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Sample records for cultured cancer cell

  1. Principles of cancer cell culture.

    Science.gov (United States)

    Cree, Ian A

    2011-01-01

    The basics of cell culture are now relatively common, though it was not always so. The pioneers of cell culture would envy our simple access to manufactured plastics, media and equipment for such studies. The prerequisites for cell culture are a well lit and suitably ventilated laboratory with a laminar flow hood (Class II), CO(2) incubator, benchtop centrifuge, microscope, plasticware (flasks and plates) and a supply of media with or without serum supplements. Not only can all of this be ordered easily over the internet, but large numbers of well-characterised cell lines are available from libraries maintained to a very high standard allowing the researcher to commence experiments rapidly and economically. Attention to safety and disposal is important, and maintenance of equipment remains essential. This chapter should enable researchers with little prior knowledge to set up a suitable laboratory to do basic cell culture, but there is still no substitute for experience within an existing well-run laboratory.

  2. Contributions of 3D Cell Cultures for Cancer Research.

    Science.gov (United States)

    Ravi, Maddaly; Ramesh, Aarthi; Pattabhi, Aishwarya

    2017-10-01

    Cancer cell lines have contributed immensely in understanding the complex physiology of cancers. They are excellent material for studies as they offer homogenous samples without individual variations and can be utilised with ease and flexibility. Also, the number of assays and end-points one can study is almost limitless; with the advantage of improvising, modifying or altering several variables and methods. Literally, a new dimension to cancer research has been achieved by the advent of 3Dimensional (3D) cell culture techniques. This approach increased many folds the ways in which cancer cell lines can be utilised for understanding complex cancer biology. 3D cell culture techniques are now the preferred way of using cancer cell lines to bridge the gap between the 'absolute in vitro' and 'true in vivo'. The aspects of cancer biology that 3D cell culture systems have contributed include morphology, microenvironment, gene and protein expression, invasion/migration/metastasis, angiogenesis, tumour metabolism and drug discovery, testing chemotherapeutic agents, adaptive responses and cancer stem cells. We present here, a comprehensive review on the applications of 3D cell culture systems for these aspects of cancers. J. Cell. Physiol. 232: 2679-2697, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  3. 5-Fluorouracil-induced apoptosis in cultured oral cancer cells.

    Science.gov (United States)

    Tong, D; Poot, M; Hu, D; Oda, D

    2000-03-01

    Chemotherapy is commonly used to treat advanced oral squamous cell carcinoma (SCC) and is known to kill cancer cells through apoptosis. Our hypothesis states that 5-fluorouracil (5FU) also kills cultured oral epithelial cells through programmed cell death or apoptosis. Cultured oral cancer cells were exposed to an optimum dose of 20 mg/ml of 5FU. Cells were analyzed for changes in cell cycle distribution and induction of cell death including apoptosis. Normal control, human papilloma virus-immortalized (PP), ATCC SCC cell line (CA1) and two primary oral SCC cell lines (CA3 and -4) were studied. Inhibition of apoptosis by a pan-caspase inhibitor was used. SYTO 11 flow cytometry showed increased apoptosis in all 5FU-treated cell cultures compared to untreated controls. The results show biological variation in apoptotic response. CA1 had the lowest apoptotic rate of the cancer cell lines at 1.5%. Next lowest was CA3, followed by CA4 and PP. In addition, alteration in the G1 and S phase fractions were found. Untreated CA1 showed 28% G1, 53% S compared to 43% G1, and 40% S of treated. We investigated the pathway of apoptosis using the pan-caspase inhibitor IDN-1529 by methylthiazolyl diphenyl tetrazolium bromide (MTT) colorimetric analysis. Results showed mild inhibition of cell death when cells were incubated with 50 microM IDN-1529 for 24 h. This suggests a probable caspase-dependent apoptotic pathway. In conclusion, our data suggest that 5FU induces oral cancer cell death through apoptosis and that biological variation exists between normal and cancer cells and between different types of cancer cells themselves. Our data indicate that cultures of a useful in vitro model for chemosensitivity assays are possible. Our results also suggest a caspase-dependent pathway for chemocytotoxicity in oral SCC.

  4. Sulphur XANES Analysis of Cultured Human Prostate Cancer Cells

    International Nuclear Information System (INIS)

    Kwiatek, W.M.; Podgorczyk, M.; Paluszkiewicz, Cz.; Balerna, A.; Kisiel, A.

    2008-01-01

    Prostate cancer is one of the most commonly diagnosed cancers in men throughout the world. It is believed that changes to the structure of protein binding sites, altering its metabolism, may play an important role in carcinogenesis. Sulphur, often present in binding sites, can influence such changes through its chemical speciation. Hence there is a need for precise investigation of coordination environment of sulphur. X-ray absorption near edge structure spectroscopy offers such possibility. Cell culture samples offer histologically well defined areas of good homogeneity, suitable for successful and reliable X-ray absorption near edge structure analysis. This paper presents sulphur speciation data collected from three different human prostate cancer cell lines (PC-3, LNCaP and DU-145). Sulphur X-ray absorption near edge structure analysis was performed on K-edge structure. The spectra of cells were compared with those of cancerous tissue and with organic substances as well as inorganic compounds. (authors)

  5. Establishment of Cancer Stem Cell Cultures from Human Conventional Osteosarcoma.

    Science.gov (United States)

    Palmini, Gaia; Zonefrati, Roberto; Mavilia, Carmelo; Aldinucci, Alessandra; Luzi, Ettore; Marini, Francesca; Franchi, Alessandro; Capanna, Rodolfo; Tanini, Annalisa; Brandi, Maria Luisa

    2016-10-14

    The current improvements in therapy against osteosarcoma (OS) have prolonged the lives of cancer patients, but the survival rate of five years remains poor when metastasis has occurred. The Cancer Stem Cell (CSC) theory holds that there is a subset of tumor cells within the tumor that have stem-like characteristics, including the capacity to maintain the tumor and to resist multidrug chemotherapy. Therefore, a better understanding of OS biology and pathogenesis is needed in order to advance the development of targeted therapies to eradicate this particular subset and to reduce morbidity and mortality among patients. Isolating CSCs, establishing cell cultures of CSCs, and studying their biology are important steps to improving our understanding of OS biology and pathogenesis. The establishment of human-derived OS-CSCs from biopsies of OS has been made possible using several methods, including the capacity to create 3-dimensional stem cell cultures under nonadherent conditions. Under these conditions, CSCs are able to create spherical floating colonies formed by daughter stem cells; these colonies are termed "cellular spheres". Here, we describe a method to establish CSC cultures from primary cell cultures of conventional OS obtained from OS biopsies. We clearly describe the several passages required to isolate and characterize CSCs.

  6. High-Throughput Cancer Cell Sphere Formation for 3D Cell Culture.

    Science.gov (United States)

    Chen, Yu-Chih; Yoon, Euisik

    2017-01-01

    Three-dimensional (3D) cell culture is critical in studying cancer pathology and drug response. Though 3D cancer sphere culture can be performed in low-adherent dishes or well plates, the unregulated cell aggregation may skew the results. On contrary, microfluidic 3D culture can allow precise control of cell microenvironments, and provide higher throughput by orders of magnitude. In this chapter, we will look into engineering innovations in a microfluidic platform for high-throughput cancer cell sphere formation and review the implementation methods in detail.

  7. Effect of primarily cultured human lung cancer-associated fibroblasts on radiosensitivity of lung cancer cells

    International Nuclear Information System (INIS)

    Ji Xiaoqin; Ji Jiang; Chen Yongbing; Shan Fang; Lu Xueguan

    2014-01-01

    Objective: To investigate the effect of human lung cancer-associated fibroblasts (CAF) on the radiosensitivity of lung cancer cells when CAF is placed in direct contact co-culture with lung cancer cells. Methods: Human lung CAF was obtained from fresh human lung adenocarcinoma tissue specimens by primary culture and subculture and was then identified by immunofluorescence staining. The CAF was placed in direct contact co-culture with lung cancer A 549 and H 1299 cells, and the effects of CAF on the radiosensitivity of A 549 and H 1299 cells were evaluated by colony-forming assay. Results: The human lung CAF obtained by adherent culture could stably grow and proliferate, and it had specific expression of α-smooth muscle actin, vimentin, and fibroblast activation protein,but without expression of cytokeratin-18. The plating efficiency (PE, %) of A 549 cells at 0 Gy irradiation was (20.0 ± 3.9)% when cultured alone versus (32.3 ± 5.5)% when co-cultured with CAF (t=3.16, P<0.05), and the PE of H 1299 cells at 0 Gy irradiation was (20.6 ± 3.1)% when cultured alone versus (35.2 ± 2.3)% when co-cultured with CAF (t=6.55, P<0.05). The cell survival rate at 2 Gy irradiation (SF 2 ) of A 549 cells was 0.727 ±0.061 when cultured alone versus 0.782 ± 0.089 when co-cultured with CAF (t=0.88, P>0.05), and the SF 2 of H 1299 cells was 0.692 ±0.065 when cultured alone versus 0.782 ± 0.037 when co-cultured with CAF (t=2.08, P>0.05). The protection enhancement ratios of human lung CAF for A 549 cells and H 1299 cells were 1.29 and 1.25, respectively. Conclusions: Human lung CAF reduces the radiosensitivity of lung cancer cells when placed in direct contact co-culture with them, and the radioprotective effect may be attributed to CAF promoting the proliferation of lung cancer cells. (authors)

  8. Sequential cancer mutations in cultured human intestinal stem cells

    NARCIS (Netherlands)

    Drost, Jarno; van Jaarsveld, Richard H.; Ponsioen, Bas; Zimberlin, Cheryl; van Boxtel, Ruben; Buijs, Arjan; Sachs, Norman; Overmeer, René M.; Offerhaus, G. Johan; Begthel, Harry; Korving, Jeroen; van de Wetering, Marc; Schwank, Gerald; Logtenberg, Meike; Cuppen, Edwin; Snippert, Hugo J.; Medema, Jan Paul; Kops, Geert J. P. L.; Clevers, Hans

    2015-01-01

    Crypt stem cells represent the cells of origin for intestinal neoplasia. Both mouse and human intestinal stem cells can be cultured in medium containing the stem-cell-niche factors WNT, R-spondin, epidermal growth factor (EGF) and noggin over long time periods as epithelial organoids that remain

  9. Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture

    Directory of Open Access Journals (Sweden)

    Magnusson Magnus K

    2010-07-01

    Full Text Available Abstract Background Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D co-culture assay. Methods Breast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy. Results In co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors. Conclusion Breast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.

  10. Peptide Hydrogelation and Cell Encapsulation for 3D Culture of MCF-7 Breast Cancer Cells

    Science.gov (United States)

    Sun, Xiuzhi S.; Nguyen, Thu A.

    2013-01-01

    Three-dimensional (3D) cell culture plays an invaluable role in tumor biology by providing in vivo like microenviroment and responses to therapeutic agents. Among many established 3D scaffolds, hydrogels demonstrate a distinct property as matrics for 3D cell culture. Most of the existing pre-gel solutions are limited under physiological conditions such as undesirable pH or temperature. Here, we report a peptide hydrogel that shows superior physiological properties as an in vitro matrix for 3D cell culture. The 3D matrix can be accomplished by mixing a self-assembling peptide directly with a cell culture medium without any pH or temperature adjustment. Results of dynamic rheological studies showed that this hydrogel can be delivered multiple times via pipetting without permanently destroying the hydrogel architecture, indicating the deformability and remodeling ability of the hydrogel. Human epithelial cancer cells, MCF-7, are encapsulated homogeneously in the hydrogel matrix during hydrogelation. Compared with two-dimensional (2D) monolayer culture, cells residing in the hydrogel matrix grow as tumor-like clusters in 3D formation. Relevant parameters related to cell morphology, survival, proliferation, and apoptosis were analyzed using MCF-7 cells in 3D hydrogels. Interestingly, treatment of cisplatin, an anti-cancer drug, can cause a significant decrease of cell viability of MCF-7 clusters in hydrogels. The responses to cisplatin were dose- and time-dependent, indicating the potential usage of hydrogels for drug testing. Results of confocal microscopy and Western blotting showed that cells isolated from hydrogels are suitable for downstream proteomic analysis. The results provided evidence that this peptide hydrogel is a promising 3D cell culture material for drug testing. PMID:23527204

  11. The usefulness of three-dimensional cell culture in induction of cancer stem cells from esophageal squamous cell carcinoma cell lines

    International Nuclear Information System (INIS)

    Fujiwara, Daisuke; Kato, Kazunori; Nohara, Shigeo; Iwanuma, Yoshimi; Kajiyama, Yoshiaki

    2013-01-01

    Highlights: •Spheroids were created from esophageal carcinoma cells using NanoCulture® Plates. •The proportion of strongly ALDH-positive cells increased in 3-D culture. •Expression of cancer stem cell-related genes was enhanced in 3-D culture. •CA-9 expression was enhanced, suggesting hypoxia had been induced in 3-D culture. •Drug resistance was increased. 3-D culture is useful for inducing cancer stem cells. -- Abstract: In recent years, research on resistance to chemotherapy and radiotherapy in cancer treatment has come under the spotlight, and researchers have also begun investigating the relationship between resistance and cancer stem cells. Cancer stem cells are assumed to be present in esophageal cancer, but experimental methods for identification and culture of these cells have not yet been established. To solve this problem, we created spheroids using a NanoCulture® Plate (NCP) for 3-dimensional (3-D) cell culture, which was designed as a means for experimentally reproducing the 3-D structures found in the body. We investigated the potential for induction of cancer stem cells from esophageal cancer cells. Using flow cytometry we analyzed the expression of surface antigen markers CD44, CD133, CD338 (ABCG2), CD318 (CDCP1), and CD326 (EpCAM), which are known cancer stem cell markers. None of these surface antigen markers showed enhanced expression in 3-D cultured cells. We then analyzed aldehyde dehydrogenase (ALDH) enzymatic activity using the ALDEFLUOR reagent, which can identify immature cells such as stem cells and precursor cells. 3-D-cultured cells were strongly positive for ALDH enzyme activity. We also analyzed the expression of the stem cell-related genes Sox-2, Nanog, Oct3/4, and Lin28 using RT-PCR. Expression of Sox-2, Nanog, and Lin28 was enhanced. Analysis of expression of the hypoxic surface antigen marker carbonic anhydrase-9 (CA-9), which is an indicator of cancer stem cell induction and maintenance, revealed that CA-9 expression

  12. Metabolic cooperation between co-cultured lung cancer cells and lung fibroblasts.

    Science.gov (United States)

    Koukourakis, Michael I; Kalamida, Dimitra; Mitrakas, Achilleas G; Liousia, Maria; Pouliliou, Stamatia; Sivridis, Efthimios; Giatromanolaki, Alexandra

    2017-11-01

    Cooperation of cancer cells with stromal cells, such as cancer-associated fibroblasts (CAFs), has been revealed as a mechanism sustaining cancer cell survival and growth. In the current study, we focus on the metabolic interactions of MRC5 lung fibroblasts with lung cancer cells (A549 and H1299) using co-culture experiments and studying changes of the metabolic protein expression profile and of their growth and migration abilities. Using western blotting, confocal microscopy and RT-PCR, we observed that in co-cultures MRC5 respond by upregulating pyruvate dehydrogenase (PDH) and the monocarboxylate transporter MCT1. In contrast, cancer cells increase the expression of glucose transporters (GLUT1), LDH5, PDH kinase and the levels of phosphorylated/inactivated pPDH. H1299 cells growing in the same culture medium with fibroblasts exhibit a 'metastasis-like' phenomenon by forming nests within the fibroblast area. LDH5 and pPDH were drastically upregulated in these nests. The growth rate of both MRC5 and cancer cells increased in co-cultures. Suppression of LDHA or PDK1 in cancer cells abrogates the stimulatory signal from cancer cells to fibroblasts. Incubation of MRC5 fibroblasts with lactate resulted in an increase of LDHB and of PDH expression. Silencing of PDH gene in fibroblasts, or silencing of PDK1 or LDHA gene in tumor cells, impedes cancer cell's migration ability. Overall, a metabolic cooperation between lung cancer cells and fibroblasts has been confirmed in the context of direct Warburg effect, thus the fibroblasts reinforce aerobic metabolism to support the intensified anaerobic glycolytic pathways exploited by cancer cells.

  13. Electrospinning PCL Scaffolds Manufacture for Three-Dimensional Breast Cancer Cell Culture

    Directory of Open Access Journals (Sweden)

    Marc Rabionet

    2017-08-01

    Full Text Available In vitro cell culture is traditionally performed within two-dimensional (2D environments, providing a quick and cheap way to study cell properties in a laboratory. However, 2D systems differ from the in vivo environment and may not mimic the physiological cell behavior realistically. For instance, 2D culture models are thought to induce cancer stem cells (CSCs differentiation, a rare cancer cell subpopulation responsible for tumor initiation and relapse. This fact hinders the development of therapeutic strategies for tumors with a high relapse percentage, such as triple negative breast cancer (TNBC. Thus, three-dimensional (3D scaffolds have emerged as an attractive alternative to monolayer culture, simulating the extracellular matrix structure and maintaining the differentiation state of cells. In this work, scaffolds were fabricated through electrospinning different poly(ε-caprolactone-acetone solutions. Poly(ε-caprolactone (PCL meshes were seeded with triple negative breast cancer (TNBC cells and 15% PCL scaffolds displayed significantly (p < 0.05 higher cell proliferation and elongation than the other culture systems. Moreover, cells cultured on PCL scaffolds exhibited higher mammosphere forming capacity and aldehyde dehydrogenase activity than 2D-cultured cells, indicating a breast CSCs enrichment. These results prove the powerful capability of electrospinning technology in terms of poly(ε-caprolactone nanofibers fabrication. In addition, this study has demonstrated that electrospun 15% PCL scaffolds are suitable tools to culture breast cancer cells in a more physiological way and to expand the niche of breast CSCs. In conclusion, three-dimensional cell culture using PCL scaffolds could be useful to study cancer stem cell behavior and may also trigger the development of new specific targets against such malignant subpopulation.

  14. Suspension culture combined with chemotherapeutic agents for sorting of breast cancer stem cells

    International Nuclear Information System (INIS)

    Li, Hai-zhi; Yi, Tong-bo; Wu, Zheng-yan

    2008-01-01

    Cancer stem cell (CSC) hypothesis has not been well demonstrated by the lack of the most convincing evidence concerning a single cell capable of giving rise to a tumor. The scarcity in quantity and improper approaches for isolation and purification of CSCs have become the major obstacles for great development in CSCs. Here we adopted suspension culture combined with anticancer regimens as a strategy for screening breast cancer stem cells (BrCSCs). BrCSCs could survive and be highly enriched in non-adherent suspension culture while chemotherapeutic agents could destroy most rapidly dividing cancer cells and spare relatively quiescent BrCSCs. TM40D murine breast cancer cells were cultured in serum-free medium. The expression of CD44 + CD24 - was measured by flow cytometry. Cells of passage 10 were treated in combination with anticancer agents pacilitaxel and epirubicin at different peak plasma concentrations for 24 hours, and then maintained under suspension culture. The rate of apoptosis was examined by flow cytometry with Annexin-V fluorescein isothiocyanate (FITC)/propidium iodide (PI) double staining method. Selected cells in different amounts were injected subcutaneously into BALB/C mice to observe tumor formation. Cells of passage 10 in suspension culture had the highest percentage of CD44 + CD24 - (about 77 percent). A single tumor cell in 0.35 PPC could generate tumors in 3 of 20 BALB/C mice. Suspension culture combined with anticancer regimens provides an effective means of isolating, culturing and purifying BrCSCs

  15. Breast Cancer Stem Cell Culture and Enrichment Using Poly(ε-Caprolactone Scaffolds

    Directory of Open Access Journals (Sweden)

    Sònia Palomeras

    2016-04-01

    Full Text Available The cancer stem cell (CSC population displays self-renewal capabilities, resistance to conventional therapies, and a tendency to post-treatment recurrence. Increasing knowledge about CSCs’ phenotype and functions is needed to investigate new therapeutic strategies against the CSC population. Here, poly(ε-caprolactone (PCL, a biocompatible polymer free of toxic dye, has been used to fabricate scaffolds, solid structures suitable for 3D cancer cell culture. It has been reported that scaffold cell culture enhances the CSCs population. A RepRap BCN3D+ printer and 3 mm PCL wire were used to fabricate circular scaffolds. PCL design and fabrication parameters were first determined and then optimized considering several measurable variables of the resulting scaffolds. MCF7 breast carcinoma cell line was used to assess scaffolds adequacy for 3D cell culture. To evaluate CSC enrichment, the Mammosphere Forming Index (MFI was performed in 2D and 3D MCF7 cultures. Results showed that the 60° scaffolds were more suitable for 3D culture than the 45° and 90° ones. Moreover, 3D culture experiments, in adherent and non-adherent conditions, showed a significant increase in MFI compared to 2D cultures (control. Thus, 3D cell culture with PCL scaffolds could be useful to improve cancer cell culture and enrich the CSCs population.

  16. Simultaneous Expression of Cancer Stem Cell-Like Properties and Cancer-Associated Fibroblast-Like Properties in a Primary Culture of Breast Cancer Cells

    Energy Technology Data Exchange (ETDEWEB)

    Ishikawa, Mami; Inoue, Takahiro; Shirai, Takuma; Takamatsu, Kazuhiko; Kunihiro, Shiori; Ishii, Hirokazu [Frontiers of Innovative Research in Science and Technology (FIRST), Konan University, Kobe 650-0047 (Japan); Nishikata, Takahito, E-mail: nisikata@konan-u.ac.jp [Frontiers of Innovative Research in Science and Technology (FIRST), Konan University, Kobe 650-0047 (Japan); Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, Kobe 650-0047 (Japan)

    2014-07-31

    The importance of cancer-associated fibroblasts (CAFs) in cancer biology has been recently highlighted owing to their critical roles in cancer growth, progression, metastasis, and therapeutic resistance. We have previously established a primary culture of breast cancer cells, which showed epithelial-mesenchymal transition and cancer stem cell-like properties. In this study, we found that the primary culture also showed CAF-like properties. For example, hypoxia inducible factor 1α (HIF1A) and its downstream genes, nuclear factor-kappa B2 (NF-κB2) and BCL2/adenovirus E1B 19 kd-interacting protein 3 (BNIP3), and many enzymes involved in glycolysis, such as GAPDH, LDH, PGAM1, and PKM2, were highly overexpressed in the primary culture. Moreover, media conditioned with the primary culture cells enhanced the growth of breast cancer cells. Similar to previous CAF studies, this enhancement suggested to be occurred through fibroblast growth factor signaling. This MCKH primary culture cell, which showed simultaneous expression of tumorigenic and CAF properties, offers a unique experimental system for studying the biology of CAFs.

  17. Optimizing structural and mechanical properties of cryogel scaffolds for use in prostate cancer cell culturing

    International Nuclear Information System (INIS)

    Cecilia, A.; Baecker, A.; Hamann, E.; Rack, A.; Kamp, T. van de; Gruhl, F.J.; Hofmann, R.; Moosmann, J.; Hahn, S.; Kashef, J.; Bauer, S.; Farago, T.; Helfen, L.

    2017-01-01

    Prostate cancer (PCa) currently is the second most diagnosed cancer in men and the second most cause of cancer death after lung cancer in Western societies. This sets the necessity of modelling prostatic disorders to optimize a therapy against them. The conventional approach to investigating prostatic diseases is based on two-dimensional (2D) cell culturing. This method, however, does not provide a three-dimensional (3D) environment, therefore impeding a satisfying simulation of the prostate gland in which the PCa cells proliferate. Cryogel scaffolds represent a valid alternative to 2D culturing systems for studying the normal and pathological behavior of the prostate cells thanks to their 3D pore architecture that reflects more closely the physiological environment in which PCa cells develop. In this work the 3D morphology of three potential scaffolds for PCa cell culturing was investigated by means of synchrotron X-ray computed micro tomography (SXCμT) fitting the according requirements of high spatial resolution, 3D imaging capability and low dose requirements very well. In combination with mechanical tests, the results allowed identifying an optimal cryogel architecture, meeting the needs for a well-suited scaffold to be used for 3D PCa cell culture applications. The selected cryogel was then used for culturing prostatic lymph node metastasis (LNCaP) cells and subsequently, the presence of multi-cellular tumor spheroids inside the matrix was demonstrated again by using SXCμT. - Highlights: • Synthesis of cryogel scaffolds for prostate cancer cell culturing. • Study of cryogel morphology by synchrotron X-ray computed micro tomography. • Analysis of cryogel mechanical properties with laboratory techniques. • Culturing of prostate cancer cell in the optimal cryogel composition for 21 days. • 3D visualization of the cells by synchrotron X-ray computed micro tomography.

  18. Optimizing structural and mechanical properties of cryogel scaffolds for use in prostate cancer cell culturing

    Energy Technology Data Exchange (ETDEWEB)

    Cecilia, A. [Institute for Photon Science and Synchrotron Radiation (IPS), Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, D-76344 Eggenstein-Leopoldshafen (Germany); Baecker, A. [Institute of Microstructure Technology (IMT), Karlsruhe Institute of Technology (KIT), Hermann-von-Helmholtz-Platz 1 Bldg 329, Eggenstein-Leopoldshafen, Karlsruhe D-76344 (Germany); Hamann, E. [Institute for Photon Science and Synchrotron Radiation (IPS), Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, D-76344 Eggenstein-Leopoldshafen (Germany); Rack, A. [European Synchrotron Radiation Facility (ESRF), 6 rue Jules Horowitz, 38000 Grenoble (France); Kamp, T. van de [Laboratory for Applications of Synchrotron Radiation (LAS), Karlsruhe Institute of Technology, 6980, D-76128 Karlsruhe (Germany); Gruhl, F.J. [Institute of Microstructure Technology (IMT), Karlsruhe Institute of Technology (KIT), Hermann-von-Helmholtz-Platz 1 Bldg 329, Eggenstein-Leopoldshafen, Karlsruhe D-76344 (Germany); Hofmann, R. [Institute for Photon Science and Synchrotron Radiation (IPS), Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, D-76344 Eggenstein-Leopoldshafen (Germany); Moosmann, J. [Institute of Materials Research, Helmholtz-Zentrum Geesthacht (HZG), Max-Planck-Str. 1, D-21502 Geesthacht (Germany); Hahn, S.; Kashef, J.; Bauer, S.; Farago, T. [Institute for Photon Science and Synchrotron Radiation (IPS), Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, D-76344 Eggenstein-Leopoldshafen (Germany); Helfen, L. [Institute for Photon Science and Synchrotron Radiation (IPS), Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, D-76344 Eggenstein-Leopoldshafen (Germany); European Synchrotron Radiation Facility (ESRF), 6 rue Jules Horowitz, 38000 Grenoble (France); and others

    2017-02-01

    Prostate cancer (PCa) currently is the second most diagnosed cancer in men and the second most cause of cancer death after lung cancer in Western societies. This sets the necessity of modelling prostatic disorders to optimize a therapy against them. The conventional approach to investigating prostatic diseases is based on two-dimensional (2D) cell culturing. This method, however, does not provide a three-dimensional (3D) environment, therefore impeding a satisfying simulation of the prostate gland in which the PCa cells proliferate. Cryogel scaffolds represent a valid alternative to 2D culturing systems for studying the normal and pathological behavior of the prostate cells thanks to their 3D pore architecture that reflects more closely the physiological environment in which PCa cells develop. In this work the 3D morphology of three potential scaffolds for PCa cell culturing was investigated by means of synchrotron X-ray computed micro tomography (SXCμT) fitting the according requirements of high spatial resolution, 3D imaging capability and low dose requirements very well. In combination with mechanical tests, the results allowed identifying an optimal cryogel architecture, meeting the needs for a well-suited scaffold to be used for 3D PCa cell culture applications. The selected cryogel was then used for culturing prostatic lymph node metastasis (LNCaP) cells and subsequently, the presence of multi-cellular tumor spheroids inside the matrix was demonstrated again by using SXCμT. - Highlights: • Synthesis of cryogel scaffolds for prostate cancer cell culturing. • Study of cryogel morphology by synchrotron X-ray computed micro tomography. • Analysis of cryogel mechanical properties with laboratory techniques. • Culturing of prostate cancer cell in the optimal cryogel composition for 21 days. • 3D visualization of the cells by synchrotron X-ray computed micro tomography.

  19. Myofibroblast androgen receptor expression determines cell survival in co-cultures of myofibroblasts and prostate cancer cells in vitro.

    Science.gov (United States)

    Palethorpe, Helen M; Leach, Damien A; Need, Eleanor F; Drew, Paul A; Smith, Eric

    2018-04-10

    Fibroblasts express androgen receptor (AR) in the normal prostate and during prostate cancer development. We have reported that loss of AR expression in prostate cancer-associated fibroblasts is a poor prognostic indicator. Here we report outcomes of direct and indirect co-cultures of immortalised AR-positive (PShTert-AR) or AR-negative (PShTert) myofibroblasts with prostate cancer cells. In the initial co-cultures the AR-negative PC3 cell line was used so AR expression and signalling were restricted to the myofibroblasts. In both direct and indirect co-culture with PShTert-AR myofibroblasts, paracrine signalling to the PC3 cells slowed proliferation and induced apoptosis. In contrast, PC3 cells proliferated with PShTert myofibroblasts irrespective of the co-culture method. In direct co-culture PC3 cells induced apoptosis in and destroyed PShTerts by direct signalling. Similar results were seen in direct co-cultures with AR-negative DU145 and AR-positive LNCaP and C4-2B prostate cancer cell lines. The AR ligand 5α-dihydrotestosterone (DHT) inhibited the proliferation of the PShTert-AR myofibroblasts, thereby reducing the extent of their inhibitory effect on cancer cell growth. These results suggest loss of stromal AR would favour prostate cancer cell growth in vivo , providing an explanation for the clinical observation that reduced stromal AR is associated with a poorer outcome.

  20. Polymer microfilters with nanostructured surfaces for the culture of circulating cancer cells

    International Nuclear Information System (INIS)

    Makarova, Olga V.; Adams, Daniel L.; Divan, Ralu; Rosenmann, Daniel; Zhu, Peixuan; Li, Shuhong; Amstutz, Platte; Tang, Cha-Mei

    2016-01-01

    There is a critical need to improve the accuracy of drug screening and testing through the development of in vitro culture systems that more effectively mimic the in vivo environment. Surface topographical features on the nanoscale level, in short nanotopography, effect the cell growth patterns, and hence affect cell function in culture. We report the preliminary results on the fabrication, and subsequent cellular growth, of nanoscale surface topography on polymer microfilters using cell lines as a precursor to circulating tumor cells (CTCs). To create various nanoscale features on the microfilter surface, we used reactive ion etching (RIE) with and without an etching mask. An anodized aluminum oxide (AAO) membrane fabricated directly on the polymer surface served as an etching mask. Polymer filters with a variety of modified surfaces were used to compare the effects on the culture of cancer cell lines in blank culture wells, with untreated microfilters or with RIE-treated microfilters. We then report the differences of cell shape, phenotype and growth patterns of bladder and glioblastoma cancer cell lines after isolation on the various types of material modifications. Our data suggest that RIE modified polymer filters can isolate model cell lines while retaining ell viability, and that the RIE filter modification allows T24 monolayering cells to proliferate as a structured cluster. - Highlights: • Surface topographical effects the growth patterns and cell function of cancer cells • Nanoscale surface topography on polymer filters for circulating tumor cell culture • Membrane fabricated directly on polymer surfaces utilized for polymer etching • Nanotopography alters cell shape, phenotype and growth patterns of cancer cells • Nanoscale surface topography dictates monolayering or 3D structured cell culture

  1. Polymer microfilters with nanostructured surfaces for the culture of circulating cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Makarova, Olga V. [Creatv MicroTech, Inc., 2242 West Harrison St., Chicago 60612, IL (United States); Adams, Daniel L., E-mail: dan@creatvmicrotech.com [Creatv MicroTech, Inc., 1 Deer Park Drive, Monmouth Junction, NJ 08852 (United States); Divan, Ralu; Rosenmann, Daniel [Center for Nanoscale Materials, Argonne National Laboratory, 9700 South Cass Ave., Argonne 60439, IL (United States); Zhu, Peixuan; Li, Shuhong; Amstutz, Platte; Tang, Cha-Mei [Creatv MicroTech, Inc., 11609 Lake Potomac Drive, Potomac 20854, MD (United States)

    2016-09-01

    There is a critical need to improve the accuracy of drug screening and testing through the development of in vitro culture systems that more effectively mimic the in vivo environment. Surface topographical features on the nanoscale level, in short nanotopography, effect the cell growth patterns, and hence affect cell function in culture. We report the preliminary results on the fabrication, and subsequent cellular growth, of nanoscale surface topography on polymer microfilters using cell lines as a precursor to circulating tumor cells (CTCs). To create various nanoscale features on the microfilter surface, we used reactive ion etching (RIE) with and without an etching mask. An anodized aluminum oxide (AAO) membrane fabricated directly on the polymer surface served as an etching mask. Polymer filters with a variety of modified surfaces were used to compare the effects on the culture of cancer cell lines in blank culture wells, with untreated microfilters or with RIE-treated microfilters. We then report the differences of cell shape, phenotype and growth patterns of bladder and glioblastoma cancer cell lines after isolation on the various types of material modifications. Our data suggest that RIE modified polymer filters can isolate model cell lines while retaining ell viability, and that the RIE filter modification allows T24 monolayering cells to proliferate as a structured cluster. - Highlights: • Surface topographical effects the growth patterns and cell function of cancer cells • Nanoscale surface topography on polymer filters for circulating tumor cell culture • Membrane fabricated directly on polymer surfaces utilized for polymer etching • Nanotopography alters cell shape, phenotype and growth patterns of cancer cells • Nanoscale surface topography dictates monolayering or 3D structured cell culture.

  2. Short-term spheroid culture of primary colorectal cancer cells as an in vitro model for personalizing cancer medicine

    DEFF Research Database (Denmark)

    Jeppesen, Maria; Hagel, Grith; Glenthoj, Anders

    2017-01-01

    Chemotherapy treatment of cancer remains a challenge due to the molecular and functional heterogeneity displayed by tumours originating from the same cell type. The pronounced heterogeneity makes it difficult for oncologists to devise an effective therapeutic strategy for the patient. One approac...... and combinations most commonly used for treatment of colorectal cancer. In summary, short-term spheroid culture of primary colorectal adenocarcinoma cells represents a promising in vitro model for use in personalized medicine....... for increasing treatment efficacy is to test the chemosensitivity of cancer cells obtained from the patient's tumour. 3D culture represents a promising method for modelling patient tumours in vitro. The aim of this study was therefore to evaluate how closely short-term spheroid cultures of primary colorectal...... cancer cells resemble the original tumour. Colorectal cancer cells were isolated from human tumour tissue and cultured as spheroids. Spheroid cultures were established with a high success rate and remained viable for at least 10 days. The spheroids exhibited significant growth over a period of 7 days...

  3. Phenotypic characterization of prostate cancer LNCaP cells cultured within a bioengineered microenvironment.

    Directory of Open Access Journals (Sweden)

    Shirly Sieh

    Full Text Available Biophysical and biochemical properties of the microenvironment regulate cellular responses such as growth, differentiation, morphogenesis and migration in normal and cancer cells. Since two-dimensional (2D cultures lack the essential characteristics of the native cellular microenvironment, three-dimensional (3D cultures have been developed to better mimic the natural extracellular matrix. To date, 3D culture systems have relied mostly on collagen and Matrigel™ hydrogels, allowing only limited control over matrix stiffness, proteolytic degradability, and ligand density. In contrast, bioengineered hydrogels allow us to independently tune and systematically investigate the influence of these parameters on cell growth and differentiation. In this study, polyethylene glycol (PEG hydrogels, functionalized with the Arginine-glycine-aspartic acid (RGD motifs, common cell-binding motifs in extracellular matrix proteins, and matrix metalloproteinase (MMP cleavage sites, were characterized regarding their stiffness, diffusive properties, and ability to support growth of androgen-dependent LNCaP prostate cancer cells. We found that the mechanical properties modulated the growth kinetics of LNCaP cells in the PEG hydrogel. At culture periods of 28 days, LNCaP cells underwent morphogenic changes, forming tumor-like structures in 3D culture, with hypoxic and apoptotic cores. We further compared protein and gene expression levels between 3D and 2D cultures upon stimulation with the synthetic androgen R1881. Interestingly, the kinetics of R1881 stimulated androgen receptor (AR nuclear translocation differed between 2D and 3D cultures when observed by immunofluorescent staining. Furthermore, microarray studies revealed that changes in expression levels of androgen responsive genes upon R1881 treatment differed greatly between 2D and 3D cultures. Taken together, culturing LNCaP cells in the tunable PEG hydrogels reveals differences in the cellular responses to

  4. FGF1 and IGF1-conditioned 3D culture system promoted the amplification and cancer stemness of lung cancer cells.

    Science.gov (United States)

    Liu, Pengpeng; Zhang, Rui; Yu, Wenwen; Ye, Yingnan; Cheng, Yanan; Han, Lei; Dong, Li; Chen, Yongzi; Wei, Xiyin; Yu, Jinpu

    2017-12-01

    Lung cancer stem cells (LCSCs) are considered as the cellular origins of metastasis and relapse of lung cancer. However, routine two-dimensional culture system (2D-culture) hardly mimics the growth and functions of LCSCs in vivo and therefore significantly decreases the stemness activity of LCSCs. In this study, we constructed a special BME-based three-dimensional culture system (3D-culture) to amplify LCSCs in human lung adenocarcinoma cell line A549 cells and found 3D-culture promoted the enrichment and amplification of LCSCs in A549 cells displaying higher proliferation potential and invasion activity, but lower apoptosis. The expression and secretion levels of FGF1 and IGF1 were dramatically elevated in 3D-culture compared to 2D-culture. After growing in FGF1 and IGF1-conditioned 3D-culture, the proportion of LCSCs with specific stemness phenotypes in A549 cells significantly increased compared to that in conventional 3D suspension culture system. Further results indicated that FGF1 and IGF1 promoted the amplification and cancer stemness of LCSCs dependent on MAPK signaling pathway. Our data firstly established a growth factors-conditioned 3D-culture for LCSCs and demonstrated the effects of FGF1 and IGF1 in promoting the enrichment and amplification of LCSCs which might provide a feasible cell model in vitro for both mechanism study and translational research on lung cancer. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Polymer microfilters with nanostructured surfaces for the culture of circulating cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Makarova, Olga V.; Adams, Daniel L.; Divan, Ralu; Rosenmann, Daniel; Zhu, Peixuan; Li, Shuhong; Amstutz, Platte; Tang, Cha-Mei

    2016-09-01

    There is a critical need to improve the accuracy of drug screening and testing through the development of in vitro culture systems that more effectively mimic the in vivo environment. Surface topographical features on the nanoscale level, in short nanotopography, effect the cell growth patterns, and hence affect cell function in culture. We report the preliminary results on the fabrication, and subsequent cellular growth, of nanoscale surface topography on polymer microfilters using cell lines as a precursor to circulating tumor cells (CTCs). To create various nanoscale features on the microfilter surface, we used reactive ion etching (RIE) with and without an etching mask. An anodized aluminum oxide (AAO) membrane fabricated directly on the polymer surface served as an etching mask. Polymer filters with a variety of modified surfaces were used to compare the effects on the culture of cancer cell lines in blank culture wells, with untreated microfilters or with RIE-treated microfilters. We then report the differences of cell shape, phenotype and growth patterns of bladder and glioblastoma cancer cell lines after isolation on the various types of material modifications. Our data suggest that RIE modified polymer filters can isolate model cell lines while retaining ell viability, and that the RIE filter modification allows T24 monolayering cells to proliferate as a structured cluster. Copyright 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  6. Identification of cancer stem-like side population cells in purified primary cultured human laryngeal squamous cell carcinoma epithelia.

    Directory of Open Access Journals (Sweden)

    Chun-Ping Wu

    Full Text Available Cancer stem-like side population (SP cells have been identified in many solid tumors; however, most of these investigations are performed using established cancer cell lines. Cancer cells in tumor tissue containing fibroblasts and many other types of cells are much more complex than any cancer cell line. Although SP cells were identified in the laryngeal squamous cell carcinoma (LSCC cell line Hep-2 in our pilot study, it is unknown whether the LSCC tissue contains SP cells. In this study, LSCC cells (LSCCs were primary cultured and purified from a surgically resected LSCC specimen derived from a well-differentiated epiglottic neoplasm of a Chinese male. This was followed by the verification of epithelium-specific characteristics, such as ultrastructure and biomarkers. A distinct SP subpopulation (4.45±1.07% was isolated by Hoechst 33342 efflux analysis from cultured LSCCs by using a flow cytometer. Cancer stem cell (CSC-associated assays, including expression of self-renewal and CSC marker genes, proliferation, differentiation, spheroid formation, chemotherapy resistance, and tumorigenicity were then conducted between SP and non-SP (NSP LSCCs. In vitro and in vivo assays revealed that SP cells manifested preferential expression of self-renewal and CSC marker genes, higher capacity for proliferation, differentiation, and spheroid formation; enhanced resistance to chemotherapy; and greater xenograft tumorigenicity in immunodeficient mice compared with NSP cells. These findings suggest that the primary cultured and purified LSCCs contain cancer stem-like SP cells, which may serve as a valuable model for CSC research in LSCC.

  7. Biosynthesis of 14C-phytoene from tomato cell suspension cultures (Lycopersicon esculentum) for utilization in prostate cancer cell culture studies.

    Science.gov (United States)

    Campbell, Jessica K; Rogers, Randy B; Lila, Mary Ann; Erdman, John W

    2006-02-08

    This work describes the development and utilization of a plant cell culture production approach to biosynthesize and radiolabel phytoene and phytofluene for prostate cancer cell culture studies. The herbicide norflurazon was added to established cell suspension cultures of tomato (Lycopersicon esculentum cv. VFNT cherry), to induce the biosynthesis and accumulation of the lycopene precursors, phytoene and phytofluene, in their natural isomeric forms (15-cis-phytoene and two cis-phytofluene isomers). Norflurazon concentrations, solvent carrier type and concentration, and duration of culture exposure to norflurazon were screened to optimize phytoene and phytofluene synthesis. Maximum yields of both phytoene and phytofluene were achieved after 7 days of treatment with 0.03 mg norflurazon/40 mL fresh medium, provided in 0.07% solvent carrier. Introduction of 14C-sucrose to the tomato cell culture medium enabled the production of 14C-labeled phytoene for subsequent prostate tumor cell uptake studies. In DU 145 prostate tumor cells, it was determined that 15-cis-phytoene and an oxidized product of phytoene were taken up and partially metabolized by the cells. The ability to biosynthesize, radiolabel, and isolate these carotenoids from tomato cell cultures is a novel, valuable methodology for further in vitro and in vivo investigations into the roles of phytoene and phytofluene in cancer chemoprevention.

  8. Three-dimensional organoid culture reveals involvement of Wnt/β-catenin pathway in proliferation of bladder cancer cells.

    Science.gov (United States)

    Yoshida, Takahiro; Sopko, Nikolai A; Kates, Max; Liu, Xiaopu; Joice, Gregory; McConkey, David J; Bivalacqua, Trinity J

    2018-02-16

    There has been increasing awareness of the importance of three-dimensional culture of cancer cells. Tumor cells growing as multicellular spheroids in three-dimensional culture, alternatively called organoids, are widely believed to more closely mimic solid tumors in situ . Previous studies concluded that the Wnt/β-catenin pathway is required for regeneration of the normal urothelium after injury and that β-catenin is upregulated in human bladder cancers, but no clear evidence has been advanced to support the idea that the Wnt/β-catenin pathway is directly involved in deregulated proliferation and the other malignant characteristics of bladder cancer cells. Here we report that the Wnt/β-catenin pathway activator, CHIR99021, promoted proliferation of established human bladder cancer cell lines when they were grown in organoid culture but not when they were grown in conventional adherent cultures. CHIR99021 activated Wnt/β-catenin pathway in bladder cancer cell lines in organoid culture. CHIR99021 also stimulated proliferation and the Wnt/b-catenin pathway in primary human bladder cancer organoids. RNAi-mediated knockdown of β-catenin blocked growth of organoids. The effects of CHIR99021 were associated with decreased expression of the urothelial terminal differentiation marker, cytokeratin 20. Our data suggest that the Wnt/β-catenin pathway is required for the proliferation of bladder cancer cells in three-dimensional organoid culture and provide a concrete example of why organoid culture is important for cancer research.

  9. The Effect of Primary Cancer Cell Culture Models on the Results of Drug Chemosensitivity Assays: The Application of Perfusion Microbioreactor System as Cell Culture Vessel

    Science.gov (United States)

    Chen, Yi-Dao; Huang, Shiang-Fu; Wang, Hung-Ming

    2015-01-01

    To precisely and faithfully perform cell-based drug chemosensitivity assays, a well-defined and biologically relevant culture condition is required. For the former, a perfusion microbioreactor system capable of providing a stable culture condition was adopted. For the latter, however, little is known about the impact of culture models on the physiology and chemosensitivity assay results of primary oral cavity cancer cells. To address the issues, experiments were performed. Results showed that minor environmental pH change could significantly affect the metabolic activity of cells, demonstrating the importance of stable culture condition for such assays. Moreover, the culture models could also significantly influence the metabolic activity and proliferation of cells. Furthermore, the choice of culture models might lead to different outcomes of chemosensitivity assays. Compared with the similar test based on tumor-level assays, the spheroid model could overestimate the drug resistance of cells to cisplatin, whereas the 2D and 3D culture models might overestimate the chemosensitivity of cells to such anticancer drug. In this study, the 3D culture models with same cell density as that in tumor samples showed comparable chemosensitivity assay results as the tumor-level assays. Overall, this study has provided some fundamental information for establishing a precise and faithful drug chemosensitivity assay. PMID:25654105

  10. High-Throughput Cancer Cell Sphere Formation for Characterizing the Efficacy of Photo Dynamic Therapy in 3D Cell Cultures

    Science.gov (United States)

    Chen, Yu-Chih; Lou, Xia; Zhang, Zhixiong; Ingram, Patrick; Yoon, Euisik

    2015-07-01

    Photodynamic therapy (PDT), wherein light sensitive non-toxic agents are locally and selectively activated using light, has emerged as an appealing alternative to traditional cancer chemotherapy. Yet to date, PDT efficacy has been mostly characterized using 2D cultures. Compared to 2D cultures, 3D sphere culture generates unique spatial distributions of nutrients and oxygen for the cells that better mimics the in-vivo conditions. Using a novel polyHEMA (non-adherent polymer) fabrication process, we developed a microfluidic sphere formation platform that can (1) generate 1,024 uniform (size variation successfully characterized the different responses in 2D and 3D cell culture to PDT. Furthermore, we investigated the treatment resistance effect in cancer cells induced by tumor associated fibroblasts (CAF). Although the CAFs can enhance the resistance to traditional chemotherapy agents, no significant difference in PDT was observed. The preliminary results suggest that the PDT can be an attractive alternative cancer therapy, which is less affected by the therapeutic resistance induced by cancer associated cells.

  11. An imbalance in progenitor cell populations reflects tumour progression in breast cancer primary culture models

    LENUS (Irish Health Repository)

    Donatello, Simona

    2011-04-26

    Abstract Background Many factors influence breast cancer progression, including the ability of progenitor cells to sustain or increase net tumour cell numbers. Our aim was to define whether alterations in putative progenitor populations could predict clinicopathological factors of prognostic importance for cancer progression. Methods Primary cultures were established from human breast tumour and adjacent non-tumour tissue. Putative progenitor cell populations were isolated based on co-expression or concomitant absence of the epithelial and myoepithelial markers EPCAM and CALLA respectively. Results Significant reductions in cellular senescence were observed in tumour versus non-tumour cultures, accompanied by a stepwise increase in proliferation:senescence ratios. A novel correlation between tumour aggressiveness and an imbalance of putative progenitor subpopulations was also observed. Specifically, an increased double-negative (DN) to double-positive (DP) ratio distinguished aggressive tumours of high grade, estrogen receptor-negativity or HER2-positivity. The DN:DP ratio was also higher in malignant MDA-MB-231 cells relative to non-tumourogenic MCF-10A cells. Ultrastructural analysis of the DN subpopulation in an invasive tumour culture revealed enrichment in lipofuscin bodies, markers of ageing or senescent cells. Conclusions Our results suggest that an imbalance in tumour progenitor subpopulations imbalances the functional relationship between proliferation and senescence, creating a microenvironment favouring tumour progression.

  12. Mammalian Cell Culture Simplified.

    Science.gov (United States)

    Moss, Robert; Solomon, Sondra

    1991-01-01

    A tissue culture experiment that does not require elaborate equipment and that can be used to teach sterile technique, the principles of animal cell line maintenance, and the concept of cell growth curves is described. The differences between cancerous and normal cells can be highlighted. The procedure is included. (KR)

  13. Culture in embryonic kidney serum and xeno-free media as renal cell carcinoma and renal cell carcinoma cancer stem cells research model.

    Science.gov (United States)

    Krawczyk, Krzysztof M; Matak, Damian; Szymanski, Lukasz; Szczylik, Cezary; Porta, Camillo; Czarnecka, Anna M

    2018-04-01

    The use of fetal bovine serum hinders obtaining reproducible experimental results and should also be removed in hormone and growth factor studies. In particular hormones found in FBS act globally on cancer cell physiology and influence transcriptome and metabolome. The aim of our study was to develop a renal carcinoma serum free culture model optimized for (embryonal) renal cells in order to select the best study model for downstream auto-, para- or endocrine research. Secondary aim was to verify renal carcinoma stem cell culture for this application. In the study, we have cultured renal cell carcinoma primary tumour cell line (786-0) as well as human kidney cancer stem cells in standard 2D monolayer cultures in Roswell Park Memorial Institute Medium or Dulbecco's Modified Eagle's Medium and Complete Human Kidney Cancer Stem Cell Medium, respectively. Serum-free, animal-component free Human Embryonic Kidney 293 media were tested. Our results revealed that xeno-free embryonal renal cells optimized culture media provide a useful tool in RCC cancer biology research and at the same time enable effective growth of RCC. We propose bio-mimic RCC cell culture model with specific serum-free and xeno-free medium that promote RCC cell viability.

  14. Data of a fluorescent imaging-based analysis of anti-cancer drug effects on three-dimensional cultures of breast cancer cells

    Directory of Open Access Journals (Sweden)

    Junji Itou

    2015-12-01

    Full Text Available Three-dimensional (3D cell culture is a powerful tool to study cell growth under 3D condition. To perform a simple test for anti-cancer drugs in 3D culture, visualization of non-proliferated cells is required. We propose a fluorescent imaging-based assay to analyze cancer cell proliferation in 3D culture. We used a pulse-labeling technique with a photoconvertible fluorescent protein Kaede to identify non-proliferated cells. This assay allows us to observe change in cell proliferation in 3D culture by simple imaging. Using this assay, we obtained the data of the effects of anti-cancer drugs, 5-fluorouracil and PD0332991 in a breast cancer cell line, MCF-7.

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

    Directory of Open Access Journals (Sweden)

    Mehdi Shakibaei

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

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

    Science.gov (United States)

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

    2014-01-01

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

  17. Pronounced radiosensitization of cultured human cancer cells by COX inhibitor under acidic microenvironment

    International Nuclear Information System (INIS)

    Shah, Tushar; Ryu, Samuel; Lee, Ho Jun; Brown, Stephen; Kim, Jae Ho

    2002-01-01

    Purpose: To demonstrate the influence of pH on the cytotoxicity and radiosensitization by COX (cyclooxygenase) -1 and -2 inhibitors using established human cancer cells in culture. Methods and Materials: Nonselective COX inhibitor, ibuprofen (IB), and selective COX-2 inhibitor, SC-236, were used to determine the cytotoxicity and radiosensitization at varying pH of culture media. Human colon carcinoma cell line (HT-29) was exposed to the drug alone and in combination with radiation at different pH of the cell culture media. The end point was clonogenic ability of the single-plated cells after the treatment. Results: Cytotoxicity and radiosensitization of IB increased with higher drug concentration and longer exposure time. The most significant radiosensitization was seen with IB (1.5 mM) for 2-h treatment at pH 6.7 before irradiation. The dose-modifying factor as defined by the ratio of radiation doses required to achieve the same effect on cell survival was 1.8 at 10% survival level. In contrast, SC-236 (50 μM for 2-8 h) showed no pH-dependent cytotoxicity. There was modest increase in the cell killing at lower doses of radiation. Conclusion: An acidic pH was an important factor affecting the increased cytotoxicity and radiosensitization by ibuprofen. Radiation response was enhanced at shoulder portion of the cell survival curve by selective COX-2 inhibitor

  18. 3D culture of Her2+ breast cancer cells promotes AKT to MAPK switching and a loss of therapeutic response.

    Science.gov (United States)

    Gangadhara, Sharath; Smith, Chris; Barrett-Lee, Peter; Hiscox, Stephen

    2016-06-01

    The Her2 receptor is overexpressed in up to 25 % of breast cancers and is associated with a poor prognosis. Around half of Her2+ breast cancers also express the estrogen receptor and treatment for such tumours can involve both endocrine and Her2-targeted therapies. However, despite preclinical data supporting the effectiveness of these agents, responses can vary widely in the clinical setting. In light of the increasing evidence pointing to the interplay between the tumour and its extracellular microenvironment as a significant determinant of therapeutic sensitivity and response here we investigated the impact of 3D matrix culture of breast cancer cells on their therapeutic sensitivity. A 3D Matrigel-based culture system was established and optimized for the growth of ER+/Her2+ breast cancer cell models. Growth of cells in response to trastuzumab and endocrine agents in 3D culture versus routine monolayer culture were assessed using cell counting and Ki67 staining. Endogenous and trastuzumab-modulated signalling pathway activity in 2D and 3D cultures were assessed using Western blotting. Breast cancer cells in 3D culture displayed an attenuated response to both endocrine agents and trastuzumab compared with cells cultured in traditional 2D monolayers. Underlying this phenomenon was an apparent matrix-induced shift from AKT to MAPK signalling; consequently, suppression of MAPK in 3D cultures restores therapeutic response. These data suggest that breast cancer cells in 3D culture display a reduced sensitivity to therapeutic agents which may be mediated by internal MAPK-mediated signalling. Targeting of adaptive pathways that maintain growth in 3D culture may represent an effective strategy to improve therapeutic response clinically.

  19. Differences of statin activity in 2D and 3D pancreatic cancer cell cultures

    Directory of Open Access Journals (Sweden)

    Paškevičiūtė M

    2017-11-01

    Full Text Available Miglė Paškevičiūtė,1 Vilma Petrikaitė1,21Department of Drug Chemistry, Faculty of Pharmacy, Lithuanian University of Health Sciences, Kaunas, Lithuania; 2Department of Biothermodynamics and Drug Design, Vilnius University Institute of Biotechnology, Vilnius, LithuaniaPurpose: To evaluate the anticancer activity of lovastatin (LOVA, mevastatin (MEVA, pitavastatin (PITA, and simvastatin (SIMVA in 2D and 3D models of three human pancreatic cancer cell lines (BxPC-3, MIA PaCa-2, and PANC-1.Methods: The effect of statins on cell viability was estimated by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide test. The activity of statins in 3D pancreatic cancer cell cultures was examined by measuring the size change of spheroids. The type of cell death was identified by cell staining with Hoechst 33342 and propidium iodide. The activity of statins on the clonogenicity of cancer cells was tested by evaluating the effect on the colony-forming ability of cells.Results: The rank order of the activity of tested statins on cell viability was as follows: PITA > SIMVA > LOVA > MEVA. Among the tested statins, PITA had the greatest effect on cell viability (half maximal effective concentration values after 72 h on BxPC-3, MIA PaCa-2, and PANC-1 cells were 1.4±0.4 µM, 1.0±0.2 µM, and 1.0±0.5 µM, respectively. PITA also showed the strongest effect on tumor spheroid growth. Statins suppressed the colony formation of cancer cells. PITA demonstrated the greatest reduction in colony size and number. Apoptosis and necrosis assay results showed that at lower concentrations statins mostly induced cell death through apoptosis, whereas higher concentrations of compounds activated also necrotic processes.Conclusion: Statins, especially PITA, demonstrate an anticancer activity against pancreatic cancer cell lines BxPC-3, MIA PaCa-2, and PANC-1 in both 2D and 3D models.Keywords: HMG-CoA reductase, cell viability, spheroid, apoptosis

  20. Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells.

    Science.gov (United States)

    Gerashchenko, O L; Zhuravel, E V; Skachkova, O V; Khranovska, N N; Filonenko, V V; Pogrebnoy, P V; Soldatkina, M A

    2013-06-01

    The aim of the study was in vitro analysis of biological activity of recombinant human beta-defensin-4 (rec-hBD-4). hBD-4 cDNA was cloned into pGEX-2T vector, and recombinant plasmid was transformed into E. coli BL21(DE3) cells. To purify soluble fusion GST-hBD-4 protein, affinity chromatography was applied. Rec-hBD-4 was cleaved from the fusion protein with thrombin, and purified by reverse phase chromatography on Sep-Pack C18. Effects of rec-hBD-4 on proliferation, viability, cell cycle distribution, substrate-independent growth, and mobility of cultured human cancer cells of A431, A549, and TPC-1 lines were analyzed by direct cell counting technique, MTT assay, flow cytofluorometry, colony forming assay in semi-soft medium, and wound healing assay. Rec-hBD-4 was expressed in bacterial cells as GST-hBD-4 fusion protein, and purified by routine 3-step procedure (affine chromatography on glutathione-agarose, cleavage of fusion protein by thrombin, and reverse phase chromatography). Analysis of in vitro activity of rec-hBD-4 toward three human cancer cell lines has demonstrated that the defensin is capable to affect cell behaviour in concentration-dependent manner. In 1-100 nM concentrations rec-hBD-4 significantly stimulates cancer cell proliferation and viability, and promotes cell cycle progression through G2/M checkpoint, greatly enhances colony-forming activity and mobility of the cells. Treatment of the cells with 500 nM of rec-hBD-4 resulted in opposite effects: significant suppression of cell proliferation and viability, blockage of cell cycle in G1/S checkpoint, significant inhibition of cell migration and colony forming activity. Recombinant human beta-defensin-4 is biologically active peptide capable to cause oppositely directed effects toward biologic features of cancer cells in vitro dependent on its concentration.

  1. The microenvironment determines the breast cancer cells' phenotype: organization of MCF7 cells in 3D cultures

    International Nuclear Information System (INIS)

    Krause, Silva; Maffini, Maricel V; Soto, Ana M; Sonnenschein, Carlos

    2010-01-01

    Stromal-epithelial interactions mediate breast development, and the initiation and progression of breast cancer. In the present study, we developed 3-dimensional (3D) in vitro models to study breast cancer tissue organization and the role of the microenvironment in phenotypic determination. The human breast cancer MCF7 cells were grown alone or co-cultured with primary human breast fibroblasts. Cells were embedded in matrices containing either type I collagen or a combination of reconstituted basement membrane proteins and type I collagen. The cultures were carried out for up to 6 weeks. For every time point (1-6 weeks), the gels were fixed and processed for histology, and whole-mounted for confocal microscopy evaluation. The epithelial structures were characterized utilizing immunohistochemical techniques; their area and proliferation index were measured using computerized morphometric analysis. Statistical differences between groups were analyzed by ANOVA, Dunnett's T3 post-hoc test and chi-square. Most of the MCF7 cells grown alone within a collagen matrix died during the first two weeks; those that survived organized into large, round and solid clusters. The presence of fibroblasts in collagen gels reduced MCF7 cell death, induced cell polarity, and the formation of round and elongated epithelial structures containing a lumen. The addition of reconstituted basement membrane to collagen gels by itself had also survival and organizational effects on the MCF7 cells. Regardless of the presence of fibroblasts, the MCF7 cells both polarized and formed a lumen. The addition of fibroblasts to the gel containing reconstituted basement membrane and collagen induced the formation of elongated structures. Our results indicate that a matrix containing both type I collagen and reconstituted basement membrane, and the presence of normal breast fibroblasts constitute the minimal permissive microenvironment to induce near-complete tumor phenotype reversion. These human

  2. Clonogenic growth of human breast cancer cells co-cultured in direct contact with serum-activated fibroblasts

    International Nuclear Information System (INIS)

    Samoszuk, Michael; Tan, Jenny; Chorn, Guillaume

    2005-01-01

    Accumulating evidence suggests that fibroblasts play a pivotal role in promoting the growth of breast cancer cells. The objective of the present study was to characterize and validate an in vitro model of the interaction between small numbers of human breast cancer cells and human fibroblasts. We measured the clonogenic growth of small numbers of human breast cancer cells co-cultured in direct contact with serum-activated, normal human fibroblasts. Using DNA microarrays, we also characterized the gene expression profile of the serum-activated fibroblasts. In order to validate the in vivo relevance of our experiments, we then analyzed clinical samples of metastatic breast cancer for the presence of myofibroblasts expressing α-smooth muscle actin. Clonogenic growth of human breast cancer cells obtained directly from in situ and invasive tumors was dramatically and consistently enhanced when the tumor cells were co-cultured in direct contact with serum-activated fibroblasts. This effect was abolished when the cells were co-cultured in transwells separated by permeable inserts. The fibroblasts in our experimental model exhibited a gene expression signature characteristic of 'serum response' (i.e. myofibroblasts). Immunostaining of human samples of metastatic breast cancer tissue confirmed that myofibroblasts are in direct contact with breast cancer cells. Serum-activated fibroblasts promote the clonogenic growth of human breast cancer cells in vitro through a mechanism that involves direct physical contact between the cells. This model shares many important molecular and phenotypic similarities with the fibroblasts that are naturally found in breast cancers

  3. Chitosan-hyaluronan based 3D co-culture platform for studying the crosstalk of lung cancer cells and mesenchymal stem cells.

    Science.gov (United States)

    Han, Hao-Wei; Hsu, Shan-Hui

    2016-09-15

    The controversial roles of mesenchymal stem cells (MSCs) in lung cancer development are not yet resolved because of the lack of an extracellular environment that mimics the tumor microenvironment. Three-dimensional (3D) culture system is an emerging research tool for biomedical applications such as drug screening. In this study, MSCs and human non-small cell lung carcinoma cells (A549) were co-cultured on a thin biomaterial-based substratum (hyaluronan-grafted chitosan, CS-HA; ∼2μm), and they were self-organized into the 3D tumor co-spheroids with core-shell structure. The gene expression levels of tumorigenicity markers in cancer cells associated with cancer stemness, epithelial-mesenchymal transition (EMT) property, and cell mobility were up-regulated for more than twofold in the MSC-tumor co-spheroids, through the promoted expression of certain tumor enhancers and the direct cell-cell interaction. To verify the different extents of tumorigenicity, A549 cells or those co-cultured with MSCs were transplanted into zebrafish embryos for evaluation in vivo. The tumorigenicity obtained from the zebrafish xenotransplantation model was consistent with that observed in vitro. These evidences suggest that the CS-HA substrate-based 3D co-culture platform for cancer cells and MSCs may be a convenient tool for studying the cell-cell interaction in a tumor-like microenvironment and potentially for cancer drug testing. Mesenchymal stem cells (MSCs) have been found in several types of tumor tissues. However, the controversial roles of MSCs in cancer development are still unsolved. Chitosan and hyaluronan are commonly used materials in the biomedical field. In the current study, we co-cultured lung cancer cells and MSCs on the planar hyaluronan-grafted chitosan (CS-HA) hybrid substrates, and discovered that lung cancer cells and MSCs were rapidly self-assembled into 3D tumor spheroids with core-shell structure on the substrates after only two days in culture. Therefore, CS

  4. Development of an innovative 3D cell culture system to study tumour--stroma interactions in non-small cell lung cancer cells.

    Directory of Open Access Journals (Sweden)

    Arno Amann

    Full Text Available INTRODUCTION: We describe a novel 3D co-culture model using non-small cell lung cancer (NSCLC cell lines in combination with lung fibroblasts. This model allows the investigation of tumour-stroma interactions and addresses the importance of having a more in vivo like cell culture model. METHODS: Automation-compatible multi-well hanging drop microtiter plates were used for the production of 3D mono- and co-cultures. In these hanging drops the two NSCLC cell lines A549 and Colo699 were cultivated either alone or co-cultured with lung fibroblasts. The viability of tumour spheroids was confirmed after five and ten days by using Annexin V/Propidium Iodide staining for flow-cytometry. Tumour fibroblast spheroid formation was characterized by scanning electron microscope (SEM, semi-thin sections, fluorescence microscope and immunohistochemistry (IHC. In addition to conventional histology, protein expression of E-Cadherin, vimentin, Ki67, fibronectin, cytokeratin 7 and α-smooth muscle actin (α-SMA was investigated by IHC. RESULTS: Lower viability was observed in A549 monocultures compared to co-cultures, whereas Colo699 monocultures showed better viability compared to co-cultures. Ki67 expression varied significantly between mono- and co-cultures in both tumour cell lines. An increase of vimentin and decreased E-Cadherin expression could be detected during the course of the cultivation suggesting a transition to a more mesenchymal phenotype. Furthermore, the fibroblast cell line showed an expression of α-SMA only in co-culture with the cancer cell line A549, thereby indicating a mesenchymal to mesenchymal shift to an even more myofibroblast phenotype. CONCLUSION: We demonstrate that our method is a promising tool for the generation of tumour spheroid co-cultures. Furthermore, these spheroids allow the investigation of tumour-stroma interactions and a better reflection of in vivo conditions of cancer cells in their microenvironment. Our method holds

  5. An imbalance in progenitor cell populations reflects tumour progression in breast cancer primary culture models.

    LENUS (Irish Health Repository)

    Donatello, Simona

    2011-01-01

    Many factors influence breast cancer progression, including the ability of progenitor cells to sustain or increase net tumour cell numbers. Our aim was to define whether alterations in putative progenitor populations could predict clinicopathological factors of prognostic importance for cancer progression.

  6. Activated human primary NK cells efficiently kill colorectal cancer cells in 3D spheroid cultures irrespectively of the level of PD-L1 expression.

    Science.gov (United States)

    Lanuza, Pilar M; Vigueras, Alan; Olivan, Sara; Prats, Anne C; Costas, Santiago; Llamazares, Guillermo; Sanchez-Martinez, Diego; Ayuso, José María; Fernandez, Luis; Ochoa, Ignacio; Pardo, Julián

    2018-01-01

    Haploidentical Natural Killer (NK) cells have been shown as an effective and safe alternative for the treatment of haematological malignancies with poor prognosis for which traditional therapies are ineffective. In contrast to haematological cancer cells, that mainly grow as single suspension cells, solid carcinomas are characterised by a tridimensional (3D) architecture that provide specific surviving advantages and resistance against chemo- and radiotherapy. However, little is known about the impact of 3D growth on solid cancer immunotherapy especially adoptive NK cell transfer. We have recently developed a protocol to activate ex vivo human primary NK cells using B lymphoblastic cell lines, which generates NK cells able to overcome chemoresistance in haematological cancer cells. Here we have analysed the activity of these allogeneic NK cells against colorectal (CRC) human cell lines growing in 3D spheroid culture and correlated with the expression of some of the main ligands regulating NK cell activity. Our results indicate that activated NK cells efficiently kill colorectal tumour cell spheroids in both 2D and 3D cultures. Notably, although 3D CRC cell cultures favoured the expression of the inhibitory immune checkpoint PD-L1, it did not correlate with increased resistance to NK cells. Finally, we have analysed in detail the infiltration of NK cells in 3D spheroids by microscopy and found that at low NK cell density, cell death is not observed although NK cells are able to infiltrate into the spheroid. In contrast, higher densities promote tumoural cell death before infiltration can be detected. These findings show that highly dense activated human primary NK cells efficiently kill colorectal carcinoma cells growing in 3D cultures independently of PD-L1 expression and suggest that the use of allogeneic activated NK cells could be beneficial for the treatment of colorectal carcinoma.

  7. The chalcone butein from Rhus verniciflua Stokes inhibits clonogenic growth of human breast cancer cells co-cultured with fibroblasts

    Directory of Open Access Journals (Sweden)

    Tan Jenny

    2005-03-01

    Full Text Available Abstract Background Butein (3,4,2',4'-tetrahydroxychalone, a plant polyphenol, is a major biologically active component of the stems of Rhus verniciflua Stokes. It has long been used as a food additive in Korea and as an herbal medicine throughout Asia. Recently, butein has been shown to suppress the functions of fibroblasts. Because fibroblasts are believed to play an important role in promoting the growth of breast cancer cells, we investigated the ability of butein to inhibit the clonogenic growth of small numbers of breast cancer cells co-cultured with fibroblasts in vitro. Methods We first measured the clonogenic growth of small numbers of the UACC-812 human breast cancer cell line co-cultured on monolayers of serum-activated, human fibroblasts in the presence of butein (2 μg/mL or various other modulators of fibroblast function (troglitazone-1 μg/mL; GW9662-1 μM; meloxican-1 μM; and 3,4 dehydroproline-10 μg/mL. In a subsequent experiment, we measured the dose-response effect on the clonogenic growth of UACC-812 breast cancer cells by pre-incubating the fibroblasts with varying concentrations of butein (10 μg/ml-1.25 μg/mL. Finally, we measured the clonogenic growth of primary breast cancer cells obtained from 5 clinical specimens with normal fibroblasts and with fibroblasts that had been pre-treated with a fixed dose of butein (2.5 μg/mL. Results Of the five modulators of fibroblast function that we tested, butein was by far the most potent inhibitor of clonogenic growth of UACC-812 breast cancer cells co-cultured with fibroblasts. Pre-treatment of fibroblasts with concentrations of butein as low as 2.5 μg/mL nearly abolished subsequent clonogenic growth of UACC-812 breast cancer cells co-cultured with the fibroblasts. A similar dose of butein had no effect on the clonogenic growth of breast cancer cells cultured in the absence of fibroblasts. Significantly, clonogenic growth of the primary breast cancer cells was also

  8. Morphological analysis of human umbilical vein endothelial cells co-cultured with ovarian cancer cells in 3D: An oncogenic angiogenesis assay.

    Directory of Open Access Journals (Sweden)

    Xiao Wan

    Full Text Available Antiangiogenic therapy for cancer is a strategy targeted at tumour vasculature, often in combination with conventional cytotoxicity treatments. Animal testing is still the most common method used for evaluating the efficacy of new drugs but tissue-engineered in vitro models are becoming more acceptable for replacing and reducing the use of animals in anti-cancer drug screening. In this study, a 3D co-culture model of human endothelial cells and ovarian cancer cells was developed. This model has the potential to mimic the interactions between endothelial cells and ovarian cancer cells. The feasibility of applying this model in drug testing was explored here. The complex morphology of the co-culture system, which features development of both endothelial tubule-like structures and tumour structures, was analysed quantitatively by an image analysis method. The co-culture morphology integrity was maintained for 10 days and the potential of the model for anti-cancer drug testing was evaluated using Paclitaxel and Cisplatin, two common anti-tumour drugs with different mechanisms of action. Both traditional cell viability assays and quantitative morphological analyses were applied in the drug testing. Cisplatin proved a good example showing the advantages of morphological analysis of the co-culture model when compared with mono-culture of endothelial cells, which did not reveal an inhibitory effect of Cisplatin on the tubule-like endothelial structures. Thus, the tubule areas of the co-culture reflected the anti-angiogenesis potential of Cisplatin. In summary, in vitro cancer models can be developed using a tissue engineering approach to more closely mimic the characteristics of tumours in vivo. Combined with the image analysis technique, this developed 3D co-culture angiogenesis model will provide more reproducible and reliably quantified results and reveal further information of the drug's effects on both tumour cell growth and tumour angiogenesis.

  9. Co-culture with NK-92MI cells enhanced the anti-cancer effect of bee venom on NSCLC cells by inactivation of NF-κB.

    Science.gov (United States)

    Kollipara, Pushpa Saranya; Kim, Jung Hyun; Won, Dohee; Lee, Sang Min; Sung, Ha Chang; Chang, Hyun Sok; Lee, Kang Tae; Lee, Kang Sik; Park, Mi Hee; Song, Min Jong; Song, Ho Sueb; Hong, Jin Tae

    2014-03-01

    In the present study we experimented on a multimodal therapeutic approach, such as combining chemotherapy agent (Bee venom) with cellular (NK-92MI) immunotherapy. Previously bee venom has been found to show anti-cancer effect in various cancer cell lines. In lung cancer cells bee venom showed an IC(50) value of 3 μg/ml in both cell lines. The co-culture of NK-92MI cell lines with lung cancer cells also show a decrease in viability upto 50 % at 48 h time point. Hence we used bee venom treated NK-92MI cells to co-culture with NSCLC cells and found that there is a further decrease in cell viability upto 70 and 75 % in A549 and NCI-H460 cell lines respectively. We further investigated the expression of various apoptotic and anti-apoptotic proteins and found that Bax, cleaved caspase-3 and -8 were increasing where as Bcl-2 and cIAP-2 was decreasing. The expression of various death receptor proteins like DR3, DR6 and Fas was also increasing. Concomitantly the expression of various death receptor ligands (TNFalpha, Apo3L and FasL) was also increasing of NK-92MI cells after co-culture. Further the DNA binding activity and luciferase activity of NF-κB was also inhibited after co-culture with bee venom treated NK-92MI cell lines. The knock down of death receptors with si-RNA has reversed the decrease in cell viability and NF-κB activity after co-culture with bee venom treated NK-92MI cells. Thus this new approach can enhance the anti-cancer effect of bee venom at a much lower concentration.

  10. Monitoring Dynamic Interactions between Breast Cancer Cells and Human Bone Tissue in a Co-Culture Model

    Science.gov (United States)

    Contag, Christopher H.; Lie, Wen-Rong; Bammer, Marie C.; Hardy, Jonathan W.; Schmidt, Tobi L.; Maloney, William J.; King, Bonnie L.

    2015-01-01

    Purpose Bone is a preferential site of breast cancer metastasis and models are needed to study this process at the level of the microenvironment. We have used bioluminescence imaging (BLI) and multiplex biomarker immunoassays to monitor dynamic breast cancer cell behaviors in co-culture with human bone tissue. Procedures Femur tissue fragments harvested from hip replacement surgeries were co-cultured with luciferase-positive MDA-MB-231-fLuc cells. BLI was performed to quantify breast cell division and track migration relative to bone tissue. Breast cell colonization of bone tissues was assessed with immunohistochemistry. Biomarkers in co-culture supernatants were profiled with MILLIPLEX® immunoassays. Results BLI demonstrated increased MDA-MB-231-fLuc proliferation (pbones, and revealed breast cell migration toward bone. Immunohistochemistry illustrated MDA-MB-231-fLuc colonization of bone, and MILLIPLEX® profiles of culture supernatants suggested breast/bone crosstalk. Conclusions Breast cell behaviors that facilitate metastasis occur reproducibly in human bone tissue co-cultures and can be monitored and quantified using BLI and multiplex immunoassays. PMID:24008275

  11. Breast cancer subtypes: two decades of journey from cell culture to patients.

    Science.gov (United States)

    Zhao, Xiangshan; Gurumurthy, Channabasavaiah Basavaraju; Malhotra, Gautam; Mirza, Sameer; Mohibi, Shakur; Bele, Aditya; Quinn, Meghan G; Band, Hamid; Band, Vimla

    2011-01-01

    Recent molecular profiling has identified six major subtypes of breast cancers that exhibit different survival outcomes for patients. To address the origin of different subtypes of breast cancers, we have now identified, isolated, and immortalized (using hTERT) mammary stem/progenitor cells which maintain their stem/progenitor properties even after immortalization. Our decade long research has shown that these stem/progenitor cells are highly susceptible to oncogenesis. Given the emerging evidence that stem/progenitor cells are precursors of cancers and that distinct subtypes of breast cancer have different survival outcome, these cellular models provide novel tools to understand the oncogenic process leading to various subtypes of breast cancers and for future development of novel therapeutic strategies to treat different subtypes of breast cancers.

  12. Differences in growth properties of endometrial cancer in three dimensional (3D) culture and 2D cell monolayer

    International Nuclear Information System (INIS)

    Chitcholtan, Kenny; Asselin, Eric; Parent, Sophie; Sykes, Peter H.; Evans, John J.

    2013-01-01

    Three-dimensional (3D) in vitro models have an invaluable role in understanding the behaviour of tumour cells in a well defined microenvironment. This is because some aspects of tumour characteristics cannot be fully recapitulated in a cell monolayer (2D). In the present study, we compared growth patterns, expression of signalling molecules, and metabolism-associated proteins of endometrial cancer cell lines in 3D and 2D cell cultures. Cancer cells formed spherical structures in 3D reconstituted basement membrane (3D rBM), and the morphological appearance was cell line dependent. Cell differentiation was observed after 8 days in the 3D rBM. There was reduced proliferation, detected by less expression of PCNA in 3D rBM than in 2D cell monolayers. The addition of exogenous epidermal growth factor (EGF) to cancer cells induced phosphorylation of EGFR and Akt in both cell culture conditions. The uptake of glucose was selectively altered in the 3D rBM, but there was a lack of association with Glut-1 expression. The secretion of vascular endothelial growth factor (VEGF) and prostaglandin E 2 (PGE 2 ) was selectively altered in 3D rBM, and it was cell line dependent. Our data demonstrated that 3D rBM as an in vitro model can influence proliferation and metabolism of endometrial cancer cell behaviour compared to 2D cell monolayer. Changes are specific to individual cell types. The use of 3D rBM is, therefore, important in the in vitro study of targeted anticancer therapies.

  13. Differences in growth properties of endometrial cancer in three dimensional (3D) culture and 2D cell monolayer

    Energy Technology Data Exchange (ETDEWEB)

    Chitcholtan, Kenny, E-mail: kenny.chitcholtan@otago.ac.nz [Department of Obstetrics and Gynaecology, University of Otago, Christchurch, 2 Riccarton Avenue, Christchurch 8011 (New Zealand); Asselin, Eric, E-mail: Eric.Asselin@uqtr.ca [Department of Chemistry and Biology, University of Quebec, at Trois-Rivières, C.P. 500, Trois-Rivières, Quebec, Canada G9A 5H7 (Canada); Parent, Sophie, E-mail: Sophie.Parent@uqtr.ca [Department of Chemistry and Biology, University of Quebec, at Trois-Rivières, C.P. 500, Trois-Rivières, Quebec, Canada G9A 5H7 (Canada); Sykes, Peter H., E-mail: peter.sykes@otago.ac.nz [Department of Obstetrics and Gynaecology, University of Otago, Christchurch, 2 Riccarton Avenue, Christchurch 8011 (New Zealand); Evans, John J., E-mail: john.evans@otago.ac.nz [Department of Obstetrics and Gynaecology, University of Otago, Christchurch, 2 Riccarton Avenue, Christchurch 8011 (New Zealand); Centre of Neuroendocrinology and The MacDiarmid Institute of Advanced Materials and Nanotechnology, University of Otago, Christchurch, 2 Riccarton Avenue, Christchurch 8011 (New Zealand)

    2013-01-01

    Three-dimensional (3D) in vitro models have an invaluable role in understanding the behaviour of tumour cells in a well defined microenvironment. This is because some aspects of tumour characteristics cannot be fully recapitulated in a cell monolayer (2D). In the present study, we compared growth patterns, expression of signalling molecules, and metabolism-associated proteins of endometrial cancer cell lines in 3D and 2D cell cultures. Cancer cells formed spherical structures in 3D reconstituted basement membrane (3D rBM), and the morphological appearance was cell line dependent. Cell differentiation was observed after 8 days in the 3D rBM. There was reduced proliferation, detected by less expression of PCNA in 3D rBM than in 2D cell monolayers. The addition of exogenous epidermal growth factor (EGF) to cancer cells induced phosphorylation of EGFR and Akt in both cell culture conditions. The uptake of glucose was selectively altered in the 3D rBM, but there was a lack of association with Glut-1 expression. The secretion of vascular endothelial growth factor (VEGF) and prostaglandin E{sub 2} (PGE{sub 2}) was selectively altered in 3D rBM, and it was cell line dependent. Our data demonstrated that 3D rBM as an in vitro model can influence proliferation and metabolism of endometrial cancer cell behaviour compared to 2D cell monolayer. Changes are specific to individual cell types. The use of 3D rBM is, therefore, important in the in vitro study of targeted anticancer therapies.

  14. Evaluation of peroxidative stress of cancer cells in vitro by real-time quantification of volatile aldehydes in culture headspace.

    Science.gov (United States)

    Shestivska, Violetta; Rutter, Abigail V; Sulé-Suso, Josep; Smith, David; Španěl, Patrik

    2017-08-30

    Peroxidation of lipids in cellular membranes results in the release of volatile organic compounds (VOCs), including saturated aldehydes. The real-time quantification of trace VOCs produced by cancer cells during peroxidative stress presents a new challenge to non-invasive clinical diagnostics, which as described here, we have met with some success. A combination of selected ion flow tube mass spectrometry (SIFT-MS), a technique that allows rapid, reliable quantification of VOCs in humid air and liquid headspace, and electrochemistry to generate reactive oxygen species (ROS) in vitro has been used. Thus, VOCs present in the headspace of CALU-1 cancer cell line cultures exposed to ROS have been monitored and quantified in real time using SIFT-MS. The CALU-1 lung cancer cells were cultured in 3D collagen to mimic in vivo tissue. Real-time SIFT-MS analyses focused on the volatile aldehydes: propanal, butanal, pentanal, hexanal, heptanal and malondialdehyde (propanedial), that are expected to be products of cellular membrane peroxidation. All six aldehydes were identified in the culture headspace, each reaching peak concentrations during the time of exposure to ROS and eventually reducing as the reactants were depleted in the culture. Pentanal and hexanal were the most abundant, reaching concentrations of a few hundred parts-per-billion by volume, ppbv, in the culture headspace. The results of these experiments demonstrate that peroxidation of cancer cells in vitro can be monitored and evaluated by direct real-time analysis of the volatile aldehydes produced. The combination of adopted methodology potentially has value for the study of other types of VOCs that may be produced by cellular damage. Copyright © 2017 John Wiley & Sons, Ltd.

  15. The gene expression profiles of canine mammary cancer cells grown with carcinoma-associated fibroblasts (CAFs as a co-culture in vitro

    Directory of Open Access Journals (Sweden)

    Król Magdalena

    2012-03-01

    Full Text Available Abstract Background It is supposed that fibroblasts present in tumour microenvironment increase cancer invasiveness and its ability to metastasize but the mechanisms have not been clearly defined yet. Thus, the current study was designed to assess changes in gene expression in five various cancer cell lines grown as a co-culture with the carcinoma-associated fibroblasts (CAFs in vitro. Results A carcinoma-associated fibroblast cell line was isolated from a canine mammary cancer. Then, a co-culture of cancer cells with the CAFs was established and maintained for 72 hrs. Having sorted the cells, a global gene expression in cancer cells using DNA microarrays was examined. The analysis revealed an up-regulation of 100 genes and a down-regulation of 106 genes in the cancer cells grown as a co-culture with the CAFs in comparison to control conditions. The PANTHER binomial statistics tool was applied to determine statistically over-manifested pathways (p Conclusion The results of the current study showed that the co-culturing of cancer cells and the CAFs caused significant changes to the cancer gene expression. The presence of the CAFs in a microenvironment of cancer cells promotes adhesion, angiogenesis and EMT.

  16. Protein kinase C-delta inactivation inhibits the proliferation and survival of cancer stem cells in culture and in vivo

    International Nuclear Information System (INIS)

    Chen, Zhihong; Forman, Lora W; Williams, Robert M; Faller, Douglas V

    2014-01-01

    CSCs in culture. PKCδ inhibition efficiently prevents tumor sphere outgrowth from tumor cell cultures, with exposure times as short as six hours. Small-molecule PKCδ inhibitors also inhibit human CSC growth in vivo in a mouse xenograft model. These findings suggest that the novel PKC isozyme PKCδ may represent a new molecular target for cancer stem cell populations

  17. Altering the Microenvironment to Promote Dormancy of Metastatic Breast Cancer Cell in a 3D Bone Culture System

    Science.gov (United States)

    2015-12-01

    of the collagen -rich extracellular matrix . Further addition of metastatic breast cancer cells to the co-culture mimicked the vicious cycle; there was...biodegradable polymers to type I collagen or matrix derived from native extracellular matrix following decellularization. Papadimitropoulos et al., presents a...metastasis, 3D model, osteoblasts, cytokines, extracellular matrix 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 18. NUMBER OF PAGES 19a

  18. Curcumin Chemosensitizes 5-Fluorouracil Resistant MMR-Deficient Human Colon Cancer Cells in High Density Cultures

    OpenAIRE

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

    2014-01-01

    OBJECTIVE: Treatment of colorectal cancer (CRC) remains a clinical challenge, as more than 15% of patients are resistant to 5-Fluorouracil (5-FU)-based chemotherapeutic regimens, and tumor recurrence rates can be as high as 50-60%. Cancer stem cells (CSC) are capable of surviving conventional chemotherapies that permits regeneration of original tumors. Therefore, we investigated the effectiveness of 5-FU and plant polyphenol (curcumin) in context of DNA mismatch repair (MMR) status and CSC ac...

  19. Insect Cell Culture

    NARCIS (Netherlands)

    Oers, van M.M.; Lynn, D.E.

    2010-01-01

    Insect cell cultures are widely used in studies on insect cell physiology, developmental biology and microbial pathology. In particular, insect cell culture is an indispensable tool for the study of insect viruses. The first continuously growing insect cell cultures were established from

  20. Inhibition of growth of human breast cancer cells in culture by neutron capture using liposomes containing 10B.

    Science.gov (United States)

    Yanagië, H; Kobayashi, H; Takeda, Y; Yoshizaki, I; Nonaka, Y; Naka, S; Nojiri, A; Shinnkawa, H; Furuya, Y; Niwa, H; Ariki, K; Yasuhara, H; Eriguchi, M

    2002-03-01

    Cell destruction in boron neutron capture therapy is effected by nuclear reaction between 10B and thermal neutrons with the release of alpha-particles (4He) and lithium-7 ions (7Li). 4He kills cells within 10 microm of the site of 4He generation, therefore it is theoretically possible to destroy tumour cells without affecting adjacent healthy tissue, given selective delivery of compounds containing 10B. Liposomes wore prepared by vortex dispersion of solutions containing 10B compounds with dried lipid films and the effects of those compounds on human breast cancer cells in culture were examined after thermal neutral irradiation. [3H]-TdR incorporation by MRKnu/nu-1 cells treated with 10B-containing liposomes showed 40% suppression compared with liposomes without 10B, at 2 x 1012 n/cm2 thermal neutron fluence. Inhibition of tumour cell growth with liposomes prepared with 100 mm 10B-compound was as significant as with those made with 500 ppm 10B solution. The concentration of 10B in liposomes was 76.5 +/- 3.4 microg/mL. Boronated liposomes can thus deliver sufficient 10B atoms to this line of breast cancer cells in culture to effect cytotoxicity and suppression of growth after thermal neutron irradiation.

  1. Cancer cells mimic in vivo spatial-temporal cell-cycle phase distribution and chemosensitivity in 3-dimensional Gelfoam® histoculture but not 2-dimensional culture as visualized with real-time FUCCI imaging.

    Science.gov (United States)

    Yano, Shuya; Miwa, Shinji; Mii, Sumiyuki; Hiroshima, Yukihiko; Uehara, Fuminaru; Kishimoto, Hiroyuki; Tazawa, Hiroshi; Zhao, Ming; Bouvet, Michael; Fujiwara, Toshiyoshi; Hoffman, Robert M

    2015-01-01

    The phase of the cell cycle can determine whether a cancer cell can respond to a given drug. We previously reported monitoring of real-time cell cycle dynamics of cancer cells throughout a live tumor, intravitally in live mice, using a fluorescence ubiquitination-based cell-cycle indicator (FUCCI). Approximately 90% of cancer cells in the center and 80% of total cells of an established tumor are in G0/G1 phase. Longitudinal real-time imaging demonstrated that cytotoxic agents killed only proliferating cancer cells at the surface and, in contrast, had little effect on quiescent cancer cells, which are the vast majority of an established tumor. Moreover, resistant quiescent cancer cells restarted cycling after cessation of chemotherapy. These results suggested why most drugs currently in clinical use, which target cancer cells in S/G2/M, are mostly ineffective on solid tumors. In the present report, we used FUCCI imaging and Gelfoam® collagen-sponge-gel histoculture, to demonstrate in real time, that the cell-cycle phase distribution of cancer cells in Gelfoam® and in vivo tumors is highly similar, whereby only the surface cells proliferate and interior cells are quiescent in G0/G1. This is in contrast to 2D culture where most cancer cells cycle. Similarly, the cancer cells responded similarly to toxic chemotherapy in Gelfoam® culture as in vivo, and very differently than cancer cells in 2D culture which were much more chemosensitive. Gelfoam® culture of FUCCI-expressing cancer cells offers the opportunity to image the cell cycle of cancer cells continuously and to screen for novel effective therapies to target quiescent cells, which are the majority in a tumor and which would have a strong probability to be effective in vivo.

  2. Adrenomedullin is a key Protein Mediating Rotary Cell Culture System that Induces the Effects of Simulated Microgravity on Human Breast Cancer Cells

    Science.gov (United States)

    Chen, Li; Yang, Xi; Cui, Xiang; Jiang, Minmin; Gui, Yu; Zhang, Yanni; Luo, Xiangdong

    2015-11-01

    Microgravity or simulated microgravity promotes stem cell proliferation and inhibits differentiation. But, researchers have not yet been able to understand the underlying mechanism through which microgravity or simulated microgravity brings about stem cell proliferation and inhibition of differentiation. In this study, we investigated the effect of simulated microgravity (SMG) on MDA-MB-231 and MCF-7 human breast cancer cells using rotary cell culture system (RCCS). SMG induced a significant accumulation of these cancer cells in S phase of the cell cycle. But, compared with the static group, there was no effect on the overall growth rate of cells in the RCCS group. Furthermore, the expression of cyclin D1 was inhibited in the RCCS group, indicating that RCCS induced cell cycle arrest. In addition, RCCS also induced glycolytic metabolism by increasing the expression of adrenomedullin (ADM), but not HIF1 a. The addition of ADM further enhanced the effects of SMG, which was induced by RCCS. But, the addition of adrenomedullin antagonist (AMA) reversed these effects of SMG. Finally, our results proved that RCCS, which induced cells cycle arrest of breast cancer cells, enhanced glycolysis and upregulated the expression of ADM. But, this did not lead to an increase in hypoxia-inducible factor-1 a (HIF1 a) expression. Thus, we have uncovered a new mechanism for understanding the Warburg effect in breast cancer cells, this mechanism is not the same as hypoxia induced glycolysis.

  3. Tumorigenic Heterogeneity in Cancer Stem Cells Evolved from Long-term Cultures of Telomerase-Immortalized

    DEFF Research Database (Denmark)

    Burns, Jorge S; Abdallah, Basem M; Guldberg, Per

    2005-01-01

    Long-term cultures of telomerase-transduced adult human mesenchymal stem cells (hMSC) may evolve spontaneous genetic changes leading to tumorigenicity in immunodeficient mice (e.g., hMSC-TERT20). We wished to clarify whether this unusual phenotype reflected a rare but dominant subpopulation or if...

  4. The Effect of VPA on Increasing Radiosensitivity in Osteosarcoma Cells and Primary-Culture Cells from Chemical Carcinogen-Induced Breast Cancer in Rats.

    Science.gov (United States)

    Liu, Guochao; Wang, Hui; Zhang, Fengmei; Tian, Youjia; Tian, Zhujun; Cai, Zuchao; Lim, David; Feng, Zhihui

    2017-05-10

    This study explored whether valproic acid (VPA, a histone deacetylase inhibitor) could radiosensitize osteosarcoma and primary-culture tumor cells, and determined the mechanism of VPA-induced radiosensitization. The working system included osteosarcoma cells (U2OS) and primary-culture cells from chemical carcinogen (DMBA)-induced breast cancer in rats; and clonogenic survival, immunofluorescence, fluorescent in situ hybridization (FISH) for chromosome aberrations, and comet assays were used in this study. It was found that VPA at the safe or critical safe concentration of 0.5 or 1.0 mM VPA could result in the accumulation of more ionizing radiation (IR)-induced DNA double strand breaks, and increase the cell radiosensitivity. VPA-induced radiosensitivity was associated with the inhibition of DNA repair activity in the working systems. In addition, the chromosome aberrations including chromosome breaks, chromatid breaks, and radial structures significantly increased after the combination treatment of VPA and IR. Importantly, the results obtained by primary-culture cells from the tissue of chemical carcinogen-induced breast cancer in rats further confirmed our findings. The data in this study demonstrated that VPA at a safe dose was a radiosensitizer for osteosarcoma and primary-culture tumor cells through suppressing DNA-double strand breaks repair function.

  5. A study on volatile organic compounds emitted by in-vitro lung cancer cultured cells using gas sensor array and SPME-GCMS.

    Science.gov (United States)

    Thriumani, Reena; Zakaria, Ammar; Hashim, Yumi Zuhanis Has-Yun; Jeffree, Amanina Iymia; Helmy, Khaled Mohamed; Kamarudin, Latifah Munirah; Omar, Mohammad Iqbal; Shakaff, Ali Yeon Md; Adom, Abdul Hamid; Persaud, Krishna C

    2018-04-02

    Volatile organic compounds (VOCs) emitted from exhaled breath from human bodies have been proven to be a useful source of information for early lung cancer diagnosis. To date, there are still arguable information on the production and origin of significant VOCs of cancer cells. Thus, this study aims to conduct in-vitro experiments involving related cell lines to verify the capability of VOCs in providing information of the cells. The performances of e-nose technology with different statistical methods to determine the best classifier were conducted and discussed. The gas sensor study has been complemented using solid phase micro-extraction-gas chromatography mass spectrometry. For this purpose, the lung cancer cells (A549 and Calu-3) and control cell lines, breast cancer cell (MCF7) and non-cancerous lung cell (WI38VA13) were cultured in growth medium. This study successfully provided a list of possible volatile organic compounds that can be specific biomarkers for lung cancer, even at the 24th hour of cell growth. Also, the Linear Discriminant Analysis-based One versus All-Support Vector Machine classifier, is able to produce high performance in distinguishing lung cancer from breast cancer cells and normal lung cells. The findings in this work conclude that the specific VOC released from the cancer cells can act as the odour signature and potentially to be used as non-invasive screening of lung cancer using gas array sensor devices.

  6. Chemo-radionuclide therapy for thyroid cancer. Initial experimental study with cultured cells

    International Nuclear Information System (INIS)

    Misaki, Takashi; Iwata, Masahiro; Iida, Yasuhiro; Kasagi, Kanji; Konishi, Junji

    2002-01-01

    Radioiodine therapy has long been used for distant metastases of thyroid cancer. Although partially effective in most cases, it can render a complete cure only in a limited number of patients. One way to enhance its efficacy would be to combine it with antineoplastic agents. Here we describe an initial in vitro evaluation with 4 thyroid cancer cell lines. Cells were sparsely seeded in microtiter plates and allowed to grow for 2 days; then they were exposed to sublethal concentrations of cisplatin (CDDP), doxorubicin (Dox), or 5-fluorouracil (5-FU), followed by treatment with I-131 for 48 hr. Cell survival was measured with a commercial kit based on the colorimetry of succinate dehydrogenase activity. Chemotherapeutic drugs exerted similar concentration-dependent cytotoxic effects in all 4 cell lines. The doses necessary to reduce the surviving fraction to half of the control were about 3 μg/ml for CDDP, 0.3 μg/ml for Dox, and 3 μg/ml for 5-FU (when used continuously for 48 hours). On the other hand, sensitivity to I-131 irradiation differed among the lines; same doses (7.4-14.8 MBq/ml) caused the greatest damage in FRO cells, a modest effect in NPA and WRO, and only minimal change in B-CPAP. The combined effect was most demonstrable in wells treated with Dox and radioiodine, whereas the addition of CDDP or 5-FU had marginal or insignificant merit, respectively. In FRO cells, half-lethal doses of the above mentioned CDDP, Dox, and 5-FU, when used together with 14.8 MBq/ml I-131, reduced cell survival to 54.5%, 29.4% and 33.4%, respectively, vs. 60.2% with radioiodine alone. In vitro, clinical concentrations of Dox can accelerate the killing of thyroid cancer cells by radioiodine. These favorable experimental results warrant future studies to evaluate whether this new bidisciplinary approach is clinically relevant and feasible. (author)

  7. Chemo-radionuclide therapy for thyroid cancer. Initial experimental study with cultured cells

    Energy Technology Data Exchange (ETDEWEB)

    Misaki, Takashi; Iwata, Masahiro; Iida, Yasuhiro; Kasagi, Kanji; Konishi, Junji [Kyoto Univ. (Japan). Graduate School of Medicine

    2002-09-01

    Radioiodine therapy has long been used for distant metastases of thyroid cancer. Although partially effective in most cases, it can render a complete cure only in a limited number of patients. One way to enhance its efficacy would be to combine it with antineoplastic agents. Here we describe an initial in vitro evaluation with 4 thyroid cancer cell lines. Cells were sparsely seeded in microtiter plates and allowed to grow for 2 days; then they were exposed to sublethal concentrations of cisplatin (CDDP), doxorubicin (Dox), or 5-fluorouracil (5-FU), followed by treatment with I-131 for 48 hr. Cell survival was measured with a commercial kit based on the colorimetry of succinate dehydrogenase activity. Chemotherapeutic drugs exerted similar concentration-dependent cytotoxic effects in all 4 cell lines. The doses necessary to reduce the surviving fraction to half of the control were about 3 {mu}g/ml for CDDP, 0.3 {mu}g/ml for Dox, and 3 {mu}g/ml for 5-FU (when used continuously for 48 hours). On the other hand, sensitivity to I-131 irradiation differed among the lines; same doses (7.4-14.8 MBq/ml) caused the greatest damage in FRO cells, a modest effect in NPA and WRO, and only minimal change in B-CPAP. The combined effect was most demonstrable in wells treated with Dox and radioiodine, whereas the addition of CDDP or 5-FU had marginal or insignificant merit, respectively. In FRO cells, half-lethal doses of the above mentioned CDDP, Dox, and 5-FU, when used together with 14.8 MBq/ml I-131, reduced cell survival to 54.5%, 29.4% and 33.4%, respectively, vs. 60.2% with radioiodine alone. In vitro, clinical concentrations of Dox can accelerate the killing of thyroid cancer cells by radioiodine. These favorable experimental results warrant future studies to evaluate whether this new bidisciplinary approach is clinically relevant and feasible. (author)

  8. Boswellia sacra essential oil induces tumor cell-specific apoptosis and suppresses tumor aggressiveness in cultured human breast cancer cells

    Science.gov (United States)

    2011-01-01

    Background Gum resins obtained from trees of the Burseraceae family (Boswellia sp.) are important ingredients in incense and perfumes. Extracts prepared from Boswellia sp. gum resins have been shown to possess anti-inflammatory and anti-neoplastic effects. Essential oil prepared by distillation of the gum resin traditionally used for aromatic therapy has also been shown to have tumor cell-specific anti-proliferative and pro-apoptotic activities. The objective of this study was to optimize conditions for preparing Boswellea sacra essential oil with the highest biological activity in inducing tumor cell-specific cytotoxicity and suppressing aggressive tumor phenotypes in human breast cancer cells. Methods Boswellia sacra essential oil was prepared from Omani Hougari grade resins through hydrodistillation at 78 or 100 oC for 12 hours. Chemical compositions were identified by gas chromatography-mass spectrometry; and total boswellic acids contents were quantified by high-performance liquid chromatography. Boswellia sacra essential oil-mediated cell viability and death were studied in established human breast cancer cell lines (T47D, MCF7, MDA-MB-231) and an immortalized normal human breast cell line (MCF10-2A). Apoptosis was assayed by genomic DNA fragmentation. Anti-invasive and anti-multicellular tumor properties were evaluated by cellular network and spheroid formation models, respectively. Western blot analysis was performed to study Boswellia sacra essential oil-regulated proteins involved in apoptosis, signaling pathways, and cell cycle regulation. Results More abundant high molecular weight compounds, including boswellic acids, were present in Boswellia sacra essential oil prepared at 100 oC hydrodistillation. All three human breast cancer cell lines were sensitive to essential oil treatment with reduced cell viability and elevated cell death, whereas the immortalized normal human breast cell line was more resistant to essential oil treatment. Boswellia sacra

  9. γ-ray hypersensitivity and faulty DNA repair in cultured cells from humans exhibiting familial cancer proneness

    International Nuclear Information System (INIS)

    Paterson, M.C.; Smith, P.J.; Bech-Hansen, N.T.; Smith, B.P.; Sell, B.M.

    1979-01-01

    The most significant danger to irradiated individuals is the induction of cancer. Ataxia telangiectasia (AT) is known as a disorder linking radiosensitivity with cancer proneness, and AT is a rare inherited disorder. This is the degenerative multisystem affliction that is transmitted as a simple autosomal recessive trait. Cell culture studies disclosed the relationship between the cellular hypersensitivity to γ-ray inactivation in vitro and the propensity to develop cancer in vivo. The molecular evidence for the defects in the repair of radiogenic DNA damage has as yet been obtained only for AT, and it seems likely that anomalous DNA repair may not be the key causal factor in the development of some of the clinical abnormalities associated with the disease, including the tendency to develop lymphoproliferative cancer. Nevertheless, AT, Rothmund-Thomson syndrome (RTS), and acute myelogenous leukemia (AML) family show promise as the models for elucidating the importance of cellular radiosensitivity and imperfect DNA repair in the induction of cancer by radiation and radiomimetic carcinogens in the biosphere. Expanded efforts are required to identify heterozygosity for the AT genes in general population and to assess the role of the interaction between this genetic make-up and environmental carcinogens in the occurrence of common cancers. (Yamashita, S.)

  10. Three-dimensional culture and interaction of cancer cells and dendritic cells in an electrospun nano-submicron hybrid fibrous scaffold

    Science.gov (United States)

    Kim, Tae-Eon; Kim, Chang Gun; Kim, Jin Soo; Jin, Songwan; Yoon, Sik; Bae, Hae-Rahn; Kim, Jeong-Hwa; Jeong, Young Hun; Kwak, Jong-Young

    2016-01-01

    An artificial three-dimensional (3D) culture system that mimics the tumor microenvironment in vitro is an essential tool for investigating the cross-talk between immune and cancer cells in tumors. In this study, we developed a 3D culture system using an electrospun poly(ε-caprolactone) (PCL) nanofibrous scaffold (NFS). A hybrid NFS containing an uninterrupted network of nano- and submicron-scale fibers (400 nm to 2 µm) was generated by deposition onto a stainless steel mesh instead of an aluminum plate. The hybrid NFS contained multiplanar pores in a 3D structure. Surface-seeded mouse CT26 colon cancer cells and bone marrow-derived dendritic cells (BM-DCs) were able to infiltrate the hybrid NFS within several hours. BM-DCs cultured on PCL nanofibers showed a baseline inactive form, and lipopolysaccharide (LPS)-activated BM-DCs showed increased expression of CD86 and major histocompatibility complex Class II. Actin and phosphorylated FAK were enriched where unstimulated and LPS-stimulated BM-DCs contacted the fibers in the 3D hybrid NFS. When BM-DCs were cocultured with mitoxantrone-treated CT26 cells in a 3D hybrid NFS, BM-DCs sprouted cytoplasm to, migrated to, synapsed with, and engulfed mitoxantrone-treated CT26 cancer cells, which were similar to the naturally occurring cross-talk between these two types of cells. The 3D hybrid NFS developed here provides a 3D structure for coculture of cancer and immune cells. PMID:27042051

  11. In-air scanning transmission ion microscopy of cultured cancer cells

    International Nuclear Information System (INIS)

    Ortega, R.; Deves, G.; Moretto, Ph.

    2001-01-01

    Scanning transmission ion microscopy (STIM) imaging of living cultured cells has been carried out using a proton external-beam with the nuclear microprobe of Bordeaux-Gradignan. STIM could be performed in air atmosphere after passage of a focused proton beam through a 150 nm thick silicon nitride window. Energy loss STIM images were obtained with a spatial resolution in the micrometer range and enabled the identification of sub-cellular ultrastructures

  12. Genome Wide Expression Profiling of Cancer Cell Lines Cultured in Microgravity Reveals Significant Dysregulation of Cell Cycle and MicroRNA Gene Networks.

    Directory of Open Access Journals (Sweden)

    Prasanna Vidyasekar

    Full Text Available Zero gravity causes several changes in metabolic and functional aspects of the human body and experiments in space flight have demonstrated alterations in cancer growth and progression. This study reports the genome wide expression profiling of a colorectal cancer cell line-DLD-1, and a lymphoblast leukemic cell line-MOLT-4, under simulated microgravity in an effort to understand central processes and cellular functions that are dysregulated among both cell lines. Altered cell morphology, reduced cell viability and an aberrant cell cycle profile in comparison to their static controls were observed in both cell lines under microgravity. The process of cell cycle in DLD-1 cells was markedly affected with reduced viability, reduced colony forming ability, an apoptotic population and dysregulation of cell cycle genes, oncogenes, and cancer progression and prognostic markers. DNA microarray analysis revealed 1801 (upregulated and 2542 (downregulated genes (>2 fold in DLD-1 cultures under microgravity while MOLT-4 cultures differentially expressed 349 (upregulated and 444 (downregulated genes (>2 fold under microgravity. The loss in cell proliferative capacity was corroborated with the downregulation of the cell cycle process as demonstrated by functional clustering of DNA microarray data using gene ontology terms. The genome wide expression profile also showed significant dysregulation of post transcriptional gene silencing machinery and multiple microRNA host genes that are potential tumor suppressors and proto-oncogenes including MIR22HG, MIR17HG and MIR21HG. The MIR22HG, a tumor-suppressor gene was one of the highest upregulated genes in the microarray data showing a 4.4 log fold upregulation under microgravity. Real time PCR validated the dysregulation in the host gene by demonstrating a 4.18 log fold upregulation of the miR-22 microRNA. Microarray data also showed dysregulation of direct targets of miR-22, SP1, CDK6 and CCNA2.

  13. Three-dimensional culture and interaction of cancer cells and dendritic cells in an electrospun nano-submicron hybrid fibrous scaffold

    Directory of Open Access Journals (Sweden)

    Kim TE

    2016-03-01

    Full Text Available Tae-Eon Kim,1–3,* Chang Gun Kim,1–3,* Jin Soo Kim,4 Songwan Jin,4 Sik Yoon,5 Hae-Rahn Bae,6 Jeong-Hwa Kim,7,8 Young Hun Jeong,7,8 Jong-Young Kwak1–3 1Department of Pharmacology, School of Medicine, 2Department of Biomedical Sciences, The Graduate School, Ajou University, Suwon, South Korea; 3Immune Network Pioneer Research Center, Ajou University Medical Center, Suwon, South Korea; 4Department of Mechanical Engineering, Korea Polytechnic University, Gyeonggi, South Korea; 5Department of Anatomy, School of Medicine, Pusan National University, Yangsan, South Korea; 6Department of Physiology, College of Medicine, Dong-A University, Busan, South Korea; 7School of Mechanical Engineering, 8Department of Mechanical Engineering, Graduate School, Kyungpook National University, Daegu, South Korea *These authors contributed equally to this work Abstract: An artificial three-dimensional (3D culture system that mimics the tumor microenvironment in vitro is an essential tool for investigating the cross-talk between immune and cancer cells in tumors. In this study, we developed a 3D culture system using an electrospun poly(ε-caprolactone (PCL nanofibrous scaffold (NFS. A hybrid NFS containing an uninterrupted network of nano- and submicron-scale fibers (400 nm to 2 µm was generated by deposition onto a stainless steel mesh instead of an aluminum plate. The hybrid NFS contained multiplanar pores in a 3D structure. Surface-seeded mouse CT26 colon cancer cells and bone marrow-derived dendritic cells (BM-DCs were able to infiltrate the hybrid NFS within several hours. BM-DCs cultured on PCL nanofibers showed a baseline inactive form, and lipopolysaccharide (LPS-activated BM-DCs showed increased expression of CD86 and major histocompatibility complex Class II. Actin and phosphorylated FAK were enriched where unstimulated and LPS-stimulated BM-DCs contacted the fibers in the 3D hybrid NFS. When BM-DCs were cocultured with mitoxantrone-treated CT26 cells in

  14. Detection of atomic scale changes in the free volume void size of three-dimensional colorectal cancer cell culture using positron annihilation lifetime spectroscopy.

    Science.gov (United States)

    Axpe, Eneko; Lopez-Euba, Tamara; Castellanos-Rubio, Ainara; Merida, David; Garcia, Jose Angel; Plaza-Izurieta, Leticia; Fernandez-Jimenez, Nora; Plazaola, Fernando; Bilbao, Jose Ramon

    2014-01-01

    Positron annihilation lifetime spectroscopy (PALS) provides a direct measurement of the free volume void sizes in polymers and biological systems. This free volume is critical in explaining and understanding physical and mechanical properties of polymers. Moreover, PALS has been recently proposed as a potential tool in detecting cancer at early stages, probing the differences in the subnanometer scale free volume voids between cancerous/healthy skin samples of the same patient. Despite several investigations on free volume in complex cancerous tissues, no positron annihilation studies of living cancer cell cultures have been reported. We demonstrate that PALS can be applied to the study in human living 3D cell cultures. The technique is also capable to detect atomic scale changes in the size of the free volume voids due to the biological responses to TGF-β. PALS may be developed to characterize the effect of different culture conditions in the free volume voids of cells grown in vitro.

  15. Cell Culture Made Easy.

    Science.gov (United States)

    Dye, Frank J.

    1985-01-01

    Outlines steps to generate cell samples for observation and experimentation. The procedures (which use ordinary laboratory equipment) will establish a short-term primary culture of normal mammalian cells. Information on culture vessels and cell division and a list of questions to generate student interest and involvement in the topics are…

  16. mRNA expression profile in DLD-1 and MOLT-4 cancer cell lines cultured under Microgravity

    Data.gov (United States)

    National Aeronautics and Space Administration — DLD-1 and MOLT-4 cell lines were cultured in a Rotating cell culture system to simulate microgravity and mRNA expression profile was observed in comparison to Static...

  17. Tumor-cytolytic human macrophages cultured as nonadherent cells: potential for the adoptive immunotherapy of cancer.

    Science.gov (United States)

    Helinski, E H; Hurley, E L; Streck, R J; Bielat, K L; Pauly, J L

    1990-01-01

    Tumor-cytolytic lymphokine (e.g., interleukin-2; IL-2)-activated killer cells are currently being evaluated in IL-2/LAK cell adoptive immunotherapy regimens for the treatment of cancer. Monocyte-derived macrophages (M phi) are also known to be efficient tumor killer cells; accordingly, M phi that have been activated in vitro may also be of therapeutic merit. However, attempts to cultivate M phi for morphological and functional studies have often been compromised because M phi adhere rapidly and tenaciously to cultureware. Studies that we have conducted to address this problem have proven successful in developing procedures for the long-term cultivation of non-adherent immunocompetent M phi in serum-free medium using petri dishes containing a thin Teflon liner. The utility of this technology is documented by the results of studies presented herein in which light and scanning electron microscopy was used to analyze tumor-cytolytic human M phi. In these experiments, we demonstrated that nonadherent immunocompetent human M phi can be prepared for detailed examinations of their pleomorphic membrane architecture. Moreover, nonadherent human M phi could readily be collected for preparing conjugates of M phi and tumor cells. It is anticipated that this technology should prove useful for future structure-function studies defining the topographical location and spatial distribution of antigens and receptors on M phi membrane ultrastructures, particularly the microvilli-like projections that bridge together an immunocompetent effector M phi and target cell (e.g., tumor cells and microbial pathogens) and which provide the physical interaction required for the initial phases of a cellular immune response that includes antigen recognition and cell-to-cell adhesion.

  18. Determination of free cisplatin in medium by differential pulse polarography after ultrasound and cisplatin treatment of a cancer cell culture

    International Nuclear Information System (INIS)

    Bernard, Vladan; Skorpikova, Jirina; Mornstein, Vojtech; Fojt, Lukas

    2011-01-01

    The in vitro study was carried out for detection of the cisplatin in free form and in culture medium, depending on various conditions of sonodynamic human ovarian cancer cells A2780 treatment by differential pulse polarography (DPP). For sonodynamic treatment, we used cisplatin alone and combined cisplatin/ultrasound treatments. The ultrasound exposure intensity of 1.0 and 2.0 Wcm 2 in far field for incubation periods 1, 24 and 48 h was used. The parameters of DPP measurements were - 1 s drop time, 5 mV.s -1 voltage scan rate, 50 mV modulation amplitude and negative scanning direction; platinum wire served as counter electrode and Ag|AgCl|3 M KCI as reference electrode. The results showed the dependence of free platinum quantities in culture medium on incubation time and treatment protocol. We found difference in concentration of free cisplatin between conventional application of cisplatin and sonodynamic treatment. The sonodynamic combined treatment of cisplatin and ultrasound field showed a higher cisplatin content in the culture medium than cisplatin treatment alone; a difference of 20% was observed for incubation time 48 h. The results also showed the influence of a time sequence of ultrasound and cytostatics in the sonodynamic treatment. The highest amount of free cisplatin in the solution was found for primary application of cisplatin and the subsequent ultrasound exposure. The quantity of free cisplatin increased with time, namely for time intervals 1-24 h. There was no difference between the DPP signal of cisplatin in reaction mixture containing cells in small quantities and micro-filtered mixture without cells. Thus, the DPP method is suitable for the detection and quantification of free cisplatin in the culture medium of cell suspension. Ultrasound field can be important factor during cytostatic therapy. (author)

  19. Molecular predictors of 3D morphogenesis by breast cancer cell lines in 3D culture.

    Directory of Open Access Journals (Sweden)

    Ju Han

    2010-02-01

    Full Text Available Correlative analysis of molecular markers with phenotypic signatures is the simplest model for hypothesis generation. In this paper, a panel of 24 breast cell lines was grown in 3D culture, their morphology was imaged through phase contrast microscopy, and computational methods were developed to segment and represent each colony at multiple dimensions. Subsequently, subpopulations from these morphological responses were identified through consensus clustering to reveal three clusters of round, grape-like, and stellate phenotypes. In some cases, cell lines with particular pathobiological phenotypes clustered together (e.g., ERBB2 amplified cell lines sharing the same morphometric properties as the grape-like phenotype. Next, associations with molecular features were realized through (i differential analysis within each morphological cluster, and (ii regression analysis across the entire panel of cell lines. In both cases, the dominant genes that are predictive of the morphological signatures were identified. Specifically, PPARgamma has been associated with the invasive stellate morphological phenotype, which corresponds to triple-negative pathobiology. PPARgamma has been validated through two supporting biological assays.

  20. Molecular Predictors of 3D Morphogenesis by Breast Cancer Cell Lines in 3D Culture

    Energy Technology Data Exchange (ETDEWEB)

    Han, Ju; Chang, Hang; Giricz, Orsi; Lee, Genee; Baehner, Frederick; Gray, Joe; Bissell, Mina; Kenny, Paraic; Parvin, Bahram

    2010-02-01

    Correlative analysis of molecular markers with phenotypic signatures is the simplest model for hypothesis generation. In this paper, a panel of 24 breast cell lines was grown in 3D culture, their morphology was imaged through phase contrast microscopy, and computational methods were developed to segment and represent each colony at multiple dimensions. Subsequently, subpopulations from these morphological responses were identified through consensus clustering to reveal three clusters of round, grape-like, and stellate phenotypes. In some cases, cell lines with particular pathobiological phenotypes clustered together (e.g., ERBB2 amplified cell lines sharing the same morphometric properties as the grape-like phenotype). Next, associations with molecular features were realized through (i) differential analysis within each morphological cluster, and (ii) regression analysis across the entire panel of cell lines. In both cases, the dominant genes that are predictive of the morphological signatures were identified. Specifically, PPAR? has been associated with the invasive stellate morphological phenotype, which corresponds to triple-negative pathobiology. PPAR? has been validated through two supporting biological assays.

  1. Lung cancer - small cell

    Science.gov (United States)

    Cancer - lung - small cell; Small cell lung cancer; SCLC ... About 15% of all lung cancer cases are SCLC. Small cell lung cancer is slightly more common in men than women. Almost all cases of SCLC are ...

  2. Matrix-assisted laser desorption/ionization mass spectrometry imaging of cell cultures for the lipidomic analysis of potential lipid markers in human breast cancer invasion.

    Science.gov (United States)

    Wang, Shujuan; Chen, Xiaowu; Luan, Hemi; Gao, Dan; Lin, Shuhai; Cai, Zongwei; Liu, Jianjun; Liu, Hongxia; Jiang, Yuyang

    2016-02-28

    Breast cancer is the leading cause of cancer death among women worldwide. Identification of lipid targets that play a role in breast cancer invasion may advance our understanding of the rapid progression of cancer and may lead to the development of new biomarkers for the disease. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) was applied for the lipidomic profiling of two poorly invasive and two highly invasive breast cancer cell lines to identify the differentially accumulated lipids related to the invasive phenotype. The four cell lines were individually grown on indium tin oxide (ITO)-coated glass slides, analyzed as cell cultures. The raster width and matrix for detection were optimized to improve detection sensitivity. Optimized MSI measurements were performed directly on the cell culture with 9-aminoacridine as matrix, resulting in 215 endogenous compounds detected in positive ion mode and 267 endogenous compounds in negative ion mode in all the four cell lines, representing the largest group of analytes that have been analyzed from cells by a single MSI study. In highly invasive cell lines, 31 lipids including phosphatidylglycerol (PG) and phosphatidic acids were found upregulated and eight lipids including sphingomyelin (SM) downregulated in negative ion mode. The products of de novo fatty acid synthesis incorporated into membrane phospholipids, like oleic-acid-containing PG, may be involved in mitochondrial dysfunction and thus affect the invasion of breast cancer cells. The deficiency of SM may be related to the disruption of apoptosis in highly invasive cancer cells. This work uncovered more analytes in cells by MSI than previous reports, providing a better visualization and novel insights to advance our understanding of the relationship between rapid progression of breast cancer and lipid metabolism. The most altered lipids may aid the discovery of diagnostic markers and therapeutic targets of breast cancer. Copyright

  3. AlgiMatrix™-Based 3D Cell Culture System as an In Vitro Tumor Model: An Important Tool in Cancer Research.

    Science.gov (United States)

    Godugu, Chandraiah; Singh, Mandip

    2016-01-01

    Routinely used two-dimensional cell culture-based models often fail while translating the observations into in vivo models. This setback is more common in cancer research, due to several reasons. The extracellular matrix and cell-to-cell interactions are not present in two-dimensional (2D) cell culture models. Diffusion of drug molecules into cancer cells is hindered by barriers of extracellular components in in vivo conditions, these barriers are absent in 2D cell culture models. To better mimic or simulate the in vivo conditions present in tumors, the current study used the alginate based three-dimensional cell culture (AlgiMatrix™) model, which resembles close to the in vivo tumor models. The current study explains the detailed protocols involved in AlgiMatrix™ based in vitro non-small-cell lung cancer (NSCLC) models. The suitability of this model was studied by evaluating, cytotoxicity, apoptosis, and penetration of nanoparticles into the in vitro tumor spheroids. This study also demonstrated the effect of EphA2 receptor targeted docetaxel-loaded nanoparticles on MDA-MB-468 TNBC cell lines. The methods section is subdivided into three subsections such as (1) preparation of AlgiMatrix™-based 3D in vitro tumor models and cytotoxicity assays, (2) free drug and nanoparticle uptake into spheroid studies, and (3) western blot, IHC, and RT-PCR studies.

  4. Co-culture of apoptotic breast cancer cells with immature dendritic cells: a novel approach for DC-based vaccination in breast cancer

    International Nuclear Information System (INIS)

    Zheng, Jin; Liu, Qiang; Yang, Jiandong; Ren, Qinyou; Cao, Wei; Yang, Jingyue; Yu, Zhaocai; Yu, Fang; Wu, Yanlan; Shi, Hengjun; Liu, Wenchao

    2012-01-01

    A dendritic cell (DC)-based vaccine strategy could reduce the risk of recurrence and improve the survival of breast cancer patients. However, while therapy-induced apoptosis of hepatocellular and colorectal carcinoma cells can enhance maturation and antigen presentation of DCs, whether this effect occurs in breast cancer is currently unknown. In the present study, we investigated the effect of doxorubicin (ADM)-induced apoptotic MCF-7 breast cancer cells on the activation of DCs. ADM-induced apoptotic MCF-7 cells could effectively induce immature DC (iDC) maturation. The mean fluorescence intensity (MFI) of DC maturity marker CD83 was 23.3 in the ADM-induced apoptotic MCF-7 cell group compared with 8.5 in the MCF-7 cell group. The MFI of DC co-stimulatory marker CD86 and HLA-DR were also increased after iDCs were treated with ADM-induced apoptotic MCF-7 cells. Furthermore, the proliferating autologous T-lymphocytes increased from 14.2 to 40.3% after incubated with DCs induced by apoptotic MCF-7 cells. The secretion of interferon-γ by these T-lymphocytes was also increased. In addition, cell-cell interaction between apoptotic MCF-7 cells and iDCs, but not soluble factors released by apoptotic MCF-7 cells, was crucial for the maturation of iDCs. These findings constitute a novel in vitro DC-based vaccine strategy for the treatment of breast cancer by ADM-induced apoptotic MCF-7 cells

  5. Co-culture of apoptotic breast cancer cells with immature dendritic cells: a novel approach for DC-based vaccination in breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Jin [Department of Oncology, State Key Discipline of Cell Biology, Xijing Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Department of Traditional Chinese and Western Medicine of Oncology, Tangdu Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Liu, Qiang [Department of Hematology, Tangdu Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Yang, Jiandong [Department of Hepatobiliary Surgery, Xijing Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Ren, Qinyou [Department of Traditional Chinese and Western Medicine of Oncology, Tangdu Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Cao, Wei [Department of Interventional Radiology, Tangdu Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Yang, Jingyue; Yu, Zhaocai [Department of Oncology, State Key Discipline of Cell Biology, Xijing Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Yu, Fang [Department of Gastrointestinal Surgery, Xijing Hospital of Digestive Diseases, the Fourth Military Medical University, Xi' an, Shaanxi (China); Wu, Yanlan [Department of Infectious Diseases, Tangdu Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Shi, Hengjun [Department of Traditional Chinese and Western Medicine of Oncology, Tangdu Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China); Liu, Wenchao [Department of Oncology, State Key Discipline of Cell Biology, Xijing Hospital, the Fourth Military Medical University, Xi' an, Shaanxi (China)

    2012-04-27

    A dendritic cell (DC)-based vaccine strategy could reduce the risk of recurrence and improve the survival of breast cancer patients. However, while therapy-induced apoptosis of hepatocellular and colorectal carcinoma cells can enhance maturation and antigen presentation of DCs, whether this effect occurs in breast cancer is currently unknown. In the present study, we investigated the effect of doxorubicin (ADM)-induced apoptotic MCF-7 breast cancer cells on the activation of DCs. ADM-induced apoptotic MCF-7 cells could effectively induce immature DC (iDC) maturation. The mean fluorescence intensity (MFI) of DC maturity marker CD83 was 23.3 in the ADM-induced apoptotic MCF-7 cell group compared with 8.5 in the MCF-7 cell group. The MFI of DC co-stimulatory marker CD86 and HLA-DR were also increased after iDCs were treated with ADM-induced apoptotic MCF-7 cells. Furthermore, the proliferating autologous T-lymphocytes increased from 14.2 to 40.3% after incubated with DCs induced by apoptotic MCF-7 cells. The secretion of interferon-γ by these T-lymphocytes was also increased. In addition, cell-cell interaction between apoptotic MCF-7 cells and iDCs, but not soluble factors released by apoptotic MCF-7 cells, was crucial for the maturation of iDCs. These findings constitute a novel in vitro DC-based vaccine strategy for the treatment of breast cancer by ADM-induced apoptotic MCF-7 cells.

  6. In vitro culture and characterization of human lung cancer circulating tumor cells isolated by size exclusion from an orthotopic nude-mouse model expressing fluorescent protein.

    Science.gov (United States)

    Kolostova, Katarina; Zhang, Yong; Hoffman, Robert M; Bobek, Vladimir

    2014-09-01

    In the present study, we demonstrate an animal model and recently introduced size-based exclusion method for circulating tumor cells (CTCs) isolation. The methodology enables subsequent in vitro CTC-culture and characterization. Human lung cancer cell line H460, expressing red fluorescent protein (H460-RFP), was orthotopically implanted in nude mice. CTCs were isolated by a size-based filtration method and successfully cultured in vitro on the separating membrane (MetaCell®), analyzed by means of time-lapse imaging. The cultured CTCs were heterogeneous in size and morphology even though they originated from a single tumor. The outer CTC-membranes were blebbing in general. Abnormal mitosis resulting in three daughter cells was frequently observed. The expression of RFP ensured that the CTCs originated from lung tumor. These readily isolatable, identifiable and cultivable CTCs can be used to characterize individual patient cancers and for screening of more effective treatment.

  7. Bacterial cell culture

    OpenAIRE

    sprotocols

    2014-01-01

    ### Materials 1. Glass culture tubes with metal caps and labels - Growth medium, from media room or customized - Glass pipette tubes - Parafilm ### Equipment 1. Vortexer - Fireboy or Bunsen burner - Motorized pipette - Micropipettes and sterile tips ### Procedure For a typical liquid culture, use 5 ml of appropriate medium. The amount in each tube does not have to be exact if you are just trying to culture cells for their precious DNA. 1. Streak an a...

  8. Gene Delivery for Metastatic Prostate Cancer Cells

    National Research Council Canada - National Science Library

    Pang, Shen

    2001-01-01

    .... Enhanced by the bystander effect, the specific expression of the DTA gene causes significant cell death in prostate cancer cell cultures, with very low background cell eradication in control cell lines...

  9. Squamous cell cancer (image)

    Science.gov (United States)

    Squamous cell cancer involves cancerous changes to the cells of the middle portion of the epidermal skin layer. It is ... malignant tumor, and is more aggressive than basal cell cancer, but still may be relatively slow-growing. It ...

  10. Liver Cell Culture Devices

    NARCIS (Netherlands)

    Andria, B.; Bracco, A.; Cirino, G.; Chamuleau, R. A. F. M.

    2010-01-01

    In the last 15 years many different liver cell culture devices, consisting of functional liver cells and artificial materials, have been developed. They have been devised for numerous different applications, such as temporary organ replacement (a bridge to liver transplantation or native liver

  11. Culture on 3D Chitosan-Hyaluronic Acid Scaffolds Enhances Stem Cell Marker Expression and Drug Resistance in Human Glioblastoma Cancer Stem Cells.

    Science.gov (United States)

    Wang, Kui; Kievit, Forrest M; Erickson, Ariane E; Silber, John R; Ellenbogen, Richard G; Zhang, Miqin

    2016-12-01

    The lack of in vitro models that support the growth of glioblastoma (GBM) stem cells (GSCs) that underlie clinical aggressiveness hinders developing new, effective therapies for GBM. While orthotopic patient-derived xenograft models of GBM best reflect in vivo tumor behavior, establishing xenografts is a time consuming, costly, and frequently unsuccessful endeavor. To address these limitations, a 3D porous scaffold composed of chitosan and hyaluronic acid (CHA) is synthesized. Growth and expression of the cancer stem cell (CSC) phenotype of the GSC GBM6 taken directly from fresh xenogratfs grown on scaffolds or as adherent monolayers is compared. While 2D adherent cultures grow as monolayers of flat epitheliod cells, GBM6 cells proliferate within pores of CHA scaffolds as clusters of self-adherent ovoid cells. Growth on scaffolds is accompanied by greater expression of genes that mediate epithelial-mesenchymal transition and maintain a primitive, undifferentiated phenotype, hallmarks of CSCs. Scaffold-grown cells also display higher expression of genes that promote resistance to hypoxia-induced oxidative stress. In accord, scaffold-grown cells show markedly greater resistance to clinically utilized alkylating agents compared to adherent cells. These findings suggest that our CHA scaffolds better mimic in vivo biological and clinical behavior and provide insights for developing novel individualized treatments. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Differential effect of TGFβ on the proteome of cancer associated fibroblasts and cancer epithelial cells in a co-culture approach - a short report.

    Science.gov (United States)

    Koczorowska, Maria Magdalena; Friedemann, Charlotte; Geiger, Klaus; Follo, Marie; Biniossek, Martin Lothar; Schilling, Oliver

    2017-12-01

    Solid tumors contain various components that together form the tumor microenvironment. Cancer associated fibroblasts (CAFs) are capable of secreting and responding to signaling molecules and growth factors. Due to their role in tumor development, CAFs are considered as potential therapeutic targets. A prominent tumor-associated signaling molecule is transforming growth factor β (TGFβ), an inducer of epithelial-to-mesenchymal transition (EMT). The differential action of TGFβ on CAFs and ETCs (epithelial tumor cells) has recently gained interest. Here, we aimed to investigate the effects of TGFβ on CAFs and ETCs at the proteomic level. We established a 2D co-culture system of differentially fluorescently labeled CAFs and ETCs and stimulated this co-culture system with TGFβ. The respective cell types were separated using FACS and subjected to quantitative analyses of individual proteomes using mass spectrometry. We found that TGFβ treatment had a strong impact on the proteome composition of CAFs, whereas ETCs responded only marginally to TGFβ. Quantitative proteomic analyses of the different cell types revealed up-regulation of extracellular matrix (ECM) proteins in TGFβ treated CAFs. In addition, we found that the TGFβ treated CAFs exhibited increased N-cadherin levels. From our data we conclude that CAFs respond to TGFβ treatment by changing their proteome composition, while ETCs appear to be rather resilient.

  13. Breast Cancer Cells in Three-dimensional Culture Display an Enhanced Radioresponse after Coordinate Targeting of Integrin ?5?1 and Fibronectin

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Jin-Min; Onodera, Yasuhito; Bissell, Mina J; Park, Catherine C

    2010-04-07

    Tactics to selectively enhance cancer radioresponse are of great interest. Cancer cells actively elaborate and remodel their extracellular matrix (ECM) to aid in survival and progression. Previous work has shown that {beta}1-integrin inhibitory antibodies can enhance the growth-inhibitory and apoptotic responses of human breast cancer cell lines to ionizing radiation, either when cells are cultured in three-dimensional laminin-rich ECM (3D lrECM) or grown as xenografts in mice. Here, we show that a specific {alpha} heterodimer of {beta}1-integrin preferentially mediates a prosurvival signal in human breast cancer cells that can be specifically targeted for therapy. 3D lrECM culture conditions were used to compare {alpha}-integrin heterodimer expression in malignant and nonmalignant cell lines. Under these conditions, we found that expression of {alpha}5{beta}1-integrin was upregulated in malignant cells compared with nonmalignant breast cells. Similarly, we found that normal and oncofetal splice variants of fibronectin, the primary ECM ligand of {alpha}5{beta}1-integrin, were also strikingly upregulated in malignant cell lines compared with nonmalignant acini. Cell treatment with a peptide that disrupts the interactions of {alpha}5{beta}1-integrin with fibronectin promoted apoptosis in malignant cells and further heightened the apoptotic effects of radiation. In support of these results, an analysis of gene expression array data from breast cancer patients revealed an association of high levels of {alpha}5-integrin expression with decreased survival. Our findings offer preclinical validation of fibronectin and {alpha}5{beta}1-integrin as targets for breast cancer therapy.

  14. Auto-catalysed progression of aneuploidy explains the Hayflick limit of cultured cells, carcinogen-induced tumours in mice, and the age distribution of human cancer.

    Science.gov (United States)

    Rasnick, D

    2000-06-15

    Evidence continues to accumulate that aneuploidy, an imbalance in the number of chromosomes, is responsible for the characteristic phenotypes of cancer, including the abnormal cellular size and morphology of cancer cells, the appearance of tumour-associated antigens, as well as the high levels of membrane-bound and secreted proteins responsible for invasiveness and loss of contact inhibition. Aneuploidy has also been demonstrated to be the self-perpetuating source of the karyotypic instability of cancer cells. Here it is shown that the auto-catalysed progression of aneuploidy explains the kinetics of the finite lifetime of diploid cells in culture, the time course of the appearance of papillomas and carcinomas in benzo[a]pyrene-treated mice, and the age-dependence of human cancers. Modelling studies indicate that the ease of spontaneous transformation of mouse cells in culture may be due to a chaotic progression of aneuploidy. Conversely, the strong preference towards senescence and resistance to transformation of human cells in culture may be the result of a non-chaotic progression of aneuploidy. Finally, a method is proposed for quantifying the aneuploidogenic potencies of carcinogens.

  15. Squamous cell skin cancer

    Science.gov (United States)

    ... that reflect light more, such as water, sand, concrete, and areas that are painted white. The higher ... - skin - squamous cell; Skin cancer - squamous cell; Nonmelanoma skin cancer - squamous ...

  16. N-Alkynyl Derivatives of 5-Fluorouracil: Susceptibility to Palladium-Mediated Dealkylation and Toxigenicity in Cancer Cell Culture

    Directory of Open Access Journals (Sweden)

    Jason T Weiss

    2014-07-01

    Full Text Available Palladium-activated prodrug therapy is an experimental therapeutic approach that relies on the unique chemical properties and biocompatibility of heterogeneous palladium catalysis to enable the spatially-controlled in vivo conversion of a biochemically-stable prodrug into its active form. This strategy, which would allow inducing local activation of systemically administered drug precursors by mediation of an implantable activating device made of Pd(0, has been proposed by our group as a way to reduce drug’s systemic toxicity while reaching therapeutic levels of the active drug in the affected tissue / organ. In the seminal study of such an approach, we reported that propargylation of the N1 position of 5-fluorouracil suppressed the drug’s cytotoxic properties, showed high stability in cell culture and facilitated the bioorthogonal restoration of the drug’s pharmacological activity in the presence of extracellular Pd(0-functionalized resins. To provide additional insight on the properties of this system, we have investigated different N1-alkynyl derivatives of 5-fluorouracil and shown that the presence of substituents near the triple bond influence negatively on its sensitivity to palladium catalysis under biocompatible conditions. Comparative studies of the N1- versus the N3-propargyl derivatives of 5-fluorouracil revealed that masking each or both positions equally led to inactive derivatives (>200-fold reduction of cytotoxicity relative to the unmodified drug, whereas the depropargylation process occurred faster at the N1 position than at the N3, thus resulting in greater toxigenic properties in cancer cell culture.

  17. Cell Culturing of Cytoskeleton

    Science.gov (United States)

    2004-01-01

    Biomedical research offers hope for a variety of medical problems, from diabetes to the replacement of damaged bone and tissues. Bioreactors, which are used to grow cells and tissue cultures, play a major role in such research and production efforts. Cell culturing, such as this bone cell culture, is an important part of biomedical research. The BioDyn payload includes a tissue engineering investigation. The commercial affiliate, Millenium Biologix, Inc., has been conducting bone implant experiments to better understand how synthetic bone can be used to treat bone-related illnesses and bone damaged in accidents. On STS-95, the BioDyn payload will include a bone cell culture aimed to help develop this commercial synthetic bone product. Millenium Biologix, Inc., is exploring the potential for making human bone implantable materials by seeding its proprietary artificial scaffold material with human bone cells. The product of this tissue engineering experiment using the Bioprocessing Modules (BPMs) on STS-95 is space-grown bone implants, which could have potential for dental implants, long bone grafts, and coating for orthopedic implants such as hip replacements.

  18. Oscillating Cell Culture Bioreactor

    Science.gov (United States)

    Freed, Lisa E.; Cheng, Mingyu; Moretti, Matteo G.

    2010-01-01

    To better exploit the principles of gas transport and mass transport during the processes of cell seeding of 3D scaffolds and in vitro culture of 3D tissue engineered constructs, the oscillatory cell culture bioreactor provides a flow of cell suspensions and culture media directly through a porous 3D scaffold (during cell seeding) and a 3D construct (during subsequent cultivation) within a highly gas-permeable closed-loop tube. This design is simple, modular, and flexible, and its component parts are easy to assemble and operate, and are inexpensive. Chamber volume can be very low, but can be easily scaled up. This innovation is well suited to work with different biological specimens, particularly with cells having high oxygen requirements and/or shear sensitivity, and different scaffold structures and dimensions. The closed-loop changer is highly gas permeable to allow efficient gas exchange during the cell seeding/culturing process. A porous scaffold, which may be seeded with cells, is fixed by means of a scaffold holder to the chamber wall with scaffold/construct orientation with respect to the chamber determined by the geometry of the scaffold holder. A fluid, with/without biological specimens, is added to the chamber such that all, or most, of the air is displaced (i.e., with or without an enclosed air bubble). Motion is applied to the chamber within a controlled environment (e.g., oscillatory motion within a humidified 37 C incubator). Movement of the chamber induces relative motion of the scaffold/construct with respect to the fluid. In case the fluid is a cell suspension, cells will come into contact with the scaffold and eventually adhere to it. Alternatively, cells can be seeded on scaffolds by gel entrapment prior to bioreactor cultivation. Subsequently, the oscillatory cell culture bioreactor will provide efficient gas exchange (i.e., of oxygen and carbon dioxide, as required for viability of metabolically active cells) and controlled levels of fluid

  19. Plant cell culture initiation

    NARCIS (Netherlands)

    Hall, R.D.

    2000-01-01

    The use of cultured plant cells in either organized or unorganized form has increased vey considerably in the last 10-15 yr. Many new technologies have been developed and applications in both fundamental and applied research have led to the development of some powerful tools for improving our

  20. Organoid Culture of Isolated Cells from Patient-derived Tissues with Colorectal Cancer

    Directory of Open Access Journals (Sweden)

    Bing-Ying Xie

    2016-01-01

    Conclusions: We successfully constructed a CRC organoid model that grew robustly over 25 days and demonstrated that 2000 cells/well in 96-well plate was a prime seeding density for cells to form organoids. The results confirmed that organoid model can be used for agent screening and personalized medicine.

  1. Paracrine interactions between LNCaP prostate cancer cells and bioengineered bone in 3D in vitro culture reflect molecular changes during bone metastasis.

    Science.gov (United States)

    Sieh, Shirly; Taubenberger, Anna V; Lehman, Melanie L; Clements, Judith A; Nelson, Colleen C; Hutmacher, Dietmar W

    2014-06-01

    As microenvironmental factors such as three-dimensionality and cell-matrix interactions are increasingly being acknowledged by cancer biologists, more complex 3D in vitro models are being developed to study tumorigenesis and cancer progression. To better understand the pathophysiology of bone metastasis, we have established and validated a 3D indirect co-culture model to investigate the paracrine interactions between prostate cancer (PCa) cells and human osteoblasts. Co-culture of the human PCa, LNCaP cells embedded within polyethylene glycol hydrogels with human osteoblasts in the form of a tissue engineered bone construct (TEB), resulted in reduced proliferation of LNCaP cells. LNCaP cells in both monoculture and co-culture were responsive to the androgen analog, R1881, as indicated by an increase in the expression (mRNA and/or protein induction) of androgen-regulated genes including prostate specific antigen and fatty acid synthase. Microarray gene expression analysis further revealed an up-regulation of bone markers and other genes associated with skeletal and vasculature development and a significant activation of transforming growth factor β1 downstream genes in LNCaP cells after co-culture with TEB. LNCaP cells co-cultured with TEB also unexpectedly showed similar changes in classical androgen-responsive genes under androgen-deprived conditions not seen in LNCaP monocultures. The molecular changes of LNCaP cells after co-culturing with TEBs suggest that osteoblasts exert a paracrine effect that may promote osteomimicry and modulate the expression of androgen-responsive genes in LNCaP cells. Taken together, we have presented a novel 3D in vitro model that allows the study of cellular and molecular changes occurring in PCa cells and osteoblasts that are relevant to metastatic colonization of bone. This unique in vitro model could also facilitate cancer biologists to dissect specific biological hypotheses via extensive genomic or proteomic assessments to

  2. Differential effects of selective frankincense (Ru Xiang) essential oil versus non-selective sandalwood (Tan Xiang) essential oil on cultured bladder cancer cells: a microarray and bioinformatics study

    Science.gov (United States)

    2014-01-01

    Background Frankincense (Boswellia carterii, known as Ru Xiang in Chinese) and sandalwood (Santalum album, known as Tan Xiang in Chinese) are cancer preventive and therapeutic agents in Chinese medicine. Their biologically active ingredients are usually extracted from frankincense by hydrodistillation and sandalwood by distillation. This study aims to investigate the anti-proliferative and pro-apoptotic activities of frankincense and sandalwood essential oils in cultured human bladder cancer cells. Methods The effects of frankincense (1,400–600 dilutions) (v/v) and sandalwood (16,000–7,000 dilutions) (v/v) essential oils on cell viability were studied in established human bladder cancer J82 cells and immortalized normal human bladder urothelial UROtsa cells using a colorimetric XTT cell viability assay. Genes that responded to essential oil treatments in human bladder cancer J82 cells were identified using the Illumina Expression BeadChip platform and analyzed for enriched functions and pathways. The chemical compositions of the essential oils were determined by gas chromatography–mass spectrometry. Results Human bladder cancer J82 cells were more sensitive to the pro-apoptotic effects of frankincense essential oil than the immortalized normal bladder UROtsa cells. In contrast, sandalwood essential oil exhibited a similar potency in suppressing the viability of both J82 and UROtsa cells. Although frankincense and sandalwood essential oils activated common pathways such as inflammatory interleukins (IL-6 signaling), each essential oil had a unique molecular action on the bladder cancer cells. Heat shock proteins and histone core proteins were activated by frankincense essential oil, whereas negative regulation of protein kinase activity and G protein-coupled receptors were activated by sandalwood essential oil treatment. Conclusion The effects of frankincense and sandalwood essential oils on J82 cells and UROtsa cells involved different mechanisms leading to

  3. Different apoptotic effects of [Pt(O,O'-acac)(γ-acac)(DMS)] and cisplatin on normal and cancerous human epithelial breast cells in primary culture.

    Science.gov (United States)

    Vetrugno, Carla; Muscella, Antonella; Fanizzi, Francesco Paolo; Cossa, Luca Giulio; Migoni, Danilo; De Pascali, Sandra Angelica; Marsigliante, Santo

    2014-11-01

    The aim of this study was to determine whether [platinum (Pt)(O,O'-acetylacetonate (acac))(γ-acac)(dimethylsulphide (DMS))] is differentially cytotoxic in normal and cancer cells, and to measure comparative levels of cytotoxicity compared with cisplatin in the same cells. We performed experiments on cancerous and normal epithelial breast cells in primary culture obtained from the same patients. The apoptotic effects [Pt(O,O'-acac)(γ-acac)(DMS)] and cisplatin in cancerous and normal breast cells were compared. Cancer cells were more sensitive to [Pt(O,O'-acac)(γ-acac)(DMS)] (IC50 = 5.22 ± 1.2 μmol·L(-1)) than normal cells (IC50 = 116.9 ± 8.8 μmol·L(-1)). However, the difference was less strong when cisplatin was used (IC50 = 96.0 ± 6.9 and 61.9 ± 6.1 μmol·L(-1) for cancer and normal cells respectively). Both compounds caused reactive oxygen species (ROS) production with different mechanisms: [Pt(O,O'-acac)(γ-acac)(DMS)] quickly activated NAD(P)H oxidase while cisplatin caused a slower formation of mitochondrial ROS. Cisplatin and [Pt(O,O'-acac)(γ-acac)(DMS)] caused activation of caspases, proteolysis of PARP and modulation of Bcl-2, Bax and Bid. [Pt(O,O'-acac)(γ-acac)(DMS)] also caused leakage of cytochrome c from the mitochondria. Overall, these processes proceeded more quickly in cells treated with [Pt(O,O'-acac)(γ-acac)(DMS)] compared with cisplatin. [Pt(O,O'-acac)(γ-acac)(DMS)] effects were faster and quantitatively greater in cancer than in normal cells. [Pt(O,O'-acac)(γ-acac)(DMS)] caused a fast decrease of mitochondrial membrane potential, especially in cancer cells. [Pt(O,O'-acac)(γ-acac)(DMS)] was specific to breast cancer cells in primary culture, and this observation makes this compound potentially more interesting than cisplatin. © 2014 The British Pharmacological Society.

  4. Different apoptotic effects of [Pt(O,O′-acac)(γ-acac)(DMS)] and cisplatin on normal and cancerous human epithelial breast cells in primary culture

    Science.gov (United States)

    Vetrugno, Carla; Muscella, Antonella; Fanizzi, Francesco Paolo; Cossa, Luca Giulio; Migoni, Danilo; De Pascali, Sandra Angelica; Marsigliante, Santo

    2014-01-01

    Background and Purpose The aim of this study was to determine whether [platinum (Pt)(O,O′-acetylacetonate (acac))(γ-acac)(dimethylsulphide (DMS))] is differentially cytotoxic in normal and cancer cells, and to measure comparative levels of cytotoxicity compared with cisplatin in the same cells. Experimental Approach We performed experiments on cancerous and normal epithelial breast cells in primary culture obtained from the same patients. The apoptotic effects [Pt(O,O′-acac)(γ-acac)(DMS)] and cisplatin in cancerous and normal breast cells were compared. Key Results Cancer cells were more sensitive to [Pt(O,O′-acac)(γ-acac)(DMS)] (IC50 = 5.22 ± 1.2 μmol·L−1) than normal cells (IC50 = 116.9 ± 8.8 μmol·L−1). However, the difference was less strong when cisplatin was used (IC50 = 96.0 ± 6.9 and 61.9 ± 6.1 μmol·L−1 for cancer and normal cells respectively). Both compounds caused reactive oxygen species (ROS) production with different mechanisms: [Pt(O,O′-acac)(γ-acac)(DMS)] quickly activated NAD(P)H oxidase while cisplatin caused a slower formation of mitochondrial ROS. Cisplatin and [Pt(O,O′-acac)(γ-acac)(DMS)] caused activation of caspases, proteolysis of PARP and modulation of Bcl-2, Bax and Bid. [Pt(O,O′-acac)(γ-acac)(DMS)] also caused leakage of cytochrome c from the mitochondria. Overall, these processes proceeded more quickly in cells treated with [Pt(O,O′-acac)(γ-acac)(DMS)] compared with cisplatin. [Pt(O,O′-acac)(γ-acac)(DMS)] effects were faster and quantitatively greater in cancer than in normal cells. [Pt(O,O′-acac)(γ-acac)(DMS)] caused a fast decrease of mitochondrial membrane potential, especially in cancer cells. Conclusions and Implications [Pt(O,O′-acac)(γ-acac)(DMS)] was specific to breast cancer cells in primary culture, and this observation makes this compound potentially more interesting than cisplatin. PMID:24990093

  5. Spheroid Culture of Head and Neck Cancer Cells Reveals an Important Role of EGFR Signalling in Anchorage Independent Survival.

    Science.gov (United States)

    Braunholz, Diana; Saki, Mohammad; Niehr, Franziska; Öztürk, Merve; Borràs Puértolas, Berta; Konschak, Robert; Budach, Volker; Tinhofer, Ingeborg

    2016-01-01

    In solid tumours millions of cells are shed into the blood circulation each day. Only a subset of these circulating tumour cells (CTCs) survive, many of them presumable because of their potential to form multi-cellular clusters also named spheroids. Tumour cells within these spheroids are protected from anoikis, which allows them to metastasize to distant organs or re-seed at the primary site. We used spheroid cultures of head and neck squamous cell carcinoma (HNSCC) cell lines as a model for such CTC clusters for determining the role of the epidermal growth factor receptor (EGFR) in cluster formation ability and cell survival after detachment from the extra-cellular matrix. The HNSCC cell lines FaDu, SCC-9 and UT-SCC-9 (UT-SCC-9P) as well as its cetuximab (CTX)-resistant sub-clone (UT-SCC-9R) were forced to grow in an anchorage-independent manner by coating culture dishes with the anti-adhesive polymer poly-2-hydroxyethylmethacrylate (poly-HEMA). The extent of apoptosis, clonogenic survival and EGFR signalling under such culture conditions was evaluated. The potential of spheroid formation in suspension culture was found to be positively correlated with the proliferation rate of HNSCC cell lines as well as their basal EGFR expression levels. CTX and gefitinib blocked, whereas the addition of EGFR ligands promoted anchorage-independent cell survival and spheroid formation. Increased spheroid formation and growth were associated with persistent activation of EGFR and its downstream signalling component (MAPK/ERK). Importantly, HNSCC cells derived from spheroid cultures retained their clonogenic potential in the absence of cell-matrix contact. Addition of CTX under these conditions strongly inhibited colony formation in CTX-sensitive cell lines but not their resistant subclones. Altogether, EGFR activation was identified as crucial factor for anchorage-independent survival of HNSCC cells. Targeting EGFR in CTC cluster formation might represent an attractive anti

  6. Epithelial Cell Cultures

    Directory of Open Access Journals (Sweden)

    Imran S. Chaudhry

    2011-01-01

    Full Text Available The biological effects of only a finite number of tobacco toxins have been studied. Here, we describe exposure of cultures of human bronchial epithelial cells to low concentrations of tobacco carcinogens: nickel sulphate, benzo(bfluoranthene, N-nitrosodiethylamine, and 4-(methylnitrosamino-1-(3-pyridyl-1-butanone (NNK. After a 24-hour exposure, EGFR was expressed in cell membrane and cytoplasm, BCL-2 was expressed only in the irregular nuclei of large atypical cells, MKI67 was expressed in nuclei with no staining in larger cells, cytoplasmic BIRC5 with stronger nuclear staining was seen in large atypical cells, and nuclear TP53 was strongly expressed in all cells. After only a 24-hour exposure, cells exhibited atypical nuclear and cytoplasmic features. After a 48-hour exposure, EGFR staining was localized to the nucleus, BCL-2 was slightly decreased in intensity, BIRC5 was localized to the cytoplasm, and TP53 staining was increased in small and large cells. BCL2L1 was expressed in both the cytoplasm and nuclei of cells at 24- and 48-hour exposures. We illustrate that short-termexposure of a bronchial epithelial cell line to smoking-equivalent concentrations of tobacco carcinogens alters the expression of key proliferation regulatory genes, EGFR, BCL-2, BCL2L1, BIRC5, TP53, and MKI67, similar to that reported in biopsy specimens of pulmonary epithelium described to be preneoplastic lesions.

  7. Human Organ Culture: Updating the Approach to Bridge the Gap from In Vitro to In Vivo in Inflammation, Cancer, and Stem Cell Biology

    Directory of Open Access Journals (Sweden)

    Rafia S. Al-Lamki

    2017-09-01

    Full Text Available Human studies, critical for developing new diagnostics and therapeutics, are limited by ethical and logistical issues, and preclinical animal studies are often poor predictors of human responses. Standard human cell cultures can address some of these concerns but the absence of the normal tissue microenvironment can alter cellular responses. Three-dimensional cultures that position cells on synthetic matrices, or organoid or organ-on-a-chip cultures, in which different cell spontaneously organize contacts with other cells and natural matrix only partly overcome this limitation. Here, we review how human organ cultures (HOCs can more faithfully preserve in vivo tissue architecture and can better represent disease-associated changes. We will specifically describe how HOCs can be combined with both traditional and more modern morphological techniques to reveal how anatomic location can alter cellular responses at a molecular level and permit comparisons among different cells and different cell types within the same tissue. Examples are provided involving use of HOCs to study inflammation, cancer, and stem cell biology.

  8. Human mammary fibroblasts stimulate invasion of breast cancer cells in a three-dimensional culture and increase stroma development in mouse xenografts

    International Nuclear Information System (INIS)

    Olsen, Charlotta J; Moreira, José; Lukanidin, Eugene M; Ambartsumian, Noona S

    2010-01-01

    Tumour phenotype is regulated in a complex fashion as a result of interactions between malignant cells and the tumour stroma. Fibroblasts are the most abundant and perhaps most active part of the tumour stroma. A better understanding of the changes that occur in fibroblasts in response to the presence of malignant cells may lead to the development of new strategies for cancer treatment. We explored the effects of fibroblasts on the growth and invasion of mammary carcinoma tumour cells in vitro and in vivo. In order to analyse secreted factors that affect invasive abilities of breast cancer cells we co-cultured human mammary fibroblasts (HMF3s) and cancer cells (MCF7S1) in three-dimensional (3D) growth conditions devoid of heterogeneous cell-cell contact. To study the possible influence of fibroblasts on MCF7S1 cancer cell growth in vivo we co-injected HMF3s and MCF7S1 cells in Balb/c nu/nu mice. In 3D co-culture both HMF3s and MCF7S1 cells demonstrated enhanced invasion into a Matrigel matrix. This was correlated with enhanced expression of the metastasis promoting S100A4 protein in fibroblasts, stimulation of the matrix metalloproteinase (MMP)-2 activity, and enhanced secretion of a range of different cytokines. Orthotopic injection of oestrogen-dependent MCF7S1 cancer cells together with fibroblasts showed stimulation of tumour growth in mice without an external oestrogen supply. The resulting tumours were characterized by increased development of extracellular matrix, as well as an increase of murine S100A4 concentration and activity of MMP-2 in the tumour interstitial fluid. Stimulation of the invasive phenotype of tumour cells in 3D co-cultures with fibroblasts could be correlated with increased production of S100A4 and MMP-2. We propose that enhanced development of mouse host-derived tumour stroma in a MCF7S1 co-injection xenograft model leads to oestrogen independency and is triggered by the initial presence of human fibroblasts

  9. Perfusion based cell culture chips

    DEFF Research Database (Denmark)

    Heiskanen, Arto; Emnéus, Jenny; Dufva, Martin

    2010-01-01

    Performing cell culture in miniaturized perfusion chambers gives possibilities to experiment with cells under near in vivo like conditions. In contrast to traditional batch cultures, miniaturized perfusion systems provide precise control of medium composition, long term unattended cultures...... and tissue like structuring of the cultures. However, as this chapter illustrates, many issues remain to be identified regarding perfusion cell culture such as design, material choice and how to use these systems before they will be widespread amongst biomedical researchers....

  10. Microfluidic Cell Culture Device

    Science.gov (United States)

    Takayama, Shuichi (Inventor); Cabrera, Lourdes Marcella (Inventor); Heo, Yun Seok (Inventor); Smith, Gary Daniel (Inventor)

    2014-01-01

    Microfluidic devices for cell culturing and methods for using the same are disclosed. One device includes a substrate and membrane. The substrate includes a reservoir in fluid communication with a passage. A bio-compatible fluid may be added to the reservoir and passage. The reservoir is configured to receive and retain at least a portion of a cell mass. The membrane acts as a barrier to evaporation of the bio-compatible fluid from the passage. A cover fluid may be added to cover the bio-compatible fluid to prevent evaporation of the bio-compatible fluid.

  11. An in vitro 3D bone metastasis model by using a human bone tissue culture and human sex-related cancer cells.

    Science.gov (United States)

    Salamanna, Francesca; Borsari, Veronica; Brogini, Silvia; Giavaresi, Gianluca; Parrilli, Annapaola; Cepollaro, Simona; Cadossi, Matteo; Martini, Lucia; Mazzotti, Antonio; Fini, Milena

    2016-11-22

    One of the main limitations, when studying cancer-bone metastasis, is the complex nature of the native bone environment and the lack of reliable, simple, inexpensive models that closely mimic the biological processes occurring in patients and allowing the correct translation of results. To enhance the understanding of the mechanisms underlying human bone metastases and in order to find new therapies, we developed an in vitro three-dimensional (3D) cancer-bone metastasis model by culturing human breast or prostate cancer cells with human bone tissue isolated from female and male patients, respectively. Bone tissue discarded from total hip replacement surgery was cultured in a rolling apparatus system in a normoxic or hypoxic environment. Gene expression profile, protein levels, histological, immunohistochemical and four-dimensional (4D) micro-CT analyses showed a noticeable specificity of breast and prostate cancer cells for bone colonization and ingrowth, thus highlighting the species-specific and sex-specific osteotropism and the need to widen the current knowledge on cancer-bone metastasis spread in human bone tissues. The results of this study support the application of this model in preclinical studies on bone metastases and also follow the 3R principles, the guiding principles, aimed at replacing/reducing/refining (3R) animal use and their suffering for scientific purposes.

  12. 3D Hanging Drop Culture to Establish Prostate Cancer Organoids.

    Science.gov (United States)

    Eder, Theresa; Eder, Iris E

    2017-01-01

    Three-dimensional (3D) cell culture enables the growth of cells in a multidimensional and multicellular manner compared to conventional cell culture techniques. Especially in prostate cancer research there is a big need for more tissue-recapitulating models to get a better understanding of the mechanisms driving prostate cancer as well as to screen for more efficient drugs that can be used for treatment. In this chapter we describe a 3D hanging drop system that can be used to culture prostate cancer organoids as tumor epithelial monocultures and as epithelial-stromal cocultures.

  13. In a patient with biclonal Waldenstrom macroglobulinemia only one clone expands in three-dimensional culture and includes putative cancer stem cells.

    Science.gov (United States)

    Kirshner, Julia; Thulien, Kyle J; Kriangkum, Jitra; Motz, Sarah; Belch, Andrew R; Pilarski, Linda M

    2011-02-01

    A small percentage of cases of Waldenstrom macroglobulinemia (WM) present with biclonality, defined here as the rearrangement of two distinct VDJ gene segments. Here we investigated the expansion of two clones from a patient with WM expressing molecularly detectable clonotypic gene rearrangements, one V(H)3 and one V(H)4. Biclonality was determined in blood and bone marrow mononuclear cells using real-time quantitative PCR (RQ-PCR). V(H)4 expressing cells but not V(H)3 expressing cells underwent clonal expansion in 3-D culture of reconstructed WM bone marrow. After 3-D culture, secondary culture in a colony forming unit assay, and RQ-PCR, only the V(H)4 clone was shown to harbor a subpopulation with characteristics of cancer stem cells, including proliferative quiescence, self-regeneration, and the ability to generate clonotypic progeny, suggesting that the V(H)4, but not the V(H)3, clone is clinically significant. Enrichment of potential WM stem cells in 3-D cultures holds promise for monitoring their response to treatment and for testing new therapies.

  14. In vitro co-culture experiments on prostate cancer and small intestine cells irradiated with carbon ions and x-rays

    International Nuclear Information System (INIS)

    Neubeck, C. von; Weyrather, W.-K.; Durante, M.

    2009-01-01

    Intensity modulated radiotherapy (IMRT) delivers the dose in many small irradiation fields of different beam direction to achieve a 3 dimensional tumour conformal dose overlapping with a maximum of normal tissue protection. In 2006 a study was started at GSI to treat prostate cancer patients with a boost irradiation of carbon ions in combination with an IMRT treatment administered at the Uniklinikum Heidelberg. The carbon ions are delivered in two opposing fields. So IMRT irradiation includes more normal tissue than carbon ion treatment but even here parts of the rectum and the bladder are in the irradiated field. This raises the question whether the irradiated tumor cells influence the normal cells (irradiated/ unirradiated) but also whether the normal irradiated cells influences normal tissue in a different way for carbon and photon irradiation. To study this problem, we established an in vitro co-culture model of prostate cancer and small intestine cells of the rat to simulate the patient treatment situation for analyzing tissue reaction exemplary. For characterization of the cells lines the parameters alpha and beta (linear quadratic model) for clonogenic survival were determined for x-rays and for carbon ions of different energies. For co-culture experiments unirradiated and irradiated cells were seeded together and the survival was analyzed

  15. miR-17-5p targets the p300/CBP-associated factor and modulates androgen receptor transcriptional activity in cultured prostate cancer cells

    International Nuclear Information System (INIS)

    Gong, Ai-Yu; Eischeid, Alex N; Xiao, Jing; Zhao, Jian; Chen, Dongqing; Wang, Zhao-Yi; Young, Charles YF; Chen, Xian-Ming

    2012-01-01

    Androgen receptor (AR) signalling is critical to the initiation and progression of prostate cancer (PCa). Transcriptional activity of AR involves chromatin recruitment of co-activators, including the p300/CBP-associated factor (PCAF). Distinct miRNA expression profiles have been identified in PCa cells during the development and progression of the disease. Whether miRNAs regulate PCAF expression in PCa cells to regulate AR transcriptional activity is still unclear. Expression of PCAF was investigated in several PCa cell lines by qRT-PCR, Western blot, and immunocytochemistry. The effects of PCAF expression on AR-regulated transcriptional activity and cell growth in PCa cells were determined by chromatin immunoprecipitation, reporter gene construct analysis, and MTS assay. Targeting of PCAF by miR-17-5p was evaluated using the luciferase reporter assay. PCAF was upregulated in several PCa cell lines. Upregulation of PCAF promoted AR transcriptional activation and cell growth in cultured PCa cells. Expression of PCAF in PCa cells was associated with the downregulation of miR-17-5p. Targeting of the 3’-untranslated region of PCAF mRNA by miR-17-5p caused translational suppression and RNA degradation, and, consequently, modulation of AR transcriptional activity in PCa cells. PCAF is upregulated in cultured PCa cells, and upregulation of PCAF is associated with the downregulation of miR-17-5p. Targeting of PCAF by miR-17-5p modulates AR transcriptional activity and cell growth in cultured PCa cells

  16. Differences in radiosensitivity among cells in culture and in experimental tumours: Significance for the effectiveness of human cancer therapy

    International Nuclear Information System (INIS)

    Barendsen, G.W.; Amsterdam Univ.

    1987-01-01

    Problems in the application of radiobiological data on various types of models, cell in vitro, experimental tumours, and clinical models, to the prediction of tumour radiocurability are discussed. On the basis of observations on cells in culture and experimental tumours it is suggested that heterogeneity in responsiveness of tumours in patients is caused in a large part by differences in intrinsic cellular radiosensitivity. Methods and developments are reviewed, which may yield better assays for the prediction of tumour responsiveness to treatments. (Auth.)

  17. Effect of aluminium on migratory and invasive properties of MCF-7 human breast cancer cells in culture.

    Science.gov (United States)

    Darbre, Philippa D; Bakir, Ayse; Iskakova, Elzira

    2013-11-01

    Aluminium (Al) has been measured in human breast tissue, nipple aspirate fluid and breast cyst fluid, and recent studies have shown that at tissue concentrations, aluminium can induce DNA damage and suspension growth in human breast epithelial cells. This paper demonstrates for the first time that exposure to aluminium can also increase migratory and invasive properties of MCF-7 human breast cancer cells. Long-term (32 weeks) but not short-term (1 week) exposure of MCF-7 cells to 10(-4) M aluminium chloride or 10(-4) M aluminium chlorohydrate increased motility of the cells as measured by live cell imaging (cumulative length moved by individual cells), by a wound healing assay and by migration in real time through 8 μm pores of a membrane using xCELLigence technology. Long-term exposure (37 weeks) to 10(-4) M aluminium chloride or 10(-4) M aluminium chlorohydrate also increased the ability of MCF-7 cells to invade through a matrigel layer as measured in real time using the xCELLigence system. Although molecular mechanisms remain to be characterized, the ability of aluminium salts to increase migratory and invasive properties of MCF-7 cells suggests that the presence of aluminium in the human breast could influence metastatic processes. This is important because mortality from breast cancer arises mainly from tumour spread rather than from the presence of a primary tumour in the breast. © 2013.

  18. Cell phones and cancer

    Science.gov (United States)

    Cancer and cell phones; Do cell phones cause cancer? ... Several major studies show no link between cell phones and cancer at this time. However, since the information available is based on short-term studies, the impact of many years of ...

  19. The Importance of Caveolin-1 as Key-Regulator of Three-Dimensional Growth in Thyroid Cancer Cells Cultured under Real and Simulated Microgravity Conditions

    Directory of Open Access Journals (Sweden)

    Stefan Riwaldt

    2015-11-01

    Full Text Available We recently demonstrated that the CAV1 gene was down-regulated, when poorly differentiated thyroid FTC-133 cancer cells formed spheroids under simulated microgravity conditions. Here, we present evidence that the caveolin-1 protein is involved in the inhibition of spheroid formation, when confluent monolayers are exposed to microgravity. The evidence is based on proteins detected in cells and their supernatants of the recent spaceflight experiment: “NanoRacks-CellBox-Thyroid Cancer”. The culture supernatant had been collected in a special container adjacent to the flight hardware incubation chamber and stored at low temperature until it was analyzed by Multi-Analyte Profiling (MAP technology, while the cells remaining in the incubation chamber were fixed by RNAlater and examined by mass spectrometry. The soluble proteins identified by MAP were investigated in regard to their mutual interactions and their influence on proteins, which were associated with the cells secreting the soluble proteins and had been identified in a preceding study. A Pathway Studio v.11 analysis of the soluble and cell-associated proteins together with protein kinase C alpha (PRKCA suggests that caveolin-1 is involved, when plasminogen enriched in the extracellular space is not activated and the vascular cellular adhesion molecule (VCAM-1 mediated cell–cell adhesion is simultaneously strengthened and activated PRKCA is recruited in caveolae, while the thyroid cancer cells do not form spheroids.

  20. Effect of DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine on radiosensitivity of the human lung cancer cells in three-dimensional culture

    International Nuclear Information System (INIS)

    Pan Dong; Xue Gang; Li Xiaoman; Chen Yaxiong; Ren Zhenxin; Du Yarong; Hu Burong

    2014-01-01

    5-Aza-CdR is a specific inhibitor of DNMTs which could suppress tumor growth by demethylation of genomic DNA. There have only few studies thus far concerning it as radiosensitizers in three-dimensional (3D) cells. The principal aim of this study is to evaluate the effects of 5-Aza-CdR on the radiosensitivity of A549 cells in monolayer (2D) and 3D cultures in an attempt to find out a new combination treatments with radiotherapy. The cell proliferation was detected by MTT assay after pretreated with different doses of 5-Aza-CdR for 72 h. A549 cells were treated with or without 5-Aza-CdR (2, 5 μmol/L) for 72 h before be exposed to X-rays of 1, 2, 4, 6 Gy, respectively. The DNA damage was evaluated by micronucleus assay and clonogenic assays. Pretreatment with 5-Aza-CdR inhibited the A549 cell proliferation significantly. More micronucleus were observed after irradiation in 3D cells pretreated with 2 and 5 μmol/L concentration of drug than those without treatment. The survival fractions of cells pretreated by both 2 and 5 μmol/L drug reduced significantly in 3D cultures after irradiation. These significances, however, were found in 2D cells pretreated by only 5 μmol/L drug. Our results suggest that 5-Aza-CdR can inhibit the A549 cells proliferation and apparently enhance the radiosensitivity of cells in 3D cultures. Using of the low dose 5-Aza-CdR in clinical radiotherapy may reduce side effects and enhance effectively the cancer target therapy. (authors)

  1. Inter-experiment variation and dependence on culture conditions in assaying the chemosensitivity of human small cell lung cancer cell lines

    DEFF Research Database (Denmark)

    Roed, H; Christensen, I B; Vindeløv, L L

    1987-01-01

    by a logarithmic function. Even after correction for lack of proportionality the two assay systems provided significantly different dose-response curves. The stability of the chemosensitivity was tested after 25-30 weeks continuous in vitro culture or prolonged storage in liquid nitrogen. One cell line underwent...... significant changes after continuous in vitro culture whereas the cell lines tested after prolonged storage in liquid nitrogen showed only minor changes. It is concluded that instead of considering the concentration necessary to achieve a certain degree of cell kill (e.g. ID50) in one experiment on one cell...

  2. The sensitivity of radiation and its combined effect with anticancer drugs on cultured human lung cancer cells

    International Nuclear Information System (INIS)

    Okada, Shinichiro

    1986-01-01

    The cultured lung tumor cells were irradiated with 60 Co γ-rays, with and without combinations of adriamycin (ADR) and 5-Fluorouracil (5-FU). Cell survival was assayed by colony formation in vitro in soft agar and in plate. Most of small cell lung carcinoma cells used here were radiosensitive, while one of them showed radioresistent comparable to non-small cell lung tumors and other tumor cells. In the treatment with drugs (ADR or 5-FJ) following 60 Co γ-irradiation (postirradiation treatment), it proved to be twice as effective as drug treatment preceding irradiation (preirradiation treatment). Preirradiation treatment demonstrated that 5-FU treated cell were more radiosensitive than ADR treated ones. In the preirradiation treatment, the most enhanced killing effect was observed when cells were irradiated twenty four hours after 5-FU treatment, on the other hand, in the ADR treatment, treatment interval showed no differrece. In the postirradiation treatment, the greatest enhancement was observed when cells were irradiated twenty hours after ADR exposure, while most effective forty eight hours after 5-FU treatment. (author)

  3. Frankincense essential oil prepared from hydrodistillation of Boswellia sacra gum resins induces human pancreatic cancer cell death in cultures and in a xenograft murine model

    Science.gov (United States)

    2012-01-01

    Background Regardless of the availability of therapeutic options, the overall 5-year survival for patients diagnosed with pancreatic cancer remains less than 5%. Gum resins from Boswellia species, also known as frankincense, have been used as a major ingredient in Ayurvedic and Chinese medicine to treat a variety of health-related conditions. Both frankincense chemical extracts and essential oil prepared from Boswellia species gum resins exhibit anti-neoplastic activity, and have been investigated as potential anti-cancer agents. The goals of this study are to identify optimal condition for preparing frankincense essential oil that possesses potent anti-tumor activity, and to evaluate the activity in both cultured human pancreatic cancer cells and a xenograft mouse cancer model. Methods Boswellia sacra gum resins were hydrodistilled at 78°C; and essential oil distillate fractions were collected at different durations (Fraction I at 0–2 h, Fraction II at 8–10 h, and Fraction III at 11–12 h). Hydrodistillation of the second half of gum resins was performed at 100°C; and distillate was collected at 11–12 h (Fraction IV). Chemical compositions were identified by gas chromatography–mass spectrometry (GC-MS); and total boswellic acids contents were quantified by high-performance liquid chromatography (HPLC). Frankincense essential oil-modulated pancreatic tumor cell viability and cytotoxicity were determined by colorimetric assays. Levels of apoptotic markers, signaling molecules, and cell cycle regulators expression were characterized by Western blot analysis. A heterotopic (subcutaneous) human pancreatic cancer xenograft nude mouse model was used to evaluate anti-tumor capability of Fraction IV frankincense essential oil in vivo. Frankincense essential oil-induced tumor cytostatic and cytotoxic activities in animals were assessed by immunohistochemistry. Results Longer duration and higher temperature hydrodistillation produced more abundant high molecular

  4. Tissue culture of osteogenic sarcoma in rats, induced by radioactive phosphorus P-32 and the effect of the anti-cancerous agents on these tumor cells under tissue culture

    International Nuclear Information System (INIS)

    Osaka, Shunzo

    1976-01-01

    Small pieces of osteogenic sarcoma, induced into albino rats of the C.F. Wistar strain by injection of radioactive phosphorus 32 P, were cultured in mixtures of Eagle's minimum essential medium and 20% calf serum. The tumor cells cultured in this way were transplanted into the subcutaneous tissue or the intraabdominal cavity to healthy albino rats. The effect of the anticancerous agents was evaluated by the decrease of nucleic acid composition in these cultured tumor cells. As anti-cancerous agents, cyclophosphamide (CPA), mitomycin C(MMC), and 5-fluorouracil(5-FU) were put into contact with the tumor cells in cultures for two hours under the following dilutions: CPA; 10 -6 , 10 -5 , 10 -4 g/ml. MMC; 2 x 10 -8 , 2 x 10 -7 , 2 x 10 -6 g/ml. 5-FU; 2 x 10 -6 , 2 x 10 -5 , 2 x 10 -4 g/ml. The results are as follows: Three of the seven osteogenic sarcomas in rats were successfully cultured, one of them through more than eighteen generations. After about five hundred thousand cultured cells had been transplanted into the subcutaneous tissues or abdominal cavities of rats, tumors grew in all of them. The histological findings of the tumors in the second generation were quite similar to those of the original tumor. The same process was repeated three times and the tumor showed histogical findings similar to those of the original ones. The capability of nucleic acid synthesis in these cells was decreased at twenty fours after CPA contact and at forty eight hours after MMC. (J.P.N.)

  5. Breast cancer cell lines: friend or foe?

    International Nuclear Information System (INIS)

    Burdall, Sarah E; Hanby, Andrew M; Lansdown, Mark RJ; Speirs, Valerie

    2003-01-01

    The majority of breast cancer research is conducted using established breast cancer cell lines as in vitro models. An alternative is to use cultures established from primary breast tumours. Here, we discuss the pros and cons of using both of these models in translational breast cancer research

  6. Identification of a unique hepatocellular carcinoma line, Li-7, with CD13(+) cancer stem cells hierarchy and population change upon its differentiation during culture and effects of sorafenib.

    Science.gov (United States)

    Yamada, Takeshi; Abei, Masato; Danjoh, Inaho; Shirota, Ryoko; Yamashita, Taro; Hyodo, Ichinosuke; Nakamura, Yukio

    2015-04-11

    Cancer stem cell (CSC) research has highlighted the necessity of developing drugs targeting CSCs. We investigated a hepatocellular carcinoma (HCC) cell line that not only has CSC hierarchy but also shows phenotypic changes (population changes) upon differentiation of CSC during culture and can be used for screening drugs targeting CSC. Based on a hypothesis that the CSC proportion should decrease upon its differentiation into progenitors (population change), we tested HCC cell lines (HuH-7, Li-7, PLC/PRF/5, HLF, HLE) before and after 2 months culture for several markers (CD13, EpCAM, CD133, CD44, CD90, CD24, CD166). Tumorigenicity was tested using nude mice. To evaluate the CSC hierarchy, we investigated reconstructivity, proliferation, ALDH activity, spheroid formation, chemosensitivity and microarray analysis of the cell populations sorted by FACS. Only Li-7 cells showed a population change during culture: the proportion of CD13 positive cells decreased, while that of CD166 positive cells increased. The high tumorigenicity of the Li-7 was lost after the population change. CD13(+)/CD166(-) cells showed slow growth and reconstructed the bulk Li-7 populations composed of CD13(+)/CD166(-), CD13(-)/CD166(-) and CD13(-)/CD166(+) fractions, whereas CD13(-)/CD166(+) cells showed rapid growth but could not reproduce any other population. CD13(+)/CD166(-) cells showed high ALDH activity, spheroid forming ability and resistance to 5-fluorouracil. Microarray analysis demonstrated higher expression of stemness-related genes in CD166(-) than CD166(+) fraction. These results indicated a hierarchy in Li-7 cells, in which CD13(+)/CD166(-) and CD13(-)/CD166(+) cells serve as slow growing CSCs and rapid growing progenitors, respectively. Sorafenib selectively targeted the CD166(-) fraction, including CD13(+) CSCs, which exhibited higher mRNA expression for FGF3 and FGF4, candidate biomarkers for sorafenib. 5-fluorouracil followed by sorafenib inhibited the growth of bulk Li-7

  7. Organoid culture systems for prostate epithelial and cancer tissue

    NARCIS (Netherlands)

    Drost, Jarno; Karthaus, Wouter R; Gao, Dong; Driehuis, Else; Sawyers, Charles L; Chen, Yu; Clevers, Hans

    This protocol describes a strategy for the generation of 3D prostate organoid cultures from healthy mouse and human prostate cells (either bulk or FACS-sorted single luminal and basal cells), metastatic prostate cancer lesions and circulating tumor cells. Organoids derived from healthy material

  8. Cytotoxicity of cancer HeLa cells sensitivity to normal MCF10A cells in cultivations with cell culture medium treated by microwave-excited atmospheric pressure plasmas

    Science.gov (United States)

    Takahashi, Yohei; Taki, Yusuke; Takeda, Keigo; Hashizume, Hiroshi; Tanaka, Hiromasa; Ishikawa, Kenji; Hori, Masaru

    2018-03-01

    Cytotoxic effects of human epithelial carcinoma HeLa cells sensitivity to human mammary epithelial MCF10A cells appeared in incubation with the plasma-activated medium (PAM), where the cell culture media were irradiated with the hollow-shaped contact of a continuously discharged plasma that was sustained by application of a microwave power under Ar gas flow at atmospheric pressure. The discharged plasma had an electron density of 7  ×  1014 cm-3. As the nozzle exit to the plasma source was a distance of 5 mm to the medium, concentrations of 180 µM for H2O2 and 77 µM for NO2- were generated in the PAM for 30 s irradiation, resulting in the control of irradiation periods for aqueous H2O2 with a generation rate of 6.0 µM s-1, and nitrite ion (NO2- ) with a rate of 2.2 µM s-1. Effective concentrations of H2O2 and NO2- for the antitumor effects were revealed in the microwave-excited PAM, with consideration of the complicated reactions at the plasma-liquid interfaces.

  9. The presence of c-erbB-2 gene product-related protein in culture medium conditioned by breast cancer cell line SK-BR-3

    International Nuclear Information System (INIS)

    Alper, O.; Yamaguchi, K.; Hitomi, J.; Honda, S.; Matsushima, T.; Abe, K.

    1990-01-01

    The Mr 185,000 glycoprotein encoded by human c-erbB-2/neu/HER2 gene, termed c-erbB-2 gene product, shows a close structural similarity with epidermal growth factor receptor and is now regarded to be a growth factor receptor for an as yet unidentified ligand. Abundant c-erbB-2 mRNA was demonstrated by Northern blot studies in the human breast cancer cell line SK-BR-3. Cellular radiolabeling experiments followed by immunoprecipitation with three different anti-c-erbB-2 gene product antibodies, recognizing extracellular domain, kinase domain, and carboxyl-terminal portion, respectively, demonstrated the production of a large amount of c-erbB-2 gene product which had the capacity to be phosphorylated. Immunization of mice with concentrated culture medium conditioned by SK-BR-3 cells always generated antibodies against c-erbB-2 gene product, demonstrating that this culture medium contained substance(s) immunologically indistinguishable from c-erbB-2 gene product. This observation was supported by the successful development of a monoclonal antibody against c-erbB-2 gene product, GFD-OA-p185-1, by immunizing mice with this culture medium. The biochemical nature of the substance(s) present in the culture medium was further characterized. When the culture medium conditioned by [35S]cysteine-labeled SK-BR-3 cells was immunoprecipitated by three different anti-c-erbB-2 gene product antibodies, only the antibody recognizing extracellular domain precipitated the [35S]-labeled protein with a molecular weight of 110,000, namely p110. The newly developed monoclonal antibody also immunoprecipitated this protein

  10. Cytotoxicity and apoptotic inducibility of Vitex agnus-castus fruit extract in cultured human normal and cancer cells and effect on growth.

    Science.gov (United States)

    Ohyama, Kunio; Akaike, Takenori; Hirobe, Chieko; Yamakawa, Toshio

    2003-01-01

    A crude extract was prepared with ethanol from dried ripened Vitex agnus-castus fruits growing in Israel (Vitex extract). Cytotoxicity of the extract against human uterine cervical canal fibroblast (HCF), human embryo fibroblast (HE-21), ovarian cancer (MCF-7), cervical carcinoma (SKG-3a), breast carcinoma (SKOV-3), gastric signet ring carcinoma (KATO-III), colon carcinoma (COLO 201), and small cell lung carcinoma (Lu-134-A-H) cells was examined. After culture for 24 h (logarithmic growth phase) or 72 h (stationary growth phase), the cells were treated with various concentrations of Vitex extract. In both growth phases, higher growth activity of cells and more cytotoxic activity of Vitex extract were seen. The cytotoxic activity against stationary growth-phase cells was less than that against logarithmic growth-phase cells. DNA fragmentation of Vitex extract-treated cells was seen in SKOV-3, KATO-III, COLO 201, and Lu-134-A-H cells. The DNA fragmentation in Vitex extract-treated KATO-III cells was inhibited by the presence of the antioxidative reagent pyrrolidine dithiocarbamate or N-acetyl-L-cysteine (NAC). Western blotting analysis showed that in Vitex extract-treated KATO-III cells, the presence of NAC also inhibited the expression of heme oxygenase-1 and the active forms of caspases-3, -8 and -9. It is concluded that the cytotoxic activity of Vitex extract may be attributed to the effect on cell growth, that cell death occurs through apoptosis, and that this apoptotic cell death may be attributed to increased intracellular oxidation by Vitex extract treatment.

  11. Utility and translatability of mathematical modeling, cell culture and small and large animal models in magnetic nanoparticle hyperthermia cancer treatment research

    Science.gov (United States)

    Hoopes, P. J.; Petryk, Alicia A.; Misra, Adwiteeya; Kastner, Elliot J.; Pearce, John A.; Ryan, Thomas P.

    2015-03-01

    For more than 50 years, hyperthermia-based cancer researchers have utilized mathematical models, cell culture studies and animal models to better understand, develop and validate potential new treatments. It has been, and remains, unclear how and to what degree these research techniques depend on, complement and, ultimately, translate accurately to a successful clinical treatment. In the past, when mathematical models have not proven accurate in a clinical treatment situation, the initiating quantitative scientists (engineers, mathematicians and physicists) have tended to believe the biomedical parameters provided to them were inaccurately determined or reported. In a similar manner, experienced biomedical scientists often tend to question the value of mathematical models and cell culture results since those data typically lack the level of biologic and medical variability and complexity that are essential to accurately study and predict complex diseases and subsequent treatments. Such quantitative and biomedical interdependence, variability, diversity and promise have never been greater than they are within magnetic nanoparticle hyperthermia cancer treatment. The use of hyperthermia to treat cancer is well studied and has utilized numerous delivery techniques, including microwaves, radio frequency, focused ultrasound, induction heating, infrared radiation, warmed perfusion liquids (combined with chemotherapy), and, recently, metallic nanoparticles (NP) activated by near infrared radiation (NIR) and alternating magnetic field (AMF) based platforms. The goal of this paper is to use proven concepts and current research to address the potential pathobiology, modeling and quantification of the effects of treatment as pertaining to the similarities and differences in energy delivered by known external delivery techniques and iron oxide nanoparticles.

  12. Stimulation of MMP-11 (stromelysin-3) expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

    International Nuclear Information System (INIS)

    Selvey, Saxon; Haupt, Larisa M; Thompson, Erik W; Matthaei, Klaus I; Irving, Michael G; Griffiths, Lyn R

    2004-01-01

    Matrix metalloproteinases (MMPs) are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14) and stromelysin-3 (MMP-11) are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs) were: a) treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b) grown on collagens I, IV and V; c) treated with fibronectin, con-A and matrigel; and d) co-cultured with a range of HBC (human breast cancer) cell lines of varied invasive and metastatic potential. Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms

  13. Stimulation of MMP-11 (stromelysin-3 expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

    Directory of Open Access Journals (Sweden)

    Matthaei Klaus I

    2004-07-01

    Full Text Available Abstract Background Matrix metalloproteinases (MMPs are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14 and stromelysin-3 (MMP-11 are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. Methods To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs were: a treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b grown on collagens I, IV and V; c treated with fibronectin, con-A and matrigel; and d co-cultured with a range of HBC (human breast cancer cell lines of varied invasive and metastatic potential. Results Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. Conclusion We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms.

  14. Proteomic response to 5,6-dimethylxanthenone 4-acetic acid (DMXAA, vadimezan in human non-small cell lung cancer A549 cells determined by the stable-isotope labeling by amino acids in cell culture (SILAC approach

    Directory of Open Access Journals (Sweden)

    Pan ST

    2015-02-01

    Full Text Available Shu-Ting Pan,1,* Zhi-Wei Zhou,2,3,* Zhi-Xu He,3 Xueji Zhang,4 Tianxin Yang,5 Yin-Xue Yang,6 Dong Wang,7 Jia-Xuan Qiu,1 Shu-Feng Zhou2 1Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, People’s Republic of China; 2Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA; 3Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, 4Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 5Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA; 6Department of Colorectal Surgery, General Hospital of Ningxia Medical University, Yinchuan, 7Cancer Center, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, People’s Republic of China *These two authors contributed equally to this work Abstract: 5,6-Dimethylxanthenone 4-acetic acid (DMXAA, also known as ASA404 and vadimezan, is a potent tumor blood vessel-disrupting agent and cytokine inducer used alone or in combination with other cytotoxic agents for the treatment of non-small cell lung cancer (NSCLC and other cancers. However, the latest Phase III clinical trial has shown frustrating outcomes in the treatment of NSCLC, since the therapeutic targets and underlying mechanism for the anticancer effect of DMXAA are not yet fully understood. This study aimed to examine the proteomic response to DMXAA and unveil the global molecular targets and possible mechanisms for the anticancer effect of DMXAA in NSCLC A549 cells using a stable-isotope labeling by amino acids in cell culture (SILAC approach. The proteomic data showed that treatment with DMXAA

  15. Colorectal cancer stem cells.

    Science.gov (United States)

    Salama, Paul; Platell, Cameron

    2009-10-01

    Somatic stem cells reside at the base of the crypts throughout the colonic mucosa. These cells are essential for the normal regeneration of the colonic epithelium. The stem cells reside within a special 'niche' comprised of intestinal sub-epithelial myofibroblasts that tightly control their function. It has been postulated that mutations within these adult colonic stem cells may induce neoplastic changes. Such cells can then dissociate from the epithelium and travel into the mesenchyme and thus form invasive cancers. This theory is based on the observation that within a colon cancer, less than 1% of the neoplastic cells have the ability to regenerate the tumour. It is this group of cells that exhibits characteristics of colonic stem cells. Although anti-neoplastic agents can induce remissions by inhibiting cell division, the stem cells appear to be remarkably resistant to both standard chemotherapy and radiotherapy. These stem cells may therefore persist after treatment and form the nucleus for cancer recurrence. Hence, future treatment modalities should focus specifically on controlling the cancer stem cells. In this review, we discuss the biology of normal and malignant colonic stem cells.

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

    International Nuclear Information System (INIS)

    Hamada, Shin; Masamune, Atsushi; Takikawa, Tetsuya; Suzuki, Noriaki; Kikuta, Kazuhiro; Hirota, Morihisa; Hamada, Hirofumi; Kobune, Masayoshi; Satoh, Kennichi; Shimosegawa, Tooru

    2012-01-01

    Highlights: ► Pancreatic stellate cells (PSCs) promote the progression of pancreatic cancer. ► Pancreatic cancer cells co-cultured with PSCs showed enhanced spheroid formation. ► Expression of stem cell-related genes ABCG2, Nestin and LIN28 was increased. ► Co-injection of PSCs enhanced tumorigenicity of pancreatic cancer cells in vivo. ► 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.

  17. In Vitro Production of Echioidinin, 7-O-Methywogonin from Callus Cultures of Andrographis lineata and Their Cytotoxicity on Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Arifullah Mohammed

    Full Text Available Andrographis lineata is an herbal medicinal plant used in traditional medicine as a substitute for Andrographis paniculata. Here, using mature leaf explants of A. lineata we demonstrate for the first time the callus induction established on MS medium containing 1.0 mg l-1 IAA. Dried callus was subjected to solvent extraction with acetone. Further the acetone residue was separated by silica gel column chromatography, crystallized and characterized on the basis of nuclear magnetic resonance (proton and c13 and liquid chromatographic mass spectroscopy. This analysis revealed the occurrence of two known flavones namely, 7-O-methylwogonin (MW and Echioidinin (ED. Furthermore, these compounds were tested for their cytotoxicity against leukemic cell line, CEM. We identify that ED and MW induced cytotoxicity in a time- and concentration-dependent manner. Further increase in the LDH release upon treatment with ED and MW further confirmed our cytotoxicity results against leukemic cell line. Strikingly, MW was more potent than ED when compared by trypan blue and MTT assays. Our results recapitulate the utility of callus cultures for the production of plant specific bioactive secondary metabolites instead of using wild plants. Together, our in vitro studies provide new insights of A. lineata callus cultures serving as a source for cancer chemotherapeutic agents.

  18. In Vitro Production of Echioidinin, 7-O-Methywogonin from Callus Cultures of Andrographis lineata and Their Cytotoxicity on Cancer Cells

    Science.gov (United States)

    Mohammed, Arifullah; Chiruvella, Kishore K.; Rao, Yerra Koteswara; Geethangili, Madamanchi; Raghavan, Sathees C.; Ghanta, Rama Gopal

    2015-01-01

    Andrographis lineata is an herbal medicinal plant used in traditional medicine as a substitute for Andrographis paniculata. Here, using mature leaf explants of A. lineata we demonstrate for the first time the callus induction established on MS medium containing 1.0 mg l–1 IAA. Dried callus was subjected to solvent extraction with acetone. Further the acetone residue was separated by silica gel column chromatography, crystallized and characterized on the basis of nuclear magnetic resonance (proton and c13) and liquid chromatographic mass spectroscopy. This analysis revealed the occurrence of two known flavones namely, 7-O-methylwogonin (MW) and Echioidinin (ED). Furthermore, these compounds were tested for their cytotoxicity against leukemic cell line, CEM. We identify that ED and MW induced cytotoxicity in a time- and concentration-dependent manner. Further increase in the LDH release upon treatment with ED and MW further confirmed our cytotoxicity results against leukemic cell line. Strikingly, MW was more potent than ED when compared by trypan blue and MTT assays. Our results recapitulate the utility of callus cultures for the production of plant specific bioactive secondary metabolites instead of using wild plants. Together, our in vitro studies provide new insights of A. lineata callus cultures serving as a source for cancer chemotherapeutic agents. PMID:26488879

  19. Stages of Renal Cell Cancer

    Science.gov (United States)

    ... Tumors Treatment Genetics of Kidney Cancer Research Renal Cell Cancer Treatment (PDQ®)–Patient Version General Information About Renal Cell Cancer Go to Health Professional Version Key Points Renal ...

  20. In vitro and in vivo MMP gene expression localisation by In Situ-RT-PCR in cell culture and paraffin embedded human breast cancer cell line xenografts

    International Nuclear Information System (INIS)

    Haupt, Larisa M; Thompson, Erik W; Trezise, Ann EO; Irving, Rachel E; Irving, Michael G; Griffiths, Lyn R

    2006-01-01

    Members of the matrix metalloproteinase (MMP) family of proteases are required for the degradation of the basement membrane and extracellular matrix in both normal and pathological conditions. In vitro, MT1-MMP (MMP-14, membrane type-1-MMP) expression is higher in more invasive human breast cancer (HBC) cell lines, whilst in vivo its expression has been associated with the stroma surrounding breast tumours. MMP-1 (interstitial collagenase) has been associated with MDA-MB-231 invasion in vitro, while MMP-3 (stromelysin-1) has been localised around invasive cells of breast tumours in vivo. As MMPs are not stored intracellularly, the ability to localise their expression to their cells of origin is difficult. We utilised the unique in situ-reverse transcription-polymerase chain reaction (IS-RT-PCR) methodology to localise the in vitro and in vivo gene expression of MT1-MMP, MMP-1 and MMP-3 in human breast cancer. In vitro, MMP induction was examined in the MDA-MB-231 and MCF-7 HBC cell lines following exposure to Concanavalin A (Con A). In vivo, we examined their expression in archival paraffin embedded xenografts derived from a range of HBC cell lines of varied invasive and metastatic potential. Mouse xenografts are heterogenous, containing neoplastic human parenchyma with mouse stroma and vasculature and provide a reproducible in vivo model system correlated to the human disease state. In vitro, exposure to Con A increased MT1-MMP gene expression in MDA-MB-231 cells and decreased MT1-MMP gene expression in MCF-7 cells. MMP-1 and MMP-3 gene expression remained unchanged in both cell lines. In vivo, stromal cells recruited into each xenograft demonstrated differences in localised levels of MMP gene expression. Specifically, MDA-MB-231, MDA-MB-435 and Hs578T HBC cell lines are able to influence MMP gene expression in the surrounding stroma. We have demonstrated the applicability and sensitivity of IS-RT-PCR for the examination of MMP gene expression both in vitro and in

  1. Cultural aspects of communication in cancer care.

    Science.gov (United States)

    Surbone, Antonella

    2008-03-01

    Cultural competence in oncology requires the acquisition of specific knowledge, clinical skills, and attitudes that facilitate effective cross-cultural negotiation in the clinical setting, thus, leading to improved therapeutic outcomes and decreased disparities in cancer care. Cultural competence in oncology entails a basic knowledge of different cultural attitudes and practices of communication of the truth and of decision-making styles throughout the world. Cultural competence always presupposes oncology professionals' awareness of their own cultural beliefs and values. To be able to communicate with cancer patients in culturally sensitive ways, oncologists should have knowledge of the concept of culture in its complexity and of the risks of racism, classism, sexism, ageism, and stereotyping that must be avoided in clinical practice. Oncologists should develop a sense of appreciation for differences in health care values, based on the recognition that no culture can claim hegemony over others and that cultures are evolving under their reciprocal influence on each other. Medical schools and oncology training can teach communication skills and cultural competence, while fostering in all students and young doctors those attitudes of humility, empathy, curiosity, respect, sensitivity, and awareness that are needed to deliver effective and culturally sensitive cancer care.

  2. Mutation in cultured mammalian cells

    International Nuclear Information System (INIS)

    Nakamura, N.; Okada, S.

    1982-01-01

    Mammalian cell cultures were exposed to gamma-rays at various dose rates. Dose-rate effects were observed in cultured somatic cells of the mouse for cell killing and mutations resistant to 6-thioguanine (TGsup(r)) and to methotrexate (MTXsup(r)). Linear quadratic model may be applied to cell killing and TGsup(r) mutations in some cases but can not explain the whole data. Results at low doses with far low dose-rate were not predictable from data at high doses with acute or chronic irradiation. Radioprotective effects of dimethyl sulfoxide were seen only after acute exposure but not after chronic one, suggesting that damages by indirect action of radiations may be potentially reparable by cells. TGsup(r) mutations seem to contain gross structural changes whereas MTXsup(r) ones may have smaller alterations. (Namekawa, K.)

  3. Fingerprints in cancer cells

    International Nuclear Information System (INIS)

    Servomaa, K.

    1994-01-01

    Gene research has shown that factors causing cancer, or carcinogens, may leave marks typical of each particular carcinogen (fingerprints) in the genotype of the cell. Radiation, for instance, may leave such fingerprints in a cancer cell. In particular, the discovery of a gene called p53 has yielded much new information on fingerprints. It has been discovered, for example, that toxic fungus and UV-radiation each leave fingerprints in the p53 gene. Based on the detection of fingerprints, it may be possible in the future to tell a cancer patient what factor had trigged the maglinancy

  4. Cell culture compositions

    Science.gov (United States)

    Dunn-Coleman, Nigel; Goedegebuur, Frits; Ward, Michael; Yiao, Jian

    2014-03-18

    The present invention provides a novel endoglucanase nucleic acid sequence, designated egl6 (SEQ ID NO:1 encodes the full length endoglucanase; SEQ ID NO:4 encodes the mature form), and the corresponding endoglucanase VI amino acid sequence ("EGVI"; SEQ ID NO:3 is the signal sequence; SEQ ID NO:2 is the mature sequence). The invention also provides expression vectors and host cells comprising a nucleic acid sequence encoding EGVI, recombinant EGVI proteins and methods for producing the same.

  5. Stem cells and cancer: A review

    Directory of Open Access Journals (Sweden)

    Najeeb Ullah

    2016-05-01

    Full Text Available Stem cells are the small units of multicellular creature. Regeneration and self-renewal are the ability of the stem cells. Each tissue is having particular stem cells, specific to it. These normal stem cells are converted into cancer stem cells through mutations in it. Although the expression of oncogenes is enhanced a lot, the tumor-supressing gene is lessened. Cancer stem cells are isolated and visualized through different techniques like immunocytochemical staining, spectral karyotyping, immunohistochemistry, induction method and dissection measures, then are performed histological procedures which include fascination, immunohistochemistry, dispensation, in situ hybridization and also quantitative examination of tissue flow cytometric analysis. For the analysis of quantization, statistical tests are also performed as two-sample t-test, Chi-square test, SD and arithmetic mean. Tumor cells generate glioma spheres. These are used in cancer study. Axin 1 is the gene suppressing cancer. Its removal causes the generation of liver cancer. Curcumin is the most effective for suppressing cancer as it increases the normal stem cell function and decreases the cancer stem cell function. Brahma-related gene 1 is crucial for the safeguarding of the stem cell residents in tissue-specific comportment. Different types of cancers originate through genetic mutation, tissue disorganization and cell proliferation. Tumor configuration is produced by the alteration in original cell culture having stem cells and progenitor cell populations. The developmental facets about cancer cells and cancer stem cells as well as their personal natal functions sustain an intricate steadiness to settle on their personal donations to the efficacy or harmfulness of the biological organization.

  6. Cultural aspects of communication in cancer care.

    Science.gov (United States)

    Surbone, A

    2006-01-01

    Cancer is increasing in incidence and prevalence worldwide, and the WHO has recently included cancer and its treatments as a health priority in developed and developing countries. The cultural diversity of oncology patients is bound to increase, and cultural sensitivity and competence are now required of all oncology professionals. A culturally competent cancer care leads to improved therapeutic outcome and it may decrease disparities in medical care. Cultural competence in medicine is a complex multilayered accomplishment, requiring knowledge, skills and attitudes whose acquisition is needed for effective cross-cultural negotiation in the clinical setting. Effective cultural competence is based on knowledge of the notion of culture; on awareness of possible biases and prejudices related to stereotyping, racism, classism, sexism; on nurturing appreciation for differences in health care values; and on fostering the attitudes of humility, empathy, curiosity, respect, sensitivity and awareness. Cultural competence in healthcare relates to individual professionals, but also to organizations and systems. A culturally competent healthcare system must consider in their separateness and yet in there reciprocal influences social, racial and cultural factors. By providing a framework of reference to interpret the external world and relate to it, culture affects patients' perceptions of disease, disability and suffering; degrees and expressions of concern about them; their responses to treatments and their relationship to individual physicians and to the healthcare system. Culture also influences the interpretation of ethical norms and principles, and especially of individual autonomy, which can be perceived either as synonymous with freedom or with isolation depending on the cultural context. This, in turn, determines the variability of truth-telling attitudes and practices worldwide as well as the different roles of family in the information and decision-making process of

  7. Collagen gel droplet-embedded culture drug sensitivity test for adjuvant chemotherapy after complete resection of non-small-cell lung cancer.

    Science.gov (United States)

    Inoue, Masayoshi; Maeda, Hajime; Takeuchi, Yukiyasu; Fukuhara, Kenjiro; Shintani, Yasushi; Funakoshi, Yasunobu; Funaki, Soichiro; Nojiri, Takashi; Kusu, Takashi; Kusumoto, Hidenori; Kimura, Toru; Okumura, Meinoshin

    2018-04-01

    We conducted a prospective clinical study to individualize adjuvant chemotherapy after complete resection of non-small-cell lung cancer (NSCLC), based on the drug sensitivity test. Patients with resectable c-stage IB-IIIA NSCLC were registered between 2005 and 2010. We performed the collagen gel droplet-embedded culture drug sensitivity test (CD-DST) on a fresh surgical specimen to assess in vitro chemosensitivity and evaluated the prognostic outcome after adjuvant chemotherapy with carboplatin/paclitaxel based on the CD-DST. Among 92 registered patients, 87 were eligible for inclusion in the analysis. The success rate of CD-DST was 86% and chemosensitivity to carboplatin and/or paclitaxel was evident in 57 (76%) of the 75 patients. Adjuvant chemotherapy was completed in 22 (73%) of 30 patients. The 5-year overall survival rates were 71, 73, and 75% for all, CD-DST success, and chemosensitive patients, respectively. The 5-year disease-free survival and overall survival rates of the chemosensitive patients who completed adjuvant chemotherapy using carboplatin/paclitaxel were 68 and 82%, respectively. The 5-year disease-free survival and overall survival rates of the patients with stage II-IIIA chemosensitive NSCLC were 58 and 75%, respectively. Comparative analyses of the chemosensitive and non-chemosensitive/CD-DST failure groups showed no significant survival difference. CD-DST can be used to evaluate chemosensitivity after lung cancer surgery; however, its clinical efficacy for assessing individualized treatment remains uncertain.

  8. Dextran sulphate crowding and sodium deoxycholate lysis of primary breast fibroblast cells achieve extracellular matrix deposition and decellularization for breast cancer stem cell culture

    Directory of Open Access Journals (Sweden)

    Aroem Naruni

    2016-01-01

    .Extracellular matrix provides tissue integrity, acts as a native scaffold for cell attachment and interaction and alsoserves as reservoir for growth factors. The aim of this experiment w asto achieve the deposition and decellularizationof ECM.Methods: Cells crowder have been developed to increase ECM deposit in the primary breast fibroblast cells layerobtained from isolation of single cell from breast mammoplasty specimen. Five hundred kDa dextran was addedinto DMEM medium containing 0.5% fetal bovine serum (FBS and 100 μm of L-ascorbic acid 2-phosphate. Afterseven days, cells were lysed by sodium deoxycholate (DOC. Results were observed in a fluorescence microscope.Results: Extracellular matrix deposition and decellularization of primary breast fibroblast cells were detected byusing extracellular matrix protein, fibronectin via rabbit anti human fibronectin and anti-rabbit IgG conjugated withAlexa Fluor 488.Conclusion: Dextran sulphate increased extracellular matrix deposit in primary breast fibroblast cell layer and thetreatment with sodium deoxycolate lysed cells resulted in extracellular matrix decellularization. (Health ScienceJournal of Indonesia 2015;6:43-7Keywords: extracellular matrix, breast cancer stem cell, breast fibroblast cell

  9. Extragonadal Germ Cell Cancer (EGC)

    Science.gov (United States)

    The Testicular Cancer Resource Center Extragonadal Germ Cell Cancer (EGC) 95% of all testicular tumors are germ cell tumors. That is, the tumors originate in the sperm forming cells in the testicles ( ...

  10. Cancer stem cells revisited

    NARCIS (Netherlands)

    Batlle, Eduard; Clevers, Hans

    2017-01-01

    The cancer stem cell (CSC) concept was proposed four decades ago, and states that tumor growth, analogous to the renewal of healthy tissues, is fueled by small numbers of dedicated stem cells. It has gradually become clear that many tumors harbor CSCs in dedicated niches, and yet their

  11. 9 CFR 101.6 - Cell cultures.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Cell cultures. 101.6 Section 101.6..., SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS DEFINITIONS § 101.6 Cell cultures. When used in conjunction with or in reference to cell cultures, which may be referred to as tissue cultures...

  12. Youth Culture and Cell Phone

    Directory of Open Access Journals (Sweden)

    mohammad saeed zokaei

    2009-11-01

    Full Text Available Iranian youth’s leisure culture has been immediately affected by the digital media culture. As a communicative media, cell phone has crossed borders of youth norms and identity; and in addition to facilitating their communication, has changed its patterns. Applying Bourdieu’s concepts of habitus and field, and relied on the qualitative and quantitative data gathered from the mobile youth users, the present study argues that mobile has produced a new field in which youth’s opportunities for leisure, entertainment, communication, and independence have extended. In addition, cell phone has facilitated and compensated for some defects in public sphere, and therefore empowered youth agency, individuality, and power. Despite this strengthening, cell phone does not cross borders of gender and class differences, or the levels of social capital.

  13. Cultural beliefs and values in cancer patients.

    Science.gov (United States)

    Daher, M

    2012-04-01

    In 2008, the International Agency for Research on Cancer (IARC) released its World Cancer Report, which indicated that cancer accounts for approximately 12% of all-cause mortality worldwide. IARC estimated that globally 7.6 million people died from cancer and that 12.4 million new cases were diagnosed in 2008. The report went on to project that, due to increases in life expectancy, improvements in clinical diagnostics, and shifting trends in health behaviors (e.g. increases in smoking and sedentary lifestyles), in the absence of significant efforts to improve global cancer control, cancer mortality could increase to 12.9 million and cancer incidence to 20 million by the year 2030. Looking deeper into the data, it becomes clear that cancer-related stigma and myths about cancer are important problems that must be addressed, although different from a country to another. Stigmas about cancer present significant challenges to cancer control: stigma can have a silencing effect, whereby efforts to increase cancer awareness are negatively affected. The social, emotional, and financial devastation that all too often accompanies a diagnosis of cancer is, in large part, due to the cultural myths and taboos surrounding the disease. Combating stigma, myths, taboos, and overcoming silence will play important roles in changing this provisional trajectory. There are several reasons that cancer is stigmatized. Many people in our area perceived cancer to be a fatal disease. Cancer symptoms or body parts affected by the disease can cultivate stigma. Fears about treatment can also fuel stigma. There was evidence of myths associated with cancer, such as the belief that cancer is contagious, or cancer may be seen as a punishment. After reviewing these different examples of cultural myths and taboos met in cancer care, we can report these lessons learned: 1. Around the world, cancer continues to carry a significant amount of stigma, myths, and taboos; however, there are opportunities to

  14. The Use of Normal Colon Cell Culture to Assess Toxicities and Cancer Molecular Pathway Alterations Induced by Disinfection Byproducts.

    Science.gov (United States)

    Recent Epidemiological studies have linked the consumption of disinfected surface waters to an increased risk of colorectal cancer (Bove, GE, Jr et al., Int. J. Health Geogr., 6:18, 2007). Approximately 600 disinfection byproducts (DBP) have been identified. Because it would be...

  15. Cancer Stem Cells in Pancreatic Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Bao, Qi; Zhao, Yue; Renner, Andrea; Niess, Hanno; Seeliger, Hendrik; Jauch, Karl-Walter; Bruns, Christiane J., E-mail: christiane.bruns@med.uni-muenchen.de [Department of Surgery, Ludwig Maximilian University of Munich, Klinikum Grosshadern, Marchioninistr. 15, D-81377, Munich (Germany)

    2010-08-19

    Pancreatic cancer is an aggressive malignant solid tumor well-known by early metastasis, local invasion, resistance to standard chemo- and radiotherapy and poor prognosis. Increasing evidence indicates that pancreatic cancer is initiated and propagated by cancer stem cells (CSCs). Here we review the current research results regarding CSCs in pancreatic cancer and discuss the different markers identifying pancreatic CSCs. This review will focus on metastasis, microRNA regulation and anti-CSC therapy in pancreatic cancer.

  16. Cancer Stem Cells in Pancreatic Cancer

    Science.gov (United States)

    Bao, Qi; Zhao, Yue; Renner, Andrea; Niess, Hanno; Seeliger, Hendrik; Jauch, Karl-Walter; Bruns, Christiane J.

    2010-01-01

    Pancreatic cancer is an aggressive malignant solid tumor well-known by early metastasis, local invasion, resistance to standard chemo- and radiotherapy and poor prognosis. Increasing evidence indicates that pancreatic cancer is initiated and propagated by cancer stem cells (CSCs). Here we review the current research results regarding CSCs in pancreatic cancer and discuss the different markers identifying pancreatic CSCs. This review will focus on metastasis, microRNA regulation and anti-CSC therapy in pancreatic cancer. PMID:24281178

  17. Cancer Stem Cells in Pancreatic Cancer

    Directory of Open Access Journals (Sweden)

    Karl-Walter Jauch

    2010-08-01

    Full Text Available Pancreatic cancer is an aggressive malignant solid tumor well-known by early metastasis, local invasion, resistance to standard chemo- and radiotherapy and poor prognosis. Increasing evidence indicates that pancreatic cancer is initiated and propagated by cancer stem cells (CSCs. Here we review the current research results regarding CSCs in pancreatic cancer and discuss the different markers identifying pancreatic CSCs. This review will focus on metastasis, microRNA regulation and anti-CSC therapy in pancreatic cancer.

  18. Long term imaging of living brain cancer cells

    Science.gov (United States)

    Farias, Patricia M. A.; Galembeck, André; Milani, Raquel; Andrade, Arnaldo C. D. S.; Stingl, Andreas

    2018-02-01

    QDs synthesized in aqueous medium and functionalized with polyethylene glycol were used as fluorescent probes. They label and monitor living healthy and cancer brain glial cells in culture. Physical-chemical characterization was performed. Toxicological studies were performed by in vivo short and long-term inhalation in animal models. Healthy and cancer glial living cells were incubated in culture media with highly controlled QDs. Specific features of glial cancer cells were enhanced by QD labelling. Cytoplasmic labelling pattern was clearly distinct for healthy and cancer cells. Labelled cells kept their normal activity for same period as non-labelled control samples.

  19. Determination of in vitro free radical scavenging and antiproliferative effect of Pennisetum alopecuroides on cultured A549 human lung cancer cells

    Directory of Open Access Journals (Sweden)

    Githa Elizabeth Mathew

    2015-01-01

    Conclusions: This is the 1 st time a pharmacological exploration of P. alopecuroides grasses has been conducted. We have shown that P. alopecuroides exhibits good free radical scavenging and strong in vitro cytotoxic activities against human lung cancer cell lines.

  20. Human breast cancer histoid: an in vitro 3-dimensional co-culture model that mimics breast cancer tissue.

    Science.gov (United States)

    Kaur, Pavinder; Ward, Brenda; Saha, Baisakhi; Young, Lillian; Groshen, Susan; Techy, Geza; Lu, Yani; Atkinson, Roscoe; Taylor, Clive R; Ingram, Marylou; Imam, S Ashraf

    2011-12-01

    Progress in our understanding of heterotypic cellular interaction in the tumor microenvironment, which is recognized to play major roles in cancer progression, has been hampered due to unavailability of an appropriate in vitro co-culture model. The aim of this study was to generate an in vitro 3-dimensional human breast cancer model, which consists of cancer cells and fibroblasts. Breast cancer cells (UACC-893) and fibroblasts at various densities were co-cultured in a rotating suspension culture system to establish co-culture parameters. Subsequently, UACC-893, BT.20, or MDA.MB.453 were co-cultured with fibroblasts for 9 days. Co-cultures resulted in the generation of breast cancer histoid (BCH) with cancer cells showing the invasion of fibroblast spheroids, which were visualized by immunohistochemical (IHC) staining of sections (4 µm thick) of BCH. A reproducible quantitative expression of C-erbB.2 was detected in UACC-893 cancer cells in BCH sections by IHC staining and the Automated Cellular Imaging System. BCH sections also consistently exhibited qualitative expression of pancytokeratins, p53, Ki-67, or E-cadherin in cancer cells and that of vimentin or GSTPi in fibroblasts, fibronectin in the basement membrane and collagen IV in the extracellular matrix. The expression of the protein analytes and cellular architecture of BCH were markedly similar to those of breast cancer tissue.

  1. Bioenergetic and antiapoptotic properties of mitochondria from cultured human prostate cancer cell lines PC-3, DU145 and LNCaP.

    Directory of Open Access Journals (Sweden)

    Alexander Panov

    Full Text Available The purpose of this work was to reveal the metabolic features of mitochondria that might be essential for inhibition of apoptotic potential in prostate cancer cells. We studied mitochondria isolated from normal prostate epithelial cells (PrEC, metastatic prostate cancer cell lines LNCaP, PC-3, DU145; and non-prostate cancer cells - human fibrosarcoma HT1080 cells; and normal human lymphoblastoid cells. PrEC cells contained 2 to 4 times less mitochondria per gram of cells than the three PC cell lines. Respiratory activities of PrEC cell mitochondria were 5-20-fold lower than PC mitochondria, depending on substrates and the metabolic state, due to lower content and lower activity of the respiratory enzyme complexes. Mitochondria from the three metastatic prostate cancer cell lines revealed several features that are distinctive only to these cells: low affinity of Complex I for NADH, 20-30 mV higher electrical membrane potential (ΔΨ. Unprotected with cyclosporine A (CsA the PC-3 mitochondria required 4 times more Ca²⁺ to open the permeability transition pore (mPTP when compared with the PrEC mitochondria, and they did not undergo swelling even in the presence of alamethicin, a large pore forming antibiotic. In the presence of CsA, the PC-3 mitochondria did not open spontaneously the mPTP. We conclude that the low apoptotic potential of the metastatic PC cells may arise from inhibition of the Ca²⁺-dependent permeability transition due to a very high ΔΨ and higher capacity to sequester Ca²⁺. We suggest that due to the high ΔΨ, mitochondrial metabolism of the metastatic prostate cancer cells is predominantly based on utilization of glutamate and glutamine, which may promote development of cachexia.

  2. c-Myc-Dependent Cell Competition in Human Cancer Cells.

    Science.gov (United States)

    Patel, Manish S; Shah, Heta S; Shrivastava, Neeta

    2017-07-01

    Cell Competition is an interaction between cells for existence in heterogeneous cell populations of multicellular organisms. This phenomenon is involved in initiation and progression of cancer where heterogeneous cell populations compete directly or indirectly for the survival of the fittest based on differential gene expression. In Drosophila, cells having lower dMyc expression are eliminated by cell competition through apoptosis when present in the milieu of cells having higher dMyc expression. Thus, we designed a study to develop c-Myc (human homolog) dependent in vitro cell competition model of human cancer cells. Cells with higher c-Myc were transfected with c-myc shRNA to prepare cells with lower c-Myc and then co-cultured with the same type of cells having a higher c-Myc in equal ratio. Cells with lower c-Myc showed a significant decrease in numbers when compared with higher c-Myc cells, suggesting "loser" and "winner" status of cells, respectively. During microscopy, engulfment of loser cells by winner cells was observed with higher expression of JNK in loser cells. Furthermore, elimination of loser cells was prevented significantly, when co-cultured cells were treated with the JNK (apoptosis) inhibitor. Above results indicate elimination of loser cells in the presence of winner cells by c-Myc-dependent mechanisms of cell competition in human cancer cells. This could be an important mechanism in human tumors where normal cells are eliminated by c-Myc-overexpressed tumor cells. J. Cell. Biochem. 118: 1782-1791, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  3. Selection of radioresistant cells by vitamin A deficiency in a small cell lung cancer cell line

    International Nuclear Information System (INIS)

    Terasaki, Takeo; Shimosato, Yukio; Wada, Makio; Yokota, Jun; Terada, Masaaki

    1990-01-01

    Radiation sensitivity of a human small cell lung cancer cell line, Lu-134-B cells, cultured in serum-supplemented medium and of cells transferred to and cultured in delipidized serum-supplemented (vitamin A-deficient) medium was studied. The cells cultured in serum-supplemented medium showed the phenotype of classic small cell lung cancer sensitive to radiation, while cells transferred to delipidized serum-supplemented medium showed partial squamous cell differentiation and became resistant to radiation. These results suggest that some small cell lung cancer cells in vitro change their morphology and radiosensitivity depending on the culture conditions. The change in radiosensitivity was reproducible, and was not reversible by culture of the radioresistant cells in delipidized serum-supplemented medium with addition of retinoic acid (vitamin A-sufficient medium) for two months, although squamous cells disappeared. Acquisition of radioresistancy was considered to occur as the result of clonal selective growth in delipidized medium of a minor cell population in the original cell culture, based on a study of chromosome number. It was also found that there was no association of myc-family oncogenes with the changes of radiosensitivity in this cell line. (author)

  4. Melphalan metabolism in cultured cells

    International Nuclear Information System (INIS)

    Seagrave, J.C.; Valdez, J.G.; Tobey, R.A.; Gurley, L.R.

    1985-06-01

    Procedures are presented for the adaptation of reversed-phase-HPLC methods to accomplish separation and isolation of the cancer therapeutic drug melphalan (L-phenylalanine mustard) and its metabolic products from whole cells. Five major degradation products of melphalan were observed following its hydrolysis in phosphate buffer in vitro. The two most polar of these products (or modifications of them) were also found in the cytosol of Chinese hamster CHO cells. The amounts of these two polar products (shown not to be mono- or dihydroxymelphalan) were significantly changed by the pretreatment of cells with ZnC1 2 , one being increased in amount while the other was reduced to an insignificant level. In ZnC1 2 -treated cells, there was also an increased binding of melphalan (or its derivatives) to one protein fraction resolved by gel filtration-HPLC. These observations suggest that changes in polar melphalan products, and perhaps their interaction with a protein, may by involved in the reduction of melphalan cytotoxicity observed in ZnC1 2 -treated cells. While ZnC1 2 is also known to increase the level of glutathione in cells, no significant amounts of glutathione-melphalan derivatives of the type formed non-enzymatically in vitro could be detected in ZnC1 2 -treated or untreated cells. Formation of derivatives of melphalan with glutathione catabolic products in ZnC1 2 -treated cells has not yet been eliminated, however. 17 refs., 5 figs., 1 tab

  5. Primary Patient-Derived Cancer Cells and Their Potential for Personalized Cancer Patient Care

    Directory of Open Access Journals (Sweden)

    David P. Kodack

    2017-12-01

    Full Text Available Personalized cancer therapy is based on a patient’s tumor lineage, histopathology, expression analyses, and/or tumor DNA or RNA analysis. Here, we aim to develop an in vitro functional assay of a patient’s living cancer cells that could complement these approaches. We present methods for developing cell cultures from tumor biopsies and identify the types of samples and culture conditions associated with higher efficiency of model establishment. Toward the application of patient-derived cell cultures for personalized care, we established an immunofluorescence-based functional assay that quantifies cancer cell responses to targeted therapy in mixed cell cultures. Assaying patient-derived lung cancer cultures with this method showed promise in modeling patient response for diagnostic use. This platform should allow for the development of co-clinical trial studies to prospectively test the value of drug profiling on tumor-biopsy-derived cultures to direct patient care.

  6. Radiation-induced DNA Double Strand Breaks and Their Modulations by Treatments with Moringa oleifera Lam. Leaf Extracts: A Cancer Cell Culture Model

    International Nuclear Information System (INIS)

    Boonsirichai, K.; Jetawattana, S.

    2014-01-01

    Gamma radiation brings deleterious effects upon human cells by inducing oxidative stress and DNA damages. Antioxidants have been shown to confer protective effects on irradiated normal cells. Moringa oleifera Lam. is a widely used nutritional supplement with antioxidant activities. This report showed that antioxidant-containing supplements, in addition to protecting normal cells, could protect cancer cells against genotoxic effects of gamma radiation. γ-H2AX immunofluorescent foci were utilized as an indicator of radiation-induced DNA double strand breaks. MCF-7 human breast adenocarcinoma cells were irradiated with 2-8 Gy gamma radiation. A linear relationship between the formation of γ-H2AX foci and radiation dose was observed with an average of 10 foci per cell per Gy. A 30-minute pretreatment of the cells with either the aqueous or the ethanolic extract of M. oleifera leaves could partially protect the cells from radiation-induced DNA double strand breaks. A pretreatment with 500 µg/mL aqueous extract reduced the number of foci formed by 15% when assayed at 30 minutes post-irradiation. The ethanolic extract was more effective; 500 µg/mL of its concentration reduced the number of foci among irradiated cells by 30%. The results indicated that irradiated cancer cells responded similarly to nutritional supplements containing antioxidants as irradiated normal cells. These natural antioxidants could confer protective effects upon cancer cells against gamma radiation. (author)

  7. Radiation-induced DNA Double Strand Breaks and Their Modulations by Treatments with Moringa oleifera Lam. Leaf Extracts: A Cancer Cell Culture Model

    Directory of Open Access Journals (Sweden)

    K. Boonsirichai

    2014-04-01

    Full Text Available Gamma radiation brings deleterious effects upon human cells by inducing oxidative stress and DNA damages. Antioxidants have been shown to confer protective effects on irradiated normal cells. Moringa oleifera Lam. is a widely used nutritional supplement with antioxidant activities. This report showed that antioxidant-containing supplements, in addition to protecting normal cells, could protect cancer cells against genotoxic effects of gamma radiation. -H2AX immunofluorescent foci were utilized as an indicator of radiation-induced DNA double strand breaks. MCF-7 human breast adenocarcinoma cells were irradiated with 2-8 Gy gamma radiation. A linear relationship between the formation of -H2AX foci and radiation dose was observed with an average of 10 foci per cell per Gy. A 30-minute pretreatment of the cells with either the aqueous or the ethanolic extract of M. oleifera leaves could partially protect the cells from radiation-induced DNA double strand breaks. A pretreatment with 500 µg/mL aqueous extract reduced the number of foci formed by 15% when assayed at 30 minutes post-irradiation. The ethanolic extract was more effective; 500 µg/mL of its concentration reduced the number of foci among irradiated cells by 30%. The results indicated that irradiated cancer cells responded similarly to nutritional supplements containing antioxidants as irradiated normal cells. These natural antioxidants could confer protective effects upon cancer cells against gamma radiation

  8. Multistage carcinogenesis in cell culture.

    Science.gov (United States)

    Rubin, H

    2001-01-01

    Rodent fibroblasts explanted from embryos to culture undergo a period of declining growth rate in serial passages leading to crisis, followed by the appearance of variants which can multiply indefinitely. If the "immortal" cell line was established by low density passage, i.e., 3T3 cells, it has a low saturation density and is non-tumorigenic. If it was established by high density passage, it has a high saturation density and is tumorigenic. The establishment of cells goes through successive stages, including increased capacity to multiply in low serum concentration, growth to high saturation density, growth in suspension, assisted tumour formation in susceptible hosts and unassisted tumour formation. Chromosome aberrations and aneuploidy occur long before the capacity to produce tumours appears. Contrary to conventional belief, human fibroblast populations also undergo a continuous loss of capacity to multiply from the time of explantation, with only the longest surviving clone reaching the Hayflick limit. Neoplastic transformation of rodent cells is strongly favoured by maintaining them in a quiescent state at confluence for prolonged periods, which results in genetic damage to the cells. It also produces a large variety of chromosomal aberrations in human cells and extends their replicative lifespan. Individual clones are more susceptible to spontaneous transformation than their heterogeneous parental cultures. The implications of these results for tumour development in vivo are that oncogenic genetic changes may be common under stressful conditions which restrict replication, and that such changes are maximized when a rogue clone reaches a critical size that reduces stabilizing interactions with neighbouring clones. An alternative explanation, described in the Addendum, which we retrospectively favor is that the easily transformed clones are a minority in the uncloned parental population. The reason they transform before the parental population is that when

  9. Bags versus flasks: a comparison of cell culture systems for the production of dendritic cell-based immunotherapies.

    Science.gov (United States)

    Fekete, Natalie; Béland, Ariane V; Campbell, Katie; Clark, Sarah L; Hoesli, Corinne A

    2018-04-19

    In recent years, cell-based therapies targeting the immune system have emerged as promising strategies for cancer treatment. This review summarizes manufacturing challenges related to production of antigen presenting cells as a patient-tailored cancer therapy. Understanding cell-material interactions is essential because in vitro cell culture manipulations to obtain mature antigen-producing cells can significantly alter their in vivo performance. Traditional antigen-producing cell culture protocols often rely on cell adhesion to surface-treated hydrophilic polystyrene flasks. More recent commercial and investigational cancer immunotherapy products were manufactured using suspension cell culture in closed hydrophobic fluoropolymer bags. The shift to closed cell culture systems can decrease risks of contamination by individual operators, as well as facilitate scale-up and automation. Selecting closed cell culture bags over traditional open culture systems entails different handling procedures and processing controls, which can affect product quality. Changes in culture vessels also entail changes in vessel materials and geometry, which may alter the cell microenvironment and resulting cell fate decisions. Strategically designed culture systems will pave the way for the generation of more sophisticated and highly potent cell-based cancer vaccines. As an increasing number of cell-based therapies enter the clinic, the selection of appropriate cell culture vessels and materials becomes a critical consideration that can impact the therapeutic efficacy of the product, and hence clinical outcomes and patient quality of life. © 2018 The Authors Transfusion published by Wiley Periodicals, Inc. on behalf of AABB.

  10. [Cytotoxic effect of physalis peruviana in cell culture of colorectal and prostate cancer and chronic myeloid leukemia].

    Science.gov (United States)

    Quispe-Mauricio, Angel; Callacondo, David; Rojas, José; Zavala, David; Posso, Margarita; Vaisberg, Abraham

    2009-01-01

    The plants have been used as drugs for centuries. However, limited research has been done on its great potential as sources of new therapeutic agents. The purpose of this study was to evaluate Physalis peruviana cytotoxic activity on cell lines HT-29, PC-3, K-562 and VERO. The HT-29 cell lines, PC-3, K-562 and VERO, were exposed to four concentrations of P. peruviana ethanolic leave and stem extracts, also at different concentrations of cisplatin and 5-fluorouracil (5-FU), which were used as positive controls. We found rates of growth within 48 hours, then we determined the inhibitory concentration 50 (IC50) using linear regression analysis and the index of selectivity of each sample. The P. peruviana ethanolic leave and stem extracts showed cytotoxic activity. The IC50 in g/mL in leaves and stems were, 0.35 (r =-0.95 p peruviana leaves and steams ethanolic extracts were more cytotoxic than cisplatin and 5 FU, on the lines HT-29, PC-3 and K562. Furthermore the P. peruviana cytotoxic effects were less than cisplatin and 5-FU for VERO control cells lines.

  11. Dynamized Preparations in Cell Culture

    Directory of Open Access Journals (Sweden)

    Ellanzhiyil Surendran Sunila

    2009-01-01

    Full Text Available Although reports on the efficacy of homeopathic medicines in animal models are limited, there are even fewer reports on the in vitro action of these dynamized preparations. We have evaluated the cytotoxic activity of 30C and 200C potencies of ten dynamized medicines against Dalton's Lymphoma Ascites, Ehrlich's Ascites Carcinoma, lung fibroblast (L929 and Chinese Hamster Ovary (CHO cell lines and compared activity with their mother tinctures during short-term and long-term cell culture. The effect of dynamized medicines to induce apoptosis was also evaluated and we studied how dynamized medicines affected genes expressed during apoptosis. Mother tinctures as well as some dynamized medicines showed significant cytotoxicity to cells during short and long-term incubation. Potentiated alcohol control did not produce any cytotoxicity at concentrations studied. The dynamized medicines were found to inhibit CHO cell colony formation and thymidine uptake in L929 cells and those of Thuja, Hydrastis and Carcinosinum were found to induce apoptosis in DLA cells. Moreover, dynamized Carcinosinum was found to induce the expression of p53 while dynamized Thuja produced characteristic laddering pattern in agarose gel electrophoresis of DNA. These results indicate that dynamized medicines possess cytotoxic as well as apoptosis-inducing properties.

  12. A method for establishing human primary gastric epithelial cell culture from fresh surgical gastric tissues.

    Science.gov (United States)

    Aziz, Faisal; Yang, Xuesong; Wen, Qingping; Yan, Qiu

    2015-08-01

    At present, biopsy specimens, cancer cell lines and tissues obtained by gastric surgery are used in the study and analysis of gastric cancer, including the molecular mechanisms and proteomics. However, fibroblasts and other tissue components may interfere with these techniques. Therefore, the present study aimed to develop a procedure for the isolation of viable human gastric epithelial cells from gastric surgical tissues. A method was developed to culture human gastric epithelial cells using fresh, surgically excised tissues and was evaluated using immunocytochemistry, periodic acid-Schiff (PAS) staining and cell viability assays. Low cell growth was observed surrounding the gastric tissue on the seventh day of tissue explant culture. Cell growth subsequently increased, and at 12 days post-explant a high number of pure epithelial cells were detected. The gastric cancer cells exhibited rapid growth with a doubling time of 13-52 h, as compared to normal cells, which had a doubling time of 20-53 h. Immunocytochemical analyses of primary gastric cells revealed positive staining for cytokeratin 18 and 19, which indicated that the culture was comprised of pure epithelial cells and contained no fibroblasts. Furthermore, PAS staining demonstrated that the cultured gastric cells produced neutral mucin. Granulin and carbohydrate antigen 724 staining confirmed the purity of gastric cancer and normal cells in culture. This method of cell culture indicated that the gastric cells in primary culture consisted of mucin-secreting gastric epithelial cells, which may be useful for the study of gastric infection with Helicobacter pylori and gastric cancer.

  13. Transcriptional profiling of human breast cancer cells cultured under microgravity conditions revealed the key role of genetic gravity sensors previously detected in Drosophila melanogaster

    Science.gov (United States)

    Valdivia-Silva, Julio E.; Lavan, David; Diego Orihuela-Tacuri, M.; Sanabria, Gabriela

    2016-07-01

    Currently, studies in Drosophila melanogaster has shown emerging evidence that microgravity stimuli can be detected at the genetic level. Analysis of the transcriptome in the pupal stage of the fruit flies under microgravity conditions versus ground controls has suggested the presence of a few candidate genes as "gravity sensors" which are experimentally validated. Additionally, several studies have shown that microgravity causes inhibitory effects in different types of cancer cells, although the genes involved and responsible for these effects are still unknown. Here, we demonstrate that the genes suggested as the sensors of gravitational waves in Drosophila melanogaster and their human counterpart (orthologous genes) are highly involved in carcinogenesis, proliferation, anti-apoptotic signals, invasiveness, and metastatic potential of breast cancer cell tumors. The transcriptome analyses suggested that the observed inhibitory effect in cancer cells could be due to changes in the genetic expression of these candidates. These results encourage the possibility of new therapeutic targets managed together and not in isolation.

  14. Selective eradication of cancer cells in vitro

    International Nuclear Information System (INIS)

    Schneiderman, M.H.; Schneiderman, G.S.

    1984-01-01

    A simple system consisting of cultured HeLa (human cancer) and WI38 (normal human fetal lung) cells and the control cultures of the individual cells were set up to test and compare the effects of the cell cycle-active agents /sup 125/I-iododeoxyuridine (/sup 125/IUdR) and hydroxyurea (HU) on cell survival. The presence of cells and growth after treatment were used as a positive indication of survival. The experimental cultures were first seeded with WI38 cells and allowed to grow to confluency before adding 1.0 x 10/sup 5/ HeLa cells. After two days of treatment-free growth, the co-cultures were continuously treated with /sup 125/IUdR (0.5-2.0 μCi/ml, carrier free) or HU (1.0 x 10/sup -9/ and 1.0 x 10/sup -3/M). At the termination of treatment the co-cultures were split 3 to 1 and incubated for seven days. As expected, there was little or no detectable effect on the growth of WI38 cells treated with HU or /sup 125/IUdR while the cells were confluent. However, HeLa cells were reduced by 1.0 x 10/sup -3/M HU and were eradicated after all concentrations of /sup 125/IUdR

  15. Modelos experimentais em oncologia: O contributo da cultura de células para o conhecimento da biologia do cancro Experimental models in oncology: Contribution of cell culture on understanding the biology of cancer

    Directory of Open Access Journals (Sweden)

    Mariana Cruz

    2009-08-01

    Full Text Available A cultura de tecidos surgiu no século xx (Harrison, 1907 para estudar o comportamento das células animais em ambiente homeostático e em situações de stress. A capacidade de estudar as células a nível molecular relaciona-se com a forma como as células podem crescer e ser manipuladas em laboratório. A cultura de células in vitro permitiu estudar crescimento, diferenciação e morte celular e efectuar manipulações genéticas necessárias ao perfeito conhecimento da estrutura e funções dos genes. A cultura de células estaminais humanas veio colmatar algumas limitações inerentes aos restantes modelos de cultura. Ao que parece, as células estaminais cancerígenas mantêm-se quiescentes nos locais metastáticos até serem activadas por sinais apropriados do microambiente. Vários estudos revelaram que diferentes tipos de cancros podem surgir da transformação maligna de células estaminais. A eliminação destas células progenitoras tumorais é essencial para o desenvolvimento de novas abordagens terapêuticas mais eficazes em cancros agressivos. Por outro lado, a utilização de células dendríticas modificadas em cultura poderá contribuir para a produção de uma potencial vacina terapêutica eficaz para obter a regressão tumoral.In the beginning of the 20th century, tissue culture was started with the aim of studying the behaviour of animal cells in normal and stress conditions. The cell study at molecular level depends on their capacity of growing and how they can be manipulated in laboratory. In vitro cell culture allows us the possibility of studying biological key processes, such as growth, differentiation and cell death, and also to do genetic manipulations essential to the knowledge of structure and genes function. Human stem cells culture provides strategies to circumvent other models’ deficiencies. It seems that cancer stem cells remain quiescent until activation by appropriated micro-environmental stimulation

  16. Culture supernatants of oral cancer cells induce impaired IFN-α production of pDCs partly through the down-regulation of TLR-9 expression.

    Science.gov (United States)

    Han, Nannan; Zhang, Zun; Jv, Houyu; Hu, Jingzhou; Ruan, Min; Zhang, Chenping

    2018-06-05

    The aim of the present study was to investigate whether tumor-derived supernatants down-regulate the immune function of plasmacytoid dendritic cells (pDCs) in oral cancer and the potential molecular mechanisms of this effect. Immunohistochemistry (IHC) and flow cytometry were used to detect tumor-infiltrating and peripheral blood pDCs. MTS and flow cytometry were employed to evaluate the immune response of CD4 + T cells. Real-time PCR and ELISA assays were used to identify TLR-7 and TLR-9 expression, IFN-α production and tumor-secreted soluble cytokines. The proportion of pDCs (0.121%±0.043%) was significantly higher in Oral squamous cell carcinoma (OSCC) samples than in normal tissue (0.023%±0.016%) (P = 0.021). TLR9 mRNA was significantly lower in tumor-infiltrating pDCs and positively correlated to low IFN-α production (r = 0.956; Poral cancer cells negatively regulated TLR9 mRNA expression and the subsequent IFN-α production of pDCs, which inhibited the immune response of CD4 + T cells. The neutralizing antibodies blocking assay showed that the specific inhibitory effect of pDC functionality was associated with the soluble fraction of the oral cancer environment, which is mainly mediated by IL-10 and TGF-β cooperation. Tumor-derived supernatants may impair the function of tumor-infiltrating pDCs, which subsequently decreases the immune response of CD4 + T cells in human oral cancer through TGF-β- and IL-10- dependent mechanisms. Careful manipulation of these impaired pDCs may help develop an important alternative immunotherapy for the treatment of oral cancer. Copyright © 2018 Elsevier Ltd. All rights reserved.

  17. Lung cancer - non-small cell

    Science.gov (United States)

    Cancer - lung - non-small cell; Non-small cell lung cancer; NSCLC; Adenocarcinoma - lung; Squamous cell carcinoma - lung ... Research shows that smoking marijuana may help cancer cells grow. But there is no direct link between ...

  18. Cancer stem cells and personalized cancer nanomedicine.

    Science.gov (United States)

    Gener, Petra; Rafael, Diana Fernandes de Sousa; Fernández, Yolanda; Ortega, Joan Sayós; Arango, Diego; Abasolo, Ibane; Videira, Mafalda; Schwartz, Simo

    2016-02-01

    Despite the progress in cancer treatment over the past years advanced cancer is still an incurable disease. Special attention is pointed toward cancer stem cell (CSC)-targeted therapies, because this minor cell population is responsible for the treatment resistance, metastatic growth and tumor recurrence. The recently described CSC dynamic phenotype and interconversion model of cancer growth hamper even more the possible success of current cancer treatments in advanced cancer stages. Accordingly, CSCs can be generated through dedifferentiation processes from non-CSCs, in particular, when CSC populations are depleted after treatment. In this context, the use of targeted CSC nanomedicines should be considered as a promising tool to increase CSC sensitivity and efficacy of specific anti-CSC therapies.

  19. Microfluidic cell culture systems for drug research.

    Science.gov (United States)

    Wu, Min-Hsien; Huang, Song-Bin; Lee, Gwo-Bin

    2010-04-21

    In pharmaceutical research, an adequate cell-based assay scheme to efficiently screen and to validate potential drug candidates in the initial stage of drug discovery is crucial. In order to better predict the clinical response to drug compounds, a cell culture model that is faithful to in vivo behavior is required. With the recent advances in microfluidic technology, the utilization of a microfluidic-based cell culture has several advantages, making it a promising alternative to the conventional cell culture methods. This review starts with a comprehensive discussion on the general process for drug discovery and development, the role of cell culture in drug research, and the characteristics of the cell culture formats commonly used in current microfluidic-based, cell-culture practices. Due to the significant differences in several physical phenomena between microscale and macroscale devices, microfluidic technology provides unique functionality, which is not previously possible by using traditional techniques. In a subsequent section, the niches for using microfluidic-based cell culture systems for drug research are discussed. Moreover, some critical issues such as cell immobilization, medium pumping or gradient generation in microfluidic-based, cell-culture systems are also reviewed. Finally, some practical applications of microfluidic-based, cell-culture systems in drug research particularly those pertaining to drug toxicity testing and those with a high-throughput capability are highlighted.

  20. Induction of cancer stem cell properties in colon cancer cells by defined factors.

    Directory of Open Access Journals (Sweden)

    Nobu Oshima

    Full Text Available Cancer stem cells (CSCs are considered to be responsible for the dismal prognosis of cancer patients. However, little is known about the molecular mechanisms underlying the acquisition and maintenance of CSC properties in cancer cells because of their rarity in clinical samples. We herein induced CSC properties in cancer cells using defined factors. We retrovirally introduced a set of defined factors (OCT3/4, SOX2 and KLF4 into human colon cancer cells, followed by culture with conventional serum-containing medium, not human embryonic stem cell medium. We then evaluated the CSC properties in the cells. The colon cancer cells transduced with the three factors showed significantly enhanced CSC properties in terms of the marker gene expression, sphere formation, chemoresistance and tumorigenicity. We designated the cells with CSC properties induced by the factors, a subset of the transduced cells, as induced CSCs (iCSCs. Moreover, we established a novel technology to isolate and collect the iCSCs based on the differences in the degree of the dye-effluxing activity enhancement. The xenografts derived from our iCSCs were not teratomas. Notably, in contrast to the tumors from the parental cancer cells, the iCSC-based tumors mimicked actual human colon cancer tissues in terms of their immunohistological findings, which showed colonic lineage differentiation. In addition, we confirmed that the phenotypes of our iCSCs were reproducible in serial transplantation experiments. By introducing defined factors, we generated iCSCs with lineage specificity directly from cancer cells, not via an induced pluripotent stem cell state. The novel method enables us to obtain abundant materials of CSCs that not only have enhanced tumorigenicity, but also the ability to differentiate to recapitulate a specific type of cancer tissues. Our method can be of great value to fully understand CSCs and develop new therapies targeting CSCs.

  1. Lingual Epithelial Stem Cells and Organoid Culture of Them

    Directory of Open Access Journals (Sweden)

    Hiroko Hisha

    2016-01-01

    Full Text Available As tongue cancer is one of the major malignant cancers in the world, understanding the mechanism of maintenance of lingual epithelial tissue, which is known to be the origin of tongue cancer, is unquestionably important. However, the actual stem cells that are responsible for the long-term maintenance of the lingual epithelium have not been identified. Moreover, a simple and convenient culture method for lingual epithelial stem cells has not yet been established. Recently, we have shown that Bmi1-positive cells, residing at the second or third layer of the epithelial cell layer at the base of the interpapillary pit (IPP, were slow-cycling and could supply keratinized epithelial cells for over one year, indicating that Bmi1-positive cells are long-term lingual epithelial stem cells. In addition, we have developed a novel lingual epithelium organoid culture system using a three-dimensional matrix and growth factors. Here, we discuss current progress in the identification of lingual stem cells and future applications of the lingual culture system for studying the regulatory mechanisms of the lingual epithelium and for regenerative medicine.

  2. Cell Culture as an Alternative in Education.

    Science.gov (United States)

    Nardone, Roland M.

    1990-01-01

    Programs that are intended to inform and provide "hands-on" experience for students and to facilitate the introduction of cell culture-based laboratory exercises into the high school and college laboratory are examined. The components of the CellServ Program and the Cell Culture Toxicology Training Programs are described. (KR)

  3. Cell culture techniques in honey bee research

    Science.gov (United States)

    Cell culture techniques are indispensable in most if not all life science disciplines to date. Wherever cell culture models are lacking scientific development is hampered. Unfortunately this has been and still is the case in honey bee research because permanent honey bee cell lines have not yet been...

  4. Utility of a Novel Three-Dimensional and Dynamic (3DD Cell Culture System for PK/PD Studies: Evaluation of a Triple Combination Therapy at Overcoming Anti-HER2 Treatment Resistance in Breast Cancer

    Directory of Open Access Journals (Sweden)

    Anusha Ande

    2018-05-01

    Full Text Available Background: Emergence of Human epidermal growth factor receptor 2 (HER2 therapy resistance in HER2-positive (HER2+ breast cancer (BC poses a major clinical challenge. Mechanisms of resistance include the over-activation of the PI3K/mTOR and Src pathways. This work aims to investigate a novel combination therapy that employs paclitaxel (PAC, a mitotic inhibitor, with everolimus (EVE, an mTOR inhibitor, and dasatinib (DAS, an Src kinase inhibitor, as a modality to overcome resistance.Methods: Static (two dimensional, 2D and three-dimensional dynamic (3DD cell culture studies were conducted using JIMT-1 cells, a HER2+ BC cell line refractory to HER2 therapies. Cell viability and caspase-3 expression were examined after JIMT-1 cell exposure to agents as monotherapy or in combination using a 2D setting. A pharmacokinetic/pharmacodynamic (PK/PD combination study with PAC+DAS+EVE was conducted over 3 weeks in a 3DD setting. PAC was administered into the system via a 3 h infusion followed by the addition of a continuous infusion of EVE+DAS 24 h post-PAC dosing. Cell counts and caspase-3 expression were quantified every 2 days. A semi-mechanistic PK/PD model was developed using the 2D data and scaled up to capture the 3DD data. The final model integrated active caspase-3 as a biomarker to bridge between drug exposures and cancer cell dynamics. Model fittings were performed using Monolix software.Results: The triple combination significantly induced caspase-3 activity in the 2D cell culture setting. In the 3DD cell culture setting, sequential dosing of PAC then EVE+DAS showed a 5-fold increase in caspase-3 activity and 8.5-fold decrease in the total cell number compared to the control. The semi-mechanistic PK/PD models fit the data well, capturing the time-course profiles of drug concentrations, caspase-3 expression, and cell counts in the 2D and 3DD settings.Conclusion: A novel, sequential triple combination therapeutic regimen was successfully evaluated

  5. Dysfunctions at human intestinal barrier by water-borne protozoan parasites: lessons from cultured human fully differentiated colon cancer cell lines.

    Science.gov (United States)

    Liévin-Le Moal, Vanessa

    2013-06-01

    Some water-borne protozoan parasites induce diseases through their membrane-associated functional structures and virulence factors that hijack the host cellular molecules and signalling pathways leading to structural and functional lesions in the intestinal barrier. In this Microreview we analyse the insights on the mechanisms of pathogenesis of Entamoeba intestinalis, Giardia and Cryptosporidium observed in the human colon carcinoma fully differentiated colon cancer cell lines, cell subpopulations and clones expressing the structural and functional characteristics of highly specialized fully differentiated epithelial cells lining the intestinal epithelium and mimicking structurally and functionally an intestinal barrier. © 2013 John Wiley & Sons Ltd.

  6. Protein biosynthesis in cultured human hair follicle cells.

    Science.gov (United States)

    Weterings, P J; Vermorken, A J; Bloemendal, H

    1980-10-31

    A new technique has been used for culturing human keratinocytes. The cells grow on the basement membrane-like capsules of bovine lenses. Lens cells were removed from the capsules by rigid trypsinization. In order to exclude any contamination with remaining living cells the isolated capsules were irradiated with X-rays at a dose of 10,000 rad. In this way human epithelial cells can be brought in culture from individual hair follicles. Since feeder cells are not used in this culture technique, the biosynthesis of keratinocyte proteins can be studied in these cultures. The newly synthesized proteins can be separated into a water-soluble, a urea-soluble, and a urea-insoluble fraction. Product analysis has been performed on the first two fractions revealing protein patterns identical to those of intact hair follicles. Product analysis of the urea-soluble fractions of microdissected hair follicles shows that the protein pattern of the cultured keratinocytes resembles the protein pattern of the hair follicle sheath. Studies on the metabolism of benzo(a)pyrene revealed that the enzyme aryl hydrocarbon hydroxylase (AHH) is present in cultured hair follicle cells. A possible use of our culture system for eventual detection of inherited predisposition for smoking-dependent lung cancer is discussed.

  7. Cancer stem cells, cancer cell plasticity and radiation therapy.

    Science.gov (United States)

    Vlashi, Erina; Pajonk, Frank

    2015-04-01

    Since the first prospective identification of cancer stem cells in solid cancers the cancer stem cell hypothesis has reemerged as a research topic of increasing interest. It postulates that solid cancers are organized hierarchically with a small number of cancer stem cells driving tumor growth, repopulation after injury and metastasis. They give rise to differentiated progeny, which lack these features. The model predicts that for any therapy to provide cure, all cancer stem cells have to be eliminated while the survival of differentiated progeny is less critical. In this review we discuss recent reports challenging the idea of a unidirectional differentiation of cancer cells. These reports provide evidence supporting the idea that non-stem cancer cells exhibit a remarkable degree of plasticity that allows them to re-acquire cancer stem cell traits, especially in the context of radiation therapy. We summarize conditions under which differentiation is reversed and discuss the current knowledge of the underlying mechanisms. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Effect of hypoxia on the uptake of [methyl-3H]choline, [1-14C] acetate and [18F]FDG in cultured prostate cancer cells

    International Nuclear Information System (INIS)

    Hara, Toshihiko; Bansal, Aditya; DeGrado, Timothy R.

    2006-01-01

    Introduction: Choline, acetate and glucose ([2- 18 F]fluoro-2-deoxyglucose, [ 18 F]FDG) analogs are under investigation as positron emission tomography (PET) tracers for the imaging of prostate cancer; however, their response to tumor hypoxia has not been clarified. Methods: The uptake of [methyl- 3 H]choline, [1- 14 C]acetate and [ 18 F]FDG was monitored in androgen-independent PC-3 cells and androgen-sensitive LNCaP cells under aerobic or anoxic conditions. The effect of androgen depletion was also examined. Results: PC-3 cells exhibited aerobic choline and acetate uptake five to six times higher than FDG uptake, whereas LNCaP cells showed choline uptake 2.2-fold higher than acetate uptake and 10-fold higher than FDG uptake. After 4 h of anoxia, PC-3 cells showed an 85% increase in FDG uptake, a 15% decrease in choline uptake and a 36% increase in acetate uptake, whereas LNCaP cells showed a 212% increase in FDG uptake, a 28% decrease in choline uptake and no change in acetate uptake. Androgen depletion resulted in a marked decrease in the uptake of all tracers in LNCaP cells but no changes in PC-3 cells. Conclusion: In aerobic conditions, both PC-3 and LNCaP cells exhibited an order of uptake preference as follows: choline>acetate>FDG. In hypoxic cells, the order is reversed, reflecting diverse biochemical responses to hypoxia. These findings may help to explain PET imaging findings of the diverse responses of these tracers in different stages and locations of prostate cancer. Androgen depletion markedly suppressed the uptake of all three tracers in LNCaP cells, which suggests the potential for underestimation of the disease state when PET imaging is performed subsequent to antiandrogen therapy

  9. General Information about Small Cell Lung Cancer

    Science.gov (United States)

    ... Lung Cancer Prevention Lung Cancer Screening Research Small Cell Lung Cancer Treatment (PDQ®)–Patient Version General Information About Small Cell Lung Cancer Go to Health Professional Version Key Points Small ...

  10. Stages of Small Cell Lung Cancer

    Science.gov (United States)

    ... Lung Cancer Prevention Lung Cancer Screening Research Small Cell Lung Cancer Treatment (PDQ®)–Patient Version General Information About Small Cell Lung Cancer Go to Health Professional Version Key Points Small ...

  11. Local advanced transitional cell cancer and squamous cell cancer of ...

    African Journals Online (AJOL)

    Case report: A 51-year-old man presented with a locally advanced squamous cell cancer of the periurethral tissues as well as an underlying isolated transitional cell cancer of the urethra. Chemotherapy with Gemcitabin and Cisplatinum together with local radiation to the pelvis and the perineum was given. There was ...

  12. A new prospect in cancer therapy: targeting cancer stem cells to eradicate cancer.

    Science.gov (United States)

    Chen, Li-Sha; Wang, An-Xin; Dong, Bing; Pu, Ke-Feng; Yuan, Li-Hua; Zhu, Yi-Min

    2012-12-01

    According to the cancer stem cell theory, cancers can be initiated by cancer stem cells. This makes cancer stem cells prime targets for therapeutic intervention. Eradicating cancer stem cells by efficient targeting agents may have the potential to cure cancer. In this review, we summarize recent breakthroughs that have improved our understanding of cancer stem cells, and we discuss the therapeutic strategy of targeting cancer stem cells, a promising future direction for cancer stem cell research.

  13. Establishment and Characterization of Primary Cultures from Iranian Oral Squamous Cell Carcinoma Patients by Enzymatic Method and Explant Culture

    Directory of Open Access Journals (Sweden)

    Meysam Ganjibakhsh

    2017-10-01

    Full Text Available Objectives: Oral Squamous Cell Carcinoma (OSCC is the most frequent oral cancer worldwide. It is known as the eighth most common cancer in men and as the fifth most common cancer in women. Cytogenetic and biochemical studies in recent decades have emphasized the necessity of providing an appropriate tool for such researches. Cancer cell culture is a useful tool for investigations on biochemical, genetic, molecular and immunological characteristics of different cancers, including oral cancer. Here, we explain the establishment process of five primary oral cancer cells derived from an Iranian population.Materials and Methods: The specimens were obtained from five oral cancer patients. Enzymatic, explant culture and magnetic-activated cell sorting (MACS methods were used for cell isolation. After quality control tests, characterization and authentication of primary oral cancer cells were performed by short tandem repeats (STR profiling, chromosome analysis, species identification, and monitoring the growth, morphology and the expression of CD326 and CD133 markers.Results: Five primary oral cancer cells were established from an Iranian population. The flow cytometry results showed that the isolated cells were positive for CD326 and CD133 markers. Furthermore, the cells were free from mycoplasma, bacterial and fungal contamination. No misidentified or cross-contaminated cells were detected by STR analysis.Conclusions: Human primary oral cancer cells provide an extremely useful platform for studying carcinogenesis pathways of oral cancer in Iranian population. They may be helpful in explaining the ethnic differences in cancer biology and the individuality in anticancer drug response in future studies.

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

    International Nuclear Information System (INIS)

    Kikuta, Kazuhiro; Masamune, Atsushi; Watanabe, Takashi; Ariga, Hiroyuki; Itoh, Hiromichi; Hamada, Shin; Satoh, Kennichi; Egawa, Shinichi; Unno, Michiaki; Shimosegawa, Tooru

    2010-01-01

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

  15. Mesenchymal stem cells enhance the metastasis of 3D-cultured hepatocellular carcinoma cells

    International Nuclear Information System (INIS)

    Liu, Chang; Liu, Yang; Xu, Xiao-xi; Guo, Xin; Sun, Guang-wei; Ma, Xiao-jun

    2016-01-01

    Accumulating evidences have demonstrated that mesenchymal stem cells (MSC) could be recruited to the tumor microenvironment. Umbilical cord mesenchymal stem cells (UCMSC) were attractive vehicles for delivering therapeutic agents against cancer. Nevertheless, the safety of UCMSC in the treatment of tumors including hepatocellular carcinoma (HCC) was still undetermined. In this study, an in vitro co-culture system was established to evaluate the effect of UCMSC on the cell growth, cancer stem cell (CSC) characteristics, drug resistance, metastasis of 3D-cultured HCC cells, and the underlying mechanism was also investigated. It was found that after co-cultured with UCMSC, the metastatic ability of 3D-cultured HCC cells was significantly enhanced as indicated by up-regulation of matrix metalloproteinase (MMP), epithelial-mesenchymal transition (EMT)-related genes, and migration ability. However, cell growth, drug resistance and CSC-related gene expression of HCC cells were not affected by UCMSC. Moreover, EMT was reversed, MMP-2 expression was down-regulated, and migration ability of HCC cell was significantly inhibited when TGF-β receptor inhibitor SB431542 was added into the co-culture system. Therefore, these data indicated that UCMSC could significantly enhance the tumor cell metastasis, which was due to the EMT of HCC cells induced by TGF-β. The online version of this article (doi:10.1186/s12885-016-2595-4) contains supplementary material, which is available to authorized users

  16. NSAIDs and Cell Proliferation in Colorectal Cancer

    Directory of Open Access Journals (Sweden)

    Raj Ettarh

    2010-06-01

    Full Text Available Colon cancer is common worldwide and accounts for significant morbidity and mortality in patients. Fortunately, epidemiological studies have demonstrated that continuous therapy with NSAIDs offers real promise of chemoprevention and adjunct therapy for colon cancer patients. Tumour growth is the result of complex regulation that determines the balance between cell proliferation and cell death. How NSAIDs affect this balance is important for understanding and improving treatment strategies and drug effectiveness. NSAIDs inhibit proliferation and impair the growth of colon cancer cell lines when tested in culture in vitro and many NSAIDs also prevent tumorigenesis and reduce tumour growth in animal models and in patients, but the relationship to inhibition of tumour cell proliferation is less convincing, principally due to gaps in the available data. High concentrations of NSAIDs are required in vitro to achieve cancer cell inhibition and growth retardation at varying time-points following treatment. However, the results from studies with colon cancer cell xenografts are promising and, together with better comparative data on anti-proliferative NSAID concentrations and doses (for in vitro and in vivo administration, could provide more information to improve our understanding of the relationships between these agents, dose and dosing regimen, and cellular environment.

  17. Epigenetics in cancer stem cells.

    Science.gov (United States)

    Toh, Tan Boon; Lim, Jhin Jieh; Chow, Edward Kai-Hua

    2017-02-01

    Compelling evidence have demonstrated that bulk tumors can arise from a unique subset of cells commonly termed "cancer stem cells" that has been proposed to be a strong driving force of tumorigenesis and a key mechanism of therapeutic resistance. Recent advances in epigenomics have illuminated key mechanisms by which epigenetic regulation contribute to cancer progression. In this review, we present a discussion of how deregulation of various epigenetic pathways can contribute to cancer initiation and tumorigenesis, particularly with respect to maintenance and survival of cancer stem cells. This information, together with several promising clinical and preclinical trials of epigenetic modulating drugs, offer new possibilities for targeting cancer stem cells as well as improving cancer therapy overall.

  18. Chlorogenic acid complex (CGA7, standardized extract from green coffee beans exerts anticancer effects against cultured human colon cancer HCT-116 cells

    Directory of Open Access Journals (Sweden)

    K. Gouthamchandra

    2017-09-01

    Full Text Available Coffee is commonly consumed beverage in the world and it has been suggested to have beneficial effect. Chlorogenic acids (CGAs are main ingredient of coffee beans which has been extensively used in nutraceuticals and medicine. Recently, various therapeutic effects of chlorogenic acids have been investigated. However, there are limited studies to investigate its anticancer properties. In the present study, we have used chlorogenic acid complex (CGA7 a decaffeinated water soluble green coffee bean extract to evaluate its cytotoxic effect on human and mouse cancer cell lines by using different approaches. From our results we found CGA7 treatment induces cell death in a dose and time dependent manner in different cancer cell lines. Further, CGA7 induced apoptosis was characterized by DNA fragmentation, PARP-1 cleavage, caspase-9 activation, and down regulation of Bcl-2, an anti-apoptotic protein and up regulation of pro-apoptotic protein BAX. Overall findings indicated that CGA7 complex a potent anticancer molecule found in green coffee beans could be a safe bioactive ingredient for prevention of cancer.

  19. Drug Treatment of Cancer Cell Lines: A Way to Select for Cancer Stem Cells?

    International Nuclear Information System (INIS)

    Chiodi, Ilaria; Belgiovine, Cristina; Donà, Francesca; Scovassi, A. Ivana; Mondello, Chiara

    2011-01-01

    Tumors are generally composed of different cell types. In recent years, it has been shown that in many types of cancers a subset of cells show peculiar characteristics, such as the ability to induce tumors when engrafted into host animals, self-renew and being immortal, and give rise to a differentiated progeny. These cells have been defined as cancer stem cells (CSCs) or tumor initiating cells. CSCs can be isolated both from tumor specimens and established cancer cell lines on the basis of their ability to exclude fluorescent dyes, express specific cell surface markers or grow in particular culture conditions. A key feature of CSCs is their resistance to chemotherapeutic agents, which could contribute to the remaining of residual cancer cells after therapeutic treatments. It has been shown that CSC-like cells can be isolated after drug treatment of cancer cell lines; in this review, we will describe the strategies so far applied to identify and isolate CSCs. Furthermore, we will discuss the possible use of these selected populations to investigate CSC biology and develop new anticancer drugs

  20. Replication of cultured lung epithelial cells

    International Nuclear Information System (INIS)

    Guzowski, D.; Bienkowski, R.

    1986-01-01

    The authors have investigated the conditions necessary to support replication of lung type 2 epithelial cells in culture. Cells were isolated from mature fetal rabbit lungs (29d gestation) and cultured on feeder layers of mitotically inactivated 3T3 fibroblasts. The epithelial nature of the cells was demonstrated by indirect immunofluorescent staining for keratin and by polyacid dichrome stain. Ultrastructural examination during the first week showed that the cells contained myofilaments, microvilli and lamellar bodies (markers for type 2 cells). The following changes were observed after the first week: increase in cell size; loss of lamellar bodies and appearance of multivesicular bodies; increase in rough endoplasmic reticulum and golgi; increase in tonafilaments and well-defined junctions. General cell morphology was good for up to 10 wk. Cells cultured on plastic surface degenerated after 1 wk. Cell replication was assayed by autoradiography of cultures exposed to ( 3 H)-thymidine and by direct cell counts. The cells did not replicate during the first week; however, between 2-10 wk the cells incorporated the label and went through approximately 6 population doublings. They have demonstrated that lung alveolar epithelial cells can replicate in culture if they are maintained on an appropriate substrate. The coincidence of ability to replicate and loss of markers for differentiation may reflect the dichotomy between growth and differentiation commonly observed in developing systems

  1. A novel three-dimensional cell culture method enhances antiviral drug screening in primary human cells.

    Science.gov (United States)

    Koban, Robert; Neumann, Markus; Daugs, Aila; Bloch, Oliver; Nitsche, Andreas; Langhammer, Stefan; Ellerbrok, Heinz

    2018-02-01

    Gefitinib is a specific inhibitor of the epidermal growth factor receptor (EGFR) and FDA approved for treatment of non-small cell lung cancer. In a previous study we could show the in vitro efficacy of gefitinib for treatment of poxvirus infections in monolayer (2D) cultivated cell lines. Permanent cell lines and 2D cultures, however, are known to be rather unphysiological; therefore it is difficult to predict whether determined effective concentrations or the drug efficacy per se are transferable to the in vivo situation. 3D cell cultures, which meanwhile are widely distributed across all fields of research, are a promising tool for more predictive in vitro investigations of antiviral compounds. In this study the spreading of cowpox virus and the antiviral efficacy of gefitinib were analyzed in primary human keratinocytes (NHEK) grown in a novel 3D extracellular matrix-based cell culture model and compared to the respective monolayer culture. 3D-cultivated NHEK grew in a polarized and thus a more physiological manner with altered morphology and close cell-cell contact. Infected cultures showed a strongly elevated sensitivity towards gefitinib. EGFR phosphorylation, cell proliferation, and virus replication were significantly reduced in 3D cultures at gefitinib concentrations which were at least 100-fold lower than those in monolayer cultures and well below the level of cytotoxicity. Our newly established 3D cell culture model with primary human cells is an easy-to-handle alternative to conventional monolayer cell cultures and previously described more complex 3D cell culture systems. It can easily be adapted to other cell types and a broad spectrum of viruses for antiviral drug screening and many other aspects of virus research under more in vivo-like conditions. In consequence, it may contribute to a more targeted realization of necessary in vivo experiments. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Zeolite scaffolds for cultures of human breast cancer cells. Part II: Effect of pure and hybrid zeolite membranes on neoplastic and metastatic activity control.

    Science.gov (United States)

    Tavolaro, Palmira; Martino, Guglielmo; Andò, Sebastiano; Tavolaro, Adalgisa

    2016-11-01

    This work is focused on the response of two invasive phenotypes of human breast cancer cells, MCF-7 and MDA-MB-231, grown on synthesized zeolite scaffolds in order to study the influence of those biomaterials in controlled conditions with and without anti-tumoral drug treatments. Our research was directed to the use of doxorubicin (DOX) and bergapten (5-MOP). The former is broadly considered the most active single agent available for the treatment of breast cancer, the second is a natural psoralen with an apoptotic effect. The results indicate that both drugs inhibit the cell viability of all cell lines grown on all zeolite scaffolds and that all Pure Zeolite Membranes are more responsive with respect to all Mixed Matrix Membranes. Moreover, the results after treatment with DOX at a concentration of 7.4μM for 24h, show that the expression of the matrix metalloproteinases (MMP-2 and MMP-9) is greatly reduced in both cell lines, especially in those adherent on Pure Zeolite Scaffolds. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Cancer stem cell markers in common cancers - therapeutic implications

    DEFF Research Database (Denmark)

    Klonisch, Thomas; Wiechec, Emilia; Hombach-Klonisch, Sabine

    2008-01-01

    Rapid advance in the cancer stem cell field warrants optimism for the development of more reliable cancer therapies within the next 2-3 decades. Below, we characterize and compare the specific markers that are present on stem cells, cancer cells and cancer stem cells (CSC) in selected tissues...

  4. 3D Cell Culture in Alginate Hydrogels

    Directory of Open Access Journals (Sweden)

    Therese Andersen

    2015-03-01

    Full Text Available This review compiles information regarding the use of alginate, and in particular alginate hydrogels, in culturing cells in 3D. Knowledge of alginate chemical structure and functionality are shown to be important parameters in design of alginate-based matrices for cell culture. Gel elasticity as well as hydrogel stability can be impacted by the type of alginate used, its concentration, the choice of gelation technique (ionic or covalent, and divalent cation chosen as the gel inducing ion. The use of peptide-coupled alginate can control cell–matrix interactions. Gelation of alginate with concomitant immobilization of cells can take various forms. Droplets or beads have been utilized since the 1980s for immobilizing cells. Newer matrices such as macroporous scaffolds are now entering the 3D cell culture product market. Finally, delayed gelling, injectable, alginate systems show utility in the translation of in vitro cell culture to in vivo tissue engineering applications. Alginate has a history and a future in 3D cell culture. Historically, cells were encapsulated in alginate droplets cross-linked with calcium for the development of artificial organs. Now, several commercial products based on alginate are being used as 3D cell culture systems that also demonstrate the possibility of replacing or regenerating tissue.

  5. Angular-dependent light scattering from cancer cells in different phases of the cell cycle.

    Science.gov (United States)

    Lin, Xiaogang; Wan, Nan; Weng, Lingdong; Zhou, Yong

    2017-10-10

    Cancer cells in different phases of the cell cycle result in significant differences in light scattering properties. In order to harvest cancer cells in particular phases of the cell cycle, we cultured cancer cells through the process of synchronization. Flow cytometric analysis was applied to check the results of cell synchronization and prepare for light scattering measurements. Angular-dependent light scattering measurements of cancer cells arrested in the G1, S, and G2 phases have been performed. Based on integral calculations for scattering intensities from 5° to 10° and from 110° to 150°, conclusions have been reached. Clearly, the sizes of the cancer cells in different phases of the cell cycle dominated the forward scatter. Accompanying the increase of cell size with the progression of the cell cycle, the forward scattering intensity also increased. Meanwhile, the DNA content of cancer cells in every phase of the cell cycle is responsible for light scattering at large scatter angles. The higher the DNA content of cancer cells was, the greater the positive effect on the high-scattering intensity. As expected, understanding the relationships between the light scattering from cancer cells and cell cycles will aid in the development of cancer diagnoses. Also, it may assist in the guidance of antineoplastic drugs clinically.

  6. A Seat at the Table: Culturally based cancer research

    Science.gov (United States)

    NCI supports research to address cancer disparities among American Indian and Alaska Native populations. In this video, two researchers advocate for more culturally sensitive practices to help people who are most disproportionately affected by cancer disparities.

  7. Advances in cell culture: anchorage dependence

    Science.gov (United States)

    Merten, Otto-Wilhelm

    2015-01-01

    Anchorage-dependent cells are of great interest for various biotechnological applications. (i) They represent a formidable production means of viruses for vaccination purposes at very large scales (in 1000–6000 l reactors) using microcarriers, and in the last decade many more novel viral vaccines have been developed using this production technology. (ii) With the advent of stem cells and their use/potential use in clinics for cell therapy and regenerative medicine purposes, the development of novel culture devices and technologies for adherent cells has accelerated greatly with a view to the large-scale expansion of these cells. Presently, the really scalable systems—microcarrier/microcarrier-clump cultures using stirred-tank reactors—for the expansion of stem cells are still in their infancy. Only laboratory scale reactors of maximally 2.5 l working volume have been evaluated because thorough knowledge and basic understanding of critical issues with respect to cell expansion while retaining pluripotency and differentiation potential, and the impact of the culture environment on stem cell fate, etc., are still lacking and require further studies. This article gives an overview on critical issues common to all cell culture systems for adherent cells as well as specifics for different types of stem cells in view of small- and large-scale cell expansion and production processes. PMID:25533097

  8. General Information about Renal Cell Cancer

    Science.gov (United States)

    ... Tumors Treatment Genetics of Kidney Cancer Research Renal Cell Cancer Treatment (PDQ®)–Patient Version General Information About Renal Cell Cancer Go to Health Professional Version Key Points Renal ...

  9. Treatment Option Overview (Renal Cell Cancer)

    Science.gov (United States)

    ... Tumors Treatment Genetics of Kidney Cancer Research Renal Cell Cancer Treatment (PDQ®)–Patient Version General Information About Renal Cell Cancer Go to Health Professional Version Key Points Renal ...

  10. Drugs Approved for Kidney (Renal Cell) Cancer

    Science.gov (United States)

    ... Your Treatment Research Drugs Approved for Kidney (Renal Cell) Cancer This page lists cancer drugs approved by the ... not listed here. Drugs Approved for Kidney (Renal Cell) Cancer Afinitor (Everolimus) Aldesleukin Avastin (Bevacizumab) Axitinib Bevacizumab Cabometyx ( ...

  11. ACVP-05: Virus Genetic Analysis from Cell-Free Plasma, Virally Infected Cells or Tissues and Cultured Supernatant Via Single Genome Amplification and Direct Sequencing | Frederick National Laboratory for Cancer Research

    Science.gov (United States)

    The Viral Evolution Core within the AIDS and Cancer Virus Program will extract viral RNA/DNA from cell-free or cell-associated samples. Complementary (cDNA) will be generated as needed, and cDNA or DNA will be diluted to a single copy prior to nested

  12. Culture conditions defining glioblastoma cells behavior: what is the impact for novel discoveries?

    Science.gov (United States)

    Ledur, Pítia Flores; Onzi, Giovana Ravizzoni; Zong, Hui; Lenz, Guido

    2017-09-15

    In cancer research, the use of established cell lines has gradually been replaced by primary cell cultures due to their better representation of in vivo cancer cell behaviors. However, a major challenge with primary culture involves the finding of growth conditions that minimize alterations in the biological state of the cells. To ensure reproducibility and translational potentials for research findings, culture conditions need to be chosen so that the cell population in culture best mimics tumor cells in vivo . Glioblastoma (GBM) is one of the most aggressive and heterogeneous tumor types and the GBM research field would certainly benefit from culture conditions that could maintain the original plethora of phenotype of the cells. Here, we review culture media and supplementation options for GBM cultures, the rationale behind their use, and how much those choices affect drug-screening outcomes. We provide an overview of 120 papers that use primary GBM cultures and discuss the current predominant conditions. We also show important primary research data indicating that "mis-cultured" glioma cells can acquire unnatural drug sensitivity, which would have devastating effects for clinical translations. Finally, we propose the concurrent test of four culture conditions to minimize the loss of cell coverage in culture.

  13. Localization of thymosin ß10 in breast cancer cells

    DEFF Research Database (Denmark)

    Mælan, A.ase Elisabeth; Rasmussen, Trine Kring; Larsson, Lars-Inge

    2007-01-01

    as in cell motility and spreading. We have studied the distribution of endogenously expressed thymosin ß10 in cultured human breast cancer cell lines. Both unperturbed monolayer cultures and wound-healing models were examined using double-staining for thymosin ß10 and polymerized (F-) actin. Our findings...... show that thymosin ß10 is expressed in all three-cancer cell lines (SK-BR-3, MCF-7 and MDA-MB-231) studied. No or little staining was detected in confluent cells, whereas strong staining occurred in semiconfluent cells and in cells populating monolayer wounds. Importantly, the distribution of staining...... for thymosin ß10 was inverse of staining for F-actin. These data support a physiological role for thymosin ß10 in sequestration of G-actin as well as in cancer cell motility....

  14. Ursodeoxycholic acid inhibits the proliferation of colon cancer cells by regulating oxidative stress and cancer stem-like cell growth

    Science.gov (United States)

    Kim, EuiJoo

    2017-01-01

    Introduction The regulation of reactive oxygen species (ROS) exists as a therapeutic target for cancer treatments. Previous studies have shown that ursodeoxycholic acid (UDCA) suppresses the proliferation of colon cancer cells. The aim of this study was to evaluate the effect of UDCA upon the proliferation of colon cancer cells as a direct result of the regulation of ROS. Method Colon cancer cell lines (HT29 and HCT116) were treated with UDCA. The total number of cells and the number of dead cells were determined using cell counters. A fluorescein isothiocyanate-bromodeoxyuridine flow kit was used to analyze cell cycle variations. Upon exposure to UDCA, the protein levels of p27, p21, CDK2, CDK4 and CDK6 were determined using western blotting, and qRT-PCR was used to determine levels of mRNA. We preformed dichlorofluorescindiacetate (DCF-DA) staining to detect alteration of intracellular ROS using fluorescence activated cell sorting (FACS). Colon cancer stem-like cell lines were generated by tumorsphere culture and treated with UDCA for seven days. The total number of tumorspheres was determined using microscopy. Results We found that UDCA reduced the total number of colon cancer cells, but did not increase the number of dead cells. UDCA inhibited the G1/S and G2/M transition phases in colon cancer cells. UDCA induced expression of cell cycle inhibitors such as p27 and p21. However, it was determined that UDCA suppressed levels of CDK2, CDK4, and CDK6. UDCA regulated intracellular ROS generation in colon cancer cells, and induced activation of Erk1/2. Finally, UDCA inhibited formation of colon cancer stem-like cells. Conclusion Our results indicate that UDCA suppresses proliferation through regulation of oxidative stress in colon cancer cells, as well as colon cancer stem-like cells. PMID:28708871

  15. Advances in 3D neuronal cell culture

    NARCIS (Netherlands)

    Frimat, Jean Philippe; Xie, Sijia; Bastiaens, Alex; Schurink, Bart; Wolbers, Floor; Den Toonder, Jaap; Luttge, Regina

    2015-01-01

    In this contribution, the authors present our advances in three-dimensional (3D) neuronal cell culture platform technology contributing to controlled environments for microtissue engineering and analysis of cellular physiological and pathological responses. First, a micromachined silicon sieving

  16. Cell and Molecular Biology of Ataxia Telangiectasia Heterozygous Human Mammary Epithelial Cells Irradiated in Culture

    Science.gov (United States)

    Richmond, Robert C.

    2001-01-01

    Autologous isolates of cell types from obligate heterozygotes with the autosomal disorder ataxia-telangiectasia (A-T)were used to begin a tissue culture model for assessing pathways of radiation-induced cancer formation in this target tissue. This was done by establishing cultures of stromal fibroblasts and long-term growth human mammary epithelial cells (HMEC) in standard 2-dimensional tissue culture in order to establish expression of markers detailing early steps of carcinogenesis. The presumptive breast cancer susceptibility of A-T heterozygotes as a sequel to damage caused by ionizing radiation provided reason to study expression of markers in irradiated HMEC. Findings from our study with HMEC have included determination of differences in specific protein expression amongst growth phase (e.g., log vs stationary) and growth progression (e.g., pass 7 vs pass 9), as well as differences in morphologic markers within populations of irradiated HMEC (e.g., development of multinucleated cells).

  17. Metabolic characterization of invaded cells of the pancreatic cancer cell line, PANC?1

    OpenAIRE

    Fujita, Mayumi; Imadome, Kaori; Imai, Takashi

    2017-01-01

    We previously reported that about 0.4% of cells in the cultured human pancreatic cancer cell line, PANC?1, can invade matrigel during the transwell invasion assay, suggesting that these invaded PANC?1 cells may have specific characteristics to keep their invasive potential. To identify the metabolic characterization specific in the invaded PANC?1 cells, metabolome analysis of the invaded PANC?1 compared with the whole cultured PANC?1 was performed using CE?TOFMS, and concentrations of 110 met...

  18. Biocompatibility of Tygon® tubing in microfluidic cell culture.

    Science.gov (United States)

    Jiang, Xiao; Jeffries, Rex E; Acosta, Miguel A; Tikunov, Andrey P; Macdonald, Jeffrey M; Walker, Glenn M; Gamcsik, Michael P

    2015-02-01

    Growth of the MDA-MB-231 breast cancer cell line in microfluidic channels was inhibited when culture media was delivered to the channels via microbore Tygon® tubing. Culture media incubated within this tubing also inhibited growth of these cells in conventional 96-well plates. These detrimental effects were not due to depletion of critical nutrients due to adsorption of media components onto the tubing surface. A pH change was also ruled out as a cause. Nuclear magnetic resonance spectroscopy of the cell growth media before and after incubation in the tubing confirmed no detectable loss of media components but did detect the presence of additional unidentified signals in the aliphatic region of the spectrum. These results indicate leaching of a chemical species from microbore Tygon® tubing that can affect cell growth in microfluidic devices.

  19. Migration assay on primary culture isolated from patient's primary breast cancer tissue

    Directory of Open Access Journals (Sweden)

    ED Yuliana

    2014-12-01

    Full Text Available Background: Migration is an essential component of breast cancer metastasis, which studyhas been concentrated on culture of established breast cancer cell lines that do not accuratelyrepresent the sophistication and heterogeneity of patient's breast cancer. An attempt toperform migration assay using Boyden Chamber Assay (BCA on primary culture originatingfrom patient's breast cancer tissue was developed to accommodate upcoming study of breastcancer migration in lndonesian patients.Methods: Pathologically proven primary breast cancer tissue samples were obtained fromCiptomangunkusumo Hospital during core (n=4 and incisional (n=3 biopsies of stage llAup to stage lllA breast cancer patients. Following biopsy, the breast cancer tissue samplesunderwent processings to isolate the cancer cells. These cancer cells were -then resuspendedwithin Dulbecco's modified Eagle's medium (DMEM ahd cultured in 12-well plate. The growthof primary culture were observed and compared between the core biopsy and the incisionalbiopsy specimens. Optimization of BCA method was later performed to investigate themigration of the breast cancer primary culture towards different experirnental conditions, whichwere control, Fetal Bovine Serum (FBS, and Stromal Derived Factor-l (SDF-1. Two differentnumber of breast cancer cells were tested for the optimization of the BCA, which were 1 x 105and3x105cells.Results: None of the culture performed on core biopsy specimens grew, while one out ofthree incisional biopsy specimens grew until confluence. The one primary culture that grewwas later assesed using BCA to assess its migration index towards different experimentalconditions. Using 1 x 10s breast cancer cells in the BCA , the result of the absorbance level ofmigrated cells showed that the migration towards SDF-1 (0.529 nearly doubled the migrationtowards controlmedium (0.239 and FBS (0.209. Meanwhile, the absorbance levelwas simiiarbetween the control medium (1.050, FBS (1 .103

  20. Culture of Mouse Neural Stem Cell Precursors

    OpenAIRE

    Currle, D. Spencer; Hu, Jia Sheng; Kolski-Andreaco, Aaron; Monuki, Edwin S.

    2007-01-01

    Primary neural stem cell cultures are useful for studying the mechanisms underlying central nervous system development. Stem cell research will increase our understanding of the nervous system and may allow us to develop treatments for currently incurable brain diseases and injuries. In addition, stem cells should be used for stem cell research aimed at the detailed study of mechanisms of neural differentiation and transdifferentiation and the genetic and environmental signals that direct the...

  1. 3-Dimensional culture systems for anti-cancer compound profiling and high-throughput screening reveal increases in EGFR inhibitor-mediated cytotoxicity compared to monolayer culture systems.

    Science.gov (United States)

    Howes, Amy L; Richardson, Robyn D; Finlay, Darren; Vuori, Kristiina

    2014-01-01

    3-dimensional (3D) culture models have the potential to bridge the gap between monolayer cell culture and in vivo studies. To benefit anti-cancer drug discovery from 3D models, new techniques are needed that enable their use in high-throughput (HT) screening amenable formats. We have established miniaturized 3D culture methods robust enough for automated HT screens. We have applied these methods to evaluate the sensitivity of normal and tumorigenic breast epithelial cell lines against a panel of oncology drugs when cultured as monolayers (2D) and spheroids (3D). We have identified two classes of compounds that exhibit preferential cytotoxicity against cancer cells over normal cells when cultured as 3D spheroids: microtubule-targeting agents and epidermal growth factor receptor (EGFR) inhibitors. Further improving upon our 3D model, superior differentiation of EC50 values in the proof-of-concept screens was obtained by co-culturing the breast cancer cells with normal human fibroblasts and endothelial cells. Further, the selective sensitivity of the cancer cells towards chemotherapeutics was observed in 3D co-culture conditions, rather than as 2D co-culture monolayers, highlighting the importance of 3D cultures. Finally, we examined the putative mechanisms that drive the differing potency displayed by EGFR inhibitors. In summary, our studies establish robust 3D culture models of human cells for HT assessment of tumor cell-selective agents. This methodology is anticipated to provide a useful tool for the study of biological differences within 2D and 3D culture conditions in HT format, and an important platform for novel anti-cancer drug discovery.

  2. Sponge cell culture? A molecular identification method for sponge cells

    NARCIS (Netherlands)

    Sipkema, D.; Heilig, G.H.J.; Akkermans, A.D.L.; Osinga, R.; Tramper, J.; Wijffels, R.H.

    2003-01-01

    Dissociated sponge cells are easily confused with unicellular organisms. This has been an obstacle in the development of sponge-cell lines. We developed a molecular detection method to identify cells of the sponge Dysidea avara in dissociated cell cultures. The 18S ribosomal RNA gene from a Dysidea

  3. Multifaceted Interpretation of Colon Cancer Stem Cells.

    Science.gov (United States)

    Hatano, Yuichiro; Fukuda, Shinya; Hisamatsu, Kenji; Hirata, Akihiro; Hara, Akira; Tomita, Hiroyuki

    2017-07-05

    Colon cancer is one of the leading causes of cancer-related deaths worldwide, despite recent advances in clinical oncology. Accumulating evidence sheds light on the existence of cancer stem cells and their role in conferring therapeutic resistance. Cancer stem cells are a minor fraction of cancer cells, which enable tumor heterogeneity and initiate tumor formation. In addition, these cells are resistant to various cytotoxic factors. Therefore, elimination of cancer stem cells is difficult but essential to cure the malignant foci completely. Herein, we review the recent evidence for intestinal stem cells and colon cancer stem cells, methods to detect the tumor-initiating cells, and clinical significance of cancer stem cell markers. We also describe the emerging problems of cancer stem cell theory, including bidirectional conversion and intertumoral heterogeneity of stem cell phenotype.

  4. Establishment and Analysis of Cancer Stem-Like and Non-Cancer Stem-Like Clone Cells from the Human Colon Cancer Cell Line SW480.

    Directory of Open Access Journals (Sweden)

    Akari Takaya

    Full Text Available 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.

  5. Establishment and Analysis of Cancer Stem-Like and Non-Cancer Stem-Like Clone Cells from the Human Colon Cancer Cell Line SW480.

    Science.gov (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

    2016-01-01

    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.

  6. Cell culture supernatants for detection perforin ELISA

    African Journals Online (AJOL)

    Najwa

    2014-02-19

    Feb 19, 2014 ... Leukemia is a cancer originating in any of hematopoietic cell that tends to ... treatment of children (Borek and Jaskolski, 2001). The current study was .... which led to the best results at 48 h of exposure than after 72 h of cells ...

  7. Inhibition of DNA synthesis in cultured lymphocytes and tumor cells by extracts of betel nut, tobacco, and miang leaf, plant substances associated with cancer of the ororespiratory epithelium.

    Science.gov (United States)

    Yang, J A; Huber, S A; Lucas, Z J

    1979-12-01

    The high incidence of oropharyngeal, esophageal, and laryngeal cancers in certain parts of the world has been ascribed to conjugated tannins found in certain folk medicinal herbs. We extracted miang leaf and betel nut with phosphate-buffered saline (0.14 M NaCl, 0.15 M potassium phosphate buffer, pH 7.4) and found that the extracts inhibited [3H]thymidine incorporation by phytohemagglutinin-stimulated human lymphocytes and by rat mammary tumor and mouse L-cells in logarithmic growth. Pretreating the lymphocytes for 1 or 4 hr with the extracts inhibited phytohemagglutinin-induced thymidine incorporation 72 hr later. At concentrations of 2.5 volumes % or lower, miang and betel nut extracts inhibited thymidine incorporation by 40 to 98% without any apparent signs of toxicity as demonstrated by the 66Rb equilibrium assay. In addition, neither extract inhibited cytotoxicity of rat mammary tumor cells by immune syngeneic spleen cells. The molecular weights of the inhibitory factors were between 1,000 and 10,000 daltons as determined by ultrafiltration and were unaffected by boiling for 3 min or by treatment with alcohol and, therefore, are probably not proteins. This in vitro demonstration of inhibition of DNA synthesis by these plant extracts presumably enriched for conjugated tannins may relate to inhibition of growth of rats and chicks fed conjugated tanin-contaminated sorghum feed. The carcinogenic potential of either these extracts or conjugated tannins is not yet established.

  8. Effects of 12-O-tetradecanoylphorbol-13-acetate in combination with gemcitabine on Panc-1 pancreatic cancer cells cultured in vitro or Panc-1 tumors grown in immunodeficient mice.

    Science.gov (United States)

    Zheng, Xi; Cui, Xiao-Xing; Gao, Zhi; Verano, Michael; Huang, Mou-Tuan; Liu, Yue; Rabson, Arnold B; Conney, Allan H

    2012-12-01

    In the present study, the effects of 12-O-tetra-decanoylphorbol-13-acetate (TPA) alone or in combination with gemcitabine on the growth of Panc-1 pancreatic cancer cells cultured in vitro or grown in NCr immunodeficient nude mice were investigated. Combinations of TPA and gemcitabine synergi-stically inhibited the growth and induced apoptosis in Panc-1 cells. The combination of TPA (0.16 nM) and gemcitabine (0.5 µM) induced a marked increase in phosphorylated c-Jun NH2-terminal kinase (JNK) in the Panc-1 cells. In animal experiments, NCr nude mice with established Panc-1 tumors received daily intraperitoneal (i.p.) injections of TPA (50 ng/g body weight/day) or gemcitabine (0.5 µg/g body weight/day) alone or in combination for 26 days. Treatment with daily i.p. injections of low doses of TPA or gemcitabine alone had a modest inhibitory effect on the growth of the tumors. However, the combination of low doses of TPA and gemcitabine more potently inhibited the growth of Panc-1 tumors than either agent used individually. Treatment with TPA or gemcitabine alone or in combination did not affect the body weight of the animals. Clinical trials with TPA alone or in combination with gemcitabine on patients with pancreatic cancer are warranted in order to confirm our results.

  9. Flux analysis of mammalian cell culture

    NARCIS (Netherlands)

    Martens, D.E.; Tramper, J.

    2010-01-01

    Animal cells are used for the production of vaccines and pharmaceutical proteins. The increase in demand for these products requires an increase in volumetric productivity of animal cell culture processes, which can be attained through an increase in biomass concentration and/or specific

  10. Quantitative volumetric Raman imaging of three dimensional cell cultures

    KAUST Repository

    Kallepitis, Charalambos; Bergholt, Mads S.; Mazo, Manuel M.; Leonardo, Vincent; Skaalure, Stacey C.; Maynard, Stephanie A.; Stevens, Molly M.

    2017-01-01

    in conventional cell culture systems and mesenchymal stem cells inside biomimetic hydrogels that supplied a 3D cell culture environment. We demonstrate visualization and quantification of fine details in cell shape, cytoplasm, nucleus, lipid bodies

  11. Organoid cultures derived from patients with advanced prostate cancer

    NARCIS (Netherlands)

    Gao, Dong; Vela, Ian; Sboner, Andrea; Iaquinta, Phillip J; Karthaus, Wouter R; Gopalan, Anuradha; Dowling, Catherine; Wanjala, Jackline N; Undvall, Eva A; Arora, Vivek K; Wongvipat, John; Kossai, Myriam; Ramazanoglu, Sinan; Barboza, Luendreo P; Di, Wei; Cao, Zhen; Zhang, Qi Fan; Sirota, Inna; Ran, Leili; MacDonald, Theresa Y; Beltran, Himisha; Mosquera, Juan-Miguel; Touijer, Karim A; Scardino, Peter T; Laudone, Vincent P; Curtis, Kristen R; Rathkopf, Dana E; Morris, Michael J; Danila, Daniel C; Slovin, Susan F; Solomon, Stephen B; Eastham, James A; Chi, Ping; Carver, Brett; Rubin, Mark A; Scher, Howard I; Clevers, Hans; Sawyers, Charles L; Chen, Yu

    2014-01-01

    The lack of in vitro prostate cancer models that recapitulate the diversity of human prostate cancer has hampered progress in understanding disease pathogenesis and therapy response. Using a 3D organoid system, we report success in long-term culture of prostate cancer from biopsy specimens and

  12. Hanging drop cultures of human testis and testis cancer samples

    DEFF Research Database (Denmark)

    Jørgensen, Anne; Young, J; Nielsen, J E

    2014-01-01

    cultured in 'hanging drops' and effects of activin A and follistatin treatment were investigated in seminoma cultures. RESULTS: Testis fragments with normal spermatogenesis or CIS cells were cultured for 14 days with sustained proliferation of germ cells and CIS cells and without increased apoptosis....... Seminoma cultures survived 7 days, with proliferating cells detectable during the first 5 days. Activin A treatment significantly reduced KIT transcript and protein levels in seminoma cultures, thereby demonstrating a specific treatment response. CONCLUSIONS: Hanging drop cultures of human testis...

  13. Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

    Science.gov (United States)

    Stampfer, Martha R; Garbe, James C

    2015-02-24

    Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

  14. Establishment of three-dimensional cultures of human pancreatic duct epithelial cells

    International Nuclear Information System (INIS)

    Gutierrez-Barrera, Angelica M.; Menter, David G.; Abbruzzese, James L.; Reddy, Shrikanth A.G.

    2007-01-01

    Three-dimensional (3D) cultures of epithelial cells offer singular advantages for studies of morphogenesis or the role of cancer genes in oncogenesis. In this study, as part of establishing a 3D culture system of pancreatic duct epithelial cells, we compared human pancreatic duct epithelial cells (HPDE-E6E7) with pancreatic cancer cell lines. Our results show, that in contrast to cancer cells, HPDE-E6E7 organized into spheroids with what appeared to be apical and basal membranes and a luminal space. Immunostaining experiments indicated that protein kinase Akt was phosphorylated (Ser473) and CTMP, a negative Akt regulator, was expressed in both HPDE-E6E7 and cancer cells. However, a nuclear pool of CTMP was detectable in HPDE-E6E7 cells that showed a dynamic concentrated expression pattern, a feature that further distinguished HPDE-E637 cells from cancer cells. Collectively, these data suggest that 3D cultures of HPDE-E6E7 cells are useful for investigating signaling and morphological abnormalities in pancreatic cancer cells

  15. Isolation and propagation of colon cancer stem cells

    NARCIS (Netherlands)

    Prasetyanti, Pramudita R.; Zimberlin, Cheryl; de Sousa E Melo, Felipe; Medema, Jan Paul

    2013-01-01

    The design of tissue culture conditions that faithfully reproduce the characteristics of cells in their native environment remains one of the main challenges of cancer stem cell (CSC) biology. Here we describe a detailed methodology for the isolation and expansion of both human colon CSCs and mouse

  16. Substrate utilisation by plant-cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Fowler, M W

    1982-01-01

    Plant cell cultures have been grown on a wide range of carbon sources in addition to the traditional ones of sucrose and glucose. Biomass yields and growth rates vary greatly between the different carbon sources and there is a variation in response between different cell cultures to individual carbon sources. Some attempts have been made to grow cell cultures on 'waste' and related carbon sources, such as lactose, maltose, starch, molasses and milk whey. Only maltose was found to support growth to anything near the levels observed with glucose and sucrose. In the case of molasses carbon source cell growth was either non-existent or only just measurable. All the data point to glucose as being the most suitable carbon source, principally on the grounds of biomass yield and growth rate. It should be noted, however, that other carbon sources do appear to have a major (positive) influence on natural product synthesis. Uptake into the cell is an important aspect of carbohydrate utilisation. There is strong evidence that from disaccharides upwards, major degradation to smaller units occurs before uptake. In some cases the necessary enzymes appear to be excreted into the culture broth, in others they may be located within the cell wall; invertase that hydrolyses sucrose is a good example. Once the products of carbohydrate degradation and mobilisation enter the cell they may suffer one of two fates, oxidation or utilisation for biosynthesis. The precise split between these two varies depending on such factors as cell growth rate, cell size, nutrient broth composition and carbohydrate status of the cells. In general rapidly growing cells have a high rate of oxidation, whereas cells growing more slowly tend to be more directed towards biosynthesis. Carbohydrate utilisation is a key area of study, underpinning as it does both biomass yield and natural product synthesis. (Refs. 13).

  17. Sensor Access to the Cellular Microenvironment Using the Sensing Cell Culture Flask.

    Science.gov (United States)

    Kieninger, Jochen; Tamari, Yaara; Enderle, Barbara; Jobst, Gerhard; Sandvik, Joe A; Pettersen, Erik O; Urban, Gerald A

    2018-04-26

    The Sensing Cell Culture Flask (SCCF) is a cell culture monitoring system accessing the cellular microenvironment in 2D cell culture using electrochemical microsensors. The system is based on microfabricated sensor chips embedded in standard cell culture flasks. Ideally, the sensor chips could be equipped with any electrochemical sensor. Its transparency allows optical inspection of the cells during measurement. The surface of the sensor chip is in-plane with the flask surface allowing undisturbed cell growth on the sensor chip. A custom developed rack system allows easy usage of multiple flasks in parallel within an incubator. The presented data demonstrates the application of the SCCF with brain tumor (T98G) and breast cancer (T-47D) cells. Amperometric oxygen sensors were used to monitor cellular respiration with different incubation conditions. Cellular acidification was accessed with potentiometric pH sensors using electrodeposited iridium oxide films. The system itself provides the foundation for electrochemical monitoring systems in 3D cell culture.

  18. Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins

    Science.gov (United States)

    2014-01-01

    Background A limiting factor in performing proteomics analysis on cancerous cells is the difficulty in obtaining sufficient amounts of starting material. Cell lines can be used as a simplified model system for studying changes that accompany tumorigenesis. This study used two-dimensional gel electrophoresis (2DE) to compare the whole cell proteome of oral cancer cell lines vs normal cells in an attempt to identify cancer associated proteins. Results Three primary cell cultures of normal cells with a limited lifespan without hTERT immortalization have been successfully established. 2DE was used to compare the whole cell proteome of these cells with that of three oral cancer cell lines. Twenty four protein spots were found to have changed in abundance. MALDI TOF/TOF was then used to determine the identity of these proteins. Identified proteins were classified into seven functional categories – structural proteins, enzymes, regulatory proteins, chaperones and others. IPA core analysis predicted that 18 proteins were related to cancer with involvements in hyperplasia, metastasis, invasion, growth and tumorigenesis. The mRNA expressions of two proteins – 14-3-3 protein sigma and Stress-induced-phosphoprotein 1 – were found to correlate with the corresponding proteins’ abundance. Conclusions The outcome of this analysis demonstrated that a comparative study of whole cell proteome of cancer versus normal cell lines can be used to identify cancer associated proteins. PMID:24422745

  19. Cell culture experiments planned for the space bioreactor

    Science.gov (United States)

    Morrison, Dennis R.; Cross, John H.

    1987-01-01

    Culturing of cells in a pilot-scale bioreactor remains to be done in microgravity. An approach is presented based on several studies of cell culture systems. Previous and current cell culture research in microgravity which is specifically directed towards development of a space bioprocess is described. Cell culture experiments planned for a microgravity sciences mission are described in abstract form.

  20. T cell resistance to activation by dendritic cells requires long-term culture in simulated microgravity

    Science.gov (United States)

    Bradley, Jillian H.; Stein, Rachel; Randolph, Brad; Molina, Emily; Arnold, Jennifer P.; Gregg, Randal K.

    2017-11-01

    Immune impairment mediated by microgravity threatens the success of space exploration requiring long-duration spaceflight. The cells of most concern, T lymphocytes, coordinate the host response against microbial and cancerous challenges leading to elimination and long-term protection. T cells are activated upon recognition of specific microbial peptides bound on the surface of antigen presenting cells, such as dendritic cells (DC). Subsequently, this engagement results in T cell proliferation and differentiation into effector T cells driven by autocrine interleukin-2 (IL-2) and other cytokines. Finally, the effector T cells acquire the weaponry needed to destroy microbial invaders and tumors. Studies conducted on T cells during spaceflight, or using Earth-based culture systems, have shown reduced production of cytokines, proliferation and effector functions as compared to controls. This may account for the cases of viral reactivation events and opportunistic infections associated with astronauts of numerous missions. This work has largely been based upon the outcome of T cell activation by stimulatory factors that target select T cell signaling pathways rather than the complex, signaling events related to the natural process of antigen presentation by DC. This study tested the response of an ovalbumin peptide-specific T cell line, OT-II TCH, to activation by DC when the T cells were cultured 24-120 h in a simulated microgravity (SMG) environment generated by a rotary cell culture system. Following 72 h culture of T cells in SMG (SMG-T) or control static (Static-T) conditions, IL-2 production by the T cells was reduced in SMG-T cells compared to Static-T cells upon stimulation by phorbol 12-myristate 13-acetate (PMA) and ionomycin. However, when the SMG-T cells were stimulated with DC and peptide, IL-2 was significantly increased compared to Static-T cells. Such enhanced IL-2 production by SMG-T cells peaked at 72 h SMG culture time and decreased thereafter. When

  1. T cell resistance to activation by dendritic cells requires long-term culture in simulated microgravity.

    Science.gov (United States)

    Bradley, Jillian H; Stein, Rachel; Randolph, Brad; Molina, Emily; Arnold, Jennifer P; Gregg, Randal K

    2017-11-01

    Immune impairment mediated by microgravity threatens the success of space exploration requiring long-duration spaceflight. The cells of most concern, T lymphocytes, coordinate the host response against microbial and cancerous challenges leading to elimination and long-term protection. T cells are activated upon recognition of specific microbial peptides bound on the surface of antigen presenting cells, such as dendritic cells (DC). Subsequently, this engagement results in T cell proliferation and differentiation into effector T cells driven by autocrine interleukin-2 (IL-2) and other cytokines. Finally, the effector T cells acquire the weaponry needed to destroy microbial invaders and tumors. Studies conducted on T cells during spaceflight, or using Earth-based culture systems, have shown reduced production of cytokines, proliferation and effector functions as compared to controls. This may account for the cases of viral reactivation events and opportunistic infections associated with astronauts of numerous missions. This work has largely been based upon the outcome of T cell activation by stimulatory factors that target select T cell signaling pathways rather than the complex, signaling events related to the natural process of antigen presentation by DC. This study tested the response of an ovalbumin peptide-specific T cell line, OT-II TCH, to activation by DC when the T cells were cultured 24-120 h in a simulated microgravity (SMG) environment generated by a rotary cell culture system. Following 72 h culture of T cells in SMG (SMG-T) or control static (Static-T) conditions, IL-2 production by the T cells was reduced in SMG-T cells compared to Static-T cells upon stimulation by phorbol 12-myristate 13-acetate (PMA) and ionomycin. However, when the SMG-T cells were stimulated with DC and peptide, IL-2 was significantly increased compared to Static-T cells. Such enhanced IL-2 production by SMG-T cells peaked at 72 h SMG culture time and decreased thereafter

  2. Role of differential physical properties in emergent behavior of 3D cell co-cultures

    Science.gov (United States)

    Kolbman, Dan; Das, Moumita

    2015-03-01

    The biophysics of binary cell populations is of great interest in many biological processes, whether the formation of embryos or the initiation of tumors. During these processes, cells are surrounded by other cell types with different physical properties, often with important consequences. For example, recent experiments on a co-culture of breast cancer cells and healthy breast epithelial cells suggest that the mechanical mismatch between the two cell types may contribute to enhanced migration of the cancer cells. Here we explore how the differential physical properties of different cell types may influence cell-cell interaction, aggregation, and migration. To this end, we study a proof of concept model- a three-dimensional binary system of interacting, active, and deformable particles with different physical properties such as elastic stiffness, contractility, and particle-particle adhesion, using Langevin Dynamics simulations. Our results may provide insights into emergent behavior such as segregation and differential migration in cell co-cultures in three dimensions.

  3. In vivo biomarker expression patterns are preserved in 3D cultures of Prostate Cancer

    International Nuclear Information System (INIS)

    Windus, Louisa C.E.; Kiss, Debra L.; Glover, Tristan; Avery, Vicky M.

    2012-01-01

    Here we report that Prostate Cancer (PCa) cell-lines DU145, PC3, LNCaP and RWPE-1 grown in 3D matrices in contrast to conventional 2D monolayers, display distinct differences in cell morphology, proliferation and expression of important biomarker proteins associated with cancer progression. Consistent with in vivo growth rates, in 3D cultures, all PCa cell-lines were found to proliferate at significantly lower rates in comparison to their 2D counterparts. Moreover, when grown in a 3D matrix, metastatic PC3 cell-lines were found to mimic more precisely protein expression patterns of metastatic tumour formation as found in vivo. In comparison to the prostate epithelial cell-line RWPE-1, metastatic PC3 cell-lines exhibited a down-regulation of E-cadherin and α6 integrin expression and an up-regulation of N-cadherin, Vimentin and β1 integrin expression and re-expressed non-transcriptionally active AR. In comparison to the non-invasive LNCaP cell-lines, PC3 cells were found to have an up-regulation of chemokine receptor CXCR4, consistent with a metastatic phenotype. In 2D cultures, there was little distinction in protein expression between metastatic, non-invasive and epithelial cells. These results suggest that 3D cultures are more representative of in vivo morphology and may serve as a more biologically relevant model in the drug discovery pipeline. -- Highlights: ► We developed and optimised 3D culturing techniques for Prostate Cancer cell-lines. ► We investigated biomarker expression in 2D versus 3D culture techniques. ► Metastatic PC3 cells re-expressed non-transcriptionally active androgen receptor. ► Metastatic PCa cell lines retain in vivo-like antigenic profiles in 3D cultures

  4. Hedgehog Signals Mediate Anti-Cancer Drug Resistance in Three-Dimensional Primary Colorectal Cancer Organoid Culture

    Directory of Open Access Journals (Sweden)

    Tatsuya Usui

    2018-04-01

    Full Text Available Colorectal cancer is one of the most common causes of cancer death worldwide. In patients with metastatic colorectal cancer, combination treatment with several anti-cancer drugs is employed and improves overall survival in some patients. Nevertheless, most patients with metastatic disease are not cured owing to the drug resistance. Cancer stem cells are known to regulate resistance to chemotherapy. In the previous study, we established a novel three-dimensional organoid culture model from tumor colorectal tissues of human patients using an air–liquid interface (ALI method, which contained numerous cancer stem cells and showed resistance to 5-fluorouracil (5-FU and Irinotecan. Here, we investigate which inhibitor for stem cell-related signal improves the sensitivity for anti-cancer drug treatment in tumor ALI organoids. Treatment with Hedgehog signal inhibitors (AY9944, GANT61 decreases the cell viability of organoids compared with Notch (YO-01027, DAPT and Wnt (WAV939, Wnt-C59 signal inhibitors. Combination treatment of AY9944 or GANT61 with 5-FU, Irinotecan or Oxaliplatin decreases the cell viability of tumor organoids compared with each anti-cancer drug alone treatment. Treatment with AY9944 or GANT61 inhibits expression of stem cell markers c-Myc, CD44 and Nanog, likely through the decrease of their transcription factor, GLI-1 expression. Combination treatment of AY9944 or GANT61 with 5-FU or Irinotecan also prevents colony formation of colorectal cancer cell lines HCT116 and SW480. These findings suggest that Hedgehog signals mediate anti-cancer drug resistance in colorectal tumor patient-derived ALI organoids and that the inhibitors are useful as a combinational therapeutic strategy against colorectal cancer.

  5. small Cell Lung Cancer

    African Journals Online (AJOL)

    Blood samples were analyzed for CTC count before and after chemotherapy. Clinical relevance of. CTCs with ... reduction (p < 0.001) in CTC count was also observed after one cycle of chemotherapy. Conclusion: Patients with low CTC ... type of cancer in China with 21.7 % of males and. 14.3 % of females. The incidence of ...

  6. Rapid selection and proliferation of CD133+ cells from cancer cell lines: chemotherapeutic implications.

    Directory of Open Access Journals (Sweden)

    Sarah E Kelly

    2010-04-01

    Full Text Available Cancer stem cells (CSCs are considered a subset of the bulk tumor responsible for initiating and maintaining the disease. Several surface cellular markers have been recently used to identify CSCs. Among those is CD133, which is expressed by hematopoietic progenitor cells as well as embryonic stem cells and various cancers. We have recently isolated and cultured CD133 positive [CD133+] cells from various cancer cell lines using a NASA developed Hydrodynamic Focusing Bioreactor (HFB (Celdyne, Houston, TX. For comparison, another bioreactor, the rotary cell culture system (RCCS manufactured by Synthecon (Houston, TX was used. Both the HFB and the RCCS bioreactors simulate aspects of hypogravity. In our study, the HFB increased CD133+ cell growth from various cell lines compared to the RCCS vessel and to normal gravity control. We observed a +15-fold proliferation of the CD133+ cellular fraction with cancer cells that were cultured for 7-days at optimized conditions. The RCCS vessel instead yielded a (-4.8-fold decrease in the CD133+cellular fraction respect to the HFB after 7-days of culture. Interestingly, we also found that the hypogravity environment of the HFB greatly sensitized the CD133+ cancer cells, which are normally resistant to chemo treatment, to become susceptible to various chemotherapeutic agents, paving the way to less toxic and more effective chemotherapeutic treatment in patients. To be able to test the efficacy of cytotoxic agents in vitro prior to their use in clinical setting on cancer cells as well as on cancer stem cells may pave the way to more effective chemotherapeutic strategies in patients. This could be an important advancement in the therapeutic options of oncologic patients, allowing for more targeted and personalized chemotherapy regimens as well as for higher response rates.

  7. Culture conditions tailored to the cell of origin are critical for maintaining native properties and tumorigenicity of glioma cells.

    Science.gov (United States)

    Ledur, Pítia F; Liu, Chong; He, Hua; Harris, Alexandra R; Minussi, Darlan C; Zhou, Hai-Yan; Shaffrey, Mark E; Asthagiri, Ashok; Lopes, Maria Beatriz S; Schiff, David; Lu, Yi-Cheng; Mandell, James W; Lenz, Guido; Zong, Hui

    2016-10-01

    Cell culture plays a pivotal role in cancer research. However, culture-induced changes in biological properties of tumor cells profoundly affect research reproducibility and translational potential. Establishing culture conditions tailored to the cancer cell of origin could resolve this problem. For glioma research, it has been previously shown that replacing serum with defined growth factors for neural stem cells (NSCs) greatly improved the retention of gene expression profile and tumorigenicity. However, among all molecular subtypes of glioma, our laboratory and others have previously shown that the oligodendrocyte precursor cell (OPC) rather than the NSC serves as the cell of origin for the proneural subtype, raising questions regarding the suitability of NSC-tailored media for culturing proneural glioma cells. OPC-originated mouse glioma cells were cultured in conditions for normal OPCs or NSCs, respectively, for multiple passages. Gene expression profiles, morphologies, tumorigenicity, and drug responsiveness of cultured cells were examined in comparison with freshly isolated tumor cells. OPC media-cultured glioma cells maintained tumorigenicity, gene expression profiles, and morphologies similar to freshly isolated tumor cells. In contrast, NSC-media cultured glioma cells gradually lost their OPC features and most tumor-initiating ability and acquired heightened sensitivity to temozolomide. To improve experimental reproducibility and translational potential of glioma research, it is important to identify the cell of origin, and subsequently apply this knowledge to establish culture conditions that allow the retention of native properties of tumor cells. © The Author(s) 2016. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  8. Targeting cancer cells using 3-bromopyruvate for selective cancer treatment

    Directory of Open Access Journals (Sweden)

    Hussam H Baghdadi

    2017-01-01

    Full Text Available Cancer treatment deserves more research efforts despite intensive conventional treatment modalities for many types of malignancies. Metastasis and resistance to chemotherapy and radiotherapy receive a lot of global research efforts. The current advances in cancer biology may improve targeting the critical metabolic differences that distinguish cancer cells from normal cells. Cancer cells are highly glycolytic for energy production, exhibit the Warburg effect, establish aggressive acidic microenvironment, maintain cancer stem cells, exhibit resistance to chemotherapy, have low antioxidant systems but different ΔΨm (delta psi, mitochondrial transmembrane potential, express P-glycoprotein for multidrug resistance, upregulate glucose transporters and monocarboxylate transporters and are under high steady-state reactive oxygen species conditions. Normal cells differ in all these aspects. Lactate produced through the Warburg effect helps cancer metastasis. Targeting glycolysis reactions for energy production in cancer cells seems promising in decreasing the proliferation and metastasis of cancer cells. 3-bromopyruvate makes use of cancer biology in treating cancer cells, cancer stem cells and preventing metastasis in human cancer as discussed in this review. Updated advances are analyzed here, which include research analysis of background, experience, readings in the field of cancer biology, oncology and biochemistry.

  9. Reprogramming mediated radio-resistance of 3D-grown cancer cells

    International Nuclear Information System (INIS)

    Xue Gang; Ren Zhenxin; Chen Yaxiong; Zhu Jiayun; Du Yarong; Pan Dong; Li Xiaoman; Hu Burong; Grabham, Peter W.

    2015-01-01

    In vitro 3D growth of tumors is a new cell culture model that more closely mimics the features of the in vivo environment and is being used increasingly in the field of biological and medical research. It has been demonstrated that cancer cells cultured in 3D matrices are more radio-resistant compared with cells in monolayers. However, the mechanisms causing this difference remain unclear. Here we show that cancer cells cultured in a 3D microenvironment demonstrated an increase in cells with stem cell properties. This was confirmed by the finding that cells in 3D cultures upregulated the gene and protein expression of the stem cell reprogramming factors such as OCT4, SOX2, NANOG, LIN28 and miR-302a, compared with cells in monolayers. Moreover, the expression of β-catenin, a regulating molecule of reprogramming factors, also increased in 3D-grown cancer cells. These findings suggest that cancer cells were reprogrammed to become stem cell-like cancer cells in a 3D growth culture microenvironment. Since cancer stem cell-like cells demonstrate an increased radio-resistance and chemo-resistance, our results offer a new perspective as to why. Our findings shed new light on understanding the features of the 3D growth cell model and its application in basic research into clinical radiotherapy and medicine. (author)

  10. Stages of Non-Small Cell Lung Cancer

    Science.gov (United States)

    ... Common Cancer Types Recurrent Cancer Common Cancer Types Bladder Cancer Breast Cancer Colorectal Cancer Kidney (Renal Cell) Cancer ... certain genes, such as the epidermal growth factor receptor (EGFR) gene or the anaplastic lymphoma kinase (ALK) ...

  11. Treatment Options by Stage (Non-Small Cell Lung Cancer)

    Science.gov (United States)

    ... Common Cancer Types Recurrent Cancer Common Cancer Types Bladder Cancer Breast Cancer Colorectal Cancer Kidney (Renal Cell) Cancer ... certain genes, such as the epidermal growth factor receptor (EGFR) gene or the anaplastic lymphoma kinase (ALK) ...

  12. Treatment Option Overview (Non-Small Cell Lung Cancer)

    Science.gov (United States)

    ... Common Cancer Types Recurrent Cancer Common Cancer Types Bladder Cancer Breast Cancer Colorectal Cancer Kidney (Renal Cell) Cancer ... certain genes, such as the epidermal growth factor receptor (EGFR) gene or the anaplastic lymphoma kinase (ALK) ...

  13. General Information about Non-Small Cell Lung Cancer

    Science.gov (United States)

    ... Common Cancer Types Recurrent Cancer Common Cancer Types Bladder Cancer Breast Cancer Colorectal Cancer Kidney (Renal Cell) Cancer ... certain genes, such as the epidermal growth factor receptor (EGFR) gene or the anaplastic lymphoma kinase (ALK) ...

  14. Miniature Dielectric Barrier Discharge Nonthermal Plasma Induces Apoptosis in Lung Cancer Cells and Inhibits Cell Migration.

    Science.gov (United States)

    Karki, Surya B; Yildirim-Ayan, Eda; Eisenmann, Kathryn M; Ayan, Halim

    2017-01-01

    Traditional cancer treatments like radiotherapy and chemotherapy have drawbacks and are not selective for killing only cancer cells. Nonthermal atmospheric pressure plasmas with dielectric barrier discharge (DBD) can be applied to living cells and tissues and have emerged as novel tools for localized cancer therapy. The purpose of this study was to investigate the different effects caused by miniature DBD (mDBD) plasma to A549 lung cancer cells. In this study, A549 lung cancer cells cultured in 12 well plates were treated with mDBD plasma for specified treatment times to assess the changes in the size of the area of cell detachment, the viability of attached or detached cells, and cell migration. Furthermore, we investigated an innovative mDBD plasma-based therapy for localized treatment of lung cancer cells through apoptotic induction. Our results indicate that plasma treatment for 120 sec causes apoptotic cell death in 35.8% of cells, while mDBD plasma treatment for 60 sec, 30 sec, or 15 sec causes apoptotic cell death in 20.5%, 14.1%, and 6.3% of the cell population, respectively. Additionally, we observed reduced A549 cell migration in response to mDBD plasma treatment. Thus, mDBD plasma system can be a viable platform for localized lung cancer therapy.

  15. Miniature Dielectric Barrier Discharge Nonthermal Plasma Induces Apoptosis in Lung Cancer Cells and Inhibits Cell Migration

    Directory of Open Access Journals (Sweden)

    Surya B. Karki

    2017-01-01

    Full Text Available Traditional cancer treatments like radiotherapy and chemotherapy have drawbacks and are not selective for killing only cancer cells. Nonthermal atmospheric pressure plasmas with dielectric barrier discharge (DBD can be applied to living cells and tissues and have emerged as novel tools for localized cancer therapy. The purpose of this study was to investigate the different effects caused by miniature DBD (mDBD plasma to A549 lung cancer cells. In this study, A549 lung cancer cells cultured in 12 well plates were treated with mDBD plasma for specified treatment times to assess the changes in the size of the area of cell detachment, the viability of attached or detached cells, and cell migration. Furthermore, we investigated an innovative mDBD plasma-based therapy for localized treatment of lung cancer cells through apoptotic induction. Our results indicate that plasma treatment for 120 sec causes apoptotic cell death in 35.8% of cells, while mDBD plasma treatment for 60 sec, 30 sec, or 15 sec causes apoptotic cell death in 20.5%, 14.1%, and 6.3% of the cell population, respectively. Additionally, we observed reduced A549 cell migration in response to mDBD plasma treatment. Thus, mDBD plasma system can be a viable platform for localized lung cancer therapy.

  16. How Can We Treat Cancer Disease Not Cancer Cells?

    Science.gov (United States)

    Kim, Kyu-Won; Lee, Su-Jae; Kim, Woo-Young; Seo, Ji Hae; Lee, Ho-Young

    2017-01-01

    Since molecular biology studies began, researches in biological science have centered on proteins and genes at molecular level of a single cell. Cancer research has also focused on various functions of proteins and genes that distinguish cancer cells from normal cells. Accordingly, most contemporary anticancer drugs have been developed to target abnormal characteristics of cancer cells. Despite the great advances in the development of anticancer drugs, vast majority of patients with advanced cancer have shown grim prognosis and high rate of relapse. To resolve this problem, we must reevaluate our focuses in current cancer research. Cancer should be considered as a systemic disease because cancer cells undergo a complex interaction with various surrounding cells in cancer tissue and spread to whole body through metastasis under the control of the systemic modulation. Human body relies on the cooperative interaction between various tissues and organs, and each organ performs its specialized function through tissue-specific cell networks. Therefore, investigation of the tumor-specific cell networks can provide novel strategy to overcome the limitation of current cancer research. This review presents the limitations of the current cancer research, emphasizing the necessity of studying tissue-specific cell network which could be a new perspective on treating cancer disease, not cancer cells.

  17. Invasive cancer cells and metastasis

    Science.gov (United States)

    Mierke, Claudia Tanja

    2013-12-01

    The physics of cancer is a relatively new emerging field of cancer research. In the last decade it has become a focus of biophysical research as well as becoming a novel focus for classical cancer research. This special section of Physical Biology focusing on invasive cancer cells and metastasis (physical oncology) will give greater insight into the different subfields where physical approaches are being applied to cancer research. This focus on the physical aspects of cancer is necessary because novel approaches in the field of genomics and proteomics have not altered the field of cancer research dramatically, due to the fact that few breakthroughs have been made. It is still not understood why some primary tumors metastasize and thus have a worse outcome compared to others that do not metastasize. As biophysicists, we and others suggest that the mechanical properties of the cancer cells, which possess the ability to transmigrate, are quite different compared to non-metastatic and non-invasive cancer cells. Furthermore, we hypothesize that these cancer cells undergo a selection process within the primary tumor that enables them to weaken their cell-cell adhesions and to alter their cell-matrix adhesions in order to be able to cross the outermost boundary of the primary tumor, as well as the surrounding basement membrane, and to invade the connective tissue. This prerequisite may also help the cancer cells to enter blood or lymph vessels, get transported with the vessel flow and form secondary tumors either within the vessel, directly on the endothelium, or in a different organ after crossing the endothelial lining a second time. This special section begins with a paper by Mark F Coughlin and Jeffrey J Fredberg on the changes in cytoskeletal dynamics and nonlinear rheology due to the metastatic capability of cancer cells from different cancer tissue types such as skin, bladder, prostate and kidney [1]. The hypothesis was that the metastatic outcome is impacted by

  18. Adoptive T cell cancer therapy

    Science.gov (United States)

    Dzhandzhugazyan, Karine N.; Guldberg, Per; Kirkin, Alexei F.

    2018-06-01

    Tumour heterogeneity and off-target toxicity are current challenges of cancer immunotherapy. Karine Dzhandzhugazyan, Per Guldberg and Alexei Kirkin discuss how epigenetic induction of tumour antigens in antigen-presenting cells may form the basis for multi-target therapies.

  19. Transfection in Primary Cultured Neuronal Cells.

    Science.gov (United States)

    Marwick, Katie F M; Hardingham, Giles E

    2017-01-01

    Transfection allows the introduction of foreign nucleic acid into eukaryotic cells. It is an important tool in understanding the roles of NMDARs in neurons. Here, we describe using lipofection-mediated transfection to introduce cDNA encoding NMDAR subunits into postmitotic rodent primary cortical neurons maintained in culture.

  20. Cell Culture Microfluidic Biochips: Experimental Throughput Maximization

    DEFF Research Database (Denmark)

    Minhass, Wajid Hassan; Pop, Paul; Madsen, Jan

    2011-01-01

    Microfluidic biochips offer a promising alternative to a conventional biochemical laboratory, integrating all necessary functionalities on-chip in order to perform biochemical applications. Researchers have started to propose computer-aided design tools for the synthesis of such biochips. Our focus...... metaheuristic for experimental design generation for the cell culture microfluidic biochips, and we have evaluated our approach using multiple experimental setups....

  1. Plant Cell Culture Initiation: practical tips

    NARCIS (Netherlands)

    Hall, R.D.

    2001-01-01

    The use of cultured plant cells in either organized or unorganized form has increased vey considerably in the last 10-15 yr. Many new technologies have been developed and applications in both fundamental and applied research have led to the development of some powerful tools for improving our

  2. Cell culture from sponges: pluripotency and immortality

    NARCIS (Netherlands)

    Caralt Bosch, de S.; Uriz, M.J.; Wijffels, R.H.

    2007-01-01

    Sponges are a source of compounds with potential pharmaceutical applications. In this article, methods of sponge cell culture for production of these bioactive compounds are reviewed, and new approaches for overcoming the problem of metabolite supply are examined. The use of embryos is proposed as a

  3. Nanotechnology, Cell Culture and Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Kazutoshi Haraguchi

    2011-01-01

    Full Text Available We have fabricated new types of polymer hydrogels and polymer nanocomposites, i.e., nanocomposite gels (NC gels and soft, polymer nanocomposites (M-NCs: solid, with novel organic/inorganic network structures. Both NC gels and M-NCs were synthesized by in-situ free-radical polymerization in the presence of exfoliated clay platelets in aqueous systems and were obtained in various forms such as film, sheet, tube, coating, etc. and sizes with a wide range of clay contents. Here, disk-like inorganic clay nanoparticles act as multi-functional crosslinkers to form new types of network systems. Both NC gels and M-NCs have extraordinary optical and mechanical properties including ultra-high reversible extensibility, as well as a number of new characteristics relating to optical anisotropy, polymer/clay morphology, biocompatibility, stimuli-sensitive surfaces, micro-patterning, etc. For examples, the biological testing of medical devices, comprised of a sensitization test, an irritation test, an intracutaneous test and an in vitro cytotoxicity test,was carried out for NC gels and M-NCs. The safety of NC gels and M-NCs was confirmed in all tests. Also, the interaction of living tissue with NC gel was investigated in vivo by implantation in live goats; neither inflammation nor concrescence occurred around the NC gels. Furthermore, it was found that both N-NC gels consisting of poly(N-isopropylacrylamide(PNIPA/clay network and M-NCs consisting of poly(2-methoxyethyacrylate(PMEA/clay network show characteristic cell culture and subsequent cell detachment on their surfaces, although it was almost impossible to culture cells on conventional, chemically-crosslinked PNIPA hydrogels and chemically crossslinked PMEA, regardless of their crosslinker concentration. Various kinds of cells, such ashumanhepatoma cells (HepG2, normal human dermal fibroblast (NHDF, and human umbilical vein endothelial cells (HUVEC, could be cultured to be confluent on the surfaces of N

  4. Gastric stem cells and gastric cancer stem cells

    OpenAIRE

    Han, Myoung-Eun; Oh, Sae-Ock

    2013-01-01

    The gastric epithelium is continuously regenerated by gastric stem cells, which give rise to various kinds of daughter cells, including parietal cells, chief cells, surface mucous cells, mucous neck cells, and enteroendocrine cells. The self-renewal and differentiation of gastric stem cells need delicate regulation to maintain the normal physiology of the stomach. Recently, it was hypothesized that cancer stem cells drive the cancer growth and metastasis. In contrast to conventional clonal ev...

  5. Altered characteristics of cancer stem/initiating cells in a breast cancer cell line treated with persistent 5-FU chemotherapy

    OpenAIRE

    LÜ, XINQUAN; DENG, QING; LI, HUIXIANG; SUO, ZHENHE

    2011-01-01

    Drug resistance of cancer stem/initiating cells has been considered to be one of the main reasons for tumor relapse. However, knowledge concerning the changes in stem/ initiating cells during chemotherapy is limited. In the present study, the breast cancer cell line MDA-MB-468 was cultured with 5-fluorouracil and serially passaged. Six cell generations were collected. Semi-quantitative RT-PCR and flow cytometric techniques were used to evaluate the protein and mRNA expression of stem/initiati...

  6. Cell Culture Assay for Human Noroviruses [response

    Energy Technology Data Exchange (ETDEWEB)

    Straub, Tim M.; Honer Zu Bentrup, Kerstin; Orosz Coghlan, Patricia; Dohnalkova, Alice; Mayer, Brooke K.; Bartholomew, Rachel A.; Valdez, Catherine O.; Bruckner-Lea, Cindy J.; Gerba, Charles P.; Abbaszadegan, Morteza A.; Nickerson, Cheryl A.

    2007-07-01

    We appreciate the comments provided by Leung et al., in response to our recently published article “In Vitro Cell Culture Infectivity Assay for Human Noroviruses” by Straub et al. (1). The specific aim of our project was to develop an in vitro cell culture infectivity assay for human noroviruses (hNoV) to enhance risk assessments when they are detected in water supplies. Reverse transcription (RT) qualitative or quantitative PCR are the primary assays for waterborne NoV monitoring. However, these assays cannot distinguish between infectious vs. non-infectious virions. When hNoV is detected in water supplies, information provided by our infectivity assay will significantly improve risk assessment models and protect human health, regardless of whether we are propagating NoV. Indeed, in vitro cell culture infectivity assays for the waterborne pathogen Cryptosporidium parvum that supplement approved fluorescent microscopy assays, do not result in amplification of the environmentally resistant hard-walled oocysts (2). However, identification of life cycle stages in cell culture provides evidence of infectious oocysts in a water supply. Nonetheless, Leung et al.’s assertion regarding the suitability of our method for the in vitro propagation of high titers of NoV is valid for the medical research community. In this case, well-characterized challenge pools of virus would be useful for developing and testing diagnostics, therapeutics, and vaccines. As further validation of our published findings, we have now optimized RT quantitative PCR to assess the level of viral production in cell culture, where we are indeed finding significant increases in viral titer. The magnitude and time course of these increases is dependent on both virus strain and multiplicity of infection. We are currently preparing a manuscript that will discuss these findings in greater detail, and the implications this may have for creating viral challenge pools

  7. Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells

    Science.gov (United States)

    2017-10-01

    AWARD NUMBER: W81XWH-15-1-0644 TITLE: Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells PRINCIPAL INVESTIGATOR: Chun-Ju...Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-15-1-0644 5c. PROGRAM ELEMENT...Unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT Cancer stem cells (CSCs), a cell population with acquired perpetuating self-renewal properties which

  8. Prostate Cancer Stem-Like Cells | Center for Cancer Research

    Science.gov (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

  9. Cell Culture in Microgravity: Opening the Door to Space Cell Biology

    Science.gov (United States)

    Pellis, Neal R.; Dawson, David L. (Technical Monitor)

    1999-01-01

    Adaptational response of human cell populations to microgravity is investigated using simulation, short-term Shuttle experiments, and long-term microgravity. Simulation consists of a clinostatically-rotated cell culture system. The system is a horizontally-rotated cylinder completely filled with culture medium. Low speed rotation results in continuous-fall of the cells through the fluid medium. In this setting, cells: 1) aggregate, 2) propagate in three dimensions, 3) synthesize matrix, 4) differentiate, and 5) form sinusoids that facilitate mass transfer. Space cell culture is conducted in flight bioreactors and in static incubators. Cells grown in microgravity are: bovine cartilage, promyelocytic leukemia, kidney proximal tubule cells, adrenal medulla, breast and colon cancer, and endothelium. Cells were cultured in space to test specific hypotheses. Cartilage cells were used to determine structural differences in cartilage grown in space compared to ground-based bioreactors. Results from a 130-day experiment on Mir revealed that cartilage grown in space was substantially more compressible due to insufficient glycosaminoglycan in the matrix. Interestingly, earth-grown cartilage conformed better to the dimensions of the scaffolding material, while the Mir specimens were spherical. The other cell populations are currently being analyzed for cell surface properties, gene expression, and differentiation. Results suggest that some cells spontaneously differentiate in microgravity. Additionally, vast changes in gene expression may occur in response to microgravity. In conclusion, the transition to microgravity may constitute a physical perturbation in cells resulting in unique gene expressions, the consequences of which may be useful in tissue engineering, disease modeling, and space cell biology.

  10. Glutathione in Cancer Cell Death

    Energy Technology Data Exchange (ETDEWEB)

    Ortega, Angel L. [Department of Physiology, Faculty of Medicine and Odontology, University of Valencia, 17 Av. Blasco Ibanez, 46010 Valencia (Spain); Mena, Salvador [Green Molecular SL, Pol. Ind. La Coma-Parc Cientific, 46190 Paterna, Valencia (Spain); Estrela, Jose M., E-mail: jose.m.estrela@uv.es [Department of Physiology, Faculty of Medicine and Odontology, University of Valencia, 17 Av. Blasco Ibanez, 46010 Valencia (Spain)

    2011-03-11

    Glutathione (L-γ-glutamyl-L-cysteinyl-glycine; GSH) in cancer cells is particularly relevant in the regulation of carcinogenic mechanisms; sensitivity against cytotoxic drugs, ionizing radiations, and some cytokines; DNA synthesis; and cell proliferation and death. The intracellular thiol redox state (controlled by GSH) is one of the endogenous effectors involved in regulating the mitochondrial permeability transition pore complex and, in consequence, thiol oxidation can be a causal factor in the mitochondrion-based mechanism that leads to cell death. Nevertheless GSH depletion is a common feature not only of apoptosis but also of other types of cell death. Indeed rates of GSH synthesis and fluxes regulate its levels in cellular compartments, and potentially influence switches among different mechanisms of death. How changes in gene expression, post-translational modifications of proteins, and signaling cascades are implicated will be discussed. Furthermore, this review will finally analyze whether GSH depletion may facilitate cancer cell death under in vivo conditions, and how this can be applied to cancer therapy.

  11. Use of an adaptable cell culture kit for performing lymphocyte and monocyte cell cultures in microgravity

    Science.gov (United States)

    Hatton, J. P.; Lewis, M. L.; Roquefeuil, S. B.; Chaput, D.; Cazenave, J. P.; Schmitt, D. A.

    1998-01-01

    The results of experiments performed in recent years on board facilities such as the Space Shuttle/Spacelab have demonstrated that many cell systems, ranging from simple bacteria to mammalian cells, are sensitive to the microgravity environment, suggesting gravity affects fundamental cellular processes. However, performing well-controlled experiments aboard spacecraft offers unique challenges to the cell biologist. Although systems such as the European 'Biorack' provide generic experiment facilities including an incubator, on-board 1-g reference centrifuge, and contained area for manipulations, the experimenter must still establish a system for performing cell culture experiments that is compatible with the constraints of spaceflight. Two different cell culture kits developed by the French Space Agency, CNES, were recently used to perform a series of experiments during four flights of the 'Biorack' facility aboard the Space Shuttle. The first unit, Generic Cell Activation Kit 1 (GCAK-1), contains six separate culture units per cassette, each consisting of a culture chamber, activator chamber, filtration system (permitting separation of cells from supernatant in-flight), injection port, and supernatant collection chamber. The second unit (GCAK-2) also contains six separate culture units, including a culture, activator, and fixation chambers. Both hardware units permit relatively complex cell culture manipulations without extensive use of spacecraft resources (crew time, volume, mass, power), or the need for excessive safety measures. Possible operations include stimulation of cultures with activators, separation of cells from supernatant, fixation/lysis, manipulation of radiolabelled reagents, and medium exchange. Investigations performed aboard the Space Shuttle in six different experiments used Jurkat, purified T-cells or U937 cells, the results of which are reported separately. We report here the behaviour of Jurkat and U937 cells in the GCAK hardware in ground

  12. A biocompatible micro cell culture chamber (mu CCC) for the culturing and on-line monitoring of eukaryote cells

    DEFF Research Database (Denmark)

    Stangegaard, Michael; Petronis, Sarunas; Jørgensen, Anders Michael

    2006-01-01

    culture chip compared to cell culture flasks. The cell culture chip could without further modification support cell growth of two other cell lines. Light coming from the microscope lamp during optical recordings of the cells was the only external factor identified, that could have a negative effect...... on cell survival. Low grade light exposure was however compatible with optical recordings as well as cell viability. These results strongly indicate that a cell culture chip could be constructed that allowed for on-line optical recording of cellular events without affecting the cell culturing condition...

  13. A biocompatible micro cell culture chamber (microCCC) for the culturing and on-line monitoring of eukaryote cells

    DEFF Research Database (Denmark)

    Stangegaard, Michael; Petronis, Sarunas; Jørgensen, A M

    2006-01-01

    culture chip compared to cell culture flasks. The cell culture chip could without further modification support cell growth of two other cell lines. Light coming from the microscope lamp during optical recordings of the cells was the only external factor identified, that could have a negative effect...... on cell survival. Low grade light exposure was however compatible with optical recordings as well as cell viability. These results strongly indicate that a cell culture chip could be constructed that allowed for on-line optical recording of cellular events without affecting the cell culturing condition...

  14. Cancer stem cells and differentiation therapy.

    Science.gov (United States)

    Jin, Xiong; Jin, Xun; Kim, Hyunggee

    2017-10-01

    Cancer stem cells can generate tumors from only a small number of cells, whereas differentiated cancer cells cannot. The prominent feature of cancer stem cells is its ability to self-renew and differentiate into multiple types of cancer cells. Cancer stem cells have several distinct tumorigenic abilities, including stem cell signal transduction, tumorigenicity, metastasis, and resistance to anticancer drugs, which are regulated by genetic or epigenetic changes. Like normal adult stem cells involved in various developmental processes and tissue homeostasis, cancer stem cells maintain their self-renewal capacity by activating multiple stem cell signaling pathways and inhibiting differentiation signaling pathways during cancer initiation and progression. Recently, many studies have focused on targeting cancer stem cells to eradicate malignancies by regulating stem cell signaling pathways, and products of some of these strategies are in preclinical and clinical trials. In this review, we describe the crucial features of cancer stem cells related to tumor relapse and drug resistance, as well as the new therapeutic strategy to target cancer stem cells named "differentiation therapy."

  15. Lipoprotein receptors in cultured bovine endothelial cells

    International Nuclear Information System (INIS)

    Struempfer, A.E.M.

    1983-07-01

    In this study, receptors that may be involved in the uptake of low density lipoproteins (LDL) and low density lipoproteins which have been modified by acetylation (AcLDL), were characterized. Aortic epithelial cells were used and a cell culture system which closely resembled the in vivo monolayer was established. Endothelial cell and lipoprotein interactions were examined by incubating the cells with 125 l-labelled lipoproteins under various conditions. The receptor affinity of bovine aortic endothelial cells was higher for AcLDL than that for LDL. Competition studies demonstrated that there were two distinct receptors for LDL and AcLDL on the endothelial cells. AcLDL did not compete with LDL for the LDL receptor, and conversely LDL did not compete with AcLDL for the AcLDL receptor. The receptor activities for LDL and AcLDL were examined as a function of culture age. Whereas the LDL receptor could be regulated, the AcLDL receptor was not as susceptible to regulation. Upon exposing endothelial cells for 72 h to either LDL or AcLDL, it was found that the total amount of cellular cholesterol increased by about 50%. However, the increase of total cholesterol was largely in the form of free cholesterol. This is in contrast to macrophages, where the increase in total cholesterol upon exposure to AcLDL is largely in the form cholesteryl esters

  16. Cell in situ zymography: an in vitro cytotechnology for localization of enzyme activity in cell culture.

    Science.gov (United States)

    Chhabra, Aastha; Jaiswal, Astha; Malhotra, Umang; Kohli, Shrey; Rani, Vibha

    2012-09-01

    In situ zymography is a unique technique for detection and localization of enzyme-substrate interactions majorly in histological sections. Substrate with quenched fluorogenic molecule is incorporated in gel over which tissue sections are mounted and then incubated in buffer. The enzymatic activity is observed in the form of fluorescent signal. With the advancements in the field of biological research, use of in vitro cell culture has become very popular and holds great significance in multiple fields including inflammation, cancer, stem cell biology and the still emerging 3-D cell cultures. The information on analysis of enzymatic activity in cell lines is inadequate presently. We propose a single-step methodology that is simple, sensitive, cost-effective, and functional to perform and study the 'in position' activity of enzyme on substrate for in vitro cell cultures. Quantification of enzymatic activity to carry out comparative studies on cells has also been illustrated. This technique can be applied to a variety of enzyme classes including proteases, amylases, xylanases, and cellulases in cell cultures.

  17. Dynamic cell culture system (7-IML-1)

    Science.gov (United States)

    Cogoli, Augusto

    1992-01-01

    This experiment is one of the Biorack experiments being flown on the International Microgravity Laboratory 1 (MIL-1) mission as part of an investigation studying cell proliferation and performance in space. One of the objectives of this investigation is to assess the potential benefits of bioprocessing in space with the ultimate goal of developing a bioreactor for continuous cell cultures in space. This experiment will test the operation of an automated culture chamber that was designed for use in a Bioreactor in space. The device to be tested is called the Dynamic Cell Culture System (DCCS). It is a simple device in which media are renewed or chemicals are injected automatically, by means of osmotic pumps. This experiment uses four Type I/O experiment containers. One DCCS unit, which contains a culture chamber with renewal of medium and a second chamber without a medium supply fits in each container. Two DCCS units are maintained under zero gravity conditions during the on-orbit period. The other two units are maintained under 1 gh conditions in a 1 g centrifuge. The schedule for incubator transfer is given.

  18. Identification and characterization of cells with cancer stem cell properties in human primary lung cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Ping Wang

    Full Text Available Lung cancer (LC with its different subtypes is generally known as a therapy resistant cancer with the highest morbidity rate worldwide. Therapy resistance of a tumor is thought to be related to cancer stem cells (CSCs within the tumors. There have been indications that the lung cancer is propagated and maintained by a small population of CSCs. To study this question we established a panel of 15 primary lung cancer cell lines (PLCCLs from 20 fresh primary tumors using a robust serum-free culture system. We subsequently focused on identification of lung CSCs by studying these cell lines derived from 4 representative lung cancer subtypes such as small cell lung cancer (SCLC, large cell carcinoma (LCC, squamous cell carcinoma (SCC and adenocarcinoma (AC. We identified a small population of cells strongly positive for CD44 (CD44(high and a main population which was either weakly positive or negative for CD44 (CD44(low/-. Co-expression of CD90 further narrowed down the putative stem cell population in PLCCLs from SCLC and LCC as spheroid-forming cells were mainly found within the CD44(highCD90(+ sub-population. Moreover, these CD44(highCD90(+ cells revealed mesenchymal morphology, increased expression of mesenchymal markers N-Cadherin and Vimentin, increased mRNA levels of the embryonic stem cell related genes Nanog and Oct4 and increased resistance to irradiation compared to other sub-populations studied, suggesting the CD44(highCD90(+ population a good candidate for the lung CSCs. Both CD44(highCD90(+ and CD44(highCD90(- cells in the PLCCL derived from SCC formed spheroids, whereas the CD44(low/- cells were lacking this potential. These results indicate that CD44(highCD90(+ sub-population may represent CSCs in SCLC and LCC, whereas in SCC lung cancer subtype, CSC potentials were found within the CD44(high sub-population.

  19. Identification and Characterization of Cells with Cancer Stem Cell Properties in Human Primary Lung Cancer Cell Lines

    Science.gov (United States)

    Suo, Zhenhe; Munthe, Else; Solberg, Steinar; Ma, Liwei; Wang, Mengyu; Westerdaal, Nomdo Anton Christiaan; Kvalheim, Gunnar; Gaudernack, Gustav

    2013-01-01

    Lung cancer (LC) with its different subtypes is generally known as a therapy resistant cancer with the highest morbidity rate worldwide. Therapy resistance of a tumor is thought to be related to cancer stem cells (CSCs) within the tumors. There have been indications that the lung cancer is propagated and maintained by a small population of CSCs. To study this question we established a panel of 15 primary lung cancer cell lines (PLCCLs) from 20 fresh primary tumors using a robust serum-free culture system. We subsequently focused on identification of lung CSCs by studying these cell lines derived from 4 representative lung cancer subtypes such as small cell lung cancer (SCLC), large cell carcinoma (LCC), squamous cell carcinoma (SCC) and adenocarcinoma (AC). We identified a small population of cells strongly positive for CD44 (CD44high) and a main population which was either weakly positive or negative for CD44 (CD44low/−). Co-expression of CD90 further narrowed down the putative stem cell population in PLCCLs from SCLC and LCC as spheroid-forming cells were mainly found within the CD44highCD90+ sub-population. Moreover, these CD44highCD90+ cells revealed mesenchymal morphology, increased expression of mesenchymal markers N-Cadherin and Vimentin, increased mRNA levels of the embryonic stem cell related genes Nanog and Oct4 and increased resistance to irradiation compared to other sub-populations studied, suggesting the CD44highCD90+ population a good candidate for the lung CSCs. Both CD44highCD90+ and CD44highCD90− cells in the PLCCL derived from SCC formed spheroids, whereas the CD44low/− cells were lacking this potential. These results indicate that CD44highCD90+ sub-population may represent CSCs in SCLC and LCC, whereas in SCC lung cancer subtype, CSC potentials were found within the CD44high sub-population. PMID:23469181

  20. 21 CFR 864.2280 - Cultured animal and human cells.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cultured animal and human cells. 864.2280 Section... Cultured animal and human cells. (a) Identification. Cultured animal and human cells are in vitro cultivated cell lines from the tissue of humans or other animals which are used in various diagnostic...

  1. Breast Cancer Screening: Cultural Beliefs and Diverse Populations

    Science.gov (United States)

    Simon, Cassandra E.

    2006-01-01

    This article addresses the role of culture in breast cancer screening behavior among African American, American Indian/Alaskan Native, Asian American/Pacific Islander, and Hispanic/Latina women. It reviews cultural beliefs, attitudes, and knowledge and their relative influence on women's decisions regarding health tests. The article explores how…

  2. Application of Synthetic Polymeric Scaffolds in Breast Cancer 3D Tissue Cultures and Animal Tumor Models

    Directory of Open Access Journals (Sweden)

    Girdhari Rijal

    2017-01-01

    Full Text Available Preparation of three-dimensional (3D porous scaffolds from synthetic polymers is a challenge to most laboratories conducting biomedical research. Here, we present a handy and cost-effective method to fabricate polymeric hydrogel and porous scaffolds using poly(lactic-co-glycolic acid (PLGA or polycaprolactone (PCL. Breast cancer cells grown on 3D polymeric scaffolds exhibited distinct survival, morphology, and proliferation compared to those on 2D polymeric surfaces. Mammary epithelial cells cultured on PLGA- or PCL-coated slides expressed extracellular matrix (ECM proteins and their receptors. Estrogen receptor- (ER- positive T47D breast cancer cells are less sensitive to 4-hydroxytamoxifen (4-HT treatment when cultured on the 3D porous scaffolds than in 2D cultures. Finally, cancer cell-laden polymeric scaffolds support consistent tumor formation in animals and biomarker expression as seen in human native tumors. Our data suggest that the porous synthetic polymer scaffolds satisfy the basic requirements for 3D tissue cultures both in vitro and in vivo. The scaffolding technology has appealing potentials to be applied in anticancer drug screening for a better control of the progression of human cancers.

  3. Road for understanding cancer stem cells

    DEFF Research Database (Denmark)

    Serakinci, Nedime; Erzik, Can

    2007-01-01

    There is increasing evidence suggesting that stem cells are susceptive to carcinogenesis and, consequently, can be the origin of many cancers. Recently, the neoplastic potential of stem cells has been supported by many groups showing the existence of subpopulations with stem cell characteristics...... in tumor biopsies such as brain and breast. Evidence supporting the cancer stem cell hypothesis has gained impact due to progress in stem cell biology and development of new models to validate the self-renewal potential of stem cells. Recent evidence on the possible identification of cancer stem cells may...... offer an opportunity to use these cells as future therapeutic targets. Therefore, model systems in this field have become very important and useful. This review will focus on the state of knowledge on cancer stem cell research, including cell line models for cancer stem cells. The latter will, as models...

  4. Epigenetic modulation of cancer-germline antigen gene expression in tumorigenic human mesenchymal stem cells: implications for cancer therapy

    DEFF Research Database (Denmark)

    Gjerstorff, Morten; Burns, Jorge S; Nielsen, Ole

    2009-01-01

    Cancer-germline antigens are promising targets for cancer immunotherapy, but whether such therapies will also eliminate the primary tumor stem cell population remains undetermined. We previously showed that long-term cultures of telomerized adult human bone marrow mesenchymal stem cells can...... spontaneously evolve into tumor-initiating, mesenchymal stem cells (hMSC-TERT20), which have characteristics of clinical sarcoma cells. In this study, we used the hMSC-TERT20 tumor stem cell model to investigate the potential of cancer-germline antigens to serve as tumor stem cell targets. We found...... of cancer-germline antigens in hMSC-TERT20 cells, while their expression levels in primary human mesenchymal stem cells remained unaffected. The expression pattern of cancer-germline antigens in tumorigenic mesenchymal stem cells and sarcomas, plus their susceptibility to enhancement by epigenetic...

  5. Alginate foam-based three-dimensional culture to investigate drug sensitivity in primary leukaemia cells.

    Science.gov (United States)

    Karimpoor, Mahroo; Yebra-Fernandez, Eva; Parhizkar, Maryam; Orlu, Mine; Craig, Duncan; Khorashad, Jamshid S; Edirisinghe, Mohan

    2018-04-01

    The development of assays for evaluating the sensitivity of leukaemia cells to anti-cancer agents is becoming an important aspect of personalized medicine. Conventional cell cultures lack the three-dimensional (3D) structure of the bone marrow (BM), the extracellular matrix and stromal components which are crucial for the growth and survival of leukaemia stem cells. To accurately predict the sensitivity of the leukaemia cells in an in vitro assay a culturing system containing the essential components of BM is required. In this study, we developed a porous calcium alginate foam-based scaffold to be used for 3D culture. The new 3D culture was shown to be cell compatible as it supported the proliferation of both normal haematopoietic and leukaemia cells. Our cell differential assay for myeloid markers showed that the porous foam-based 3D culture enhanced myeloid differentiation in both leukaemia and normal haematopoietic cells compared to two-dimensional culture. The foam-based scaffold reduced the sensitivity of the leukaemia cells to the tested antileukaemia agents in K562 and HL60 leukaemia cell line model and also primary myeloid leukaemia cells. This observation supports the application of calcium alginate foams as scaffold components of the 3D cultures for investigation of sensitivity to antileukaemia agents in primary myeloid cells. © 2018 The Author(s).

  6. Comparison of submerged and unsubmerged printing of ovarian cancer cells.

    Science.gov (United States)

    Davidoff, Sherry N; Au, David; Smith, Samuel; Brooks, Amanda E; Brooks, Benjamin D

    2015-01-01

    A high-throughput cell based assay would greatly aid in the development and screening of ovarian cancer drug candidates. Previously, a three-dimensional microfluidic printer that is not only capable of controlling the location of cell deposition, but also of maintaining a liquid, nutrient rich environment to preserve cellular phenotype has been developed (Wasatch Microfluidics). In this study, we investigated the impact (i.e., viability, density, and phenotype) of depositing cells on a surface submerged in cell culture media. It was determined that submersion of the microfluidic print head in cell media did not alter the cell density, viability, or phenotype.. This article describes an in depth study detailing the impact of one of the fundamental components of a 3D microfluidic cell printer designed to mimic the in vivo cell environment. Development of such a tool holds promise as a high-throughput drug-screening platform for new cancer therapeutics.

  7. STAMP alters the growth of transformed and ovarian cancer cells

    International Nuclear Information System (INIS)

    He, Yuanzheng; Blackford, John A Jr; Kohn, Elise C; Simons, S Stoney Jr

    2010-01-01

    Steroid receptors play major roles in the development, differentiation, and homeostasis of normal and malignant tissue. STAMP is a novel coregulator that not only enhances the ability of p160 coactivator family members TIF2 and SRC-1 to increase gene induction by many of the classical steroid receptors but also modulates the potency (or EC 50 ) of agonists and the partial agonist activity of antisteroids. These modulatory activities of STAMP are not limited to gene induction but are also observed for receptor-mediated gene repression. However, a physiological role for STAMP remains unclear. The growth rate of HEK293 cells stably transfected with STAMP plasmid and overexpressing STAMP protein is found to be decreased. We therefore asked whether different STAMP levels might also contribute to the abnormal growth rates of cancer cells. Panels of different stage human cancers were screened for altered levels of STAMP mRNA. Those cancers with the greatest apparent changes in STAMP mRNA were pursued in cultured cancer cell lines. Higher levels of STAMP are shown to have the physiologically relevant function of reducing the growth of HEK293 cells but, unexpectedly, in a steroid-independent manner. STAMP expression was examined in eight human cancer panels. More extensive studies of ovarian cancers suggested the presence of higher levels of STAMP mRNA. Lowering STAMP mRNA levels with siRNAs alters the proliferation of several ovarian cancer tissue culture lines in a cell line-specific manner. This cell line-specific effect of STAMP is not unique and is also seen for the conventional effects of STAMP on glucocorticoid receptor-regulated gene transactivation. This study indicates that a physiological function of STAMP in several settings is to modify cell growth rates in a manner that can be independent of steroid hormones. Studies with eleven tissue culture cell lines of ovarian cancer revealed a cell line-dependent effect of reduced STAMP mRNA on cell growth rates. This

  8. Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

    Energy Technology Data Exchange (ETDEWEB)

    Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing, E-mail: caijingmmm@hotmail.com; Wang, Zehua, E-mail: zehuawang@163.net

    2015-09-10

    Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs.

  9. Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

    International Nuclear Information System (INIS)

    Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing; Wang, Zehua

    2015-01-01

    Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs

  10. A microfluidic cell culture array with various oxygen tensions.

    Science.gov (United States)

    Peng, Chien-Chung; Liao, Wei-Hao; Chen, Ying-Hua; Wu, Chueh-Yu; Tung, Yi-Chung

    2013-08-21

    Oxygen tension plays an important role in regulating various cellular functions in both normal physiology and disease states. Therefore, drug testing using conventional in vitro cell models under normoxia often possesses limited prediction capability. A traditional method of setting an oxygen tension in a liquid medium is by saturating it with a gas mixture at the desired level of oxygen, which requires bulky gas cylinders, sophisticated control, and tedious interconnections. Moreover, only a single oxygen tension can be tested at the same time. In this paper, we develop a microfluidic cell culture array platform capable of performing cell culture and drug testing under various oxygen tensions simultaneously. The device is fabricated using an elastomeric material, polydimethylsiloxane (PDMS) and the well-developed multi-layer soft lithography (MSL) technique. The prototype device has 4 × 4 wells, arranged in the same dimensions as a conventional 96-well plate, for cell culture. The oxygen tensions are controlled by spatially confined oxygen scavenging chemical reactions underneath the wells using microfluidics. The platform takes advantage of microfluidic phenomena while exhibiting the combinatorial diversities achieved by microarrays. Importantly, the platform is compatible with existing cell incubators and high-throughput instruments (liquid handling systems and plate readers) for cost-effective setup and straightforward operation. Utilizing the developed platform, we successfully perform drug testing using an anti-cancer drug, triapazamine (TPZ), on adenocarcinomic human alveolar basal epithelial cell line (A549) under three oxygen tensions ranging from 1.4% to normoxia. The developed platform is promising to provide a more meaningful in vitro cell model for various biomedical applications while maintaining desired high throughput capabilities.

  11. Mouse cell culture - Methods and protocols

    Directory of Open Access Journals (Sweden)

    CarloAlberto Redi

    2010-12-01

    Full Text Available The mouse is, out of any doubt, the experimental animal par excellence for many many colleagues within the scientific community, notably for those working in mammalian biology (in a broad sense, from basic genetic to modeling human diseases, starting at least from 1664 Robert Hooke experiments on air’s propertyn. Not surprising then that mouse cell cultures is a well established field of research itself and that there are several handbooks devoted to this discipline. Here, Andrew Ward and David Tosh provide a necessary update of the protocols currently needed. In fact, nearly half of the book is devoted to stem cells culture protocols, mainly embryonic, from a list of several organs (kidney, lung, oesophagus and intestine, pancreas and liver to mention some........

  12. Advantages and challenges of microfluidic cell culture in polydimethylsiloxane devices.

    Science.gov (United States)

    Halldorsson, Skarphedinn; Lucumi, Edinson; Gómez-Sjöberg, Rafael; Fleming, Ronan M T

    2015-01-15

    Culture of cells using various microfluidic devices is becoming more common within experimental cell biology. At the same time, a technological radiation of microfluidic cell culture device designs is currently in progress. Ultimately, the utility of microfluidic cell culture will be determined by its capacity to permit new insights into cellular function. Especially insights that would otherwise be difficult or impossible to obtain with macroscopic cell culture in traditional polystyrene dishes, flasks or well-plates. Many decades of heuristic optimization have gone into perfecting conventional cell culture devices and protocols. In comparison, even for the most commonly used microfluidic cell culture devices, such as those fabricated from polydimethylsiloxane (PDMS), collective understanding of the differences in cellular behavior between microfluidic and macroscopic culture is still developing. Moving in vitro culture from macroscopic culture to PDMS based devices can come with unforeseen challenges. Changes in device material, surface coating, cell number per unit surface area or per unit media volume may all affect the outcome of otherwise standard protocols. In this review, we outline some of the advantages and challenges that may accompany a transition from macroscopic to microfluidic cell culture. We focus on decisive factors that distinguish macroscopic from microfluidic cell culture to encourage a reconsideration of how macroscopic cell culture principles might apply to microfluidic cell culture. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  13. Investigation of cancer cell behavior on nanofibrous scaffolds

    Energy Technology Data Exchange (ETDEWEB)

    Szot, Christopher S.; Buchanan, Cara F. [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 (United States); Gatenholm, Paul [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 (United States); Department of Chemical and Biological Engineering, Chalmers University of Technology, SE-412 96 Goeteborg (Sweden); Rylander, Marissa Nichole [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 (United States); Freeman, Joseph W., E-mail: jwfreeman@vt.edu [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 (United States)

    2011-01-01

    Tissue engineering and the use of nanofibrous biomaterial scaffolds offer a unique perspective for studying cancer development in vitro. Current in vitro models of tumorigenesis are limited by the use of static, two-dimensional (2D) cell culture monolayers that lack the structural architecture necessary for cell-cell interaction and three-dimensional (3D) scaffolds that are too simplistic for studying basic pathological mechanisms. In this study, two nanofibrous biomaterials that mimic the structure of the extracellular matrix, bacterial cellulose and electrospun polycaprolactone (PCL)/collagen I, were investigated as potential 3D scaffolds for an in vitro cancer model. Multiple cancer cell lines were cultured on each scaffold material and monitored for cell viability, proliferation, adhesion, infiltration, and morphology. Both bacterial cellulose and electrospun PCL/collagen I, which have nano-scale structures on the order of 100-500 nm, have been used in many diverse tissue engineering applications. Cancer cell adhesion and growth were limited on bacterial cellulose, while all cellular processes were enhanced on the electrospun scaffolds. This initial analysis has demonstrated the potential of electrospun PCL/collagen I scaffolds toward the development of an improved 3D in vitro cancer model.

  14. Differentiation of mammalian skeletal muscle cells cultured on microcarrier beads in a rotating cell culture system

    Science.gov (United States)

    Torgan, C. E.; Burge, S. S.; Collinsworth, A. M.; Truskey, G. A.; Kraus, W. E.

    2000-01-01

    The growth and repair of adult skeletal muscle are due in part to activation of muscle precursor cells, commonly known as satellite cells or myoblasts. These cells are responsive to a variety of environmental cues, including mechanical stimuli. The overall goal of the research is to examine the role of mechanical signalling mechanisms in muscle growth and plasticity through utilisation of cell culture systems where other potential signalling pathways (i.e. chemical and electrical stimuli) are controlled. To explore the effects of decreased mechanical loading on muscle differentiation, mammalian myoblasts are cultured in a bioreactor (rotating cell culture system), a model that has been utilised to simulate microgravity. C2C12 murine myoblasts are cultured on microcarrier beads in a bioreactor and followed throughout differentiation as they form a network of multinucleated myotubes. In comparison with three-dimensional control cultures that consist of myoblasts cultured on microcarrier beads in teflon bags, myoblasts cultured in the bioreactor exhibit an attenuation in differentiation. This is demonstrated by reduced immunohistochemical staining for myogenin and alpha-actinin. Western analysis shows a decrease, in bioreactor cultures compared with control cultures, in levels of the contractile proteins myosin (47% decrease, p < 0.01) and tropomyosin (63% decrease, p < 0.01). Hydrodynamic measurements indicate that the decrease in differentiation may be due, at least in part, to fluid stresses acting on the myotubes. In addition, constraints on aggregate size imposed by the action of fluid forces in the bioreactor affect differentiation. These results may have implications for muscle growth and repair during spaceflight.

  15. Increased hydrostatic pressure enhances motility of lung cancer cells.

    Science.gov (United States)

    Kao, Yu-Chiu; Lee, Chau-Hwang; Kuo, Po-Ling

    2014-01-01

    Interstitial fluid pressures within most solid tumors are significantly higher than that in the surrounding normal tissues. Therefore, cancer cells must proliferate and migrate under the influence of elevated hydrostatic pressure while a tumor grows. In this study, we developed a pressurized cell culture device and investigated the influence of hydrostatic pressure on the migration speeds of lung cancer cells (CL1-5 and A549). The migration speeds of lung cancer cells were increased by 50-60% under a 20 mmHg hydrostatic pressure. We also observed that the expressions of aquaporin in CL1-5 and A549 cells were increased under the hydrostatic pressure. Our preliminary results indicate that increased hydrostatic pressure plays an important role in tumor metastasis.

  16. A POX on Renal Cancer Cells | Center for Cancer Research

    Science.gov (United States)

    Proline oxidase, or POX, is an enzyme responsible for metabolizing the amino acid proline. POX contributes to the regulation of cell death that occurs when cellular systems malfunction, a process called apoptosis. Previous studies have determined that levels of POX are reduced in several types of human cancer. Likewise, many cancer cells become resistant to apoptosis, suggesting a link between POX and cancer cell survival.

  17. In silico characterization of cell-cell interactions using a cellular automata model of cell culture.

    Science.gov (United States)

    Kihara, Takanori; Kashitani, Kosuke; Miyake, Jun

    2017-07-14

    Cell proliferation is a key characteristic of eukaryotic cells. During cell proliferation, cells interact with each other. In this study, we developed a cellular automata model to estimate cell-cell interactions using experimentally obtained images of cultured cells. We used four types of cells; HeLa cells, human osteosarcoma (HOS) cells, rat mesenchymal stem cells (MSCs), and rat smooth muscle A7r5 cells. These cells were cultured and stained daily. The obtained cell images were binarized and clipped into squares containing about 10 4 cells. These cells showed characteristic cell proliferation patterns. The growth curves of these cells were generated from the cell proliferation images and we determined the doubling time of these cells from the growth curves. We developed a simple cellular automata system with an easily accessible graphical user interface. This system has five variable parameters, namely, initial cell number, doubling time, motility, cell-cell adhesion, and cell-cell contact inhibition (of proliferation). Within these parameters, we obtained initial cell numbers and doubling times experimentally. We set the motility at a constant value because the effect of the parameter for our simulation was restricted. Therefore, we simulated cell proliferation behavior with cell-cell adhesion and cell-cell contact inhibition as variables. By comparing growth curves and proliferation cell images, we succeeded in determining the cell-cell interaction properties of each cell. Simulated HeLa and HOS cells exhibited low cell-cell adhesion and weak cell-cell contact inhibition. Simulated MSCs exhibited high cell-cell adhesion and positive cell-cell contact inhibition. Simulated A7r5 cells exhibited low cell-cell adhesion and strong cell-cell contact inhibition. These simulated results correlated with the experimental growth curves and proliferation images. Our simulation approach is an easy method for evaluating the cell-cell interaction properties of cells.

  18. Metabolic characterization of invaded cells of the pancreatic cancer cell line, PANC-1.

    Science.gov (United States)

    Fujita, Mayumi; Imadome, Kaori; Imai, Takashi

    2017-05-01

    We previously reported that about 0.4% of cells in the cultured human pancreatic cancer cell line, PANC-1, can invade matrigel during the transwell invasion assay, suggesting that these invaded PANC-1 cells may have specific characteristics to keep their invasive potential. To identify the metabolic characterization specific in the invaded PANC-1 cells, metabolome analysis of the invaded PANC-1 compared with the whole cultured PANC-1 was performed using CE-TOFMS, and concentrations of 110 metabolites were measured. In contrast to the whole cultured cells, the invaded PANC-1 was characterized as a population with reduced levels of amino acids and TCA cycle intermediates, and decreased and increased intermediates in glycolysis and nucleic acid metabolism. In particular, the ratio of both adenosine and guanosine energy charge was reduced in the invaded cells, revealing that the consumption of ATP and GTP was high in the invaded cells, and thus suggesting that ATP- or GTP-generating pathways are stimulated. In addition, the GSH/GSSG ratio was low in the invaded cells, but these cells had a higher surviving fraction after exposure to hydrogen peroxide. Thus, the invaded cells were the population resistant to oxidative stress. Furthermore, reduction in intracellular GSH content inhibited PANC-1 invasiveness, indicated that GSH has an important role in PANC-1 invasiveness. Overall, we propose the invaded cells have several unique metabolic profiles. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  19. Pericellular oxygen monitoring with integrated sensor chips for reproducible cell culture experiments.

    Science.gov (United States)

    Kieninger, J; Aravindalochanan, K; Sandvik, J A; Pettersen, E O; Urban, G A

    2014-04-01

    Here we present an application, in two tumour cell lines, based on the Sensing Cell Culture Flask system as a cell culture monitoring tool for pericellular oxygen sensing. T-47D (human breast cancer) and T98G (human brain cancer) cells were cultured either in atmospheric air or in a glove-box set at 4% oxygen, in both cases with 5% CO2 in the gas phase. Pericellular oxygen tension was measured with the help of an integrated sensor chip comprising oxygen sensor arrays. Obtained results illustrate variation of pericellular oxygen tension in attached cells covered by stagnant medium. Independent of incubation conditions, low pericellular oxygen concentration levels, usually associated with hypoxia, were found in dense cell cultures. Respiration alone brought pericellular oxygen concentration down to levels which could activate hypoxia-sensing regulatory processes in cultures believed to be aerobic. Cells in culture believed to experience conditions of mild hypoxia may, in reality, experience severe hypoxia. This would lead to incorrect assumptions and suggests that pericellular oxygen concentration readings are of great importance to obtain reproducible results when dealing with hypoxic and normoxic (aerobic) incubation conditions. The Sensing Cell Culture Flask system allows continuous monitoring of pericellular oxygen concentration with outstanding long-term stability and no need for recalibration during cell culture experiments. The sensor is integrated into the flask bottom, thus in direct contact with attached cells. No additional equipment needs to be inserted into the flask during culturing. Transparency of the electrochemical sensor chip allows optical inspection of cells attached on top of the sensor. © 2014 John Wiley & Sons Ltd.

  20. Extinction models for cancer stem cell therapy

    Science.gov (United States)

    Sehl, Mary; Zhou, Hua; Sinsheimer, Janet S.; Lange, Kenneth L.

    2012-01-01

    Cells with stem cell-like properties are now viewed as initiating and sustaining many cancers. This suggests that cancer can be cured by driving these cancer stem cells to extinction. The problem with this strategy is that ordinary stem cells are apt to be killed in the process. This paper sets bounds on the killing differential (difference between death rates of cancer stem cells and normal stem cells) that must exist for the survival of an adequate number of normal stem cells. Our main tools are birth–death Markov chains in continuous time. In this framework, we investigate the extinction times of cancer stem cells and normal stem cells. Application of extreme value theory from mathematical statistics yields an accurate asymptotic distribution and corresponding moments for both extinction times. We compare these distributions for the two cell populations as a function of the killing rates. Perhaps a more telling comparison involves the number of normal stem cells NH at the extinction time of the cancer stem cells. Conditioning on the asymptotic time to extinction of the cancer stem cells allows us to calculate the asymptotic mean and variance of NH. The full distribution of NH can be retrieved by the finite Fourier transform and, in some parameter regimes, by an eigenfunction expansion. Finally, we discuss the impact of quiescence (the resting state) on stem cell dynamics. Quiescence can act as a sanctuary for cancer stem cells and imperils the proposed therapy. We approach the complication of quiescence via multitype branching process models and stochastic simulation. Improvements to the τ-leaping method of stochastic simulation make it a versatile tool in this context. We conclude that the proposed therapy must target quiescent cancer stem cells as well as actively dividing cancer stem cells. The current cancer models demonstrate the virtue of attacking the same quantitative questions from a variety of modeling, mathematical, and computational perspectives

  1. Targetless T cells in cancer immunotherapy

    DEFF Research Database (Denmark)

    thor Straten, Eivind Per; Garrido, Federico

    2016-01-01

    Attention has recently focused on new cancer immunotherapy protocols aiming to activate T cell mediated anti-tumor responses. To this end, administration of antibodies that target inhibitory molecules regulating T-cell cytotoxicity has achieved impressive clinical responses, as has adoptive cell...... infiltrate tumor tissues and destroy HLA class I positive tumor cells expressing the specific antigen. In fact, current progress in the field of cancer immune therapy is based on the capacity of T cells to kill cancer cells that present tumor antigen in the context on an HLA class I molecule. However......, it is also well established that cancer cells are often characterized by loss or down regulation of HLA class I molecules, documented in a variety of human tumors. Consequently, immune therapy building on CD8 T cells will be futile in patients harboring HLA class-I negative or deficient cancer cells...

  2. Evaluation of Stem Cell Markers, CD44/CD24 in Breast Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Masoud Hashemi Arabi

    2014-05-01

    Four breast cancer cell lines, MCF-7 ، T47D ، MDA-MB231 and MDA-MB468 were purchased from National cell Bank of Iran based in Iran Pasture Institute and were cultured in high glucose DMEM supplemented with 10% FCS. Cells were stained with antiCD44-PE and antiCD24-FITC antibodies and Status of CD44 and CD24 as markers of breast cancer stem cells were evaluated using flow cytometer and fluorescent microscopy.Evaluation of CD44 and CD24 as markers of breast cancer stem cells showed that MDA-MB231 with 97±1.2% CD44+/CD24-/low cells is significantly different from the others that they were mainly CD44 and CD24 positive cells(p

  3. C-C motif ligand 5 promotes migration of prostate cancer cells in the prostate cancer bone metastasis microenvironment.

    Science.gov (United States)

    Urata, Satoko; Izumi, Kouji; Hiratsuka, Kaoru; Maolake, Aerken; Natsagdorj, Ariunbold; Shigehara, Kazuyoshi; Iwamoto, Hiroaki; Kadomoto, Suguru; Makino, Tomoyuki; Naito, Renato; Kadono, Yoshifumi; Lin, Wen-Jye; Wufuer, Guzailinuer; Narimoto, Kazutaka; Mizokami, Atsushi

    2018-03-01

    Chemokines and their receptors have key roles in cancer progression. The present study investigated chemokine activity in the prostate cancer bone metastasis microenvironment. Growth and migration of human prostate cancer cells were assayed in cocultures with bone stromal cells. The migration of LNCaP cells significantly increased when co-cultured with bone stromal cells isolated from prostate cancer bone metastases. Cytokine array analysis of conditioned medium from bone stromal cell cultures identified CCL5 as a concentration-dependent promoter of LNCaP cell migration. The migration of LNCaP cells was suppressed when C-C motif ligand 5 (CCL5) neutralizing antibody was added to cocultures with bone stromal cells. Knockdown of androgen receptor with small interfering RNA increased the migration of LNCaP cells compared with control cells, and CCL5 did not promote the migration of androgen receptor knockdown LNCaP. Elevated CCL5 secretion in bone stromal cells from metastatic lesions induced prostate cancer cell migration by a mechanism consistent with CCL5 activity upstream of androgen receptor signaling. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  4. A novel role of EMMPRIN/CD147 in transformation of quiescent fibroblasts to cancer-associated fibroblasts by breast cancer cells

    Science.gov (United States)

    Xu, Jing; Lu, Yang; Qiu, Songbo; Chen, Zhi-Nan; Fan, Zhen

    2013-01-01

    We tested the novel hypothesis that EMMPRIN/CD147, a transmembrane glycoprotein overexpressed in breast cancer cells, has a previously unknown role in transforming fibroblasts to cancer-associated fibroblasts, and that cancer-associated fibroblasts in turn induce epithelial-to-mesenchymal transition of breast cancer cells. Co-culture of fibroblasts with breast cancer cells or treatment of fibroblasts with breast cancer cell conditioned culture medium or recombinant EMMPRIN/CD147 induced expression of α-SMA in the fibroblasts in an EMMPRIN/CD147-dependent manner and promoted epithelial-to-mesenchymal transition of breast cancer cells and enhanced cell migration potential. These findings support a novel role of EMMPRIN/CD147 in regulating the interaction between cancer and stroma. PMID:23474495

  5. Effect of microenviroment hypoxia on glioma cells radiosensitivity through cancer stem cell pathway

    International Nuclear Information System (INIS)

    Tong Liumei; Feng Libo; Lu Xueguan; Chen Liesong; Guo Xinwei; Tian Ye

    2010-01-01

    Objective: To investigate the effect of microenviroment hypoxia on glioma cells radiosensitivity through cancer stem pathway, and to explore the related mechanism. Methods: Glioma cell lines SHG44 and U251 were cultured in normoxia (20% O 2 ) or continuous hypoxia (1% O 2 ) for 12 and 24 h. The fraction of glioma cells with positive expression of CD133 was assayed by flow cytometry. The radiosensitivity of glioma cells was determined by clonogenic cell assay. Western blotting was used to investigate the expressions of HIF-1 α and its downstream gene Notch 1. Results: The fraction of glioma cells with positive expression of CD133 was higher after hypoxic culture for 12 and 24 h than that of the corresponding cells cultured in normoxia. Compared to the cells cultured in normoxia, SF 2 (survival fraction at 2 Gy) were enhanced significantly in SHG44 and U251 cells cultured in hypoxia for 12 and 24 h. The OER (oxygen-enhancement ratio) of SHG44 cells in hypoxia for 12 and 24 h was 1.54 and 1.38, respectively. The OER of U251 cells was 1.44 and 1.23, respectively. The radiosensitivity of these two cell line was decreased in hypoxia. The protein expressions of HIF-1 α and Notch 1 genes were elevated more significantly for cells cultured in hypoxia for 12 and 24 h than for those in normoxia. Conclusions: Microenviroment hypoxia could increase the radioresistance of glioma cells through enrichment of cancer stem cells, and HIF-1 α-Notch 1 signal pathway may play an important role in this process. (authors)

  6. How do culture media influence in vitro perivascular cell behavior?

    Science.gov (United States)

    Huber, Birgit; Volz, Ann-Cathrin; Kluger, Petra Juliane

    2015-12-01

    Perivascular cells are multilineage cells located around the vessel wall and important for wall stabilization. In this study, we evaluated a stem cell media and a perivascular cell-specific media for the culture of primary perivascular cells regarding their cell morphology, doubling time, stem cell properties, and expression of cell type-specific markers. When the two cell culture media were compared to each other, perivascular cells cultured in the stem cell medium had a more elongated morphology and a faster doubling rate and cells cultured in the pericyte medium had a more typical morphology, with several filopodia, and a slower doubling rate. To evaluate stem cell properties, perivascular cells, CD146(-) cells, and mesenchymal stem cells (MSCs) were differentiated into the adipogenic, osteogenic, and chondrogenic lineages. It was seen that perivascular cells, as well as CD146(-) cells and MSCs, cultured in stem cell medium showed greater differentiation than cells cultured in pericyte-specific medium. The expression of pericyte-specific markers CD146, neural/glial antigen 2 (NG2), platelet-derived growth factor receptor-β (PDGFR-β), myosin, and α-smooth muscle actin (α-SMA) could be found in both pericyte cultures, as well as to varying amounts in CD146(-) cells, MSCs, and endothelial cells. The here presented work shows that perivascular cells can adapt to their in vitro environment and cell culture conditions influence cell functionality, such as doubling rate or differentiation behavior. Pericyte-specific markers were shown to be expressed also from cells other than perivascular cells. We can further conclude that CD146(+) perivascular cells are inhomogeneous cell population probably containing stem cell subpopulations, which are located perivascular around capillaries. © 2015 International Federation for Cell Biology.

  7. Genomic instability of osteosarcoma cell lines in culture: impact on the prediction of metastasis relevant genes.

    Science.gov (United States)

    Muff, Roman; Rath, Prisni; Ram Kumar, Ram Mohan; Husmann, Knut; Born, Walter; Baudis, Michael; Fuchs, Bruno

    2015-01-01

    Osteosarcoma is a rare but highly malignant cancer of the bone. As a consequence, the number of established cell lines used for experimental in vitro and in vivo osteosarcoma research is limited and the value of these cell lines relies on their stability during culture. Here we investigated the stability in gene expression by microarray analysis and array genomic hybridization of three low metastatic cell lines and derivatives thereof with increased metastatic potential using cells of different passages. The osteosarcoma cell lines showed altered gene expression during in vitro culture, and it was more pronounced in two metastatic cell lines compared to the respective parental cells. Chromosomal instability contributed in part to the altered gene expression in SAOS and LM5 cells with low and high metastatic potential. To identify metastasis-relevant genes in a background of passage-dependent altered gene expression, genes involved in "Pathways in cancer" that were consistently regulated under all passage comparisons were evaluated. Genes belonging to "Hedgehog signaling pathway" and "Wnt signaling pathway" were significantly up-regulated, and IHH, WNT10B and TCF7 were found up-regulated in all three metastatic compared to the parental cell lines. Considerable instability during culture in terms of gene expression and chromosomal aberrations was observed in osteosarcoma cell lines. The use of cells from different passages and a search for genes consistently regulated in early and late passages allows the analysis of metastasis-relevant genes despite the observed instability in gene expression in osteosarcoma cell lines during culture.

  8. Continuous Release of Tumor-Derived Factors Improves the Modeling of Cachexia in Muscle Cell Culture

    Directory of Open Access Journals (Sweden)

    Robert W. Jackman

    2017-09-01

    Full Text Available Cachexia is strongly associated with a poor prognosis in cancer patients but the biological trigger is unknown and therefore no therapeutics exist. The loss of skeletal muscle is the most deleterious aspect of cachexia and it appears to depend on secretions from tumor cells. Models for studying wasting in cell culture consist of experiments where skeletal muscle cells are incubated with medium conditioned by tumor cells. This has led to candidates for cachectic factors but some of the features of cachexia in vivo are not yet well-modeled in cell culture experiments. Mouse myotube atrophy measured by myotube diameter in response to medium conditioned by mouse colon carcinoma cells (C26 is consistently less than what is seen in muscles of mice bearing C26 tumors with moderate to severe cachexia. One possible reason for this discrepancy is that in vivo the C26 tumor and skeletal muscle share a circulatory system exposing the muscle to tumor factors in a constant and increasing way. We have applied Transwell®-adapted cell culture conditions to more closely simulate conditions found in vivo where muscle is exposed to the ongoing kinetics of constant tumor secretion of active factors. C26 cells were incubated on a microporous membrane (a Transwell® insert that constitutes the upper compartment of wells containing plated myotubes. In this model, myotubes are exposed to a constant supply of cancer cell secretions in the medium but without direct contact with the cancer cells, analogous to a shared circulation of muscle and cancer cells in tumor-bearing animals. The results for myotube diameter support the idea that the use of Transwell® inserts serves as a more physiological model of the muscle wasting associated with cancer cachexia than the bolus addition of cancer cell conditioned medium. The Transwell® model supports the notion that the dose and kinetics of cachectic factor delivery to muscle play a significant role in the extent of pathology.

  9. Reversible gelling culture media for in-vitro cell culture in three-dimensional matrices

    Science.gov (United States)

    An, Yuehuei H.; Mironov, Vladimir A.; Gutowska, Anna

    2000-01-01

    A gelling cell culture medium useful for forming a three dimensional matrix for cell culture in vitro is prepared by copolymerizing an acrylamide derivative with a hydrophilic comonomer to form a reversible (preferably thermally reversible) gelling linear random copolymer in the form of a plurality of linear chains having a plurality of molecular weights greater than or equal to a minimum gelling molecular weight cutoff, mixing the copolymer with an aqueous solvent to form a reversible gelling solution and adding a cell culture medium to the gelling solution to form the gelling cell culture medium. Cells such as chondrocytes or hepatocytes are added to the culture medium to form a seeded culture medium, and temperature of the medium is raised to gel the seeded culture medium and form a three dimensional matrix containing the cells. After propagating the cells in the matrix, the cells may be recovered by lowering the temperature to dissolve the matrix and centrifuging.

  10. Fluorescent polymer-based post-translational differentiation and subtyping of breast cancer cells.

    Science.gov (United States)

    Scott, Michael D; Dutta, Rinku; Haldar, Manas K; Wagh, Anil; Gustad, Thomas R; Law, Benedict; Friesner, Daniel L; Mallik, Sanku

    2012-12-07

    Herein, we report the application of synthesized fluorescent, water soluble polymers for post-translational subtyping and differentiation of breast cancer cells in vitro. The fluorescence emission spectra from these polymers were modulated differently in the presence of conditioned cell culture media from various breast cancer cells. These polymers differentiate at a post-translation level possibly due to their ability to interact with extracellular enzymes that are over-expressed in cancerous conditions.

  11. Targeting Stromal Recruitment by Prostate Cancer Cells

    Science.gov (United States)

    2006-03-01

    Ensinger, C., Tumer , Z., Tommerup, N. et al.: Hedgehog signaling in small-cell lung cancer : frequent in vivo but a rare event in vitro. Lung Cancer , 52...W81XWH-04-1-0157 TITLE: Targeting Stromal Recruitment by Prostate Cancer Cells PRINCIPAL INVESTIGATOR: Jingxian Zhang, Ph.D...DATES COVERED (From - To) 15 Feb 2004 – 14 Feb 2006 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Targeting Stromal Recruitment by Prostate Cancer

  12. Mesenchymal stem cells derived from normal gingival tissue inhibit the proliferation of oral cancer cells in vitro and in vivo.

    Science.gov (United States)

    Ji, Xiaoli; Zhang, Zhihui; Han, Ying; Song, Jiangyuan; Xu, Xiangliang; Jin, Jianqiu; Su, Sha; Mu, Dongdong; Liu, Xiaodan; Xu, Si; Cui, Hongwei; Zhao, Zhongfang; Wang, Yixiang; Liu, Hongwei

    2016-11-01

    The interplay between tumor cells and mesenchymal stem cells (MSCs) within tumor microenvironment plays a significant role in tumor development, and thus might be exploited for therapeutic intervention. In this study, we isolated MSCs from normal gingival tissue (GMSCs), and detected the effect of GMSCs on oral cancer cells via direct co-culture and indirect co-culture systems. The cell proliferation assay of direct co-culture showed that GMSCs could inhibit the growth of oral cancer cells. Conditioned medium derived from GMSCs (GMSCs-CM) also exerted an anticancer effect, which indicates that soluble factors in GMSCs-CM played a dominant role in GMSCs-induced cancer cell growth inhibition. To investigate the mechanism, we performed apoptosis assay by flow cytometry, and confirmed that cancer cell apoptosis induced by GMSCs could be a reason for the effect of GMSCs on the growth of oral cancer cells. Western blotting also confirmed that GMSCs could upregulate expression of pro-apoptotic genes including p-JNK, cleaved PARP, cleaved caspase-3, Bax expression and downregulate proliferation- and anti-apoptosis-related gene expression such as p-ERK1/2, Bcl-2, CDK4, cyclin D1, PCNA and survivin. Importantly, the inhibitory effect of GMSCs on cancer cells can partially be restored by blockade of JNK pathway. Moreover, animal studies showed that GMSCs exerted an anticancer effect after oral cancer cells and GMSCs were co-injected with oral cancer cells. Taken together, our data suggest that GMSCs can suppress oral cancer cell growth in vitro and in vivo via altering the surrounding microenvironment of oral cancer cells, which indicates that GMSCs have a potential use in the management of oral dysplasia and oral cancer in future.

  13. Traditional and Modern Cell Culture in Virus Diagnosis.

    Science.gov (United States)

    Hematian, Ali; Sadeghifard, Nourkhoda; Mohebi, Reza; Taherikalani, Morovat; Nasrolahi, Abbas; Amraei, Mansour; Ghafourian, Sobhan

    2016-04-01

    Cell cultures are developed from tissue samples and then disaggregated by mechanical, chemical, and enzymatic methods to extract cells suitable for isolation of viruses. With the recent advances in technology, cell culture is considered a gold standard for virus isolation. This paper reviews the evolution of cell culture methods and demonstrates why cell culture is a preferred method for identification of viruses. In addition, the advantages and disadvantages of both traditional and modern cell culture methods for diagnosis of each type of virus are discussed. Detection of viruses by the novel cell culture methods is considered more accurate and sensitive. However, there is a need to include some more accurate methods such as molecular methods in cell culture for precise identification of viruses.

  14. Cell of Origin and Cancer Stem Cell Phenotype in Medulloblastomas

    Science.gov (United States)

    2017-09-01

    AWARD NUMBER: W81XWH-14-1-0115 TITLE: Cell of Origin and Cancer Stem Cell Phenotype in Medulloblastomas PRINCIPAL INVESTIGATOR: Kyuson Yun...CA130273 - Cell of Origin and Cancer Stem Cell Phenotype in Medulloblastomas 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-14-1-0115 5c. PROGRAM...hypothesis, we originally proposed to transform neural stem cells (NSCs) and neural progenitor cells (NPCs) in vivo by expressing an activated form

  15. Impact of Mesenchymal Stem Cell secreted PAI-1 on colon cancer cell migration and proliferation

    International Nuclear Information System (INIS)

    Hogan, Niamh M.; Joyce, Myles R.; Murphy, J. Mary; Barry, Frank P.; O’Brien, Timothy; Kerin, Michael J.; Dwyer, Roisin M.

    2013-01-01

    Highlights: •MSCs were directly co-cultured with colorectal cancer (CRC) cells on 3D scaffolds. •MSCs influence CRC protein/gene expression, proliferation and migration. •We report a significant functional role of MSC-secreted PAI-1 in colon cancer. -- Abstract: Mesenchymal Stem Cells are known to engraft and integrate into the architecture of colorectal tumours, with little known regarding their fate following engraftment. This study aimed to investigate mediators of Mesenchymal Stem Cell (MSC) and colon cancer cell (CCC) interactions. Mesenchymal Stem Cells and colon cancer cells (HT29 and HCT-116) were cultured individually or in co-culture on 3-dimensional scaffolds. Conditioned media containing all secreted factors was harvested at day 1, 3 and 7. Chemokine secretion and expression were analyzed by Chemi-array, ELISA (Macrophage migration inhibitory factor (MIF), plasminogen activator inhibitor type 1 (PAI-1)) and RQ-PCR. Colon cancer cell migration and proliferation in response to recombinant PAI-1, MSCs and MSCs + antibody to PAI-1 was analyzed using Transwell inserts and an MTS proliferation assay respectively. Chemi-array revealed secretion of a wide range of factors by each cell population, including PAI-1and MIF. ELISA analysis revealed Mesenchymal Stem Cells to secrete the highest levels of PAI-1 (MSC mean 10.6 ng/mL, CCC mean 1.01 ng/mL), while colon cancer cells were the principal source of MIF. MSC-secreted PAI-1 stimulated significant migration of both CCC lines, with an antibody to the chemokine shown to block this effect (67–88% blocking,). A cell-line dependant effect on CCC proliferation was shown for Mesenchymal Stem Cell-secreted PAI-1 with HCT-116 cells showing decreased proliferation at all concentrations, and HT29 cells showing increased proliferation in the presence of higher PAI-1 levels. This is the first study to identify PAI-1 as an important mediator of Mesenchymal Stem Cell/colon cancer cell interactions and highlights the

  16. Impact of Mesenchymal Stem Cell secreted PAI-1 on colon cancer cell migration and proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Hogan, Niamh M. [Discipline of Surgery, School of Medicine, National University of Ireland, Galway (Ireland); Joyce, Myles R. [Department of Colorectal Surgery, University College Hospital, Galway (Ireland); Murphy, J. Mary; Barry, Frank P.; O’Brien, Timothy [Regenerative Medicine Institute, National University of Ireland, Galway (Ireland); Kerin, Michael J. [Discipline of Surgery, School of Medicine, National University of Ireland, Galway (Ireland); Dwyer, Roisin M., E-mail: roisin.dwyer@nuigalway.ie [Discipline of Surgery, School of Medicine, National University of Ireland, Galway (Ireland)

    2013-06-14

    Highlights: •MSCs were directly co-cultured with colorectal cancer (CRC) cells on 3D scaffolds. •MSCs influence CRC protein/gene expression, proliferation and migration. •We report a significant functional role of MSC-secreted PAI-1 in colon cancer. -- Abstract: Mesenchymal Stem Cells are known to engraft and integrate into the architecture of colorectal tumours, with little known regarding their fate following engraftment. This study aimed to investigate mediators of Mesenchymal Stem Cell (MSC) and colon cancer cell (CCC) interactions. Mesenchymal Stem Cells and colon cancer cells (HT29 and HCT-116) were cultured individually or in co-culture on 3-dimensional scaffolds. Conditioned media containing all secreted factors was harvested at day 1, 3 and 7. Chemokine secretion and expression were analyzed by Chemi-array, ELISA (Macrophage migration inhibitory factor (MIF), plasminogen activator inhibitor type 1 (PAI-1)) and RQ-PCR. Colon cancer cell migration and proliferation in response to recombinant PAI-1, MSCs and MSCs + antibody to PAI-1 was analyzed using Transwell inserts and an MTS proliferation assay respectively. Chemi-array revealed secretion of a wide range of factors by each cell population, including PAI-1and MIF. ELISA analysis revealed Mesenchymal Stem Cells to secrete the highest levels of PAI-1 (MSC mean 10.6 ng/mL, CCC mean 1.01 ng/mL), while colon cancer cells were the principal source of MIF. MSC-secreted PAI-1 stimulated significant migration of both CCC lines, with an antibody to the chemokine shown to block this effect (67–88% blocking,). A cell-line dependant effect on CCC proliferation was shown for Mesenchymal Stem Cell-secreted PAI-1 with HCT-116 cells showing decreased proliferation at all concentrations, and HT29 cells showing increased proliferation in the presence of higher PAI-1 levels. This is the first study to identify PAI-1 as an important mediator of Mesenchymal Stem Cell/colon cancer cell interactions and highlights the

  17. Isolation and Characterization of Cancer Stem Cells of the Non-Small-Cell Lung Cancer (A549) Cell Line.

    Science.gov (United States)

    Halim, Noor Hanis Abu; Zakaria, Norashikin; Satar, Nazilah Abdul; Yahaya, Badrul Hisham

    2016-01-01

    Cancer is a major health problem worldwide. The failure of current treatments to completely eradicate cancer cells often leads to cancer recurrence and dissemination. Studies have suggested that tumor growth and spread are driven by a minority of cancer cells that exhibit characteristics similar to those of normal stem cells, thus these cells are called cancer stem cells (CSCs). CSCs are believed to play an important role in initiating and promoting cancer. CSCs are resistant to currently available cancer therapies, and understanding the mechanisms that control the growth of CSCs might have great implications for cancer therapy. Cancer cells are consist of heterogeneous population of cells, thus methods of identification, isolation, and characterisation of CSCs are fundamental to obtain a pure CSC populations. Therefore, this chapter describes in detail a method for isolating and characterizing a pure population of CSCs from heterogeneous population of cancer cells and CSCs based on specific cell surface markers.

  18. Intersections of lung progenitor cells, lung disease and lung cancer.

    Science.gov (United States)

    Kim, Carla F

    2017-06-30

    The use of stem cell biology approaches to study adult lung progenitor cells and lung cancer has brought a variety of new techniques to the field of lung biology and has elucidated new pathways that may be therapeutic targets in lung cancer. Recent results have begun to identify the ways in which different cell populations interact to regulate progenitor activity, and this has implications for the interventions that are possible in cancer and in a variety of lung diseases. Today's better understanding of the mechanisms that regulate lung progenitor cell self-renewal and differentiation, including understanding how multiple epigenetic factors affect lung injury repair, holds the promise for future better treatments for lung cancer and for optimising the response to therapy in lung cancer. Working between platforms in sophisticated organoid culture techniques, genetically engineered mouse models of injury and cancer, and human cell lines and specimens, lung progenitor cell studies can begin with basic biology, progress to translational research and finally lead to the beginnings of clinical trials. Copyright ©ERS 2017.

  19. Intersections of lung progenitor cells, lung disease and lung cancer

    Directory of Open Access Journals (Sweden)

    Carla F. Kim

    2017-06-01

    Full Text Available The use of stem cell biology approaches to study adult lung progenitor cells and lung cancer has brought a variety of new techniques to the field of lung biology and has elucidated new pathways that may be therapeutic targets in lung cancer. Recent results have begun to identify the ways in which different cell populations interact to regulate progenitor activity, and this has implications for the interventions that are possible in cancer and in a variety of lung diseases. Today's better understanding of the mechanisms that regulate lung progenitor cell self-renewal and differentiation, including understanding how multiple epigenetic factors affect lung injury repair, holds the promise for future better treatments for lung cancer and for optimising the response to therapy in lung cancer. Working between platforms in sophisticated organoid culture techniques, genetically engineered mouse models of injury and cancer, and human cell lines and specimens, lung progenitor cell studies can begin with basic biology, progress to translational research and finally lead to the beginnings of clinical trials.

  20. CANCER PATIENT’S EXPERIENCE CROSSING THE HEALTH CULTURE

    Directory of Open Access Journals (Sweden)

    Maura G. Felea

    2014-12-01

    Full Text Available Cognitive anthropology does not predict human behavior, but tries to access principles that rule behavior. Cross-cultural communication is a skill acquired through a learning process, and it can improve doctor-patient relationship and enhance the outcomes of care. The unfulfilled expectations of a patient may influence the patient self-esteem and his perceived role in the society. For some patients living with cancer, it was found as an unforeseen benefit of learning to be closer to God. Based on a narrative communication, we tried to underline cross-cultural differences in cancer patients from different countries with various backgrounds. We described the patient reactions, his way of interpreting the things that happened to him, and his actions regarding adaptive changes in behavior. The originality of the study resides in understanding cross-cultural patterns of cancer patients. The innovative element is the use of qualitative research and its application in health care.

  1. PHYTOCHEMICAL STUDY OF CELL CULTURE JATROPHA CURCAS

    Directory of Open Access Journals (Sweden)

    KOMAR RUSLAN

    2011-01-01

    Full Text Available Jatropha curcas belongs to the Euphorbiaceae family which has potential economically. This plant has been reported to contain toxic compounds such as curcin and phorbol ester and its derivatives. These compounds may become a problem if J. curcas will be explored as a source of biofuel. In order to provide safety plants, the research on the study of phytochemical and initiation of cell and organ culture have been carried out. J curcas which has been collected from different regions in Indonesia showed to contain relatively the same profile of chemical contents. Dominant compounds that were detected by GCMS are hidrocarbon such as 2-heptenal, decadienal, hexsadecane, pentadecane, cyclooctane etc, fatty acid such as oktadecanoate acid, etthyl linoleate, ethyl stearate, heksadecanoate acid and steroid such as stigmasterol, fucosterol, sitosterol. No phorbol ester and its derivatives have been detected yet by the GCMS method. Callus and suspension cultures of J. curcas have been established to be used for further investigation.

  2. Investigating the effect of cell substrate on cancer cell stiffness by optical tweezers.

    Science.gov (United States)

    Yousafzai, Muhammad Sulaiman; Coceano, Giovanna; Bonin, Serena; Niemela, Joseph; Scoles, Giacinto; Cojoc, Dan

    2017-07-26

    The mechanical properties of cells are influenced by their microenvironment. Here we report cell stiffness alteration by changing the cell substrate stiffness for isolated cells and cells in contact with other cells. Polydimethylsiloxane (PDMS) is used to prepare soft substrates with three different stiffness values (173, 88 and 17kPa respectively). Breast cancer cells lines, namely HBL-100, MCF-7 and MDA-MB-231 with different level of aggressiveness are cultured on these substrates and their local elasticity is investigated by vertical indentation of the cell membrane. Our preliminary results show an unforeseen behavior of the MDA-MB-231 cells. When cultured on glass substrate as isolated cells, they are less stiff than the other two types of cells, in agreement with the general statement that more aggressive and metastatic cells are softer. However, when connected to other cells the stiffness of MDA-MB-231 cells becomes similar to the other two cell lines. Moreover, the stiffness of MDA-MB-231 cells cultured on soft PDMS substrates is significantly higher than the stiffness of the other cell types, demonstrating thus the strong influence of the environmental conditions on the mechanical properties of the cells. Copyright © 2017. Published by Elsevier Ltd.

  3. Good cell culture practices &in vitro toxicology.

    Science.gov (United States)

    Eskes, Chantra; Boström, Ann-Charlotte; Bowe, Gerhard; Coecke, Sandra; Hartung, Thomas; Hendriks, Giel; Pamies, David; Piton, Alain; Rovida, Costanza

    2017-12-01

    Good Cell Culture Practices (GCCP) is of high relevance to in vitro toxicology. The European Society of Toxicology In Vitro (ESTIV), the Center for Alternatives for Animal Testing (CAAT) and the In Vitro Toxicology Industrial Platform (IVTIP) joined forces to address by means of an ESTIV 2016 pre-congress session the different aspects and applications of GCCP. The covered aspects comprised the current status of the OECD guidance document on Good In Vitro Method Practices, the importance of quality assurance for new technological advances in in vitro toxicology including stem cells, and the optimized implementation of Good Manufacturing Practices and Good Laboratory Practices for regulatory testing purposes. General discussions raised the duality related to the difficulties in implementing GCCP in an academic innovative research framework on one hand, and on the other hand, the need for such GCCP principles in order to ensure reproducibility and robustness of in vitro test methods for toxicity testing. Indeed, if good cell culture principles are critical to take into consideration for all uses of in vitro test methods for toxicity testing, the level of application of such principles may depend on the stage of development of the test method as well as on the applications of the test methods, i.e., academic innovative research vs. regulatory standardized test method. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Apoptosis induced by GanoPoly in human gastric cancer cell line ...

    African Journals Online (AJOL)

    In order to investigate polysaccharide effect on the cultured human gastric cancer cells (SGC7901), DNA ladder, flow cytometry and western blot were used to examine the morpholog, proliferation and apoptosis of human gastric cancer SGC-7901 cells when they were affected by polysaccharide. Results show that ...

  5. The stem cell division theory of cancer.

    Science.gov (United States)

    López-Lázaro, Miguel

    2018-03-01

    All cancer registries constantly show striking differences in cancer incidence by age and among tissues. For example, lung cancer is diagnosed hundreds of times more often at age 70 than at age 20, and lung cancer in nonsmokers occurs thousands of times more frequently than heart cancer in smokers. An analysis of these differences using basic concepts in cell biology indicates that cancer is the end-result of the accumulation of cell divisions in stem cells. In other words, the main determinant of carcinogenesis is the number of cell divisions that the DNA of a stem cell has accumulated in any type of cell from the zygote. Cell division, process by which a cell copies and separates its cellular components to finally split into two cells, is necessary to produce the large number of cells required for living. However, cell division can lead to a variety of cancer-promoting errors, such as mutations and epigenetic mistakes occurring during DNA replication, chromosome aberrations arising during mitosis, errors in the distribution of cell-fate determinants between the daughter cells, and failures to restore physical interactions with other tissue components. Some of these errors are spontaneous, others are promoted by endogenous DNA damage occurring during quiescence, and others are influenced by pathological and environmental factors. The cell divisions required for carcinogenesis are primarily caused by multiple local and systemic physiological signals rather than by errors in the DNA of the cells. As carcinogenesis progresses, the accumulation of DNA errors promotes cell division and eventually triggers cell division under permissive extracellular environments. The accumulation of cell divisions in stem cells drives not only the accumulation of the DNA alterations required for carcinogenesis, but also the formation and growth of the abnormal cell populations that characterize the disease. This model of carcinogenesis provides a new framework for understanding the

  6. Radiometric detection of yeasts in blood cultures of cancer patients

    International Nuclear Information System (INIS)

    Hopfer, R.L.; Orengo, A.; Chesnut, S.; Wenglar, M.

    1980-01-01

    During a 12-month period, 19,457 blood cultures were collected. Yeasts were isolated from 193 cultures derived from 76 cancer patients. Candida albicans or Candida tropicalis accounted for 79% of isolates. Of the three methods compared, the radiometric method required 2.9 days to become positive, blind subculture required 2.6 days, and Gram stains required 1 day. However, the radiometric method was clearly superior in detecting positive cultures, since 73% of all cultures were first detected radiometrically, 22% were detected by subculture, and only 5% were detected by Gram stain. Although 93% of the isolates were detected by aerobic culture, five (7%) isolates were obtained only from anaerobic cultures. Seven days of incubation appear to be sufficient for the radiometric detection of yeasts

  7. Radiation induced damage to the lipid contents of bacteria and cultured mammalian cells

    International Nuclear Information System (INIS)

    Gholipour Khalili, K.

    1993-01-01

    In this study, exponentially growing phase of E. Coli. K12-N167 and cultured mouse leukemic L5178Y were used to study the effect of gamma irradiation on phospholipid contents. Following irradiation, both bacteria and cultured cells were incubated with either 14 C or 32 P labelled precursors for periods of cell division time. Phospholipid composition and their contents were detected in both the bacteria and cultured cells by using liquid scintillation counting and autoradiography methods. In contrast, as radiation dose increased, the Phospholipid contents were decreased in the both bacteria and cultured cells. It was concluded that the changes of phospholipid contents may result to altered activities of phospholipid pathway enzymes damaged by a radiation dose. The results of this investigation would be helpful in control of induced radiation damages in cell killings in radiation workers and radiation treatment of human cancer in the clinics. (author). 35 refs, 3 figs, 4 tabs

  8. Enhanced infectivity of bluetongue virus in cell culture by centrifugation.

    OpenAIRE

    Sundin, D R; Mecham, J O

    1989-01-01

    The effects of centrifugation of the infection of cell culture with bluetongue virus (BTV) were investigated. Baby hamster kidney cells were infected with BTV with or without centrifugation. Viral antigen was detected by immunofluorescence at 24 h in both centrifuged and noncentrifuged cultures. However, after 24 h of infection, the production of PFU in centrifuged cell cultures was 10- to 20-fold greater than that seen in cultures not centrifuged. In addition, centrifugation enhanced the dir...

  9. Differences in gene expression of cells growing in conventional 2D versus 3D cell culture

    International Nuclear Information System (INIS)

    Zschenker, Oliver; Cordes, Nils; Streichert, Thomas

    2009-01-01

    Full text: Telomeres are DNA protein complexes on the ends of chromosomes that distinguish the ends of chromosomes from double strand breaks and prevent degradation or fusion by nonhomologous end-joining. The loss of telomeres is associated with a loss of heterochromatic features leading to a less compact chromatin structure which allows e.g. DNA repair proteins to get better access to the site of the DNA damage and facilitate chromosome fusions. Telomerase is an enzyme that can counteract the loss of telomeres by adding telomeric repeats on the ends of chromosomes. Since telomerase is active in most tumor cells, telomerase is suggested to be the reason for the unlimited number of cell divisions of cancer cells. TRF2 is one of the most important proteins of the Shelterin complex protecting the telomeres from shortening by inhibiting ATM which is up-stream of the DNA repair mechanisms. Thus, we are concentrating on TRF2 and telomerase to investigate the differences in DNA repair in telomeric (heterochromatic) versus euchromatic regions. Human cancer cells with differences in status of p53 and telomerase like A549, UT-SCC15 and FaDu cells are used. Without any treatment, FaDu cells express high levels of telomerase and TRF2 in conventional 2D cell culture which is in contrast to e.g. A549. We found that telomerase is even higher expressed in 3D than in 2D cell culture. To connect telomere associated processes to both repair of radiogenic DNA damage/lesions and to cell-extracellular matrix interactions, we performed whole genome microarray analysis. By comparing the differential expression of genes associated with these three cell functions, we intend to yield new molecular insight into radiotherapy relevant tumor characteristics, particularly radioresistance and DNA damage response network processing. (author)

  10. Cytologic studies on irradiated gestric cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Isono, S; Takeda, T; Amakasu, H; Asakawa, H; Yamada, S [Miyagi Prefectural Adult Disease Center, Natori (Japan)

    1981-06-01

    The smears of the biopsy and resected specimens obtained from 74 cases of irradiated gastric cancer were cytologically analyzed for effects of irradiation. Irradiation increased the amount of both necrotic materials and neutrophils in the smears. Cancer cells were decreased in number almost in inverse proportion to irradiation dose. Clusters of cancer cells shrank in size and cells were less stratified after irradiation. Irradiated cytoplasms were swollen, vacuolated and stained abnormally. Irradiation with less than 3,000 rads gave rise to swelling of cytoplasms in almost all cases. Nuclei became enlarged, multiple, pyknotic and/or stained pale after irradiation. Nuclear swelling was more remarkable in cancer cells of differentiated adenocarcinomas.

  11. Cultural Effects on Cancer Prevention Behaviors: Fatalistic Cancer Beliefs and Risk Optimism Among Asians in Singapore.

    Science.gov (United States)

    Kim, Hye Kyung; Lwin, May O

    2017-10-01

    Although culture is acknowledged as an important factor that influences health, little is known about cultural differences pertaining to cancer-related beliefs and prevention behaviors. This study examines two culturally influenced beliefs-fatalistic beliefs about cancer prevention, and optimistic beliefs about cancer risk-to identify reasons for cultural disparity in the engagement of cancer prevention behaviors. We utilized data from national surveys of European Americans in the United States (Health Information National Trends Survey 4, Cycle3; N = 1,139) and Asians in Singapore (N = 1,200) to make cultural comparisons. The odds of an Asian adhering to prevention recommendations were less than half the odds of a European American, with the exception of smoking avoidance. Compared to European Americans, Asians were more optimistic about their cancer risk both in an absolute and a comparative sense, and held stronger fatalistic beliefs about cancer prevention. Mediation analyses revealed that fatalistic beliefs and absolute risk optimism among Asians partially explain their lower engagement in prevention behaviors, whereas comparative risk optimism increases their likelihood of adhering to prevention behaviors. Our findings underscore the need for developing culturally targeted interventions in communicating cancer causes and prevention.

  12. Cardiac Cells Beating in Culture: A Laboratory Exercise

    Science.gov (United States)

    Weaver, Debora

    2007-01-01

    This article describes how to establish a primary tissue culture, where cells are taken directly from an organ of a living animal. Cardiac cells are taken from chick embryos and transferred to culture dishes. These cells are not transformed and therefore have a limited life span. However, the unique characteristics of cardiac cells are maintained…

  13. Induction of apoptosis by eugenol in human breast cancer cells

    International Nuclear Information System (INIS)

    Vidhya, N.; Niranjali Devaraj, S.

    2011-01-01

    In the present study, potential anticancer effect of eugenol on inhibition of cell proliferation and induction of apoptosis in human MCF-7 breast cancer cells was investigated. Induction of cell death by eugenol was evaluated following MTT assay and monitoring lactate dehydrogenase released into the culture medium for cell viability and cytotoxicity, giemsa staining for morphological alterations, fluorescence microscopy analysis of cells using ethidium bromide and acridine orange and quantitation of DNA fragments for induction of apoptosis. Effect of eugenol on intracellular redox status of the human breast cancer cells was assessed by determining the level of glutathione and lipid peroxidation products (TBARS). Eugenol treatment inhibited the growth and proliferation of human MCF-7 breast cancer cells through induction of cell death, which was dose and time dependent. Microscopic examination of eugenol treated cells showed cell shrinkage, membrane blebbing and apoptotic body formation. Further, eugenol treatment also depleted the level of intracellular glutathione and increased the level of lipid peroxidation. The dose dependent increase in the percentage of apoptotic cells and DNA fragments suggested that apoptosis was involved in eugenol induced cell death and apoptosis might have played a role in the chemopreventive action of eugenol. (author)

  14. Culture models of human mammary epithelial cell transformation

    Energy Technology Data Exchange (ETDEWEB)

    Stampfer, Martha R.; Yaswen, Paul

    2000-11-10

    Human pre-malignant breast diseases, particularly ductal carcinoma in situ (DCIS)3 already display several of the aberrant phenotypes found in primary breast cancers, including chromosomal abnormalities, telomerase activity, inactivation of the p53 gene and overexpression of some oncogenes. Efforts to model early breast carcinogenesis in human cell cultures have largely involved studies in vitro transformation of normal finite lifespan human mammary epithelial cells (HMEC) to immortality and malignancy. We present a model of HMEC immortal transformation consistent with the know in vivo data. This model includes a recently described, presumably epigenetic process, termed conversion, which occurs in cells that have overcome stringent replicative senescence and are thus able to maintain proliferation with critically short telomeres. The conversion process involves reactivation of telomerase activity, and acquisition of good uniform growth in the absence and presence of TFGB. We propose th at overcoming the proliferative constraints set by senescence, and undergoing conversion, represent key rate-limiting steps in human breast carcinogenesis, and occur during early stage breast cancer progression.

  15. 3D cell cultures of human head and neck squamous cell carcinoma cells are radiosensitized by the focal adhesion kinase inhibitor TAE226

    International Nuclear Information System (INIS)

    Hehlgans, Stephanie; Lange, Inga; Eke, Iris; Cordes, Nils

    2009-01-01

    Background and purpose: Focal adhesion kinase (FAK), a main player in integrin signaling and survival, is frequently overexpressed in human cancers and therefore postulated as potential target in cancer therapy. The aim of this study was to evaluate the radiosensitizing potential of the FAK inhibitor TAE226 in three-dimensional (3D) tumor cell cultures. Materials and methods: Head and neck squamous cell carcinoma (HNSCC) cells (FaDu, UT-SCC15, UT-SCC45), lung cancer cells (A549), colorectal carcinoma cells (DLD-1, HCT-116) and pancreatic tumor cells (MiaPaCa2, Panc1) were treated with different concentrations of TAE226 (0-1 μm; 1 or 24 h) without or in combination with irradiation (0-6 Gy, X-ray, single dose). Subsequently, 3D clonogenic survival assays (laminin-rich extracellular matrix) and Western blotting (expression/phosphorylation, e.g. FAK, Akt, ERK1/2) were performed. Results: All investigated 3D cell cultures showed a dose-dependent reduction in clonogenic survival by TAE226. Intriguingly, TAE226 only significantly radiosensitized 3D HNSCC cell cultures accompanied by a pronounced dephosphorylation of FAK, Akt and ERK1/2. Conclusions: Our data demonstrate TAE226 as potent FAK inhibitor that enhances the cellular radiosensitivity particularly of HNSCC cells grown in a 3D cell culture model. Future in vitro and in vivo investigations will clarify, to which extent this approach might be clinically relevant for radiotherapy of HNSCC.

  16. DNA MUTAGENESIS IN PANAX GINSENG CELL CULTURES

    Directory of Open Access Journals (Sweden)

    Kiselev K.V.

    2012-08-01

    Full Text Available At the present time, it is well documented that plant tissue culture induces a number of mutations and chromosome rearrangements termed “somaclonal variations”. However, little is known about the nature and the molecular mechanisms of the tissue culture-induced mutagenesis and the effects of long-term subculturing on the rate and specific features of the mutagenesis. The aim of the present study was to investigate and compare DNA mutagenesis in different genes of Panax ginseng callus cultures of different age. It has previously been shown that the nucleotide sequences of the Agrobacterium rhizogenes rolC locus and the selective marker nptII developed mutations during long-term cultivation of transgenic cell cultures of P. ginseng. In the present work, we analyzed nucleotide sequences of selected plant gene families in a 2-year-old and 20-year-old P. ginseng 1c cell culture and in leaves of cultivated P. ginseng plants. We analysed sequence variability between the Actin genes, which are a family of house-keeping genes; the phenylalanine ammonia-lyase (PAL and dammarenediol synthase (DDS genes, which actively participate in the biosynthesis of ginsenosides; and the somatic embryogenesis receptor kinase (SERK genes, which control plant development. The frequency of point mutations in the Actin, PAL, DDS, and SERK genes in the 2-year-old callus culture was markedly higher than that in cultivated plants but lower than that in the 20-year-old callus culture of P. ginseng. Most of the mutations in the 2- and 20-year-old P. ginseng calli were A↔G and T↔C transitions. The number of nonsynonymous mutations was higher in the 2- and 20-year-old callus cultures than the number of nonsynonymous mutations in the cultivated plants of P. ginseng. Interestingly, the total number of N→G or N→C substitutions in the analyzed genes was 1.6 times higher than the total number of N→A or N→T substitutions. Using methylation-sensitive DNA fragmentation

  17. Cultured circulating tumor cells and their derived xenografts for personalized oncology

    Directory of Open Access Journals (Sweden)

    Ruoxiang Wang

    2016-10-01

    Full Text Available Recent cancer research has demonstrated the existence of circulating tumor cells (CTCs in cancer patient's blood. Once identified, CTC biomarkers will be invaluable tools for clinical diagnosis, prognosis and treatment. In this review, we propose ex vivo culture as a rational strategy for large scale amplification of the limited numbers of CTCs from a patient sample, to derive enough CTCs for accurate and reproducible characterization of the biophysical, biochemical, gene expressional and behavioral properties of the harvested cells. Because of tumor cell heterogeneity, it is important to amplify all the CTCs in a blood sample for a comprehensive understanding of their role in cancer metastasis. By analyzing critical steps and technical issues in ex vivo CTC culture, we developed a cost-effective and reproducible protocol directly culturing whole peripheral blood mononuclear cells, relying on an assumed survival advantage in CTCs and CTC-like cells over the normal cells to amplify this specified cluster of cancer cells.

  18. T-cell receptor v-alpha and v-Beta gene usage in interleukin-2-cultured tumor-infiltrating lymphocytes from patients with breast-cancer

    DEFF Research Database (Denmark)

    Andersen, E; Scholler, J; Straten, P

    1994-01-01

    Tumor-infiltrating lymphocytes (TIL) are often found in malignant breast tumors, and have been claimed to be of prognostic value. It has been proposed that TIL may represent an enriched population of tumor-specific cytotoxic lymphocytes, reacting with antigenic determinants on the tumor cell...

  19. Turbulent Dynamics of Epithelial Cell Cultures

    Science.gov (United States)

    Blanch-Mercader, C.; Yashunsky, V.; Garcia, S.; Duclos, G.; Giomi, L.; Silberzan, P.

    2018-05-01

    We investigate the large length and long time scales collective flows and structural rearrangements within in vitro human bronchial epithelial cell (HBEC) cultures. Activity-driven collective flows result in ensembles of vortices randomly positioned in space. By analyzing a large population of vortices, we show that their area follows an exponential law with a constant mean value and their rotational frequency is size independent, both being characteristic features of the chaotic dynamics of active nematic suspensions. Indeed, we find that HBECs self-organize in nematic domains of several cell lengths. Nematic defects are found at the interface between domains with a total number that remains constant due to the dynamical balance of nucleation and annihilation events. The mean velocity fields in the vicinity of defects are well described by a hydrodynamic theory of extensile active nematics.

  20. Natural Killer T Cells in Cancer Immunotherapy

    Science.gov (United States)

    Nair, Shiny; Dhodapkar, Madhav V.

    2017-01-01

    Natural killer T (NKT) cells are specialized CD1d-restricted T cells that recognize lipid antigens. Following stimulation, NKT cells lead to downstream activation of both innate and adaptive immune cells in the tumor microenvironment. This has impelled the development of NKT cell-targeted immunotherapies for treating cancer. In this review, we provide a brief overview of the stimulatory and regulatory functions of NKT cells in tumor immunity as well as highlight preclinical and clinical studies based on NKT cells. Finally, we discuss future perspectives to better harness the potential of NKT cells for cancer therapy. PMID:29018445

  1. 3D cellular structures and co-cultures formed through the contactless magnetic manipulation of cells on adherent surfaces.

    Science.gov (United States)

    Abdel Fattah, Abdel Rahman; Mishriki, Sarah; Kammann, Tobias; Sahu, Rakesh P; Geng, Fei; Puri, Ishwar K

    2018-02-27

    A magnet array is employed to manipulate diamagnetic cells that are contained in paramagnetic medium to demonstrate for the first time the contactless bioprinting of three-dimensional (3D) cellular structures and co-cultures of breast cancer MCF-7 and endothelial HUVEC at prescribed locations on tissue culture treated well plates. Sequential seeding of different cell lines and the spatial displacement of the magnet array creates co-cultured cellular structures within a well without using physically intrusive well inserts. Both monotypic and co-culture experiments produce morphologically rich 3D cell structures that are otherwise absent in regular monolayer cell cultures. The magnetic contactless bioprinting of cells provides further insight into cell behaviour, invasion strategies and transformations that are useful for potential applications in drug screening, 3D cell culture formation and tissue engineering.

  2. Prostate cancer cells induce osteoblastic differentiation via semaphorin 3A.

    Science.gov (United States)

    Liu, Fuzhou; Shen, Weiwei; Qiu, Hao; Hu, Xu; Zhang, Chao; Chu, Tongwei

    2015-03-01

    Prostate cancer metastasis to bone is the second most commonly diagnosed malignant disease among men worldwide. Such metastatic disease is characterized by the presence of osteoblastic bone lesions, and is associated with high rates of mortality. However, the various mechanisms involved in prostate cancer-induced osteoblastic differentiation have not been fully explored. Semaphorin 3A (Sema 3A) is a newly identified regulator of bone metabolism which stimulates differentiation of pre-osteoblastic cells under physiological conditions. We investigated in this study whether prostate cancer cells can mediate osteoblastic activity through Sema 3A. We cultured osteoprogenitor MC3T3-E1 cells in prostate cancer-conditioned medium, and analyzed levels of Sema 3A protein in diverse prostate cancer cell lines to identify cell lines in which Sema 3A production showed a positive correlation with osteo-stimulation. C4-2 cells were stably transfected with Sema 3A short hairpin RNA to further determine whether Sema 3A contributes to the ability of C4-2 cells to induce osteoblastic differentiation. Down-regulation of Sema 3A expression decreased indicators of C4-2 CM-induced osteoblastic differentiation, including alkaline phosphatase production and mineralization. Additionally, silencing or neutralizing Sema 3A in C4-2 cells resulted in diminished β-catenin expression in osteogenitor MC3T3-E1 cells. Our results suggest that prostate cancer-induced osteoblastic differentiation is at least partially mediated by Sema 3A, and may be regulated by the β-catenin signalling pathway. Sema 3A may represent a novel target for treatment of prostate cancer-induced osteoblastic lesions. © 2014 Wiley Periodicals, Inc.

  3. Low Temperature Plasma for the Treatment of Epithelial Cancer Cells

    Science.gov (United States)

    Mohades, Soheila

    Biomedical applications of low temperature plasmas (LTP) may lead to a paradigm shift in treating various diseases by conducting fundamental research on the effects of LTP on cells, tissues, organisms (plants, insects, and microorganisms). This is a rapidly growing interdisciplinary research field that involves engineering, physics, life sciences, and chemistry to find novel solutions for urgent medical needs. Effects of different LTP sources have shown the anti-tumor properties of plasma exposure; however, there are still many unknowns about the interaction of plasma with eukaryotic cells which must be elucidated in order to evaluate the practical potential of plasma in cancer treatment. Plasma, the fourth state of matter, is composed of electrons, ions, reactive molecules (radicals and non-radicals), excited species, radiation, and heat. A sufficient dose (time) of plasma exposure can induce death in cancer cells. The plasma pencil is employed to study the anti-tumor properties of this treatment on epithelial cells. The plasma pencil has been previously used for the inactivation of bacteria, destroying amyloid fibrils, and the killing of various cancer cells. Bladder cancer is the 9th leading cause of cancer. In this dissertation, human urinary bladder tissue with the squamous cell carcinoma disease (SCaBER cells) is treated with LTP utilizing two different approaches: direct plasma exposure and Plasma Activated Media (PAM) as an advancement to the treatment. PAM is produced by exposing a liquid cell culture medium to the plasma pencil. Direct LTP treatment of cancer cells indicates a dose-dependent killing effect at post-treatment times. Similarly, PAM treatment shows an anti-cancer effect by inducing substantial cell death. Reactive oxygen species (ROS) and reactive nitrogen species (RNS) have an important role in the biomedical effects of LTP treatment. This study demonstrates the capability of the plasma pencil to transport ROS/RNS into cell culture media

  4. A Rapid Culture Technique Produces Functional Dendritic-Like Cells from Human Acute Myeloid Leukemia Cell Lines

    Directory of Open Access Journals (Sweden)

    Jian Ning

    2011-01-01

    Full Text Available Most anti-cancer immunotherapeutic strategies involving dendritic cells (DC as vaccines rely upon the adoptive transfer of DC loaded with exogenous tumour-peptides. This study utilized human acute myeloid leukemia (AML cells as progenitors from which functional dendritic-like antigen presenting cells (DLC were generated, that constitutively express tumour antigens for recognition by CD8+ T cells. DLC were generated from AML cell lines KG-1 and MUTZ-3 using rapid culture techniques and appropriate cytokines. DLC were evaluated for their cell-surface phenotype, antigen uptake and ability to stimulate allogeneic responder cell proliferation, and production of IFN-γ; compared with DC derived from normal human PBMC donors. KG-1 and MUTZ-3 DLC increased expression of CD80, CD83, CD86, and HLA-DR, and MUTZ-3 DLC downregulated CD14 and expressed CD1a. Importantly, both KG-1 and MUTZ-3-derived DLC promoted proliferation of allogeneic responder cells more efficiently than unmodified cells; neither cells incorporated FITC-labeled dextran, but both stimulated IFN-γ production from responding allogeneic CD8+ T cells. Control DC produced from PBMC using the FastDC culture also expressed high levels of critical cell surface ligands and demonstrated good APC function. This paper indicates that functional DLC can be cultured from the AML cell lines KG-1 and MUTZ-3, and FastDC culture generates functional KG-1 DLC.

  5. Recombinant Protein Production and Insect Cell Culture and Process

    Science.gov (United States)

    Spaulding, Glenn F. (Inventor); Goodwin, Thomas J. (Inventor); OConnor, Kim C. (Inventor); Francis, Karen M. (Inventor); Andrews, Angela D. (Inventor); Prewett, Tracey L. (Inventor)

    1997-01-01

    A process has been developed for recombinant production of selected polypeptides using transformed insect cells cultured in a horizontally rotating culture vessel modulated to create low shear conditions. A metabolically transformed insect cell line is produced using the culture procedure regardless of genetic transformation. The recombinant polypeptide can be produced by an alternative process using virtually infected or stably transformed insect cells containing a gene encoding the described polypeptide. The insect cells can also be a host for viral production.

  6. Changes in the gene expression of co-cultured human fibroblast cells and osteosarcoma cells: the role of microenvironment.

    Science.gov (United States)

    Salvatore, Viviana; Focaroli, Stefano; Teti, Gabriella; Mazzotti, Antonio; Falconi, Mirella

    2015-10-06

    The progression of malignant tumors does not depend exclusively on the autonomous properties of cancer cells; it is also influenced by tumor stroma reactivity and is under strict microenvironmental control. By themselves, stromal cells are not malignant, and they maintain normal tissue structure and function. However, through intercellular interactions or by paracrine secretions from cancer cells, normal stromal cells acquire abnormal phenotypes that sustain cancer cell growth and tumor progression. In their dysfunctional state, fibroblast and immune cells produce chemokines and growth factors that stimulate cancer cell growth and invasion. In our previous work, we established an in vitro model based on a monolayer co-culture system of healthy human fibroblasts (HFs) and human osteosarcoma cells (the MG-63 cell line) that simulates the microenvironment of tumor cells and healthy cells. The coexistence between MG-63 cells and HFs allowed us to identify the YKL-40 protein as the main marker for verifying the influence of tumor cells grown in contact with healthy cells. In this study, we evaluated the interactions of HFs and MG-63 cells in a transwell co-culture system over 24 h, 48 h, 72 h, and 96 h. We analyzed the contributions of these populations to the tumor microenvironment during cancer progression, as measured by multiple markers. We examined the effect of siRNA knockdown of YKL-40 by tracking the subsequent changes in gene expression within the co-culture. We validated the expression of several genes, focusing on those involved in cancer cell invasion, inflammatory responses, and angiogenesis: TNF alpha, IL-6, MMP-1, MMP-9, and VEGF. We compared the results to those from a transwell co-culture without the YKL-40 knockdown. In a pro-inflammatory environment promoted by TNF alpha and IL-6, siRNA knockdown of YKL-40 caused a down-regulation of VEGF and MMP-1 expression in HFs. These findings demonstrated that the tumor microenvironment has an influence on the

  7. Fabrication of cell-benign inverse opal hydrogels for three-dimensional cell culture.

    Science.gov (United States)

    Im, Pilseon; Ji, Dong Hwan; Kim, Min Kyung; Kim, Jaeyun

    2017-05-15

    Inverse opal hydrogels (IOHs) for cell culture were fabricated and optimized using calcium-crosslinked alginate microbeads as sacrificial template and gelatin as a matrix. In contrast to traditional three-dimensional (3D) scaffolds, the gelatin IOHs allowed the utilization of both the macropore surface and inner matrix for cell co-culture. In order to remove templates efficiently for the construction of 3D interconnected macropores and to maintain high cell viability during the template removal process using EDTA solution, various factors in fabrication, including alginate viscosity, alginate concentration, alginate microbeads size, crosslinking calcium concentration, and gelatin network density were investigated. Low viscosity alginate, lower crosslinking calcium ion concentration, and lower concentration of alginate and gelatin were found to obtain high viability of cells encapsulated in the gelatin matrix after removal of the alginate template by EDTA treatment by allowing rapid dissociation and diffusion of alginate polymers. Based on the optimized fabrication conditions, gelatin IOHs showed good potential as a cell co-culture system, applicable to tissue engineering and cancer research. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. The Implications of Cancer Stem Cells for Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Wenjing Jiang

    2012-12-01

    Full Text Available Surgery, radiotherapy and chemotherapy are universally recognized as the most effective anti-cancer therapies. Despite significant advances directed towards elucidating molecular mechanisms and developing clinical trials, cancer still remains a major public health issue. Recent studies have showed that cancer stem cells (CSCs, a small subpopulation of tumor cells, can generate bulk populations of nontumorigenic cancer cell progeny through the self-renewal and differentiation processes. As CSCs are proposed to persist in tumors as a distinct population and cause relapse and metastasis by giving rise to new tumors, development of CSC-targeted therapeutic strategies holds new hope for improving survival and quality of life in patients with cancer. Therapeutic innovations will emerge from a better understanding of the biology and environment of CSCs, which, however, are largely unexplored. This review summarizes the characteristics, evidences and development of CSCs, as well as implications and challenges for cancer treatment.

  9. Effect of radiation on microtubule structure in cancer cells

    International Nuclear Information System (INIS)

    Tripath, Shambhoo Sharan; Panda, Dulal; Jayakumar, S.; Maikho, Thoh; Sandur, Santosh Kumar

    2017-01-01

    Microtubules (MT) are dynamic structural cellular components. In proliferating cells, they are essential components in cell division through the formation of the mitotic spindle. Radiotherapy is an integral part of cancer treatment for most of the solid cancers. Scanty data exists in the literature related to how ionizing radiation affects microtubule reorganization in tumor cells. In the present study, breast cancer cell line (MCF-7 cells) was exposed to different doses of radiation (2-10Gy). Cells were cultured for 24 h, fixed and stained with antitubulin antibody and subjected to immunofluorescence microscopy. In another experiment, cells were subjected to cold treatment for 5 min or 30 min for studying the disassembly of microtubules after 24 h of irradiation. Further, these cells were incubated at 37°C for 20 min for studying the reassembly of microtubules. Acetylation of microtubule was also examined after exposure of cells to radiation. Experiments were also performed by combining radiation with low concentration of CXI-Benzo 84 (MT destabilizing agent 1 and 2.5 uM). Exposure of MCF-7 cells to radiation lead to destabilization of microtubules. Interestingly, destabilization of microtubule was faster upon cold treatment in irradiated group as compared to control group. These cells failed to re-stabilize at 37°C. Radiation also reduced the acetylation level of microtubule. Combination treatment of CXI-Benzo 84 with radiation exhibited additive effect in terms of depolymerization of MT. Our results suggest that ionizing radiation indeed modulates microtubule dynamics. (author)

  10. Low white blood cell count and cancer

    Science.gov (United States)

    ... gov/ency/patientinstructions/000675.htm Low white blood cell count and cancer To use the sharing features on this page, please enable JavaScript. White blood cells (WBCs) fight infections from bacteria, viruses, fungi, and ...

  11. Cancer stem cells of the digestive system.

    Science.gov (United States)

    Colvin, Hugh S; Nishida, Naohiro; Koseki, Jun; Konno, Masamitsu; Kawamoto, Koichi; Tsunekuni, Kenta; Doki, Yuichiro; Mori, Masaki; Ishii, Hideshi

    2014-12-01

    Stem cells of the digestive system are ideal in many ways for research, given they are abundant, highly proliferative and have a uniform structural arrangement. This in turn has enormously aided the research of cancer stem cells of the digestive system, which is now shaping our understanding of cancer stem cells. In this review, the recent advances in the understanding of cancer stem cells of the digestive system have been summarized, including aspects such as their identification, origin, cell-cycle dormancy, relationship with epithelial-mesenchymal transition, cellular metabolism and the underlying molecular mechanisms. Newly acquired knowledge concerning cancer stem cells have led to the development of novel cancer therapeutics with provisional yet encouraging results. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  12. Embryonal carcinoma cell induction of miRNA and mRNA changes in co-cultured prostate stromal fibromuscular cells

    Science.gov (United States)

    VÊNCIO, ENEIDA F.; PASCAL, LAURA E.; PAGE, LAURA S.; DENYER, GARETH; WANG, AMY J.; RUOHOLA-BAKER, HANNELE; ZHANG, SHILE; WANG, KAI; GALAS, DAVID J.; LIU, ALVIN Y.

    2014-01-01

    The prostate stromal mesenchyme controls organ-specific development. In cancer, the stromal compartment shows altered gene expression compared to non-cancer. The lineage relationship between cancer-associated stromal cells and normal tissue stromal cells is not known. Nor is the cause underlying the expression difference. Previously, the embryonal carcinoma (EC) cell line, NCCIT, was used by us to study the stromal induction property. In the current study, stromal cells from non-cancer (NP) and cancer (CP) were isolated from tissue specimens and co-cultured with NCCIT cells in a trans-well format to preclude heterotypic cell contact. After 3 days, the stromal cells were analyzed by gene arrays for microRNA (miRNA) and mRNA expression. In co-culture, NCCIT cells were found to alter the miRNA and mRNA expression of NP stromal cells to one like that of CP stromal cells. In contrast, NCCIT had no significant effect on the gene expression of CP stromal cells. We conclude that the gene expression changes in stromal cells can be induced by diffusible factors synthesized by EC cells, and suggest that cancer-associated stromal cells represent a more primitive or less differentiated stromal cell type. PMID:20945389

  13. Differential marker expression by cultures rich in mesenchymal stem cells

    Science.gov (United States)

    2013-01-01

    Background Mesenchymal stem cells have properties that make them amenable to therapeutic use. However, the acceptance of mesenchymal stem cells in clinical practice requires standardized techniques for their specific isolation. To date, there are no conclusive marker (s) for the exclusive isolation of mesenchymal stem cells. Our aim was to identify markers differentially expressed between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. We compared and contrasted the phenotype of tissue cultures in which mesenchymal stem cells are rich and rare. By initially assessing mesenchymal stem cell differentiation, we established that bone marrow and breast adipose cultures are rich in mesenchymal stem cells while, in our hands, foreskin fibroblast and olfactory tissue cultures contain rare mesenchymal stem cells. In particular, olfactory tissue cells represent non-stem cell mesenchymal cells. Subsequently, the phenotype of the tissue cultures were thoroughly assessed using immuno-fluorescence, flow-cytometry, proteomics, antibody arrays and qPCR. Results Our analysis revealed that all tissue cultures, regardless of differentiation potential, demonstrated remarkably similar phenotypes. Importantly, it was also observed that common mesenchymal stem cell markers, and fibroblast-associated markers, do not discriminate between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. Examination and comparison of the phenotypes of mesenchymal stem cell and non-stem cell mesenchymal cell cultures revealed three differentially expressed markers – CD24, CD108 and CD40. Conclusion We indicate the importance of establishing differential marker expression between mesenchymal stem cells and non-stem cell mesenchymal cells in order to determine stem cell specific markers. PMID:24304471

  14. Radiosensitivity of primary tumour cultures as a determinant of curability of human head and neck cancers

    International Nuclear Information System (INIS)

    Peters, L.J.; Tofilon, P.J.; Goepfert, H.; Brock, W.A.

    1989-01-01

    Between November 1985 and November 1987, 31 patients with squamous cell carcinomas of the head and neck who were treated on protocol by surgery and post-operative radiotherapy at the University of Texas M. D. Anderson Cancer Center had radiosensitivity measurements made on primary cultures of the surgical specimens using the Adhesive Tumour Cell Culture System. The parameter of cell survival at 2 Gy (S 2 ) was correlated with the clinical outcome independently of pathological risk factors. All five recurrences have been in patients with S 2 values >0.3 (p = 0.08). Evidence of significant intratumoral heterogeneity of cellular radiosensitivity in vitro was demonstrated in one of four cultures tested. Mathematical modelling suggests that in the absence of marked heterogeneity, the S 2 parameter is likely to be more robust than other radiobiologically based assays in predicting clinical treatment outcome. (author)

  15. Promoting Culturally Respectful Cancer Education Through Digital Storytelling.

    Science.gov (United States)

    Cueva, Melany; Kuhnley, Regina; Lanier, Anne; Dignan, Mark; Revels, Laura; Schoenberg, Nancy E; Cueva, Katie

    Cancer is the leading cause of mortality among Alaska Native people. Over half of Alaska Native people live in rural communities where specially trained community members called Community Health Aides/Practitioners (CHA/Ps) provide health care. In response to CHA/Ps' expressed desire to learn more about cancer, four 5-day cancer education and digital storytelling courses were provided in 2014. Throughout each course, participants explored cancer information, reflected on their personal experiences, and envisioned how they might apply their knowledge within their communities. Each course participant also created a personal and authentic digital story, a methodology increasingly embraced by Indigenous communities as a way to combine storytelling traditions with modern technology to promote both individual and community health. Opportunities to learn of CHA/Ps' experiences with cancer and digital storytelling included a 3-page end-of-course written evaluation, a weekly story-showing log kept for 4 weeks post-course, a group teleconference held 1-2 weeks post-course, and a survey administered 6 months post-course. Participants described digital storytelling as a culturally respectful way to support cancer awareness and education. Participants described the process of creating digital stories as supporting knowledge acquisition, encouraging personal reflection, and sparking a desire to engage in cancer risk reduction activities for themselves and with their families and patients. As a result of creating a personalized digital story, CHA/Ps reported feeling differently about cancer, noting an increase in cancer knowledge and comfort to talk about cancer with clients and family. Indigenous digital stories have potential for broad use as a culturally appropriate health messaging tool.

  16. Protection of cultured mammalian cells by rebamipide

    Energy Technology Data Exchange (ETDEWEB)

    Antoku, Shigetoshi; Aramaki, Ryoji [Kyushu Univ., Fukuoka (Japan). Faculty of Medicine; Tanaka, Hisashi; Kusumoto, Naotoshi

    1997-06-01

    Rebamipide which is used as a drug for gastritis and stomach ulcer has large capability for OH radical scavenging. It is expected that rebamipide has protective effect against ionizing radiations. The present paper deals with protective effect of rebamipide for cultured mammalian cells exposed to ionizing radiations. As rebamipide is insoluble in water, three solvents were used to dissolve. Rebamipide dissolved in dimethyl sulfoxide (DMSO), dimethyl formamide (DMFA) and 0.02 N NaOH was added to the cells in Eagle`s minimum essential medium (MEM) supplemented with 10% fetal calf serum and the cells were irradiated with X-rays. After irradiation, the cells were trypsinized, plated in MEM with 10% fetal calf serum and incubated for 7 days in a CO{sub 2} incubator to form colonies. Rebamipide dissolved in 0.02 N NaOH exhibited the protective effect expected its OH radical scavenging capability. However, the protective effect of rebamipide dissolved in DMSO was about half of that expected by its radical scavenging capability and that of rebamipide dissolved in DMFA was not observed. Uptake of rebamipide labeled with {sup 14}C increased with increasing contact time with rebamipide. These rebamipide mainly distributed in nucleus rather than cytoplasm. (author)

  17. Protection of cultured mammalian cells by rebamipide

    International Nuclear Information System (INIS)

    Antoku, Shigetoshi; Aramaki, Ryoji; Tanaka, Hisashi; Kusumoto, Naotoshi.

    1997-01-01

    Rebamipide which is used as a drug for gastritis and stomach ulcer has large capability for OH radical scavenging. It is expected that rebamipide has protective effect against ionizing radiations. The present paper deals with protective effect of rebamipide for cultured mammalian cells exposed to ionizing radiations. As rebamipide is insoluble in water, three solvents were used to dissolve. Rebamipide dissolved in dimethyl sulfoxide (DMSO), dimethyl formamide (DMFA) and 0.02 N NaOH was added to the cells in Eagle's minimum essential medium (MEM) supplemented with 10% fetal calf serum and the cells were irradiated with X-rays. After irradiation, the cells were trypsinized, plated in MEM with 10% fetal calf serum and incubated for 7 days in a CO 2 incubator to form colonies. Rebamipide dissolved in 0.02 N NaOH exhibited the protective effect expected its OH radical scavenging capability. However, the protective effect of rebamipide dissolved in DMSO was about half of that expected by its radical scavenging capability and that of rebamipide dissolved in DMFA was not observed. Uptake of rebamipide labeled with 14 C increased with increasing contact time with rebamipide. These rebamipide mainly distributed in nucleus rather than cytoplasm. (author)

  18. Adipocyte activation of cancer stem cell signaling in breast cancer

    Institute of Scientific and Technical Information of China (English)

    Benjamin; Wolfson; Gabriel; Eades; Qun; Zhou

    2015-01-01

    Signaling within the tumor microenvironment has a critical role in cancer initiation and progression. Adipocytes, one of the major components of the breast microenvironment,have been shown to provide pro-tumorigenic signals that promote cancer cell proliferation and invasiveness in vitro and tumorigenicity in vivo. Adipocyte secreted factors such as leptin and interleukin-6(IL-6) have a paracrine effect on breast cancer cells. In adipocyte-adjacent breast cancer cells, the leptin and IL-6 signaling pathways activate janus kinase 2/signal transducer and activatorof transcription 5, promoting the epithelial-mesenchymal transition, and upregulating stemness regulators such as Notch, Wnt and the Sex determining region Y-box 2/octamer binding transcription factor 4/Nanog signaling axis. In this review we will summarize the major signaling pathways that regulate cancer stem cells in breast cancer and describe the effects that adipocyte secreted IL-6 and leptin have on breast cancer stem cell signaling. Finally we will introduce a new potential treatment paradigm of inhibiting the adipocyte-breast cancer cell signaling via targeting the IL-6 or leptin pathways.

  19. Ethnicity and cultural models of recovery from breast cancer.

    Science.gov (United States)

    Coreil, Jeannine; Corvin, Jaime A; Nupp, Rebecca; Dyer, Karen; Noble, Charlotte

    2012-01-01

    Recovery narratives describe the culturally shared understandings about the ideal or desirable way to recover from an illness experience. This paper examines ethnic differences in recovery narratives among women participating in breast cancer support groups in Central Florida, USA. It compares groups serving African-American, Latina, and European American women, with the objective of better understanding the appeal of ethnic-specific illness support groups for culturally diverse populations. A mixed-method study design combined qualitative and quantitative measures, including in-depth interviews, participant observation at support group meetings, collection of printed documents, and a structured survey. Core elements of the recovery narrative drew from the dominant societal cancer discourse of optimism and personal transformation through adversity; however, important ethnic differences were evident in the meaning assigned to these themes. Groups gave distinctive salience to themes of faith and spirituality, empowerment through the migration experience, and becoming a better person through the journey of recovery. The findings suggest that ethnic cancer support groups draw upon dominant societal discourses about cancer, but they espouse distinctive recovery narratives that are consonant with the groups' cultural models of illness. Similarity between ethnic members' individual recovery narratives and that of the group may contribute to the appeal of ethnic illness support groups for culturally diverse populations.

  20. The evolution of chicken stem cell culture methods.

    Science.gov (United States)

    Farzaneh, M; Attari, F; Mozdziak, P E; Khoshnam, S E

    2017-12-01

    1. The avian embryo is an excellent model for studying embryology and the production of pharmaceutical proteins in transgenic chickens. Furthermore, chicken stem cells have the potential for proliferation and differentiation and emerged as an attractive tool for various cell-based technologies. 2. The objective of these studies is the derivation and culture of these stem cells is the production of transgenic birds for recombinant biomaterials and vaccine manufacture, drug and cytotoxicity testing, as well as to gain insight into basic science, including cell tracking. 3. Despite similarities among the established chicken stem cell lines, fundamental differences have been reported between their culture conditions and applications. Recent conventional protocols used for expansion and culture of chicken stem cells mostly depend on feeder cells, serum-containing media and static culture. 4. Utilising chicken stem cells for generation of cell-based transgenic birds and a variety of vaccines requires large-scale cell production. However, scaling up the conventional adherent chicken stem cells is challenging and labour intensive. Development of a suspension cell culture process for chicken embryonic stem cells (cESCs), chicken primordial germ cells (PGCs) and chicken induced pluripotent stem cells (ciPSCs) will be an important advance for increasing the growth kinetics of these cells. 6. This review describes various approaches and suggestions to achieve optimal cell growth for defined chicken stem cells cultures and use in future manufacturing applications.

  1. Pancreatic Cancer-Derived Exosomes Cause Paraneoplastic β-cell Dysfunction.

    Science.gov (United States)

    Javeed, Naureen; Sagar, Gunisha; Dutta, Shamit K; Smyrk, Thomas C; Lau, Julie S; Bhattacharya, Santanu; Truty, Mark; Petersen, Gloria M; Kaufman, Randal J; Chari, Suresh T; Mukhopadhyay, Debabrata

    2015-04-01

    Pancreatic cancer frequently causes diabetes. We recently proposed adrenomedullin as a candidate mediator of pancreatic β-cell dysfunction in pancreatic cancer. How pancreatic cancer-derived adrenomedullin reaches β cells remote from the cancer to induce β-cell dysfunction is unknown. We tested a novel hypothesis that pancreatic cancer sheds adrenomedullin-containing exosomes into circulation, which are transported to β cells and impair insulin secretion. We characterized exosomes from conditioned media of pancreatic cancer cell lines (n = 5) and portal/peripheral venous blood of patients with pancreatic cancer (n = 20). Western blot analysis showed the presence of adrenomedullin in pancreatic cancer-exosomes. We determined the effect of adrenomedullin-containing pancreatic cancer exosomes on insulin secretion from INS-1 β cells and human islets, and demonstrated the mechanism of exosome internalization into β cells. We studied the interaction between β-cell adrenomedullin receptors and adrenomedullin present in pancreatic cancer-exosomes. In addition, the effect of adrenomedullin on endoplasmic reticulum (ER) stress response genes and reactive oxygen/nitrogen species generation in β cells was shown. Exosomes were found to be the predominant extracellular vesicles secreted by pancreatic cancer into culture media and patient plasma. Pancreatic cancer-exosomes contained adrenomedullin and CA19-9, readily entered β cells through caveolin-mediated endocytosis or macropinocytosis, and inhibited insulin secretion. Adrenomedullin in pancreatic cancer exosomes interacted with its receptor on β cells. Adrenomedullin receptor blockade abrogated the inhibitory effect of exosomes on insulin secretion. β cells exposed to adrenomedullin or pancreatic cancer exosomes showed upregulation of ER stress genes and increased reactive oxygen/nitrogen species. Pancreatic cancer causes paraneoplastic β-cell dysfunction by shedding adrenomedullin(+)/CA19-9(+) exosomes into

  2. Extracellular Molecules Involved in Cancer Cell Invasion

    International Nuclear Information System (INIS)

    Stivarou, Theodora; Patsavoudi, Evangelia

    2015-01-01

    Nowadays it is perfectly clear that understanding and eradicating cancer cell invasion and metastasis represent the crucial, definitive points in cancer therapeutics. During the last two decades there has been a great interest in the understanding of the extracellular molecular mechanisms involved in cancer cell invasion. In this review, we highlight the findings concerning these processes, focusing in particular on extracellular molecules, including extracellular matrix proteins and their receptors, growth factors and their receptors, matrix metalloproteinases and extracellular chaperones. We report the molecular mechanisms underlying the important contribution of this pool of molecules to the complex, multi-step phenomenon of cancer cell invasion

  3. Extracellular Molecules Involved in Cancer Cell Invasion

    Directory of Open Access Journals (Sweden)

    Theodora Stivarou

    2015-01-01

    Full Text Available Nowadays it is perfectly clear that understanding and eradicating cancer cell invasion and metastasis represent the crucial, definitive points in cancer therapeutics. During the last two decades there has been a great interest in the understanding of the extracellular molecular mechanisms involved in cancer cell invasion. In this review, we highlight the findings concerning these processes, focusing in particular on extracellular molecules, including extracellular matrix proteins and their receptors, growth factors and their receptors, matrix metalloproteinases and extracellular chaperones. We report the molecular mechanisms underlying the important contribution of this pool of molecules to the complex, multi-step phenomenon of cancer cell invasion.

  4. Cultural considerations for South Asian women with breast cancer.

    Science.gov (United States)

    Bedi, Manveen; Devins, Gerald M

    2016-02-01

    Cultural values shape a woman's experience of disease and introduce novel stressors that influence psychosocial needs and adaptation. This literature review examines the psychosocial impact of breast cancer in South Asian women, a large group that has received little attention in this regard. We conducted a comprehensive review of the literature published before April 2014 using Ovid MEDLINE, PsychINFO, PubMED, CINHAL, EMBASE, and Sociological Abstracts. We searched for articles about the psychosocial impact of breast cancer in South Asian women. We retained 23 studies for review. The literature concerning South Asian women's experiences identified culturally linked themes that play significant roles in shaping the illness experience; e.g., stigma and breast cancer, low priority of women's health, collective experience of disease, and religion and spirituality. There is a growing need for culturally sensitive care for South Asian women. By understanding the core cultural values and integrating them into clinical practice, Western healthcare providers may improve the quality of care they deliver and help women to extract the maximum benefit. Developing culturally competent support services may enhance effectiveness in addressing the healthcare needs of South Asian women and may serve other ethnic minorities in North America.

  5. Targeting senescence cells in pancreatic cancer | IDRC ...

    International Development Research Centre (IDRC) Digital Library (Canada)

    Targeting senescence cells in pancreatic cancer. Cellular senescence is a programmed response to oncogenic (tumour-causing) stress that aims to halt the expansion of cells with malignant potential. It does this by stopping the proliferation of pre-cancerous lesions and recruitment of the immune system for their elimination.

  6. Chemo Resistance of Breast Cancer Stem Cells

    Science.gov (United States)

    2007-05-01

    165-72. 60. Vestergaard J, Pedersen MW, Pedersen N, Ensinger C, Tumer Z, Tommerup N, et al. Hedgehog signaling in small-cell lung cancer : frequent......NUMBER Chemo Resistance of Breast Cancer Stem Cells 5b. GRANT NUMBER W81XWH-04-1-0471 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d

  7. Engineered T cells for pancreatic cancer treatment

    Science.gov (United States)

    Katari, Usha L; Keirnan, Jacqueline M; Worth, Anna C; Hodges, Sally E; Leen, Ann M; Fisher, William E; Vera, Juan F

    2011-01-01

    Objective Conventional chemotherapy and radiotherapy produce marginal survival benefits in pancreatic cancer, underscoring the need for novel therapies. The aim of this study is to develop an adoptive T cell transfer approach to target tumours expressing prostate stem cell antigen (PSCA), a tumour-associated antigen that is frequently expressed by pancreatic cancer cells. Methods Expression of PSCA on cell lines and primary tumour samples was confirmed by immunohistochemistry. Healthy donor- and patient-derived T cells were isolated, activated in vitro using CD3/CD28, and transduced with a retroviral vector encoding a chimeric antigen receptor (CAR) targeting PSCA. The ability of these cells to kill tumour cells was analysed by chromium-51 (Cr51) release. Results Prostate stem cell antigen was expressed on >70% of the primary tumour samples screened. Activated, CAR-modified T cells could be readily generated in clinically relevant numbers and were specifically able to kill PSCA-expressing pancreatic cancer cell lines with no non-specific killing of PSCA-negative target cells, thus indicating the potential efficacy and safety of this approach. Conclusions Prostate stem cell antigen is frequently expressed on pancreatic cancer cells and can be targeted for immune-mediated destruction using CAR-modified, adoptively transferred T cells. The safety and efficacy of this approach indicate that it deserves further study and may represent a promising novel treatment for patients with pancreatic cancer. PMID:21843265

  8. An immunosurveillance mechanism controls cancer cell ploidy.

    Science.gov (United States)

    Senovilla, Laura; Vitale, Ilio; Martins, Isabelle; Tailler, Maximilien; Pailleret, Claire; Michaud, Mickaël; Galluzzi, Lorenzo; Adjemian, Sandy; Kepp, Oliver; Niso-Santano, Mireia; Shen, Shensi; Mariño, Guillermo; Criollo, Alfredo; Boilève, Alice; Job, Bastien; Ladoire, Sylvain; Ghiringhelli, François; Sistigu, Antonella; Yamazaki, Takahiro; Rello-Varona, Santiago; Locher, Clara; Poirier-Colame, Vichnou; Talbot, Monique; Valent, Alexander; Berardinelli, Francesco; Antoccia, Antonio; Ciccosanti, Fabiola; Fimia, Gian Maria; Piacentini, Mauro; Fueyo, Antonio; Messina, Nicole L; Li, Ming; Chan, Christopher J; Sigl, Verena; Pourcher, Guillaume; Ruckenstuhl, Christoph; Carmona-Gutierrez, Didac; Lazar, Vladimir; Penninger, Josef M; Madeo, Frank; López-Otín, Carlos; Smyth, Mark J; Zitvogel, Laurence; Castedo, Maria; Kroemer, Guido

    2012-09-28

    Cancer cells accommodate multiple genetic and epigenetic alterations that initially activate intrinsic (cell-autonomous) and extrinsic (immune-mediated) oncosuppressive mechanisms. Only once these barriers to oncogenesis have been overcome can malignant growth proceed unrestrained. Tetraploidization can contribute to oncogenesis because hyperploid cells are genomically unstable. We report that hyperploid cancer cells become immunogenic because of a constitutive endoplasmic reticulum stress response resulting in the aberrant cell surface exposure of calreticulin. Hyperploid, calreticulin-exposing cancer cells readily proliferated in immunodeficient mice and conserved their increased DNA content. In contrast, hyperploid cells injected into immunocompetent mice generated tumors only after a delay, and such tumors exhibited reduced DNA content, endoplasmic reticulum stress, and calreticulin exposure. Our results unveil an immunosurveillance system that imposes immunoselection against hyperploidy in carcinogen- and oncogene-induced cancers.

  9. Are cancer cells really softer than normal cells?

    Science.gov (United States)

    Alibert, Charlotte; Goud, Bruno; Manneville, Jean-Baptiste

    2017-05-01

    Solid tumours are often first diagnosed by palpation, suggesting that the tumour is more rigid than its surrounding environment. Paradoxically, individual cancer cells appear to be softer than their healthy counterparts. In this review, we first list the physiological reasons indicating that cancer cells may be more deformable than normal cells. Next, we describe the biophysical tools that have been developed in recent years to characterise and model cancer cell mechanics. By reviewing the experimental studies that compared the mechanics of individual normal and cancer cells, we argue that cancer cells can indeed be considered as softer than normal cells. We then focus on the intracellular elements that could be responsible for the softening of cancer cells. Finally, we ask whether the mechanical differences between normal and cancer cells can be used as diagnostic or prognostic markers of cancer progression. © 2017 Société Française des Microscopies and Société de Biologie Cellulaire de France. Published by John Wiley & Sons Ltd.

  10. Dual Functional Capability of Dendritic Cells - Cytokine-Induced Killer Cells in Improving Side Effects of Colorectal Cancer Therapy.

    Science.gov (United States)

    Mosińska, Paula; Gabryelska, Agata; Zasada, Malwina; Fichna, Jakub

    2017-01-01

    The aim of cancer therapy is to eradicate cancer without affecting healthy tissues. Current options available for treating colorectal cancer (CRC), including surgery, chemotherapy or radiotherapy, usually elicit multiple adverse effects and frequently fail to completely remove the tumor cells. Thus, there is a constant need for seeking cancer cell-specific therapeutics to improve the course of cancer therapy and reduce the risk of relapse. In this review we elaborate on the mechanisms underlying the immunotherapy with dendritic cells (DCs) and cytokine-induced killer (CIK) cells, and summarize their effectiveness and tolerability available clinical studies. Finally, we discuss the up-to-date combinatorial adoptive anti-cancer immunotherapy with CIK cells co-cultured with DCs that recently showed encouraging efficacy and usefulness in treating malignant disease, including CRC.

  11. Triiodothyronine regulates cell growth and survival in renal cell cancer.

    Science.gov (United States)

    Czarnecka, Anna M; Matak, Damian; Szymanski, Lukasz; Czarnecka, Karolina H; Lewicki, Slawomir; Zdanowski, Robert; Brzezianska-Lasota, Ewa; Szczylik, Cezary

    2016-10-01

    Triiodothyronine plays an important role in the regulation of kidney cell growth, differentiation and metabolism. Patients with renal cell cancer who develop hypothyreosis during tyrosine kinase inhibitor (TKI) treatment have statistically longer survival. In this study, we developed cell based model of triiodothyronine (T3) analysis in RCC and we show the different effects of T3 on renal cell cancer (RCC) cell growth response and expression of the thyroid hormone receptor in human renal cell cancer cell lines from primary and metastatic tumors along with human kidney cancer stem cells. Wild-type thyroid hormone receptor is ubiquitously expressed in human renal cancer cell lines, but normalized against healthy renal proximal tube cell expression its level is upregulated in Caki-2, RCC6, SKRC-42, SKRC-45 cell lines. On the contrary the mRNA level in the 769-P, ACHN, HKCSC, and HEK293 cells is significantly decreased. The TRβ protein was abundant in the cytoplasm of the 786-O, Caki-2, RCC6, and SKRC-45 cells and in the nucleus of SKRC-42, ACHN, 769-P and cancer stem cells. T3 has promoting effect on the cell proliferation of HKCSC, Caki-2, ASE, ACHN, SK-RC-42, SMKT-R2, Caki-1, 786-0, and SK-RC-45 cells. Tyrosine kinase inhibitor, sunitinib, directly inhibits proliferation of RCC cells, while thyroid hormone receptor antagonist 1-850 (CAS 251310‑57-3) has less significant inhibitory impact. T3 stimulation does not abrogate inhibitory effect of sunitinib. Renal cancer tumor cells hypostimulated with T3 may be more responsive to tyrosine kinase inhibition. Moreover, some tumors may be considered as T3-independent and present aggressive phenotype with thyroid hormone receptor activated independently from the ligand. On the contrary proliferation induced by deregulated VHL and or c-Met pathways may transgress normal T3 mediated regulation of the cell cycle.

  12. Taraxacum officinale dandelion extract efficiently inhibited the breast cancer stem cell proliferation

    OpenAIRE

    Ngu Van Trinh; Nghi Doan-Phuong Dang; Diem Hong Tran; Phuc Van Pham

    2016-01-01

    Background: Breast cancer stem cells (BCSCs) play an important role in breast cancer initiation, metastasis, recurrence, and drug resistance. Therefore, targeting BCSCs is an essential strategy to suppress cancer growth. This study aimed to evaluate the effects of dandelion Taraxacum officinale extracts on BCSC proliferation in vitro in 2D and 3D cell culture platforms. Materials and Methods: The BCSCs were maintained under standard conditions, verified for expression of CD44 and CD24 surface...

  13. X-ray microanalysis of single and cultured cells

    International Nuclear Information System (INIS)

    Wroblewski, J.; Roomans, G.M.

    1984-01-01

    X-ray microanalysis of single or cultured cells is often a useful alternative or complement to the analysis of the corresponding tissue. It also allows the analysis of individual cells in a cell population. Preparation for X-ray microanalysis poses a number of typical problems. Suspensions of single cells can be prepared by either of two pathways: (1) washing - mounting - drying, or (2) centrifugation - freezing or fixation - sectioning. The washing step in the preparation of single or cultured cells presents the most severe problems. Cultured cells are generally grown on a substrate that is compatible with both the analysis and the culture, washed and dried. In some cases, sectioning of cultured cell monolayers has been performed. Special problems in quantitative analysis occur in those cases where the cells are analyzed on a thick substrate, since the substrate contributes to the spectral background

  14. Establishment and characterization of American elm cell suspension cultures

    Science.gov (United States)

    Steven M. Eshita; Joseph C. Kamalay; Vicki M. Gingas; Daniel A. Yaussy

    2000-01-01

    Cell suspension cultures of Dutch elm disease (DED)-tolerant and DED-susceptible American elms clones have been established and characterized as prerequisites for contrasts of cellular responses to pathogen-derived elicitors. Characteristics of cultured elm cell growth were monitored by A700 and media conductivity. Combined cell growth data for all experiments within a...

  15. A 3D Microfluidic Model to Recapitulate Cancer Cell Migration and Invasion

    Directory of Open Access Journals (Sweden)

    Yi-Chin Toh

    2018-04-01

    Full Text Available We have developed a microfluidic-based culture chip to simulate cancer cell migration and invasion across the basement membrane. In this microfluidic chip, a 3D microenvironment is engineered to culture metastatic breast cancer cells (MX1 in a 3D tumor model. A chemo-attractant was incorporated to stimulate motility across the membrane. We validated the usefulness of the chip by tracking the motilities of the cancer cells in the system, showing them to be migrating or invading (akin to metastasis. It is shown that our system can monitor cell migration in real time, as compare to Boyden chambers, for example. Thus, the chip will be of interest to the drug-screening community as it can potentially be used to monitor the behavior of cancer cell motility, and, therefore, metastasis, in the presence of anti-cancer drugs.

  16. Metformin is synthetically lethal with glucose withdrawal in cancer cells.

    Science.gov (United States)

    Menendez, Javier A; Oliveras-Ferraros, Cristina; Cufí, Sílvia; Corominas-Faja, Bruna; Joven, Jorge; Martin-Castillo, Begoña; Vazquez-Martin, Alejandro

    2012-08-01

    Glucose deprivation is a distinctive feature of the tumor microecosystem caused by the imbalance between poor supply and an extraordinarily high consumption rate. The metabolic reprogramming from mitochondrial respiration to aerobic glycolysis in cancer cells (the "Warburg effect") is linked to oncogenic transformation in a manner that frequently implies the inactivation of metabolic checkpoints such as the energy rheostat AMP-activated protein kinase (AMPK). Because the concept of synthetic lethality in oncology can be applied not only to genetic and epigenetic intrinsic differences between normal and cancer cells but also to extrinsic ones such as altered microenvironment, we recently hypothesized that stress-energy mimickers such as the AMPK agonist metformin should produce metabolic synthetic lethality in a glucose-starved cell culture milieu imitating the adverse tumor growth conditions in vivo. Under standard high-glucose conditions, metformin supplementation mostly caused cell cycle arrest without signs of apoptotic cell death. Under glucose withdrawal stress, metformin supplementation circumvented the ability of oncogenes (e.g., HER2) to protect breast cancer cells from glucose-deprivation apoptosis. Significantly, representative cell models of breast cancer heterogeneity underwent massive apoptosis (by >90% in some cases) when glucose-starved cell cultures were supplemented with metformin. Our current findings may uncover crucial issues regarding the cell-autonomous metformin's anti-cancer actions: (1) The offently claimed clinically irrelevant, non-physiological concentrations needed to observe the metformin's anti-cancer effects in vitro merely underlie the artifactual interference of erroneous glucose-rich experimental conditions that poorly reflect glucose-starved in vivo conditions; (2) the preferential killing of cancer stem cells (CSC) by metformin may simply expose the best-case scenario for its synthetically lethal activity because an increased

  17. Cancer stem cells in head and neck cancer

    Directory of Open Access Journals (Sweden)

    Trapasso S

    2012-11-01

    Full Text Available Eugenia Allegra, Serena TrapassoOtolaryngology – Head and Neck Surgery, University Magna Graecia of Catanzaro, Catanzaro, ItalyAbstract: Cancer stem cells (CSCs, also called "cells that start the tumor," represent in themselves one of the most topical and controversial issues in the field of cancer research. Tumor stem cells are able to self-propagate in vitro (self-renewal, giving rise both to other tumor stem cells and most advanced cells in the line of differentiation (asymmetric division. A final characteristic is tumorigenicity, a fundamental property, which outlines the tumor stem cell as the only cell able to initiate the formation of a tumor when implanted in immune-deficient mice. The hypothesis of a hierarchical organization of tumor cells dates back more than 40 years, but only in 1997, thanks to the work of John Dick and Dominique Bonnet, was there the formal proof of such an organization in acute myeloid leukemia. Following this, many other research groups were able to isolate CSCs, by appropriate selection markers, in various malignancies, such as breast, brain, colon, pancreas, and liver cancers and in melanoma. To date, however, it is not possible to isolate stem cells from all types of neoplasia, particularly in solid tumors. From a therapeutic point of view, the concept of tumor stem cells implies a complete revision of conventional antineoplastic treatment. Conventional cytotoxic agents are designed to target actively proliferating cells. In the majority of cases, this is not sufficient to eliminate the CSCs, which thanks to their reduced proliferative activity and/or the presence of proteins capable of extruding chemotherapeutics from the cell are not targeted. Therefore, the theory of cancer stem cells can pose new paradigms in terms of cancer treatment. Potential approaches, even in the very early experimental stages, relate to the selective inhibition of pathways connected with self-renewal, or more specifically based on

  18. The influence of micronutrients in cell culture: a reflection on viability and genomic stability.

    Science.gov (United States)

    Arigony, Ana Lúcia Vargas; de Oliveira, Iuri Marques; Machado, Miriana; Bordin, Diana Lilian; Bergter, Lothar; Prá, Daniel; Henriques, João Antonio Pêgas

    2013-01-01

    Micronutrients, including minerals and vitamins, are indispensable to DNA metabolic pathways and thus are as important for life as macronutrients. Without the proper nutrients, genomic instability compromises homeostasis, leading to chronic diseases and certain types of cancer. Cell-culture media try to mimic the in vivo environment, providing in vitro models used to infer cells' responses to different stimuli. This review summarizes and discusses studies of cell-culture supplementation with micronutrients that can increase cell viability and genomic stability, with a particular focus on previous in vitro experiments. In these studies, the cell-culture media include certain vitamins and minerals at concentrations not equal to the physiological levels. In many common culture media, the sole source of micronutrients is fetal bovine serum (FBS), which contributes to only 5-10% of the media composition. Minimal attention has been dedicated to FBS composition, micronutrients in cell cultures as a whole, or the influence of micronutrients on the viability and genetics of cultured cells. Further studies better evaluating micronutrients' roles at a molecular level and influence on the genomic stability of cells are still needed.

  19. Transitions from mono- to co- to tri-culture uniquely affect gene expression in breast cancer, stromal, and immune compartments.

    Science.gov (United States)

    Regier, Mary C; Maccoux, Lindsey J; Weinberger, Emma M; Regehr, Keil J; Berry, Scott M; Beebe, David J; Alarid, Elaine T

    2016-08-01

    Heterotypic interactions in cancer microenvironments play important roles in disease initiation, progression, and spread. Co-culture is the predominant approach used in dissecting paracrine interactions between tumor and stromal cells, but functional results from simple co-cultures frequently fail to correlate to in vivo conditions. Though complex heterotypic in vitro models have improved functional relevance, there is little systematic knowledge of how multi-culture parameters influence this recapitulation. We therefore have employed a more iterative approach to investigate the influence of increasing model complexity; increased heterotypic complexity specifically. Here we describe how the compartmentalized and microscale elements of our multi-culture device allowed us to obtain gene expression data from one cell type at a time in a heterotypic culture where cells communicated through paracrine interactions. With our device we generated a large dataset comprised of cell type specific gene-expression patterns for cultures of increasing complexity (three cell types in mono-, co-, or tri-culture) not readily accessible in other systems. Principal component analysis indicated that gene expression was changed in co-culture but was often more strongly altered in tri-culture as compared to mono-culture. Our analysis revealed that cell type identity and the complexity around it (mono-, co-, or tri-culture) influence gene regulation. We also observed evidence of complementary regulation between cell types in the same heterotypic culture. Here we demonstrate the utility of our platform in providing insight into how tumor and stromal cells respond to microenvironments of varying complexities highlighting the expanding importance of heterotypic cultures that go beyond conventional co-culture.

  20. Ionizing radiation induces stemness in cancer cells.

    Directory of Open Access Journals (Sweden)

    Laura Ghisolfi

    Full Text Available The cancer stem cell (CSC model posits the presence of a small number of CSCs in the heterogeneous cancer cell population that are ultimately responsible for tumor initiation, as well as cancer recurrence and metastasis. CSCs have been isolated from a variety of human cancers and are able to generate a hierarchical and heterogeneous cancer cell population. CSCs are also resistant to conventional chemo- and radio-therapies. Here we report that ionizing radiation can induce stem cell-like properties in heterogeneous cancer cells. Exposure of non-stem cancer cells to ionizing radiation enhanced spherogenesis, and this was accompanied by upregulation of the pluripotency genes Sox2 and Oct3/4. Knockdown of Sox2 or Oct3/4 inhibited radiation-induced spherogenesis and increased cellular sensitivity to radiation. These data demonstrate that ionizing radiation can activate stemness pathways in heterogeneous cancer cells, resulting in the enrichment of a CSC subpopulation with higher resistance to radiotherapy.

  1. Electrospinning of microbial polyester for cell culture

    Energy Technology Data Exchange (ETDEWEB)

    Kwon, Oh Hyeong [Department of Polymer Science and Engineering, Kumoh National Institute of Technology, 1 Yangho-dong, Gumi, Gyeongbuk 730-701 (Korea, Republic of); Lee, Ik Sang [Department of Polymer Science and Engineering, Kumoh National Institute of Technology, 1 Yangho-dong, Gumi, Gyeongbuk 730-701 (Korea, Republic of); Ko, Young-Gwang [Department of Polymer Science and Engineering, Kumoh National Institute of Technology, 1 Yangho-dong, Gumi, Gyeongbuk 730-701 (Korea, Republic of); Meng, Wan [Department of Polymer Science, Kyungpook National University, 1370 Sankyuk-dong, Buk-gu, Daegu 702-701 (Korea, Republic of); Jung, Kyung-Hye [Department of Polymer Science, Kyungpook National University, 1370 Sankyuk-dong, Buk-gu, Daegu 702-701 (Korea, Republic of); Kang, Inn-Kyu [Department of Polymer Science, Kyungpook National University, 1370 Sankyuk-dong, Buk-gu, Daegu 702-701 (Korea, Republic of); Ito, Yoshihiro [Kanagawa Academy of Science and Technology, KSP East 309, Sakado 3-2-1, Takatsu-ku, Kawasaki 213-0012 (Japan)

    2007-03-01

    Biodegradable and biocompatible poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), a copolymer of microbial polyester, was fabricated as a nanofibrous mat by electrospinning. The specific surface area and the porosity of electrospun PHBV nanofibrous mat were determined. When the mechanical properties of flat film and electrospun PHBV nanofibrous mats were investigated, both the tensile modulus and strength of electrospun PHBV were less than those of cast PHBV film. However, the elongation ratio of nanofiber mat was higher than that of the cast film. The structure of electrospun nanofibers using PHBV-trifluoroethanol solutions depended on the solution concentrations. When x-ray diffraction patterns of bulk PHBV before and after electrospinning were compared, the crystallinity of PHBV was not significantly affected by the electrospinning process. Chondrocytes adhered and grew on the electrospun PHBV nanofibrous mat better than on the cast PHBV film. Therefore, the electrospun PHBV was considered to be suitable for cell culture.

  2. Drug-resistant colon cancer cells produce high carcinoembryonic antigen and might not be cancer-initiating cells

    Science.gov (United States)

    Lee, Hsin-chung; Ling, Qing-Dong; Yu, Wan-Chun; Hung, Chunh-Ming; Kao, Ta-Chun; Huang, Yi-Wei; Higuchi, Akon

    2013-01-01

    Purpose We evaluated the higher levels of carcinoembryonic antigen (CEA) secreted by the LoVo human colon carcinoma cells in a medium containing anticancer drugs. Drug-resistant LoVo cells were analyzed by subcutaneously xenotransplanting them into mice. The aim of this study was to evaluate whether the drug-resistant cells isolated in this study were cancer-initiating cells, known also as cancer stem cells (CSCs). Methods The production of CEA was investigated in LoVo cells that were cultured with 0–10 mM of anticancer drugs, and we evaluated the increase in CEA production by the LoVo cells that were stimulated by anticancer drug treatment. The expression of several CSC markers in LoVo cells treated with anticancer drugs was also evaluated. Following anticancer drug treatment, LoVo cells were injected subcutaneously into the flanks of severe combined immunodeficiency mice in order to evaluate the CSC fraction. Results Production of CEA by LoVo cells was stimulated by the addition of anticancer drugs. Drug-resistant LoVo cells expressed lower levels of CSC markers, and LoVo cells treated with any of the anticancer drugs tested did not generate tumors within 8 weeks from when the cells were injected subcutaneously into severe combined immunodeficiency mice. These results suggest that the drug-resistant LoVo cells have a smaller population of CSCs than the untreated LoVo cells. Conclusion Production of CEA by LoVo cells can be stimulated by the addition of anticancer drugs. The drug-resistant subpopulation of LoVo colon cancer cells could stimulate the production of CEA, but these cells did not act as CSCs in in vivo tumor generation experiments. PMID:23818760

  3. A stem cell medium containing neural stimulating factor induces a pancreatic cancer stem-like cell-enriched population

    Science.gov (United States)

    WATANABE, YUSAKU; YOSHIMURA, KIYOSHI; YOSHIKAWA, KOICHI; TSUNEDOMI, RYOICHI; SHINDO, YOSHITARO; MATSUKUMA, SOU; MAEDA, NORIKO; KANEKIYO, SHINSUKE; SUZUKI, NOBUAKI; KURAMASU, ATSUO; SONODA, KOUHEI; TAMADA, KOJI; KOBAYASHI, SEI; SAYA, HIDEYUKI; HAZAMA, SHOICHI; OKA, MASAAKI

    2014-01-01

    Cancer stem cells (CSCs) have been studied for their self-renewal capacity and pluripotency, as well as their resistance to anticancer therapy and their ability to metastasize to distant organs. CSCs are difficult to study because their population is quite low in tumor specimens. To overcome this problem, we established a culture method to induce a pancreatic cancer stem-like cell (P-CSLC)-enriched population from human pancreatic cancer cell lines. Human pancreatic cancer cell lines established at our department were cultured in CSC-inducing media containing epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), leukemia inhibitory factor (LIF), neural cell survivor factor-1 (NSF-1), and N-acetylcysteine. Sphere cells were obtained and then transferred to a laminin-coated dish and cultured for approximately two months. The surface markers, gene expression, aldehyde dehydrogenase (ALDH) activity, cell cycle, and tumorigenicity of these induced cells were examined for their stem cell-like characteristics. The population of these induced cells expanded within a few months. The ratio of CD24high, CD44high, epithelial specific antigen (ESA) high, and CD44variant (CD44v) high cells in the induced cells was greatly enriched. The induced cells stayed in the G0/G1 phase and demonstrated mesenchymal and stemness properties. The induced cells had high tumorigenic potential. Thus, we established a culture method to induce a P-CSLCenriched population from human pancreatic cancer cell lines. The CSLC population was enriched approximately 100-fold with this method. Our culture method may contribute to the precise analysis of CSCs and thus support the establishment of CSC-targeting therapy. PMID:25118635

  4. Stimulation of the proliferation of hemopoietic stem cells in irradiated bone marrow cell culture

    International Nuclear Information System (INIS)

    Mori, K.J.; Izumi, H.; Seto, A.

    1981-01-01

    Long-term hemopoiesis was established in bone marrow cell culture in vitro. This culture was shown to support the recovery proliferation of hemopoietic stem cells completely in vitro after irradiation. Hemopoietic stem cells were stimulated into proliferation in culture when normal bone marrow cells were overlayed on top of the irradiated adherent cell colonies. These results indicate that proliferation and differentiation of hemopoietic stem cells in vitro are also supported by stromahemopoietic cell interactions

  5. Sphere-forming cell subpopulations with cancer stem cell properties in human hepatoma cell lines

    Directory of Open Access Journals (Sweden)

    Chen Lei

    2011-06-01

    Full Text Available Abstract Background Cancer stem cells (CSCs are regarded as the cause of tumor formation and recurrence. The isolation and identification of CSCs could help to develop novel therapeutic strategies specifically targeting CSCs. Methods Human hepatoma cell lines were plated in stem cell conditioned culture system allowed for sphere forming. To evaluate the stemness characteristics of spheres, the self-renewal, proliferation, chemoresistance, tumorigenicity of the PLC/PRF/5 sphere-forming cells, and the expression levels of stem cell related proteins in the PLC/PRF/5 sphere-forming cells were assessed, comparing with the parental cells. The stem cell RT-PCR array was performed to further explore the biological properties of liver CSCs. Results The PLC/PRF/5, MHCC97H and HepG2 cells could form clonal nonadherent 3-D spheres and be serially passaged. The PLC/PRF/5 sphere-forming cells possessed a key criteria that define CSCs: persistent self-renewal, extensive proliferation, drug resistance, overexpression of liver CSCs related proteins (Oct3/4, OV6, EpCAM, CD133 and CD44. Even 500 sphere-forming cells were able to form tumors in NOD/SCID mice, and the tumor initiating capability was not decreased when spheres were passaged. Besides, downstream proteins DTX1 and Ep300 of the CSL (CBF1 in humans, Suppressor of hairless in Drosophila and LAG1 in C. elegans -independent Notch signaling pathway were highly expressed in the spheres, and a gamma-secretase inhibitor MRK003 could significantly inhibit the sphere formation ability. Conclusions Nonadherent tumor spheres from hepatoma cell lines cultured in stem cell conditioned medium possess liver CSC properties, and the CSL-independent Notch signaling pathway may play a role in liver CSCs.

  6. Biomimetic brain tumor niche regulates glioblastoma cells towards a cancer stem cell phenotype.

    Science.gov (United States)

    Liu, Yung-Chiang; Lee, I-Chi; Chen, Pin-Yuan

    2018-05-01

    Glioblastoma (GBM) is the most malignant primary brain tumor and contains tumorigenic cancer stem cells (CSCs), which support the progression of tumor growth. The selection of CSCs and facilitation of the brain tumor niches may assist the development of novel therapeutics for GBM. Herein, hydrogel materials composed of agarose and hydroxypropyl methyl cellulose (HMC) in different concentrations were established and compared to emulate brain tumor niches and CSC microenvironments within a label-free system. Human GBM cell line, U-87 MG, was cultured on a series of HMC-agarose based culture system. Cell aggregation and spheroids formation were investigated after 4 days of culture, and 2.5% HMC-agarose based culture system demonstrated the largest spheroids number and size. Moreover, CD133 marker expression of GBM cells after 6 days of culture in 2.5% HMC-agarose based culture system was 60%, relatively higher than the control group at only 15%. Additionally, cells on 2.5% HMC-agarose based culture system show the highest chemoresistance, even at the high dose of 500 µM temozolomide for 72 h, the live cell ratio was still > 80%. Furthermore, the results also indicate that the expression of ABCG2 gene was up-regulated after culture in 2.5% HMC-agarose based culture system. Therefore, our results demonstrated that biomimetic brain tumor microenvironment may regulate GBM cells towards the CSC phenotype and expression of CSC characteristics. The microenvironment selection and spheroids formation in HMC-agarose based culture system may provide a label-free CSC selection strategy and drug testing model for future biomedical applications.

  7. Acetylcholine release by human colon cancer cells mediates autocrine stimulation of cell proliferation.

    Science.gov (United States)

    Cheng, Kunrong; Samimi, Roxana; Xie, Guofeng; Shant, Jasleen; Drachenberg, Cinthia; Wade, Mark; Davis, Richard J; Nomikos, George; Raufman, Jean-Pierre

    2008-09-01

    Most colon cancers overexpress M3 muscarinic receptors (M3R), and post-M3R signaling stimulates human colon cancer cell proliferation. Acetylcholine (ACh), a muscarinic receptor ligand traditionally regarded as a neurotransmitter, may be produced by nonneuronal cells. We hypothesized that ACh release by human colon cancer cells results in autocrine stimulation of proliferation. H508 human colon cancer cells, which have robust M3R expression, were used to examine effects of muscarinic receptor antagonists, acetylcholinesterase inhibitors, and choline transport inhibitors on cell proliferation. A nonselective muscarinic receptor antagonist (atropine), a selective M3R antagonist (p-fluorohexahydro-sila-difenidol hydrochloride), and a choline transport inhibitor (hemicholinum-3) all inhibited unstimulated H508 colon cancer cell proliferation by approximately 40% (P<0.005). In contrast, two acetylcholinesterase inhibitors (eserine-hemisulfate and bis-9-amino-1,2,3,4-tetrahydroacridine) increased proliferation by 2.5- and 2-fold, respectively (P<0.005). By using quantitative real-time PCR, expression of choline acetyltransferase (ChAT), a critical enzyme for ACh synthesis, was identified in H508, WiDr, and Caco-2 colon cancer cells. By using high-performance liquid chromatography-electrochemical detection, released ACh was detected in H508 and Caco-2 cell culture media. Immunohistochemistry in surgical specimens revealed weak or no cytoplasmic staining for ChAT in normal colon enterocytes (n=25) whereas half of colon cancer specimens (n=24) exhibited moderate to strong staining (P<0.005). We conclude that ACh is an autocrine growth factor in colon cancer. Mechanisms that regulate colon epithelial cell production and release of ACh warrant further investigation.

  8. Regulatory T Cells in Human Ovarian Cancer

    Directory of Open Access Journals (Sweden)

    Dong-Jun Peng

    2012-01-01

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

  9. Cancer Cell Metabolism: One Hallmark, Many Faces

    OpenAIRE

    Cantor, Jason R.; Sabatini, David M.

    2012-01-01

    Cancer cells must rewire cellular metabolism to satisfy the demands of growth and proliferation. Although many of the metabolic alterations are largely similar to those in normal proliferating cells, they are aberrantly driven in cancer by a combination of genetic lesions and nongenetic factors such as the tumor microenvironment. However, a single model of altered tumor metabolism does not describe the sum of metabolic changes that can support cell growth. Instead, the diversity of such chang...

  10. T cell recognition of breast cancer antigens

    DEFF Research Database (Denmark)

    Petersen, Nadia Viborg; Andersen, Sofie Ramskov; Andersen, Rikke Sick

    Recent studies are encouraging research of breast cancer immunogenicity to evaluate the applicability ofimmunotherapy as a treatment strategy. The epitope landscape in breast cancer is minimally described, thus it is necessary to identify T cell targets to develop immune mediated therapies.......This project investigates four proteins commonly upregulated in breast cancer and thus probable tumor associated antigens (TAAs). Aromatase, prolactin, NEK3, and PIAS3 contribute to increase growth, survival, and motility of malignant cells. Aspiring to uncover novel epitopes for cytotoxic T cells, a reverse...... recognition utilizing DNA barcode labeled MHC multimers to screen peripheral blood lymphocytes from breast cancer patients and healthy donor samples. Signif-icantly more TAA specific T cell responses were detected in breast cancer patients than healthy donors for both HLA-A*0201 (P

  11. Particle Trajectories in Rotating Wall Cell Culture Devices

    Science.gov (United States)

    Ramachandran N.; Downey, J. P.

    1999-01-01

    Cell cultures are extremely important to the medical community since such cultures provide an opportunity to perform research on human tissue without the concerns inherent in experiments on individual humans. Development of cells in cultures has been found to be greatly influenced by the conditions of the culture. Much work has focused on the effect of the motions of cells in the culture relative to the solution. Recently rotating wall vessels have been used with success in achieving improved cellular cultures. Speculation and limited research have focused on the low shear environment and the ability of rotating vessels to keep cells suspended in solution rather than floating or sedimenting as the primary reasons for the improved cellular cultures using these devices. It is widely believed that the cultures obtained using a rotating wall vessel simulates to some degree the effect of microgravity on cultures. It has also been speculated that the microgravity environment may provide the ideal acceleration environment for culturing of cellular tissues due to the nearly negligible levels of sedimentation and shear possible. This work predicts particle trajectories of cells in rotating wall vessels of cylindrical and annular design consistent with the estimated properties of typical cellular cultures. Estimates of the shear encountered by cells in solution and the interactions with walls are studied. Comparisons of potential experiments in ground and microgravity environments are performed.

  12. Cancer stem cells in colorectal cancer: a review.

    Science.gov (United States)

    Munro, Matthew J; Wickremesekera, Susrutha K; Peng, Lifeng; Tan, Swee T; Itinteang, Tinte

    2018-02-01

    Colorectal cancer (CRC) is the second most common cancer in women and the third most common in men. Adenocarcinoma accounts for 90% of CRC cases. There has been accumulating evidence in support of the cancer stem cell (CSC) concept of cancer which proposes that CSCs are central in the initiation of cancer. CSCs have been the focus of study in a range of cancers, including CRC. This has led to the identification and understanding of genes involved in the induction and maintenance of pluripotency of stem cells, and markers for CSCs, including those investigated specifically in CRC. Knowledge of the expression pattern of CSCs in CRC has been increasing in recent years, revealing a heterogeneous population of cells within CRC ranging from pluripotent to differentiated cells, with overlapping and sometimes unique combinations of markers. This review summarises current literature on the understanding of CSCs in CRC, including evidence of the presence of CSC subpopulations, and the stem cell markers currently used to identify and localise these CSC subpopulations. Future research into this field may lead to improved methods for early detection of CRC, novel therapy and monitoring of treatment for CRC and other cancer types. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  13. Microfluidic engineered high cell density three-dimensional neural cultures

    Science.gov (United States)

    Cullen, D. Kacy; Vukasinovic, Jelena; Glezer, Ari; La Placa, Michelle C.

    2007-06-01

    Three-dimensional (3D) neural cultures with cells distributed throughout a thick, bioactive protein scaffold may better represent neurobiological phenomena than planar correlates lacking matrix support. Neural cells in vivo interact within a complex, multicellular environment with tightly coupled 3D cell-cell/cell-matrix interactions; however, thick 3D neural cultures at cell densities approaching that of brain rapidly decay, presumably due to diffusion limited interstitial mass transport. To address this issue, we have developed a novel perfusion platform that utilizes forced intercellular convection to enhance mass transport. First, we demonstrated that in thick (>500 µm) 3D neural cultures supported by passive diffusion, cell densities =104 cells mm-3), continuous medium perfusion at 2.0-11.0 µL min-1 improved viability compared to non-perfused cultures (p death and matrix degradation. In perfused cultures, survival was dependent on proximity to the perfusion source at 2.00-6.25 µL min-1 (p 90% viability in both neuronal cultures and neuronal-astrocytic co-cultures. This work demonstrates the utility of forced interstitial convection in improving the survival of high cell density 3D engineered neural constructs and may aid in the development of novel tissue-engineered systems reconstituting 3D cell-cell/cell-matrix interactions.

  14. Matrix rigidity regulates cancer cell growth and cellular phenotype.

    Directory of Open Access Journals (Sweden)

    Robert W Tilghman

    2010-09-01

    Full Text Available The mechanical properties of the extracellular matrix have an important role in cell growth and differentiation. However, it is unclear as to what extent cancer cells respond to changes in the mechanical properties (rigidity/stiffness of the microenvironment and how this response varies among cancer cell lines.In this study we used a recently developed 96-well plate system that arrays extracellular matrix-conjugated polyacrylamide gels that increase in stiffness by at least 50-fold across the plate. This plate was used to determine how changes in the rigidity of the extracellular matrix modulate the biological properties of tumor cells. The cell lines tested fall into one of two categories based on their proliferation on substrates of differing stiffness: "rigidity dependent" (those which show an increase in cell growth as extracellular rigidity is increased, and "rigidity independent" (those which grow equally on both soft and stiff substrates. Cells which grew poorly on soft gels also showed decreased spreading and migration under these conditions. More importantly, seeding the cell lines into the lungs of nude mice revealed that the ability of cells to grow on soft gels in vitro correlated with their ability to grow in a soft tissue environment in vivo. The lung carcinoma line A549 responded to culture on soft gels by expressing the differentiated epithelial marker E-cadherin and decreasing the expression of the mesenchymal transcription factor Slug.These observations suggest that the mechanical properties of the matrix environment play a significant role in regulating the proliferation and the morphological properties of cancer cells. Further, the multiwell format of the soft-plate assay is a useful and effective adjunct to established 3-dimensional cell culture models.

  15. Matrix Rigidity Regulates Cancer Cell Growth and Cellular Phenotype

    Science.gov (United States)

    Tilghman, Robert W.; Cowan, Catharine R.; Mih, Justin D.; Koryakina, Yulia; Gioeli, Daniel; Slack-Davis, Jill K.; Blackman, Brett R.; Tschumperlin, Daniel J.; Parsons, J. Thomas

    2010-01-01

    Background The mechanical properties of the extracellular matrix have an important role in cell growth and differentiation. However, it is unclear as to what extent cancer cells respond to changes in the mechanical properties (rigidity/stiffness) of the microenvironment and how this response varies among cancer cell lines. Methodology/Principal Findings In this study we used a recently developed 96-well plate system that arrays extracellular matrix-conjugated polyacrylamide gels that increase in stiffness by at least 50-fold across the plate. This plate was used to determine how changes in the rigidity of the extracellular matrix modulate the biological properties of tumor cells. The cell lines tested fall into one of two categories based on their proliferation on substrates of differing stiffness: “rigidity dependent” (those which show an increase in cell growth as extracellular rigidity is increased), and “rigidity independent” (those which grow equally on both soft and stiff substrates). Cells which grew poorly on soft gels also showed decreased spreading and migration under these conditions. More importantly, seeding the cell lines into the lungs of nude mice revealed that the ability of cells to grow on soft gels in vitro correlated with their ability to grow in a soft tissue environment in vivo. The lung carcinoma line A549 responded to culture on soft gels by expressing the differentiated epithelial marker E-cadherin and decreasing the expression of the mesenchymal transcription factor Slug. Conclusions/Significance These observations suggest that the mechanical properties of the matrix environment play a significant role in regulating the proliferation and the morphological properties of cancer cells. Further, the multiwell format of the soft-plate assay is a useful and effective adjunct to established 3-dimensional cell culture models. PMID:20886123

  16. Matrigel Basement Membrane Matrix influences expression of microRNAs in cancer cell lines

    International Nuclear Information System (INIS)

    Price, Karina J.; Tsykin, Anna; Giles, Keith M.; Sladic, Rosemary T.; Epis, Michael R.; Ganss, Ruth; Goodall, Gregory J.; Leedman, Peter J.

    2012-01-01

    Highlights: ► Matrigel alters cancer cell line miRNA expression relative to culture on plastic. ► Many identified Matrigel-regulated miRNAs are implicated in cancer. ► miR-1290, -210, -32 and -29b represent a Matrigel-induced miRNA signature. ► miR-32 down-regulates Integrin alpha 5 (ITGA5) mRNA. -- Abstract: Matrigel is a medium rich in extracellular matrix (ECM) components used for three-dimensional cell culture and is known to alter cellular phenotypes and gene expression. microRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression and have roles in cancer. While miRNA profiles of numerous cell lines cultured on plastic have been reported, the influence of Matrigel-based culture on cancer cell miRNA expression is largely unknown. This study investigated the influence of Matrigel on the expression of miRNAs that might facilitate ECM-associated cancer cell growth. We performed miRNA profiling by microarray using two colon cancer cell lines (SW480 and SW620), identifying significant differential expression of miRNAs between cells cultured in Matrigel and on plastic. Many of these miRNAs have previously been implicated in cancer-related processes. A common Matrigel-induced miRNA signature comprised of up-regulated miR-1290 and miR-210 and down-regulated miR-29b and miR-32 was identified using RT-qPCR across five epithelial cancer cell lines (SW480, SW620, HT-29, A549 and MDA-MB-231). Experimental modulation of these miRNAs altered expression of their known target mRNAs involved in cell adhesion, proliferation and invasion, in colon cancer cell lines. Furthermore, ITGA5 was identified as a novel putative target of miR-32 that may facilitate cancer cell interactions with the ECM. We propose that culture of cancer cell lines in Matrigel more accurately recapitulates miRNA expression and function in cancer than culture on plastic and thus is a valuable approach to the in vitro study of miRNAs.

  17. Usability and Applicability of Microfluidic Cell Culture Systems

    DEFF Research Database (Denmark)

    Hemmingsen, Mette

    possibilities for, for example, precise control of the chemical environment, 3D cultures, controlled co-culture of different cell types or automated, individual control of up to 96 cell culture chambers in one integrated system. Despite the great new opportunities to perform novel experimental designs......Microfluidic cell culture has been a research area with great attention the last decade due to its potential to mimic the in vivo cellular environment more closely compared to what is possible by conventional cell culture methods. Many exciting and complex devices have been presented providing......, these devices still lack general implementation into biological research laboratories. In this project, the usability and applicability of microfluidic cell culture systems have been investigated. The tested systems display good properties regarding optics and compatibility with standard laboratory equipment...

  18. Cardiac Glycoside Glucoevatromonoside Induces Cancer Type-Specific Cell Death

    Directory of Open Access Journals (Sweden)

    Naira F. Z. Schneider

    2018-03-01

    Full Text Available Cardiac glycosides (CGs are natural compounds used traditionally to treat congestive heart diseases. Recent investigations repositioned CGs as potential anticancer agents. To discover novel cytotoxic CG scaffolds, we selected the cardenolide glucoevatromonoside (GEV out of 46 CGs for its low nanomolar anti-lung cancer activity. GEV presented reduced toxicity toward non-cancerous cell types (lung MRC-5 and PBMC and high-affinity binding to the Na+/K+-ATPase α subunit, assessed by computational docking. GEV-induced cell death was caspase-independent, as investigated by a multiparametric approach, and culminates in severe morphological alterations in A549 cells, monitored by transmission electron microscopy, live cell imaging and flow cytometry. This non-canonical cell death was not preceded or accompanied by exacerbation of autophagy. In the presence of GEV, markers of autophagic flux (e.g. LC3I-II conversion were impacted, even in presence of bafilomycin A1. Cell death induction remained unaffected by calpain, cathepsin, parthanatos, or necroptosis inhibitors. Interestingly, GEV triggered caspase-dependent apoptosis in U937 acute myeloid leukemia cells, witnessing cancer-type specific cell death induction. Differential cell cycle modulation by this CG led to a G2/M arrest, cyclin B1 and p53 downregulation in A549, but not in U937 cells. We further extended the anti-cancer potential of GEV to 3D cell culture using clonogenic and spheroid formation assays and validated our findings in vivo by zebrafish xenografts. Altogether, GEV shows an interesting anticancer profile with the ability to exert cytotoxic effects via induction of different cell death modalities.

  19. THE ALKALOID CYTISINE IN THE CELL CULTURE

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    Gazaliev A.M.

    2012-08-01

    Full Text Available Alkaloids are vegetative establishments of complex and original structure with nitrous heterocycles in the basis. For a long time they drew researchers’ attention because of their unique and specific physiological effect on alive organisms. Not all the representatives of the globe’s flora contain these unique substances. Alkaloid cytisine is to be found mainly in the plants of the fabaceous family - Fabaceae. For the cytisine production the seeds of Thermopsis lanceolata R.Br (T. lanceolata R.Br and Cytisus laburnum (C. laburnum are used as a raw material. The object of the research is T. lanceolata cell culture. Sterile sprouts are used at the first stage of the experiment. Callus genesis is accompanied with dedifferentiation. It leads to the cellular organization simplification. Based on an important property of a plant cell, such as totipotency, there appears the formation of the “de novo” biosynthetic device. The cultivation algorithm consists of two basic stages: (i the cultivation conditions optimization of callus with a high level of the primary metabolites biosynthesis (Aspartat – lysine; (ii the research of cultivation chemical and physical factors influence on the secondary metabolite (cytisine biosynthesis and accumulation. During the cultivation the Murashige and Skoog classical recipe of nutrient medium will be used. Optimization of the cultivation conditions will concern the phytohormones, macro- and micronutrients content, as the purpose of optimization is the production of the determined high-level competence embriogenical callus. The main problem is genetic heterogeneity of a cellular population and instability of morpho-physiological processes. The correct management of higher plants cells population is possible at the synchronization of a cellular cycle phases. The references analysis has shown that it is almost impossible to synchronize cellular cycles in the culture of plant tissue. The application of chemical

  20. Isolation and culture of larval cells from C. elegans.

    Directory of Open Access Journals (Sweden)

    Sihui Zhang

    Full Text Available Cell culture is an essential tool to study cell function. In C. elegans the ability to isolate and culture cells has been limited to embryonically derived cells. However, cells or blastomeres isolated from mixed stage embryos terminally differentiate within 24 hours of culture, thus precluding post-embryonic stage cell culture. We have developed an efficient and technically simple method for large-scale isolation and primary culture of larval-stage cells. We have optimized the treatment to maximize cell number and minimize cell death for each of the four larval stages. We obtained up to 7.8×10(4 cells per microliter of packed larvae, and up to 97% of adherent cells isolated by this method were viable for at least 16 hours. Cultured larval cells showed stage-specific increases in both cell size and multinuclearity and expressed lineage- and cell type-specific reporters. The majority (81% of larval cells isolated by our method were muscle cells that exhibited stage-specific phenotypes. L1 muscle cells developed 1 to 2 wide cytoplasmic processes, while L4 muscle cells developed 4 to 14 processes of various thicknesses. L4 muscle cells developed bands of myosin heavy chain A thick filaments at the cell center and spontaneously contracted ex vivo. Neurons constituted less than 10% of the isolated cells and the majority of neurons developed one or more long, microtubule-rich protrusions that terminated in actin-rich growth cones. In addition to cells such as muscle and neuron that are high abundance in vivo, we were also able to isolate M-lineage cells that constitute less than 0.2% of cells in vivo. Our novel method of cell isolation extends C. elegans cell culture to larval developmental stages, and allows use of the wealth of cell culture tools, such as cell sorting, electrophysiology, co-culture, and high-resolution imaging of subcellular dynamics, in investigation of post-embryonic development and physiology.

  1. Efficient generation of patient-matched malignant and normal primary cell cultures from clear cell renal cell carcinoma patients: clinically relevant models for research and personalized medicine

    International Nuclear Information System (INIS)

    Lobo, Nazleen C.; Gedye, Craig; Apostoli, Anthony J.; Brown, Kevin R.; Paterson, Joshua; Stickle, Natalie; Robinette, Michael; Fleshner, Neil; Hamilton, Robert J.; Kulkarni, Girish; Zlotta, Alexandre; Evans, Andrew; Finelli, Antonio; Moffat, Jason; Jewett, Michael A. S.; Ailles, Laurie

    2016-01-01

    Patients with clear cell renal cell carcinoma (ccRCC) have few therapeutic options, as ccRCC is unresponsive to chemotherapy and is highly resistant to radiation. Recently targeted therapies have extended progression-free survival, but responses are variable and no significant overall survival benefit has been achieved. Commercial ccRCC cell lines are often used as model systems to develop novel therapeutic approaches, but these do not accurately recapitulate primary ccRCC tumors at the genomic and transcriptional levels. Furthermore, ccRCC exhibits significant intertumor genetic heterogeneity, and the limited cell lines available fail to represent this aspect of ccRCC. Our objective was to generate accurate preclinical in vitro models of ccRCC using tumor tissues from ccRCC patients. ccRCC primary single cell suspensions were cultured in fetal bovine serum (FBS)-containing media or defined serum-free media. Established cultures were characterized by genomic verification of mutations present in the primary tumors, expression of renal epithelial markers, and transcriptional profiling. The apparent efficiency of primary cell culture establishment was high in both culture conditions, but genotyping revealed that the majority of cultures contained normal, not cancer cells. ccRCC characteristically shows biallelic loss of the von Hippel Lindau (VHL) gene, leading to accumulation of hypoxia-inducible factor (HIF) and expression of HIF target genes. Purification of cells based on expression of carbonic anhydrase IX (CA9), a cell surface HIF target, followed by culture in FBS enabled establishment of ccRCC cell cultures with an efficiency of >80 %. Culture in serum-free conditions selected for growth of normal renal proximal tubule epithelial cells. Transcriptional profiling of ccRCC and matched normal cell cultures identified up- and down-regulated networks in ccRCC and comparison to The Cancer Genome Atlas confirmed the clinical validity of our cell cultures. The ability

  2. Agarose hydrogel induced MCF-7 and BMG-1 cell line progressive 3D and 3D revert cultures.

    Science.gov (United States)

    Subramaniyan, Aishwarya; Ravi, Maddaly

    2018-04-01

    3D culture systems have enhanced the utility of cancer cell lines as they are considered closer to the in vivo systems. A variety of changes are induced in cells cultured in 3D systems; an apparent and striking feature being the spontaneous acquisition of distinct morphological entities. 3D reverts (3DRs) can be obtained by introducing 3D aggregates in scaffold/matrix-free culture units. It could be seen that the two cell lines used in this study exhibited differences in 3DR structures, though both were cultured on agarose hydrogels. Also, differences in 3DR formation, growth and survival were different. While 3D aggregates of several cell lines have been reported for a variety of studies, there are no studies that describe or utilize 3DRs. 3DRs can provide insights into complex events that can occur in cancer cells; especially as material to study metastasis, migration, and invasion. © 2017 Wiley Periodicals, Inc.

  3. Apoptotic induction of skin cancer cell death by plant extracts.

    Science.gov (United States)

    Thuncharoen, Walairat; Chulasiri, Malin; Nilwarangkoon, Sirinun; Nakamura, Yukio; Watanapokasin, Ramida

    2013-01-01

    The aim of the present study was to investigate the effects of plant extracts on cancer apoptotic induction. Human epidermoid carcinoma A431 cell line, obtained from the American Type Culture Collection (ATCC, Manassas, VA), was maintained in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) at 37 degrees C, 5% carbon dioxide (CO2). Plant extract solutions were obtained from S & J international enterprises public company limited. These plant extracts include 50% hydroglycol extracts from Etlingera elatior (Jack) R.M.Smith (torch ginger; EE), Rosa damascene (damask rose; DR) and Rafflesia kerrii Meijer (bua phut; RM). The cell viability, time and dose dependency were determined by MTT (3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assay. A431 cells were treated with the plant extracts and stained with Hoechst 33342 fluorescent staining dye. Cell viability was demonstrated by the inhibitory concentration 50% (IC50). The anti-proliferative effects were shown to be dependent on time and dose. Typical characteristics of apoptosis which are cell morphological changes and chromatin condensation were clearly observed. The plant extracts was shown to be effective for anti-proliferation and induction of apoptosis cell death in skin cancer cells. Therefore, mechanisms underlying the cell death and its potential use for treatment of skin cancer will be further studied.

  4. Physical View on the Interactions Between Cancer Cells and the Endothelial Cell Lining During Cancer Cell Transmigration and Invasion

    Science.gov (United States)

    Mierke, Claudia T.

    There exist many reviews on the biological and biochemical interactions of cancer cells and endothelial cells during the transmigration and tissue invasion of cancer cells. For the malignant progression of cancer, the ability to metastasize is a prerequisite. In particular, this means that certain cancer cells possess the property to migrate through the endothelial lining into blood or lymph vessels, and are possibly able to transmigrate through the endothelial lining into the connective tissue and follow up their invasion path in the targeted tissue. On the molecular and biochemical level the transmigration and invasion steps are well-defined, but these signal transduction pathways are not yet clear and less understood in regards to the biophysical aspects of these processes. To functionally characterize the malignant transformation of neoplasms and subsequently reveal the underlying pathway(s) and cellular properties, which help cancer cells to facilitate cancer progression, the biomechanical properties of cancer cells and their microenvironment come into focus in the physics-of-cancer driven view on the metastasis process of cancers. Hallmarks for cancer progression have been proposed, but they still lack the inclusion of specific biomechanical properties of cancer cells and interacting surrounding endothelial cells of blood or lymph vessels. As a cancer cell is embedded in a special environment, the mechanical properties of the extracellular matrix also cannot be neglected. Therefore, in this review it is proposed that a novel hallmark of cancer that is still elusive in classical tumor biological reviews should be included, dealing with the aspect of physics in cancer disease such as the natural selection of an aggressive (highly invasive) subtype of cancer cells displaying a certain adhesion or chemokine receptor on their cell surface. Today, the physical aspects can be analyzed by using state-of-the-art biophysical methods. Thus, this review will present

  5. In Situ Gelation-Induced Death of Cancer Cells Based on Proteinosomes.

    Science.gov (United States)

    Zhou, Yuting; Song, Jianmin; Wang, Lei; Xue, Xuting; Liu, Xiaoman; Xie, Hui; Huang, Xin

    2017-08-14

    Hydrogels are an excellent type of material that can be utilized as a platform for cell culture. However, when a bulky hydrogel forms on the inside of cancer cells, the result would be different. In this study, we demonstrate a method for in situ gelation inside cancer cells that can efficiently induce cell death. Glutathione-responsive proteinosomes with good biocompatibility were prepared as carriers for sodium alginate to be endocytosed by cancer cells, where the chelation between sodium alginate and free calcium ions in the culture medium occurs during the diffusion process. The uptake of the hydrogel-loaded proteinosomes into the cancer cells, and then the triggered release of hydrogel with concomitant aggregation, was well-confirmed by monitoring the change of the Young's modulus of the cells based on AFM force measurements. Accordingly, when a large amount of hydrogel formed in cells, the cell viability would be inhibited by ∼90% by MTT assay at a concentration of 5.0 μM of hydrogel-loaded proteinosomes after 48 h incubation, which clearly proves the feasibility of the demonstrated method for killing cancer cells. Although more details regarding the mechanism of cell death should be conducted in the near future, such a demonstrated method of in situ gelation inside cells provides another choice for killing cancer cells.

  6. Biogelx: Cell Culture on Self-Assembling Peptide Gels.

    Science.gov (United States)

    Harper, Mhairi M; Connolly, Michael L; Goldie, Laura; Irvine, Eleanore J; Shaw, Joshua E; Jayawarna, Vineetha; Richardson, Stephen M; Dalby, Matthew J; Lightbody, David; Ulijn, Rein V

    2018-01-01

    Aromatic peptide amphiphiles can form self-supporting nanostructured hydrogels with tunable mechanical properties and chemical compositions. These hydrogels are increasingly applied in two-dimensional (2D) and three-dimensional (3D) cell culture, where there is a rapidly growing need to store, grow, proliferate, and manipulate naturally derived cells within a hydrated, 3D matrix. Biogelx Limited is a biomaterials company, created to commercialize these bio-inspired hydrogels to cell biologists for a range of cell culture applications. This chapter describes methods of various characterization and cell culture techniques specifically optimized for compatibility with Biogelx products.

  7. Development of a microfluidic perfusion 3D cell culture system

    Science.gov (United States)

    Park, D. H.; Jeon, H. J.; Kim, M. J.; Nguyen, X. D.; Morten, K.; Go, J. S.

    2018-04-01

    Recently, 3-dimensional in vitro cell cultures have gained much attention in biomedical sciences because of the closer relevance between in vitro cell cultures and in vivo environments. This paper presents a microfluidic perfusion 3D cell culture system with consistent control of long-term culture conditions to mimic an in vivo microenvironment. It consists of two sudden expansion reservoirs to trap incoming air bubbles, gradient generators to provide a linear concentration, and microchannel mixers. Specifically, the air bubbles disturb a flow in the microfluidic channel resulting in the instability of the perfusion cell culture conditions. For long-term stable operation, the sudden expansion reservoir is designed to trap air bubbles by using buoyancy before they enter the culture system. The performance of the developed microfluidic perfusion 3D cell culture system was examined experimentally and compared with analytical results. Finally, it was applied to test the cytotoxicity of cells infected with Ewing’s sarcoma. Cell death was observed for different concentrations of H2O2. For future work, the developed microfluidic perfusion 3D cell culture system can be used to examine the behavior of cells treated with various drugs and concentrations for high-throughput drug screening.

  8. Slug/SNAI2 regulates cell proliferation and invasiveness of metastatic prostate cancer cell lines.

    Science.gov (United States)

    Emadi Baygi, Modjtaba; Soheili, Zahra-Soheila; Essmann, Frank; Deezagi, Abdolkhaleg; Engers, Rainer; Goering, Wolfgang; Schulz, Wolfgang A

    2010-08-01

    Many metastatic cancers recapitulate the epithelial-to-mesenchymal transition (EMT) resulting in enhanced cell motility and invasiveness. The EMT is regulated by several transcription factors, including the zinc finger protein SNAI2, also named Slug, which appears to exert additional functions during development and cancer progression. We have studied the function of SNAI2 in prostate cancer cells. Quantitative RT-PCR analysis showed strong SNAI2 expression particularly in the PC-3 and PC3-16 prostate carcinoma cell lines. Knockdown of SNAI2 by specific siRNA induced changes in EMT markers and inhibited invasion of both cell lines into a matrigel matrix. SNAI2 siRNA-treated cells did not tolerate detachment from the culture plates, likely at least in part due to downregulation of integrin alpha6beta4. SNAI2 knockdown disturbed the microtubular and actin cytoskeletons, especially severely in PC-3 cells, resulting in grossly enlarged, flattened, and sometimes multinuclear cells. Knockdown also decreased cell proliferation, with a prominent G0/G1 arrest in PC3-16. Together, our data imply that SNAI2 exerts strong effects on the cytoskeleton and adhesion of those prostate cancer cells that express it and is necessary for their proliferation and invasiveness.

  9. The calcimimetic R-568 induces apoptotic cell death in prostate cancer cells

    Directory of Open Access Journals (Sweden)

    Cheng Guangming

    2009-07-01

    Full Text Available Abstract Background Increased serum level of parathyroid hormone (PTH was found in metastatic prostate cancers. Calcimimetic R-568 was reported to reduce PTH expression, to suppress cell proliferation and to induce apoptosis in parathyroid cells. In this study, we investigated the effect of R-568 on cellular survival of prostate cancer cells. Methods Prostate cancer cell lines LNCaP and PC-3 were used in this study. Cellular survival was determined with MTT, trypan blue exclusion and fluorescent Live/Death assays. Western blot assay was utilized to assess apoptotic events induced by R-568 treatment. JC-1 staining was used to evaluate mitochondrial membrane potential. Results In cultured prostate cancer LNCaP and PC-3 cells, R-568 treatment significantly reduced cellular survival in a dose- and time-dependent manner. R-568-induced cell death was an apoptotic event, as evidenced by caspase-3 processing and PARP cleavage, as well as JC-1 color change in mitochondria. Knocking down calcium sensing receptor (CaSR significantly reduced R-568-induced cytotoxicity. Enforced expression of Bcl-xL gene abolished R-568-induced cell death, while loss of Bcl-xL expression led to increased cell death in R-568-treated LNCaP cells,. Conclusion Taken together, our data demonstrated that calcimimetic R-568 triggers an intrinsic mitochondria-related apoptotic pathway, which is dependent on the CaSR and is modulated by Bcl-xL anti-apoptotic pathway.

  10. Plasma-activated medium (PAM) kills human cancer-initiating cells.

    Science.gov (United States)

    Ikeda, Jun-Ichiro; Tanaka, Hiromasa; Ishikawa, Kenji; Sakakita, Hajime; Ikehara, Yuzuru; Hori, Masaru

    2018-01-01

    Medical non-thermal plasma (NTP) treatments for various types of cancers have been reported. Cells with tumorigenic potential (cancer-initiating cells; CICs) are few in number in many types of tumors. CICs efficiently eliminate anti-cancer chemicals and exhibit high-level aldehyde dehydrogenase (ALDH) activity. We previously examined the effects of direct irradiation via NTP on cancer cells; even though we targeted CICs expressing high levels of ALDH, such treatment affected both non-CICs and CICs. Recent studies have shown that plasma-activated medium (PAM) (culture medium irradiated by NTP) selectively induces apoptotic death of cancer but not normal cells. Therefore, we explored the anti-cancer effects of PAM on CICs among endometrioid carcinoma and gastric cancer cells. PAM reduced the viability of cells expressing both low and high levels of ALDH. Combined PAM/cisplatin appeared to kill cancer cells more efficiently than did PAM or cisplatin alone. In a mouse tumor xenograft model, PAM exerted an anti-cancer effect on CICs. Thus, our results suggest that PAM effectively kills both non-CICs and CICs, as does NTP. Therefore, PAM may be a useful new anti-cancer therapy, targeting various cancer cells including CICs. © 2017 Japanese Society of Pathology and John Wiley & Sons Australia, Ltd.

  11. Magnetic Nanowires as Materials for Cancer Cell Destruction

    KAUST Repository

    Contreras, Maria F.

    2015-12-01

    Current cancer therapies are highly cytotoxic and their delivery to exclusively the affected site is poorly controlled, resulting in unavoidable and often severe side effects. In an effort to overcome such issues, magnetic nanoparticles have been recently gaining relevance in the areas of biomedical applications and therapeutics, opening pathways to alternative methods. This led to the concept of magnetic particle hyperthermia in which magnetic nano beads are heated by a high power magnetic field. The increase in temperature kills the cancer cells, which are more susceptible to heat in comparison to healthy cells. In this dissertation, the possibility to kill cancer cells with magnetic nanowires is evaluated. The idea is to exploit a magnetomechanical effect, where nanowires cause cancer cell death through vibrating in a low power magnetic field. Specifically, the magnetic nanowires effects to cells in culture and their ability to induce cancer cell death, when combined with an alternating magnetic field, was investigated. Nickel and iron nanowires of 35 nm diameter and 1 to 5 μm long were synthesized by electrodeposition into nanoporous alumina templates, which were prepared using a two-step anodization process on highly pure aluminum substrates. For the cytotoxicity studies, the nanowires were added to cancer cells in culture, varying the incubation time and the concentration. The cell-nanowire interaction was thoroughly studied at the cellular level (mitochondrial metabolic activity, cell membrane integrity and, apoptosis/necrosis assay), and optical level (transmission electron and confocal microscopy). Furthermore, to investigate their therapeutic potential, an alternating magnetic field was applied varying its intensity and frequency. After the magnetic field application, cells health was measured at the mitochondrial activity level. Cytotoxicity results shed light onto the cellular tolerance to the nanowires, which helped in establishing the appropriate

  12. Radiolabeling and in vitro evaluation of 99mTc-methotrexate on breast cancer cell line

    International Nuclear Information System (INIS)

    Emre Ozgenc; Meliha Ekinci; Derya Ilem-Ozdemir; Evren Gundogdu; Makbule Asikoglu

    2016-01-01

    In the present study 99m Tc-MTX was prepared with high labeling yield by a new simple and easy formulation method. According to cell culture studies, 99m Tc- MTX incorporated with both MCF-7 and CRL8798 cells, with significant differences in the uptake percentages. Since 99m Tc-MTX highly uptake in cancer cell line, the results demonstrated that radiolabeled MTX may be promising for breast cancer diagnosis of oncological patients. (author)

  13. Systems Biology for Organotypic Cell Cultures

    Energy Technology Data Exchange (ETDEWEB)

    Grego, Sonia [RTI International, Research Triangle Park, NC (United States); Dougherty, Edward R. [Texas A & M Univ., College Station, TX (United States); Alexander, Francis J. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Auerbach, Scott S. [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Berridge, Brian R. [GlaxoSmithKline, Research Triangle Park, NC (United States); Bittner, Michael L. [Translational Genomics Research Inst., Phoenix, AZ (United States); Casey, Warren [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Cooley, Philip C. [RTI International, Research Triangle Park, NC (United States); Dash, Ajit [HemoShear Therapeutics, Charlottesville, VA (United States); Ferguson, Stephen S. [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Fennell, Timothy R. [RTI International, Research Triangle Park, NC (United States); Hawkins, Brian T. [RTI International, Research Triangle Park, NC (United States); Hickey, Anthony J. [RTI International, Research Triangle Park, NC (United States); Kleensang, Andre [Johns Hopkins Univ., Baltimore, MD (United States). Center for Alternatives to Animal Testing; Liebman, Michael N. [IPQ Analytics, Kennett Square, PA (United States); Martin, Florian [Phillip Morris International, Neuchatel (Switzerland); Maull, Elizabeth A. [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Paragas, Jason [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Qiao, Guilin [Defense Threat Reduction Agency, Ft. Belvoir, VA (United States); Ramaiahgari, Sreenivasa [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Sumner, Susan J. [RTI International, Research Triangle Park, NC (United States); Yoon, Miyoung [The Hamner Inst. for Health Sciences, Research Triangle Park, NC (United States); ScitoVation, Research Triangle Park, NC (United States)

    2016-08-04

    Translating in vitro biological data into actionable information related to human health holds the potential to improve disease treatment and risk assessment of chemical exposures. While genomics has identified regulatory pathways at the cellular level, translation to the organism level requires a multiscale approach accounting for intra-cellular regulation, inter-cellular interaction, and tissue/organ-level effects. Tissue-level effects can now be probed in vitro thanks to recently developed systems of three-dimensional (3D), multicellular, “organotypic” cell cultures, which mimic functional responses of living tissue. However, there remains a knowledge gap regarding interactions across different biological scales, complicating accurate prediction of health outcomes from molecular/genomic data and tissue responses. Systems biology aims at mathematical modeling of complex, non-linear biological systems. We propose to apply a systems biology approach to achieve a computational representation of tissue-level physiological responses by integrating empirical data derived from organotypic culture systems with computational models of intracellular pathways to better predict human responses. Successful implementation of this integrated approach will provide a powerful tool for faster, more accurate and cost-effective screening of potential toxicants and therapeutics. On September 11, 2015, an interdisciplinary group of scientists, engineers, and clinicians gathered for a workshop in Research Triangle Park, North Carolina, to discuss this ambitious goal. Participants represented laboratory-based and computational modeling approaches to pharmacology and toxicology, as well as the pharmaceutical industry, government, non-profits, and academia. Discussions focused on identifying critical system perturbations to model, the computational tools required, and the experimental approaches best suited to generating key data. This consensus report summarizes the discussions held.

  14. Mechanisms of Cancer Cell Dormancy--Another Hallmark of Cancer?

    Science.gov (United States)

    Yeh, Albert C; Ramaswamy, Sridhar

    2015-12-01

    Disease relapse in cancer patients many years after clinical remission, often referred to as cancer dormancy, is well documented but remains an incompletely understood phenomenon on the biologic level. Recent reviews have summarized potential models that can explain this phenomenon, including angiogenic, immunologic, and cellular dormancy. We focus on mechanisms of cellular dormancy as newer biologic insights have enabled better understanding of this process. We provide a historical context, synthesize current advances in the field, and propose a mechanistic framework that treats cancer cell dormancy as a dynamic cell state conferring a fitness advantage to an evolving malignancy under stress. Cellular dormancy appears to be an active process that can be toggled through a variety of signaling mechanisms that ultimately downregulate the RAS/MAPK and PI(3)K/AKT pathways, an ability that is preserved even in cancers that constitutively depend on these pathways for their growth and survival. Just as unbridled proliferation is a key hallmark of cancer, the ability of cancer cells to become quiescent may be critical to evolving malignancies, with implications for understanding cancer initiation, progression, and treatment resistance. ©2015 American Association for Cancer Research.

  15. A method for culturing human hair follicle cells.

    Science.gov (United States)

    Weterings, P J; Vermorken, A J; Bloemendal, H

    1981-01-01

    For the first time a method for culturing human hair follicle cells is described. The bovine eye lens capsule, a basement membrane-like structure, is used as the substrate for the cultures. In a culture medium supplemented with hydrocortisone and insulin about 70% of the original follicles will form growing colonies of diploid keratinocytes.

  16. Are Mast Cells MASTers in Cancer?

    Science.gov (United States)

    Varricchi, Gilda; Galdiero, Maria Rosaria; Loffredo, Stefania; Marone, Giancarlo; Iannone, Raffaella; Marone, Gianni; Granata, Francescopaolo

    2017-01-01

    Prolonged low-grade inflammation or smoldering inflammation is a hallmark of cancer. Mast cells form a heterogeneous population of immune cells with differences in their ultra-structure, morphology, mediator content, and surface receptors. Mast cells are widely distributed throughout all tissues and are stromal components of the inflammatory microenvironment that modulates tumor initiation and development. Although canonically associated with allergic disorders, mast cells are a major source of pro-tumorigenic (e.g., angiogenic and lymphangiogenic factors) and antitumorigenic molecules (e.g., TNF-α and IL-9), depending on the milieu. In certain neoplasias (e.g., gastric, thyroid and Hodgkin's lymphoma) mast cells play a pro-tumorigenic role, in others (e.g., breast cancer) a protective role, whereas in yet others they are apparently innocent bystanders. These seemingly conflicting results suggest that the role of mast cells and their mediators could be cancer specific. The microlocalization (e.g., peritumoral vs intratumoral) of mast cells is another important aspect in the initiation/progression of solid and hematologic tumors. Increasing evidence in certain experimental models indicates that targeting mast cells and/or their mediators represent a potential therapeutic target in cancer. Thus, mast cells deserve focused consideration also as therapeutic targets in different types of tumors. There are many unanswered questions that should be addressed before we understand whether mast cells are an ally, adversary, or innocent bystanders in human cancers.

  17. Characterization of aldehyde dehydrogenase isozymes in ovarian cancer tissues and sphere cultures

    Directory of Open Access Journals (Sweden)

    Saw Yu-Ting

    2012-08-01

    Full Text Available Abstract Background Aldehyde dehydrogenases belong to a superfamily of detoxifying enzymes that protect cells from carcinogenic aldehydes. Of the superfamily, ALDH1A1 has gained most attention because current studies have shown that its expression is associated with human cancer stem cells. However, ALDH1A1 is only one of the 19 human ALDH subfamilies currently known. The purpose of the present study was to determine if the expression and activities of other major ALDH isozymes are associated with human ovarian cancer and ovarian cancer sphere cultures. Methods Immunohistochemistry was used to delineate ALDH isozyme localization in clinical ovarian tissues. Western Blot analyses were performed on lysates prepared from cancer cell lines and ovarian cancer spheres to confirm the immunohistochemistry findings. Quantitative reverse transcription-polymerase chain reactions were used to measure the mRNA expression levels. The Aldefluor® assay was used to measure ALDH activity in cancer cells from the four tumor subtypes. Results Immunohistochemical staining showed significant overexpression of ALDH1A3, ALDH3A2, and ALDH7A1 isozymes in ovarian tumors relative to normal ovarian tissues. The expression and activity of ALDH1A1 is tumor type-dependent, as seen from immunohistochemisty, Western blot analysis, and the Aldefluor® assay. The expression was elevated in the mucinous and endometrioid ovarian epithelial tumors than in serous and clear cell tumors. In some serous and most clear cell tumors, ALDH1A1 expression was found in the stromal fibroblasts. RNA expression of all studied ALDH isozymes also showed higher expression in endometrioid and mucinous tumors than in the serous and clear cell subtypes. The expression of ALDH enzymes showed tumor type-dependent induction in ovarian cancer cells growing as sphere suspensions in serum-free medium. Conclusions The results of our study indicate that ALDH enzyme expression and activity may be associated

  18. Characterization of aldehyde dehydrogenase isozymes in ovarian cancer tissues and sphere cultures

    International Nuclear Information System (INIS)

    Saw, Yu-Ting; Thompson, David; Vasiliou, Vasilis; Berkowitz, Ross S; Ng, Shu-Wing; Yang, Junzheng; Ng, Shu-Kay; Liu, Shubai; Singh, Surendra; Singh, Margit; Welch, William R; Tsuda, Hiroshi; Fong, Wing-Ping

    2012-01-01

    Aldehyde dehydrogenases belong to a superfamily of detoxifying enzymes that protect cells from carcinogenic aldehydes. Of the superfamily, ALDH1A1 has gained most attention because current studies have shown that its expression is associated with human cancer stem cells. However, ALDH1A1 is only one of the 19 human ALDH subfamilies currently known. The purpose of the present study was to determine if the expression and activities of other major ALDH isozymes are associated with human ovarian cancer and ovarian cancer sphere cultures. Immunohistochemistry was used to delineate ALDH isozyme localization in clinical ovarian tissues. Western Blot analyses were performed on lysates prepared from cancer cell lines and ovarian cancer spheres to confirm the immunohistochemistry findings. Quantitative reverse transcription-polymerase chain reactions were used to measure the mRNA expression levels. The Aldefluor® assay was used to measure ALDH activity in cancer cells from the four tumor subtypes. Immunohistochemical staining showed significant overexpression of ALDH1A3, ALDH3A2, and ALDH7A1 isozymes in ovarian tumors relative to normal ovarian tissues. The expression and activity of ALDH1A1 is tumor type-dependent, as seen from immunohistochemisty, Western blot analysis, and the Aldefluor® assay. The expression was elevated in the mucinous and endometrioid ovarian epithelial tumors than in serous and clear cell tumors. In some serous and most clear cell tumors, ALDH1A1 expression was found in the stromal fibroblasts. RNA expression of all studied ALDH isozymes also showed higher expression in endometrioid and mucinous tumors than in the serous and clear cell subtypes. The expression of ALDH enzymes showed tumor type-dependent induction in ovarian cancer cells growing as sphere suspensions in serum-free medium. The results of our study indicate that ALDH enzyme expression and activity may be associated with specific cell types in ovarian tumor tissues and vary according to

  19. Sensor Access to the Cellular Microenvironment Using the Sensing Cell Culture Flask

    Directory of Open Access Journals (Sweden)

    Jochen Kieninger

    2018-04-01

    Full Text Available The Sensing Cell Culture Flask (SCCF is a cell culture monitoring system accessing the cellular microenvironment in 2D cell culture using electrochemical microsensors. The system is based on microfabricated sensor chips embedded in standard cell culture flasks. Ideally, the sensor chips could be equipped with any electrochemical sensor. Its transparency allows optical inspection of the cells during measurement. The surface of the sensor chip is in-plane with the flask surface allowing undisturbed cell growth on the sensor chip. A custom developed rack system allows easy usage of multiple flasks in parallel within an incubator. The presented data demonstrates the application of the SCCF with brain tumor (T98G and breast cancer (T-47D cells. Amperometric oxygen sensors were used to monitor cellular respiration with different incubation conditions. Cellular acidification was accessed with potentiometric pH sensors using electrodeposited iridium oxide films. The system itself provides the foundation for electrochemical monitoring systems in 3D cell culture.

  20. Cell cycle regulation in human embryonic stem cells: links to adaptation to cell culture.

    Science.gov (United States)

    Barta, Tomas; Dolezalova, Dasa; Holubcova, Zuzana; Hampl, Ales

    2013-03-01

    Cell cycle represents not only a tightly orchestrated mechanism of cell replication and cell division but it also plays an important role in regulation of cell fate decision. Particularly in the context of pluripotent stem cells or multipotent progenitor cells, regulation of cell fate decision is of paramount importance. It has been shown that human embryonic stem cells (hESCs) show unique cell cycle characteristics, such as short doubling time due to abbreviated G1 phase; these properties change with the onset of differentiation. This review summarizes the current understanding of cell cycle regulation in hESCs. We discuss cell cycle properties as well as regulatory machinery governing cell cycle progression of undifferentiated hESCs. Additionally, we provide evidence that long-term culture of hESCs is accompanied by changes in cell cycle properties as well as configuration of several cell cycle regulatory molecules.

  1. Cancer cell adaptation to chemotherapy

    International Nuclear Information System (INIS)

    Di Nicolantonio, Federica; Johnson, Penny; Somers, Shaw S; Toh, Simon; Higgins, Bernie; Lamont, Alan; Gulliford, Tim; Hurren, Jeremy; Yiangou, Constantinos; Cree, Ian A; Mercer, Stuart J; Knight, Louise A; Gabriel, Francis G; Whitehouse, Pauline A; Sharma, Sanjay; Fernando, Augusta; Glaysher, Sharon; Di Palma, Silvana

    2005-01-01

    Tumor resistance to chemotherapy may be present at the beginning of treatment, develop during treatment, or become apparent on re-treatment of the patient. The mechanisms involved are usually inferred from experiments with cell lines, as studies in tumor-derived cells are difficult. Studies of human tumors show that cells adapt to chemotherapy, but it has been largely assumed that clonal selection leads to the resistance of recurrent tumors. Cells derived from 47 tumors of breast, ovarian, esophageal, and colorectal origin and 16 paired esophageal biopsies were exposed to anticancer agents (cisplatin; 5-fluorouracil; epirubicin; doxorubicin; paclitaxel; irinotecan and topotecan) in short-term cell culture (6 days). Real-time quantitative PCR was used to measure up- or down-regulation of 16 different resistance/target genes, and when tissue was available, immunohistochemistry was used to assess the protein levels. In 8/16 paired esophageal biopsies, there was an increase in the expression of multi-drug resistance gene 1 (MDR1) following epirubicin + cisplatin + 5-fluorouracil (ECF) chemotherapy and this was accompanied by increased expression of the MDR-1 encoded protein, P-gp. Following exposure to doxorubicin in vitro, 13/14 breast carcinomas and 9/12 ovarian carcinomas showed >2-fold down-regulation of topoisomerase IIα (TOPOIIα). Exposure to topotecan in vitro, resulted in >4-fold down-regulation of TOPOIIα in 6/7 colorectal tumors and 8/10 ovarian tumors. This study suggests that up-regulation of resistance genes or down-regulation in target genes may occur rapidly in human solid tumors, within days of the start of treatment, and that similar changes are present in pre- and post-chemotherapy biopsy material. The molecular processes used by each tumor appear to be linked to the drug used, but there is also heterogeneity between individual tumors, even those with the same histological type, in the pattern and magnitude of response to the same drugs. Adaptation

  2. Rabbit uterine epithelial cells: Co-culture with spermatozoa

    International Nuclear Information System (INIS)

    Boice, M.L.

    1988-01-01

    A primary culture of rabbit uterine epithelial cells was established and their effects on sperm function were examined in vitro. Epithelial cells were isolated from uteri of estrous rabbits and cultured on floating collagen gels in phenol red-free medium supplemented with 5% fetal bovine serum. Light microscopy and keratin staining showed that the epithelial cell population established in culture had morphological characteristics similar to that seen in the intact endometrium. Cells were cultured with 3 H-leucine and uptake of label by cells and its incorporation into cellular and secretory proteins determined. When compared to cells cultured for 24-48 h, incorporation of label into cellular protein was lower at 72-96 h, but secretion increased. Estradiol 17-β did not affect label uptake or incorporation, but did enhance proliferation of cells as judged by total DNA content of the cell population. Analysis of proteins in media by sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorography suggested that epithelial and stromal cells synthesis proteins that may be secretory in nature during 72-96 h culture. Twenty-nine to thirty-one h after initiation of epithelial cultures, 1-2 x 10 6 sperm were co-incubated with cells and sperm viability, motility, loss of acrosome and fertilizing ability determined

  3. Fibroblast-Derived Extracellular Matrices: An Alternative Cell Culture System That Increases Metastatic Cellular Properties.

    Directory of Open Access Journals (Sweden)

    Michael T Scherzer

    Full Text Available Poor survival rates from lung cancer can largely be attributed to metastatic cells that invade and spread throughout the body. The tumor microenvironment (TME is composed of multiple cell types, as well as non-cellular components. The TME plays a critical role in the development of metastatic cancers by providing migratory cues and changing the properties of the tumor cells. The Extracellular Matrix (ECM, a main component of the TME, has been shown to change composition during tumor progression, contributing to cancer cell invasion and survival away from the primary cancer site. Although the ECM is well-known to influence the fate of tumor progression, little is known about the molecular mechanisms that are affected by the cancer cell-ECM interactions. It is imperative that these mechanisms are elucidated in order to properly understand and prevent lung cancer dissemination. However, common in vitro studies do not incorporate these interactions into everyday cell culture assays. We have adopted a model that examines decellularized human fibroblast-derived ECM as a 3-dimensional substrate for growth of lung adenocarcinoma cell lines. Here, we have characterized the effect of fibroblast-derived matrices on the properties of various lung-derived epithelial cell lines, including cancerous and non-transformed cells. This work highlights the significance of the cell-ECM interaction and its requirement for incorporation into in vitro experiments. Implementation of a fibroblast-derived ECM as an in vitro technique will provide researchers with an important factor to manipulate to better recreate and study the TME.

  4. Metabolic cooperation between cancer and non-cancerous stromal cells is pivotal in cancer progression.

    Science.gov (United States)

    Lopes-Coelho, Filipa; Gouveia-Fernandes, Sofia; Serpa, Jacinta

    2018-02-01

    The way cancer cells adapt to microenvironment is crucial for the success of carcinogenesis, and metabolic fitness is essential for a cancer cell to survive and proliferate in a certain organ/tissue. The metabolic remodeling in a tumor niche is endured not only by cancer cells but also by non-cancerous cells that share the same microenvironment. For this reason, tumor cells and stromal cells constitute a complex network of signal and organic compound transfer that supports cellular viability and proliferation. The intensive dual-address cooperation of all components of a tumor sustains disease progression and metastasis. Herein, we will detail the role of cancer-associated fibroblasts, cancer-associated adipocytes, and inflammatory cells, mainly monocytes/macrophages (tumor-associated macrophages), in the remodeling and metabolic adaptation of tumors.

  5. The Antimetastatic and Antiangiogenesis Effects of Kefir Water on Murine Breast Cancer Cells.

    Science.gov (United States)

    Zamberi, Nur Rizi; Abu, Nadiah; Mohamed, Nurul Elyani; Nordin, Noraini; Keong, Yeap Swee; Beh, Boon Kee; Zakaria, Zuki Abu Bakar; Nik Abdul Rahman, Nik Mohd Afizan; Alitheen, Noorjahan Banu

    2016-12-01

    Kefir is a unique cultured product that contains beneficial probiotics. Kefir culture from other parts of the world exhibits numerous beneficial qualities such as anti-inflammatory, immunomodulation, and anticancer effects. Nevertheless, kefir cultures from different parts of the world exert different effects because of variation in culture conditions and media. Breast cancer is the leading cancer in women, and metastasis is the major cause of death associated with breast cancer. The antimetastatic and antiangiogenic effects of kefir water made from kefir grains cultured in Malaysia were studied in 4T1 breast cancer cells. 4T1 cancer cells were treated with kefir water in vitro to assess its antimigration and anti-invasion effects. BALB/c mice were injected with 4T1 cancer cells and treated orally with kefir water for 28 days. Kefir water was cytotoxic toward 4T1 cells at IC 50 (half-maximal inhibitory concentration) of 12.5 and 8.33 mg/mL for 48 and 72 hours, respectively. A significant reduction in tumor size and weight (0.9132 ± 0.219 g) and a substantial increase in helper T cells (5-fold) and cytotoxic T cells (7-fold) were observed in the kefir water-treated group. Proinflammatory and proangiogenic markers were significantly reduced in the kefir water-treated group. Kefir water inhibited tumor proliferation in vitro and in vivo mainly through cancer cell apoptosis, immunomodulation by stimulating T helper cells and cytotoxic T cells, and anti-inflammatory, antimetastatic, and antiangiogenesis effects. This study brought out the potential of the probiotic beverage kefir water in cancer treatment. © The Author(s) 2016.

  6. Vitamin D compounds inhibit cancer stem-like cells and induce differentiation in triple negative breast cancer.

    Science.gov (United States)

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

    2017-10-01

    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) 2 D 3 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) 2 D 3 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

  7. Metformin and Ara-a Effectively Suppress Brain Cancer by Targeting Cancer Stem/Progenitor Cells

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    Tarek H. Mouhieddine

    2015-11-01

    Full Text Available Background: Gliomas and neuroblastomas pose a great health burden worldwide with a poor and moderate prognosis, respectively. Many studies have tried to find effective treatments for these primary malignant brain tumors. Of interest, the AMP-activated protein kinase (AMPK pathway was found to be associated with tumorigenesis and tumor survival, leading to many studies on AMPK drugs, especially Metformin, and their potential role as anti-cancer treatments. Cancer stem cells (CSCs are a small population of slowly-dividing, treatment-resistant, undifferentiated cancer cells that are being discovered in a multitude of cancers. They are thought to be responsible for replenishing the tumor with highly proliferative cells and increasing the risk of recurrence. Methods: Metformin and 9-β-d-Arabinofuranosyl Adenine (Ara-a were used to study the role of the AMPK pathway in vitro on U251 (glioblastoma and SHSY-5Y (neuroblastoma cell lines.Results: We found that both drugs are able to decrease the survival of U251 and SH-SY5Y cell lines in a 2D as well as a 3D culture model. Metformin and Ara-a significantly decreased the invasive ability of these cancer cell lines. Treatment with these drugs decreased the sphere-forming units (SFU of U251 cells, with Ara-a being more efficient, signifying the extinction of the CSC population. However, if treatment is withdrawn before all SFUs are extinguished, the CSCs regain some of their sphere-forming capabilities in the case of Metformin but not Ara-a treatment. Conclusion: Metformin and Ara-a have proved to be effective in the treatment of glioblastomas and neuroblastomas, in vitro, by targeting their cancer stem/progenitor cell population, which prevents recurrence.

  8. Long-term maintenance of human induced pluripotent stem cells by automated cell culture system.

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    Konagaya, Shuhei; Ando, Takeshi; Yamauchi, Toshiaki; Suemori, Hirofumi; Iwata, Hiroo

    2015-11-17

    Pluripotent stem cells, such as embryonic stem cells and induced pluripotent stem (iPS) cells, are regarded as new sources for cell replacement therapy. These cells can unlimitedly expand under undifferentiated conditions and be differentiated into multiple cell types. Automated culture systems enable the large-scale production of cells. In addition to reducing the time and effort of researchers, an automated culture system improves the reproducibility of cell cultures. In the present study, we newly designed a fully automated cell culture system for human iPS maintenance. Using an automated culture system, hiPS cells maintained their undifferentiated state for 60 days. Automatically prepared hiPS cells had a potency of differentiation into three germ layer cells including dopaminergic neurons and pancreatic cells.

  9. Culture and Drug Profiling of Patient Derived Malignant Pleural Effusions for Personalized Cancer Medicine.

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    Ruiz, Christian; Kustermann, Stefan; Pietilae, Elina; Vlajnic, Tatjana; Baschiera, Betty; Arabi, Leila; Lorber, Thomas; Oeggerli, Martin; Savic, Spasenija; Obermann, Ellen; Singer, Thomas; Rothschild, Sacha I; Zippelius, Alfred; Roth, Adrian B; Bubendorf, Lukas

    2016-01-01

    The use of patients' own cancer cells for in vitro selection of the most promising treatment is an attractive concept in personalized medicine. Human carcinoma cells from malignant pleural effusions (MPEs) are suited for this purpose since they have already adapted to the liquid environment in the patient and do not depend on a stromal cell compartment. Aim of this study was to develop a systematic approach for the in-vitro culture of MPEs to analyze the effect of chemotherapeutic as well as targeted drugs. MPEs from patients with solid tumors were selected for this study. After morphological and molecular characterization, they were cultured in medium supplemented with patient-derived sterile-filtered effusion supernatant. Growth characteristics were monitored in real-time using the xCELLigence system. MPEs were treated with a targeted therapeutic (erlotinib) according to the mutational status or chemotherapeutics based on the recommendation of the oncologists. We have established a robust system for the ex-vivo culture of MPEs and the application of drug tests in-vitro. The use of an antibody based magnetic cell separation system for epithelial cells before culture allowed treatment of effusions with only moderate tumor cell proportion. Experiments using drugs and drug-combinations revealed dose-dependent and specific growth inhibitory effects of targeted drugs. We developed a new approach for the ex-vivo culture of MPEs and the application of drug tests in-vitro using real-time measuring of cell growth, which precisely reproduced the effect of clinically established treatments by standard chemotherapy and targeted drugs. This sets the stage for future studies testing agents against specific targets from genomic profiling of metastatic tumor cells and multiple drug-combinations in a personalized manner.

  10. Culture and Drug Profiling of Patient Derived Malignant Pleural Effusions for Personalized Cancer Medicine.

    Directory of Open Access Journals (Sweden)

    Christian Ruiz

    Full Text Available The use of patients' own cancer cells for in vitro selection of the most promising treatment is an attractive concept in personalized medicine. Human carcinoma cells from malignant pleural effusions (MPEs are suited for this purpose since they have already adapted to the liquid environment in the patient and do not depend on a stromal cell compartment. Aim of this study was to develop a systematic approach for the in-vitro culture of MPEs to analyze the effect of chemotherapeutic as well as targeted drugs.MPEs from patients with solid tumors were selected for this study. After morphological and molecular characterization, they were cultured in medium supplemented with patient-derived sterile-filtered effusion supernatant. Growth characteristics were monitored in real-time using the xCELLigence system. MPEs were treated with a targeted therapeutic (erlotinib according to the mutational status or chemotherapeutics based on the recommendation of the oncologists.We have established a robust system for the ex-vivo culture of MPEs and the application of drug tests in-vitro. The use of an antibody based magnetic cell separation system for epithelial cells before culture allowed treatment of effusions with only moderate tumor cell proportion. Experiments using drugs and drug-combinations revealed dose-dependent and specific growth inhibitory effects of targeted drugs.We developed a new approach for the ex-vivo culture of MPEs and the application of drug tests in-vitro using real-time measuring of cell growth, which precisely reproduced the effect of clinically established treatments by standard chemotherapy and targeted drugs. This sets the stage for future studies testing agents against specific targets from genomic profiling of metastatic tumor cells and multiple drug-combinations in a personalized manner.

  11. Macrolide Antibiotics Exhibit Cytotoxic Effect under Amino Acid-Depleted Culture Condition by Blocking Autophagy Flux in Head and Neck Squamous Cell Carcinoma Cell Lines

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    Hirasawa, Kazuhiro; Moriya, Shota; Miyahara, Kana; Kazama, Hiromi; Hirota, Ayako; Takemura, Jun; Abe, Akihisa; Inazu, Masato; Hiramoto, Masaki; Tsukahara, Kiyoaki

    2016-01-01

    Autophagy, a self-digestive system for cytoplasmic components, is required to maintain the amino acid pool for cellular homeostasis. We previously reported that the macrolide antibiotics azithromycin (AZM) and clarithromycin (CAM) have an inhibitory effect on autophagy flux, and they potently enhance the cytocidal effect of various anticancer reagents in vitro. This suggests that macrolide antibiotics can be used as an adjuvant for cancer chemotherapy. Since cancer cells require a larger metabolic demand than normal cells because of their exuberant growth, upregulated autophagy in tumor cells has now become the target for cancer therapy. In the present study, we examined whether macrolides exhibit cytotoxic effect under an amino acid-starving condition in head and neck squamous cancer cell lines such as CAL 27 and Detroit 562 as models of solid tumors with an upregulated autophagy in the central region owing to hypovascularity. AZM and CAM induced ce