-like cells; HIWI- and HILI-positive spermatocyte-like cells; and haploid cells expressing acrosin, transition protein 1, and protamine 1 (proteins that are uniquely found in spermatids and/or sperm. These spermatids show uniparental genomic imprints similar to those of human sperm on two loci: H19 and IGF2. These results demonstrate that male PSCs have the ability to differentiate directly into advanced germ cell lineages and may represent a novel strategy for studying spermatogenesis in vitro.

  5. Microenvironment promotes tumor cell reprogramming in human breast cancer cell lines.

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    Fabrizio D'Anselmi

    Full Text Available The microenvironment drives mammary gland development and function, and may influence significantly both malignant behavior and cell growth of mammary cancer cells. By restoring context, and forcing cells to properly interpret native signals from the microenvironment, the cancer cell aberrant behavior can be quelled, and organization re-established. In order to restore functional and morphological differentiation, human mammary MCF-7 and MDA-MB-231 cancer cells were allowed to grow in a culture medium filled with a 10% of the albumen (EW, Egg White from unfertilized chicken egg. That unique microenvironment behaves akin a 3D culture and induces MCF-7 cells to produce acini and branching duct-like structures, distinctive of mammary gland differentiation. EW-treated MDA-MB-231 cells developed buds of acini and duct-like structures. Both MCF-7 and MDA-MB-231 cells produced β-casein, a key milk component. Furthermore, E-cadherin expression was reactivated in MDA-MB-231 cells, as a consequence of the increased cdh1 expression; meanwhile β-catenin - a key cytoskeleton component - was displaced behind the inner cell membrane. Such modification hinders the epithelial-mesenchymal transition in MDA-MB-231 cells. This differentiating pathway is supported by the contemporary down-regulation of canonical pluripotency markers (Klf4, Nanog. Given that egg-conditioned medium behaves as a 3D-medium, it is likely that cancer phenotype reversion could be ascribed to the changed interactions between cells and their microenvironment.

  6. Maturing human pluripotent stem cell-derived cardiomyocytes in human engineered cardiac tissues.

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    Feric, Nicole T; Radisic, Milica

    2016-01-15

    Engineering functional human cardiac tissue that mimics the native adult morphological and functional phenotype has been a long held objective. In the last 5 years, the field of cardiac tissue engineering has transitioned from cardiac tissues derived from various animal species to the production of the first generation of human engineered cardiac tissues (hECTs), due to recent advances in human stem cell biology. Despite this progress, the hECTs generated to date remain immature relative to the native adult myocardium. In this review, we focus on the maturation challenge in the context of hECTs, the present state of the art, and future perspectives in terms of regenerative medicine, drug discovery, preclinical safety testing and pathophysiological studies. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Laser printing of skin cells and human stem cells.

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    Koch, Lothar; Kuhn, Stefanie; Sorg, Heiko; Gruene, Martin; Schlie, Sabrina; Gaebel, Ralf; Polchow, Bianca; Reimers, Kerstin; Stoelting, Stephanie; Ma, Nan; Vogt, Peter M; Steinhoff, Gustav; Chichkov, Boris

    2010-10-01

    Laser printing based on laser-induced forward transfer (LIFT) is a new biofabrication technique for the arrangement of biological materials or living cells in well-defined patterns. In the current study, skin cell lines (fibroblasts/keratinocytes) and human mesenchymal stem cells (hMSC) were chosen for laser printing experiments due to their high potential in regeneration of human skin and new application possibilities of stem cell therapy. To evaluate the influence of LIFT on the cells, their survival rate, their proliferation and apoptotic activity, and the DNA damages and modifications of their cell surface markers were assessed and statistically evaluated over several days. The cells survived the transfer procedure with a rate of 98%  +/- 1% standard error of the mean (skin cells) and 90%  +/- 10% (hMSC), respectively. All used cell types maintain their ability to proliferate after LIFT. Further, skin cells and hMSC did not show an increase of apoptosis or DNA fragmentation. In addition, the hMSC keep their phenotype as proven by fluorescence activated cell sorting (FACS) analysis. This study demonstrates LIFT as a suitable technique for unharmed computer-controlled positioning of different cell types and a promising tool for future applications in the ex vivo generation of tissue replacements.

  8. Dynamics of sleep stage transitions in healthy humans and patients with chronic fatigue syndrome.

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    Kishi, Akifumi; Struzik, Zbigniew R; Natelson, Benjamin H; Togo, Fumiharu; Yamamoto, Yoshiharu

    2008-06-01

    Physiological and/or pathological implications of the dynamics of sleep stage transitions have not, to date, been investigated. We report detailed duration and transition statistics between sleep stages in healthy subjects and in others with chronic fatigue syndrome (CFS); in addition, we also compare our data with previously published results for rats. Twenty-two healthy females and 22 female patients with CFS, characterized by complaints of unrefreshing sleep, underwent one night of polysomnographic recording. We find that duration of deep sleep (stages III and IV) follows a power-law probability distribution function; in contrast, stage II sleep durations follow a stretched exponential and stage I, and REM sleep durations follow an exponential function. These stage duration distributions show a gradually increasing departure from the exponential form with increasing depth of sleep toward a power-law type distribution for deep sleep, suggesting increasing complexity of regulation of deeper sleep stages. We also find a substantial number of REM to non-REM sleep transitions in humans, while this transition is reported to be virtually nonexistent in rats. The relative frequency of this REM to non-REM sleep transition is significantly lower in CFS patients than in controls, resulting in a significantly greater relative transition frequency of moving from both REM and stage I sleep to awake. Such an alteration in the transition pattern suggests that the normal continuation of sleep in light or REM sleep is disrupted in CFS. We conclude that dynamic transition analysis of sleep stages is useful for elucidating yet-to-be-determined human sleep regulation mechanisms with pathophysiological implications.

  9. Canine Mammary Cancer Stem Cells are Radio- and Chemo- Resistant and Exhibit an Epithelial-Mesenchymal Transition Phenotype.

    Science.gov (United States)

    Pang, Lisa Y; Cervantes-Arias, Alejandro; Else, Rod W; Argyle, David J

    2011-03-30

    Canine mammary carcinoma is the most common cancer among female dogs and is often fatal due to the development of distant metastases. In humans, solid tumors are made up of heterogeneous cell populations, which perform different roles in the tumor economy. A small subset of tumor cells can hold or acquire stem cell characteristics, enabling them to drive tumor growth, recurrence and metastasis. In veterinary medicine, the molecular drivers of canine mammary carcinoma are as yet undefined. Here we report that putative cancer stem cells (CSCs) can be isolated form a canine mammary carcinoma cell line, REM134. We show that these cells have an increased ability to form tumorspheres, a characteristic of stem cells, and that they express embryonic stem cell markers associated with pluripotency. Moreover, canine CSCs are relatively resistant to the cytotoxic effects of common chemotherapeutic drugs and ionizing radiation, indicating that failure of clinical therapy to eradicate canine mammary cancer may be due to the survival of CSCs. The epithelial to mesenchymal transition (EMT) has been associated with cancer invasion, metastasis, and the acquisition of stem cell characteristics. Our results show that canine CSCs predominantly express mesenchymal markers and are more invasive than parental cells, indicating that these cells have a mesenchymal phenotype. Furthermore, we show that canine mammary cancer cells can be induced to undergo EMT by TGFβ and that these cells have an increased ability to form tumorspheres. Our findings indicate that EMT induction can enrich for cells with CSC properties, and provide further insight into canine CSC biology.

  10. Canine Mammary Cancer Stem Cells are Radio- and Chemo-Resistant and Exhibit an Epithelial-Mesenchymal Transition Phenotype

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    Pang, Lisa Y., E-mail: lisa.pang@ed.ac.uk; Cervantes-Arias, Alejandro; Else, Rod W.; Argyle, David J. [Royal (Dick) School of Veterinary Studies and Roslin Institute, The University of Edinburgh, Easter Bush, Midlothian, EH25 9RG (United Kingdom)

    2011-03-30

    Canine mammary carcinoma is the most common cancer among female dogs and is often fatal due to the development of distant metastases. In humans, solid tumors are made up of heterogeneous cell populations, which perform different roles in the tumor economy. A small subset of tumor cells can hold or acquire stem cell characteristics, enabling them to drive tumor growth, recurrence and metastasis. In veterinary medicine, the molecular drivers of canine mammary carcinoma are as yet undefined. Here we report that putative cancer stem cells (CSCs) can be isolated form a canine mammary carcinoma cell line, REM134. We show that these cells have an increased ability to form tumorspheres, a characteristic of stem cells, and that they express embryonic stem cell markers associated with pluripotency. Moreover, canine CSCs are relatively resistant to the cytotoxic effects of common chemotherapeutic drugs and ionizing radiation, indicating that failure of clinical therapy to eradicate canine mammary cancer may be due to the survival of CSCs. The epithelial to mesenchymal transition (EMT) has been associated with cancer invasion, metastasis, and the acquisition of stem cell characteristics. Our results show that canine CSCs predominantly express mesenchymal markers and are more invasive than parental cells, indicating that these cells have a mesenchymal phenotype. Furthermore, we show that canine mammary cancer cells can be induced to undergo EMT by TGFβ and that these cells have an increased ability to form tumorspheres. Our findings indicate that EMT induction can enrich for cells with CSC properties, and provide further insight into canine CSC biology.

  11. Human spleen and red blood cells

    Science.gov (United States)

    Pivkin, Igor; Peng, Zhangli; Karniadakis, George; Buffet, Pierre; Dao, Ming

    2016-11-01

    Spleen plays multiple roles in the human body. Among them is removal of old and altered red blood cells (RBCs), which is done by filtering cells through the endothelial slits, small micron-sized openings. There is currently no experimental technique available that allows us to observe RBC passage through the slits. It was previously noticed that people without a spleen have less deformable red blood cells, indicating that the spleen may play a role in defining the size and shape of red blood cells. We used detailed RBC model implemented within the Dissipative Particle Dynamics (DPD) simulation framework to study the filter function of the spleen. Our results demonstrate that spleen indeed plays major role in defining the size and shape of the healthy human red blood cells.

  12. Rotating cell culture systems for human cell culture: human trophoblast cells as a model.

    Science.gov (United States)

    Zwezdaryk, Kevin J; Warner, Jessica A; Machado, Heather L; Morris, Cindy A; Höner zu Bentrup, Kerstin

    2012-01-18

    The field of human trophoblast research aids in understanding the complex environment established during placentation. Due to the nature of these studies, human in vivo experimentation is impossible. A combination of primary cultures, explant cultures and trophoblast cell lines support our understanding of invasion of the uterine wall and remodeling of uterine spiral arteries by extravillous trophoblast cells (EVTs), which is required for successful establishment of pregnancy. Despite the wealth of knowledge gleaned from such models, it is accepted that in vitro cell culture models using EVT-like cell lines display altered cellular properties when compared to their in vivo counterparts. Cells cultured in the rotating cell culture system (RCCS) display morphological, phenotypic, and functional properties of EVT-like cell lines that more closely mimic differentiating in utero EVTs, with increased expression of genes mediating invasion (e.g. matrix metalloproteinases (MMPs)) and trophoblast differentiation. The Saint Georges Hospital Placental cell Line-4 (SGHPL-4) (kindly donated by Dr. Guy Whitley and Dr. Judith Cartwright) is an EVT-like cell line that was used for testing in the RCCS. The design of the RCCS culture vessel is based on the principle that organs and tissues function in a three-dimensional (3-D) environment. Due to the dynamic culture conditions in the vessel, including conditions of physiologically relevant shear, cells grown in three dimensions form aggregates based on natural cellular affinities and differentiate into organotypic tissue-like assemblies. The maintenance of a fluid orbit provides a low-shear, low-turbulence environment similar to conditions found in vivo. Sedimentation of the cultured cells is countered by adjusting the rotation speed of the RCCS to ensure a constant free-fall of cells. Gas exchange occurs through a permeable hydrophobic membrane located on the back of the bioreactor. Like their parental tissue in vivo, RCCS

  13. Autophagy in human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Thien Tra

    Full Text Available Autophagy (macroautophagy is a degradative process that involves the sequestration of cytosolic material including organelles into double membrane vesicles termed autophagosomes for delivery to the lysosome. Autophagy is essential for preimplantation development of mouse embryos and cavitation of embryoid bodies. The precise roles of autophagy during early human embryonic development, remain however largely uncharacterized. Since human embryonic stem cells constitute a unique model system to study early human embryogenesis we investigated the occurrence of autophagy in human embryonic stem cells. We have, using lentiviral transduction, established multiple human embryonic stem cell lines that stably express GFP-LC3, a fluorescent marker for the autophagosome. Each cell line displays both a normal karyotype and pluripotency as indicated by the presence of cell types representative of the three germlayers in derived teratomas. GFP expression and labelling of autophagosomes is retained after differentiation. Baseline levels of autophagy detected in cultured undifferentiated hESC were increased or decreased in the presence of rapamycin and wortmannin, respectively. Interestingly, autophagy was upregulated in hESCs induced to undergo differentiation by treatment with type I TGF-beta receptor inhibitor SB431542 or removal of MEF secreted maintenance factors. In conclusion we have established hESCs capable of reporting macroautophagy and identify a novel link between autophagy and early differentiation events in hESC.

  14. Reduced Number of Transitional and Naive B Cells in Addition to Decreased BAFF Levels in Response to the T Cell Independent Immunogen Pneumovax®23.

    Science.gov (United States)

    Roth, Alena; Glaesener, Stephanie; Schütz, Katharina; Meyer-Bahlburg, Almut

    2016-01-01

    Protective immunity against T cell independent (TI) antigens such as Streptococcus pneumoniae is characterized by antibody production of B cells induced by the combined activation of T cell independent type 1 and type 2 antigens in the absence of direct T cell help. In mice, the main players in TI immune responses have been well defined as marginal zone (MZ) B cells and B-1 cells. However, the existence of human equivalents to these B cell subsets and the nature of the human B cell compartment involved in the immune reaction remain elusive. We therefore analyzed the effect of a TI antigen on the B cell compartment through immunization of healthy individuals with the pneumococcal polysaccharide (PnPS)-based vaccine Pneumovax®23, and subsequent characterization of B cell subpopulations. Our data demonstrates a transient decrease of transitional and naïve B cells, with a concomitant increase of IgA+ but not IgM+ or IgG+ memory B cells and a predominant generation of PnPS-specific IgA+ producing plasma cells. No alterations could be detected in T cells, or proposed human B-1 and MZ B cell equivalents. Consistent with the idea of a TI immune response, antigen-specific memory responses could not be observed. Finally, BAFF, which is supposed to drive class switching to IgA, was unexpectedly found to be decreased in serum in response to Pneumovax®23. Our results demonstrate that a characteristic TI response induced by Pneumovax®23 is associated with distinct phenotypical and functional changes within the B cell compartment. Those modulations occur in the absence of any modulations of T cells and without the development of a specific memory response.

  15. Reduced Number of Transitional and Naive B Cells in Addition to Decreased BAFF Levels in Response to the T Cell Independent Immunogen Pneumovax®23.

    Directory of Open Access Journals (Sweden)

    Alena Roth

    Full Text Available Protective immunity against T cell independent (TI antigens such as Streptococcus pneumoniae is characterized by antibody production of B cells induced by the combined activation of T cell independent type 1 and type 2 antigens in the absence of direct T cell help. In mice, the main players in TI immune responses have been well defined as marginal zone (MZ B cells and B-1 cells. However, the existence of human equivalents to these B cell subsets and the nature of the human B cell compartment involved in the immune reaction remain elusive. We therefore analyzed the effect of a TI antigen on the B cell compartment through immunization of healthy individuals with the pneumococcal polysaccharide (PnPS-based vaccine Pneumovax®23, and subsequent characterization of B cell subpopulations. Our data demonstrates a transient decrease of transitional and naïve B cells, with a concomitant increase of IgA+ but not IgM+ or IgG+ memory B cells and a predominant generation of PnPS-specific IgA+ producing plasma cells. No alterations could be detected in T cells, or proposed human B-1 and MZ B cell equivalents. Consistent with the idea of a TI immune response, antigen-specific memory responses could not be observed. Finally, BAFF, which is supposed to drive class switching to IgA, was unexpectedly found to be decreased in serum in response to Pneumovax®23. Our results demonstrate that a characteristic TI response induced by Pneumovax®23 is associated with distinct phenotypical and functional changes within the B cell compartment. Those modulations occur in the absence of any modulations of T cells and without the development of a specific memory response.

  16. Human embryonic stem cells for neuronal repair.

    Science.gov (United States)

    Ben-Hur, Tamir

    2006-02-01

    Human embryonic stem cells may serve as a potentially endeless source of transplantable cells to treat various neurologic disorders. Accumulating data have shown the therapeutic value of various neural precursor cell types in experimental models of neurologic diseases. Tailoring cell therapy for specific disorders requires the generation of cells that are committed to specific neural lineages. To this end, protocols were recently developed for the derivation of dopaminergic neurons, spinal motor neurons and oligodendrocytes from hESC. These protocols recapitulate normal development in culture conditions. However, a novel concept emerging from these studies is that the beneficial effect of transplanted stem cells is not only via cell replacement in damaged host tissue, but also by trophic and protective effects, as well as by an immunomodulatory effect that down-regulates detrimental brain inflammation.

  17. Transcriptional regulation is a major controller of cell cycle transition dynamics

    DEFF Research Database (Denmark)

    Romanel, Alessandro; Jensen, Lars Juhl; Cardelli, Luca

    2012-01-01

    in various organisms showed the importance of positive feedbacks in other transitions as well. Here we investigate if a universal control system with transcriptional regulation(s) and post-translational positive feedback(s) can be proposed for the regulation of all cell cycle transitions. Through...... computational modeling, we analyze the transition dynamics in all possible combinations of transcriptional and post-translational regulations. We find that some combinations lead to 'sloppy' transitions, while others give very precise control. The periodic transcriptional regulation through the activator...... or the inhibitor leads to radically different dynamics. Experimental evidence shows that in cell cycle transitions of organisms investigated for cell cycle dependent periodic transcription, only the inhibitor OR the activator is under cyclic control and never both of them. Based on these observations, we propose...

  18. Novel agents inhibit human leukemic cells

    Institute of Scientific and Technical Information of China (English)

    Wei-ping YU; Juan LI

    2012-01-01

    Ouabain (OUA) and pyrithione zinc (PZ) have been proved as the potential drugs for treating acute myeloid leukemia (AML).Selected from a screening among 1040 Food and Drug Administration-approved pharmacological agents,both drugs showability to induce apoptosis of the culturing AML cells,exhibiting the poisoning effect on the cells.Studies also reveal the efficiency of the drugs in inhibiting the growth of human AML cells injected into the mice lacking of immunity and killing primary AML cells from the peripheral blood of AML patients[1].

  19. 3 CFR - Guidelines for Human Stem Cell Research

    Science.gov (United States)

    2010-01-01

    ... 3 The President 1 2010-01-01 2010-01-01 false Guidelines for Human Stem Cell Research Presidential Documents Other Presidential Documents Memorandum of July 30, 2009 Guidelines for Human Stem Cell Research..., scientifically worthy human stem cell research, including human embryonic stem cell research, to the extent...

  20. Myristoylation profiling in human cells and zebrafish

    Directory of Open Access Journals (Sweden)

    Malgorzata Broncel

    2015-09-01

    Full Text Available Human cells (HEK 293, HeLa, MCF-7 and zebrafish embryos were metabolically tagged with an alkynyl myristic acid probe, lysed with an SDS buffer and tagged proteomes ligated to multifunctional capture reagents via copper-catalyzed alkyne azide cycloaddition (CuAAC. This allowed for affinity enrichment and high-confidence identification, by delivering direct MS/MS evidence for the modification site, of 87 and 61 co-translationally myristoylated proteins in human cells and zebrafish, respectively. The data have been deposited to ProteomeXchange Consortium (Vizcaíno et al., 2014 Nat. Biotechnol., 32, 223–6 (PXD001863 and PXD001876 and are described in detail in Multifunctional reagents for quantitative proteome-wide analysis of protein modification in human cells and dynamic protein lipidation during vertebrate development׳ by Broncel et al., Angew. Chem. Int. Ed.

  1. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins

    Science.gov (United States)

    Fukusumi, Hayato; Shofuda, Tomoko; Bamba, Yohei; Yamamoto, Atsuyo; Kanematsu, Daisuke; Handa, Yukako; Okita, Keisuke; Nakamura, Masaya; Yamanaka, Shinya; Okano, Hideyuki; Kanemura, Yonehiro

    2016-01-01

    Human neural progenitor cells (hNPCs) have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC) clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB) formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi). Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes. PMID:27212953

  2. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins.

    Science.gov (United States)

    Fukusumi, Hayato; Shofuda, Tomoko; Bamba, Yohei; Yamamoto, Atsuyo; Kanematsu, Daisuke; Handa, Yukako; Okita, Keisuke; Nakamura, Masaya; Yamanaka, Shinya; Okano, Hideyuki; Kanemura, Yonehiro

    2016-01-01

    Human neural progenitor cells (hNPCs) have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC) clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB) formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi). Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes.

  3. The human fetal lymphocyte lineage: identification by CD27 and LIN28B expression in B cell progenitors

    Science.gov (United States)

    McWilliams, Laurie; Su, Kuei-Ying; Liang, Xiaoe; Liao, Dongmei; Floyd, Serina; Amos, Joshua; Moody, M. Anthony; Kelsoe, Garnett; Kuraoka, Masayuki

    2013-01-01

    CD27, a member of the TNFR superfamily, is used to identify human memory B cells. Nonetheless, CD27+ B cells are present in patients with HIGM1 syndrome who are unable to generate GCs or memory B cells. CD27+IgD+ fetal B cells are present in umbilical cord blood, and CD27 may also be a marker of the human B1-like B cells. To define the origin of naïve CD27+IgD+ human B cells, we studied B cell development in both fetal and adult tissues. In human FL, most CD19+ cells coexpressed CD10, a marker of human developing B cells. Some CD19+CD10+ B cells expressed CD27, and these fetal CD27+ cells were present in the pro-B, pre-B, and immature/transitional B cell compartments. Lower frequencies of phenotypically identical cells were also identified in adult BM. CD27+ pro-B, pre-B, and immature/transitional B cells expressed recombination activating gene-1, terminal deoxynucleotidyl transferase and Vpre-B mRNA comparably to their CD27− counterparts. CD27+ and CD27− developing B cells showed similar Ig heavy chain gene usage with low levels of mutations, suggesting that CD27+ developing B cells are distinct from mutated memory B cells. Despite these similarities, CD27+ developing B cells differed from CD27− developing B cells by their increased expression of LIN28B, a transcription factor associated with the fetal lymphoid lineages of mice. Furthermore, CD27+ pro-B cells efficiently generated IgM+IgD+ immature/transitional B cells in vitro. Our observations suggest that CD27 expression during B cell development identifies a physiologic state or lineage for human B cell development distinct from the memory B cell compartment. PMID:23901121

  4. Massachusetts Fuel Cell Bus Project: Demonstrating a Total Transit Solution for Fuel Cell Electric Buses in Boston

    Energy Technology Data Exchange (ETDEWEB)

    2017-05-22

    The Federal Transit Administration's National Fuel Cell Bus Program focuses on developing commercially viable fuel cell bus technologies. Nuvera is leading the Massachusetts Fuel Cell Bus project to demonstrate a complete transit solution for fuel cell electric buses that includes one bus and an on-site hydrogen generation station for the Massachusetts Bay Transportation Authority (MBTA). A team consisting of ElDorado National, BAE Systems, and Ballard Power Systems built the fuel cell electric bus, and Nuvera is providing its PowerTap on-site hydrogen generator to provide fuel for the bus.

  5. CCND1-CDK4-mediated cell cycle progression provides a competitive advantage for human hematopoietic stem cells in vivo.

    Science.gov (United States)

    Mende, Nicole; Kuchen, Erika E; Lesche, Mathias; Grinenko, Tatyana; Kokkaliaris, Konstantinos D; Hanenberg, Helmut; Lindemann, Dirk; Dahl, Andreas; Platz, Alexander; Höfer, Thomas; Calegari, Federico; Waskow, Claudia

    2015-07-27

    Maintenance of stem cell properties is associated with reduced proliferation. However, in mouse hematopoietic stem cells (HSCs), loss of quiescence results in a wide range of phenotypes, ranging from functional failure to extensive self-renewal. It remains unknown whether the function of human HSCs is controlled by the kinetics of cell cycle progression. Using human HSCs and human progenitor cells (HSPCs), we report here that elevated levels of CCND1-CDK4 complexes promoted the transit from G0 to G1 and shortened the G1 cell cycle phase, resulting in protection from differentiation-inducing signals in vitro and increasing human leukocyte engraftment in vivo. Further, CCND1-CDK4 overexpression conferred a competitive advantage without impacting HSPC numbers. In contrast, accelerated cell cycle progression mediated by elevated levels of CCNE1-CDK2 led to the loss of functional HSPCs in vivo. Collectively, these data suggest that the transition kinetics through the early cell cycle phases are key regulators of human HSPC function and important for lifelong hematopoiesis. © 2015 Mende et al.

  6. Efficient derivation and genetic modifications of human pluripotent stem cells on engineered human feeder cell lines.

    Science.gov (United States)

    Zou, Chunlin; Chou, Bin-Kuan; Dowey, Sarah N; Tsang, Kitman; Huang, Xiaosong; Liu, Cyndi F; Smith, Cory; Yen, Jonathan; Mali, Prashant; Zhang, Yu Alex; Cheng, Linzhao; Ye, Zhaohui

    2012-08-10

    Derivation of pluripotent stem cells (iPSCs) induced from somatic cell types and the subsequent genetic modifications of disease-specific or patient-specific iPSCs are crucial steps in their applications for disease modeling as well as future cell and gene therapies. Conventional procedures of these processes require co-culture with primary mouse embryonic fibroblasts (MEFs) to support self-renewal and clonal growth of human iPSCs as well as embryonic stem cells (ESCs). However, the variability of MEF quality affects the efficiencies of all these steps. Furthermore, animal sourced feeders may hinder the clinical applications of human stem cells. In order to overcome these hurdles, we established immortalized human feeder cell lines by stably expressing human telomerase reverse transcriptase, Wnt3a, and drug resistance genes in adult mesenchymal stem cells. Here, we show that these immortalized human feeders support efficient derivation of virus-free, integration-free human iPSCs and long-term expansion of human iPSCs and ESCs. Moreover, the drug-resistance feature of these feeders also supports nonviral gene transfer and expression at a high efficiency, mediated by piggyBac DNA transposition. Importantly, these human feeders exhibit superior ability over MEFs in supporting homologous recombination-mediated gene targeting in human iPSCs, allowing us to efficiently target a transgene into the AAVS1 safe harbor locus in recently derived integration-free iPSCs. Our results have great implications in disease modeling and translational applications of human iPSCs, as these engineered human cell lines provide a more efficient tool for genetic modifications and a safer alternative for supporting self-renewal of human iPSCs and ESCs.

  7. Resveratrol Induces Apoptosis in Human Osteosarcoma MG63 Cells

    Institute of Scientific and Technical Information of China (English)

    Yan Liu; Xin Wang; Yuxin Xie; Jingui Zhang; Qingshan Wang; Xianhui Xu

    2008-01-01

    OBJECTIVE To investigate apoptosis in human osteosarcoma MG63 cells induced by resveratrol and the molecular mechanism involved.METHODS MG63 cells were treated with different concentrations of resveratrol and transmission electron microscopy was used to observe morphological changes occurring in apoptosis.The MTT method was used to determine the inhibitory rate and flow cytometry was used to assess apoptosis and to analyze the expression of the p21ciP1/WAF1 and survivin proteins;the expression of p21ciP1/WAF1 and survivin mRNAs was analyzed by the reverse transcriptase polymerase chain reaction (RT-PCR).RESULTS After resveratrol treatment,the growth of the MG63 cells was significantly inhibited in a time- and dose-dependent fashion.By transmission electron microscopy,the cells displayed morphological changes characteristic of apoptosis,including formation of cytoplasmic vacuoles,chromatin condensation and margination.Flow cytometry showed that the growth of the cells was inhibited after resveratrol (10 mg/L and 20 mg/L) treatment.The inhibitory rates were (11.9 ±0.63)% and (19.7 ± 0.88)%respectively.The quantity of treated cells in G0/G1 transition was increased,but the number in the S phase and G2/M transition was decreased.A subdiploid peak was observed.The expression of p21ciP1/WAF1 was up-regulated while survivin was down-regulated.CONCLUSION Resveratrol can inhibit growth and induce apoptosis of MG63 cells.Its molecular mechanism might be related to modulation of survivin and p21ciP1/WAF1 expression.

  8. [Human pluripotent stem cell and neural differentiation].

    Science.gov (United States)

    Wataya, Takafumi; Muguruma, Keiko; Sasai, Yoshiki

    2008-10-01

    Recovery of lost brain function is an important issue in medical studies because neurons of the central nervous system (CNS) have poor potential for regeneration. Since few CNS diseases can be treated completely by medicines, regenerative therapy by using stem cells should be studied as a new type of therapeutic intervention. The efficacy of cell replacement therapy in Parkinson's disease has been well investigated. Several studies on fetal tissue transplantation have revealed that quantity and purity of transplanted cells are necessary for recovery of symptoms. SFEB (Serum-free floating culture of embryoid body-like aggregates) method is capable of inducing multi-potential CNS progenitors that can be steered to differentiate into region-specific tissues. On the basis of the existing knowledge of embryology, we have succeeded in the generating of various types of neurons such as telencephalic, cerebeller (Purkinje and granule cells), retinal (photoreceptor cells) and hypothalamic neurons. Application of this culture method to human ES (hES) cells is necessary for clinical purpose: however, poor survival of hES cells in SFEB culture might limit the possibility of using these cells for future medical applications. We found that a selective Rho-associated kinase (ROCK) inhibitor, Y-27632, markedly diminished the dissociation-induced apoptosis of hES cells and enabled the cells to form aggregates in SFEB culture. For both mouse and human ES cells, SFEB culture is a favorable method that can generate large amounts of region-specific neurons. However, stem cell-based therapy continues to face several obstacles. It is important that researchers in the basic sciences and clinical medicine should discuss these problems together to overcome both scientific and ethical issues related to stem cells.

  9. The origins of the Acheulean: past and present perspectives on a major transition in human evolution.

    Science.gov (United States)

    de la Torre, Ignacio

    2016-07-01

    The emergence of the Acheulean from the earlier Oldowan constitutes a major transition in human evolution, the theme of this special issue. This paper discusses the evidence for the origins of the Acheulean, a cornerstone in the history of human technology, from two perspectives; firstly, a review of the history of investigations on Acheulean research is presented. This approach introduces the evolution of theories throughout the development of the discipline, and reviews the way in which cumulative knowledge led to the prevalent explanatory framework for the emergence of the Acheulean. The second part presents the current state of the art in Acheulean origins research, and reviews the hard evidence for the appearance of this technology in Africa around 1.7 Ma, and its significance for the evolutionary history of Homo erectusThis article is part of the themed issue 'Major transitions in human evolution'.

  10. T-cell response in human leishmaniasis

    DEFF Research Database (Denmark)

    Kharazmi, A; Kemp, K; Ismail, A

    1999-01-01

    In the present communication we provide evidence for the existence of a Th1/Th2 dichotomy in the T-cell response to Leishmania antigens in human leishmaniasis. Our data suggest that the pattern of IL-4 and IFN-gamma response is polarised in these patients. Lymphocytes from individuals recovered......+. Furthermore, IL-10 plays an important role in the development of post kala azar dermal leishmaniasis (PKDL) from VL. The balance between the parasitic-specific T-cell response plays an important regulatory role in determining the outcome of Leishmania infections in humans....

  11. Characterization of human pluripotent stem cells.

    Science.gov (United States)

    Gokhale, Paul J; Andrews, Peter W

    2013-12-18

    Human pluripotent stem cells (PSCs), whether embryonic stem cells or induced PSCs, offer enormous opportunities for regenerative medicine and other biomedical applications once we have developed the ability to harness their capacity for extensive differentiation. Central to this is our ability to identify and characterize such PSCs, but this is fraught with potential difficulties that arise from a tension between functional definitions of pluripotency and the more convenient use of 'markers', a problem exacerbated by ethical issues, our lack of knowledge of early human embryonic development, and differences from the mouse paradigm.

  12. Grape seed proanthocyanidins inhibit the invasive potential of head and neck cutaneous squamous cell carcinoma cells by targeting EGFR expression and epithelial-to-mesenchymal transition

    Directory of Open Access Journals (Sweden)

    Sun Qian

    2011-12-01

    Full Text Available Abstract Background Head and neck squamous cell carcinoma (HNSCC is responsible for over 20,000 deaths every year in United States. Most of the deaths are due, in large part, to its propensity to metastasize. We have examined the effect of bioactive component grape seed proanthocyanidins (GSPs on human cutaneous HNSCC cell invasion and the molecular mechanisms underlying these effects using SCC13 cell line as an in vitro model. Methods The therapeutic effects of GSPs on cancer cell invasion were studied using Boyden chamber and wound healing assays. The effects of GSPs on the levels of various proteins related with cancer cell invasion were determined using western blot analysis. Results Using in vitro cell invasion assays, we observed that treatment of SCC13 cells with GSPs resulted in a concentration-dependent inhibition of cell invasion of these cells, which was associated with a reduction in the levels of epidermal growth factor receptor (EGFR. Treatment of cells with gefitinib and erlotinib, inhibitors of EGFR, or transient transfection of SCC13 cells with EGFR small interfering RNA, also inhibited invasion of these cells. The inhibition of cell invasion by GSPs was associated with the inhibition of the phosphorylation of ERK1/2, a member of mitogen-activated protein kinase family. Treatment of cells with UO126, an inhibitor of MEK, also inhibited the invasion potential of SCC13 cells. Additionally, inhibition of human cutaneous HNSCC cell invasion by GSPs was associated with reversal of epithelial-to-mesenchymal transition (EMT process, which resulted in an increase in the levels of epithelial biomarker (E-cadherin while loss of mesenchymal biomarkers (vimentin, fibronectin and N-cadherin in cells. Similar effect on EMT biomarkers was also observed when cells were treated with erlotinib. Conclusion The results obtained from this study indicate that grape seed proanthocyanidins have the ability to inhibit the invasion of human cutaneous

  13. Grape seed proanthocyanidins inhibit the invasive potential of head and neck cutaneous squamous cell carcinoma cells by targeting EGFR expression and epithelial-to-mesenchymal transition.

    Science.gov (United States)

    Sun, Qian; Prasad, Ram; Rosenthal, Eben; Katiyar, Santosh K

    2011-12-21

    Head and neck squamous cell carcinoma (HNSCC) is responsible for over 20,000 deaths every year in United States. Most of the deaths are due, in large part, to its propensity to metastasize. We have examined the effect of bioactive component grape seed proanthocyanidins (GSPs) on human cutaneous HNSCC cell invasion and the molecular mechanisms underlying these effects using SCC13 cell line as an in vitro model. The therapeutic effects of GSPs on cancer cell invasion were studied using Boyden chamber and wound healing assays. The effects of GSPs on the levels of various proteins related with cancer cell invasion were determined using western blot analysis. Using in vitro cell invasion assays, we observed that treatment of SCC13 cells with GSPs resulted in a concentration-dependent inhibition of cell invasion of these cells, which was associated with a reduction in the levels of epidermal growth factor receptor (EGFR). Treatment of cells with gefitinib and erlotinib, inhibitors of EGFR, or transient transfection of SCC13 cells with EGFR small interfering RNA, also inhibited invasion of these cells. The inhibition of cell invasion by GSPs was associated with the inhibition of the phosphorylation of ERK1/2, a member of mitogen-activated protein kinase family. Treatment of cells with UO126, an inhibitor of MEK, also inhibited the invasion potential of SCC13 cells. Additionally, inhibition of human cutaneous HNSCC cell invasion by GSPs was associated with reversal of epithelial-to-mesenchymal transition (EMT) process, which resulted in an increase in the levels of epithelial biomarker (E-cadherin) while loss of mesenchymal biomarkers (vimentin, fibronectin and N-cadherin) in cells. Similar effect on EMT biomarkers was also observed when cells were treated with erlotinib. The results obtained from this study indicate that grape seed proanthocyanidins have the ability to inhibit the invasion of human cutaneous HNSCC cells by targeting the EGFR expression and reversing the

  14. Fuel Cell Transit Bus Evaluations: Joint Evaluation Plan for the U.S. Department of Energy and the Federal Transit Administration (Report and Appendix)

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, L.; Chandler, K.

    2010-11-01

    This document describes the fuel cell transit bus evaluations performed by the National Renewable Energy Laboratory (NREL) and funded by the U.S. Department of Energy (DOE) and the U.S. Department of Transportation's Federal Transit Administration (FTA). This document provides a description of the demonstration sites, funding sources, and data collection activities for fuel cell transit bus evaluations currently planned from FY10 through FY12.

  15. CLOSTRIDIUM SPORE ATTACHMENT TO HUMAN CELLS

    Energy Technology Data Exchange (ETDEWEB)

    PANESSA-WARREN,B.; TORTORA,G.; WARREN,J.

    1997-08-10

    This paper uses high resolution scanning electron microscopy (SEM) with a LaB6 gun and the newest commercial field emission guns, to obtain high magnification images of intact clostridial spores throughout the activation/germination/outgrowth process. By high resolution SEM, the clostridial exosporial membrane can be seen to produce numerous delicate projections (following activation), that extend from the exosporial surface to a nutritive substrate (agar), or cell surface when anaerobically incubated in the presence of human cells (embryonic fibroblasts and colon carcinoma cells). Magnifications of 20,000 to 200,000Xs at accelerating voltages low enough to minimize or eliminate specimen damage (1--5 kV) have permitted the entire surface of C.sporogenes and C.difficile endospores to be examined during all stages of germination. The relationships between the spore and the agar or human cell surface were also clearly visible.

  16. Human pluripotent stem cells in contemporary medicine

    Directory of Open Access Journals (Sweden)

    S. A. Rodin

    2015-01-01

    Full Text Available Human pluripotent stem cells (hPSCs are capable of indefinite proliferation and can be differentiated into any cell type of the human body. Therefore, they are a promising source of cells for treatment of numerous degenerative diseases and injuries. Pluripotent stem cells are also associated with a number of ethical, safety and technological issues. In this review, we describe various types of hPSCs, safety issues that concern all or some types of hPSCs and methods of clinical-grade hPSC line development. Also, we discuss current and past clinical trials involving hPSCs, their outcomes and future perspectives of hPSC-based therapy. 

  17. Platelets alter tumor cell attributes to propel metastasis: programming in transit.

    Science.gov (United States)

    Gay, Laurie J; Felding-Habermann, Brunhilde

    2011-11-15

    Metastasis of epithelial tumors critically depends on acquisition of a disseminating phenotype that allows tumor cells to colonize distant organs. In this issue of Cancer Cell, Labelle et al. demonstrate that an epithelial-mesenchymal-like transition can be induced by interaction between platelets and tumor cells.

  18. Cell-Cycle-Dependent Reconfiguration of the DNA Methylome during Terminal Differentiation of Human B Cells into Plasma Cells.

    Science.gov (United States)

    Caron, Gersende; Hussein, Mourad; Kulis, Marta; Delaloy, Céline; Chatonnet, Fabrice; Pignarre, Amandine; Avner, Stéphane; Lemarié, Maud; Mahé, Elise A; Verdaguer-Dot, Núria; Queirós, Ana C; Tarte, Karin; Martín-Subero, José I; Salbert, Gilles; Fest, Thierry

    2015-11-03

    Molecular mechanisms underlying terminal differentiation of B cells into plasma cells are major determinants of adaptive immunity but remain only partially understood. Here we present the transcriptional and epigenomic landscapes of cell subsets arising from activation of human naive B cells and differentiation into plasmablasts. Cell proliferation of activated B cells was linked to a slight decrease in DNA methylation levels, but followed by a committal step in which an S phase-synchronized differentiation switch was associated with an extensive DNA demethylation and local acquisition of 5-hydroxymethylcytosine at enhancers and genes related to plasma cell identity. Downregulation of both TGF-?1/SMAD3 signaling and p53 pathway supported this final step, allowing the emergence of a CD23-negative subpopulation in transition from B cells to plasma cells. Remarkably, hydroxymethylation of PRDM1, a gene essential for plasma cell fate, was coupled to progression in S phase, revealing an intricate connection among cell cycle, DNA (hydroxy)methylation, and cell fate determination.

  19. Cell-Cycle-Dependent Reconfiguration of the DNA Methylome during Terminal Differentiation of Human B Cells into Plasma Cells

    Directory of Open Access Journals (Sweden)

    Gersende Caron

    2015-11-01

    Full Text Available Molecular mechanisms underlying terminal differentiation of B cells into plasma cells are major determinants of adaptive immunity but remain only partially understood. Here we present the transcriptional and epigenomic landscapes of cell subsets arising from activation of human naive B cells and differentiation into plasmablasts. Cell proliferation of activated B cells was linked to a slight decrease in DNA methylation levels, but followed by a committal step in which an S phase-synchronized differentiation switch was associated with an extensive DNA demethylation and local acquisition of 5-hydroxymethylcytosine at enhancers and genes related to plasma cell identity. Downregulation of both TGF-β1/SMAD3 signaling and p53 pathway supported this final step, allowing the emergence of a CD23-negative subpopulation in transition from B cells to plasma cells. Remarkably, hydroxymethylation of PRDM1, a gene essential for plasma cell fate, was coupled to progression in S phase, revealing an intricate connection among cell cycle, DNA (hydroxymethylation, and cell fate determination.

  20. General Information about Transitional Cell Cancer of the Renal Pelvis and Ureter

    Science.gov (United States)

    ... Renal Pelvis and Ureter Treatment (PDQ®)–Patient Version General Information About Transitional Cell Cancer of the Renal ... through the urethra and leaves the body. Enlarge Anatomy of the male urinary system (left panel) and ...

  1. Vascular endothelial cell membranes differentiate between stretch and shear stress through transitions in their lipid phases.

    Science.gov (United States)

    Yamamoto, Kimiko; Ando, Joji

    2015-10-01

    Vascular endothelial cells (ECs) respond to the hemodynamic forces stretch and shear stress by altering their morphology, functions, and gene expression. However, how they sense and differentiate between these two forces has remained unknown. Here we report that the plasma membrane itself differentiates between stretch and shear stress by undergoing transitions in its lipid phases. Uniaxial stretching and hypotonic swelling increased the lipid order of human pulmonary artery EC plasma membranes, thereby causing a transition from the liquid-disordered phase to the liquid-ordered phase in some areas, along with a decrease in membrane fluidity. In contrast, shear stress decreased the membrane lipid order and increased membrane fluidity. A similar increase in lipid order occurred when the artificial lipid bilayer membranes of giant unilamellar vesicles were stretched by hypotonic swelling, indicating that this is a physical phenomenon. The cholesterol content of EC plasma membranes significantly increased in response to stretch but clearly decreased in response to shear stress. Blocking these changes in the membrane lipid order by depleting membrane cholesterol with methyl-β-cyclodextrin or by adding cholesterol resulted in a marked inhibition of the EC response specific to stretch and shear stress, i.e., phosphorylation of PDGF receptors and phosphorylation of VEGF receptors, respectively. These findings indicate that EC plasma membranes differently respond to stretch and shear stress by changing their lipid order, fluidity, and cholesterol content in opposite directions and that these changes in membrane physical properties are involved in the mechanotransduction that activates membrane receptors specific to each force.

  2. Cell pattern in adult human corneal endothelium.

    Directory of Open Access Journals (Sweden)

    Carlos H Wörner

    Full Text Available A review of the current data on the cell density of normal adult human endothelial cells was carried out in order to establish some common parameters appearing in the different considered populations. From the analysis of cell growth patterns, it is inferred that the cell aging rate is similar for each of the different considered populations. Also, the morphology, the cell distribution and the tendency to hexagonallity are studied. The results are consistent with the hypothesis that this phenomenon is analogous with cell behavior in other structures such as dry foams and grains in polycrystalline materials. Therefore, its driving force may be controlled by the surface tension and the mobility of the boundaries.

  3. Merkel cell distribution in the human eyelid

    Directory of Open Access Journals (Sweden)

    C.A. May

    2013-10-01

    Full Text Available Although Merkel cell carcinoma of the eye lid is reported frequently in the literature, only limited information exists about the distribution of Merkel cells in this tissue. Therefore, serial sections of 18 human cadaver eye lids (donors ages ranging between 63 and 97 years were stained for cytokeratin 20 in various planes. The overall appearance of Merkel cells in these samples was low and mainly located in the outer root layer of the cilia hair follicles. Merkel cells were more frequent in the middle, and almost not detectable at the nasal and temporal edges. The localization is in accordance with that of Merkel cell carcinoma, but concerning the scarce appearance within this adulthood group, a specific physiological role of these cells in the eye lid is difficult to establish.

  4. Natural killer cells in human autoimmune disorders

    Science.gov (United States)

    2013-01-01

    Natural killer (NK) cells are innate lymphocytes that play a critical role in early host defense against viruses. Through their cytolytic capacity and generation of cytokines and chemokines, NK cells modulate the activity of other components of the innate and adaptive immune systems and have been implicated in the initiation or maintenance of autoimmune responses. This review focuses on recent research elucidating a potential immunoregulatory role for NK cells in T-cell and B-cell-mediated autoimmune disorders in humans, with a particular focus on multiple sclerosis, rheumatoid arthritis, and systemic lupus erythematous. A better understanding of the contributions of NK cells to the development of autoimmunity may lead to novel therapeutic targets in these diseases. PMID:23856014

  5. Human Colon Cancer Cells Cultivated in Space

    Science.gov (United States)

    1995-01-01

    Within five days, bioreactor cultivated human colon cancer cells (shown) grown in Microgravity on the STS-70 mission in 1995, had grown 30 times the volume of the control specimens on Earth. The samples grown in space had a higher level of cellular organization and specialization. Because they more closely resemble tumors found in the body, microgravity grown cell cultures are ideal for research purposes.

  6. Quercetin Inhibits Cell Migration and Invasion in Human Osteosarcoma Cells

    Directory of Open Access Journals (Sweden)

    Haifeng Lan

    2017-09-01

    Full Text Available Background/Aims: Osteosarcoma is a malignant tumor associated with high mortality; however, no effective therapies for the disease have been developed. Several studies have focused on elucidating the pathogenesis of osteosarcoma and have aimed to develop novel therapies for the disease. Quercetin is a vital dietary flavonoid that has been shown to have a variety of anticancer effects, as it induces cell cycle arrest, apoptosis, and differentiation and is involved in cell adhesion, metastasis and angiogenesis. Herein, we aimed to investigate the effects of quercetin on osteosarcoma migration and invasion in vitro and in vivo and to explore the molecular mechanisms underlying its effects on osteosarcoma migration and invasion. Methods: Cell viability, cell cycle activity and cell apoptosis were measured using CCK-8 assay and flow cytometry, and cell migration and invasion were evaluated by wound healing and transwell assays, respectively. The mRNA and protein expression levels of several proteins of interest were assessed by real-time quantitative PCR and western blotting, respectively. Moreover, a nude mouse model of human osteosarcoma lung metastasis was established to assess the anti-metastatic effects of quercetin in vivo. Results: We noted no significant differences in cell cycle activity and apoptosis between HOS and MG63 cells and control cells. Treatment with quercetin significantly attenuated cell migration and invasion in HOS and MG63 cells compared with treatment with control medium. Moreover HIF-1α, VEGF, MMP2, and MMP9 mRNA and protein expression levels were significantly downregulated in HOS cells treated with quercetin compared with HOS cells treated with controls. Additionally, treatment with quercetin attenuated metastatic lung tumor formation and growth in the nude mouse model of osteosarcoma compared with treatment with controls. Conclusion: Our findings regarding the inhibitory effects of quercetin on cell migration and

  7. Human Mammary Luminal Epithelial Cells Contain Progenitors to Myoepithelial Cells

    Energy Technology Data Exchange (ETDEWEB)

    Pechoux, Christine; Gudjonsson, Thorarinn; Ronnov-Jessen, Lone; Bissell, Mina J; Petersen, Ole

    1999-02-01

    The origin of the epithelial and myoepithelial cells in the human breast has not been delineated. In this study we have addressed whether luminal epithelial cells and myoepithelial cells are vertically connected, i.e., whether one is the precursor for the other. We used a primary culture assay allowing preservation of basic phenotypic traits of luminal epithelial and myoepithelial cells in culture. The two cell types were then separated immunomagnetically using antibodies directed against lineage-specific cell surface antigens into at best 100% purity. The cellular identity was ascertained by cytochemistry, immunoblotting, and 2-D gel electrophoresis. Luminal epithelial cells were identified by strong expression of cytokeratins 18 and 19 while myoepithelial cells were recognized by expression of vimentin and {alpha}-smooth muscle actin. We used a previously devised culture medium (CDM4) that allows vigorous expansion of proliferative myoepithelial cells and also devised a medium (CDM6) that allowed sufficient expansion of differentiated luminal epithelial cells based on addition of hepatocyte growth factor/scatter factor. The two different culture media supported each lineage for at least five passages without signs of interconversion. We used parallel cultures where we switched culture media, thus testing the ability of each lineage to convert to the other. Whereas the myoepithelial lineage showed no signs of interconversion, a subset of luminal epithelial cells, gradually, but distinctly, converted to myoepithelial cells. We propose that in the mature human breast, it is the luminal epithelial cell compartment that gives rise to myoepithelial cells rather than the other way around.

  8. Genetic Manipulation of Human Embryonic Stem Cells.

    Science.gov (United States)

    Eiges, Rachel

    2016-01-01

    One of the great advantages of embryonic stem (ES) cells over other cell types is their accessibility to genetic manipulation. They can easily undergo genetic modifications while remaining pluripotent, and can be selectively propagated, allowing the clonal expansion of genetically altered cells in culture. Since the first isolation of ES cells in mice, many effective techniques have been developed for gene delivery and manipulation of ES cells. These include transfection, electroporation, and infection protocols, as well as different approaches for inserting, deleting, or changing the expression of genes. These methods proved to be extremely useful in mouse ES cells, for monitoring and directing differentiation, discovering unknown genes, and studying their function, and are now being extensively implemented in human ES cells (HESCs). This chapter describes the different approaches and methodologies that have been applied for the genetic manipulation of HESCs and their applications. Detailed protocols for generating clones of genetically modified HESCs by transfection, electroporation, and infection will be described, with special emphasis on the important technical details that are required for this purpose. All protocols are equally effective in human-induced pluripotent stem (iPS) cells.

  9. Notch-1 induces epithelial-mesenchymal transition consistent with cancer stem cell phenotype in pancreatic cancer cells.

    Science.gov (United States)

    Bao, Bin; Wang, Zhiwei; Ali, Shadan; Kong, Dejuan; Li, Yiwei; Ahmad, Aamir; Banerjee, Sanjeev; Azmi, Asfar S; Miele, Lucio; Sarkar, Fazlul H

    2011-08-01

    Activation of Notch-1 is known to be associated with the development and progression of human malignancies including pancreatic cancer. Emerging evidence suggest that the acquisition of epithelial-mesenchymal transition (EMT) phenotype and induction of cancer stem cell (CSC) or cancer stem-like cell phenotype are interrelated and contributes to tumor recurrence and drug resistance. The molecular mechanism(s) by which Notch-1 contributes to the acquisition of EMT phenotype and CSC self-renewal capacity has not been fully elucidated. Here we show that forced over-expression of Notch-1 leads to increased cell growth, clonogenicity, migration and invasion of AsPC-1 cells. Moreover, over-expression of Notch-1 led to the induction of EMT phenotype by activation of mesenchymal cell markers such as ZEB1, CD44, EpCAM, and Hes-1. Here we also report, for the first time, that over-expression of Notch-1 leads to increased expression of miR-21, and decreased expression of miR-200b, miR-200c, let-7a, let-7b, and let-7c. Re-expression of miR-200b led to decreased expression of ZEB1, and vimentin, and increased expression of E-cadherin. Over-expression of Notch-1 also increased the formation of pancreatospheres consistent with expression of CSC surface markers CD44 and EpCAM. Finally, we found that genistein, a known natural anti-tumor agent inhibited cell growth, clonogenicity, migration, invasion, EMT phenotype, formation of pancreatospheres and expression of CD44 and EpCAM. These results suggest that the activation of Notch-1 signaling contributes to the acquisition of EMT phenotype, which is in part mediated through the regulation of miR-200b and CSC self-renewal capacity, and these processes could be attenuated by genistein treatment.

  10. Polypyridyl transition metal complexes with application in water oxidation catalysis and dye-sensitised solar cells

    OpenAIRE

    Rudd, Jennifer A.

    2012-01-01

    This thesis contains complementary synthetic and computational studies of transition metal complexes with polypyridyl ligands for use either as water oxidation catalysts or for application in dye-sensitised solar cells (DSSCs). Chapter 1 introduces the reasons for researching water splitting catalysts and describes a number of current techniques used to do so; from photoelectrochemical cells to the use of transition metal polypyridyl complexes. It also introduces three commercially avail...

  11. Epithelial-mesenchymal transition (EMT): A biological process in the development, stem cell differentiation, and tumorigenesis.

    Science.gov (United States)

    Chen, Tong; You, Yanan; Jiang, Hua; Wang, Zack Z

    2017-12-01

    The lineage transition between epithelium and mesenchyme is a process known as epithelial-mesenchymal transition (EMT), by which polarized epithelial cells lose their adhesion property and obtain mesenchymal cell phenotypes. EMT is a biological process that is often involved in embryogenesis and diseases, such as cancer invasion and metastasis. The EMT and the reverse process, mesenchymal-epithelial transition (MET), also play important roles in stem cell differentiation and de-differentiation (or reprogramming). In this review, we will discuss current research progress of EMT in embryonic development, cellular differentiation and reprogramming, and cancer progression, all of which are representative models for researches of stem cell biology in normal and in diseases. Understanding of EMT and MET may help to identify specific markers to distinguish normal stem cells from cancer stem cells in future. © 2017 Wiley Periodicals, Inc.

  12. FHOD1, a formin upregulated in epithelial-mesenchymal transition, participates in cancer cell migration and invasion.

    Directory of Open Access Journals (Sweden)

    Maria Gardberg

    Full Text Available Cancer cells can obtain their ability to invade and metastasise by undergoing epithelial-to-mesenchymal transition (EMT. Exploiting this mechanism of cellular plasticity, malignant cells can remodel their actin cytoskeleton and down-regulate proteins needed for cell-cell contacts. The mechanisms of cytoskeletal reorganisation resulting in mesenchymal morphology and increased invasive potential are poorly understood. Actin nucleating formins have been implicated as key players in EMT. Here, we analysed which formins are altered in squamous cell carcinoma related EMT. FHOD1, a poorly studied formin, appeared to be markedly upregulated upon EMT. In human tissues FHOD1 was primarily expressed in mesenchymal cells, with little expression in epithelia. However, specimens from oral squamous cell cancers demonstrated consistent FHOD1 upregulation in mesenchymally transformed cells at the invasive edge. This upregulation was confirmed in an oral squamous carcinoma model, where FHOD1 expression was markedly increased upon EMT in a PI3K signalling dependent manner. In the EMT cells FHOD1 contributed to the spindle-shaped morphology and mesenchymal F-actin organization. Furthermore, functional assays demonstrated that FHOD1 contributes to cell migration and invasion. Finally, FHOD1 depletion reduced the ability of EMT cancer cells to form invadopodia and to degrade extracellular matrix. Our results indicate that FHOD1 participates in cytoskeletal changes in EMT. In addition, we show that FHOD1 upregulation occurs during cancer cell EMT in vivo, which indicates that FHOD1 may contribute to tumour progression.

  13. FHOD1, a Formin Upregulated in Epithelial-Mesenchymal Transition, Participates in Cancer Cell Migration and Invasion

    Science.gov (United States)

    Gardberg, Maria; Kaipio, Katja; Lehtinen, Laura; Mikkonen, Piia; Heuser, Vanina D.; Talvinen, Kati; Iljin, Kristiina; Kampf, Caroline; Uhlen, Mathias; Grénman, Reidar; Koivisto, Mari; Carpén, Olli

    2013-01-01

    Cancer cells can obtain their ability to invade and metastasise by undergoing epithelial-to-mesenchymal transition (EMT). Exploiting this mechanism of cellular plasticity, malignant cells can remodel their actin cytoskeleton and down-regulate proteins needed for cell-cell contacts. The mechanisms of cytoskeletal reorganisation resulting in mesenchymal morphology and increased invasive potential are poorly understood. Actin nucleating formins have been implicated as key players in EMT. Here, we analysed which formins are altered in squamous cell carcinoma related EMT. FHOD1, a poorly studied formin, appeared to be markedly upregulated upon EMT. In human tissues FHOD1 was primarily expressed in mesenchymal cells, with little expression in epithelia. However, specimens from oral squamous cell cancers demonstrated consistent FHOD1 upregulation in mesenchymally transformed cells at the invasive edge. This upregulation was confirmed in an oral squamous carcinoma model, where FHOD1 expression was markedly increased upon EMT in a PI3K signalling dependent manner. In the EMT cells FHOD1 contributed to the spindle-shaped morphology and mesenchymal F-actin organization. Furthermore, functional assays demonstrated that FHOD1 contributes to cell migration and invasion. Finally, FHOD1 depletion reduced the ability of EMT cancer cells to form invadopodia and to degrade extracellular matrix. Our results indicate that FHOD1 participates in cytoskeletal changes in EMT. In addition, we show that FHOD1 upregulation occurs during cancer cell EMT in vivo, which indicates that FHOD1 may contribute to tumour progression. PMID:24086398

  14. T. brucei infection reduces B lymphopoiesis in bone marrow and truncates compensatory splenic lymphopoiesis through transitional B-cell apoptosis.

    Directory of Open Access Journals (Sweden)

    Viki Bockstal

    2011-06-01

    Full Text Available African trypanosomes of the Trypanosoma brucei species are extracellular protozoan parasites that cause the deadly disease African trypanosomiasis in humans and contribute to the animal counterpart, Nagana. Trypanosome clearance from the bloodstream is mediated by antibodies specific for their Variant Surface Glycoprotein (VSG coat antigens. However, T. brucei infection induces polyclonal B cell activation, B cell clonal exhaustion, sustained depletion of mature splenic Marginal Zone B (MZB and Follicular B (FoB cells, and destruction of the B-cell memory compartment. To determine how trypanosome infection compromises the humoral immune defense system we used a C57BL/6 T. brucei AnTat 1.1 mouse model and multicolor flow cytometry to document B cell development and maturation during infection. Our results show a more than 95% reduction in B cell precursor numbers from the CLP, pre-pro-B, pro-B, pre-B and immature B cell stages in the bone marrow. In the spleen, T. brucei induces extramedullary B lymphopoiesis as evidenced by significant increases in HSC-LMPP, CLP, pre-pro-B, pro-B and pre-B cell populations. However, final B cell maturation is abrogated by infection-induced apoptosis of transitional B cells of both the T1 and T2 populations which is not uniquely dependent on TNF-, Fas-, or prostaglandin-dependent death pathways. Results obtained from ex vivo co-cultures of living bloodstream form trypanosomes and splenocytes demonstrate that trypanosome surface coat-dependent contact with T1/2 B cells triggers their deletion. We conclude that infection-induced and possibly parasite-contact dependent deletion of transitional B cells prevents replenishment of mature B cell compartments during infection thus contributing to a loss of the host's capacity to sustain antibody responses against recurring parasitemic waves.

  15. Enriched retinal ganglion cells derived from human embryonic stem cells

    Science.gov (United States)

    Gill, Katherine P.; Hung, Sandy S. C.; Sharov, Alexei; Lo, Camden Y.; Needham, Karina; Lidgerwood, Grace E.; Jackson, Stacey; Crombie, Duncan E.; Nayagam, Bryony A.; Cook, Anthony L.; Hewitt, Alex W.; Pébay, Alice; Wong, Raymond C. B.

    2016-01-01

    Optic neuropathies are characterised by a loss of retinal ganglion cells (RGCs) that lead to vision impairment. Development of cell therapy requires a better understanding of the signals that direct stem cells into RGCs. Human embryonic stem cells (hESCs) represent an unlimited cellular source for generation of human RGCs in vitro. In this study, we present a 45-day protocol that utilises magnetic activated cell sorting to generate enriched population of RGCs via stepwise retinal differentiation using hESCs. We performed an extensive characterization of these stem cell-derived RGCs by examining the gene and protein expressions of a panel of neural/RGC markers. Furthermore, whole transcriptome analysis demonstrated similarity of the hESC-derived RGCs to human adult RGCs. The enriched hESC-RGCs possess long axons, functional electrophysiological profiles and axonal transport of mitochondria, suggestive of maturity. In summary, this RGC differentiation protocol can generate an enriched population of functional RGCs from hESCs, allowing future studies on disease modeling of optic neuropathies and development of cell therapies. PMID:27506453

  16. Human ES cells: starting culture from frozen cells.

    Science.gov (United States)

    Trish, Erin; Dimos, John; Eggan, Kevin

    2006-11-09

    Here we demonstrate how our lab begins a HuES human embryonic stem cell line culture from a frozen stock. First, a one to two day old ten cm plate of approximately one (to two) million irradiated mouse embryonic fibroblast feeder cells is rinsed with HuES media to remove residual serum and cell debris, and then HuES media added and left to equilibrate in the cell culture incubator. A frozen vial of cells from long term liquid nitrogen storage or a -80 C freezer is sourced and quickly submerged in a 37 C water bath for quick thawing. Cells in freezing media are then removed from the vial and placed in a large volume of HuES media. The large volume of HuES media facilitates removal of excess serum and DMSO, which can cause HuES human embryonic stem cells to differentiate. Cells are gently spun out of suspension, and then re-suspended in a small volume of fresh HuES media that is then used to seed the MEF plate. It is considered important to seed the MEF plate by gently adding the HuES cells in a drop wise fashion to evenly disperse them throughout the plate. The newly established HuES culture plate is returned to the incubator for 48 hrs before media is replaced, then is fed every 24 hours thereafter.

  17. Sodium Valproate Induces Cell Senescence in Human Hepatocarcinoma Cells

    Directory of Open Access Journals (Sweden)

    Hong-Mei An

    2013-12-01

    Full Text Available Hepatocarcinogenesis is associated with epigenetic changes, including histone deacetylases (HDACs. Epigenetic modulation by HDAC inhibition is a potentially valuable approach for hepatocellular carcinoma treatment. In present study, we evaluated the anticancer effects of sodium valproate (SVP, a known HDAC inhibitor, in human hepatocarcinoma cells. The results showed SVP inhibited the proliferation of Bel-7402 cells in a dose-dependent manner. Low dose SVP treatment caused a large and flat morphology change, positive SA-β-gal staining, and G0/G1 phase cell cycle arrest in human hepatocarcinoma cells. Low dose SVP treatment also increased acetylation of histone H3 and H4 on p21 promoter, accompanied by up-regulation of p21 and down-regulation of RB phosphorylation. These observations suggested that a low dose of SVP could induce cell senescence in hepatocarcinoma cells, which might correlate with hyperacetylation of histone H3 and H4, up-regulation of p21, and inhibition of RB phosphorylation. Since the effective concentration inducing cell senescence in hepatocarcinoma cells is clinically available, whether a clinical dose of SVP could induce cell senescence in clinical hepatocarcinoma is worthy of further study.

  18. Human embryonic stem cell lines model experimental human cytomegalovirus latency.

    Science.gov (United States)

    Penkert, Rhiannon R; Kalejta, Robert F

    2013-05-28

    Herpesviruses are highly successful pathogens that persist for the lifetime of their hosts primarily because of their ability to establish and maintain latent infections from which the virus is capable of productively reactivating. Human cytomegalovirus (HCMV), a betaherpesvirus, establishes latency in CD34(+) hematopoietic progenitor cells during natural infections in the body. Experimental infection of CD34(+) cells ex vivo has demonstrated that expression of the viral gene products that drive productive infection is silenced by an intrinsic immune defense mediated by Daxx and histone deacetylases through heterochromatinization of the viral genome during the establishment of latency. Additional mechanistic details about the establishment, let alone maintenance and reactivation, of HCMV latency remain scarce. This is partly due to the technical challenges of CD34(+) cell culture, most notably, the difficulty in preventing spontaneous differentiation that drives reactivation and renders them permissive for productive infection. Here we demonstrate that HCMV can establish, maintain, and reactivate in vitro from experimental latency in cultures of human embryonic stem cells (ESCs), for which spurious differentiation can be prevented or controlled. Furthermore, we show that known molecular aspects of HCMV latency are faithfully recapitulated in these cells. In total, we present ESCs as a novel, tractable model for studies of HCMV latency.

  19. Salinomycin as a Drug for Targeting Human Cancer Stem Cells

    Directory of Open Access Journals (Sweden)

    Cord Naujokat

    2012-01-01

    Full Text Available Cancer stem cells (CSCs represent a subpopulation of tumor cells that possess self-renewal and tumor initiation capacity and the ability to give rise to the heterogenous lineages of malignant cells that comprise a tumor. CSCs possess multiple intrinsic mechanisms of resistance to chemotherapeutic drugs, novel tumor-targeted drugs, and radiation therapy, allowing them to survive standard cancer therapies and to initiate tumor recurrence and metastasis. Various molecular complexes and pathways that confer resistance and survival of CSCs, including expression of ATP-binding cassette (ABC drug transporters, activation of the Wnt/β-catenin, Hedgehog, Notch and PI3K/Akt/mTOR signaling pathways, and acquisition of epithelial-mesenchymal transition (EMT, have been identified recently. Salinomycin, a polyether ionophore antibiotic isolated from Streptomyces albus, has been shown to kill CSCs in different types of human cancers, most likely by interfering with ABC drug transporters, the Wnt/β-catenin signaling pathway, and other CSC pathways. Promising results from preclinical trials in human xenograft mice and a few clinical pilote studies reveal that salinomycin is able to effectively eliminate CSCs and to induce partial clinical regression of heavily pretreated and therapy-resistant cancers. The ability of salinomycin to kill both CSCs and therapy-resistant cancer cells may define the compound as a novel and an effective anticancer drug.

  20. Fuel Cell Buses in U.S. Transit Fleets: Current Status 2015

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, Leslie [National Renewable Energy Lab. (NREL), Golden, CO (United States); Post, Matthew [National Renewable Energy Lab. (NREL), Golden, CO (United States); Gikakis, Christina [Federal Transit Administration, Washington, DC (United States)

    2015-12-11

    This report, published annually, summarizes the progress of fuel cell electric bus (FCEB) development in the United States and discusses the achievements and challenges of introducing fuel cell propulsion in transit. Various stakeholders, including FCEB developers, transit agencies, and system integrators, have expressed the value of this annual status report, which provides a summary of results from evaluations performed by the National Renewable Energy Laboratory. The annual status report tracks the progress of the FCEB industry toward meeting technical targets, documents the lessons learned, and discusses the path forward for commercial viability of fuel cell technology for transit buses. The 2015 summary results primarily focus on the most recent year for each demonstration, from August 2014 through July 2015. The results for these buses account for more than 1,045,000 miles traveled and 83,000 hours of fuel cell power system operation. The primary results presented in the report are from two demonstrations of fuel-cell-dominant bus designs: the Zero Emission Bay Area Demonstration Group led by Alameda-Contra Costa Transit District (AC Transit) in California and the American Fuel Cell Bus Project at SunLine Transit Agency in California.

  1. [Immune system evolution. (From cells to humans)].

    Science.gov (United States)

    Belek, A S

    1992-01-01

    The great variety of cells and molecules observed in the mammalian immune system can be explained by stepwise acquisition of them during phylogeny. Self/nonself discrimination and cell-mediated immunity have been present since the early stages of evolution. Although some inducible antimicrobial molecules have been demonstrated in invertebrates, immunoglobulins appear in vertebrates. T and B cell diversity, development of the lymphoid organs, MHC molecules, complement and cytokines are the characteristics that appear through the evolution of vertebrates. Further knowledge that will be obtained from phylogenetic studies will improve our understanding of the immune system of human.

  2. Centre for human development, stem cells & regeneration.

    Science.gov (United States)

    Oreffo, Richard O C

    2014-01-01

    The Centre for Human Development, Stem Cells and Regeneration (CHDSCR) was founded in 2004 as a cross-disciplinary research and translational program within the Faculty of Medicine at the University of Southampton. The Centre undertakes fundamental research into early development and stem cells together with applied translational research for patient benefit. The Centre has vibrant and thriving multidisciplinary research programs that harness the translational strength of the Faculty together with an innovative Stem Cell PhD program, outstanding clinical infrastructure and enterprise to deliver on this vision.

  3. Biobanking human embryonic stem cell lines

    DEFF Research Database (Denmark)

    Holm, Søren

    2016-01-01

    Stem cell banks curating and distributing human embryonic stem cells have been established in a number of countries and by a number of private institutions. This paper identifies and critically discusses a number of arguments that are used to justify the importance of such banks in policy...... are curiously absent from the particular stem cell banking policy discourse. This to some extent artificially isolates this discourse from the broader discussions about the flows of reproductive materials and tissues in modern society, and such isolation may lead to the interests of important actors being...

  4. Advances in human B cell phenotypic profiling

    Directory of Open Access Journals (Sweden)

    Denise A Kaminski

    2012-10-01

    Full Text Available To advance our understanding and treatment of disease, research immunologists have been called-upon to place more centralized emphasis on impactful human studies. Such endeavors will inevitably require large-scale study execution and data management regulation (Big Biology, necessitating standardized and reliable metrics of immune status and function. A well-known example setting this large-scale effort in-motion is identifying correlations between eventual disease outcome and T lymphocyte phenotype in large HIV-patient cohorts using multiparameter flow cytometry. However, infection, immunodeficiency, and autoimmunity are also characterized by correlative and functional contributions of B lymphocytes, which to-date have received much less attention in the human Big Biology enterprise. Here, we review progress in human B cell phenotyping, analysis, and bioinformatics tools that constitute valuable resources for the B cell research community to effectively join in this effort.

  5. Human plasma cells express granzyme B.

    Science.gov (United States)

    Xu, Wei; Narayanan, Priya; Kang, Ning; Clayton, Sandra; Ohne, Yoichiro; Shi, Peiqing; Herve, Marie-Cecile; Balderas, Robert; Picard, Capucine; Casanova, Jean-Laurent; Gorvel, Jean-Pierre; Oh, Sangkon; Pascual, Virginia; Banchereau, Jacques

    2014-01-01

    While studying the plasma cell (PC) compartment in human tonsils, we identified that immunoglobulin kappa or lambda chain-expressing PCs are the main cells expressing granzyme B (GrzB). In vitro studies revealed that activated B cells differentiated into GrzB-expressing PCs when co-cultured with macrophages and follicular helper T cells. This effect could be reproduced on combined stimulation of IL-15 (produced by macrophages) and IL-21 (produced by T follicular helper cells) in a STAT3-dependent manner. Whereas IL-21 triggers the transcription of mRNA of GrzB, IL-15 synergizes the translation of GrzB proteins. The precise role of GrzB in PC biology remains to be understood and studies in mice will not help as their PCs do not express GrzB.

  6. Human CD56bright NK Cells

    DEFF Research Database (Denmark)

    Michel, Tatiana; Poli, Aurélie; Cuapio, Angelica

    2016-01-01

    Human NK cells can be subdivided into various subsets based on the relative expression of CD16 and CD56. In particular, CD56(bright)CD16(-/dim) NK cells are the focus of interest. They are considered efficient cytokine producers endowed with immunoregulatory properties, but they can also become...... cytotoxic upon appropriate activation. These cells were shown to play a role in different disease states, such as cancer, autoimmunity, neuroinflammation, and infection. Although their phenotype and functional properties are well known and have been extensively studied, their lineage relationship with other...... NK cell subsets is not fully defined, nor is their precise hematopoietic origin. In this article, we summarize recent studies about CD56(bright) NK cells in health and disease and briefly discuss the current controversies surrounding them....

  7. Twist基因逆转录病毒载体的构建及其诱导正常乳腺上皮细胞发生上皮间质转化%Construction of retroviral vector carrying Twist gene and its induction of epithelialmesenchymal transition in human mammary epithelial cells

    Institute of Scientific and Technical Information of China (English)

    杨佳佳; 胡萍; 周明莉; 黄杰涛; 柳满然

    2013-01-01

    Objective To construct a retroviral vector carrying Twist gene and investigate its effect on human mammary MCF10A epithelial cells.Methods Myc-Twist was digested from pcDNA3/myc-Twist and subcloned into the retroviral vector pBABE-puro to construct a recombinant plasmid (pBABE-myc-Twist).The inserted Twist gene was confirmed by restriction enzyme digestion and DNA sequencing.The plasmid pBABE-myc-Twist and the packaging plasmid pAmpho were co-transfected into HEK293T cells for packaging of retrovirus.Meanwhile,the control plasmid pBABE-puro and the packaging plasmid were co-transfected into the other HEK293T cells as a control group.Human mammary MCF10A epithelial cells were infected with the retroviruses carrying Twist gene or the controls,and selected by puromycin.The expression of Twist in the MCF10A-Twist and MCF10A-Vector cells was determined by RT-PCR and Westem blotting.The expressions of epithelial-mesenchymal transition (EMT) marker proteins induced by Twist in MCF10A cells were detected using immunofluorescence cytochemistry and Westem blotting.Cell migration and invasion abilities were analyzed by Transwell(R) assay.Results The myc-tagged Twist gene was correctly inserted into the retroviral expression vector as a recombinant plasmid (pBABE-myc-Twist) as identified by restriction analysis and DNA sequencing.The Twist gene was efficiently delivered into human mammary MCF10A epithelial cells by the retrovirus,resulting in the stable expression of Twist mRNA and myc-tagged Twist protein as shown by RT-PCR and Western blotting,respectively.The expression of the epithelial biomarker E-cadherin was downregulated whereas,the mesenchymal marker vimentin upregulated in MCF10A-Twist cells as shown by immunofluorescence cytochemistry and Western blotting.Cell migration and invasion abilities were enhanced notably in MCF10A-Twist cells as compared with MCF10A-Vector control cells (P <0.01).Conclusion Twist induces EMT of MCF10A cells and plays an important role in

  8. Alterations in voltage-sensing of the mitochondrial permeability transition pore in ANT1-deficient cells.

    Science.gov (United States)

    Doczi, Judit; Torocsik, Beata; Echaniz-Laguna, Andoni; Mousson de Camaret, Bénédicte; Starkov, Anatoly; Starkova, Natalia; Gál, Aniko; Molnár, Mária J; Kawamata, Hibiki; Manfredi, Giovanni; Adam-Vizi, Vera; Chinopoulos, Christos

    2016-05-25

    The probability of mitochondrial permeability transition (mPT) pore opening is inversely related to the magnitude of the proton electrochemical gradient. The module conferring sensitivity of the pore to this gradient has not been identified. We investigated mPT's voltage-sensing properties elicited by calcimycin or H2O2 in human fibroblasts exhibiting partial or complete lack of ANT1 and in C2C12 myotubes with knocked-down ANT1 expression. mPT onset was assessed by measuring in situ mitochondrial volume using the 'thinness ratio' and the 'cobalt-calcein' technique. De-energization hastened calcimycin-induced swelling in control and partially-expressing ANT1 fibroblasts, but not in cells lacking ANT1, despite greater losses of mitochondrial membrane potential. Matrix Ca(2+) levels measured by X-rhod-1 or mitochondrially-targeted ratiometric biosensor 4mtD3cpv, or ADP-ATP exchange rates did not differ among cell types. ANT1-null fibroblasts were also resistant to H2O2-induced mitochondrial swelling. Permeabilized C2C12 myotubes with knocked-down ANT1 exhibited higher calcium uptake capacity and voltage-thresholds of mPT opening inferred from cytochrome c release, but intact cells showed no differences in calcimycin-induced onset of mPT, irrespective of energization and ANT1 expression, albeit the number of cells undergoing mPT increased less significantly upon chemically-induced hypoxia than control cells. We conclude that ANT1 confers sensitivity of the pore to the electrochemical gradient.

  9. The transition from stem cell to progenitor spermatogonia and male fertility requires the SHP2 protein tyrosine phosphatase.

    Science.gov (United States)

    Puri, Pawan; Phillips, Bart T; Suzuki, Hitomi; Orwig, Kyle E; Rajkovic, Aleksandar; Lapinski, Philip E; King, Philip D; Feng, Gen-Sheng; Walker, William H

    2014-03-01

    SHP2 is a widely expressed protein tyrosine phosphatase required for signal transduction from multiple cell surface receptors. Gain and loss of function SHP2 mutations in humans are known to cause Noonan and LEOPARD syndromes, respectively, that are characterized by numerous pathological conditions including male infertility. Using conditional gene targeting in the mouse, we found that SHP2 is required for maintaining spermatogonial stem cells (SSCs) and the production of germ cells required for male fertility. After deleting SHP2, spermatogenesis was halted at the initial step during which transit-amplifying undifferentiated spermatogonia are produced from SSCs. In the absence of SHP2, proliferation of SSCs and undifferentiated spermatogonia was inhibited, thus germ cells cannot be replenished and SSCs cannot undergo renewal. However, germ cells beyond the undifferentiated spermatogonia stage of development at the time of SHP2 knockout were able to complete their maturation to become sperm. In cultures of SSCs and their progeny, inhibition of SHP2 activity reduced growth factor-mediated intracellular signaling that regulates SSC proliferation and cell fate. Inhibition of SHP2 also decreased the number of SSCs present in culture and caused SSCs to detach from supporting cells. Injection of mice with an SHP2 inhibitor blocked the production of germ cells from SSCs. Together, our studies show that SHP2 is essential for SSCs to maintain fertility and indicates that the pathogenesis of infertility in humans with SHP2 mutations is due to compromised SSC functions that block spermatogenesis. © AlphaMed Press.

  10. Intra-urban human mobility and activity transition: evidence from social media check-in data.

    Science.gov (United States)

    Wu, Lun; Zhi, Ye; Sui, Zhengwei; Liu, Yu

    2014-01-01

    Most existing human mobility literature focuses on exterior characteristics of movements but neglects activities, the driving force that underlies human movements. In this research, we combine activity-based analysis with a movement-based approach to model the intra-urban human mobility observed from about 15 million check-in records during a yearlong period in Shanghai, China. The proposed model is activity-based and includes two parts: the transition of travel demands during a specific time period and the movement between locations. For the first part, we find the transition probability between activities varies over time, and then we construct a temporal transition probability matrix to represent the transition probability of travel demands during a time interval. For the second part, we suggest that the travel demands can be divided into two classes, locationally mandatory activity (LMA) and locationally stochastic activity (LSA), according to whether the demand is associated with fixed location or not. By judging the combination of predecessor activity type and successor activity type we determine three trip patterns, each associated with a different decay parameter. To validate the model, we adopt the mechanism of an agent-based model and compare the simulated results with the observed pattern from the displacement distance distribution, the spatio-temporal distribution of activities, and the temporal distribution of travel demand transitions. The results show that the simulated patterns fit the observed data well, indicating that these findings open new directions for combining activity-based analysis with a movement-based approach using social media check-in data.

  11. Intra-urban human mobility and activity transition: evidence from social media check-in data.

    Directory of Open Access Journals (Sweden)

    Lun Wu

    Full Text Available Most existing human mobility literature focuses on exterior characteristics of movements but neglects activities, the driving force that underlies human movements. In this research, we combine activity-based analysis with a movement-based approach to model the intra-urban human mobility observed from about 15 million check-in records during a yearlong period in Shanghai, China. The proposed model is activity-based and includes two parts: the transition of travel demands during a specific time period and the movement between locations. For the first part, we find the transition probability between activities varies over time, and then we construct a temporal transition probability matrix to represent the transition probability of travel demands during a time interval. For the second part, we suggest that the travel demands can be divided into two classes, locationally mandatory activity (LMA and locationally stochastic activity (LSA, according to whether the demand is associated with fixed location or not. By judging the combination of predecessor activity type and successor activity type we determine three trip patterns, each associated with a different decay parameter. To validate the model, we adopt the mechanism of an agent-based model and compare the simulated results with the observed pattern from the displacement distance distribution, the spatio-temporal distribution of activities, and the temporal distribution of travel demand transitions. The results show that the simulated patterns fit the observed data well, indicating that these findings open new directions for combining activity-based analysis with a movement-based approach using social media check-in data.

  12. Knockdown of Collagen Triple Helix Repeat Containing-1 Inhibits the Proliferation and Epithelial-to-Mesenchymal Transition in Renal Cell Carcinoma Cells.

    Science.gov (United States)

    Jin, Xue-Fei; Li, Hai; Zong, Shi; Li, Hong-Yan

    2016-10-27

    Collagen triple helix repeat containing-1 (CTHRC1), a secreted glycoprotein, is frequently upregulated in human cancers. However, the functional role of CTHRC1 in renal cell carcinoma (RCC) remains unclear. Thus, the aim of this study was to explore the role of CTHRC1 in RCC. Our results demonstrated that CTHRC1 was upregulated in RCC tissues and cell lines. Knockdown of CTHRC1 significantly inhibits the proliferation in RCCs. Furthermore, knockdown of CTHRC1 significantly inhibited the epithelial-to-mesenchymal transition (EMT) process in RCCs, as well as suppressed RCC cell migration and invasion. Mechanistically, knockdown of CTHRC1 inhibited the expression of β-catenin, c-Myc, and cyclin D1 in RCC cells. In conclusion, the results of the present study indicated that CTHRC1 downregulation inhibited proliferation, migration, EMT, and β-catenin expression in RCC cells. Therefore, CTHRC1 may be a potential therapeutic target for the treatment of RCC.

  13. Fuel Cell Buses in U.S. Transit Fleets: Current Status 2016

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, Leslie [National Renewable Energy Lab. (NREL), Golden, CO (United States); Post, Matthew [National Renewable Energy Lab. (NREL), Golden, CO (United States); Jeffers, Matthew [National Renewable Energy Lab. (NREL), Golden, CO (United States)

    2016-11-01

    This report, published annually, summarizes the progress of fuel cell electric bus development in the United States and discusses the achievements and challenges of introducing fuel cell propulsion in transit. The report provides a summary of results from evaluations performed by the National Renewable Energy Laboratory. Funding for this effort is provided by the U.S. Department of Energy's Fuel Cell Technologies Office within the Office of Energy Efficiency and Renewable Energy and by the U.S. Department of Transportation's Federal Transit Administration. The 2016 summary results primarily focus on the most recent year for each demonstration, from August 2015 through July 2016. The results for these buses account for more than 550,000 miles traveled and 59,500 hours of fuel cell power system operation. The primary results presented in the report are from three demonstrations of two different fuel-cell-dominant bus designs: Zero Emission Bay Area Demonstration Group led by Alameda-Contra Costa Transit District (AC Transit) in California; American Fuel Cell Bus Project at SunLine Transit Agency in California; and American Fuel Cell Bus Project at the University of California at Irvine.

  14. Stem cell factor and c-Kit in human primordial germ cells and fetal ovaries

    DEFF Research Database (Denmark)

    Høyer, Poul Erik; Byskov, Anne Grete; Møllgård, Kjeld

    2005-01-01

    Prenatal ovary (human), Primordial germ cells, Folliculogenesis, c-Kit, Stem cell factor, immunohistochemistry......Prenatal ovary (human), Primordial germ cells, Folliculogenesis, c-Kit, Stem cell factor, immunohistochemistry...

  15. JNK对TGF-β1诱导的人肺上皮-间质转分化的调控作用%Influence of JNK Signaling Pathway in the Epithelial-mesenchymal Transition Process of Human Alveolar Epithelial Cells Induced by TGF-β1

    Institute of Scientific and Technical Information of China (English)

    邹勇; 曾玉兰

    2014-01-01

    目的:探讨c‐Jun氨基末端激酶(JNK)在转化生长因子‐β1(TGF‐β1)诱导的人肺上皮细胞A549转分化中的调控作用。方法将体外培养的人肺上皮细胞(A549)随机分成3组:正常对照组、TGF‐β1组(加入10 ng/mL TGF‐β1)及抑制剂组(加入10 ng/mL TGF‐β1和20μmol/L JNK的特异性抑制剂Sp600125),培养于3%的血清培养液中,光镜下观察3组细胞形态的变化,并通过RT‐PCT检测各组A549细胞的上皮标志物E‐钙黏蛋白(E‐cadherin ,E‐cad)及间充质标志物α‐平滑肌肌动蛋白(α‐smooth muscle actin ,α‐SMA)和胶原纤维Ⅰ(collagen fibersⅠ,ColⅠ)的表达的变化,West‐ern blot检测JNK磷酸化(p‐JNK)水平的变化。结果正常对照组体外培养的A549细胞光镜下为鹅卵石样紧密排列生长,有E‐cad表达及微量的α‐SMA、ColⅠ及p‐JNK表达。TGF‐β1组培养72 h后细胞基本长成梭形、纺锤形,E‐cad表达下调,α‐SMA、ColⅠ及p‐JNK表达上调。与 TGF‐β1组比较,抑制剂组培养72 h后细胞梭形有所逆转,E‐cad表达上调,α‐SM A、ColⅠ及p‐JNK表达明显抑制;与正常对照组比较,细胞形态较扁长,E‐cad、α‐SM A、ColⅠ及p‐JNK表达差异无统计学意义。结论 JNK信号通路参与 TGF‐β1介导的人肺上皮‐间质转分化过程,JNK的特异性抑制剂Sp600125可有效抑制该过程。%Objective To explore the role of JNK signaling pathway in epithelial mesenchymal transition (EMT)process of human alveolar epithelial cells A549 induced by TGF‐β1 in vitro.Methods Human alveolar epithelial cells (A549)cultured in vitro were divided randomly into three groups :normal group ,TGF‐β1 group ( treated by TGF‐β1 with 10 ng/mL)and inhibitor group (treated by 10 ng/mL TGF‐β1 and 20 μmol/L Sp600125).Morphological observation on the cells was performed under light microscope after

  16. Spontaneous quaternary and tertiary T-R transitions of human hemoglobin in molecular dynamics simulation.

    Directory of Open Access Journals (Sweden)

    Jochen S Hub

    2010-05-01

    Full Text Available We present molecular dynamics simulations of unliganded human hemoglobin (Hb A under physiological conditions, starting from the R, R2, and T state. The simulations were carried out with protonated and deprotonated HC3 histidines His(beta146, and they sum up to a total length of 5.6 micros. We observe spontaneous and reproducible T-->R quaternary transitions of the Hb tetramer and tertiary transitions of the alpha and beta subunits, as detected from principal component projections, from an RMSD measure, and from rigid body rotation analysis. The simulations reveal a marked asymmetry between the alpha and beta subunits. Using the mutual information as correlation measure, we find that the beta subunits are substantially more strongly linked to the quaternary transition than the alpha subunits. In addition, the tertiary populations of the alpha and beta subunits differ substantially, with the beta subunits showing a tendency towards R, and the alpha subunits showing a tendency towards T. Based on the simulation results, we present a transition pathway for coupled quaternary and tertiary transitions between the R and T conformations of Hb.

  17. Differences in the microrheology of human embryonic stem cells and human induced pluripotent stem cells.

    Science.gov (United States)

    Daniels, Brian R; Hale, Christopher M; Khatau, Shyam B; Kusuma, Sravanti; Dobrowsky, Terrence M; Gerecht, Sharon; Wirtz, Denis

    2010-12-01

    Embryonic and adult fibroblasts can be returned to pluripotency by the expression of reprogramming genes. Multiple lines of evidence suggest that these human induced pluripotent stem (hiPS) cells and human embryonic stem (hES) cells are behaviorally, karyotypically, and morphologically similar. Here we sought to determine whether the physical properties of hiPS cells, including their micromechanical properties, are different from those of hES cells. To this end, we use the method of particle tracking microrheology to compare the viscoelastic properties of the cytoplasm of hES cells, hiPS cells, and the terminally differentiated parental human fibroblasts from which our hiPS cells are derived. Our results indicate that although the cytoplasm of parental fibroblasts is both viscous and elastic, the cytoplasm of hiPS cells does not exhibit any measurable elasticity and is purely viscous over a wide range of timescales. The viscous phenotype of hiPS cells is recapitulated in parental cells with disassembled actin filament network. The cytoplasm of hES cells is predominantly viscous but contains subcellular regions that are also elastic. This study supports the hypothesis that intracellular elasticity correlates with the degree of cellular differentiation and reveals significant differences in the mechanical properties of hiPS cells and hES cells. Because mechanical stimuli have been shown to mediate the precise fate of differentiating stem cells, our results support the concept that stem cell "softness" is a key feature of force-mediated differentiation of stem cells and suggest there may be subtle functional differences between force-mediated differentiation of hiPS cells and hES cells.

  18. Relocalization of cell adhesion molecules during neoplastic transformation of human fibroblasts.

    Science.gov (United States)

    Belgiovine, Cristina; Chiodi, Ilaria; Mondello, Chiara

    2011-11-01

    Studying neoplastic transformation of telomerase immortalized human fibroblasts (cen3tel), we found that the transition from normal to tumorigenic cells was associated with the loss of growth contact inhibition, the acquisition of an epithelial-like morphology and a change in actin organization, from stress fibers to cortical bundles. We show here that these variations were paralleled by an increase in N-cadherin expression and relocalization of different adhesion molecules, such as N-cadherin, α-catenin, p-120 and β-catenin. These proteins presented a clear membrane localization in tumorigenic cells compared to a more diffuse, cytoplasmic distribution in primary fibroblasts and non-tumorigenic immortalized cells, suggesting that tumorigenic cells could form strong cell-cell contacts and cell contacts did not induce growth inhibition. The epithelial-like appearance of tumorigenic cells did not reflect a mesenchymal-epithelial transition; in fact, cen3tel cells expressed vimentin and did not express cytokeratins at all transformation stages. Moreover, they did not express epithelial proteins such as occluding and claudin-1. In contrast, ZO-1 showed higher levels and a more defined membrane localization in tumorigenic cells compared to non-tumorigenic cells; this confirms its role in adherens junction formation in mesenchymal cells and is in agreement with the strong cell-cell contact formation by neoplastically transformed cells. Finally, we found α-catenin and ZO-1 nuclear localization in non-transformed cells, suggestive of possible additional roles of these proteins besides cell junction formation.

  19. Generation of mature hematopoietic cells from human pluripotent stem cells.

    Science.gov (United States)

    Togarrati, Padma Priya; Suknuntha, Kran

    2012-06-01

    A number of malignant and non-malignant hematological disorders are associated with the abnormal production of mature blood cells or primitive hematopoietic precursors. Their capacity for continuous self-renewal without loss of pluripotency and the ability to differentiate into adult cell types from all three primitive germ layers make human embryonic stem cells and induced pluripotent stem cells (hiPSCs) attractive complementary cell sources for large-scale production of transfusable mature blood cell components in cell replacement therapies. The generation of patient-specific hematopoietic stem/precursor cells from iPSCs by the regulated manipulation of various factors involved in reprograming to ensure complete pluripotency, and developing innovative differentiation strategies for generating unlimited supply of clinically safe, transplantable, HLA-matched cells from hiPSCs to outnumber the inadequate source of hematopoietic stem cells obtained from cord blood, bone marrow and peripheral blood, would have a major impact on the field of regenerative and personalized medicine leading to translation of these results from bench to bedside.

  20. Cripto is essential to capture mouse epiblast stem cell and human embryonic stem cell pluripotency.

    Science.gov (United States)

    Fiorenzano, Alessandro; Pascale, Emilia; D'Aniello, Cristina; Acampora, Dario; Bassalert, Cecilia; Russo, Francesco; Andolfi, Gennaro; Biffoni, Mauro; Francescangeli, Federica; Zeuner, Ann; Angelini, Claudia; Chazaud, Claire; Patriarca, Eduardo J; Fico, Annalisa; Minchiotti, Gabriella

    2016-09-02

    Known molecular determinants of developmental plasticity are mainly transcription factors, while the extrinsic regulation of this process has been largely unexplored. Here we identify Cripto as one of the earliest epiblast markers and a key extracellular determinant of the naive and primed pluripotent states. We demonstrate that Cripto sustains mouse embryonic stem cell (ESC) self-renewal by modulating Wnt/β-catenin, whereas it maintains mouse epiblast stem cell (EpiSC) and human ESC pluripotency through Nodal/Smad2. Moreover, we provide unprecedented evidence that Cripto controls the metabolic reprogramming in ESCs to EpiSC transition. Remarkably, Cripto deficiency attenuates ESC lineage restriction in vitro and in vivo, and permits ESC transdifferentiation into trophectoderm lineage, suggesting that Cripto has earlier functions than previously recognized. All together, our studies provide novel insights into the current model of mammalian pluripotency and contribute to the understanding of the extrinsic regulation of the first cell lineage decision in the embryo.

  1. Effects of calcitriol, seocalcitol, and medium-chain triglyceride on a canine transitional cell carcinoma cell line

    DEFF Research Database (Denmark)

    Kaewsakhorn, T.; Kisseberth, W.C.; Capen, C.C.

    2005-01-01

    Background: Transitional cell carcinoma (TCC) in dogs is associated with high morbidity and mortality. Calcitriol and its analog seocalcitol, combined with medium-chain triglyceride (MCT), have potential for the treatment of this disease. Materials and Methods: TCC cells were treated...... with calcitriol or seocalcitol, alone or combined with MCT. Cell growth, cell cycle kinetics, vitamin D receptor (VDR) localization and expression, and Bcl-2 expression were measured. Results: Canine TCC expresses high levels of nuclear VDR. Furthermore, calcitriol and seocalcitol significantly inhibited cell...... inhibited TCC cell growth via induction of cell cycle arrest and MCT enhanced this effect. Therefore, calcitriol and seocalcitol with MCT may have therapeutic potential for canine bladder cancer....

  2. Human colostral cells. I. Separation and characterization.

    Science.gov (United States)

    Crago, S S; Prince, S J; Pretlow, T G; McGhee, J R; Mestecky, J

    1979-12-01

    Analyses of the cells present in human colostrum obtained from fifty-four healthy donors during the first four days of lactation revealed that there were 3.3 x 10(6) (range 1.1 x 10(5)--1.2 x 10(7)) cells per ml of colostrum. Based on histochemical examinations, it was found that this population consisted of 30--47% macrophages, 40--60% polymorphonuclear leucocytes, 5.2--8.9% lymphocytes, and 1.3--2.8% colostral corpuscles; epithelial cells were rarely encountered. The identity of various cell types was confirmed by Wright's stain and by a series of histochemical techniques which disclosed the presence of non-specific esterase, peroxidase, and lipids. For further characterization, the different types of cells were separated by various methods, such as Ficoll-Hypaque density centrifugation, isokinetic centrifugation on a linear Ficoll gradient, adherence to glass or plastic, and phagocytosis of carbonyl iron. Immunohistochemical staining with FITC- and/or TRITC-labelled reagents to IgA, IgM, IgG, K- and lambda-chains, secretory component, lactoferrin, and alpha-lactalbumin were applied to unseparated as well as separated colostral cells. Polymorphonuclear leucocytes (staining for peroxidase) as well as macrophages and colostral corpuscles (staining for non-specific esterase) exhibited numerous intracellular vesicles that contained lipids as well as various combinations of milk proteins. Lymphoid cells did not stain with any of these reagents and plasma cells were not detected among the colostral cells. Individual phagocytic cells contained immunoglobulins of the IgA and IgM classes, both K and lambda light chains, secretory component, lactoferrin, and alpha-lactalbumin. The coincidental appearance of these proteins in single, phagocytic cells but not in lymphoid cells indicate that the cells acquired these proteins by ingestion from the environment. Markers commonly used for the identification of B lymphocytes (surface immunoglobulins) and T lymphocytes (receptors

  3. 21 CFR 864.2280 - Cultured animal and human cells.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cultured animal and human cells. 864.2280 Section... (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture Products § 864.2280 Cultured animal and human cells. (a) Identification. Cultured animal and human cells are in vitro...

  4. Human red blood cells deformed under thermal fluid flow.

    Science.gov (United States)

    Foo, Ji-Jinn; Chan, Vincent; Feng, Zhi-Qin; Liu, Kuo-Kang

    2006-03-01

    The flow-induced mechanical deformation of a human red blood cell (RBC) during thermal transition between room temperature and 42.0 degrees C is interrogated by laser tweezer experiments. Based on the experimental geometry of the deformed RBC, the surface stresses are determined with the aid of computational fluid dynamics simulation. It is found that the RBC is more deformable while heating through 37.0 degrees C to 42.0 degrees C, especially at a higher flow velocity due to a thermal-fluid effect. More importantly, the degree of RBC deformation is irreversible and becomes softer, and finally reaches a plateau (at a uniform flow velocity U > 60 microm s(-1)) after the heat treatment, which is similar to a strain-hardening dominated process. In addition, computational simulated stress is found to be dependent on the progression of thermotropic phase transition. Overall, the current study provides new insights into the highly coupled temperature and hydrodynamic effects on the biomechanical properties of human erythrocyte in a model hydrodynamic flow system.

  5. Management of transitional cell carcinoma of the urinary bladder in dogs: a review.

    Science.gov (United States)

    Fulkerson, Christopher M; Knapp, Deborah W

    2015-08-01

    Transitional cell carcinoma (TCC), also referred to as urothelial carcinoma, is the most common form of urinary bladder cancer in dogs, affecting tens of thousands of dogs worldwide each year. Canine TCC is usually a high grade invasive cancer. Problems associated with TCC include urinary tract obstruction, distant metastases in >50% of affected dogs, and clinical signs that are troubling both to the dogs and to their owners. Risk factors for TCC include exposure to older types of flea control products and lawn chemicals, obesity, female sex, and a very strong breed-associated risk. This knowledge is allowing pet owners to take steps to reduce the risk of TCC in their dog. The diagnosis of TCC is made by histopathology of tissue biopsies obtained by cystoscopy, surgery, or catheter. Percutaneous aspirates and biopsies should be avoided due to the risk of tumor seeding. TCC is most commonly located in the trigone region of the bladder precluding complete surgical resection. Medical treatment is the mainstay for TCC therapy in dogs. Although TCC is not usually curable in dogs, multiple drugs have activity against it. Approximately 75% of dogs respond favorably to TCC treatment and can enjoy several months to a year or more of good quality life. Many promising new therapies for TCC are emerging and with the close similarity between TCC in dogs and high grade invasive bladder cancer in humans, new treatment strategies found to be successful in canine studies are expected to help dogs and to be subsequently translated to humans.

  6. DNA repair responses in human skin cells

    Energy Technology Data Exchange (ETDEWEB)

    Hanawalt, P.C.; Liu, S.C.; Parsons, C.S.

    1981-07-01

    Sunlight and some environmental chemical agents produce lesions in the DNA of human skin cells that if unrepaired may interfere with normal functioning of these cells. The most serious outcome of such interactions may be malignancy. It is therefore important to develop an understanding of mechanisms by which the lesions may be repaired or tolerated without deleterious consequences. Our models for the molecular processing of damaged DNA have been derived largely from the study of bacterial systems. Some similarities but significant differences are revealed when human cell responses are tested against these models. It is also of importance to learn DNA repair responses of epidermal keratinocytes for comparison with the more extensive studies that have been carried out with dermal fibroblasts. Our experimental results thus far indicate similarities for the excision-repair of ultraviolet-induced pyrimidine dimers in human keratinocytes and fibroblasts. Both the monoadducts and the interstrand crosslinks produced in DNA by photoactivated 8-methoxypsoralen (PUVA) can be repaired in normal human fibroblasts but not in those from xeroderma pigmentosum patients. The monoadducts, like pyrimidine dimers, are probably the more mutagenic/carcinogenic lesions while the crosslinks are less easily repaired and probably result in more effective blocking of DNA function. It is suggested that a split-dose protocol that maximizes the production of crosslinks while minimizing the yield of monoadducts may be more effective and potentially less carcinogenic than the single ultraviolet exposure regimen in PUVA therapy for psoriasis.

  7. Human T Cell Memory: A Dynamic View

    Directory of Open Access Journals (Sweden)

    Derek C. Macallan

    2017-02-01

    Full Text Available Long-term T cell-mediated protection depends upon the formation of a pool of memory cells to protect against future pathogen challenge. In this review we argue that looking at T cell memory from a dynamic viewpoint can help in understanding how memory populations are maintained following pathogen exposure or vaccination. For example, a dynamic view resolves the apparent paradox between the relatively short lifespans of individual memory cells and very long-lived immunological memory by focussing on the persistence of clonal populations, rather than individual cells. Clonal survival is achieved by balancing proliferation, death and differentiation rates within and between identifiable phenotypic pools; such pools correspond broadly to sequential stages in the linear differentiation pathway. Each pool has its own characteristic kinetics, but only when considered as a population; single cells exhibit considerable heterogeneity. In humans, we tend to concentrate on circulating cells, but memory T cells in non-lymphoid tissues and bone marrow are increasingly recognised as critical for immune defence; their kinetics, however, remain largely unexplored. Considering vaccination from this viewpoint shifts the focus from the size of the primary response to the survival of the clone and enables identification of critical system pinch-points and opportunities to improve vaccine efficacy.

  8. Biological impact of human embryonic stem cells.

    Science.gov (United States)

    Martín, Miguel; Menéndez, Pablo

    2012-01-01

    Research on human embryonic stem cells (hESCs) and induced pluripotent (iPS) stem cells is currently a field of great potential in biomedicine. These cells represent a highly valuable tool for developmental biology studies, disease models, and drug screening and toxicity. The ultimate goal of hESCs and iPS cell research is the treatment of diseases or disorders for which there is currently no treatment or existing therapies are only partially effective. Despite the disproportionate short-term hopes generated, which are putting too much pressure on scientists, the international scientific community is making rapid progress in understanding hESCs and iPS cells. Nonetheless, great efforts have to be made to provide an answer to still quite basic questions concerning their biology. Moreover, translation to clinical applications in cell replacement therapy requires prior solution to ethical barriers. The recent development of iPS cells has provided a strong alternative to overcome ethical issues concerning hESCs. However, an in-depth characterization of their genetic and epigenetic features, as well as their differentiation potential still remains to be undertaken. This chapter will describe, precisely, what the critical issues are, where scientific and ethical barriers stand, and how we are to overcome them. Only then, we shall finally discover whether hESCs and iPS cells will allow building reproducible disease models, and whether they really are a safe tool, with great potential for regenerative medicine.

  9. How institutions shaped the last major evolutionary transition to large-scale human societies.

    Science.gov (United States)

    Powers, Simon T; van Schaik, Carel P; Lehmann, Laurent

    2016-02-05

    What drove the transition from small-scale human societies centred on kinship and personal exchange, to large-scale societies comprising cooperation and division of labour among untold numbers of unrelated individuals? We propose that the unique human capacity to negotiate institutional rules that coordinate social actions was a key driver of this transition. By creating institutions, humans have been able to move from the default 'Hobbesian' rules of the 'game of life', determined by physical/environmental constraints, into self-created rules of social organization where cooperation can be individually advantageous even in large groups of unrelated individuals. Examples include rules of food sharing in hunter-gatherers, rules for the usage of irrigation systems in agriculturalists, property rights and systems for sharing reputation between mediaeval traders. Successful institutions create rules of interaction that are self-enforcing, providing direct benefits both to individuals that follow them, and to individuals that sanction rule breakers. Forming institutions requires shared intentionality, language and other cognitive abilities largely absent in other primates. We explain how cooperative breeding likely selected for these abilities early in the Homo lineage. This allowed anatomically modern humans to create institutions that transformed the self-reliance of our primate ancestors into the division of labour of large-scale human social organization.

  10. Human fetal liver stromal cells expressing erythropoietin promote hematopoietic development from human embryonic stem cells.

    Science.gov (United States)

    Yang, Chao; Ji, Lei; Yue, Wen; Shi, Shuang-Shuang; Wang, Ruo-Yong; Li, Yan-Hua; Xie, Xiao-Yan; Xi, Jia-Fei; He, Li-Juan; Nan, Xue; Pei, Xue-Tao

    2012-02-01

    Blood cells transfusion and hematopoietic stem cells (HSCs) transplantation are important methods for cell therapy. They are widely used in the treatment of incurable hematological disorder, infectious diseases, genetic diseases, and immunologic deficiency. However, their availability is limited by quantity, capacity of proliferation and the risk of blood transfusion complications. Recently, human embryonic stem cells (hESCs) have been shown to be an alternative resource for the generation of hematopoietic cells. In the current study, we describe a novel method for the efficient production of hematopoietic cells from hESCs. The stable human fetal liver stromal cell lines (hFLSCs) expressing erythropoietin (EPO) were established using the lentiviral system. We observed that the supernatant from the EPO transfected hFLSCs could induce the hESCs differentiation into hematopoietic cells, especially erythroid cells. They not only expressed fetal and embryonic globins but also expressed the adult-globin chain on further maturation. In addition, these hESCs-derived erythroid cells possess oxygen-transporting capacity, which indicated hESCs could generate terminally mature progenies. This should be useful for ultimately developing an animal-free culture system to generate large numbers of erythroid cells from hESCs and provide an experimental model to study early human erythropoiesis.

  11. Delayed transition to new cell fates during cellular reprogramming.

    Science.gov (United States)

    Cheng, Xianrui; Lyons, Deirdre C; Socolar, Joshua E S; McClay, David R

    2014-07-15

    In many embryos specification toward one cell fate can be diverted to a different cell fate through a reprogramming process. Understanding how that process works will reveal insights into the developmental regulatory logic that emerged from evolution. In the sea urchin embryo, cells at gastrulation were found to reprogram and replace missing cell types after surgical dissections of the embryo. Non-skeletogenic mesoderm (NSM) cells reprogrammed to replace missing skeletogenic mesoderm cells and animal caps reprogrammed to replace all endomesoderm. In both cases evidence of reprogramming onset was first observed at the early gastrula stage, even if the cells to be replaced were removed earlier in development. Once started however, the reprogramming occurred with compressed gene expression dynamics. The NSM did not require early contact with the skeletogenic cells to reprogram, but the animal cap cells gained the ability to reprogram early in gastrulation only after extended contact with the vegetal halves prior to that time. If the entire vegetal half was removed at early gastrula, the animal caps reprogrammed and replaced the vegetal half endomesoderm. If the animal caps carried morpholinos to either hox11/13b or foxA (endomesoderm specification genes), the isolated animal caps failed to reprogram. Together these data reveal that the emergence of a reprogramming capability occurs at early gastrulation in the sea urchin embryo and requires activation of early specification components of the target tissues.

  12. Dewetting transition assisted clearance of (NFGAILS) amyloid fibrils from cell membranes by graphene

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Jiajia; Yang, Zaixing; Gu, Zonglin [Institute of Quantitative Biology and Medicine, SRMP and RAD-X, Collaborative Innovation Center of Radiation Medicine of Jiangsu Higher Education Institutions, and Jiangsu Provincial Key Laboratory of Radiation Medicine and Protection, Soochow University, Suzhou 215123 (China); Li, Haotian [Bio-X Lab, Department of Physics, Zhejiang University, Hangzhou 310027 (China); Garate, Jose Antonio [IBM Thomas J. Watson Research Center, Yorktown Heights, New York 10598 (United States); Zhou, Ruhong, E-mail: ruhongz@us.ibm.com [Institute of Quantitative Biology and Medicine, SRMP and RAD-X, Collaborative Innovation Center of Radiation Medicine of Jiangsu Higher Education Institutions, and Jiangsu Provincial Key Laboratory of Radiation Medicine and Protection, Soochow University, Suzhou 215123 (China); IBM Thomas J. Watson Research Center, Yorktown Heights, New York 10598 (United States); Department of Chemistry, Columbia University, New York, New York 10027 (United States)

    2014-12-14

    Clearance of partially ordered oligomers and monomers deposited on cell membrane surfaces is believed to be an effective route to alleviate many potential protein conformational diseases (PCDs). With large-scale all-atom molecular dynamics simulations, here we show that graphene nanosheets can easily and quickly win a competitive adsorption of human islet amyloid polypeptides (hIAPP{sub 22-28}) NFGAILS and associated fibrils against cell membrane, due to graphene's unique two-dimensional, highly hydrophobic surface with its all-sp{sup 2} hybrid structure. A nanoscale dewetting transition was observed at the interfacial region between the fibril (originally deposited on the membrane) and the graphene nanosheet, which significantly assisted the adsorption of fibrils onto graphene from the membrane. The π–π stacking interaction between Phe23 and graphene played a crucial role, providing the driving force for the adsorption at the graphene surface. This study renders new insight towards the importance of water during the interactions between amyloid peptides, the phospholipidic membrane, and graphene, which might shed some light on future developments of graphene-based nanomedicine for preventing/curing PCDs like type II diabetes mellitus.

  13. Transition zone assembly and its contribution to axoneme formation in Drosophila male germ cells.

    Science.gov (United States)

    Vieillard, Jennifer; Paschaki, Marie; Duteyrat, Jean-Luc; Augière, Céline; Cortier, Elisabeth; Lapart, Jean-André; Thomas, Joëlle; Durand, Bénédicte

    2016-09-26

    The ciliary transition zone (TZ) is a complex structure found at the cilia base. Defects in TZ assembly are associated with human ciliopathies. In most eukaryotes, three protein complexes (CEP290, NPHP, and MKS) cooperate to build the TZ. We show that in Drosophila melanogaster, mild TZ defects are observed in the absence of MKS components. In contrast, Cby and Azi1 cooperate to build the TZ by acting upstream of Cep290 and MKS components. Without Cby and Azi1, centrioles fail to form the TZ, precluding sensory cilia assembly, and no ciliary membrane cap associated with sperm ciliogenesis is made. This ciliary cap is critical to recruit the tubulin-depolymerizing kinesin Klp59D, required for regulation of axonemal growth. Our results show that Drosophila TZ assembly in sensory neurons and male germ cells involves cooperative actions of Cby and Dila. They further reveal that temporal control of membrane cap assembly by TZ components and microtubule elongation by kinesin-13 is required for axoneme formation in male germ cells.

  14. Transcutaneous carbon dioxide suppresses epithelial-mesenchymal transition in oral squamous cell carcinoma.

    Science.gov (United States)

    Iwata, Eiji; Hasegawa, Takumi; Takeda, Daisuke; Ueha, Takeshi; Kawamoto, Teruya; Akisue, Toshihiro; Sakai, Yoshitada; Komori, Takahide

    2016-04-01

    Oral squamous cell carcinoma (OSCC) is the most common form of oral cancers. Recent studies have shown that the malignant transformation of various carcinomas, including OSCC, is associated with epithelial-mesenchymal transition (EMT), and that expression of the EMT factors are significantly associated with tumor invasion, tumor metastasis, and survival rates in OSCC patients. Hence, there is a possibility that EMT suppression may improve the prognosis of OSCC patients. Hypoxia inducible factor-1α (HIF-1α) is a crucial microenvironmental factor in tumor progression, which induces the expression of EMT factors. We previously reported that transcutaneous CO2 suppresses both human OSCC tumor growth and metastasis to the regional lymph nodes by improving hypoxia in treated tissue. According to this background, we hypothesized that increased EMT with HIF-1α expression may increase the progression and the metastatic potential of OSCC, and that decreased hypoxia by transcutaneous CO2 could suppress EMT. In the present study, in vitro studies showed that hypoxic conditions increased the expression of HIF-1α and EMT factors in OSCC cells. In addition, in vivo studies revealed that transcutaneous CO2 increased E-cadherin expression with the decreased expression of HIF-1α, Snail, Slug, N-cadherin, and Vimentin in tumor treatment. These results suggest that transcutaneous CO2 could suppress EMT by improving hypoxia, resulting in the reduction of metastatic potential of OSCC. The findings indicate that transcutaneous CO2 may be able to improve the prognosis of OSCC patients through the suppression of EMT.

  15. Cell phoney: human cloning after Quintavalle.

    Science.gov (United States)

    Morgan, Derek; Ford, Mary

    2004-12-01

    Reproductive cloning has thrown up new scientific possibilities, ethical conundrums, and legal challenges. An initial question, considered by the English courts in 2003, was whether the technique presently available, that of cell nucleus replacement, falls outside the provisions of the Human Fertilisation and Embryology Act 1990. If it does, the creation and use, including use in research protocols, of human embryos would be unregulated, disclosing a need to consider remedial legislation. The resolution by the courts of this legal question dramatically engages them in a resolution of fundamental ethical dilemmas, and discloses the possibilities and limitation of negotiating science policy through the processes of litigation.

  16. Regulation of matrix stiffness on the epithelial-mesenchymal transition of breast cancer cells under hypoxia environment

    Science.gov (United States)

    Lv, Yonggang; Chen, Can; Zhao, Boyuan; Zhang, Xiaomei

    2017-06-01

    Substrate stiffness and hypoxia are associated with tumor development and progression, respectively. However, the synergy of them on the biological behavior of human breast cancer cell is still largely unknown. This study explored how substrate stiffness regulates the cell phenotype, viability, and epithelial-mesenchymal transition (EMT) of human breast cancer cells MCF-7 under hypoxia (1% O2). TRITC-phalloidin staining showed that MCF-7 cells transformed from round to irregular polygon with stiffness increase either in normoxia or hypoxia. While being accompanied with the upward tendency from a 0.5- to a 20-kPa substrate, the percentage of cell apoptosis was significantly higher in hypoxia than that in normoxia, especially on the 20-kPa substrate. Additionally, it was hypoxia, but not normoxia, that promoted the EMT of MCF-7 by upregulating hypoxia-inducible factor-1α (HIF-1α), vimentin, Snail 1, and matrix metalloproteinase 2 (MMP 2) and 9 (MMP 9), and downregulating E-cadherin simultaneously regardless of the change of substrate stiffness. In summary, this study discovered that hypoxia and stiffer substrate (20 kPa) could synergistically induce phenotype change, apoptosis, and EMT of MCF-7 cells. Results of this study have an important significance on further exploring the synergistic effect of stiffness and hypoxia on the EMT of breast cancer cells and its molecular mechanism.

  17. Bay Area Transit Agencies Propel Fuel Cell Buses Toward Commercialization (Fact Sheet)

    Energy Technology Data Exchange (ETDEWEB)

    2010-07-01

    This fact sheet describes the Zero Emission Bay Area (ZEBA) demonstration of the next generation of fuel cells buses. Several transit agencies in the San Francisco Bay Area are participating in demonstrating the largest single fleet of fuel cell buses in the United States.

  18. Fuel Cell Buses in U.S. Transit Fleets: Current Status 2012

    Energy Technology Data Exchange (ETDEWEB)

    Eudy, Leslie [National Renewable Energy Lab. (NREL), Golden, CO (United States); Chandler, Kevin [Battelle, Columbus, OH (United States); Gikakis, Christina [Federal Transit Administration, Washington, DC (United States)

    2012-11-01

    This report is the sixth in an annual series of reports that summarize the progress of fuel cell electric bus (FCEB) development in the United States and discuss the achievements and challenges of introducing fuel cell propulsion in transit. The report also provides a snapshot of current FCEB performance results over the last year.

  19. Chronic inorganic arsenic exposure in vitro induces a cancer cell phenotype in human peripheral lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Person, Rachel J.; Olive Ngalame, Ntube N.; Makia, Ngome L.; Bell, Matthew W.; Waalkes, Michael P.; Tokar, Erik J., E-mail: tokare@niehs.nih.gov

    2015-07-01

    Inorganic arsenic is a human lung carcinogen. We studied the ability of chronic inorganic arsenic (2 μM; as sodium arsenite) exposure to induce a cancer phenotype in the immortalized, non-tumorigenic human lung peripheral epithelial cell line, HPL-1D. After 38 weeks of continuous arsenic exposure, secreted matrix metalloproteinase-2 (MMP2) activity increased to over 200% of control, levels linked to arsenic-induced cancer phenotypes in other cell lines. The invasive capacity of these chronic arsenic-treated lung epithelial (CATLE) cells increased to 320% of control and colony formation increased to 280% of control. CATLE cells showed enhanced proliferation in serum-free media indicative of autonomous growth. Compared to control cells, CATLE cells showed reduced protein expression of the tumor suppressor gene PTEN (decreased to 26% of control) and the putative tumor suppressor gene SLC38A3 (14% of control). Morphological evidence of epithelial-to-mesenchymal transition (EMT) occurred in CATLE cells together with appropriate changes in expression of the EMT markers vimentin (VIM; increased to 300% of control) and e-cadherin (CDH1; decreased to 16% of control). EMT is common in carcinogenic transformation of epithelial cells. CATLE cells showed increased KRAS (291%), ERK1/2 (274%), phosphorylated ERK (p-ERK; 152%), and phosphorylated AKT1 (p-AKT1; 170%) protein expression. Increased transcript expression of metallothioneins, MT1A and MT2A and the stress response genes HMOX1 (690%) and HIF1A (247%) occurred in CATLE cells possibly in adaptation to chronic arsenic exposure. Thus, arsenic induced multiple cancer cell characteristics in human peripheral lung epithelial cells. This model may be useful to assess mechanisms of arsenic-induced lung cancer. - Highlights: • Chronic arsenic exposure transforms a human peripheral lung epithelia cell line. • Cells acquire characteristics in common with human lung adenocarcinoma cells. • These transformed cells provide a

  20. Cell surface glycan alterations in epithelial mesenchymal transition process of Huh7 hepatocellular carcinoma cell.

    Directory of Open Access Journals (Sweden)

    Shan Li

    Full Text Available BACKGROUND AND OBJECTIVE: Due to recurrence and metastasis, the mortality of Hepatocellular carcinoma (HCC is high. It is well known that the epithelial mesenchymal transition (EMT and glycan of cell surface glycoproteins play pivotal roles in tumor metastasis. The goal of this study was to identify HCC metastasis related differential glycan pattern and their enzymatic basis using a HGF induced EMT model. METHODOLOGY: HGF was used to induce HCC EMT model. Lectin microarray was used to detect the expression of cell surface glycan and the difference was validated by lectin blot and fluorescence cell lectin-immunochemistry. The mRNA expression levels of glycotransferases were determined by qRT-PCR. RESULTS: After HGF treatment, the Huh7 cell lost epithelial characteristics and obtained mesenchymal markers. These changes demonstrated that HGF could induce a typical cell model of EMT. Lectin microarray analysis identified a decreased affinity in seven lectins ACL, BPL, JAC, MPL, PHA-E, SNA, and SBA to the glycan of cell surface glycoproteins. This implied that glycan containing T/Tn-antigen, NA2 and bisecting GlcNAc, Siaα2-6Gal/GalNAc, terminal α or βGalNAc structures were reduced. The binding ability of thirteen lectins, AAL, LCA, LTL, ConA, NML, NPL, DBA, HAL, PTL II, WFL, ECL, GSL II and PHA-L to glycan were elevated, and a definite indication that glycan containing terminal αFuc and ± Sia-Le, core fucose, α-man, gal-β(α GalNAc, β1,6 GlcNAc branching and tetraantennary complex oligosaccharides structures were increased. These results were further validated by lectin blot and fluorescence cell lectin-immunochemistry. Furthermore, the mRNA expression level of Mgat3 decreased while that of Mgat5, FucT8 and β3GalT5 increased. Therefore, cell surface glycan alterations in the EMT process may coincide with the expression of glycosyltransferase. CONCLUSIONS: The findings of this study systematically clarify the alterations of cell surface

  1. Hypoxia induces a phase transition within a kinase signaling network in cancer cells.

    Science.gov (United States)

    Wei, Wei; Shi, Qihui; Remacle, Francoise; Qin, Lidong; Shackelford, David B; Shin, Young Shik; Mischel, Paul S; Levine, R D; Heath, James R

    2013-04-09

    Hypoxia is a near-universal feature of cancer, promoting glycolysis, cellular proliferation, and angiogenesis. The molecular mechanisms of hypoxic signaling have been intensively studied, but the impact of changes in oxygen partial pressure (pO2) on the state of signaling networks is less clear. In a glioblastoma multiforme (GBM) cancer cell model, we examined the response of signaling networks to targeted pathway inhibition between 21% and 1% pO2. We used a microchip technology that facilitates quantification of a panel of functional proteins from statistical numbers of single cells. We find that near 1.5% pO2, the signaling network associated with mammalian target of rapamycin (mTOR) complex 1 (mTORC1)--a critical component of hypoxic signaling and a compelling cancer drug target--is deregulated in a manner such that it will be unresponsive to mTOR kinase inhibitors near 1.5% pO2, but will respond at higher or lower pO2 values. These predictions were validated through experiments on bulk GBM cell line cultures and on neurosphere cultures of a human-origin GBM xenograft tumor. We attempt to understand this behavior through the use of a quantitative version of Le Chatelier's principle, as well as through a steady-state kinetic model of protein interactions, both of which indicate that hypoxia can influence mTORC1 signaling as a switch. The Le Chatelier approach also indicates that this switch may be thought of as a type of phase transition. Our analysis indicates that certain biologically complex cell behaviors may be understood using fundamental, thermodynamics-motivated principles.

  2. Lymphoid Cell-Glioma Cell Interaction Enhances Cell Coat Production by Human Gliomas: Novel Suppressor Mechanism

    Science.gov (United States)

    Dick, Steven J.; Macchi, Beatrice; Papazoglou, Savvas; Oldfield, Edward H.; Kornblith, Paul L.; Smith, Barry H.; Gately, Maurice K.

    1983-05-01

    Certain human glioma lines produce mucopolysaccharide coats that impair the generation of cytolytic lymphocytes in response to these lines in vitro. Coat production is substantially enhanced by the interaction of glioma cells with a macromolecular factor released by human peripheral blood mononuclear cells in culture. This interaction thus constitutes an unusual mechanism by which inflammatory cells may nonspecifically suppress the cellular immune response to at least one class of solid tumors in humans.

  3. Human induced pluripotent stem cell-derived models to investigate human cytomegalovirus infection in neural cells.

    Directory of Open Access Journals (Sweden)

    Leonardo D'Aiuto

    Full Text Available Human cytomegalovirus (HCMV infection is one of the leading prenatal causes of congenital mental retardation and deformities world-wide. Access to cultured human neuronal lineages, necessary to understand the species specific pathogenic effects of HCMV, has been limited by difficulties in sustaining primary human neuronal cultures. Human induced pluripotent stem (iPS cells now provide an opportunity for such research. We derived iPS cells from human adult fibroblasts and induced neural lineages to investigate their susceptibility to infection with HCMV strain Ad169. Analysis of iPS cells, iPS-derived neural stem cells (NSCs, neural progenitor cells (NPCs and neurons suggests that (i iPS cells are not permissive to HCMV infection, i.e., they do not permit a full viral replication cycle; (ii Neural stem cells have impaired differentiation when infected by HCMV; (iii NPCs are fully permissive for HCMV infection; altered expression of genes related to neural metabolism or neuronal differentiation is also observed; (iv most iPS-derived neurons are not permissive to HCMV infection; and (v infected neurons have impaired calcium influx in response to glutamate.

  4. Mediation of transitional B cell maturation in the absence of functional Bruton’s tyrosine kinase

    Science.gov (United States)

    Tanwar, Shalini; Dhar, Atika; Varanasi, Vineeth; Mukherjee, Tapas; Boppana, Ramanamurthy; Basak, Soumen; Bal, Vineeta; George, Anna; Rath, Satyajit

    2017-01-01

    X-linked immune-deficient (Xid) mice, carrying a mutation in Bruton’s tyrosine kinase (Btk), have multiple B cell lineage differentiation defects. We now show that, while Xid mice showed only mild reduction in the frequency of the late transitional (T2) stage of peripheral B cells, the defect became severe when the Xid genotype was combined with either a CD40-null, a TCRbeta-null or an MHC class II (MHCII)-null genotype. Purified Xid T1 and T2 B cells survived poorly in vitro compared to wild-type (WT) cells. BAFF rescued WT but not Xid T1 and T2 B cells from death in culture, while CD40 ligation equivalently rescued both. Xid transitional B cells ex vivo showed low levels of the p100 protein substrate for non-canonical NF-kappaB signalling. In vitro, CD40 ligation induced equivalent activation of the canonical but not of the non-canonical NF-kappaB pathway in Xid and WT T1 and T2 B cells. CD40 ligation efficiently rescued p100-null T1 B cells from neglect-induced death in vitro. These data indicate that CD40-mediated signals, likely from CD4 T cells, can mediate peripheral transitional B cell maturation independent of Btk and the non-canonical NF-kappaB pathway, and thus contribute to the understanding of the complexities of peripheral B cell maturation. PMID:28378771

  5. Mediation of transitional B cell maturation in the absence of functional Bruton's tyrosine kinase.

    Science.gov (United States)

    Tanwar, Shalini; Dhar, Atika; Varanasi, Vineeth; Mukherjee, Tapas; Boppana, Ramanamurthy; Basak, Soumen; Bal, Vineeta; George, Anna; Rath, Satyajit

    2017-04-05

    X-linked immune-deficient (Xid) mice, carrying a mutation in Bruton's tyrosine kinase (Btk), have multiple B cell lineage differentiation defects. We now show that, while Xid mice showed only mild reduction in the frequency of the late transitional (T2) stage of peripheral B cells, the defect became severe when the Xid genotype was combined with either a CD40-null, a TCRbeta-null or an MHC class II (MHCII)-null genotype. Purified Xid T1 and T2 B cells survived poorly in vitro compared to wild-type (WT) cells. BAFF rescued WT but not Xid T1 and T2 B cells from death in culture, while CD40 ligation equivalently rescued both. Xid transitional B cells ex vivo showed low levels of the p100 protein substrate for non-canonical NF-kappaB signalling. In vitro, CD40 ligation induced equivalent activation of the canonical but not of the non-canonical NF-kappaB pathway in Xid and WT T1 and T2 B cells. CD40 ligation efficiently rescued p100-null T1 B cells from neglect-induced death in vitro. These data indicate that CD40-mediated signals, likely from CD4 T cells, can mediate peripheral transitional B cell maturation independent of Btk and the non-canonical NF-kappaB pathway, and thus contribute to the understanding of the complexities of peripheral B cell maturation.

  6. Human somatic cell nuclear transfer and cloning.

    Science.gov (United States)

    2012-10-01

    This document presents arguments that conclude that it is unethical to use somatic cell nuclear transfer (SCNT) for infertility treatment due to concerns about safety; the unknown impact of SCNT on children, families, and society; and the availability of other ethically acceptable means of assisted reproduction. This document replaces the ASRM Ethics Committee report titled, "Human somatic cell nuclear transfer (cloning)," last published in Fertil Steril 2000;74:873-6. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  7. Research of TGF-beta1 Inducing Lung Adencarcinoma PC9 Cells to Mesenchymal Cells Transition

    Directory of Open Access Journals (Sweden)

    Xiaofeng CHEN

    2010-01-01

    Full Text Available Background and objective It has been proven that epithelial-mesenchymal transition (EMT not only correlated with embryonic development but also could promote tumor invasion and metastasis. Transforming growth factor beta-1 (TGF-β1 has been identified as the main inducer of tumor EMT. The aim of this study was to investigate the effects of TGF-β1 on EMT and PI3K/AKT signaling pathway in lung adencarcinoma PC9 cells. Methods Cultured PC9 cells were treated with different concentrations of TGF-β1 for 48 h. The morphological changes were observed under phase-contrast microscopy; EMT relative marker protein changes were assessed by Western blot and immunoflurescence staining. In addition, the expression of AKT and P-AKT were also measured by Western blot. Results The data showed that TGF-β1 could induce PC9 morphological alteration from epithelial to mesenchymal and upregulate the expression of mesenchymal maker protein Fibronectin. Obviously, the expression of P-AKT was downregulated by TGF-β1 treatment for 48 h. Conclusion TGF-β1 might induce EMT of PC9 cells , accompanied by the changes of PI3K/AKT signaling pathway.

  8. Dopamine receptor repertoire of human granulosa cells

    Directory of Open Access Journals (Sweden)

    Kunz Lars

    2007-10-01

    Full Text Available Abstract Background High levels of dopamine (DA were described in human ovary and recently evidence for DA receptors in granulosa and luteal cells has been provided, as well. However, neither the full repertoire of ovarian receptors for DA, nor their specific role, is established. Human granulosa cells (GCs derived from women undergoing in vitro fertilization (IVF are an adequate model for endocrine cells of the follicle and the corpus luteum and were therefore employed in an attempt to decipher their DA receptor repertoire and functionality. Methods Cells were obtained from patients undergoing IVF and examined using cDNA-array, RT-PCR, Western blotting and immunocytochemistry. In addition, calcium measurements (with FLUO-4 were employed. Expression of two DA receptors was also examined by in-situ hybridization in rat ovary. Effects of DA on cell viability and cell volume were studied by using an ATP assay and an electronic cell counter system. Results We found members of the two DA receptor families (D1- and D2 -like associated with different signaling pathways in human GCs, namely D1 (as expected and D5 (both are Gs coupled and linked to cAMP increase and D2, D4 (Gi/Gq coupled and linked to IP3/DAG. D3 was not found. The presence of the trophic hormone hCG (10 IU/ml in the culture medium for several days did not alter mRNA (semiquantitative RT-PCR or protein levels (immunocytochemistry/Western blotting of D1,2,4,5 DA receptors. Expression of prototype receptors for the two families, D1 and D2, was furthermore shown in rat granulosa and luteal cells by in situ hybridization. Among the DA receptors found in human GCs, D2 expression was marked both at mRNA and protein levels and it was therefore further studied. Results of additional RT-PCR and Western blots showed two splice variants (D2L, D2S. Irrespective of these variants, D2 proved to be functional, as DA raised intracellular calcium levels. This calcium mobilizing effect of DA was observed

  9. JDP2 inhibits the epithelial-to-mesenchymal transition in pancreatic cancer BxPC3 cells.

    Science.gov (United States)

    Liu, Zhe; Du, Ruixia; Long, Jin; Dong, Anbing; Fan, Jianpeng; Guo, Kejian; Xu, Yuanhong

    2012-10-01

    Pancreatic carcinoma is one of the most malignant and aggressive cancers. Increased motility and invasiveness of pancreatic cancer cells are believed to be associated with epithelial-to-mesenchymal transition (EMT). However, the molecular basis of EMT in pancreatic cancer cells is poorly understood. In this study, we examined the relationship between Jun dimerization protein 2 (JDP2), which is an AP-1 inhibitor, and EMT in human pancreatic carcinoma cells. We demonstrated that transforming growth factor-β1 (TGF-β1) promoted epidermal growth factor (EGF)-induced EMT in co-treated human pancreatic BxPC3 cells and that JDP2 overexpression reversed the EMT that was induced by co-treatment with TGF-β1 and EGF. These results suggest that EGF plays a principal role in EMT through its association with TGF-β1 in human pancreatic BxPC3 cells and that JDP2 may be a molecular target for pancreatic carcinoma intervention.

  10. Rapid G0/1 transition and cell cycle progression in CD8(+) T cells compared to CD4(+) T cells following in vitro stimulation.

    Science.gov (United States)

    Mishima, Takuya; Fukaya, Shotaro; Toda, Shoko; Ando, Yoshiaki; Matsunaga, Tsukasa; Inobe, Manabu

    2017-04-01

    T cell population consists of two major subsets, CD4(+) T cells and CD8(+) T cells, which can be distinguished by the expression of CD4 or CD8 molecules, respectively. Although they play quite different roles in an immune system, many of their basic cellular processes such as proliferation following stimulation are presumably common. In this study, we have carefully analyzed time course of G0/1 transition as well as cell cycle progression in the two subsets of quiescent T cell population following in vitro growth stimulation. We found that CD8(+) T cells promote G0/1 transition more rapidly and drive their cell cycle progression faster compared to CD4(+) T cells. In addition, expression of CD25 and effects of its blockade revealed that IL-2 is implicated in the rapid progression, but not the earlier G0/1 transition, of CD8(+) T cells.

  11. Endothelial adhesion of synchronized gastric tumor cells changes during cell cycle transit and correlates with the expression level of CD44 splice variants

    Institute of Scientific and Technical Information of China (English)

    Anton Oertl; Jens Castein; Tobias Engl; Wolf-Dietrich Beecken; Dietger Jonas; Richard Melamed; Roman A. Blaheta

    2005-01-01

    AIM: To study adhesion capacity and CD44 expression of human gastric adenocarcinoma MKN45 cells at different stages of a first cell cycle.METHODS: MKN45 cells were synchronized by aphidicolin and assayed for adhesion to an endothelial cell (HUVEC)monolayer. Surface expression of CD44 and CD44 splice variants on MKN45 cells was evaluated by flow cytometry.Functional relevance of CD44 adhesion receptors was investigated by blocking studies using anti CD44 monoclonal antibodies or by hyaluronan digestion.RESULTS: Adhesion of MKN45 to HUVEC was increased during G2/M transit, after which adhesion returned to baseline levels with cell cycle completion. In parallel, CD44splice variants CD44v4, CD44v5, and CD44v7 were all upregulated on MKN45 during cell cycle progression with a maximum effect in G2/M. The function of CD44 surface receptors was assessed with specific receptor blocking monodonal antibodies or removal of hyaluronan by digestion with hyaluronidase. Both strategies inhibited tumor cell adhesion to HUVEC by nearly 50%, which indicates that MKN45-HUVEC-interaction is CD44 dependent.CONCLUSION: CD44 expression level is linked to the cell cycle in gastrointestinal tumor cells, which in turn leads to cell cyde dependent alterations of their adhesion behaviour to endothelium.

  12. Heat Capacity Changes for Transition-State Analogue Binding and Catalysis with Human 5'-Methylthioadenosine Phosphorylase.

    Science.gov (United States)

    Firestone, Ross S; Cameron, Scott A; Karp, Jerome M; Arcus, Vickery L; Schramm, Vern L

    2017-02-17

    Human 5'-methylthioadenosine phosphorylase (MTAP) catalyzes the phosphorolysis of 5'-methylthioadenosine (MTA). Its action regulates cellular MTA and links polyamine synthesis to S-adenosylmethionine (AdoMet) salvage. Transition state analogues with picomolar dissociation constants bind to MTAP in an entropically driven process at physiological temperatures, suggesting increased hydrophobic character or dynamic structure for the complexes. Inhibitor binding exhibits a negative heat capacity change (-ΔCp), and thus the changes in enthalpy and entropy upon binding are strongly temperature-dependent. The ΔCp of inhibitor binding by isothermal titration calorimetry does not follow conventional trends and is contrary to that expected from the hydrophobic effect. Thus, ligands of increasing hydrophobicity bind with increasing values of ΔCp. Crystal structures of MTAP complexed to transition-state analogues MT-DADMe-ImmA, BT-DADMe-ImmA, PrT-ImmA, and a substrate analogue, MT-tubercidin, reveal similar active site contacts and overall protein structural parameters, despite large differences in ΔCp for binding. In addition, ΔCp values are not correlated with Kd values. Temperature dependence of presteady state kinetics revealed the chemical step for the MTAP reaction to have a negative heat capacity for transition state formation (-ΔCp(‡)). A comparison of the ΔCp(‡) for MTAP presteady state chemistry and ΔCp for inhibitor binding revealed those transition-state analogues most structurally and thermodynamically similar to the transition state. Molecular dynamics simulations of MTAP apoenzyme and complexes with MT-DADMe-ImmA and MT-tubercidin show small, but increased dynamic motion in the inhibited complexes. Variable temperature CD spectroscopy studies for MTAP-inhibitor complexes indicate remarkable protein thermal stability (to Tm = 99 °C) in complexes with transition-state analogues.

  13. TGFβ Pathway Inhibition Redifferentiates Human Pancreatic Islet β Cells Expanded In Vitro.

    Directory of Open Access Journals (Sweden)

    Ginat Toren-Haritan

    Full Text Available In-vitro expansion of insulin-producing cells from adult human pancreatic islets could provide an abundant cell source for diabetes therapy. However, proliferation of β-cell-derived (BCD cells is associated with loss of phenotype and epithelial-mesenchymal transition (EMT. Nevertheless, BCD cells maintain open chromatin structure at β-cell genes, suggesting that they could be readily redifferentiated. The transforming growth factor β (TGFβ pathway has been implicated in EMT in a range of cell types. Here we show that human islet cell expansion in vitro involves upregulation of the TGFβ pathway. Blocking TGFβ pathway activation using short hairpin RNA (shRNA against TGFβ Receptor 1 (TGFBR1, ALK5 transcripts inhibits BCD cell proliferation and dedifferentiation. Treatment of expanded BCD cells with ALK5 shRNA results in their redifferentiation, as judged by expression of β-cell genes and decreased cell proliferation. These effects, which are reproducible in cells from multiple human donors, are mediated, at least in part, by AKT-FOXO1 signaling. ALK5 inhibition synergizes with a soluble factor cocktail to promote BCD cell redifferentiation. The combined treatment may offer a therapeutically applicable way for generating an abundant source of functional insulin-producing cells following ex-vivo expansion.

  14. Withaferin A Inhibits Experimental Epithelial-Mesenchymal Transition in MCF-10A Cells and Suppresses Vimentin Protein Level in Vivo in Breast Tumors

    OpenAIRE

    Lee, Joomin; Hahm, Eun-Ryeong; Marcus, Adam I.; Singh, Shivendra V.

    2013-01-01

    We have shown previously that withaferin A (WA), a bioactive component of the medicinal plant Withania somnifera, inhibits growth of cultured and xenografted human breast cancer cells and prevents breast cancer development and pulmonary metastasis incidence in a transgenic mouse model. The present study was undertaken to determine if the anticancer effect of WA involved inhibition of epithelial-mesenchymal transition (EMT). Experimental EMT induced by exposure of MCF-10A cells to tumor necros...

  15. TALEN-Induced Translocations in Human Cells.

    Science.gov (United States)

    Piganeau, Marion; Renouf, Benjamin; Ghezraoui, Hind; Brunet, Erika

    2016-01-01

    Induction of chromosomal translocations in human cells is of a great interest to study tumorigenesis and genome instability. Here, we explain in detail a method to induce translocations using the transcription activator-like effector nucleases (TALENs). We describe how to detect translocation formation by PCR, calculate translocation frequency by 96-well PCR screen, and analyze breakpoint junctions. When inducing cancer translocations, it is also possible to detect the fusion gene by FISH analysis or western blot.

  16. Intermediate band solar cells. The transition metal supersaturated Silicon approach

    OpenAIRE

    González Díaz, Germán; García Hemme, Eric; García Hernansanz, Rodrigo; Olea Ariza, Javier; Pastor Pastor, David; Prado Millán, Alvaro del; Mártil de la Plaza, Ignacio; Wahnón Benarroch, Perla

    2013-01-01

    Within the framework of the third solar cell generation some new ideas to enlarge the spectral response of the solar cells toward the infrared have been proposed. Among them the inclusion of an Intermediate Band (IB) seems to be very promising. This paper will deal with one of the ways to generate the IB namely the deep level center approach. We will discuss not only its existence but also the carriers lifetime recovery which is necessary to obtain the expected increase of the solar cell effi...

  17. The core regulatory network in human cells.

    Science.gov (United States)

    Kim, Man-Sun; Kim, Dongsan; Kang, Nam Sook; Kim, Jeong-Rae

    2017-03-04

    In order to discover the common characteristics of various cell types in the human body, many researches have been conducted to find the set of genes commonly expressed in various cell types and tissues. However, the functional characteristics of a cell is determined by the complex regulatory relationships among the genes rather than by expressed genes themselves. Therefore, it is more important to identify and analyze a core regulatory network where all regulatory relationship between genes are active across all cell types to uncover the common features of various cell types. Here, based on hundreds of tissue-specific gene regulatory networks constructed by recent genome-wide experimental data, we constructed the core regulatory network. Interestingly, we found that the core regulatory network is organized by simple cascade and has few complex regulations such as feedback or feed-forward loops. Moreover, we discovered that the regulatory links from genes in the core regulatory network to genes in the peripheral regulatory network are much more abundant than the reverse direction links. These results suggest that the core regulatory network locates at the top of regulatory network and plays a role as a 'hub' in terms of information flow, and the information that is common to all cells can be modified to achieve the tissue-specific characteristics through various types of feedback and feed-forward loops in the peripheral regulatory networks. We also found that the genes in the core regulatory network are evolutionary conserved, essential and non-disease, non-druggable genes compared to the peripheral genes. Overall, our study provides an insight into how all human cells share a common function and generate tissue-specific functional traits by transmitting and processing information through regulatory network.

  18. Inhibition of RAB1A suppresses epithelial-mesenchymal transition and proliferation of triple-negative breast cancer cells.

    Science.gov (United States)

    Xu, Hui; Qian, Mingping; Zhao, Bingkun; Wu, Chenyang; Maskey, Niraj; Song, Hongming; Li, Dengfeng; Song, Jialu; Hua, Kaiyao; Fang, Lin

    2017-03-01

    RAB1A acts as an oncogene in various cancers, and emerging evidence has verified that RAB1A is an mTORC1 activator in hepatocellular and colorectal cancer, but the role of RAB1A in breast cancer remains unclear. In this investigation, RAB1A siRNA was successfully transfected in MDA-MB-231 and BT-549 human triple-negative breast cancer cells, and verified by real‑time quantitative polymerase chain reaction and western blotting. Then, MTT cell proliferation, colony formation, cell invasion and wound healing assays were performed to characterize the function of RAB1A in the breast cancer cell lines. Downregulation of RAB1A inhibited cellular growth, cell migration, cell invasion and cell epithelial-mesenchymal transition. Furthermore, compared with NC siRNA transfected cells, RAB1A siRNA transfected breast cancer cells inhibited the phosphorylation of S6K1, the effector molecular of mTORC1. Collectively, our data suggested that RAB1A acts as an oncogene by regulating cellular proliferation, growth, invasion and metastasis via activation of mTORC1 pathway in triple-negative breast cancer.

  19. Cell Culture Assay for Human Noroviruses [response

    Energy Technology Data Exchange (ETDEWEB)

    Straub, Tim M.; Honer Zu Bentrup, Kerstin; Orosz Coghlan, Patricia; Dohnalkova, Alice; Mayer, Brooke K.; Bartholomew, Rachel A.; Valdez, Catherine O.; Bruckner-Lea, Cindy J.; Gerba, Charles P.; Abbaszadegan, Morteza A.; Nickerson, Cheryl A.

    2007-07-01

    We appreciate the comments provided by Leung et al., in response to our recently published article “In Vitro Cell Culture Infectivity Assay for Human Noroviruses” by Straub et al. (1). The specific aim of our project was to develop an in vitro cell culture infectivity assay for human noroviruses (hNoV) to enhance risk assessments when they are detected in water supplies. Reverse transcription (RT) qualitative or quantitative PCR are the primary assays for waterborne NoV monitoring. However, these assays cannot distinguish between infectious vs. non-infectious virions. When hNoV is detected in water supplies, information provided by our infectivity assay will significantly improve risk assessment models and protect human health, regardless of whether we are propagating NoV. Indeed, in vitro cell culture infectivity assays for the waterborne pathogen Cryptosporidium parvum that supplement approved fluorescent microscopy assays, do not result in amplification of the environmentally resistant hard-walled oocysts (2). However, identification of life cycle stages in cell culture provides evidence of infectious oocysts in a water supply. Nonetheless, Leung et al.’s assertion regarding the suitability of our method for the in vitro propagation of high titers of NoV is valid for the medical research community. In this case, well-characterized challenge pools of virus would be useful for developing and testing diagnostics, therapeutics, and vaccines. As further validation of our published findings, we have now optimized RT quantitative PCR to assess the level of viral production in cell culture, where we are indeed finding significant increases in viral titer. The magnitude and time course of these increases is dependent on both virus strain and multiplicity of infection. We are currently preparing a manuscript that will discuss these findings in greater detail, and the implications this may have for creating viral challenge pools

  20. Preliminary study on human fibroblasts as feeder layer for human embryonic stem cells culture in vitro

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    To avoid the direct contact with mouse cells and possible heterogeneous pathogen in future application, we need to replace mouse embryonic fibroblastswith human fibroblasts as the feeder layer to maintain human embryonic stem cells growth in the undifferentiated state. We successfully use human fibroblasts derived from aborted fetus and adult prepuce as feeder layer to maintain human embryonic stem cells growth. During the passage and growth on this feeder layer, the human embryonic stem cells can keep their undifferentiated state.

  1. Fuel oxidation at the walk-to-run-transition in humans.

    Science.gov (United States)

    Ganley, Kathleen J; Stock, Anthony; Herman, Richard M; Santello, Marco; Willis, Wayne T

    2011-05-01

    Multiple factors (including anthropometric, kinetic, mechanical, kinematic, perceptual, and energetic factors) are likely to play a role in the walk-to-run transition in humans. The primary purpose of the present study was to consider an additional factor, the metabolic fuel source. Indirect calorimetry was used to measure fuel oxidation, and perception of effort was recorded as 10 overnight-fasted adults locomoted on a level treadmill at speeds progressing from 1.56 to 2.46 m s(-1) in increments of 0.11 m s(-1) and 10.0 minutes under 3 conditions: (1) unconstrained choice of gait, (2) walking at all speeds, and (3) running at all speeds. The preferred transition speed was 2.08 ± 0.03 m s(-1). Gait transition from walking to running increased oxygen consumption rate, decreased the perception of effort, and decreased the rate of carbohydrate oxidation. We propose that, in an evolutionary context, gait transition, guided by the perception of effort, can be viewed as a carbohydrate-sparing strategy.

  2. Proteomic and protein interaction network analysis of human T lymphocytes during cell-cycle entry

    Science.gov (United States)

    Orr, Stephen J; Boutz, Daniel R; Wang, Rong; Chronis, Constantinos; Lea, Nicholas C; Thayaparan, Thivyan; Hamilton, Emma; Milewicz, Hanna; Blanc, Eric; Mufti, Ghulam J; Marcotte, Edward M; Thomas, N Shaun B

    2012-01-01

    Regulating the transition of cells such as T lymphocytes from quiescence (G0) into an activated, proliferating state involves initiation of cellular programs resulting in entry into the cell cycle (proliferation), the growth cycle (blastogenesis, cell size) and effector (functional) activation. We show the first proteomic analysis of protein interaction networks activated during entry into the first cell cycle from G0. We also provide proof of principle that blastogenesis and proliferation programs are separable in primary human T cells. We employed a proteomic profiling method to identify large-scale changes in chromatin/nuclear matrix-bound and unbound proteins in human T lymphocytes during the transition from G0 into the first cell cycle and mapped them to form functionally annotated, dynamic protein interaction networks. Inhibiting the induction of two proteins involved in two of the most significantly upregulated cellular processes, ribosome biogenesis (eIF6) and hnRNA splicing (SF3B2/SF3B4), showed, respectively, that human T cells can enter the cell cycle without growing in size, or increase in size without entering the cell cycle. PMID:22415777

  3. Primitive cardiac cells from human embryonic stem cells.

    Science.gov (United States)

    Hudson, James; Titmarsh, Drew; Hidalgo, Alejandro; Wolvetang, Ernst; Cooper-White, Justin

    2012-06-10

    Pluripotent stem cell-derived cardiomyocytes are currently being investigated for in vitro human heart models and as potential therapeutics for heart failure. In this study, we have developed a differentiation protocol that minimizes the need for specific human embryonic stem cell (hESC) line optimization. We first reduced the heterogeneity that exists within the starting population of bulk cultured hESCs by using cells adapted to single-cell passaging in a 2-dimensional (2D) culture format. Compared with bulk cultures, single-cell cultures comprised larger fractions of TG30(hi)/OCT4(hi) cells, corresponding to an increased expression of pluripotency markers OCT4 and NANOG, and reduced expression of early lineage-specific markers. A 2D temporal differentiation protocol was then developed, aimed at reducing the inherent heterogeneity and variability of embryoid body-based protocols, with induction of primitive streak cells using bone morphogenetic protein 4 and activin A, followed by cardiogenesis via inhibition of Wnt signaling using the small molecules IWP-4 or IWR-1. IWP-4 treatment resulted in a large percentage of cells expressing low amounts of cardiac myosin heavy chain and expression of early cardiac progenitor markers ISL1 and NKX2-5, thus indicating the production of large numbers of immature cardiomyocytes (~65,000/cm(2) or ~1.5 per input hESC). This protocol was shown to be effective in HES3, H9, and, to a lesser, extent, MEL1 hESC lines. In addition, we observed that IWR-1 induced predominantly atrial myosin light chain (MLC2a) expression, whereas IWP-4 induced expression of both atrial (MLC2a) and ventricular (MLC2v) forms. The intrinsic flexibility and scalability of this 2D protocol mean that the output population of primitive cardiomyocytes will be particularly accessible and useful for the investigation of molecular mechanisms driving terminal cardiomyocyte differentiation, and potentially for the future treatment of heart failure.

  4. Effects of calcitriol, seocalcitol, and medium-chain triglyceride on a canine transitional cell carcinoma cell line

    DEFF Research Database (Denmark)

    Kaewsakhorn, T.; Kisseberth, W.C.; Capen, C.C.

    2005-01-01

    Background: Transitional cell carcinoma (TCC) in dogs is associated with high morbidity and mortality. Calcitriol and its analog seocalcitol, combined with medium-chain triglyceride (MCT), have potential for the treatment of this disease. Materials and Methods: TCC cells were treated with calcitr......Background: Transitional cell carcinoma (TCC) in dogs is associated with high morbidity and mortality. Calcitriol and its analog seocalcitol, combined with medium-chain triglyceride (MCT), have potential for the treatment of this disease. Materials and Methods: TCC cells were treated...... inhibited TCC cell growth via induction of cell cycle arrest and MCT enhanced this effect. Therefore, calcitriol and seocalcitol with MCT may have therapeutic potential for canine bladder cancer....

  5. Influences of MDM2 on epithelial mesenchymal transition of human breast cancer cells and its molecular mechanism%MDM2对人乳腺癌细胞上皮间质转化的影响及其机制研究

    Institute of Scientific and Technical Information of China (English)

    闫彩云; 仇金荣; 卢建磊; 殷咏梅

    2014-01-01

    目的:探讨鼠双微粒体2( MDM2)在上皮间质转化( EMT )过程中的作用及其分子机制。方法采用Western blotting检测人乳腺癌 MCF-7、MDA-MB-231和 MDA-MB-435细胞株中 MDM2及 EMT相关标记分子( E-cadherin、N-cadherin及Vimentin)水平;分别于MCF-7细胞中瞬时转染MDM2空质粒pcmv、MDM2过表达质粒pcmv-MDM2,MDA-MB-231细胞中瞬时转染针对MDM2三个不同靶点的干扰质粒36h后观察细胞形态,并采用Western blotting和免疫荧光检测EMT相关标记分子及MDM2的表达情况;采用Western blotting及实时定量PCR检测在MCF-7、MDA-MB-231细胞过表达MDM2后对促EMT转录因子Snail1、Twist 蛋白和mRNA水平的影响。结果 MCF-7细胞过表达MDM2后形态由鹅卵石形变为纺锤形, E-cadherin蛋白随MDM2表达水平的上调表达降低;MDA-MB-231细胞敲低MDM2后形态由长梭形变为卵圆形,Vimentin、N-cadherin蛋白随MDM2表达水平的下调表达依次降低;MCF-7及MDA-MB-231细胞过表达MDM2后Snail1、Twist的蛋白及mRNA水平均升高。结论在乳腺癌细胞株中MDM2可促进 EMT发生,其可能通过上调转录因子 Snail1和 Twist表达来实现。%Objective To explore the influence of murine double minute 2 ( MDM2) on epithelial mesenchymal transition ( EMT) of human breast cancer cells and its molecular mechanism. Methods Western blotting method was performed to examine the protein expressions of MDM2 and EMT-related proteins ( E-cadherin, N-cadherin and Vimentin) in human breast cancer cell lines MCF-7, MDA-MB-231 and MDA-MB-435. The optical microscope was used to observe the morphological changes at 36th hour after transiently transfection of MCF-7 cells with empty plasmid ( pcmv) or MDM2 overexpression plasmid ( pcmv-MDM2) and MDA-MB-231 cells with three different interference plasmids. Meanwhile, the expressions of MDM2 and EMT related molecular markers were measured by Western blotting and immunofluorescence assays

  6. Proteomic analysis of proton beam irradiated human melanoma cells.

    Directory of Open Access Journals (Sweden)

    Sylwia Kedracka-Krok

    Full Text Available Proton beam irradiation is a form of advanced radiotherapy providing superior distributions of a low LET radiation dose relative to that of photon therapy for the treatment of cancer. Even though this clinical treatment has been developing for several decades, the proton radiobiology critical to the optimization of proton radiotherapy is far from being understood. Proteomic changes were analyzed in human melanoma cells treated with a sublethal dose (3 Gy of proton beam irradiation. The results were compared with untreated cells. Two-dimensional electrophoresis was performed with mass spectrometry to identify the proteins. At the dose of 3 Gy a minimal slowdown in proliferation rate was seen, as well as some DNA damage. After allowing time for damage repair, the proteomic analysis was performed. In total 17 protein levels were found to significantly (more than 1.5 times change: 4 downregulated and 13 upregulated. Functionally, they represent four categories: (i DNA repair and RNA regulation (VCP, MVP, STRAP, FAB-2, Lamine A/C, GAPDH, (ii cell survival and stress response (STRAP, MCM7, Annexin 7, MVP, Caprin-1, PDCD6, VCP, HSP70, (iii cell metabolism (TIM, GAPDH, VCP, and (iv cytoskeleton and motility (Moesin, Actinin 4, FAB-2, Vimentin, Annexin 7, Lamine A/C, Lamine B. A substantial decrease (2.3 x was seen in the level of vimentin, a marker of epithelial to mesenchymal transition and the metastatic properties of melanoma.

  7. Appearance of Human Plasma Cells Following Differentiation of Human B Cells in NOD/SCID Mouse Spleen

    OpenAIRE

    2003-01-01

    Relatively little is known for the differentiation and maturation process of human B cells to plasma cells. This is particularly important in reconstitution work involving transfer of autoantibodies. To address this issue, we transplanted human peripheral blood mononuclear cells (PBMC) directly into the spleen of irradiated NOD/SCID mice depleted of natural killer cell activity. Within 6 weeks, naïve B cells differentiated into memory B cells and, importantly, the numbers of human CD138+ plas...

  8. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins

    Directory of Open Access Journals (Sweden)

    Hayato Fukusumi

    2016-01-01

    Full Text Available Human neural progenitor cells (hNPCs have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi. Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes.

  9. Human microglial cells synthesize albumin in brain.

    Directory of Open Access Journals (Sweden)

    Sung-Min Ahn

    Full Text Available Albumin, an abundant plasma protein with multifunctional properties, is mainly synthesized in the liver. Albumin has been implicated in Alzheimer's disease (AD since it can bind to and transport amyloid beta (Abeta, the causative agent of AD; albumin is also a potent inhibitor of Abeta polymerization. Despite evidence of non-hepatic transcription of albumin in many tissues including kidney and pancreas, non-hepatic synthesis of albumin at the protein level has been rarely confirmed. In a pilot phase study of Human Brain Proteome Project, we found evidence that microglial cells in brain may synthesize albumin. Here we report, for the first time, the de novo synthesis of albumin in human microglial cells in brain. Furthermore, we demonstrate that the synthesis and secretion of albumin from microglial cells is enhanced upon microglial activation by Abeta(1-42- or lipopolysaccharide (LPS-treatment. These data indicate that microglial cells may play a beneficial role in AD by secreting albumin that not only inhibits Abeta polymerization but also increases its clearance.

  10. STIM1 accelerates cell senescence in a remodeled microenvironment but enhances the epithelial-to-mesenchymal transition in prostate cancer

    Science.gov (United States)

    Xu, Yingxi; Zhang, Shu; Niu, Haiying; Ye, Yujie; Hu, Fen; Chen, Si; Li, Xuefei; Luo, Xiaohe; Jiang, Shan; Liu, Yanhua; Chen, Yanan; Li, Junying; Xiang, Rong; Li, Na

    2015-01-01

    The importance of store-operated Ca2+ entry (SOCE) and the role of its key molecular regulators, STIM1 and ORAI1, in the development of cancer are emerging. Here, we report an unexpected dual function of SOCE in prostate cancer progression by revealing a decrease in the expression of STIM1 in human hyperplasia and tumor tissues of high histological grade and by demonstrating that STIM1 and ORAI1 inhibit cell growth by arresting the G0/G1 phase and enhancing cell senescence in human prostate cancer cells. In addition, STIM1 and ORAI1 inhibited NF-κB signaling and remodeled the tumor microenvironment by reducing the formation of M2 phenotype macrophages, possibly creating an unfavorable tumor microenvironment and inhibiting cancer development. However, STIM1 also promoted cell migration and the epithelial-to-mesenchymal transition by activating TGF-β, Snail and Wnt/β-Catenin pathways. Thus, our study revealed novel regulatory effects and the mechanisms by which STIM1 affects cell senescence, tumor migration and the tumor microenvironment, revealing that STIM1 has multiple functions in prostate cancer cells. PMID:26257076

  11. Neocortical glial cell numbers in human brains

    DEFF Research Database (Denmark)

    Pelvig, D.P.; Pakkenberg, H.; Stark, A.K.

    2008-01-01

    and neurons and counting were done in each of the four lobes. The study showed that the different subpopulations of glial cells behave differently as a function of age; the number of oligodendrocytes showed a significant 27% decrease over adult life and a strong correlation to the total number of neurons...... while the total astrocyte number is constant through life; finally males have a 28% higher number of neocortical glial cells and a 19% higher neocortical neuron number than females. The overall total number of neocortical neurons and glial cells was 49.3 billion in females and 65.2 billion in males......, a difference of 24% with a high biological variance. These numbers can serve as reference values in quantitative studies of the human neocortex. (C) 2007 Elsevier Inc. All rights reserved Udgivelsesdato: 2008/11...

  12. Characterizing motility dynamics in human RPE cells

    Science.gov (United States)

    Liu, Zhuolin; Kurokawa, Kazuhiro; Zhang, Furu; Miller, Donald T.

    2017-02-01

    Retinal pigment epithelium (RPE) cells are vital to health of the outer retina, however, are often compromised in ageing and ocular diseases that lead to blindness. Early manifestation of RPE disruption occurs at the cellular level, but while in vivo biomarkers at this scale hold considerable promise, RPE cells have proven extremely challenging to image in the living human eye. Recently we addressed this problem by using organelle motility as a novel contrast agent to enhance the RPE cell in conjunction with 3D resolution of adaptive optics-optical coherence tomography (AO-OCT) to section the RPE layer. In this study, we expand on the central novelty of our method - organelle motility - by characterizing the dynamics of the motility in individual RPE cells, important because of its direct link to RPE physiology. To do this, AO-OCT videos of the same retinal patch were acquired at approximately 1 min intervals or less, time stamped, and registered in 3D with sub-cellular accuracy. Motility was quantified by an exponential decay time constant, the time for motility to decorrelate the speckle field across an RPE cell. In two normal subjects, we found the decay time constant to be just 3 seconds, thus indicating rapid motility in normal RPE cells.

  13. Human cell culture in a space bioreactor

    Science.gov (United States)

    Morrison, Dennis R.

    1988-01-01

    Microgravity offers new ways of handling fluids, gases, and growing mammalian cells in efficient suspension cultures. In 1976 bioreactor engineers designed a system using a cylindrical reactor vessel in which the cells and medium are slowly mixed. The reaction chamber is interchangeable and can be used for several types of cell cultures. NASA has methodically developed unique suspension type cell and recovery apparatus culture systems for bioprocess technology experiments and production of biological products in microgravity. The first Space Bioreactor was designed for microprocessor control, no gaseous headspace, circulation and resupply of culture medium, and slow mixing in very low shear regimes. Various ground based bioreactors are being used to test reactor vessel design, on-line sensors, effects of shear, nutrient supply, and waste removal from continuous culture of human cells attached to microcarriers. The small Bioreactor is being constructed for flight experiments in the Shuttle Middeck to verify systems operation under microgravity conditions and to measure the efficiencies of mass transport, gas transfer, oxygen consumption and control of low shear stress on cells.

  14. Redifferentiation of adult human β cells expanded in vitro by inhibition of the WNT pathway.

    Directory of Open Access Journals (Sweden)

    Ayelet Lenz

    Full Text Available In vitro expansion of adult human islet β cells is an attractive solution for the shortage of tissue for cell replacement therapy of type 1 diabetes. Using a lineage tracing approach we have demonstrated that β-cell-derived (BCD cells rapidly dedifferentiate in culture and can proliferate for up to 16 population doublings. Dedifferentiation is associated with changes resembling epithelial-mesenchymal transition (EMT. The WNT pathway has been shown to induce EMT and plays key roles in regulating replication and differentiation in many cell types. Here we show that BCD cell dedifferentiation is associated with β-catenin translocation into the nucleus and activation of the WNT pathway. Inhibition of β-catenin expression in expanded BCD cells using short hairpin RNA resulted in growth arrest, mesenchymal-epithelial transition, and redifferentiation, as judged by activation of β-cell gene expression. Furthermore, inhibition of β-catenin expression synergized with redifferentiation induced by a combination of soluble factors, as judged by an increase in the number of C-peptide-positive cells. Simultaneous inhibition of the WNT and NOTCH pathways also resulted in a synergistic effect on redifferentiation. These findings, which were reproducible in cells derived from multiple human donors, suggest that inhibition of the WNT pathway may contribute to a therapeutically applicable way for generation of functional insulin-producing cells following ex-vivo expansion.

  15. Noise-induced phase transition in the model of human virtual stick balancing

    CERN Document Server

    Zgonnikov, Arkady

    2016-01-01

    Humans face the task of balancing dynamic systems near an unstable equilibrium repeatedly throughout their lives. Much research has been aimed at understanding the mechanisms of intermittent control in the context of human balance control. The present paper deals with one of the recent developments in the theory of human intermittent control, namely, the double-well model of noise-driven control activation. We demonstrate that the double-well model can reproduce the whole range of experimentally observed distributions under different conditions. Moreover, we show that a slight change in the noise intensity parameter leads to a sudden shift of the action point distribution shape, that is, a phase transition is observed.

  16. Tumor Budding Cells, Cancer Stem Cells and Epithelial-Mesenchymal Transition-type Cells in Pancreatic Cancer

    Directory of Open Access Journals (Sweden)

    Eva eKaramitopoulou

    2013-01-01

    Full Text Available Pancreatic ductal adenocarcinoma (PDAC is one of the most lethal cancers with a 5-year survival rate of less than 5%. Moreover, PDAC escapes early detection and resists treatment. Multiple combinations of genetic alterations are known to occur in PDAC including mutational activation of KRAS, inactivation of p16/CDKN2A and SMAD4 (DPC4 and dysregulation of PTEN/PI3K/AKT signaling. Through their interaction with WNT pathway, the downstream molecules of these pathways have been implicated in the promotion of epithelial-mesenchymal transition (EMT. Emerging evidence has demonstrated that cancer stem cells (CSCs, small populations of which have been identified in PDAC, and EMT-type cells play critical roles in drug resistance, invasion and metastasis in pancreatic cancer. EMT may be histologically represented by the presence of tumor budding which is described as the occurrence of single tumor cells or small clusters (<5 of dedifferentiated cells at the invasive front of gastrointestinal (including colorectal, oesophageal, gastric and ampullary carcinomas and is linked to poor prognosis. Tumor budding has recently been shown to occur frequently in PDAC and to be associated with adverse clinicopathological features and decreased disease-free and overall survival. The aim of this review is to present a short overview on the morphological and molecular aspects that underline the relationship between tumor budding cells, CSCs and EMT-type cells in PDAC.

  17. Canine Mammary Cancer Stem Cells are Radio- and Chemo- Resistant and Exhibit an Epithelial-Mesenchymal Transition Phenotype

    Directory of Open Access Journals (Sweden)

    David J. Argyle

    2011-03-01

    Full Text Available Canine mammary carcinoma is the most common cancer among female dogs and is often fatal due to the development of distant metastases. In humans, solid tumors are made up of heterogeneous cell populations, which perform different roles in the tumor economy. A small subset of tumor cells can hold or acquire stem cell characteristics, enabling them to drive tumor growth, recurrence and metastasis. In veterinary medicine, the molecular drivers of canine mammary carcinoma are as yet undefined. Here we report that putative cancer stem cells (CSCs can be isolated form a canine mammary carcinoma cell line, REM134. We show that these cells have an increased ability to form tumorspheres, a characteristic of stem cells, and that they express embryonic stem cell markers associated with pluripotency. Moreover, canine CSCs are relatively resistant to the cytotoxic effects of common chemotherapeutic drugs and ionizing radiation, indicating that failure of clinical therapy to eradicate canine mammary cancer may be due to the survival of CSCs. The epithelial to mesenchymal transition (EMT has been associated with cancer invasion, metastasis, and the acquisition of stem cell characteristics. Our results show that canine CSCs predominantly express mesenchymal markers and are more invasive than parental cells, indicating that these cells have a mesenchymal phenotype. Furthermore, we show that canine mammary cancer cells can be induced to undergo EMT by TGFβ and that these cells have an increased ability to form tumorspheres. Our findings indicate that EMT induction can enrich for cells with CSC properties, and provide further insight into canine CSC biology.

  18. Appearance of Human Plasma Cells Following Differentiation of Human B Cells in NOD/SCID Mouse Spleen

    Directory of Open Access Journals (Sweden)

    Kentaro Kikuchi

    2003-01-01

    Full Text Available Relatively little is known for the differentiation and maturation process of human B cells to plasma cells. This is particularly important in reconstitution work involving transfer of autoantibodies. To address this issue, we transplanted human peripheral blood mononuclear cells (PBMC directly into the spleen of irradiated NOD/SCID mice depleted of natural killer cell activity. Within 6 weeks, naïve B cells differentiated into memory B cells and, importantly, the numbers of human CD138+ plasma cells in spleen increased by 100 fold after transplantation. Plasma cell numbers correlated with the detection of human IgM and IgG in serum, indicating that human B cells had differentiated into mature plasma cells in the murine spleen. In addition to CD19+ plasma cells, a distinct CD19- plasma cell population was detected, suggesting that downregulation of CD19 associated with maturation of plasma cells occurred. When purified human B cells were transplanted, those findings were not observed. Our results indicate that differentiation and maturation of human B cells and plasma cells can be investigated by transplantation of human PBMC into the spleen of NOD/SCID mice. The model will be useful for studying the differentiation of human B cells and generation of plasma cells.

  19. Staphylococcus aureus Lpl Lipoproteins Delay G2/M Phase Transition in HeLa Cells.

    Science.gov (United States)

    Nguyen, Minh-Thu; Deplanche, Martine; Nega, Mulugeta; Le Loir, Yves; Peisl, Loulou; Götz, Friedrich; Berkova, Nadia

    2016-01-01

    The cell cycle is an ordered set of events, leading to cell growth and division into two daughter cells. The eukaryotic cell cycle consists of interphase (G1, S, and G2 phases), followed by the mitotic phase and G0 phase. Many bacterial pathogens secrete cyclomodulins that interfere with the host cell cycle. In Staphylococcus aureus four cyclomodulins have been described so far that all represent toxins and are secreted into the culture supernatant. Here we show that the membrane-anchored lipoprotein-like proteins (Lpl), encoded on a genomic island called νSaα, interact with the cell cycle of HeLa cells. By comparing wild type and lpl deletion mutant it turned out that the lpl cluster is causative for the G2/M phase transition delay and also contributes to increased invasion frequency. The lipoprotein Lpl1, a representative of the lpl cluster, also caused G2/M phase transition delay. Interestingly, the lipid modification, which is essential for TLR2 signaling and activation of the immune system, is not necessary for cyclomodulin activity. Unlike the other staphylococcal cyclomodulins Lpl1 shows no cytotoxicity even at high concentrations. As all Lpl proteins are highly conserved there might be a common function that is accentuated by their multiplicity in a tandem gene cluster. The cell surface localized Lpls' suggests a correlation between G2/M phase transition delay and host cell invasion.

  20. Staphylococcus aureus Lpl Lipoproteins Delay G2/M Phase Transition in HeLa Cells

    Science.gov (United States)

    Nguyen, Minh-Thu; Deplanche, Martine; Nega, Mulugeta; Le Loir, Yves; Peisl, Loulou; Götz, Friedrich; Berkova, Nadia

    2016-01-01

    The cell cycle is an ordered set of events, leading to cell growth and division into two daughter cells. The eukaryotic cell cycle consists of interphase (G1, S, and G2 phases), followed by the mitotic phase and G0 phase. Many bacterial pathogens secrete cyclomodulins that interfere with the host cell cycle. In Staphylococcus aureus four cyclomodulins have been described so far that all represent toxins and are secreted into the culture supernatant. Here we show that the membrane-anchored lipoprotein-like proteins (Lpl), encoded on a genomic island called νSaα, interact with the cell cycle of HeLa cells. By comparing wild type and lpl deletion mutant it turned out that the lpl cluster is causative for the G2/M phase transition delay and also contributes to increased invasion frequency. The lipoprotein Lpl1, a representative of the lpl cluster, also caused G2/M phase transition delay. Interestingly, the lipid modification, which is essential for TLR2 signaling and activation of the immune system, is not necessary for cyclomodulin activity. Unlike the other staphylococcal cyclomodulins Lpl1 shows no cytotoxicity even at high concentrations. As all Lpl proteins are highly conserved there might be a common function that is accentuated by their multiplicity in a tandem gene cluster. The cell surface localized Lpls' suggests a correlation between G2/M phase transition delay and host cell invasion. PMID:28083519

  1. Primary retroperitoneal transitional cell carcinoma presenting as a dumb-bell tumour.

    Science.gov (United States)

    Basu, S; Ansari, M; Gupta, S; Kumar, A

    2009-11-01

    We report a retroperitoneal transitional cell carcinoma arising from the primitive urogenital remnants of a 56-year-old married Indian woman. She presented with a huge cystic mass in the hypogastrium and right iliac fossa, which extended into the right thigh as a massive dumb-bell tumour. On exploration, it was found not to be arising from any known retroperitoneal structure. The mass was excised, and the histopathology confirmed transitional cell carcinoma with positive margins. Though she received postoperative chemotherapy with cyclophosphamide, adriamycin and cisplatin, she developed extensive local recurrence and hepatic secondaries, and succumbed to the disease after ten months of follow-up. We highlight the rarity of the disease, its atypical presentation as a cystic dumb-bell lump, its diagnostic challenges and aggressive behaviour, and review the literature on primary retroperitoneal transitional cell carcinomas.

  2. An unusual Case of Transitional Cell Carcinoma of Renal Pelvis Presenting with Brain Metastases

    Directory of Open Access Journals (Sweden)

    MR Razzaghi

    2009-04-01

    Full Text Available ABSTRACT: Introduction & Objective: Transitional cell carcinoma of renal pelvis presenting with brain metastases is a very rare case which should be diagnosed and treated in order to prevent further damages. Case: We report a rare case, who had presented with a constellation of neurological symptoms (due to multiple brain metastases, but without any urological symptoms. During evaluation of patient, we found transitional cell carcinoma (TCC of left renal pelvis, for which palliative radical nephroureterectomy was performed . Conclusion: Although transitional cell carcinoma of renal pelvis presenting with brain metastases is a very rare case, but the patient was managed with gamma knife stereotactic radiosurgery for the metastatic lesions. Afterward he received four cycles of paclitaxel and carboplatin chemotherapy. The patient is alive with stable disease at 32- months’ follow-up.

  3. Direct detection and quantification of transition metal ions in human atherosclerotic plaques

    DEFF Research Database (Denmark)