Sample records for cells enhance stem-like

  1. Astrocytes enhance the invasion potential of glioblastoma stem-like cells.

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    Barbara H Rath

    Full Text Available Glioblastomas (GBMs are characterized as highly invasive; the contribution of GBM stem-like cells (GSCs to the invasive phenotype, however, has not been completely defined. Towards this end, we have defined the invasion potential of CD133+ GSCs and their differentiated CD133- counterparts grown under standard in vitro conditions and in co-culture with astrocytes. Using a trans-well assay, astrocytes or astrocyte conditioned media in the bottom chamber significantly increased the invasion of GSCs yet had no effect on CD133- cells. In addition, a monolayer invasion assay showed that the GSCs invaded farther into an astrocyte monolayer than their differentiated progeny. Gene expression profiles were generated from two GSC lines grown in trans-well culture with astrocytes in the bottom chamber or directly in contact with astrocyte monolayers. In each co-culture model, genes whose expression was commonly increased in both GSC lines involved cell movement and included a number of genes that have been previously associated with tumor cell invasion. Similar gene expression modifications were not detected in CD133- cells co-cultured under the same conditions with astrocytes. Finally, evaluation of the secretome of astrocytes grown in monolayer identified a number of chemokines and cytokines associated with tumor cell invasion. These data suggest that astrocytes enhance the invasion of CD133+ GSCs and provide additional support for a critical role of brain microenvironment in the regulation of GBM biology.

  2. Interferon-α/β enhances temozolomide activity against MGMT-positive glioma stem-like cells. (United States)

    Shen, Dong; Guo, Cheng-Cheng; Wang, Jing; Qiu, Zhi-Kun; Sai, Ke; Yang, Qun-Ying; Chen, Yin-Sheng; Chen, Fu-Rong; Wang, Jie; Panasci, Lawrence; Chen, Zhong-Ping


    Glioma is one of the most common primary tumors of the central nervous system in adults. Glioblastoma (GBM) is the most lethal type of glioma, whose 5-year survival is 9.8% at best. Glioma stem-like cells (GSCs) play an important role in recurrence and treatment resistance. MGMT is a DNA repair protein that removes DNA adducts and therefore attenuates treatment efficiency. It has been reported that interferon-α/β (IFN-α/β) downregulates the level of MGMT and sensitizes glioma cells to temozolomide. In the present study, we assessed whether IFN-α/β is able to sensitize GSCs to temozolomide by modulating MGMT expression. Upon the treatment of IFN-α/β, the efficacy of temozolomide against MGMT‑positive GSCs was markedly enhanced by combination treatment with IFN-α/β when compared with the temozolomide single agent group, and MGMT expression was markedly decreased at the same time. Further mechanistic study showed that IFN-α/β suppressed the NF-κB activity, which further mediated the sensitization of MGMT‑positive GSCs to temozolomide. Our data therefore demonstrated that the application of IFN-α/β is a promising agent with which to enhance temozolomide efficiency and reduce drug resistance, and our findings shed light on improving clinical outcomes and prolonging the survival of patients with malignant gliomas.

  3. Enhanced MGMT expression contributes to temozolomide resistance in glioma stem-like cells

    Institute of Scientific and Technical Information of China (English)

    Zhi-Kun Qiu; Dong Shen; Yin-Sheng Chen; Qun-Ying Yang; Cheng-Cheng Guo; Bing-Hong Feng; Zhong-Ping Chen


    O6-methylguanine DNA methyltransferase (MGMT) can remove DNA alkylation adducts, thereby repairing damaged DNA and contributing to the drug resistance of gliomas to alkylating agents. In addition, glioma stem-like cells (GSCs) have been demonstrated to be involved in the recurrence and treatment resistance of gliomas. In this study, we aimed to investigate MGMT expression and regulatory mechanisms in GSCs and the association of MGMT with temozolomide (TMZ) sensitivity. GSCs were enriched from one MGMT-positive cellline (SF-767) and 7 MGMT-negative celllines (U251, SKMG-4, SKMG-1, SF295, U87, MGR1, and MGR2) through serum-free clone culture. GSCs from the U251G, SKMG-4G, SF295G, and SKMG-1G cell lines became MGMT-positive, but those from the U87G, MGR1G, and MGR2G cell lines remained MGMT-negative. However, al the GSCs and their parental glioma celllines were positive for nuclear factor-κB (NF-κB). In addition, GSCs were more resistant to TMZ than their parental glioma cell lines (P 0.05). When we treated the MGMT-positive GSCs with TMZ plus MG-132 (an NF-κB inhibitor), the antitumor activity was significantly enhanced compared to that of GSCs treated with TMZ alone (P < 0.05). Furthermore, we found that MGMT expression decreased through the down-regulation of NF-κB expression by MG-132. Our results show that MG-132 may inhibit NF-κB expression and further decrease MGMT expression, resulting in a synergistic effect on MGMT-positive GSCs. These results indicate that enhanced MGMT expression contributes to TMZ resistance in MGMT-positive GSCs.

  4. The Brain Microenvironment Preferentially Enhances the Radioresistance of CD133+ Glioblastoma Stem-like Cells

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    Muhammad Jamal


    Full Text Available Brain tumor xenografts initiated from glioblastoma (GBM CD133+ tumor stem-like cells (TSCs are composed of TSC and non-TSC subpopulations, simulating the phenotypic heterogeneity of GBMs in situ. Given that the discrepancies between the radiosensitivity of GBM cells in vitro and the treatment response of patients suggest a role for the microenvironment in GBM radioresistance, we compared the response of TSCs and non-TSCs irradiated under in vitro and orthotopic conditions. As a measure of radioresponse determined at the individual cell level, γH2AX and 53BP1 foci were quantified in CD133+ cells and their differentiated (CD133- progeny. Under in vitro conditions, no difference was detected between CD133+ and CD133- cells in foci induction or dispersal after irradiation. However, irradiation of orthotopic xenografts initiated from TSCs resulted in the induction of fewer γH2AX and 53BP1 foci in CD133+ cells compared to their CD133- counterparts within the same tumor. Xenograft irradiation resulted in a tumor growth delay of approximately 7 days with a corresponding increase in the percentage of CD133+ cells at 7 days after radiation, which persisted to the onset of neurologic symptoms. These results suggest that, although the radioresponse of TSCs and non-TSCs does not differ under in vitro growth conditions, CD133+ cells are relatively radioresistant under intracerebral growth conditions. Whereas these findings are consistent with the suspected role for TSCs as a determinant of GBM radioresistance, these data also illustrate the dependence of the cellular radioresistance on the brain microenvironment.

  5. Chronic exposure to combined carcinogens enhances breast cell carcinogenesis with mesenchymal and stem-like cell properties.

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    Lenora Ann Pluchino

    Full Text Available Breast cancer is the most common type of cancer affecting women in North America and Europe. More than 85% of breast cancers are sporadic and attributable to long-term exposure to small quantities of multiple carcinogens. To understand how multiple carcinogens act together to induce cellular carcinogenesis, we studied the activity of environmental carcinogens 4-(methylnitrosamino-1-(3-pyridyl-1-butanone (NNK and benzo[a]pyrene (B[a]P, and dietary carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP using our breast cell carcinogenesis model. Our study revealed, for the first time, that combined NNK and B[a]P enhanced breast cell carcinogenesis chronically induced by PhIP in both non-cancerous and cancerous breast cells. Co-exposure was more potent than sequential exposure to combined NNK and B[a]P followed by PhIP in inducing carcinogenesis. Initiation of carcinogenesis was measured by transient endpoints induced in a single exposure, while progression of carcinogenesis was measured by acquisition of constitutive endpoints in cumulative exposures. Transient endpoints included DNA damage, Ras-Erk-Nox pathway activation, reactive oxygen species elevation, and increased cellular proliferation. Constitutive endpoints included various cancer-associated properties and signaling modulators, as well as enrichment of cancer stem-like cell population and activation of the epithelial-to-mesenchymal transition program. Using transient and constitutive endpoints as targets, we detected that a combination of the green tea catechins ECG and EGCG, at non-cytotoxic levels, was more effective than individual agents in intervention of cellular carcinogenesis induced by combined NNK, B[a]P, and PhIP. Thus, use of combined ECG and EGCG should be seriously considered for early intervention of breast cell carcinogenesis associated with long-term exposure to environmental and dietary carcinogens.

  6. Activation of Wnt signaling pathway by AF1q enriches stem-like population and enhance mammosphere formation of breast cells. (United States)

    Tse, Charlotte Olivia; Kim, Soojin; Park, Jino


    Wnt signaling pathway is believed to be responsible for control over various types of stem cells and may act as a niche factor to maintain stem cells in a self-renewing state. Moreover, dysregulated Wnt signaling pathway is strongly associated with several diseases including cancer. Previously, we have shown that AF1q associates with a poor prognosis in leukemia, myelodysplastic syndromes, multiple myeloid, ovarian cancer, and breast cancer. Also, AF1q plays a pivotal role as an oncogene and metastasis enhancer in breast cancer via activation of Wnt signaling pathway. AF1q is highly expressed in stem cells, and this expression is diminished by differentiation. To understand the role of AF1q in stem-like population, we examined stem-like cells derived from breast cells which dysregulated Wnt signaling pathway by alteration of AF1q expression. The effect of Wnt signaling pathway by AF1q on EMT marker expression, stem cell marker expression, and sphere formation was determined. Activated Wnt signaling pathway by AF1q enriched stem-like population showed enhanced sphere formation ability. Interestingly, Wnt signaling pathway inhibitor, Quercetin, decreased the sphere formation in these cells. These results suggest that AF1q would have a role as an enhancer in generation of stem-like population through activation of Wnt signaling pathway.

  7. Adult stem-like cells in kidney

    Institute of Scientific and Technical Information of China (English)

    Keiichi Hishikawa; Osamu Takase; Masahiro Yoshikawa; Taro Tsujimura; Masaomi Nangaku; Tsuyoshi Takato


    Human pluripotent cells are promising for treatmentfor kidney diseases, but the protocols for derivationof kidney cell types are still controversial. Kidneytissue regeneration is well confirmed in several lowervertebrates such as fish, and the repair of nephronsafter tubular damages is commonly observed after renalinjury. Even in adult mammal kidney, renal progenitorcell or system is reportedly presents suggesting thatadult stem-like cells in kidney can be practical clinicaltargets for kidney diseases. However, it is still unclearif kidney stem cells or stem-like cells exist or not. Ingeneral, stemness is defined by several factors suchas self-renewal capacity, multi-lineage potency andcharacteristic gene expression profiles. The definiteuse of stemness may be obstacle to understand kidneyregeneration, and here we describe the recent broadfindings of kidney regeneration and the cells thatcontribute regeneration.

  8. Preferential Iron Trafficking Characterizes Glioblastoma Stem-like Cells

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    Schonberg, David L; Miller, Tyler E; Wu, Qiulian


    Glioblastomas display hierarchies with self-renewing cancer stem-like cells (CSCs). RNA sequencing and enhancer mapping revealed regulatory programs unique to CSCs causing upregulation of the iron transporter transferrin, the top differentially expressed gene compared with tissue-specific progeni...

  9. Target irradiation induced bystander effects between stem-like and non stem-like cancer cells

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    Liu, Yu [State Key Laboratory of Nuclear Physics and Technology, School of Physics, Peking University, Beijing 100871 (China); Space Radiation Research Unit, International Open Laboratory, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Kobayashi, Alisa [Space Radiation Research Unit, International Open Laboratory, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Department of Technical Support and Development, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Maeda, Takeshi [Department of Technical Support and Development, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Fu, Qibin [State Key Laboratory of Nuclear Physics and Technology, School of Physics, Peking University, Beijing 100871 (China); Oikawa, Masakazu [Department of Technical Support and Development, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Yang, Gen, E-mail: [State Key Laboratory of Nuclear Physics and Technology, School of Physics, Peking University, Beijing 100871 (China); Space Radiation Research Unit, International Open Laboratory, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Konishi, Teruaki, E-mail: [Space Radiation Research Unit, International Open Laboratory, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Department of Technical Support and Development, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Uchihori, Yukio [Space Radiation Research Unit, International Open Laboratory, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); Department of Technical Support and Development, National Institute of Radiological Sciences, 4-9-1 Inage-ku, Chiba 263-8555 (Japan); and others


    Highlights: • Existence of radiation induced bystander effects (RIBE) between cancer stem-like cells (CSCs) and non stem-like cancer cells (NSCCs) in human fibrosarcoma HT1080 cells. • Existence of significant difference in generation and response of bystander signals between CSCs and NSCCs. • CSCs are significantly less sensitive to NO scavenger than that of NSCCs in terms of DNA double strand breaks induced by RIBE. - Abstract: Tumors are heterogeneous in nature and consist of multiple cell types. Among them, cancer stem-like cells (CSCs) are suggested to be the principal cause of tumor metastasis, resistance and recurrence. Therefore, understanding the behavior of CSCs in direct and indirect irradiations is crucial for clinical radiotherapy. Here, the CSCs and their counterpart non stem-like cancer cells (NSCCs) in human HT1080 fibrosarcoma cell line were sorted and labeled, then the two cell subtypes were mixed together and chosen separately to be irradiated via a proton microbeam. The radiation-induced bystander effect (RIBE) between the CSCs and NSCCs was measured by imaging 53BP1 foci, a widely used indicator for DNA double strand break (DSB). CSCs were found to be less active than NSCCs in both the generation and the response of bystander signals. Moreover, the nitric oxide (NO) scavenger c-PTIO can effectively alleviate the bystander effect in bystander NSCCs but not in bystander CSCs, indicating a difference of the two cell subtypes in NO signal response. To our knowledge, this is the first report shedding light on the RIBE between CSCs and NSCCs, which might contribute to a further understanding of the out-of-field effect in cancer radiotherapy.

  10. HDAC inhibitors enhance the lethality of low dose salinomycin in parental and stem-like GBM cells. (United States)

    Booth, Laurence; Roberts, Jane L; Conley, Adam; Cruickshanks, Nichola; Ridder, Thomas; Grant, Steven; Poklepovic, Andrew; Dent, Paul


    The present studies determined whether the antibiotic salinomycin interacted with HDAC inhibitors to kill primary human GBM cells. Regardless of PTEN, ERBB1, or p53 mutational status salinomycin interacted with HDAC inhibitors in a synergistic fashion to kill GBM cells. Inhibition of CD95/Caspase 8 or of CD95/RIP-1/AIF signaling suppressed killing by the drug combination. Salinomycin increased the levels of autophagosomes that correlated with increased p62 and LC3II levels; valproate co-treatment correlated with reduced LC3II and p62 expression, and increased caspase 3 cleavage. Molecular inhibition of autophagosome formation was protective against drug exposure. The drug combination enhanced eIF2α phosphorylation and decreased expression of MCL-1 and phosphorylation of mTOR and p70 S6K. Activation of p70 S6K or mTOR promoted cell survival in the face of combined drug exposure. Overexpression of BCL-XL or c-FLIP-s was protective. Collectively our data demonstrate that the lethality of low nanomolar concentrations of salinomycin are enhanced by HDAC inhibitors in GBM cells and that increased death receptor signaling together with reduced mitochondrial function are causal in the combinatorial drug necro-apoptotic killing effect.

  11. Celecoxib Suppresses the Phosphorylation of STAT3 Protein and Can Enhance the Radiosensitivity of Medulloblastoma-Derived Cancer Stem-Like Cells

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    Meng-Yin Yang


    Full Text Available Medulloblastoma (MB is a malignant primary brain tumor with poor prognosis. MB-derived CD133/Nestin double-positive cells (MB-DPs exhibit cancer stem-like cell (CSC-like properties that may contribute to chemoradioresistance, tumorigenesis and recurrence. In various tumors, signal transducer and activator of transcription 3 (STAT3 upregulation including MB which can regulate the expression of Nestin. Celecoxib, a selective COX-2 inhibitor, has been shown to potentially reduce STAT3 phosphorylation. The aim of the present study was to investigate the role of celecoxib in enhancing the effects of ionizing radiotherapy (IR on MB-DP. MB-DPs and MB-derived CD133/Nestin double-negative cells (MB-DNs were isolated from medulloblastoma cell line Daoy. Then, both of them were treated with celecoxib in different concentrations, and cell viability was assessed. The assays of cell survival, sphere formation, radiosensitivity, colony formation, apoptotic activity and mouse xenografting experiments in MB-DPs and MB-DNs treated with celecoxib alone, radiation alone, or celecoxib combined with radiation were further evaluated. We isolated MB-DPs from MB cell line Daoy, which exhibited typical CSC-like characteristics. Microarray analysis and Western blotting both indicated the upregulation of Janus kinase (JAK-STAT cascade and STAT3 phosphorylation. Incubation with celecoxib dose-dependently suppressed the CSC-like properties and enhanced the IR effect on the induction of apoptosis, as detected by TUNEL assay and staining for Caspase 3 and Annexin V. Finally, celecoxib also enhanced the IR effect to suppress tumorigenesis and synergistically improve the recipient survival in orthotopic MB-derived CD133/Nestin double-positive cells (MB-DP cells bearing mice.

  12. Target irradiation induced bystander effects between stem-like and non stem-like cancer cells. (United States)

    Liu, Yu; Kobayashi, Alisa; Maeda, Takeshi; Fu, Qibin; Oikawa, Masakazu; Yang, Gen; Konishi, Teruaki; Uchihori, Yukio; Hei, Tom K; Wang, Yugang


    Tumors are heterogeneous in nature and consist of multiple cell types. Among them, cancer stem-like cells (CSCs) are suggested to be the principal cause of tumor metastasis, resistance and recurrence. Therefore, understanding the behavior of CSCs in direct and indirect irradiations is crucial for clinical radiotherapy. Here, the CSCs and their counterpart non stem-like cancer cells (NSCCs) in human HT1080 fibrosarcoma cell line were sorted and labeled, then the two cell subtypes were mixed together and chosen separately to be irradiated via a proton microbeam. The radiation-induced bystander effect (RIBE) between the CSCs and NSCCs was measured by imaging 53BP1 foci, a widely used indicator for DNA double strand break (DSB). CSCs were found to be less active than NSCCs in both the generation and the response of bystander signals. Moreover, the nitric oxide (NO) scavenger c-PTIO can effectively alleviate the bystander effect in bystander NSCCs but not in bystander CSCs, indicating a difference of the two cell subtypes in NO signal response. To our knowledge, this is the first report shedding light on the RIBE between CSCs and NSCCs, which might contribute to a further understanding of the out-of-field effect in cancer radiotherapy. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Enhancement of cancer stem-like and epithelial−mesenchymal transdifferentiation property in oral epithelial cells with long-term nicotine exposure: Reversal by targeting SNAIL

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    Yu, Cheng-Chia [Institute of Oral Science, Chung Shan Medical University, Taichung, Taiwan (China); School of Dentistry, Chung Shan Medical University, Taichung, Taiwan (China); Department of Dentistry, Chung Shan Medical University Hospital, Taichung, Taiwan (China); Chang, Yu-Chao, E-mail: [School of Dentistry, Chung Shan Medical University, Taichung, Taiwan (China); Department of Dentistry, Chung Shan Medical University Hospital, Taichung, Taiwan (China)


    Cigarette smoking is one of the major risk factors in the development and further progression of tumorigenesis, including oral squamous cell carcinoma (OSCC). Recent studies suggest that interplay cancer stem-like cells (CSCs) and epithelial−mesenchymal transdifferentiation (EMT) properties are responsible for the tumor maintenance and metastasis in OSCC. The aim of the present study was to investigate the effects of long-term exposure with nicotine, a major component in cigarette, on CSCs and EMT characteristics. The possible reversal regulators were further explored in nicotine-induced CSCs and EMT properties in human oral epithelial (OE) cells. Long-term exposure with nicotine was demonstrated to up-regulate ALDH1 population in normal gingival and primary OSCC OE cells dose-dependently. Moreover, long-term nicotine treatment was found to enhance the self-renewal sphere-forming ability and stemness gene signatures expression and EMT regulators in OE cells. The migration/cell invasiveness/anchorage independent growth and in vivo tumor growth by nude mice xenotransplantation assay was enhanced in long-term nicotine-stimulated OE cells. Knockdown of Snail in long-term nicotine-treated OE cells was found to reduce their CSCs properties. Therapeutic delivery of Si-Snail significantly blocked the xenograft tumorigenesis of long-term nicotine-treated OSCC cells and largely significantly improved the recipient survival. The present study demonstrated that the enrichment of CSCs coupled EMT property in oral epithelial cells induced by nicotine is critical for the development of OSCC tumorigenesis. Targeting Snail might offer a new strategy for the treatment of OSCC patients with smoking habit. -- Highlights: ► Sustained nicotine treatment induced CSCs properties of oral epithelial cells. ► Long-term nicotine treatment enhance EMT properties of oral epithelial cells. ► Long-term nicotine exposure increased tumorigenicity of oral epithelial cells. ► Si

  14. Prostate cancer stem-like cells proliferate slowly and resist etoposide-induced cytotoxicity via enhancing DNA damage response

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Judy [Division of Nephrology, Department of Medicine, McMaster University, Juravinski Innovation Tower, Room T3310, St. Joseph' s Hospital, 50 Charlton Ave East, Hamilton, Ontario, Canada L8S 4L8 (Canada); Father Sean O' Sullivan Research Institute, Hamilton, Ontario, Canada L8N 4A6 (Canada); The Hamilton Centre for Kidney Research (HCKR), St. Joseph' s Hamilton Healthcare, Hamilton, Ontario, Canada L8N 4A6 (Canada); Tang, Damu, E-mail: [Division of Nephrology, Department of Medicine, McMaster University, Juravinski Innovation Tower, Room T3310, St. Joseph' s Hospital, 50 Charlton Ave East, Hamilton, Ontario, Canada L8S 4L8 (Canada); Father Sean O' Sullivan Research Institute, Hamilton, Ontario, Canada L8N 4A6 (Canada); The Hamilton Centre for Kidney Research (HCKR), St. Joseph' s Hamilton Healthcare, Hamilton, Ontario, Canada L8N 4A6 (Canada)


    Despite the development of chemoresistance as a major concern in prostate cancer therapy, the underlying mechanisms remain elusive. In this report, we demonstrate that DU145-derived prostate cancer stem cells (PCSCs) progress slowly with more cells accumulating in the G1 phase in comparison to DU145 non-PCSCs. Consistent with the important role of the AKT pathway in promoting G1 progression, DU145 PCSCs were less sensitive to growth factor-induced activation of AKT in comparison to non-PCSCs. In response to etoposide (one of the most commonly used chemotherapeutic drugs), DU145 PCSCs survived significantly better than non-PCSCs. In addition to etoposide, PCSCs demonstrated increased resistance to docetaxel, a taxane drug that is commonly used to treat castration-resistant prostate cancer. Etoposide produced elevated levels of γH2AX and triggered a robust G2/M arrest along with a coordinated reduction of the G1 population in PCSCs compared to non-PCSCs, suggesting that elevated γH2AX plays a role in the resistance of PCSCs to etoposide-induced cytotoxicity. We have generated xenograft tumors from DU145 PCSCs and non-PCSCs. Consistent with the knowledge that PCSCs produce xenograft tumors with more advanced features, we were able to demonstrate that PCSC-derived xenograft tumors displayed higher levels of γH2AX and p-CHK1 compared to non-PCSC-produced xenograft tumors. Collectively, our research suggests that the elevation of DNA damage response contributes to PCSC-associated resistance to genotoxic reagents. - Highlights: • Increased survival in DU145 PCSCs following etoposide-induced cytotoxicity. • PCSCs exhibit increased sensitivity to etoposide-induced DDR. • Resistance to cytotoxicity may be due to slower proliferation in PCSCs. • Reduced kinetics to growth factor induced activation of AKT in PCSCs.

  15. Dual drug-loaded biofunctionalized amphiphilic chitosan nanoparticles: Enhanced synergy between cisplatin and demethoxycurcumin against multidrug-resistant stem-like lung cancer cells. (United States)

    Huang, Wei-Ting; Larsson, Mikael; Lee, Yi-Chi; Liu, Dean-Mo; Chiou, Guang-Yuh


    Lung cancer kills more humans than any other cancer and multidrug resistance (MDR) in cancer stem-like cells (CSC) is emerging as a reason for failed treatments. One concept that addresses this root cause of treatment failure is the utilization of nanoparticles to simultaneously deliver dual drugs to cancer cells with synergistic performance, easy to envision - hard to achieve. (1) It is challenging to simultaneously load drugs of highly different physicochemical properties into one nanoparticle, (2) release kinetics may differ between drugs and (3) general requirements for biomedical nanoparticles apply. Here self-assembled nanoparticles of amphiphilic carboxymethyl-hexanoyl chitosan (CHC) were shown to present nano-microenvironments enabling simultaneous loading of hydrophilic and hydrophobic drugs. This was expanded into a dual-drug nano-delivery system to treat lung CSC. CHC nanoparticles were loaded/chemically modified with the anticancer drug cisplatin and the MDR-suppressing Chinese herbal extract demethoxycurcumin, followed by biofunctionalization with CD133 antibody for enhanced uptake by lung CSC, all in a feasible one-pot preparation. The nanoparticles were characterized with regard to chemistry, size, zeta potential and drug loading/release. Biofunctionalized and non-functionalized nanoparticles were investigated for uptake by lung CSC. Subsequently the cytotoxicity of single and dual drugs, free in solution or in nanoparticles, was evaluated against lung CSC at different doses. From the dose response at different concentrations the degree of synergy was determined through Chou-Talalay's Plot. The biofunctionalized nanoparticles promoted synergistic effects between the drugs and were highly effective against MDR lung CSC. The efficacy and feasible one-pot preparation suggests preclinical studies using relevant disease models to be justified. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. EGFR Amplification and Glioblastoma Stem-Like Cells

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    Katrin Liffers


    Full Text Available Glioblastoma (GBM, the most common malignant brain tumor in adults, contains a subpopulation of cells with a stem-like phenotype (GS-cells. GS-cells can be maintained in vitro using serum-free medium supplemented with epidermal growth factor, basic fibroblast growth factor-2, and heparin. However, this method does not conserve amplification of the Epidermal Growth Factor Receptor (EGFR gene, which is present in over 50% of all newly diagnosed GBM cases. GS-cells with retained EGFR amplification could overcome the limitations of current in vitro model systems and contribute significantly to preclinical research on EGFR-targeted therapy. This review recapitulates recent methodological approaches to expand stem-like cells from GBM with different EGFR status in order to maintain EGFR-dependent intratumoral heterogeneity in vitro. Further, it will summarize the current knowledge about the impact of EGFR amplification and overexpression on the stem-like phenotype of GBM-derived GS-cells and different approaches to target the EGFR-dependent GS-cell compartment of GBM.

  17. Harnessing the apoptotic programs in cancer stem-like cells. (United States)

    Wang, Ying-Hua; Scadden, David T


    Elimination of malignant cells is an unmet challenge for most human cancer types even with therapies targeting specific driver mutations. Therefore, a multi-pronged strategy to alter cancer cell biology on multiple levels is increasingly recognized as essential for cancer cure. One such aspect of cancer cell biology is the relative apoptosis resistance of tumor-initiating cells. Here, we provide an overview of the mechanisms affecting the apoptotic process in tumor cells emphasizing the differences in the tumor-initiating or stem-like cells of cancer. Further, we summarize efforts to exploit these differences to design therapies targeting that important cancer cell population. © 2015 The Authors.

  18. Breast cancer stem-like cells and breast cancer therapy

    Institute of Scientific and Technical Information of China (English)

    Niansong Qian; Nobuko Kawaguchi-Sakita; Masakazu Toi


    @@ Until the early 1990s, human cancers were considered a morphologically heterogeneous population of cells. In 1997, Bonnet et al[1] demonstrated that a small population of leukemia cells was able to differentiate in vivo into leukemic blasts, indicating that the leukemic clone was organized as a hierarchy; this was subsequently denoted as cancer stem like cells (CSCs). CSCs are cancer cells that possess characteristics associated with normal stem cells and have the specific ability to give rise to all cell types found in a particular cancer. One reason for the failure of traditional anti tumor therapies might be their inability to eradicate CSCs. Therefore, therapies must identify and destroy CSCs in both primary and metastatic tumors.

  19. Characterization of glioma stem-like cells from human glioblastomas. (United States)

    Yamamuro, Shun; Okamoto, Yutaka; Sano, Emiko; Ochiai, Yushi; Ogino, Akiyoshi; Ohta, Takashi; Hara, Hiroyuki; Ueda, Takuya; Nakayama, Tomohiro; Yoshino, Atsuo; Katayama, Yoichi


    Glioma stem-like cells (GSCs) could have potential for tumorigenesis, treatment resistance, and tumor recurrence (GSC hypothesis). However, the mechanisms underlying such potential has remained elusive and few ultrastructural features of the cells have been reported in detail. We therefore undertook observations of the antigenic characteristics and ultrastructural features of GSCs isolated from human glioblastomas. Tumor spheres formed by variable numbers of cells, exhibiting a variable appearance in both their size and shape, were frequently seen in GSCs expressing the stem cell surface markers CD133 and CD15. Increased cell nucleus atypia, mitochondria, rough endoplasmic reticulum, coated vesicles, and microvilli, were noted in the GSCs. Furthermore, cells at division phases and different phases of the apoptotic process were occasionally observed. These findings could imply that GSCs have certain relations with human neural stem cells (NSCs) but are primitively different from undifferentiated NSCs. The data may provide support for the GSC hypothesis, and also facilitate the establishment of future glioblastoma treatments targeting GSCs.

  20. Delayed cell death associated with mitotic catastrophe in γ-irradiated stem-like glioma cells

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    Esser Norbert


    Full Text Available Abstract Background and Purpose Stem-like tumor cells are regarded as highly resistant to ionizing radiation (IR. Previous studies have focused on apoptosis early after irradiation, and the apoptosis resistance observed has been attributed to reduced DNA damage or enhanced DNA repair compared to non-stem tumor cells. Here, early and late radioresponse of patient-derived stem-like glioma cells (SLGCs and differentiated cells directly derived from them were examined for cell death mode and the influence of stem cell-specific growth factors. Materials and methods Primary SLGCs were propagated in serum-free medium with the stem-cell mitogens epidermal growth factor (EGF and fibroblast growth factor-2 (FGF-2. Differentiation was induced by serum-containing medium without EGF and FGF. Radiation sensitivity was evaluated by assessing proliferation, clonogenic survival, apoptosis, and mitotic catastrophe. DNA damage-associated γH2AX as well as p53 and p21 expression were determined by Western blots. Results SLGCs failed to apoptose in the first 4 days after irradiation even at high single doses up to 10 Gy, but we observed substantial cell death later than 4 days postirradiation in 3 of 6 SLGC lines treated with 5 or 10 Gy. This delayed cell death was observed in 3 of the 4 SLGC lines with nonfunctional p53, was associated with mitotic catastrophe and occurred via apoptosis. The early apoptosis resistance of the SLGCs was associated with lower γH2AX compared to differentiated cells, but we found that the stem-cell culture cytokines EGF plus FGF-2 strongly reduce γH2AX levels. Nonetheless, in two p53-deficient SLGC lines examined γIR-induced apoptosis even correlated with EGF/FGF-induced proliferation and mitotic catastrophe. In a line containing CD133-positive and -negative stem-like cells, the CD133-positive cells proliferated faster and underwent more γIR-induced mitotic catastrophe. Conclusions Our results suggest the importance of delayed

  1. Gene transfection in primary stem-like cells of giant cell tumor of bone. (United States)

    Singh, Shalini; Mak, Isabella; Power, Patricia; Cunningham, Melissa; Cunnigham, Melissa; Turcotte, Robert; Ghert, Michelle


    The neoplastic stem-like stromal cell of giant cell tumor of bone (GCT) survives for multiple passages in primary culture with a stable phenotype, and exhibits multipotent characteristics. The pathophysiology of this tumor has been studied through the primary culture of these cells. However, successful gene transfer of these cells has not been reported to date. In this short report, we describe the development of the first reported technique that results in efficient gene transfection in primary stem-like cells of GCT.

  2. Cisplatin selects for stem-like cells in osteosarcoma by activating Notch signaling. (United States)

    Yu, Ling; Fan, Zhengfu; Fang, Shuo; Yang, Jian; Gao, Tian; Simões, Bruno M; Eyre, Rachel; Guo, Weichun; Clarke, Robert B


    Notch signaling regulates normal stem cells and is also thought to regulate cancer stem cells (CSCs). Recent data indicate that Notch signaling plays a role in the development and progression of osteosarcoma, however the regulation of Notch in chemo-resistant stem-like cells has not yet been fully elucidated. In this study we generated cisplatin-resistant osteosarcoma cells by treating them with sub-lethal dose of cisplatin, sufficient to induce DNA damage responses. Cisplatin-resistant osteosarcoma cells exhibited lower proliferation, enhanced spheroid formation and more mesenchymal characteristics than cisplatin-sensitive cells, were enriched for Stro-1+/CD117+ cells and showed increased expression of stem cell-related genes. A similar effect was observed in vivo, and in addition in vivo tumorigenicity was enhanced during serial transplantation. Using several publicly available datasets, we identified that Notch expression was closely associated with osteosarcoma stem cells and chemotherapy resistance. We confirmed that cisplatin-induced enrichment of osteosarcoma stem cells was mediated through Notch signaling in vitro, and immunohistochemistry showed that cleaved Notch1 (NICD1) positive cells were significantly increased in a relapsed xenograft which had received cisplatin treatment. Furthermore, pretreatment with a γ-secretase inhibitor (GSI) to prevent Notch signalling inhibited cisplatin-enriched osteosarcoma stem cell activity in vitro, including Stro-1+/CD117+ double positive cells and spheroid formation capacity. The Notch inhibitor DAPT also prevented tumor recurrence in resistant xenograft tumors. Overall, our results show that cisplatin induces the enrichment of osteosarcoma stem-like cells through Notch signaling, and targeted inactivation of Notch may be useful for the elimination of CSCs and overcoming drug resistance.

  3. Cancer stem-like cells in Epstein-Barr virus-associated nasopharyngeal carcinoma

    Institute of Scientific and Technical Information of China (English)

    Samantha Wei-Man Lun; Siu-Tim Cheung; Kwok-Wai Lo


    Although the Epstein-Barr virus (EBV) has spread to all populations in the world, EBV-associated nasopharyngeal carcinoma (NPC) is prevalent only in South China and Southeast Asia. The role of EBV in the malignant transformation of nasopharyngeal epithelium is the main focus of current researches. Radiotherapy and chemoradiotherapy have been successful in treating early stage NPC, but the recurrence rates remain high. Unfortunately, local relapse and metastasis are commonly unresponsive to conventional treatments. These recurrent and metastatic lesions are believed to arise from residual or surviving cells that have the properties of cancer stem cels. These cancer stem-like cels (CSCs) have the ability to self-renew, differentiate, and sustain propagation. They are also chemo-resistant and can form spheres in anchorage-independent environments. This review summarizes recent researches on the CSCs in EBV-associated NPC, including the findings regarding cell surface markers, stem cell-related transcription factors, and various signaling pathways. In particular, the review focuses on the roles of EBV latent genes [latent membrane protein 1 (LMP1) and latent membrane protein 2A (LMP2A)], cellular microRNAs, and adenosine triphosphate (ATP)-binding cassette chemodrug transporters in contributing to the properties of CSCs, including the epithelial-mesenchymal transition, stem-like transition, and chemo-resistance. Novel therapeutics that enhance the efficacy of radiotherapy and chemoradiotherapy and inhibitors that suppress the properties of CSCs are also discussed.

  4. Isolation and Identification of Cancer Stem-Like Cells from Murine Melanoma Cell Lines

    Institute of Scientific and Technical Information of China (English)

    Jun Dou; Kai Hu; Ning Gu; Meng Pan; Ping Wen; Yating Li; Quan Tang; Lili Chu; Fengshu Zhao; Chuilian Jiang; Weihua Hu


    In current study, cancer stem-like cells in the murine melanoma B16F10 cells were investigated. CD phenotypes of the B16F10 cells were analyzed by flow cytometry, and the specific CD phenotype cells from the B16F10 cells were isolated by MACS. Then we used colony formation assay in soft agar media, the cell growth assay in serum-free culture media as well as the tumorigenicity investigation of the specific CD phenotype cells in C57BL/6 mice,respectively, to identify cancer stem-like cells in the B16F10 cells. The results showed that the B16F10 cells could form spherical clones in serum-free culture media, and the rate of clonegenesis of CD133+, CD44+ and CD44+CD133+ cells was higher than that of CD133-, CD44- and CD44+CD133- cells in soft agar media, respectively.The tumorigenic potential of CD133+, CD44+, CD44+CD133+ cells and CD44+CD133+CD24+ cells was stronger than that of CD133-, CD44-, CD44+CD133- cells and CD44+CD133+CD24- cells in mice, respectively. In conclusion, the CD44+CD133+CD24+ cells have some biological properties of cancer stem-like cells or are highly similar to the characteristics of cancer stem cells (CSC). These results provide an important method for identifying cancer stem-like cells in B16F10 cells and for further cancer target therapy.

  5. Transcriptional profiling of adult neural stem-like cells from the human brain. (United States)

    Sandberg, Cecilie Jonsgar; Vik-Mo, Einar O; Behnan, Jinan; Helseth, Eirik; Langmoen, Iver A


    There is a great potential for the development of new cell replacement strategies based on adult human neural stem-like cells. However, little is known about the hierarchy of cells and the unique molecular properties of stem- and progenitor cells of the nervous system. Stem cells from the adult human brain can be propagated and expanded in vitro as free floating neurospheres that are capable of self-renewal and differentiation into all three cell types of the central nervous system. Here we report the first global gene expression study of adult human neural stem-like cells originating from five human subventricular zone biopsies (mean age 42, range 33-60). Compared to adult human brain tissue, we identified 1,189 genes that were significantly up- and down-regulated in adult human neural stem-like cells (1% false discovery rate). We found that adult human neural stem-like cells express stem cell markers and have reduced levels of markers that are typical of the mature cells in the nervous system. We report that the genes being highly expressed in adult human neural stem-like cells are associated with developmental processes and the extracellular region of the cell. The calcium signaling pathway and neuroactive ligand-receptor interactions are enriched among the most differentially regulated genes between adult human neural stem-like cells and adult human brain tissue. We confirmed the expression of 10 of the most up-regulated genes in adult human neural stem-like cells in an additional sample set that included adult human neural stem-like cells (n = 6), foetal human neural stem cells (n = 1) and human brain tissues (n = 12). The NGFR, SLITRK6 and KCNS3 receptors were further investigated by immunofluorescence and shown to be heterogeneously expressed in spheres. These receptors could potentially serve as new markers for the identification and characterisation of neural stem- and progenitor cells or as targets for manipulation of cellular fate.

  6. The senescent microenvironment promotes the emergence of heterogeneous cancer stem-like cells. (United States)

    Castro-Vega, Luis Jaime; Jouravleva, Karina; Ortiz-Montero, Paola; Liu, Win-Yan; Galeano, Jorge Luis; Romero, Martha; Popova, Tatiana; Bacchetti, Silvia; Vernot, Jean Paul; Londoño-Vallejo, Arturo


    There is a well-established association between aging and the onset of metastasis. Although the mechanisms through which age impinges upon the malignant phenotype remain uncharacterized, the role of a senescent microenvironment has been emphasized. We reported previously that human epithelial cells that undergo telomere-driven chromosome instability (T-CIN) display global microRNA (miR) deregulation and develop migration and invasion capacities. Here, we show that post-crisis cells are not able to form tumors unless a senescent microenvironment is provided. The characterization of cell lines established from such tumors revealed that these cells have acquired cell autonomous tumorigenicity, giving rise to heterogeneous tumors. Further experiments demonstrate that explanted cells, while displaying differences in cell differentiation markers, are all endowed of enhanced stem cell properties including self-renewal and multilineage differentiation capacity. Treatments of T-CIN+ cells with senescence-conditioned media induce sphere formation exclusively in cells with senescence-associated tumorigenicity, a capacity that depends on miR-145 repression. These results indicate that the senescent microenvironment, while promoting further transdifferentiations in cells with genome instability, is able to propel the progression of premalignant cells towards a malignant, cell stem-like state.

  7. HSP DNAJB8 Controls Tumor-Initiating Ability in Renal Cancer Stem-like Cells

    NARCIS (Netherlands)

    Nishizawa, Satoshi; Hirohashi, Yoshihiko; Torigoe, Toshihiko; Takahashi, Akari; Tamura, Yasuaki; Mori, Takashi; Kanaseki, Takayuki; Kamiguchi, Kenjiro; Asanuma, Hiroko; Morita, Rena; Sokolovskaya, Alice; Matsuzaki, Junichi; Yamada, Ren; Fujii, Reona; Kampinga, Harm H.; Kondo, Toru; Hasegawa, Tadashi; Hara, Isao; Sato, Noriyuki


    Cancer stem-like cells (CSC) are a small population of cancer cells with superior tumor initiating, self-renewal, and differentiation properties. In this study, we show that the cancer-testis antigen and HSP40 family member DNAJB8 contributes to the CSC phenotype in renal cell carcinoma (RCC). DNAJB

  8. Cytotoxic effect of disulfiram/copper on human glioblastoma cell lines and ALDH-positive cancer-stem-like cells (United States)

    Liu, P; Brown, S; Goktug, T; Channathodiyil, P; Kannappan, V; Hugnot, J-P; Guichet, P-O; Bian, X; Armesilla, A L; Darling, J L; Wang, W


    Background: Glioblastoma multiforme (GBM) cells are resistant to anticancer drugs. Cancer stem cells (CSCs) are a key mediator of chemoresistance. We have reported that disulfiram (DS), an aldehyde dehydrogenase (ALDH) inhibitor, targets breast CSC-like cells. In this study, the effect of DS and combination of DS and gemcitabine (dFdC) on GBM cells and GBM stem-like cells was investigated. Methods: 1-(4,5-Dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT), combination index (CI)-isobologram, western blot, luciferase reporter gene assay, electrophoretic mobility-shift assay and ALDH analysis were used in this study. Results: Disulfiram is cytotoxic in GBM cell lines in a copper (Cu)-dependent manner. Disulfiram/copper enhances the cytotoxicity of dFdC. Combination index-isobologram analysis indicates a synergistic effect between DS/Cu and dFdC. Disulfiram/copper induces reactive oxygen species (ROS), activates JNK and p38 pathways and inhibits nuclear factor-kappa B activity in GBM cell lines. Disulfiram/copper may trigger intrinsic apoptotic pathway via modulation of the Bcl2 family. Disulfiram/copper abolishes stem-like cell population in GBM cell lines. Conclusion: Our findings indicate that the cytotoxicity of DS/Cu and the enhancing effect of DS/Cu on the cytotoxicity of dFdC in GBM stem-like cells may be caused by induction of ROS and inhibition of both ALDH and the NFkB pathway. Both DS and dFdC can traverse the blood–brain barrier. Further study may lead them into GBM chemotherapy. PMID:23033007

  9. Two-stage induced differentiation of OCT4+/Nanog+ stem-like cells in lung adenocarcinoma. (United States)

    Li, Rong; Huang, Jinsu; Ma, Meili; Lou, Yuqing; Zhang, Yanwei; Wu, Lixia; Chang, David W; Zhao, Picheng; Dong, Qianggang; Wu, Xifeng; Han, Baohui


    Stem-like cells in solid tumors are purported to contribute to cancer development and poor treatment outcome. The abilities to self-renew, differentiate, and resist anticancer therapies are hallmarks of these rare cells, and steering them into lineage commitment may be one strategy to curb cancer development or progression. Vitamin D is a prohormone that can alter cell growth and differentiation and may induce the differentiation cancer stem-like cells. In this study, octamer-binding transcription factor 4 (OCT4)-positive/Nanog homeobox (Nanog)- positive lung adenocarcinoma stem-like cells (LACSCs) were enriched from spheroid cultured SPC-A1 cells and differentiated by a two-stage induction (TSI) method, which involved knockdown of hypoxia-inducible factor 1-alpha (HIF1α) expression (first stage) followed by sequential induction with 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3, VD3) and suberoylanilide hydroxamic acid (SAHA) treatment (second stage). The results showed the HIF1α-knockdowned cells displayed diminished cell invasion and clonogenic activities. Moreover, the TSI cells highly expressed tumor suppressor protein p63 (P63) and forkhead box J1 (FOXJ1) and lost stem cell characteristics, including absent expression of OCT4 and Nanog. These cells regained sensitivity to cisplatin in vitro while losing tumorigenic capacity and decreased tumor cell proliferation in vivo. Our results suggest that induced transdifferentiation of LACSCs by vitamin D and SAHA may become novel therapeutic avenue to alter tumor cell phenotypes and improve patient outcome.The development and progression of lung cancer may involve rare population of stem-like cells that have the ability to grow, differentiate, and resist drug treatment. However, current therapeutic strategies have mostly focused on tumor characteristics and neglected the potential source of cells that may contribute to poor clinical outcome. We generated lung adenocarcinoma stem-like cells from spheroid culture and

  10. Prostate Cancer Stem-Like Cells | Center for Cancer Research (United States)

    Prostate cancer is the third leading cause of cancer-related death among men, killing an estimated 27,000 men each year in the United States. Men with advanced prostate cancer often become resistant to conventional therapies. Many researchers speculate that the emergence of resistance is due to the presence of cancer stem cells, which are believed to be a small subpopulation of tumor cells that can self-renew and give rise to more differentiated tumor cells. It is thought that these stem cells survive initial therapies (such as chemotherapy and hormone therapy) and then generate new tumor cells that are resistant to these standard treatments. If prostate cancer stem cells could be identified and characterized, it might be possible to design treatments that prevent resistance.

  11. Stem-Like Cells in Bone Sarcomas: Implications for Tumorigenesis

    Directory of Open Access Journals (Sweden)

    C. Parker Gibbs


    Full Text Available Bone sarcomas are a clinically and molecularly heterogeneous group of malignancies characterized by varying degrees of mesenchymal differentiation. Despite advances in medical and surgical management, survival rates for high-grade tumors have remained static at 50% to 70%. Tumor stem cells have been recently implicated in the pathogenesis of other heterogeneous, highly malignant tumors. We demonstrate here the existence of a small subpopulation of self-renewing bone sarcoma cells that are capable of forming suspended spherical, clonal colonies, also called “sarcospheres,” in anchorage-independent, serum-starved conditions. These bone sarcoma cells as well as tissue specimens express activated STAT3 and the marker genes of pluripotent embryonic stem (ES cells, Oct 3/4 and Nanog. Expression levels of Oct 3/4 and Nanog are greater in sarcospheres than in adherent cultures. A subset of bone sarcoma cells displays several surface markers of mesenchymal stem cells (Stro-1, CD105, and CD44 as well as attributes of mesodermal, ectodermal, and endodermal differentiation. Although previously documented in brain and breast tumors, our results support the extension of the cancer stem cell hypothesis to include tumors of mesenchymal lineage. Furthermore, they suggest the participation of ES cell homeobox proteins in non-germ cell tumorigenesis.

  12. Generation of Novel Thyroid Cancer Stem-Like Cell Clones: Effects of Resveratrol and Valproic Acid. (United States)

    Hardin, Heather; Yu, Xiao-Min; Harrison, April D; Larrain, Carolina; Zhang, Ranran; Chen, Jidong; Chen, Herbert; Lloyd, Ricardo V


    Anaplastic thyroid cancer is an aggressive and highly lethal cancer for which conventional therapies have proved ineffective. Cancer stem-like cells (CSCs) represent a small fraction of cells in the cancer that are resistant to chemotherapy and radiation therapy and are responsible for tumor reoccurrence and metastasis. We characterized CSCs in thyroid carcinomas and generated clones of CSC lines. Our study showed that anaplastic thyroid cancers had significantly more CSCs than well-differentiated thyroid cancers. We also showed that Aldefluor-positive cells revealed significantly higher expression of stem cell markers, self-renewal properties, thyrosphere formation, and enhanced tumorigenicity. In vivo passaging of Aldefluor-positive cells resulted in the growth of larger, more aggressive tumors. We isolated and generated two clonal spheroid CSC lines derived from anaplastic thyroid cancer that were even more enriched with stem cell markers and more tumorigenic than the freshly isolated Aldefluor-positive cells. Resveratrol and valproic acid treatment of one of the CSC lines resulted in a significant decrease in stem cell markers, Aldefluor expression, proliferation, and invasiveness, with an increase in apoptosis and thyroid differentiation markers, suggesting that these cell lines may be useful for discovering new adjuvant therapies for aggressive thyroid cancers. For the first time, we have two thyroid CSC lines that will be useful tools for the study of thyroid CSC targeted therapies.

  13. LGR5 positivity defines stem-like cells in colorectal cancer

    NARCIS (Netherlands)

    Hirsch, Daniela; Barker, Nick; McNeil, Nicole; Hu, Yue; Camps, Jordi; McKinnon, Katherine; Clevers, Hans; Ried, Thomas; Gaiser, Timo


    Like normal colorectal epithelium, colorectal carcinomas (CRCs) are organized hierarchically and include populations of cells with stem-like properties. Leucine-rich-repeat-containing G-protein-coupled receptor 5 (LGR5) is associated with these stem cells in normal colorectal epithelium; however, th

  14. Glioblastoma stem-like cells give rise to tumour endothelium

    NARCIS (Netherlands)

    R. Wang; K. Chadalavada; J. Wilshire; U. Kowalik; K.E. Hovinga; A. Geber; B. Fligelman; M. Leversha; C. Brennan; V. Tabar


    Glioblastoma (GBM) is among the most aggressive of human cancers(1). A key feature of GBMs is the extensive network of abnormal vasculature characterized by glomeruloid structures and endothelial hyperplasia(2). Yet the mechanisms of angiogenesis and the origin of tumour endothelial cells remain poo

  15. Secondary Data Analytics of Aquaporin Expression Levels in Glioblastoma Stem-Like Cells. (United States)

    Isokpehi, Raphael D; Wollenberg Valero, Katharina C; Graham, Barbara E; Pacurari, Maricica; Sims, Jennifer N; Udensi, Udensi K; Ndebele, Kenneth


    Glioblastoma is the most common brain tumor in adults in which recurrence has been attributed to the presence of cancer stem cells in a hypoxic microenvironment. On the basis of tumor formation in vivo and growth type in vitro, two published microarray gene expression profiling studies grouped nine glioblastoma stem-like (GS) cell lines into one of two groups: full (GSf) or restricted (GSr) stem-like phenotypes. Aquaporin-1 (AQP1) and aquaporin-4 (AQP4) are water transport proteins that are highly expressed in primary glial-derived tumors. However, the expression levels of AQP1 and AQP4 have not been previously described in a panel of 92 glioma samples. Therefore, we designed secondary data analytics methods to determine the expression levels of AQP1 and AQP4 in GS cell lines and glioblastoma neurospheres. Our investigation also included a total of 2,566 expression levels from 28 Affymetrix microarray probe sets encoding 13 human aquaporins (AQP0-AQP12); CXCR4 (the receptor for stromal cell derived factor-1 [SDF-1], a potential glioma stem cell therapeutic target]); and PROM1 (gene encoding CD133, the widely used glioma stem cell marker). Interactive visual representation designs for integrating phenotypic features and expression levels revealed that inverse expression levels of AQP1 and AQP4 correlate with distinct phenotypes in a set of cell lines grouped into full and restricted stem-like phenotypes. Discriminant function analysis further revealed that AQP1 and AQP4 expression are better predictors for tumor formation and growth types in glioblastoma stem-like cells than are CXCR4 and PROM1. Future investigations are needed to characterize the molecular mechanisms for inverse expression levels of AQP1 and AQP4 in the glioblastoma stem-like neurospheres.

  16. mir-300 promotes self-renewal and inhibits the differentiation of glioma stem-like cells

    KAUST Repository

    Zhang, Daming


    MicroRNAs (miRNAs) are small noncoding RNAs that have been critically implicated in several human cancers. miRNAs are thought to participate in various biological processes, including proliferation, cell cycle, apoptosis, and even the regulation of the stemness properties of cancer stem cells. In this study, we explore the potential role of miR-300 in glioma stem-like cells (GSLCs). We isolated GSLCs from glioma biopsy specimens and identified the stemness properties of the cells through neurosphere formation assays, multilineage differentiation ability analysis, and immunofluorescence analysis of glioma stem cell markers. We found that miR-300 is commonly upregulated in glioma tissues, and the expression of miR-300 was higher in GSLCs. The results of functional experiments demonstrated that miR-300 can enhance the self-renewal of GSLCs and reduce differentiation toward both astrocyte and neural fates. In addition, LZTS2 is a direct target of miR-300. In conclusion, our results demonstrate the critical role of miR-300 in GSLCs and its functions in LZTS2 inhibition and describe a new approach for the molecular regulation of tumor stem cells. © 2014 Springer Science+Business Media.

  17. Angiogenesis-independent tumor growth mediated by stem-like cancer cells.

    NARCIS (Netherlands)

    Sakariassen, P.; Prestegarden, L.; Wang, J.; Skaftnesmo, K.O.; Mahesparan, R.; Molthoff, C.F.M.; Sminia, P.; Sundlisaeter, E.; Misra, A.; Tysnes, B.B.; Chekenya, M.; Peters, H.; Lende, G.; Kalland, K.H.; Oyan, A.M.; Petersen, K.; Jonassen, I.; Kogel, A.J. van der; Feuerstein, B.G.; Terzis, A.J.; Bjerkvig, R.; Enger, P.O.


    In this work, highly infiltrative brain tumors with a stem-like phenotype were established by xenotransplantation of human brain tumors in immunodeficient nude rats. These tumors coopted the host vasculature and presented as an aggressive disease without signs of angiogenesis. The malignant cells ex

  18. Aberrant mesenchymal differentiation of glioma stem-like cells : implications for therapeutic targeting

    NARCIS (Netherlands)

    Balasubramaniyan, Veerakumar; Vaillant, Brian; Wang, Shuzhen; Gumin, Joy; Butalid, M. Elena; Sai, Ke; Mukheef, Farah; Kim, Se Hoon; Boddeke, H. W. G. M.; Lang, Frederick; Aldape, Kenneth; Sulman, Erik P.; Bhat, Krishna P.; Colman, Howard


    Differentiation has been proposed as a therapeutic strategy for glioblastoma (GBM) in part due to observations of stem-like cells in GBM that have been shown to undergo terminal differentiation in response to growth factor withdrawal and BMP activation. However, the effects of long term exposure to

  19. Characterization of stem-like cells in a new astroblastoma cell line. (United States)

    Coban, Esra Aydemir; Kasikci, Ezgi; Karatas, Omer Faruk; Suakar, Oznur; Kuskucu, Aysegul; Altunbek, Mine; Türe, Uğur; Sahin, Fikrettin; Bayrak, Omer Faruk


    Cell lines established from tumors are the most commonly used models in cancer research, and their use in recent years has enabled a greater understanding of the biology of cancer and the means to develop effective treatment strategies. Astroblastomas are uncommon neuroepithelial tumors of glial origin, predominantly affecting young people, mainly teenagers and children, predominantly females. To date, only a single study has reported that astroblastomas contain a large number of neural stem-like cells, which had only a partial proliferation capacity and differentiation. Our objective was to establish an astroblastoma cell line to investigate the presence of astroblastic cells and cancer stem-like cells. The migratory and invasion abilities of the cells were quantified with invasion and migration assays and compared to a glioblastoma cell line. The presence of stem cells was detected with surface-marker analysis by using flow cytometry, and measuring the differentiation ability with a differentiation assay and the self-renewal capacity with a sphere-forming assay. These characteristics may determine whether this novel cell line is a model for astroblastomas that may have stem-cell characteristics. With this novel cell line, scientists can investigate the molecular pathways underlying astroblastomas and develop new therapeutic strategies for patients with these tumors. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Polysome Profiling Links Translational Control to the Radioresponse of Glioblastoma Stem-like Cells. (United States)

    Wahba, Amy; Rath, Barbara H; Bisht, Kheem; Camphausen, Kevin; Tofilon, Philip J


    Changes in polysome-bound mRNA (translatome) are correlated closely with changes in the proteome in cells. Therefore, to better understand the processes mediating the response of glioblastoma to ionizing radiation (IR), we used polysome profiling to define the IR-induced translatomes of a set of human glioblastoma stem-like cell (GSC) lines. Although cell line specificity accounted for the largest proportion of genes within each translatome, there were also genes that were common to the GSC lines. In particular, analyses of the IR-induced common translatome identified components of the DNA damage response, consistent with a role for the translational control of gene expression in cellular radioresponse. Moreover, translatome analyses suggested that IR enhanced cap-dependent translation processes, an effect corroborated by the finding of increased eIF4F-cap complex formation detected after irradiation in all GSC lines. Translatome analyses also predicted that Golgi function was affected by IR. Accordingly, Golgi dispersal was detected after irradiation of each of the GSC lines. In addition to the common responses seen, translatome analyses predicted cell line-specific changes in mitochondria, as substantiated by changes in mitochondrial mass and DNA content. Together, these results suggest that analysis of radiation-induced translatomes can provide new molecular insights concerning the radiation response of cancer cells. More specifically, they suggest that the translational control of gene expression may provide a source of molecular targets for glioblastoma radiosensitization. Cancer Res; 76(10); 3078-87. ©2016 AACR.

  1. Establishment and Analysis of Cancer Stem-Like and Non-Cancer Stem-Like Clone Cells from the Human Colon Cancer Cell Line SW480. (United States)

    Takaya, Akari; Hirohashi, Yoshihiko; Murai, Aiko; Morita, Rena; Saijo, Hiroshi; Yamamoto, Eri; Kubo, Terufumi; Nakatsugawa, Munehide; Kanaseki, Takayuki; Tsukahara, Tomohide; Tamura, Yasuaki; Takemasa, Ichiro; Kondo, Toru; Sato, Noriyuki; Torigoe, Toshihiko


    Human cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) can be isolated as side population (SP) cells, aldehyde dehydrogenase high (ALDHhigh) cells or cell surface marker-positive cells including CD44+ cells and CD133+ cells. CSCs/CICs and non-CSCs/CICs are unstable in in vitro culture, and CSCs/CICs can differentiate into non-CSCs/CICs and some non-CSCs/CICs can dedifferentiate into CSCs/CICs. Therefore, experiments using a large amount of CSCs/CICs are technically very difficult. In this study, we isolated single cell clones from SP cells and main population (MP) cells derived from the human colon cancer cell line SW480. SP analysis revealed that SP clone cells had relatively high percentages of SP cells, whereas MP clone cells showed very few SP cells, and the phenotypes were sustainable for more than 2 months of in vitro culture. Xenograft transplantation revealed that SP clone cells have higher tumor-initiating ability than that of MP clone cells and SP clone cell showed higher chemo-resistance compared with MP clone cells. These results indicate that SP clone cells derived from SW480 cells are enriched with CSCs/CICs, whereas MP clone cells are pure non-CSCs/CICs. SP clone cells and MP clone cells are a very stable in vitro CSC/CIC-enriched and non-CSC/CIC model for further analysis.

  2. The Expression of Connexins and SOX2 Reflects the Plasticity of Glioma Stem-Like Cells

    Directory of Open Access Journals (Sweden)

    Joana Balça-Silva


    Full Text Available Glioblastoma (GBM is the most malignant primary brain tumor, with an average survival rate of 15 months. GBM is highly refractory to therapy, and such unresponsiveness is due, primarily, but not exclusively, to the glioma stem-like cells (GSCs. This subpopulation express stem-like cell markers and is responsible for the heterogeneity of GBM, generating multiple differentiated cell phenotypes. However, how GBMs maintain the balance between stem and non-stem populations is still poorly understood. We investigated the GBM ability to interconvert between stem and non-stem states through the evaluation of the expression of specific stem cell markers as well as cell communication proteins. We evaluated the molecular and phenotypic characteristics of GSCs derived from differentiated GBM cell lines by comparing their stem-like cell properties and expression of connexins. We showed that non-GSCs as well as GSCs can undergo successive cycles of gain and loss of stem properties, demonstrating a bidirectional cellular plasticity model that is accompanied by changes on connexins expression. Our findings indicate that the interconversion between non-GSCs and GSCs can be modulated by extracellular factors culminating on differential expression of stem-like cell markers and cell-cell communication proteins. Ultimately, we observed that stem markers are mostly expressed on GBMs rather than on low-grade astrocytomas, suggesting that the presence of GSCs is a feature of high-grade gliomas. Together, our data demonstrate the utmost importance of the understanding of stem cell plasticity properties in a way to a step closer to new strategic approaches to potentially eliminate GSCs and, hopefully, prevent tumor recurrence.

  3. JNK contributes to temozolomide resistance of stem-like glioblastoma cells via regulation of MGMT expression. (United States)

    Okada, Masashi; Sato, Atsushi; Shibuya, Keita; Watanabe, Eriko; Seino, Shizuka; Suzuki, Shuhei; Seino, Manabu; Narita, Yoshitaka; Shibui, Soichiro; Kayama, Takamasa; Kitanaka, Chifumi


    While elimination of the cancer stem cell population is increasingly recognized as a key to successful treatment of cancer, the high resistance of cancer stem cells to conventional chemoradiotherapy remains a therapeutic challenge. O6-methylguanine DNA methyltransferase (MGMT), which is frequently expressed in cancer stem cells of glioblastoma, has been implicated in their resistance to temozolomide, the first-line chemotherapeutic agent against newly diagnosed glioblastoma. However, much remains unknown about the molecular regulation that underlies MGMT expression and temozolomide resistance of glioblastoma cancer stem cells. Here, we identified JNK as a novel player in the control of MGMT expression and temozolomide resistance of glioblastoma cancer stem cells. We showed that inhibition of JNK, either pharmacologically or by RNA interference, in stem-like glioblastoma cells derived directly from glioblastoma tissues reduces their MGMT expression and temozolomide resistance. Importantly, sensitization of stem-like glioblastoma cells to temozolomide by JNK inhibition was dependent on MGMT expression, implying that JNK controls temozolomide resistance of stem-like glioblastoma cells through MGMT expression. Our findings suggest that concurrent use of JNK inhibitors with temozolomide may be a rational therapeutic approach to effectively target the cancer stem cell population in the treatment of glioblastoma.

  4. Transforming Growth Factor-beta signal responding in hepatic stem-like cells

    Institute of Scientific and Technical Information of China (English)

    CUI Wei


    Objective To investigate the effects of TGF-β on the expressions and distribution of phosphorated Smad2/3 and Smad7 in hepatic stem-like cells. Methods Using immunogold transmission electron microscopy, we observed the expressions and distribution of phosphorated Smad2/3, and Smad7 before and after TGF-β1 (5 ng·mL-1) treatment for 4, 8, and 24 hours in hepatic stem-like cells (WB cells). In addition, we also detected the apoptosis status after TGF-β1 stimulation by transmission electron microscopy. Results TGF-β1 stimulation can result in expression increasing of phosphorated Smad2/3 in WB cells, and reach the peak at 8 h, especially in the nuclear. After treatment with TGF-β1 for 24 h, the nuclear expression of phosphorated Smad2/3 gradually decreased. Additionally, we found that TGF-β1 treatment also contributed to increasing in protein level and alteration in cellular distribution of Smad7 (translocation from the nucleus to the cytoplasm) in WB cells. Furthermore, we observed apoptotic body in WB cells after TGF-β1 treatment for 8 h. Conclusions These results indicate that TGF-β stimulation can alter the expression and cellular distribution of phosphorated Srnad2/3 and Smad7 which are its downstream molecular, suggesting hepatic stem-like cells own intact responding to TGF-β.

  5. Epigallocatechin-3-gallate inhibits stem-like inflammatory breast cancer cells.

    Directory of Open Access Journals (Sweden)

    Nora D Mineva

    Full Text Available Inflammatory Breast Cancer (IBC is a highly aggressive form of cancer characterized by high rates of proliferation, lymphangiogenesis and metastasis, and an overall poor survival. As regular green tea consumption has been associated with improved prognosis of breast cancer patients, including decreased risk of recurrence, here the effects of the green tea polyphenol epigallocatechin-3-gallate (EGCG were tested on two IBC lines: SUM-149 and SUM-190. EGCG decreased expression of genes that promote proliferation, migration, invasion, and survival. Consistently, growth, invasive properties, and survival of IBC cells were reduced by EGCG treatment. EGCG also reduced lymphangiogenesis-promoting genes, in particular VEGF-D. Conditioned media from EGCG-treated IBC cells displayed decreased VEGF-D secretion and reduced ability to promote lymphangiogenesis in vitro as measured by hTERT-HDLEC lymphatic endothelial cell migration and tube formation. Tumorsphere formation by SUM-149 cells was robustly inhibited by EGCG, suggesting effects on self-renewal ability. Stem-like SUM-149 cells with high aldehyde dehydrogenase (ALDH activity, previously implicated in poor patient prognosis, were isolated. EGCG treatment reduced growth and induced apoptosis of the stem-like SUM-149 cells in culture. In an orthotopic mouse model, EGCG decreased growth of pre-existing tumors derived from ALDH-positive stem-like SUM-149 cells and their expression of VEGF-D, which correlated with a significant decrease in peritumoral lymphatic vessel density. Thus, EGCG inhibits the overall aggressive IBC phenotype. Reduction of the stem-like cell compartment by EGCG may explain the decreased risk of breast cancer recurrence among green tea drinkers. Recent clinical trials demonstrate the efficacy of green tea polyphenol extracts in treatment of prostate cancer and lymphocytic leukemia with low toxicity. Given the poor prognosis of IBC patients, our findings suggest further exploration

  6. Differentiation of glioblastoma multiforme stem-like cells leads to downregulation of EGFR and EGFRvIII and decreased tumorigenic and stem-like cell potential

    DEFF Research Database (Denmark)

    Stockhausen, Marie-Thérése; Kristoffersen, Karina; Stobbe Olsen, Marie-Louise;


    Glioblastoma multiforme (GBM) is the most common and devastating primary brain tumor among adults. Despite recent treatment progress, most patients succumb to their disease within 2 years of diagnosis. Current research has highlighted the importance of a subpopulation of cells, assigned brain...... cancer stem-like cells (bCSC), to play a pivotal role in GBM malignancy. bCSC are identified by their resemblance to normal neural stem cells (NSC), and it is speculated that the bCSC have to be targeted in order to improve treatment outcome for GBM patients. One hallmark of GBM is aberrant expression...... and activation of the epidermal growth factor receptor (EGFR) and expression of a deletion variant EGFRvIII. In the normal brain, EGFR is expressed in neurogenic areas where also NSC are located and it has been shown that EGFR is involved in regulation of NSC proliferation, migration, and differentiation...

  7. Novel anticancer activity of phloroglucinol against breast cancer stem-like cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Rae-Kwon; Uddin, Nizam [Department of Life Science, Research Institute for Natural Sciences, Hanyang University, Seoul 133-791 (Korea, Republic of); Hyun, Jin-Won [College of Medicine and Applied Radiological Science Research Institute, Jeju National University, Jeju-si 690-756 (Korea, Republic of); Kim, Changil [Department of Biotechnology, Konkuk University, Chungju 380-701 (Korea, Republic of); Suh, Yongjoon, E-mail: [Department of Life Science, Research Institute for Natural Sciences, Hanyang University, Seoul 133-791 (Korea, Republic of); Lee, Su-Jae, E-mail: [Department of Life Science, Research Institute for Natural Sciences, Hanyang University, Seoul 133-791 (Korea, Republic of)


    Poor prognosis of breast cancer patients is closely associated with metastasis and relapse. There is substantial evidence supporting that cancer stem-like cells (CSCs) are primarily responsible for relapse in breast cancer after anticancer treatment. However, there is a lack of suitable drugs that target breast cancer stem-like cells (BCSCs). Here, we report that phloroglucinol (PG), a natural phlorotannin component of brown algae, suppresses sphere formation, anchorage-independent colony formation and in vivo tumorigenicity. In line with these observations, treatment with PG also decreased CD44{sup +} cancer cell population as well as expression of CSC regulators such as Sox2, CD44, Oct4, Notch2 and β-catenin. Also, treatment with PG sensitized breast cancer cells to anticancer drugs such as cisplatin, etoposide, and taxol as well as to ionizing radiation. Importantly, PG inhibited KRAS and its downstream PI3K/AKT and RAF-1/ERK signaling pathways that regulate the maintenance of CSCs. Taken together, our findings implicate PG as a good candidate to target BCSCs and to prevent the disease relapse. - Highlights: • Phloroglucinol suppresses in vivo tumor formation. • Phloroglucinol sensitizes breast cancer cells to anticancer agents. • Phloroglucinol inhibits breast cancer stem-like cells. • Phloroglucinol inhibits PI3K/AKT and KRAS/RAF/ERK signaling pathways.

  8. Isolation of stem-like cells from spontaneous feline mammary carcinomas: Phenotypic characterization and tumorigenic potential

    Energy Technology Data Exchange (ETDEWEB)

    Barbieri, Federica; Wurth, Roberto [Section of Pharmacology, Dept. of Internal Medicine Di.M.I., and Center of Excellence for Biomedical Research - University of Genova, Viale Benedetto XV, 2, 16132 Genova (Italy); Ratto, Alessandra; Campanella, Chiara; Vito, Guendalina [Istituto Zooprofilattico Sperimentale del Piemonte, Liguria e Valle D' Aosta, National Reference Center of Veterinary and Comparative Oncology (CEROVEC), Piazza Borgo Pila, 16129, Genova (Italy); Thellung, Stefano [Section of Pharmacology, Dept. of Internal Medicine Di.M.I., and Center of Excellence for Biomedical Research - University of Genova, Viale Benedetto XV, 2, 16132 Genova (Italy); Daga, Antonio [Laboratory of Translational Oncology, IRCCS Azienda Ospedaliera Universitaria San Martino - IST- Istituto Nazionale Ricerca sul Cancro, L.go R. Benzi, 10, 16132 Genova Italy (Italy); Cilli, Michele [Animal Facility, IRCCS Azienda Ospedaliera Universitaria San Martino - IST- Istituto Nazionale Ricerca sul Cancro, L.go R. Benzi, 10, 16132 Genova Italy (Italy); Ferrari, Angelo [Istituto Zooprofilattico Sperimentale del Piemonte, Liguria e Valle D' Aosta, National Reference Center of Veterinary and Comparative Oncology (CEROVEC), Piazza Borgo Pila, 16129, Genova (Italy); Florio, Tullio, E-mail: [Section of Pharmacology, Dept. of Internal Medicine Di.M.I., and Center of Excellence for Biomedical Research - University of Genova, Viale Benedetto XV, 2, 16132 Genova (Italy)


    Current carcinogenesis theory states that only a small subset of tumor cells, the cancer stem cells or tumor initiating cells (TICs), are responsible for tumor formation and progression. Human breast cancer-initiating cells have been identified as CD44-expressing cells, which retain tumorigenic activity and display stem cell-like properties. Spontaneous feline mammary carcinoma (FMC) is an aggressive cancer, which shows biological similarities to the human tumor counterpart. We report the isolation and phenotypic characterization of FMC-derived stem/progenitor cells, showing in vitro self-renewal, long-lasting proliferation and in vivo tumorigenicity. Twenty-one FMC samples were collected, histologically classified and characterized for the expression of Ki67, EGFR, ER-{alpha} and CD44, by immunohistochemistry. By culture in stem cell permissive conditions, we isolated, from 13 FMCs, a CD44-positive subpopulation able to survive and proliferate in vitro as mammospheres of different sizes and morphologies. When injected in NOD/SCID mice, FMC stem-like cells initiate tumors, generating cell heterogeneity and recapitulating the original histotype. In serum-containing medium, spheroid cells showed differentiation properties as shown by morphological changes, the loss of CD44 expression and tumorigenic potential. These data show that stem-defined culture of FMC enriches for TICs and validate the use of these cells as a suitable model for comparative oncology studies of mammary biology and testing therapeutic strategies aimed at eradicating TICs. -- Highlights: Black-Right-Pointing-Pointer Feline mammary carcinoma contain a sub-population of stem-like cells expressing CD44 Black-Right-Pointing-Pointer These grow as spheres in serum-free medium and self-renew Black-Right-Pointing-Pointer Isolated stem-like cancer cells initiate tumor in immunodeficient mice Black-Right-Pointing-Pointer Xenografted tumors are phenotypically similar to the original tumor Black

  9. Fluorouracil selectively enriches stem-like cells in the lung adenocarcinoma cell line SPC. (United States)

    Shi, Mu-mu; Xiong, Yan-lei; Jia, Xin-shan; Li, Xin; Zhang, Li; Li, Xiao-lei; Wang, En-Hua


    Most adult stem cells are in the G0 or quiescent phase of the cell cycle and account for only a small percentage of the cells in the tissue. Thus, isolation of stem cells from tissues for further study represents a major challenge. This study sought to enrich cancer stem cells and explore cancer stem-like cell clones using 5-fluorouracil (5-FU) in the lung adenocarcinoma cell line, SPC. Proliferation inhibition was analyzed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays, according to which half maximal inhibitory concentration values were calculated. Expression levels of stem cell markers after treatment with 5-FU were examined using immunofluorescence and Western blotting. Additionally, side population (SP) cells were sorted using FACS. Properties of SP cells were evaluated by using Transwell, colony-forming assays, and tumor formation experiments. 5-FU greatly inhibits proliferation, especially of cells in S phase. SP cells possess greater invasive potential, higher clone-forming potential, and greater tumor-forming ability than non-SP cells. Treatment with 5-FU enriches the SP cells with stem cell properties in human lung adenocarcinoma cell lines.

  10. Natural compounds' activity against cancer stem-like or fast-cycling melanoma cells.

    Directory of Open Access Journals (Sweden)

    Malgorzata Sztiller-Sikorska

    Full Text Available BACKGROUND: Accumulating evidence supports the concept that melanoma is highly heterogeneous and sustained by a small subpopulation of melanoma stem-like cells. Those cells are considered as responsible for tumor resistance to therapies. Moreover, melanoma cells are characterized by their high phenotypic plasticity. Consequently, both melanoma stem-like cells and their more differentiated progeny must be eradicated to achieve durable cure. By reevaluating compounds in heterogeneous melanoma populations, it might be possible to select compounds with activity not only against fast-cycling cells but also against cancer stem-like cells. Natural compounds were the focus of the present study. METHODS: We analyzed 120 compounds from The Natural Products Set II to identify compounds active against melanoma populations grown in an anchorage-independent manner and enriched with cells exerting self-renewing capacity. Cell viability, cell cycle arrest, apoptosis, gene expression, clonogenic survival and label-retention were analyzed. FINDINGS: Several compounds efficiently eradicated cells with clonogenic capacity and nanaomycin A, streptonigrin and toyocamycin were effective at 0.1 µM. Other anti-clonogenic but not highly cytotoxic compounds such as bryostatin 1, siomycin A, illudin M, michellamine B and pentoxifylline markedly reduced the frequency of ABCB5 (ATP-binding cassette, sub-family B, member 5-positive cells. On the contrary, treatment with maytansine and colchicine selected for cells expressing this transporter. Maytansine, streptonigrin, toyocamycin and colchicine, even if highly cytotoxic, left a small subpopulation of slow-dividing cells unaffected. Compounds selected in the present study differentially altered the expression of melanocyte/melanoma specific microphthalmia-associated transcription factor (MITF and proto-oncogene c-MYC. CONCLUSION: Selected anti-clonogenic compounds might be further investigated as potential adjuvants

  11. Evodiamine selectively targets cancer stem-like cells through the p53-p21-Rb pathway

    Energy Technology Data Exchange (ETDEWEB)

    Han, Seula [The Research Center for Cell Fate Control, College of Pharmacy, Sookmyung Women' s University, Seoul (Korea, Republic of); Woo, Jong Kyu [College of Pharmacy, Gachon University, Incheon (Korea, Republic of); Jung, Yuchae; Jeong, Dawoon; Kang, Minsook; Yoo, Young-Ji; Lee, Hani [The Research Center for Cell Fate Control, College of Pharmacy, Sookmyung Women' s University, Seoul (Korea, Republic of); Oh, Seung Hyun [College of Pharmacy, Gachon University, Incheon (Korea, Republic of); Ryu, Jae-Ha [The Research Center for Cell Fate Control, College of Pharmacy, Sookmyung Women' s University, Seoul (Korea, Republic of); Kim, Woo-Young, E-mail: [The Research Center for Cell Fate Control, College of Pharmacy, Sookmyung Women' s University, Seoul (Korea, Republic of)


    In spite of the recent improvements, the resistance to chemotherapy/radiotherapy followed by relapse is the main hurdle for the successful treatment of breast cancer, a leading cause of death in women. A small population of breast cancer cells that have stem-like characteristics (cancer stem-like cells; CSLC) may contribute to this resistance and relapse. Here, we report on a component of a traditional Chinese medicine, evodiamine, which selectively targets CSLC of breast cancer cell lines MCF7 and MDAMB 231 at a concentration that does show a little or no cytotoxic effect on bulk cancer cells. While evodiamine caused the accumulation of bulk cancer cells at the G2/M phase, it did not hold CSLC in a specific cell cycle phase but instead, selectively killed CSLC. This was not due to the culture of CSLC in suspension or without FBS. A proteomic analysis and western blotting revealed that evodiamine changed the expression of cell cycle regulating molecules more efficiently in CSLC cells than in bulk cancer cells. Surprisingly, evodiamine selectively activated p53 and p21 and decreased inactive Rb, the master molecules in G1/S checkpoint. These data collectively suggest a novel mechanism involving CSLC-specific targeting by evodiamine and its possible use to the therapy of breast cancer. - Highlights: • Evodiamine selectively kills breast cancer stem like cells at G1 phase. • Evodiamine utilizes different mechanism of cell cycle modulation in CSLC and in bulk cancer cells. • Evodiamine activate the p53, p21 and Rb pathway.

  12. Evodiamine selectively targets cancer stem-like cells through the p53-p21-Rb pathway. (United States)

    Han, Seula; Woo, Jong Kyu; Jung, Yuchae; Jeong, Dawoon; Kang, Minsook; Yoo, Young-Ji; Lee, Hani; Oh, Seung Hyun; Ryu, Jae-Ha; Kim, Woo-Young


    In spite of the recent improvements, the resistance to chemotherapy/radiotherapy followed by relapse is the main hurdle for the successful treatment of breast cancer, a leading cause of death in women. A small population of breast cancer cells that have stem-like characteristics (cancer stem-like cells; CSLC) may contribute to this resistance and relapse. Here, we report on a component of a traditional Chinese medicine, evodiamine, which selectively targets CSLC of breast cancer cell lines MCF7 and MDAMB 231 at a concentration that does show a little or no cytotoxic effect on bulk cancer cells. While evodiamine caused the accumulation of bulk cancer cells at the G2/M phase, it did not hold CSLC in a specific cell cycle phase but instead, selectively killed CSLC. This was not due to the culture of CSLC in suspension or without FBS. A proteomic analysis and western blotting revealed that evodiamine changed the expression of cell cycle regulating molecules more efficiently in CSLC cells than in bulk cancer cells. Surprisingly, evodiamine selectively activated p53 and p21 and decreased inactive Rb, the master molecules in G1/S checkpoint. These data collectively suggest a novel mechanism involving CSLC-specific targeting by evodiamine and its possible use to the therapy of breast cancer.

  13. Isolation and characterization of cancer stem-like cells from MHCC97H Cell Lines

    Institute of Scientific and Technical Information of China (English)

    Shanyong Yi; Kejun Nan; Aihua Yuan; Chuangxin Lu


    Objective:To identify and isolate CD133 positive cancer stem-like cells (CD133+ cells) from the highly invasive human hepatocellular carcinoma cell line(MHCC97H), and examine their potential for clonogenicity and tumorigenicity. Methods: CD133+ and CD133- cells were isolated from MHCC97H cell line by magnetic bead cell sorting(MACS), and the potentials of CD133+ cells for colony formation and tumorigenicity were evaluated by soft agar cloning and tumor formation following nude mice inoculation. Results:CD133+ cells represent a minority(0.5-2.0%) of the tumor cell population with a greater colony-forming efficiency and greater tumor production ability. The colony-forming efficiency of CD133+ cells in soft agar was significantly higher than CD133- cells(36.8±1.4 vs 12.9±0.8, P<0.05).After 6 weeks, 3/5 mice inoculated with 1 × 103 CD133+ cells, 4/5 with 1 × 104 CD133+ cells and 5/5 with 1 × 105 CD133+ cells developed detectable tumors at the injection site, while only one tumor was found in mice treated with same numbers of CD133- cells. Conclusion: CD133 may be a hallmark of liver cancer stem cells (CSC) in human hepatocellular carcinoma(HCC), because the CD133+ cells identified and isolated with anti-CD133 labeled magnetic beads from MHCC97H cell line exhibit high potentials for clonogenicity and tumorigenicity. These CD133+ cells might contribute to hepatocarcinogenesis, as well as the growth and recurrence of human HCC, and therefore may be a useful target for anti-cancer therapy.

  14. The chemokine receptor CCR7 promotes mammary tumorigenesis through amplification of stem-like cells. (United States)

    Boyle, S T; Ingman, W V; Poltavets, V; Faulkner, J W; Whitfield, R J; McColl, S R; Kochetkova, M


    The chemokine receptor CCR7 is widely implicated in breast cancer pathobiology. Although recent reports correlated high CCR7 levels with more advanced tumor grade and poor prognosis, limited in vivo data are available regarding its specific function in mammary gland neoplasia and the underlying mechanisms involved. To address these questions we generated a bigenic mouse model of breast cancer combined with CCR7 deletion, which revealed that CCR7 ablation results in a considerable delay in tumor onset as well as significantly reduced tumor burden. Importantly, CCR7 was found to exert its function by regulating mammary cancer stem-like cells in both murine and human tumors. In vivo experiments showed that loss of CCR7 activity either through deletion or pharmacological antagonism significantly decreased functional pools of stem-like cells in mouse primary mammary tumors, providing a mechanistic explanation for the tumor-promoting role of this chemokine receptor. These data characterize the oncogenic properties of CCR7 in mammary epithelial neoplasia and point to a new route for therapeutic intervention to target evasive cancer stem cells.

  15. Combination therapy targeting both cancer stem-like cells and bulk tumor cells for improved efficacy of breast cancer treatment. (United States)

    Wang, Tao; Narayanaswamy, Radhika; Ren, Huilan; Torchilin, Vladimir P


    Many types of tumors are organized in a hierarchy of heterogeneous cell populations. The cancer stem-like cells (CSCs) hypothesis suggests that tumor development and metastasis are driven by a minority population of cells, which are responsible for tumor initiation, growth and recurrences. The inability to efficiently eliminate CSCs during chemotherapy, together with CSCs being highly tumorigenic and invasive, may result in treatment failure due to cancer relapse and metastases. CSCs are emerging as a promising target for the development of translational cancer therapies. Ideal panacea for cancer would kill all malignant cells, including CSCs and bulk tumor cells. Since both chemotherapy and CSCs-specific therapy are insufficient to cure cancer, we propose combination therapy with CSCs-targeted agents and chemotherapeutics for improved breast cancer treatment. We generated in vitro mammosphere of 2 breast cancer cell lines, and demonstrated ability of mammospheres to grow and enrich cancer cells with stem-like properties, including self-renewal, multilineage differentiation and enrichment of cells expressing breast cancer stem-like cell biomarkers CD44(+)/CD24(-/low). The formation of mammospheres was significantly inhibited by salinomycin, validating its pharmacological role against the cancer stem-like cells. In contrast, paclitaxel showed a minimal effect on the proliferation and growth of breast cancer stem-like cells. While combination therapies of salinomycin with conventional chemotherapy (paclitaxel or lipodox) showed a potential to improve tumor cell killing, different subtypes of breast cancer cells showed different patterns in response to the combination therapies. While optimization of combination therapy is warranted, the design of combination therapy should consider phenotypic attributes of breast cancer types.

  16. Ursolic acid inhibits the proliferation of human ovarian cancer stem-like cells through epithelial-mesenchymal transition. (United States)

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


    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.

  17. Characterization of cancer stem-like cells in the side population cells of human gastric cancer cell line MKN-45

    Institute of Scientific and Technical Information of China (English)

    Hai-hong ZHANG; Ai-zhen CAI; Xue-ming WEI; Li DING; Feng-zhi LI; Ai-ming ZHENG; Da-jiang DAI


    Objective:Side population (SP) cells may play a crucial role in tumorigenesis and the recurrence of cancer.Many kinds of cell lines and tissues have demonstrated the presence of SP cells,including several gastric cancer cell lines.This study is aimed to identify the cancer stem-like cells in the SP of gastric cancer cell line MKN-45.Methods:We used fluorescence activated cell sorting (FACS) to sort SP cells in the human gastric carcinoma cell line MKN-45 (cells labeled with Hoechst 33342) and then characterized the cancer stem-like properties of SP cells.Results:This study found that the SP cells had higher clone formation efficiency than major population (MP) cells.Five stemness-related gene expression profiles,including OCT-4,SOX-2,NANOG,CD44,and adenosine triphosphate (ATP)-binding cassette transporters gene ABCG2,were tested in SP and MP cells using quantitative real-time reverse transcription polymerase chain reaction (RT-PCR).Western blot was used to show the difference of protein expression between SP and MP cells.Both results show that there was significantly higher protein expression in SP cells than in MP cells.When inoculated into non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice,SP cells show higher tumorigenesis tendency than MP cells.Conclusions:These results indicate that SP cells possess cancer stem cell properties and prove that SP cells from MKN-45 are gastric cancer stem-like cells.

  18. Notch4+ cancer stem-like cells promote the metastatic and invasive ability of melanoma. (United States)

    Lin, Xian; Sun, Baocun; Zhu, Dongwang; Zhao, Xiulan; Sun, Ran; Zhang, Yanhui; Zhang, Danfang; Dong, Xueyi; Gu, Qiang; Li, Yanlei; Liu, Fang


    Sphere formation in conditioned serum-free culture medium supplemented with epidermal growth factor and basic fibroblast growth factor (tumorospheres) is considered useful for the enrichment of cancer stem-like cells, also known as tumor-initiating cells. We used a gene expression microarray to investigate the gene expression profile of melanoma cancer stem-like cells (MCSLCs). The results showed that MCSLCs highly expressed the following Notch signaling pathway molecules: Notch3 (NM_008716), Notch4 (NM_010929), Dtx4 (NM_172442), and JAG2 (NM_010588). Immunofluorescence staining showed tumorosphere cells highly expressed Notch4. Notch4(high) B16F10 cells were isolated by FACS, and Western blotting showed that high Notch4 expression is related to the expression of epithelial-mesenchymal transition (EMT)-associated proteins. Reduced invasive and migratory properties concomitant with the downregulation of the EMT markers Twist1, vimentin, and VE-cadherin and the overexpression of E-cadherin was observed in human melanoma A375 and MUM-2B cells. In these cells, Notch4 was also downregulated, both by Notch4 gene knockdown and by application of the γ-secretase inhibitor, DAPT. Mechanistically, the re-overexpression of Twist1 by the transfection of cells with a Twist1 expression plasmid led to an increase in VE-cadherin expression and a decrease in E-cadherin expression. Immunohistochemical analysis of 120 human melanoma tissues revealed a significant correlation between the high expression of Notch4 and the metastasis of melanoma. Taken together, our findings indicate that Notch4+ MCSLCs trigger EMT and promote the metastasis of melanoma cells.

  19. Canine mammary tumors contain cancer stem-like cells and form spheroids with an embryonic stem cell signature. (United States)

    Ferletta, Maria; Grawé, Jan; Hellmén, Eva


    We have investigated the presence of tentative stem-like cells in the canine mammary tumor cell line CMT-U229. This cell line is established from an atypical benign mixed mammary tumor, which has the property of forming duct-like structures in collagen gels. Stem cells in mammary glands are located in the epithelium; therefore we thought that the CMT-U229 cell line would be suitable for detection of tentative cancer stem-like cells. Side population (SP) analyses by flow cytometry were performed with cells that formed spheroids and with cells that did not. Flow cytometric, single sorted cells were expanded and re-cultured as spheroids. The spheroids were paraffin embedded and characterized by immunohistochemistry. SP analyses showed that spheroid forming cells (retenate) as well as single cells (filtrate) contained SP cells. Sca1 positive cells were single cell sorted and thereafter the SP population increased with repeated SP analyses. The SP cells were positively labeled with the cell surface-markers CD44 and CD49f (integrin alpha6); however the expression of CD24 was low or negative. The spheroids expressed the transcription factor and stem cell marker Sox2, as well as Oct4. Interestingly, only peripheral cells of the spheroids and single cells were positive for Oct4 expression. SP cells are suggested to correspond to stem cells and in this study, we have enriched for tentative tumor stem-like cells derived from a canine mammary tumor. All the used markers indicate that the studied CMT-U229 cell line contains SP cells, which in particular have cancer stem-like cell characteristics.

  20. Nanomedicine to overcome radioresistance in glioblastoma stem-like cells and surviving clones. (United States)

    Séhédic, Delphine; Cikankowitz, Annabelle; Hindré, François; Davodeau, François; Garcion, Emmanuel


    Radiotherapy is one of the standard treatments for glioblastoma, but its effectiveness often encounters the phenomenon of radioresistance. This resistance was recently attributed to distinct cell contingents known as glioblastoma stem-like cells (GSCs) and dominant clones. It is characterized in particular by the activation of signaling pathways and DNA repair mechanisms. Recent advances in the field of nanomedicine offer new possibilities for radiosensitizing these cell populations. Several strategies have been developed in this direction, the first consisting of encapsulating a contrast agent or synthesizing metal-based nanocarriers to concentrate the dose gradient at the level of the target tissue. In the second strategy the physicochemical properties of the vectors are used to encapsulate a wide range of pharmacological agents which act in synergy with the ionizing radiation to destroy the cancerous cells. This review reports on the various molecular anomalies present in GSCs and the predominant role of nanomedicines in the development of radiosensitization strategies.

  1. Podocalyxin-like protein is expressed in glioblastoma multiforme stem-like cells and is associated with poor outcome.

    Directory of Open Access Journals (Sweden)

    Zev A Binder

    Full Text Available Glioblastoma multiforme (GBM is the most common primary malignant adult brain tumor and is associated with poor survival. Recently, stem-like cell populations have been identified in numerous malignancies including GBM. To identify genes whose expression is changed with differentiation, we compared transcript profiles from a GBM oncosphere line before and after differentiation. Bioinformatic analysis of the gene expression profiles identified podocalyxin-like protein (PODXL, a protein highly expressed in human embryonic stem cells, as a potential marker of undifferentiated GBM stem-like cells. The loss of PODXL expression upon differentiation of GBM stem-like cell lines was confirmed by quantitative real-time PCR and flow cytometry. Analytical flow cytometry of numerous GBM oncosphere lines demonstrated PODXL expression in all lines examined. Knockdown studies and flow cytometric cell sorting experiments demonstrated that PODXL is involved in GBM stem-like cell proliferation and oncosphere formation. Compared to PODXL-negative cells, PODXL-positive cells had increased expression of the progenitor/stem cell markers Musashi1, SOX2, and BMI1. Finally, PODXL expression directly correlated with increasing glioma grade and was a marker for poor outcome in patients with GBM. In summary, we have demonstrated that PODXL is expressed in GBM stem-like cells and is involved in cell proliferation and oncosphere formation. Moreover, high PODXL expression correlates with increasing glioma grade and decreased overall survival in patients with GBM.

  2. Drug delivery using nanoparticles for cancer stem-like cell targeting

    Directory of Open Access Journals (Sweden)

    Bing eLu


    Full Text Available The theory of cancer stem-like cell (or cancer stem cell, CSC has been established to explain how tumor heterogeneity arises and contributes to tumor progression in diverse cancer types. CSCs are believed to drive tumor growth and elicit resistance to conventional therapeutics. Therefore, CSCs are becoming novel target in both medical researches and clinical studies. Emerging evidences showed that nanoparticles effectively inhibit many types of CSCs by targeting various specific markers (aldehyde dehydrogenases, CD44, CD90, and CD133 and signaling pathways (Notch, Hedgehog, and TGF-β, which are critically involved in CSC function and maintenance. In this review, we briefly summarize the current status of CSC research and review a number of state-of-the-art nanomedicine approaches targeting CSC. In addition, we discuss emerging therapeutic strategies using epigenetic drugs to eliminate CSCs and inhibit cancer cell reprogramming.

  3. Hypoxia in the glioblastoma microenvironment: shaping the phenotype of cancer stem-like cells. (United States)

    Colwell, Nicole; Larion, Mioara; Giles, Amber J; Seldomridge, Ashlee N; Sizdahkhani, Saman; Gilbert, Mark R; Park, Deric M


    Glioblastoma is the most common and aggressive malignant primary brain tumor. Cellular heterogeneity is a characteristic feature of the disease and contributes to the difficulty in formulating effective therapies. Glioma stem-like cells (GSCs) have been identified as a subpopulation of tumor cells that are thought to be largely responsible for resistance to treatment. Intratumoral hypoxia contributes to maintenance of the GSCs by supporting the critical stem cell traits of multipotency, self-renewal, and tumorigenicity. This review highlights the interaction of GSCs with the hypoxic tumor microenvironment, exploring the mechanisms underlying the contribution of GSCs to tumor vessel dynamics, immune modulation, and metabolic alteration. Published by Oxford University Press on behalf of the Society for Neuro-Oncology 2017. This work is written by (a) US Government employee(s) and is in the public domain in the US.

  4. Myeloid-Derived Suppressor Cells Endow Stem-like Qualities to Breast Cancer Cells through IL6/STAT3 and NO/NOTCH Cross-talk Signaling. (United States)

    Peng, Dongjun; Tanikawa, Takashi; Li, Wei; Zhao, Lili; Vatan, Linda; Szeliga, Wojciech; Wan, Shanshan; Wei, Shuang; Wang, Yin; Liu, Yan; Staroslawska, Elzbieta; Szubstarski, Franciszek; Rolinski, Jacek; Grywalska, Ewelina; Stanisławek, Andrzej; Polkowski, Wojciech; Kurylcio, Andrzej; Kleer, Celina; Chang, Alfred E; Wicha, Max; Sabel, Michael; Zou, Weiping; Kryczek, Ilona


    Myeloid-derived suppressor cells (MDSC) contribute to immune suppression in cancer, but the mechanisms through which they drive metastatic progression are not fully understood. In this study, we show how MDSC convey stem-like qualities to breast cancer cells that coordinately help enable immune suppression and escape. We found that MDSC promoted tumor formation by enhancing breast cancer cell stem-like properties as well as by suppressing T-cell activation. Mechanistic investigations indicated that these effects relied upon cross-talk between the STAT3 and NOTCH pathways in cancer cells, with MDSC inducing IL6-dependent phosphorylation of STAT3 and activating NOTCH through nitric oxide leading to prolonged STAT3 activation. In clinical specimens of breast cancer, the presence of MDSC correlated with the presence of cancer stem-like cells (CSC) and independently predicted poor survival outcomes. Collectively, our work revealed an immune-associated mechanism that extrinsically confers cancer cell stemness properties and affects patient outcome. We suggest that targeting STAT3-NOTCH cross-talk between MDSC and CSC could offer a unique locus to improve cancer treatment, by coordinately targeting a coupled mechanism that enables cancer stemness and immune escape. Cancer Res; 76(11); 3156-65. ©2016 AACR.

  5. Isolation and characterization of lymphoid enhancer factor-1-positive deciduous dental pulp stem-like cells after transfection with a piggyBac vector containing LEF1 promoter-driven selection markers. (United States)

    Murakami, Tomoya; Saitoh, Issei; Sato, Masahiro; Inada, Emi; Soda, Miki; Oda, Masataka; Domon, Hisanori; Iwase, Yoko; Sawami, Tadashi; Matsueda, Kazunari; Terao, Yutaka; Ohshima, Hayato; Noguchi, Hirofumi; Hayasaki, Haruaki


    Lymphoid enhancer-binding factor-1 (LEF1) is a 48-kD nuclear protein that is expressed in pre-B and T cells. LEF1 is also an important member of the Wnt/β-catenin signaling pathway that plays important roles in the self-renewal and differentiation of embryonic stem cells. We speculated that LEF1 might function in the stem cells from human exfoliated deciduous teeth (SHED). In this study, we attempted to isolate such LEF1-positive cells from human deciduous dental pulp cells (HDDPCs) by genetic engineering technology, using the human LEF1 promoter. A piggyBac transposon plasmid (pTA-LEN) was introduced into HDDPCs, using the Neon(®) transfection system. After G418 selection, the emerging colonies were assessed for EGFP-derived fluorescence by fluorescence microscopy. Reverse transcription polymerase chain reaction (RT-PCR) analysis was performed using RNA isolated from these colonies to examine stem cell-specific transcript expression. Osteoblastic or neuronal differentiation was induced by cultivating the LEF1-positive cells with differentiation-inducing medium. RT-PCR analysis confirmed the expression of several stem cell markers, including OCT3/4, SOX2, REX1, and NANOG, in LEF1-positive HDDPCs, which could be differentiated into osteoblasts and neuronal cells. The isolated LEF1-positive HDDPCs exhibited the properties of stem cells, suggesting that LEF1 might serve as a marker for SHED. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  6. The Impact of the Tumor Microenvironment on the Properties of Glioma Stem-Like Cells. (United States)

    Audia, Alessandra; Conroy, Siobhan; Glass, Rainer; Bhat, Krishna P L


    Glioblastoma is the most common and highly malignant primary brain tumor, and patients affected with this disease exhibit a uniformly dismal prognosis. Glioma stem-like cells (GSCs) are a subset of cells within the bulk tumor that possess self-renewal and multi-lineage differentiation properties similar to somatic stem cells. These cells also are at the apex of the cellular hierarchy and cause tumor initiation and expansion after chemo-radiation. These traits make them an attractive target for therapeutic development. Because GSCs are dependent on the brain microenvironment for their growth, and because non-tumorigenic cell types in the microenvironment can influence GSC phenotypes and treatment response, a better understanding of these cell types is needed. In this review, we provide a focused overview of the contributions from the microenvironment to GSC homing, maintenance, phenotypic plasticity, and tumor initiation. The interaction of GSCs with the vascular compartment, mesenchymal stem cells, immune system, and normal brain cell types are discussed. Studies that provide mechanistic insight into each of these GSC-microenvironment interactions are warranted in the future.

  7. Notch signals in the endothelium and cancer "stem-like" cells: opportunities for cancer therapy

    Directory of Open Access Journals (Sweden)

    Gu Jian-Wei


    Full Text Available Abstract Anti-angiogenesis agents and the identification of cancer stem-like cells (CSC are opening new avenues for targeted cancer therapy. Recent evidence indicates that angiogenesis regulatory pathways and developmental pathways that control CSC fate are intimately connected, and that endothelial cells are a key component of the CSC niche. Numerous anti-angiogenic therapies developed so far target the VEGF pathway. However, VEGF-targeted therapy is hindered by clinical resistance and side effects, and new approaches are needed. One such approach may be direct targeting of tumor endothelial cell fate determination. Interfering with tumor endothelial cells growth and survival could inhibit not only angiogenesis but also the self-replication of CSC, which relies on signals from surrounding endothelial cells in the tumor microenvironment. The Notch pathway is central to controlling cell fate both during angiogenesis and in CSC from several tumors. A number of investigational Notch inhibitors are being developed. Understanding how Notch interacts with other factors that control endothelial cell functions and angiogenesis in cancers could pave the way to innovative therapeutic strategies that simultaneously target angiogenesis and CSC.

  8. Natural killer cell-based adoptive immunotherapy eradicates and drives differentiation of chemoresistant bladder cancer stem-like cells. (United States)

    Ferreira-Teixeira, Margarida; Paiva-Oliveira, Daniela; Parada, Belmiro; Alves, Vera; Sousa, Vitor; Chijioke, Obinna; Münz, Christian; Reis, Flávio; Rodrigues-Santos, Paulo; Gomes, Célia


    High-grade non-muscle invasive bladder cancer (NMIBC) has a high risk of recurrence and progression to muscle-invasive forms, which seems to be largely related to the presence of tumorigenic stem-like cell populations that are refractory to conventional therapies. Here, we evaluated the therapeutic potential of Natural Killer (NK) cell-based adoptive immunotherapy against chemoresistant bladder cancer stem-like cells (CSCs) in a pre-clinical relevant model, using NK cells from healthy donors and NMIBC patients. Cytokine-activated NK cells from healthy donors and from high-grade NMIBC patients were phenotypically characterized and assayed in vitro against stem-like and bulk differentiated bladder cancer cells. Stem-like cells were isolated from two bladder cancer cell lines using the sphere-forming assay. The in vivo therapeutic efficacy was evaluated in mice bearing a CSC-induced orthotopic bladder cancer. Animals were treated by intravesical instillation of interleukin-activated NK cells. Tumor response was evaluated longitudinally by non-invasive bioluminescence imaging. NK cells from healthy donors upon activation with IL-2 and IL-15 kills indiscriminately both stem-like and differentiated tumor cells via stress ligand recognition. In addition to cell killing, NK cells shifted CSCs towards a more differentiated phenotype, rendering them more susceptible to cisplatin, highlighting the benefits of a possible combined therapy. On the contrary, NK cells from NMIBC patients displayed a low density on NK cytotoxicity receptors, adhesion molecules and a more immature phenotype, losing their ability to kill and drive differentiation of CSCs. The local administration, via the transurethral route, of activated NK cells from healthy donors provides an efficient tumor infiltration and a subsequent robust tumoricidal activity against bladder cancer with high selective cytolytic activity against CSCs, leading to a dramatic reduction in tumor burden from 80 % to complete

  9. Embryonic stem-like cells derived from in vitro produced bovine blastocysts

    Directory of Open Access Journals (Sweden)

    Erika Regina Leal de Freitas


    Full Text Available The aim of this work was to study the derivation of bovine embryonic stem-like (ES-like cells from the inner cell mass (ICM of in vitro produced blastocysts. The ICMs were mechanically isolated and six out of seventeen (35% ICMs could attach to a monolayer of murine embryonic fibroblasts (MEF. Ten days after, primary outgrowths were mechanically dissected into several small clumps and transferred to a new MEF layer. Cells were further propagated and passaged by physical dissociation over a 60 days period. The pluripotency of the bovine ES-like cells was confirmed by RT-PCR of Oct-4 and STAT-3 gene markers. The colonies were weakly stained for alkaline phosphatase and the mesoderm and endoderm differentiation gene markers such as GATA-4 and Flk-1, respectively, were not expressed. Embryoid bodies were spontaneously formed at the seventh passage. Results showed that bovine ES-like cells could be obtained and passaged by mechanical procedures from the fresh in vitro produced blastocysts.

  10. Survival, proliferation, and migration of human meningioma stem-like cells in a nanopeptide scaffold

    Directory of Open Access Journals (Sweden)

    Sajad Sahab Negah


    Full Text Available Objective(s: In order to grow cells in a three-dimensional (3D microenvironment, self-assembling peptides, such as PuraMatrix, have emerged with potential to mimic the extracellular matrix. The aim of the present study was to investigate the influence of the self-assembling peptide on the morphology, survival, proliferation rate, migration potential, and differentiation of human meningioma stem-like cells (hMgSCs. Materials and Methods: The efficacy of a novel method for placing hMgSCs in PuraMatrix (the injection approach was compared to the encapsulation and surface plating methods. In addition, we designed a new method for measurement of migration distance in 3D cultivation of hMgSCs in PuraMatrix. Results: Our results revealed that hMgSCs have the ability to form spheres in stem cell culture condition. These meningioma cells expressed GFAP, CD133, vimentin, and nestin. Using the injection method, a higher proliferation rate of the hMgSCs was observed after seven days of culture. Furthermore, the novel migration assay was able to measure the migration of a single cell alone in 3D environment. Conclusion: The results indicate the injection method as an efficient technique for culturing hMgSCs in PuraMatrix. Furthermore, the novel migration assay enables us to evaluate the migration of hMgSCs.

  11. Identification of quiescent, stem-like cells in the distal female reproductive tract.

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    Yongyi Wang

    Full Text Available In fertile women, the endometrium undergoes regular cycles of tissue build-up and regression. It is likely that uterine stem cells are involved in this remarkable turn over. The main goal of our current investigations was to identify slow-cycling (quiescent endometrial stem cells by means of a pulse-chase approach to selectively earmark, prospectively isolate, and characterize label-retaining cells (LRCs. To this aim, transgenic mice expressing histone2B-GFP (H2B-GFP in a Tet-inducible fashion were administered doxycycline (pulse which was thereafter withdrawn from the drinking water (chase. Over time, dividing cells progressively loose GFP signal whereas infrequently dividing cells retain H2B-GFP expression. We evaluated H2B-GFP retaining cells at different chase time points and identified long-term (LT; >12 weeks LRCs. The LT-LRCs are negative for estrogen receptor-α and express low levels of progesterone receptors. LRCs sorted by FACS are able to form spheroids capable of self-renewal and differentiation. Upon serum stimulation spheroid cells are induced to differentiate and form glandular structures which express markers of mature műllerian epithelial cells. Overall, the results indicate that quiescent cells located in the distal oviduct have stem-like properties and can differentiate into distinct cell lineages specific of endometrium, proximal and distal oviduct. Future lineage-tracing studies will elucidate the role played by these cells in homeostasis, tissue injury and cancer of the female reproductive tract in the mouse and eventually in man.

  12. Targeting cancer stem-like cells in glioblastoma and colorectal cancer through metabolic pathways. (United States)

    Kahlert, U D; Mooney, S M; Natsumeda, M; Steiger, H-J; Maciaczyk, J


    Cancer stem-like cells (CSCs) are thought to be the main cause of tumor occurrence, progression and therapeutic resistance. Strong research efforts in the last decade have led to the development of several tailored approaches to target CSCs with some very promising clinical trials underway; however, until now no anti-CSC therapy has been approved for clinical use. Given the recent improvement in our understanding of how onco-proteins can manipulate cellular metabolic networks to promote tumorigenesis, cancer metabolism research may well lead to innovative strategies to identify novel regulators and downstream mediators of CSC maintenance. Interfering with distinct stages of CSC-associated metabolics may elucidate novel, more efficient strategies to target this highly malignant cell population. Here recent discoveries regarding the metabolic properties attributed to CSCs in glioblastoma (GBM) and malignant colorectal cancer (CRC) were summarized. The association between stem cell markers, the response to hypoxia and other environmental stresses including therapeutic insults as well as developmentally conserved signaling pathways with alterations in cellular bioenergetic networks were also discussed. The recent developments in metabolic imaging to identify CSCs were also summarized. This summary should comprehensively update basic and clinical scientists on the metabolic traits of CSCs in GBM and malignant CRC. © 2016 UICC.

  13. Automatic cell cloning assay for determining the clonogenic capacity of cancer and cancer stem-like cells. (United States)

    Fedr, Radek; Pernicová, Zuzana; Slabáková, Eva; Straková, Nicol; Bouchal, Jan; Grepl, Michal; Kozubík, Alois; Souček, Karel


    The clonogenic assay is a well-established in vitro method for testing the survival and proliferative capability of cells. It can be used to determine the cytotoxic effects of various treatments including chemotherapeutics and ionizing radiation. However, this approach can also characterize cells with different phenotypes and biological properties, such as stem cells or cancer stem cells. In this study, we implemented a faster and more precise method for assessing the cloning efficiency of cancer stem-like cells that were characterized and separated using a high-speed cell sorter. Cell plating onto a microplate using an automatic cell deposition unit was performed in a single-cell or dilution rank mode by the fluorescence-activated cell sorting method. We tested the new automatic cell-cloning assay (ACCA) on selected cancer cell lines and compared it with the manual approach. The obtained results were also compared with the results of the limiting dilution assay for different cell lines. We applied the ACCA to analyze the cloning capacity of different subpopulations of prostate and colon cancer cells based on the expression of the characteristic markers of stem (CD44 and CD133) and cancer stem cells (TROP-2, CD49f, and CD44). Our results revealed that the novel ACCA is a straightforward approach for determining the clonogenic capacity of cancer stem-like cells identified in both cell lines and patient samples. Copyright © 2013 International Society for Advancement of Cytometry.

  14. Adoptive transfer of osteoclast-expanded natural killer cells for immunotherapy targeting cancer stem-like cells in humanized mice. (United States)

    Kozlowska, Anna K; Kaur, Kawaljit; Topchyan, Paytsar; Jewett, Anahid


    Based on data obtained from oral, pancreatic and lung cancers, glioblastoma, and melanoma, we have established that natural killer (NK) cells target cancer stem-like cells (CSCs). CSCs displaying low MHC class I, CD54, and PD-L1 are killed by cytotoxic NK cells and are differentiated by split anergized NK cells through both membrane bound and secreted forms of TNF-α and IFN-γ. NK cells select and differentiate both healthy and transformed stem-like cells, resulting in target cell maturation and shaping of their microenvironment. In our recent studies, we have observed that oral, pancreatic, and melanoma CSCs were capable of forming large tumors in humanized bone marrow, liver, thymus (hu-BLT) mice with fully reconstituted human immune system. In addition, major human immune subsets including NK cells, T cells, B cells, and monocytes were present in the spleen, bone marrow, peripheral blood, and tumor microenvironment. Similar to our previously published in vitro data, CSCs differentiated with split anergized NK cells prior to implantation in mice formed smaller tumors. Intravenous injection of functionally potent osteoclast-expanded NK cells inhibited tumor growth through differentiation of CSCs in humanized mice. In this review, we present current approaches, advances, and existing limitations in studying interactions of the immune system with the tumor, in particular NK cells with CSCs, using in vivo preclinical hu-BLT mouse model. In addition, we discuss the use of osteoclast-expanded NK cells in targeting cancer stem-like tumors in humanized mice-a strategy that provides a much-needed platform to develop effective cancer immunotherapies.

  15. Accumulation efficiency of cancer stem-like cells post {gamma}-ray and proton irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Quan Yi; WangWeikang; Fu Qibin; Mei Tao; Wu Jingwen; Li Jia [State Key Laboratory of Nuclear Physics and Technology, Peking University, Beijing 100871 (China); Yang, Gen, E-mail: [State Key Laboratory of Nuclear Physics and Technology, Peking University, Beijing 100871 (China); Wang Yugang [State Key Laboratory of Nuclear Physics and Technology, Peking University, Beijing 100871 (China)


    Ionizing radiation (IR) has been proven to be a powerful medical treatment in cancer therapy. Rational and effective use of its killing power depends on understanding IR-mediated responses at the molecular, cellular and tissue levels. Increasing evidence supports that cancer stem-like cells (CSCs) play an important role in tumor regrowth and spread post radiotherapy, for they are resistant to various therapy methods including radiation. Presently, SW620 colon carcinoma monolayer culture cells were irradiated with {gamma}-rays and protons of 2 Gy. Then apoptosis, clonogenic survival and the expression of CD133{sup +} protein were examined. The results showed that there was no significantly difference either on long-term clonogenic survival or on short-term apoptosis ratio. However, compared with {gamma}-rays, irradiation with protons was less efficient to accumulate CSCs at the same dose, although both protons and {gamma}-rays can significantly accumulate the CD133{sup +} CSCs subpopulation. In addition, the results of sphere formation assay also confirmed that proton irradiation is less efficient in CSCs accumulation, suggesting proton irradiation might have higher efficiency in CSCs elimination for cancer radiotherapy.

  16. Prostate Cancer Stem-like Cells Contribute to the Development of Castration-Resistant Prostate Cancer

    Directory of Open Access Journals (Sweden)

    Diane Ojo


    Full Text Available Androgen deprivation therapy (ADT has been the standard care for patients with advanced prostate cancer (PC since the 1940s. Although ADT shows clear benefits for many patients, castration-resistant prostate cancer (CRPC inevitably occurs. In fact, with the two recent FDA-approved second-generation anti-androgens abiraterone and enzalutamide, resistance develops rapidly in patients with CRPC, despite their initial effectiveness. The lack of effective therapeutic solutions towards CRPC largely reflects our limited understanding of the underlying mechanisms responsible for CRPC development. While persistent androgen receptor (AR signaling under castration levels of serum testosterone (<50 ng/mL contributes to resistance to ADT, it is also clear that CRPC evolves via complex mechanisms. Nevertheless, the physiological impact of individual mechanisms and whether these mechanisms function in a cohesive manner in promoting CRPC are elusive. In spite of these uncertainties, emerging evidence supports a critical role of prostate cancer stem-like cells (PCSLCs in stimulating CRPC evolution and resistance to abiraterone and enzalutamide. In this review, we will discuss the recent evidence supporting the involvement of PCSLC in CRPC acquisition as well as the pathways and factors contributing to PCSLC expansion in response to ADT.

  17. Transforming growth factor-beta1 promotes the migration and invasion of sphere-forming stem-like cell subpopulations in esophageal cancer

    Energy Technology Data Exchange (ETDEWEB)

    Yue, Dongli; Zhang, Zhen; Li, Jieyao; Chen, Xinfeng [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); Department of Oncology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Ping, Yu; Liu, Shasha [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); School of Life Sciences, Zhengzhou University, Zhengzhou 450000 (China); Shi, Xiaojuan; Li, Lifeng [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); Department of Oncology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Wang, Liping [Department of Oncology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Huang, Lan [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); Zhang, Bin [Biotherapy Center, the First Affiliated Hospital of Zhengzhou University, No. 1 Jianshe Road, Zhengzhou 450052, Henan, PR China (China); Robert H. Lurie Comprehensive Cancer Center, Department of Medicine-Division of Hematology/Oncology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611 (United States); Sun, Yan [Department of Oncology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Department of Medical Oncology, Cancer Hospital, Chinese Academy of Medical Sciences (China); and others


    Esophageal cancer is one of the most lethal solid malignancies. Mounting evidence demonstrates that cancer stem cells (CSCs) are able to cause tumor initiation, metastasis and responsible for chemotherapy and radiotherapy failures. As CSCs are thought to be the main reason of therapeutic failure, these cells must be effectively targeted to elicit long-lasting therapeutic responses. We aimed to enrich and identify the esophageal cancer cell subpopulation with stem-like properties and help to develop new target therapy strategies for CSCs. Here, we found esophageal cancer cells KYSE70 and TE1 could form spheres in ultra low attachment surface culture and be serially passaged. Sphere-forming cells could redifferentiate and acquire morphology comparable to parental cells, when return to adherent culture. The sphere-forming cells possessed the key criteria that define CSCs: persistent self-renewal, overexpression of stemness genes (SOX2, ALDH1A1 and KLF4), reduced expression of differentiation marker CK4, chemoresistance, strong invasion and enhanced tumorigenic potential. SB525334, transforming growth factor-beta 1(TGF-β1) inhibitor, significantly inhibited migration and invasion of sphere-forming stem-like cells and had no effect on sphere-forming ability. In conclusion, esophageal cancer sphere-forming cells from KYSE70 and TE1 cultured in ultra low attachment surface possess cancer stem cell properties, providing a model for CSCs targeted therapy. TGF-β1 promotes the migration and invasion of sphere-forming stem-like cells, which may guide future studies on therapeutic strategies targeting these cells. - Highlights: • Esophageal cancer sphere-forming cells possess cancer stem cell properties. • Sphere-forming cells enhance TGF-β1 pathway activity. • TGF-β 1 inhibitor suppresses the migration and invasion of sphere-forming cells.

  18. Acetate supplementation as a means of inducing glioblastoma stem-like cell growth arrest. (United States)

    Long, Patrick M; Tighe, Scott W; Driscoll, Heather E; Fortner, Karen A; Viapiano, Mariano S; Jaworski, Diane M


    Glioblastoma (GBM), the most common primary adult malignant brain tumor, is associated with a poor prognosis due, in part, to tumor recurrence mediated by chemotherapy and radiation resistant glioma stem-like cells (GSCs). The metabolic and epigenetic state of GSCs differs from their non-GSC counterparts, with GSCs exhibiting greater glycolytic metabolism and global hypoacetylation. However, little attention has been focused on the potential use of acetate supplementation as a therapeutic approach. N-acetyl-l-aspartate (NAA), the primary storage form of brain acetate, and aspartoacylase (ASPA), the enzyme responsible for NAA catalysis, are significantly reduced in GBM tumors. We recently demonstrated that NAA supplementation is not an appropriate therapeutic approach since it increases GSC proliferation and pursued an alternative acetate source. The FDA approved food additive Triacetin (glyceryl triacetate, GTA) has been safely used for acetate supplementation therapy in Canavan disease, a leukodystrophy due to ASPA mutation. This study characterized the effects of GTA on the proliferation and differentiation of six primary GBM-derived GSCs relative to established U87 and U251 GBM cell lines, normal human cerebral cortical astrocytes, and murine neural stem cells. GTA reduced proliferation of GSCs greater than established GBM lines. Moreover, GTA reduced growth of the more aggressive mesenchymal GSCs greater than proneural GSCs. Although sodium acetate induced a dose-dependent reduction of GSC growth, it also reduced cell viability. GTA-mediated growth inhibition was not associated with differentiation, but increased protein acetylation. These data suggest that GTA-mediated acetate supplementation is a novel therapeutic strategy to inhibit GSC growth.

  19. Sphere-forming tumor cells possess stem-like properties in human fibrosarcoma primary tumors and cell lines (United States)



    Fibrosarcoma is a malignant soft tissue tumor of mesenchymal origin. Despite advances in medical and surgical treatment, patient survival rates have remained poor. According to the cancer stem cell hypothesis, tumors are comprised of heterogeneous cell populations that have different roles in tumor formation and growth. Cancer stem cells are a small cell subpopulation that exhibits stem-like properties to gain aggressiveness and recurrence. These cells have been identified in a variety of cancerous tumors, but not in human fibrosarcoma. In this study, we observed that HT1080 cells and primary fibrosarcoma cells formed spheres and showed higher self-renewal capacity, invasiveness and drug resistance compared with their adherent counterparts. Moreover, we demonstrated that the cells showed higher expression of the embryonic stem cell-related genes Nanog, Oct3/4, Sox2, Sox10 and their encoding proteins, as well as greater tumorigenic capacity in nude mice. In conclusion, our data suggest the presence of a stem-like cell population in human fibrosarcoma tumors, which provides more evidence for the cancer stem cell hypothesis and assistance in designing new therapeutic strategies against human fibrosarcoma. PMID:23205129

  20. β-Elemene-Attenuated Tumor Angiogenesis by Targeting Notch-1 in Gastric Cancer Stem-Like Cells

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    Bing Yan


    Full Text Available Emerging evidence suggests that cancer stem cells are involved in tumor angiogenesis. The Notch signaling pathway is one of the most important regulators of these processes. β-Elemene, a naturally occurring compound extracted from Curcumae Radix, has been used as an antitumor drug for various cancers in China. However, its underlying mechanism in the treatment of gastric cancer remains largely unknown. Here, we report that CD44+ gastric cancer stem-like cells (GCSCs showed enhanced proliferation capacity compared to their CD44− counterparts, and this proliferation was accompanied by the high expression of Notch-1 (in vitro. These cells were also more superior in spheroid colony formation (in vitro and tumorigenicity (in vivo and positively associated with microvessel density (in vivo. β-Elemene was demonstrated to effectively inhibit the viability of GCSCs in a dose-dependent manner, most likely by suppressing Notch-1 (in vitro. β-Elemene also contributed to growth suppression and attenuated the angiogenesis capacity of these cells (in vivo most likely by interfering with the expression of Notch-1 but not with Dll4. Our findings indicated that GCSCs play an important role in tumor angiogenesis, and Notch-1 is one of the most likely mediators involved in these processes. β-Elemene was effective at attenuating angiogenesis by targeting the GCSCs, which could be regarded as a potential mechanism for its efficacy in gastric cancer management in the future.

  1. p21-activated kinase 1 determines stem-like phenotype and sunitinib resistance via NF-κB/IL-6 activation in renal cell carcinoma. (United States)

    Zhu, Y; Liu, H; Xu, L; An, H; Liu, W; Liu, Y; Lin, Z; Xu, J


    The p21-activated kinase 1 (PAK1), a serine/threonine kinase that orchestrates cytoskeletal remodeling and cell motility, has been shown to function as downstream node for various oncogenic signaling pathways to promote cell proliferation, regulate apoptosis and accelerate mitotic abnormalities, resulting in tumor formation and invasiveness. Although alterations in PAK1 expression and activity have been detected in various human malignancies, its potential biological and clinical significance in renal cell carcinoma (RCC) remains obscure. In this study, we found increased PAK1 and phosphorylated PAK1 levels in tumor tissues according to TNM stage progression. Elevated phosphorylated PAK1 levels associated with progressive features and indicated unfavorable overall survival (OS) as an independent adverse prognosticator for patients with RCC. Moreover, PAK1 kinase activation with constitutive active PAK1 mutant T423E promoted growth, colony formation, migration, invasion and stem-like phenotype of RCC cells, and vice versa, in PAK1 inhibition by PAK1 kinase inactivation with specific PAK1 shRNA, dead kinase PAK1 mutant K299R or allosteric inhibitor IPA3. Stem-like phenotype due to sunitinib administration via increased PAK1 kinase activation could be ameliorated by PAK1 shRNA, PAK1 mutant K299R and IPA3. Furthermore, nuclear factor-κB (NF-κB)/interleukin-6 (IL-6) activation was found to be responsible for PAK1-mediated stem-like phenotype following sunitinib treatment. Both IL-6 neutralizing antibody and IPA3 administration enhanced tumor growth inhibition effect of sunitinib treatment on RCC cells in vitro and in vivo. Our results unraveled that oncogenic activation of PAK1 defines an important mechanism for maintaining stem-like phenotype and sunitinib resistance through NF-κB/IL-6 activation in RCC, lending PAK1-mediated NF-κB/IL-6 activation considerable appeal as novel pharmacological therapeutic targets against sunitinib resistance.

  2. Tumour-initiating stem-like cells in human prostate cancer exhibit increased NF-κB signalling



    Androgen depletion is a key strategy for treating human prostate cancer, but the presence of hormone-independent cells escaping treatment remains a major therapeutic challenge. Here, we identify a minor subset of stem-like human prostate tumour-initiating cells (TICs) that do not express prostate cancer markers, such as androgen receptor or prostate specific antigen. These TICs possess stem cell characteristics and multipotency as demonstrated by in vitro sphere-formation and in vivo tumour-i...

  3. Proteomic analysis of human Sonic Hedgehog (SHH) medulloblastoma stem-like cells. (United States)

    Ronci, Maurizio; Catanzaro, Giuseppina; Pieroni, Luisa; Po, Agnese; Besharat, Zein Mersini; Greco, Viviana; Levi Mortera, Stefano; Screpanti, Isabella; Ferretti, Elisabetta; Urbani, Andrea


    Human medulloblastoma (MB) is a malignant brain tumor that comprises four distinct molecular subgroups including the Sonic Hedgehog (SHH)-MB group. A leading cause of the SHH subgroup is an aberrant activation of the SHH pathway, a developmental signaling that regulates postnatal development of the cerebellum by promoting the mitotic expansion of granule neural precursors (GNPs) in the external granule layer (EGL). The abnormal SHH signaling pathway drives not only SHH-MB but also its cancer stem-like cells (SLCs), which represent a fraction of the tumor cell population that maintain cancer growth and have been associated with high grade tumors. Here, we report the first proteomic analysis of human SHH-MB SLCs before and after Retinoic Acid (RA)-induced differentiation. A total of 994 nLC-MS buckets were statistically analysed returning 68 modulated proteins between SLCs and their differentiated counterparts. Heat Shock Protein 70 (Hsp70) was one of the proteins that characterized the protein profile of SLCs. By means of Ingenuity Pathway Analysis (IPA), Genomatix analysis and extending the network obtained using the differentially expressed proteins we found a correlation between Hsp70 and the NF-κB complex. A key driver of the SHH-MB group is cMET whose downstream proliferation/survival signalling is indeed via PI3K/Akt/NF-κB. We confirmed the results of the proteomic analysis by western blot, underlining that a P-p65/NF-κB activatory complex is highly expressed in SLCs. Taking together these results we define a new protein feature of SHH-MB SLCs.

  4. B7-H4 expression is elevated in human U251 glioma stem-like cells and is inducible in monocytes cultured with U251 stem-like cell conditioned medium. (United States)

    Mo, Lian-Jie; Ye, Hong-Xing; Mao, Ying; Yao, Yu; Zhang, Jian-Min


    Previous studies indicated that B7-H4, the youngest B7 family, negatively regulates T cell-mediated immunity and is significantly overexpressed in many human tumors. Tumor stem cells are purported to play a role in tumor renewal and resistance to radiation and chemotherapy. However, the link between B7-H4 and tumor stem cells is unclear. In this study, we investigated B7-H4 expression in the medium of human glioma U251 cell cultures. Immunofluorescence results showed that U251 cells cultured in serum-free medium (supplemented with 2% B27, 20 ng/mL epidermal growth factor, 20 ng/mL basic fibroblast growth factor) maintained stem-like cell characteristics, including expression of stem cell marker CD133 and the neural progenitor cell markers nestin and SOX2. In contrast, U251 cells cultured in serum-containing medium highly expressed differentiation marker glial fibrillary acidic protein. Flow cytometry analysis showed serum-free medium-cultured U251 cells expressed higher intracellular B7-H4 than serum-containing medium-cultured U251 cells (24%-35% vs. 8%-11%, P cells revealed moderate expression of B7-H4 after stimulation with conditioned medium from U251 cells cultured in serum-containing medium. Moreover, conditioned medium from U251 stem-like cells had a significant stimulation effect on B7-H4 expression compared with serum-containing conditioned medium (P cells, and conditioned medium from these cells more effectively induced monocytes to express B7-H4 than conditioned medium from U251 cells cultured in the presence of serum. Our results show that U251 stem-like cells may play a more crucial role in tumor immunoloregulation with high expression of B7-H4.

  5. Comparison of efficacy of Salmonella typhimurium A1-R and chemotherapy on stem-like and non-stem human pancreatic cancer cells


    Hiroshima, Yukihiko; Zhao, Ming; Zhang, Yong,; Maawy, Ali; Hassanein, Mohamed K.; Uehara, Fuminari; MIWA, SHINJI; Yano, Shuya; Momiyama, Masashi; Suetsugu, Atsushi; Chishima, Takashi; Tanaka, Kuniya; Bouvet, Michael; Endo, Itaru; Hoffman, Robert M.


    The XPA1 human pancreatic cancer cell line is dimorphic, with spindle stem-like cells and round non-stem cells. We report here the in vitro IC50 values of stem-like and non-stem XPA1 human pancreatic cells cells for: (1) 5-fluorouracil (5-FU), (2) cisplatinum (CDDP), (3) gemcitabine (GEM), and (4) tumor-targeting Salmonella typhimurium A1-R (A1-R). IC50 values of stem-like XPA1 cells were significantly higher than those of non-stem XPA1 cells for 5-FU (P = 0.007) and CDDP (P = 0.012). In cont...

  6. A novel model for evaluating therapies targeting human tumor vasculature and human cancer stem-like cells. (United States)

    Burgos-Ojeda, Daniela; McLean, Karen; Bai, Shoumei; Pulaski, Heather; Gong, Yusong; Silva, Ines; Skorecki, Karl; Tzukerman, Maty; Buckanovich, Ronald J


    Human tumor vessels express tumor vascular markers (TVM), proteins that are not expressed in normal blood vessels. Antibodies targeting TVMs could act as potent therapeutics. Unfortunately, preclinical in vivo studies testing anti-human TVM therapies have been difficult to do due to a lack of in vivo models with confirmed expression of human TVMs. We therefore evaluated TVM expression in a human embryonic stem cell-derived teratoma (hESCT) tumor model previously shown to have human vessels. We now report that in the presence of tumor cells, hESCT tumor vessels express human TVMs. The addition of mouse embryonic fibroblasts and human tumor endothelial cells significantly increases the number of human tumor vessels. TVM induction is mostly tumor-type-specific with ovarian cancer cells inducing primarily ovarian TVMs, whereas breast cancer cells induce breast cancer specific TVMs. We show the use of this model to test an anti-human specific TVM immunotherapeutics; anti-human Thy1 TVM immunotherapy results in central tumor necrosis and a three-fold reduction in human tumor vascular density. Finally, we tested the ability of the hESCT model, with human tumor vascular niche, to enhance the engraftment rate of primary human ovarian cancer stem-like cells (CSC). ALDH(+) CSC from patients (n = 6) engrafted in hESCT within 4 to 12 weeks whereas none engrafted in the flank. ALDH(-) ovarian cancer cells showed no engraftment in the hESCT or flank (n = 3). Thus, this model represents a useful tool to test anti-human TVM therapy and evaluate in vivo human CSC tumor biology.

  7. Chitosan-Decorated Doxorubicin-Encapsulated Nanoparticle Targets and Eliminates Tumor Reinitiating Cancer Stem-like Cells. (United States)

    Rao, Wei; Wang, Hai; Han, Jianfeng; Zhao, Shuting; Dumbleton, Jenna; Agarwal, Pranay; Zhang, Wujie; Zhao, Gang; Yu, Jianhua; Zynger, Debra L; Lu, Xiongbin; He, Xiaoming


    Tumor reinitiating cancer stem-like cells are responsible for cancer recurrence associated with conventional chemotherapy. We developed a doxorubicin-encapsulated polymeric nanoparticle surface-decorated with chitosan that can specifically target the CD44 receptors of these cells. This nanoparticle system was engineered to release the doxorubicin in acidic environments, which occurs when the nanoparticles are localized in the acidic tumor microenvironment and when they are internalized and localized in the cellular endosomes/lysosomes. This nanoparticle design strategy increases the cytotoxicity of the doxorubicin by six times in comparison to the use of free doxorubicin for eliminating CD44(+) cancer stem-like cells residing in 3D mammary tumor spheroids (i.e., mammospheres). We further show these nanoparticles reduced the size of tumors in an orthotopic xenograft tumor model with no evident systemic toxicity. The development of nanoparticle system to target cancer stem-like cells with low systemic toxicity provides a new treatment arsenal for improving the survival of cancer patients.

  8. Therapeutic implications of an enriched cancer stem-like cell population in a human osteosarcoma cell line

    Directory of Open Access Journals (Sweden)

    Martins-Neves Sara R


    Full Text Available Abstract Background Osteosarcoma is a bone-forming tumor of mesenchymal origin that presents a clinical pattern that is consistent with the cancer stem cell model. Cells with stem-like properties (CSCs have been identified in several tumors and hypothesized as the responsible for the relative resistance to therapy and tumor relapses. In this study, we aimed to identify and characterize CSCs populations in a human osteosarcoma cell line and to explore their role in the responsiveness to conventional therapies. Methods CSCs were isolated from the human MNNG/HOS cell line using the sphere formation assay and characterized in terms of self-renewal, mesenchymal stem cell properties, expression of pluripotency markers and ABC transporters, metabolic activity and tumorigenicity. Cell's sensitivity to conventional chemotherapeutic agents and to irradiation was analyzed and related with cell cycle-induced alterations and apoptosis. Results The isolated CSCs were found to possess self-renewal and multipotential differentiation capabilities, express markers of pluripotent embryonic stem cells Oct4 and Nanog and the ABC transporters P-glycoprotein and BCRP, exhibit low metabolic activity and induce tumors in athymic mice. Compared with parental MNNG/HOS cells, CSCs were relatively more resistant to both chemotherapy and irradiation. None of the treatments have induced significant cell-cycle alterations and apoptosis in CSCs. Conclusions MNNG/HOS osteosarcoma cells contain a stem-like cell population relatively resistant to conventional chemotherapeutic agents and irradiation. This resistant phenotype appears to be related with some stem features, namely the high expression of the drug efflux transporters P-glycoprotein and BCRP and their quiescent nature, which may provide a biological basis for resistance to therapy and recurrence commonly observed in osteosarcoma.

  9. Cell-surface Vimentin: A mislocalized protein for isolating csVimentin(+) CD133(-) novel stem-like hepatocellular carcinoma cells expressing EMT markers. (United States)

    Mitra, Abhisek; Satelli, Arun; Xia, Xueqing; Cutrera, Jeffrey; Mishra, Lopa; Li, Shulin


    Recent advances in cancer stem cell biology have shown that cancer stem-like cells with epithelial-mesenchymal transition (EMT) phenotypes are more aggressive and cause relapse; however, absence of a specific marker to isolate these EMT stem-like cells hampers research in this direction. Cell surface markers have been identified for isolating cancer stem-like cells, but none has been identified for isolating cancer stem-like cells with EMT phenotype. Recently, we discovered that Vimentin, an intracellular EMT tumor cell marker, is present on the surface of colon metastatic tumor nodules in the liver. In our study, we examined the potential of targeting cell surface Vimentin (CSV) to isolate stem-like cancer cells with EMT phenotype, by using a specific CSV-binding antibody, 84-1. Using this antibody, we purified the CSV-positive, CD133-negative (csVim(+) CD133(-) ) cell population from primary liver tumor cell suspensions and characterized for stem cell properties. The results of sphere assays and staining for the stem cell markers Sox2 and Oct4A demonstrated that csVim(+) CD133(-) cells have stem-like properties similar to csVim(-) CD133(+) population. Our investigation further revealed that the csVim(+) CD133(-) cells had EMT phenotypes, as evidenced by the presence of Twist and Slug in the nucleus, the absence of EpCAM on the cell surface and basal level of expression of epithelial marker E-cadherin. The csVimentin-negative CD133-positive stem cells do not have any EMT phenotypes. csVim(+) CD133(-) cells exhibited more aggressively metastatic in livers than csVim(-) CD133(+) cells. Our findings indicate that csVim(+) CD133(-) cells are promising targets for treatment and prevention of metastatic hepatocellular carcinoma. © 2014 UICC.

  10. Identification and analysis of CD133(+) melanoma stem-like cells conferring resistance to taxol: An insight into the mechanisms of their resistance and response. (United States)

    El-Khattouti, Abdelouahid; Selimovic, Denis; Haïkel, Youssef; Megahed, Mosaad; Gomez, Christian R; Hassan, Mohamed


    The presence and the involvement of cancer stem-like cells (CSCs) in tumor initiation and progression, and chemo-resistance are documented. Herein, we functionally analyzed melanoma stem-like cells (MSC)/CD133(+) cells on their resistance and response to taxol-induced apoptosis. Besides being taxol resistant, the CD133(+) cells demonstrated a growth advantage over the CD133(-) subpopulation. Taxol induced apoptosis on CD133(-) cells, but not on CD133(+) cells. In the CD133(-) subpopulation, the exposure to taxol induced the activation of apoptosis signal-regulating kinase1 (ASK1)/c-jun-N-terminal kinase (JNK), p38, extracellular signal regulated kinase (ERK) pathways and Bax expression, while in CD133(+) cells taxol was able only to enhance the activity of the ERK pathway. In CD133(+) cells, the direct gene transfer of Bax overcame the acquired resistance to taxol. Taken together, our data provide an insight into the mechanistic cascade of melanoma resistance to taxol and suggest Bax gene transfer as a complementary approach to chemotherapy.

  11. Isolation and phenotypic characterization of cancer stem-like side population cells in colon cancer. (United States)

    Feng, Long; Wu, Jian-Bing; Yi, Feng-Ming


    Previous studies in cancer biology suggest that chemotherapeutic drug resistance and tumor relapse are driven by cells within a tumor termed 'cancer stem cells'. In the present study, a Hoechst 33342 dye exclusion technique was used to identify cancer stem‑like side population (SP) cells in colon carcinoma, which accounted for 3.4% of the total cell population. Following treatment with verapamil, the population of SP cells was reduced to 0.6%. In addition, the sorted SP cells exhibited marked multidrug resistance and enhanced cell survival rates compared with non‑SP cells. The SP cells were able to generate more tumor spheres and were CD133 positive. Subsequent biochemical analysis revealed that the levels of the adenosine triphosphate‑binding cassette sub‑family G member 2 transporter protein, B‑cell lymphoma anti‑apoptotic factor and autocrine production of interleukin‑4 were significantly enhanced in the colon cancer SP cells, which contributed to drug resistance, protection of the cells from apoptosis and tumor recurrence. Therefore, the findings suggested that treatment failure and colon tumorigenesis is dictated by a small population of SP cells, which indicate a potential target in future therapies.

  12. Notch inhibition suppresses nasopharyngeal carcinoma by depleting cancer stem-like side population cells. (United States)

    Yu, Shudong; Zhang, Ruxin; Liu, Fenye; Wang, Hong; Wu, Jing; Wang, Yanqing


    The cancer stem cell (CSC) is responsible for the initiation, proliferation and radiation resistance. Side population (SP) cells are a rare subset of cells enriched with CSCs. The targeting of key signaling pathways that are active in CSCs is a therapeutic approach to treating cancer. Notch signaling is important for the self-renewal and maintenance of stem cells. Our previous studies demonstrated that downregulation of Notch signaling could enhance radiosensitivity of nasopharyngeal carcinoma (NPC) cells. In this study, we found that Notch signaling was highly activated in SP cells compared with that of non-SP (NSP) cells of NPC. Therefore, Notch inhibition could reduce the proportion of SP cells. As SP cells decreased, proliferation, anti-apoptosis and tumorigenesis were also decreased. This study shows that Notch inhibition may be a promising clinical approach in CSC-targeting therapy for NPC.

  13. Vitamin D compounds inhibit cancer stem-like cells and induce differentiation in triple negative breast cancer. (United States)

    Shan, Naing Lin; Wahler, Joseph; Lee, Hong Jin; Bak, Min Ji; Gupta, Soumyasri Das; Maehr, Hubert; Suh, Nanjoo


    Triple-negative breast cancer is one of the least responsive breast cancer subtypes to available targeted therapies due to the absence of hormonal receptors, aggressive phenotypes, and the high rate of relapse. Early breast cancer prevention may therefore play an important role in delaying the progression of triple-negative breast cancer. Cancer stem cells are a subset of cancer cells that are thought to be responsible for tumor progression, treatment resistance, and metastasis. We have previously shown that vitamin D compounds, including a Gemini vitamin D analog BXL0124, suppress progression of ductal carcinoma in situ in vivo and inhibit cancer stem-like cells in MCF10DCIS mammosphere cultures. In the present study, the effects of vitamin D compounds in regulating breast cancer stem-like cells and differentiation in triple-negative breast cancer were assessed. Mammosphere cultures, which enriches for breast cancer cells with stem-like properties, were used to assess the effects of 1α,25(OH)2D3 and BXL0124 on cancer stem cell markers in the triple-negative breast cancer cell line, SUM159. Vitamin D compounds significantly reduced the mammosphere forming efficiency in primary, secondary and tertiary passages of mammospheres compared to control groups. Key markers of cancer stem-like phenotype and pluripotency were analyzed in mammospheres treated with 1α,25(OH)2D3 and BXL0124. As a result, OCT4, CD44 and LAMA5 levels were decreased. The vitamin D compounds also down-regulated the Notch signaling molecules, Notch1, Notch2, Notch3, JAG1, JAG2, HES1 and NFκB, which are involved in breast cancer stem cell maintenance. In addition, the vitamin D compounds up-regulated myoepithelial differentiating markers, cytokeratin 14 and smooth muscle actin, and down-regulated the luminal marker, cytokeratin 18. Cytokeratin 5, a biomarker associated with basal-like breast cancer, was found to be significantly down-regulated by the vitamin D compounds. These results suggest that

  14. Hypoxia and oxygenation induce a metabolic switch between pentose phosphate pathway and glycolysis in glioma stem-like cells. (United States)

    Kathagen, Annegret; Schulte, Alexander; Balcke, Gerd; Phillips, Heidi S; Martens, Tobias; Matschke, Jakob; Günther, Hauke S; Soriano, Robert; Modrusan, Zora; Sandmann, Thomas; Kuhl, Carsten; Tissier, Alain; Holz, Mareike; Krawinkel, Lutz A; Glatzel, Markus; Westphal, Manfred; Lamszus, Katrin


    Fluctuations in oxygen tension during tissue remodeling impose a major metabolic challenge in human tumors. Stem-like tumor cells in glioblastoma, the most common malignant brain tumor, possess extraordinary metabolic flexibility, enabling them to initiate growth even under non-permissive conditions. We identified a reciprocal metabolic switch between the pentose phosphate pathway (PPP) and glycolysis in glioblastoma stem-like (GS) cells. Expression of PPP enzymes is upregulated by acute oxygenation but downregulated by hypoxia, whereas glycolysis enzymes, particularly those of the preparatory phase, are regulated inversely. Glucose flux through the PPP is reduced under hypoxia in favor of flux through glycolysis. PPP enzyme expression is elevated in human glioblastomas compared to normal brain, especially in highly proliferative tumor regions, whereas expression of parallel preparatory phase glycolysis enzymes is reduced in glioblastomas, except for strong upregulation in severely hypoxic regions. Hypoxia stimulates GS cell migration but reduces proliferation, whereas oxygenation has opposite effects, linking the metabolic switch to the "go or grow" potential of the cells. Our findings extend Warburg's observation that tumor cells predominantly utilize glycolysis for energy production, by suggesting that PPP activity is elevated in rapidly proliferating tumor cells but suppressed by acute severe hypoxic stress, favoring glycolysis and migration to protect cells against hypoxic cell damage.

  15. Autocrine VEGF-VEGFR2-Neuropilin-1 signaling promotes glioma stem-like cell viability and tumor growth

    DEFF Research Database (Denmark)

    Hamerlik, Petra; Lathia, Justin D; Rasmussen, Rikke;


    glioma stem-like cells (GSCs), whose viability, self-renewal, and tumorigenicity rely, at least in part, on signaling through the VEGF-VEGFR2-Neuropilin-1 (NRP1) axis. We find that the limited impact of bevacizumab-mediated VEGF blockage may reflect ongoing autocrine signaling through VEGF-VEGFR2-NRP1......, which is associated with VEGFR2-NRP1 recycling and a pool of active VEGFR2 within a cytosolic compartment of a subset of human GBM cells. Whereas bevacizumab failed to inhibit prosurvival effects of VEGFR2-mediated signaling, GSC viability under unperturbed or radiation-evoked stress conditions...

  16. Substrate-mediated reprogramming of human fibroblasts into neural crest stem-like cells and their applications in neural repair. (United States)

    Tseng, Ting-Chen; Hsieh, Fu-Yu; Dai, Niann-Tzyy; Hsu, Shan-Hui


    Cell- and gene-based therapies have emerged as promising strategies for treating neurological diseases. The sources of neural stem cells are limited while the induced pluripotent stem (iPS) cells have risk of tumor formation. Here, we proposed the generation of self-renewable, multipotent, and neural lineage-related neural crest stem-like cells by chitosan substrate-mediated gene transfer of a single factor forkhead box D3 (FOXD3) for the use in neural repair. A simple, non-toxic, substrate-mediated method was applied to deliver the naked FOXD3 plasmid into human fibroblasts. The transfection of FOXD3 increased cell proliferation and up-regulated the neural crest marker genes (FOXD3, SOX2, and CD271), stemness marker genes (OCT4, NANOG, and SOX2), and neural lineage-related genes (Nestin, β-tubulin and GFAP). The expression levels of stemness marker genes and neural crest maker genes in the FOXD3-transfected fibroblasts were maintained until the fifth passage. The FOXD3 reprogrammed fibroblasts based on the new method significantly rescued the neural function of the impaired zebrafish. The chitosan substrate-mediated delivery of naked plasmid showed feasibility in reprogramming somatic cells. Particularly, the FOXD3 reprogrammed fibroblasts hold promise as an easily accessible cellular source with neural crest stem-like behavior for treating neural diseases in the future.

  17. Transplantation of rat hepatic stem-like (HSL) cells with collagen matrices. (United States)

    Ueno, Yasuharu; Nagai, Hirokazu; Watanabe, Go; Ishikawa, Kiyoshi; Yoshikawa, Kiwamu; Koizumi, Yukio; Kameda, Takashi; Sugiyama, Toshihiro


    Organ restitution using somatic stem cells is of great clinical interest. Recent advances in the field of tissue engineering have demonstrated that the use of collagen matrices as scaffolds facilitates tissue reconstruction. Here, we examine the efficacy of transplantation of HSL cells, a previously established liver epithelial cell line with a potential for differentiation, using collagen scaffolds. To this end, HSL cells were transplanted into Nagase's analbuminemic rat with spongy or gelatinous type I collagen matrices. Consequently, immunohistochemical analyses and genomic PCR experiments revealed engraftment of the transplanted cells. Furthermore, the levels of serum albumin in recipient rats were found to increase up to 2.5-fold relative to controls after transplantation. These findings suggest that HSL cells are able to differentiate into functional hepatocytes in vivo, and that biodegradable collagen matrices enhance this phenomenon by providing an appropriate microenvironment for hepatocytic repopulation.

  18. Overexpression of Large-Conductance Calcium-Activated Potassium Channels in Human Glioblastoma Stem-Like Cells and Their Role in Cell Migration. (United States)

    Rosa, Paolo; Sforna, Luigi; Carlomagno, Silvia; Mangino, Giorgio; Miscusi, Massimo; Pessia, Mauro; Franciolini, Fabio; Calogero, Antonella; Catacuzzeno, Luigi


    Glioblastomas (GBMs) are brain tumors characterized by diffuse invasion of cancer cells into the healthy brain parenchyma, and establishment of secondary foci. GBM cells abundantly express large-conductance, calcium-activated potassium (BK) channels that are thought to promote cell invasion. Recent evidence suggests that the GBM high invasive potential mainly originates from a pool of stem-like cells, but the expression and function of BK channels in this cell subpopulation have not been studied. We investigated the expression of BK channels in GBM stem-like cells using electrophysiological and immunochemical techniques, and assessed their involvement in the migratory process of this important cell subpopulation. In U87-MG cells, BK channel expression and function were markedly upregulated by growth conditions that enriched the culture in GBM stem-like cells (U87-NS). Cytofluorimetric analysis further confirmed the appearance of a cell subpopulation that co-expressed high levels of BK channels and CD133, as well as other stem cell markers. A similar association was also found in cells derived from freshly resected GBM biopsies. Finally, transwell migration tests showed that U87-NS cells migration was much more sensitive to BK channel block than U87-MG cells. Our data show that BK channels are highly expressed in GBM stem-like cells, and participate to their high migratory activity. J. Cell. Physiol. 232: 2478-2488, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  19. Survivin Modulates Squamous Cell Carcinoma-Derived Stem-Like Cell Proliferation, Viability and Tumor Formation in Vivo

    Directory of Open Access Journals (Sweden)

    Roberta Lotti


    Full Text Available Squamous Cell Carcinoma-derived Stem-like Cells (SCC-SC originate from alterations in keratinocyte stem cells (KSC gene expression and sustain tumor development, invasion and recurrence. Since survivin, a KSC marker, is highly expressed in SCC-SC, we evaluate its role in SCC-SC cell growth and SCC models. Survivin silencing by siRNA decreases clonal growth of SCC keratinocytes and viability of total, rapidly adhering (RAD and non-RAD (NRAD cells from primary SCC. Similarly, survivin silencing reduces the expression of stem cell markers (OCT4, NOTCH1, CD133, β1-integrin, while it increases the level of differentiation markers (K10, involucrin. Moreover, survivin silencing improves the malignant phenotype of SCC 3D-reconstruct, as demonstrated by reduced epidermal thickness, lower Ki-67 positive cell number, and decreased expression of MMP9 and psoriasin. Furthermore, survivin depletion by siRNA in RasG12V-IκBα-derived tumors leads to smaller tumor formation characterized by lower mitotic index and reduced expression of the tumor-associated marker HIF1α, VEGF and CD51. Therefore, our results indicate survivin as a key gene in regulating SCC cancer stem cell formation and cSCC development.

  20. High-Throughput Single-Cell Derived Sphere Formation for Cancer Stem-Like Cell Identification and Analysis (United States)

    Chen, Yu-Chih; Ingram, Patrick N.; Fouladdel, Shamileh; McDermott, Sean P.; Azizi, Ebrahim; Wicha, Max S.; Yoon, Euisik


    Considerable evidence suggests that many malignancies are driven by a cellular compartment that displays stem cell properties. Cancer stem-like cells (CSCs) can be identified by expression of cell surface markers or enzymatic activity, but these methods are limited by phenotypic heterogeneity and plasticity of CSCs. An alternative phenotypic methodology based on in-vitro sphere formation has been developed, but it is typically labor-intensive and low-throughput. In this work, we present a 1,024-microchamber microfluidic platform for single-cell derived sphere formation. Utilizing a hydrodynamic capturing scheme, more than 70% of the microchambers capture only one cell, allowing for monitoring of sphere formation from heterogeneous cancer cell populations for identification of CSCs. Single-cell derived spheres can be retrieved and dissociated for single-cell analysis using a custom 96-gene panel to probe heterogeneity within the clonal CSC spheres. This microfluidic platform provides reliable and high-throughput sphere formation for CSC identification and downstream clonal analysis.

  1. Generation and characterisation of cisplatin-resistant non-small cell lung cancer cell lines displaying a stem-like signature.

    Directory of Open Access Journals (Sweden)

    Martin P Barr

    Full Text Available INTRODUCTION: Inherent and acquired cisplatin resistance reduces the effectiveness of this agent in the management of non-small cell lung cancer (NSCLC. Understanding the molecular mechanisms underlying this process may result in the development of novel agents to enhance the sensitivity of cisplatin. METHODS: An isogenic model of cisplatin resistance was generated in a panel of NSCLC cell lines (A549, SKMES-1, MOR, H460. Over a period of twelve months, cisplatin resistant (CisR cell lines were derived from original, age-matched parent cells (PT and subsequently characterized. Proliferation (MTT and clonogenic survival assays (crystal violet were carried out between PT and CisR cells. Cellular response to cisplatin-induced apoptosis and cell cycle distribution were examined by FACS analysis. A panel of cancer stem cell and pluripotent markers was examined in addition to the EMT proteins, c-Met and β-catenin. Cisplatin-DNA adduct formation, DNA damage (γH2AX and cellular platinum uptake (ICP-MS was also assessed. RESULTS: Characterisation studies demonstrated a decreased proliferative capacity of lung tumour cells in response to cisplatin, increased resistance to cisplatin-induced cell death, accumulation of resistant cells in the G0/G1 phase of the cell cycle and enhanced clonogenic survival ability. Moreover, resistant cells displayed a putative stem-like signature with increased expression of CD133+/CD44+cells and increased ALDH activity relative to their corresponding parental cells. The stem cell markers, Nanog, Oct-4 and SOX-2, were significantly upregulated as were the EMT markers, c-Met and β-catenin. While resistant sublines demonstrated decreased uptake of cisplatin in response to treatment, reduced cisplatin-GpG DNA adduct formation and significantly decreased γH2AX foci were observed compared to parental cell lines. CONCLUSION: Our results identified cisplatin resistant subpopulations of NSCLC cells with a putative stem-like

  2. Derivation of Stromal (Skeletal, Mesenchymal) Stem-like cells from Human Embryonic Stem Cells

    DEFF Research Database (Denmark)

    Mahmood, Amer; Harkness, Linda; Abdallah, Basem


    stromal (mesenchymal, skeletal) stem cell (hMSC)-like population, known to be osteoblastic cell precursors and to test their osteoblastic differentiation capacity in ex vivo cultures and in vivo. We cultured hESC in a feeder-free environment using serum replacement and as suspension aggregates (embryoid...

  3. Ginsenoside Rh2 Inhibits Cancer Stem-Like Cells in Skin Squamous Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Shunli Liu


    Full Text Available Background/Aims: Treatments targeting cancer stem cells (CSCs are most effective cancer therapy, whereas determination of CSCs is challenging. We have recently reported that Lgr5-positive cells are cancer stem cells (CSCs in human skin squamous cell carcinoma (SCC. Ginsenoside Rh2 (GRh2 has been shown to significantly inhibit growth of some types of cancers, whereas its effects on the SCC have not been examined. Methods: Here, we transduced human SCC cells with lentivirus carrying GFP reporter under Lgr5 promoter. The transduced SCC cells were treated with different doses of GRh2, and then analyzed cell viability by CCK-8 assay and MTT assay. The effects of GRh2 on Lgr5-positive CSCs were determined by fow cytometry and by tumor sphere formation. Autophagy-associated protein and β-catenin were measured by Western blot. Expression of short hairpin small interfering RNA (shRNA for Atg7 and β-catenin were used to inhibit autophagy and β-catenin signaling pathway, respectively, as loss-of-function experiments. Results: We found that GRh2 dose-dependently reduced SCC viability, possibly through reduced the number of Lgr5-positive CSCs. GRh2 increased autophagy and reduced β-catenin signaling in SCC cells. Inhibition of autophagy abolished the effects of GRh2 on β-catenin and cell viability, while increasing β-catenin abolished the effects of GRh2 on autophagy and cell viability. Conclusion: Taken together, our data suggest that GRh2 inhibited SCC growth, possibly through reduced the number of Lgr5-positive CSCs. This may be conducted through an interaction between autophagy and β-catenin signaling.

  4. Hepatocyte growth factor-induced proliferation of hepatic stem-like cells depends on activation of NF-κB

    Institute of Scientific and Technical Information of China (English)

    PengYao; YiqunZhan; WangxiangXu; ChangyanLi; PeibinYue; ChengwangXu; DarongHU; ChengkuiQu; XiaomingYang


    Background/Aims: Hepatocyte growth factor (HGF) regulates proliferation of hepatic stem cells. Transcription factor nuclear factor kappa B (NF-κB) has been demonstrated as a key mediator for cell growth regulation. We investigated the role of NF-κB in HGF-mediated cellular proliferation responses in a rat liver.derived hepatic stem-like cell line WB.F344. Methods: Cell proliferation was determined by incorporation of [3H]thymidine. Phosphorylation of ERK1/2, p38 MAPK, Akt and IκBα by HGF stimulation was detected by Western blotting. NF-κB activation was determined by electrophoretic mobility shift assay and NF-κB.mediated SEAP reporter assay. NF-κB activation was inhibited by treatment with an IκBα dominant-negative vector or inhibitor BAY-11-7082. Results: We found that stimulation of WB-F344 cells with HGF promoted cell proliferation and effectively protected WB-F344 cells from apoptosis induced by TNF-α. We also observed activation of ERK1/2, p38 MAPK, Akt and NF-κB signaling pathways by HGF in WB-F344 cells. HGF-induced cell proliferation was partly blocked by pre-treatment of the cells with inhibitors against MEK1 or p38 MAPK, and completely blocked using an inhibitor for NF-κB activity.Furthermore, it was demonstrated that IκB mutant that suppressed NF-κB activity completely blocked HGF-induced cell proliferation. Conclusions: NF-κB activity is required for HGF-induced proliferation in hepatic stem-like cell line WB-F344, and this activity requires ERK1/2 and p38 MAPK pathways.

  5. The nutritional phenome of EMT-induced cancer stem-like cells (United States)

    Cuyàs, Elisabet; Corominas-Faja, Bruna; Menendez, Javier A.


    The metabolic features of cancer stem (CS) cells and the effects of specific nutrients or metabolites on CS cells remain mostly unexplored. A preliminary study to delineate the nutritional phenome of CS cells exploited the landmark observation that upon experimental induction into an epithelial-to-mesenchymal (EMT) transition, the proportion of CS-like cells drastically increases within a breast cancer cell population. EMT-induced CS-like cells (HMLERshEcad) and isogenic parental cells (HMLERshCntrol) were simultaneously screened for their ability to generate energy-rich NADH when cultured in a standardized high-throughput metabolic phenotyping platform comprising >350 wells that were pre-loaded with different carbohydrates/starches, alcohols, fatty acids, ketones, carboxylic acids, amino acids, and bi-amino acids. The generation of “phenetic maps” of the carbon and nitrogen utilization patterns revealed that the acquisition of a CS-like cellular state provided an enhanced ability to utilize additional catabolic fuels, especially under starvation conditions. Crucially, the acquisition of cancer stemness activated a metabolic infrastructure that enabled the vectorial transfer of high-energy nutrients such as glycolysis end products (pyruvate, lactate) and bona fide ketone bodies (β-hydroxybutyrate) from the extracellular microenvironment to support mitochondrial energy production in CS-like cells. Metabolic reprogramming may thus constitute an efficient adaptive strategy through which CS-like cells would rapidly obtain an advantage in hostile conditions such as nutrient starvation following the inhibition of tumor angiogenesis. By understanding how specific nutrients could bioenergetically boost EMT-CS-like phenotypes, “smart foods” or systemic “metabolic nichotherapies” may be tailored to specific nutritional CSC phenomes, whereas high-resolution heavy isotope-labeled nutrient tracking may be developed to monitor the spatiotemporal distribution and

  6. Differentially expressed miRNAs in cancer-stem-like cells: markers for tumor cell aggressiveness of pancreatic cancer. (United States)

    Bao, Bin; Ali, Shadan; Ahmad, Aamir; Li, Yiwei; Banerjee, Sanjeev; Kong, Dejuan; Aboukameel, Amro; Mohammad, Ramzi; Van Buren, Eric; Azmi, Asfar S; Sarkar, Fazlul H


    Pancreatic cancer (PC) is one of the most deadly cancers. The higher mortality is in part due to treatment resistance and early onset of metastasis. The existence of cancer-stem-like cells (CSLCs) has been widely accepted to be responsible for tumor aggressiveness in PC. Emerging evidence suggests that CSLCs have the capacity for increased cell growth, cell migration/invasion, metastasis, and treatment resistance, which leads to poor clinical outcome. However, the molecular role of CSLCs in tumor development and progression is poorly understood. Therefore, mechanistic understanding, and targeted killing of CSLCs may provide a newer therapeutic strategy for the treatment of PC. It has been well accepted that microRNAs (miRNAs) play critical roles during tumor development and progression through deregulation of multiple genes. Moreover, deregulated expression of miRNAs may also play a key role in the regulation of CSLC characteristics and functions. Here we show that isolated CD44(+)/CD133(+)/EpCAM(+) cells (triple-marker-positive cells) from human PC cell lines, MiaPaCa-2 and L3.6pl cells, display aggressive characteristics, such as increased cell growth, clonogenicity, cell migration, and self-renewal capacity, which is consistent with overexpression of CSLC signatures/markers. We also found deregulated expression of over 400 miRNAs, including let-7, miR-30, miR-125b, and miR-335, in CSLCs. As a proof-of-concept, knockdown of miR-125b resulted in the inhibition of tumor cell aggressiveness of CSLCs (triple-marker-positive cells), consistent with the downregulation of CD44, EpCAM, EZH2, and snail. These results clearly suggest the importance of miRNAs in the regulation of CSLC characteristics, and may serve as novel targets for therapy.

  7. Genome-wide microarray expression and genomic alterations by array-CGH analysis in neuroblastoma stem-like cells.

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    Raquel Ordóñez

    Full Text Available Neuroblastoma has a very diverse clinical behaviour: from spontaneous regression to a very aggressive malignant progression and resistance to chemotherapy. This heterogeneous clinical behaviour might be due to the existence of Cancer Stem Cells (CSC, a subpopulation within the tumor with stem-like cell properties: a significant proliferation capacity, a unique self-renewal capacity, and therefore, a higher ability to form new tumors. We enriched the CSC-like cell population content of two commercial neuroblastoma cell lines by the use of conditioned cell culture media for neurospheres, and compared genomic gains and losses and genome expression by array-CGH and microarray analysis, respectively (in CSC-like versus standard tumor cells culture. Despite the array-CGH did not show significant differences between standard and CSC-like in both analyzed cell lines, the microarray expression analysis highlighted some of the most relevant biological processes and molecular functions that might be responsible for the CSC-like phenotype. Some signalling pathways detected seem to be involved in self-renewal of normal tissues (Wnt, Notch, Hh and TGF-β and contribute to CSC phenotype. We focused on the aberrant activation of TGF-β and Hh signalling pathways, confirming the inhibition of repressors of TGF-β pathway, as SMAD6 and SMAD7 by RT-qPCR. The analysis of the Sonic Hedgehog pathway showed overexpression of PTCH1, GLI1 and SMO. We found overexpression of CD133 and CD15 in SIMA neurospheres, confirming that this cell line was particularly enriched in stem-like cells. This work shows a cross-talk among different pathways in neuroblastoma and its importance in CSC-like cells.

  8. Oct-4 expression maintained cancer stem-like properties in lung cancer-derived CD133-positive cells.

    Directory of Open Access Journals (Sweden)

    Yu-Chih Chen

    Full Text Available CD133 (prominin-1, a 5-transmembrane glycoprotein, has recently been considered to be an important marker that represents the subset population of cancer stem-like cells. Herein we report the isolation of CD133-positive cells (LC-CD133(+ and CD133-negative cells (LC-CD133(- from tissue samples of ten patients with non-small cell lung cancer (LC and five LC cell lines. LC-CD133(+ displayed higher Oct-4 expressions with the ability to self-renew and may represent a reservoir with proliferative potential for generating lung cancer cells. Furthermore, LC-CD133(+, unlike LC-CD133(-, highly co-expressed the multiple drug-resistant marker ABCG2 and showed significant resistance to chemotherapy agents (i.e., cisplatin, etoposide, doxorubicin, and paclitaxel and radiotherapy. The treatment of Oct-4 siRNA with lentiviral vector can specifically block the capability of LC-CD133(+ to form spheres and can further facilitate LC-CD133(+ to differentiate into LC-CD133(-. In addition, knock-down of Oct-4 expression in LC-CD133(+ can significantly inhibit the abilities of tumor invasion and colony formation, and increase apoptotic activities of caspase 3 and poly (ADP-ribose polymerase (PARP. Finally, in vitro and in vivo studies further confirm that the treatment effect of chemoradiotherapy for LC-CD133(+ can be improved by the treatment of Oct-4 siRNA. In conclusion, we demonstrated that Oct-4 expression plays a crucial role in maintaining the self-renewing, cancer stem-like, and chemoradioresistant properties of LC-CD133(+. Future research is warranted regarding the up-regulated expression of Oct-4 in LC-CD133(+ and malignant lung cancer.

  9. δ-Tocotrienol, a natural form of vitamin E, inhibits pancreatic cancer stem-like cells and prevents pancreatic cancer metastasis (United States)

    Husain, Kazim; Centeno, Barbara A; Coppola, Domenico; Trevino, Jose; Sebti, Said M; Malafa, Mokenge P


    The growth, metastasis, and chemotherapy resistance of pancreatic ductal adenocarcinoma (PDAC) is characterized by the activation and growth of tumor-initiating cells in distant organs that have stem-like properties. Thus, inhibiting growth of these cells may prevent PDAC growth and metastases. We have demonstrated that δ-tocotrienol, a natural form of vitamin E (VEDT), is bioactive against cancer, delays progression, and prevents metastases in transgenic mouse models of PDAC. In this report, we provide the first evidence that VEDT selectively inhibits PDAC stem-like cells. VEDT inhibited the viability, survival, self-renewal, and expression of Oct4 and Sox2 transcription factors in 3 models of PDAC stem-like cells. In addition, VEDT inhibited the migration, invasion, and several biomarkers of epithelial-to-mesenchymal transition and angiogenesis in PDAC cells and tumors. These processes are critical for tumor metastases. Furthermore, in the L3.6pl orthotopic model of PDAC metastases, VEDT significantly inhibited growth and metastases of these cells. Finally, in an orthotopic xenograft model of human PDAC stem-like cells, we showed that VEDT significantly retarded the growth and metastases of gemcitabine-resistant PDAC human stem-like cells. Because VEDT has been shown to be safe and to reach bioactive levels in humans, this work supports investigating VEDT for chemoprevention of PDAC metastases. PMID:28404939

  10. In vitro identification and characterization of CD133(pos cancer stem-like cells in anaplastic thyroid carcinoma cell lines.

    Directory of Open Access Journals (Sweden)

    Giovanni Zito

    they might represent putative thyroid cancer stem-like cells. Our in vitro findings might provide new insights for novel therapeutic approaches.

  11. Chemical Library Screening and Structure-Function Relationship Studies Identify Bisacodyl as a Potent and Selective Cytotoxic Agent Towards Quiescent Human Glioblastoma Tumor Stem-Like Cells.

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    Maria Zeniou

    Full Text Available Cancer stem-like cells reside in hypoxic and slightly acidic tumor niches. Such microenvironments favor more aggressive undifferentiated phenotypes and a slow growing "quiescent state" which preserves them from chemotherapeutic agents that essentially target proliferating cells. Our objective was to identify compounds active on glioblastoma stem-like cells, including under conditions that mimick those found in vivo within this most severe and incurable form of brain malignancy. We screened the Prestwick Library to identify cytotoxic compounds towards glioblastoma stem-like cells, either in a proliferating state or in more slow-growing "quiescent" phenotype resulting from non-renewal of the culture medium in vitro. Compound effects were assessed by ATP-level determination using a cell-based assay. Twenty active molecules belonging to different pharmacological classes have thus been identified. Among those, the stimulant laxative drug bisacodyl was the sole to inhibit in a potent and specific manner the survival of quiescent glioblastoma stem-like cells. Subsequent structure-function relationship studies led to identification of 4,4'-dihydroxydiphenyl-2-pyridyl-methane (DDPM, the deacetylated form of bisacodyl, as the pharmacophore. To our knowledge, bisacodyl is currently the only known compound targeting glioblastoma cancer stem-like cells in their quiescent, more resistant state. Due to its known non-toxicity in humans, bisacodyl appears as a new potential anti-tumor agent that may, in association with classical chemotherapeutic compounds, participate in tumor eradication.

  12. Chemical Library Screening and Structure-Function Relationship Studies Identify Bisacodyl as a Potent and Selective Cytotoxic Agent Towards Quiescent Human Glioblastoma Tumor Stem-Like Cells. (United States)

    Zeniou, Maria; Fève, Marie; Mameri, Samir; Dong, Jihu; Salomé, Christophe; Chen, Wanyin; El-Habr, Elias A; Bousson, Fanny; Sy, Mohamadou; Obszynski, Julie; Boh, Alexandre; Villa, Pascal; Assad Kahn, Suzana; Didier, Bruno; Bagnard, Dominique; Junier, Marie-Pierre; Chneiweiss, Hervé; Haiech, Jacques; Hibert, Marcel; Kilhoffer, Marie-Claude


    Cancer stem-like cells reside in hypoxic and slightly acidic tumor niches. Such microenvironments favor more aggressive undifferentiated phenotypes and a slow growing "quiescent state" which preserves them from chemotherapeutic agents that essentially target proliferating cells. Our objective was to identify compounds active on glioblastoma stem-like cells, including under conditions that mimick those found in vivo within this most severe and incurable form of brain malignancy. We screened the Prestwick Library to identify cytotoxic compounds towards glioblastoma stem-like cells, either in a proliferating state or in more slow-growing "quiescent" phenotype resulting from non-renewal of the culture medium in vitro. Compound effects were assessed by ATP-level determination using a cell-based assay. Twenty active molecules belonging to different pharmacological classes have thus been identified. Among those, the stimulant laxative drug bisacodyl was the sole to inhibit in a potent and specific manner the survival of quiescent glioblastoma stem-like cells. Subsequent structure-function relationship studies led to identification of 4,4'-dihydroxydiphenyl-2-pyridyl-methane (DDPM), the deacetylated form of bisacodyl, as the pharmacophore. To our knowledge, bisacodyl is currently the only known compound targeting glioblastoma cancer stem-like cells in their quiescent, more resistant state. Due to its known non-toxicity in humans, bisacodyl appears as a new potential anti-tumor agent that may, in association with classical chemotherapeutic compounds, participate in tumor eradication.

  13. FoxM1 Drives a Feed-forward STAT3-activation Signaling Loop that Promotes the Self-renewal and Tumorigenicity of Glioblastoma Stem-like Cells (United States)

    Gong, Ai-hua; Wei, Ping; Zhang, Sicong; Yao, Jun; Yuan, Ying; Zhou, Ai-dong; Lang, Frederick F.; Heimberger, Amy B.; Rao, Ganesh; Huang, Suyun


    The growth factor PDGF controls the development of glioblastoma (GBM) but its contribution to the function of GBM stem-like cells (GSC) has been little studied. Here we report that the transcription factor FoxM1 promotes PDGFA-STAT3 signaling to drive GSC self-renewal and tumorigenicity. In GBM we found a positive correlation between expression of FoxM1 and PDGF-A. In GSC and mouse neural stem cells, FoxM1 bound to the PDGF-A promoter to upregulate PDGF-A expression, acting to maintain the stem-like qualities of GSC in part through this mechanism. Analysis of the human cancer genomic database TCGA revealed that GBM express higher levels of STAT3, a PDGF-A effector signaling molecule, as compared with normal brain. FoxM1 regulated STAT3 transcription through interactions with the β-catenin/TCF4 complex. FoxM1 deficiency inhibited PDGF-A and STAT3 expression in neural stem cells and GSC, abolishing their stem-like and tumorigenic properties. Further mechanistic investigations defined a FoxM1-PDGFA-STAT3 feed-forward pathway that was sufficient to confer stem-like properties to glioma cells. Collectively, our findings showed how FoxM1 activates expression of PDGF-A and STAT3 in a pathway required to maintain the self-renewal and tumorigenicity of glioma stem-like cells. PMID:25832656

  14. Kinomic and phospho-proteomic analysis of breast cancer stem-like cells

    DEFF Research Database (Denmark)

    Leth-Larsen, Rikke; Christensen, Anne Geske Lindhard; Ehmsen, Sidse

    cell death, while the bulk of a tumor lacks these capacities. The resistance mechanisms may cause these cells to survive and become the source of later tumor recurrence, highlighting the need for therapeutic strategies that specifically target pathways central to these cancer stem cells. The CD44hi....../CD24-/low compartment of human breast cancer is enriched in tumor-initiating cells; however the functional heterogeneity within this subpopulation remains poorly defined. From a triple-negative breast cancer cell line we isolated and cloned CD44hi single-cells that exhibited functional heterogeneity....... Transcriptomic analysis using Affymetrix Human Gene 1.0 ST Arrays revealed similar alterations at the gene expression level. We used these single cell clones to further investigate the molecular mechanisms underlying the association between selected proteins, some of them phosphorylated, and cancer stem cell...

  15. Kinomic and phospho-proteomic analysis of breast cancer stem-like cells

    DEFF Research Database (Denmark)

    Leth-Larsen, Rikke; Christensen, Anne Geske Lindhard; Ehmsen, Sidse

    /CD24-/low compartment of human breast cancer is enriched in tumor-initiating cells; however the functional heterogeneity within this subpopulation remains poorly defined. From a triple-negative breast cancer cell line we isolated and cloned CD44hi single-cells that exhibited functional heterogeneity...

  16. ER-mitochondria contacts control surface glycan expression and sensitivity to killer lymphocytes in glioma stem-like cells. (United States)

    Bassoy, Esen Yonca; Kasahara, Atsuko; Chiusolo, Valentina; Jacquemin, Guillaume; Boydell, Emma; Zamorano, Sebastian; Riccadonna, Cristina; Pellegatta, Serena; Hulo, Nicolas; Dutoit, Valérie; Derouazi, Madiha; Dietrich, Pierre Yves; Walker, Paul R; Martinvalet, Denis


    Glioblastoma is a highly heterogeneous aggressive primary brain tumor, with the glioma stem-like cells (GSC) being more sensitive to cytotoxic lymphocyte-mediated killing than glioma differentiated cells (GDC). However, the mechanism behind this higher sensitivity is unclear. Here, we found that the mitochondrial morphology of GSCs modulates the ER-mitochondria contacts that regulate the surface expression of sialylated glycans and their recognition by cytotoxic T lymphocytes and natural killer cells. GSCs displayed diminished ER-mitochondria contacts compared to GDCs. Forced ER-mitochondria contacts in GSCs increased their cell surface expression of sialylated glycans and reduced their susceptibility to cytotoxic lymphocytes. Therefore, mitochondrial morphology and dynamism dictate the ER-mitochondria contacts in order to regulate the surface expression of certain glycans and thus play a role in GSC recognition and elimination by immune effector cells. Targeting the mitochondrial morphology, dynamism, and contacts with the ER could be an innovative strategy to deplete the cancer stem cell compartment to successfully treat glioblastoma. © 2017 The Authors.

  17. Stem-Like Memory T Cells Are Discovered | Center for Cancer Research (United States)

    T cells are the white blood cells that are the body’s first line of attack against foreign invaders.  When designing immunotherapies to treat cancer the goal is to prolong the immune response of T cells a bit beyond what the body normally does when a bacterium or a virus is encountered.   Nicholas P. Restifo, M.D., working with Luca Gattinoni, M.D., and other colleagues in CCR’s Surgery Branch recently discovered a subset within the human T cell population that may help clinicians to do just  this.

  18. Blockade of Rho-associated protein kinase (ROCK) inhibits the contractility and invasion potential of cancer stem like cells. (United States)

    Srinivasan, Srisathya; Ashok, Vandhana; Mohanty, Sagarajit; Das, Alakesh; Das, Sreya; Kumar, Sushant; Sen, Shamik; Purwar, Rahul


    Recent studies have implicated the roles of cancer stem like cells (CSCs) in cancer metastasis. However, very limited knowledge exists at the molecular and cellular level to target CSCs for prevention of cancer metastasis. In this study, we examined the roles of contractile dynamics of CSCs in cell invasion and delineated the underlying molecular mechanisms of their distinct cell invasion potential. Using de-adhesion assay and atomic force microscopy, we show that CSCs derived from melanoma and breast cancer cell lines exhibit increased contractility compared to non-CSCs across all tumor types. In addition, CSCs possess increased ECM remodeling capacity as quantified by collagen degradation assay. More importantly, pharmacological blockade of Rho-associated protein kinase completely abolished the contractility and collagen degradation capacity of both CSCs and non-CSCs. In conclusion, our study demonstrates the importance of cell contractility in regulating invasiveness of CSCs and suggests that pharmacological targeting of ROCK pathway represents a novel strategy for targeting both CSCs and bulk population for the treatment of cancer metastasis.

  19. Autophagy promotes resistance to photodynamic therapy-induced apoptosis selectively in colorectal cancer stem-like cells. (United States)

    Wei, Ming-Feng; Chen, Min-Wei; Chen, Ke-Cheng; Lou, Pei-Jen; Lin, Susan Yun-Fan; Hung, Shih-Chieh; Hsiao, Michael; Yao, Cheng-Jung; Shieh, Ming-Jium


    Recent studies have indicated that cancer stem-like cells (CSCs) exhibit a high resistance to current therapeutic strategies, including photodynamic therapy (PDT), leading to the recurrence and progression of colorectal cancer (CRC). In cancer, autophagy acts as both a tumor suppressor and a tumor promoter. However, the role of autophagy in the resistance of CSCs to PDT has not been reported. In this study, CSCs were isolated from colorectal cancer cells using PROM1/CD133 (prominin 1) expression, which is a surface marker commonly found on stem cells of various tissues. We demonstrated that PpIX-mediated PDT induced the formation of autophagosomes in PROM1/CD133(+) cells, accompanied by the upregulation of autophagy-related proteins ATG3, ATG5, ATG7, and ATG12. The inhibition of PDT-induced autophagy by pharmacological inhibitors and silencing of the ATG5 gene substantially triggered apoptosis of PROM1/CD133(+) cells and decreased the ability of colonosphere formation in vitro and tumorigenicity in vivo. In conclusion, our results revealed a protective role played by autophagy against PDT in CSCs and indicated that targeting autophagy could be used to elevate the PDT sensitivity of CSCs. These findings would aid in the development of novel therapeutic approaches for CSC treatment.

  20. Bovine mammary epithelial cells retain stem-like phenotype in long-term cultures. (United States)

    Cravero, Diego; Diego, Cravero; Martignani, Eugenio; Eugenio, Martignani; Miretti, Silvia; Silvia, Miretti; Macchi, Elisabetta; Elisabetta, Macchi; Accornero, Paolo; Paolo, Accornero; Baratta, Mario; Mario, Baratta


    The detection and characterization of bovine mammary stem cells may give a better understanding of the cyclic characteristic of mammary gland development. In turn, this could potentially offer techniques to manipulate lactation yield and for regenerative medicine. We previously demonstrated that adult stem cells reside in the bovine mammary gland and possess an intrinsic regenerative potential. In vitro maintenance and expansion of this primitive population is a challenging task that could make easier the study of adult mammary stem cells. The aim of this study is to investigate this possibility. Different subpopulations of mammary epithelial cells emerge when they are cultured in two defined culture conditions. Specific cell differentiation markers as cytokeratin 18 (CK18) and cytokeratin 14 (CK14) were expressed with significant differences according to culture conditions. Vimentin, a well-known fibroblast marker was observed to increase significantly (P day 20. In both conditions, after prolonged culture (25 days) a subset of cells still retained regenerative capabilities. These cells were able to form organized pseudo-alveoli when transplanted in immunodeficient mice as shown by the expression of cytokeratin 14 (CK14), cytokeratin 18 (CK18), p63 (a mammary basal cell layer marker) and Epithelial Cell Adhesion Molecule (EpCAM). We also were able to observe the presence of milk proteins signal in these regenerated structures, which is a specific marker of functional mammary alveoli. Progenitor content was also analyzed in vitro through Colony-Forming Cell (CFC) assays with no substantial differences among culture conditions and time points. These results demonstrate that long-term culture of a multipotent cell subpopulation with intrinsic regenerative potential is possible.

  1. Targeting Netrin-1 in glioblastoma stem-like cells inhibits growth, invasion, and angiogenesis. (United States)

    Sanvoranart, Tanwarat; Supokawej, Aungkura; Kheolamai, Pakpoom; U-Pratya, Yaowalak; Poungvarin, Niphon; Sathornsumetee, Sith; Issaragrisil, Surapol


    Glioblastoma (GBM) is an aggressive malignant brain tumor that still lacks effective therapy. Glioblastoma stem cells (GBM-SCs) were identified to contribute to aggressive phenotypes and poor clinical outcomes for GBM. Netrin-1, an axon guidance molecule, has been found in several tumors in adults. However, the role of Netrin-1 in GBM-SCs remains largely unknown. In this study, CD133-positive U251 GBM cells were used as a putative GBM-SC population to identify the functions of Netrin-1. Using lentiviral transduction, Netrin-1 miR RNAi vectors were transduced into CD133-positive U251 cells. We demonstrated that cell proliferation and survival were decreased following targeted deletion of Netrin-1. Cell invasion was dramatically diminished in Netrin-1 knockdown GBM-SCs. Moreover, Netrin-1 knockdown GBM-SCs exhibited less proangiogenic activity. In conclusion, Netrin-1 may represent a therapeutic target in glioblastoma.

  2. A Distinct Slow-Cycling Cancer Stem-like Subpopulation of Pancreatic Adenocarcinoma Cells is maintained in Vivo

    Energy Technology Data Exchange (ETDEWEB)

    Dembinski, Jennifer L., E-mail:; Krauss, Stefan [Cellular and Genetic Therapy, Department of Microbiology, Cancer Stem Cell Innovation Center (CAST), Oslo University Hospital, Rikshospitalet, Oslo (Norway)


    Pancreatic adenocarcinoma has the worst prognosis of any major malignancy, with <5% of patients surviving five years. This can be contributed to the often late diagnosis, lack of sufficient treatment and metastatic spread. Heterogeneity within tumors is increasingly becoming a focus in cancer research, as novel therapies are required to target the most aggressive subpopulations of cells that are frequently termed cancer stem cells (CSCs). In the current study, we describe the identification of a slow-cycling cancer stem-like population of cells in vivo in BxPC-3 and Panc03.27 xenografts. A distinct slow-cycling label-retaining population of cells (DiI+/SCC) was found both at the edge of tumors, and in small circumscribed areas within the tumors. DiI+/SCC in these areas display an epithelial-to-mesenchymal transition (EMT) fingerprint, including an upregulation of the mesenchymal markers vimentin and N-cadherin and a loss of the epithelial marker E-cadherin. DiI+/SCC also displayed a critical re-localization of beta-catenin from the membrane to the nucleus. Additionally, the DiI+/SCC population was found to express the developmental signaling molecule sonic hedgehog. This study represents a novel step in defining the biological activities of a tumorigenic subpopulation within the heterogeneous tumor microenvironment in vivo. Understanding the interactions and functions of a CSC population within the context of the tumor microenvironment is critical to design targeted therapeutics.

  3. Autocrine Secretion of Progastrin Promotes the Survival and Self-Renewal of Colon Cancer Stem-like Cells. (United States)

    Giraud, Julie; Failla, Laura M; Pascussi, Jean-Marc; Lagerqvist, Ebba L; Ollier, Jérémy; Finetti, Pascal; Bertucci, François; Ya, Chu; Gasmi, Imène; Bourgaux, Jean-François; Prudhomme, Michel; Mazard, Thibault; Ait-Arsa, Imade; Houhou, Leila; Birnbaum, Daniel; Pélegrin, André; Vincent, Charles; Ryall, James G; Joubert, Dominique; Pannequin, Julie; Hollande, Frédéric


    Subpopulations of cancer stem-like cells (CSC) are thought to drive tumor progression and posttreatment recurrence in multiple solid tumors. However, the mechanisms that maintain stable proportions of self-renewing CSC within heterogeneous tumors under homeostatic conditions remain poorly understood. Progastrin is a secreted peptide that exhibits tumor-forming potential in colorectal cancer, where it regulates pathways known to modulate colon CSC behaviors. In this study, we investigated the role of progastrin in regulating CSC phenotype in advanced colorectal cancer. Progastrin expression and secretion were highly enriched in colon CSC isolated from human colorectal cancer cell lines and colon tumor biopsies. Progastrin expression promoted CSC self-renewal and survival, whereas its depletion by RNA interference-mediated or antibody-mediated strategies altered the homeostatic proportions of CSC cells within heterogeneous colorectal cancer tumors. Progastrin downregulation also decreased the frequency of ALDH(high) cells, impairing their tumor-initiating potential, and inhibited the high glycolytic activity of ALDH(high) CSC to limit their self-renewal capability. Taken together, our results show how colorectal CSC maintain their tumor-initiating and self-renewal capabilities by secreting progastrin, thereby contributing to the tumor microenvironment to support malignancy. Cancer Res; 76(12); 3618-28. ©2016 AACR.

  4. CD133 positive embryonal rhabdomyosarcoma stem-like cell population is enriched in rhabdospheres.

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    Dagmar Walter

    Full Text Available Cancer stem cells (CSCs have been identified in a number of solid tumors, but not yet in rhabdomyosarcoma (RMS, the most frequently occurring soft tissue tumor in childhood. Hence, the aim of this study was to identify and characterize a CSC population in RMS using a functional approach. We found that embryonal rhabdomyosarcoma (eRMS cell lines can form rhabdomyosarcoma spheres (short rhabdospheres in stem cell medium containing defined growth factors over several passages. Using an orthotopic xenograft model, we demonstrate that a 100 fold less sphere cells result in faster tumor growth compared to the adherent population suggesting that CSCs were enriched in the sphere population. Furthermore, stem cell genes such as oct4, nanog, c-myc, pax3 and sox2 are significantly upregulated in rhabdospheres which can be differentiated into multiple lineages such as adipocytes, myocytes and neuronal cells. Surprisingly, gene expression profiles indicate that rhabdospheres show more similarities with neuronal than with hematopoietic or mesenchymal stem cells. Analysis of these profiles identified the known CSC marker CD133 as one of the genes upregulated in rhabdospheres, both on RNA and protein levels. CD133(+ sorted cells were subsequently shown to be more tumorigenic and more resistant to commonly used chemotherapeutics. Using a tissue microarray (TMA of eRMS patients, we found that high expression of CD133 correlates with poor overall survival. Hence, CD133 could be a prognostic marker for eRMS. These experiments indicate that a CD133(+ CSC population can be enriched from eRMS which might help to develop novel targeted therapies against this pediatric tumor.

  5. Chemoresistance and chemotherapy targeting stem-like cells in malignant glioma

    DEFF Research Database (Denmark)

    Sørensen, Mia Dahl; Fosmark, Sigurd; Hellwege, Sofie


    Glioblastoma remains a tumor with a dismal prognosis because of failure of current treatment. Glioblastoma cells with stem cell (GSC) properties survive chemotherapy and give rise to tumor recurrences that invariably result in the death of the patients. Here we summarize the current knowledge...... by extrinsic factors like hypoxia increasing MGMT expression and thereby resistance to alkylating chemotherapy. The search of new biomarkers helping to predict the tumor response to chemotherapy is ongoing and will complement the already known markers like MGMT....

  6. In Vitro Anticancer Effect of Gedunin on Human Teratocarcinomal (NTERA-2 Cancer Stem-Like Cells

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    Luxmiga Tharmarajah


    Full Text Available Gedunin is one of the major compounds found in the neem tree (Azadirachta indica. In the present study, antiproliferative potential of gedunin was evaluated in human embryonal carcinoma cells (NTERA-2, a cancer stem cell model and peripheral blood mononuclear cells (PBMCs, using Sulforhodamine (SRB and WST-1 assays, respectively. The effects of gedunin on expression of heat shock protein 90 (HSP90, its cochaperone Cdc37, and HSP client proteins (AKT, ErbB2, and HSF1 were evaluated by real-time PCR. Effects of gedunin on apoptosis were evaluated by (a apoptosis associated morphological changes, (b caspase 3/7 expression, (c DNA fragmentation, (d TUNEL assay, and (e real-time PCR of apoptosis related genes (Bax, p53, and survivin. Gedunin showed a promising antiproliferative effect in NTERA-2 cells with IC50 values of 14.59, 8.49, and 6.55 μg/mL at 24, 48, and 72 h after incubations, respectively, while exerting a minimal effect on PBMCs. Expression of HSP90, its client proteins, and survivin was inhibited and Bax and p53 were upregulated by gedunin. Apoptosis related morphological changes, DNA fragmentation, and increased caspase 3/7 activities confirmed the proapoptotic effects of gedunin. Collectively, results indicate that gedunin may be a good drug lead for treatment of chemo and radiotherapy resistant cancer stem cells.

  7. Silencing of E6/E7 expression in cervical cancer stem-like cells. (United States)

    Gu, Wenyi; McMillan, Nigel; Yu, Chengzhong


    Accumulating evidence supports the concept that cancer stem cells (CSCs) are responsible for the tumor recurrence and metastasis, the two major causes of cancer-related death. Therefore, CSC-targeted cancer therapy is important for the future development of more effective and advanced cancer therapy. One of the approaches is to specifically silence oncogene expression in CSCs and inhibit their growth. The significance of this approach is its specificity and ability to avoid multi-drug resistance of CSCs. In this chapter, we will describe a method of silencing HPV oncogenes E6/E7 in human cervical CSCs using HeLa cells as a model system.

  8. Non-coding RNAs as epigenetic regulator of glioma stem-like cell differentiation

    Directory of Open Access Journals (Sweden)

    Keisuke eKatsushima


    Full Text Available Glioblastomas show heterogeneous histological features. These distinct phenotypic states are thought to be associated with the presence of glioma stem cells (GSCs, which are highly tumorigenic and self-renewing sub-population of tumor cells that have different functional characteristics. Differentiation of GSCs may be regulated by multi-tiered epigenetic mechanisms that orchestrate the expression of thousands of genes. One such regulatory mechanism involves functional non-coding RNAs (ncRNAs, such as microRNAs (miRNAs; a large number of ncRNAs have been identified and shown to regulate the expression of genes associated with cell differentiation programs. Given the roles of miRNAs in cell differentiation, it is possible they are involved in the regulation of gene expression networks in GSCs that are important for the maintenance of the pluripotent state and for directing differentiation. Here, we review recent findings on ncRNAs associated with GSC differentiation and discuss how these ncRNAs contribute to the establishment of tissue heterogeneity during glioblastoma tumor formation.

  9. Sulforaphane and TRAIL induce a synergistic elimination of advanced prostate cancer stem-like cells. (United States)

    Labsch, Sabrina; Liu, Li; Bauer, Nathalie; Zhang, Yiyao; Aleksandrowicz, Ewa; Gladkich, Jury; Schönsiegel, Frank; Herr, Ingrid


    Advanced androgen-independent prostate cancer (AIPC) is an aggressive malignancy with a poor prognosis. Apoptosis-resistant cancer stem cells (CSCs) have been identified in AIPC and are not eliminated by current therapeutics. Novel therapeutic options, which are currently being evaluated in patient studies, include TRAIL and the broccoli-derived isothiocyanate sulforaphane. Although neither agent targets normal cells, TRAIL induces apoptosis in most cancer cells, and sulforaphane eliminates CSCs. In this study, the established AIPC cell lines DU145 and PC3, with enriched CSC features, and primary patient-derived prostate CSCs were treated with sulforaphane and recombinant soluble TRAIL. We examined the effects of these drugs on NF-κB activity, self-renewal and differentiation potential, and stem cell signaling via spheroid- and colony-forming assays, FACS and western blot analyses, immunohistochemistry, and an antibody protein array in vitro and after xenotransplantation. We largely found a stronger effect of sulforaphane on CSC properties compared to TRAIL, though the agents acted synergistically when applied in combination. This was associated with the inhibition of TRAIL-induced NF-κB binding; CXCR4, Jagged1, Notch 1, SOX 2, and Nanog expression; ALDH1 activity inhibition; and the elimination of differentiation and self-renewal potential. In vivo, tumor engraftment and tumor growth were strongly inhibited, without the induction of liver necrosis or other obvious side effects. These findings suggest that sulforaphane shifts the balance from TRAIL-induced survival signals to apoptosis and thus explains the observed synergistic effect. A nutritional strategy for high sulforaphane intake may target the cancer-specific activity of TRAIL in CSCs.

  10. Cancer stem-like sphere cells induced from de-differentiated hepatocellular carcinoma-derived cell lines possess the resistance to anti-cancer drugs. (United States)

    Hashimoto, Noriaki; Tsunedomi, Ryouichi; Yoshimura, Kiyoshi; Watanabe, Yusaku; Hazama, Shoichi; Oka, Masaaki


    Cancer stem cells (CSCs) are thought to play important roles in therapy-resistance. In this study, we induced cancer stem-like cells from hepatocellular carcinoma (HCC) cell lines using a unique medium, and examined their potential for resistance to anti-cancer drugs. The human HCC cell lines SK-HEP-1 (SK), HLE, Hep 3B, and HuH-7 were used to induce cancer stem-like cells with our sphere induction medium supplemented with neural survival factor-1. NANOG and LIN28A were examined as stemness markers. Several surface markers for CSC such as CD24, CD44, CD44 variant, and CD90 were analyzed by flow-cytometry. To assess the resistance to anti-cancer drugs, the MTS assay, cell cycle analysis, and reactive oxygen species (ROS) activity assay were performed. Poorly differentiated HCC derived SK and undifferentiated HCC derived HLE cell lines efficiently formed spheres of cells (SK-sphere and HLE-sphere), but well-differentiated HCC-derived HuH-7 and Hep 3B cells did not. SK-spheres showed increased NANOG, LIN28A, and ALDH1A1 mRNA levels compared to parental cells. We observed more CD44 variant-positive cells in SK-spheres than in parental cells. The cell viability of SK-spheres was significantly higher than that of SK cells in the presence of several anti-cancer drugs except sorafenib (1.7- to 7.3-fold, each P < 0.05). The cell cycle of SK-spheres was arrested at the G0/G1 phase compared to SK cells. SK-spheres showed higher ABCG2 and HIF1A mRNA expression and lower ROS production compared to parental cells. Our novel method successfully induced cancer stem-like cells, which possessed chemoresistance that was related to the cell cycle, drug efflux, and ROS.

  11. Intermittent Hypoxia Regulates Stem-like Characteristics and Differentiation of Neuroblastoma Cells


    Vasantha Kumar Bhaskara; Indra Mohanam; Jasti S Rao; Sanjeeva Mohanam


    BACKGROUND: Neuroblastomas are the most common extracranial solid tumors in children. Neuroblastomas are derived from immature cells of the sympathetic nervous system and are characterized by clinical and biological heterogeneity. Hypoxia has been linked to tumor progression and increased malignancy. Intermittent hypoxia or repeated episodes of hypoxia followed by re-oxygenation is a common phenomenon in solid tumors including neuroblastoma and it has a significant influence on the outcome of...

  12. A novel mouse model of human breast cancer stem-like cells with high CD44+CD24-/lower phenotype metastasis to human bone

    Institute of Scientific and Technical Information of China (English)

    LING Li-jun; WANG Feng; WANG Shui; LIU Xiao-an; SHEN En-chao; DING Qiang; LU Chao; XU Jian; CAO Qin-hong; ZHU Hai-qing


    Background A satisfactory animal model of breast cancer metastasizing to bone is unavailable. In this study, we used human breast cancer stem-like cells and human bone to build a novel "human-source" model of human breast cancer skeletal metastasis.Methods Human breast cancer stem-like cells, the CD44+/CD24-/lower subpopulation, was separated and cultured. Before injection with the stem-like cells, mice were implanted with human bone in the right or left dorsal flanks. Animals in Groups A, B, and C were injected with 1x105, 1x106 human breast cancer stem-like cells, and 1x106 parental MDA-MB-231 cells, respectively. A positive control group (D) without implantation of human bone was also injected with 1x106 MDA-MB-231 cells. Immunohistochemistry was performed for determination of CD34, CD105, smooth muscle antibody, CD44, CD24, cytokine, CXC chemokine receptor-4 (CXCR4), and osteopontin (OPN). mRNA levels of CD44, CD24, CXCR4, and OPN in bone metastasis tissues were analyzed by real-time quantitative polymerase chain reaction (PCR). Results Our results demonstrated that cells in implanted human bones of group B, which received 1x106 cancer stem-like cells, stained strongly positive for CD44, CXCR4, and OPN, whereas those of other groups showed no or minimum staining. Moreover, group B had the highest incidence of human bone metastasis (77.8%, P=0.0230) and no accompaniment of other tissue metastasis. The real-time PCR showed an increase of CD44, CXCR4, and OPN mRNA in metastatic bone tissues in group B compared with those of groups C and D, however the expression of CD24 mRNA in group B were the lowest. Conclusions In the novel "human source" model of breast cancer, breast cancer stem-like cells demonstrated a higher human bone-seeking ability. Its mechanism might be related to the higher expressions of CD44, CXCR4, and OPN, and the lower expression of CD24 in breast cancer stem-like cells.

  13. Reactive astrocytes promote the metastatic growth of breast cancer stem-like cells by activating Notch signalling in brain. (United States)

    Xing, Fei; Kobayashi, Aya; Okuda, Hiroshi; Watabe, Misako; Pai, Sudha K; Pandey, Puspa R; Hirota, Shigeru; Wilber, Andrew; Mo, Yin-Yuan; Moore, Brian E; Liu, Wen; Fukuda, Koji; Iiizumi, Megumi; Sharma, Sambad; Liu, Yin; Wu, Kerui; Peralta, Elizabeth; Watabe, Kounosuke


    Brain metastasis of breast cancer profoundly affects the cognitive and sensory functions as well as morbidity of patients, and the 1 year survival rate among these patients remains less than 20%. However, the pathological mechanism of brain metastasis is as yet poorly understood. In this report, we found that metastatic breast tumour cells in the brain highly expressed IL-1β which then 'activated' surrounding astrocytes. This activation significantly augmented the expression of JAG1 in the astrocytes, and the direct interaction of the reactivated astrocytes and cancer stem-like cells (CSCs) significantly stimulated Notch signalling in CSCs. We also found that the activated Notch signalling in CSCs up-regulated HES5 followed by promoting self-renewal of CSCs. Furthermore, we have shown that the blood-brain barrier permeable Notch inhibitor, Compound E, can significantly suppress the brain metastasis in vivo. These results represent a novel paradigm for the understanding of how metastatic breast CSCs re-establish their niche for their self-renewal in a totally different microenvironment, which opens a new avenue to identify a novel and specific target for the brain metastatic disease.

  14. YAP1 Regulates OCT4 Activity and SOX2 Expression to Facilitate Self-Renewal and Vascular Mimicry of Stem-Like Cells. (United States)

    Bora-Singhal, Namrata; Nguyen, Jonathan; Schaal, Courtney; Perumal, Deepak; Singh, Sandeep; Coppola, Domenico; Chellappan, Srikumar


    Non-small cell lung cancer (NSCLC) is highly correlated with smoking and has very low survival rates. Multiple studies have shown that stem-like cells contribute to the genesis and progression of NSCLC. Our results show that the transcriptional coactivator yes-associated protein 1 (YAP1), which is the oncogenic component of the Hippo signaling pathway, is elevated in the stem-like cells from NSCLC and contributes to their self-renewal and ability to form angiogenic tubules. Inhibition of YAP1 by a small molecule or depletion of YAP1 by siRNAs suppressed self-renewal and vascular mimicry of stem-like cells. These effects of YAP1 were mediated through the embryonic stem cell transcription factor, Sox2. YAP1 could transcriptionally induce Sox2 through a physical interaction with Oct4; Sox2 induction occurred independent of TEAD2 transcription factor, which is the predominant mediator of YAP1 functions. The binding of Oct4 to YAP1 could be detected in cell lines as well as tumor tissues; the interaction was elevated in NSCLC samples compared to normal tissue as seen by proximity ligation assays. YAP1 bound to Oct4 through the WW domain, and a peptide corresponding to this region could disrupt the interaction. Delivery of the WW domain peptide to stem-like cells disrupted the interaction and abrogated Sox2 expression, self-renewal, and vascular mimicry. Depleting YAP1 reduced the expression of multiple epithelial-mesenchymal transition genes and prevented the growth and metastasis of tumor xenografts in mice; overexpression of Sox2 in YAP1 null cells rescued these functions. These results demonstrate a novel regulation of stem-like functions by YAP1, through the modulation of Sox2 expression. © 2015 AlphaMed Press.

  15. Metformin inhibits the proliferation, metastasis, and cancer stem-like sphere formation in osteosarcoma MG63 cells in vitro. (United States)

    Chen, Xu; Hu, Chuanzhen; Zhang, Weibin; Shen, Yuhui; Wang, Jun; Hu, Fangqiong; Yu, Pei


    Metformin is an oral drug that has been widely used to treat type 2 diabetes mellitus. Interestingly, accumulated evidence indicate that metformin may reduce the risk of cancer in patients with type 2 diabetes and inhibit tumor cell growth and survival in numerous malignancies, including osteosarcoma (OS) cells. In the present study, we aimed to investigate the effects of metformin on the proliferation, migration, invasion, and sphere formation in OS MG63 cells in vitro. Metformin suppressed OS MG63 cell proliferation in a dose- and time-dependent manner and markedly blocked anti-metastatic potentials, migration, and invasion, by downregulating matrix metalloproteinase 2 (MMP2) and MMP9. Besides, we established OS cancer stem-like cell (CSC) model with sarcosphere formation assay and demonstrated that metformin posed damage on CSCs in OS by inhibiting sphere formation and by inducing their stemness loss. The stemness of CSCs in OS such as self-renewal and differentiation potentials was both impaired with a significant decrease of Oct-4 and Nanog activation. Consistent with this, the positive rates of CD90, CD133, and stage-specific embryonic antigen-4 (SSEA-4) were all observed with reductions in response to metformin exposure. In addition, Western blot showed that metformin activated AMPKα at Tyr172, followed by a downregulated phosphorylation of mammalian target of rapamycin (mTOR)/S6 and feedback activation of p-AKT Ser(473) in both OS MG63 cells and CSCs. This indicates that AMPK/mTOR/S6 signaling pathway might be involved in the growth inhibition of both OS MG63 cells and CSCs. These results suggest that metformin, a potential anti-neoplastic agent, might make it a novel therapeutic choice for the treatment of OS in the future.

  16. Bladder Cancer Stem-Like Cells: Their Origin and Therapeutic Perspectives

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    Tomokazu Ohishi


    Full Text Available Bladder cancer (BC, the most common cancer arising from the human urinary tract, consists of two major clinicopathological phenotypes: muscle-invasive bladder cancer (MIBC and non-muscle-invasive bladder cancer (NMIBC. MIBC frequently metastasizes and is associated with an unfavorable prognosis. A certain proportion of patients with metastatic BC can achieve a remission with systemic chemotherapy; however, the disease relapses in most cases. Evidence suggests that MIBC comprises a small population of cancer stem cells (CSCs, which may be resistant to these treatments and may be able to form new tumors in the bladder or other organs. Therefore, the unambiguous identification of bladder CSCs and the development of targeted therapies are urgently needed. Nevertheless, it remains unclear where bladder CSCs originate and how they are generated. We review recent studies on bladder CSCs, specifically focusing on their proposed origin and the possible therapeutic options based on the CSC theory.

  17. Active targeting docetaxel-PLA nanoparticles eradicate circulating lung cancer stem-like cells and inhibit liver metastasis. (United States)

    Yang, Nan; Jiang, Yao; Zhang, Huifeng; Sun, Bo; Hou, Chunying; Zheng, Ji; Liu, Yanyong; Zuo, Pingping


    Lung cancer is the major cause of cancer related lethality worldwide, and metastasis to distant organs is the pivotal cause of death for the vast majority of lung cancer patients. Accumulated evidence indicates that lung cancer stem-like cells (CSLCs) play important roles in metastagenesis, and these circulating CSLCs may be important targets to inhibit the subsequent metastasis. The present study was aimed at establishing CSLC-targeting polylactic acid (PLA) encapsulated docetaxel nanoparticles for antimetastatic therapy. Cyclic binding peptides were screened on CSLCs in vitro and the peptide CVKTPAQSC exhibiting high specific binding ability to pulmonary adenocarcinoma tissue was subsequently conjugated to the nanoparticles loaded with docetaxel (NDTX). Antimetastatic effect of CSLC-targeting nanoparticles loaded with docetaxel (TNDTX) was evaluated in a nude mouse model of liver metastasis. Results showed that, in the absence of targeting peptide, NDTX hardly exhibited any antimetastatic effect. However, TNDTX treatment significantly decreased the metastatic tumor area in the nude mouse liver. Histopathological and serological results also confirmed the antimetastatic efficacy of TNDTX. To our knowledge, this is the first report on establishing a CSLC-based strategy for lung cancer metastatic treatment, and we hope this will offer a potential therapeutic approach for management of metastatic lung cancer.

  18. Normal and malignant epithelial cells with stem-like properties have an extended G2 cell cycle phase that is associated with apoptotic resistance

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    Biddle Adrian


    Full Text Available Abstract Background Subsets of cells with stem-like properties have been previously isolated from human epithelial cancers and their resistance to apoptosis-inducing stimuli has been related to carcinoma recurrence and treatment failure. The aim of this study was to investigate the mechanisms of resistance to apoptosis-inducing agents of cells with stem-like properties in both normal and malignant human epithelia. Methods Cells isolated from fresh human head and neck carcinomas (n = 11, cell lines derived from head and neck, prostate and breast human carcinomas (n = 7, and from normal human oral mucosa (n = 5, were exposed to various apoptosis-inducing stimuli (UV, Tumour Necrosis Factor, Cisplatin, Etoposide, and Neocarzinostatin. Flow cytometry for CD44 and epithelial-specific antigen (ESA expression, colony morphology, tumour sphere formation and rapid adherence assays were used to identify the subset of cells with stem-like properties. Apoptosis, cell cycle and expression of various cell cycle checkpoint proteins were assessed (Western Blot, qPCR. The role of G2-checkpoint regulators Chk1 and Chk2 was investigated by use of debromohymenialdisine (DBH and siRNA. Results In both cancer biopsies and carcinoma cell lines a subset of CD44high cells showed increased clonogenicity, a significantly lower rate of apoptosis, and a significantly higher proportion of cells in the G2-phase of the cell cycle. An inverse correlation between the percentage of cells in G2-phase and the rate of apoptosis was found. Pulse-chase with iododeoxyuridine (IdU demonstrated that CD44high carcinoma cells spent longer time in G2, even in un-treated controls. These cells expressed higher levels of G2 checkpoint proteins, and their release from G2 with BDH or Chk1 siRNA increased their rate of apoptosis. Low passage cultures of normal keratinocytes were also found to contain a subset of CD44high cells showing increased clonogenicity, and a similar pattern of G2-block

  19. Chemo-predictive assay for targeting cancer stem-like cells in patients affected by brain tumors.

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    Sarah E Mathis

    Full Text Available Administration of ineffective anticancer therapy is associated with unnecessary toxicity and development of resistant clones. Cancer stem-like cells (CSLCs resist chemotherapy, thereby causing relapse of the disease. Thus, development of a test that identifies the most effective chemotherapy management offers great promise for individualized anticancer treatments. We have developed an ex vivo chemotherapy sensitivity assay (ChemoID, which measures the sensitivity of CSLCs as well as the bulk of tumor cells to a variety of chemotherapy agents. Two patients, a 21-year old male (patient 1 and a 5-month female (patient 2, affected by anaplastic WHO grade-III ependymoma were screened using the ChemoID assay. Patient 1 was found sensitive to the combination of irinotecan and bevacizumab, which resulted in a prolonged disease progression free period of 18 months. Following recurrence, the combination of various chemotherapy drugs was tested again with the ChemoID assay. We found that benzyl isothiocyanate (BITC greatly increased the chemosensitivity of the ependymoma cells to the combination of irinotecan and bevacizumab. After patient 1 was treated for two months with irinotecan, bevacizumab and supplements of cruciferous vegetable extracts containing BITC, we observed over 50% tumoral regression in comparison with pre-ChemoID scan as evidenced by MRI. Patient 2 was found resistant to all treatments tested and following 6 cycles of vincristine, carboplatin, cyclophosphamide, etoposide, and cisplatin in various combinations, the tumor of this patient rapidly progressed and proton beam therapy was recommended. As expected animal studies conducted with patient derived xenografts treated with ChemoID screened drugs recapitulated the clinical observation. This assay demonstrates that patients with the same histological stage and grade of cancer may vary considerably in their clinical response, suggesting that ChemoID testing which measures the sensitivity

  20. Selective targeting of human colon cancer stem-like cells by the mTOR inhibitor Torin-1. (United States)

    Francipane, Maria Giovanna; Lagasse, Eric


    Metastatic colorectal cancer (CRC) is incurable for most patients. Since mammalian target of rapamycin (mTOR) has been suggested as a crucial modulator of tumor biology, we aimed at evaluating the effectiveness of mTOR targeting for CRC therapy. To this purpose, we analyzed mTOR expression and the effect of mTOR inhibition in cancer stem-like cells isolated from three human metastatic CRCs (CoCSCs). CoCSCs exhibited a strong mTOR complex 2 (mTORC2) expression, and a rare expression of mTOR complex 1 (mTORC1). This latter correlated with differentiation, being expressed in CoCSC-derived xenografts. We indicate Serum/glucocorticoid-regulated kinase 1 (SGK1) as the possible main mTORC2 effector in CoCSCs, as highlighted by the negative effect on cancer properties following its knockdown. mTOR inhibitors affected CoCSCs differently, resulting in proliferation, autophagy as well as apoptosis induction. The apoptosis-inducing mTOR inhibitor Torin-1 hindered growth, motility, invasion, and survival of CoCSCs in vitro, and suppressed tumor growth in vivo with a concomitant reduction in vessel formation. Torin-1 also affected the expression of markers for cell proliferation, angio-/lympho-genesis, and stemness in vivo, including Ki67, DLL1, DLL4, Notch, Lgr5, and CD44. Importantly, Torin-1 did not affect the survival of normal colon stem cells in vivo, suggesting its selectivity towards cancer cells. Thus, we propose Torin-1 as a powerful drug candidate for metastatic CRC therapy.

  1. EGFR/Src/Akt signaling modulates Sox2 expression and self-renewal of stem-like side-population cells in non-small cell lung cancer

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    Singh Sandeep


    Full Text Available Abstract Background Cancer stem cells are thought to be responsible for the initiation and progression of cancers. In non-small cell lung cancers (NSCLCs, Hoechst 33342 dye effluxing side population (SP cells are shown to have stem cell like properties. The oncogenic capacity of cancer stem-like cells is in part due to their ability to self-renew; however the mechanistic correlation between oncogenic pathways and self-renewal of cancer stem-like cells has remained elusive. Here we characterized the SP cells at the molecular level and evaluated its ability to generate tumors at the orthotopic site in the lung microenvironment. Further, we investigated if the self-renewal of SP cells is dependent on EGFR mediated signaling. Results SP cells were detected and isolated from multiple NSCLC cell lines (H1650, H1975, A549, as well as primary human tumor explants grown in nude mice. SP cells demonstrated stem-like properties including ability to self-renew and grow as spheres; they were able to generate primary and metastatic tumors upon orthotopic implantation into the lung of SCID mice. In vitro study revealed elevated expression of stem cell associated markers like Oct4, Sox2 and Nanog as well as demonstrated intrinsic epithelial to mesenchymal transition features in SP cells. Further, we show that abrogation of EGFR, Src and Akt signaling through pharmacological or genetic inhibitors suppresses the self-renewal growth and expansion of SP-cells and resulted in specific downregulation of Sox2 protein expression. siRNA mediated depletion of Sox2 significantly blocked the SP phenotype as well as its self-renewal capacity; whereas other transcription factors like Oct4 and Nanog played a relatively lesser role in regulating self-renewal. Interestingly, Sox2 was elevated in metastatic foci of human NSCLC samples. Conclusions Our findings suggest that Sox2 is a novel target of EGFR-Src-Akt signaling in NSCLCs that modulates self-renewal and expansion of

  2. Low adherent cancer cell subpopulations are enriched in tumorigenic and metastatic epithelial-to-mesenchymal transition-induced cancer stem-like cells. (United States)

    Morata-Tarifa, Cynthia; Jiménez, Gema; García, María A; Entrena, José M; Griñán-Lisón, Carmen; Aguilera, Margarita; Picon-Ruiz, Manuel; Marchal, Juan A


    Cancer stem cells are responsible for tumor progression, metastasis, therapy resistance and cancer recurrence, doing their identification and isolation of special relevance. Here we show that low adherent breast and colon cancer cells subpopulations have stem-like properties. Our results demonstrate that trypsin-sensitive (TS) breast and colon cancer cells subpopulations show increased ALDH activity, higher ability to exclude Hoechst 33342, enlarged proportion of cells with a cancer stem-like cell phenotype and are enriched in sphere- and colony-forming cells in vitro. Further studies in MDA-MB-231 breast cancer cells reveal that TS subpopulation expresses higher levels of SLUG, SNAIL, VIMENTIN and N-CADHERIN while show a lack of expression of E-CADHERIN and CLAUDIN, being this profile characteristic of the epithelial-to-mesenchymal transition (EMT). The TS subpopulation shows CXCL10, BMI-1 and OCT4 upregulation, differing also in the expression of several miRNAs involved in EMT and/or cell self-renewal such as miR-34a-5p, miR-34c-5p, miR-21-5p, miR-93-5p and miR-100-5p. Furthermore, in vivo studies in immunocompromised mice demonstrate that MDA-MB-231 TS cells form more and bigger xenograft tumors with shorter latency and have higher metastatic potential. In conclusion, this work presents a new, non-aggressive, easy, inexpensive and reproducible methodology to isolate prospectively cancer stem-like cells for subsequent biological and preclinical studies.

  3. Cathepsin K cleavage of SDF-1α inhibits its chemotactic activity towards glioblastoma stem-like cells. (United States)

    Hira, Vashendriya V V; Verbovšek, Urška; Breznik, Barbara; Srdič, Matic; Novinec, Marko; Kakar, Hala; Wormer, Jill; der Swaan, Britt Van; Lenarčič, Brigita; Juliano, Luiz; Mehta, Shwetal; Van Noorden, Cornelis J F; Lah, Tamara T


    Glioblastoma (GBM) is the most aggressive primary brain tumor with poor patient survival that is at least partly caused by malignant and therapy-resistant glioma stem-like cells (GSLCs) that are protected in GSLC niches. Previously, we have shown that the chemo-attractant stromal-derived factor-1α (SDF-1α), its C-X-C receptor type 4 (CXCR4) and the cysteine protease cathepsin K (CatK) are localized in GSLC niches in glioblastoma. Here, we investigated whether SDF-1α is a niche factor that through its interactions with CXCR4 and/or its second receptor CXCR7 on GSLCs facilitates their homing to niches. Furthermore, we aimed to prove that SDF-1α cleavage by CatK inactivates SDF-1α and inhibits the invasion of GSLCs. We performed mass spectrometric analysis of cleavage products of SDF-1α after proteolysis by CatK. We demonstrated that CatK cleaves SDF-1α at 3 sites in the N-terminus, which is the region of SDF-1α that binds to its receptors. Confocal imaging of human GBM tissue sections confirmed co-localization of SDF-1α and CatK in GSLC niches. In accordance, 2D and 3D invasion experiments using CXCR4/CXCR7-expressing GSLCs and GBM cells showed that SDF-1α had chemotactic activity whereas CatK cleavage products of SDF-1α did not. Besides, CXCR4 inhibitor plerixafor inhibited invasion of CXCR4/CXCR7-expressing GSLCs. In conclusion, CatK can cleave and inactivate SDF-1α. This implies that CatK activity facilitates migration of GSLCs out of niches. We propose that activation of CatK may be a promising strategy to prevent homing of GSLCs in niches and thus render these cells sensitive to chemotherapy and radiation.

  4. Cisplatin-selected resistance is associated with increased motility and stem-like properties via activation of STAT3/Snail axis in atypical teratoid/rhabdoid tumor cells (United States)

    Liu, Wei-Hsiu; Wang, Mong-Lien; Chiou, Guang-Yuh; Chien, Chian-Shiu; Huang, Pin-I; Chen, Yi-Wei; Huang, Ming-Chao; Chiou, Shih-Hwa; Shih, Yang-Hsin; Ma, Hsin-I


    Atypical teratoid/rhabdoid tumor (ATRT) is a malignant pediatric brain tumor with great recurrence after complete surgery and chemotherapy. Here, we demonstrate that cisplatin treatment selects not only for resistance but also for a more oncogenic phenotype characterized by high self-renewal and invasive capabilities. These phenomena are likely due to STAT3 upregulatoin which occurred simultaneously with higher expression of Snail, an activator of epithelial–mesenchymal transition (EMT), in ATRT-CisR cells. STAT3 knockdown effectively suppressed Snail expression and blocked motility and invasion in ATRT-CisR cells, while overexpressing Snail reversed these effects. Chromatin immunoprecipitation assay indicated that STAT3 directly bound to Snail promoter. Moreover, STAT3 knockdown effectively suppressed cancer stem-like properties, synergistically enhanced the chemotherapeutic effect, and significantly improved survival rate in ATRT-CisR-transplanted immunocompromised mice. Finally, immunohistochemistrical analysis showed that STAT3 and Snail were coexpressed at high levels in recurrent ATRT tissues. Thus, the STAT3/Snail pathway plays an important role in oncogenic resistance, rendering cells not only drug-resistant but also increasingly oncogenic (invasion, EMT and recurrence). Therefore, the STAT3/Snail could be a target for ATRT treatment. PMID:25638155

  5. γ-Secretase Inhibitor, DAPT Inhibits Self-renewal and Stemness Maintenance of Ovarian Cancer Stem-like Cells In Vitro

    Institute of Scientific and Technical Information of China (English)

    Li-yu Jiang; Xiao-lei Zhang; Ping Du; Jian-hua Zheng


    Objective: The Notch signaling pathway plays an important role in the stem cell signaling network and contributes to tumorigenesis. However, the functions of Notch signaling in ovarian cancer stem cells (OCSCs) are not well understood. We aimed to investigate the effects of Notch blockade on self-renewal and stemness maintenance of OCSCs. Methods: Ovarian cancer stem-like cells were enriched from ovarian cancer cell lines in serum-free medium. A y-secretase inhibitor, (DAPT), was used to block Notch signaling. MTT assays were performed to assess self-renewal and proliferation inhibition, flow cytometry was performed to analyze cell surface marker and immunofluorescence,Western Blot and Real-time RT-PCR assays were performed to detect Oct4 and Sox2 protein and mRNA expression of the Ovarian cancer stem-like cells treated with DAPT. Results: Notch blockade markedly inhibits self-renewal and proliferation of ovarian cancer stem-like cells,significantly downregulates the expression of OCSCs-specific surface markers, and reduces protein and mRNA expression of Oct4 and Sox2 in OCSC-like cells. Conclusion: Our results suggest that Notch signaling is not only critical for the self-renewal and proliferation of OCSCs, but also for the stemness maintenance of OCSCs. The γ-secretase inhibitor is a promising treatment targeting OCSCs.

  6. Oxaliplatin-incorporated micelles eliminate both cancer stem-like and bulk cell populations in colorectal cancer

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    Wang K


    Full Text Available Ke Wang1,*, Lina Liu2,*, Tao Zhang1, Yong-liang Zhu3, Fuming Qiu4, Xian-guo Wu1, Xiao-lei Wang1, Fu-qiang Hu5, Jian Huang1,41Cancer Institute (Key Laboratory of Cancer Prevention and Intervention, National Ministry of Education; Provincial Key Laboratory of Molecular Biology in Medical Sciences, The Second Affiliated Hospital, Zhejiang University School of Medicine; 2Department of Pharmacy, Second Affiliated Hospital (Binjiang Branch, Zhejiang University School of Medicine; 3Department of Gastroenterology; 4Department of Oncology, Second Affiliated Hospital, Zhejiang University School of Medicine; 5College of Pharmaceutical Science, Zhejiang University, Hangzhou, China, , *These authors contributed equally to this workPurpose: The failure of cancer treatments is partly due to the enrichment of cancer stem-like cells (CSLCs that are resistant to conventional chemotherapy. A novel micelle formulation of oxaliplatin (OXA encapsulated in chitosan vesicle was developed. The authors postulate that micelle encapsulation of OXA would eliminate both CSLCs and bulk cancer cells in colorectal cancer (CRC.Experimental design: In this study, using stearic acid-g-chitosan oligosaccharide (CSO-SA polymeric micelles as a drug-delivery system, OXA-loaded CSO-SA micelles (CSO-SA/OXA were prepared. Intracellular uptake of CSO-SA/OXA micelles was assessed by confocal microscope. The effects of free OXA, the empty carrier, and CSO-SA/OXA micelles were tested using human CRC cell lines in vitro and in vivo.Results: The micelles showed excellent internalization ability that increased OXA accumulation both in CRC cells and tissues. Furthermore, CSO-SA/OXA micelles could either increase the cytotoxicity of OXA against the bulk cancer cells or reverse chemoresistance of CSLC subpopulations in vitro. Intravenous administration of CSO-SA/OXA micelles effectively suppressed the tumor growth and reduced CD133+/CD24+ cell (putative CRC CSLC markers compared with free OXA

  7. Linking of mPGES-1 and iNOS activates stem-like phenotype in EGFR-driven epithelial tumor cells. (United States)

    Terzuoli, Erika; Finetti, Federica; Costanza, Filomena; Giachetti, Antonio; Ziche, Marina; Donnini, Sandra


    Inflammatory prostaglandin E-2 (PGE-2) favors cancer progression in epithelial tumors characterized by persistent oncogene input. However, its effects on tumor cell stemness are poorly understood at molecular level. Here we describe two epithelial tumor cells A431 and A459, originating from human lung and skin tumors, in which epithelial growth factor (EGF) induces sequential up-regulation of mPGES-1 and iNOS enzymes, producing an inflammatory intracellular milieu. We demonstrated that concerted action of EGF, mPGES-1 and iNOS causes sharp changes in cell phenotype demonstrated by acquisition of stem-cell features and activation of the epithelial-mesenchymal transition (EMT). When primed with EGF, epithelial tumor cells transfected with mPGES-1 or iNOS to ensure steady enzyme levels display major stem-like and EMT markers, such as reduction in E-cadherin with a concomitant rise in vimentin, ALDH-1, CD133 and ALDH activity. Tumorsphere studies with these cells show increased sphere number and size, enhanced migratory and clonogenic capacity and sharp changes in EMT markers, indicating activation of this process. The concerted action of the enzymes forms a well-orchestrated cascade where expression of iNOS depends on overexpression of mPGES-1. Indeed, we show that through its downstream effectors (PGE-2, PKA, PI3K/Akt), mPGES-1 recruits non-canonical transcription factors, thus facilitating iNOS production. In conclusion, we propose that the initial event leading to tumor stem-cell activation may be a leveraged intrinsic mechanism in which all players are either inherent constituents (EGF) or highly inducible proteins (mPGES-1, iNOS) of tumor cells. We suggest that incipient tumor aggressiveness may be moderated by reducing pivotal input of mPGES-1.

  8. Salinomycin Promotes Anoikis and Decreases the CD44+/CD24- Stem-Like Population via Inhibition of STAT3 Activation in MDA-MB-231 Cells.

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    Hyunsook An

    Full Text Available Triple-negative breast cancer (TNBC is an aggressive tumor subtype with an enriched CD44+/CD24- stem-like population. Salinomycin is an antibiotic that has been shown to target cancer stem cells (CSC; however, the mechanisms of action involved have not been well characterized. The objective of the present study was to investigate the effect of salinomycin on cell death, migration, and invasion, as well as CSC-like properties in MDA-MB-231 breast cancer cells. Salinomycin significantly induced anoikis-sensitivity, accompanied by caspase-3 and caspase-8 activation and PARP cleavage, during anchorage-independent growth. Salinomycin treatment also caused a marked suppression of cell migration and invasion with concomitant downregulation of MMP-9 and MMP-2 mRNA levels. Notably, salinomycin inhibited the formation of mammospheres and effectively reduced the CD44+/CD24- stem-like population during anchorage-independent growth. These observations were associated with the inhibition of STAT3 phosphorylation (Tyr705. Furthermore, interleukin-6 (IL-6-induced STAT3 activation was strongly suppressed by salinomycin challenge. These findings support the notion that salinomycin may be potentially efficacious for targeting breast cancer stem-like cells through the inhibition of STAT3 activation.

  9. Salinomycin Promotes Anoikis and Decreases the CD44+/CD24- Stem-Like Population via Inhibition of STAT3 Activation in MDA-MB-231 Cells. (United States)

    An, Hyunsook; Kim, Ji Young; Oh, Eunhye; Lee, Nahyun; Cho, Youngkwan; Seo, Jae Hong


    Triple-negative breast cancer (TNBC) is an aggressive tumor subtype with an enriched CD44+/CD24- stem-like population. Salinomycin is an antibiotic that has been shown to target cancer stem cells (CSC); however, the mechanisms of action involved have not been well characterized. The objective of the present study was to investigate the effect of salinomycin on cell death, migration, and invasion, as well as CSC-like properties in MDA-MB-231 breast cancer cells. Salinomycin significantly induced anoikis-sensitivity, accompanied by caspase-3 and caspase-8 activation and PARP cleavage, during anchorage-independent growth. Salinomycin treatment also caused a marked suppression of cell migration and invasion with concomitant downregulation of MMP-9 and MMP-2 mRNA levels. Notably, salinomycin inhibited the formation of mammospheres and effectively reduced the CD44+/CD24- stem-like population during anchorage-independent growth. These observations were associated with the inhibition of STAT3 phosphorylation (Tyr705). Furthermore, interleukin-6 (IL-6)-induced STAT3 activation was strongly suppressed by salinomycin challenge. These findings support the notion that salinomycin may be potentially efficacious for targeting breast cancer stem-like cells through the inhibition of STAT3 activation.

  10. Analysis of microRNA expression in canine mammary cancer stem-like cells indicates epigenetic regulation of transforming growth factor-beta signaling. (United States)

    Rybicka, A; Mucha, J; Majchrzak, K; Taciak, B; Hellmen, E; Motyl, T; Krol, M


    Cancer stem cells (CSCs) display both unique self-renewal ability as well as the ability to differentiate into many kinds of cancer cells. They are supposed to be responsible for cancer initiation, recurrence and drug resistance. Despite the fact that a variety of methods are currently employed in order to target CSCs, little is known about the regulation of their phenotype and biology by miRNAs. The aim of our study was to assess miRNA expression in canine mammary cancer stem-like cells (expressing stem cell antigen 1, Sca-1; CD44 and EpCAM) sorted from canine mammary tumour cell lines (CMT-U27, CMT-309 and P114). In order to prove their stem-like phenotype, we conducted a colony formation assay that confirmed their ability to form colonies from a single cell. Profiles of miRNA expression were investigated using Agilent custom-designed microarrays. The results were further validated by real-time rt-PCR analysis of expression of randomly selected miRNAs. Target genes were indicated and analysed using Kioto Encyclopedia of Genes and Genomes (KEGG) and BioCarta databases. The results revealed 24 down-regulated and nine up-regulated miRNAs in cancer stem-like cells compared to differentiated tumour cells. According to KEGG and BioCarta databases, target genes (n=240) of significantly down-regulated miRNAs were involved in transforming growth factor-beta signaling, mitogen-activated protein kinases (MAPK) signaling pathway, anaplastic lymphoma receptor tyrosine kinase (ALK) and peroxisome proliferator-activated receptor gamma, coactivator 1 alpha (PGC1A) pathways. The analysis of single-gene overlapping with different pathways showed that the most important genes were: TGFBR1, TGFBR2, SOS1, CHUK, PDGFRA, SMAD2, MEF2A, MEF2C and MEF2D. All of them are involved in tumor necrosis factor-beta signaling and may indicate its important role in cancer stem cell biology. Increased expression of TGFBR2, SMAD2, MEF2A and MEF2D in canine mammary cancer stem-like cells was further

  11. O6-Methylguanine-Methyltransferase (MGMT Promoter Methylation Status in Glioma Stem-Like Cells is Correlated to Temozolomide Sensitivity Under Differentiation-Promoting Conditions

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    Lucie Karayan-Tapon


    Full Text Available Glioblastoma (GBM is the most malignant type of primary brain tumor with a very poor prognosis. The actual standard protocol of treatment for GBM patients consists of radiotherapy and concomitant temozolomide (TMZ. However, the therapeutic efficacy of this treatment is limited due to tumor recurrence and TMZ resistance. Recently isolated, glioma stem-like cells (GSCs are thought to represent the population of tumorigenic cells responsible for GBM resistance and recurrence following surgery and chemotherapy. In addition, MGMT (O6-methylguanine-methyltransferase methylation is considered as one of the principal mechanisms contributing to TMZ sensitivity of GBM. In this study we have isolated GSCs from 10 adult GBM patients and investigated the relationship between MGMT methylation status and Temozolomide (TMZ sensitivity of these lines grown either in stem-like or differentiation promoting conditions. Sensitivity to TMZ was significantly associated with MGMT methylation status in cells committed to differentiation but not in stem-like cells. In addition, patients harboring highly methylated MGMT promoters had a longer overall survival. These results reveal the importance of the differentiation process when considering the predictive value of MGMT status in GSCs for clinical response to TMZ.

  12. MiR-21-5p Links Epithelial-Mesenchymal Transition Phenotype with Stem-Like Cell Signatures via AKT Signaling in Keloid Keratinocytes (United States)

    Yan, Li; Cao, Rui; Liu, Yuanbo; Wang, Lianzhao; Pan, Bo; Lv, Xiaoyan; Jiao, Hu; Zhuang, Qiang; Sun, Xuejian; Xiao, Ran


    Keloid is the abnormal wound healing puzzled by the aggressive growth and high recurrence rate due to its unrevealed key pathogenic mechanism. MicroRNAs contribute to a series of biological processes including epithelial-mesenchymal transition (EMT) and cells stemness involved in fibrotic disease. Here, using microRNAs microarray analysis we found mir-21-5p was significantly up-regulated in keloid epidermis. To investigate the role of miR-21-5p in keloid pathogenesis, we transfected miR-21-5p mimic or inhibitor in keloid keratinocytes and examined the abilities of cell proliferation, apoptosis, migration and invasion, the expressions of EMT-related markers vimentin and E-cadherin and stem-like cells-associated markers CD44 and ALDH1, and the involvement of PTEN and the signaling of AKT and ERK. Our results demonstrated that up-regulation or knockdown of miR-21-5p significantly increased or decreased the migration, invasion and sphere-forming abilities of keloid keratinocytes, and the phenotype of EMT and cells stemness were enhanced or reduced as well. Furthermore, PTEN and p-AKT were shown to participate in the regulation of miR-21-5p on EMT phenotypes and stemness signatures of keloid keratinocytes, which might account for the invasion and recurrence of keloids. This molecular mechanism of miR-21-5p on keloid keratinocytes linked EMT with cells stemness and implicated novel therapeutic targets for keloids.

  13. The effectiveness of an anti-human IL-6 receptor monoclonal antibody combined with chemotherapy to target colon cancer stem-like cells. (United States)

    Ying, Jin; Tsujii, Masahiko; Kondo, Jumpei; Hayashi, Yoshito; Kato, Motohiko; Akasaka, Tomofumi; Inoue, Takuta; Shiraishi, Eri; Inoue, Tahahiro; Hiyama, Satoshi; Tsujii, Yoshiki; Maekawa, Akira; Kawai, Shoichiro; Fujinaga, Tetsuji; Araki, Maekawa; Shinzaki, Shinichiro; Watabe, Kenji; Nishida, Tsutomu; Iijima, Hideki; Takehara, Tetsuo


    Recent studies have demonstrated that cancer stem cells (CSCs) can initiate and sustain tumor growth and exhibit resistance to clinical cytotoxic therapies. Therefore, CSCs represent the main target of anticancer therapy. Interleukin-6 (IL-6) promotes cellular proliferation and drug resistance in colorectal cancer, and its serum levels correlate with patient survival. Therefore, IL-6 and its downstream signaling molecule the signal transducer and activator of transcription-3 (STAT3) represent potential molecular targets. In the present study, we investigated the effects of IL-6 and its downstream signaling components on stem cell biology, particularly the chemoresistance of CSCs, to explore potential molecular targets for cancer therapy. The colon cancer cell line WiDr was cultured in serum-free, non-adherent, and three-dimensional spheroid-forming conditions to enrich the stem cell-like population. Spheroid-forming cells slowly proliferated and expressed high levels of Oct-4, Klf4, Bmi-1, Lgr5, IL-6, and Notch 3 compared with adherent cells. Treatment with an anti-human IL-6 receptor monoclonal antibody reduced spheroid formation, stem cell-related gene expression, and 5-fluorouracil (5-FU) resistance. In addition, IL-6 treatment enhanced the levels of p-STAT3 (Tyr705), the expression of Oct-4, Klf4, Lgr5, and Notch 3, and chemoresistance to 5-FU. siRNA targeting Notch 3 suppressed spheroid formation, Oct-4 and Lgr5 expression, and 5-FU chemoresistance, whereas STAT3 inhibition enhanced Oct-4, Klf4, Lgr5, and Notch 3 expression and 5-FU chemoresistance along with reduced spheroid growth. Taken together, these results indicate that IL-6 functions in dichotomous pathways involving Notch 3 induction and STAT3 activation. The former pathway is involved in cancer stem-like cell biology and enhanced chemoresistance, and the latter pathway leads to accelerated proliferation and reduced chemoresistance. Thus, an anti-human IL-6 receptor monoclonal antibody or Notch 3

  14. Endothelial cells create a stem cell niche in glioblastoma by providing NOTCH ligands that nurture self-renewal of cancer stem-like cells. (United States)

    Zhu, Thant S; Costello, Mark A; Talsma, Caroline E; Flack, Callie G; Crowley, Jessica G; Hamm, Lisa L; He, Xiaobing; Hervey-Jumper, Shawn L; Heth, Jason A; Muraszko, Karin M; DiMeco, Francesco; Vescovi, Angelo L; Fan, Xing


    One important function of endothelial cells in glioblastoma multiforme (GBM) is to create a niche that helps promote self-renewal of cancer stem-like cells (CSLC). However, the underlying molecular mechanism for this endothelial function is not known. Since activation of NOTCH signaling has been found to be required for propagation of GBM CSLCs, we hypothesized that the GBM endothelium may provide the source of NOTCH ligands. Here, we report a corroboration of this concept with a demonstration that NOTCH ligands are expressed in endothelial cells adjacent to NESTIN and NOTCH receptor-positive cancer cells in primary GBMs. Coculturing human brain microvascular endothelial cells (hBMEC) or NOTCH ligand with GBM neurospheres promoted GBM cell growth and increased CSLC self-renewal. Notably, RNAi-mediated knockdown of NOTCH ligands in hBMECs abrogated their ability to induce CSLC self-renewal and GBM tumor growth, both in vitro and in vivo. Thus, our findings establish that NOTCH activation in GBM CSLCs is driven by juxtacrine signaling between tumor cells and their surrounding endothelial cells in the tumor microenvironment, suggesting that targeting both CSLCs and their niche may provide a novel strategy to deplete CSLCs and improve GBM treatment.

  15. T-Bet and Eomes Regulate the Balance between the Effector/Central Memory T Cells versus Memory Stem Like T Cells.

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    Gang Li

    Full Text Available Memory T cells are composed of effector, central, and memory stem cells. Previous studies have implicated that both T-bet and Eomes are involved in the generation of effector and central memory CD8 T cells. The exact role of these transcription factors in shaping the memory T cell pool is not well understood, particularly with memory stem T cells. Here, we demonstrate that both T-bet or Eomes are required for elimination of established tumors by adoptively transferred CD8 T cells. We also examined the role of T-bet and Eomes in the generation of tumor-specific memory T cell subsets upon adoptive transfer. We showed that combined T-bet and Eomes deficiency resulted in a severe reduction in the number of effector/central memory T cells but an increase in the percentage of CD62L(highCD44(low Sca-1(+ T cells which were similar to the phenotype of memory stem T cells. Despite preserving large numbers of phenotypic memory stem T cells, the lack of both of T-bet and Eomes resulted in a profound defect in antitumor memory responses, suggesting T-bet and Eomes are crucial for the antitumor function of these memory T cells. Our study establishes that T-bet and Eomes cooperate to promote the phenotype of effector/central memory CD8 T cell versus that of memory stem like T cells.

  16. Elimination of Cancer Stem-Like “Side Population” Cells in Hepatoma Cell Lines by Chinese Herbal Mixture “Tien-Hsien Liquid”

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    Chih-Jung Yao


    Full Text Available There are increasing pieces of evidence suggesting that the recurrence of cancer may result from a small subpopulation of cancer stem cells, which are resistant to the conventional chemotherapy and radiotherapy. We investigated the effects of Chinese herbal mixture Tien-Hsien Liquid (THL on the cancer stem-like side population (SP cells isolated from human hepatoma cells. After sorting and subsequent culture, the SP cells from Huh7 hepatoma cells appear to have higher clonogenicity and mRNA expressions of stemness genes such as SMO, ABCG2, CD133, β-catenin, and Oct-4 than those of non-SP cells. At dose of 2 mg/mL, THL reduced the proportion of SP cells in HepG2, Hep3B, and Huh7 cells from 1.33% to 0.49%, 1.55% to 0.43%, and 1.69% to 0.27%, respectively. The viability and colony formation of Huh7 SP cells were effectively suppressed by THL dose-dependently, accompanied with the inhibition of stemness genes, e.g., ABCG2, CD133, and SMO. The tumorigenicity of THL-treated Huh7 SP cells in NOD/SCID mice was also diminished. Moreover, combination with THL could synergize the effect of doxorubicin against Huh7 SP cells. Our data indicate that THL may act as a cancer stem cell targeting therapeutics and be regarded as complementary and integrative medicine in the treatment of hepatoma.

  17. Isolation and Characterization of Squamous Cell Carcinoma-Derived Stem-like Cells: Role in Tumor Formation

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    Carlo Pincelli


    Full Text Available In human epidermis, keratinocyte stem cells (KSC are characterized by high levels of β1-integrin, resulting in the rapid adhesion to type IV collagen. Since epithelial tumors originate from KSC, we evaluated the features of rapidly adhering (RAD keratinocytes derived from primary human squamous cell carcinoma of the skin (cSCC. RAD cells expressed higher levels of survivin, a KSC marker, as compared to non-rapidly adhering (NRAD cells. Moreover, RAD cells proliferated to a greater extent and were more efficient in forming colonies than NRAD cells. RAD cells also migrated significantly better than NRAD cells. When seeded in a silicone chamber and grafted onto the back skin of NOD SCID mice, RAD cells formed tumors 2–4 fold bigger than those derived from NRAD cells. In tumors derived from RAD cells, the mitotic index was significantly higher than in those derived from NRAD cells, while Ki-67 and survivin expression were more pronounced in RAD tumors. This study suggests that SCC RAD stem cells play a critical role in the formation and development of epithelial tumors.

  18. Isolation and characterization of squamous cell carcinoma-derived stem-like cells: role in tumor formation. (United States)

    Dallaglio, Katiuscia; Petrachi, Tiziana; Marconi, Alessandra; Truzzi, Francesca; Lotti, Roberta; Saltari, Annalisa; Morandi, Paolo; Puviani, Mario; Maiorana, Antonino; Roop, Dennis R; Pincelli, Carlo


    In human epidermis, keratinocyte stem cells (KSC) are characterized by high levels of β1-integrin, resulting in the rapid adhesion to type IV collagen. Since epithelial tumors originate from KSC, we evaluated the features of rapidly adhering (RAD) keratinocytes derived from primary human squamous cell carcinoma of the skin (cSCC). RAD cells expressed higher levels of survivin, a KSC marker, as compared to non-rapidly adhering (NRAD) cells. Moreover, RAD cells proliferated to a greater extent and were more efficient in forming colonies than NRAD cells. RAD cells also migrated significantly better than NRAD cells. When seeded in a silicone chamber and grafted onto the back skin of NOD SCID mice, RAD cells formed tumors 2-4 fold bigger than those derived from NRAD cells. In tumors derived from RAD cells, the mitotic index was significantly higher than in those derived from NRAD cells, while Ki-67 and survivin expression were more pronounced in RAD tumors. This study suggests that SCC RAD stem cells play a critical role in the formation and development of epithelial tumors.

  19. Prostaglandin E2 receptor EP4 as the common target on cancer cells and macrophages to abolish angiogenesis, lymphangiogenesis, metastasis, and stem-like cell functions. (United States)

    Majumder, Mousumi; Xin, Xiping; Liu, Ling; Girish, Gannareddy V; Lala, Peeyush K


    We previously established that COX-2 overexpression promotes breast cancer progression and metastasis. As long-term use of COX-2 inhibitors (COX-2i) can promote thrombo-embolic events, we tested an alternative target, prostaglandin E2 receptor EP4 subtype (EP4), downstream of COX-2. Here we used the highly metastatic syngeneic murine C3L5 breast cancer model to test the role of EP4-expressing macrophages in vascular endothelial growth factor (VEGF)-C/D production, angiogenesis, and lymphangiogenesis in situ, the role of EP4 in stem-like cell (SLC) functions of tumor cells, and therapeutic effects of an EP4 antagonist RQ-15986 (EP4A). C3L5 cells expressed all EP receptors, produced VEGF-C/D, and showed high clonogenic tumorsphere forming ability in vitro, functions inhibited with COX-2i or EP4A. Treating murine macrophage RAW 264.7 cell line with COX-2i celecoxib and EP4A significantly reduced VEGF-A/C/D production in vitro, measured with quantitative PCR and Western blots. Orthotopic implants of C3L5 cells in C3H/HeJ mice showed rapid tumor growth, angiogenesis, lymphangiogenesis (CD31/LYVE-1 and CD31/PROX1 immunostaining), and metastasis to lymph nodes and lungs. Tumors revealed high incidence of EP4-expressing, VEGF-C/D producing macrophages identified with dual immunostaining of F4/80 and EP4 or VEGF-C/D. Celecoxib or EP4A therapy at non-toxic doses abrogated tumor growth, lymphangiogenesis, and metastasis to lymph nodes and lungs. Residual tumors in treated mice revealed markedly reduced VEGF-A/C/D and phosphorylated Akt/ERK proteins, VEGF-C/D positive macrophage infiltration, and proliferative/apoptotic cell ratios. Knocking down COX-2 or EP4 in C3L5 cells or treating cells in vitro with celecoxib or EP4A and treating tumor-bearing mice in vivo with the same drug reduced SLC properties of tumor cells including preferential co-expression of COX-2 and SLC markers ALDH1A, CD44, OCT-3/4, β-catenin, and SOX-2. Thus, EP4 is an excellent therapeutic target to block

  20. [Spheres isolated from Colo205 cell line possess cancer stem-like cells under serum-free culture condition]. (United States)

    Li, Ying-fei; Xiao, Bing; Lai, Zhuo-sheng; Tu, San-fang; Wang, Yuan-yuan; Zhang, Xiao-lan


    Isolation and expansion tumor spheres from colorectal cancer cell line Colo205 cultured in serum-free medium(SFM) supplemented with human recombinant EGF and bFGF. Colo205 cells were cultivated in SFM,while cells cultivated in serum-supplemented medium(SSM) served as the control. Cells morphology were observed by optical microscope, and expression of intestinal stem cells marker Musashi-1 was detected by immunocytochemical. To induce cell differentiation, tumour spheres were cultivated without EGF and bFGF in the presence of 10% serum. Then we analysed expressions of stem cell surface markers CD133 and CD44 among undifferentiated cell, post-differentiated cells and routine Colo205 cells under serum-supplemented culture condition by flow cytometry. At last we compared cell cycle and spectral karyotype between two groups. In SFM consisting of EGF and bFGF, a minority of Colo205 cells could survive, proliferate and form the suspended tumor spheres. We detected high Musashi-1 expression in these cells. Compared with the SSM group and the post-differentiation SFM group, the expressions of CD133 and CD44 were significantly increased in the undifferentiated SFM group (Pstatistical difference in the expression of CD133 and CD44 between the post-differentiation SFM group and the SSM group (P>0.05). Cell cycle analysis indicated that tumor spheres were of a high proliferation state.We could not find any noticeable difference in the number of chromatosomes between the SFM group and the SSM group. Tumor spheres in which enriched cancer stem cells can be generated under serum-free culture condition with EGF and bFGF.

  1. Astrocytes derived from trisomic human embryonic stem cells express markers of astrocytic cancer cells and premalignant stem-like progenitors

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    Iverson Linda E


    Full Text Available Abstract Background Trisomic variants of human embryonic stem cells (hESCs arise spontaneously in culture. Although trisomic hESCs share many properties with diploid hESCs, they also exhibit features of cancer stem cells. Since most hESC-based therapies will utilize differentiated derivatives, it is imperative to investigate the potential of trisomic hESCs to undergo malignant transformation during differentiation prior to their use in the clinical setting. Methods Diploid and trisomic hESCs were differentiated into astrocytic progenitors cells (APCs, RNA extracted and hybridized to human exon-specific microarrays. Global gene expression profiles of diploid and trisomic APCs were compared to that of an astrocytoma cell line and glioblastoma samples, analyzed by others, using the same microarray platform. Results Bioinformatic analysis of microarray data indicates that differentiated trisomic APCs exhibit global expression profiles with similarities to the malignant astrocytoma cell line. An analogous trend is observed in comparison to glioblastoma samples indicating that trisomic APCs express markers of astrocytic cancer cells. The analysis also allowed identification of transcripts predicted to be differentially expressed in brain tumor stem cells. These data indicate that in vitro differentiation of trisomic hESCs along astrocytic pathways give rise to cells exhibiting properties of premalignant astrocytic stem/progenitor cells. Conclusions Given their occult nature, opportunities to study premalignant stem/progenitor cells in human have been few. The ability to propagate and direct the differentiation of aneuploid hESCs provides a powerful in vitro system for investigating biological properties of human cells exhibiting features of premalignant stem cells. This in vitro culture system can be used to elucidate changes in gene expression occurring enroute to malignant transformation and to identify molecular markers of cancer stem

  2. Single cell dual adherent-suspension co-culture micro-environment for studying tumor-stromal interactions with functionally selected cancer stem-like cells. (United States)

    Chen, Yu-Chih; Zhang, Zhixiong; Fouladdel, Shamileh; Deol, Yadwinder; Ingram, Patrick N; McDermott, Sean P; Azizi, Ebrahim; Wicha, Max S; Yoon, Euisik


    Considerable evidence suggests that cancer stem-like cells (CSCs) are critical in tumor pathogenesis, but their rarity and transience has led to much controversy about their exact nature. Although CSCs can be functionally identified using dish-based tumorsphere assays, it is difficult to handle and monitor single cells in dish-based approaches; single cell-based microfluidic approaches offer better control and reliable single cell derived sphere formation. However, like normal stem cells, CSCs are heavily regulated by their microenvironment, requiring tumor-stromal interactions for tumorigenic and proliferative behaviors. To enable single cell derived tumorsphere formation within a stromal microenvironment, we present a dual adherent/suspension co-culture device, which combines a suspension environment for single-cell tumorsphere assays and an adherent environment for co-culturing stromal cells in close proximity by selectively patterning polyHEMA in indented microwells. By minimizing dead volume and improving cell capture efficiency, the presented platform allows for the use of small numbers of cells (cells). As a proof of concept, we co-cultured single T47D (breast cancer) cells and primary cancer associated fibroblasts (CAF) on-chip for 14 days to monitor sphere formation and growth. Compared to mono-culture, co-cultured T47D have higher tumorigenic potential (sphere formation rate) and proliferation rates (larger sphere size). Furthermore, 96-multiplexed single-cell transcriptome analyses were performed to compare the gene expression of co-cultured and mono-cultured T47D cells. Phenotypic changes observed in co-culture correlated with expression changes in genes associated with proliferation, apoptotic suppression, tumorigenicity and even epithelial-to-mesechymal transition. Combining the presented platform with single cell transcriptome analysis, we successfully identified functional CSCs and investigated the phenotypic and transcriptome effects induced by

  3. CEH-20/Pbx and UNC-62/Meis function upstream of rnt-1/Runx to regulate asymmetric divisions of the C. elegans stem-like seam cells

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    Samantha Hughes


    Caenorhabditis elegans seam cells divide in the stem-like mode throughout larval development, with the ability to both self-renew and produce daughters that differentiate. Seam cells typically divide asymmetrically, giving rise to an anterior daughter that fuses with the hypodermis and a posterior daughter that proliferates further. Previously we have identified rnt-1 (a homologue of the mammalian cancer-associated stem cell regulator Runx as being an important regulator of seam development, acting to promote proliferation; rnt-1 mutants have fewer seam cells whereas overexpressing rnt-1 causes seam cell hyperplasia. We isolated the interacting CEH-20/Pbx and UNC-62/Meis TALE-class transcription factors during a genome-wide RNAi screen for novel regulators of seam cell number. Animals lacking wild type CEH-20 or UNC-62 display seam cell hyperplasia, largely restricted to the anterior of the worm, whereas double mutants have many additional seam cells along the length of the animal. The cellular basis of the hyperplasia involves the symmetrisation of normally asymmetric seam cell divisions towards the proliferative stem-like fate. The hyperplasia is completely suppressed in rnt-1 mutants, and rnt-1 is upregulated in ceh-20 and unc-62 mutants, suggesting that CEH-20 and UNC-62 function upstream of rnt-1 to limit proliferative potential to the appropriate daughter cell. In further support of this we find that CEH-20 is asymmetrically localised in seam daughters following an asymmetric division, being predominantly restricted to anterior nuclei whose fate is to differentiate. Thus, ceh-20 and unc-62 encode crucial regulators of seam cell division asymmetry, acting via rnt-1 to regulate the balance between proliferation and differentiation.

  4. Curcumin improves the efficacy of cisplatin by targeting cancer stem-like cells through p21 and cyclin D1-mediated tumour cell inhibition in non-small cell lung cancer cell lines. (United States)

    Baharuddin, Puteri; Satar, Nazilah; Fakiruddin, Kamal Shaik; Zakaria, Norashikin; Lim, Moon Nian; Yusoff, Narazah Mohd; Zakaria, Zubaidah; Yahaya, Badrul Hisham


    Natural compounds such as curcumin have the ability to enhance the therapeutic effectiveness of common chemotherapy agents through cancer stem-like cell (CSC) sensitisation. In the present study, we showed that curcumin enhanced the sensitivity of the double-positive (CD166+/EpCAM+) CSC subpopulation in non-small cell lung cancer (NSCLC) cell lines (A549 and H2170) to cisplatin-induced apoptosis and inhibition of metastasis. Our results revealed that initial exposure of NSCLC cell lines to curcumin (10-40 µM) markedly reduced the percentage of viability to an average of ~51 and ~54% compared to treatment with low dose cisplatin (3 µM) with only 94 and 86% in both the A549 and H2170 cells. Moreover, sensitisation of NSCLC cell lines to curcumin through combined treatment enhanced the single effect induced by low dose cisplatin on the apoptosis of the double-positive CSC subpopulation by 18 and 20% in the A549 and H2170 cells, respectively. Furthermore, we found that curcumin enhanced the inhibitory effects of cisplatin on the highly migratory CD166+/EpCAM+ subpopulation, marked by a reduction in cell migration to 9 and 21% in the A549 and H2170 cells, respectively, indicating that curcumin may increase the sensitivity of CSCs to cisplatin-induced migratory inhibition. We also observed that the mRNA expression of cyclin D1 was downregulated, while a substantial increased in p21 expression was noted, followed by Apaf1 and caspase-9 activation in the double-positive (CD166+/EpCAM+) CSC subpopulation of A549 cells, suggested that the combined treatments induced cell cycle arrest, therefore triggering CSC growth inhibition via the intrinsic apoptotic pathway. In conclusion, we provided novel evidence of the previously unknown therapeutic effects of curcumin, either alone or in combination with cisplatin on the inhibition of the CD166+/EpCAM+ subpopulation of NSCLC cell lines. This finding demonstrated the potential therapeutic approach of using curcumin that may

  5. Temozolomide in combination with metformin act synergistically to inhibit proliferation and expansion of glioma stem-like cells (United States)



    Glioblastoma is the most common and most aggressive brain tumor in adults. The introduction of temozolomide (TMZ) has advanced chemotherapy for malignant gliomas, but it is not curative. The difficulties in treating glioblastoma may be as a result of the presence of glioma stem cells (GSCs), which are a source of relapse and chemoresistance. Another reason may be that endogenous Akt kinase activity may be activated in response to clinically relevant concentrations of TMZ. Akt activation is correlated with the increased tumorigenicity, invasiveness and stemness of cancer cells and overexpression of an active form of Akt increases glioma cell resistance to TMZ. Mounting evidence has demonstrated that cancer stem cells are preferentially sensitive to an inhibitor of Akt and down-regulation of the PI3K/Akt pathway may enhance the cytotoxicity of TMZ. Metformin (MET), the first-line drug for treating diabetes, it has been proved that it reduces AKT activation and selectively kills cancer stem cells, but whether it can potentiate the cytotoxicity of TMZ for GSCs remains unknown. In the present study, the GSCs isolated from human glioma cell line U87 and Rat glioma cell line C6, in vitro treatment with TMZ either alone or with MET. The present study demonstrates that MET acts synergistically with TMZ in inhibiting GSCs proliferation and generating the highest apoptotic rates when compared to either drug alone. These findings implicate that GSCs cytotoxicity mediated by TMZ may be stimulated by MET, have a synergistic effect, but the definite mechanisms remain elusive. PMID:27073554

  6. Identification of T cell target antigens in glioblastoma stem-like cells using an integrated proteomics-based approach in patient specimens. (United States)

    Rapp, Carmen; Warta, Rolf; Stamova, Slava; Nowrouzi, Ali; Geisenberger, Christoph; Gal, Zoltan; Roesch, Saskia; Dettling, Steffen; Juenger, Simone; Bucur, Mariana; Jungk, Christine; DaoTrong, Philip; Ahmadi, Rezvan; Sahm, Felix; Reuss, David; Fermi, Valentina; Herpel, Esther; Eckstein, Volker; Grabe, Niels; Schramm, Christoph; Weigand, Markus A; Debus, Juergen; von Deimling, Andreas; Unterberg, Andreas; Abdollahi, Amir; Beckhove, Philipp; Herold-Mende, Christel


    Glioblastoma (GBM) is a highly aggressive brain tumor and still remains incurable. Among others, an immature subpopulation of self-renewing and therapy-resistant tumor cells-often referred to as glioblastoma stem-like cells (GSCs)-has been shown to contribute to disease recurrence. To target these cells personalized immunotherapy has gained a lot of interest, e.g. by reactivating pre-existing anti-tumor immune responses against GSC antigens. To identify T cell targets commonly presented by GSCs and their differentiated counterpart, we used a proteomics-based separation of GSC proteins in combination with a T cell activation assay. Altogether, 713 proteins were identified by LC-ESI-MS/MS mass spectrometry. After a thorough filtering process, 32 proteins were chosen for further analyses. Immunogenicity of corresponding peptides was tested ex vivo. A considerable number of these antigens induced T cell responses in GBM patients but not in healthy donors. Moreover, most of them were overexpressed in primary GBM and also highly expressed in recurrent GBM tissues. Interestingly, expression of the most frequent T cell target antigens could also be confirmed in quiescent, slow-cycling GSCs isolated in high purity by the DEPArray technology. Finally, for a subset of these T cell target antigens, an association between expression levels and higher T cell infiltration as well as an increased expression of positive immune modulators was observed. In summary, we identified novel immunogenic proteins, which frequently induce tumor-specific T cell responses in GBM patients and were also detected in vitro in therapy-resistant quiescent, slow-cycling GSCs. Stable expression of these T cell targets in primary and recurrent GBM support their suitability for future clinical use.

  7. Protein phosphatase 2A mediates dormancy of glioblastoma multiforme-derived tumor stem-like cells during hypoxia.

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    Christoph P Hofstetter

    Full Text Available PURPOSE: The hypoxic microenvironment of glioblastoma multiforme (GBM is thought to increase resistance to cancer therapies. Recent evidence suggests that hypoxia induces protein phosphatase 2A (PP2A, a regulator of cell cycle and cell death. The effects of PP2A on GBM tumor cell proliferation and survival during hypoxic conditions have not been studied. EXPERIMENTAL DESIGN: Expression of PP2A subunits and HIF-α proteins was measured in 65 high-grade astrocytoma and 18 non-neoplastic surgical brain specimens by western blotting. PP2A activity was measured by an immunoprecipitation assay. For in vitro experiments, GBM-derived tumor stem cell-like cells (TSCs were exposed to severe hypoxia produced by either CoCl₂ or 1% O₂. PP2A activity was inhibited either by okadaic acid or by shRNA depletion of the PP2A C subunit. Effects of PP2A activity on cell cycle progression and cell survival during hypoxic conditions were assessed using flow cytometry. RESULTS: In our patient cohort, PP2A activity was positively correlated with HIF-1∝ protein expression (P = 0.002. Patients with PP2A activity levels above 160 pMP had significantly worse survival compared to patients with levels below this threshold (P = 0.002. PP2A activity was an independent predictor of survival on multivariable analysis (P = 0.009. In our in vitro experiments, we confirmed that severe hypoxia induces PP2A activity in TSCs 6 hours after onset of exposure. PP2A activity mediated G1/S phase growth inhibition and reduced cellular ATP consumption in hypoxic TSCs. Conversely, inhibition of PP2A activity led to increased cell proliferation, exhaustion of intracellular ATP, and accelerated P53-independent cell death of hypoxic TSCs. CONCLUSIONS: Our results suggest that PP2A activity predicts poor survival in GBM. PP2A appears to reduce the metabolic demand of hypoxic TSCs and enhances tumor cell survival. Modulation of PP2A may be a potential target for cancer therapy.

  8. Synergistic inhibition of characteristics of liver cancer stem-like cells with a combination of sorafenib and 8-bromo-7-methoxychrysin in SMMC-7721 cell line. (United States)

    Zou, Hui; Cao, Xiaozheng; Xiao, Qiao; Sheng, Xifeng; Ren, Kaiqun; Quan, Meifang; Song, Zhengwei; Li, Duo; Zheng, Yu; Zeng, Wenbin; Cao, Jianguo; Peng, Yaojin


    Sorafenib, a multi-kinase inhibitor, has shown its promising antitumor effect in a series of clinical trials, and has been approved as the current standard treatment for advanced hepatocellular carcinoma (HCC). 8-Bromo‑7-methoxychrysin (BrMC) is a novel chrysin synthetic analogue that has been reported to inhibit the growth of various tumor cells and possess properties for targeting liver cancer stem cells (LCSCs) . The present study investigated the synergistic targeting effects on the properties of liver cancer stem-like cells (LCSLCs) by a combination of sorafenib and BrMC in SMMC-7721 cell line. We also investigated whether this effect involves regulation of HIF-1α, Twist and NF-κB protein. We found that the sphere-forming cells (SFCs) from the SMMC‑7721 cells possessed the properties of LCSLCs. Sorafenib diminished the self-renewal capacity and downregulated the expression of stem cell biomarkers (CD133, CD44 and ALDH1) in a dose-dependent manner, while BrMC cooperated with sorafenib to strengthen this inhibition. Moreover, the combination of sorafenib and BrMC led to a remarkable decrease in the cellular migration and invasion, the downregulation of N-cadherin protein and upregulation of E-cadherin protein, and increase of cell apoptosis in LCSLCs. BrMC has a remarkable antagonistic effect on the upregulation of protein expression and DNA binding activity of NF-κB (p65) induced by sorafenib. In addition, our results indicated that the synergistic inhibition of sorafenib and BrMC on the characteristics of LCSLCs involves the downregulated expression of HIF-1α and EMT regulator Twist1. Collectively, the combination therapy of sorafenib and BrMC could be a new and promising therapeutic approach in the treatment of HCC.

  9. Genome-wide CRISPR-Cas9 Screens Reveal Loss of Redundancy between PKMYT1 and WEE1 in Glioblastoma Stem-like Cells

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    Chad M. Toledo


    Full Text Available To identify therapeutic targets for glioblastoma (GBM, we performed genome-wide CRISPR-Cas9 knockout (KO screens in patient-derived GBM stem-like cells (GSCs and human neural stem/progenitors (NSCs, non-neoplastic stem cell controls, for genes required for their in vitro growth. Surprisingly, the vast majority GSC-lethal hits were found outside of molecular networks commonly altered in GBM and GSCs (e.g., oncogenic drivers. In vitro and in vivo validation of GSC-specific targets revealed several strong hits, including the wee1-like kinase, PKMYT1/Myt1. Mechanistic studies demonstrated that PKMYT1 acts redundantly with WEE1 to inhibit cyclin B-CDK1 activity via CDK1-Y15 phosphorylation and to promote timely completion of mitosis in NSCs. However, in GSCs, this redundancy is lost, most likely as a result of oncogenic signaling, causing GBM-specific lethality.

  10. Arsenic trioxide depletes cancer stem-like cells and inhibits repopulation of neurosphere derived from glioblastoma by downregulation of Notch pathway. (United States)

    Wu, Jianing; Ji, Zhiyong; Liu, Huailei; Liu, Yaohua; Han, Dayong; Shi, Chen; Shi, Changbin; Wang, Chunlei; Yang, Guang; Chen, Xiaofeng; Shen, Chen; Li, Huadong; Bi, Yunke; Zhang, Dongzhi; Zhao, Shiguang


    Notch signaling has been demonstrated to have a central role in cancer stem-like cells (CSLCs) in glioblastoma multiforme (GBM). We have recently demonstrated the inhibitory effect of arsenic trioxide (ATO) on CSLCs in glioblastoma cell lines. In this study we used neurosphere recovery assay that measured neurosphere formation at three time points to assess the capacity of the culture to repopulate after ATO treatment. Our results provided strong evidence that ATO depleted CSLCs in GBM, and inhibited neurosphere recovery and secondary neurosphere formation. ATO inhibited the phosphorylation and activation of AKT and STAT3 through Notch signaling blockade. These data show that the ATO is a promising new approach to decrease glioblastoma proliferation and recurrence by downregulation of Notch pathway.

  11. Genome-wide CRISPR-Cas9 Screens Reveal Loss of Redundancy between PKMYT1 and WEE1 in Glioblastoma Stem-like Cells. (United States)

    Toledo, Chad M; Ding, Yu; Hoellerbauer, Pia; Davis, Ryan J; Basom, Ryan; Girard, Emily J; Lee, Eunjee; Corrin, Philip; Hart, Traver; Bolouri, Hamid; Davison, Jerry; Zhang, Qing; Hardcastle, Justin; Aronow, Bruce J; Plaisier, Christopher L; Baliga, Nitin S; Moffat, Jason; Lin, Qi; Li, Xiao-Nan; Nam, Do-Hyun; Lee, Jeongwu; Pollard, Steven M; Zhu, Jun; Delrow, Jeffery J; Clurman, Bruce E; Olson, James M; Paddison, Patrick J


    To identify therapeutic targets for glioblastoma (GBM), we performed genome-wide CRISPR-Cas9 knockout (KO) screens in patient-derived GBM stem-like cells (GSCs) and human neural stem/progenitors (NSCs), non-neoplastic stem cell controls, for genes required for their in vitro growth. Surprisingly, the vast majority GSC-lethal hits were found outside of molecular networks commonly altered in GBM and GSCs (e.g., oncogenic drivers). In vitro and in vivo validation of GSC-specific targets revealed several strong hits, including the wee1-like kinase, PKMYT1/Myt1. Mechanistic studies demonstrated that PKMYT1 acts redundantly with WEE1 to inhibit cyclin B-CDK1 activity via CDK1-Y15 phosphorylation and to promote timely completion of mitosis in NSCs. However, in GSCs, this redundancy is lost, most likely as a result of oncogenic signaling, causing GBM-specific lethality.

  12. Human glioblastoma stem-like cells accumulate protoporphyrin IX when subjected to exogenous 5-aminolaevulinic acid, rendering them sensitive to photodynamic treatment. (United States)

    Schimanski, Adrian; Ebbert, Lara; Sabel, Michael C; Finocchiaro, Gaetano; Lamszus, Katrin; Ewelt, Christian; Etminan, Nima; Fischer, Johannes C; Sorg, Rüdiger V


    Glioblastoma (GBM) is the most frequent and lethal primary brain tumor in adults. Despite multimodal therapy combining resection, radio- and alkylating chemotherapy, disease recurrence is universal and prognosis of patients is poor. Glioblastoma stem-like cells (GSC), which can be grown as neurospheres from primary tumors in vitro, appear to be resistant to the established therapies and are suspected to be the driving force for disease recurrence. Thus, efficacy of emerging therapies may depend on targeting GSC. 5-aminolaevulinic acid-mediated photodynamic therapy (5-ALA/PDT) is a promising therapeutic approach in GBM. It utilizes the selective accumulation of the photosensitizer protoporphyrin IX (PPIX) in GBM cells after application of 5-ALA. When exposed to laser light of 635nm wavelength, PPIX initiates a photochemical reaction resulting in the generation of reactive oxygen species, which kill the tumor cells. Whether GSC accumulate PPIX and are sensitive to 5-ALA/PDT is currently unknown. Therefore, human GSC were derived from primary tumors and grown as neurospheres under serum free conditions. When subjected to exogenous 5-ALA, a dose- and time-dependent accumulation of PPIX in GSC was observed by flow cytometry, which varied between individual GSC preparations. Subsequent exposure to laser light of 635nm wavelength substantially killed GSC, whereas treatment with 5-ALA or exposure to laser light only had no effect. LD50 values differed between GSC preparations, but were negatively correlated with PPIX accumulation in GSC. In summary, we report for the first time that glioblastoma stem-like cells accumulate PPIX when subjected to 5-aminolaevulinic acid and are sensitive to 5-aminolaevulinc acid based photodynamic therapy. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. The Caenorhabditis elegans GATA factor ELT-1 works through the cell proliferation regulator BRO-1 and the Fusogen EFF-1 to maintain the seam stem-like fate. (United States)

    Brabin, Charles; Appleford, Peter J; Woollard, Alison


    Seam cells in Caenorhabditis elegans provide a paradigm for the stem cell mode of division, with the ability to both self-renew and produce daughters that differentiate. The transcription factor RNT-1 and its DNA binding partner BRO-1 (homologues of the mammalian cancer-associated stem cell regulators RUNX and CBFβ, respectively) are known rate-limiting regulators of seam cell proliferation. Here, we show, using a combination of comparative genomics and DNA binding assays, that bro-1 expression is directly regulated by the GATA factor ELT-1. elt-1(RNAi) animals display similar seam cell lineage defects to bro-1 mutants, but have an additional phenotype in which seam cells lose their stem cell-like properties and differentiate inappropriately by fusing with the hyp7 epidermal syncytium. This phenotype is dependent on the fusogen EFF-1, which we show is repressed by ELT-1 in seam cells. Overall, our data suggest that ELT-1 has dual roles in the stem-like seam cells, acting both to promote proliferation and prevent differentiation.

  14. The Caenorhabditis elegans GATA factor ELT-1 works through the cell proliferation regulator BRO-1 and the Fusogen EFF-1 to maintain the seam stem-like fate.

    Directory of Open Access Journals (Sweden)

    Charles Brabin


    Full Text Available Seam cells in Caenorhabditis elegans provide a paradigm for the stem cell mode of division, with the ability to both self-renew and produce daughters that differentiate. The transcription factor RNT-1 and its DNA binding partner BRO-1 (homologues of the mammalian cancer-associated stem cell regulators RUNX and CBFβ, respectively are known rate-limiting regulators of seam cell proliferation. Here, we show, using a combination of comparative genomics and DNA binding assays, that bro-1 expression is directly regulated by the GATA factor ELT-1. elt-1(RNAi animals display similar seam cell lineage defects to bro-1 mutants, but have an additional phenotype in which seam cells lose their stem cell-like properties and differentiate inappropriately by fusing with the hyp7 epidermal syncytium. This phenotype is dependent on the fusogen EFF-1, which we show is repressed by ELT-1 in seam cells. Overall, our data suggest that ELT-1 has dual roles in the stem-like seam cells, acting both to promote proliferation and prevent differentiation.

  15. Sulforaphane inhibits cancer stem-like cell properties and cisplatin resistance through miR-214-mediated downregulation of c-MYC in non-small cell lung cancer (United States)

    Wu, Yue; Li, Lin-Lin; Liu, Ying; Miao, Xiao-Bo; Liu, Qiu-Zhen; Yao, Kai-Tai; Xiao, Guang-Hui


    We herein report that sulforaphane (SFN), a potent anti-cancer and well-tolerated dietary compound, inhibits cancer stem-like cell (CSC) properties and enhances therapeutic efficacy of cisplatin in human non-small cell lung cancer (NSCLC). SFN exerted these functions through upregulation of miR-214, which in turn targets the coding region of c-MYC. This finding was further corroborated by our observations that plasmid or lentiviral vector-mediated expression of 3'UTR-less c-MYC cDNA and cisplatin- or doxorubicin-induced endogenous c-MYC accumulation was similarly suppressed by either SFN or miR-214. Further, we showed that the reported inhibitory effects of SFN on β-catenin are also mediated by miR-214. SFN/miR-214 signaling inhibited CSC properties and enhanced the cytotoxicity of chemotherapeutic drugs in vitro. Experiments with nude mice carrying xenograft tumors showed that SFN sensitized NSCLC cells to cisplatin's efficacy, which is accompanied by inhibition of cisplatin-induced c-MYC accumulation in tumor tissues. Our results provided strong evidence and mechanisms to support consideration of SFN or synthetic derivatives as a therapeutic agent in combination with cisplatin for the treatment of patients with NSCLC and, potentially, other types of c-MYC-addicted tumors. PMID:28076844

  16. Loss of fructose-1,6-bisphosphatase induces glycolysis and promotes apoptosis resistance of cancer stem-like cells: an important role in hexavalent chromium-induced carcinogenesis. (United States)

    Dai, Jin; Ji, Yanli; Wang, Wei; Kim, Donghern; Fai, Leonard Yenwong; Wang, Lei; Luo, Jia; Zhang, Zhuo


    Hexavalent chromium (Cr(VI)) compounds are confirmed human carcinogens for lung cancer. Our previous studies has demonstrated that chronic exposure of human bronchial epithelial BEAS-2B cells to low dose of Cr(VI) causes malignant cell transformation. The acquisition of cancer stem cell-like properties is involved in the initiation of cancers. The present study has observed that a small population of cancer stem-like cells (BEAS-2B-Cr-CSC) exists in the Cr(VI)-transformed cells (BEAS-2B-Cr). Those BEAS-2B-Cr-CSC exhibit extremely reduced capability of generating reactive oxygen species (ROS) and apoptosis resistance. BEAS-2B-Cr-CSC are metabolic inactive as evidenced by reductions in oxygen consumption, glucose uptake, ATP production, and lactate production. Most importantly, BEAS-2B-Cr-CSC are more tumorigenic with high levels of cell self-renewal genes, Notch1 and p21. Further study has found that fructose-1,6-bisphosphatase (FBP1), an rate-limiting enzyme driving glyconeogenesis, was lost in BEAS-2B-Cr-CSC. Forced expression of FBP1 in BEAS-2B-Cr-CSC restored ROS generation, resulting in increased apoptosis, leading to inhibition of tumorigenesis. In summary, the present study suggests that loss of FBP1 is a critical event in tumorigenesis of Cr(VI)-transformed cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. The Low Chamber Pancreatic Cancer Cells Had Stem-Like Characteristics in Modified Transwell System: Is It a Novel Method to Identify and Enrich Cancer Stem-Like Cells?

    Directory of Open Access Journals (Sweden)

    Dongqing Wang


    Full Text Available Cancer stem cells (CSCs or cancer-initiating cells (CICs play an important role in tumor initiation, progression, metastasis, chemoresistance, and recurrence. It is important to construct an effective method to identify and isolate CSCs for biotherapy of cancer. During the past years, many researchers had paid more attention to it; however, this method was still on seeking. Therefore, compared to the former methods that were used to isolate the cancer stem cell, in the present study, we tried to use modified transwell system to isolate and enrich CSCs from human pancreatic cancer cell lines (Panc-1. Our results clearly showed that the lower chamber cells in modified transwell system were easily forming spheres; furthermore, these spheres expressed high levels of stem cell markers (CD133/CD44/CD24/Oct-4/ESA and exhibited chemoresistance, underwent epithelial-to-mesenchymal transition (EMT, and possessed the properties of self-renewal in vitro and tumorigenicity in vivo. Therefore, we speculated that modified transwell assay system, as a rapid and effective method, can be used to isolate and enrich CSCs.

  18. Notch signaling induces retinal stem-like properties in perinatal neural retina progenitors and promotes symmetric divisions in adult retinal stem cells. (United States)

    Balenci, Laurent; van der Kooy, Derek


    Understanding the mechanisms regulating retinal stem cell (RSC) activity is fundamental for future stem cell-based therapeutic purposes. By combining gain and loss of function approaches, we addressed whether Notch signaling may play a selective role in retinal stem versus retinal progenitor cells in both developing and adult eyes. Inhibition of either Notch or fibroblast growth factor signaling reduced proliferation of retinal stem and retinal progenitor cells, and inhibited RSC self-renewal. Conversely, exogenous Delta-like 3 and direct intrinsic Notch activation stimulated expansionary symmetric divisions in adult RSCs with the concomitant upregulation of Hes5. Knocking down Hes5 expression specifically decreased the numbers, but not the diameters, of adult RSC primary spheres, indicating that HES5 is the downstream effector of Notch receptor in controlling adult RSC proliferation. In addition, constitutive Notch activation induced retinal stem-like asymmetric self-renewal properties, with no expansion (no symmetrical division) in perinatal neural retina progenitor cells. These findings highlight central roles of Notch signaling activity in regulating the modes of division of retinal stem and retinal progenitor cells.

  19. CD133+ subpopulation of the HT1080 human fibrosarcoma cell line exhibits cancer stem-like characteristics. (United States)

    Feng, Bao-Hua; Liu, Ai-Guo; Gu, Wen-Guang; Deng, Liang; Cheng, Xian-Gyang; Tong, Tie-Jun; Zhang, Hong-Zhi


    The cancer stem cell (CSC) theory holds that a minority population within tumors possesses stem cell properties of self-renewal and multilineage differentiation capacity and provides the initiating cells from which tumors are derived and sustained. However, verifying the existence of these CSCs has been a significant challenge. The CD133 antigen is a pentaspan membrane glycoprotein proposed to be a CSC marker for cancer-initiating subpopulations in the brain, colon and various other tissues. Here, CD133+ cells were obtained and characterized from the HT1080 cell line to determine the utility of this marker for isolating CSCs from human fibrosarcoma cells. In this study, CD133+ cells were separated from HT1080 cells using magnetic beads and characterized for their proliferation rate and resistance to chemotherapeutic drugs, cisplatin and doxorubicin, by MTS assay. Relative expression of tumor-associated genes Sox2, Oct3/4, Nanog, c-Myc, Bmi-1 and ABCG2 was measured by real-time polymerase chain reaction (PCR). Clonal sphere formation and the ability of CD133+ cells to initiate tumors in BALB/c nude mice was also evaluated. We found that CD133+ cells showed a high proliferation rate, increased resistance to chemotherapy drugs and overexpression of tumor-associated genes compared with these features in CD133- cells. Additionally, CD133+ cells were able to form spherical clusters in serum-free medium with high clonogenic efficiency, indicating a significantly greater tumor-initiating potential when compared with CD133- cells. These findings indicate that CD133+ cells identified within the HT1080 human fibrosarcoma cell line possess many CSC properties and may facilitate the development of improved therapies for fibrosarcoma.

  20. Cancer stem-like cells enriched with CD29 and CD44 markers exhibit molecular characteristics with epithelial-mesenchymal transition in squamous cell carcinoma. (United States)

    Geng, Songmei; Guo, Yuanyuan; Wang, Qianqian; Li, Lan; Wang, Jianli


    Increasing evidences have indicated that only a phenotypic subset of cancer cells, termed as the cancer stem cells (CSCs), is capable of initiating tumor growth and provide a reservoir of cells that cause tumor recurrence after therapy. Epithelial-mesenchymal transition (EMT), a cell type change from an epithelial cobblestone phenotype to an elongated fibroblastic phenotype, plays a critical role not only in tumor metastasis but also in tumor recurrence and contributes to drug resistance. Accumulating evidence has shown that cells with an EMT phenotype are rich sources for CSCs, suggesting a biological link between EMT and CSCs; thus study on the link will help understand the cellular and molecular mechanisms of tumor metastasis and drug resistance. CD29 is involved in EMT through cross-talk with cadherins and CD44 has been reported as a successful used marker for CSCs. Here, we try to address whether combination of CD29 and CD44 could be used to identify cancer stem-like cells undergoing EMT in squamous cell carcinoma (SCC) and compare the molecular differences between CD29high/CD44high and CD29low/CD44low cells in SCC. Expression pattern of CD29 and CD44 was analyzed in tissues of skin SCC and cultured A431 cells by immunostaining. Subtype cells of CD29high/CD44high and CD29low/CD44low A431 were sorted by fluorescence-activated cell sorting and proliferating abilities were assayed by cell counting, colony forming and tumorigenicity in NOD/SCID mice. Finally, to probe more deeply into the molecular differences between CD29high/CD44high and CD29low/CD44low A431 cells, gene microarray analysis was applied to compare gene expression profiling. Staining of CD29 and CD44 showed similar heterogeneous expression pattern with positive cells located in the invasion front of SCC tissue as well as in cultured A431 cells. Sorted CD29high/CD44high A431 cells had higher proliferating ability in vitro and in NOD/SCID mice as compared with CD29low/CD44low cells. Gene profiling

  1. Epithelial-mesenchymal transition associates with maintenance of stemness in spheroid-derived stem-like colon cancer cells.

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    Xiao-Yan Han

    Full Text Available Despite earlier studies demonstrating characteristics of colon cancer stem cells (CCSCs and the role of epithelial-mesenchymal transition (EMT in tumor development, it remains controversial as to the relationship between CCSCs and EMT. In this study, in order to present an insight into this relationship in colon cancer, we developed HCT116 and HT29 sphere models, which are known to be the cells enriching cancer stem cells. Compared to their parental counterparts, spheroid cells displayed lower homotypic/heterotypic adhesion but higher in vitro migratory/invasive capacity, as well as higher tumorigenic and metastatic potential in vivo. The spheroid cells also demonstrated down-regulated E-cadherin and up-regulated α-SMA and Vimentin expression, which is the typical phenotype of EMT. In order to explore whether this phenomenon is associated to activation of Wnt/β-catenin pathway, we detected several key signaling molecules. Compared with their parental cells, HCT116 and HT29 spheroid cells demonstrated down-regulated expression of GSK3β, but up-regulated expression of Slug and Snail. And also, the up-regulation of nucleus β-catenin in spheroid cells indicated that the free β-catenin transferred from cytoplasm to cell nucleus. Our findings indicate that spheroid cells have the characteristics of colon cancer stem cells, and EMT may account for their stemness and malignancy. And persistent activation of Wnt/β-catenin pathway may play an important role in the EMT of CCSCs.

  2. High linear-energy-transfer radiation can overcome radioresistance of glioma stem-like cells to low linear-energy-transfer radiation (United States)

    Hirota, Yuki; Masunaga, Shin-Ichiro; Kondo, Natsuko; Kawabata, Shinji; Hirakawa, Hirokazu; Yajima, Hirohiko; Fujimori, Akira; Ono, Koji; Kuroiwa, Toshihiko; Miyatake, Shin-Ichi


    Ionizing radiation is applied as the standard treatment for glioblastoma multiforme (GBM). However, radiotherapy remains merely palliative, not curative, because of the existence of glioma stem cells (GSCs), which are regarded as highly radioresistant to low linear-energy-transfer (LET) photons. Here we analyzed whether or not high-LET particles can overcome the radioresistance of GSCs. Glioma stem-like cells (GSLCs) were induced from the GBM cell line A172 in stem cell culture medium. The phenotypes of GSLCs and wild-type cells were confirmed using stem cell markers. These cells were irradiated with 60Co gamma rays or reactor neutron beams. Under neutron-beam irradiation, high-LET proton particles can be produced through elastic scattering or nitrogen capture reaction. Radiosensitivity was assessed by a colony-forming assay, and the DNA double-strand breaks (DSBs) were assessed by a histone gamma-H2AX focus detection assay. In stem cell culture medium, GSLCs could form neurosphere-like cells and express neural stem cell markers (Sox2 and Musashi) abundantly in comparison with their parental cells. GSLCs were significantly more radioresistant to gamma rays than their parental cells, but neutron beams overcame this resistance. There were significantly fewer gamma-H2AX foci in the A172 GSLCs 24 h after irradiation with gamma rays than in their parental cultured cells, while there was no apparent difference following neutron-beam irradiation. High-LET radiation can overcome the radioresistance of GSLCs by producing unrepairable DNA DSBs. High-LET radiation therapy might have the potential to overcome GBM's resistance to X-rays in a clinical setting. PMID:23955054

  3. GP73-regulated oncolytic adenoviruses possess potent killing effect on human liver cancer stem-like cells (United States)

    Zhang, Rong; Ma, Buyun; Liu, Tao; Yang, Yu; Xie, Wenjie; Liu, Xianglei; Huang, Fang; Liu, Tao; Zhou, Xiumei; Liu, Xinyuan; Wang, Yigang


    Cancer stem cells (CSCs), also known as tumor-initiating cells, are highly metastatic, chemo-resistant and tumorigenic, and are critical for cancer development, maintenance and recurrence. Oncolytic adenovirus could targetedly kill CSCs and has been acted as a promising anticancer agent. Currently, a novel GP73-regulated oncolytic adenovirus GD55 was constructed to specifically treat liver cancer and exhibited obvious cytotoxicity effect. However, there remains to be confirmed that whether GD55 could effectively eliminate liver CSCs. We first utilized the suspension culture to enrich the liver CSCs-like cells, which acquires the properties of liver CSCs in self-renewal, differentiation, quiescence, chemo-resistance and tumorigenicity. The results indicated that GD55 elicited more significant cytotoxicity and stronger oncolytic effect in liver CSC-like cells compared to common oncolytic virus ZD55. Additionally, GD55 possessed the greater efficacy in suppressing the growth of implanted tumors derived from liver CSC-like cells than ZD55. Furthermore, GD55 induced remarkable apoptosis of liver CSC-like cells in vitro and in vivo, and inhibited the propogation of cells and angiogenesis in xenograft tumor tissues. Thus, GD55 may virtually represent an attractive therapeutic agent for targeting liver CSCs to achieve better clinical outcomes for HCC patients. PMID:27121064

  4. Breast cancer stem-like cells are inhibited by a non-toxic aryl hydrocarbon receptor agonist.

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    Gérald J Prud'homme

    Full Text Available BACKGROUND: Cancer stem cells (CSCs have increased resistance to cancer chemotherapy. They can be enriched as drug-surviving CSCs (D-CSCs by growth with chemotherapeutic drugs, and/or by sorting of cells expressing CSC markers such as aldehyde dehydrogenase-1 (ALDH. CSCs form colonies in agar, mammospheres in low-adherence cultures, and tumors following xenotransplantation in Scid mice. We hypothesized that tranilast, a non-toxic orally active drug with anti-cancer activities, would inhibit breast CSCs. METHODOLOGY/FINDINGS: We examined breast cancer cell lines or D-CSCs generated by growth of these cells with mitoxantrone. Tranilast inhibited colony formation, mammosphere formation and stem cell marker expression. Mitoxantrone-selected cells were enriched for CSCs expressing stem cell markers ALDH, c-kit, Oct-4, and ABCG2, and efficient at forming mammospheres. Tranilast markedly inhibited mammosphere formation by D-CSCs and dissociated formed mammospheres, at pharmacologically relevant concentrations. It was effective against D-CSCs of both HER-2+ and triple-negative cell lines. Tranilast was also effective in vivo, since it prevented lung metastasis in mice injected i.v. with triple-negative (MDA-MB-231 mitoxantrone-selected cells. The molecular targets of tranilast in cancer have been unknown, but here we demonstrate it is an aryl hydrocarbon receptor (AHR agonist and this plays a key role. AHR is a transcription factor activated by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, polycyclic aromatic hydrocarbons and other ligands. Tranilast induced translocation of the AHR to the nucleus and stimulated CYP1A1 expression (a marker of AHR activation. It inhibited binding of the AHR to CDK4, which has been linked to cell-cycle arrest. D-CSCs expressed higher levels of the AHR than other cells. Knockdown of the AHR with siRNA, or blockade with an AHR antagonist, entirely abrogated the anti-proliferative and anti-mammosphere activity of tranilast

  5. Evolution of Cancer Stem-like Cells in Endocrine-Resistant Metastatic Breast Cancers Is Mediated by Stromal Microvesicles. (United States)

    Sansone, Pasquale; Berishaj, Marjan; Rajasekhar, Vinagolu K; Ceccarelli, Claudio; Chang, Qing; Strillacci, Antonio; Savini, Claudia; Shapiro, Lauren; Bowman, Robert L; Mastroleo, Chiara; De Carolis, Sabrina; Daly, Laura; Benito-Martin, Alberto; Perna, Fabiana; Fabbri, Nicola; Healey, John H; Spisni, Enzo; Cricca, Monica; Lyden, David; Bonafé, Massimiliano; Bromberg, Jacqueline


    The hypothesis that microvesicle-mediated miRNA transfer converts noncancer stem cells into cancer stem cells (CSC) leading to therapy resistance remains poorly investigated. Here we provide direct evidence supporting this hypothesis, by demonstrating how microvesicles derived from cancer-associated fibroblasts (CAF) transfer miR-221 to promote hormonal therapy resistance (HTR) in models of luminal breast cancer. We determined that CAF-derived microvesicles horizontally transferred miR-221 to tumor cells and, in combination with hormone therapy, activated an ER(lo)/Notch(hi) feed-forward loop responsible for the generation of CD133(hi) CSCs. Importantly, microvesicles from patients with HTR metastatic disease expressed high levels of miR-221. We further determined that the IL6-pStat3 pathway promoted the biogenesis of onco-miR-221(hi) CAF microvesicles and established stromal CSC niches in experimental and patient-derived breast cancer models. Coinjection of patient-derived CAFs from bone metastases led to de novo HTR tumors, which was reversed with IL6R blockade. Finally, we generated patient-derived xenograft (PDX) models from patient-derived HTR bone metastases and analyzed tumor cells, stroma, and microvesicles. Murine and human CAFs were enriched in HTR tumors expressing high levels of CD133(hi) cells. Depletion of murine CAFs from PDX restored sensitivity to HT, with a concurrent reduction of CD133(hi) CSCs. Conversely, in models of CD133(neg), HT-sensitive cancer cells, both murine and human CAFs promoted de novo HT resistance via the generation of CD133(hi) CSCs that expressed low levels of estrogen receptor alpha. Overall, our results illuminate how microvesicle-mediated horizontal transfer of genetic material from host stromal cells to cancer cells triggers the evolution of therapy-resistant metastases, with potentially broad implications for their control. Cancer Res; 77(8); 1927-41. ©2017 AACR. ©2017 American Association for Cancer Research.

  6. Phenethyl isothiocyanate upregulates death receptors 4 and 5 and inhibits proliferation in human cancer stem-like cells


    Wang, Dan; Upadhyaya, Bijaya; Liu, Yi; Knudsen, David; Dey, Moul


    Background The cytokine TRAIL (tumor necrotic factor-related apoptosis-inducing ligand) selectively induces apoptosis in cancer cells, but cancer stem cells (CSCs) that contribute to cancer-recurrence are frequently TRAIL-resistant. Here we examined hitherto unknown effects of the dietary anti-carcinogenic compound phenethyl isothiocyanate (PEITC) on attenuation of proliferation and tumorigenicity and on up regulation of death receptors and apoptosis in human cervical CSC. Methods Cancer stem...

  7. CD133 Modulate HIF-1α Expression under Hypoxia in EMT Phenotype Pancreatic Cancer Stem-Like Cells

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    Koki Maeda


    Full Text Available Although CD133 is a known representative cancer stem cell marker, its function in tumor aggressiveness under hypoxia is not fully known. The aim of this study is to demonstrate that CD133 regulates hypoxia inducible factor (HIF-1α expression with tumor migration. The CD133+ pancreatic cancer cell line, Capan1M9, was compared with the CD133− cell line, shCD133M9, under hypoxia. HIF-1α expression levels were compared by Western blot, HIF-1α nucleus translocation assay and real-time (RT-PCR. The hypoxia responsive element (HRE was observed by luciferase assay. The migration ability was analyzed by migration and wound healing assays. Epithelial mesenchymal transition (EMT related genes were analyzed by real-time RT-PCR. HIF-1α was highly expressed in Capan1M9 compared to shCD133M9 under hypoxia because of the high activation of HRE. Furthermore, the migration ability of Capan1M9 was higher than that of shCD133M9 under hypoxia, suggesting higher expression of EMT related genes in Capan1M9 compared to shCD133M9. Conclusion: HIF-1α expression under hypoxia in CD133+ pancreatic cancer cells correlated with tumor cell migration through EMT gene expression. Understanding the function of CD133 in cancer aggressiveness provides a novel therapeutic approach to eradicate pancreatic cancer stem cells.

  8. p38γ MAPK Is a Therapeutic Target for Triple-Negative Breast Cancer by Stimulation of Cancer Stem-Like Cell Expansion. (United States)

    Qi, Xiaomei; Yin, Ning; Ma, Shao; Lepp, Adrienne; Tang, Jun; Jing, Weiqing; Johnson, Bryon; Dwinell, Michael B; Chitambar, Christopher R; Chen, Guan


    Triple-negative breast cancer (TNBC) is highly progressive and lacks established therapeutic targets. p38γ mitogen-activated protein kinase (MAPK) (gene name: MAPK12) is overexpressed in TNBC but how overexpressed p38γ contributes to TNBC remains unknown. Here, we show that p38γ activation promotes TNBC development and progression by stimulating cancer stem-like cell (CSC) expansion and may serve as a novel therapeutic target. p38γ silencing in TNBC cells reduces mammosphere formation and decreases expression levels of CSC drivers including Nanog, Oct3/4, and Sox2. Moreover, p38γ MAPK-forced expression alone is sufficient to stimulate CSC expansion and to induce epithelial cell transformation in vitro and in vivo. Furthermore, p38γ depends on its activity to stimulate CSC expansion and breast cancer progression, indicating a therapeutic opportunity by application of its pharmacological inhibitor. Indeed, the non-toxic p38γ specific pharmacological inhibitor pirfenidone selectively inhibits TNBC growth in vitro and/or in vivo and significantly decreases the CSC population. Mechanistically, p38γ stimulates Nanog transcription through c-Jun/AP-1 via a multi-protein complex formation. These results together demonstrate that p38γ can drive TNBC development and progression and may be a novel therapeutic target for TNBC by stimulating CSC expansion. Inhibiting p38γ activity with pirfenidone may be a novel strategy for the treatment of TNBC.

  9. Inhibition of telomerase activity preferentially targets aldehyde dehydrogenase-positive cancer stem-like cells in lung cancer

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    Iniesta Pilar


    Full Text Available Abstract Background Mortality rates for advanced lung cancer have not declined for decades, even with the implementation of novel chemotherapeutic regimens or the use of tyrosine kinase inhibitors. Cancer Stem Cells (CSCs are thought to be responsible for resistance to chemo/radiotherapy. Therefore, targeting CSCs with novel compounds may be an effective approach to reduce lung tumor growth and metastasis. We have isolated and characterized CSCs from non-small cell lung cancer (NSCLC cell lines and measured their telomerase activity, telomere length, and sensitivity to the novel telomerase inhibitor MST312. Results The aldehyde dehydrogenase (ALDH positive lung cancer cell fraction is enriched in markers of stemness and endowed with stem cell properties. ALDH+ CSCs display longer telomeres than the non-CSC population. Interestingly, MST312 has a strong antiproliferative effect on lung CSCs and induces p21, p27 and apoptosis in the whole tumor population. MST312 acts through activation of the ATM/pH2AX DNA damage pathway (short-term effect and through decrease in telomere length (long-term effect. Administration of this telomerase inhibitor (40 mg/kg in the H460 xenograft model results in significant tumor shrinkage (70% reduction, compared to controls. Combination therapy consisting of irradiation (10Gy plus administration of MST312 did not improve the therapeutic efficacy of the telomerase inhibitor alone. Treatment with MST312 reduces significantly the number of ALDH+ CSCs and their telomeric length in vivo. Conclusions We conclude that antitelomeric therapy using MST312 mainly targets lung CSCs and may represent a novel approach for effective treatment of lung cancer.

  10. Nucleolin overexpression in breast cancer cell sub-populations with different stem-like phenotype enables targeted intracellular delivery of synergistic drug combination. (United States)

    Fonseca, Nuno A; Rodrigues, Ana S; Rodrigues-Santos, Paulo; Alves, Vera; Gregório, Ana C; Valério-Fernandes, Ângela; Gomes-da-Silva, Lígia C; Rosa, Manuel Santos; Moura, Vera; Ramalho-Santos, João; Simões, Sérgio; Moreira, João Nuno


    Breast cancer stem cells (CSC) are thought responsible for tumor growth and relapse, metastization and active evasion to standard chemotherapy. The recognition that CSC may originate from non-stem cancer cells (non-SCC) through plastic epithelial-to-mesenchymal transition turned these into relevant cell targets. Of crucial importance for successful therapeutic intervention is the identification of surface receptors overexpressed in both CSC and non-SCC. Cell surface nucleolin has been described as overexpressed in cancer cells as well as a tumor angiogenic marker. Herein we have addressed the questions on whether nucleolin was a common receptor among breast CSC and non-SCC and whether it could be exploited for targeting purposes. Liposomes functionalized with the nucleolin-binding F3 peptide, targeted simultaneously, nucleolin-overexpressing putative breast CSC and non-SCC, which was paralleled by OCT4 and NANOG mRNA levels in cells from triple negative breast cancer (TNBC) origin. In murine embryonic stem cells, both nucleolin mRNA levels and F3 peptide-targeted liposomes cellular association were dependent on the stemness status. An in vivo tumorigenic assay suggested that surface nucleolin overexpression per se, could be associated with the identification of highly tumorigenic TNBC cells. This proposed link between nucleolin expression and the stem-like phenotype in TNBC, enabled 100% cell death mediated by F3 peptide-targeted synergistic drug combination, suggesting the potential to abrogate the plasticity and adaptability associated with CSC and non-SCC. Ultimately, nucleolin-specific therapeutic tools capable of simultaneous debulk multiple cellular compartments of the tumor microenvironment may pave the way towards a specific treatment for TNBC patient care.

  11. Salinomycin possesses anti-tumor activity and inhibits breast cancer stem-like cells via an apoptosis-independent pathway. (United States)

    An, Hyunsook; Kim, Ji Young; Lee, Nahyun; Cho, Youngkwan; Oh, Eunhye; Seo, Jae Hong


    Cancer stem cells (CSCs) play important roles in the formation, growth and recurrence of tumors, particularly following therapeutic intervention. Salinomycin has received recent attention for its ability to target breast cancer stem cells (BCSCs), but the mechanisms of action involved are not fully understood. In the present study, we sought to investigate the mechanisms responsible for salinomycin's selective targeting of BCSCs and its anti-tumor activity. Salinomycin suppressed cell viability, concomitant with the downregulation of cyclin D1 and increased p27(kip1) nuclear accumulation. Mammosphere formation assays revealed that salinomycin suppresses self-renewal of ALDH1-positive BCSCs and downregulates the transcription factors Nanog, Oct4 and Sox2. TUNEL analysis of MDA-MB-231-derived xenografts revealed that salinomycin administration elicited a significant reduction in tumor growth with a marked downregulation of ALDH1 and CD44 levels, but seemingly without the induction of apoptosis. Our findings shed further light on the mechanisms responsible for salinomycin's effects on BCSCs.

  12. Combination Treatment with PPARγ Ligand and Its Specific Inhibitor GW9662 Downregulates BIS and 14-3-3 Gamma, Inhibiting Stem-Like Properties in Glioblastoma Cells

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    Chang-Nim Im


    Full Text Available PPARγ is a nuclear receptor that regulates differentiation and proliferation and is highly expressed in many cancer cells. Its synthetic ligands, such as rosiglitazone and ciglitazone, and its inhibitor GW9662, were shown to induce cellular differentiation, inhibit proliferation, and lead to apoptosis. Glioblastoma is a common brain tumor with poor survival prospects. Recently, glioblastoma stem cells (GSCs have been examined as a potential target for anticancer therapy; however, little is known about the combined effect of various agents on GSCs. In this study, we found that cotreatment with PPARγ ligands and GW9662 inhibited stem-like properties in GSC-like spheres, which significantly express SOX2. In addition, this treatment decreased the activation of STAT3 and AKT and decreased the amounts of 14-3-3 gamma and BIS proteins. Moreover, combined administration of small-interfering RNA (siRNA transfection with PPARγ ligands induced downregulation of SOX2 and MMP2 activity together with inhibition of sphere-forming activity regardless of poly(ADP-ribose polymerase (PARP cleavage. Taken together, our findings suggest that a combination therapy using PPARγ ligands and its inhibitor could be a potential therapeutic strategy targeting GSCs.

  13. Kinome-wide shRNA Screen Identifies the Receptor Tyrosine Kinase AXL as a Key Regulator for Mesenchymal Glioblastoma Stem-like Cells

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    Peng Cheng


    Full Text Available Glioblastoma is a highly lethal cancer for which novel therapeutics are urgently needed. Two distinct subtypes of glioblastoma stem-like cells (GSCs were recently identified: mesenchymal (MES and proneural (PN. To identify mechanisms to target the more aggressive MES GSCs, we combined transcriptomic expression analysis and kinome-wide short hairpin RNA screening of MES and PN GSCs. In comparison to PN GSCs, we found significant upregulation and phosphorylation of the receptor tyrosine kinase AXL in MES GSCs. Knockdown of AXL significantly decreased MES GSC self-renewal capacity in vitro and inhibited the growth of glioblastoma patient-derived xenografts. Moreover, inhibition of AXL with shRNA or pharmacologic inhibitors also increased cell death significantly more in MES GSCs. Clinically, AXL expression was elevated in the MES GBM subtype and significantly correlated with poor prognosis in multiple cancers. In conclusion, we identified AXL as a potential molecular target for novel approaches to treat glioblastoma and other solid cancers.

  14. Cannabidiol stimulates Aml-1a-dependent glial differentiation and inhibits glioma stem-like cells proliferation by inducing autophagy in a TRPV2-dependent manner. (United States)

    Nabissi, Massimo; Morelli, Maria Beatrice; Amantini, Consuelo; Liberati, Sonia; Santoni, Matteo; Ricci-Vitiani, Lucia; Pallini, Roberto; Santoni, Giorgio


    Glioma stem-like cells (GSCs) correspond to a tumor cell subpopulation, involved in glioblastoma multiforme (GBM) tumor initiation and acquired chemoresistance. Currently, drug-induced differentiation is considered as a promising approach to eradicate this tumor-driving cell population. Recently, the effect of cannabinoids (CBs) in promoting glial differentiation and inhibiting gliomagenesis has been evidenced. Herein, we demonstrated that cannabidiol (CBD) by activating transient receptor potential vanilloid-2 (TRPV2) triggers GSCs differentiation activating the autophagic process and inhibits GSCs proliferation and clonogenic capability. Above all, CBD and carmustine (BCNU) in combination overcome the high resistance of GSCs to BCNU treatment, by inducing apoptotic cell death. Acute myeloid leukemia (Aml-1) transcription factors play a pivotal role in GBM proliferation and differentiation and it is known that Aml-1 control the expression of several nociceptive receptors. So, we evaluated the expression levels of Aml-1 spliced variants (Aml-1a, b and c) in GSCs and during their differentiation. We found that Aml-1a is upregulated during GSCs differentiation, and its downregulation restores a stem cell phenotype in differentiated GSCs. Since it was demonstrated that CBD induces also TRPV2 expression and that TRPV2 is involved in GSCs differentiation, we evaluated if Aml-1a interacted directly with TRPV2 promoters. Herein, we found that Aml-1a binds TRPV2 promoters and that Aml-1a expression is upregulated by CBD treatment, in a TRPV2 and PI3K/AKT dependent manner. Altogether, these results support a novel mechanism by which CBD inducing TRPV2-dependent autophagic process stimulates Aml-1a-dependent GSCs differentiation, abrogating the BCNU chemoresistance in GSCs.

  15. The long non-coding RNA – HIF1A-AS2 facilitates the maintenance of mesenchymal glioblastoma stem-like cells in hypoxic niches (United States)

    Mineo, Marco; Ricklefs, Franz; Rooj, Arun K.; Lyons, Shawn M.; Ivanov, Pavel; Ansari, Khairul I.; Nakano, Ichiro; Chiocca, E. Antonio; Godlewski, Jakub; Bronisz, Agnieszka


    Long-non-coding RNAs (lncRNAs) have an undefined role in the pathobiology of glioblastoma multiforme (GBM). These tumors are genetically and phenotypically heterogeneous with transcriptome subtype-specific GBM stem-like cells (GSCs) that adapt to the brain tumor microenvironment, including hypoxic niches. We identified hypoxia inducible factor 1 alpha-antisense RNA 2 (HIF1A-AS2) as a subtype-specific hypoxia inducible lncRNA, up-regulated in mesenchymal GSCs. Its deregulation affects GSC growth, self-renewal and hypoxia-dependent molecular reprogramming. Amongst the HIF1A-AS2 interactome, IGF2BP2 and DHX9 were identified as direct partners. This association was needed for maintenance of expression of their target gene, HMGA1. Down-regulation of HIF1A-AS2 led to delayed growth of mesenchymal GSC tumors, survival benefits, and impaired expression of HMGA1 in vivo. Our data demonstrate that HIF1A-AS2 contributes to GSCs’ speciation and adaptation to hypoxia within the tumor microenvironment, acting directly through its interactome/targets and indirectly by modulating responses to hypoxic stress depending on the subtype-specific genetic context. PMID:27264189

  16. The Long Non-coding RNA HIF1A-AS2 Facilitates the Maintenance of Mesenchymal Glioblastoma Stem-like Cells in Hypoxic Niches

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    Marco Mineo


    Full Text Available Long non-coding RNAs (lncRNAs have an undefined role in the pathobiology of glioblastoma multiforme (GBM. These tumors are genetically and phenotypically heterogeneous with transcriptome subtype-specific GBM stem-like cells (GSCs that adapt to the brain tumor microenvironment, including hypoxic niches. We identified hypoxia-inducible factor 1 alpha-antisense RNA 2 (HIF1A-AS2 as a subtype-specific hypoxia-inducible lncRNA, upregulated in mesenchymal GSCs. Its deregulation affects GSC growth, self-renewal, and hypoxia-dependent molecular reprogramming. Among the HIF1A-AS2 interactome, IGF2BP2 and DHX9 were identified as direct partners. This association was needed for maintenance of expression of their target gene, HMGA1. Downregulation of HIF1A-AS2 led to delayed growth of mesenchymal GSC tumors, survival benefits, and impaired expression of HMGA1 in vivo. Our data demonstrate that HIF1A-AS2 contributes to GSCs’ speciation and adaptation to hypoxia within the tumor microenvironment, acting directly through its interactome and targets and indirectly by modulating responses to hypoxic stress depending on the subtype-specific genetic context.

  17. Garcinol from Garcinia indica Downregulates Cancer Stem-like Cell Biomarker ALDH1A1 in Nonsmall Cell Lung Cancer A549 Cells through DDIT3 Activation. (United States)

    Wang, Jinhan; Wang, Liwen; Ho, Chi-Tang; Zhang, Kunsheng; Liu, Qiang; Zhao, Hui


    Nonsmall cell lung cancer (NSCLC) is the predominant type of lung cancer. Patients with NSCLC show high mortality rates because of failure to clean up cancer stem cells (CSCs). The anticancer activity of phytochemical garcinol has been identified in various cancer cell models. However, the effect of garcinol on NSCLC cell lines is still lacking. Of the NSCLC cell lines we tested, A549 cells were the most sensitive to garcinol. Interestingly, Aldehyde Dehydrogenase 1 Family Member A1 (ALDH1A1) was preferentially expressed in A549 cells and downregulated by the addition of garcinol. We also found that garcinol enriched DNA damage-inducible transcript 3 (DDIT3) and then altered DDIT3-CCAAT-enhancer-binding proteins beta (C/EBPβ) interaction resulting in a decreased binding of C/EBPβ to the endogenous ALDH1A1 promoter. Furthermore, garcinol's inhibition of ALDH1A1 was identified in a xenograft mice model. Garcinol repressed ALDH1A1 transcription in A549 cells through alterations in the interaction between DDIT3 and C/EBPβ. Garcinol could be a potential dietary phytochemical candidate for NSCLCs patients whose tumors harbored high ALDH1A1 expression.

  18. Comparative expression study of the endo-G protein coupled receptor (GPCR repertoire in human glioblastoma cancer stem-like cells, U87-MG cells and non malignant cells of neural origin unveils new potential therapeutic targets.

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    Marie Fève

    Full Text Available Glioblastomas (GBMs are highly aggressive, invasive brain tumors with bad prognosis and unmet medical need. These tumors are heterogeneous being constituted by a variety of cells in different states of differentiation. Among these, cells endowed with stem properties, tumor initiating/propagating properties and particularly resistant to chemo- and radiotherapies are designed as the real culprits for tumor maintenance and relapse after treatment. These cells, termed cancer stem-like cells, have been designed as prominent targets for new and more efficient cancer therapies. G-protein coupled receptors (GPCRs, a family of membrane receptors, play a prominent role in cell signaling, cell communication and crosstalk with the microenvironment. Their role in cancer has been highlighted but remains largely unexplored. Here, we report a descriptive study of the differential expression of the endo-GPCR repertoire in human glioblastoma cancer stem-like cells (GSCs, U-87 MG cells, human astrocytes and fetal neural stem cells (f-NSCs. The endo-GPCR transcriptome has been studied using Taqman Low Density Arrays. Of the 356 GPCRs investigated, 138 were retained for comparative studies between the different cell types. At the transcriptomic level, eight GPCRs were specifically expressed/overexpressed in GSCs. Seventeen GPCRs appeared specifically expressed in cells with stem properties (GSCs and f-NSCs. Results of GPCR expression at the protein level using mass spectrometry and proteomic analysis are also presented. The comparative GPCR expression study presented here gives clues for new pathways specifically used by GSCs and unveils novel potential therapeutic targets.

  19. Enrichment of prostate cancer stem-like cells from human prostate cancer cell lines by culture in serum-free medium and chemoradiotherapy. (United States)

    Wang, Lei; Huang, Xing; Zheng, Xinmin; Wang, Xinghuan; Li, Shiwen; Zhang, Lin; Yang, Zhonghua; Xia, Zhiping


    The discovery of rare subpopulations of cancer stem cells (CSCs) has created a new focus in cancer research. As CSCs demonstrate resistance to chemoradiation therapy relative to other cancer cells, this allows the enrichment of CSC populations by killing apoptosis-susceptible cancer cells. In this study, three commonly used human prostate cancer (PCa) cell lines (DU145, PC-3 and LNCaP) were examined for their expression of the putative stem cell markers CD133 and CD44 via flow cytometric analysis. Under normal culture conditions, CD133(+)/CD44(+) cells were only present in the DU145 cell line, and comprised only a minor percentage (0.1% ± 0.01%) of the total population. However, the proportion of these CD133(+)/CD44(+) prostate CSCs could be increased in these cell lines via culture in serum-free medium (SFM), or through chemotherapy or radiotherapy. Indeed, after culture in SFM, the proportion of CD133(+)/CD44(+) cells in DU145 and PC-3 had increased to 10.3% and 3.0%, respectively. Moreover, the proportion had increased to 9.8% enriched by chemotherapy and 3.5% by radiotherapy in DU145. Colony-formation tests, cell invasion assays, and tumor xenografts in BALB/c nude mice were used to evaluate the stem cell properties of CD133(+)/CD44(+) PCa cells that were isolated via fluorescence-activated cell sorting (FACS). CD133(+)/CD44(+) cells had an enhanced colony-formation capability and invasive ability in vitro, and displayed greater tumorigenic properties in vivo. These results demonstrate the presence of CD133(+)/CD44(+) prostate CSCs in established PCa cell lines and that populations of these cells can be enriched by culture in SFM or chemoradiotherapy. Finding novel therapies to override chemoradiation resistance in the prostate CSCs is the key to improve long-term results in PCa management.

  20. Acquired resistance to metformin in breast cancer cells triggers transcriptome reprogramming toward a degradome-related metastatic stem-like profile (United States)

    Oliveras-Ferraros, Cristina; Vazquez-Martin, Alejandro; Cuyàs, Elisabet; Corominas-Faja, Bruna; Rodríguez-Gallego, Esther; Fernández-Arroyo, Salvador; Martin-Castillo, Begoña; Joven, Jorge; Menendez, Javier A


    Therapeutic interventions based on metabolic inhibitor-based therapies are expected to be less prone to acquired resistance. However, there has not been any study assessing the possibility that the targeting of the tumor cell metabolism may result in unforeseeable resistance. We recently established a pre-clinical model of estrogen-dependent MCF-7 breast cancer cells that were chronically adapted to grow (> 10 months) in the presence of graded, millimolar concentrations of the anti-diabetic biguanide metformin, an AMPK agonist/mTOR inhibitor that has been evaluated in multiple in vitro and in vivo cancer studies and is now being tested in clinical trials. To assess what impact the phenomenon of resistance might have on the metformin-like “dirty” drugs that are able to simultaneously hit several metabolic pathways, we employed the ingenuity pathway analysis (IPA) software to functionally interpret the data from Agilent whole-human genome arrays in the context of biological processes, networks, and pathways. Our findings establish, for the first time, that a “global” targeting of metabolic reprogramming using metformin certainly imposes a great selective pressure for the emergence of new breast cancer cellular states. Intriguingly, acquired resistance to metformin appears to trigger a transcriptome reprogramming toward a metastatic stem-like profile, as many genes encoding the components of the degradome (KLK11, CTSF, FREM1, BACE-2, CASP, TMPRSS4, MMP16, HTRA1), cancer cell migration and invasion factors (TP63, WISP2, GAS3, DKK1, BCAR3, PABPC1, MUC1, SPARCL1, SEMA3B, SEMA6A), stem cell markers (DCLK1, FAK), and key pro-metastatic lipases (MAGL and Cpla2) were included in the signature. Because this convergent activation of pathways underlying tumor microenvironment interactions occurred in low-proliferative cancer cells exhibiting a notable downregulation of the G2/M DNA damage checkpoint regulators that maintain genome stability (CCNB1, CCNB2, CDC20, CDC25C

  1. Disulfiram targets cancer stem-like cells and reverses resistance and cross-resistance in acquired paclitaxel-resistant triple-negative breast cancer cells (United States)

    Liu, P; Kumar, I S; Brown, S; Kannappan, V; Tawari, P E; Tang, J Z; Jiang, W; Armesilla, A L; Darling, J L; Wang, W


    Background: Triple-negative breast cancer (TNBC) has significantly worse prognosis. Acquired chemoresistance remains the major cause of therapeutic failure of TNBC. In clinic, the relapsed TNBC is commonly pan-resistant to various drugs with completely different resistant mechanisms. Investigation of the mechanisms and development of new drugs to target pan-chemoresistance will potentially improve the therapeutic outcomes of TNBC patients. Methods: In this study, 1-(4,5-Dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT), combination index (CI)–isobologram, western blot, ALDEFLUOR analysis, clonogenic assay and immunocytochemistry were used. Results: The chemoresistant MDA-MB-231PAC10 cells are highly cross-resistant to paclitaxel (PAC), cisplatin (CDDP), docetaxel and doxorubicin. The MDA-MB-231PAC10 cells are quiescent with significantly longer doubling time (64.9 vs 31.7 h). This may be caused by high expression of p21Waf1. The MDA-MB-231PAC10 cells express high aldehyde dehydrogenase (ALDH) activity and a panel of embryonic stem cell-related proteins, for example, Oct4, Sox2, Nanog and nuclealisation of HIF2α and NF-κBp65. We have previously reported that disulfiram (DS), an antialcoholism drug, targets cancer stem cells (CSCs) and enhances cytotoxicity of anticancer drugs. Disulfiram abolished CSC characters and completely reversed PAC and CDDP resistance in MDA-MB-231PAC10 cells. Conclusion: Cancer stem cells may be responsible for acquired pan-chemoresistance. As a drug used in clinic, DS may be repurposed as a CSC inhibitor to reverse the acquired pan-chemoresistance. PMID:24008666

  2. Sphere-forming-like cells (squamospheres) with cancer stem-like cell traits from VX2 rabbit buccal squamous cell carcinoma. (United States)

    Chen, Yuk-Kwan; Huang, Anderson Hsien-Cheng; Lin, Li-Min


    Previous studies have demonstrated that spheroid type cells grown under suspension culture conditions have cancer stem cell (CSC) traits in a number of cancers, but this phenomenon has not yet been reported in the VX2 rabbit oral cancer model. Hence, this study aimed to study the spheroid cells from VX2 rabbit buccal squamous cell carcinomas (SCCs) and assess their CSC characteristics. Five adult male New Zealand white outbred rabbits were used to generate VX2 rabbit buccal SCC. Sphere-forming cell culture was performed for the VX2 rabbit buccal SCC specimens. The self-renewal capability; cluster of designation (CD) 44, CD133, acetaldehyde dehydrogenase 1 (ALDH1), B cell-specific Moloney murine leukemia virus integration site 1 (Bmi-1), Nestin, octamer-binding transcription factor 4 (Oct4) and reduced expression protein-1 (Rex-1) expression with reverse transcription-polymerase chain reaction (RT-PCR); chemoresistance to cisplatin and 5-fluorouracil; and in vivo tumorigenicity of spheroid cell transplantation in nude mice were evaluated to determine the CSC characteristics of the resulting spheroid cells. We successfully obtained spheroid cells from the VX2 rabbit OSCC tissues. The spheroid cells exhibited CSC traits, including the expression of CSC and stem cell markers (CD44, Bmi-1, Nestin, Oct4 and Rex-1), capacity to generate new spheroid colonies within 1 week of reseeding from single-dissociated spheroid cells, chemoresistance capacity and generation of tumour xenografts (with histological features resembling those of the original VX2 rabbit buccal SCC) from the transplantation of 10(3) undifferentiated spheroid cells into nude mice. In summary, we demonstrated that spheroid cells with CSC cell traits can be derived from VX2 rabbit buccal SCCs, indicating that this animal cancer model is applicable for studying CSCs in human oral cancers.

  3. Sphere-forming-like cells (squamospheres) with cancer stem-like cell traits from VX2 rabbit buccal squamous cell carcinoma

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    Yuk-Kwan Chen; Anderson Hsien-Cheng Huang; Li-Min Lin


    Previous studies have demonstrated that spheroid type cells grown under suspension culture conditions have cancer stem cell (CSC) traits in a number of cancers, but this phenomenon has not yet been reported in the VX2 rabbit oral cancer model. Hence, this study aimed to study the spheroid cells from VX2 rabbit buccal squamous cell carcinomas (SCCs) and assess their CSC characteristics. Five adult male New Zealand white outbred rabbits were used to generate VX2 rabbit buccal SCC. Sphere-forming cell culture was performed for the VX2 rabbit buccal SCC specimens. The self-renewal capability;cluster of designation (CD) 44, CD133, acetaldehyde dehydrogenase 1 (ALDH1), B cell-specific Moloney murine leukemia virus integration site 1 (Bmi-1), Nestin, octamer-binding transcription factor 4 (Oct4) and reduced expression protein-1 (Rex-1) expression with reverse transcription-polymerase chain reaction (RT-PCR);chemoresistance to cisplatin and 5-fluorouracil;and in vivo tumorigenicity of spheroid cell transplantation in nude mice were evaluated to determine the CSC characteristics of the resulting spheroid cells. We successfully obtained spheroid cells from the VX2 rabbit OSCC tissues. The spheroid cells exhibited CSC traits, including the expression of CSC and stem cell markers (CD44, Bmi-1, Nestin, Oct4 and Rex-1), capacity to generate new spheroid colonies within 1 week of reseeding from single-dissociated spheroid cells, chemoresistance capacity and generation of tumour xenografts (with histological features resembling those of the original VX2 rabbit buccal SCC) from the transplantation of 103 undifferentiated spheroid cells into nude mice. In summary, we demonstrated that spheroid cells with CSC cell traits can be derived from VX2 rabbit buccal SCCs, indicating that this animal cancer model is applicable for studying CSCs in human oral cancers.

  4. Transforming growth factor-β1 regulates epithelial-mesenchymal transition in association with cancer stem-like cells in a breast cancer cell line. (United States)

    Jia, Yongfeng; Wu, Di; Yun, Fen; Shi, Lin; Luo, Nianrong; Liu, Zhiyue; Shi, Yonghong; Sun, Qinnuan; Jiang, Lili; Wang, Shiqi; Du, Maolin


    Epithelial-mesenchymal transition (EMT) is associated with altered connection and junctions between cells and changes in abilities of invasion and migration. In this study, we investigated whether SK-BR-3 breast cancer cells induced to undergo EMT exhibit changes in morphological and invasion abilities after Transforming growth factor β1 (TGF-β1) treatment. Serum-deprived SK-BR-3 cells were treated with TGF-β1 (0, 10 ng/mL) for 24 h. The cells morphological changes were observed and imaged using inverted phase contrast microscope. Scratch experiment and invasion experiment were employed to detect changes of invasion ability, cell-flow experiment was used to assess cell cycle, immunohistochemistry technique was used to detect epithelial and mesenchymal markers after the crawling cells were fixed. Our research reveal that SK-BR-3 cells become larger and more messy, the elongated cells extend pseudopodia, the link of the cells became more loosely and cell gap widened after TGF-β1 treatment. SK-BR-3 cells showed faster growing and improved invasion abilities after TGF-β1 treatment, and reduced G1 phase cells proportion in the total number of cells after the conversion, in contrast the S phase cells accounted for the proportion of the total number of cells increased. These findings indicate that TGF-β1-induced EMT in breast cancer cells may be associated with major alterations in morphological and invasion abilities.

  5. miR-92a-3p Exerts Various Effects in Glioma and Glioma Stem-Like Cells Specifically Targeting CDH1/β-Catenin and Notch-1/Akt Signaling Pathways

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    Hang Song


    Full Text Available MicroRNAs (miRNAs are implicated in the regulation of tumor progression and stemness of cancer stem-like cells. Recently, miR-92a-3p was reported to be up-regulated in human glioma samples. Nevertheless, the precise role of miR-92a-3p in glioma cells and glioma stem-like cells (GSCs has not been fully elucidated. It is necessary to clarify the function of miR-92a-3p in glioma and GSCs to develop novel therapeutic approaches for glioma patients. In the present study, we applied methyl-thiazolyl-tetrazolium (MTT assay and Transwell assay to measure the proliferation rate and metastatic potential of glioma cells. Meanwhile, the self-renewal ability of GSCs was detected by tumor sphere formation assay. The results revealed that down-regulation of miR-92a-3p suppressed the glioma cell malignancy in vitro. Moreover, knockdown of miR-92a-3p led to a reduction of tumorgenesis in vivo. Interestingly, we also observed that up-regulation of miR-92a-3p could inhibit the stemness of GSCs. Subsequent mechanistic investigation indicated that cadherin 1 (CDH1/β-catenin signaling and Notch-1/Akt signaling were the downstream pathways of miR-92a-3p in glioma cells and GSCs, respectively. These results suggest that miR-92a-3p plays different roles in glioma cells and GSCs through regulating different signaling pathways.

  6. A novel berbamine derivative inhibits cell viability and induces apoptosis in cancer stem-like cells of human glioblastoma, via up-regulation of miRNA-4284 and JNK/AP-1 signaling.

    Directory of Open Access Journals (Sweden)

    Fan Yang

    Full Text Available Glioblastoma (GBM is the most common primary brain tumor, accounting for approximately 40% of all central nervous system malignancies. Despite standard treatment consisting of surgical resection, radiotherapy and/or chemotherapy, the prognosis for GBM is poor; with a median survival of 14.6 months. The cancer stem cell or cancer-initiating cell model has provided a new paradigm for understanding development and recurrence of GBM following treatment. Berbamine (BBM is a natural compound derived from the Berberis amurensis plant, and along with its derivatives, has been shown to exhibit antitumor activity in several cancers. Here, we reported that a novel synthetic Berbamine derivative, BBMD3, inhibits cell viability and induces apoptosis of cancer stem-like cells (CSCs in a time- and dose-dependent manner when the CSCs from four GBM patients (PBT003, PBT008, PBT022, and PBT030 were cultured. These CSCs grew in neurospheres and expressed CD133 and nestin as markers. Treatment with BBMD3 destroyed the neurosphere morphology, and led to the induction of apoptosis in the CSCs. Induction of apoptosis in these CSCs is dependent upon activation of caspase-3 and cleavage of poly (ADP-ribose polymerase (PARP. MicroRNA-4284 (miR-4284 was shown to be over-expressed about 4-fold in the CSCs following BBMD3 treatment. Furthermore, transfection of synthetic anti-sense oligonucleotide against human miR-4284 partially blocked the anticancer effects of BBMD3 on the GBM derived CSCs. BBMD3 also increased phosphorylation of the c-Jun N-terminal kinase (JNK/stress-activated protein kinase (SAPK, resulting in an increase expression of phosphorylated c-Jun and total c-Fos; the major components of transcriptional factor AP-1. The JNK-c-Jun/AP-1 signaling pathway plays an important role in the induction of apoptosis in response to UV irradiation and some drug treatments. Targeting glioblastoma stem-like cells with BBMD3 is therefore novel, and may have promise as an

  7. Secreted frizzled-related protein 4 inhibits glioma stem-like cells by reversing epithelial to mesenchymal transition, inducing apoptosis and decreasing cancer stem cell properties.

    Directory of Open Access Journals (Sweden)

    G Bhuvanalakshmi

    Full Text Available The Wnt pathway is integrally involved in regulating self-renewal, proliferation, and maintenance of cancer stem cells (CSCs. We explored the effect of the Wnt antagonist, secreted frizzled-related protein 4 (sFRP4, in modulating epithelial to mesenchymal transition (EMT in CSCs from human glioblastoma cells lines, U87 and U373. sFRP4 chemo-sensitized CSC-enriched cells to the most commonly used anti-glioblastoma drug, temozolomide (TMZ, by the reversal of EMT. Cell movement, colony formation, and invasion in vitro were suppressed by sFRP4+TMZ treatment, which correlated with the switch of expression of markers from mesenchymal (Twist, Snail, N-cadherin to epithelial (E-cadherin. sFRP4 treatment elicited activation of the Wnt-Ca2(+ pathway, which antagonizes the Wnt/ß-catenin pathway. Significantly, the chemo-sensitization effect of sFRP4 was correlated with the reduction in the expression of drug resistance markers ABCG2, ABCC2, and ABCC4. The efficacy of sFRP4+TMZ treatment was demonstrated in vivo using nude mice, which showed minimum tumor engraftment using CSCs pretreated with sFRP4+TMZ. These studies indicate that sFRP4 treatment would help to improve response to commonly used chemotherapeutics in gliomas by modulating EMT via the Wnt/ß-catenin pathway. These findings could be exploited for designing better targeted strategies to improve chemo-response and eventually eliminate glioblastoma CSCs.

  8. New approaches of PARP-1 inhibitors in human lung cancer cells and cancer stem-like cells by some selected anthraquinone-derived small molecules.

    Directory of Open Access Journals (Sweden)

    Yu-Ru Lee

    Full Text Available Poly (ADP-ribose polymerase-1 (PARP-1 and telomerase, as well as DNA damage response pathways are targets for anticancer drug development, and specific inhibitors are currently under clinical investigation. The purpose of this work is to evaluate anticancer activities of anthraquinone-derived tricyclic and tetracyclic small molecules and their structure-activity relationships with PARP-1 inhibition in non-small cell lung cancer (NSCLC and NSCLC-overexpressing Oct4 and Nanog clone, which show high-expression of PARP-1 and more resistance to anticancer drug. We applied our library selected compounds to NCI's 60 human cancer cell-lines (NCI-60 in order to generate systematic profiling data. Based on our analysis, it is hypothesized that these drugs might be, directly and indirectly, target components to induce mitochondrial permeability transition and the release of pro-apoptotic factors as potential anti-NSCLC or PARP inhibitor candidates. Altogether, the most active NSC747854 showed its cytotoxicity and dose-dependent PARP inhibitory manner, thus it emerges as a promising structure for anti-cancer therapy with no significant negative influence on normal cells. Our studies present evidence that telomere maintenance should be taken into consideration in efforts not only to overcome drug resistance, but also to optimize the use of telomere-based therapeutics. These findings will be of great value to facilitate structure-based design of selective PARP inhibitors, in general, and telomerase inhibitors, in particular. Together, the data presented here expand our insight into the PARP inhibitors and support the resource-demanding lead optimization of structurally related small molecules for human cancer therapy.

  9. Analysis of cancer stem like cell characteristic in MKN-45 side population%MKN-45侧群细胞的肿瘤干细胞干性特征分析

    Institute of Scientific and Technical Information of China (English)


    Objective We tried to ascertain gastric cancer cell line MKN-45 contain SP cells, and learned about wether gastric cancer SP cells possess cancer stem-like characteristic. Method We used fluorescence activated cell sorting (FACS) to sort SP cells in human gastric carcinoma (GC) cell line MKN-45 cells labeled with Hoechst 33342, and then characterized the cancer stem-like properties of SP cells. Result The SP cells had higher clone formation efficiency than major population (MP) cells. Five stemness–related genes expression profiles, including OCT-4, SOX-2, NANOG, CD44 and ATP-binding cassette transporters gene ABCG-2, were tested in SP and MP cells using quantitative real-time RT-PCR. Western blot was chosen to show the difference of protein expression between SP and MP cells. Both results showed that there was significantly higher expression in SP cells than in MP cells. When inoculated into non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, SP cells showed higher tumorigenesis tendency than MP cells. Conclusion SP cells possess cancer stem cell properties and proved that SP cells from MKN-45 were gastric cancer stem-like cells.%  目的探讨胃癌细胞系中是否存在SP细胞及SP细胞的生物学特征。方法采用荧光活化的细胞分选技术(FACS)从胃癌MKN-45细胞株获得SP细胞,进而分析这些SP细胞有无肿瘤干细胞特性;采用实时定量的RT-PCR检测5个干性相关基因OCT-4, SOX-2, NANOG, CD44和ABCG-2的mRNA在SP和MP 细胞中的表达水平,并进行统计学差异分析。结果发现二者之间具有显著差异。进一步使用蛋白印迹法(Western blot)定性分析发现:5个干性相关基因在SP和MP细胞中的蛋白表达水平亦有差异;体内实验结果显示,当SP细胞和MP细胞注射非糖尿病联合重度免疫缺陷(NOD/SCID)的小鼠,SP 细胞较MP细胞具有更强的成瘤性。结论综合分析本实验相关结果,可以推测SP细胞具有

  10. Melanoma dormancy in a mouse model is linked to GILZ/FOXO3A-dependent quiescence of disseminated stem-like cells. (United States)

    Touil, Yasmine; Segard, Pascaline; Ostyn, Pauline; Begard, Severine; Aspord, Caroline; El Machhour, Raja; Masselot, Bernadette; Vandomme, Jerome; Flamenco, Pilar; Idziorek, Thierry; Figeac, Martin; Formstecher, Pierre; Quesnel, Bruno; Polakowska, Renata


    Metastatic cancer relapses following the reactivation of dormant, disseminated tumour cells; however, the cells and factors involved in this reactivation are just beginning to be identified. Using an immunotherapy-based syngeneic model of melanoma dormancy and GFP-labelled dormant cell-derived cell lines, we determined that vaccination against melanoma prevented tumour growth but did not prevent tumour cell dissemination or eliminate all tumour cells. The persistent disseminated melanoma tumour cells were quiescent and asymptomatic for one year. The quiescence/activation of these cells in vitro and the dormancy of melanoma in vivo appeared to be regulated by glucocorticoid-induced leucine zipper (GILZ)-mediated immunosuppression. GILZ expression was low in dormant cell-derived cultures, and re-expression of GILZ inactivated FOXO3A and its downstream target, p21CIP1. The ability of dormancy-competent cells to re-enter the cell cycle increased after a second round of cellular dormancy in vivo in association with shortened tumour dormancy period and faster and more aggressive melanoma relapse. Our data indicate that future cancer treatments should be adjusted according to the stage of disease progression.

  11. Evidence for embryonic stem-like signature and epithelial-mesenchymal transition features in the spheroid cells derived from lung adenocarcinoma. (United States)

    Roudi, Raheleh; Madjd, Zahra; Ebrahimi, Marzieh; Najafi, Ali; Korourian, Alireza; Shariftabrizi, Ahmad; Samadikuchaksaraei, Ali


    Identification of the cellular and molecular aspects of lung cancer stem cells (LCSCs) that are suggested to be the main culprit of tumor initiation, maintenance, drug resistance, and relapse is a prerequisite for targeted therapy of lung cancer. In the current study, LCSCs subpopulation of A549 cells was enriched, and after characterization of the spheroid cells, complementary DNA (cDNA) microarray analysis was applied to identify differentially expressed genes (DEGs) between the spheroid and parental cells. Microarray results were validated using quantitative real-time reverse transcription-PCR (qRT-PCR), flow cytometry, and western blotting. Our results showed that spheroid cells had higher clonogenic potential, up-regulation of stemness gene Sox2, loss of CD44 expression, and gain of CD24 expression compared to parental cells. Among a total of 160 genes that were differentially expressed between the spheroid cells and the parental cells, 104 genes were up-regulated and 56 genes were down-regulated. Analysis of cDNA microarray revealed an embryonic stem cell-like signature and over-expression of epithelial-mesenchymal transition (EMT)-associated genes in the spheroid cells. cDNA microarray results were validated at the gene expression level using qRT-PCR, and further validation was performed at the protein level by flow cytometry and western blotting. The embryonic stem cell-like signature in the spheroid cells supports two important notions: maintenance of CSCs phenotype by dedifferentiating mechanisms activated through oncogenic pathways and the origination of CSCs from embryonic stem cells (ESCs). PI3/AKT3, as the most common up-regulated pathway, and other pathways related to aggressive tumor behavior and EMT process can confer to the spheroid cells' high potential for metastasis and distant seeding.

  12. Comparison of the transcriptomic profile of hepatic human induced pluripotent stem like cells cultured in plates and in a 3D microscale dynamic environment. (United States)

    Leclerc, Eric; Kimura, Keiichi; Shinohara, Marie; Danoy, Mathieu; Le Gall, Morgane; Kido, Taketomo; Miyajima, Atsushi; Fujii, Teruo; Sakai, Yasuyuki


    We have compared the transcriptomic profiles of human induced pluripotent stem cells after their differentiation in hepatocytes like cells in plates and microfluidic biochips. The biochips provided a 3D and dynamic support during the cell differentiation when compared to the 2D static cultures in plates. The microarray have demonstrated the up regulation of important pathway related to liver development and maturation during the culture in biochips. Furthermore, the results of the transcriptomic profile, coupled with immunostaining, and RTqPCR analysis have shown typical biomarkers illustrating the presence of responders of biliary like cells, hepatocytes like cells, and endothelial like cells. However, the overall tissue still presented characteristic of immature and foetal patterns. Nevertheless, the biochip culture provided a specific micro-environment in which a complex multicellular differentiation toward liver could be oriented.

  13. Role of adenosine A2B receptor signaling in contribution of cardiac mesenchymal stem-like cells to myocardial scar formation. (United States)

    Ryzhov, Sergey; Sung, Bong Hwan; Zhang, Qinkun; Weaver, Alissa; Gumina, Richard J; Biaggioni, Italo; Feoktistov, Igor


    Adenosine levels increase in ischemic hearts and contribute to the modulation of that pathological environment. We previously showed that A2B adenosine receptors on mouse cardiac Sca1(+)CD31(-) mesenchymal stromal cells upregulate secretion of paracrine factors that may contribute to the improvement in cardiac recovery seen when these cells are transplanted in infarcted hearts. In this study, we tested the hypothesis that A2B receptor signaling regulates the transition of Sca1(+)CD31(-) cells, which occurs after myocardial injury, into a myofibroblast phenotype that promotes myocardial repair and remodeling. In vitro, TGFβ1 induced the expression of the myofibroblast marker α-smooth muscle actin (αSMA) and increased collagen I generation in Sca1(+)CD31(-) cells. Stimulation of A2B receptors attenuated TGFβ1-induced collagen I secretion but had no effect on αSMA expression. In vivo, myocardial infarction resulted in a rapid increase in the numbers of αSMA-positive cardiac stromal cells by day 5 followed by a gradual decline. Genetic deletion of A2B receptors had no effect on the initial accumulation of αSMA-expressing stromal cells but hastened their subsequent decline; the numbers of αSMA-positive cells including Sca1(+)CD31(-) cells remained significantly higher in wild type compared with A2B knockout hearts. Thus, our study revealed a significant contribution of cardiac Sca1(+)CD31(-) cells to the accumulation of αSMA-expressing cells after infarction and implicated A2B receptor signaling in regulation of myocardial repair and remodeling by delaying deactivation of these cells. It is plausible that this phenomenon may contribute to the beneficial effects of transplantation of these cells to the injured heart.

  14. Efficient enrichment of hepatic cancer stem-like cells from a primary rat HCC model via a density gradient centrifugation-centered method.

    Directory of Open Access Journals (Sweden)

    Wei-hui Liu

    Full Text Available BACKGROUND: Because few definitive markers are available for hepatic cancer stem cells (HCSCs, based on physical rather than immunochemical properties, we applied a novel method to enrich HCSCs. METHODOLOGY: After hepatic tumor cells (HTCs were first isolated from diethylinitrosamine-induced F344 rat HCC model using percoll discontinuous gradient centrifugation (PDGC and purified via differential trypsinization and differential attachment (DTDA, they were separated into four fractions using percoll continuous gradient centrifugation (PCGC and sequentially designated as fractions I-IV (FI-IV. Morphological characteristics, mRNA and protein levels of stem cell markers, proliferative abilities, induced differentiation, in vitro migratory capacities, in vitro chemo-resistant capacities, and in vivo malignant capacities were determined for the cells of each fraction. FINDINGS: As the density of cells increased, 22.18%, 11.62%, 4.73% and 61.47% of primary cultured HTCs were segregated in FI-FIV, respectively. The cells from FIII (density between 1.041 and 1.062 g/ml displayed a higher nuclear-cytoplasmic ratio and fewer organelles and expressed higher levels of stem cell markers (AFP, EpCAM and CD133 than cells from other fractions (P<0.01. Additionally, in vitro, the cells from FIII showed a greater capacity to self-renew, differentiate into mature HTCs, transit across membranes, close scratches, and carry resistance to chemotherapy than did cells from any other fraction; in vivo, injection of only 1×10(4 cells from FIII could generate tumors not only in subcutaneous tissue but also in the livers of nude mice. CONCLUSIONS: Through our novel method, HCSC-like cells were successfully enriched in FIII. This study will greatly contribute to two important areas of biological interest: CSC isolation and HCC therapy.

  15. In vitro and in vivo antiproliferative activity of metformin on stem-like cells isolated from spontaneous canine mammary carcinomas: translational implications for human tumors


    Barbieri, Federica; Thellung, Stefano; Ratto, Alessandra; Carra, Elisa; Marini, Valeria; Fucile, Carmen; Bajetto, Adriana; Pattarozzi, Alessandra; Würth, Roberto; Gatti, Monica; Campanella, Chiara; Vito, Guendalina; Mattioli, Francesca; Pagano, Aldo; Daga, Antonio


    Background Cancer stem cells (CSCs) are considered the cell subpopulation responsible for breast cancer (BC) initiation, growth, and relapse. CSCs are identified as self-renewing and tumor-initiating cells, conferring resistance to chemo- and radio-therapy to several neoplasias. Nowadays, th (about 10mM)e pharmacological targeting of CSCs is considered an ineludible therapeutic goal. The antidiabetic drug metformin was reported to suppress in vitro and in vivo CSC survival in different tumors...

  16. A novel spontaneous model of epithelial-mesenchymal transition (EMT) using a primary prostate cancer derived cell line demonstrating distinct stem-like characteristics. (United States)

    Harner-Foreman, Naomi; Vadakekolathu, Jayakumar; Laversin, Stéphanie A; Mathieu, Morgan G; Reeder, Stephen; Pockley, A Graham; Rees, Robert C; Boocock, David J


    Cells acquire the invasive and migratory properties necessary for the invasion-metastasis cascade and the establishment of aggressive, metastatic disease by reactivating a latent embryonic programme: epithelial-to-mesenchymal transition (EMT). Herein, we report the development of a new, spontaneous model of EMT which involves four phenotypically distinct clones derived from a primary tumour-derived human prostate cancer cell line (OPCT-1), and its use to explore relationships between EMT and the generation of cancer stem cells (CSCs) in prostate cancer. Expression of epithelial (E-cadherin) and mesenchymal markers (vimentin, fibronectin) revealed that two of the four clones were incapable of spontaneously activating EMT, whereas the others contained large populations of EMT-derived, vimentin-positive cells having spindle-like morphology. One of the two EMT-positive clones exhibited aggressive and stem cell-like characteristics, whereas the other was non-aggressive and showed no stem cell phenotype. One of the two EMT-negative clones exhibited aggressive stem cell-like properties, whereas the other was the least aggressive of all clones. These findings demonstrate the existence of distinct, aggressive CSC-like populations in prostate cancer, but, importantly, that not all cells having a potential for EMT exhibit stem cell-like properties. This unique model can be used to further interrogate the biology of EMT in prostate cancer.

  17. A functional study of EGFR and Notch signaling in brain cancer stem-like cells from glioblastoma multiforme (Ph.d.)

    DEFF Research Database (Denmark)

    Kristoffersen, Karina


    on their resemblance to normal neural stem cells (NSC) and their tumorigenic potential. Like for NSC, the epidermal growth factor receptor (EGFR) and Notch receptor signaling pathways are believed to be important for the maintenance of bCSC. These pathways as such present promising targets in a future anti-bCSC GBM...... treatment. The overall aim of the present PhD project has been to study the functional role of EGFR and Notch activity in bCSCs stem cell-like features and tumorigenic potential with the purpose of deepen our knowledge about the significance of these pathways in the bCSC population in GBM. By establishing...... and culturing human derived GBM xenograft cells under NSC conditions we obtained neurosphere cultures that contained cells with stem cell-like and tumorigenic properties. We moreover characterized the different cultures based on their expression level of the EGFR and Notch receptor as well as the expression...

  18. Nuclear reprogramming of luminal-like breast cancer cells generates Sox2-overexpressing cancer stem-like cellular states harboring transcriptional activation of the mTOR pathway (United States)

    Corominas-Faja, Bruna; Cufí, Sílvia; Oliveras-Ferraros, Cristina; Cuyàs, Elisabet; López-Bonet, Eugeni; Lupu, Ruth; Alarcón, Tomás; Vellon, Luciano; Iglesias, Juan Manuel; Leis, Olatz; Martín, Ángel G; Vazquez-Martin, Alejandro; Menendez, Javier A


    Energy metabolism plasticity enables stemness programs during the reprogramming of somatic cells to an induced pluripotent stem cell (iPSC) state. This relationship may introduce a new era in the understanding of Warburg’s theory on the metabolic origin of cancer at the level of cancer stem cells (CSCs). Here, we used Yamanaka’s stem cell technology in an attempt to create stable CSC research lines in which to dissect the transcriptional control of mTOR—the master switch of cellular catabolism and anabolism—in CSC-like states. The rare colonies with iPSC-like morphology, obtained following the viral transduction of the Oct4, Sox2, Klf4, and c-Myc (OSKM) stemness factors into MCF-7 luminal-like breast cancer cells (MCF-7/Rep), demonstrated an intermediate state between cancer cells and bona fide iPSCs. MCF-7/Rep cells notably overexpressed SOX2 and stage-specific embryonic antigen (SSEA)-4 proteins; however, other stemness-related markers (OCT4, NANOG, SSEA-1, TRA-1–60, and TRA-1–81) were found at low to moderate levels. The transcriptional analyses of OSKM factors confirmed the strong but unique reactivation of the endogenous Sox2 stemness gene accompanied by the silencing of the exogenous Sox2 transgene in MCF-7/Rep cells. Some but not all MCF-7/Rep cells acquired strong alkaline phosphatase (AP) activity compared with MCF-7 parental cells. SOX2-overexpressing MCF-7/Rep cells contained drastically higher percentages of CD44+ and ALDEFLUOR-stained ALDHbright cells than MCF-7 parental cells. The overlap between differentially expressed mTOR signaling-related genes in 3 different SOX2-overexpressing CSC-like cell lines revealed a notable downregulation of 3 genes, PRKAA1 (which codes for the catalytic α 1 subunit of AMPK), DDIT4/REDD1 (a stress response gene that operates as a negative regulator of mTOR), and DEPTOR (a naturally occurring endogenous inhibitor of mTOR activity). The insulin-receptor gene (INSR) was differentially upregulated in MCF-7/Rep


    Directory of Open Access Journals (Sweden)

    Roberto eWurth


    Full Text Available Chemokines are crucial autocrine and paracrine players in tumor development. In particular, CXCL12, through its receptors CXCR4 and CXCR7, affects tumor progression by controlling cancer cell survival, proliferation and migration, and, indirectly, via angiogenesis or recruiting immune cells.Glioblastoma (GBM is the most prevalent primary malignant brain tumor in adults and despite current multimodal therapies it remains almost incurable. The aggressive and recurrent phenotype of GBM is ascribed to high growth rate, invasiveness to normal brain, marked angiogenesis, ability to escape the immune system and resistance to standard of care therapies. Tumor molecular and cellular heterogeneity severely hinders GBM therapeutic improvement. In particular, a subpopulation of chemo- and radio-therapy resistant tumorigenic cancer stem–like cells (CSCs is believed to be the main responsible for tumor cell dissemination to the brain.GBM cells display heterogeneous expression levels of CXCR4 and CXCR7 that are overexpressed in CSCs, representing a molecular correlate for the invasive potential of GBM. The microenvironment contribution in GBM development is increasingly emphasized. An interplay exists between CSCs, differentiated GBM cells, and the microenvironment, mainly through secreted chemokines (e.g. CXCL12 causing recruitment of fibroblasts, endothelial, mesenchymal and inflammatory cells to the tumor, via specific receptors such as CXCR4.This review covers recent developments on the role of CXCL12/CXCR4-CXCR7 networks in GBM progression and the potential translational impact of their targeting. The biological and molecular understanding of the heterogeneous GBM cell behavior, phenotype and signaling is still limited. Progress in the identification of chemokine-dependent mechanisms that affect GBM cell survival, trafficking and chemo-attractive functions, opens new perspectives for development of more specific therapeutic approaches that include

  20. Oncogenic signaling pathways and origins of tumor-initiating stem-like cells of hepatocellular carcinomas induced by hepatitis C virus, alcohol and/or obesity. (United States)

    Chen, Chia-Lin; Tsukamoto, Hidekazu; Machida, Keigo


    This review article discusses the importance and oncogenic signaling pathways of tumor-initiating cells (TICs) in several etiologies of hepatocellular carcinomas (HCCs) induced by hepatitis C virus (HCV), alcohol, obesity and/or chemicals. Stem cells may be present in cancer tissue, and a hierarchy of cells is formed, as is the case for normal tissue. Tumor formation, growth and propagation are maintained by a small proportion of cells with stem cell-like properties. TICs are present in alcohol-fed HCV transgenic mice, diethylnitrosamine/phenobarbital-treated mice (chemical carcinogenesis) and Spnb2 +/- mice (defective TGF-β signal). Alcohol/obesity-associated endotoxemia induces the stem cell marker Nanog through TLR4 signaling to generate TICs and liver tumors in several HCC models. The oncogenic pathway (such as the STAT3 and TLR4-NANOG pathway) and mechanism of generation of TICs of HCCs associated with HCV, alcohol and obesity are discussed. Understanding the molecular stemness signaling and cellular hierarchy and defining key TIC-specific genes will accelerate the development of novel biomarkers and treatment strategies. This review highlights recent advances in understanding the pathogenesis of liver TICs and discusses unanswered questions about the concept of liver TICs. (This project was supported by NIH grants 1R01AA018857 and P50AA11999).

  1. Transient TNF regulates the self-renewing capacity of stem-like label-retaining cells in sphere and skin equivalent models of melanoma. (United States)

    Ostyn, Pauline; El Machhour, Raja; Begard, Severine; Kotecki, Nuria; Vandomme, Jerome; Flamenco, Pilar; Segard, Pascaline; Masselot, Bernadette; Formstecher, Pierre; Touil, Yasmine; Polakowska, Renata


    It is well established that inflammation promotes cancer, including melanoma, although the exact mechanisms involved are less known. In this study, we tested the hypothesis that inflammatory factors affect the cancer stem cell (CSC) compartment responsible for tumor development and relapse. Using an inducible histone 2B-GFP fusion protein as a tracer of cell divisional history, we determined that tumor necrosis factor (TNF), which is a classical pro-inflammatory cytokine, enlarged the CSC pool of GFP-positive label-retaining cells (LRCs) in tumor-like melanospheres. Although these cells acquired melanoma stem cell markers, including ABCB5 and CD271, and self-renewal ability, they lost their capacity to differentiate, as evidenced by the diminished MelanA expression in melanosphere cells and the loss of pigmentation in a skin equivalent model of human melanoma. The undifferentiated cell phenotype could be reversed by LY294002, which is an inhibitor of the PI3K/AKT signaling pathway, and this reversal was accompanied by a significant reduction in CSC phenotypic markers and functional properties. Importantly, the changes induced by a transient exposure to TNF were long-lasting and observed for many generations after TNF withdrawal. We conclude that pro-inflammatory TNF targets the quiescent/slow-cycling melanoma SC compartment and promotes PI3K/AKT-driven expansion of melanoma SCs most likely by preventing their asymmetrical self-renewal. This TNF effect is maintained and transferred to descendants of LRC CSCs and is manifested in the absence of TNF, suggesting that a transient exposure to inflammatory factors imprints long-lasting molecular and/or cellular changes with functional consequences long after inflammatory signal suppression. Clinically, these results may translate into an inflammation-triggered accumulation of quiescent/slow-cycling CSCs and a post-inflammatory onset of an aggressive tumor.

  2. Leptin stimulates migration and invasion and maintains cancer stem-like properties in ovarian cancer cells: an explanation for poor outcomes in obese women. (United States)

    Kato, Sumie; Abarzua-Catalan, Lorena; Trigo, César; Delpiano, Ana; Sanhueza, Cristobal; García, Karen; Ibañez, Carolina; Hormazábal, Katherine; Diaz, Daniela; Brañes, Jorge; Castellón, Enrique; Bravo, Erasmo; Owen, Gareth; Cuello, Mauricio A


    The evidence linking obesity with ovarian cancer remains controversial. Leptin is expressed at higher levels in obese women and stimulates cell migration in other epithelial cancers. Here, we explored the clinical impact of overweight/obesity on patient prognosis and leptin's effects on the metastatic potential of ovarian cancer cells. We assessed clinical outcomes in 70 ovarian cancer patients (33 healthy weight and 37 overweight) that were validated with an external cohort from The Cancer Genome Atlas (TCGA) database. Progression-free and overall survival rates were significantly decreased in overweight patients. Similarly, a worse overall survival rate was found in TCGA patients expressing higher leptin/OB-Rb levels. We explored serum and ascites leptin levels and OB-Rb expression in our cohort. Serum and ascites leptin levels were higher in overweight patients experiencing worse survival. OB-Rb was more highly expressed in ascites and metastases than in primary tumors. Leptin exposure increased cancer cell migration/invasion through leptin-mediated activation of JAK/STAT3, PI3/AKT and RhoA/ROCK and promoted new lamellipodial, stress-fiber and focal adhesion formation. Leptin also contributed to the maintenance of stemness and the mesenchymal phenotype in ovarian cancer cells. Our findings demonstrate that leptin stimulated ovarian cancer cell migration and invasion, offering a potential explanation for the poor prognosis among obese women.

  3. MicroRNA-7 inhibits the stemness of prostate cancer stem-like cells and tumorigenesis by repressing KLF4/PI3K/Akt/p21 pathway. (United States)

    Chang, Yun-Li; Zhou, Pei-Jie; Wei, Lianzi; Li, Wang; Ji, Zhongzhong; Fang, Yu-Xiang; Gao, Wei-Qiang


    Up to now, the molecular mechanisms underlying the stemness of prostate cancer stem cells (PCSCs) are still poorly understood. In this study, we demonstrated that microRNA-7 (miR-7) appears to be a novel tumor-suppressor miRNA, which abrogates the stemness of PCSCs and inhibits prostate tumorigenesis by suppressing a key stemness factor KLF4. MicroRNA-7 is down-regulated in prostate cancer cells compared to non-tumorigenic prostate epithelial cells. Restoration of miR-7 suppresses the expression of the stemness factor KLF4 in PCSCs and inhibits prostate tumorigenesis both in vitro and in vivo. Interestingly, the suppression of the stemness of PCSCs by miR-7 is sustained for generations in xenografts. Analysis of clinical samples also revealed a negative correlation between miR-7 expression and prostate tumor progression. Mechanistically, overexpression of miR-7 may lead to a cell cycle arrest but not apoptosis, which seems achieved via suppressing the KLF4/PI3K/Akt/p21 pathway. This study identifies miR-7 as a suppressor of PCSCs' stemness and implicates its potential application for PCa therapy.

  4. The isolation and cloning of sheep embryonic stem-like cells%绵羊类胚胎干细胞的分离与克隆

    Institute of Scientific and Technical Information of China (English)

    任卫青; 陈亮; 和小娥; 何俊丹; 王新庄


    从胚胎发育阶段、饲养层和培养体系等方面对影响绵羊类ES细胞分离、克隆效率的因素进行探讨。结果显示:致密桑葚胚和囊胚的ICM增殖率高于囊胚和孵化囊胚。绵羊类ES细胞在同源绵羊胎儿成纤维细胞(SEF)上生长比较缓慢,最终传代次数也低于小鼠胎儿成纤维细胞(MEF)组。培养液中同时添加胎牛血清(FBS)和Knock-out血清替代品(KSR),绵羊类ES传至7代,添加了碱性成纤维细胞生长因子(bFGF)后,最高可传至8代,而单纯添加KSR或FBS,分别传至4代和5代。对类ES细胞进行AKP染色、核型分析、体外分化试验,证实分离的类ES细胞符合ES细胞的主要特征,而且表达多潜能性细胞因子Nanog。由此认为,致密桑葚胚和囊胚更适合绵羊类ES细胞的体外分离和培养,而且MEF更适合于绵羊类ES细胞的分离传代,培养液中添加5%FBS和15%KSR,比较适合类ES细胞的分离传代,bFGF对绵羊类ES细胞的增殖具有促进作用。%Experiment were performed to determine the effect factor of solation,cloning and passage of embryonic stem cells(ES)in sheep,including developmental stage of embryos,feeder cells and growth factor.The result showed that The ICM growing rate of compacted morula and blastocysts were better than that of morula and hatching embryos.The growth rate and the capability of maintained undifferentiation of sheep ES-like cells which were cultured on MEF,are better than those on SEF.We applied basic medium with 15% KSR and 5% FBS as a new culture system to isolate sheep ES-like cells,the ES-like cell line maintained undifferentiating for7 passages.Added bFGF at the same time,it maintained undifferentiating for 8 passages,but when applied only 15% KSR or 15% FBS,it only could remain undifferentiated for 4 passages or 5 passages.Sheep ES-like cells isolated from ICMs had a normal karyotype and highly expressed alkaline phosphatase.Multiple differentiation potency

  5. Disulfiram targets cancer stem-like properties and the HER2/Akt signaling pathway in HER2-positive breast cancer. (United States)

    Kim, Ji Young; Cho, Youngkwan; Oh, Eunhye; Lee, Nahyun; An, Hyunsook; Sung, Daeil; Cho, Tae-Min; Seo, Jae Hong


    HER2-positive breast tumors are known to harbor cancer stem-like cell populations and are associated with an aggressive tumor phenotype and poor clinical outcomes. Disulfiram (DSF), an anti-alcoholism drug, is known to elicit cytotoxicity in many cancer cell types in the presence of copper (Cu). The objective of the present study was to investigate the mechanism of action responsible for the induction of apoptosis by DSF/Cu and its effect on cancer stem cell properties in HER2-positive breast cancers in vitro and in vivo. DSF/Cu treatment induced apoptosis, associated with a marked decrease in HER2, truncated p95HER2, phospho-HER2, HER3, phospho-HER3 and phospho-Akt levels, and p27 nuclear accumulation. This was accompanied by the eradication of cancer stem-like populations, concomitant with the suppression of aldehyde dehydrogenase 1 (ALDH1) activity and mammosphere formation. DSF administration resulted in a significant reduction in tumor growth and an enhancement of apoptosis, as well as HER2 intracellular domain (ICD) and ALDH1A1 downregulation. Our results demonstrate that DSF/Cu induces apoptosis and eliminates cancer stem-like cells via the suppression of HER2/Akt signaling, suggesting that DSF may be potentially effective for the treatment of HER2-positive cancers.

  6. Matrix metalloproteinase-10 regulates stemness of ovarian cancer stem-like cells by activation of canonical Wnt signaling and can be a target of chemotherapy-resistant ovarian cancer (United States)

    Mariya, Tasuku; Hirohashi, Yoshihiko; Torigoe, Toshihiko; Tabuchi, Yuta; Asano, Takuya; Saijo, Hiroshi; Kuroda, Takafumi; Yasuda, Kazuyo; Mizuuchi, Masahito; Saito, Tsuyoshi; Sato, Noriyuki


    Epithelial ovarian cancer (EOC) is one of the most lethal cancers in females. Cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) have been reported to be origin of primary and recurrent cancers and to be resistant to several treatments. In this study, we identified matrix metalloproteinase-10 (MMP10) is expressed in CSCs/CICs of EOC. An immunohistochemical study revealed that a high expression level of MMP10 is a marker for poor prognosis and platinum resistance in multivariate analysis. MMP10 gene overexpression experiments and MMP10 gene knockdown experiments using siRNAs revealed that MMP10 has a role in the maintenance of CSCs/CICs in EOC and resistance to platinum reagent. Furthermore, MMP10 activate canonical Wnt signaling by inhibiting noncanonical Wnt signaling ligand Wnt5a. Therefore, MMP10 is a novel marker for CSCs/CICs in EOC and that targeting MMP10 is a novel promising approach for chemotherapy-resistant CSCs/CICs in EOC. PMID:27072580

  7. Neuropilin-1 is expressed by breast cancer stem-like cells and is linked to NF-κB activation and tumor sphere formation. (United States)

    Glinka, Yelena; Mohammed, Nada; Subramaniam, Venkateswaran; Jothy, Serge; Prud'homme, Gérald J


    Cancer stem cells (CSCs) initiate tumors and have a high resistance to conventional cancer therapy. Tranilast is an orally active drug of low toxicity that exerts inhibitory effects on breast CSCs. This appears to depend on its aryl hydrocarbon receptor (AHR) agonistic activity, but this receptor has diverse functions and it is unclear how CSCs are inhibited. CSCs generate tumor spheres in low-adherence cultures, and we employed the mammosphere-forming assay as a functional test for breast CSCs. Because NF-κB has a key role in mammosphere formation and CSC-mediated tumor initiation, we examined that pathway. We also examined the role of neuropilin-1 (Nrp1), which is a growth factor coreceptor linked to the tumorigenicity of some CSCs. We found that tranilast concurrently suppressed mammosphere formation, Nrp1 expression and constitutive NF-κB activation. Flow cytometric analysis revealed that a subpopulation of breast cancer cells bearing breast CSC markers also expressed Nrp1. A blocking anti-Nrp1 antibody suppressed mammosphere formation. We examined whether there was a link between Nrp1 and NF-κB activation. The siRNA knockdown of Nrp1 severely suppressed NF-κB activation and mammosphere formation. The phosphorylation of Akt and ERK1/2 was also reduced, but to a lesser extent. We conclude that Nrp1 plays a key role in mammosphere formation and this activity is linked to NF-κB activation. Thus, Nrp1 might be a target for therapy against breast CSCs, and the anticancer drug tranilast suppresses its expression. Copyright © 2012 Elsevier Inc. All rights reserved.

  8. The Immuno-therapy with Dendritic Cells Sensitized with mRNA from Breast tumor Stem-like Cells for Breast Carcinoma%乳腺癌干细胞样细胞mRNA致敏的树突状细胞疫苗免疫治疗研究

    Institute of Scientific and Technical Information of China (English)

    郑秋红; 谢云青; 刘健; 应敏刚


    Objective To investigate the induced immuno-reaction in vitro of dendritic cell vaccine sensitized with nucleic-acid antigen from breast tumor stem-like cells. Method The breast tumor MCF-7 stem-like cells were cultured and enriched in serum-free medium containing defined growth factors, and were confirmed by the identification of cellular surface markers by flow cytometer analysis, by clone-forming capability test with a soft agar assay, and by tumorigenic ability test in vivo using an NOD-SCID mouse model. The mRNAs were amplified by using T7 mMessage mMACHINE kit with total RNA template extracted from MCF-7 stem-like cells. The peripheral blood dendritic cells were transfected with the mRNA and their activities of inducing CTL were measured by MTT method. Result The breast tumor stem-like cells with CD44/CD24 phenotype could be cultured and enriched (90. 17%) in the defined serum-free medium and demonstrate strong tumorigenicity after being challenged into NOD/SCID mouse. The dendritic cells, when loaded with nucleic-acid antigen, could express specific cellular surface markers of CD80/CD83/CD86 and HLA-DR at high level. The dendritic cells, which were transfected with mRNA from the suspension, low-differentiation breast tumor stem-like cells, in comparison with the adhesive, differentiated breast tumor stem-like cells, possessed stronger capability to induce TCL mediated killing of target cells (P<0. 01). Conclusion The dendritic cells, which were sensitized with mRNA derived from breast tumor stem-like cells, could induce strong CTL targeting breast tumor stem-like cells in vitro. It may provide a new choice of clinical immuno-therapy for breast carcinoma.%目的 观察乳腺癌干细胞样细胞核酸抗原致敏的树突状细胞疫苗体外诱导免疫反应作用.方法 利用含有生长因子的无血清培养基悬浮培养法富集MCF-7乳腺癌干细胞样细胞,经单克隆形成、表面标记检测、NOD-SCID小鼠成瘤等实验鉴定后,采用T7

  9. Osteopontin Overexpression Induced Tumor Progression and Chemoresistance to Oxaliplatin through Induction of Stem-Like Properties in Human Colorectal Cancer

    Directory of Open Access Journals (Sweden)

    Lui Ng


    Full Text Available Colorectal cancer (CRC is one of the most common and fatal malignancies worldwide. The poor prognosis of colorectal cancer patients is due to development of chemoresistance and cancer metastasis. Recently osteopontin (OPN has been associated with stem-like properties in colorectal cancer. This study further examined the clinicopathological significance of OPN in CRC and its effect on chemoresistance and transcription of stem cell markers. We examined the transcription level of OPN in 84 CRC patients and correlated the expression with their clinicopathological parameters. The associations of OPN overexpression with transcription of stem cell markers and response to chemotherapy in DLD1-OPN overexpressing clones and CRC patients were also investigated. Our results showed that OPN was significantly overexpressed in CRC, and its overexpression correlated with tumor stage and poor prognosis. Overexpression of CRC induced OCT4 and SOX2 expression in vitro and correlated with SOX2 overexpression in CRC patients. In addition, DLD1-OPN overexpressing cells showed enhanced ability to survive upon oxaliplatin treatment, and OPN expression was higher in CRC patients who were resistant to oxaliplatin-involved chemotherapy treatment. Thus, CRC cells overexpressing OPN demonstrated stem-like properties and OPN inhibition is a potential therapeutic approach to combat CRC progression and chemoresistance.

  10. Activation of c-MET induces a stem-like phenotype in human prostate cancer.

    Directory of Open Access Journals (Sweden)

    Geert J L H van Leenders

    Full Text Available Prostate cancer consists of secretory cells and a population of immature cells. The function of immature cells and their mutual relation with secretory cells are still poorly understood. Immature cells either have a hierarchical relation to secretory cells (stem cell model or represent an inducible population emerging upon appropriate stimulation of differentiated cells. Hepatocyte Growth Factor (HGF receptor c-MET is specifically expressed in immature prostate cells. Our objective is to determine the role of immature cells in prostate cancer by analysis of the HGF/c-MET pathway.Gene-expression profiling of DU145 prostate cancer cells stimulated with HGF revealed induction of a molecular signature associated with stem cells, characterized by up-regulation of CD49b, CD49f, CD44 and SOX9, and down-regulation of CD24 ('stem-like signature'. We confirmed the acquisition of a stem-like phenotype by quantitative PCR, FACS analysis and Western blotting. Further, HGF led to activation of the stem cell related Notch pathway by up-regulation of its ligands Jagged-1 and Delta-like 4. Small molecules SU11274 and PHA665752 targeting c-MET activity were both able to block the molecular and biologic effects of HGF. Knock-down of c-MET by shRNA infection resulted in significant reduction and delay of orthotopic tumour-formation in male NMRI mice. Immunohistochemical analysis in prostatectomies revealed significant enrichment of c-MET positive cells at the invasive front, and demonstrated co-expression of c-MET with stem-like markers CD49b and CD49f.In conclusion, activation of c-MET in prostate cancer cells induced a stem-like phenotype, indicating a dynamic relation between differentiated and stem-like cells in this malignancy. Its mediation of efficient tumour-formation in vivo and predominant receptor expression at the invasive front implicate that c-MET regulates tumour infiltration in surrounding tissues putatively by acquisition of a stem-like phenotype.


    Institute of Scientific and Technical Information of China (English)

    华进联; 刘雨潇; 董武子


    @@ Stem cells are classified into embryonic stem (ES) cells and adult stem cells, which are the progenitors of any cell types of the body or tissues, and have the ability to self-renew and produce one or more differentiated cell types.

  12. Repression of mammosphere formation of human breast cancer cells by soy isoflavone genistein and blueberry polyphenolic acids suggests diet-mediated targeting of cancer stem-like/progenitor cells (United States)

    Mammary stem cells are undifferentiated epithelial cells which initiate mammary tumors and render them resistant to anticancer therapies, when deregulated. Diets rich in fruits and vegetables are implicated in breast cancer risk reduction, yet underlying mechanisms are poorly understood. Here, we ad...

  13. Integrative analyses of gene expression and DNA methylation profiles in breast cancer cell line models of tamoxifen-resistance indicate a potential role of cells with stem-like properties

    DEFF Research Database (Denmark)

    Lin, Xue; Li, Jian; Yin, Guangliang


    Development of resistance to tamoxifen is an important clinical issue in the treatment of breast cancer. Tamoxifen resistance may be the result of acquisition of epigenetic regulation within breast cancer cells, such as DNA methylation, resulting in changed mRNA expression of genes pivotal...

  14. Signaling Crosstalk in the Regulation of Epithelial-Mesenchymal Transition and Cancer Stem-Like Cells Properties Acquisition%调控肿瘤上皮-间质转化及肿瘤干细胞样特性的信号通路串话

    Institute of Scientific and Technical Information of China (English)

    张婷; 崔戈; 邵圣文; 叶家辉; 黄斌; 潘臻


    Epithelial-mesenchymal transition (EMT) is an important biological process that epithelial tumor cells can obtain the ability of invasion and metastasis. Cancer stem-like cells (CSLCs)/tumor-initiatmg cells (TICs) play a key role in tumorigenesis, tumor invasion, metastasis and recurrence. In recent years, it was found that EMT had a close correlation with the acquisition of CSLCs properties, they promote tumorigenesis, tumor invasion and metastasis by complicated interaction through signaling crosstalk between TGF-β, Wnt/p-catenin, Notch, Hedgehog, FGF, PI3k/Akt and other signaling pathways. Understanding the functions and interactions of key molecules within the context of EMT/CSLCs signaling is critical to design targeted therapeutics.%上皮-间质转化(epithelial-mesenchymal transition,EMT)是上皮来源肿瘤细胞获得侵袭和转移能力的重要生物学过程.肿瘤干细胞样细胞(cancer stem-like cells,CSLCs)在肿瘤发生、侵袭、转移和复发中亦起着关键作用.近年发现,EMT与肿瘤干细胞样特性获得存在密切关联,二者通过TGF-β、Wnt/β-catenin、Notch、Hedgehog、FGF、PI3k/Akt等多种信号通路及通路间的信号串话而交互作用,共同影响着肿瘤发生、侵袭及转移,了解调控EMT/CSLCs关键信号分子的功能及相互作用对于肿瘤靶向治疗具有重要意义.

  15. Isolation and cultivation of keratinocyte stem-like cells from human skin dermis%人皮肤真皮组织中角朊干细胞样细胞的分离与培养*☆

    Institute of Scientific and Technical Information of China (English)

    池光范; 李美英; 赵贵芳; 邓继鸿; 刘晋宇; 李玉林


      背景:皮肤真皮组织是皮肤损伤修复时除表皮基底层以外可形成表皮层结构的重要干细胞来源。目的:探索从人皮肤真皮组织中分离培养角朊干细胞样细胞的新方法。方法:将一定大小的成人腹部全层皮肤组织用 Dispase 过夜消化后去除表皮层结构,然后用0.1%Ⅰ型胶原酶过夜消化,离心分离以获取真皮来源总细胞。将分离的细胞用含有体积分数1% N2,2% B27,20μg/L表皮生长因子和40μg/L 碱性成纤维细胞生长因子的DMEM/F12(1∶1)无血清培养液悬浮后以低密度接种于培养皿进行培养。采用RT-PCR和免疫细胞荧光染色法对其进行分析。结果与结论:RT-PCR分析结果显示所分离的真皮细胞中含有表达Lgr5和Lirg1等特异基因的毛囊来源干细胞;免疫细胞荧光分析结果显示其表达细胞角蛋白8和细胞角蛋白14。在无血清条件下培养6 d时,可观察到表达细胞角蛋白14和整合素α6的“铺路石”样细胞克隆。进行传代培养后P1细胞表达角朊干细胞标志物CD29、细胞角蛋白14和P63。传代培养至P3,4时大部分细胞分化,不能继续传代。结果证实,以无血清培养液结合低密度培养法可直接从人真皮组织中有效地培养增殖活跃、细胞角蛋白14和P63阳性的角朊干细胞样细胞。%  BACKGROUND: Skin dermis is an essential stem cel source, except basal layer of epidermis, for forming epidermis structure during the process of skin wound healing. OBJECTIVE: To explore a new method for isolating and culturing keratinocyte-stem like cel s from human skin dermis. METHODS: Definitive sizes of ful skins from adult abdomen were digested with dispase overnight and the epidermal structure was discarded. The remaining dermis were digested with 0.1% type I col agenase overnight. Thereafter, the digested cel s were col ected by centrifugation, suspended in serum-free Dulbecco’s modified Eagle

  16. Bruton's tyrosine kinase (Btk) inhibitor ibrutinib suppresses stem-like traits in ovarian cancer. (United States)

    Zucha, Muhammad Ary; Wu, Alexander T H; Lee, Wei-Hwa; Wang, Liang-Shun; Lin, Wan-Wan; Yuan, Chiou-Chung; Yeh, Chi-Tai


    According to a Prognoscan database, upregulation of Bruton's tyrosine kinase (Btk) is associated with low overall survival in ovarian cancer patients. We found that spheroids-forming ovarian cancer cell, which highly expressed cancer stem-like cell (CSC) markers and Btk, were cisplatin resistant. We next treated CSCs and non-CSCs by a combination of ibrutinib and cisplatin. We found that chemoresistance was dependent on Btk and JAK2/STAT3, which maintained CSC by inducing Sox-2 and prosurvival genes. We suggest that addition of ibrutinib to cisplatin may improve treatment outcome in ovarian cancer.

  17. Paclitaxel-Fe3O4 nanoparticles inhibit growth of CD138–  CD34– tumor stem-like cells in multiple myeloma-bearing mice

    Directory of Open Access Journals (Sweden)

    Yang C


    Full Text Available Cuiping Yang,1,3,* Jing Wang,2,* Dengyu Chen,1,* Junsong Chen,1 Fei Xiong,4 Hongyi Zhang,1 Yunxia Zhang,2 Ning Gu,4 Jun Dou11Department of Pathogenic Biology and Immunology, Medical School, 2Department of Gynecology and Obstetrics, Zhongda Hospital, Southeast University, Nanjing, 3Department of Pathogenic Biology and Immunology, School of Basic Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, 4School of Biological Science and Medical Engineering, Southeast University, Nanjing, People’s Republic of China*These authors contributed equally to this workBackground: There is growing evidence that CD138– CD34– cells may actually be tumor stem cells responsible for initiation and relapse of multiple myeloma. However, effective drugs targeted at CD138– CD34– tumor stem cells are yet to be developed. The purpose of this study was to investigate the inhibitory effect of paclitaxel-loaded Fe3O4 nanoparticles (PTX-NPs on CD138– CD34– tumor stem cells in multiple myeloma-bearing mice.Methods: CD138– CD34– cells were isolated from a human U266 multiple myeloma cell line using an immune magnetic bead sorting method and then subcutaneously injected into mice with nonobese diabetic/severe combined immunodeficiency to develop a multiple myeloma-bearing mouse model. The mice were treated with Fe3O4 nanoparticles 2 mg/kg, paclitaxel 4.8 mg/kg, and PTX-NPs 0.64 mg/kg for 2 weeks. Tumor growth, pathological changes, serum and urinary interleukin-6 levels, and molecular expression of caspase-3, caspase-8, and caspase-9 were evaluated.Results: CD138– CD34– cells were found to have tumor stem cell characteristics. All the mice developed tumors in 40 days after injection of 1 × 106 CD138– CD34– tumor stem cells. Tumor growth in mice treated with PTX-NPs was significantly inhibited compared with the controls (P <  0.005, and the groups that received nanoparticles alone (P < 0.005 or paclitaxel alone (P < 0.05. In addition

  18. Isolation and identiffcation of cancer stem-like side population cells in pancreatic cancer%胰腺癌肿瘤干细胞样侧群细胞的分离鉴定

    Institute of Scientific and Technical Information of China (English)

    周静; 王春友; 周峰; 刘涛; 吴河水; 熊炯圻; 赵刚; 陶京; 杨明; 周蒙滔


    目的 分离鉴定胰腺癌中的侧群(SP)细胞亚群.方法 应用Hoechst33342染色,流式细胞仪检测5个胰腺癌细胞系及3个原代培养的临床胰腺癌标本中sP细胞的含量.以PANC-1为例,通过平板克隆形成试验和NOD-SCID小鼠异种移植成瘤实验比较SP细胞与non-SP细胞的克隆形成能力及成瘤能力,通过对体外培养的SP细胞和SP细胞衍生肿瘤的HoechsG3342复染SP再分析判断其是否具有分化潜能.结果 除了BXPC-3,其他胰腺癌细胞系及原代培养标本都存在Verapamil敏感的SP细胞.SP细胞与non-SP细胞比较具有较高的克隆形成能力[(43.67±3.10)%比(8.33±1.63)%,P<0.01],并且能够分化产生non-SP细胞并维持自身SP细胞的比例在一个较稳定的水平.SP细胞的成瘤能力是non-SP细胞的100倍以上.而且SP细胞在体内亦可发生不对称分裂生成SP细胞和non-SP细胞.结论 SP细胞可能是胰腺癌干细胞的候选细胞之一.%Objective To isolate and identify side population(SP)cells in pancreatic cancer.Methods We detected the number of SP cells in 5 pancreatic cancer cell lines and 3 clinical samples by Hoechst33342 dyeing and FACS analysis.The clone formation efficiency and tumorigenicity were compared between SP cels and non-SP ceils by clone formation assay and NOD/SCID xenograft transplantation experiment.The differentiation ability of SP ceils was studied by flow cytometry reanalysis of SP-derived tumors and cultured SP ceils.Results All cell lines and clinical samples were found to exhibit verapamilsensitive SP cells except BXPC-3.Using SP cells sorted from PANC-1 as a model.SP ceUs were demonstratcd to exhibit higher colony-formation ability than non-SP cells [(43.67±3.10)% vs (8.33±1.63)%.P<0.01].And flow cytometry reanalysis indicated SP ceHs could generate both SP and non-SP cells with a fraction size comparable with the original population.In addition.SP cells were enriched in tumorigenicity by at least 100-fold

  19. Impact of Microenvironment and Stem-Like Plasticity in Cholangiocarcinoma

    DEFF Research Database (Denmark)

    Raggi, Chiara; Invernizzi, Pietro; Andersen, Jesper Bøje


    Clinical complexity, anatomic diversity and molecular heterogeneity of cholangiocarcinoma (CCA) represent a major challenge in the assessment of effective targeted therapies. Molecular and cellular mechanisms underlying diversity of CCA growth patterns remain a key issue and a clinical concern...... or tumor microenvironment (TME) likely promotes initiation and progression of this malignancy contributing to its heterogeneity. This review will emphasize the dynamic interplay between stem-like intrinsic and TME-extrinsic pathways, which may represent novel options for multi-targeted therapies in CCA....

  20. COMP-angiopoietin 1 increases proliferation, differentiation, and migration of stem-like cells through Tie-2-mediated activation of p38 MAPK and PI3K/Akt signal transduction pathways

    Energy Technology Data Exchange (ETDEWEB)

    Kook, Sung-Ho [Cluster for Craniofacial Development and Regeneration Research, Institute of Oral Biosciences and School of Dentistry, Chonbuk National University, Jeonju (Korea, Republic of); Lim, Shin-Saeng [School of Dentistry and Dental Research Institute, Seoul National University, Seoul (Korea, Republic of); Cho, Eui-Sic; Lee, Young-Hoon; Han, Seong-Kyu; Lee, Kyung-Yeol [Cluster for Craniofacial Development and Regeneration Research, Institute of Oral Biosciences and School of Dentistry, Chonbuk National University, Jeonju (Korea, Republic of); Kwon, Jungkee [College of Veterinary Medicine, Chonbuk National University, Jeonju (Korea, Republic of); Hwang, Jae-Won; Bae, Cheol-Hyeon [Cluster for Craniofacial Development and Regeneration Research, Institute of Oral Biosciences and School of Dentistry, Chonbuk National University, Jeonju (Korea, Republic of); Seo, Young-Kwon [Research Institute of Biotechnology, Dongguk University, Seoul (Korea, Republic of); Lee, Jeong-Chae, E-mail: [Cluster for Craniofacial Development and Regeneration Research, Institute of Oral Biosciences and School of Dentistry, Chonbuk National University, Jeonju (Korea, Republic of)


    Highlights: • COMP-Ang1 induces Tie-2 activation in BMMSCs, but not in primary osteoblasts. • Tie-2 knockdown inhibits COMP-Ang1-stimulated proliferation and osteoblastogenesis. • Tie-2 knockdown prevents COMP-Ang1-induced activation of PI3K/Akt and p38 MAPK. • COMP-Ang1 induces migration of cells via activation of PI3K/Akt and CXCR4 pathways. • COMP-Ang1 stimulates in vivo migration of PDLSCs into a calvarial defect site of rats. - Abstract: Recombinant COMP-Ang1, a chimera of angiopoietin-1 (Ang1) and a short coiled-coil domain of cartilage oligomeric matrix protein (COMP), is under consideration as a therapeutic agent capable of inducing the homing of cells with increased angiogenesis. However, the potentials of COMP-Ang1 to stimulate migration of mesenchymal stem cells (MSCs) and the associated mechanisms are not completely understood. We examined the potential of COMP-Ang1 on bone marrow (BM)-MSCs, human periodontal ligament stem cells (PDLSCs), and calvarial osteoblasts. COMP-Ang1 augmented Tie-2 induction at protein and mRNA levels and increased proliferation and expression of runt-related transcription factor 2 (Runx2), osterix, and CXCR4 in BMMSCs, but not in osteoblasts. The COMP-Ang1-mediated increases were inhibited by Tie-2 knockdown and by treating inhibitors of phosphoinositide 3-kinase (PI3K), LY294002, or p38 mitogen-activated protein kinase (MAPK), SB203580. Phosphorylation of p38 MAPK and Akt was prevented by siRNA-mediated silencing of Tie-2. COMP-Ang1 also induced in vitro migration of BMMSCs and PDLSCs. The induced migration was suppressed by Tie-2 knockdown and by CXCR4-specific peptide antagonist or LY294002, but not by SB203580. Furthermore, COMP-Ang1 stimulated the migration of PDLSCs into calvarial defect site of rats. Collectively, our results demonstrate that COMP-Ang1-stimulated proliferation, differentiation, and migration of progenitor cells may involve the Tie-2-mediated activation of p38 MAPK and PI3K/Akt pathways.

  1. Cell Growth Enhancement (United States)


    Exogene Corporation uses advanced technologies to enhance production of bio-processed substances like proteins, antibiotics and amino acids. Among them are genetic modification and a genetic switch. They originated in research for Jet Propulsion Laboratory. Extensive experiments in cell growth through production of hemoglobin to improve oxygen supply to cells were performed. By improving efficiency of oxygen use by cells, major operational expenses can be reduced. Greater product yields result in decreased raw material costs and more efficient use of equipment. A broad range of applications is cited.

  2. Disulfiram inhibits TGF-β-induced epithelial-mesenchymal transition and stem-like features in breast cancer via ERK/NF-κB/Snail pathway. (United States)

    Han, Dan; Wu, Gang; Chang, Chan; Zhu, Fang; Xiao, Yin; Li, Qiuhui; Zhang, Tao; Zhang, Liling


    Disulfiram (DSF), an anti-alcoholism drug, has been reported as an inhibitor of NF-κB. NF-κB is involved in epithelial-mesenchymal transition (EMT) and self-renewal of breast cancer stem cells (CSCs). In this study, we treated MCF-7 and MDA-MB-231 breast cancer cells with TGF-β to induce EMT and cancer stem-like features and studied whether DSF can reverse this process. We found that DSF inhibited TGF-β induced EMT in breast cancer cells in a dose-dependent manner. Also, DSF inhibited EMT-associated stem-like features, migration and invasion of tumor cells as well as tumor growth in xenograft model. The activation of NF-κB was linked with EMT and stem-like cells. We conclude that DSF can suppress NF-κB activity and downregulate ERK/NF-κB/Snail pathway, leading to reverse EMT and stem-like features. Our data suggest that DSF inhibits EMT and stem-like properties in breast cancer cells associated with inhibition of the ERK/NF-κB/Snail pathway.

  3. Exploring the clonal evolution of CD133/aldehyde-dehydrogenase-1 (ALDH1)-positive cancer stem-like cells from primary to recurrent high-grade serous ovarian cancer (HGSOC). A study of the Ovarian Cancer Therapy-Innovative Models Prolong Survival (OCTIPS) Consortium. (United States)

    Ruscito, Ilary; Cacsire Castillo-Tong, Dan; Vergote, Ignace; Ignat, Iulia; Stanske, Mandy; Vanderstichele, Adriaan; Ganapathi, Ram N; Glajzer, Jacek; Kulbe, Hagen; Trillsch, Fabian; Mustea, Alexander; Kreuzinger, Caroline; Benedetti Panici, Pierluigi; Gourley, Charlie; Gabra, Hani; Kessler, Mirjana; Sehouli, Jalid; Darb-Esfahani, Silvia; Braicu, Elena Ioana


    High-grade serous ovarian cancer (HGSOC) causes 80% of all ovarian cancer (OC) deaths. In this setting, the role of cancer stem-like cells (CSCs) is still unclear. In particular, the evolution of CSC biomarkers from primary (pOC) to recurrent (rOC) HGSOCs is unknown. Aim of this study was to investigate changes in CD133 and aldehyde dehydrogenase-1 (ALDH1) CSC biomarker expression in pOC and rOC HGSOCs. Two-hundred and twenty-four pOC and rOC intrapatient paired tissue samples derived from 112 HGSOC patients were evaluated for CD133 and ALDH1 expression using immunohistochemistry (IHC); pOCs and rOCs were compared for CD133 and/or ALDH1 levels. Expression profiles were also correlated with patients' clinicopathological and survival data. Some 49.1% of the patient population (55/112) and 37.5% (42/112) pOCs were CD133+ and ALDH1+ respectively. CD133+ and ALDH1+ samples were detected in 33.9% (38/112) and 36.6% (41/112) rOCs. CD133/ALDH1 coexpression was observed in 23.2% (26/112) and 15.2% (17/112) of pOCs and rOCs respectively. Pairwise analysis showed a significant shift of CD133 staining from higher (pOCs) to lower expression levels (rOCs) (p cancer cells, providing also a first evidence that there is no correlation between CSCs and BRCA status. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. 人肝癌PLC/PRF-5细胞中干细胞样细胞的分离及其特异性miRNAs的筛选%Isolation of stem-like cells from human hepatocellular carcinoma PLC/PRF-5 cells and screening for their specific miRNAs

    Institute of Scientific and Technical Information of China (English)

    郑秋红; 许扬梅; 魏植强; 龚福生; 杨建伟; 谢云青; 应敏刚


    Objective: To isolate stem-like cells from hepatocellular carcinoma cell line PLC/PRF-5 and to study their miRNA profile. Methods:ABCG2 + and ABCG2- PLC/PRF-5 cells were isolated from the PLC/PRF-5 cell line by magnetic activated cell sorting (MACS) method, and further identified by flow cytometry. The colony formation ability in soft agar and tumor formation ability in NOD/SCID mice of ABCG2+ and ABCG2- PLC/PRF-5 cells were observed. miRNA chip was adopted to screen the differentially expressed miRNAs between ABCG2 + and ABCG2 - PLC/PRF-5 cells; and real-time PCR assay was used to confirm the results of miRNA chip. Results:The purity of ABCG2 + PLC/PRF-5 cells isolated by MACS method was ( 84.20 ± 4.52 ) %. The colony number and size formed by ABCG2 + PLC/PRF-5 cells were more and larger than those formed by ABCG2 - cells (47.17 ± 10.50 vs 23.33 ±7.31, P <0.05). 1 × 104 ABCG2 + cells could form tumors with at least 5 × 105 cells needed for ABCG2 - cells. The size of tumors generated by 5 × 105 ABCG2 + cells was larger than that by the ABCG2 - cells ( [ 3.73 ± 1.19 ]cm3 vs [0.72 ± 0.57 ] cm3, P < O. Ol ). There are 20 miRNAs differentially expressed between ABCG2 + and ABCG2- cells, with 13 up-regulated and 7 down-regulated. Realtime PCR assay was applied to further verify the differential expression of hsa-miR-30a and hsa-miR-630, and the results were in accordance with those of miRNA chip. Conclusion: ABCG2+ PLC/PRF-5 cells in hepatocellular carcinoma cell line PLC/PRF-5 have the properties of cancer stem cells. Twenty miRNAs are differentially expressed between ABCG2 + and ABCG2 - PLC/PRF-5 cells, which might play important roles in the carcinogenesis of hepatocellular carcinoma.%目的:分选及鉴定人肝癌PLC/PRF-5细胞中的肝癌干细胞样细胞,研究其microRNAs(miRNAs)表达谱.方法:以ABCG2为表面标志,免疫磁珠法分选、流式细胞术检测ABCG2+和ABCG2-PLC/PRF-5细胞,观察ABCG2+与ABCG2-PLC/PRF-5细胞的琼脂克隆

  5. Remission of invasive, cancer stem-like glioblastoma xenografts using lentiviral vector-mediated suicide gene therapy.

    Directory of Open Access Journals (Sweden)

    Peter C Huszthy

    Full Text Available BACKGROUND: Glioblastoma is the most frequent and most malignant primary brain tumor with a poor prognosis. The translation of therapeutic strategies for glioblastoma from the experimental phase into the clinic has been limited by insufficient animal models, which lack important features of human tumors. Lentiviral gene therapy is an attractive therapeutic option for human glioblastoma, which we validated in a clinically relevant animal model. METHODOLOGY/PRINCIPAL FINDINGS: We used a rodent xenograft model that recapitulates the invasive and angiogenic features of human glioblastoma to analyze the transduction pattern and therapeutic efficacy of lentiviral pseudotyped vectors. Both, lymphocytic choriomeningitis virus glycoprotein (LCMV-GP and vesicular stomatitis virus glycoprotein (VSV-G pseudotyped lentiviral vectors very efficiently transduced human glioblastoma cells in vitro and in vivo. In contrast, pseudotyped gammaretroviral vectors, similar to those evaluated for clinical therapy of glioblastoma, showed inefficient gene transfer in vitro and in vivo. Both pseudotyped lentiviral vectors transduced cancer stem-like cells characterized by their CD133-, nestin- and SOX2-expression, the ability to form spheroids in neural stem cell medium and to express astrocytic and neuronal differentiation markers under serum conditions. In a therapeutic approach using the suicide gene herpes simplex virus thymidine kinase (HSV-1-tk fused to eGFP, both lentiviral vectors mediated a complete remission of solid tumors as seen on MRI resulting in a highly significant survival benefit (p<0.001 compared to control groups. In all recurrent tumors, surviving eGFP-positive tumor cells were found, advocating prodrug application for several cycles to even enhance and prolong the therapeutic effect. CONCLUSIONS/SIGNIFICANCE: In conclusion, lentiviral pseudotyped vectors are promising candidates for gene therapy of glioma in patients. The inefficient gene delivery

  6. TNFα enhances cancer stem cell-like phenotype via Notch-Hes1 activation in oral squamous cell carcinoma cells. (United States)

    Lee, Sung Hee; Hong, Hannah S; Liu, Zi Xiao; Kim, Reuben H; Kang, Mo K; Park, No-Hee; Shin, Ki-Hyuk


    Cancer stem-like cell (CSC; also known as tumor initiating cell) is defined as a small subpopulation of cancer cells within a tumor and isolated from various primary tumors and cancer cell lines. CSCs are highly tumorigenic and resistant to anticancer treatments. In this study, we found that prolonged exposure to tumor necrosis factor alpha (TNFα), a major proinflammatory cytokine, enhances CSC phenotype of oral squamous cell carcinoma (OSCC) cells, such as an increase in tumor sphere-forming ability, stem cell-associated genes expression, chemo-radioresistance, and tumorigenicity. Moreover, activation of Notch1 signaling was detected in the TNFα-exposed cells, and suppression of Notch1 signaling inhibited CSC phenotype. Furthermore, we demonstrated that inhibition of a Notch downstream target, Hes1, led to suppression of CSC phenotype in the TNFα-exposed cells. We also found that Hes1 expression is commonly upregulated in OSCC lesions compared to precancerous dysplastic lesions, suggesting the possible involvement of Hes1 in OSCC progression and CSC in vivo. In conclusion, inflammatory cytokine exposure may enhance CSC phenotype of OSCC, in part by activating the Notch-Hes1 pathway.

  7. MiR-214 targets β-catenin pathway to suppress invasion, stem-like traits and recurrence of human hepatocellular carcinoma.

    Directory of Open Access Journals (Sweden)

    Hongping Xia

    Full Text Available The down-regulation of miR-214 has previously been observed in human hepatocellular carcinoma (HCC. Here, we demonstrated the down-regulation of miR-214 is associated with cell invasion, stem-like traits and early recurrence of HCC. Firstly, we validated the suppression of miR-214 in human HCC by real-time quantitative RT-PCR (qRT-PCR in 20 paired tumor and non-tumor liver tissues of HCC patients and 10 histologically normal liver tissues from colorectal cancer patients with liver metastases. Further qRT-PCR analysis of 50 HCC tissues from an independent cohort of HCC patients of whom 29 with early recurrent disease (<2 years and 21 with late recurrent disease demonstrated that the suppression of miR-214 was significantly more suppressed in samples from HCC patients with early recurrent disease compared those from patients with no recurrence. Re-expression of miR-214 significantly suppressed the growth of HCC cells in vitro and reduced their tumorigenicity in vivo. The enhancer of zeste homologue 2 (EZH2 and β-catenin (CTNNB1 was identified as two potential direct downstream targets of miR-214 through bioinformatics analysis and experimentally validated the miRNA-target interactions with a dual-firefly luciferase reporter assay. In corroborate with this, both EZH2 and CTNNB1 are found to be significantly overexpressed in human HCC biopsies. Since EZH2 can regulate CTNNB1, CTNNB1 can also be an indirect target of miR-214 through EZH2. Silencing EZH2 or CTNNB1 expression suppressed the growth and invasion of HCC cells and induced E-cadherin (CDH1, known to inhibit cell invasion and metastasis. Furthermore, the silencing of miR-214 or overexpression of EZH2 increased EpCAM(+ stem-like cells through the activation of CTNNB1. Interestingly, the up-regulation of EZH2, CTNNB1 and the down-regulation of CDH1 in HCC patients correlated with early recurrent disease and can be an independent predictor of poor survival. Therefore, miR-214 can directly or

  8. Ovarian cancer plasticity and epigenomics in the acquisition of a stem-like phenotype

    Directory of Open Access Journals (Sweden)

    Berry Nicholas B


    Full Text Available Abstract Aggressive epithelial ovarian cancer (EOC is genetically and epigenetically distinct from normal ovarian surface epithelial cells (OSE and early neoplasia. Co-expression of epithelial and mesenchymal markers in EOC suggests an involvement of epithelial-mesenchymal transition (EMT in cancer initiation and progression. This phenomenon is often associated with acquisition of a stem cell-like phenotype and chemoresistance that correlate with the specific gene expression patterns accompanying transformation, revealing a plasticity of the ovarian cancer cell genome during disease progression. Differential gene expressions between normal and transformed cells reflect the varying mechanisms of regulation including genetic changes like rearrangements within the genome, as well as epigenetic changes such as global genomic hypomethylation with localized promoter CpG island hypermethylation. The similarity of gene expression between ovarian cancer cells and the stem-like ovarian cancer initiating cells (OCIC are surprisingly also correlated with epigenetic mechanisms of gene regulation in normal stem cells. Both normal and cancer stem cells maintain genetic flexibility by co-placement of activating and/or repressive epigenetic modifications on histone H3. The co-occupancy of such opposing histone marks is believed to maintain gene flexibility and such bivalent histones have been described as being poised for transcriptional activation or epigenetic silencing. The involvement of both-microRNA (miRNA mediated epigenetic regulation, as well as epigenetic-induced changes in miRNA expression further highlight an additional complexity in cancer stem cell epigenomics. Recent advances in array-based whole-genome/epigenome analyses will continue to further unravel the genomes and epigenomes of cancer and cancer stem cells. In order to illuminate phenotypic signatures that delineate ovarian cancer from their associated cancer stem cells, a priority must lie

  9. Antiproliferative Effect of Androgen Receptor Inhibition in Mesenchymal Stem-Like Triple-Negative Breast Cancer

    Directory of Open Access Journals (Sweden)

    Aiyu Zhu


    Full Text Available Background/Aims: Androgen receptor (AR, a steroid hormone receptor, has recently emerged as prognostic and treatment-predictive marker in breast cancer. Previous studies have shown that AR is widely expressed in up to one-third of triple-negative breast cancer (TNBC. However, the role of AR in TNBC is still not fully understood, especially in mesenchymal stem-like (MSL TNBC cells. Methods: MSL TNBC MDA-MB-231 and Hs578T breast cancer cells were exposed to various concentration of agonist 5-α-dihydrotestosterone (DHT or nonsteroidal antagonist bicalutamide or untreated. The effects of AR on cell viability and apoptosis were determined by MTT assay, cell counting, flow cytometry analysis and protein expression of p53, p73, p21 and Cyclin D1 were analyzed by western blotting. The bindings of AR to p73 and p21 promoter were detected by ChIP assay. MDA-MB-231 cells were transplanted into nude mice and the tumor growth curves were determined and expression of AR, p73 and p21 were detected by Immunohistochemistry (IHC staining after treatment of DHT or bicalutamide. Results: We demonstrate that AR agonist DHT induces MSL TNBC breast cancer cells proliferation and inhibits apoptosis in vitro. Similarly, activated AR significantly increases viability of MDA-MB-231 xenografts in vivo. On the contrary, AR antagonist, bicalutamide, causes apoptosis and exerts inhibitory effects on the growth of breast cancer. Moreover, DHT-dependent activation of AR involves regulation in the cell cycle related genes, including p73, p21 and Cyclin D1. Further investigations indicate the modulation of AR on p73 and p21 mediated by direct binding of AR to their promoters, and DHT could make these binding more effectively. Conclusions: Our study demonstrates the tumorigenesis role of AR and the inhibitory effect of bicalutamide in AR-positive MSL TNBC both in vitro and in vivo, suggesting that AR inhibition could be a potential therapeutic approach for AR-positive TNBC

  10. Characterization of cancer stem-like cells in a novel STI571-resistant chronic myeloid leukemia cell line%K562多药耐药细胞系中肿瘤干细胞样细胞对伊马替尼耐药机制的初步研究

    Institute of Scientific and Technical Information of China (English)

    Baijun Fang; Yongping Song; Yanli Zhang; Quande Lin; Xudong Wei


    Objective: To characterize a novel chronic myeloid leukemia (CML) cell line and to further elucidate the mechanisms of resistance to STI571. Methods: A novel K562 cell line (K562NP16) was achieved after exposure of the K562 cells to VP16. A small subpopulation (K562NP16 SP) that was capable of excluding Hoechst 33342 in the K562NP16 cell line was isolated by flow cytometry sorting. The rest of the K562NP16 cells were classified as non-SP K562NP16. The mechanisms involved in K562NP16 SP cells which became resistant to STI571 were studied. Results: The levels of Bcr-Abl and Abl proteins were similar in the K562 cell line and in non-SP K562NP16 and K562NP16 SP cells. The multidrug-resistant gene 1 (MDR1) expression of the 170 kDa P-glycoprotein (P-gp) was detected in K562NP16 non-SP and K562NP16 SP cells but not in K562 cells. The expression levels of P-gp in the two K562NP16 cell lines were similar. Compared with non-SP K562/VP16, the K562NP16 SP cells were more resistant to STI571. This resistance could hardly be reversed by many multidrug resistance inhibitors. In addition, in vivo study showed that the K562NP16 SP cells induced tumorigenesis in mice, while the K562NP16 non-SP cells failed to do so. Conclusion: A novel K562 cell line, K562NP16, was generated. A small side population K562NP16 SP cells, had high resistance to STI571 treatment and more tumorigenic than the K562 cells. It may represent the cancer stem cells of the K562NP16 cell line.

  11. ΔNp63 promotes stem cell activity in mammary gland development and basal-like breast cancer by enhancing Fzd7 expression and Wnt signalling. (United States)

    Chakrabarti, Rumela; Wei, Yong; Hwang, Julie; Hang, Xiang; Andres Blanco, Mario; Choudhury, Abrar; Tiede, Benjamin; Romano, Rose-Anne; DeCoste, Christina; Mercatali, Laura; Ibrahim, Toni; Amadori, Dino; Kannan, Nagarajan; Eaves, Connie J; Sinha, Satrajit; Kang, Yibin


    Emerging evidence suggests that cancer is populated and maintained by tumour-initiating cells (TICs) with stem-like properties similar to those of adult tissue stem cells. Despite recent advances, the molecular regulatory mechanisms that may be shared between normal and malignant stem cells remain poorly understood. Here we show that the ΔNp63 isoform of the Trp63 transcription factor promotes normal mammary stem cell (MaSC) activity by increasing the expression of the Wnt receptor Fzd7, thereby enhancing Wnt signalling. Importantly, Fzd7-dependent enhancement of Wnt signalling by ΔNp63 also governs tumour-initiating activity of the basal subtype of breast cancer. These findings establish ΔNp63 as a key regulator of stem cells in both normal and malignant mammary tissues and provide direct evidence that breast cancer TICs and normal MaSCs share common regulatory mechanisms.

  12. Direct contact with perivascular tumor cells enhances integrin αvβ3 signaling and migration of endothelial cells (United States)

    Burgett, Monica E.; Lathia, Justin D.; Roth, Patrick; Nowacki, Amy S.; Galileo, Deni S.; Pugacheva, Elena; Huang, Ping; Vasanji, Amit; Li, Meizhang; Byzova, Tatiana; Mikkelsen, Tom; Bao, Shideng; Rich, Jeremy N.; Weller, Michael; Gladson, Candece L.


    The secretion of soluble pro-angiogenic factors by tumor cells and stromal cells in the perivascular niche promotes the aggressive angiogenesis that is typical of glioblastoma (GBM). Here, we show that angiogenesis also can be promoted by a direct interaction between brain tumor cells, including tumor cells with cancer stem-like properties (CSCs), and endothelial cells (ECs). As shown in vitro, this direct interaction is mediated by binding of integrin αvβ3 expressed on ECs to the RGD-peptide in L1CAM expressed on CSCs. It promotes both EC network formation and enhances directed migration toward basic fibroblast growth factor. Activation of αvβ3 and bone marrow tyrosine kinase on chromosome X (BMX) is required for migration stimulated by direct binding but not for migration stimulated by soluble factors. RGD-peptide treatment of mice with established intracerebral GBM xenografts significantly reduced the percentage of Sox2-positive tumor cells and CSCs in close proximity to ECs, decreased integrin αvβ3 and BMX activation and p130CAS phosphorylation in the ECs, and reduced the vessel surface area. These results reveal a previously unrecognized aspect of the regulation of angiogenesis in GBM that can impact therapeutic anti-angiogenic targeting. PMID:27270311

  13. Completely Enhanced Cell Phone Keypad

    CERN Document Server

    Sharmeen, Rezwana; Ahmad, Shabbir; Kamruzzaman, S M


    The enhanced frequency based keypad is designed to speed up the typing process. This paper will show that the proposed layout will increase the typing speed and be flexible for thumb. Traditional cell phone keypad is not a scientific keypad from the frequency point of view. Approaches have been explored to speed up the typing process. We found that no manufacturer has considered the frequency of the alphabet. The current architecture does not provide flexibility although the users are accustomed to the currently available multi-tapping keypad. Since the currently available keypad layouts are not best suited for users, this paper will suggest a keypad for cell phone and other cellular device based on the frequency of the alphabet in English language and also with the view of structure of human finger movements to provide a flexible and fast cell phone keypad. It also takes into consideration the key jamming problem that was available in typewriter. At first we identified those keys of cell phone, which are eas...

  14. The NSL chromatin-modifying complex subunit KANSL2 regulates cancer stem-like properties in glioblastoma that contribute to tumorigenesis (United States)

    Ferreyra-Solari, Nazarena; Belforte, Fiorella S.; Canedo, Lucía; Videla-Richardson, Guillermo A.; Espinosa, Joaquín M.; Rossi, Mario; Serna, Eva; Riudavets, Miguel A.; Martinetto, Horacio; Sevlever, Gustavo; Perez-Castro, Carolina


    KANSL2 is an integral subunit of the Non-Specific Lethal (NSL) chromatin-modifying complex which contributes to epigenetic programs in embryonic stem cells. In this study, we report a role for KANSL2 in regulation of stemness in glioblastoma (GBM), which is characterized by heterogeneous tumor stem-like cells associated with therapy resistance and disease relapse. KANSL2 expression is upregulated in cancer cells, mainly at perivascular regions of tumors. RNAi-mediated silencing of KANSL2 in GBM cells impairs their tumorigenic capacity in mouse xenograft models. In clinical specimens, we found that expression levels of KANSL2 correlate with stemness markers in GBM stem-like cell populations. Mechanistic investigations showed that KANSL2 regulates cell self-renewal, which correlates with effects on expression of the stemness transcription factor POU5F1. RNAi-mediated silencing of POU5F1 reduced KANSL2 levels, linking these two genes to stemness control in GBM cells. Together, our findings indicate that KANSL2 acts to regulate the stem cell population in GBM, defining it as a candidate GBM biomarker for clinical use. PMID:27406830

  15. 高表达β1,6分支N-糖链的骨肉瘤类肿瘤干细胞诱导巨噬细胞M2表型分化%Cancer stem-like cells from osteosarcoma with highly expressed beta 1,6 GlcNAc branched N-glycan induce a phenotypic switch in polarization of bone marrow-derived macrophages

    Institute of Scientific and Technical Information of China (English)

    张思胜; 刘世清


    Objective To investigate the effect of cancer stem-like cells (CSCs) from osteosarcoma cell line on the polarization of macrophages.Methods CSCs were pre-treated with different doses of swainsonine and co-cultured with macrophages,and the phenotypic specific markers on macrophages were detected respectively.Results Compared to the control group,the expression of Arg-1 [(12.0 ± 3.1) % vs.(40.0±2.6)%,P<0.05] and interleukin (IL)-10 [(90.0±4.4) ng/Lvs.(150.0±6.8) ng/L,P <0.05] in macrophages co-cultured with CSCs pre-treated with swainsonine (1 mg/L) was decreased,whereas inducible nitric oxide synthase (iNOS) [(50.0 ±2.1)% vs.(12.0 ± 1.3)%,P<0.05] and tumor necrosis factor (TNF)-α [(240.0 ± 8.1) ng/L vs.(50.0 ± 3.3) ng/L,P < 0.05] increased.Conclusion Increased expression of beta-1,6-oligosaccharide in CSCs derived from osteosarcoma cell line induced the differentiation of bone marrow-derived macrophages into anti-inflammatory M2 macrophages.%目的 检测骨肉瘤类肿瘤干细胞对巨噬细胞表型转换的影响.方法 苦马豆素预处理的CD133+ CD44+骨肉瘤细胞LM8与骨髓巨噬细胞共培养后检测巨噬细胞表型相关标志分子的表达.结果 与未处理组比较,1 mg/L苦马豆素处理组巨噬细胞精氨酸酶(Arg)-1[(12.0±3.1)%比(40.0±2.6)%,P<0.05]和白细胞介素(IL)-10[(90.0±4.4) ng/L比(150.0 ±6.8) ng/L,P<0.05]表达下降,诱导型一氧化氮合酶(iNOS)[(50.0±2.1)%比(12.0±1.3)%,P<0.05]与肿瘤坏死因子(TNF)-α[(240.0 ±8.1)ng/L比(50.0±3.3) ng/L,P<0.05]表达上升.结论 骨肉瘤类肿瘤干细胞高表达1,6分支N-糖链诱导巨噬细胞M2表型转换.

  16. Isolation and characterization of cardiogenic, stem-like cardiac precursors from heart samples of patients with congenital heart disease. (United States)

    Ghazizadeh, Zaniar; Vahdat, Sadaf; Fattahi, Faranak; Fonoudi, Hananeh; Omrani, Gholamreza; Gholampour, Maziar; Aghdami, Nasser


    Regenerative therapies based on resident human cardiac progenitor cells (hCPCs) are a promising alternative to medical treatments for patients with myocardial infarction. However, hCPCs are rare in human heart and finding efficient source and proper surface marker for isolation of these cells would make them a good candidate for therapy. We have isolated 5.34∗10(6)±2.04∗10(5)/g viable cells from 35 heart tissue samples of 23 patients with congenital heart disease obtained during their heart surgery along with 6 samples from 3 normal subjects during cardiac biopsy. According to FACS analysis, younger ages, atrial specimen and disease with increased pulmonary vascular resistance were associated with higher percentage of c-kit(+) (CD117) hCPCs. Analysis for other stemness markers revealed increased CD133(+) cells in the hearts of patients with congenital heart disease. By using both immune-labeling and PCR, we demonstrated that these cells express key cardiac lineage and endothelial transcription factors and structural proteins during in vitro differentiation and do express stemness transcription factors in undifferentiated state. Another novel datum of potentially relevant interest is their ability in promoting greater myocardial regeneration and better survival in rat model of myocardial infarction following transplantation. Our results could provide evidence for conditions associated with enriched hCPCs in patients with congenital heart disease. Moreover, we showed presence of a significant number of CD133 expressing cardiogenic stem-like cardiac precursors in the heart of patients with congenital heart disease, which could be isolated and stored for future regenerative therapies in these patients. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Enhancement of cancer stem cell susceptibility to conventional treatments through complementary yoga therapy: possible cellular and molecular mechanisms. (United States)

    Bhargav, Hemant; Metri, Kashinath; Raghuram, Nagarathna; Ramarao, Nagendra Hongasandra; Koka, Prasad S


    Cancer stem cells (CSCs) are stem-like tumor populations that are reported to contribute towards tumor growth, maintenance and recurrence after therapy. Hypoxia increases CSC fraction and promotes acquisition of a stem-cell-like state. Cancer stem cells are critically dependant on the hypoxia-inducible factor-1 (HIF-1) for survival, self-renewal, tumor growth and maintenance of their undifferentiated phenotype. Recent researches show that stage of differentiation of the tumor cells is predictive of their susceptibility to natural killer cell (NK) cell mediated cytotoxicity and cancer stem cells are significant targets of NK cell cytotoxicity. Studies also show that reversion of tumor cells to a less-differentiated phenotype can be achieved by blocking NFκB. Yoga therapy (yogic lifestyle modifications encompassing physical postures, breathing practices, relaxation techniques and meditations) is known to modulate neural, endocrine and immune functions at the cellular level through influencing cell cycle control, aging, oxidative stress, apoptosis and several pathways of stress signaling molecules. Yoga therapy has also been shown to enhance natural killer cell activity and modulate stress and DNA damage in breast cancer patients receiving radiotherapy. Recent study found that brief daily yogic meditation may reverse the pattern of increased NFκB-related transcription of pro-inflammatory cytokines in leukocytes. Thus, yoga therapy has the potential to reduce cancer stem cell survival, self -renewal and tumor growth by modifying the tumor micro-environment through various mechanisms such as; 1) reducing HIF-1 activity by enhanced oxygenation, 2) promoting NK cell activity directly (or indirectly through down regulating NFκB expression), thereby enhancing NK cell mediated CSC lysis, and 3) by minimizing the aberrant expressions or activities of various hormones, cytokines, chemokines and tumor signaling pathways. Yoga therapy may have a synergistic effect with

  18. Current-Enhanced Quantum Well Solar Cells

    Institute of Scientific and Technical Information of China (English)

    LOU Chao-Gang; SUN Qiang; XU Jun; ZHANG Xiao-Bing; LEI Wei; WANG Bao-Ping; CHEN Wen-Jun; QIAO Zai-Xiang


    We present the experimental results that demonstrate the enhancement of the short-circuit current of quantum well solar cells. The spectral response shows that the introduction of quantum wells extends the absorption spectrum of solar cells. The current densities under different truncated spectrums significantly increase, showing that quantum well solar cells are suitable to be the middle cells of GaInP/GaAs/Ge triple-junction solar cells to increase their overall conversion efficiency.

  19. Inhibition of Sonic Hedgehog Signaling Pathway by Thiazole Antibiotic Thiostrepton Attenuates the CD44+/CD24-Stem-Like Population and Sphere-Forming Capacity in Triple-Negative Breast Cancer

    Directory of Open Access Journals (Sweden)

    Na Yang


    Full Text Available Background/Aim: Triple-negative breast cancer (TNBC represents a particular clinical challenge because these cancers do not respond to endocrine therapy or other available targeted agents. The lack of effective agents and obvious targets are major challenges in treating TNBC. In this study we explored the cytostatic effect of thiazole ring containing antibiotic drug thiostrepton on TNBC cell lines and investigated the molecular mechanism. Methods: Cell viability was measured by MTT assay. Cell surface marker was monitored by FCM. Western blot was applied to assess the protein expression levels of target genes. Results: We found that thiostrepton remarkably suppressed the CD44+/CD24- stem-like population and sphere forming capacity of TNBC cell lines. Notably, we showed for the first time that thiostrepton exerted its pharmacological action by targeting sonic hedgehog (SHH signaling pathway. Thiostrepton repressed SHH ligand expression and reduced Gli-1 nuclear localization in TNBC cell line. Furthermore, the downstream target of SHH signaling undergone dose-dependent, rapid, and sustained loss of mRNA transcript level after thiostrepton treatment. Finally, we showed that SHH ligand was essential for maintaining CD44+/CD24- stem-like population in TNBC cell line. Conclusion: We conclude that thiostrepton suppresses the CD44+/CD24- stem-like population through inhibition of SHH signaling pathway. Our results give a new insight into the mechanism of thiostrepton anti-tumor activity and suggest thiostrepton as a promising agent that targets hedgehog signaling pathway in TNBC.

  20. Autophagy positively regulates the CD44(+) CD24(-/low) breast cancer stem-like phenotype. (United States)

    Cufí, Sílvia; Vazquez-Martin, Alejandro; Oliveras-Ferraros, Cristina; Martin-Castillo, Begoña; Vellon, Luciano; Menendez, Javier A


    The molecular mechanisms used by breast cancer stem cells (BCSCs) to survive and/or maintain their undifferentiated CD44(+) CD24(-/low ) mesenchymal-like antigenic state remains largely unexplored. Autophagy, a key homeostatic process of cytoplasmic degradation and recycling evolved to respond to stress conditions, might be causally fundamental in the biology of BCSCs. Stable & specific knockdown of autophagy-regulatory genes by lentiviral-delivered small hairpin (sh) RNA drastically decreased the number of JIMT-1 epithelial BC cells bearing CD44(+) CD24(-/low) cell-surface antigens from ~75% in parental and control (-) shRNA-transduced cells to 26% and 7% in ATG8/LC3 shRNA- and ATG12 shRNA-transduced cells, respectively. Autophagy inhibition notably enhanced transcriptional activation of CD24 gene, potentiating the epithelial-like phenotype of CD44(+) CD24(+) cells versus the mesenchymal CD44(+) CD24(-/low ) progeny. EMT-focused Real Time RT-PCR profiling revealed that genetic ablation of autophagy transcriptionally repressed the gene coding for the mesenchymal filament vimentin (VIM). shRNA-driven silencing of the ATG12 gene and disabling the final step in the autophagy pathway by the antimalarial drug chloroquine both prevented TGFb1-induced accumulation of vimentin in JIMT-1 cells. Knockdown of autophagy-specific genes was sufficient also to increase by up to 11-times the number of CD24(+) cells in MDA-MB-231 cells, a BC model of mesenchymal origin that is virtually composed of CD44(+) CD24(-/low ) cells. Chloroquine treatment augmented the number of CD24(+) cells and concomitantly reduced constitutive overexpression of vimentin in MDA-MB-231 cells. This is the first report demonstrating that autophagy is mechanistically linked to the maintenance of tumor cells expressing high levels of CD44 and low levels of CD24, which are typical of BCSCs.

  1. Osteopontin-CD44 signaling in the glioma perivascular niche enhances cancer stem cell phenotypes and promotes aggressive tumor growth (United States)

    Pietras, Alexander; Katz, Amanda M.; Ekström, Elin J.; Wee, Boyoung; Halliday, John J.; Pitter, Kenneth L.; Werbeck, Jillian L.; Amankulor, Nduka M.; Huse, Jason T.; Holland, Eric C.


    Summary Stem-like glioma cells reside within a perivascular niche and display hallmark radiation resistance. Understanding of the mechanisms underlying these properties will be vital for the development of effective therapies. Here we show that the stem cell marker CD44 promotes cancer stem cell phenotypes and radiation resistance. In a mouse model of glioma, Cd44−/− and Cd44+/− animals showed improved survival compared to controls. The CD44 ligand Osteopontin shared a perivascular expression pattern with CD44 and promoted glioma stem cell-like phenotypes. These effects were mediated via the γ-secretase regulated intracellular domain of CD44, which promoted aggressive glioma growth in vivo and stem cell-like phenotypes via CBP/p300-dependent enhancement of HIF-2α activity. In human glioblastoma multiforme, expression of CD44 correlated with hypoxia-induced gene signatures and poor survival. Together, these data suggest that in the glioma perivascular niche, Osteopontin promotes stem cell-like properties and radiation resistance in adjacent tumor cells via activation of CD44 signaling. PMID:24607407

  2. Enhanced live cell imaging via photonic crystal enhanced fluorescence microscopy. (United States)

    Chen, Weili; Long, Kenneth D; Yu, Hojeong; Tan, Yafang; Choi, Ji Sun; Harley, Brendan A; Cunningham, Brian T


    We demonstrate photonic crystal enhanced fluorescence (PCEF) microscopy as a surface-specific fluorescence imaging technique to study the adhesion of live cells by visualizing variations in cell-substrate gap distance. This approach utilizes a photonic crystal surface incorporated into a standard microscope slide as the substrate for cell adhesion, and a microscope integrated with a custom illumination source as the detection instrument. When illuminated with a monochromatic light source, angle-specific optical resonances supported by the photonic crystal enable efficient excitation of surface-confined and amplified electromagnetic fields when excited at an on-resonance condition, while no field enhancement occurs when the same photonic crystal is illuminated in an off-resonance state. By mapping the fluorescence enhancement factor for fluorophore-tagged cellular components between on- and off-resonance states and comparing the results to numerical calculations, the vertical distance of labelled cellular components from the photonic crystal substrate can be estimated, providing critical and quantitative information regarding the spatial distribution of the specific components of cells attaching to a surface. As an initial demonstration of the concept, 3T3 fibroblast cells were grown on fibronectin-coated photonic crystals with fluorophore-labelled plasma membrane or nucleus. We demonstrate that PCEF microscopy is capable of providing information about the spatial distribution of cell-surface interactions at the single-cell level that is not available from other existing forms of microscopy, and that the approach is amenable to large fields of view, without the need for coupling prisms, coupling fluids, or special microscope objectives.

  3. MYC activates stem-like cell potential in hepatocarcinoma by a p53-dependent mechanism

    DEFF Research Database (Denmark)

    Akita, Hirofumi; Marquardt, Jens U; Durkin, Marian E;


    Activation of c-MYC is an oncogenic hallmark of many cancers including liver cancer, and is associated with a variety of adverse prognostic characteristics. Despite a causative role during malignant transformation and progression in hepatocarcinogenesis, consequences of c-MYC activation for the b...

  4. ES micro-environment enhances stemness and inhibits apoptosis in human limbal stem cells via the maintenance of telomerase activity.

    Directory of Open Access Journals (Sweden)

    Zhiping Liu

    Full Text Available Our previous work had found that telomerase rejuvenated in the cytoplasm of corneal epithelial cells cultured in embryonic stem cell-conditioned medium, the functional properties of stem-like corneal epithelial cells can be enhanced by co-culturing with embryonic stem cells (ESCs via activation of the integrinβ1-FAK-PI3K/Akt signaling pathway. The goal of this study was to explore the potential molecular mechanisms of the ES micro-environment that enhance the stem cell-like phenotype and inhibit apoptosis in human limbal stem cells (LSC. The LSC were cultured in different media, either CnT-20 medium or CnT-20 +20% ES culture supernatant (ESC-CM. We observed that LSC cultured in ESC-CM had an increased proliferative capacity, greater serial passage capacity, higher colony-forming efficiency (CFE and higher levels of stem cell-associated marker than those cultured in CnT-20. Compared with CnT-20, ESC-CM enhanced the undifferentiated status and inhibited apoptosis in the LSC by promoting the maintenance of telomerase activity, which could reduce the generation of reactive oxygen species (ROS, maintain the membrane potential (Δψm at higher levels and reduce the expression of the p21 protein. Our findings indicated that ESC-CM system induced LSC to maintain a stem cell phenotype and inhibit the process of apoptosis. These effects might partially be achieved via the telomerase-p21-mitochondrial axis and the activation of the FAK/Wnt signaling pathways. This study may have high impact and clinic implication on the expansion of LSC in regenerative medicine, especially for ocular surface reconstruction.

  5. Neural Progenitor Cells Undergoing Yap/Tead-Mediated Enhanced Self-Renewal Form Heterotopias More Easily in the Diencephalon than in the Telencephalon. (United States)

    Saito, Kanako; Kawasoe, Ryotaro; Sasaki, Hiroshi; Kawaguchi, Ayano; Miyata, Takaki


    Spatiotemporally ordered production of cells is essential for brain development. Normally, most undifferentiated neural progenitor cells (NPCs) face the apical (ventricular) surface of embryonic brain walls. Pathological detachment of NPCs from the apical surface and their invasion of outer neuronal territories, i.e., formation of NPC heterotopias, can disrupt the overall structure of the brain. Although NPC heterotopias have previously been observed in a variety of experimental contexts, the underlying mechanisms remain largely unknown. Yes-associated protein 1 (Yap1) and the TEA domain (Tead) proteins, which act downstream of Hippo signaling, enhance the stem-like characteristics of NPCs. Elevated expression of Yap1 or Tead in the neural tube (future spinal cord) induces massive NPC heterotopias, but Yap/Tead-induced expansion of NPCs in the developing brain has not been previously reported to produce NPC heterotopias. To determine whether NPC heterotopias occur in a regionally characteristic manner, we introduced the Yap1-S112A or Tead-VP16 into NPCs of the telencephalon and diencephalon, two neighboring but distinct forebrain regions, of embryonic day 10 mice by in utero electroporation, and compared NPC heterotopia formation. Although NPCs in both regions exhibited enhanced stem-like behaviors, heterotopias were larger and more frequent in the diencephalon than in the telencephalon. This result, the first example of Yap/Tead-induced NPC heterotopia in the forebrain, reveals that Yap/Tead-induced NPC heterotopia is not specific to the neural tube, and also suggests that this phenomenon depends on regional factors such as the three-dimensional geometry and assembly of these cells.

  6. Enhanced cell growth and tumorigenicity of rat glioma cells by stable expression of human CD133 through multiple molecular actions. (United States)

    Fang, Kuan-Min; Lin, Tzu-Chien; Chan, Ti-Chun; Ma, Shi-Zhang; Tzou, Bo-Cheng; Chang, Wen-Ruei; Liu, Jun-Jen; Chiou, Shih-Hwa; Yang, Chung-Shi; Tzeng, Shun-Fen


    CD133 (Prominin-1/AC133) is generally treated as a cell surface marker found on multipotent stem cells and tumor stem-like cells, and its biological function remains debated. Genetically modified rat glioma cell lines were generated by lentiviral gene delivery of human CD133 into rat C6 glioma cells (hCD133(+) -C6) or by infection of C6 cells with control lentivirus (mock-C6). Stable hCD133 expression promoted the self-renewal ability of C6-formed spheres with an increase in the expression of the stemness markers, Bmi-1 and SOX2. Akt phosphorylation, Notch-1 activation, and Notch-1 target gene expression (Hes-1, Hey1 and Hey2) were increased in hCD133(+) -C6 when compared to mock-C6. The inhibition of Akt phosphorylation, Notch-1 activation, and Hes-1 in hCD133(+) -C6 cells effectively suppressed their clonogenic ability, indicating that these factors are involved in expanding the growth of hCD133(+) -C6. An elevated expression of GTPase-activating protein 27 (Arhgap27) was detected in hCD133(+) -C6. A decline in the invasion of hCD133(+) -C6 by knockdown of Arhgap27 expression indicated the critical role of Arhgap27 in promoting cell migration of hCD133(+) -C6. In vivo study further showed that hCD133(+) -C6 formed aggressive tumors in vivo compared to mock-C6. Exposure of hCD133(+) -C6 to arsenic trioxide not only reduced Akt phosphorylation, Notch-1 activation and Hes-1 expression in vitro, but also inhibited their tumorigenicity in vivo. The results show that C6 glioma cells with stable hCD133 expression enhanced their stemness properties with increased Notch-1/Hes-1 signaling, Akt activation, and Arhgap27 action, which contribute to increased cell proliferation and migration of hCD133(+) -C6 in vitro, as well as progressive tumor formation in vivo.

  7. Activation of c-MET induces a Stem-Like phenotype in human prostate cancer

    NARCIS (Netherlands)

    G.J.H.L. Leenders (Geert); R. Sookhlall (Rajesh); W.J. Teubel (Wilma); C.M.A. Ridder (Corrina); S. Reneman (Suzanne); A. Sacchetti (Andrea); K.J. Vissers (Kees); W.M. van Weerden (Wytske); G.W. Jenster (Guido)


    textabstractProstate cancer consists of secretory cells and a population of immature cells. The function of immature cells and their mutual relation with secretory cells are still poorly understood. Immature cells either have a hierarchical relation to secretory cells (stem cell model) or represent

  8. Radiation Enhances Regulatory T Cell Representation

    Energy Technology Data Exchange (ETDEWEB)

    Kachikwu, Evelyn L.; Iwamoto, Keisuke S.; Liao, Yu-Pei; DeMarco, John J.; Agazaryan, Nzhde [Department of Radiation Oncology, David Geffen School of Medicine at UCLA, Los Angeles, CA (United States); Economou, James S. [Department of Surgical Oncology, David Geffen School of Medicine at UCLA, Los Angeles, CA (United States); McBride, William H. [Department of Radiation Oncology, David Geffen School of Medicine at UCLA, Los Angeles, CA (United States); Schaue, Doerthe, E-mail: [Department of Radiation Oncology, David Geffen School of Medicine at UCLA, Los Angeles, CA (United States)


    Purpose: Immunotherapy could be a useful adjunct to standard cytotoxic therapies such as radiation in patients with micrometastatic disease, although successful integration of immunotherapy into treatment protocols will require further understanding of how standard therapies affect the generation of antitumor immune responses. This study was undertaken to evaluate the impact of radiation therapy (RT) on immunosuppressive T regulatory (Treg) cells. Methods and Materials: Treg cells were identified as a CD4{sup +}CD25{sup hi}Foxp3{sup +} lymphocyte subset, and their fate was followed in a murine TRAMP C1 model of prostate cancer in mice with and without RT. Results: CD4{sup +}CD25{sup hi}Foxp3{sup +} Treg cells increased in immune organs after local leg or whole-body radiation. A large part, but not all, of this increase after leg-only irradiation could be ascribed to radiation scatter and Treg cells being intrinsically more radiation resistant than other lymphocyte subpopulations, resulting in their selection. Their functional activity on a per-cell basis was not affected by radiation exposure. Similar findings were made with mice receiving local RT to murine prostate tumors growing in the leg. The importance of the Treg cell population in the response to RT was shown by systemic elimination of Treg cells, which greatly enhanced radiation-induced tumor regression. Conclusions: We conclude that Treg cells are more resistant to radiation than other lymphocytes, resulting in their preferential increase. Treg cells may form an important homeostatic mechanism for tissues injured by radiation, and in a tumor context, they may assist in immune evasion during therapy. Targeting this population may allow enhancement of radiotherapeutic benefit through immune modulation.

  9. Mast cells enhance T cell activation: Importance of mast cell-derived TNF (United States)

    Nakae, Susumu; Suto, Hajime; Kakurai, Maki; Sedgwick, Jonathon D.; Tsai, Mindy; Galli, Stephen J.


    Mast cells are not only important effector cells in immediate hypersensitivity reactions and immune responses to pathogens but also can contribute to T cell-mediated disorders. However, the mechanisms by which mast cells might influence T cells in such settings are not fully understood. We find that mast cells can enhance proliferation and cytokine production in multiple T cell subsets. Mast cell-dependent enhancement of T cell activation can be promoted by FcRI-dependent mast cell activation, TNF production by both mast cells and T cells, and mast cell-T cell contact. However, at high concentrations of cells, mast cells can promote T cell activation independent of IgE or TNF. Finally, mast cells also can promote T cell activation by means of soluble factors. These findings identify multiple mechanisms by which mast cells can influence T cell proliferation and cytokine production. allergy | asthma | autoimmunity | cytokines | immune response

  10. Imprinting localized plasmons for enhanced solar cells. (United States)

    Dunbar, Ricky B; Pfadler, Thomas; Lal, Niraj N; Baumberg, Jeremy J; Schmidt-Mende, Lukas


    Imprinted silver nanovoid arrays are investigated via angle-resolved reflectometry to demonstrate their suitability for plasmonic light trapping. Both wavelength- and subwavelength-scale nanovoids are imprinted into standard solar cell architectures to achieve nanostructured metallic electrodes which provide enhanced absorption for improving solar cell performance. The technique is versatile, low-cost and scalable and can be applied to a wide range of organic semiconductors. Absorption features which are independent of incident polarization and weakly dependent on incident angle reveal localized plasmonic modes at the structured interface. Metallic nanostructure-PCPDTBT:PCBM samples demonstrate absorption enhancements of up to 40%. The structured interface provides light trapping, which boosts absorption at wavelengths where the semiconductors absorb poorly.

  11. The selection and function of cell type-specific enhancers. (United States)

    Heinz, Sven; Romanoski, Casey E; Benner, Christopher; Glass, Christopher K


    The human body contains several hundred cell types, all of which share the same genome. In metazoans, much of the regulatory code that drives cell type-specific gene expression is located in distal elements called enhancers. Although mammalian genomes contain millions of potential enhancers, only a small subset of them is active in a given cell type. Cell type-specific enhancer selection involves the binding of lineage-determining transcription factors that prime enhancers. Signal-dependent transcription factors bind to primed enhancers, which enables these broadly expressed factors to regulate gene expression in a cell type-specific manner. The expression of genes that specify cell type identity and function is associated with densely spaced clusters of active enhancers known as super-enhancers. The functions of enhancers and super-enhancers are influenced by, and affect, higher-order genomic organization.

  12. Enhancing endothelial progenitor cell for clinical use

    Institute of Scientific and Technical Information of China (English)


    Circulating endothelial progenitor cells (EPCs) havebeen demonstrated to correlate negatively with vascularendothelial dysfunction and cardiovascular risk factors.However, translation of basic research into the clinicalpractice has been limited by the lack of unambiguousand consistent definitions of EPCs and reduced EPCcell number and function in subjects requiring them forclinical use. This article critically reviews the definitionof EPCs based on commonly used protocols, their valueas a biomarker of cardiovascular risk factor in subjectswith cardiovascular disease, and strategies to enhanceEPCs for treatment of ischemic diseases.

  13. Activation of the Lin28/let-7 Axis by Loss of ESE3/EHF Promotes a Tumorigenic and Stem-like Phenotype in Prostate Cancer. (United States)

    Albino, Domenico; Civenni, Gianluca; Dallavalle, Cecilia; Roos, Martina; Jahns, Hartmut; Curti, Laura; Rossi, Simona; Pinton, Sandra; D'Ambrosio, Gioacchino; Sessa, Fausto; Hall, Jonathan; Catapano, Carlo V; Carbone, Giuseppina M


    Although cancer stem-like cells (CSC) are thought to be the most tumorigenic, metastatic, and therapy-resistant cell subpopulation within human tumors, current therapies target bulk tumor cells while tending to spare CSC. In seeking to understand mechanisms needed to acquire and maintain a CSC phenotype in prostate cancer, we investigated connections between the ETS transcription factor ESE3/EHF, the Lin28/let-7 microRNA axis, and the CSC subpopulation in this malignancy. In normal cells, we found that ESE3/EHF bound and repressed promoters for the Lin28A and Lin28B genes while activating transcription and maturation of the let-7 microRNAs. In cancer cells, reduced expression of ESE3/EHF upregulated Lin28A and Lin28B and downregulated the let-7 microRNAs. Notably, we found that deregulation of the Lin28/let-7 axis with reduced production of let-7 microRNAs was critical for cell transformation and expansion of prostate CSC. Moreover, targeting Lin28A/Lin28B in cell lines and tumor xenografts mimicked the effects of ESE3/EHF and restrained tumor-initiating and self-renewal properties of prostate CSC both in vitro and in vivo These results establish that tight control by ESE3/EHF over the Lin28/let-7 axis is a critical barrier to malignant transformation, and they also suggest new strategies to antagonize CSC in human prostate cancer for therapeutic purposes. Cancer Res; 76(12); 3629-43. ©2016 AACR. ©2016 American Association for Cancer Research.

  14. Cholangiocarcinoma stem-like subset shapes tumor-initiating niche by educating associated macrophages

    DEFF Research Database (Denmark)

    Raggi, Chiara; Correnti, Margherita; Sica, Antonio


    providing a rationale for a synergistic therapeutic strategy for CCA-disease. LAY SUMMARY: Immune plasticity represents an important hallmark of tumor outcome. Since cancer stem cells are able to manipulate stromal cells to their needs, a better definition of key deregulated immune subtype responsible...

  15. Synaptic Contacts Enhance Cell-to-Cell Tau Pathology Propagation

    Directory of Open Access Journals (Sweden)

    Sara Calafate


    Full Text Available Accumulation of insoluble Tau protein aggregates and stereotypical propagation of Tau pathology through the brain are common hallmarks of tauopathies, including Alzheimer’s disease (AD. Propagation of Tau pathology appears to occur along connected neurons, but whether synaptic contacts between neurons are facilitating propagation has not been demonstrated. Using quantitative in vitro models, we demonstrate that, in parallel to non-synaptic mechanisms, synapses, but not merely the close distance between the cells, enhance the propagation of Tau pathology between acceptor hippocampal neurons and Tau donor cells. Similarly, in an artificial neuronal network using microfluidic devices, synapses and synaptic activity are promoting neuronal Tau pathology propagation in parallel to the non-synaptic mechanisms. Our work indicates that the physical presence of synaptic contacts between neurons facilitate Tau pathology propagation. These findings can have implications for synaptic repair therapies, which may turn out to have adverse effects by promoting propagation of Tau pathology.

  16. Synaptic Contacts Enhance Cell-to-Cell Tau Pathology Propagation. (United States)

    Calafate, Sara; Buist, Arjan; Miskiewicz, Katarzyna; Vijayan, Vinoy; Daneels, Guy; de Strooper, Bart; de Wit, Joris; Verstreken, Patrik; Moechars, Diederik


    Accumulation of insoluble Tau protein aggregates and stereotypical propagation of Tau pathology through the brain are common hallmarks of tauopathies, including Alzheimer's disease (AD). Propagation of Tau pathology appears to occur along connected neurons, but whether synaptic contacts between neurons are facilitating propagation has not been demonstrated. Using quantitative in vitro models, we demonstrate that, in parallel to non-synaptic mechanisms, synapses, but not merely the close distance between the cells, enhance the propagation of Tau pathology between acceptor hippocampal neurons and Tau donor cells. Similarly, in an artificial neuronal network using microfluidic devices, synapses and synaptic activity are promoting neuronal Tau pathology propagation in parallel to the non-synaptic mechanisms. Our work indicates that the physical presence of synaptic contacts between neurons facilitate Tau pathology propagation. These findings can have implications for synaptic repair therapies, which may turn out to have adverse effects by promoting propagation of Tau pathology.

  17. Pancreatic stellate cells enhance stem cell-like phenotypes in pancreatic cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Hamada, Shin [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Masamune, Atsushi, E-mail: [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Takikawa, Tetsuya; Suzuki, Noriaki; Kikuta, Kazuhiro; Hirota, Morihisa [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Hamada, Hirofumi [Laboratory of Oncology, Department of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Hachioji (Japan); Kobune, Masayoshi [Fourth Department of Internal Medicine, Sapporo Medical University School of Medicine, Sapporo (Japan); Satoh, Kennichi [Division of Cancer Stem Cell, Miyagi Cancer Center Research Institute, Natori (Japan); Shimosegawa, Tooru [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan)


    Highlights: Black-Right-Pointing-Pointer Pancreatic stellate cells (PSCs) promote the progression of pancreatic cancer. Black-Right-Pointing-Pointer Pancreatic cancer cells co-cultured with PSCs showed enhanced spheroid formation. Black-Right-Pointing-Pointer Expression of stem cell-related genes ABCG2, Nestin and LIN28 was increased. Black-Right-Pointing-Pointer Co-injection of PSCs enhanced tumorigenicity of pancreatic cancer cells in vivo. Black-Right-Pointing-Pointer This study suggested a novel role of PSCs as a part of the cancer stem cell niche. -- 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. Recent studies have identified that a portion of cancer cells, called 'cancer stem cells', within the entire cancer tissue harbor highly tumorigenic and chemo-resistant phenotypes, which lead to the recurrence after surgery or re-growth of the tumor. The mechanisms that maintain the 'stemness' of these cells remain largely unknown. We hypothesized that PSCs might enhance the cancer stem cell-like phenotypes in pancreatic cancer cells. Indirect co-culture of pancreatic cancer cells with PSCs enhanced the spheroid-forming ability of cancer cells and induced the expression of cancer stem cell-related genes ABCG2, Nestin and LIN28. In addition, co-injection of PSCs enhanced tumorigenicity of pancreatic cancer cells in vivo. These results suggested a novel role of PSCs as a part of the cancer stem cell niche.

  18. Sulforaphane Inhibits c-Myc-Mediated Prostate Cancer Stem-Like Traits. (United States)

    Vyas, Avani R; Moura, Michelle B; Hahm, Eun-Ryeong; Singh, Krishna Beer; Singh, Shivendra V


    Preventive and therapeutic efficiencies of dietary sulforaphane (SFN) against human prostate cancer have been demonstrated in vivo, but the underlying mechanism(s) by which this occurs is poorly understood. Here, we show that the prostate cancer stem cell (pCSC)-like traits, such as accelerated activity of aldehyde dehydrogenase 1 (ALDH1), enrichment of CD49f+ fraction, and sphere forming efficiency, are attenuated by SFN treatment. Interestingly, the expression of c-Myc, an oncogenic transcription factor that is frequently deregulated in prostate cancer cells, was markedly suppressed by SFN both in vitro and in vivo. This is biologically relevant, because the lessening of pCSC-like phenotypes mediated by SFN was attenuated when c-Myc was overexpressed. Naturally occurring thio, sulfinyl, and sulfonyl analogs of SFN were also effective in causing suppression of c-Myc protein level. However, basal glycolysis, a basic metabolic pathway that can also be promoted by c-Myc overexpression, was not largely suppressed by SFN, implying that, in addition to c-Myc, there might be another SFN-sensitive cellular factor, which is not directly involved in basal glycolysis, but cooperates with c-Myc to sustain pCSC-like phenotypes. Our study suggests that oncogenic c-Myc is a target of SFN to prevent and eliminate the onset of human prostate cancer. J. Cell. Biochem. 117: 2482-2495, 2016. © 2016 Wiley Periodicals, Inc.

  19. Plasmonic-enhanced organic solar cells (United States)

    Shahin, Shiva; Gangopadhyay, Palash; Norwood, Robert


    Organic bulk-heterojunction solar cells have several good characteristics, such as ease of fabrication, and low-cost materials. However, the bottleneck in their adoption is their much lower efficiency as compared with their silicon counterparts. In our previous work, we demonstrated that by appropriately inserting AuNPs in the OPV device, the efficiency can be increased by 30% and that silanization of ITO positively impacts device performance, where we identified the field enhancement due to AuNPs as the main reason for the increase in the efficiency of the device. In this work, we further investigate the impact of self-assembly of the gold nanoparticles on the efficiency by also considering two other factors which can possibly contribute to the improvement of our structure's performance. One is the change in the substrate's workfunction after silanization, and the other factor is the variations in PEDOT: PSS characteristics due to the AuNPs' plasmonic resonance. We conclude that the AuNPs not only increase the photon absorption efficiency but also increase the conductivity of the surrounding medium (PEDOT: PSS) thereby facilitating charge transport through PEDOT: PSS.

  20. Bystander cells enhance NK cytotoxic efficiency by reducing search time. (United States)

    Zhou, Xiao; Zhao, Renping; Schwarz, Karsten; Mangeat, Matthieu; Schwarz, Eva C; Hamed, Mohamed; Bogeski, Ivan; Helms, Volkhard; Rieger, Heiko; Qu, Bin


    Natural killer (NK) cells play a central role during innate immune responses by eliminating pathogen-infected or tumorigenic cells. In the microenvironment, NK cells encounter not only target cells but also other cell types including non-target bystander cells. The impact of bystander cells on NK killing efficiency is, however, still elusive. In this study we show that the presence of bystander cells, such as P815, monocytes or HUVEC, enhances NK killing efficiency. With bystander cells present, the velocity and persistence of NK cells were increased, whereas the degranulation of lytic granules remained unchanged. Bystander cell-derived H2O2 was found to mediate the acceleration of NK cell migration. Using mathematical diffusion models, we confirm that local acceleration of NK cells in the vicinity of bystander cells reduces their search time to locate target cells. In addition, we found that integrin β chains (β1, β2 and β7) on NK cells are required for bystander-enhanced NK migration persistence. In conclusion, we show that acceleration of NK cell migration in the vicinity of H2O2-producing bystander cells reduces target cell search time and enhances NK killing efficiency.

  1. Identification of biologically relevant enhancers in human erythroid cells. (United States)

    Su, Mack Y; Steiner, Laurie A; Bogardus, Hannah; Mishra, Tejaswini; Schulz, Vincent P; Hardison, Ross C; Gallagher, Patrick G


    Identification of cell type-specific enhancers is important for understanding the regulation of programs controlling cellular development and differentiation. Enhancers are typically marked by the co-transcriptional activator protein p300 or by groups of cell-expressed transcription factors. We hypothesized that a unique set of enhancers regulates gene expression in human erythroid cells, a highly specialized cell type evolved to provide adequate amounts of oxygen throughout the body. Using chromatin immunoprecipitation followed by massively parallel sequencing, genome-wide maps of candidate enhancers were constructed for p300 and four transcription factors, GATA1, NF-E2, KLF1, and SCL, using primary human erythroid cells. These data were combined with gene expression analyses, and candidate enhancers were identified. Consistent with their predicted function as candidate enhancers, there was statistically significant enrichment of p300 and combinations of co-localizing erythroid transcription factors within 1-50 kb of the transcriptional start site (TSS) of genes highly expressed in erythroid cells. Candidate enhancers were also enriched near genes with known erythroid cell function or phenotype. Candidate enhancers exhibited moderate conservation with mouse and minimal conservation with nonplacental vertebrates. Candidate enhancers were mapped to a set of erythroid-associated, biologically relevant, SNPs from the genome-wide association studies (GWAS) catalogue of NHGRI, National Institutes of Health. Fourteen candidate enhancers, representing 10 genetic loci, mapped to sites associated with biologically relevant erythroid traits. Fragments from these loci directed statistically significant expression in reporter gene assays. Identification of enhancers in human erythroid cells will allow a better understanding of erythroid cell development, differentiation, structure, and function and provide insights into inherited and acquired hematologic disease.

  2. Interleukin-8 enhances the effect of colchicine on cell death. (United States)

    Yokoyama, Chikako; Yajima, Chika; Machida, Tetsuro; Kawahito, Yuji; Uchida, Marie; Hisatomi, Hisashi


    Pro-inflammatory cytokines are known to be generated in tumors and play important roles in angiogenesis, mitosis, and tumor progression. However, few studies have investigated the synergistic effects of pro-inflammatory cytokines and anticancer drugs on cell death. In the present study, we examined the combined effects of pro-inflammatory cytokines and colchicine on cell death of cancer cells. Colchicine induces G2/M arrest in the cell cycle by binding to tubulin, one of the main constituents of microtubules. SUIT-2 human pancreatic cancer cell line cells overexpressing pro-inflammatory cytokines, including interleukin (IL)-1β, IL-8, and tumor necrosis factor (TNF)-α, were treated with colchicine. The effect of colchicine on cell death was enhanced in cells overexpressing IL-8. Moreover, the effect of colchicine on cell death was enhanced in cells overexpressing two IL-8 up-regulators, NF-κB and IL-6, but not in cells overexpressing an IL-8 down-regulator, splicing factor proline/glutamine-rich (SFPQ). Synergistic effects of IL-8 and colchicine were also observed in cells overexpressing IL-8 isoforms lacking the signal peptide. Therefore, IL-8 appeared to function as an enhancer of cell death in cancer cells treated with colchicine. The present results suggest a new role for IL-8 related to cell death of cancer cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Enhanced casein kinase II activity in human tumour cell cultures

    DEFF Research Database (Denmark)

    Prowald, K; Fischer, H; Issinger, O G


    Casein kinase II (CKII) activity is enhanced as much as 2-3 fold in established and 4-5-fold in transformed human cell lines when compared to that of fibroblasts and primary human tumour cell cultures where CKII activity never exceeded a basic level. The high activity of CKII in transformed cells...

  4. Fuel cell energy storage for Space Station enhancement (United States)

    Stedman, J. K.


    Viewgraphs on fuel cell energy storage for space station enhancement are presented. Topics covered include: power profile; solar dynamic power system; photovoltaic battery; space station energy demands; orbiter fuel cell power plant; space station energy storage; fuel cell system modularity; energy storage system development; and survival power supply.

  5. Lymphoid Cell-Glioma Cell Interaction Enhances Cell Coat Production by Human Gliomas: Novel Suppressor Mechanism (United States)

    Dick, Steven J.; Macchi, Beatrice; Papazoglou, Savvas; Oldfield, Edward H.; Kornblith, Paul L.; Smith, Barry H.; Gately, Maurice K.


    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.

  6. Microgravity-Enhanced Stem Cell Selection (United States)

    Claudio, Pier Paolo; Valluri, Jagan


    Stem cells, both embryonic and adult, promise to revolutionize the practice of medicine in the future. In order to realize this potential, a number of hurdles must be overcome. Most importantly, the signaling mechanisms necessary to control the differentiation of stem cells into tissues of interest remain to be elucidated, and much of the present research on stem cells is focused on this goal. Nevertheless, it will also be essential to achieve large-scale expansion and, in many cases, assemble cells in 3D as transplantable tissues. To this end, microgravity analog bioreactors can play a significant role. Microgravity bioreactors were originally conceived as a tool to study the cellular responses to microgravity. However, the technology can address some of the shortcomings of conventional cell culture systems; namely, the deficiency of mass transport in static culture and high mechanical shear forces in stirred systems. Unexpectedly, the conditions created in the vessel were ideal for 3D cell culture. Recently, investigators have demonstrated the capability of the microgravity bioreactors to expand hematopoietic stem cells compared to static culture, and facilitate the differentiation of umbilical cord stem cells into 3D liver aggregates. Stem cells are capable of differentiating into functional cells. However, there are no reliable methods to induce the stem cells to form specific cells or to gain enough cells for transplantation, which limits their application in clinical therapy. The aim of this study is to select the best experimental setup to reach high proliferation levels by culturing these cells in a microgravity-based bioreactor. In typical cell culture, the cells sediment to the bottom surface of their container and propagate as a one-cell-layer sheet. Prevention of such sedimentation affords the freedom for self-assembly and the propagation of 3D tissue arrays. Suspension of cells is easily achievable using stirred technologies. Unfortunately, in

  7. Photovoltaic enhancement of Si solar cells by assembled carbon nanotubes

    Institute of Scientific and Technical Information of China (English)

    Y. F. Zhang; Y. F. Wang; N. Chen; Y. Y. Wang; Y. Z. Zhang; Z. H. Zhou; L. M. Wei


    Photovoltaic conversion was enhanced by directly assemble of a network of single-walled carbon nanotubes (SWNTs) onto the surface of n-p junction silicon solar cells. When the density of SWNTs increased from 50 to 400 tubes µm-2, an enhancement of 3.92%in energy conversion efficiency was typically obtained. The effect of the SWNTs network is proposed for trapping incident photons and assisting electronic transportation at the interface of silicon solar cells.

  8. Efficiency Enhancement in Plasmonic IBC Solar Cells


    Christian Chaverri-Ramos; J. Ayúcar; L. Bellières; Guillermo Sánchez Plaza; James Connolly


    Silicon solar cells dominate photovoltaics but suffer from poor interaction with light. This work reports on progress regarding both spectral conversion and improved light interaction with the LIMA design [1]. This combines an efficient interdigitated back-contact (IBC) solar cell [2] with a silicon quantum dot (Si-QD) [3] to optimize the spectral distribution of the incident spectrum, and finally a front-side plasmon layer to optimize light interaction. Reflectivity after thickness and proce...

  9. Patterned Poly(dopamine) Films for Enhanced Cell Adhesion. (United States)

    Chen, Xi; Cortez-Jugo, Christina; Choi, Gwan H; Björnmalm, Mattias; Dai, Yunlu; Yoo, Pil J; Caruso, Frank


    Engineered materials that promote cell adhesion and cell growth are important in tissue engineering and regenerative medicine. In this work, we produced poly(dopamine) (PDA) films with engineered patterns for improved cell adhesion. The patterned films were synthesized via the polymerization of dopamine at the air-water interface of a floating bed of spherical particles. Subsequent dissolution of the particles yielded free-standing PDA films with tunable geometrical patterns. Our results show that these patterned PDA films significantly enhance the adhesion of both cancer cells and stem cells, thus showing promise as substrates for cell attachment for various biomedical applications.

  10. Gamma-irradiation enhances transgene expression in leukemic cells. (United States)

    Vereecque, R; Saudemont, A; Wickham, T J; Gonzalez, R; Hetuin, D; Fenaux, P; Quesnel, B


    The majority of immunotherapy-based gene therapy protocols consist of ex vivo gene transfer in tumor cells. To prevent further in vivo growth, modified cells must be irradiated before reinjection into patients. The present study examines the effects of gamma-irradiation on transgene expression in transduced leukemic cells. Human and murine leukemic cells were transfected with retroviral vectors or plasmids carrying beta-galactosidase, GM-CSF or CD80 genes. Fresh leukemic cells from patients with acute myeloid leukemia (AML) were transfected with AdZ.F(pK7) adenoviral vector. gamma-irradiation at various lethal doses enhanced transgene expression in leukemic cell lines and fresh AML cells when the gene of interest was under CMV promoter but not when SV40 promoter was used. Oxidative stress also enhanced transgene expression and both irradiation and oxidative stress effects were inhibited by addition of N-acetyl-L-cysteine, a thiol anti-oxidant, indicating the involvement of reactive oxygen species. Transgene expression was also enhanced in vivo 48 and 120 h after subcutaneous injection of irradiated leukemic cells in syngeneic mice. These results show that a cell vaccine protocol using ex vivo gene transfer of transduced cells might be feasible in acute leukemia even if leukemic cells must be irradiated at lethal doses prior to reinjection to patients.

  11. Surface-enhanced Raman spectroscopy of the endothelial cell membrane.

    Directory of Open Access Journals (Sweden)

    Simon W Fogarty

    Full Text Available We applied surface-enhanced Raman spectroscopy (SERS to cationic gold-labeled endothelial cells to derive SERS-enhanced spectra of the bimolecular makeup of the plasma membrane. A two-step protocol with cationic charged gold nanoparticles followed by silver-intensification to generate silver nanoparticles on the cell surface was employed. This protocol of post-labelling silver-intensification facilitates the collection of SERS-enhanced spectra from the cell membrane without contribution from conjugated antibodies or other molecules. This approach generated a 100-fold SERS-enhancement of the spectral signal. The SERS spectra exhibited many vibrational peaks that can be assigned to components of the cell membrane. We were able to carry out spectral mapping using some of the enhanced wavenumbers. Significantly, the spectral maps suggest the distribution of some membrane components are was not evenly distributed over the cells plasma membrane. These results provide some possible evidence for the existence of lipid rafts in the plasma membrane and show that SERS has great potential for the study and characterization of cell surfaces.

  12. Acetylcholine is released from taste cells, enhancing taste signalling. (United States)

    Dando, Robin; Roper, Stephen D


    Acetylcholine (ACh), a candidate neurotransmitter that has been implicated in taste buds, elicits calcium mobilization in Receptor (Type II) taste cells. Using RT-PCR analysis and pharmacological interventions, we demonstrate that the muscarinic acetylcholine receptor M3 mediates these actions. Applying ACh enhanced both taste-evoked Ca2+ responses and taste-evoked afferent neurotransmitter (ATP) secretion from taste Receptor cells. Blocking muscarinic receptors depressed taste-evoked responses in Receptor cells, suggesting that ACh is normally released from taste cells during taste stimulation. ACh biosensors confirmed that, indeed, taste Receptor cells secrete acetylcholine during gustatory stimulation. Genetic deletion of muscarinic receptors resulted in significantly diminished ATP secretion from taste buds. The data demonstrate a new role for acetylcholine as a taste bud transmitter. Our results imply specifically that ACh is an autocrine transmitter secreted by taste Receptor cells during gustatory stimulation, enhancing taste-evoked responses and afferent transmitter secretion.

  13. Gold nanoparticles enhanced electroporation for mammalian cell transfection. (United States)

    Zu, Yingbo; Huang, Shuyan; Liao, Wei-Ching; Lu, Yang; Wang, Shengnian


    Electroporation figured prominently as an effective nonviral gene delivery approach for its balance on the transfection efficiency and cell viability, no restrictions of probe or cell type, and operation simplicity. The commercial electroporation systems have been widely adopted in the past two decades while still carry drawbacks associated with the high applied electric voltage, unsatisfied delivery efficiency, and/or low cell viability. By adding highly conductive gold nanoparticles (AuNPs) in electroporation solution, we demonstrated enhanced electroporation performance (i.e., better DNA delivery efficiency and higher cell viability) on mammalian cells from two different aspects: the free, naked AuNPs reduce the resistance of the electroporation solution so that the local pulse strength on cells was enhanced; targeting AuNPs (e.g., Tf-AuNPs) were brought to the cell membrane to work as virtual microelectrodes to porate cells with limited area from many different sites. The enhancement was confirmed with leukemia cells in both a commercial batch electroporation system and a home-made flow-through system using pWizGFP plasmid DNA probes. Such enhancement depends on the size, concentration, and the mixing ratio of free AuNPs/Tf-AuNPs. An equivalent mixture of free AuNPs and Tf-AuNPs exhibited the best enhancement with the transfection efficiency increased 2-3 folds at minimum sacrifice of cell viability. This new delivery concept, the combination of nanoparticles and electroporation technologies, may stimulate various in vitro and in vivo biomedical applications which rely on the efficient delivery of nucleic acids, anticancer drugs, or other therapeutic materials.

  14. Side population rather than CD133+ cells distinguishes enriched tumorigenicity in hTERT-immortalized primary prostate cancer cells

    Directory of Open Access Journals (Sweden)

    Wolcott Karen


    Full Text Available Abstract Background Subpopulations of cancer cells with the capacity of generating solid tumors have been characterized. In various cancer types, including prostate cancer cells, a side population (SP and CD133-expressing cells have been proposed as containing a population cancer cells with stem-like ability. Therefore the aim of this work was to determine, in prostate cancer cell lines, the frequency and tumorigenic potential of SP and CD133+ cells. Results In vitro 2D colony-forming assay and sphere-forming assay, Flow cytometry analysis and magnetic cell sorting were utilized to sort CD133+, CD133- and Side population (SP cells. Our findings indicate that CD44 and integrin α-6 are uniformly expressed in the hTERT cell lines; however, CD133 is expressed only in a small population (in vitro and in vivo. Additionally, for the hTERT cells, SP rather than CD133 expression showed an 8-fold enhanced tumorigenic potential. The data suggest that SP cells, rather than those with CD133 marker, contain the rare population of CSC capable of producing prostate tumors. Conclusion Collectively, our data suggest that although CD133 is expressed only in a small population of hTERT-immortalized prostate cancer cells, it is not likely to be associated with stem cells, as CD133- and CD133+ cells exhibited similar tumorigenicity. However, SP isolated cells, appear to be enriched with tumorigenic stem-like cells capable of generating palpable tumors.

  15. Enhancing Solar Cell Efficiencies through 1-D Nanostructures

    Directory of Open Access Journals (Sweden)

    Yu Kehan


    Full Text Available Abstract The current global energy problem can be attributed to insufficient fossil fuel supplies and excessive greenhouse gas emissions resulting from increasing fossil fuel consumption. The huge demand for clean energy potentially can be met by solar-to-electricity conversions. The large-scale use of solar energy is not occurring due to the high cost and inadequate efficiencies of existing solar cells. Nanostructured materials have offered new opportunities to design more efficient solar cells, particularly one-dimensional (1-D nanomaterials for enhancing solar cell efficiencies. These 1-D nanostructures, including nanotubes, nanowires, and nanorods, offer significant opportunities to improve efficiencies of solar cells by facilitating photon absorption, electron transport, and electron collection; however, tremendous challenges must be conquered before the large-scale commercialization of such cells. This review specifically focuses on the use of 1-D nanostructures for enhancing solar cell efficiencies. Other nanostructured solar cells or solar cells based on bulk materials are not covered in this review. Major topics addressed include dye-sensitized solar cells, quantum-dot-sensitized solar cells, and p-n junction solar cells.

  16. Stem cell transplantation enhances endogenous brain repair after experimental stroke. (United States)

    Horie, Nobutaka; Hiu, Takeshi; Nagata, Izumi


    Stem cell transplantation for stroke treatment has been a promising therapy in small and large animal models, and many clinical trials are ongoing to establish this strategy in a clinical setting. However, the mechanism underlying functional recovery after stem cell transplantation has not been fully established and there is still a need to determine the ideal subset of stem cells for such therapy. We herein reviewed the recent evidences showing the underlying mechanism of functional recovery after cell transplantation, focusing on endogenous brain repair. First, angiogenesis/neovascularization is promoted by trophic factors including vascular endothelial growth factor secreted from stem cells, and stem cells migrated to the lesion along with the vessels. Second, axonal sprouting, dendritic branching, and synaptogenesis were enhanced altogether in the both ipsilateral and contralateral hemisphere remapping the pyramidal tract across the board. Finally, endogenous neurogenesis was also enhanced although little is known how much these neurogenesis contribute to the functional recovery. Taken together, it is clear that stem cell transplantation provides functional recovery via endogenous repair enhancement from multiple ways. This is important to maximize the effect of stem cell therapy after stroke, although it is still undetermined which repair mechanism is mostly contributed.

  17. Identification of a population of epidermal squamous cell carcinoma cells with enhanced potential for tumor formation.

    Directory of Open Access Journals (Sweden)

    Gautam Adhikary

    Full Text Available Epidermal squamous cell carcinoma is among the most common cancers in humans. These tumors are comprised of phenotypically diverse populations of cells that display varying potential for proliferation and differentiation. An important goal is identifying cells from this population that drive tumor formation. To enrich for tumor-forming cells, cancer cells were grown as spheroids in non-attached conditions. We show that spheroid-selected cells form faster growing and larger tumors in immune-compromised mice as compared to non-selected cells. Moreover, spheroid-selected cells gave rise to tumors following injection of as few as one hundred cells, suggesting these cells have enhanced tumor-forming potential. Cells isolated from spheroid-selected tumors retain an enhanced ability to grow as spheroids when grown in non-attached culture conditions. Thus, these tumor-forming cells retain their phenotype following in vivo passage as tumors. Detailed analysis reveals that spheroid-selected cultures are highly enriched for expression of epidermal stem cell and embryonic stem cell markers, including aldehyde dehydrogenase 1, keratin 15, CD200, keratin 19, Oct4, Bmi-1, Ezh2 and trimethylated histone H3. These studies indicate that a subpopulation of cells that possess stem cell-like properties and express stem cell markers can be derived from human epidermal cancer cells and that these cells display enhanced ability to drive tumor formation.

  18. Dynamic regulation of Schwann cell enhancers after peripheral nerve injury. (United States)

    Hung, Holly A; Sun, Guannan; Keles, Sunduz; Svaren, John


    Myelination of the peripheral nervous system is required for axonal function and long term stability. After peripheral nerve injury, Schwann cells transition from axon myelination to a demyelinated state that supports neuronal survival and ultimately remyelination of axons. Reprogramming of gene expression patterns during development and injury responses is shaped by the actions of distal regulatory elements that integrate the actions of multiple transcription factors. We used ChIP-seq to measure changes in histone H3K27 acetylation, a mark of active enhancers, to identify enhancers in myelinating rat peripheral nerve and their dynamics after demyelinating nerve injury. Analysis of injury-induced enhancers identified enriched motifs for c-Jun, a transcription factor required for Schwann cells to support nerve regeneration. We identify a c-Jun-bound enhancer in the gene for Runx2, a transcription factor induced after nerve injury, and we show that Runx2 is required for activation of other induced genes. In contrast, enhancers that lose H3K27ac after nerve injury are enriched for binding sites of the Sox10 and early growth response 2 (Egr2/Krox20) transcription factors, which are critical determinants of Schwann cell differentiation. Egr2 expression is lost after nerve injury, and many Egr2-binding sites lose H3K27ac after nerve injury. However, the majority of Egr2-bound enhancers retain H3K27ac, indicating that other transcription factors maintain active enhancer status after nerve injury. The global epigenomic changes in H3K27ac deposition pinpoint dynamic changes in enhancers that mediate the effects of transcription factors that control Schwann cell myelination and peripheral nervous system responses to nerve injury.

  19. Estrogen enhanced cell-cell signalling in breast cancer cells exposed to targeted irradiation

    Directory of Open Access Journals (Sweden)

    Held Kathryn D


    Full Text Available Abstract Background Radiation-induced bystander responses, where cells respond to their neighbours being irradiated are being extensively studied. Although evidence shows that bystander responses can be induced in many types of cells, it is not known whether there is a radiation-induced bystander effect in breast cancer cells, where the radiosensitivity may be dependent on the role of the cellular estrogen receptor (ER. This study investigated radiation-induced bystander responses in estrogen receptor-positive MCF-7 and estrogen receptor-negative MDA-MB-231 breast cancer cells. Methods The influence of estrogen and anti-estrogen treatments on the bystander response was determined by individually irradiating a fraction of cells within the population with a precise number of helium-3 using a charged particle microbeam. Damage was scored as chromosomal damage measured as micronucleus formation. Results A bystander response measured as increased yield of micronucleated cells was triggered in both MCF-7 and MDA-MB-231 cells. The contribution of the bystander response to total cell damage in MCF-7 cells was higher than that in MDA-MB-231 cells although the radiosensitivity of MDA-MB-231 was higher than MCF-7. Treatment of cells with 17β-estradiol (E2 increased the radiosensitivity and the bystander response in MCF-7 cells, and the effect was diminished by anti-estrogen tamoxifen (TAM. E2 also increased the level of intracellular reactive oxygen species (ROS in MCF-7 cells in the absence of radiation. In contrast, E2 and TAM had no influence on the bystander response and ROS levels in MDA-MB-231 cells. Moreover, the treatment of MCF-7 cells with antioxidants eliminated both the E2-induced ROS increase and E2-enhanced bystander response triggered by the microbeam irradiation, which indicates that ROS are involved in the E2-enhanced bystander micronuclei formation after microbeam irradiation. Conclusion The observation of bystander responses in breast

  20. Myeloid derived suppressor cells enhance IgE-mediated mast cell responses (United States)

    We previously demonstrated that enhanced development of myeloid derived suppressor cells (MDSC) in ADAM10 transgenic mice yielded resistance to infection with Nippostrongylus brasiliensis infection, and that co-culturing MDSC with IgE-activated mast cells enhanced cytokine production. In the current...

  1. CIITA enhances HIV-1 attachment to CD4+ T cells leading to enhanced infection and cell depletion. (United States)

    Porter, Kristen A; Kelley, Lauren N; Nekorchuk, Michael D; Jones, James H; Hahn, Amy B; de Noronha, Carlos M C; Harton, Jonathan A; Duus, Karen M


    Activated CD4(+) T cells are more susceptible to HIV infection than resting T cells; the reason for this remains unresolved. Induction of CIITA and subsequent expression of the MHC class II isotype HLA-DR are hallmarks of CD4(+) T cell activation; therefore, we investigated the role of CIITA expression in T cells during HIV infection. CIITA-expressing SupT1 cells display enhanced virion attachment in a gp160/CD4-dependent manner, which results in increased HIV infection, virus release, and T cell depletion. Although increased attachment and infection of T cells correlated with HLA-DR surface expression, Ab blocking, transient expression of HLA-DR without CIITA, and short hairpin RNA knockdown demonstrate that HLA-DR does not directly enhance susceptibility of CIITA-expressing cells to HIV infection. Further analysis of the remaining MHC class II isotypes, HLA-DP and HLA-DQ, MHC class I isotypes, HLA-A, HLA-B, and HLA-C, and the class II Ag presentation genes, invariant chain and HLA-DM, demonstrate that these proteins likely do not contribute to CIITA enhancement of HIV infection. Finally, we demonstrate that in activated primary CD4(+) T cells as HLA-DR/CIITA expression increases there is a corresponding increase in virion attachment. Overall, this work suggests that induction of CIITA expression upon CD4(+) T cell activation contributes to enhanced attachment, infection, virus release, and cell death through an undefined CIITA transcription product that may serve as a new antiviral target.

  2. Mast Cells Can Enhance Resistance to Snake and Honeybee Venoms (United States)

    Metz, Martin; Piliponsky, Adrian M.; Chen, Ching-Cheng; Lammel, Verena; Åbrink, Magnus; Pejler, Gunnar; Tsai, Mindy; Galli, Stephen J.


    Snake or honeybee envenomation can cause substantial morbidity and mortality, and it has been proposed that the activation of mast cells by snake or insect venoms can contribute to these effects. We show, in contrast, that mast cells can significantly reduce snake-venom-induced pathology in mice, at least in part by releasing carboxypeptidase A and possibly other proteases, which can degrade venom components. Mast cells also significantly reduced the morbidity and mortality induced by honeybee venom. These findings identify a new biological function for mast cells in enhancing resistance to the morbidity and mortality induced by animal venoms.

  3. Thermodynamics of photon-enhanced thermionic emission solar cells

    DEFF Research Database (Denmark)

    Reck, Kasper; Hansen, Ole


    Photon-enhanced thermionic emission (PETE) cells in which direct photon energy as well as thermal energy can be harvested have recently been suggested as a new candidate for high efficiency solar cells. Here, we present an analytic thermodynamical model for evaluation of the efficiency of PETE...... solar cells including an analysis of the entropy production due to thermionic emission of general validity. The model is applied to find the maximum efficiency of a PETE cell for given cathode and anode work functions and temperatures. ©...

  4. Photovoltaic cell with light trapping for enhanced efficiency (United States)

    Brener, Igal; Fofang, Nche Tumasang; Luk, Ting S.


    The efficiency of a photovoltaic cell is enhanced by light trapping using Mie-scattering nanostructures. In one embodiment, an array of nanocylinders is formed on the front surface of a silicon film to enhance forward scattering into the film, and an array of nanocylinders is formed on the back surface to enhance backscattering so that more light is absorbed within the silicon film. In an alternate embodiment, a mirror layer is formed on the back surface of the silicon film to reflect light within the film back toward the front-surface nanocylinder array.

  5. Gold nanoparticle-enhanced electroporation for leukemia cell transfection. (United States)

    Huang, Shuyan; Zu, Yingbo; Wang, Shengnian


    Electroporation serves as an attractive nonviral gene delivery approach for its effectiveness, operational simplicity, and no restrictions of probe or cell type. The commercial electroporation systems have been widely adopted in research and clinics with protocols usually compromising appropriate transfection efficiency and cell viability. By introducing gold nanoparticles (AuNPs), we demonstrated greatly enhanced performance of electroporation from two aspects: the highly conductive, naked AuNPs help reduce the potential drop consumed by the electroporation solution so that the majority of the applied voltage of an electric pulse is truly imposed on cells with enhanced field strength; AuNPs with targeting ligands (e.g., transferrin-AuNPs or Tf-AuNPs) are bound to the cell membrane, working as virtual microelectrodes to create pores on cells with limited opening area while from many different sites. The addition of AuNPs during electroporation therefore benefits not only quicker recovery and better survival of cells but also more efficient uptake of the subjected probes. Such enhancement was successfully confirmed on a chronic myeloid leukemia cell line (i.e., K562 cells) in both a commercial batch electroporation system and a homemade flow system with pWizGFP plasmid DNA probes. The efficiency was found to be dependent on the size, concentration, and mixing ratio of free AuNPs/Tf-AuNPs. An equivalent mixture of free AuNPs and Tf-AuNPs exhibited the best enhancement with the transfection efficiency increase of two to threefold at a minimum sacrifice of the cell viability.

  6. CCL5 activation of CCR5 regulates cell metabolism to enhance proliferation of breast cancer cells. (United States)

    Gao, Darrin; Rahbar, Ramtin; Fish, Eleanor N


    In earlier studies, we showed that CCL5 enhances proliferation and survival of MCF-7 breast cancer cells in an mTOR-dependent manner and we provided evidence that, for T cells, CCL5 activation of CCR5 results in increased glycolysis and enhanced ATP production. Increases in metabolic activity of cancer cells, specifically increased glycolytic activity and increased expression of glucose transporters, are associated with tumour progression. In this report, we provide evidence that CCL5 enhances the proliferation of human breast cancer cell lines (MDA-MB-231, MCF-7) and mouse mammary tumour cells (MMTV-PyMT), mediated by CCR5 activation. Concomitant with enhanced proliferation we show that CCL5 increases cell surface expression of the glucose transporter GLUT1, and increases glucose uptake and ATP production by these cells. Blocking CCL5-inducible glucose uptake abrogates the enhanced proliferation induced by CCL5. We provide evidence that increased glucose uptake is associated with enhanced glycolysis, as measured by extracellular acidification. Moreover, CCL5 enhances the invasive capacity of these breast cancer cells. Using metabolomics, we demonstrate that the metabolic signature of CCL5-treated primary mouse mammary tumour cells reflects increased anabolic metabolism. The implications are that CCL5-CCR5 interactions in the tumour microenvironment regulate metabolic events, specifically glycolysis, to promote tumour proliferation and invasion.

  7. Materials That Enhance Efficiency and Radiation Resistance of Solar Cells (United States)

    Sun, Xiadong; Wang, Haorong


    A thin layer (approximately 10 microns) of a novel "transparent" fluorescent material is applied to existing solar cells or modules to effectively block and convert UV light, or other lower solar response waveband of solar radiation, to visible or IR light that can be more efficiently used by solar cells for additional photocurrent. Meanwhile, the layer of fluorescent coating material remains fully "transparent" to the visible and IR waveband of solar radiation, resulting in a net gain of solar cell efficiency. This innovation alters the effective solar spectral power distribution to which an existing cell gets exposed, and matches the maximum photovoltaic (PV) response of existing cells. By shifting a low PV response waveband (e.g., UV) of solar radiation to a high PV response waveband (e.g. Vis-Near IR) with novel fluorescent materials that are transparent to other solar-cell sensitive wavebands, electrical output from solar cells will be enhanced. This approach enhances the efficiency of solar cells by converting UV and high-energy particles in space that would otherwise be wasted to visible/IR light. This innovation is a generic technique that can be readily implemented to significantly increase efficiencies of both space and terrestrial solar cells, without incurring much cost, thus bringing a broad base of economical, social, and environmental benefits. The key to this approach is that the "fluorescent" material must be very efficient, and cannot block or attenuate the "desirable" and unconverted" waveband of solar radiation (e.g. Vis-NIR) from reaching the cells. Some nano-phosphors and novel organometallic complex materials have been identified that enhance the energy efficiency on some state-of-the-art commercial silicon and thin-film-based solar cells by over 6%.

  8. Acidosis Promotes Metastasis Formation by Enhancing Tumor Cell Motility. (United States)

    Riemann, A; Schneider, B; Gündel, D; Stock, C; Gekle, M; Thews, O


    The tumor microenvironment is characterized by hypoxia, acidosis as well as other metabolic and biochemical alterations. Its role in cancer progression is increasingly appreciated especially on invasive capacity and the formation of metastasis. The effect of acidosis on metastasis formation of two rat carcinoma cell lines was studied in the animal model. In order to analyze the pH dependency of different steps of metastasis formation, invasiveness, cell adhesion and migration of AT-1 prostate cancer cells as well as possible underlying cell signaling pathways were studied in vitro. Acidosis significantly increased the formation of lung metastases of both tumor cell lines in vivo. In vitro, extracellular acidosis neither enhanced invasiveness nor affected cell adhesion to a plastic or to an endothelial layer. However, cellular motility was markedly elevated at pH 6.6 and this effect was sustained even when extracellular pH was switched back to pH 7.4. When analyzing the underlying mechanism, a prominent role of ROS in the induction of migration was observed. Signaling through the MAP kinases ERK1/2 and p38 as well as Src family kinases was not involved. Thus, cancer cells in an acidic microenvironment can acquire enhanced motility, which is sustained even if the tumor cells leave their acidic microenvironment e.g. by entering the blood stream. This increase depended on elevated ROS production and may contribute to the augmented formation of metastases of acidosis-primed tumor cells in vivo.

  9. Resveratrol and piperine enhance radiosensitivity of tumor cells

    Directory of Open Access Journals (Sweden)

    Jean Kyoung Tak


    Full Text Available The use of ionizing radiation (IR is essential for treating manyhuman cancers. However, radioresistance markedly impairsthe efficacy of tumor radiotherapy. IR enhances the productionof reactive oxygen species (ROS in a variety of cells which aredeterminant components in the induction of apoptosis. Muchinterest has developed to augment the effect of radiation in tumorsby combining it with radiosensitizers to improve the therapeuticratio. In the current study, the radiosensitizing effectsof resveratrol and piperine on cancer cells were evaluated.Cancer cell lines treated with these natural products exhibitedsignificantly augmented IR-induced apoptosis and loss of mitochondrialmembrane potential, presumably through enhancedROS generation. Applying natural products as sensitizersfor IR-induced apoptotic cell death offers a promisingtherapeutic approach to treat cancer. [BMB reports 2012;45(4: 242-246

  10. Nanoantennas for enhanced light trapping in transparent organic solar cells

    CERN Document Server

    Voroshilov, Pavel M; Belov, Pavel A


    We propose a light-trapping structure offering a significant enhancement of photovoltaic absorption in transparent organic solar cells operating at infrared while the visible light transmission keeps sufficiently high. The main mechanism of light trapping is related with the excitation of collective oscillations of the metal nanoantenna arrays, characterized by advantageous field distribution in the volume of the solar cell. It allows more than triple increase of infrared photovoltaic absorption.

  11. Light enhancements in nano-structured solar cells


    Pastorelli, Francesco


    In this century some of our main issues are energy shortage and pollution. This work will briefly describe these problems, proposing a plan of action combining energy saving and different sustainable energy sources. Within different types of renewable energy sources, solar energy is the most abundant one. To make solar energy a more sustainable and cost effective technology we focus on enhancing the optical characteristics of thin film solar cells. In this category, organic solar cells are go...

  12. Enhanced methanol utilization in direct methanol fuel cell (United States)

    Ren, Xiaoming; Gottesfeld, Shimshon


    The fuel utilization of a direct methanol fuel cell is enhanced for improved cell efficiency. Distribution plates at the anode and cathode of the fuel cell are configured to distribute reactants vertically and laterally uniformly over a catalyzed membrane surface of the fuel cell. A conductive sheet between the anode distribution plate and the anodic membrane surface forms a mass transport barrier to the methanol fuel that is large relative to a mass transport barrier for a gaseous hydrogen fuel cell. In a preferred embodiment, the distribution plate is a perforated corrugated sheet. The mass transport barrier may be conveniently increased by increasing the thickness of an anode conductive sheet adjacent the membrane surface of the fuel cell.

  13. Vascular endothelial growth factor enhances macrophage clearance of apoptotic cells (United States)

    Dalal, Samay; Horstmann, Sarah A.; Richens, Tiffany R.; Tanaka, Takeshi; Doe, Jenna M.; Boe, Darren M.; Voelkel, Norbert F.; Taraseviciene-Stewart, Laimute; Janssen, William J.; Lee, Chun G.; Elias, Jack A.; Bratton, Donna; Tuder, Rubin M.; Henson, Peter M.; Vandivier, R. William


    Efficient clearance of apoptotic cells from the lung by alveolar macrophages is important for the maintenance of tissue structure and function. Lung tissue from humans with emphysema contains increased numbers of apoptotic cells and decreased levels of vascular endothelial growth factor (VEGF). Mice treated with VEGF receptor inhibitors have increased numbers of apoptotic cells and develop emphysema. We hypothesized that VEGF regulates apoptotic cell clearance by alveolar macrophages (AM) via its interaction with VEGF receptor 1 (VEGF R1). Our data show that the uptake of apoptotic cells by murine AMs and human monocyte-derived macrophages is inhibited by depletion of VEGF and that VEGF activates Rac1. Antibody blockade or pharmacological inhibition of VEGF R1 activity also decreased apoptotic cell uptake ex vivo. Conversely, overexpression of VEGF significantly enhanced apoptotic cell uptake by AMs in vivo. These results indicate that VEGF serves a positive regulatory role via its interaction with VEGF R1 to activate Rac1 and enhance AM apoptotic cell clearance. PMID:22307908

  14. Solid oxide cell oxygen electrode with enhanced durability

    DEFF Research Database (Denmark)


    The present invention provides various methods of producing a solid oxide cell with an enhanced durability. Dopants are provided at the interface of the electrode comprising LSM and electrolyte and prevent especially the diffusion of Mn from the electrode layer into the electrolyte, stabilizing...

  15. Enhanced 3D fluorescence live cell imaging on nanoplasmonic substrate

    Energy Technology Data Exchange (ETDEWEB)

    Gartia, Manas Ranjan [Department of Nuclear, Plasma and Radiological Engineering, University of Illinois, Urbana, IL 61801 (United States); Hsiao, Austin; Logan Liu, G [Department of Bioengineering, University of Illinois, Urbana, IL 61801 (United States); Sivaguru, Mayandi [Institute for Genomic Biology, University of Illinois, Urbana, IL 61801 (United States); Chen Yi, E-mail: [Department of Electrical and Computer Engineering, University of Illinois, Urbana, IL 61801 (United States)


    We have created a randomly distributed nanocone substrate on silicon coated with silver for surface-plasmon-enhanced fluorescence detection and 3D cell imaging. Optical characterization of the nanocone substrate showed it can support several plasmonic modes (in the 300-800 nm wavelength range) that can be coupled to a fluorophore on the surface of the substrate, which gives rise to the enhanced fluorescence. Spectral analysis suggests that a nanocone substrate can create more excitons and shorter lifetime in the model fluorophore Rhodamine 6G (R6G) due to plasmon resonance energy transfer from the nanocone substrate to the nearby fluorophore. We observed three-dimensional fluorescence enhancement on our substrate shown from the confocal fluorescence imaging of chinese hamster ovary (CHO) cells grown on the substrate. The fluorescence intensity from the fluorophores bound on the cell membrane was amplified more than 100-fold as compared to that on a glass substrate. We believe that strong scattering within the nanostructured area coupled with random scattering inside the cell resulted in the observed three-dimensional enhancement in fluorescence with higher photostability on the substrate surface.

  16. Plasmonic versus dielectric enhancement in thin-film solar cells

    DEFF Research Database (Denmark)

    Dühring, Maria Bayard; Mortensen, N. Asger; Sigmund, Ole


    to its metallic counterpart. We show that the enhanced normalized short-circuit current for a cell with silicon strips can be increased 4 times compared to the best performance for strips of silver, gold, or aluminium. For this particular case, the simple dielectric grating may outperform its plasmonic...

  17. Prokaryotic arsenate reductase enhances arsenate resistance in Mammalian cells. (United States)

    Wu, Dan; Tao, Xuanyu; Wu, Gaofeng; Li, Xiangkai; Liu, Pu


    Arsenic is a well-known heavy metal toxicant in the environment. Bioremediation of heavy metals has been proposed as a low-cost and eco-friendly method. This article described some of recent patents on transgenic plants with enhanced heavy metal resistance. Further, to test whether genetic modification of mammalian cells could render higher arsenic resistance, a prokaryotic arsenic reductase gene arsC was transfected into human liver cancer cell HepG2. In the stably transfected cells, the expression level of arsC gene was determined by quantitative real-time PCR. Results showed that arsC was expressed in HepG2 cells and the expression was upregulated by 3 folds upon arsenate induction. To further test whether arsC has function in HepG2 cells, the viability of HepG2-pCI-ArsC cells exposed to arsenite or arsenate was compared to that of HepG2-pCI cells without arsC gene. The results indicated that arsC increased the viability of HepG2 cells by 25% in arsenate, but not in arsenite. And the test of reducing ability of stably transfected cells revealed that the concentration of accumulated trivalent arsenic increased by 25% in HepG2-pCI-ArsC cells. To determine the intracellular localization of ArsC, a fusion vector with fluorescent marker pEGFP-N1-ArsC was constructed and transfected into.HepG2. Laser confocal microscopy showed that EGFP-ArsC fusion protein was distributed throughout the cells. Taken together, these results demonstrated that prokaryotic arsenic resistant gene arsC integrated successfully into HepG2 genome and enhanced arsenate resistance of HepG2, which brought new insights of arsenic detoxification in mammalian cells.

  18. Functional Overload Enhances Satellite Cell Properties in Skeletal Muscle

    Directory of Open Access Journals (Sweden)

    Shin Fujimaki


    Full Text Available Skeletal muscle represents a plentiful and accessible source of adult stem cells. Skeletal-muscle-derived stem cells, termed satellite cells, play essential roles in postnatal growth, maintenance, repair, and regeneration of skeletal muscle. Although it is well known that the number of satellite cells increases following physical exercise, functional alterations in satellite cells such as proliferative capacity and differentiation efficiency following exercise and their molecular mechanisms remain unclear. Here, we found that functional overload, which is widely used to model resistance exercise, causes skeletal muscle hypertrophy and converts satellite cells from quiescent state to activated state. Our analysis showed that functional overload induces the expression of MyoD in satellite cells and enhances the proliferative capacity and differentiation potential of these cells. The changes in satellite cell properties coincided with the inactivation of Notch signaling and the activation of Wnt signaling and likely involve modulation by transcription factors of the Sox family. These results indicate the effects of resistance exercise on the regulation of satellite cells and provide insight into the molecular mechanism of satellite cell activation following physical exercise.

  19. Reprogramming of non-genomic estrogen signaling by the stemness factor SOX2 enhances the tumor-initiating capacity of breast cancer cells (United States)

    Vazquez-Martin, Alejandro; Cufí, Sílvia; López-Bonet, Eugeni; Corominas-Faja, Bruna; Cuyàs, Elisabet; Vellon, Luciano; Iglesias, Juan Manuel; Leis, Olatz; Martín, Ángel G; Menendez, Javier A


    The restoration of pluripotency circuits by the reactivation of endogenous stemness factors, such as SOX2, may provide a new paradigm in cancer development. The tumoral stem cell reprogramming hypothesis, i.e., the ability of stemness factors to redirect normal and differentiated tumor cells toward a less-differentiated and stem-like state, adds new layers of complexity to cancer biology, because the effects of such reprogramming may remain dormant until engaged later in response to (epi)genetic and/or (micro)environmental events. To test this hypothesis, we utilized an in vitro model of a SOX2-overexpressing cancer stem cell (CSC)-like cellular state that was recently developed in our laboratory by employing Yamanaka’s nuclear reprogramming technology in the estrogen receptor α (ERα)-positive MCF-7 breast cancer cell line. Despite the acquisition of distinct molecular features that were compatible with a breast CSC-like cellular state, such as strong aldehyde dehydrogenase activity, as detected by ALDEFLUOR, and overexpression of the SSEA-4 and CD44 breast CSC markers, the tumor growth-initiating ability of SOX2-overexpressing CSC-like MCF-7 cells solely occurred in female nude mice supplemented with estradiol when compared with MCF-7 parental cells. Ser118 phosphorylation of estrogen receptor α (ERα), which is a pivotal integrator of the genomic and nongenomic E2/ERα signaling pathways, drastically accumulated in nuclear speckles in the interphase nuclei of SOX2-driven CSC-like cell populations. Moreover, SOX2-positive CSC-like cells accumulated significantly higher numbers of actively dividing cells, and the highest levels of phospho-Ser118-ERα occurred when chromosomes lined up on a metaphase plate. The previously unrecognized link between E2/ERα signaling and SOX2-driven stem cell circuitry may significantly impact our current understanding of breast cancer initiation and progression, i.e., SOX2 can promote non-genomic E2 signaling that leads to

  20. Dynamic Enhanced Inter-Cell Interference Coordination for Realistic Networks

    DEFF Research Database (Denmark)

    Pedersen, Klaus I.; Alvarez, Beatriz Soret; Barcos, Sonia;


    Enhanced Inter-Cell Interference Coordination (eICIC) is a key ingredient to boost the performance of co-channel Heterogeneous Networks (HetNets). eICIC encompasses two main techniques: Almost Blank Subframes (ABS), during which the macro cell remains silent to reduce the interference, and biased...... and an opportunistic approach exploiting the varying cell conditions. Moreover, an autonomous fast distributed muting algorithm is presented, which is simple, robust, and well suited for irregular network deployments. Performance results for realistic network deployments show that the traditional semi-static e...

  1. Machine learning classification of cell-specific cardiac enhancers uncovers developmental subnetworks regulating progenitor cell division and cell fate specification


    Ahmad, Shaad M.; Busser, Brian W; Huang, Di; Cozart, Elizabeth J.; Michaud, Sébastien; Zhu, Xianmin; Jeffries, Neal; Aboukhalil, Anton; Bulyk, Martha L.; Ovcharenko, Ivan; Michelson, Alan M.


    The Drosophila heart is composed of two distinct cell types, the contractile cardial cells (CCs) and the surrounding non-muscle pericardial cells (PCs), development of which is regulated by a network of conserved signaling molecules and transcription factors (TFs). Here, we used machine learning with array-based chromatin immunoprecipitation (ChIP) data and TF sequence motifs to computationally classify cell type-specific cardiac enhancers. Extensive testing of predicted enhancers at single-c...

  2. Ouabain enhances ADPKD cell apoptosis via the intrinsic pathway

    Directory of Open Access Journals (Sweden)

    Gustavo eBlanco


    Full Text Available Progression of autosomal dominant polycystic kidney disease (ADPKD is highly influenced by factors circulating in blood. We have shown that the hormone ouabain enhances several characteristics of the ADPKD cystic phenotype, including the rate of cell proliferation, fluid secretion and the capacity of the cells to form cysts. In this work, we found that physiological levels of ouabain (3nM also promote programmed cell death of renal epithelial cells obtained from kidney cysts of patients with ADPKD (ADPKD cells. This was determined by Alexa Fluor 488 labeled-Annexin-V staining and TUNEL assay, both biochemical markers of apoptosis. Ouabain-induced apoptosis also takes place when ADPKD cell growth is blocked; suggesting that the effect is not secondary to the stimulatory actions of ouabain on cell proliferation. Ouabain alters the expression of BCL family of proteins, reducing BCL-2 and increasing BAX expression levels, anti- and pro-apoptotic mediators respectively. In addition, ouabain caused the release of cytochrome c from mitochondria. Moreover, ouabain activates caspase-3, a key executioner caspase in the cell apoptotic pathway, but did not affect caspase-8. This suggests that ouabain triggers ADPKD cell apoptosis by stimulating the intrinsic, but not the extrinsic pathway of programmed cell death. The apoptotic effects of ouabain are specific for ADPKD cells and do not occur in normal human kidney cells (NHK cells. Taken together with our previous observations, these results show that ouabain causes an imbalance in cell growth/death, to favor growth of the cystic cells. This event, characteristic of ADPKD, further suggests the importance of ouabain as a circulating factor that promotes ADPKD progression.

  3. Hypergravity Stimulation Enhances PC12 Neuron-Like Cell Differentiation

    Directory of Open Access Journals (Sweden)

    Giada Graziana Genchi


    Full Text Available Altered gravity is a strong physical cue able to elicit different cellular responses, representing a largely uninvestigated opportunity for tissue engineering/regenerative medicine applications. Our recent studies have shown that both proliferation and differentiation of C2C12 skeletal muscle cells can be enhanced by hypergravity treatment; given these results, PC12 neuron-like cells were chosen to test the hypothesis that hypergravity stimulation might also affect the behavior of neuronal cells, in particular promoting an enhanced differentiated phenotype. PC12 cells were thus cultured under differentiating conditions for either 12 h or 72 h before being stimulated with different values of hypergravity (50 g and 150 g. Effects of hypergravity were evaluated at transcriptional level 1 h and 48 h after the stimulation, and at protein level 48 h from hypergravity exposure, to assess its influence on neurite development over increasing differentiation times. PC12 differentiation resulted strongly affected by the hypergravity treatments; in particular, neurite length was significantly enhanced after exposure to high acceleration values. The achieved results suggest that hypergravity might induce a faster and higher neuronal differentiation and encourage further investigations on the potential of hypergravity in the preparation of cellular constructs for regenerative medicine and tissue engineering purposes.

  4. Interfacial geometry dictates cancer cell tumorigenicity (United States)

    Lee, Junmin; Abdeen, Amr A.; Wycislo, Kathryn L.; Fan, Timothy M.; Kilian, Kristopher A.


    Within the heterogeneous architecture of tumour tissue there exists an elusive population of stem-like cells that are implicated in both recurrence and metastasis. Here, by using engineered extracellular matrices, we show that geometric features at the perimeter of tumour tissue will prime a population of cells with a stem-cell-like phenotype. These cells show characteristics of cancer stem cells in vitro, as well as enhanced tumorigenicity in murine models of primary tumour growth and pulmonary metastases. We also show that interfacial geometry modulates cell shape, adhesion through integrin α5β1, MAPK and STAT activity, and initiation of pluripotency signalling. Our results for several human cancer cell lines suggest that interfacial geometry triggers a general mechanism for the regulation of cancer-cell state. Similar to how a growing tumour can co-opt normal soluble signalling pathways, our findings demonstrate how cancer can also exploit geometry to orchestrate oncogenesis.

  5. STAT3 signaling pathway is necessary for cell survival and tumorsphere forming capacity in ALDH{sup +}/CD133{sup +} stem cell-like human colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Li, E-mail: [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States); Division of Cardiology, Department of Internal Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030 (China); Fuchs, James; Li, Chenglong [Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, Columbus, OH 43210 (United States); Olson, Veronica [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States); Bekaii-Saab, Tanios [Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43210 (United States); Lin, Jiayuh, E-mail: [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States)


    Highlights: Black-Right-Pointing-Pointer The phosphorylated or activated form of STAT3 was expressed in colon cancer stem-like cells. Black-Right-Pointing-Pointer STAT3 inhibitor, FLLL32 inhibits P-STAT3 and STAT3 target genes in colon cancer stem-like cells. Black-Right-Pointing-Pointer Inhibition of STAT3 resulted in decreased cell viability and reduced numbers of tumorspheres. Black-Right-Pointing-Pointer STAT3 is required for survival and tumorsphere forming capacity in colon cancer stem-like cells. Black-Right-Pointing-Pointer Targeting STAT3 in cancer stem-like cells may offer a novel treatment approach for colon cancer. -- Abstract: Persistent activation of Signal Transducers and Activators of Transcription 3 (STAT3) is frequently detected in colon cancer. Increasing evidence suggests the existence of a small population of colon cancer stem or cancer-initiating cells may be responsible for tumor initiation, metastasis, and resistance to chemotherapy and radiation. Whether STAT3 plays a role in colon cancer-initiating cells and the effect of STAT3 inhibition is still unknown. Flow cytometry was used to isolate colon cancer stem-like cells from three independent human colon cancer cell lines characterized by both aldehyde dehydrogenase (ALDH)-positive and CD133-positive subpopulation (ALDH{sup +}/CD133{sup +}). The effects of STAT3 inhibition in colon cancer stem-like cells were examined. The phosphorylated or activated form of STAT3 was expressed in colon cancer stem-like cells and was reduced by a STAT3-selective small molecular inhibitor, FLLL32. FLLL32 also inhibited the expression of potential STAT3 downstream target genes in colon cancer stem-like cells including survivin, Bcl-XL, as well as Notch-1, -3, and -4, which may be involved in stem cell function. Furthermore, FLLL32 inhibited cell viability and tumorsphere formation as well as induced cleaved caspase-3 in colon cancer stem-like cells. FLLL32 is more potent than curcumin as evidenced with lower

  6. Flexible organic solar cells including efficiency enhancing grating structures

    DEFF Research Database (Denmark)

    Oliveira Hansen, Roana Melina de; Liu, Yinghui; Madsen, Morten


    enhancement. Since the solar cells avoid using brittle electrodes, the performance of the flexible devices is not affected by the peeling process. We have investigated three different nanostructured grating designs and conclude that gratings with a 500 nm pitch distance have the highest light......In this work, a new method for the fabrication of organic solar cells containing functional light-trapping nanostructures on flexible substrates is presented. Polyimide is spin-coated on silicon support substrates, enabling standard micro- and nanotechnology fabrication techniques......, such as photolithography and electron-beam lithography, besides the steps required for the bulk-heterojunction organic solar cell fabrication. After the production steps, the solar cells on polyimide are peeled off the silicon support substrates, resulting in flexible devices containing nanostructures for light absorption...

  7. Enhanced test methods to characterise automotive battery cells (United States)

    Mulder, Grietus; Omar, Noshin; Pauwels, Stijn; Leemans, Filip; Verbrugge, Bavo; De Nijs, Wouter; Van den Bossche, Peter; Six, Daan; Van Mierlo, Joeri

    This article evaluates the methods to characterise the behaviour of lithium ion cells of several chemistries and a nickel metal hydride cell for automotive applications like (plug-in) hybrid vehicles and battery electric vehicles. Although existing characterisation test methods are used, it was also indicated to combine test methods in order to speed up the test time and to create an improved comparability of the test results. Also, the existing capacity tests ignore that cells can be charged at several current rates. However, this is of interest for, e.g. fast charging and regenerative braking. Tests for high power and high energy application have been integrated in the enhanced method. The article explains the rationale to ameliorate the test methods. The test plan should make it possible to make an initial division in a group of cells purchased from several suppliers.

  8. Enzymes and other agents that enhance cell wall extensibility (United States)

    Cosgrove, D. J.


    Polysaccharides and proteins are secreted to the inner surface of the growing cell wall, where they assemble into a network that is mechanically strong, yet remains extensible until the cells cease growth. This review focuses on the agents that directly or indirectly enhance the extensibility properties of growing walls. The properties of expansins, endoglucanases, and xyloglucan transglycosylases are reviewed and their postulated roles in modulating wall extensibility are evaluated. A summary model for wall extension is presented, in which expansin is a primary agent of wall extension, whereas endoglucanases, xyloglucan endotransglycosylase, and other enzymes that alter wall structure act secondarily to modulate expansin action.

  9. Metal-air cell with performance enhancing additive

    Energy Technology Data Exchange (ETDEWEB)

    Friesen, Cody A; Buttry, Daniel


    Systems and methods drawn to an electrochemical cell comprising a low temperature ionic liquid comprising positive ions and negative ions and a performance enhancing additive added to the low temperature ionic liquid. The additive dissolves in the ionic liquid to form cations, which are coordinated with one or more negative ions forming ion complexes. The electrochemical cell also includes an air electrode configured to absorb and reduce oxygen. The ion complexes improve oxygen reduction thermodynamics and/or kinetics relative to the ionic liquid without the additive.

  10. Methamphetamine Enhances HIV-1 Infectivity in Monocyte Derived Dendritic Cells



    The US is currently experiencing an epidemic of methamphetamine (Meth) use as a recreational drug. Recent studies also show a high prevalence of HIV-1 infection among Meth users. We report that Meth enhances HIV-1 infectivity of dendritic cells as measured by multinuclear activation of a galactosidase indicator (MAGI) cell assay, p24 assay, and LTR-RU5 amplification. Meth induces increased HIV-1 infection in association with an increase in the HIV-1 coreceptors, CXCR4 and CCR5, and infection ...

  11. HES6 enhances the motility of alveolar rhabdomyosarcoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Wickramasinghe, Caroline M [MRC Cancer Cell Unit, Hutchison-MRC Research centre, Addenbrooke' s Hospital Cambridge, CB2 0XZ (United Kingdom); MRC Laboratory of Molecular Biology, Addenbrooke' s Hospital Cambridge, CB2 0QH (United Kingdom); Domaschenz, Renae [MRC Cancer Cell Unit, Hutchison-MRC Research centre, Addenbrooke' s Hospital Cambridge, CB2 0XZ (United Kingdom); Gene Regulation and Chromatin Group, MRC Clinical Sciences Centre, Faculty of Medicine, Imperial College, Hammersmith Campus, Du Cane Road, London W12 ONN (United Kingdom); Amagase, Yoko [MRC Cancer Cell Unit, Hutchison-MRC Research centre, Addenbrooke' s Hospital Cambridge, CB2 0XZ (United Kingdom); Department of Pathophysiology, Faculty of Pharmaceutical Sciences, Doshisha Women' s College of Liberal Arts, Kodo, Kyotanabe, Kyoto 610-0395 (Japan); Williamson, Daniel [Molecular Cytogenetics, The Institute of Cancer Research, Sutton SM2 5NG (United Kingdom); Northern Institute for Cancer Research, Paul O' Gorman Building, Medical School, Newcastle University, Framlington Place, Newcastle upon Tyne, NE2 4HH (United Kingdom); Missiaglia, Edoardo; Shipley, Janet [Molecular Cytogenetics, The Institute of Cancer Research, Sutton SM2 5NG (United Kingdom); Murai, Kasumi [MRC Cancer Cell Unit, Hutchison-MRC Research centre, Addenbrooke' s Hospital Cambridge, CB2 0XZ (United Kingdom); Jones, Philip H, E-mail: [MRC Cancer Cell Unit, Hutchison-MRC Research centre, Addenbrooke' s Hospital Cambridge, CB2 0XZ (United Kingdom)


    Absract: HES6, a member of the hairy-enhancer-of-split family of transcription factors, plays multiple roles in myogenesis. It is a direct target of the myogenic transcription factor MyoD and has been shown to regulate the formation of the myotome in development, myoblast cell cycle exit and the organization of the actin cytoskeleton during terminal differentiation. Here we investigate the expression and function of HES6 in rhabdomyosarcoma, a soft tissue tumor which expresses myogenic genes but fails to differentiate into muscle. We show that HES6 is expressed at high levels in the subset of alveolar rhabdomyosarcomas expressing PAX/FOXO1 fusion genes (ARMSp). Knockdown of HES6 mRNA in the ARMSp cell line RH30 reduces proliferation and cell motility. This phenotype is rescued by expression of mouse Hes6 which is insensitive to HES6 siRNA. Furthermore, expression microarray analysis indicates that the HES6 knockdown is associated with a decrease in the levels of Transgelin, (TAGLN), a regulator of the actin cytoskeleton. Knockdown of TAGLN decreases cell motility, whilst TAGLN overexpression rescues the motility defect resulting from HES6 knockdown. These findings indicate HES6 contributes to the pathogenesis of ARMSp by enhancing both proliferation and cell motility.

  12. Gold Nanoparticles-Enhanced Proton Exchange Membrane (PEM) Fuel Cell (United States)

    Li, Hongfei; Pan, Cheng; Liu, Ping; Zhu, Yimei; Adzic, Radoslav; Rafailovich, Miriam

    Proton exchange membrane fuel cells have drawn great attention and been taken as a promising alternated energy source. One of the reasons hamper the wider application of PEM fuel cell is the catalytic poison effect from the impurity of the gas flow. Haruta has predicted that gold nanoparticles that are platelet shaped and have direct contact with the metal oxide substrate to be the perfect catalysts of the CO oxidization, yet the synthesis method is difficult to apply in the Fuel Cell. In our approach, thiol-functionalized gold nanoparticles were synthesized through two-phase method developed by Brust et al. We deposit these Au particles with stepped surface directly onto the Nafion membrane in the PEM fuel cell by Langmuir-Blodgett method, resulting in over 50% enhancement of the efficiency of the fuel cell. DFT calculations were conducted to understand the theory of this kind of enhancement. The results indicated that only when the particles were in direct surface contact with the membrane, where AuNPs attached at the end of the Nafion side chains, it could reduce the energy barrier for the CO oxidation that could happen at T<300K.

  13. Flexible organic solar cells including efficiency enhancing grating structures (United States)

    Melina de Oliveira Hansen, Roana; Liu, Yinghui; Madsen, Morten; Rubahn, Horst-Günter


    In this work, a new method for the fabrication of organic solar cells containing functional light-trapping nanostructures on flexible substrates is presented. Polyimide is spin-coated on silicon support substrates, enabling standard micro- and nanotechnology fabrication techniques, such as photolithography and electron-beam lithography, besides the steps required for the bulk-heterojunction organic solar cell fabrication. After the production steps, the solar cells on polyimide are peeled off the silicon support substrates, resulting in flexible devices containing nanostructures for light absorption enhancement. Since the solar cells avoid using brittle electrodes, the performance of the flexible devices is not affected by the peeling process. We have investigated three different nanostructured grating designs and conclude that gratings with a 500 nm pitch distance have the highest light-trapping efficiency for the selected active layer material (P3HT:PCBM), resulting in an enhancement of about 34% on the solar cell efficiency. The presented method can be applied to a large variety of flexible nanostructured devices in future applications.

  14. Siah1 proteins enhance radiosensitivity of human breast cancer cells

    Directory of Open Access Journals (Sweden)

    Engenhart-Cabillic Rita


    Full Text Available Abstract Background Siah proteins play an important role in cancer progression. We evaluated the effect of Siah1, its splice variants Siah1L and the Siah1 mutant with the RING finger deleted (Siah1ΔR on radiosensitization of human breast cancer cells. Methods The status of Siah1 and Siah1L was analysed in five breast cancer cell lines. To establish stable cells, SKBR3 cells were transfected with Siah1, Siah-1L and Siah1ΔR. Siah1 function was suppressed by siRNA in MCF-7 cells. The impact of Siah1 overexpression and silencing on apoptosis, proliferation, survival, invasion ability and DNA repair was assessed in SKBR3 and MCF-7 cells, also in regards to radiation. Results Siah1 and Siah1L mRNA expression was absent in four of five breast cancer cells lines analysed. Overexpression of Siah1 and Siah1L enhanced radiation-induced apoptosis in stable transfected SKBR3 cells, while Siah1ΔR failed to show this effect. In addition, Siah1 and Siah1L significantly reduced cell clonogenic survival and proliferation. Siah1L sensitization enhancement ratio values were over 1.5 and 4.0 for clonogenic survival and proliferation, respectively, pointing to a highly cooperative and potentially synergistic fashion with radiation. Siah1 or Siah1L significantly reduced invasion ability of SKBR3 and suppressed Tcf/Lef factor activity. Importantly, Siah1 siRNA demonstrated opposite effects in MCF-7 cells. Siah1 and Siah1L overexpression resulted in inhibition of DNA repair as inferred by increased levels of DNA double-strand breaks in irradiated SKBR3 cells. Conclusion Our results reveal for the first time how overexpression of Siah1L and Siah1 can determine radiosensitivity of breast cancer cells. These findings suggest that development of drugs augmenting Siah1 and Siah1L activity could be a novel approach in improving tumor cell kill.

  15. Lithographic antennas for enhancement of solar cell efficiency

    Energy Technology Data Exchange (ETDEWEB)

    Kotter, D.K. [Lockheed Martin Idaho Technologies Co., Idaho Falls, ID (United States); Boreman, G. [Univ. of Central Florida, Orlando, FL (United States). Center for Research and Education in Optics and Lasers


    This report documents proof-of-concept demonstration of the use of lithographic antennas for enhancement of solar-cell efficiency. A micro-sized lithographic antenna array was theoretically modeled, designed and fabricated. Experimental research was performed to validate the ability of the antenna array to concentrate infrared and visible energy onto photovoltaic (PV) materials. The research will serve as the basis for the design of a miniature power source for remote sensors.

  16. Radiosensitivity of human colon cancer cell enhanced by immunoliposomal docetaxel

    Institute of Scientific and Technical Information of China (English)

    Qing-Wei Wang; Hui-Lan Lü; Chang-Cheng Song; Hong Liu; Cong-Gao Xu


    AIM: To enhance the radiosensitivity of human colon cancer cells by docetaxel.METHODS: Immunoliposomal docetaxel was prepared by coupling monodonal antibody against carcinoembryonic antigen to cyanuric chloride at the PEG terminus of liposome. LoVo adenocarcinoma cell line was treated with immunoliposomal docetaxel or/and irradiation. MTT colorimetric assay was used to estimate cytotoxicity of immunoliposomal docetaxel and radiotoxicity. Cell cycle redistribution and apoptosis were determined with flow cytometry. Survivin expression in LoVo cells was verified by immunohistochemistry. D801 morphologic analysis system was used to semi-quantify immunohistochemical staining of survivin.RESULTS: Cytotoxicity was induced by immunoliposomal docetaxel alone in a dose-dependent manner. Immunoliposomal docetaxel yielded a cytotoxicity effect at a low dose of 2 nmol/L. With a single dose irradiation, the relative surviving fraction of LoVo cells showed a dosedependent response, but there were no significant changes as radiation delivered from 4 to 8 Gy. Compared with liposomal docetaxel or single dose irradiation,strongly radiopotentiating effects of immunoliposomal docetaxel on LoVo cells were observed. A low dose of immunoliposomal docetaxel could yield sufficient radiosensitivity. Immunoliposomal docetaxel were achieved both specificity of the conjugated antibody and drug radiosensitization. Combined with radiation,immunoliposomal docetaxel significantly increased the percentage of G2/M cells and induced apoptosis, but significantly decreased the percentage of cells in G2/G1 and S phase by comparison with liposomal docetaxel.Immunohistochemical analysis showed that the brown stained survivin was mainly in cytoplasm of LoVo cells.Semi-quantitative analysis of the survivin immunostaining showed that the expression of survivin in LoVo cells under irradiation with immunoliposomal docetaxel was significantly decreased.CONCLUSION: Immunoliposomal docetaxel is strongly effective

  17. Distinct population of highly malignant cells in a head and neck squamous cell carcinoma cell line established by xenograft model

    Directory of Open Access Journals (Sweden)

    Jan Chia-Ing


    Full Text Available Abstract The progression and metastasis of solid tumors, including head and neck squamous cell carcinoma (HNSCC, have been related to the behavior of a small subpopulation of cancer stem cells. Here, we have established a highly malignant HNSCC cell line, SASVO3, from primary tumors using three sequential rounds of xenotransplantation. SASVO3 possesses enhanced tumorigenic ability both in vitro and in vivo. Moreover, SASVO3 exhibits properties of cancer stem cells, including that increased the abilities of sphere-forming, the number of side population cells, the potential of transplanted tumor growth and elevated expression of the stem cell marker Bmi1. Injection of SASVO3 into the tail vein of nude mice resulted in lung metastases. These results are consistent with the postulate that the malignant and/or metastasis potential of HNSCC cells may reside in a stem-like subpopulation.

  18. Dissection of thousands of cell type-specific enhancers identifies dinucleotide repeat motifs as general enhancer features. (United States)

    Yáñez-Cuna, J Omar; Arnold, Cosmas D; Stampfel, Gerald; Boryń, Lukasz M; Gerlach, Daniel; Rath, Martina; Stark, Alexander


    Gene expression is determined by genomic elements called enhancers, which contain short motifs bound by different transcription factors (TFs). However, how enhancer sequences and TF motifs relate to enhancer activity is unknown, and general sequence requirements for enhancers or comprehensive sets of important enhancer sequence elements have remained elusive. Here, we computationally dissect thousands of functional enhancer sequences from three different Drosophila cell lines. We find that the enhancers display distinct cis-regulatory sequence signatures, which are predictive of the enhancers' cell type-specific or broad activities. These signatures contain transcription factor motifs and a novel class of enhancer sequence elements, dinucleotide repeat motifs (DRMs). DRMs are highly enriched in enhancers, particularly in enhancers that are broadly active across different cell types. We experimentally validate the importance of the identified TF motifs and DRMs for enhancer function and show that they can be sufficient to create an active enhancer de novo from a nonfunctional sequence. The function of DRMs as a novel class of general enhancer features that are also enriched in human regulatory regions might explain their implication in several diseases and provides important insights into gene regulation.

  19. Covalent immobilization of p-selectin enhances cell rolling. (United States)

    Hong, Seungpyo; Lee, Dooyoung; Zhang, Huanan; Zhang, Jennifer Q; Resvick, Jennifer N; Khademhosseini, Ali; King, Michael R; Langer, Robert; Karp, Jeffrey M


    Cell rolling is an important physiological and pathological process that is used to recruit specific cells in the bloodstream to a target tissue. This process may be exploited for biomedical applications to capture and separate specific cell types. One of the most commonly studied proteins that regulate cell rolling is P-selectin. By coating surfaces with this protein, biofunctional surfaces that induce cell rolling can be prepared. Although most immobilization methods have relied on physisorption, chemical immobilization has obvious advantages, including longer functional stability and better control over ligand density and orientation. Here we describe chemical methods to immobilize P-selectin covalently on glass substrates. The chemistry was categorized on the basis of the functional groups on modified glass substrates: amine, aldehyde, and epoxy. The prepared surfaces were first tested in a flow chamber by flowing microspheres functionalized with a cell surface carbohydrate (sialyl Lewis(x)) that binds to P-selectin. Adhesion bonds between P-selectin and sialyl Lewis(x) dissociate readily under shear forces, leading to cell rolling. P-selectin immobilized on the epoxy glass surfaces exhibited enhanced long-term stability of the function and better homogeneity as compared to that for surfaces prepared by other methods and physisorbed controls. The microsphere rolling results were confirmed in vitro with isolated human neutrophils. This work is essential for the future development of devices for isolating specific cell types based on cell rolling, which may be useful for hematologic cancers and certain metastatic cancer cells that are responsive to immobilized selectins.

  20. Tumorigenic lung tumorospheres exhibit stem-like features with significantly increased expression of CD133 and ABCG2


    Zhao, Wensi; Luo, Yi; Li, Boyi; Zhang, Tao


    Accumulating evidence supports the existence of cancer stem cells (CSCs) in human tumors, and the successful certification of CSCs may lead to the identification of therapeutic targets, which are more effective for the treatment of cancer. The use of spherical cancer models has increased in popularity in cancer stem cell investigations. Tumorospheres, which are used as a model of CSCs and are established in serum-free medium supplemented with growth factors under non-adherent conditions, are ...

  1. Absorption enhancement by textured InP in solar cells (United States)

    Yun, Seokhun; Ji, Taeksoo


    III-V compound semiconductors seem to be the ideal materials for photovoltaic devices because they exhibit fast carrier velocity. III-V compound semiconductors, however, are unfavorable materials to be commercialized on large scale photovoltaic devices because of their high material cost. The textured surface shows the potential to increase the performance of solar cells because of the properties such as high absorption and longer light path length. These properties can overcome the disadvantage of the III-V compound semiconductors through thin thickness use when producing solar cells. In this study, we demonstrate that textured surfaces on InP formed by nano-sphere lithography and plasma etching process can enhance the absorption effectively in comparison with planar surface. The power conversion efficiency of InP solar cells using the textured InP and the aluminum doped zinc oxide was achieved up to 8%.

  2. Enhancement of photovoltaic cell performance using periodic triangular gratings (United States)

    Bordatchev, Evgueni; Tauhiduzzaman, Mohammed; Dey, Rajat


    The solar energy industry strives to produce more efficient and yet cost effective solar panels each consisting of an array of photovoltaic (PV) cells. The goal of this study was to enhance the performance of PV cells through increasing the cells' optical efficiency defined as a percentage of surface incident light that reaches the PV material. This was achieved through the reduction of waveguide decoupling loss and Fresnel reflection losses by integrating specific nonimaging micro-optical structures on the top surface of existing PV cells. Due to this integration, optical efficiency and performance were increased through the enhancement of light trapping, light guiding, and in-coupling functionalities. Periodic triangular gratings (PTGs) were designed, nonsequentially modeled, optimized, and fabricated in polydimethylsiloxane as proposed micro-optical structures. Then the performance of PV cells with and without integrated PTGs was evaluated and compared. Initial optical simulation results show that an original PV cell (without PTG) exhibits an average optical efficiency of 32.7% over a range of incident light angles between 15 and 90 deg. Integration of the PTG allows the capture of incoming sunlight by total internal reflection (TIR), whence it is reflected back onto the PV cell for multiple consecutive chances for absorption and PV conversion. Geometry of the PTG was optimized with respect to an angle of light incidence of {15, 30, 45, 60, 75, 90} deg. Optical efficiency of the geometrically optimized PTGs was then analyzed under the same set of incident light angles and a maximum optical efficiency of 54.1% was observed for a PV cell with integrated PTG optimized at 90 deg. This is a 53.3% relative improvement in optical performance when compared to an original PV cell. Functional PTG prototypes were then fabricated with optical surface quality (below 10 nm Ra) and integrated with PV cells demonstrating an increase in maximum power by 1.08 mW/cm (7

  3. Plasma-Sprayed Titanium Patterns for Enhancing Early Cell Responses (United States)

    Shi, Yunqi; Xie, Youtao; Pan, Houhua; Zheng, Xuebin; Huang, Liping; Ji, Fang; Li, Kai


    Titanium coating has been widely used as a biocompatible metal in biomedical applications. However, the early cell responses and long-term fixation of titanium implants are not satisfied. To obviate these defects, in this paper, micro-post arrays with various widths (150-1000 μm) and intervals (100-300 μm) were fabricated on the titanium substrate by template-assisted plasma spraying technology. In vitro cell culture experiments showed that MC3T3-E1 cells exhibited significantly higher osteogenic differentiation as well as slightly improved adhesion and proliferation on the micro-patterned coatings compared with the traditional one. The cell number on the pattern with 1000 µm width reached 130% after 6 days of incubation, and the expressions of osteopontin (OPN) as well as osteocalcin (OC) were doubled. No obvious difference was found in cell adhesion on various size patterns. The present micro-patterned coatings proposed a new modification method for the traditional plasma spraying technology to enhance the early cell responses and convenience for the bone in-growth.

  4. Pan-Bcl-2 inhibitor AT-101 enhances tumor cell killing by EGFR targeted T cells.

    Directory of Open Access Journals (Sweden)

    Archana Thakur

    Full Text Available Pancreatic cancer is a deadly disease and has the worst prognosis among almost all cancers and is in dire need of new and improved therapeutic strategies. Conditioning of tumor cells with chemotherapeutic drug has been shown to enhance the anti-tumor effects of cancer vaccines and adoptive cell therapy. In this study, we investigated the immunomodulatory effects of pan-Bcl-2 inhibitor AT-101 on pancreatic cancer (PC cell cytotoxicity by activated T cells (ATC. The effects of AT-101 on cytotoxicity, early apoptosis, and Granzyme B (GrzB and IFN-γ signaling pathways were evaluated during EGFR bispecific antibody armed ATC (aATC-mediated killing of L3.6pl and MiaPaCa-2 PC cells pre-sensitized with AT-101. We found that pretreatment of tumor cells with AT-101 enhanced susceptibility of L3.6pl and MiaPaCa-2 tumor cells to ATC and aATC-mediated cytotoxicity, which was in part mediated via enhanced release of cytolytic granule GrzB from ATC and aATC. AT-101-sensitized L3.6pl cells showed up-regulation of IFN-γ-mediated induction in the phosphorylation of Ser(727-Stat1 (pS(727-Stat1, and IFN-γ induced dephosphorylation of phospho-Tyr(705-Stat3 (pY(705-Stat3. Priming (conditioning of PC cells with AT-101 can significantly enhance the anti-tumor activity of EGFRBi armed ATC through increased IFN-γ induced activation of pS(727-Stat1 and inhibition of pY(705-Stat3 phosphorylation, and resulting in increased ratio of pro-apoptotic to anti-apoptotic proteins. Our results verify enhanced cytotoxicity after a novel chemotherapy conditioning strategy against PC that warrants further in vivo and clinical investigations.

  5. Enhancement of cell radiation sensitivity by pegylated gold nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Liu, C-J; Wang, C-H; Chen, S-T; Chen, H-H; Leng, W-H; Chien, C-C; Wang, C-L; Kempson, Ivan M; Hwu, Y [Institute of Physics, Academia Sinica, Nankang, Taipei 11529, Taiwan (China); Lai, T-C; Hsiao, Michael [Genomics Research Center, Academia Sinica, Nankang, Taipei 11529, Taiwan (China); Yang, C-S [Center for Nanomedicine, National Health Research Institutes, Miaoli 35053, Taiwan (China); Chen, Y-J [Mackay Memorial Hospital, Taipei 10449, Taiwan (China); Margaritondo, G [Ecole Polytechnique Federale de Lausanne (EPFL), CH-1015 Lausanne (Switzerland)], E-mail:


    Biocompatible Au nanoparticles with surfaces modified by PEG (polyethylene glycol) were developed in view of possible applications for the enhancement of radiotherapy. Such nanoparticles exhibit preferential deposition at tumor sites due to the enhanced permeation and retention (EPR) effect. Here, we systematically studied their effects on EMT-6 and CT26 cell survival rates during irradiation for a dose up to 10 Gy with a commercial biological irradiator (E{sub average} = 73 keV), a Cu-K{alpha}{sub 1} x-ray source (8.048 keV), a monochromatized synchrotron source (6.5 keV), a radio-oncology linear accelerator (6 MeV) and a proton source (3 MeV). The percentage of surviving cells after irradiation was found to decrease by {approx}2-45% in the presence of PEG-Au nanoparticles ([Au] = 400, 500 or 1000 {mu}M). The cell survival rates decreased as a function of the dose for all sources and nanoparticle concentrations. These results could open the way to more effective cancer irradiation therapies by using nanoparticles with optimized surface treatment. Difficulties in applying MTT assays were also brought to light, showing that this approach is not suitable for radiobiology.

  6. Aligned-Braided Nanofibrillar Scaffold with Endothelial Cells Enhances Arteriogenesis. (United States)

    Nakayama, Karina H; Hong, Guosong; Lee, Jerry C; Patel, Jay; Edwards, Bryan; Zaitseva, Tatiana S; Paukshto, Michael V; Dai, Hongjie; Cooke, John P; Woo, Y Joseph; Huang, Ngan F


    The objective of this study was to enhance the angiogenic capacity of endothelial cells (ECs) using nanoscale signaling cues from aligned nanofibrillar scaffolds in the setting of tissue ischemia. Thread-like nanofibrillar scaffolds with porous structure were fabricated from aligned-braided membranes generated under shear from liquid crystal collagen solution. Human ECs showed greater outgrowth from aligned scaffolds than from nonpatterned scaffolds. Integrin α1 was in part responsible for the enhanced cellular outgrowth on aligned nanofibrillar scaffolds, as the effect was abrogated by integrin α1 inhibition. To test the efficacy of EC-seeded aligned nanofibrillar scaffolds in improving neovascularization in vivo, the ischemic limbs of mice were treated with EC-seeded aligned nanofibrillar scaffold; EC-seeded nonpatterned scaffold; ECs in saline; aligned nanofibrillar scaffold alone; or no treatment. After 14 days, laser Doppler blood spectroscopy demonstrated significant improvement in blood perfusion recovery when treated with EC-seeded aligned nanofibrillar scaffolds, in comparison to ECs in saline or no treatment. In ischemic hindlimbs treated with scaffolds seeded with human ECs derived from induced pluripotent stem cells (iPSC-ECs), single-walled carbon nanotube (SWNT) fluorophores were systemically delivered to quantify microvascular density after 28 days. Near infrared-II (NIR-II, 1000-1700 nm) imaging of SWNT fluorophores demonstrated that iPSC-EC-seeded aligned scaffolds group showed significantly higher microvascular density than the saline or cells groups. These data suggest that treatment with EC-seeded aligned nanofibrillar scaffolds improved blood perfusion and arteriogenesis, when compared to treatment with cells alone or scaffold alone, and have important implications in the design of therapeutic cell delivery strategies.

  7. Blockade of Glutamine Synthetase Enhances Inflammatory Response in Microglial Cells (United States)

    Palmieri, Erika M.; Menga, Alessio; Lebrun, Aurore; Hooper, Douglas C.; Butterfield, D. Allan


    Abstract Aims: Microglial cells are brain-resident macrophages engaged in surveillance and maintained in a constant state of relative inactivity. However, their involvement in autoimmune diseases indicates that in pathological conditions microglia gain an inflammatory phenotype. The mechanisms underlying this change in the microglial phenotype are still unclear. Since metabolism is an important modulator of immune cell function, we focused our attention on glutamine synthetase (GS), a modulator of the response to lipopolysaccharide (LPS) activation in other cell types, which is expressed by microglia. Results: GS inhibition enhances release of inflammatory mediators of LPS-activated microglia in vitro, leading to perturbation of the redox balance and decreased viability of cocultured neurons. GS inhibition also decreases insulin-mediated glucose uptake in microglia. In vivo, microglia-specific GS ablation enhances expression of inflammatory markers upon LPS treatment. In the spinal cords from experimental autoimmune encephalomyelitis (EAE), GS expression levels and glutamine/glutamate ratios are reduced. Innovation: Recently, metabolism has been highlighted as mediator of immune cell function through the discovery of mechanisms that (behind these metabolic changes) modulate the inflammatory response. The present study shows for the first time a metabolic mechanism mediating microglial response to a proinflammatory stimulus, pointing to GS activity as a master modulator of immune cell function and thus unraveling a potential therapeutic target. Conclusions: Our study highlights a new role of GS in modulating immune response in microglia, providing insights into the pathogenic mechanisms associated with inflammation and new strategies of therapeutic intervention. Antioxid. Redox Signal. 26, 351–363. PMID:27758118

  8. Machine learning classification of cell-specific cardiac enhancers uncovers developmental subnetworks regulating progenitor cell division and cell fate specification. (United States)

    Ahmad, Shaad M; Busser, Brian W; Huang, Di; Cozart, Elizabeth J; Michaud, Sébastien; Zhu, Xianmin; Jeffries, Neal; Aboukhalil, Anton; Bulyk, Martha L; Ovcharenko, Ivan; Michelson, Alan M


    The Drosophila heart is composed of two distinct cell types, the contractile cardial cells (CCs) and the surrounding non-muscle pericardial cells (PCs), development of which is regulated by a network of conserved signaling molecules and transcription factors (TFs). Here, we used machine learning with array-based chromatin immunoprecipitation (ChIP) data and TF sequence motifs to computationally classify cell type-specific cardiac enhancers. Extensive testing of predicted enhancers at single-cell resolution revealed the added value of ChIP data for modeling cell type-specific activities. Furthermore, clustering the top-scoring classifier sequence features identified novel cardiac and cell type-specific regulatory motifs. For example, we found that the Myb motif learned by the classifier is crucial for CC activity, and the Myb TF acts in concert with two forkhead domain TFs and Polo kinase to regulate cardiac progenitor cell divisions. In addition, differential motif enrichment and cis-trans genetic studies revealed that the Notch signaling pathway TF Suppressor of Hairless [Su(H)] discriminates PC from CC enhancer activities. Collectively, these studies elucidate molecular pathways used in the regulatory decisions for proliferation and differentiation of cardiac progenitor cells, implicate Su(H) in regulating cell fate decisions of these progenitors, and document the utility of enhancer modeling in uncovering developmental regulatory subnetworks.

  9. Cell-to-cell contact of human monocytes with infected arterial smooth-muscle cells enhances growth of Chlamydia pneumoniae. (United States)

    Puolakkainen, Mirja; Campbell, Lee Ann; Lin, Tsun-Mei; Richards, Theresa; Patton, Dorothy L; Kuo, Cho-Chou


    Chlamydia pneumoniae can infect arterial cells. It has been shown that coculture of human monocytes (U937) and endothelial cells promotes infection of C. pneumoniae in endothelial cells and that the enhancement was mediated by a soluble factor (insulin-like growth factor 2) secreted by monocytes. In this study, it is shown that coculture of monocytes with C. pneumoniae enhances infection of C. pneumoniae in arterial smooth-muscle cells 5.3-fold at a monocyte-to-smooth-muscle cell ratio of 5. However, unlike endothelial cells, no enhancement was observed if monocytes were placed in cell culture inserts or if conditioned medium from monocyte cultures was used, which suggests that cell-to-cell contact is critical. The addition of mannose 6-phosphate or octyl glucoside, a nonionic detergent containing a sugar group, to cocultures inhibited the enhancement. These findings suggest that the monocyte-smooth-muscle cell interaction may be mediated by mannose 6-phosphate receptors present on monocytes.

  10. Clarithromycin Synergistically Enhances Thalidomide Cytotoxicity in Myeloma Cells. (United States)

    Qiu, Xu-Hua; Shao, Jing-Jing; Mei, Jian-Gang; Li, Han-Qing; Cao, Hong-Qin


    Clarithromycin (CAM) is a macrolide antibiotic that is widely used in the treatment of respiratory tract infections, sexually transmitted diseases and infections caused by the Helicobacter pylori and Mycobacterium avium complex. Recent studies showed that CAM was highly effective against multiple myeloma (MM) when used in combination with immunomodulatory drugs and dexamethasone. However, the related mechanism is still unknown. As 3 immunomodulatory agents are all effective in the respective regimen, we postulated that CAM might enhance the effect of immunomodulatory drugs. We evaluated the interaction effects of CAM and thalidomide on myeloma cells. Taking into consideration that thalidomide did not affect the proliferation of myeloma cells in vitro, we cocultured myeloma cells with peripheral blood monocytes and evaluated the effects of CAM and thalidomide on the cocultured cell model. Data showed that thalidomide and CAM synergistically inhibited the proliferation of the cells. On this same model, we also found that thalidomide and CAM synergistically decreased the secretion of tumor necrosis factor-α and interleukin-6. This might be caused by the effect of the 2 drugs on inhibiting the activation of ERK1/2 and AKT. These data suggest that the efficacy of CAM against MM was partly due to its synergistic action with the immunomodulatory agents.

  11. TGF-β inhibition enhances chemotherapy action against triple-negative breast cancer. (United States)

    Bhola, Neil E; Balko, Justin M; Dugger, Teresa C; Kuba, María Gabriela; Sánchez, Violeta; Sanders, Melinda; Stanford, Jamie; Cook, Rebecca S; Arteaga, Carlos L


    After an initial response to chemotherapy, many patients with triple-negative breast cancer (TNBC) have recurrence of drug-resistant metastatic disease. Studies with TNBC cells suggest that chemotherapy-resistant populations of cancer stem-like cells (CSCs) with self-renewing and tumor-initiating capacities are responsible for these relapses. TGF-β has been shown to increase stem-like properties in human breast cancer cells. We analyzed RNA expression in matched pairs of primary breast cancer biopsies before and after chemotherapy. Biopsies after chemotherapy displayed increased RNA transcripts of genes associated with CSCs and TGF-β signaling. In TNBC cell lines and mouse xenografts, the chemotherapeutic drug paclitaxel increased autocrine TGF-β signaling and IL-8 expression and enriched for CSCs, as indicated by mammosphere formation and CSC markers. The TGF-β type I receptor kinase inhibitor LY2157299, a neutralizing TGF-β type II receptor antibody, and SMAD4 siRNA all blocked paclitaxel-induced IL8 transcription and CSC expansion. Moreover, treatment of TNBC xenografts with LY2157299 prevented reestablishment of tumors after paclitaxel treatment. These data suggest that chemotherapy-induced TGF-β signaling enhances tumor recurrence through IL-8-dependent expansion of CSCs and that TGF-β pathway inhibitors prevent the development of drug-resistant CSCs. These findings support testing a combination of TGF-β inhibitors and anticancer chemotherapy in patients with TNBC.

  12. TGF-β inhibition enhances chemotherapy action against triple-negative breast cancer (United States)

    Bhola, Neil E.; Balko, Justin M.; Dugger, Teresa C.; Kuba, María Gabriela; Sánchez, Violeta; Sanders, Melinda; Stanford, Jamie; Cook, Rebecca S.; Arteaga, Carlos L.


    After an initial response to chemotherapy, many patients with triple-negative breast cancer (TNBC) have recurrence of drug-resistant metastatic disease. Studies with TNBC cells suggest that chemotherapy-resistant populations of cancer stem-like cells (CSCs) with self-renewing and tumor-initiating capacities are responsible for these relapses. TGF-β has been shown to increase stem-like properties in human breast cancer cells. We analyzed RNA expression in matched pairs of primary breast cancer biopsies before and after chemotherapy. Biopsies after chemotherapy displayed increased RNA transcripts of genes associated with CSCs and TGF-β signaling. In TNBC cell lines and mouse xenografts, the chemotherapeutic drug paclitaxel increased autocrine TGF-β signaling and IL-8 expression and enriched for CSCs, as indicated by mammosphere formation and CSC markers. The TGF-β type I receptor kinase inhibitor LY2157299, a neutralizing TGF-β type II receptor antibody, and SMAD4 siRNA all blocked paclitaxel-induced IL8 transcription and CSC expansion. Moreover, treatment of TNBC xenografts with LY2157299 prevented reestablishment of tumors after paclitaxel treatment. These data suggest that chemotherapy-induced TGF-β signaling enhances tumor recurrence through IL-8–dependent expansion of CSCs and that TGF-β pathway inhibitors prevent the development of drug-resistant CSCs. These findings support testing a combination of TGF-β inhibitors and anticancer chemotherapy in patients with TNBC. PMID:23391723

  13. Hypoxic enhancement of exosome release by breast cancer cells

    Directory of Open Access Journals (Sweden)

    King Hamish W


    Full Text Available Abstract Background Exosomes are nanovesicles secreted by tumour cells which have roles in paracrine signalling during tumour progression, including tumour-stromal interactions, activation of proliferative pathways and bestowing immunosuppression. Hypoxia is an important feature of solid tumours which promotes tumour progression, angiogenesis and metastasis, potentially through exosome-mediated signalling. Methods Breast cancer cell lines were cultured under either moderate (1% O2 or severe (0.1% O2 hypoxia. Exosomes were isolated from conditioned media and quantitated by nanoparticle tracking analysis (NTA and immunoblotting for the exosomal protein CD63 in order to assess the impact of hypoxia on exosome release. Hypoxic exosome fractions were assayed for miR-210 by real-time reverse transcription polymerase chain reaction and normalised to exogenous and endogenous control genes. Statistical significance was determined using the Student T test with a P value of  Results Exposure of three different breast cancer cell lines to moderate (1% O2 and severe (0.1% O2 hypoxia resulted in significant increases in the number of exosomes present in the conditioned media as determined by NTA and CD63 immunoblotting. Activation of hypoxic signalling by dimethyloxalylglycine, a hypoxia-inducible factor (HIF hydroxylase inhibitor, resulted in significant increase in exosome release. Transfection of cells with HIF-1α siRNA prior to hypoxic exposure prevented the enhancement of exosome release by hypoxia. The hypoxically regulated miR-210 was identified to be present at elevated levels in hypoxic exosome fractions. Conclusions These data provide evidence that hypoxia promotes the release of exosomes by breast cancer cells, and that this hypoxic response may be mediated by HIF-1α. Given an emerging role for tumour cell-derived exosomes in tumour progression, this has significant implications for understanding the hypoxic tumour phenotype, whereby hypoxic

  14. Cord blood T cells mediate enhanced antitumor effects compared with adult peripheral blood T cells. (United States)

    Hiwarkar, Prashant; Qasim, Waseem; Ricciardelli, Ida; Gilmour, Kimberly; Quezada, Sergio; Saudemont, Aurore; Amrolia, Persis; Veys, Paul


    Unrelated cord blood transplantation (CBT) without in vivo T-cell depletion is increasingly used to treat high-risk hematologic malignancies. Following T-replete CBT, naïve CB T cells undergo rapid peripheral expansion with memory-effector differentiation. Emerging data suggest that unrelated CBT, particularly in the context of HLA mismatch and a T-replete graft, may reduce leukemic relapse. To study the role of CB T cells in mediating graft-versus-tumor responses and dissect the underlying immune mechanisms for this, we compared the ability of HLA-mismatched CB and adult peripheral blood (PB) T cells to eliminate Epstein-Barr virus (EBV)-driven human B-cell lymphoma in a xenogeneic NOD/SCID/IL2rg(null) mouse model. CB T cells mediated enhanced tumor rejection compared with equal numbers of PB T cells, leading to improved survival in the CB group (P cells that were autologous vs allogeneic to the lymphoma demonstrated that this antitumor effect was mediated by alloreactive rather than EBV-specific T cells. Analysis of tumor-infiltrating lymphocytes demonstrated that CB T cells mediated this enhanced antitumor effect by rapid infiltration of the tumor with CCR7(+)CD8(+) T cells and prompt induction of cytotoxic CD8(+) and CD4(+) T-helper (Th1) T cells in the tumor microenvironment. In contrast, in the PB group, this antilymphoma effect is impaired because of delayed tumoral infiltration of PB T cells and a relative bias toward suppressive Th2 and T-regulatory cells. Our data suggest that, despite being naturally programmed toward tolerance, reconstituting T cells after unrelated T-replete CBT may provide superior Tc1-Th1 antitumor effects against high-risk hematologic malignancies.

  15. Nanoparticles for Fidgety Cell Movement and Enhanced Wound Healing. (United States)

    Jozic, Ivan; Daunert, Sylvia; Tomic-Canic, Marjana; Pastar, Irena


    Complex spatiotemporal interaction of Rho GTPases with microtubules (MTs) and MT-associated proteins drives directed cellular migration. In this issue, Charafeddine et al. describe a role for a novel MT-severing enzyme, fidgetin-like 2 (FL2), in directional migration of keratinocytes and fibroblasts. FL2 normally localizes to the leading edge of the cell cortex where it shears MTs, thus dictating the size and distribution of focal adhesions by regulating cytoskeletal remodeling. Small interfering RNA (siRNA)-directed knockdown of FL2 increases cell migration and focal adhesion area in vitro through possible interaction with Rho GTPases. Efficient FL2 knockdown in murine wounds was achieved using nanoparticles as a siRNA delivery vehicle, and this resulted in enhanced wound closure in vivo. Effective siRNA nanoparticle targeting of MT-severing enzymes offers promise of controlled and targeted delivery that may maximize therapeutic success for patients with burn wounds and chronic wound disorders.

  16. Phenotypic characterization of drug resistance and tumor initiating cancer stem cells from human bone tumor osteosarcoma cell line OS-77

    Directory of Open Access Journals (Sweden)

    Yue Zhang


    Full Text Available The cancer stem cell theory suggest that presence of small subpopulation of cancer stem cells are the major implication in the cancer treatment and also responsible for tumor recurrence. Based on Hoechst 33342 dye exclusion technique, we have identified about 3.3% of cancer stem like side population (SP cells from human osteosarcoma OS-77 cell line whose prevalence is significantly reduced to 0.3% after treatment with verapamil. The sphere formation assay revealed that osteosarcoma SP cells are highly capable to form tumor spheres (sarcospheres. Further by immunocytochemistry and RT-PCR, we show that OS-77 SP cells have enhanced expression of stem cell surface markers such as CD44, Nanog and ATP-binding cassette (ABC transporter gene (ABCG2 which contributes to self-renewal and drug resistance, respectively. Our findings help to designing a novel therapeutic drug which could effectively target the cancer stem cells and prevent the tumor relapse.

  17. Enhanced regeneration of degraded polymer solar cells by thermal annealing (United States)

    Kumar, Pankaj; Bilen, Chhinder; Feron, Krishna; Zhou, Xiaojing; Belcher, Warwick J.; Dastoor, Paul C.


    The degradation and thermal regeneration of poly(3-hexylethiophene) (P3HT):[6,6]-phenyl-C61-butyric acid methyl ester (PCBM) and P3HT:indene-C60 bisadduct (ICBA) polymer solar cells, with Ca/Al and Ca/Ag cathodes and indium tin oxide/poly(ethylene-dioxythiophene):polystyrene sulfonate anode have been investigated. Degradation occurs via a combination of three primary pathways: (1) cathodic oxidation, (2) active layer phase segregation, and (3) anodic diffusion. Fully degraded devices were subjected to thermal annealing under inert atmosphere. Degraded solar cells possessing Ca/Ag electrodes were observed to regenerate their performance, whereas solar cells having Ca/Al electrodes exhibited no significant regeneration of device characteristics after thermal annealing. Moreover, the solar cells with a P3HT:ICBA active layer exhibited enhanced regeneration compared to P3HT:PCBM active layer devices as a result of reduced changes to the active layer morphology. Devices combining a Ca/Ag cathode and P3HT:ICBA active layer demonstrated ˜50% performance restoration over several degradation/regeneration cycles.

  18. Carbon fiber enhanced bioelectricity generation in soil microbial fuel cells. (United States)

    Li, Xiaojing; Wang, Xin; Zhao, Qian; Wan, Lili; Li, Yongtao; Zhou, Qixing


    The soil microbial fuel cell (MFC) is a promising biotechnology for the bioelectricity recovery as well as the remediation of organics contaminated soil. However, the electricity production and the remediation efficiency of soil MFC are seriously limited by the tremendous internal resistance of soil. Conductive carbon fiber was mixed with petroleum hydrocarbons contaminated soil and significantly enhanced the performance of soil MFC. The maximum current density, the maximum power density and the accumulated charge output of MFC mixed carbon fiber (MC) were 10, 22 and 16 times as high as those of closed circuit control due to the carbon fiber productively assisted the anode to collect the electron. The internal resistance of MC reduced by 58%, 83% of which owed to the charge transfer resistance, resulting in a high efficiency of electron transfer from soil to anode. The degradation rates of total petroleum hydrocarbons enhanced by 100% and 329% compared to closed and opened circuit controls without the carbon fiber respectively. The effective range of remediation and the bioelectricity recovery was extended from 6 to 20cm with the same area of air-cathode. The mixed carbon fiber apparently enhanced the bioelectricity generation and the remediation efficiency of soil MFC by means of promoting the electron transfer rate from soil to anode. The use of conductively functional materials (e.g. carbon fiber) is very meaningful for the remediation and bioelectricity recovery in the bioelectrochemical remediation.

  19. Inverted Silicon Nanopencil Array Solar Cells with Enhanced Contact Structures (United States)

    Liang, Xiaoguang; Shu, Lei; Lin, Hao; Fang, Ming; Zhang, Heng; Dong, Guofa; Yip, Senpo; Xiu, Fei; Ho, Johnny C.


    Although three-dimensional nanostructured solar cells have attracted extensive research attention due to their superior broadband and omnidirectional light-harvesting properties, majority of them are still suffered from complicated fabrication processes as well as disappointed photovoltaic performances. Here, we employed our newly-developed, low-cost and simple wet anisotropic etching to fabricate hierarchical silicon nanostructured arrays with different solar cell contact design, followed by systematic investigations of their photovoltaic characteristics. Specifically, nano-arrays with the tapered tips (e.g. inverted nanopencils) are found to enable the more conformal top electrode deposition directly onto the nanostructures for better series and shunt conductance, but its insufficient film coverage at the basal plane would still restrict the charge carrier collection. In contrast, the low-platform contact design facilitates a substantial photovoltaic device performance enhancement of ~24%, as compared to the one of conventional top electrode design, due to the shortened current path and improved lateral conductance for the minimized carrier recombination and series resistance. This enhanced contact structure can not only maintain excellent photon-trapping behaviors of nanostructures, but also help to eliminate adverse impacts of these tapered nano-morphological features on the contact resistance, providing further insight into design consideration in optimizing the contact geometry for high-performance nanostructured photovoltaic devices.

  20. Nanotextured polymer substrates show enhanced cancer cell isolation and cell culture (United States)

    Islam, Muhymin; Sajid, Adeel; Arif Iftakher Mahmood, M.; Motasim Bellah, Mohammad; Allen, Peter B.; Kim, Young-Tae; Iqbal, Samir M.


    Detection of circulating tumor cells (CTCs) in the early stages of cancer is a great challenge because of their exceedingly small concentration. There are only a few approaches sensitive enough to differentiate tumor cells from the plethora of other cells in a sample like blood. In order to detect CTCs, several antibodies and aptamers have already shown high affinity. Nanotexture can be used to mimic basement membrane to further enhance this affinity. This article reports an approach to fabricate nanotextured polydimethylsiloxane (PDMS) substrates using micro reactive ion etching (micro-RIE). Three recipes were used to prepare nanotextured PDMS using oxygen and carbon tetrafluoride. Micro-RIE provided better control on surface properties. Nanotexturing improved the affinity of PDMS surfaces to capture cancer cells using surface immobilized aptamers against cell membrane overexpressed with epidermal growth factor receptors. In all cases, nanotexture of PDMS increased the effective surface area by creating nanoscale roughness on the surface. Nanotexture also enhanced the growth rate of cultured cells compared to plain surfaces. A comparison among the three nanotextured surfaces demonstrated an almost linear relationship between the surface roughness and density of captured tumor cells. The nanotextured PDMS mimicked biophysical environments for cells to grow faster. This can have many implications in microfluidic platforms used for cell handling.

  1. Stem cell and neurogenic gene-expression profiles link prostate basal cells to aggressive prostate cancer. (United States)

    Zhang, Dingxiao; Park, Daechan; Zhong, Yi; Lu, Yue; Rycaj, Kiera; Gong, Shuai; Chen, Xin; Liu, Xin; Chao, Hsueh-Ping; Whitney, Pamela; Calhoun-Davis, Tammy; Takata, Yoko; Shen, Jianjun; Iyer, Vishwanath R; Tang, Dean G


    The prostate gland mainly contains basal and luminal cells constructed as a pseudostratified epithelium. Annotation of prostate epithelial transcriptomes provides a foundation for discoveries that can impact disease understanding and treatment. Here we describe a genome-wide transcriptome analysis of human benign prostatic basal and luminal epithelial populations using deep RNA sequencing. Through molecular and biological characterizations, we show that the differential gene-expression profiles account for their distinct functional properties. Strikingly, basal cells preferentially express gene categories associated with stem cells, neurogenesis and ribosomal RNA (rRNA) biogenesis. Consistent with this profile, basal cells functionally exhibit intrinsic stem-like and neurogenic properties with enhanced rRNA transcription activity. Of clinical relevance, the basal cell gene-expression profile is enriched in advanced, anaplastic, castration-resistant and metastatic prostate cancers. Therefore, we link the cell-type-specific gene signatures to aggressive subtypes of prostate cancer and identify gene signatures associated with adverse clinical features.

  2. Glucocorticoids enhance CD163 expression in placental Hofbauer cells. (United States)

    Tang, Zhonghua; Niven-Fairchild, Tracy; Tadesse, Serkalem; Norwitz, Errol R; Buhimschi, Catalin S; Buhimschi, Irina A; Guller, Seth


    Periplacental levels of glucocorticoid (GC) peak at parturition, and synthetic GC is administered to women at risk for preterm delivery. However, little is known concerning cell-type-specific effects of GC in placenta. Hofbauer cells (HBCs) are fetal macrophages that are located adjacent to fetal capillaries in placenta. The goal of the current study was to determine whether GC treatment altered HBC gene expression and function. Western blotting and flow cytometry revealed CD163 and folate receptor-β (FR-β), markers of antiinflammatory M2 macrophages, were specifically expressed by primary cultures of HBCs immunopurified from human term placentas. GC receptor mRNA and protein levels were higher in HBCs compared with placental fibroblasts. Treatment of HBCs with cortisol or dexamethasone (DEX) markedly and specifically enhanced CD163 protein and mRNA levels, whereas expression of FR-β and CD68 were largely unresponsive to GC treatment. DEX treatment also increased hemoglobin uptake by HBCs, evidence of enhanced HBC function. The level of CD163 mRNA, but not FR-β or CD68 mRNA, was stimulated in placental explant cultures by DEX treatment, and increased CD163/FR-β and CD163/CD68 mRNA ratios sensitively reflected the response to GC. Maternal GC administration was associated with increased CD163/FR-β and CD163/CD68 mRNA ratios in placentas from women with spontaneous preterm birth. In conclusion, in vitro studies indicated that GC treatment specifically up-regulated CD163 expression in HBCs and enhanced HBC function. In addition, the observed alterations in patterns of expression of macrophage marker genes associated with maternal GC administration suggest that HBCs are in vivo targets of GC action.

  3. Titanium phosphate glass microcarriers induce enhanced osteogenic cell proliferation and human mesenchymal stem cell protein expression

    Directory of Open Access Journals (Sweden)

    Nilay J Lakhkar


    Full Text Available In this study, we have developed 50- to 100-µm-sized titanium phosphate glass microcarriers (denoted as Ti5 that show enhanced proliferation of human mesenchymal stem cells and MG63 osteosarcoma cells, as well as enhanced human mesenchymal stem cell expression of bone differentiation markers, in comparison with commercially available glass microspheres at all time points. We also demonstrate that these microcarriers provide superior human mesenchymal stem cell proliferation with conventional Dulbecco’s Modified Eagle medium than with a specially developed commercial stem cell medium. The microcarrier proliferative capacity is revealed by a 24-fold increase in MG63 cell numbers in spinner flask bioreactor studies performed over a 7-day period, versus only a 6-fold increase in control microspheres under the same conditions; the corresponding values of Ti5 and control microspheres under static culture are 8-fold and 7-fold, respectively. The capability of guided osteogenic differentiation is confirmed by ELISAs for bone morphogenetic protein-2 and osteopontin, which reveal significantly greater expression of these markers, especially osteopontin, by human mesenchymal stem cells on the Ti5 microspheres than on the control. Scanning electron microscopy and confocal laser scanning microscopy images reveal favorable MG63 and human mesenchymal stem cell adhesion on the Ti5 microsphere surfaces. Thus, the results demonstrate the suitability of the developed microspheres for use as microcarriers in bone tissue engineering applications.

  4. Establishment of human cell type-specific iPS cells with enhanced chondrogenic potential. (United States)

    Guzzo, Rosa M; Scanlon, Vanessa; Sanjay, Archana; Xu, Ren-He; Drissi, Hicham


    The propensity of induced pluripotent stem (iPS) cells to differentiate into specific lineages may be influenced by a number of factors, including the selection of the somatic cell type used for reprogramming. Herein we report the generation of new iPS cells, which we derived from human articular chondrocytes and from cord blood mononucleocytes via lentiviral-mediated delivery of Oct4, Klf4, Sox2, and cMyc. Molecular, cytochemical, and cytogenic analyses confirmed the acquisition of hallmark features of pluripotency, as well as the retention of normal karyotypes following reprogramming of both the human articular chondrocytes (AC) and the cord blood (CB) cells. In vitro and in vivo functional analyses formally established the pluripotent differentiation capacity of all cell lines. Chondrogenic differentiation assays comparing iPS cells derived from AC, CB, and a well established dermal fibroblast cell line (HDFa-Yk26) identified enhanced proteoglycan-rich matrix formation and cartilage-associated gene expression from AC-derived iPS cells. These findings suggest that the tissue of origin may impact the fate potential of iPS cells for differentiating into specialized cell types, such as chondrocytes. Thus, we generated new cellular tools for the identification of inherent features driving high chondrogenic potential of reprogrammed cells.

  5. Atg3 Overexpression Enhances Bortezomib-Induced Cell Death in SKM-1 Cell.

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    Lin Zhuang

    Full Text Available Myelodysplastic syndrome (MDS is a group of heterogeneous hematopoietic stem cell malignancies with a high risk of transformation into acute myeloid leukemia (AML. Clonal evolutions are significantly associated with transformation to AML. According to a gene expression microarray, atg3 is downregulated in MDS patients progressing to leukemia, but less is known about the function of Atg3 in the survival and death of MSD/AML cells. Moreover, the role of autophagy as a result of bortezomib treatment is controversial. The current study was designed to investigate the function of Atg3 in SKM-1 cells and to study the effect of Atg3 on cell viability and cell death following bortezomib treatment.Four leukemia cell lines (SKM-1, THP-1, NB4 and K562 and two healthy patients' bone marrow cells were analyzed for Atg3 expression via qRT-PCR and Western blotting analysis. The role of Atg3 in SKM-1 cell survival and cell death was analyzed by CCK-8 assay, trypan blue exclusion assay, DAPI staining and Annexin V/PI dual staining with or without bortezomib treatment. Western blotting analysis was used to detect proteins in autophagic and caspase signaling pathways. Electron microscopy was used to observe ultrastructural changes after Atg3 overexpression.Downregulation of Atg3 expression was detected in four leukemia cell lines compared with healthy bone marrow cells. Atg3 mRNA was significantly decreased in MDS patients' bone marrow cells. Overexpression of Atg3 in SKM-1 cells resulted in AKT-mTOR-dependent autophagy, a significant reduction in cell proliferation and increased cell death, which could be overcome by the autophagy inhibitor 3-MA. SKM-1 cells overexpressing Atg3 were hypersensitive to bortezomib treatment at different concentrations via autophagic cell death and enhanced sensitivity to apoptosis in the SKM-1 cell line. Following treatment with 3-MA, the sensitivity of Atg3-overexpressing cells to bortezomib treatment was reduced. Atg3 knockdown

  6. Cell penetrating peptide-modified poly(lactic-co-glycolic acid) nanoparticles with enhanced cell internalization. (United States)

    Steinbach, Jill M; Seo, Young-Eun; Saltzman, W Mark


    The surface modification of nanoparticles (NPs) can enhance the intracellular delivery of drugs, proteins, and genetic agents. Here we studied the effect of different surface ligands, including cell penetrating peptides (CPPs), on the cell binding and internalization of poly(lactic-co-glycolic) (PLGA) NPs. Relative to unmodified NPs, we observed that surface-modified NPs greatly enhanced cell internalization. Using one CPP, MPG (unabbreviated notation), that achieved the highest degree of internalization at both low and high surface modification densities, we evaluated the effect of two different NP surface chemistries on cell internalization. After 2h, avidin-MPG NPs enhanced cellular internalization by 5 to 26-fold relative to DSPE-MPG NP formulations. Yet, despite a 5-fold increase in MPG density on DSPE compared to Avidin NPs, both formulations resulted in similar internalization levels (48 and 64-fold, respectively) after 24h. Regardless of surface modification, all NPs were internalized through an energy-dependent, clathrin-mediated process, and became dispersed throughout the cell. Overall both Avidin- and DSPE-CPP modified NPs significantly increased internalization and offer promising delivery options for applications in which internalization presents challenges to efficacious delivery.

  7. Modeling nanostructure-enhanced light trapping in organic solar cells

    DEFF Research Database (Denmark)

    Adam, Jost

    A promising approach for improving the power conversion efficiencies of organic solar cells (OSCs) is by incorporating nanostructures in their thin film architecture to improve the light absorption in the device’s active polymer layers. Here, we present a modelling framework for the prediction...... of optical and plasmonic field enhancement by nanostructures in (or close to) the active layers and electrodes in OSCs. We incorporate finite-difference time-domain (FDTD) calculations alongside semi- analytical approaches, as the rigorous coupled-wave analysis (RCWA) and mode-coupling theory. Our simulation......-compatible method for non-periodic electrode structuring by pores of controlled dimensions, formed through anodic oxidation of sputter-deposited high-purity aluminium films [3]. [1] Kluge, C., et al. Multi-periodic nanostructures for photon control. Optics Express, 22 (S5), A1363. (2014) [2] Skigin, D., et al...

  8. Optical enhancement effects of plasmonic nanostructures on organic photovoltaic cells

    KAUST Repository

    Park, Hui Joon


    © 2015 Hui Joon Park and L. Jay Guo. Published by Elsevier B.V. on behalf of Chinese Chemical Society and Institute of Materia Medica, Chinese Academy of Medical Sciences. All rights reserved. In this article, the optical enhancement effects of plasmonic nanostructures on OPV cells were reviewed as an effective way to resolve the mismatch problems between the short exciton diffusion length in organic semiconductors (around 10 nm) and the large thickness required to fully absorb sunlight (e.g. hundreds of nanometers). Especially, the performances of OPVs with plasmonic nanoparticles in photoactive and buffer layers and with periodic nanostructures were investigated. Furthermore, nanoimprint lithography-based nanofabrication processes that can easily control the dimension and uniformity of structures for large-area and uniform plasmonic nanostructures were demonstrated.

  9. Evidence that pulsed electric field treatment enhances the cell wall porosity of yeast cells. (United States)

    Ganeva, Valentina; Galutzov, Bojidar; Teissie, Justin


    The application of rectangular electric pulses, with 0.1-2 ms duration and field intensity of 2.5-4.5 kV/cm, to yeast suspension mediates liberation of cytoplasmic proteins without cell lysis. The aim of this study was to evaluate the effect of pulsed electric field with similar parameters on cell wall porosity of different yeast species. We found that electrically treated cells become more susceptible to lyticase digestion. In dependence on the strain and the electrical conditions, cell lysis was obtained at 2-8 times lower enzyme concentration in comparison with control untreated cells. The increase of the maximal lysis rate was between two and nine times. Furthermore, when applied at low concentration (1 U/ml), the lyticase enhanced the rate of protein liberation from electropermeabilized cells without provoking cell lysis. Significant differences in the cell surface of control and electrically treated cells were revealed by scanning electron microscopy. Data presented in this study allow us to conclude that electric field pulses provoke not only plasma membrane permeabilization, but also changes in the cell wall structure, leading to increased wall porosity.

  10. Programmed cell death-10 enhances proliferation and protects malignant T cells from apoptosis

    DEFF Research Database (Denmark)

    Lauenborg, Britt; Kopp, Katharina; Krejsgaard, Thorbjørn;


    The programmed cell death-10 (PDCD10; also known as cerebral cavernous malformation-3 or CCM3) gene encodes an evolutionarily conserved protein associated with cell apoptosis. Mutations in PDCD10 result in cerebral cavernous malformations, an important cause of cerebral hemorrhage. PDCD10...... of cutaneous T-cell lymphoma (Sezary syndrome) patients. PDCD10 is associated with protein phosphatase-2A, a regulator of mitogenesis and apoptosis in malignant T cells. Inhibition of oncogenic signal pathways [Jak3, Notch1, and nuclear factor-¿B (NF-¿B)] partly inhibits the constitutive PDCD10 expression......, whereas an activator of Jak3 and NF-¿B, interleukin-2 (IL-2), enhances PDCD10 expression. Functional data show that PDCD10 depletion by small interfering RNA induces apoptosis and decreases proliferation of the sensitive cells. To our knowledge, these data provide the first functional link between PDCD10...

  11. Particle-cell contact enhances antibacterial activity of silver nanoparticles.

    Directory of Open Access Journals (Sweden)

    Olesja Bondarenko

    Full Text Available BACKGROUND: It is generally accepted that antibacterial properties of Ag nanoparticles (AgNPs are dictated by their dissolved fraction. However, dissolution-based concept alone does not fully explain the toxic potency of nanoparticulate silver compared to silver ions. METHODOLOGY/PRINCIPAL FINDINGS: Herein, we demonstrated that the direct contact between bacterial cell and AgNPs' surface enhanced the toxicity of nanosilver. More specifically, cell-NP contact increased the cellular uptake of particle-associated Ag ions - the single and ultimate cause of toxicity. To prove that, we evaluated the toxicity of three different AgNPs (uncoated, PVP-coated and protein-coated to six bacterial strains: Gram-negative Escherichia coli, Pseudomonas fluorescens, P. putida and P. aeruginosa and Gram-positive Bacillus subtilis and Staphylococcus aureus. While the toxicity of AgNO3 to these bacteria varied only slightly (the 4-h EC50 ranged from 0.3 to 1.2 mg Ag/l, the 4-h EC50 values of protein-coated AgNPs for various bacterial strains differed remarkably, from 0.35 to 46 mg Ag/l. By systematically comparing the intracellular and extracellular free Ag(+ liberated from AgNPs, we demonstrated that not only extracellular dissolution in the bacterial test environment but also additional dissolution taking place at the particle-cell interface played an essential role in antibacterial action of AgNPs. The role of the NP-cell contact in dictating the antibacterial activity of Ag-NPs was additionally proven by the following observations: (i separation of bacterial cells from AgNPs by particle-impermeable membrane (cut-off 20 kDa, ∼4 nm significantly reduced the toxicity of AgNPs and (ii P. aeruginosa cells which tended to attach onto AgNPs, exhibited the highest sensitivity to all forms of nanoparticulate Ag. CONCLUSIONS/SIGNIFICANCE: Our findings provide new insights into the mode of antibacterial action of nanosilver and explain some discrepancies in this field

  12. Knockdown of the cell cycle inhibitor p21 enhances cartilage formation by induced pluripotent stem cells. (United States)

    Diekman, Brian O; Thakore, Pratiksha I; O'Connor, Shannon K; Willard, Vincent P; Brunger, Jonathan M; Christoforou, Nicolas; Leong, Kam W; Gersbach, Charles A; Guilak, Farshid


    The limited regenerative capacity of articular cartilage contributes to progressive joint dysfunction associated with cartilage injury or osteoarthritis. Cartilage tissue engineering seeks to provide a biological substitute for repairing damaged or diseased cartilage, but requires a cell source with the capacity for extensive expansion without loss of chondrogenic potential. In this study, we hypothesized that decreased expression of the cell cycle inhibitor p21 would enhance the proliferative and chondrogenic potential of differentiated induced pluripotent stem cells (iPSCs). Murine iPSCs were directed to differentiate toward the chondrogenic lineage with an established protocol and then engineered to express a short hairpin RNA (shRNA) to reduce the expression of p21. Cells expressing the p21 shRNA demonstrated higher proliferative potential during monolayer expansion and increased synthesis of glycosaminoglycans (GAGs) in pellet cultures. Furthermore, these cells could be expanded ∼150-fold over three additional passages without a reduction in the subsequent production of GAGs, while control cells showed reduced potential for GAG synthesis with three additional passages. In pellets from extensively passaged cells, knockdown of p21 attenuated the sharp decrease in cell number that occurred in control cells, and immunohistochemical analysis showed that p21 knockdown limited the production of type I and type X collagen while maintaining synthesis of cartilage-specific type II collagen. These findings suggest that manipulating the cell cycle can augment the monolayer expansion and preserve the chondrogenic capacity of differentiated iPSCs, providing a strategy for enhancing iPSC-based cartilage tissue engineering.

  13. Forced extinction of CD24 stem-like breast cancer marker alone promotes radiation resistance through the control of oxidative stress. (United States)

    Bensimon, Julie; Biard, Denis; Paget, Vincent; Goislard, Maud; Morel-Altmeyer, Sandrine; Konge, Julie; Chevillard, Sylvie; Lebeau, Jérôme


    Along with CD44, CD24 is a key marker of breast cancer stem cells (CSCs), frequently defined by CD24(-)/CD44(+) labeling. Among all phenotypes classically attributed to breast CD24(-)/CD44(+) cancer cells, radiation resistance has been extensively described and seen as being implicated in radiotherapy failure. Our previous data indicated that CD24(-) cells constitute a radiation-resistant subpopulation transitory selected by high doses of ionizing radiation. However, little is known about the biological role of CD24 in breast cancers, and no function has been assigned to CD24 in radiation response. Here, CD24 expression was induced in CD24(-) cells or knocked-down in CD24(+) cells. We show that forced extinction of CD24 expression is associated with decreased proliferation rate, lower levels of reactive oxygen species (ROS) and decreased genomic instability. On the opposite when CD24 is artificially expressed in CD24(-) cells, proliferation rates in vitro and in vivo, ROS levels and genomic instability are enhanced. Moreover, we observe that loss of CD24 expression leads to radiation resistance, by preventing radiation-induced cell death and promoting generation of progeny in relation to lower G2/M blockade and a smaller proportion of polyploid cells. Finally, control of ROS levels appears to be the key event in the CD24-mediated radiation response. For the first time, CD24 is proposed as a direct actor in radiation response of breast cancer cells, independently of CD44 expression. These findings could have interesting applications in evaluating the intrinsic radiation response of primary tumors.

  14. Cell-Cell Connection Enhances Proliferation and Neuronal Differentiation of Rat Embryonic Neural Stem/Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Qian Jiao


    Full Text Available Cell-cell interaction as one of the niche signals plays an important role in the balance of stem cell quiescence and proliferation or differentiation. In order to address the effect and the possible mechanisms of cell-cell connection on neural stem/progenitor cells (NSCs/NPCs proliferation and differentiation, upon passaging, NSCs/NPCs were either dissociated into single cell as usual (named Group I or mechanically triturated into a mixture of single cell and small cell clusters containing direct cell-cell connections (named Group II. Then the biological behaviors including proliferation and differentiation of NSCs/NPCs were observed. Moreover, the expression of gap junction channel, neurotrophic factors and the phosphorylation status of MAPK signals were compared to investigate the possible mechanisms. Our results showed that, in comparison to the counterparts in Group I, NSCs/NPCs in Group II survived well with preferable neuronal differentiation. In coincidence with this, the expression of connexin 45 (Cx45, as well as brain derived neurotrophic factor (BDNF and neurotrophin 3 (NT-3 in Group II were significantly higher than those in Group I. Phosphorylation of ERK1/2 and JNK2 were significantly upregulated in Group II too, while no change was found about p38. Furthermore, the differences of NSCs/NPCs biological behaviors between Group I and II completely disappeared when ERK and JNK phosphorylation were inhibited. These results indicated that cell-cell connection in Group II enhanced NSCs/NPCs survival, proliferation and neuronal differentiation through upregulating the expression of gap junction and neurotrophic factors. MAPK signals- ERK and JNK might contribute to the enhancement. Efforts for maintaining the direct cell-cell connection are worth making to provide more favorable niches for NSCs/NPCs survival, proliferation and neuronal differentiation.

  15. RASSF1A expression inhibits cell growth and enhances cell chemosensitivity to mitomycin in BEL-7402 hepatocellular carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    GUAN Hong-geng; XUE Wan-jiang; QIAN Hai-xin; ZHOU Xiao-jun; QIN Lei; LAN Jing


    Background The antitumor role of Ras association domain family 1A (RASSFIA) gene and its potential molecular mechanisms are not well understood. The objective of this study was to observe the antitumor ability of RASSFIA in hepatoceliular carcinoma, and study the mechanisms of cell apoptosis induced by RASSFIA.Methods After stably transfecting a RASSF1A (wild-type or mutant) expression vector into the BEL-7402 hepatocellular carcinoma cell line, RT-PCR and Westem blotting was used to detect the RASSF1A expression levels in recombinant cells. The effects of wild-type RASSF1A on cell growth were observed in vitro by analyzing cell proliferation rate, cell colony formation, and in vivo by analyzing tumorigenesis in nude mice. In addition, the effect of RASSF1A gene expression on the chemosensitivity of human hepatocellular carcinoma cells to antitumor drugs was examined by inhibition of cell proliferation and the percentage of apoptotic cells.Results Wild-type RASSF1A, not the mutant, suppressed cell growth in vitro and in vivo. Re-expression of wild-type RASSF1A could enhance the inhibition of cell proliferation and the percentage of apoptotic cells following cell treatment with mitomycin, but had no significant effect when combined with adriamycin, etoposide, 5-fluorouracil and cisplatJn treatment.Conclusion Wild-type RASSF1A inhibits cell growth and enhances cell chemosensitivity to mitomycin in hepatocellular carcinoma, suggesting that RASSF1A may serve as a new target for gene therapy in hepatocellular carcinoma patients.

  16. Super-enhancers in the control of cell identity and disease. (United States)

    Hnisz, Denes; Abraham, Brian J; Lee, Tong Ihn; Lau, Ashley; Saint-André, Violaine; Sigova, Alla A; Hoke, Heather A; Young, Richard A


    Super-enhancers are large clusters of transcriptional enhancers that drive expression of genes that define cell identity. Improved understanding of the roles that super-enhancers play in biology would be afforded by knowing the constellation of factors that constitute these domains and by identifying super-enhancers across the spectrum of human cell types. We describe here the population of transcription factors, cofactors, chromatin regulators, and transcription apparatus occupying super-enhancers in embryonic stem cells and evidence that super-enhancers are highly transcribed. We produce a catalog of super-enhancers in a broad range of human cell types and find that super-enhancers associate with genes that control and define the biology of these cells. Interestingly, disease-associated variation is especially enriched in the super-enhancers of disease-relevant cell types. Furthermore, we find that cancer cells generate super-enhancers at oncogenes and other genes important in tumor pathogenesis. Thus, super-enhancers play key roles in human cell identity in health and in disease.

  17. Paracrine factors of mesenchymal stem cells recruit macrophages and endothelial lineage cells and enhance wound healing.

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    Liwen Chen

    Full Text Available Bone marrow derived mesenchymal stem cells (BM-MSCs have been shown to enhance wound healing; however, the mechanisms involved are barely understood. In this study, we examined paracrine factors released by BM-MSCs and their effects on the cells participating in wound healing compared to those released by dermal fibroblasts. Analyses of BM-MSCs with Real-Time PCR and of BM-MSC-conditioned medium by antibody-based protein array and ELISA indicated that BM-MSCs secreted distinctively different cytokines and chemokines, such as greater amounts of VEGF-alpha, IGF-1, EGF, keratinocyte growth factor, angiopoietin-1, stromal derived factor-1, macrophage inflammatory protein-1alpha and beta and erythropoietin, compared to dermal fibroblasts. These molecules are known to be important in normal wound healing. BM-MSC-conditioned medium significantly enhanced migration of macrophages, keratinocytes and endothelial cells and proliferation of keratinocytes and endothelial cells compared to fibroblast-conditioned medium. Moreover, in a mouse model of excisional wound healing, where concentrated BM-MSC-conditioned medium was applied, accelerated wound healing occurred compared to administration of pre-conditioned or fibroblast-conditioned medium. Analysis of cell suspensions derived from the wound by FACS showed that wounds treated with BM-MSC-conditioned medium had increased proportions of CD4/80-positive macrophages and Flk-1-, CD34- or c-kit-positive endothelial (progenitor cells compared to wounds treated with pre-conditioned medium or fibroblast-conditioned medium. Consistent with the above findings, immunohistochemical analysis of wound sections showed that wounds treated with BM-MSC-conditioned medium had increased abundance of macrophages. Our results suggest that factors released by BM-MSCs recruit macrophages and endothelial lineage cells into the wound thus enhancing wound healing.

  18. Long Noncoding RNA MALAT-1 Enhances Stem Cell-Like Phenotypes in Pancreatic Cancer Cells

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    Feng Jiao


    Full Text Available Cancer stem cells (CSCs play a vital role in tumor initiation, progression, metastasis, chemoresistance, and recurrence. The mechanisms that maintain the stemness of these cells remain largely unknown. Our previous study indicated that MALAT-1 may serve as an oncogenic long noncoding RNA in pancreatic cancer by promoting epithelial-mesenchymal transition (EMT and regulating CSCs markers expression. More significantly, there is emerging evidence that the EMT process may give rise to CSCs, or at least cells with stem cell-like properties. Therefore, we hypothesized that MALAT-1 might enhance stem cell-like phenotypes in pancreatic cancer cells. In this study, our data showed that MALAT-1 could increase the proportion of pancreatic CSCs, maintain self-renewing capacity, decrease the chemosensitivity to anticancer drugs, and accelerate tumor angiogenesis in vitro. In addition, subcutaneous nude mouse xenografts revealed that MALAT-1 could promote tumorigenicity of pancreatic cancer cells in vivo. The underlying mechanisms may involve in increased expression of self-renewal related factors Sox2. Collectively, we for the first time found the potential effects of MALAT-1 on the stem cell-like phenotypes in pancreatic cancer cells, suggesting a novel role of MALAT-1 in tumor stemness, which remains to be fully elucidated.

  19. Stem Cells Derived from Tooth Periodontal Ligament Enhance Functional Angiogenesis by Endothelial Cells (United States)

    Yeasmin, Shamima; Ceccarelli, Jacob; Vigen, Marina; Carrion, Bita; Putnam, Andrew J.; Tarle, Susan A.


    In regenerative medicine approaches involving cell therapy, selection of the appropriate cell type is important in that the cells must directly (differentiation) or indirectly (trophic effects) participate in the regenerative response. Regardless of the mode of action of the cells, angiogenesis underlies the success of these approaches. Stem cells derived from tooth tissues, specifically the periodontal ligament of teeth (periodontal ligament stem cells [PDLSCs]), have recently been identified as a good source of multipotent cells for cell therapies. PDLSCs have demonstrated properties similar to mesenchymal stem cells (MSCs), yet, unlike MSCs, their vascular potential has not been previously demonstrated. Thus, the aim of this study was to determine if PDLSCs could modulate angiogenesis. In comparison to MSCs and stem cells derived from tooth pulp tissues (SHEDs), we first determined if PDLSCs released soluble proangiogenic factors with the capacity to induce vessel formation by endothelial cells (ECs). Next, the ability of PDLSCs to modulate angiogenesis was examined through their cotransplantation with ECs in subcutaneous sites of immunocompromised mice. Finally, the stability of the PDLSC-mediated vasculature was determined through evaluation of the maturity and functionality of the vessels formed following PDLSC transplantation. It was determined that PDLSCs produced appreciable levels of vascular endothelial growth factor and basic fibroblast growth factor-2, and additionally, were able to initiate in vitro angiogenesis of ECs comparable to MSC- and SHED-mediated angiogenesis. In vivo cotransplantation of ECs with PDLSCs significantly (>50% increase) enhanced the number of blood vessels formed relative to transplantation of ECs alone. Finally, vessels formed following PDLSC cotransplantation were more mature and less permeable than those formed after transplantation of EC alone. These data demonstrate for the first time that PDLSCs have vascular potential

  20. PAX4 enhances beta-cell differentiation of human embryonic stem cells.

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    Chee Gee Liew

    Full Text Available BACKGROUND: Human embryonic stem cells (HESC readily differentiate into an apparently haphazard array of cell types, corresponding to all three germ layers, when their culture conditions are altered, for example by growth in suspension as aggregates known as embryoid bodies (EBs. However, this diversity of differentiation means that the efficiency of producing any one particular cell type is inevitably low. Although pancreatic differentiation has been reported from HESC, practicable applications for the use of beta-cells derived from HESC to treat diabetes will only be possible once techniques are developed to promote efficient differentiation along the pancreatic lineages. METHODS AND FINDINGS: Here, we have tested whether the transcription factor, Pax4 can be used to drive the differentiation of HESC to a beta-cell fate in vitro. We constitutively over-expressed Pax4 in HESCs by stable transfection, and used Q-PCR analysis, immunocytochemistry, ELISA, Ca(2+ microfluorimetry and cell imaging to assess the role of Pax4 in the differentiation and intracellular Ca(2+ homeostasis of beta-cells developing in embryoid bodies produced from such HESC. Cells expressing key beta-cell markers were isolated by fluorescence-activated cell sorting after staining for high zinc content using the vital dye, Newport Green. CONCLUSION: Constitutive expression of Pax4 in HESC substantially enhances their propensity to form putative beta-cells. Our findings provide a novel foundation to study the mechanism of pancreatic beta-cells differentiation during early human development and to help evaluate strategies for the generation of purified beta-cells for future clinical applications.

  1. Dendritic Cells Enhance HIV Infection of Memory CD4(+) T Cells in Human Lymphoid Tissues. (United States)

    Reyes-Rodriguez, Angel L; Reuter, Morgan A; McDonald, David


    Dendritic cells (DCs) play a key role in controlling infections by coordinating innate and adaptive immune responses to invading pathogens. Paradoxically, DCs can increase HIV-1 dissemination in vitro by binding and transferring infectious virions to CD4(+) T cells, a process called transinfection. Transinfection has been well characterized in cultured cell lines and circulating primary T cells, but it is unknown whether DCs enhance infection of CD4(+) T cells in vivo. In untreated HIV infection, massive CD4(+) T-cell infection and depletion occur in secondary lymphoid tissues long before decline is evident in the peripheral circulation. To study the role of DCs in HIV infection of lymphoid tissues, we utilized human tonsil tissues, cultured either as tissue blocks or as aggregate suspension cultures, in single-round infection experiments. In these experiments, addition of monocyte-derived DCs (MDDCs) to the cultures increased T-cell infection, particularly in CD4(+) T cells expressing lower levels of HLA-DR. Subset analysis demonstrated that MDDCs increased HIV-1 infection of central and effector memory T-cell populations. Depletion of endogenous myeloid DCs (myDCs) from the cultures decreased memory T-cell infection, and readdition of MDDCs restored infection to predepletion levels. Using an HIV-1 fusion assay, we found that MDDCs equally increased HIV delivery into naïve, central, and effector memory T cells in the cultures, whereas predepletion of myDCs reduced fusion into memory T cells. Together, these data suggest that resident myDCs facilitate memory T-cell infection in lymphoid tissues, implicating DC-mediated transinfection in driving HIV dissemination within these tissues in untreated HIV/AIDS.

  2. Organic solar cells: Degradation processes and approaches to enhance performance (United States)

    Fungura, Fadzai

    Organic solar cells (OSCs) have attracted a lot of attention due to their potential as flexible, lightweight, and low-cost renewable energy sources. Significant improvements have been made in increasing the devices' power conversion efficiency (PCE) and extensive efforts to understand degradation mechanisms and increase OSCs' lifetimes are ongoing. OSCs with higher than 10% efficiency have been reported. Enhanced stability and efficiency of inverted poly(3-hexylthiophene) (P3HT) solar cells with Cesium (Cs) halides were achieved by spin-coating Bphen (4,7-di(phenyl)-1,10-phenanthroline) on the halide layer and adding an 100nm polystyrene beads layer on the blank side of the OSC. To investigate photodegradation in a low-bandgap polymer, PBDTTT-EFT (benzo[1,2-b:4,5-b']dithiophene (BDT) and thieno[3,4-b]-thiophene), PBDTTT-EFT:PCBM bulk heterojunction (BHJ) solar cells were irradiated under 1X sun intensity and their electronic measurements were monitored over time. The electronic measurements revealed an increase in deep defect density in the polymer and at the donor-acceptor (D/A) interface of BHJ SCs and an increase in charge recombination as well as a decrease in external quantum efficiency, charge collection, short circuit current, open circuit voltage and hole mobility. Filtering blue and UV light resulted in drastically reduced photodegradation. Electron paramagnetic resonance measurements were performed on the photodegraded polymer and BHJ films, and revealed for the first time experimental evidence for metastable carbon dangling bonds (g=2.0029+/-0.0004) formed by blue/UV irradiation of the films. Dark EPR showed an increase in densities of other spin-active sites in the polymer, fullerene and polymer:fullerene blends in agreement with electronic measurements. The EPR results revealed that the carbon dangling bonds were at the D/A interface. These studies helped to better understand degradation mechanisms in a low-bandgap polymer, PBDTTT-EFT, ways to enhance

  3. Spaceflight enhances cell aggregation and random budding in Candida albicans.

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    Aurélie Crabbé

    Full Text Available This study presents the first global transcriptional profiling and phenotypic characterization of the major human opportunistic fungal pathogen, Candida albicans, grown in spaceflight conditions. Microarray analysis revealed that C. albicans subjected to short-term spaceflight culture differentially regulated 452 genes compared to synchronous ground controls, which represented 8.3% of the analyzed ORFs. Spaceflight-cultured C. albicans-induced genes involved in cell aggregation (similar to flocculation, which was validated by microscopic and flow cytometry analysis. We also observed enhanced random budding of spaceflight-cultured cells as opposed to bipolar budding patterns for ground samples, in accordance with the gene expression data. Furthermore, genes involved in antifungal agent and stress resistance were differentially regulated in spaceflight, including induction of ABC transporters and members of the major facilitator family, downregulation of ergosterol-encoding genes, and upregulation of genes involved in oxidative stress resistance. Finally, downregulation of genes involved in actin cytoskeleton was observed. Interestingly, the transcriptional regulator Cap1 and over 30% of the Cap1 regulon was differentially expressed in spaceflight-cultured C. albicans. A potential role for Cap1 in the spaceflight response of C. albicans is suggested, as this regulator is involved in random budding, cell aggregation, and oxidative stress resistance; all related to observed spaceflight-associated changes of C. albicans. While culture of C. albicans in microgravity potentiates a global change in gene expression that could induce a virulence-related phenotype, no increased virulence in a murine intraperitoneal (i.p. infection model was observed under the conditions of this study. Collectively, our data represent an important basis for the assessment of the risk that commensal flora could play during human spaceflight missions. Furthermore, since the

  4. NF-κB activation enhances cell death by antimitotic drugs in human prostate cancer cells

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    Parrondo Ricardo


    Full Text Available Abstract Background NF-κB is a transcription factor that promotes inhibition of apoptosis and resistance to chemotherapy. It is commonly believed that inhibition of NF-κB activity can increase sensitivity of cancer cells to chemotherapy. However, there is evidence that NF-κB activation can sensitize cells to apoptosis and that inhibition of NF-κB results in resistance to chemotherapy. In prostate cancer, it is not clear in the different cell types (androgen-dependent and castration-resistant if activation or inhibition of NF-κB is required for stimulation of apoptosis by chemotherapy. Results Our data indicate that the response of prostate cancer (PC cells to the antimitotic drugs docetaxel (Doc and 2-methoxyestradiol (2ME2 is dependent on the levels of NF-κB activity. In androgen-dependent LNCaP cells, Doc and 2ME2 treatment increased the low basal NF-κB activity, as determined by Western blot analysis of phospho-IκBα/p65, NF-κB promoter reporter assays, and p65 localization. Treatment of LNCaP cells with parthenolide, a pharmacologic inhibitor of NF-κB, or introduction of dominant-negative IκBα, or an shRNA specific for p65, a component of the NF-κB heterodimer, blocked apoptosis induced by Doc and 2ME2. In castration-resistant DU145 and PC3 cells, Doc and 2ME2 had little effect on the high basal NF-κB activity and addition of parthenolide did not enhance cell death. However, the combination of Doc or 2ME2 with betulinic acid (BA, a triterpenoid that activates NF-κB, stimulated apoptosis in LNCaP and non-apoptotic cell death in DU145 and PC3 cells. Increased sensitivity to cell death mediated by the Doc or 2ME2 + BA combination is likely due to increased NF-κB activity. Conclusions Our data suggest that the combination of antimitotic drugs with NF-κB inhibitors will have antagonistic effects in a common type of PC cell typical of LNCaP. However, combination strategies utilizing antimitotic drugs with BA, an activator of NF

  5. Enhancement of heat transfer in red cell suspensions in vitro experiments. (United States)

    Carr, R T; Tiruvaloor, N R


    New data on laminar heat convection with red cell suspensions have been gathered for both heating and cooling. When compared to data for the suspending medium alone, it is apparent that the red cells enhance laminar heat transfer when Pe greater than 4. This is probably due to particle movements. These new data disagree with earlier studies which indicated no enhancement of heat transfer for blood cell suspensions. The data do agree with previous correlations for enhanced thermal transport in sheared suspensions.

  6. Enhancement of Spectral Response of Dye-Sensitized Solar Cells (United States)

    Chang, Shuai

    ruthenium-based/organic dyes for co-sensitized DSSCs are also investigated. Another approach is to increase light utility in DSSCs exploiting surface plasmon resonance (SPR) of noble metal nanostructures (e.g. Au, Ag). In this thesis, I will show that the strong longitudinal plasmonic absorption of Au nanorods (NRs) can be used to increase the low-photon energy sunlight harvesting in DSSCs, broadening strong light response of the devices. In specific, a remarkable improvement in photocurrent generation at 600-720 nm is achieved. This enhancement mechanism is anticipated to be applied to other kind of DSSCs with various dye molecules. In another approach, AuNRs/TiO2 core-shell nanostructures are employed as scattering layer for plasmon-enhanced light harvesting in DSSCs and also obtained positive results. Evolved from DSSCs, perovskite solar cells (PSCs) now become a new favorite in the field of photovoltaics. An intrinsic problem of this kind of solar cells is the use of lead based materials, which is of high toxicity and prohibited by the European Union and some other countries. I have conducted some fundamental research for lead-free PSCs using tin-based perovskite and have observed a surface plasmon resonance absorption of the organometal perovskite film of CH3NH3SnI3, which has potential applications for IR-absorption in the future solar photovoltaics. I believe the improved understanding on the co-sensitization mechanisms and the plasmonic effect to broaden the spectral response in DSSCs are luciferous for the design and fabrication of the new generation solar cells with high-efficiency and low-cost.

  7. Spheroid culture for enhanced differentiation of human embryonic stem cells to hepatocyte-like cells. (United States)

    Subramanian, Kartik; Owens, Derek Jason; Raju, Ravali; Firpo, Meri; O'Brien, Timothy D; Verfaillie, Catherine M; Hu, Wei-Shou


    Stem cell-derived hepatocyte-like cells hold great potential for the treatment of liver disease and for drug toxicity screening. The success of these applications hinges on the generation of differentiated cells with high liver specific activities. Many protocols have been developed to guide human embryonic stem cells (hESCs) to differentiate to the hepatic lineage. Here we report cultivation of hESCs as three-dimensional aggregates that enhances their differentiation to hepatocyte-like cells. Differentiation was first carried out in monolayer culture for 20 days. Subsequently cells were allowed to self-aggregate into spheroids. Significantly higher expression of liver-specific transcripts and proteins, including Albumin, phosphoenolpyruvate carboxykinase, and asialoglycoprotein receptor 1 was observed. The differentiated phenotype was sustained for more than 2 weeks in the three-dimensional spheroid culture system, significantly longer than in monolayer culture. Cells in spheroids exhibit morphological and ultrastructural characteristics of primary hepatocytes by scanning and transmission electron microscopy in addition to mature functions, such as biliary excretion of metabolic products and cytochrome P450 activities. This three-dimensional spheroid culture system may be appropriate for generating high quality, functional hepatocyte-like cells from ESCs.

  8. Enhancement of cell performance using a gadolinium strontium cobaltite coated cathode in molten carbonate fuel cells (United States)

    Song, Shin Ae; Jang, Seong-Cheol; Han, Jonghee; Yoon, Sung Pil; Nam, Suk Woo; Oh, In-Hwan; Lim, Tae-Hoon

    To enhance cathode performance, gadolinium strontium cobaltite (Gd 0.6Sr 0.4CoO 3, GSC) is coated onto a porous Ni plate by a vacuum suction method, for use as the cathode in molten carbonate fuel cells (MCFCs). GSC is a mixed ionic and electronic conductor (MIEC) material, and thus has high electronic conductivity and catalytic activity at low temperatures. The electrode performance of the GSC-coated cathode is examined by various methods, such as single cell operation and electrochemical impedance spectroscopy (EIS). At 600 °C, the performance of a single cell using a GSC-coated cathode is 0.813 V. This result is very surprising given that the performance of an uncoated conventional cathode is 0.69 V. Impedance analysis confirms that a dramatic decrease in the charge transfer resistance after GSC coating is primarily responsible for the cell enhancement at low temperature. The reaction orders for O 2 and CO 2 at uncoated and GSC-coated cathodes are also examined via a symmetric cell test, to identify the reaction mechanism of oxygen reduction. The peroxide mechanism, which is known to be a fast reaction, is predominant for the GSC-coated cathode at low temperatures, whereas the superoxide mechanism is predominant for the uncoated cathode.

  9. Mathematical modelling of phenotypic plasticity and conversion to a stem-cell state under hypoxia (United States)

    Dhawan, Andrew; Madani Tonekaboni, Seyed Ali; Taube, Joseph H.; Hu, Stephen; Sphyris, Nathalie; Mani, Sendurai A.; Kohandel, Mohammad


    Hypoxia, or oxygen deficiency, is known to be associated with breast tumour progression, resistance to conventional therapies and poor clinical prognosis. The epithelial-mesenchymal transition (EMT) is a process that confers invasive and migratory capabilities as well as stem cell properties to carcinoma cells thus promoting metastatic progression. In this work, we examined the impact of hypoxia on EMT-associated cancer stem cell (CSC) properties, by culturing transformed human mammary epithelial cells under normoxic and hypoxic conditions, and applying in silico mathematical modelling to simulate the impact of hypoxia on the acquisition of CSC attributes and the transitions between differentiated and stem-like states. Our results indicate that both the heterogeneity and the plasticity of the transformed cell population are enhanced by exposure to hypoxia, resulting in a shift towards a more stem-like population with increased EMT features. Our findings are further reinforced by gene expression analyses demonstrating the upregulation of EMT-related genes, as well as genes associated with therapy resistance, in hypoxic cells compared to normoxic counterparts. In conclusion, we demonstrate that mathematical modelling can be used to simulate the role of hypoxia as a key contributor to the plasticity and heterogeneity of transformed human mammary epithelial cells.

  10. Disodium cromoglycate enhances ongoing immunoglobulin production in vitro in human B cells. (United States)

    Kimata, H; Yoshida, A; Ishioka, C; Mikawa, H


    The effect of disodium cromoglycate (DSCG) upon human immunoglobulin (Ig) isotypes and IgG subclasses production by purified B cells was studied. DSCG enhanced IgM, IgG1, IgG2, IgG3, IgG4 and IgA production in a dose-dependent fashion, while DSCG failed to induce IgE production at any concentrations tested by purified B cells. When B cells were separated into small resting and large activated B cells, DSCG failed to induce Ig production from small resting B cells in the presence or absence of Staphylococcus aureus Cowan strain I (SAC). In contrast, in large activated B cells DSCG significantly enhanced all types of Ig production (two-to threefold), especially IgG4 production (seven-to 11-fold), except IgE, which large B cells did not produce. The enhancement of IgG subclass production was not subclass switching, since DSCG failed to enhance IgG1 production in B cells depleted of surface IgG1+ cells (sIgG1+ cells). Similarly, DSCG did not enhance IgG2, IgG3 or IgG4 production from sIgG2-, sIgG3- or sIgG4- B cells, respectively, Interleukin-4 (IL-4) or interleukin-6 (IL-6) also enhanced Ig production except IgG4 from large activated B cells. The enhancing effect of DSCG was not mediated by IL-4 or IL-6 since anti-IL-4 or anti-IL-6 antibody failed to block the DSCG-induced enhancement. DSCG also enhanced IgG2 and IgM production from human B-cell lines GM-1500 and CBL, respectively. These results suggest that DSCG directly and preferentially stimulates activated B cells which are producing Ig and, in addition, enhances their Ig production. PMID:1904400

  11. Chloroquine enhances gefitinib cytotoxicity in gefitinib-resistant nonsmall cell lung cancer cells.

    Directory of Open Access Journals (Sweden)

    Mei-Chuan Tang

    Full Text Available Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs, including gefitinib, are effective for non-small cell lung cancer (NSCLC patients with EGFR mutations. However, these patients eventually develop resistance to EGFR-TKI. The goal of the present study was to investigate the involvement of autophagy in gefitinib resistance. We developed gefitinib-resistant cells (PC-9/gef from PC-9 cells (containing exon 19 deletion EGFR after long-term exposure in gefitinib. PC-9/gef cells (B4 and E3 were 200-fold more resistant to gefitinib than PC-9/wt cells. Compared with PC-9/wt cells, both PC-9/gefB4 and PC-9/gefE3 cells demonstrated higher basal LC3-II levels which were inhibited by 3-methyladenine (3-MA, an autophagy inhibitor and potentiated by chloroquine (CQ, an inhibitor of autophagolysosomes formation, indicating elevated autophagy in PC-9/gef cells. 3-MA and CQ concentration-dependently inhibited cell survival of both PC-9wt and PC-9/gef cells, suggesting that autophagy may be pro-survival. Furthermore, gefitinib increased LC3-II levels and autolysosome formation in both PC-9/wt cells and PC-9/gef cells. In PC-9/wt cells, CQ potentiated the cytotoxicity by low gefitinib (3 nM. Moreover, CQ overcame the acquired gefitinib resistance in PC-9/gef cells by enhancing gefitinib-induced cytotoxicity, activation of caspase 3 and poly (ADP-ribose polymerase cleavage. Using an in vivo model xenografting with PC-9/wt and PC-9/gefB4 cells, oral administration of gefitinib (50 mg/kg completely inhibited the tumor growth of PC-9/wt but not PC-9/gefB4cells. Combination of CQ (75 mg/kg, i.p. and gefitinib was more effective than gefitinib alone in reducing the tumor growth of PC-9/gefB4. Our data suggest that inhibition of autophagy may be a therapeutic strategy to overcome acquired resistance of gefitinib in EGFR mutation NSCLC patients.

  12. Active screen plasma nitriding enhances cell attachment to polymer surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Kaklamani, Georgia, E-mail: [University of Birmingham, College of Engineering and Physical Sciences, School of Metallurgy and Materials, Edgbaston, Birmingham B15 2TT (United Kingdom); Bowen, James; Mehrban, Nazia [University of Birmingham, College of Engineering and Physical Sciences, School of Chemical Engineering, Edgbaston, Birmingham B15 2TT (United Kingdom); Dong, Hanshan [University of Birmingham, College of Engineering and Physical Sciences, School of Metallurgy and Materials, Edgbaston, Birmingham B15 2TT (United Kingdom); Grover, Liam M. [University of Birmingham, College of Engineering and Physical Sciences, School of Chemical Engineering, Edgbaston, Birmingham B15 2TT (United Kingdom); Stamboulis, Artemis [University of Birmingham, College of Engineering and Physical Sciences, School of Metallurgy and Materials, Edgbaston, Birmingham B15 2TT (United Kingdom)


    Active screen plasma nitriding (ASPN) is a well-established technique used for the surface modification of materials, the result of which is often a product with enhanced functional performance. Here we report the modification of the chemical and mechanical properties of ultra-high molecular weight poly(ethylene) (UHMWPE) using 80:20 (v/v) N{sub 2}/H{sub 2} ASPN, followed by growth of 3T3 fibroblasts on the treated and untreated polymer surfaces. ASPN-treated UHMWPE showed extensive fibroblast attachment within 3 h of seeding, whereas fibroblasts did not successfully attach to untreated UHMWPE. Fibroblast-coated surfaces were maintained for up to 28 days, monitoring their metabolic activity and morphology throughout. The chemical properties of the ASPN-treated UHMWPE surface were studied using X-ray photoelectron spectroscopy, revealing the presence of C-N, C=N, and C≡N chemical bonds. The elastic modulus, surface topography, and adhesion properties of the ASPN-treated UHMWPE surface were studied over 28 days during sample storage under ambient conditions and during immersion in two commonly used cell culture media.

  13. Polylysine modification of adenoviral fiber protein enhances muscle cell transduction. (United States)

    Bouri, K; Feero, W G; Myerburg, M M; Wickham, T J; Kovesdi, I; Hoffman, E P; Clemens, P R


    Adenoviral vectors (ADVs) are used widely for gene delivery to different tissues including muscle. One particularly promising use for ADVs is in the transfer of the dystrophin gene to the muscle of patients with Duchenne muscular dystrophy (DMD). However, studies in different animal models of DMD suggest that ADVs inefficiently transduce mature skeletal muscle. In this article we test whether AdZ.F(pK7), a genetically modified ADV that expresses a polylysine moiety on the end of the fiber protein, could enhance transduction of muscle cells and circumvent the maturation-dependent loss of muscle infectivity by ADVs. The efficiency of transduction was tested at different levels of muscle maturation. In vitro, AdZ.F(pK7) showed a higher level of transduction at all stages of differentiation including myoblasts, myotubes, and single muscle fibers. In vivo, mature skeletal muscle was transduced fourfold better by AdZ.F(pK7) than by the unmodifled vector (AdZ.F). Together, these observations demonstrate improved ADV transduction of skeletal muscle by modifying ADV tropism, and provide a proof-of-principle that modification of ADVs to target muscle-specific molecules could result in tissue-specific transfer of skeletal muscle tissue as well.

  14. Super-enhancers delineate disease-associated regulatory nodes in T cells. (United States)

    Vahedi, Golnaz; Kanno, Yuka; Furumoto, Yasuko; Jiang, Kan; Parker, Stephen C J; Erdos, Michael R; Davis, Sean R; Roychoudhuri, Rahul; Restifo, Nicholas P; Gadina, Massimo; Tang, Zhonghui; Ruan, Yijun; Collins, Francis S; Sartorelli, Vittorio; O'Shea, John J


    Enhancers regulate spatiotemporal gene expression and impart cell-specific transcriptional outputs that drive cell identity. Super-enhancers (SEs), also known as stretch-enhancers, are a subset of enhancers especially important for genes associated with cell identity and genetic risk of disease. CD4(+) T cells are critical for host defence and autoimmunity. Here we analysed maps of mouse T-cell SEs as a non-biased means of identifying key regulatory nodes involved in cell specification. We found that cytokines and cytokine receptors were the dominant class of genes exhibiting SE architecture in T cells. Nonetheless, the locus encoding Bach2, a key negative regulator of effector differentiation, emerged as the most prominent T-cell SE, revealing a network in which SE-associated genes critical for T-cell biology are repressed by BACH2. Disease-associated single-nucleotide polymorphisms for immune-mediated disorders, including rheumatoid arthritis, were highly enriched for T-cell SEs versus typical enhancers or SEs in other cell lineages. Intriguingly, treatment of T cells with the Janus kinase (JAK) inhibitor tofacitinib disproportionately altered the expression of rheumatoid arthritis risk genes with SE structures. Together, these results indicate that genes with SE architecture in T cells encompass a variety of cytokines and cytokine receptors but are controlled by a 'guardian' transcription factor, itself endowed with an SE. Thus, enumeration of SEs allows the unbiased determination of key regulatory nodes in T cells, which are preferentially modulated by pharmacological intervention.

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

    Directory of Open Access Journals (Sweden)

    Cook John A


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

  16. Continual Cell Deformation Induced via Attachment to Oriented Fibers Enhances Fibroblast Cell Migration (United States)

    Qin, Sisi; Ricotta, Vincent; Simon, Marcia; Clark, Richard A. F.; Rafailovich, Miriam H.


    Fibroblast migration is critical to the wound healing process. In vivo, migration occurs on fibrillar substrates, and previous observations have shown that a significant time lag exists before the onset of granulation tissue. We therefore conducted a series of experiments to understand the impact of both fibrillar morphology and migration time. Substrate topography was first shown to have a profound influence. Fibroblasts preferentially attach to fibrillar surfaces, and orient their cytoplasm for maximal contact with the fiber edge. In the case of en-mass cell migration out of an agarose droplet, fibroblasts on flat surfaces emerged with an enhanced velocity, v = 52μm/h, that decreases to the single cell value, v = 28μm/h within 24 hours and remained constant for at least four days. Fibroblasts emerging on fibrillar surfaces emerged with the single cell velocity, which remained constant for the first 24 hours and then increased reaching a plateau with more than twice the initial velocity within the next three days. The focal adhesions were distributed uniformly in cells on flat surfaces, while on the fibrillar surface they were clustered along the cell periphery. Furthermore, the number of focal adhesions for the cells on the flat surfaces remained constant, while it decreased on the fibrillar surface during the next three days. The deformation of the cell nuclei was found to be 50% larger on the fiber surfaces for the first 24 hours. While the mean deformation remained constant on the flat surface, it increased for the next three days by 24% in cells on fibers. On the fourth day, large actin/myosin fibers formed in cells on fibrillar surfaces only and coincided with a change from the standard migration mechanism involving extension of lamellipodia, and retraction of the rear, to one involving strong contractions oriented along the fibers and centered about the nucleus. PMID:25774792

  17. Continual cell deformation induced via attachment to oriented fibers enhances fibroblast cell migration.

    Directory of Open Access Journals (Sweden)

    Sisi Qin

    Full Text Available Fibroblast migration is critical to the wound healing process. In vivo, migration occurs on fibrillar substrates, and previous observations have shown that a significant time lag exists before the onset of granulation tissue. We therefore conducted a series of experiments to understand the impact of both fibrillar morphology and migration time. Substrate topography was first shown to have a profound influence. Fibroblasts preferentially attach to fibrillar surfaces, and orient their cytoplasm for maximal contact with the fiber edge. In the case of en-mass cell migration out of an agarose droplet, fibroblasts on flat surfaces emerged with an enhanced velocity, v = 52μm/h, that decreases to the single cell value, v = 28μm/h within 24 hours and remained constant for at least four days. Fibroblasts emerging on fibrillar surfaces emerged with the single cell velocity, which remained constant for the first 24 hours and then increased reaching a plateau with more than twice the initial velocity within the next three days. The focal adhesions were distributed uniformly in cells on flat surfaces, while on the fibrillar surface they were clustered along the cell periphery. Furthermore, the number of focal adhesions for the cells on the flat surfaces remained constant, while it decreased on the fibrillar surface during the next three days. The deformation of the cell nuclei was found to be 50% larger on the fiber surfaces for the first 24 hours. While the mean deformation remained constant on the flat surface, it increased for the next three days by 24% in cells on fibers. On the fourth day, large actin/myosin fibers formed in cells on fibrillar surfaces only and coincided with a change from the standard migration mechanism involving extension of lamellipodia, and retraction of the rear, to one involving strong contractions oriented along the fibers and centered about the nucleus.

  18. Mogoltacin enhances vincristine cytotoxicity in human transitional cell carcinoma (TCC) cell line. (United States)

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


    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%.

  19. Absorption efficiency enhancement in inorganic and organic thin film solar cells via plasmonic honeycomb nanoantenna arrays. (United States)

    Tok, Rüştü Umut; Sendur, Kürşat


    We demonstrate theoretically that by embedding plasmonic honeycomb nanoantenna arrays into the active layers of inorganic (c-Si) and organic (P3HT:PCBM/PEDOT:PSS) thin film solar cells, absorption efficiency can be improved. To obtain the solar cell absorption spectrum that conforms to the solar radiation, spectral broadening is achieved by breaking the symmetry within the Wigner-Seitz unit cell on a uniform hexagonal grid. For optimized honeycomb designs, absorption efficiency enhancements of 106.2% and 20.8% are achieved for c-Si and P3HT:PCBM/PEDOT:PSS thin film solar cells, respectively. We have demonstrated that the transverse modes are responsible for the enhancement in c-Si solar cells, whereas both the longitudinal and transverse modes, albeit weaker, are the main enhancement mechanisms for P3HT:PCBM/PEDOT:PSS solar cells. For both inorganic and organic solar cells, the absorption enhancement is independent of polarization.

  20. Disruption of cell walls for enhanced lipid recovery (United States)

    Knoshaug, Eric P; Donohoe, Bryon S; Gerken, Henri; Laurens, Lieve; Van Wychen, Stefanie Rose


    Presented herein are methods of using cell wall degrading enzymes for recovery of internal lipid bodies from biomass sources such as algae. Also provided are algal cells that express at least one exogenous gene encoding a cell wall degrading enzyme and methods for recovering lipids from the cells.

  1. Identification of SSEA-1 expressing enhanced reprogramming (SEER) cells in porcine embryonic fibroblasts

    DEFF Research Database (Denmark)

    Li, Dong; Secher, Jan Ole Bertelsen; Juhl, Morten


    Previous research has shown that a subpopulation of cells within cultured human dermal fibroblasts, termed multilineage-differentiating stress enduring (Muse) cells, are preferentially reprogrammed into induced pluripotent stem cells. However, controversy exists over whether these cells are the o......Previous research has shown that a subpopulation of cells within cultured human dermal fibroblasts, termed multilineage-differentiating stress enduring (Muse) cells, are preferentially reprogrammed into induced pluripotent stem cells. However, controversy exists over whether these cells...... in mesenchymal stem cells (MSCs). We therefore termed these cells SSEA-1 Expressing Enhanced Reprogramming (SEER) cells. Interestingly, SEER cells were more effective at differentiating into osteocytes and chondrocytes in vitro. We conclude that SEER cells are more amenable for reprogramming...... and that the expression of mesenchymal stem cell genes is advantageous in the reprogramming process. This data provides evidence supporting the elite theory and helps to delineate which cell types and specific genes are important for reprogramming in the pig....

  2. Alpha beta T-cell development is not affected by inversion of TCR beta gene enhancer sequences: polar enhancement of gene expression regardless of enhancer orientation. (United States)

    Huang, Fang; Cabaud, Olivier; Verthuy, Christophe; Hueber, Anne-Odile; Ferrier, Pierre


    V(D)J recombination and expression of the T-cell receptor beta (TCRbeta) gene are required for the development of the alphabeta T lymphocyte lineage. These processes depend on a transcriptional enhancer (Ebeta) which acts preferentially on adjacent upstream sequences, and has little impact on the 5' distal and 3' proximal regions of the TCRbeta locus. Using knock-in mice, we show that alphabeta T-cell differentiation and TCRbeta gene recombination and expression are not sensitive to the orientation of Ebeta sequences. We discuss the implication of these results regarding the mode of enhancer function at this locus during T lymphocyte development.

  3. Enhancement of insulin-producing cell differentiation from embryonic stem cells using pax4-nucleofection method

    Institute of Scientific and Technical Information of China (English)

    Han-Tso Lin; Hung-Hai Ku; Chung-Lan Kao; Kun-Hsiung Lee; Yuh-Lih Chang; Shih-Hwa Chiou; Fu-Ting Tsai; Tung-Hu Tsai; Dey-Chyi Sheu; Larry LT Ho


    AIM: To enhance the differentiation of insulin producing cell (IPC) ability from embryonic stem (ES) cells in vitro.METHODS: Four-day embryoid body (EB)-formatted ES cells were dissociated as single cells for the followed plasmid DNA delivery. The use of Nucleofector- Electroporator (Amaxa biosystems, Germany) in combination with medium-contained G418 provided a high efficiency of gene delivery for advanced selection. Neucleofected cells were plated on the top of fibronectin coated Petri dishes. Addition of Ly294002 and raised the glucose in medium at 24 h before examination.The differentiation status of these cells was monitored by semi-quantitative PCR (SQ-PCR) detection of the expression of relative genes, such as oct-4, sox-17, foxa2, mixl1, pdx-1, insulin 1, glucagons and somatostatin. The percentage of IPC population on d 18 of the experiment was investigated by immunohistochemistry (IHC), and the content/secretion of insulin was estimated by ELISA assay. The mice with severe combined immunodeficiency disease (SCID) pretreated with streptozotocin (STZ) were used to eliminate plasma glucose restoration after pax4+ ES implantation.RESULTS: A high efficiency of gene delivery was demonstrated when neucleofection was used in the present study; approximately 70% cells showed DsRed expression 2 d after neucleofection. By selection of medium-contained G418, the percentage of DsRed expressing cells kept high till the end of study. The pancreatic differentiation seemed to be accelerated by pax4 nucleofection. When compared to the group of cells with mock control, foxa2, mixl1, pdx1, higher insulin and somatostatin levels were detected by SQ-PCR 4 d after nucleofection in the group of pax4 expressing plasmid delivery. Approximately 55% of neucleofected cells showed insulin expression 18 d after neucleofection, and only 18% of cells showed insulin expression in mock control. The disturbance was shown by nucleofected pax4 RNAi vector; only 8% of cells expressed insulin 18

  4. Protein hairy enhancer of split-1 expression during differentiation of muscle-derived stem cells into neuron-like cells

    Institute of Scientific and Technical Information of China (English)

    Mina Huang; Zhanpeng Guo; Kun Liu; Xifan Mei; Shiqiang Fang; Jinhao Zeng; Yansong Wang; Yajiang Yuan


    Muscle-derived stem cells were isolated from the skeletal muscle of Sprague-Dawley neonatal rats aged 3 days old.Cells at passage 5 were incubated in Dulbecco's modified Eagle's medium supplemented with 10% (v/v) fetal bovine serum,20 μg/L nerve growth factor,20 μg/L basic fibroblast growth factor and 1% (v/v) penicillin for 6 days.Cells presented with long processes, similar to nerve cells.Connections were formed between cell processes.Immunocytochemical staining with neuron specific enolase verified that cells differentiated into neuron-like cells. Immunofluorescence cytochemistry and western blot results revealed that the expression of protein hairy enhancer of split-1 was significantly reduced.These results indicate that low expression of protein hairy enhancer of split-1 participates in the differentiation of muscle-derived stem cells into neuron-like cells.

  5. CD44v6 regulates growth of brain tumor stem cells partially through the AKT-mediated pathway.

    Directory of Open Access Journals (Sweden)

    Mayumi Jijiwa

    Full Text Available Identification of stem cell-like brain tumor cells (brain tumor stem-like cells; BTSC has gained substantial attention by scientists and physicians. However, the mechanism of tumor initiation and proliferation is still poorly understood. CD44 is a cell surface protein linked to tumorigenesis in various cancers. In particular, one of its variant isoforms, CD44v6, is associated with several cancer types. To date its expression and function in BTSC is yet to be identified. Here, we demonstrate the presence and function of the variant form 6 of CD44 (CD44v6 in BTSC of a subset of glioblastoma multiforme (GBM. Patients with CD44(high GBM exhibited significantly poorer prognoses. Among various variant forms, CD44v6 was the only isoform that was detected in BTSC and its knockdown inhibited in vitro growth of BTSC from CD44(high GBM but not from CD44(low GBM. In contrast, this siRNA-mediated growth inhibition was not apparent in the matched GBM sample that does not possess stem-like properties. Stimulation with a CD44v6 ligand, osteopontin (OPN, increased expression of phosphorylated AKT in CD44(high GBM, but not in CD44(low GBM. Lastly, in a mouse spontaneous intracranial tumor model, CD44v6 was abundantly expressed by tumor precursors, in contrast to no detectable CD44v6 expression in normal neural precursors. Furthermore, overexpression of mouse CD44v6 or OPN, but not its dominant negative form, resulted in enhanced growth of the mouse tumor stem-like cells in vitro. Collectively, these data indicate that a subset of GBM expresses high CD44 in BTSC, and its growth may depend on CD44v6/AKT pathway.

  6. Enhancer identification in mouse embryonic stem cells using integrative modeling of chromatin and genomic features

    Directory of Open Access Journals (Sweden)

    Chen Chih-yu


    Full Text Available Abstract Background Epigenetic modifications, transcription factor (TF availability and differences in chromatin folding influence how the genome is interpreted by the transcriptional machinery responsible for gene expression. Enhancers buried in non-coding regions are found to be associated with significant differences in histone marks between different cell types. In contrast, gene promoters show more uniform modifications across cell types. Here we used histone modification and chromatin-associated protein ChIP-Seq data sets in mouse embryonic stem (ES cells as well as genomic features to identify functional enhancer regions. Using co-bound sites of OCT4, SOX2 and NANOG (co-OSN, validated enhancers and co-bound sites of MYC and MYCN (limited enhancer activity as enhancer positive and negative training sets, we performed multinomial logistic regression with LASSO regularization to identify key features. Results Cross validations reveal that a combination of p300, H3K4me1, MED12 and NIPBL features to be top signatures of co-OSN regions. Using a model from 10 signatures, 83% of top 1277 putative 1 kb enhancer regions (probability greater than or equal to 0.8 overlapped with at least one TF peak from 7 mouse ES cell ChIP-Seq data sets. These putative enhancers are associated with increased gene expression of neighbouring genes and significantly enriched in multiple TF bound loci in agreement with combinatorial models of TF binding. Furthermore, we identified several motifs of known TFs significantly enriched in putative enhancer regions compared to random promoter regions and background. Comparison with an active H3K27ac mark in various cell types confirmed cell type-specificity of these enhancers. Conclusions The top enhancer signatures we identified (p300, H3K4me1, MED12 and NIPBL will allow for the identification of cell type-specific enhancer regions in diverse cell types.

  7. Simulations of solar cell absorption enhancement using resonant modes of a nanosphere array



    We propose an approach for enhancing the absorption of thin-film amorphous silicon solar cells using periodic arrangements of resonant dielectric nanospheres deposited as a continuous film on top of a thin planar cell. We numerically demonstrate this enhancement using three dimensional (3D) full field, finite difference time domain simulations and 3D finite element device physics simulations of a nanosphere array above a thin-film amorphous silicon solar cell structure featuring back reflecto...

  8. Stem cell characteristics in prostate cancer cell lines.

    NARCIS (Netherlands)

    Pfeiffer, M.J.; Schalken, J.A.


    BACKGROUND: Recent studies indicate the presence of a small, stem-like cell population in several human cancers that is crucial for the tumour (re)population. OBJECTIVE: Six established prostate cancer (PCa) cell lines-DU145, DuCaP, LAPC-4, 22Rv1, LNCaP, and PC-3-were examined for their stem cell pr

  9. Hypoxic preconditioning enhances neural stem cell transplantation therapy after intracerebral hemorrhage in mice. (United States)

    Wakai, Takuma; Narasimhan, Purnima; Sakata, Hiroyuki; Wang, Eric; Yoshioka, Hideyuki; Kinouchi, Hiroyuki; Chan, Pak H


    Previous studies have shown that intraparenchymal transplantation of neural stem cells ameliorates neurological deficits in animals with intracerebral hemorrhage. However, hemoglobin in the host brain environment causes massive grafted cell death and reduces the effectiveness of this approach. Several studies have shown that preconditioning induced by sublethal hypoxia can markedly improve the tolerance of treated subjects to more severe insults. Therefore, we investigated whether hypoxic preconditioning enhances neural stem cell resilience to the hemorrhagic stroke environment and improves therapeutic effects in mice. To assess whether hypoxic preconditioning enhances neural stem cell survival when exposed to hemoglobin, neural stem cells were exposed to 5% hypoxia for 24 hours before exposure to hemoglobin. To study the effectiveness of hypoxic preconditioning on grafted-neural stem cell recovery, neural stem cells subjected to hypoxic preconditioning were grafted into the parenchyma 3 days after intracerebral hemorrhage. Hypoxic preconditioning significantly enhanced viability of the neural stem cells exposed to hemoglobin and increased grafted-cell survival in the intracerebral hemorrhage brain. Hypoxic preconditioning also increased neural stem cell secretion of vascular endothelial growth factor. Finally, transplanted neural stem cells with hypoxic preconditioning exhibited enhanced tissue-protective capability that accelerated behavioral recovery. Our results suggest that hypoxic preconditioning in neural stem cells improves efficacy of stem cell therapy for intracerebral hemorrhage.

  10. Graphene for enhanced embryonic stem cell photo-transfection efficiency

    CSIR Research Space (South Africa)

    Mthunzi, P


    Full Text Available Due to their pluripotency properties, embryonic stem (ES) cells possess great potential in regenerative therapy. Since reported a promising tissue engineering scaffold material, here, graphene is demonstrated to significantly improve the ES cell...

  11. Graphene substrates enhance optical transfection efficiency in pluripotent stem cells

    CSIR Research Space (South Africa)

    Khanyile, T


    Full Text Available . The effects of growing CHO-K1 cells on graphene coated glass as opposed to neat glass was evaluated here. By monitoring mitochondrial activity through adenosine triphosphate (ATP) luminescence, the cell viability was determined. Cytosolic lactate...

  12. Dimethyloxalylglycine may be enhance the capacity of neural-like cells in treatment of Alzheimer disease. (United States)

    Ghasemi Moravej, Fahimeh; Vahabian, Mehrangiz; Soleimani Asl, Sara


    Although using differentiated stem cells is the best proposed option for the treatment of Alzheimer disease (AD), an efficient differentiation and cell therapy require enhanced cell survival and homing and decreased apoptosis. It seems that hypoxia preconditioning via Dimethyloxalylglycine (DMOG) may increase the capacity of MSC to induce neural like stem cells (NSCs). Furthermore, it can likely improve the viability of NSCs when transplanted into the brain of AD rats. © 2016 International Federation for Cell Biology.

  13. Adipocyte secreted factors enhance aggressiveness of prostate carcinoma cells.

    Directory of Open Access Journals (Sweden)

    Ângela Moreira

    Full Text Available Obesity has been associated with increased incidence and risk of mortality of prostate cancer. One of the proposed mechanisms underlying this risk association is the change in adipokines expression that could promote the development and progression of the prostate tumor cells. The main goal of this study was to evaluate the effect of preadipocyte and adipocyte secretome in the proliferation, migration and invasion of androgen independent prostate carcinoma cells (RM1 and to assess cell proliferation in the presence of the adiposity signals leptin and insulin. RM1 cells were co-cultured in with preadipocytes, adipocytes or cultured in their respective conditioned medium. Cell proliferation was assessed by flow cytometry and XTT viability test. Cell migration was evaluated using a wound healing injury assay of RM1 cells cultured with conditioned media. Cellular invasion of RM1 cells co-cultured with adipocytes and preadipocytes was assessed using matrigel membranes. Preadipocyte conditioned medium was associated with a small increase in RM1 proliferation, while adipocytes conditioned media significantly increased RM1 cell proliferation (p<0.01. Adipocytes also significantly increased the RM1 cells proliferation in co-culture (p <0.01. Cell migration was higher in RM1 cells cultured with preadipocyte and adipocyte conditioned medium. RM1 cell invasion was significantly increased after co-culture with preadipocytes and adipocytes (p <0.05. Insulin also increased significantly the cell proliferation in contrast to leptin, which showed no effect. In conclusion, prostate carcinoma cells seem to be influenced by factors secreted by adipocytes that are able to increase their ability to proliferate, migrate and invade.

  14. Stathmin1 increases radioresistance by enhancing autophagy in non-small-cell lung cancer cells

    Directory of Open Access Journals (Sweden)

    Zhang X


    Full Text Available Xi Zhang,1,2 Jingfen Ji,3 Yu Yang,4 Juan Zhang,2 Liangfang Shen1 1Department of Oncology, Xiangya Hospital of Central South University, 2Department of Oncology, The Third Xiangya Hospital of Central South University, 3Department of General Surgery, The Second Xiangya Hospital of Central South University, 4Department of Oncology, 163 Hospital of PLA, Changsha, Hunan, People’s Republic of China Abstract: Radioresistance has been demonstrated to be involved in the poor prognosis of patients with non-small-cell lung cancer (NSCLC. However, the underlying mechanism remains largely unclear. Investigation on special therapeutic targets associated with radioresistance shows promises for the enhancement of clinical radiotherapy effect toward NSCLC. This study aimed to reveal the role of Stathmin1 (STMN1 in radioresistance in NSCLC as well as the underlying mechanism. Our data showed that the protein levels of STMN1 were significantly upregulated in NSCLC cells subjected to radiation, accompanied with the activation of autophagy. Knockdown of STMN1 expression enhanced the sensitivity of NSCLC cells to X-ray, and the radiation-induced autophagy was also inhibited. Molecular mechanism investigation showed that knockdown of STMN1 expression upregulated the activity of phosphoinositide 3-kinase (PI3K/mammalian target of rapamycin (mTOR signaling pathway in NSCLC cells. Moreover, the activation of PI3K/mTOR signaling showed an inhibitory effect on the autophagy and radioresistance induced by STMN1 in NSCLC cells. In addition, luciferase reporter assay data indicated that STMN1 was a direct target gene of miR-101, which had been reported to be an inhibitor of autophagy. Based on these data, we suggest that as a target gene of miR-101, STMN1 promotes the radioresistance by induction of autophagy through PI3K/mTOR signaling pathway in NSCLC. Therefore, STMN1 may become a potential therapeutic target for NSCLC radiotherapy. Keywords: radioresistance, non-small-cell

  15. Enhancing Solar Cell Efficiency Using Photon Upconversion Materials

    Directory of Open Access Journals (Sweden)

    Yunfei Shang


    Full Text Available Photovoltaic cells are able to convert sunlight into electricity, providing enough of the most abundant and cleanest energy to cover our energy needs. However, the efficiency of current photovoltaics is significantly impeded by the transmission loss of sub-band-gap photons. Photon upconversion is a promising route to circumvent this problem by converting these transmitted sub-band-gap photons into above-band-gap light, where solar cells typically have high quantum efficiency. Here, we summarize recent progress on varying types of efficient upconversion materials as well as their outstanding uses in a series of solar cells, including silicon solar cells (crystalline and amorphous, gallium arsenide (GaAs solar cells, dye-sensitized solar cells, and other types of solar cells. The challenge and prospect of upconversion materials for photovoltaic applications are also discussed

  16. Enhancing Solar Cell Efficiency Using Photon Upconversion Materials (United States)

    Shang, Yunfei; Hao, Shuwei; Yang, Chunhui; Chen, Guanying


    Photovoltaic cells are able to convert sunlight into electricity, providing enough of the most abundant and cleanest energy to cover our energy needs. However, the efficiency of current photovoltaics is significantly impeded by the transmission loss of sub-band-gap photons. Photon upconversion is a promising route to circumvent this problem by converting these transmitted sub-band-gap photons into above-band-gap light, where solar cells typically have high quantum efficiency. Here, we summarize recent progress on varying types of efficient upconversion materials as well as their outstanding uses in a series of solar cells, including silicon solar cells (crystalline and amorphous), gallium arsenide (GaAs) solar cells, dye-sensitized solar cells, and other types of solar cells. The challenge and prospect of upconversion materials for photovoltaic applications are also discussed. PMID:28347095

  17. Enhancement of Radiosensitivity by Roscovitine Pretreatment in Human Non-small Cell Lung Cancer A549 Cells


    Feng, Zhang; Tao, Zhang; Zhong-Ping, GU; Yong-An, Zhou; yong, Han; Xiao-Fei, LI; Xiao-Ping, WANG; Qing-Shu, CHENG; Qi-Bing, MEI; Department of Thoracic Surgery, Tangdu Hospital, The Fourth Military Medical University; Department of Pharmacology, The Fourth Military Medical University


    Roscovitine has been reported to have anti-proliferative properties and is in process of undergoing clinical trials. In addition to its intrinsic anticancer properties, it has recently been suggested that roscovitine may also enhance the activity of traditional chemo- and radio-therapies in certain cancer cell lines. The purpose of this study was to define the activity of roscovitine in increasing radiosensitivity of human non-small cell lung cancer (NSCLC) cell line A549 cells in vitro. A549...

  18. Adamantyl Retinoid-Related Molecules Induce Apoptosis in Pancreatic Cancer Cells by Inhibiting IGF-1R and Wnt/β-Catenin Pathways

    Directory of Open Access Journals (Sweden)

    Lulu Farhana


    Full Text Available Pancreatic carcinoma has a dismal prognosis as it often presents as locally advanced or metastatic. We have found that exposure to adamantyl-substituted retinoid-related (ARR compounds 3-Cl-AHPC and AHP3 resulted in growth inhibition and apoptosis induction in PANC-1, Capan-2, and MiaPaCa-2 pancreatic cancer cell lines. In addition, AHP3 and 3-Cl-AHPC inhibited growth and induced apoptosis in spheres derived from the CD44+/CD24+ (CD133+/EpCAM+ stem-like cell population isolated from the pancreatic cancer cell lines. 3-Cl-AHPC-induced apoptosis was preceded by decreasing expression of IGF-1R, cyclin D1, β-catenin, and activated Notch-1 in the pancreatic cancer cell lines. Decreased IGF-1R expression inhibited PANC-1 proliferation, enhanced 3-Cl-AHPC-mediated apoptosis, and significantly decreased sphere formation. 3-Cl-AHPC inhibited the Wnt/β-catenin pathway as indicated by decreased β-catenin nuclear localization and inhibited Wnt/β-catenin activation of transcription factor TCF/LEF. Knockdown of β-catenin using sh-RNA also induced apoptosis and inhibited growth in pancreatic cancer cells. Thus, 3-Cl-AHPC and AHP3 induce apoptosis in pancreatic cancer cells and cancer stem-like cells and may serve as an important potential therapeutic agent in the treatment of pancreatic cancer.

  19. Reversal by EGTA of the enhanced secretory responsiveness of mast cells due to treatment with ouabain

    DEFF Research Database (Denmark)

    Johansen, Torben; Knudsen, T; Bertelsen, Niels Haldor


    The effect of EGTA on the enhancement by ouabain of compound 48/80-induced secretion from mast cells was compared with the effect on the Na(+)-K+ pump activity. The time-dependent secretory enhancement by ouabain was blocked by addition of EGTA to the cell suspension concomitantly with the addition...... of ouabain, and EGTA caused a large increase in the pump activity. Addition of 10 microM EGTA to ouabain-treated cells stopped but did not reverse the enhancement. The experiments show that the effect of ouabain was due to changes in a calcium pool utilized in compound 48/80-induced secretion following...

  20. Enhancement of radiosensitivity by CpG-oligodeoxyribonucleotide-7909 in human non-small cell lung cancer A549 cells. (United States)

    Zha, Lin; Qiao, Tiankui; Yuan, Sujuan; Lei, Linjie


    CpG-oligodeoxyribonucleotides (CpG-ODNs), which induce signaling through the toll-like receptor 9, are currently under investigation as immunity stimulators against cancer. It has recently been suggested that CpG-ODNs may also enhance sensitivity to traditional therapies including chemotherapy in certain cancer-cell lines. The purpose of this study was to define the activity of CpG-ODN7909 in increasing radiosensitivity of the human non-small cell lung cancer cell line A549 in vitro. First, a dose- and time-dependent inhibitory effect on cell viability was observed after A549 cells were treated with different concentrations of CpG-ODN7909 (5, 10, 30, and 60 microg/mL). Second, decreased cell clonogenic survival, enhanced cell apoptotic index, accumulated percentage of cells in the G2/M phase, and increased tumor necrosis factor (TNF)-alpha secretion were found after combined treatments with 10 microg/mL of CpG-ODN7909 and radiation compared to either treatment alone (p CpG-ODN7909 can increase the radiosensitivity of human non-small cell lung cancer A549 cells, which may be associated with reduced cell clonogenic survival, enhanced apoptosis, prolonged cell-cycle arrest in G2/M, and stimulation of TNF-alpha secretion.

  1. Performance enhancement of PV cells through micro-channel cooling

    Directory of Open Access Journals (Sweden)

    Muzaffar Ali


    Full Text Available Efficiency of a PV cell is strongly dependent on its surface temperature. The current study is focused